Cocktail compositions containing respiratory antibacterial bacteriophages and their methods of use

Abstract

This invention relates to the field of phage therapy for the treatment and control of bacterial infections, particularly respiratory bacterial infections such as bacterial pneumonia. More specifically, this invention relates to novel phage strains and mixtures thereof, as well as variants thereof; and methods for treating and preventing bacterial infections using them, including respiratory infections caused by, for example, *Pseudomonas aeruginosa* and / or *Klebsiella pneumoniae*. The mixtures can be used alone as pharmaceutical compositions or further in combination with other therapies, such as antibiotics or other standard and non-standard therapies for respiratory infections.

Description

1. Technical Field

[0001] This invention relates to the field of phage therapy for the treatment and control of bacterial infections, particularly respiratory bacterial infections such as bacterial pneumonia. One aspect of the invention relates to a novel phage strain mixture comprising Kle_F391 / 08 (SEQ ID NO:1), Kle_F17 / 19 (SEQ ID NO:2), and Kle_F58 / 19 (SEQ ID NO:3) and variants thereof; and methods of using the mixture to treat and prevent bacterial infections, including respiratory infections caused by, for example, Klebsiella pneumoniae. This mixture can be administered in combination with other phage compositions (including mixtures) targeting the same or different pathogens (e.g., Pseudomonas aeruginosa). The mixture can be used alone as a pharmaceutical composition or further in combination with other therapies, such as antibiotics or other standard and non-standard therapies for respiratory infections. 2. Background Technology

[0002] Bacteriophages (phages) are viruses that specifically infect and lyse bacteria. Phage therapy, a method of treating bacterial infectious diseases using intact bacteriophage viruses, was proposed by Felix d'Herelle in the 1920s. However, with the development of antibiotics in the 1940s, interest in phage-based therapies declined in the Western world. One of the most significant factors contributing to this decline was the lack of standardized testing and manufacturing methods. The failure to establish industry-wide standards for testing phage therapies interfered with the compilation of research results, leading to perceived lack of efficacy and credibility issues regarding the value of phage therapies. Another problem in phage manufacturing involved the purity level of commercially available phage formulations, which sometimes contained unwanted bacterial components, such as endotoxins. Consequently, adverse events were often associated with the formulation, particularly in patients receiving the formulation intravenously.

[0003] However, in Eastern Europe and the Soviet Union, where antibiotic use was restricted, the development and use of phage therapy continued in combination with or as a substitute for antibiotics. Furthermore, with the increase in antibiotic-resistant strains of many bacteria, interest in phage-based therapies re-emerged in the Western world. That is, the prospect that bacteria would eventually develop resistance to new drugs, despite the potential development of new classes of antibiotics, led to a greater inclination to seek non-chemical treatments for controlling, preventing, and treating bacterial infections.

[0004] Phage therapy, particularly phage mixture therapy, offers an alternative to antibiotics for treating bacterial infections, especially respiratory infections, including hospital-acquired bacterial pneumonia (HABP). HABP is an acute lung infection and one of the most common types of infection acquired in intensive care unit settings, and is associated with increased mortality (ranging from 33% to 41%) (Guzman-Herrador B et al., 2014, J Hosp Infect 86(1):53-56). Hospital-acquired lung infections are usually caused by methicillin-resistant Staphylococcus aureus (MRSA), Gram-negative Enterobacteriaceae (such as Klebsiella pneumoniae), or Gram-negative non-Enterobacteriaceae (such as Pseudomonas aeruginosa and Acinetobacter species) (Quartin AA et al., 2013, BMC Infect Dis 13:561-566; and Di Pasuale M et al., 2014, Crit Care Med 42(2):303-312).

[0005] Antibiotic therapy is routinely used in HABP, but treatment options are limited for multidrug-resistant (MDR) bacteria, especially Gram-negative bacteria. No new drug classes have been introduced recently, and the few available options currently include colistin, tigecycline, and fosfomycin. For severe hospital-acquired infections, the choice of antibiotics is very limited (Orsi GB et al., 2011, Expert Rev Anti Infect Ther 9(8):653-679).

[0006] Compared with intravenous antibiotic administration, nebulization of antibiotics allows for greater delivery of antibiotics to the lung parenchyma (Luyt CE et al., 2009, Crit Care 13(6):R200). However, to date, there has been no significant clinical benefit in the use of nebulized antibiotics (such as colistin) for the treatment of pulmonary infections due to the direct toxicity of antibiotics to the airways and lung parenchyma. These side effects include, for example, mucosal irritation, and side effects resulting from the systemic absorption of antibiotics, such as nephrotoxicity of aminoglycosides and polymyxins (Luyt CE et al., 2013, Expert Rev Anti Infect Ther 11(5):511-521; and Quon BS et al., 2014, Ann Am Thorac Soc 11(3):425-434).

[0007] Different studies have attempted to treat bacterial lung infections using phages administered via various routes (Hoe S et al., 2013, J Aerosol Med Pulm Drug Deliv 26:317-335; Morello E et al., 2011, PLoSOne 6(2):e16963; and Debarbieux L et al., 2010, J Infect Dis 201(7):1096-1104). However, there is little publicly available experimental evidence that nebulized phages can cure established infections (Ryan EM et al., 2011, JPharm Pharmacol 63:1253-1264). Previous published studies have not assessed the role of phage nebulization in established infections, with most examining results only hours after infection (Wilson KR et al., 2007, Microbiology 153 (Part 4): 968-979; and Alemayehu D. et al., 2012, MBio 3(2): e00029-12). Furthermore, there are few phage mixtures with antimicrobial activity against different bacteria, likely due to the difficulty in combining the different specificities of phages while maintaining storage stability.

[0008] Therefore, there remains a need to develop novel phage products as therapeutic and / or prophylactic agents to combat pathogenic bacteria, particularly lung bacteria, in vivo. Better treatments, especially nebulized therapy, are also needed for respiratory infections. In particular, there is a need for phage mixtures capable of lysing bacteria that cause hospital-acquired respiratory infections, including *Pseudomonas aeruginosa* and / or *Klebsiella pneumoniae*. This application addresses these and other needs. 3. Summary of the Invention

[0009] This article provides information on novel Klebsiella pneumoniae bacteriophages and their use in the treatment of bacterial infections. Pharmaceutical compositions comprising a mixture of the bacteriophages disclosed herein or a combination of three or more bacteriophages described herein may be used to treat, manage, or prevent bacterial infections, particularly Pseudomonas aeruginosa and / or Klebsiella pneumoniae infections.

[0010] Furthermore, this document provides a novel Klebsiella pneumoniae phage mixture and its use in the treatment of bacterial infections. Pharmaceutical compositions comprising the phage mixture disclosed herein or a combination of three or more phages described herein can be used to treat, manage, or prevent bacterial infections, particularly Pseudomonas aeruginosa and / or Klebsiella pneumoniae infections. Such pharmaceutical compositions can be particularly used to treat, manage, or prevent respiratory infections and can be formulated for pulmonary delivery.

[0011] One aspect of this invention relates to novel Klebsiella pneumoniae phages. This document provides purified Klebsiella pneumoniae phages Kle_F17 / 19 having a genome comprising or consisting of the nucleic acid sequence SEQ ID NO:2, and purified Klebsiella pneumoniae phages Kle_F58 / 19 having a genome comprising or consisting of the nucleic acid sequence SEQ ID NO:3, as well as variants thereof that retain the lytic activity of these phages and possess antibacterial activity against Klebsiella pneumoniae. These phages have been deposited: Kle_F17 / 19 has the NCIMB 43534 accession number, and Kle_F58 / 19 has the NCIMB 43535 accession number. These phages are particularly capable of lysing Klebsiella pneumoniae strains 57 / 15 (NCIMB accession number 43536) and 237 / 14 (NCIMB accession number 43537).

[0012] In one embodiment, the present invention provides an isolated bacteriophage F17 / 19 having a genome containing the nucleic acid sequence SEQ ID NO:2 and exhibiting antibacterial activity against one or more strains of Klebsiella pneumoniae.

[0013] In one embodiment, the present invention provides an isolated bacteriophage F58 / 19 having a genome containing the nucleic acid sequence SEQ ID NO:3 and exhibiting antibacterial activity against one or more strains of Klebsiella pneumoniae.

[0014] Another aspect of the invention relates to compositions comprising three or more different purified phages in a cocktail-like combination. In one aspect, the phage mixture comprises three Klebsiella pneumoniae phages, Kle_F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 (including variants thereof that retain phage lytic activity) and has antibacterial activity against Klebsiella pneumoniae. In other aspects of the invention, such a mixture may further comprise, or be administered in combination with, a mixture of one, two, three, or more other phages (e.g., those against Pseudomonas aeruginosa). In one aspect, a mixture of Kle_F391 / 08 (accession number NCIMB 42918), Kle_F17 / 19, and Kle_F58 / 19 is formulated and / or administered in combination with a mixture of Pseudomonas aeruginosa phages Psa_F99 / 10 (accession number NCIMB 42915), Psa_F27 / 12 (accession number NCIMB 42916), and Psa_F95 / 13 (accession number NCIMB 42917) (disclosed in US2019 / 0290709). In other embodiments, the phage mixture disclosed herein is administered in combination with one or more other purified phages having antibacterial activity against Klebsiella pneumoniae, Pseudomonas aeruginosa, or bacteria other than Klebsiella pneumoniae or Pseudomonas aeruginosa. In a more preferred embodiment, the composition is formulated for administration as an aerosol and for pulmonary delivery.

[0015] Another aspect of the invention relates to pharmaceutical compositions comprising the phage or phage product of the invention and a pharmaceutically acceptable carrier. In some embodiments, the pharmaceutical composition further comprises one or more other phages or phage products having antibacterial activity against Klebsiella pneumoniae. In some embodiments, the composition is formulated into a dosage form in which the amount of phage present provides a multiple of infection (MOI) of about 1 to about 10 when the composition is administered to a subject in need. In some embodiments, each phage is administered at a dose of 10... 9 One PFU to 10 10 Administered at a dose of 10 pfu, and for example at 10 9 One pfu / ml is present in the composition. In a preferred embodiment, the composition is formulated for application as an aerosol.

[0016] Another aspect of the invention relates to a method for treating or reducing the occurrence of or managing bacterial infections in subjects with such need, comprising administering to the subject a therapeutically or preventively effective amount of the pharmaceutical composition of the invention, and relating to the use of the pharmaceutical composition in this respect. In some embodiments, the bacterial infection is caused by a strain of Klebsiella pneumoniae, and in some respects, and / or by a strain of Pseudomonas aeruginosa, including bacterial strains that exhibit resistance to one or more known antibiotics and / or are capable of forming biofilms. In a preferred embodiment, the bacterial infection to be treated or reduced is a respiratory infection, more preferably hospital-acquired bacterial pneumonia or a respiratory infection associated with cystic fibrosis. In a particularly preferred embodiment, the composition is administered to the lungs as an aerosol. In some embodiments, the composition is re-administered about 4, 6, or 8 hours after the initial administration.

[0017] Another aspect of the invention relates to a method for diagnosing pathogenic factors of bacterial infections and / or for assessing whether an infectious factor is susceptible to phage lytic activity, comprising (i) culturing a sample from a patient, such as a swab or sputum, or other sample suitable for culturing bacteria causing an infection; (ii) contacting the culture from step (i) with a phage or phage product disclosed herein; and (iii) monitoring for evidence of culture growth or lysis, wherein evidence of culture lysis indicates that the culture contains bacterial strains known to be susceptible to the phage or phage product used in step (ii). In some embodiments, the sample is a tissue biopsy or swab taken from a patient's respiratory tract. For example, the sample may include bronchoalveolar lavage fluid or bronchial secretions.

[0018] Another aspect of the invention provides a method for reducing or inhibiting bacterial colonization or growth on a surface, comprising contacting the surface with a bacteriophage or bacteriophage product of the invention. In some embodiments, the surface is a mammalian mucosa, preferably a mucosa of the human respiratory tract. In some embodiments, the surface is a non-biological surface, preferably the surface of a medical device or part of a medical device, more preferably the surface of a surgical instrument or part of a surgical device.

[0019] 3.1 Definition

[0020] As used herein, in the context of nucleic acid molecules, the term "isolated" refers to a first nucleic acid molecule that is separated from other nucleic acid molecules present in the natural source of the first nucleic acid molecule. An "isolated nucleic acid molecule," such as an "orf" or phage genome, when produced by recombinant technology, is substantially free of other cellular material or culture medium, or when chemically synthesized, is substantially free of chemical precursors or other chemicals; and may also be free of other DNA or other genomic DNA molecules, for example, when it has been purified and isolated from other clones in a nucleic acid library or from isolated phages. Furthermore, when produced by recombinant technology or isolated from phages, "isolated" genomic DNA is substantially free of other viral or cellular material or culture medium, or when chemically synthesized, is substantially free of chemical precursors or other chemicals, and may also be free of other DNA or other genomic DNA molecules, for example, when it has been purified and isolated from preparations containing other phages or cellular material.

[0021] In the context of bacteriophages, the term "purified" means that the concentration of the bacteriophage has been measurably increased through any purification process, including but not limited to isolation from the environment or culture, such as isolation from the culture after propagation and / or amplification, centrifugation, etc., thereby partially, substantially, almost completely, or completely removing impurities, such as host cells and host cell components. Those skilled in the art will understand the required level of purification for a given application. For example, purified bacteriophages intended for use in therapeutic compositions for human administration typically must have high purity, in accordance with regulatory standards and good manufacturing practices.

[0022] With respect to peptides, polypeptides, fusion proteins, or nucleic acid molecules, the term "purified" means that the concentration of the peptide, polypeptide, fusion protein, or nucleic acid molecule has been measurably increased through any purification process, including but not limited to column chromatography, HPLC, precipitation, electrophoresis, etc., thereby partially, substantially, almost completely, or completely removing impurities, such as precursors or other chemicals involved in the preparation of the peptide, polypeptide, fusion protein, or nucleic acid molecule. Those skilled in the art will understand the required level of purification for a given application. For example, genomic DNA or proteins or polypeptides isolated and purified for use in therapeutic compositions intended for human administration typically must have high purity, in accordance with regulatory standards and good manufacturing practices.

[0023] As used herein, the term “phage product” or “active phage product” refers to a protein or fragment or variant thereof, and the nucleic acid encoding them, which has been isolated or derived from the phages of the present invention, and which retain the biological functions or activities associated with the phage from which they were isolated or derived (e.g., antibacterial activities, such as lytic cell killing).

[0024] As used herein, in the context of nucleotide sequences, the term "variant" refers to a nucleotide sequence that comprises or constitutes a nucleotide sequence having at least 70%, 75%, 80%, 85%, 90%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99% sequence identity with a reference nucleic acid sequence. Variants may optionally retain one or more functions of the reference nucleic acid sequence. For example, a variant phage may exhibit at least one biological activity derived from that variant phage, such as antibacterial activity, like lytic killing activity. Those skilled in the art will understand that the accuracy of nucleic acid replication in a phage is less than 100%, such that a given phage will exhibit at least 1% variation during its replication (including during its production as an antibiotic agent). Genomic variations anticipated during the preparation and use of phages may result in offspring variants having at least about 85%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity with the parent genome. Therefore, in some embodiments, the phage of the present invention comprises or is composed of a genome having at least about 85%, 90%, 95%, 96%, 97%, 98%, or at least 99% sequence identity with the nucleotide sequence of the parent phage, while retaining antibacterial activity against the target (host) bacteria of the parent phage. In the context of phages, a "variant" is a phage whose genome has at least about 85%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity with the nucleotide sequence of the parent genome, and retains lytic activity like that of the parent phage (i.e., lysis of the same one or more bacterial strains). In some aspects, a "variant" is identified solely by its lytic activity against one, two, or three bacterial strains (e.g., as described in the embodiments herein).

[0025] For example, in some embodiments, the bacteriophage is a variant of Kle_F391 / 08, Kle_F17 / 19, or Kle_F58 / 19, and retains antimicrobial and / or lytic activity against one, two, or three of Klebsiella pneumoniae strains 57 / 17, 237 / 14, and 397 / 07.

[0026] The term “progeny” when referring to any novel phages used herein means a phage replica containing a progeny of a phage with the specific nucleic acid identified herein, or a phage produced by methods known to those skilled in the art, or a phage having a substantially equivalent RFLP (restriction fragment length polymorphism) DNA profile to that of a phage with the specific nucleic acid identified herein. The term “having substantially equivalent or equal RFLP” is used to represent variability between organisms, according to the method proposed by Tenover et al. (Tenover, FC et al. Interpreting Chromosomal DNA Restriction Patterns Produced by Pulsed-Field Gel Electrophoresis: Criteria for Bacterial Strain Typing. J. Clin. Microbiol 33:2233-2239 (1995)). Tenover et al. proposed an acceptable level of variability, provided that the genomes of equivalently proliferating organisms are defined using restriction enzymes followed by electrophoresis. According to the criteria proposed by Tenover et al., progeny with equivalent RFLP DNA profiles can be considered as phages that are substantially equivalent to the phages of the specific nucleic acids identified in this paper, that is, substantially equivalent to phages containing nucleic acid sequences having any of the nucleotide sequences in SEQ ID NO:1-3.

[0027] As used herein, the term "host cell" refers to a specific target cell transfected with a nucleic acid molecule and the offspring or potential offspring of such cells containing that nucleic acid molecule or its chromosomal integration. Such offspring may not be identical to the parent cell transfected with the nucleic acid molecule due to mutations or environmental influences that may occur in subsequent generations, or due to the integration of the nucleic acid molecule into the host cell's genome. "Host cell" also refers to a cell infected by a bacteriophage (e.g., a complete bacteriophage), such as a bacterial cell, in which the bacteriophage survives and replicates. In order to produce a bacteriophage, the host cell may or may not belong to the same species or strain from which the bacteriophage was isolated or cultured.

[0028] As used herein, the term "fragment" refers to a peptide or polypeptide comprising an amino acid sequence of at least 5 consecutive amino acid residues, at least 10 consecutive amino acid residues, at least 15 consecutive amino acid residues, at least 20 consecutive amino acid residues, at least 25 consecutive amino acid residues, at least 40 consecutive amino acid residues, at least 50 consecutive amino acid residues, at least 60 consecutive amino acid residues, at least 70 consecutive amino acid residues, at least 80 consecutive amino acid residues, at least 90 consecutive amino acid residues, at least 100 consecutive amino acid residues, at least 125 consecutive amino acid residues, at least 150 consecutive amino acid residues, at least 175 consecutive amino acid residues, at least 200 consecutive amino acid residues, or at least 250 consecutive amino acid residues. In a particular embodiment, the fragment is a functional fragment because it retains at least one function of the protein from which it is isolated, such as retaining antimicrobial activity, such as lytic cell killing.

[0029] As used herein, the terms “in combination” or “further combination” or “also combination” refer to the use of other prophylactic and / or therapeutic agents with the phages or phage products of the present invention (including phage mixtures of different phages of the present invention). The use of the term “in combination” does not limit the order in which the prophylactic and / or therapeutic agents are administered to the subject. The first prophylactic agent or therapeutic agent may be administered to the subject before (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks prior to, during, or after (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks prior to, during, or after) the administration of the second prophylactic agent or therapeutic agent (different from the first prophylactic agent or therapeutic agent).

[0030] As used herein, the terms “enhancement” or “enhancement agent” refer to the subsequent repeated use of the same or substantially the same prophylactic and / or therapeutic agent, such as repeated doses of the phage, phage product, or phage mixture of the present invention. The same or substantially the same prophylactic or therapeutic agent may be administered first before a second administration to the subject (e.g., before 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks).

[0031] As used herein, the terms “preventive agent” and “multiple preventive agents” refer to reagents / pharmaceuticals, such as the bacteriophages, bacteriophage products or bacteriophage mixtures of the present invention, which can be used to prevent, manage, control or reduce the occurrence of one or more symptoms of a disease or condition, particularly a disease or condition associated with a bacterial infection, and more specifically a disease or condition associated with a respiratory bacterial infection, such as, but not limited to, hospital-acquired bacterial pneumonia or a respiratory bacterial infection associated with cystic fibrosis.

[0032] As used herein, the terms “therapeutic agent” and “multiple therapeutic agents” refer to reagents / pharmaceuticals, such as the bacteriophages, bacteriophage products or bacteriophage mixtures of the present invention, which can be used to treat, manage or control one or more symptoms of a disease or condition, particularly a disease or condition associated with a bacterial infection, and more specifically a disease or condition associated with a bacterial infection of the respiratory system, such as, but not limited to, hospital-acquired bacterial pneumonia or a bacterial infection of the respiratory system associated with cystic fibrosis.

[0033] As used herein, the terms “treat,” “treatment,” and “treating” refer to the attainment of a therapeutic benefit in a subject receiving a pharmaceutical composition. In achieving a therapeutic benefit, the aim is to eliminate, alleviate, reduce the severity, improve, or delay the progression of symptoms or underlying causes (e.g., bacterial infection) associated with a pathological condition or symptom. A “therapeuticly effective amount” refers to an amount of therapeutic agent (e.g., a bacteriophage or phage product in the pharmaceutical composition of the present invention) sufficient to achieve at least one therapeutic benefit in a subject receiving the pharmaceutical composition.

[0034] As used herein, the terms “prevent,” “prevention,” and “preventing” refer to obtaining a preventive benefit in a subject receiving the pharmaceutical composition. With regard to achieving a preventive benefit, the aim is to delay, prevent, or reduce the occurrence of symptoms or underlying causes (e.g., bacterial infection) associated with a pathological condition or symptom. “Effective preventive amount” refers to an amount of preventive agent (e.g., a bacteriophage or bacteriophage product in the pharmaceutical composition of the present invention) sufficient to achieve at least one preventive benefit in a subject receiving the pharmaceutical composition.

[0035] As used herein, the terms “antimicrobial activity” and “antimicrobial activity” when referring to bacteriophages or bacteriophage products (e.g., bacteriophage proteins), or their variants or fragments, are used interchangeably to refer to the ability to kill and / or inhibit the growth or reproduction of microorganisms (especially species or strains of bacteria infected by bacteriophages). In some embodiments, antimicrobial activity is assessed by culturing bacteria, such as Gram-negative bacteria (e.g., *Pseudomonas aeruginosa* or *Klebsiella pneumoniae*), according to standard techniques (e.g., in liquid culture or on agar plates), contacting the culture with the bacteriophages, bacteriophage proteins, or variants thereof of the present invention, or with a mixture of bacteriophages, bacteriophage proteins, or variants thereof, and monitoring cell growth after said contact. For example, in liquid culture, bacteria may grow to an optical density (“OD”) representing the midpoint of the exponential growth of the culture; the culture may be exposed to one or more concentrations of one or more bacteriophages, bacteriophage products, or variants thereof of the present invention, and the OD value may be monitored relative to a control culture. A decrease in OD value relative to a control culture indicates that one or more phages or phage products exhibit antimicrobial activity (e.g., lytic killing activity). Similarly, bacterial colonies can be formed on an agar plate, which is then exposed to one or more of the phages or phage products of the present invention or their variants, and colony growth can subsequently be evaluated relative to a control plate. A decrease in colony size or total colony count indicates that one or more phages or phage products possess antimicrobial activity. 4. Description of the attached drawings

[0036] This patent or application document contains at least one drawing in color. A copy of the published text of this patent or application with the color drawing will be provided by the competent authority upon request and payment of the necessary fees.

[0037] Figure 1 The morphological features of bacteriophages Kle_F391 / 08(A), Kle_F17 / 19(B), and Kle_F58 / 19(C) are shown. These bacteriophages were characterized using transmission electron microscopy (Hitachi H-7650).

[0038] Figure 2a and Figure 2b A schematic diagram of the F391 / 08 genome structure is shown. Predicted open reading frames (ORFs) within approximately 113 kb of the genome are indicated by arrows and numbered in black. The direction of the arrows indicates the direction of transcription. Color coding: Black – Open reading frames (ORFs) whose products can be functionally resolved based on homologous proteins; Gray – ORFs encoding products similar to proteins with unknown functions; Empty arrows – ORFs encoding proteins that do not have significant homology with proteins in the database.

[0039] Figure 3A schematic diagram of the Kle_F17 / 19 genome is shown. Predicted ORFs in approximately 45kb of the genome are indicated by arrows and numbered in black. The direction of the arrows indicates the direction of transcription. Color coding: Black – ORFs whose products can be functionally resolved based on homologous proteins; Gray – ORFs encoding products similar to proteins with unknown functions; Empty arrows – ORFs encoding proteins that do not have significant homology with proteins in the database.

[0040] Figure 4a and Figure 4b A schematic diagram of the Kle_F58 / 19 genome is shown. Predicted ORFs within approximately 170kb of the genome are indicated by arrows and numbered in black. The direction of the arrows indicates the direction of transcription. Color coding: Black – ORFs whose products can be functionally resolved based on homologous proteins; Gray – ORFs encoding products similar to proteins with unknown functions; Empty arrows – ORFs encoding proteins that do not have significant homology with proteins in the database.

[0041] Figure 5 The single lysis curves represent the lysis curves of phages F391 / 08 and Kle_F58 / 19 (MOI 10) against Klebsiella pneumoniae 57 / 17. Viable cell counts were quantified using a 10-fold serial dilution method, monitored every 1 hour for 8 hours, and again at 24-hour ppi.

[0042] Figure 6 The single lysis curve represents the Kle_F17 / 19 phage (MOI 10) against Klebsiella pneumoniae 237 / 14. Viable cell counts were quantified by a 10-fold serial dilution method, monitored every 1 hour for 8 hours, and again at 24-hour ppi.

[0043] Figure 7 The combined lysis curves of phages F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 at an MOI of approximately 10 were plotted. Viable cell counts were quantified using a 10-fold serial dilution method, monitored every hour for 8 hours, and again at 24 hours post-infection (ppi).

[0044] Figure 8 Bacterial load at 24 h post-infection is shown, using strain 2 and phage mixture / antibiotic (combined) treatment or NaCl (simulated) treatment with nebulized purification. Bacterial load is given as CFU per finely sectioned lung slice. Columns represent the mean ± SD of two technical replicates in n = 1 experiment.

[0045] Figure 9Tissue viability at 24 h post-infection was depicted using either strain 2 treated with a mixture of nebulized purified phage / antibiotic (combined) or NaCl (simulated) treatment. Tissue viability was assessed by calcein staining and given in relative light units. Columns represent the mean ± SD of two technical replicates (each measurement in duplicate) in n = 1 experiment.

[0046] Figure 10 The efficacy of phage titers on rat PCLS treated with aerosolized phage is shown. Phage titers in PCLS were measured at 1 h and 24 h using a plaque assay and the indicated test strain. Phage titers are given in plaque-forming units (PFU) per PCLS. Columns represent the mean ± SD of 2–4 technical replicates (1–2 exposure plates, each with 2 PCLS, shown as a single point) for n = 1 experiment.

[0047] Figure 11 The efficacy of nebulized phage treatment on rat PCLS is shown. Bacterial load was measured 1 h or 24 h after treatment with a nebulized phage mixture or NaCl (medium) following infection. Bacterial load is given as CFU per PCLS. Columns represent the mean ± SD of four technical replicates (two exposure plates, each with two PCLS, with the mean for each plate shown as a dot) for n = 1 experiment.

[0048] Figure 12 The efficacy of phage titer in rat PCLS is shown by combining aerosolized phage treatment with antibiotic co-treatment (phage lysate). Phage titers were measured using a plaque formation assay at 1 and 24 h after inoculum treatment with strain 1 and the phage mixture. Phage titers are given in plaque formation units. Columns represent the mean ± SD of two technical replicates (shown as single points) of n = 3 experiments.

[0049] Figure 13 The efficacy of co-treatment with nebulized phage and antibiotics on bacterial load in rat PCLS is shown. Bacterial load at pi 24 h is presented after treatment with nebulized NaCl or a phage mixture alone or with immersion antibiotics. Bacterial load is given as CFU per PCLS. Columns represent the mean ± SD of two technical replicates (shown as single points) of n = 3 experiments. *** indicates significance of p < 0.001 compared to the control column (infection + NaCl at pi 24 h) based on one-way ANOVA and Sidak's follow-up test of multiple comparisons. ### indicates significance of p < 0.001 for direct comparison of the groups shown based on an unpaired two-tailed t-test.

[0050] Figure 14This study demonstrates the efficacy of nebulized phage treatment and co-treatment with antibiotics on tissue viability in rat PCLS. Tissue viability was assessed at 24 h post-infection and with NaCl (the medium), nebulized phage mixture alone, or with antibiotics. Tissue viability was assessed by calcein staining and given in relative light units (RLU). Columns represent the mean ± SD of two technical replicates (shown as a single point, with each measurement in duplicate) for n = 3 experiments. ** and *** indicate significance compared to the control column (infection + NaCl at 24 h post-infection) based on one-way ANOVA and Sidak's follow-up test of multiple comparisons; p < 0.01; p < 0.001. 5. Detailed Description of the Invention

[0052] This invention relates to phage therapy for the treatment and control of bacterial infections, particularly respiratory bacterial infections such as bacterial pneumonia and respiratory infections associated with cystic fibrosis. One aspect of the invention relates to novel phage strains, including Klebsiella pneumoniae phages Kle_F17 / 19 and Kle_F58 / 19, and variants thereof. Another aspect of the invention relates to cocktail compositions of one or more of the phages and / or phage products of the invention, and combinations thereof with other phages, including Kle_F391 / 08 previously disclosed in PCT / PT2011 / 000031. Yet another aspect of the invention relates to cocktail compositions of Klebsiella pneumoniae phages Kle_F391 / 08, Kle_F17 / 19, and Kle_F58 / 19, and variants thereof. Still another aspect relates to pharmaceutical compositions of phages, and methods of using them to treat and prevent bacterial infections, particularly respiratory infections caused by Klebsiella pneumoniae. Other aspects of the invention relate to the use of bacteriophages and combinations thereof as diagnostic tools and disinfectants.

[0053] 5.1 Bacteriophages and their variants

[0054] Another aspect of the invention relates to novel Klebsiella pneumoniae phages targeting numerous strains of Klebsiella pneumoniae. Klebsiella pneumoniae is a Gram-negative, non-motile rod-shaped bacterium found in the normal flora of the oral cavity, skin, and intestines. As a capsular facultative anaerobe, it also naturally occurs in soil. Clinically, it is the most important member of the genus Klebsiella in the family Enterobacteriaceae. Klebsiella infections often occur in individuals with weakened immune systems due to poor diet, such as alcoholics and diabetics. Klebsiella is also an opportunistic pathogen in patients with chronic lung disease, nasal mucosal atrophy, cystic fibrosis, and rhinoscleroderma. New antibiotic-resistant strains of Klebsiella pneumoniae are emerging, and it is increasingly being found in hospital-acquired infections, for example, due to contact with contaminated instruments.

[0055] Klebsiella pneumoniae is indeed one of the most important pathogens causing respiratory infections in humans, accounting for 25-43% of hospital-acquired pneumonia caused by Gram-negative bacteria (Chibber S et al., 2008, J Med Microbiol 57(12):1508-1513). The high incidence of multidrug-resistant bacteria has led to the limited efficacy of current antibiotics and a high probability of colonization of patients by drug-resistant strains. Capsular polysaccharides are important virulence factors of Klebsiella strains and also limiting factors for phage infection. The literature describes 78 capsule types (Hus CR et al., 2013, PLoS One 8(8):e70092), and phages infecting these species have overcome this "barrier". Virulent strains of Klebsiella pneumoniae are primarily associated with K1 and K2 capsular serotypes (Cleg S et al., 2016, Microbiol Spectr 4(1); and Lin TZ et al., 2014, J Infect Dis 210:1734-1744), for example in purulent liver abscesses; however, the K1 capsular serotype is associated with community-acquired isolates rather than hospital-acquired isolates (Tsay RW et al., 2002, Arch Intern Med 162(9):1021-1027). Nevertheless, depending on the type of infection, strains can exhibit a wide variety of capsular serotypes, and the distribution of Klebsiella pneumoniae capsular serotypes varies worldwide (Hus CR et al., 2013, PLoS One 8(8):e70092).

[0056] In one implementation, purified phage F17 / 19 (this name is interchangeable with "Kle_F17 / 19") is provided, which targets many strains of Klebsiella pneumoniae.

[0057] In another implementation, purified phage F58 / 19 (this name is interchangeable with "Kle_F58 / 19") is provided, which also targets many Klebsiella pneumoniae strains.

[0058] In some embodiments, the phage of the present invention comprises or consists of a genome having at least 85%, 90%, 95%, 96%, 97%, 98%, or at least 99% sequence identity with the nucleic acid sequence SEQ ID NO:2 or SEQ ID NO:3, and the phage exhibits at least one of one or more biological activities of phages F17 / 19 and F58 / 19, such as antimicrobial or antibacterial activity (e.g., lytic killing activity).

[0059] The bacteriophages and certain bacterial strains disclosed in this article were deposited at NCIMB (NCIMB Ltd., Ferguson Building, Craibstone Estate, Bucksburn, Aberdeen, UK) on December 13, 2019.

[0060] Table 1

[0061]

[0062] 5.2 Cocktail Composition

[0063] One specific aspect of the invention relates to cocktail compositions of different phages. This "cocktail / mixture" may contain at least two different purified phages, for example, two, three, four, five, six, seven, eight, nine, ten, or more different purified phages or variants thereof. The mixture may be used alone or in combination with other therapies (e.g., antibiotics and / or antifungals).

[0064] Phage mixtures offer advantages over using a single phage, such as enhanced lytic activity against specific bacterial species or strains and / or reduced likelihood of the emergence of bacteria resistant to a single phage. Different phages can also be mixed like a cocktail to broaden their properties, preferably producing a broader overall spectrum of antimicrobial activity. However, phage mixtures with antimicrobial activity against different bacteria are rare, likely because it is difficult to combine the different specificities of phage strains while maintaining the infectivity and / or lytic activity of each phage in the presence of different strains.

[0065] In some embodiments, the present invention provides a cocktail composition comprising at least three different purified phages having antibacterial activity against the same or different bacterial species or strains. In some specific embodiments, the present invention provides a cocktail composition comprising at least purified phages Kle_F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 or variants thereof, or consisting of purified phages Kle_F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 or variants thereof, including variants having lytic activity against one or all of Klebsiella pneumoniae 57 / 17, 237 / 14, and / or 397 / 07.

[0066] In some specific embodiments, a composition comprising at least two different purified phages or compositions thereof is provided, the phages including at least one of F17 / 19 or F58 / 19 or variants thereof.

[0067] In some preferred embodiments, the combination does not weaken or reduce (or substantially or significantly weaken or reduce) the infectivity, host range, and / or lytic activity of the various phages in the presence of different phage strains. In some particularly preferred embodiments, the potency of at least one phage in the cocktail combination is enhanced or increased due to the presence of at least one other phage in the cocktail combination, thereby producing a synergistic effect.

[0068] In some embodiments, the cocktail composition comprises at least three bacteriophages exhibiting antibacterial activity against Klebsiella pneumoniae. In some specific embodiments, the present invention provides a cocktail composition comprising at least three different purified bacteriophages or consisting of three purified bacteriophages F391 / 08, F17 / 19, and F58 / 19, or variants thereof having antibacterial activity against Klebsiella pneumoniae.

[0069] In other particularly preferred embodiments, the composition comprises or is composed of purified bacteriophages F391 / 08, F17 / 19, and F58 / 19 or variants thereof, and three purified bacteriophages F99 / 10, F27 / 12, and F95 / 13 having antibacterial activity against Pseudomonas aeruginosa. Alternatively, the composition comprising or being composed of purified bacteriophages F391 / 08, F17 / 19, and F58 / 19 may be administered in combination with a composition comprising or being composed of purified bacteriophages F99 / 10, F27 / 12, and F95 / 13. F99 / 10, F27 / 12, and F95 / 13 are disclosed in US2019 / 0290709. These bacteriophages and certain strains of Pseudomonas aeruginosa were deposited at NCIMB (NCIMB Ltd., Ferguson Building, Craibstone Estate, Bucksburn, Aberdeen, UK) on December 1, 2017.

[0070] Table 2

[0071]

[0072] In some embodiments, a cocktail composition is provided comprising at least two different purified phages, one of which is F17 / 19 or F58 / 19, and at least one purified phage selected from F92 / 15, F105 / 15, F134 / 15, and F141 / 15, which are resistant to strains of one or more Klebsiella species, more preferably including Klebsiella pneumoniae, all of which are disclosed in US 2019 / 0290709.

[0073] In some embodiments, the present invention provides a cocktail composition comprising at least F17 / 19 or F58 / 19, further combined with at least one other phage selected from the group consisting of: phage F168 / 08 (as disclosed in WO2010 / 090542) having antibiotic activity against one or more Enterococcus faecalis and / or Enterococcus faecium strains; phage F170 / 08 (as disclosed in WO 2010 / 090542) having antibiotic activity against one or more Enterococcus faecalis and / or Enterococcus faecium strains; phage F770 / 05 (as disclosed in WO 2010 / 090542) having antibacterial activity against one or more Pseudomonas aeruginosa strains; and phage F197 / 08 (as disclosed in WO 2010 / 090542) having antibacterial activity against one or more Staphylococcus aureus strains. The following bacteriophages exhibit antibacterial activity against one or more Staphylococcus aureus strains (as disclosed in WO 2010 / 090542): F86 / 06, F87s / 06, F91a / 06, F1245 / 05, F394 / 08, F394 / 08, and Escherichia coli. Phages with antibacterial activity against *Pseudomonas aeruginosa* strains (as disclosed in WO 2012 / 036580), phages with antibacterial activity against one or more *Pseudomonas aeruginosa* strains (as disclosed in WO 2012 / 036580), phages with antibacterial activity against one or more *Staphylococcus aureus* strains (as disclosed in WO 2012 / 036580), phages with antibacterial activity against one or more *Klebsiella pneumoniae* strains (as disclosed in WO 2012 / 036580), and phages with antibacterial activity against one or more *Staphylococcus aureus* strains (as disclosed in WO 2012 / 036580).

[0074] The bacteriophages of the present invention and / or those used in the cocktail compositions of the present invention can be obtained by any method known in the art and / or disclosed herein. In some embodiments, the present invention provides, for example, methods for generating and purifying bacteriophages F391 / 08, F17 / 19 and / or F58 / 19 from the preserved strains disclosed herein.

[0075] In addition, bacteriophages can be isolated from bacterial samples using any of the methods described herein or known in the art (see, for example, Carlson, “Working with bacteriophages: common techniques and methodological approaches,” in Bacteriophages: Biology and Applications, 5th edition, CRC Press (2005) by Kutter and Sulakvelidze (eds.). Specific bacterial strains that can be used include, for example, Klebsiella pneumoniae strains 57 / 17, 397 / 07, or 237 / 14 (e.g., for isolating phages F391 / 08, F17 / 19, or F58 / 19), Pseudomonas aeruginosa strains 391 / 08, 92 / 15, 105 / 115, or 121 / 15 (e.g., for isolating phages F99 / 10, F110 / 10, F27 / 12, F83 / 13, and / or F95 / 13); or phages can be isolated from any other bacterial strain susceptible to infection by one or more phages in which the phages replicate.

[0076] Skilled technicians may also use one or more methods to propagate or amplify phages, particularly purified phages F391 / 08, F17 / 19, or F58 / 19, and their variants, to obtain larger quantities of a given phage. In some embodiments, methods for generating and / or isolating other phages may include (i) obtaining a culture of *Klebsiella pneumoniae*; (ii) infecting it with phage F391 / 08, F17 / 19, or F58 / 19, or its variants; (iii) culturing until significant lysis of the culture is observed; and (iv) isolating the phage from the culture. The host cell used may be any bacterial strain, for example, any phage-susceptible *Klebsiella pneumoniae* strain that can be used to replicate the phage. In some embodiments, the host cell used may be, for example, *Klebsiella pneumoniae* strain 57 / 17, *Klebsiella pneumoniae* strain 237 / 14, or *Klebsiella pneumoniae* strain 397 / 07.

[0077] 5.3 Pharmaceutical Composition

[0078] The purified bacteriophages disclosed herein, including their variants and phage cocktail combinations, can be administered alone or incorporated into pharmaceutical compositions for the treatment or prevention of bacterial infections, such as those caused by bacteria (including, but not limited to, *Pseudomonas aeruginosa* and *Klebsiella pneumoniae*). One or more phages or phage products can be combined with pharmaceutically acceptable carriers, excipients, or stabilizers. Examples of pharmaceutically acceptable carriers, excipients, and stabilizers include, but are not limited to, buffers such as phosphates, citrates, and other organic acids; antioxidants, including ascorbic acid; low molecular weight peptides; proteins such as serum albumin and gelatin; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates, including glucose, mannose, or dextrin; chelating agents such as EDTA; sugar alcohols such as mannitol or sorbitol; salt-forming counterions such as sodium; and / or nonionic surfactants such as TWEEN. TM Polyethylene glycol (PEG) and PLUONICS TM For example, in addition to the components described above, the pharmaceutical compositions of the present invention (e.g., antibacterial compositions) may also include lubricants, wetting agents, emulsifiers, suspending agents, and preservatives. Specifically, purified bacteriophages are formulated in a NaCl solution (e.g., 0.9% NaCl).

[0079] In some embodiments, the pharmaceutical composition is formulated for administration as an aerosol. Formulations for aerosol delivery can be in the form of dry powder, fine particles, nanoparticles, solutions, lyophilized formulations, liposomes, etc. Liposome formulations can protect phages from the harsh conditions of sputum and improve penetration through biofilms and / or allow for a more sustained release of the drug within the airways. Formulations for aerosol delivery typically contain sterile water and little or no preservatives to reduce side effects such as bronchial irritation and bronchospasm. Formulations for aerosol delivery preferably have the same or substantially the same osmolality as the liquid layer on the airway surface.

[0080] The bacteriophages and bacteriophage mixtures disclosed herein can be combined with one or more other therapeutic and / or prophylactic agents (e.g., one or more other bacteriophages or antibiotics) as described herein and / or known in the art for the treatment of bacterial infections. For example, the pharmaceutical compositions of the present invention may comprise two or more purified bacteriophages disclosed herein (with antibacterial activity against the same or different bacterial species or strains) and bacteriophages known in the art. In certain embodiments, the combined therapeutic components target two or more bacterial species or strains.

[0081] The pharmaceutical compositions of the present invention can also be combined with one or more non-phage therapeutic and / or preventive agents (e.g., one or more conventional antibiotics) as described herein and / or known in the art for the treatment and / or prevention of bacterial infections. Other therapeutic and / or preventive agents that can be used in conjunction with the phages or phage products of the present invention include, but are not limited to, antibiotics, anti-inflammatory agents, antiviral agents, antifungal agents, or local anesthetics. In some preferred embodiments, the pharmaceutical compositions are formulated for the treatment and / or prevention of lung infections and contain one or more other therapeutic and / or preventive agents selected from antibiotics, antifungal agents, and local anesthetics. In some embodiments, the pharmaceutical compositions comprise the phage cocktail of the present invention, administered in the absence of standard or conventional antibiotics.

[0082] In certain embodiments, the phage mixture is administered with broad-spectrum antibiotics, particularly those with activity against Gram-negative bacteria, such as anti-pseudomonal β-lactams (e.g., piperacillin, tazobactam, ceftazidime, or meropenem) and / or aminoglycosides (e.g., amikacin), and / or with antibiotics against Gram-positive bacteria, such as vancomycin or oxazolidinones (e.g., linesolid), and / or in combination with colistin. Standard or conventional antibiotics include, but are not limited to, amikacin, gentamicin, kanamycin, neomycin, netilmicin, paromomycin, rhodostreptomycin, streptomycin, tobramycin, aporamycin, rifamycin, naphazoline, mupirocin, geldanamycin, anserine, carbapenems, imipenem, meropenem, ertapenem, and faropenem. Doripenum, Panipenum / Betamilon, Biapenem, PZ-601, Cephalosporins, Cetirizine, Cefadroxil, Cefalothin, Cefotaxime, Cefalothin, Cephalosporin, Ceftriaxone, Cefotaxime, Cefoxitin, Cefoxitin, Cefoxitin, Cefoxitin, Cefotaxime, Cefoxitin, Cefotaxime, Cefotiam, Cefotaxime, Cefotiam, Cefotiam, Cefotiam, Cefotiam, Cefotiam Cefcapene, cefodazole, cefdinir, cefotolam, cefetamet, cefixime, cefotaxime, cefodil, cefbufen, ceftiofur, cefotaxime, cefazolin, cefazolin, ceftriaxone, cefoperazone, ceftazidime, latamoxef, cefclidine, cefepime, cefotaxime Prenam), cefotaxime, cefazolin, cefpirome, cefquinoxime, fluoxetine, ceftobiprole, azithromycin, clarithromycin, erythromycin, roxithromycin, aztreonam, penicillin and penicillin derivatives, actinomycin, bacitracin, colistin, polymyxin B, sinofloxacin, flumethylquine, nalidixic acid, oxolinic acid(acid), pyrrolic acid, pipemidic acid, rosofaxin, ciprofloxacin, enoxacin, fleroxacin, lomefloxacin, naflufloxacin, norfloxacin, ofloxacin, pefloxacin, levofloxacin, balofloxacin, gatifloxacin, gaptafloxacin, levofloxacin, moxifloxacin, pazufloxacin, sofloxacin, timafloxacin, tosufloxacin, clinfloxacin, garayfloxacin, gimifloxacin, stifloxacin, trovalfloxacin, prulifloxacin, acetazolamide, benzolamide, bumetanide, celecoxib, chlorthalidone, clopamide, dichlorphenamide, dzodamine, ethoxazole Yzolamide), furosemide, hydrochlorothiazide, indapamide, mafendide, mefcitide, metolazone, probenecid, acesulfame, sulfadimethoxine, sulfadoxine, sulfonamides, sulfamethoxazole, sulfasalazine, sucralfate, sumatriptan, sipamide, tetracycline, chlorotetracycline, doxycycline, lysine, methylchlorocycline, methacycline, minocycline, rolicycline, methicillin, nafcillin, oxacillin, cloxacillin, vancomycin, teicoplanin, clindamycin, co-trimoxazole, flucloxacillin, dicloxacillin, ampicillin, amoxicillin, and any combination thereof.

[0083] In some embodiments, the pharmaceutical compositions of the present invention comprise antibiotics with antibacterial activity against *Pseudomonas aeruginosa* and / or *Klebsiella pneumoniae*. In other embodiments, the pharmaceutical compositions of the present invention comprise antibiotics with antibacterial activity against bacteria other than *Pseudomonas aeruginosa* and / or *Klebsiella pneumoniae*. In preferred embodiments, the amount of antibiotic used can effectively adjuvant or synergistically enhance the therapeutic and / or preventive effects of the bacteriophages, bacteriophage products, or bacteriophage mixtures of the present invention against a given infection.

[0084] Standard antifungal agents include amphotericin B, such as liposomal amphotericin B and non-liposomal amphotericin B.

[0085] In some preferred embodiments, the pharmaceutical compositions of the present invention are formulated for aerosol administration and further comprise one or more antibiotics also intended for aerosol delivery. Antibiotics intended for aerosol delivery include, for example, inhaled aminoglycosides such as tobramycin (e.g., tobramycin solution or tobramycin dry powder), gentamicin, amikacin, inhaled polymyxins such as colistin solution or colistin dry powder, and colistin E methanesulfonate sodium; and inhaled monocyclic β-lactams (e.g., aztreonam solution or nebulized aztreonam lysine); and aerosolized levofloxacin, ceftazidime, fosfomycin, gentamicin, vancomycin, amphotericin B, capreomycin, rifampin, isoniazid, and ciprofloxacin (Quon BS et al., 2014, Annals ATS11(3):425-434). In some embodiments, the aerosolized pharmaceutical composition of the present invention further comprises one or more antifungal agents also used for aerosol delivery, such as liposomal amphotericin B.

[0086] In some embodiments, the pharmaceutical compositions of the present invention are formulated for the treatment and / or prevention of bacterial infections caused by species of the genus Klebsiella, such as Klebsiella pneumoniae. In some such embodiments, the pharmaceutical composition comprises a cocktail composition containing one or more purified F17 / 19 or F58 / 19 bacteriophages and optionally F391 / 08 bacteriophage or variants thereof, and in one embodiment, comprising F17 / 19, F58 / 19, and F391 / 08 or variants thereof, or a combination thereof. The composition further comprises purified bacteriophages F99 / 10, F27 / 12, and F95 / 13 or variants thereof having anti-Pseudomonas aeruginosa activity, or a combination thereof. In some embodiments, the pharmaceutical composition may further comprise other agents, such as antibiotics with antibacterial activity against Klebsiella pneumoniae and / or Pseudomonas aeruginosa, and / or antibiotics with antibacterial activity against bacteria other than Klebsiella pneumoniae or Pseudomonas aeruginosa. In some embodiments, the composition is formulated into a dosage form in which the presence of the phage provides a multiple of infection (MOI) of about 1 to about 10 when administered to a subject in need. In some embodiments, each phage is present in 10 9 One PFU to 10 10 A dose of 10 pfu is administered, and for example, in 10 9 One pfu / ml is present in the composition. In a preferred embodiment, the composition is formulated for application as an aerosol.

[0087] Pharmaceutical compositions comprising the purified phage mixture of the present invention can be formulated into single-dose or multi-dose formulations. Preferred formulations are those that can be delivered as an aerosol as described above. Other suitable formulations include suspensions, emulsions, lotions, solutions, creams, ointments, or dispensing agents, or in the form of skin patches.

[0088] Alternatively or concurrently, the pharmaceutical compositions provided herein may be administered as suppositories or vaginal suppositories, orally (e.g., as tablets which may contain excipients such as starch or lactose, as capsules, beads, elixirs, solutions, or suspensions, each optionally containing flavoring agents, coloring agents, and / or excipients), or by parenteral (e.g., intravenous, intramuscular, or subcutaneous) injection. For parenteral administration, the composition may be used as a sterile aqueous solution which may contain other substances, such as sufficient salts or monosaccharides, to make the solution isotonic with blood. For buccal or sublingual administration, the composition may be administered as tablets or lozenges which may be formulated in a conventional manner. Topical formulations typically include sterile buffers, such as sterile PBS, water or saline buffers, or sterile SM buffer.

[0089] The administration methods described herein and / or known in the art can be used to deliver the desired dose of the phages, phage products, and / or phage mixtures of the present invention, in accordance with appropriate dosing regimens. Dosage and dosing regimens can vary depending on the specific formulation, route of administration, condition being treated, and other factors. For example, within the skill level of a physician of ordinary skill, animal studies can provide reliable guidance for determining effective doses in human therapy. Those skilled in the art can perform interspecies extrapolation of effective doses following the principles described below, for example, in Mordenti, J. et al., “The use of interspecies scaling in toxicokinetics” in Toxicokinetics and New Drug Development, eds. Yacobi et al., Pergamon Press, New York, 1989, pp. 42-96. For example, mouse models of acute pulmonary infection can be used to evaluate the potency of the pharmaceutical compositions of the present invention, as detailed in the following examples.

[0090] The pharmaceutical compositions of the present invention can be administered according to a dosing regimen. In some embodiments, the dosing regimen includes administering the cocktail composition of the present invention every 6 or 8 hours (e.g., a multiple-dose regimen for topical application of a phage mixture to treat diabetic skin wounds (Mendes JJ et al., 2013, Wound Repair Regen 21:595-603)). In a preferred embodiment, a second or “boost” dose follows the initial dose, involving the re-administration of the pharmaceutical composition. For example, a booster dose may be administered following the initial dose after about 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 10 hours, 12 hours, 1 day, or 2 days. In a preferred embodiment, for example, in the treatment of respiratory or pulmonary infections, including but not limited to hospital-acquired bacterial pneumonia (HABP), a booster dose may be administered about 4, about 5, about 6, or about 8 hours after the initial dose.

[0091] 5.4 Therapeutic uses

[0092] On the other hand, the use of purified bacteriophages and bacteriophage mixtures in pharmaceutical compositions for the prevention and / or treatment of bacterial infections is involved. Bacteriophages have shown great potential in the treatment of bacterial infections due to their specificity and effectiveness in lysing pathogenic bacteria, including those associated with multidrug resistance (Larche J et al., 2012, Antimicrob Agents Chemother 56(12):6175-6180), their potential efficiency against bacteria in biofilms (Phee A et al., 2013, J Endod 39(3):364-369); their lack of pathogenicity to human and animal cells (Abedon ST et al., 2011, Bacteriophage 1(2):66-85); and their activity even in microaerophilic environments with high bacterial loads (Azeredo J et al., 2008. Curr Pharm Biotechnol 9:261-266). Phage mixtures offer additional advantages compared to using a single phage, such as increased lytic activity against specific bacterial strains, broadened host range, and / or reduced likelihood of bacterial resistance to a single phage. In fact, different phages are often mixed in cocktails to broaden their properties, preferably resulting in a larger overall antimicrobial spectrum, such as a broader host range, which makes it less likely that subjects receiving the drug will develop resistance.

[0093] In certain embodiments, the subject receiving the pharmaceutical composition of the present invention is a mammal (e.g., bovids, sheep, capillaries, equines, primates (such as humans), rodents, lagomorphs, or birds (e.g., chickens, ducks, geese)). In a preferred embodiment, the subject receiving the pharmaceutical composition of the present invention is a human, particularly a patient suffering from or at risk of respiratory or lung infections, including hospital-acquired bacterial pneumonia (HABP), respiratory equipment-acquired pneumonia (VAP), healthcare-associated pneumonia (HCAP), or cystic fibrosis-related infections. In some embodiments, the infection is caused by MDR, XDR, and PDR infectious agents.

[0094] In preferred embodiments, the pharmaceutical compositions of the present invention have activity against a variety of bacterial strains. In some preferred embodiments, the pharmaceutical compositions comprise a phage cocktail combination active against a variety of *Pseudomonas aeruginosa* and / or *Klebsiella pneumoniae* strains. Accordingly, the present invention provides methods for treating and / or preventing infections associated with *Pseudomonas aeruginosa* and / or *Klebsiella pneumoniae* in humans and animals using the phages, phage products, or phage cocktail compositions of the present invention. In other aspects, the present invention provides methods for treating and / or preventing infections associated with related species or strains of these bacteria.

[0095] Pseudomonas aeruginosa and Klebsiella pneumoniae are the culprits behind many serious opportunistic infections, especially in individuals with compromised immune systems. This encompasses the use of the pharmaceutical compositions of the present invention for the treatment and / or prevention of any infection associated with Pseudomonas aeruginosa and / or Klebsiella pneumoniae or with other bacterial species or strains, including but not limited to lung and respiratory tract infections, postoperative infections, infections associated with catheters and surgical drainage tubes, and bloodstream infections. In a preferred embodiment, the pharmaceutical compositions of the present invention can be used for the treatment and / or prevention of bacterial infections associated with the lungs and respiratory tract.

[0096] Respiratory and pulmonary infections include, but are not limited to, infections associated with cystic fibrosis, such as cystic fibrotic bronchiectasis; pneumonia, including hospital-acquired bacterial pneumonia, equipment-associated pneumonia, and bronchopneumonia; non-cystic fibrotic bronchiectasis; bronchitis; chronic obstructive pulmonary disease; mycobacterial infections; post-lung transplant infections; tuberculosis-related infections; empyema with chest wall fistula; pleurisy with fistula; lung abscess; rhinitis; purulent cysts; and pulmonary sepsis. Symptoms of respiratory or pulmonary infections include, for example, cough, wheezing, sputum production, shortness of breath (dyspnea), dysarthria (speech difficulties), and a general decline in quality of life. In a particularly preferred embodiment, the respiratory or pulmonary infection is hospital-acquired bacterial pneumonia (HABP).

[0097] Regarding HABP, the time of onset during hospitalization is a risk indicator for specific pathogens and outcomes. For early onset, such as within the first 4 days of hospitalization, the most common pathogens are endogenous microbiota, such as *Streptococcus pneumoniae* and *Haemophilus influenzae*, as well as antibiotic-sensitive Gram-negative and community-acquired Staphylococcus aureus. For late onset, such as after 5 days of hospitalization, Gram-negative bacteria account for the majority of cases, many of which are antibiotic-resistant, such as certain strains of *Pseudomonas aeruginosa*, *Klebsiella pneumoniae*, *Enterobacter* species, and *Acinetobacter* species, and certain *Staphylococcus aureus* infections, particularly in neurosurgical patients, diabetic patients, and patients with chronic kidney disease (2005, *Am J Respir Crit Care Med Vol 171(4):388:416*). Strains of *Pseudomonas aeruginosa* and *Klebsiella pneumoniae* are particularly associated with late-onset HABP.

[0098] Klebsiella pneumoniae and Pseudomonas aeruginosa are also associated with infections involving other organ systems with high fluid content, and the phage mixtures of the present invention have therapeutic and / or preventative uses for such infections. For example, the pharmaceutical compositions of the present invention can be used for the prevention or treatment of cerebrospinal fluid, peritoneal fluid, and urinary tract infections.

[0099] In some embodiments, the present invention provides a method for treating and / or preventing respiratory or pulmonary infections, comprising administering a therapeutically or preventively effective amount of the pharmaceutical composition of the present invention to a subject in need. In a preferred embodiment, the administration induces respiratory improvement, for example, restoring normal breathing to those experiencing dyspnea or tachypnea.

[0100] In particularly preferred embodiments, the present invention provides methods for treating an unexpected range of Klebsiella pneumoniae strains using phage mixtures of the invention, and in some embodiments, methods for treating Pseudomonas aeruginosa strains. For example, phage mixtures comprising Klebsiella pneumoniae phages F391 / 08, F17 / 19, and F58 / 19 have shown efficacy against a highly diverse range of clinical strains of Klebsiella pneumoniae (53% infection rate) when compared to homologous Klebsiella pneumoniae phages. In other embodiments, for example, phage mixtures further comprising Pseudomonas aeruginosa phages F99 / 10, F27 / 12, and F95 / 13 have shown efficacy against a highly diverse range of clinical strains of Pseudomonas aeruginosa exhibiting multiple capsular serotypes (57%) when compared to certain other Pseudomonas aeruginosa phages.

[0101] In a preferred embodiment, administration includes delivering the pharmaceutical composition to one or more airways of the subject via an aerosol, such as by inhalation. Inhalation administration can improve the delivery of the drug to the target site of infection (i.e., the airway) and / or limit the likelihood of systemic side effects. Administration of the pharmaceutical composition in aerosol form includes, but is not limited to, administration by inhalation, intranasal instillation, endotracheal intubation, delivery to the pleural cavity of the lungs, or by bronchoscopy (Abedon ST, 2015, Bacteriophage, 5(1):e1020260-1 to e1020260-13). During aerosol administration of the pharmaceutical composition, the bacteriophage remains viable and can be contained in particles of a suitable size to reach the lower respiratory tract. For example, in a particularly preferred embodiment, most of the aerosolized particles have a diameter of less than 5 μm, for example, at least 50%, 60%, 70%, or 80% of the particles have a diameter of less than 5 μm, more preferably about 2 μm.

[0102] For intranasal or inhalation administration, the phages and / or phage products of the present invention can be delivered in the form of dry powder, fine particles, nanoparticles, solutions, lyophilized formulations, liposome formulations, etc. Typically, formulations containing the phages, phage products, and / or phage mixtures of the present invention are delivered as dry powder inhalers or aerosol sprays from a pressurized container, pump, nebulizer, or nebulizer using suitable propellants such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, hydrofluoroalkane such as 1,1,1,2-tetrafluoroethane (HFA 134A.TM) or 1,1,1,2,3,3,3-heptafluoropropane (FIFA227EA.TM), carbon dioxide, or other suitable gases.

[0103] In the case of pressurized aerosols, the dosage unit can be determined by providing a valve for delivering the measured amount. The pressurized container, pump, nebulizer, or atomizer may contain a solution or suspension of the active compound, for example, using a mixture of ethanol and a propellant as a solvent, and may additionally contain a lubricant, such as sorbitan trioleate. Capsules and canisters for inhalation or blowing (e.g., made of gelatin) may be formulated as a powder mixture containing the phages, phage products, and / or phage mixtures of the present invention, and a suitable powder base such as lactose or starch.

[0104] Nebulization can be achieved using any method known in the art or as described herein. Typically, nebulization is achieved using a jet nebulizer, which uses air or oxygen to produce an aerosol under high pressure. Other nebulizers include vibrating screen nebulizers, driven by piezoelectric actuators, which reduce dimensional variability and shorten nebulization time (see, for example, Aeroneb, Nebutec 4, Nebutec 6). Another approach involves mechanical ventilation, where the nebulizer is connected to the inspiratory branch of the breathing apparatus tubing. Yet another method uses emulsion-based "spray-drying" to transform a solution or emulsion from a fluid state into fine particles (approximately 1-5 μm) with a uniform size distribution. Two typical categories of nebulizers include AeroEclips (Trudell Medical International), a jet nebulizer that nebulizes only when the patient inhales; and Omron (Omron, MicroAir U22), a battery-powered screen nebulizer that relies on the vibration of a piezoelectric crystal to force the medication through a fine screen to produce an aerosol (Sahota et al., 2015, J. Aerosol Medicine and Pulmonary Drug Delivery 28(0):1-8). In some preferred embodiments, the SYSTAM L290 (SYSTAM, Villeneuve Sur Lot, France) nebulizer is used. This nebulizer produces an ultrasonically atomized agent in which approximately 70% of the particles have a diameter of less than 5 μm.

[0105] The pharmaceutical compositions of the present invention will comprise one or more phages or phage products as described herein that are effective for treatment and / or prevention. Effective amounts for treatment and / or prevention refer to the amounts required to produce therapeutic and / or preventive benefits, respectively, in subjects receiving said amounts. Effective amounts for treatment and / or prevention will depend on the specific formulation, route of administration, condition being treated, whether other agents or therapies are used in combination with the methods of the present invention, and other factors.

[0106] In some embodiments, the pharmaceutical composition is delivered to the subject in need to provide one or more phages in an amount corresponding to a multiple of infection (MOI) of about 1 to about 10. The MOI is determined by assessing an approximate bacterial load in the lungs, or by calculating the bacterial load in the lungs of a specific patient, or by using an estimate for a given type of respiratory infection; then, an amount of phage calculated to provide the required MOI (e.g., 2 x 10⁻⁶). 7One pfu / g lung provides an MOI of 10. The MOI can be selected based on the "multiples of 10 rule," which states that when each bacterium adsorbs an average of approximately a multiple of 10 of phage, the bacterial density decreases significantly (Abedon ST, 2009, Foodborne Pathog Dis6:807-815; and Kasman LM et al., 2002, J Virol 76:5557-5564); while low-titer phage administration (e.g., using an MOI below 10) is unlikely to be successful (Goode D et al., 2003, App Environ Microbiol 69:5032-5036; Kumari S et al., 2010, J Infect Dev Ctries 4:367-377).

[0107] In some preferred embodiments, a phage mixture comprising F391 / 08, F17 / 19, and F58 / 19 is delivered to provide an MOI of 1 to 10 for each phage, resulting in a reduction of Klebsiella pneumoniae in the lungs by approximately 80%, approximately 85%, approximately 95%, approximately 97%, approximately 98%, or up to approximately 100% (with viable cell counts reduced to zero). In some preferred embodiments, a phage mixture further comprising (or administered in conjunction with) purified phages F99 / 10, F27 / 12, and F95 / 13 is delivered to provide an MOI of 1 to 10 for each phage, resulting in a reduction of Pseudomonas aeruginosa in the lungs by approximately 80%, approximately 85%, approximately 95%, approximately 97%, approximately 98%, or up to approximately 100%. In some particularly preferred embodiments, the phage mixture unexpectedly exhibits synergistic lytic activity compared to each phage alone.

[0108] Surprisingly, in some implementations, lower doses can provide advantages over higher doses. For example, in some implementations, an MOI of 1 or approximately 1 maintains low bacterial levels in the lungs for a longer period compared to an MOI of 10 or approximately 10. For instance, in the lungs of infected animals, lower MOIs of F391 / 08, F17 / 19, and F58 / 19 resulted in lower Klebsiella pneumoniae loads over a longer period post-treatment, while lower MOIs of F99 / 10, F110 / 10, and / or F27 / 12 resulted in lower Pseudomonas aeruginosa loads, for example, at 12, 15, 18, 24, 30, 36 hours, or longer post-treatment. It is not desirable to be bound by theory, but this could be due to a delayed response to lower doses of phages leading to the development of bacterial resistance.

[0109] In some other implementations, MOIs as low as approximately 0.2 to 0.4 can produce efficacy, for example, statistically significant reductions in the load of Klebsiella pneumoniae and / or Pseudomonas aeruginosa in the lungs of infected animals. Without being bound by any particular theory, efficacy may be attributed to active therapy. That is, a phage dose with an MOI of 10 provides sufficient phage to exceed the target bacterial population to reduce bacterial load without phage replication or completion of its life cycle. Lower phage doses may rely on active therapy, which involves the phage infection / replication cycle to reduce the target bacteria (Loc Carrillo C et al., 2005, Appl Environ Microbiol 71:6554-6563; see also Cairns BJ et al., 2009, PLoSPathog 5:e1000253; and Hooton SP et al., 2011, Int J Food Microbiol 151:157-163).

[0110] In some embodiments, the purified bacteriophages or bacteriophage cocktail compositions of the present invention are used as a single agent for the treatment or prevention of infections (e.g., respiratory or lung infections) caused by *Pseudomonas aeruginosa* and / or *Klebsiella pneumoniae*. In other embodiments, the bacteriophages or bacteriophage mixtures disclosed herein are further used in combination with other agents, including standard antibiotics targeting the same or different species of bacteria, including bacteria selected from any Gram-positive bacteria, any Gram-negative bacteria, and any other bacterial group not classified as Gram-positive or Gram-negative. The compositions of the present invention can also be used in combination with any other means of treating bacterial infections known to those skilled in the art, particularly with any other means of treating respiratory infections.

[0111] In some particularly preferred embodiments, the present invention provides a method for treating and / or preventing respiratory or pulmonary infections, comprising administering the bacteriophage mixture of the present invention in combination with standard and / or non-standard therapies. Standard therapies for respiratory infections include inhaled and / or systemic antibiotics, such as tobramycin, amikacin, colistin, aztreonam, as well as levofloxacin, ceftazidime, fosfomycin, gentamicin, vancomycin, amphotericin B, capreomycin, rifampin, isoniazid, and ciprofloxacin; and inhaled and / or systemic antifungal agents, such as amphotericin B.

[0112] In some embodiments, the phage, phage product, or phage cocktail composition of the present invention is administered in the form of an aerosol, while other agents are administered systemically. For example, in some preferred embodiments, the phage cocktail composition of the present invention is administered by inhalation, while an antibiotic, such as an antibiotic with activity against Pseudomonas aeruginosa and / or Klebsiella pneumoniae, is administered systemically. In some embodiments, the phage cocktail composition of the present invention is administered by inhalation together with other agents, also administered in the form of an aerosol. For example, in some preferred embodiments, the phage mixture pharmaceutical composition of the present invention is administered to the lungs in the form of an aerosol together with another antibiotic or antifungal agent.

[0113] In some embodiments, the present invention provides methods for treating and / or preventing respiratory or lung infections, comprising administering the bacteriophage, bacteriophage product, or bacteriophage cocktail composition of the present invention in combination with non-standard therapies for respiratory infections. Non-standard therapies are typically used when one or more standard therapies are ineffective against respiratory infections.

[0114] 5.5 Disinfectants and Anti-infective Uses

[0115] Bacterial pathogens most commonly infect at the mucous membranes (e.g., through the mucous membranes of the upper or lower respiratory tract, through the urogenital system, ocular structures, etc.). Mucous membranes themselves are often reservoirs, and sometimes the only reservoirs, of pathogenic bacteria present in the environment. Although studies have shown that reducing or eliminating such reservoirs (especially in environments such as hospitals and nursing homes) significantly reduces the incidence of infection, very few anti-infective agents are designed to control these pathogenic bacterial reservoirs.

[0116] The bacteriophages, phage products, and phage mixtures of the present invention can be used in anti-infective compositions to control the growth of bacteria, particularly Klebsiella pneumoniae and Pseudomonas aeruginosa, thereby preventing or reducing the occurrence of hospital-acquired infections. The anti-infective compositions can be used to reduce or inhibit bacterial colonization or growth on surfaces in contact with them. The bacteriophages, phage products, and phage mixtures of the present invention can be incorporated into compositions to formulate applications for biological surfaces such as skin and mucous membranes, as well as for non-biological surfaces.

[0117] Anti-infective formulations that can be used on biological surfaces include, but are not limited to, gels, creams, ointments, sprays, etc. In certain embodiments, the anti-infective formulation is used to sterilize the hands and / or exposed skin of medical personnel and / or patients in surgical settings. In a preferred embodiment, the biological surface is a mammalian mucosa, more preferably a human mucosa. In a particularly preferred embodiment, the biological surface is a respiratory mucosa, such as the nasal mucosa, or the lining of the pharynx, larynx, trachea, bronchi, and / or lungs.

[0118] Anti-infective formulations that can be used on non-biological surfaces include sprays, solutions, suspensions, and wipes impregnated with solutions or suspensions. In certain embodiments, the anti-infective formulations are used on solid surfaces in hospitals, nursing homes, ambulances, etc., including, for example, instruments, work surfaces, and medical devices and hospital equipment. In a preferred embodiment, the non-biological surface is the surface of hospital instruments or some hospital equipment. In a particularly preferred embodiment, the non-biological surface is a surgical instrument or some surgical equipment.

[0119] 5.6 Diagnostic methods

[0120] The present invention also includes diagnostic methods for identifying pathogenic factors in bacterial infections. In some embodiments, the diagnosis of pathogenic factors in bacterial infections is performed by: (i) culturing a sample from a patient, such as a swab, sputum, or other sample suitable for culturing bacteria causing the infection; (ii) contacting the culture with one or more phages, phage products, and phage mixtures of the present invention; and (iii) monitoring evidence of cell growth and / or lysis of the culture. Because the activity predisposition of phages and / or their isolated products (e.g., polypeptides, their bioactive fragments or variants, or nucleic acids encoding them) is species or strain specific, susceptibility or lack of susceptibility to one or more phages, phage products, and phage mixtures of the present invention can indicate the species or strain of bacteria causing the infection.

[0121] In some implementations, test cultures are obtained from a patient and contacted with one or more of F391 / 08, F17 / 19, or F58 / 19, or their variants. Decreased growth and / or lysis of the culture can indicate that the test sample contains Klebsiella pneumoniae, particularly strains susceptible to infection by the phages or phage mixtures disclosed herein, thereby identifying the infectious agent and allowing for appropriate diagnosis and / or treatment.

[0122] The sample can be a tissue biopsy or swab taken from the patient, or a fluid sample such as blood, tears, or urine. In a preferred embodiment, a tissue sample, such as a mucus sample, sputum, or a swab from the nostril, is obtained from the patient's respiratory tract.

[0123] 6. Example

[0124] It should be understood that the following embodiments and implementations described herein are for illustrative purposes only, and those skilled in the art will make various modifications or changes based on these embodiments and implementations, which will be included within the spirit and scope of this application and the scope of the appended claims.

[0125] Unless otherwise stated, the specific phages disclosed herein were isolated, processed, and analyzed according to the methods described below. Furthermore, in accordance with Portuguese law, the research described below was approved locally by the Animal Ethics Committee of the Institute of Molecular Medicine and nationally by the Portuguese Directorate General of Veterinary Services (DGV). Geral de The study was conducted in accordance with European Guideline 86 / 609 / EC (Council Guideline 86 / 609 / EEC of 24 November 1986 concerning the harmonization and uniformity of laws, regulations and administrative provisions of the Member States relating to the protection of animals for laboratory and other scientific purposes, Off J Eur Communities L358:1-28), Portuguese law (Regulation 1005 / 92) (Portuguese Ministry of Agriculture, Regulation 1005 / 92 of 23 October concerning the protection of animals for laboratory and other scientific purposes, Diário da República I–Série B 245:4930-4942), and the Guidelines for the Management and Use of Laboratory Animals (NRC 2011) (Institute for Laboratory Animal Research. 2011. Guidelines for the Management and Use of Laboratory Animals. Washington (DC): National Academies Press.).

[0126] One objective of this study was to investigate the antimicrobial activity of a nebulized phage mixture against *Pseudomonas aeruginosa* and *Klebsiella pneumoniae* in a mouse model of acute lung infection. The aerosol form of the phage allows for direct delivery to the infected lung region, overcoming some of the side effects of nebulized antibiotics.

[0127] 6.1 Example 1 - Bacterial strains

[0128] Between 2005 and 2019, a selected group of Klebsiella pneumoniae bacterial isolates (n=36) were collected from at least seven different healthcare facilities. Overall, these isolates were collected from hospital environments (n=25), outpatients (n=9), or sources of unknown origin (n=2), and from a wide variety of biological products [urine (n=10), respiratory secretions (n=5), unknown (n=8), ascites (n=5), and others (n=8)]. The group was assessed for antibiotic susceptibility using a disc diffusion method against a selected group of clinically important antibiotics (β-lactam antibiotics—ampicillin, ceftazidime, piperacillin, tazobactam, and meropenem; fluoroquinolones—ciprofloxacin; aminoglycosides—gentamicin; and sulfonamides—trimethoprim and sulfamethoxazole). The results were interpreted according to the cutoff values ​​recommended by the European Committee for Testing Susceptibility to Antimicrobial Agents (EUCAST, http: / / mic.eucast.org / Eucast2 / ). An isolate is considered to have multidrug resistance (MDR) when it is not susceptible to three or more structurally unrelated classes of antibiotics (Magiorakos AP, Srinivasan A, Carey RB., Carmeli Y, Falagas ME et al. 2011. Clin Microbiol Infect 2012; 18:268–281).

[0129] This group of Klebsiella pneumoniae bacterial isolates (n=36) was also characterized by MLST, as previously reported, to assess genetic diversity, clonal set, and global prevalence. The generated sequences were then analyzed using Finch TV and assigned to their respective STs using tools available on the MLST website (https: / / pubmlst.org). Each bacterial isolate was streaked onto tryptone soy agar plates (TSA, Biokar Diagnostics, Pantin Cedex, France) and incubated at +37°C for 18 h. All clinical strains were stored at -70°C in tryptone soy broth containing 15% glycerol (w / v) (TSB, Biokar Diagnostics, Pantin Cedex, France) until needed.

[0130] For in vitro experiments, strains frozen at -70°C were grown overnight at 37°C for 18 hours on a TSA. Single colonies were then grown overnight in a TSB under shaking conditions at 37°C. Fresh bacterial suspensions (diluted overnight cultures) were prepared and incubated with shaking at 37°C. Bacteria were harvested when the exponential growth phase was reached (optical density of 0.3–0.5 at 600 nm). Approximately 2.0 × 10⁻⁶ cells were used in the lysis curve.6 Inoculum of cfu / mL.

[0131] 6.2 Example 2 - Phage source, amplification, and mixtures

[0132] 6.2.1 Source of bacteriophage

[0133] Vicious phages of Klebsiella pneumoniae F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 were isolated from sewage in the Lisbon area and amplified in clinical strains of Klebsiella pneumoniae 397 / 07 (F391 / 08), Klebsiella pneumoniae 237 / 14 (Kle_F17 / 19), and Klebsiella pneumoniae 57 / 17 (Kle_F58 / 19).

[0134] To isolate lysing phages resistant to Klebsiella pneumoniae, several clinical strains were used. Wastewater from various sources in the Lisbon city area was tested to determine the presence of phages based on their ability to infect clinical strains of Klebsiella pneumoniae, using a double agar overlay plaque assay (Kropinski A, Mazzocco A, Waddell TE, Lingohr E, Johnson RP. 2009. Enumeration of bacteriophages by double agaroverlay plaque assay. Methods Mol Biol 501:69-76.).

[0135] In short, bacterial strains were grown overnight in TSB at +37°C with shaking. Fresh bacterial suspensions (diluted overnight cultures) were prepared, incubated at +37°C with shaking, and harvested when the exponential growth phase (optical density at 600 nm of 0.3–0.5) was reached. Each culture was added to an aqueous sample, and 3 ml of pre-equilibrated 0.7% soft agar was added to the mixture during incubation at +37°C for 30 minutes. The agar-water-bacterial suspension was then spread onto 1.5% TSA plates, allowed to harden at room temperature, and incubated at +37°C. After 18 hours of incubation, the presence of phage plaques (clear zones) within the bacterial growth on the plates was examined, indicating the presence of phages. Phage plaques were picked up using sterile pipette tips, transferred to SM buffer, and stored at +4°C.

[0136] 6.2.2 Phage amplification

[0137] Before assessing the host range, newly isolated phages were propagated, amplified, and purified (three consecutive elutions) in indicator strains. The susceptibility of 36 Klebsiella pneumoniae bacterial isolates to specific phages was investigated using a double-layer agar plaque assay. The susceptibility of the 36 bacterial isolates to specific phages was determined by observing phage plaques within the bacterial flora. Phages with the highest percentage of infection within the host range were screened, and then subjected to amplification, high-speed centrifugation concentration, cesium chloride (CsCl) gradient purification, phage genomic DNA extraction, and restriction fragment length polymorphism analysis.

[0138] Phages with unique restriction maps and a broader host range were selected for sequencing. The whole genome sequence was determined by pyrosequencing using an Illumina HiSeq 2000 genome analyzer.

[0139] Following bioinformatics analysis, the most promising phages were selected to form the therapeutic mixture. Morphological analysis of Klebsiella pneumoniae F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 phages was performed at the Histology and Comparative Pathology Laboratory of the Institute of Molecular Medicine in Lisbon, Portugal. These data were combined with genomic analysis, and the phages were classified according to the Ackermann classification method (Ackermann HW. 2009. Phage classification and characterization. Methods Mol Biol 501:127-140).

[0140] 6.3 Example 3 Phage analysis

[0141] 6.3.1 Phenotypic Characterization

[0142] Phenotypic characterization of the collected bacteria showed that, among β-lactam antibiotics, 100% of the isolates were not susceptible to ampicillin, 97.2% were not susceptible to piperacillin plus tazobactam, 86.1% were not susceptible to ceftazidime, and 77.8% were not susceptible to meropenem. Regarding non-β-lactam antibiotics, this group of bacterial isolates showed that 91.7% were not susceptible to ciprofloxacin, 72.2% were not susceptible to gentamicin, and 88.9% were not susceptible to trimethoprim and sulfamethoxazole. Overall, 33 / 36 (91.7%) of the isolates were MDR (Table 3).

[0143] Table 3. Phage infection, phenotype, and genotype characterization of selected bacterial isolate subsets (n=36).

[0144]

[0145]

[0146]

[0147] Furthermore, interpretation of antibiotic susceptibility tests revealed the production of broad-spectrum β-lactamases (ESBLs) or carbapenemases (Table 1). Overall, most isolates showed low susceptibility to last-resource antibiotics, such as carbapenems (23 / 30, 76.6%), MDRs (n = 22 / 30, 73.3%), and produced clinically important carbapenemases or ESBLs (16 / 30, 53.3%), such as the globally prevalent VIM-2, KPC-2, and even NDM-1.

[0148] Clonal lineage identification revealed high genetic diversity represented by 19 different STs, particularly high-risk clones (ST11, ST13, ST14, ST15, or ST258) that exhibit widespread prevalence globally and have actually contributed to disease outbreaks and high morbidity and mortality (Brink, AJ, 2019. Epidemiology of carbapenem-resistant Gram-negative infections globally. Curr Opin Infect Dis 32, 609-616; Karakonstantis, S., Kritsotakis, EI, Gikas, A., 2019. Pandrug-resistant Gram-negative bacteria: a systematic review of current epidemiology, prognosis and treatment options. J Antimicrob Chemother.; Li, J., Li, Y., Song, N., Chen, Y., 2019. Risk factors for carbapenem-resistant Klebsiella pneumoniae infection: a meta-analysis. J Glob Antimicrob Resist.; Navon-Venezia, S., Kondratyeva, K., Carattoli, A., 2017. Klebsiella pneumoniae: a major worldwide source and shuttle for antibiotic resistance. FEMS Microbiol Rev 41, 252-275; Sarda, C., Fazal, F., Rello, J., 2019. Management of ventilator-associated pneumonia (VAP) caused by resistant Gram-negative bacteria: which is the best strategy to treat? Expert Rev Respir Med 13,787-798).

[0149] 6.3.2 Phage Morphology

[0150] Transmission electron microscopy was used to classify purified bacteriophages based on their viral morphology. Klebsiella pneumoniae F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 bacteriophages belong to the order Caudovirales. The F391 / 08 bacteriophage exhibits an icosahedral head and a long, non-retractable, and usually flexible tail. These characteristics, along with its genomic features, allow us to classify this bacteriophage as a member of the family Siphoviridae. The Kle_F17 / 19 and Kle_F58 / 19 bacteriophages are classified as members of the family Myoviridae, exhibiting a retractable tail and an icosahedral head (capsid), and possessing a basal plate structure and tail filaments (…). Figure 1 ).

[0151] 6.3.3 Phage Host Range

[0152] Selected phages of Klebsiella pneumoniae were tested in 36 clinical strains isolated from human clinical samples collected and identified from hospitals in the Lisbon area. Of this diverse group of clinically important Klebsiella pneumoniae isolates, 53.8% were infected with at least one phage comprising the mixture (Table 4).

[0153] Table 4. Percentage and total coverage of each Klebsiella pneumoniae phage strain within the host range.

[0154]

[0155] In fact, Klebsiella pneumoniae is a major MDR pathogen affecting humans and a leading source of hospital-acquired infections, associated with high morbidity and mortality due to limited treatment options. These strains can actually develop resistance to all available antibiotic classes, posing a significant challenge to clinicians due to the limited treatment options available (Brink, AJ, 2019. Epidemiology of carbapenem-resistant Gram-negative infections globally. Curr Opin Infect Dis 32, 609-616; Navon-Venezia, S., Kondratyeva, K., Carattoli, A., 2017. Klebsiella pneumoniae: a major worldwide source and shuttle for antibiotic resistance. FEMS Microbiol Rev 41, 252-275).

[0156] Since the first emergence of carbapenemase-producing Klebsiella pneumoniae, nearly 100 international outbreaks caused by high-risk clones have been reported. The most serious outbreaks were caused by clone group 258, accounting for 68% of all outbreaks, consisting of three STs: ST258, ST11, and ST512. The second most prevalent clone group was clone group 15, consisting of ST14 and ST15, accounting for approximately 20% of all outbreaks. Other important STs have caused outbreaks and clinically significant infections to a lesser extent (ST147, ST37, ST101, ST17) (Navon-Venezia, S., Kondratyeva, K., Carattoli, A., 2017. Klebsiella pneumoniae: a major worldwide source and shuttle for antibiotic resistance. FEMS Microbiol Rev 41, 252-275).

[0157] Overall, infections caused by high-risk carbapenem-resistant Klebsiella pneumoniae clones are a major health problem and have caused high mortality rates on numerous occasions worldwide. Klebsiella pneumoniae bacteriophages are able to effectively eliminate isolates that not only exhibit carbapenem resistance caused by acquired carbapenemases but also belong to five of the nine globally prevalent high-risk clones (ST11, ST14, ST15, ST258, and ST147).

[0158] 6.3.4 Genome Analysis

[0159] Whole-genome sequencing was performed on the genomic DNA of Klebsiella pneumoniae F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 phages. Following annotation, a circular map of each genome was plotted, showing the predicted ORFs encoding putative proteins and their putative functions (Figures 2-4). Preliminary NCBI nucleotide blast analysis (blastn) was performed on the whole-genome sequences of the selected phages. Klebsiella pneumoniae phage F391 / 08 showed high significant homology with Klebsiella pneumoniae phage vB_Kpn_IME260 (NCBI reference sequence: KX845404.2). Kle_F17 / 19 phage showed the highest similarity to Klebsiella pneumoniae phage KOX1 (NCBI reference sequence: KY780482.1). The genome sequence of phage Kle_F58 / 19 showed high homology with other sequences in the NCBI database. The highest similarity was found to Klebsiella pneumoniae phage vB_KpnM_Potts1 (NCBI reference sequence: MN013081.1).

[0160] The genome size of phage F391 / 08 is 113073 bp (Figure 2), and it shares up to 95% sequence identity with Klebsiella pneumoniae phage vB_Kpn_IME260 at 96% genome coverage. 173 ORFs were predicted, and 28% of the inferred functions were resolved, including 25 tRNA genes. One of the predicted ORFs showed no significant homology with any sequence from the NCBI Non-Redundant Protein Sequence Database. 90 of the predicted ORFs showed homology with proteins from the NCBI Non-Redundant Protein Sequence Database, but their inferred functions could not be resolved. Genome analysis of phage Kle_F17 / 19 showed a 45423 bp sequence (…) in 94% of the genome. Figure 3 The ORF showed up to 96% identity with the myotail bacteriophage KOX1 (Brown TL, Petrovski S, Hoyle D, Chan HT, Lock P, Tucci J. 2017. Characterization and formulation into solid dosage forms of a novelbacteriophage lytic against Klebsiella oxytoca. PLoS ONE 12(8):e0183510). 50% of the 64 predicted ORFs showed resolvable inferred function. Inferred function could not be resolved in 32 ORFs. Phage Kle_F58 / 19, with a 169725 bp genome (Figure 4), showed high similarity to the myotail bacteriophage vB_KpnM_Potts1. Sequence identity was as high as 97.5% in 98% of the genome sequence. 292 ORFs were predicted, and 45.9% of the inferred functions were resolved, including 7 tRNA genes. Approximately 45% of the predicted ORFs showed homology with sequences from the NCBI Non-Redundant Protein Sequence Database, but no functions were resolved. Only one ORF showed no significant homology with any sequence from the NCBI Non-Redundant Protein Sequence Database.

[0161] No significant similarity was found in the sequences of these bacteriophages to known virulence or toxin proteins or to elements typically associated with lysogenicity (integers, repressors, and antirepressors).

[0162] 6.3.5 Lytic activity of bacteriophage mixtures

[0163] To characterize the phage mixture, the lytic activity of novel Klebsiella pneumoniae F391 / 08, Kle_F17 / 19 and Kle_F58 / 19 phages on planktonic cultures was evaluated.

[0164] Plot conventional lysis profiles under controlled conditions using a pre-determined bacterial inoculum. Use approximately 2 x 10⁻⁶ bacteria. 6 Cultures were prepared from inoculum at a concentration of 1 CFU / mL. Two different Klebsiella pneumoniae strains, 57 / 17 and 237 / 14, were used because highly virulent strains simultaneously infected with all three phages were not found in our bacterial collection. Each phage was individually ( Figure 5 and Figure 6 ) and combination ( Figure 7 The test was performed with an MOI approximately equal to 10. Viable bacterial counts were monitored every hour for 8 hours and again 24 hours after phage inoculation (ppi).

[0165] Phage F391 / 08 was tested individually when the MOI was approximately 10. Figure 5 Furthermore, 1 hour after phage inoculation (ppi), the bacterial load of this culture was compared with that of the bacterial control culture (8.4 x 10⁻⁶). 6 Compared to (cfu / mL), it decreased by approximately 3 log orders (2.3 x 10⁻⁶). 3 (cfu / mL). Within the first 4 hours, F391 / 08 was able to maintain a viable bacteria count below 10. 4 CFU / mL. The bacterial load in the culture began to increase, and at 8 h ppi, the culture exceeded the initial bacterial load (3.8 x 10⁻⁶ CFU / mL). 7 (cfu / mL). However, at 24h ppi, the viable count was 8.8 x 10⁻⁶. 7 CFU / mL, compared to the control culture (1.2 x 10⁻⁶). 11 The number of CFU / mL was significantly lower than that of KLE57 / 17, indicating that F391 / 08 retained the ability to infect KLE57 / 17 even during the stationary phase.

[0166] Phage Kle_F17 / 19 was tested against Klebsiella pneumoniae 237 / 14 at an MOI of approximately 10, and within 1 hour, compared with the control culture of bacteria (1.0 x 10⁻⁶). 7 Compared to (cfu / mL), it reduced the viable bacterial count by approximately 5 log units (5.0 x 10⁻⁶ CFU / mL). 2 (cfu / mL). Until 4 hours post-pigmentation (ppi), Kle_F17 / 19 phage was able to maintain the bacterial load in the culture below 10⁻⁶ CFU / mL. 4 CFU / mL, as observed with F391 / 08 in its host. At 8 h ppi, the viable bacterial count rapidly increased to 1.1 x 10⁻⁶ CFU / mL. 8 The viable bacterial count was 3.1 x 10⁻⁶ CFU / mL and continued until the end of culture incubation (24 h ppi), at which point the viable bacterial count was 3.1 x 10⁻⁶ CFU / mL.10 One CFU / mL, almost one log order higher than the measured value of the control culture.

[0167] Kle_F58 / 19 phage ( Figure 6 The effect on strain 57 / 17 was faster than that on phage F391 / 08. At 1 h ppi, the viable bacterial count decreased by approximately 4 log units compared to the control culture, reaching 2.0 × 10⁻⁶. 2 CFU / mL. At 8 hours post-infection (ppi), viable bacteria increased, with a count of 1.8 x 10⁻⁶ at this time point. 7 CFU / mL. At 24h ppi, phage Kle_F58 / 19 was no longer as effective against the host (e.g., F391 / 08), but still achieved a 96% reduction in viable cells, which was a very significant reduction compared to the bacterial control culture.

[0168] In separate cases, although the three bacteriophages exhibited high lytic activity, their efficacy in their respective hosts varied slightly over time, which may reflect differences in adsorption rate, latency, and lysis rate (data not shown).

[0169] Figure 7 The co-lysis curves of phages F391 / 08 and Kle_F58 / 19 in Klebsiella pneumoniae 57 / 17 at an MOI of approximately 10 were plotted. At ppi 1 h (1.0 x 10⁻¹⁰), the lysis curves were obtained. 1 At 100 cfu / mL, a significant decrease in phage activity was observed. Over the next 8 hours, although the synergistic effects of the phages changed somewhat, the viable cell count remained below 10. 4 The cell count was increased to 10⁵ CFU / mL, thus prolonging the interaction time of each phage as observed in individual cases. Despite bacterial regrowth, the cell count at 24 h PPI was 5.1 x 10⁻⁶. 7 CFU / mL, compared to the bacterial load of the control culture (4.2 x 10⁻⁶). 10 Compared to (cfu / mL), it showed a significant reduction (approximately 3 log units).

[0170] In addition to the lysis curves of F391 / 08 and Kle_F58 / 19, a second co-culture (MOI of approximately 10) was prepared using three phages, F391 / 08, Kle_F58 / 19, and Kle_F17 / 19, to observe any effect of Kle_F17 / 19 on the effectiveness of F391 / 08 and Kle_F58 / 19. Although Klebsiella pneumoniae 57 / 17 is not readily infected by phage Kle_F17 / 19, its presence appeared to induce a stabilizing effect on the efficacy of phages F391 / 08 and Kle_F58 / 19 when used in combination during the first 6 hours of culture. Figure 9 At 1 h ppi, the decrease in cell count was not significant compared to the curves for F391 / 08 and Kle_F58 / 19, but the behavior over the following hours indicated an interaction with Kle_F17 / 19. At 8 h ppi, compared to 5.3 × 10⁻⁶ cells in the control culture... 9 Compared to 1 CFU / mL, the bacterial load was lower, at 5.3 × 10⁻⁶. 3 The cell count was 4.7 × 10⁶ CFU / mL at the end of the culture incubation. 7 One CFU / mL, a concentration close to the bacterial concentration introduced at the start of the culture.

[0171] As previously reported by Loc-Carrillo and Abedon in 2011 (Loc-Carrillo C, Abedon ST. 2011. Pros and cons of phage therapy. Bacteriophage 1(2):111-114), different phages can be mixed in a mixture to enhance their efficiency, generally resulting in a larger overall spectrum of antibacterial activity. This study demonstrates this point.

[0172] 6.3.6 Conclusion

[0173] The phage mixture characterized and tested in this study consisted of three Klebsiella pneumoniae phages: F391 / 08, Kle_F17 / 19, and Kle_F58 / 19. Genomic sequence analysis did not identify any known genes associated with integration, toxins, or antibiotic resistance—important characteristics for the safe use of phages. Comparative genomics analysis revealed significant similarity between Klebsiella pneumoniae phages F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 and Klebsiella pneumoniae phages vB_Kpn_IME260, KOX1, and vB_KpnM_Potts1, respectively.

[0174] Klebsiella pneumoniae phage vB_Kpn_IME260 is a T5-like virus (Xing S, Pan X, Sun Q, Pei G, An X, Mi Z, Huang Y, Zhao B, Tong Y. 2017. Complete genome sequence of a novel multidrug-resistant Klebsiella pneumoniae phage, vB_Kpn_IME260. Genome Announc 5:e00055-170). The Klebsiella oxytoca phage KOX1 showed the highest homology with Kle_F17 / 19. Brown et al. (Brown TL, Petrovski S, Hoyle D, Chan HT, Lock P, Tucci J. 2017. Characterization and formulation into solid dosage forms of a novelbacteriophage lytic against Klebsiella oxytoca. PLoS ONE 12(8):e0183510) described KOX1 as belonging to the Myotail Phage Family. Apart from morphological information, there is no information on the vB_KpnM_Potts1 phage in the literature. Transmission electron microscopy analysis confirmed that phages Kle_F58 / 19 and Kle_F17 / 19 belong to the Myotail Phage Family.

[0175] Another important selection criterion for bacteriophages is their host range, which should be as broad as possible, especially including clinically prevalent bacterial species (Gill JJ, Hyman P. 2010. Phage choice, isolation, and preparation for phage therapy. Curr Pharm Biotechnol 11(1):2-14). The lytic activity of Klebsiella pneumoniae bacteriophages F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 was tested on 36 clinical strains with high genomic diversity isolated from hospitals in the Lisbon metropolitan area over the past few years.

[0176] Prior to in vivo efficacy studies, time-kill curves are typically used to investigate the antimicrobial effects and dosing regimens of single and combined drug compounds (NCCLS. 1999. Methods for determining bactericidal activity of antimicrobial agents: approved guideline. 940 West Valley Road, Suite 1400, Wayne, Pennsylvania 19087, USA: NCCLS). In this study, lysis curves were plotted to illustrate the effect of phage lysis capacity (alone and in combination) on bacterial cultures. Single bacterial inoculum was used. The values ​​were carefully selected based on previous evidence (Mendes JJ, Leandro C, Motolla C, Barbosa R, Silva F, Oliveira M, Viela C, Melo-Cristino J, Gorski A, Pimentel M, C, Cavaco-Silva P, Garcia M. 2014. In vitro design of a novel lytic bacteriophage cocktail with therapeutic potential against organisms causing diabetic footinfections. J Med Microbiol 63:1055-1065. Between the first and second hours post-phage inoculation, all cultures showed initial bacterial reduction, followed by significant regrowth after 6 hours, and even more pronounced after 24 hours. Individually, all three phages exhibited high lytic efficacy against planktonic cells. Bacterial reductions of 95% to 99% at 24 hours of culture indicated their enhanced ability to control Klebsiella pneumoniae load (except for Kle_F17 / 19). The combined activity of two of the three phages in the mixture demonstrated synergistic lytic activity against bacterial strains. The selected MOI was chosen based on the "rule of 10," which states that if the goal is to significantly reduce bacterial density, then efforts should be made to adsorb approximately 10 multiples of phages per bacteriophage on average (Abedon ST. 2009. Kinetics of phage-mediated biocontrol of bacteria. Foodborne Pathog Dis 6:807-815; Kasman LM, Kasman A, Westwater C, Dolan J, Schmidt MG, Norris JS. 2002. Overcoming the phage replication threshold: a mathematical model with implications for phage therapy. J Virol 76:5557-5564). Regeneration was observed in planktonic cells exposed to the phages.

[0177] 6.3.7 Example 4 - In vitro interactions between Pseudomonas aeruginosa phage mixtures and antibiotics

[0178] In vitro techniques were used to evaluate the potential interactions between a mixture of bacteriophages F99 / 10, F27 / 12, and F95 / 13 and selected antibiotics frequently used as empirical treatment for hospital-acquired pneumonia. While the mixture targets Gram-negative bacteria, empirical antibiotic therapy typically includes drugs used to treat pneumonia caused by Gram-positive pathogens; therefore, molecules targeting Staphylococcus aureus were also involved.

[0179] In short, the culture microdilution method was used to determine the minimum inhibitory concentrations (MICs) against β-lactams (piperacillin, tazobactam, ceftazidime, and meropenem), aminoglycosides (amikacin), glycopeptides (vancomycin), and oxazolidinones (linezolid) at different MOIs (0.1, 1, and 10), in the presence or absence of single and combined phages. This method was developed and optimized as a proof-of-concept for *Pseudomonas aeruginosa* using a single strain infected with all three phages (PSA_1992 / 05; NCIMB accession number 42914). The selected MIC range was 0.016 to 32 mg / L, allowing for the inclusion and detection of susceptibility and resistance breakpoints for all seven antibiotics. The technique was optimized using different time points (4 h, 6 h, 12 h, and 18 h), but the final reading was determined at 18 h. All experiments were replicated. This study optimized a single phage and presents the final assay here, which includes data for the complete mixture.

[0180] In summary, the final tests conducted with the mixture (phages Psa_F99 / 10, Psa_F27 / 12, and Psa_F95 / 13) confirmed the results obtained for each individual phage (data not shown). For all antibiotics tested, as well as all three MOIs (0.1, 1, and 10) tested during this study, the potency of the mixture was sufficient to reduce the MIC of PSA_1992 / 05 from a much higher MIC value to ≤0.016 mg / L (Table 5).

[0181] Table 5. MICs of Pseudomonas aeruginosa PSA_1992 / 05 at MOIs of 0.1, 1, and 10 in the presence of a phage mixture.

[0182]

[0183]

[0184] Infection of PSA_1992 / 05 with a phage mixture (mixture activity control reading) showed no inhibitory effect on bacterial growth at 18 h. TZP, piperacillin and tazobactam; CZP, ceftazidime; MEM, meropenem; AK, amikacin; VA, vancomycin; LZD, linezolid; CS, colistin. NA, not applicable.

[0185] This data indicates that the combined potency of the mixture of F99 / 10, F27 / 12, and F95 / 13 is increased in the presence of these antibiotics. No negative effects from the addition of the phage mixture were observed.

[0186] Overall, while vancomycin and linezolid are antibiotics that do not exhibit any activity against Gram-negative bacteria, their presence did not inhibit the activity of the mixture. Major antibiotic classes used to treat hospital-acquired pneumonia have been evaluated in combination with individual phages for proof-of-concept and optimization purposes, and also in combination with phage mixtures. Overall, no interactions that could lead to phage inhibition or a significant reduction in antibiotic activity were detected.

[0187] 6.3.8 Example 5 - Inhalation phage application feasibility Research

[0188] Nebulization has been increasingly used in inhalation therapy, specifically the use of bacteriophages in animal models via powdered formulations (Chang, RYK, Chen, K., Wang, J., Wallin, M., Britton, W., Morales, S., Kutter, E., Li, J., Chan, HK, 2018. Proof-of-Principle Study in a Murine Lung Infection Model of Antipseudomonal Activity of Phage PEV20 in a Dry-Powder Formulation. Antimicrob Agents Chemother 62,2). Here, a feasibility study of inhaled bacteriophage applications is conducted using liquid formulations and various options of vibrating screen nebulizers. These medical-grade nebulizers are designed for use not only in hospital wards but also in intensive care units, and can be readily connected to any standard respiratory equipment tubing.

[0189] To compare three vibrating screen nebulizer devices (Aeroneb, Nebutec 4, and Nebutec 6), phage suspensions containing 10... were prepared using DPBS (first experiment) or 0.9% NaCl solution as diluents (second experiment). 9Mixtures of each phage (F99 / 10, F27 / 12, and F95 / 13) at pfu / ml were used. Three independent nebulization experiments were performed using all three devices for each diluent. At each nebulization, 1 ml of the phage mixture was nebulized over a 1-minute timeframe, followed by a 60-minute settling period. The phages were collected in petri dishes containing 15 ml of the corresponding collection buffer. Phage titers were determined using specific detection (titration) strains for each phage. Droplet sizes for each device were determined experimentally using nebulization and laser diffraction, consistent with manufacturer information. Furthermore, the aerosol delivery rate was experimentally determined, and the probability of lung deposition was calculated based on measured data, indicating that Aeroneb and Nebutec 4 achieved similar deposition rates, with Aeroneb outperforming Nebutec 6 (data not shown). Notably, only 75% ± 8.6% of the nebulized liquid successfully deposited on the petri dishes due to droplet adhesion to the nebulizer tube. Overall, phage recovery was higher with NaCl as the diluent compared to DPBS. Furthermore, the Aeroneb device produced the most consistent recovery rates for all three phage types (data not shown).

[0190] Based on the results, the Aeroneb device and NaCl were selected as the media. Overall, the results allowed for the selection of the Aeroneb vibrating screen nebulizer device, which was then applied to residual potency and toxicity studies conducted in a PCLS in vitro model (see Example 6). The recovery and logarithmic reduction of phage titers were consistent between the two formulations and between different phages, and were also consistent with the expected phage loss throughout the system.

[0191] 6.3.9 Example 6- In the in vitro model of precision lung slices (PCLS) efficacy and toxicity studies

[0192] To evaluate the potency and toxicity of phages F99 / 10, F27 / 12, and F95 / 13, a comprehensive in vitro study was meticulously designed in a PCLS Pseudomonas aeruginosa infection model. The system was programmed and optimized in rat PCLS, where rat PCLS were treated with a mixture of purified F99 / 10, F27 / 12, and F95 / 13 via immersion and nebulization, both with and without antibiotic co-treatment. The system was then transferred to a final human PCLS setting. To evaluate the potency and toxicity of phage inhalation applied to Pseudomonas aeruginosa-infected human lung tissue, alone or in combination with antibiotics, experiments were conducted using human PCLS Pseudomonas aeruginosa. The nebulized phage mixture was applied directly to tissue cultured at the air-liquid interface, while the antibiotic (at a subinhibitory concentration of 0.5 μg / ml) was applied via immersion (i.e., into the culture medium). Furthermore, uninfected controls were treated with the nebulized phage mixture to exclude phage cytotoxicity in human lung tissue. Similarly, for rat PCLS (data not shown), both nebulized phage treatment and antibiotics reduced bacterial load in infected human PCLS to similar degrees, and their combined treatment produced a strong synergistic effect in reducing bacteria.

[0193] Phage treatment alone reduced bacterial load by more than one log, while antibiotic treatment alone resulted in a CFU reduction of approximately four logs. In the combined treatment, a synergistic effect was observed, resulting in a bacterial load reduction of more than five logs. Figure 8 No cytotoxic effects of purified phage on uninfected tissues were observed during tissue viability analysis. Treatment of infected PCLS with purified phage completely salvaged viability loss caused by Pseudomonas aeruginosa infection. Antibiotic treatment alone or in combination with phage treatment restored tissue viability to comparable levels. Figure 9 In uninfected human PCLS, phages comparable to the inoculum quantity were detectable 24 h post-treatment. Conversely, phage replication was observed in infected samples, but not in infected samples co-treated with antibiotics, consistent with the observed differences in bacterial load (data not shown).

[0194] Overall, studies conducted in an in vitro Pseudomonas aeruginosa PCLS model demonstrated that treatment of infected rat / human slides with a mixture of F99 / 10, F27 / 12, and F95 / 13 not only showed efficacy through immersion and nebulization but also restored lung tissue viability without toxic effects, even under combined antibiotic therapy (meropenem, amikacin, and vancomycin). Analysis of uninfected slides treated with this mixture under multiple conditions (immersion vs. nebulization; rats vs. humans; phage lysates vs. purified phages) showed no negative effect on lung tissue viability, further confirming the absence of predictable toxicity, particularly cytotoxicity, in the lungs associated with phage treatment. This evidence is consistent with previous studies that have shown no adverse effects from phage treatment in other infection models and compassionate use cases (McCallin, S., Sacher, JC, Zheng, J., Chan, BK, 2019. Current State of Compassionate Phage Therapy. Viruses 11, 343; Oliveira, A., Sereno, R., Nicolau, A., Azeredo, J., 2009. In vivo toxicity study of phage lysate in chickens. Br Poult Sci 50, 558-563).

[0195] For different routes of administration, nebulized phage treatment successfully reduced the bacterial load in infected PCLS, thus demonstrating efficacy comparable to immersion treatment, despite the expected phage attrition in the nebulizer. An increase in phage titer was observed near 24 h. Phage titer determination in the inoculum indicated that the number of phages in the samples with and without the antibiotic mixture was comparable, therefore the presence of antibiotic did not affect phage titer or activity. On the other hand, it was also confirmed that the mixture did not interfere with antibiotic activity. In fact, phage / antibiotic co-treatment completely restored tissue viability. Overall, this combination was also supported by all safety endpoints in infected and uninfected rat / human lung sections.

[0196] 6.3.10 Example 7 - Precision lung section (PCLS) study

[0197] In an in vitro model of Klebsiella pneumoniae infection using precisely sectioned lung slices (PCLS), the efficacy of inhaled Klebsiella pneumoniae F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 bacteriophages against respiratory bacterial infections was investigated. This study included confirming the biological activity of the nebulized phages, establishing the Klebsiella pneumoniae infection model, and evaluating the activity of the nebulized phages in rat PCLS.

[0198] The experiment was conducted at the Fraunhofer ITEM Department of Preclinical Pharmacology (Hannover, Germany). The euthanasia of animals for organ harvesting in the preparation of PCLS was registered with the relevant authorities (Lower Saxony Federal State Office for Consumer Protection and Food Safety) and carried out in accordance with the German Animal Protection Act (German Animal Protection Act of May 18, 2006, BGBl.I S. 1206, 1313; adopted July 28, 2014, BGBl.I S. 1308) and the European Parliament and Council Guideline 2010 / 63 / EU on the Protection of Animals for Scientific Purposes.

[0199] Female rats (Wistar WU, 8-21 weeks old at the time of the experiment) were obtained from Charles River (Sulzfeld, Germany). Animals were kept under standard feeding conditions (22°C, 55% humidity, and a 12-h diurnal rhythm) prior to PCLS preparation. Rats were euthanized by exsanguination via the vena cava after administering an excess of sodium pentobarbital (Narcoren, Merial GmbH, Hallbergmoos, Germany). Whole lungs of the rats were filled with a warm, low-melting-point agarose / medium solution (final concentration 1.5% (v / v)) at 37°C. The filled lung lobes were cooled in ice-cold PBS to allow agarose polymerization. Cylindrical specimens with a diameter of 10 mm were drilled from the lung tissue and sectioned into sections approximately 300 μm thick using a Krumdieck microtome (Alabama Research and Development, Munford, AL, USA). The sections were collected in cold EBSS at 4°C and then…

[0200] Transferred to culture dishes containing PCLS medium and incubated under cell culture conditions (37°C, 5% CO2) for approximately 30 minutes. Subsequently, sections were washed four times with PCLS medium for 30 minutes each time, and finally incubated overnight under cell culture conditions before the infection assay. Klebsiella pneumoniae test and detection strains (57 / 17, 131 / 15, 130 / 14, and 25 / 14) were grown overnight in TSB medium at 37°C and 150 rpm. The overnight cultures were diluted 1:30 and incubated for another 1 h at 37°C and 150 rpm to produce cultures in the exponential growth phase. Plaque formation assays were performed using detection strains with OD600 of 0.3–0.5. Cultures of the infected strain 57 / 17 were adjusted to 2 x 10⁻⁶. 6 One CFU / ml, and the inoculum sample was plated to confirm the CFU concentration.

[0201] Phage atomization was performed using a vibrating screen nebulizer (AeroNeb). In short, phages containing 10... 9 A mixture of each phage at PFU / ml was produced by atomizing 1 mL of phage mixture over a 1-minute time interval, followed by a 60-minute settling period.

[0202] For the nebulization pretest (without PCLS), a mixture of phages F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 was prepared. The phages were nebulized and collected in petri dishes containing 15 ml of the medium as a collection buffer (0.9% NaCl). The biological activity of the phages was determined by the double-layer agar plaque assay.

[0203] The efficacy of phage atomization was evaluated in rat PCLS to assess the effectiveness of phage atomization on PCLS in a Klebsiella pneumoniae infection model. Therefore, rat PCLS were cultured at the air-liquid interface (ALI) in 12-well transwell plates (500 μl reservoir volume) and infected with Klebsiella pneumoniae strain 57 / 17 (at 1 x 10⁻⁶ mcg). 5 One CFU / PCLS was applied directly to the tissue. One hour post-infection (pi), the inoculum was removed, the PCLS were washed, and the transwell was transferred to the wells of a new mating plate containing 500 μl of fresh culture medium in the lower compartment. Prepare a solution containing 1 x 10⁻⁶ CFU / PCLS. 10Phage mixtures of each phage (diluent: 0.9% NaCl) at PFU / mL were used. 1 mL of phage mixture or medium (0.9% NaCl) was nebulized onto PCLS using an Aeroneb device and an exposure system with custom adapters for 4 transwells (nebulization time: 1 min, settling time: 60 min). Separate exposure systems and adapters were used to nebulize the phages or medium to remove contamination. The PCLS were incubated overnight at 37°C and 5% CO2, and samples were harvested and analyzed at 24 h p. 1. Furthermore, in this first experiment, separate samples were harvested directly after phage nebulization to determine the actual PFU deposited in each well.

[0204] After confirming the efficacy of phage aerosolization in the Klebsiella pneumoniae PCLS model, the experiment was expanded to test the efficacy of phage mixtures alone and in combination with antibiotic co-treatment. Rat PCLS were prepared, cultured, and infected, and phage was treated via aerosolization as described above. For co-treatment with antibiotics, meropenem, vancomycin, and amikacin were added to the culture medium of each PCLS at final concentrations of 8, 8, or 1 μg / ml, respectively. Antibiotic treatment and phage treatment were performed simultaneously at pi 1 hour. Therefore, the washed PCLS transwells were transferred to new suited plates, the outer compartment of which contained culture medium or a culture medium containing an antibiotic mixture, and the top was exposed to the phage or saline aerosol as described above.

[0205] Analyze the CFU load, tissue viability, and phage replication of PCLS.

[0206] To analyze bacterial load and phage titer, PCLS were lysed with 1% Triton X-100 / PBS at 4°C for 30 min. Tissue residue was removed, and the lysate was centrifuged at 8000 x g for 10 min at 4°C to precipitate bacteria. The clarified lysate was used to determine the phage titer. The precipitate was resuspended in 250 μl PBS / Tween and used to determine the bacterial load.

[0207] Phage titers were determined using a plaque assay, specifically detecting phages F391 / 08, Kle_F17 / 19, and Kle_F58 / 19 using Klebsiella pneumoniae strains 131 / 15, 130 / 14, and 25 / 14. Each sample treated with a phage mixture was co-plated with each detection strain. Phages were released from PCLS by tissue lysis and isolated from bacteria by centrifugation (as described above). Phage samples were prepared in Eppendorf tubes with sequential 1:10 dilutions of PBS / Tween and mixed by inversion to avoid mechanical interference. 100 μl of each phage dilution was mixed with 100 μl of each phage dilution.

[0208] The test strain cultures were added together to 3 ml of soft agar at 48°C, and the mixture was spread onto a TSA plate. At the end of each PFU experiment, all test strains were plated to exclude phage contamination from the bacterial cultures. After incubating the agar plates overnight at 37°C, phage plaques were counted. To calculate the PFU for each PCLS, the plaque count was multiplied by its respective dilution factor, and then multiplied by a factor of 2.5, reflecting the ratio of the total phage sample volume (250 μl) to the plate volume (100 μl).

[0209] To quantify the bacterial load in PCLS lysates, samples were prepared in sequential 1:10 dilutions with PBS / Tween, and 50 μl of each dilution was spread onto half of a TSA plate. After incubation overnight at 37°C, colonies were counted. To calculate the CFU for each PCLS, the colony count was multiplied by its respective dilution factor, and then multiplied by a factor of 5, reflecting the ratio of the total bacterial suspension volume (250 μl) to the plate volume (50 μl).

[0210] Viability of uninfected and infected tissues was assessed by calcein staining. Calcein AM infiltrates intact cells and hydrolyzes intracellularly to generate calcein, a fluorescent compound retained in the cytoplasm. Increased number of dead or damaged cells within PCLS leads to decreased calcein staining intensity. PCLS incubated in 70% ethanol at room temperature for 15 min were used as a reference (“death control”). All PCLS were washed once with warm medium and incubated in the dark with calcein AM staining solution (4 μM, dissolved in DMEM / F12) at 37°C and 150 rpm for 45 min. After washing three times with PBS, PCLS were lysed in the dark at 4°C with 1% Triton-X100 for 30 min. Subsequently, 50 μl of tissue lysate was transferred to black-walled 96-well plates, and fluorescence was measured using a microplate reader (excitation wavelength: 485 nm, emission wavelength: 535 nm). All samples were measured in duplicate.

[0211] 6.3.11 Example 8: Nebulization Study of PCLS Klebsiella pneumoniae Infection Model

[0212] The biological activity of the phages after atomization was determined prior to the first atomization test.

[0213] 1x10⁻⁶ was prepared in 0.9% NaCl. 6 A phage mixture of 1 PFU / ml was used. After nebulization, the recovered phage titer indicated high loss during deposition; therefore, 1×10⁻⁶ PFU / ml was used. 9New mixtures were prepared at 1 PFU / ml for each phage. Other nebulization tests were also performed to assess the expected phage titers after nebulization. Detailed results are shown in Table 6.

[0214] Table 6. Phage recovery after atomization using an Aeroneb nebulizer.

[0215]

[0216] In summary, when the initial phage mixture was 1x10⁻⁶, 9 At a concentration of 1 PFU / ml, the residual activity after nebulization is expected to be approximately 1.5 to 2.5 log-order lower (approximately 2 log-order lower for phage 391 / 08, approximately 1.5 log-order lower for phage Kle_F17 / 19, and approximately 2.5 log-order lower for phage Kle_F58 / 19). To accommodate this reduction, a concentration of 1 x 10⁻⁶ PFU / ml for each phage was determined. 10 Potency tests were conducted at higher phage mixture titers of 1 PFU / ml.

[0217] Based on these results, it can be concluded that although the activity of the phage titer is partially lost after nebulization, the confirmed phage activity after nebulization should be sufficient to demonstrate efficacy in the PCLS infection model.

[0218] For treatment with atomized phage, rat PCLS were cultured at the gas-liquid interface in a transwell, and the bacterial inoculum and atomized phage (or NaCl treated as a medium) were directly deposited onto the tissue. Nebulization was performed as described above.

[0219] The atomizer contains 1x10 10 Immediately after nebulization (pi 1h), a phage mixture of each phage at 1 PFU / ml resulted in a deposition titer of approximately 1 x 10⁻¹⁰ PFU / ml for phages 391 / 08 and Kle_F17 / 19 per well (per PCLS). 7 One PFU, while the deposition titer of phage Kle_F58 / 19 is approximately 1 x 10^6 PFU. 4 PFU / PCLS. Phages 391 / 08 and Kle_F58 / 19 replicated strongly in PCLS infected with Klebsiella pneumoniae 57 / 17, reaching approximately 3 x 10^6 PFU / PCLS. 8 The titers were 1 PFU and 2 x 10⁷ PFU, while the titer of phage Kle_F17 / 19 was 2 x 10⁷ PFU. 6 The number of PFUs even decreased slightly. Figure 10 This was expected, as phage Kle_F17 / 19 cannot infect Klebsiella pneumoniae 57 / 17. Nevertheless, it was decided to test a mixture of three phages on this strain to gather results on any toxic effects that might result from treatment with this phage.

[0220] Use 1×10 5 PCLS were infected with an inoculum of 1 CFU of Klebsiella pneumoniae for 1 hour, then the inoculum was removed, washed, and transferred to a new well to avoid overgrowth of non-tissue-attached airborne bacteria. Using this procedure, the actual bacterial load immediately following exposure to the medium (NaCl) was approximately 10-1. 4 One CFU / PCLS. If exposed to the nebulized phage mixture for 1 hour, the CFU load had already decreased to 10. 2 One CFU / PCLS indicates that the bacteria were rapidly lysed by bacteriophages. Figure 8 In PCLS with medium treatment, strong bacterial growth at pi24h produced approximately 4 × 10⁻⁶ bacteria. 8 The bacterial load was [number] CFU. Treatment with the nebulized phage mixture resulted in a reduction of approximately two logarithmic orders of magnitude in bacterial load at 24 hours post-pi. Therefore, nebulized phage treatment successfully reduced the bacterial load in PCLS infected with Klebsiella pneumoniae. Figure 11 ).

[0221] To achieve proof-of-concept for phage efficacy following inhalation alone or in combination with antibiotics, rat PCLS cultured at the air-liquid interface were infected with Klebsiella pneumoniae strain 57 / 17, and a phage mixture was nebulized directly onto the air-liquid interface cultured tissues. For co-treatment, antibiotics (at subinhibitory concentrations) were applied during phage treatment via immersion (i.e., into the culture medium in the outer compartment of the transwell substrate). One plate with four replicate wells was used for each condition. Two wells were used to determine CFU / PFU, and the other two wells were used to determine tissue viability (calcein staining). Nebulization of the phage mixture resulted in approximately 10 depositions of phages 391 / 08 and Kle_F17 / 19 immediately after nebulization (1 h pi). 7 One PFU, while the deposition for phage Kle_F58 / 19 is approximately 10. 5 PFUs ( Figure 12 ).

[0222] Subsequently, phages 391 / 08 and Kle_F58 / 19 replicated in PCLS infected with Klebsiella pneumoniae, resulting in a 2-3 logarithmic increase in phage titers at 24h pi, while the titer of phage Kle_F17 / 19 remained stable. Samples co-treated with antibiotics showed similar phage titers, which was expected given that bacterial load was observed to be unaffected.

[0223] Nebulized phage treatment successfully reduced the CFU load of infected PCLS by approximately 1.5 log units. Figure 13Treatment with sub-inhibitory doses of antibiotics (immersion) did not reduce CFU. Notably, combined treatment with a mixture of antibiotics and nebulized phages showed a strong synergistic effect, as it reduced CFU load by approximately four logarithmic orders. Figure 13 ).

[0224] Infection with Klebsiella pneumoniae resulted in a significant loss of tissue viability in PCLS at 24 hours post-infection. Figure 14 Treatment with a nebulized phage mixture significantly inhibited this infection-induced loss of tissue viability. Figure 14 Treatment with subinhibitory doses of antibiotic had no effect on tissue viability, as expected based on unchanged CFU. In phage / antibiotic co-treatment, the synergistic effect on bacterial load was not reflected in the further increase in tissue viability, since phage treatment alone already significantly improved tissue viability.

[0225] Precisely sectioned lung slices (PCLS) are fresh lung tissue sections that serve as organoid models, encompassing all cell types present in the lung and allowing for detailed study of lung tissue responses. Their high complexity provides a physiologically relevant model. Data from PCLS studies demonstrate the efficacy of nebulized phage therapy on lung tissue slices infected with Klebsiella pneumoniae. Administration of the phage mixture reduced bacterial load and restored tissue viability. An additional synergistic effect of reduced CFU against the tested antibiotic-resistant Klebsiella pneumoniae strains was observed in co-treatment with sub-inhibitory doses of antibiotics.

[0226] After reading the foregoing description, certain modifications and improvements will arise in the mind of those skilled in the art. It should be understood that all such modifications and improvements have been removed herein for the sake of brevity and readability, but they are all properly within the scope of the following claims. sequence list <110> TECHNOPHAGE, INVESTIGAÇÃO E DESENVOLVIMENTO EM BIOTECNOLOGIA, SA <120> Cocktail compositions containing respiratory antibacterial bacteriophages and their methods of use <130> US 62 / 950,850 <160> 3 <170> PatentIn version 3.5 <210> 1 <211> 113073 <212> DNA <213> Klebsiella pneumoniae phage Kle_F391 / 08 <400> 1 attachment ctaaacgaa tcatacactc cactaatta aggttgacaa catgaaacgc tttgtaatat catctaacc ggcggcaaaa ttgaacaaaa tggcaagctc ggaactggtc aaaatgcagc gcgatatggc gcggcgttac cgctcttatc agcgcggagg tcagcaaggc 180 ttaccgatga tttgggataa catgatgcgt tatctaaaat ctttacaaaa taacgtttga caatctgccg father attachment ctcaatagc aacaccgctc tttaaaaatt tggaaaagtc gattctctca gtaagggtaa ttgagttttc tgttaacact ctagcagtaa aagagtgtta accggataac ccaataaact aaactggagt aagaaacatg caaaacgtta aaaccccaat gatcggtgat tccgttttta ttccttttcgt tactggcgac gtaagtaagc 480 cgggcgaaaa tgaaaaaatc ggatatatca aaggggcggc gatgatcccg tttgataaaa ttaatgcggt atacgccgaa acggaaaaag caaaaaacag cgacgcgaag atttacagcg 660. tccgggtaga ttccggcgac gtggtgaaag tcattcgcaa agatgataaa tggttggctg tcgcttaaca gccaataagt ttaaccccgg caataatgg gatgcttccc ctatgttaga cttccgatgt atattccatg cagcgcaagc aatgttatgg tgcgcaaatt aaacttatgt 780 gctggaactt atatcatgct ttgaacggca attgtgagtt aacacgaatc tatagcgtga 840 atagacgta atagcgtaat tattaaaggc agcgtatata tacaatgccg ctttaaatag 900 ctttaaattg aaaactaaaa taaagagttt tcgttatata ttctagctag aatatagc 960 ggctaacttt ctagccttaa cccttaaata aaactggaga tatatcatga ctgctactaa 1020 aaccgaaaaa ttcgcatgga acgaaacaaa cgccgctacc gccgttgaaa tgtacgaaaa 1080 gatcattgct tccgatggtc tggaggtggc taactcgcaa ggtcttattg atattgcaaa 1140 ggctgtaggt gctgaatcgc atgttaaagt gcgctctaaa ctggttagcg caaaggtgta 1200 tcagaagtcc gataagccgc gcaaagtcgg cggcggctcc tccctgcgta aagctcacta 1260 tgtgcgcgtt ctcactcagc acgccattgc tgacggcctg attgatgatg ccgatggtct 1320 ggcaagtctg gaacaaatga agctcgacca actggacgtt atcgcccgta tggttggcgt 1380 tcaggatgaa gtaaaagaat cagcagaata attataatgc gcctccctag cggggcgcat 1440 attaaatat ttgtccccc ataactggag aaataaaaat ggctttaatt aaaggctccg 1500 ttattaaatt aacgggaacc gttgtggatg aattaatcca aacggggtat caggataata 1560 aggttatgac gcccccaagc gttaaagtac ctgaatatat tgttttgtgg gttaaccctg 1620 atgcggatac ttcgggatg gctattaacc gcgaagtctt taagccggaa atgctggaat 1680 tatcttcccg cgaaatttac cttctcaatt acgctttcag cgtagaagaa aaggaggttg 1740 taaaatgatc ttttcccta ctgaatcctt aattcttatc gctggctttt tcgccgccgc 1800 ttgtctctat ggttactata atttcatgga aataggcagc gaacaaacgg atttattacg 1860 ccgtgacttc tggttcaaga aagcgagtat ttgccgccgc tggtcaataa tctttattat 1920 tctagcggta actttcggaa tatcagcaag cattatcccg gcaatagttt agagtcgcaa 1980 attataacgc caccagcaac aaggcgttat atttggcaat tctgcctata tcccattaac 2040 tggagaattt agataatgat cattgcaatt gagaaacaaa aagctatttt aaccgctgct 2100 aacaacctca atttttgggg taaacgcctg cgcgctaaaa agctggagat ctgcgacgaa 2160 ttgagcaaag agcattacgg gacggcgaa cactctagcg aaatttgcga ttggctggat agtaacaagc ctgttaacc cgctgctgaa aaacgcgccc aacgtgttgc ggtggaagat tcccgccccg ttgccgctgg tcagcttaat tccagcgtcg aaagctggaa ggtaattccg ggccgccgtt ttctgctaac gtcgattcaa fathercct tccctcatgc taatttctgg aaaactttac aggaggccgc gaaatatctc ggcgctactc tgttagtaag caaatacgcc 2520. ggcaacgatg aactgaata tgatgatgct ttctcagatt tcatttgtga tgaaaacgtg tttttgggca accgcgaaac cggattcgca ttcatggcgg aaattaat tctgccaact gctgattttc cgctgtccgg ctttggcgag actgcgaccg cctacggcct gaaagggctg gctattggtc acgctaaaat caccgccgaa agtgtcccgg caatgaaggg cgacactgtg cgccgtatgt attcaactgg cacagcgacg ctgaaaatt atattcagca aaaagcgggc caaaaggccg aagcgctgca taactatggc gctttgctgg tagagatcga cgacaacggc aacttctttg ctcgccaaat cgaaacgatg gacgaaagcg ggatgtttta cgatcttaat cataaattta ctgttaacgg tggcgaggag 2940 gtaacgggcc acgttgccgc gctgcaatat ggtgacattc acgccgaaaa gctggatcac 3000 gctgtcgcct ttgcttcatg ggggccgtgt gatgattctc tagtgaacgt cctgcgccct 3060 cgttaccaga ttgtgcatga tgtgcatgac tttacatcac gaaaccatca taaccgcgct 3120 agtggcgttt tccttgcgaa acaatatgcc gccggacgtg acaaggtaat tgacgatctt 3180 atcgataccg ggcgcgtact agaagcaatg gaacgtgaat tcagtcaaac ggtcattgtt 3240 gaatcgaacc acgatcttgc gctttctcgc tggctggatg atagtaaagc taacattcaa 3300 caagacccgg cgaacgccca tctttattat cgcctgaatg ctgcgattta tgaagcaatc 3360 gaaaacaagg acgatacttt taacgtacta gattacgccc tgcgcaatgt tgctggctgc 3420 gattttgctg cgatcttctt aaccacggat gaatctatga aaattgcggg catcgaatgc 3480 ggttcccacg gtcacaacgg cattaacggc gctcgcggca acccgaaggg attccgcaag 3540 ctcggcaaga tgaacactgg ccatacgcat acgcccagca tttacggcgg cgtttatacc 3600 gctggcgtcg ctggttccct cgatatgggt tacaacatcg gcgcgtctag ctggtcacaa 3660 acgcatttga tcacctatga aaacgggcaa cgtaccttga ttgactttaa agacggcgtt 3720 ttctttgcat aacaaaaaag ccggggtaat tgccccggca ataaataacc cataaataaa 3780 taataagct ggagaaacaa atgaaactta cttcatcta taacaatcgt aaatctttca 3840 ccgcgtccaa cgtcgtagaa atagccttg ttatttcccg cgatagtgaa gggcgcccgc 3900 atgttagcta tcagaaagtt aacacggtgg acggcgacac tgttttaaaa gctctagccg 3960 ctatcctccc gcgccccgct gaatttaaag aatccggcat tgtgtcgcaa tgtcgccg 4020 ccgattctat tctttatgag gctgatatct gcgagatcgt agagattgac gccgccgccg 4080 ctggccttat gtttatcgtc gtaagcgaaa atgattatga tgatacctat ctgctcggcg 4140 acgtgatgga ttattcttct agcgaatata cgccgccgct ggcaattgtc ccggtaatgg 4200 ctacccggat taaaccggct gaattagccg atgccttaac tttattcttc tgataattaa 4260 caaagcgagt tataatatag ctcgcttgat taattaccaa caaccccgaa accggagaaa 4320 tgaaaaatgg cttttaataa actggcaatt aaggcaatta agttatggga tttagacgga 4380 actgttatta attcctttgc ccgcgtgttc ccgtgtatgg atgataaagg gaacttagat 4440 ttaaacatgt accgggaaaa ggcttgcgta catgatgcaa ttatgaccga cactctttta 4500 ccgctggttg aatatatgcg ggcatcgctt aacgatccca ctgtattaaa cattattgtc 4560 accgctcgtt atatgggtaa gagtgactac tatttcctac gcaaacaacg tatccgggcg 4620 gggcgcggtg gtaatatcca gatactgtcc cgcgatgtat tgcaccgata tattggcgat 4680 gctgattata aagaggtgta ctattcgaaa gatggtatct ataaaacgca ttattcgaa 4740 atgcttaaag ctgaatatcc gaacgctact atcactatga ttgacgataa tagaggcgtg 4800 ttggctgctg ctgctgctgc tggccttcaa acgatggacg ctaccgcaat taacgatatt 4860 ctatctattg gtgtgcgcct tgcgggtgag tcattcatag atgaagcgct ggatgatgat 4920 aatgattatc aatatctttg cgagcgttta gctcattgct gggaaggtat gaccgaagaa 4980 gaagaagcg actacggaat taagccgcaa caatttatc aatcgttagc catcgcatca 5040 taagaaagct aatagtaccc tagcgaaata gttagggtac taatagttag ctaggggggt 5100 aatcggacta ccataccacc ccccgaccct ctaagcccac ctccatgtgt aactttatga 5160 aatttggaga aaaggctaaa gggagtactt aggcaaactt agaacgagca gcccgaatca 5220 aagctactac tgctactgcc gctgtcgcaa taagaacttc ccgagtcata cccgctatac 5280 gttgcaactg ccgcagttgt agctatgatt gctgcagtcg acccatcatc gctccgagaa 5340 cttcccgagt tcgatgaagg acttcccgaa actttccgag aattatccga gcctgcccgt 5400 ttttgcttgc gtttcgcgac ctgtggtgtc actggaggga tgtagtacgg gttgctcggg 5460 tcattccgaa gatcgttcat gactcgttgg tgttttgaga tcgctacttg acgttctgcc 5520 cgagtttgaa ggatctcttg atgcagactt ttgttacgaa gatctgccgc tactaccgcg 5580 ttccgcaggt catgtgcccg cacaccaaac caaactgctg ccaggaaaaa gaatgttgtg 5640 aaaattaata caatagtcat gttttacacc atacccttag taatgtagat tttgtttgct 5700 gattctagtt ctgccttgca ttcagacagg tttaactgtg ccttatgcag caaacgggtt 5760 tgattgtcat aagacttctg cagcaccccg ttagtgcgtt ctagctgttt gattgcctta 5820 ccctgcttta cggtaatgac gaatagtacg attgagagta aaacaagtgc tcctataact 5880 acttccatag tggtcctcct agatggtttt aaaaaatctg gtttacttgg cggttgattt 5940 aatttataat aatattatat tataaaagtc acttgttagc aagtgagatt ttgaggtttt 6000 tatgggcttt tatgcaggaa ggataggcga caaaaaagtg ttgtctctta cctctggcaa 6060 caataaagac gttaataacc acactaatcc aggttgggat actatattcc atagcgatat 6120 gcctcacgtt gtagttttag aaactcacga aagagacctc tgggatggtg gtgattggta 6180 tcgttgtact agaatgccag acagaattat tcaggtactg tccgcagact acgacagggt 6240 tgtcttaact gaagttgagt ttgaggatgg caccagacgt ttcatttatg gtacatccct 6300 gggtgtgggc gctaaagcgt ataacgctta ctttagtaat actgtcggtt cccaggcttc 6360 tgcaggtact atggctagta tgaagactaa cgtttgtgct tctgcagact tacacatgga 6420 tattagtttc tatttcgaag agacgccagg tactattaac gagaagctac gagacggtac 6480 tgggtgtatg tacacctggg gcgttaactc agaatggga gacagagggc ctgggccgcc 6540 ggttggagct cctatacgtc ctaactttga gactattatt aaagctggat gggtgctcta 6600 tagggggct tttagtggca atatagccgg ttctgtgtct ccgcccaata gaccccttac 6660 tattggtgtt gatgctatgc gccacccgtg gatgcgtact actggtgtta acagtatatg 6720 tttgcgtggt gagactctca accgtaatat gtacggacat atggggccta gatatgggca 6780 gagctccaat cccgttggtg gtccttatgc tcataacatc cagactgagt cttatcagga 6840 ggttcagtac aaagcaggtt ttttccgtgg tccgcccaat aactttatgg ggtgggaaaa 6900 cacagataac aacaatgctg gtagtggttg gggtaacaat gccatttacc gtgacaataa 6960 cttccgtgtt cctaaacgtg ttcgctggta tattactaat atgaaataca atgggcaggg 7020 gttctacgca gagaacgtat ttggctcccg caaccaggag attaaaatca gccctagaga 7080 gttcattgta aacggtataa acctaatgaa cactgggtgg aagttcataa accagaacga 7140 tataaactac agcccaggta ataggcctga tatccgagtt attgcaacca acgtcgccag 7200 atttagtggc aaccctactg ttggtaataa tggatatgtt cactttaatc agcctctaac 7260 tcgaccagat aatggtgctg agtttggaca aggtaacgtg agtgagatgc atgtaaccac 7320 tgtaggggtt tacaatttta gatctgatgc acagtggtac gtaaaatcta acccgccgga 7380 aatcggaaac cagtggggtc cagtatggtc tgaatcaact cgacctctcc gcctagtggg 7440 cggtaccggc tctgctgata ttggagggaa cctacgtact agcggtaatg ctagtcacca 7500 cctggctacc ttgtggttag gggttaataa ctcccgaaat ggggcttgcg tcgtcactct 7560 agactggaaa aatgatgagt ggattgctgc tgcagggatt ggatgttata accctctaga 7620 agatcttacc cagtggagcg aggtggatag taggctgaga attttcggaa atcacttcca 7680 gaaacgtgtg catcaaatca tgtgtttgcc cgttaacatg tgtgtgccgt tccactttat 7740 acgcgggacc gtaacccagt gtggggttat tcctgggaac aacgccatgc aaatgaaggc 7800 tatgtgggca cctactacta ccaactctgc cactcagggc gattatgcca ttatctattg 7860 gctgatagct agggctgacg gtagtgttga agtttgggtt aacgttgaga tgagcaatat 7920 catgaatatg cgagtaattc ttcctgaggt taggattgct gtgcaaaggc ttgcctaaag 7980 gaggcaatat gagcaatgat ttaattgtac cagatacaat gtctccggaa ggcatgctag 8040 ttagaggc ctacctggag tctggcagcg acgttgcgaa agcagcgtta gctgttggca 8100 tggaggaacc taaattccgg gagattatgc gtaaacctga ggtcaaagcg taccttacgg 8160 atacttcat ggaatctggc ttccgtaacc gtgataaatt cttcggcatt ctagacactg 8220 ttctaactat gaaaatggag gaactggatg aaactggaat gggttcagag atggatatta 8280 tggatatcct caaactcatg cacaaaatga agatggatga gatgaagatg cagatcgagt 8340 atgagaaggt gaagcaagct aaagctccaa tacaccaaaa tatactcag atcaacctgg 8400 cagggggtca cgactctaat tacacggacc ttctgtcacg cattgtggga gcaggtaagt 8460 aatggaaatc tcacgtagtt atattaatac gactgacgtg gtggattttg gtgttgataa 8520 acgattcttt aaattcccgg tgtccggctt gctggccacc gaggggatcg ttccaaatgg 8580 cccgcagtgc gcaatcataa acgcactgga agacccacgt caccgtttcg ttacagcatg 8640 cgtatcacgt cgtgttggca agtcgttcat tgcgtatact ctaggcttcc ttaaactcct 8700 agagccgaac gttaaggtgc tcgtagtagc tccgaactac tccctggcaa acatcgggtg 8760 ggcgcagatc aaaggtctga ttaagaaata cgggcttcaa actgagcgtg agaacgccaa 8820 agataaagag attgaacttg ctaacggttc gctctttaag ctggcttg ctgcgcaggc 8880 tgactccgca gttggtcgtt catacgactt tattatcttt gacgaagcag cgatctccga 8940 cgttggtggg gacgctttg atattcagct ccgtcctaca ctagacaaac ctaactctaa 9000 agctctgttt atctctactc ctcgtggcgg taactggttc aagcgcttct atgaacaggg 9060 attcagagag gatcttccgc aatgggtatc aatccacggt acataccgag ataaccctcg 9120 cgtatctctt gcagatatcg aggaagcacg taagactgta tctaagaact acttcaaaca 9180 ggagtatgag gcggacttct ccgtattcga aggtcagatt tacgacacct tcagcgtttc 9240 cgagcacgta caggatcttg caggtatggg gcacttcttt gctgcggacc atgagttcga 9300 aaccattcta ggtatcgacg ttggttacag agacccgaca gcagtactta ccattaagta 9360 ccactatgac siagatgtgt actacatcct tgaggagtac cagcaggcag agagactac 9420 agcacagcat gccatgtaca tacaactg catagaccgg tataacgtag accgtactt 9480 cgttgactcc gcggctgcac agttccgaca ggacttagca tacgagcatg aaatatccctc 9540 tgctcctgcc aaaaaatctg ttctagatgg cctggcatgc ctagctgctc tattccagca 9600 gggtaagatc attgtcgacg cctcgtgcac tgcgttaatt cacgcactcc agaactacaa 9660 gtgggacttc caaagggg aggaaagct ctccagagag aaacctcgtc acgatgccaa 9720 ctctcactta tgtgacgcac tgcgttacgg catctactcg atttcccgtg gcaaataaga 9780 gctgagttgg aacggatagt agtgagtaca aattccgttc caacttata aattcactt 9840 tacaatttcg tgtgtggcag ttatata ttcatacctt gagagatctc atagacatca 9900 gggtgaatga gaggaatatt taattgctat acccgaagta gtattttccc aactaagagg 9960 aggaacactt tggctccaa tgtaaaatat aagagagatg caatctccat aatgcgtgac 10020 ggaatcaaag cccagtacaa aagaggcaac tgttgcgcca ttgtgactc acaggaaac 10080 ctggagctac accactactc gactgtggcc ttactagtta aaaactttgc taaagaattc 10140 caactggatt tcactgactc agaagtcgtc ctaagtaatc gggataagtt ctataaacat 10200 tattggcatg agctagtaga ggacacggtc accttatgtg tctttcatca tcagacctta 10260 cacaaggtct atacgaaaga acctccgttg ttttcagcta acaaacagaa gatttgggtt 10320 gagaaacaac gtgaaagatg tatgaatcca gaggcacctc gtacaagcaa cactggcgaa 10380 agatcaggct ttgcgaagtg gcttccgact gacgtcaaga ctgagaaatc aggattcgca 10440 aggttcctat aatggctatt cgtgactggt tagttactaa actaaaccgc ggacaacgca 10500 taatcaggga cttggaggat gttagtcacc gtactaacgt caagccattc acgactggca 10560 aagcctattc gtctattgag atccttaata gatccgcgaa catggtaatt gacagcgccg 10620 ctgagtgctc ctacaccgta ggtgaacagt ataaaacaat aacaacctat ggcacgatca 10680 ggagtaaaac tcttgagacg ctgcttaatg ttcgccctaa cccgtacatg gattccagta 10740 cttttagacg cctaatagtg tctgaccttc tattcgaagg gtgcgcgtat atccattggg 10800 acggttcgtc tctgtaccat ctgcctgctg ccctaatgga agtaaaagca gatgacaaaa 10860 aattcgttaa caaattcgtc tttaataata tgatcgacta tcgcgttgat gaaattatct 10920 tcatcaaaga taatggccag aatggtggta ttaactccca gattacgggt caatctcgtg 10980 tggctaccgt aattaactcg cttactaagc gtgagaaaat gcttgagttc aaggagaaat 11040 tcctggacaa tggtacggtt atcggtctta ttctagaaac agatgaaatc ctaaataaaa 11100 agctccgtga acgtaaacaa gaagagcttc agctggacta taaccctagt accggccaat 11160 caactgtgct cattctagac ggcgggatga aggctaagcc atactcccaa atctcctcct 11220 ttaaagatct cgatttcgag aacgatatcg ctcgttttaa taagacgta tgtatcgctc 11280 tcggagtccc acaactattg atcgatgggg gtaacaacgc caatattcga ccgaacattg 11340 agctgttcta ctacatgact attgtgccta tgctcaataa agtatgtagc tctcttacgt 11400 tcttctcgg ttttaaagta acgcccaata ctaaagacgt agtagcacta actcctgaca 11460 aggagaaaga gctaaatttc gtaactgcac tggtcaacaa cggtatcctt accggtaacg 11520 aaggtcgtat agagcttggt tacgaggagc tggctgacga gcagatgaag aaaattcgca 11580 tccctgccaa cgtagctgga tcagctaccg gagtaagtgg acagaaggt ggcgctccca 11640 aaagacga ggaaaaaaaaaaaaaaaaaaaaaaaagcatta tccctaatgg 11700 tctccccgag gcacataacg tgtttgcac cgttaaggca catctcactt accagattct 11760 gcgtaggag tatgggtacg ctgctactaa cagtaaaacc tgggatcagt tttaggaagc 11820 ctacgcggaa gcaactaagc cagtaccggt agctctctgtt agtatcacgg gcgctcctgc 11880 atccttagac tatactaga ctgtacagct tgccgcaact gtcctaccaa caacgcaga 11940 tataaaact gtaacgtgga agactga tgtacccta gctactgtga gctcaacagg 12000 attagtaaca gccctgtcta agcaggcac tgttaaaatt actgccactg caggtggtaa 12060 atctagtgaa gtgtctattc aggtcaggc tcctgttgta gcagttaccg gtgtcactat 12120 gtcacctaag actattacaa tcgaagcagg tagaccggc aaacttactg gtaccgtagc 12180 cccggctaac gcaaccaata agtctgtaac ttacacttct gctgatacca ccaaagctac 12240 cgtagctgcg gacggtacgg taactgttcc tgctaacctg gctgcggata gtaccgtagt 12300 tattactgtt aaaacagctg acggcaataa gaccgacact gcaatagtaa cagttaaggt 12360 tcctacagcg ggtgtgtaat atctgttaac tggggagctt cggctcccca tttttctatg 12420 ctagtgcata actattttca gtgtaaccaa tttcgactac ctactaattc cacacaccgt 12480 gtgggattt caaatatctt ttgacattcg cgatttggta tgtaataata gtatctgaaa 12540 actgaggaac aacacgaaag tgtttggagc aaacgatgca gaatattaat cttaatgctg 12600 agattaaatc tgttaaagct gtcggtgagg gtgataatcc tcctttaaaa atcaaagggt 12660 atgctaacac gattaccaaa gaccgcgccg gcgacgttat tctctccgaa gcgtggacta 12720 ccagtaacgc cctcaagaat tttatgaaga acccgatcat gcttttcggc cataaccata 12780 gccgcccaat tggtaagatc ctagacctgg taccaaccga gtccggactt atggttgagg 12840 gtgaggtaag tgctgctgat ctacagattt actcattaat acgtgatgag gttcttaaaa 12900 cgttctcggt aggcttctat attaaagacg ccgaatggga cgatatgact gaaacgttca 12960 ttattaaaga tctggagtta ctcgagatct ctgtagtctc ggttccctgc aaccaagact 13020 caacttttag cctttcgaag tctgtaaacc ataatgatta catggagctt cgtaagtctt 13080 ttgtaaaatc ttcgcaggtc cagcctgttg aacaacctga actttctaat ttagagaaat 13140 tcctagtagc cgctggttac gctaaaggat aatggagaaa tataatgtc ttacgatatc 13200 gcacaactgt ctaaagatct gggcctgggt gacattgctg agcagcttaa aggtctaacc 13260 gcctctcaga aagctgaaga agctcgcaaa tttgctgctg agcaggaagc taaagaactc 13320 aagcgtatgg aagacctggt tgctaaagca actggtgaag accgtaaaa cctggcagaa 13380 gctctggagc ttgtaaaaaa cctggatgag aaatccaaac agtccgctga agcgttcgtt 13440 aaagcaatga actcccagca ggaagaaatc actggtctga aagaagaaat caaatctctg 13500 ctggccgctc gtgaaaatgg tcgctctttc gtagctgatg gcgttgctaa ggcaatgttt 13560 ggtaagcagg aagatttcga agacgaagtt gagaaactgg ttcttctgtc ctacgtaatg 13620 cagaaagatg tattcggtac taagcgtggc gaagcccacc tgaaagctgt taacggctct 13680 tcttctatcg aagtttctac cgaagcatac gaaaccatct tctccctgcg tatcctgcgt 13740 gacattcagg ctaaactgat tatcggtacc atgttcgaag aactgccgat gtctagcaaa 13800 ttgctgacca tgatggttga gccggaagct ggtgaagcta gctgggttga cgcctctact 13860 tacggtactc ctgctactgt tggtgcggaa gacaaaacca agctgtctga aatcaccttc 13920 aagacctaca aactggctgc taaagcgtat atgaccgacg aaacagaaga agatgctatc 13980 ttcactctgc tgccgatcat gcgtcgtcgt ctgattgaag ctcacgctat cgcaatcgaa 14040 aaggcgttcc tgaccggtac tggtgctgcg ggtactccga aaggcctgat ccagttcgct 14100 aaagacgatg gtaaagtggt tgctaccact gctaaagctg acggttctgt taaagttacc 14160 gctaaagaaa ttcacaagct gcgtcgttcg ctgggccgcc acggtctaga cctgaacaaa 14220 ctggctctgg ttgtttcgat ggatgcttac tacgatctga tcgaagacga agaattccag 14280 gacgttgcac aggttaccgc taccaccgct atcaaactgc agggtcaggt tggtcgtatc 14340 tacggtctgc cggtactggt ttctgagttc ttcccagcta aggctgcaag cgctgagttc 14400 tgtgtagttg tttaccgtga caacttcatc gttcctcgtc agcgtgcaat cactgttgag 14460 aaagagcgtc aggctgaacg tcagcgcgat gcgtactacg ttactcagcg tctgaacctg 14520 atgcgtttct tcgagaacgg cgtagttgct ggtgcttacg ctgcttaatt tggctatact 14580 gccgactaaa ggagagcttc ggctctcctt ttttattggg taaaatatgc aattcatgac 14640 agattctgat tggagaacat atggaggcct taaacgtcct gacttagagt caaatatccc 14700 aatgttaatc aaagcagcca atgctctgat tactcagctt ctaggtattg acgacactgc 14760 taacgttgta gacgttctgc ctactaaacc agcacgcaaa aagtacttcc tgtcttctcc 14820 cgtgcctagc acgatcacta aaattacgat taacgatcag gagatcgata agtcgcaata 14880 taagaactac ccggatggta cgctcttgct gaaattctcc cctccagagg ggtatatgga 14940 ownttt actcagaccg gcttcacctc gatccctgac gacctggtac tagcagcctg cttcctagtt gatcactggg tcaagaagga ctaccgcgag tctcgtacat tcggcggaga 15060 aaccgttact ttcaatacca ctaatctgg cgtaccgga cacattcgta ctataatcga agtataccgg aggctgtagt ggcgttgggc gatctagctc gacagatagt taaagaacag ctggacatta tgagtggtgg tagccactct accaagaaca ccgtgatata taggcgga actatggata accacaaaga tggcaccata ggcaaggtat ccttccgatt taccaagccg gtatcggagg atttactgaa tgttcggacg tcctctattt taaaagctgt ctcgtcgtca cttaatctgg aaggtgacgt tggtgttatc gataaccttc taatagtat cactggtaaa 15480. aatccaaaa tagged acggtctacc ggtagggtag aggttaactt tggagatcct tcagatgctg acaacggtta cgctggtgct atttctggcg cttctgggcg tttcgtctca aacacaaacc ttagagcgtt acttgaactc gtagcgaaag aatacttagt caaggatatg aaaaaagctg gggcacccct taaatttaga acggggcgtt tcgctaattc cttgaagatt 15660 aaagacgtgc tgttaagaga agacgcaggg gcaaagactc ctgacctcaa catcacgtat 15720 aactacatgc ttaagcctta ctctgtgttc aaccctgccg tctctaccta ccgcggactt 15780 tccttacggc ctttccctgg tgctaggaac ccgcagaagc tgatcggcga ggcaatcgct 15840 aaggctgcaa gagaccttat ccattctaga taccggataa gagtaaatca gggtacataa 15900 tgaattacag aacaagtatt gctgatgccc tagtggaacg actgaagaag gacatggatg 15960 ggagtaatcc cacagagttc ttcactaata tgtatgggaa cgtatcccgc cagacttatt 16020 cgtttgagca gatcaatgag ttcccttaca tagcagtcca tgtgggtacc gaaactggaa 16080 actacctgcc gtctgcacag cagtgggttt accttgaaat tcctattctt atctatgata 16140 aagaaaagga tgatattaac atgcaacttg agaaactcat agcggatgta aaaacctcta 16200 ttgacactgg aggaaattta caatatacta taatgaaacc tgatggttca actatcgatt 16260 ctgaagccac tgacatgcag atcacgtcgg tgtccacaga cgagggtata ctgtccccgt 16320 ttggttttgc tcaagttaac gtaacagtcc ggtatatgcc tctgagaaga gcgctggata 16380 gataagttac agctcaggag aaatttaaat gtctgtacaa ctattacgta atacacgaat 16440 cttcgtgtca accgttacta cggggttcac taaggccaac actcaggaga tcctagttca 16500 ggatgatgtc tcctggagcc aggacagtaa ctccactgat attaccctaa atgaggctgg 16560 ccctaagccg acccgcggtt cgcagcgctt taacgattca ttgaacgctg ctgagtggag 16620 cttctccact tatatcctac cgtatgacga tgcaggtaaa cagatcctgc cggactacct 16680 actgtggcac ggactggcaa ctggagctgc cgtgaatcta gcaggtacta ctggcgtatt 16740 ccagaatgct actaacctgg ttgtcaactt taaagacaac gggtaccacg aactggccat 16800 gttgaacatc tacatcctaa ccgatagctc ttggtctgtg atccgtaact gccaggttgg 16860 gcaggctgag gttaacgtag atattgatga tatcgggcgt gtaacctggt caggtaatgg 16920 tactcgcttg gagaccctag cttctcagcc gttcgaccct aaaaccatag ggatagacga 16980 cgctctttat gctaagattc agagttctta tatcaagaac aaactgacca ttcttaagct 17040 gaaacaac gctacggcg gaaaaaccta taacatcccg atcacgggag gttctttcac 17100 tatgaacac aacgtgacgt acctgactcc taacatcatg tctcgtgttg acgttccgat 17160 cggttcattc actggttcct tcgagctgac tggttccttg acagcgtata tgaatgatgc 17220 tgccaacggc tctatccagc tgtacaaaga tctggtttcc gacctgaaag ctgtgaacga 17280 cttcgaagtt gcaatcatcc tgggtggaga gtatgatact gctcgtccgg cagctgttct 17340 ggtggctaag cacgctaacc tgaacatccc gtcaatcgaa actgatgacg tgctgggtgt 17400 gtctattgag ttcaaggcta ttccgactca gatggacgca ggggatgaag gttatctggg 17460 cttctcttcc aagtacacca agacttcgat cgcgaagctg atcagctctg gtgacggtaa 17520 ccctgtcaca ccataaggat aactaatgct atactcccta atgcgggagt ctagagtagt 17580 catcgagtac gatggcaggg cgtacggatt tgacgccctg tctgattaca ctgctgggaac 17640 gtcctacgaa gagtttaaag caaatcgtag gacgattcac agacgcagta actacgccta 17700 ttcgaagata actgctcagt ctccttcttc aatttctcta actcttaact tctctagcaa 17760 tgctctcgaa ggtctatttt tcgagttgat ggggtttata gagatagacg gaatgtatca 17820 gatgcccttg ttcagtaata atattgagcc taaaatgttc tccgtatata ttattaacaa 17880 gaacacgagc ttacgtttcg ataactgttt tgctaccacc tgcgactttt ctctagataa 17940 gagtgtcccg gtgctaaacg ttggtatcga gtcgggatac tttgaggaag taggccaccc 18000 actcaacagc tatacgcttg atcaaggtga ggtgctacca tttcattac ctcaggtatc 18060 ttcgaatggg agagtgctcc caggacttat gtcagccggt atgtcattcc agcagcaatg 18120 cgaatggcga ggtgacagaa gtttattcga tatcaataag atttataata atagaagggc 18180 aatcgttaac gaattgaact catccgcttt gatatcgatg tactatgcaa agagtcttca 18240 gatagactct acgcataaca ttaaacctga tattggccta ccggtacaaa tcagaaataa 18300 atatattgtg gtggatttcc cttccactcg aatcacaaaa cgcctagact taaccgacgt 18360 gtacaagatc gattatgacg taatacctac tgagcaatca gatcctgtcc gaatcaagct 18420 aattggagaa taaaatga gtattaacct aaaagatatt gcactggata ccaaacagat 18480 caccattgca taccctggcc taccacactt caaactgaaa gttaactacg tctccccgtaa 18540 gctctccaag aaaattctgg aagctgcaca agagaaccag tttgttaatg gtatcgctgt 18600 taaagtgcaa aacgatgaca aattcgcaga agagttcgtc aaagtggcta ttgcaggttg 18660 ggaaggtctg accgtcgcag atgttgagaa actaatgctg atcgaagttc cagaagatcg 18720 tctggagaac aaagtcgaat ttagtatcga caacgcgatg atgttggtgc gtaactccag 18780 tgccttcgag acttggatga acagcactgt cttccaccta gacacttttc gtggctcaaa 18840 atcggaacct actgcttaag gaaatagatg cctttgcaga acgatgtgtc aaaggagggg 18900 actccaaaat tacacgggag foottctga cgatgtgcga atccctcggg gaggaaccta 18960 accccgaggt acttaaacgt ttcgtagaga tccatgactt ccctgaaatc gcacagaccg 19020 ctctaacaat atataacaac ttatcggata actatatccc cggagattat ccaacctatt 19080 taggcaagga taagagtgct ttactagttt tcttcgatat ctacggagtc gaagacgctg 19140 atgagaagag ccttatactc caaatcatca atatattcga ctcacatgcc gtggcagcct 19200 ctcgtaaacg cgttgaagca gctattaaga agtctaaaat gaagtcttct agtaggtagt 19260 gagttacagt ttcctccaac gggcgttcca cgtgaggctt ggcctagggt taatgcccta 19320 ggccttttta ttggaaaaaa tcatgacaga tagactaata cgagaattac ttgtagatat 19380 caagcagcgt ggtggatcca aagccgctaa acaaatcagg gatgttgaag ctgctttaga 19440 cggggctgct cagagctcag agggtctaaa tacaagcttg ggcaaacttc ctgggtcttt 19500 cacggcgctg gaacgatctg tatcacgtac tgctaagtcg ctggagaaat tatcctcgac 19560 caccagcatt acagcattag cagcgtctat cggcatgcta agcggcaagt ttacctcgtt 19620 cgaggttgac ttggctaaat ccgtactaaa aatcaacgca aacctaaacg gggtgacttc 19680 cgccgctaac aaaatggcct ctggttttga cactgcagcc acttcttcgg ttgcggactt 19740 aaaccgcgtc aataaggctc ttcaggagtt agatgcgcac gcctcttcag tagctaaagt 19800 gttgcagacc ttgaaggcgg gggccgggtt agaatctatt agctctagtg ctgctaaggc 19860 tagtacggat cttagccacc tagtatctgg ggtggaaaag ataggtaacc aattagctag 19920 aatggcggag caagccgtgc tggcaggcag gtctcttcag gggctgaaag ccgactcctt 19980 aggggctgcc ggagagcatc tgagtaaggc tgcgtccggg atttccgtag ctgtatcttc 20040 tatgggcgaa gaggtgaaca agctaaacaa actacttctt gagttggcag taaaggctga 20100 tttagcgagt aaatctatag caaatattgc accagggacc aaactgaata gtctgggaac 20160 tgaaatccag aagattaata ctagtttagc cactgcagcc aatacctcgg tagctgagat 20220 atctaagatt aaagcagccc ttacgtcttt agtatcctct accgctacag ctgccgcttc 20280 aatgaaaacc gtaggaaccg gtagtggtct gagcaagcta atctcagaga tatcagcagc 20340 cacctcagct tccacttcgg atatctctaa agtaacagcc gctttaaaac agcttaacgt 20400 agatgctacg gcagcaggaa aagcactgca aagtattaaa gcaggcgcaa atctttcttc 20460 tgtacctaca gttgttggaa agataggtac ttcaatgacg cagttgcgtg ctcagttaga 20520 aggatctgta accggtatcg agaaaagcct aaatgatcta tctagagctt ttgccactat 20580 gggaggtacg ggaaacctga atccactggg taactccatc agaggtatga tcccgtcact 20640 cacccagctg gctaaggccg ctgtgcaagt taactccgct ctgtcaaaaa tacaggcagg 20700 caggggcgta cttcaattac ctacccaatt caaagcagta acggcctcat taaatgccct 20760 ggagactaaa ctggcctcta cgtctcaaat actagagcgc ggattctcca agggatttca 20820 ggatatggcg tctaaatcaa cctcgtcatc tacgagaatg attaacaact tccagaaagt 20880 ggtaccggag ctcaacgcta ttgaagctgc tgctatacgt tctgctgctg caatagacaa 20940 gctaatagcc aaacgtatac gcctcggaca agctggggga gggggtaacc ctgcagcgtt 21000 caatatgggt gccttagtag cggaaatgaa caggattgta acctccattg aagctatggg 21060 caacaaaatg aataccacca tggctgatat ggcacggagt acggacaagg tatctgacaa 21120 attaacagat ctaaactcgg gagttcggga tgttaatact ggtttaggtg ggttgaattc 21180 aacgttaacg ggtacgggta gcgctgctaa tagggcgtcc agagcgttgg gaaatacttc 21240 aggatctgct cgcggagcta ctaggaactt cgcagcacta gctatggtga ctggccctat 21300 gcctcttatc tacggtgcta tagcctctaa cgtatacgtg cttaaagcag cgttcgatca 21360 gctaaaactt ggagaccagc tgaaccgctt agaacagttt ggatctatcg taggagcgaa 21420 gacaggtaca cctattcagt cccttgctgt ggcactgcag gaagctaccg gccacgcggt 21480 atcctttgaa gaggcaatgc gtcaggcatc tactgcggcc gcgtatggtt tcgacgctaa 21540 acagattagc gagtttgctc tagttgcacg tagggcagcg gctactcttg gcgttgatat 21600 gaccgatgca ctcaaccgtg taatcaaggg tgtgtccaag caggaaattg agcttctaga 21660 cgaattaggt gtaaccatcc gtttaaatga tgcgtatgcc gaatacgtta aaatacttaa 21720 tgcggctaac actggtataa cgtataacat tcagggtcta acttccttcc agaagcagca 21780 ggcatatgct aatgcggtag tagcagagtc taccaaacgc ttcggctacc ttgatgaggt 21840 actacgagca acccatggg agcaatttgc ggctaatgcg gattctgcat tacgcaggt 21900 tcacaagca gctgctaaat atctaggtcc agttattgca tctaaacg cagcattcta 21960 tacgtctaag gcttcggtat ctgcagaggc agctactgcc cagcaaggt cgattaagca 22020 aatggacggt aagactcta acgcagtggt catgaacctt gaggctctc agaaaggctt 22080 ggatgatgca gtcaagcaa agaggagt aaaaaaaa ctcgcagctc ttattagga 22140 gataatggat aggaggcga agatgat gtccactgca ctggccacag ccgccacta 22200 tagtgggttc ggtaatctgc ttaccctggg agcctactaa gctacaaag atttacaca 22260 acagactgca gatatgcgta gaggcgta ttgttacag caggagttag cagattctgc 22320 gggagctatc caaaatgga aagacgccag ggactccgct ctactagg ctcagaaaga 22380 gaacccagaa ctggcgggga acttaat agggcagaac gttgaagcaa gtaatggact 22440 atacaccttt gaacgcag cattagacgg ggcagttgct ctacggaagg agttcaataa 22500 tataagaaa acttccggag atctgagcaa cgatatccag aactttgcac aggactctaa 22560 cactgcgtct cgagctactg cagcactggg tgatgcactt aaggcggttg agtcattggc 22620 gggtggatct acggaaaaag ccaatcaaat gaccaaggac cttaatttgg gctattccac 22680 22740 tgaggctaag tctaagctag atgttgagaa acgtatagcg gaggtctacg ctgccactcg 22800 taataaggat aaggcggaag aagctggtag agccctagaa atgcagcaac ttagcgcgaa 22860 gaagaggca ttgaaagctg tgctggcgac gaacaaggac aacaaggcta ttcaaaaaaga 22920 gttgaccctg ttagagacgg aagagctcaa agtgaagaac cagggcatgg aagcgactaa 22980 gaaggaaa ttctataagg atagatagt aggcatagat cgagaaatag cactcctaaa 23040 taatcgcact atgacagatt ctcagtataa cgtggcgaac cttaaattaa atctacaagt 23100 agaaagat aggttagcct tactgaagac tcaggcagat aaggaagaagg aagccgaaca 23160 gtctagacgt aacattgcct ctattgaaag ggaaatctgg aaagagcagc ttgaccgtaa 23220 tgccaaaact gctgagatgc gtaaagaaga attcgagcgc aatcaaagca tgaagcctct 23280 aatgggagag tcacagaaaa tgcaggagca gctagcgttc taccaagaaa tgaaggaatt 23340 cacgaaaggt aacgctgatg aacaggcgcg ttggagcaag gaaattgcta acactactgc 23400 gcaaatggcg gctctcaaag ctcagcgtac tgcacagatg atggatcgcg taggacagtc 23460 cttgggcgca gactatacgc ctactactgg cctggagggg gaggataaga aattcgccga 23520 catggaaaac cagatggcgt catacgatac cgctatcggt aagctctctc agctaaattc 23580 ggaggctact gctactgctc aaagtatggg gaacttagct aacgctatga tccagttctc 23640 tcaaggatct ctggatacta cgtccatgat tgcagcgggc atgcagacag ttagccagat 23700 gatcagctac ggtactaacc agcagatttc agcaattgat gcagctatag cggcagagca 23760 gaagcgtgac ggtaaatctg agcagtctaa gaacaagatc aaaaagatgg aagcggagaa 23820 gattaaactt cagcaggaat ctgctaagaa gcaaatcatt atccagacag cagtagcggt 23880 aatgcaggca gcacagctg ttccgtatcc gttctctatt cctctgatga tcgcggctgg 23940 gcttgcaggt gctctgtctc tagcacaggc gtctagtgct accgaatga ccgatatagc 24000 aggatctggt ggtgaaaccg cctcttacct aaccttaggt gagcgccaga aaacgtaga 24060 cgttagcatg ggcgcaacg caggtgaact atcttacgtt cgcggcgata aaggcatagg 24120 tagtgctaac ggatttatcc ctcgtgcaga aggtgtaat acctacccag gtgtctccta 24180 tagatgggt gaacacggaa cggaagttgc aactcctatg gtgcctacca aggtaactcc 24240 agcggataaa gtggctctg aaacttctc tggcggtgct agagaccgg ttaatctgaa 24300 catccaggca atggacgcta agagcttta ggagtacgca ttggaaacc ctgcggcatt 24360 ccaggcagct gtagagttag ccttgaatga acaagggctg agcttaaga acctgaatta 24420 actaaactaa agggaggacg taattgtcct cccttcttt tggaatttat aaattatct 24480 tgaaaatttt tcttgatatg actataatat taacattgag aggagaaac aaaacattgag 24540 attaccagat ccctttacac atcctcagta taacggcctt gggtttgata aagctacgct 24600 gatcgataat gatccagtga tcagagacga gctaccaaac ggcaaggtta acgaagttaa 24660 aacagccact cagtactggg gtcttaacat tagttatcct gtgatgtttc ccgacgagta 24720 tgctgtactt tcgtcagcaa ttctagagta taagcgtacc agaggctatc tcgacgttat 24780 actcccacat tacgagtctt accgagtaag aggggatgcg aacaactgtc gcattgccgc 24840 tggacaaaaa ggctccacac tggttatcac caatacgaat tccttatctg gagaacctaa 24900 gccaggtgat ttattccagt taaccacaca tccaaaggtg tataagatta catcttttaa 24960 aaacgtagca ggagtatgga cacttaatct ataccctgat ctgttgctca ctaccaacgg 25020 ctccgagaga ccacgtttca atgggattct tttccaaact aaattaatga acggagattc 25080 attcagcgaa gagatcacag ttgatggtgt atacgacggg gttaacctag ttctgagaga 25140 aagtctatga gacagatcct tccttctgcg aaagcctacc ttgccaacaa tgacaagata 25200 cgattagcgt atcttgtctc tatcgaactc ccggggtcca cgggtaataa cgctgtttat 25260 gcttatatga cggactatat gagagatatc aactatggtg gtatactct ccaatcagggg 25320 aaaattaaaa cattagcag ccacaaaaaaccgtacgt taaccgtcgg cagttttgagc 25380 tttagtgtta ctggtacgga tgccaacgaa gtcattaagc tcgtgcaag tggtgtatca 25440 ttttagatc gctctatttc tatatatcag gcgatcatcg acgacaatgg ggaaatcctt 25500 ccagtggacc cagacta tggccctta ctcttcttta ggggtaagat tgtaggtggt 25560 ggtatcaaag aaagtaatac agtatccgga gttggtactt ctgttataac ctggaactgt 25620 tctaatgaat tctatgattt tgagcggtt gctggacgct tcacagatga cgcttcccac 25680 cgaggacttg agatgtaaa tggagatta ctgccttctc acggtgccaa acgaccggaa 25740 taccagaag actatggatt cttccacgcc aacaagagcg ttaacttccct agctaaat 25800 caagtaaag aagaacgata caagctagaa tctagaga aattattcgg tctctccaag 25860 agctacagcc ttaaaaagta ttatgagact gttactaag agtagacct ggactcaac 25920 cttgcagcca aatttattcc tgtagtatac ggtactcaa aagttcctgg tatcccggtt 25980 ttcgcggata cagagagaaa caatccaaac gttgtgtacg tggtttacgc gttctgtgag 26040 ggtgagattg aaggattcct agacttccag tttggggacg cccccatgat ctgtactgat 26100 caaactgaca gtacatctcg tacatgcttt ggacaaaaaa gggtgtcggg agatactatg 26160 gcaagaattt ccacagggct cccatcaaca tctctctcca cgcatggtca agaatacaag 26220 tataatgacg gtaacggaga tatacgaatc tggacattcc atggcaagcc agaccaaacg 26280 gtagctacgg tactaagaga cattgctgct gctaacaatt tcttccttca aggagagaac 26340 ggtaatggtc cggagtactg ggattctagg tacaagttat tagacaccgc atacgctgtc 26400 atacgtttca ctatcacgga gaacagaact gatattcctg aagtatccgc agaattaagc 26460 ggacgcaaag tgaaagtata ccaggcagat ggttctgtta aaatggataa aacgagtcag 26520 aatggtgtgt ggcaaacatt tgactaccta acctccacca cctttggtgc aagtatcccg 26580 atagacagaa tggtgattgg tgactggaga aaagaggccg atctattaaa cattatagac 26640 acctcttatc aaactagttg gcaacctttc tggagatacg ttggatggga gagctggaca gccgaaaca gacaataat gcaaatgaac acaatcctgg ataactccaa ctctgtgttc aaaaaacgtgc aggagctatt agaatccttc caggggggcgt tgaataacct atcaggtatc ttccgcatca ccgtagagaa agattcaaaa actccgctag aacttaattt cctagatact tatggggacc tggatctatc to be ggccgtaata to be 26940 tctctgattg atcctaccct gactggaa accaactcta taacgttcta taattctag tttaagaatg aagatcgtgg agttgacaag aaacttcaac tttcttttgc taacataacc aactactaca ctgctagaag cttggcagat agagagctta agaagtctcg ttactctcgc tctctgagtt tctctttacc ttacaaattc cttggtatag aacctacga tcctgtagta ttcacctacg atcgttatgg ctggaataag aagttcttcc tagtagatga ggtggagaac acaagggatg gtaagataaa cgtaactctt caggagtatg gtgaggatgt atttattaac tcaacgcagg tggataacag tagcgaggcc gttcctgaaa tatccaata cgtcctgcca ccaagagact ttaagtacac ccctacacca ggtggaatgg taggtgatgt tggcaaaaac 27420 ggagagcttt catggcttcc tagcttgaca cctaacgtag tctattactc gatacgtaaa 27480 tccgaccgcg tggatcctta tattgtgcag cagactgctt tcacacctaa cgttagaatg 27540 ttccaagata tcgtgggtga gcctgctggc ctgactattt tcgagatccg cgccgttgat 27600 attaatggtc gtcgtagctc tccagtaaca atttctgtag atttgaactc agctaaaaac 27660 cttagtatgg tggaaaactt ccgagtgctt aaccttgccc ctgaccctgc tgaatgggta 27720 ggacctgact tagagttagg gtgggacaag ctgcaagaag aggggctgat ttcagggatt 27780 ttctacacgc ttgaaataag ggacaacact aataaactgc ttagatccgc aaagatcacc 27840 agtttatata actacagcta cctgttgggc tataataagc tcgactacaa ggccaacaac 27900 tcgaatactt tggggattta cagagcactt cagccaagaa ttcaggcgga gggaccaaag 27960 ggcgagaagt cggttgcatg ggcatatatc taaatgattt caaacatagc accagctaaa 28020 atggtactgc agaatatcgt gaccggctat acgattgcga gtattcagca ctcaatcttt 28080 tccgattacg acgtaattgg tagaactttc tggctaacta cgggtggggt aactacccgt 28140 agggacttta ccggcgttga tacattcatt gccacaatta acaatctaat cgctggcgct 28200 acctactctg cccagggtgc tttctatgac tcgatggttg atgcagagct gatggctgct 28260 aaagtaggta tgaacctctc tagcaccatt aacttcaaga tgaaaactgc tccgaagatt 28320 accaaagtgt cctcttttgc agaatctgtt gacgtgggtg tgggtgctcc tatggttgtc 28380 gtagagcttt ctggggaggc cgaatacgtt accatcgaaa tgaaacctga gggctctagc 28440 acctggacta aatactaccg tggtccaatc actgagcaga tcatctttgg gggtgttcca 28500 gttggcagat acaatatccg ggtatctggt gttgtcacta tgccagatgg tgttactgtg 28560 gatgtttctg ggtatgatac ctggccgtca ctgtttaacc tgacctacaa cttcactcca 28620 ccgtctgccc caactaacct gcgtttcaaa actgcccaca tccaagatgg tatggagcgt 28680 tttgacgttc gcctggaatg ggattggact cgtggtacgg gtgctaacgt tcgtgaattt 28740 atcatccagt atattagtaa cgatgagttt gcaaaaactg gatggactaa ggccaacaag 28800 ctaaacgtgg gtgcagctaa agctggtact atcactagct tcccttacaa aatccgccac 28860 cgcttccgtg tactatcggt tgcttggggc ccagatactc agtctataac taactctaac 28920 gaggttactt atattataga cgagagcacc actttcgaca atgcattcat taacgagacc 28980 ggtgtagaaa tgacctacgc aggtatcaag ggtaaactct ggaacctctaa caccaaacag 29040 tgggagcaga cttcttagt cgatgctgct acaggtgcag tagttcttgg tacactcgat 29100 gaaaatggta aagcgccgat ttcattcgac cctgttaata agattgtaaa cgtcgatggt 29160 aaagttatca ctaaagacat taatgctgct aacgtaattc ttactaacct gaccggtaaa 29220 gataacccgg caatcttcac tcagggtaag aagtacggta ataacgcagg tggtgtctgg 29280 atgggtgttg acaacgttga cggtaaggcc aaatttgacc tcggtaataa cactcagtat 29340 gtgcgctggg acggtgatac tcttcgcatt tctggtaacg ttgtaatcgg tactcctgga 29400 ggcgatgtag accttgaaac cggtatgcag ggtaagcaga ctgtatttgc ttataaactt 29460 ggtacatctc tgccaagtcg cccgcttgac caagtttatc caccagctgg atggtccgca 29520 ttcccgccta accgcactgc tcagaatcag aacgtgtacg ttgtgcaggg tactctggat 29580 cctaagaaga gtcctcctgc tctagtagac ggtaccaact actcagctgc atcccagtgg 29640 tctggtgtcc caggtactgg tggtactgat ggttctaacg gtgattacac tgttcagatt 29700 taccagatca gtgctagtaa gcccacgaaa ccaggcaata tcaatgaccc tagcggttgg 29760 agtcgtaccc caccgactgg aacccctctt tggatgtgtt ctggtagatt caatggcgat 29820 actaacgctc taactgttga gggttggtca gacccgatcc gagtagacgg cgagaaaggg 29880 gctactggtg ctactggtgc tactggtcct caaggtcctc aaggtccggc aggtggttca 29940 gtagaagtac agtggtctaa agatggtact actaactggc atgcaaactt tactactggt 30000 gatatctaca tgcgtcagcg tgttggtaca ggtgggtgga gttcagctat ccgtgcagtt 30060 ggggaagatg gtactaatgg tactcctggg tctaagggta actacattgg gatgcgcttc 30120 cgagtggcgg ctgagaaacc tgccactccg actggccaga ccccttcagg ttggtcagat 30180 gcacctcctc agggaaaccc tctatggatg gttaaagctg agttcaacgg tcaaaccaac 30240 gccctagtag gtacgtggtc agaaccagtt cgtattgatg gcgaaggtat tggagtaaac 30300 ctgtatccgg ttaagaaaac actggatcag tggaccggaa tgagcaacgg tactatggtt 30360 aagaacccag ataccctcag cttaccata accaacacag agagcactac ctcctctacc 30420 ggaccgggtg cacatcctgt tccgttccag ggttcacaag gtcctatagt tgagattcca 30480 gtaaagccga atacagcgta catcttcact tatgaggtta gtactgatag caccagcttt 30540 gttctgagag acctgttact agagttctct agcattaccg gaagttttac caacttccaa 30600 gagttactga ctggagccaa aggtaagcag gaagctaaga ttgtcactcg tgctgatact 30660 aagttcctga gcttccgtcc aggtgtccgc actgccggag ctacggttac ttattctaat 30720 ctaaaacttg aagaaggcat taaagctacc gcttacagcg tagaggcttc cgacagtatc 30780 ggtgagaaag gagaccaagg tactcagggt ccacaagggc ctcagggtaa tcagggtcca 30840 cagggtaatc aaggcccgca aggtgctaag ggtgataatg caaaaggatt cagcctctct 30900 tctctcggtc agacgtttac gtacgacgca gaaggtaaac tgaagtccga tgcaaccatc 30960 ctgttccagg ctttccgtca gaatactact gcaaacgtta cttggtcagc caaggacgag 31020 aaaggtggga acatcactct gacaagtact agcaatactg gtgctacact gaccgccgct 31080 aacttcaaaa catcgaagtc tgtagtagtt acagccgtct gtgatggtat caccgatcag 31140 atcactattg tgcgtctaga cgatggttcc aacgctctcg tagggcttct gaccaacgaa 31200 ggttccacag ttctagcgaa ctatgctggt tacgtgcaaa actatagtac tggttctggt 31260 gacttcaaag tattctatgg ggcgaaggat atcacctcag aatgcacctt cagtactatg 31320 gagaagaata accttgatgc tgatattact tcagcgggca aatatacttt aaaaggtatg 31380 ccggctggta ctgatgttat caacggctgg gtcgacttac gtgctgtaca ccctacttac 31440 ggtgccgtgg tccgcagagt ggcaactact aaatctatcc ttgcaaaagg ttatgatcgc 31500 gttattacca cttcattcga gaacggaaat aagggtacct ggtcgactgg tagtgtccaa 31560 ggggtttctg gtgccacaat tgtagccgca ggtttcagca aggccctagt gatctctgct 31620 agagactgta tagagatgc taacgcattc cctgtagtag ctggtcagaa atagatactg 31680 ggcatgtgga taatggctag tgagtctaaa gtcaatatca acatgggaat gcgaattgta 31740 agggcggccg acggagttgt tgattggcaa ggaaccctta tgattgcaca aggtactgta 31800 gtacctggtg gctggtccta catcgaaaaa gagtttactg ttggcagttc taataccggt 31860 atagcaatgc cgtggatcca gatggctggg tcttctggta gtgacttagg taaagcttac 31920 gttaccgata ttcacatttt cgccctagaa atggatgggg agaaaggtga tactggtgca 31980 actggtgcta caggttctca aggtccacaa ggtccacagg gtaataaagg ggacaaaggt 32040 gatactggtg caactggtgc tcaaggccct gctggtagct ctgtaaacgt ccagtggtct 32100 aggatgggt ctactaactg gcatgctggt ttccaacctg gcgatatttt catgcgtcag 32160 caggttaatg gagtttgggg cagtgctatt cgtgcagttg gcgaagacgg taaaaatggt 32220 gctgatggta ctgacggtga ctacatttca atgaagttta tcgttcagga taccaaacct 32280 ggcacaccta ctggtaacaa cccaggtagc tggagtgatg ctcctccagt tggtagccct 32340 ttatggatga ctaagggcac tatgaatgct agcggtcaac tacagggtac gtggtctaat 32400 ccggtccgcc tagacgggac tatcaaccct aacctgttcg cagtacgtaa gtggatggca 32460 gggatgactg gtacggaagc cggcacatct aagaacgata tcgagaagct agcgcatacc 32520 ttgactcgta cttccggaac tgataatacg gctccagggt gctatgcgac cccgtatcta 32580 ggtagtgggg cgttttccca tccagtgact ccaggtaaac gctacactct aacctataat 32640 atcgacgcgg ctagcgaggt ccagacccga gatactatat tctggcaggc taatccagac 32700 tcaggacaat ccacctacat agaggaactg aataccggga cgtctataaa ggttaagcgt 32760 acctttgtag ttcctacagg tatgaactac ctaaccctac gcccttctgc tctgacactt 32820 aacgtagcga ctacgtggag caagattaag ctggaagaag gcggagagaa aactgagtat 32880 caagtagaat actcagacag tatcggtata gtaggtaaat cagttttagt acagtggtct 32940 aggatagct cttttagtaa ctggcatgat accttccaaa caggtgactt gttcatgcgt 33000 cagaacgttg acggtgtttg gggtcctgcg attcgtgcta taggtgagaa aggtgagatt 33060 ggtcctgacg gtaagaaagg taactacacc aatatcatct tccgtatttc ggatactaaaa 33120 ccagctaaac ctactggtaa caaacctacg gattggtttg atgctccacc tgatggttcc 33180 cctctttgga tggcaacagc aacgtttaat ggggatacta acgctatcat tggtgcttgg 33240 tctgaccctg tgcgaatcga tgcttcggc gtaggcgaaa acttcttagc gttcaaagag 33300 tggatgatgt ctattcagag ggctgagggg acgggctctt cagttagcaa gaatccggac 33360 cagatgagat tccgggtaac agctgggcca tccagaaatg acgcgtacac taccccttac 33420 caaggaaccg gcacacattt tatagaagtg tcgcctaata ctgtgtatac tttatcattt 33480 gaaatggaaa ctgctgtgtc cactagaatg atgctgttgc agtttgataa tggtaatggg 33540 ggcacgcacg cacgtaacaa tcaggttata tcaaccagta ctggtataaa cagtctgact 33600 attactacgg gtgctaacac tacccacctg tctatgcgta tgtcaatctc taacatggga 33660 gagactaacg tactgatgaa acctaagctg gaactcgggg cgtttcctac ggcatacgtg 33720 gcgcatccta gtgatctact tggtaaagat ggtgctactg gagctactgg tcctcaaggt 33780 ccgcagggta acactggtgc tactggtgct acaggcccac aaggtagtaa aggtgacaca 33840 ggggctactg gtcctcaagg tcctcaaggt ccgaaaggta acgcaggtga aaacgctaaa 33900 ggattcgccc taacgtcaga ttatcagtca ttcgtgtatg atactgtagg ggacattaaa 33960 tccgctacta ctattctgtt caagggacta aaacagaata ctactgcagg gatcacgtgg 34020 agtgctgtaa ataatacggg agctgcagta acactgatga attctggaga taaccgtcag 34080 ctaaccgctg cgaacttcgg cgcctctaag tgggttacga taacagcaac ctgtgatggt 34140 ttatcagatc aaattactgt ggttcgtttg caggacggtg agaacgtgtt gaccgctgtt 34200 atgacaaatg aggcagctac ggtacttgct aactactccg gatattgcca gagttacgaa 34260 aacgccaaag gtcagatgag agtttggtac ggaagcaccg atgttactgg tcagtgcact 34320 ttctctgagg gcggaagaag taacgtaact cctagcatca actcagcaaa tgggaactac 34380 tccgttactg gtatgttgga tgggaccgat attaccgaag gttgggtgga cgttaaggct 34440 actcatccaa aatatggagc aattaccaag cgttttgcgg taactaaggt attcctagcc 34500 aagagctatg agatggttat caccaatacc ttcgagaatg gtaacaaagg ttcatgggca 34560 ggagctctgc agagtgtttc cggcccaaca aaccagagca tctctaaggc gctgcgtatc 34620 acagctagag ataacctaga gggtaggaac accatcccag tagcagggg gcaaaaagtc 34680 cgtatcagat tctggtacaa cccactagga ttaggagaag ccatttttag agtaggcttt 34740 attgttcacc gcaaagatgg gggcaaaggt tacccctcca gaactgtggt tacgggcccc 34800 gctcctaata gctcctgggc gtacttcgat caagagttga ctctaagtgc taacgatgag 34860 ggtattgcct ggccgtggtt ccagttagat aacaaaactt ctggttcttc attagggtat 34920 atgcttgttg ctgacataca cttcgaagat ctatccatgg atggtgcaga cggagctact 34980 ggtcctcaag gtccgcaggg taacactggt gctacaggtc ctcaaggtaa caagggggat 35040 actggtccac agggtccaca aggtcctgca ggagcttcag ttggtgttca gtggtctaaa 35100 accggcaacg cgagcgattg gcacacaaac tatgctactg gcgacattta tatgcgtcag 35160 caagttaacg gtgtgtggtc ttcggcaatc cgcgcagttg gcgaggatgg gcgggttggt 35220 gctgatggta aattacctc cctgagattc caagtagctg caactaaacc tgctagacct 35280 acagggaact ctccggctaa ctggtctgat tcacctccag aaggttcccc actctggatg 35340 gttaaaggtg agtttgactc cagcaaccaa cttcagggaa cctggtcaga tccagttcgt 35400 ctggacggtg agacggtcaa cctcaacctg tttgctaaca aagcatggat tgcgtctata 35460 acgggtgcca gtggtagtgg atctgttgta gccaaaaacc ctgacgaact acgcttacgc 35520 attaccgcag gttctggtgc aactgacgcg tataccatgc ctagtggtgg tgatggtacg 35580 ttcttcacta aagttaccgc aggtaagcgc tacactatgt cgttcgacac ggatagtgct 35640 35700 ttctcttgga tagcgtcaac aactgctggc agaactagct ggtcgttcac tgttcctgca 35760 ggatgtgata gggtatctgt ccgtgtgtca ctgaacaa acccaggtgg aaccaacgtt 35820 gtttccaata tcaagctgga agagggagat ttcgccacag cgttcattag gaacgagctg 35880 gatactattg gtgctgatgg ttcacaaggc ccacaaggtc cacagggtag taaaggtgat 35940 aaaggggaca caggagctac tggtccacaa ggtccgcagg gtccaaatgg tactagcgcc 36000 aaagcctttg ccttaacatc cgatagcctg tcctttagct tcgatactag tggtaacctg 36060 aaatctaatg gtactatcaa gatcgatagc tggagacaga ataccactgc tgcaataacc 36120 tggactgcca agaaccaagc agggagtaat ataactttag gtggcactgc tacaaaag 36180 actataacct ctgctcagtt cggaagctca gagtacgtaa cagtaaccgc gacgtgtgat 36240 ggaataaccg attccattac aatagttagg ctgcaagacg gggtgaactc tctagtcggg 36300 tacttaacta acgaagcggc taacctgtcg tgtaactcct acggttttgt gcagaattgg 36360 gatggcacta cgggtaactt taaggtgttc tacggaacgg tagatgttac cagtcagtgt 36420 accttcggag tggaggataa gagcaatctt aacggcaaca tagggtcaag cggttattat 36480 gcccctagcg ctatgccaaa cgggctggaa attacctctg ggtgggtaga ttataaggct 36540 acgcatccta agtacggaac acttattaag aggtttacac tcaaaaagag cctgcccggg 36600 attggttacg acagagtgtt cacggggtcc tttgactctg gtaacaatgg atcgtgggga 36660 cgtacggtag ttgacatcgc tacgggcagt cccggaggac acaccaaggc tatacagtgc 36720 acctctaggg acaccatgga aagtagtaac tggttcccta ctcgtaaggg gatgcgctac 36780 cgtgtaactg catgggtgaa caactctgag ggtgagtatc agctaaggtt aggcctccat 36840 acccagaact cttctggcag cgttaacact ggttacccaa ctatgctagc cgcatcagct 36900 aaggattccg agggatggaa actagtaact ggtattgtca cggtaggaga tgggtctact 36960 gcggagactg gtagggcaag accatttatc cagatgaatg gtagtgctag ccccttcggc 37020 aacgcttacg tagctgctat agcgatcgaa gacctatcta tggatggtgc agatggggca 37080 acagggccgc agggtccaca aggtaatact ggtgccactg gtccgcaagg tgataaaggg 37140 gatactggtc ctcagggtcc acagggtcct gcaggtaact ctgtaaacgt tcaatggtct 37200 aaggacgggt ctactaactg gcacagtacg ttcacttcag gtgatctgta catgcgtcag 37260 caggtgaatg gatcctgggg tcctgcgatc agagcagtag gtgagaacgg ggctaatggt 37320 acgccaggtt ctaagggtaa ctatgtgagt atgaagttcg ccgtaatggc tagcactcct 37380 agtaggcctt ctggtagtaa tccggctggc tggtcagata gccctcctcc aggtaacccg 37440 ttatggatga ttaaagcgga gtttaatggg gagactaatg ctattatagg taattggtca 37500 gatcctattc gcctagatgg ggatagtatt aacgagaacc tgttctactt taaagcctgg 37560 ttagactcga ttaccggagt tgcaggcaac ggttcatcta taggtaagaa ctacgagtta 37620 ctaagggcta ggataatcgc aggtacagga gttaccgatg cgtataccct cccatcagat 37680 ggatccgcct ccatgttcac ctaccttcct ccaagcacca catatacgat gtctttcgag 37740 actgataacg ctgtagaagt tcgttgtcac gtattttggt acgctaaggg aagcaatacc 37800 actggaggag tgctgagac tattgcatct actactgcag gtttgagcag ctttaccttc 37860 accacgccgg ctattcgga taggatatcc gttaggttct cagttaacga atctggggga 37920 aaacgttg taggaggtg taagattga aagggagcct tcgtaacgtc atatgttcgt 37980 aaccagtag acgctgtagg ggatcgtggt ccaggggttct atactcaggc gatcacaaac 38040 38100 ccggttaagt atgacgtact aactcagtat aagtcgggct ctccgcagaa ctcctggact 38160 cgtcaatgga atggctcggc atggacagct cctgcattaa ctgttcatgg tgatatgatc 38220 gtctccggtt ctatcactgc tgataagatt attgcaaca acggcgttcct ggcgcagatt 38280 ggtgttgaca tcctttacaa tagggccgcc gcactaagct ctaacccaga gggcacttac 38340 acaatgaaaa tagacctggc taacggggtac attcatataa ggtaacaaat gagcacggaa 38400 aacagagtag ttgatattat ccttgatcaa aacgtgtcat acggattgat gctacagttc 38460 atggatatcg atgactctgc gtacccagca acggaaaccc ctgtcaatct gacaggggta 38520 acccttaagt cttcaattaa agactctctg gaatccactg gggtaaaatt agcagatttc 38580 gtcgtaacag tagtaaacgc tacacaaggt caggcgtcgc taggattaac tgcggctacc 38640 gtggcaacaa tcgttagtaa agcaagtaaa gaacgagata aatataatcc tagacttcgg 38700 ttcgcaggtt actatgatgt aatcatgacc aaaggaacag gagctaccgc tacctcttat 38760 agagtcatgg aggggagcgt gtacgtcagt gatggagataa ccgcgtaatg gctattacaa 38820 38880 acccaaaatt taccgtacag ctaggatact cagttagttc tctggctgct agaattac 38940 tagatgctgc cactagatcg gcagcatccg ccgccgccgc taaaacttcg gaaaccaatg 39000 ctaaggcttc tgaaccgct agtaaaaatt cccagacagc agctaagact tcagaacta 39060 acgcggcagc atccgcccaa gtagctcaga atttggcagg taaagcgtcc ctagtaacac 39120 cactaggagt gatgaccggc tcggcagaag ctaagattgc atctataaca atagccgcaa 39180 atcagtcttc tagcgttcac gtattatttg cgctctacgc tacaggtaat ggagccaaca 39240 gggatgatat atacaatatg gagatagtat ccctggcatt acctggtcct gtaacctctg 39300 taaccgcgga caacatcggc agcttcctta gtcatcgagt aataggtccg gccaacacta 39360 atggatttat ggtaggactt aaatctacta ttgagggtag caacgtgacc tatgatgtat 39420 acctcaaatc tcgtagtagt ttcagagacc ctaagatggc attcttatcc ggctccatat 39480 ctgttactcc acctaccgga cctttggtgg acggaacatc ccctgcgtgg aaaactacag 39540 gttttgatac tgatgttatc tatgtaaata gggcgcaagt aattgatgat ggcattagtt 39600 tagcccgcat caaacaacta gctataacta acggtaaaac cgatagctct atacttcttt 39660 tatcctattt aaatgaaaca ggtatactat ctaccaataa gaagtctatt tccctccggc 39720 caggaggtac gagtgactct agtattgcag ctacggagtt tctacctaac gggaatataa 39780 ttctgcctaa tggggatact gggaaccaaa ctattagttg gttaggtggt ccccgcatac 39840 gagtcaactc caacgggtct tttgttcttt ctactaataa tcccagtaat caaactagtg 39900 ggtttataac tttcaggcca caaggtgatc aagtaacttc cactgagctt cagattaggg 39960 atgatggtaa cattaagcag acagctccac agtcatcggc aggcaatgca cttatacgcc 40020 aggatgcggc tattcaacat atcatggata aggctccggc tgccggtatt actgctaacc 40080 ccctaagcga cttgaacgta atacctacgc ctgaaggtac agatccttgg ggagcagacg 40140 gtgtacgtgt attccaatca ggggtatcaa caaaaaatac tccggacgga actactggtc 40200 ggcttggaac tatcctcaac gttaggcaca cccagtaccg tataatgcag ttcttcatgc 40260 agagtaatgc tactgcacct attctgcata ttagatcatt aagggctgat cagggtaata 40320 ccccaccggc atggtttaaa gtttatacag aatactctaa acctaacatt cagtctgaca 40380 tagcaggtat tactatagac ggcaatggtt tcgttaagaa agcctccccg atcgccaagc 40440 tcatagccga aattcctagc aaagaggatt cattcttctg gatagcgtg gaactgtgg 40500 gaggttacgt agggtgtaac gcagaggctc aaggtgtatt tgctgtaaaa actggactag 40560 gcaagtatac tattaaaggg agtttaggct ggaacacgga aggatggaa ttgagctac 40620 cagagatga taacggcaat atgttgtgct tcgtggaatc ggattggaat gaggagaga 40680 aagagctaaa catacaggtg ttcacccgta agttttgatat taacacaggt atatcatcg 40740 ctggtgaacc tatggata ccacaggtc gttggattga tctacgtctg gaatgccaa 40800 aagtggaat accagaagtg gatttccag aagatccaga agtataata aaaaaccc 40860 cagtcggagc aatccgctgg gtttttcta tttacatctt accagaactt ccgaagccac 40920 cttcgccacg atcagtgtcg gtcagactat cgacaatttc aatgtcattg gggttatat 40980 gaggtacgat caccaactga cacagacgct caagttatc aatcggttgt gtctcgttgc 41040 tcatattcaa caggttcatc ttgatggttc ctcgataatc tcatattca accctgttg 41100 tattggcaat cattagccga cgttcccta gagaacttct cggaaccacg attcctacc 41160 agccttcggg aatctcaacc gcaactccgg tatcaatcat caatgactcc ccaggaggta 41220 tggcacgcaa taggtcggaa gcacgatctc cgaagtaggc ccgtagatcc ataccggctg 41280 cttccgagct tccaacgtga ggtttacaat cggggtgaga taatttaata cgcattatac 41340 ttctcctgtg gcaatagccg taatgtcttt aacaaattga tcgtagatat cttgtcctgc 41400 tgcggcaact gcttccccac agaaacttgg gagatctact agagttaggt tacgttccat 41460 aagttcgcct gatttattca gagcctggac gaatttttga gttcctggga gcggtagtgc 41520 atcaataatg tctagcacgc taccatgttc ccggattagg ttgtagccac gtttttcgcc 41580 gataccttca acaccacgaa tgttatcccc catatcgccc ataatggctt ttaatgagat 41640 aaactgatct acagtatcta cgttatggtt gtcgaacata tctttctcgt gatattcttt 41700 acgagtagta aacgagaagc gggagatatt cggagcaagc agggtatccc agtcaccgtc 41760 tgtggagatg agccagatat gatcgtagtg atgacctatc agctgaataa tgaaagccgc 41820 catgtcatct gcttctactc cgcggattcg gaaagtaggg aattggcttg caataagatc 41880 aaacgcatcg tctaagtatt caaagaattg acgatccgcc tctacttccg cttcggagcg 41940 atcggcgtac ttagcgtctc ggttgccctt atactccggg aaaatattgg tacggaagat 42000 actcttccct ttatccccta gaacgatagt gtgcttcgca tcgtaggagt ttgccaaaga 42060 gttgatggtg tttgcaaagg atgctgcaat aggcttacca ctatctttct tgaagcggaa 42120 gcctaagttc gtaccgtcaa caatcataag attacggcgt gaggcgaggc gttgttctgc 42180 ctctcgtttc attgttcccc aggatttact catttaacta aatctccaat ttcacatgcg 42240 tttaaccacg gctcaaaaag accgataaca atttccattc cacgtttatt aactatgaaa 42300 tgtgctctac tcatgaggtt atcaatcata gggtcgctac tatccagagc gataagccat 42360 tgaccacgat ctttcttaaa gatcagggcg ggtttcatgt tcatttgctc accttcacgt 42420 gcggcttggg cccaccattt ttctagctgg gattctccaa cgtttaggat attgctatta 42480 aatttttcgt ctgcgtagtg tttgatctca aaacagtatt ggctcatttt acctacgcta 42540 ggtggcagat acacatcccc ctttagggag tgggattggc caaaagctcc tgaaccagga 42600 acacgttccc agtccaagga ggtatattta cgcagcatat cacggatttg gtattccgcg 42660 cgttttcctt tctcgcgact atctacggcc attaggcctc caaaattgat aagccagctt 42720 cgtcttgttt aacctcaatc ttatgtgcat acggatgtgt gtgcccgtga gatacgatga 42780 ttgagttcag ctgatcttct tcctgtagaa gttctacgag ggtattaatt ccatccgggt 42840 caatataact tacaacttca tcaaggaaga gcaggttgat attaacttta ctaatcgttg 42900 aaagcaacat tcgaattgct aatagcgtgg aatattgat acggctctgc tgaccagtgg 42960 agcagttgac catagaagtc ctgttaccat cattgtagat gacaacctgt agcttcgtct 43020 cgtctaactc gaaccctaga gcaaatttac cgctggtcat gatagagaga taatgattga 43080 tcatctcttc aaataccttg acactatgct ccagcttata ccctaccaga tcttttagcc 43140 cggtaattaa gatatctaga tccgctactt cttctgcaac taaggacagt tcagcctgga 43200 tcttatcaaa ctcctgctcg gccttttgaa tctgttcacg cttagcttca tatttcgcat 43260 tttcacgtac aactgattcg ttatgggttc ttgcaagttc taccgaggat ctgccttgac 43320 gaatgctaga ctcaatagct ttgatacgct cagctaaccc gctagcgtct acgatttcat 43380 cctgcgttcc attgacctca tcaaattcct tgaggttttt gcgagcctgt tccagagcgt 43440 taaacttaga gatataatcc ttaaacgccc tatcttcctt cctaagttct tcaacttctg 43500 cttccagctt ttgacgttct ttatagagcg gatcatactc ctccttagtt cgtctcatag 43560 cttcttgggc ggcggaggta tctaagtgag ttccgcacgc agaacactga gttttacctg 43620 cggcattttt gaaatcgtag taacgtttct tcaaatcatc cgctctagtt gccacgaccg 43680 ttaggttacg cgtaacactc aggagttcct cagatttcga ggtgggcgca ggtaaatttt 43740 cgaatggtgc aaaagttttt tcggcagctt gtacggccat gtccaaagag ctacgttttg 43800 caatactggc tttttgcttc tgagccagtg ctaccttgac ttttgcttcg gttaattctt 43860 cgactagtgg ctcttcatcg aaaacaggga ttgggatctc tgattgaggt gcgccaagcg 43920 aggcttttga ggaagtata gattgggcag ttcgcaggct accttccaat ccggacagtt 43980 ttgccgagac ggctttacgt cccgctttaa cacgttcaga aacttcctta tattgttcct 44040 gatcaacag gttaacgagg aaagtcttgc gctgagcatc cgtagtcttg aggaagtcga 44100 gactggatcc aactgactga tatacgagct tagtaaatgt cgtaaaatca caggcgagta 44160 cctcttctat cagcttatac gtttgggttg cggtgtgtcc gctgatatct tcaccgtttt 44220 tagtgagagt tactttggcg gttgacttca caactttctt gacgttgtac acgtcaccgt 44280 cttttgtgaa ctgaccttct agcgtatagg ccttggagcc agtatgccag ttaaacaggt 44340 cgtccttctt gataccacgt gagttcttat tataaagaag ctccttaga gctctaccaa 44400 ttgttgattt acccagcccg ttacgcccga ttaactgagt tacgcggtga ttatcaaatt 44460 ctatcaccac gttctcagca tatgacataa aatggctgat tgtcaattta ttaattacta 44520 tcgacatact cagcggctct ctttaatatg cgtgttctag acgcttcgtc tagtttctgc 44580 acgtttgtga aataagtatc cagctcctgt agcagtgaca tatcgtctag atctaactta 44640 gcatctttgg tgacacggtt attgatcttc ttatctaaca gttcgctgtt cttaacgtt 44700 ttgagttgtg atacgtcgcc ggtgacttcg tatataactc tgtcatacgc atcggcttc 44760 attggctcgc cgacttcaat agttttcgt atgagctgtg gaagatctcc cagttcaatc 44820 cacgaaaggt agtgctcatg gtctcttggc aagaccgtat cgatgataaa agctccgtta 44880 gttccttttg tcctttctct gtgaaacgaa gttgtaagcg gagatccggg atagagcagg 44940 tcaacatccc cgatcttttg gctgttcttg tagctgtgga gatcgcctgc aaatactttg 45000 gagtacccat gttcgacgaa tctttcgagg tagatttctg gtacgacatg aggcgggatt 45060 gcacctctga catgcgtgaa gcaaatgtct gaaactctgg gcttccaact agctttgtga 45120 aggctgctgt aagggatgat atcgaactct ggagatcggt agtctttgac gactttccaa 45180 agtccatctg ttgcgtcgct aattgctgct gcatagtgat ccaggcaaga aatcgtttta 45240 gatttcattt cgtggttccc ggtgtaaata ataccaggat gttggagggt agctaggaat 45300 gcaaacagta gttcaacttc ttctgaagac gggtctgaaa catccatgat gtcaccacca 45360 ataatgtgaa ggtcacatcc agtagctcca aacacttcgt ccagcttttc accgagaaga 45420 ataaaccgat ttttctgcca ttcctgagga acgttcttcg ctcctagctt gatgtgatga 45480 tctgctgaga aaagtatttt catagtcaaa ataaaaggga gccgaagctc cccttgtttg 45540 gattaatcga ggtcagaagc tgcttcgtga tcgatgttac cggagttcgc ggaaccttca 45600 ttacctttag attcgtcgcc ctcctcatct ttcttacctt ccagccatgc ctggattgct 45660 ttcttctgct catcatagga cggaaccggg taggtaacat ccagtttagg cactttttcg 45720 aagccttcga attcgtctgc ttcgttgtac agagcctcac caatcagagc aacgtcggct 45780 tcatgcaggg cggcctcttt agaacccggc tgatttttag ccatctggaa ctgcattgca 45840 gcgatctgct gtacatcgta cttggtatcg aagccagtac cagatttgga gatagtgata 45900 tcgatctcac ccggatcttc cagattcagc tgggccatga tggagtggat accggtcagg 45960 atggttgcct taacttccat cagcttcagt ttgttatcag tacgatcgat aacaacggcc 46020 aggtagtttt tcttaggacg cagcggctga gccttaccat ctttctttc cggatctttg 46080 aagcccatgt cgtgaaccgg atcggatgct ccacggatga aacgctcttt ttcacggtcg 46140 aaacgcaggc actcgaacgg agccggttta ccttccttat tctgaatcca gtacacataa 46200 cgcgggagaa cgccggaaac gatacgtaca acgttcttac cgttgttgaa cttcatgtag 46260 tccagtttgt cgccgttaga accgccagta gtagaacccc aagatttagt agccatattt 46320 aattttcctc gataaatgtc aattttgact tgttgattgc gattagtggg tggtgctcga 46380 taactgctct aggaacccac ggtggaatat attgcatatc cagtgatgta tccttcgaga 46440 agctatattc ggcatagttt ctaaaactca aaagtcctag atactcagcc aactgtcggt 46500 ctgacagctt tcgtttatta taaacgattg tatcttcgtt caagataaat gatctacccg 46560 ttaacataac atgagcttcg ggatcttcaa ccattcgtct gataatctgt acaatcagcg 46620 atgagttacc tcgtgctaat atgtaaattt tttcgtaatc ataaacaac atgattatta 46680 tacaccattt tgagagaatt tagcaactaa gtttttattt ttctgcttcg gactttccga 46740 agattcagtt gctgttccca ttaatttata cagatattat atcgtatttt tgaggactca 46800 gcaactgaaa tttttaacta tttgcttaac tcttctttgt aaatctgtct ctcagttcaa 46860 tataaatatt atatcaaact ttggggttct tagcaaacag aattttatct aagtttgcca 46920 tattcttctc gttaaagtca agaacttccc acccattgag acgatatacc gcttctctac 46980 ccgccgcctg cttaaatcct acacctccta gcttcagatc cacgaggatt gggtctaact 47040 tatcatctgc catacgttgg acacgacctg ccaactgttc gattagagat tcgttattta 47100 ttaaagagcc taacactaaa cacgataaag catttaatga tacaccctcc gagaagatgc 47160 tttgggaagc cgctaacaca cacggaccgt cattggtcac gtcaatctgt acttgctccc 47220 tatcatcgag cgaagttaca ccggtgatcg tgtaggtctt cacacccctg agttccaaag 47280 cattcgtaac tctctcaatt agatctatac gatcggacac aaacaggact ttatggcctg 47340 ccatcgaata tagctcgcac aagtcaacaa cctgttgaaa gtattctggc tgcgaatata 47400 catcgttagc tcttatcgcc cacggcacat tcatatttc agaaacctgc gtttttagcg 47460 caaacctatg gatagtaggt ggcatagtgt tatttaccgg aggactatat actttcgttc 47520 caaaatagtc cttaaacata acctgcaaac catccttacg ttttaatgta ccggatagcc 47580 cgattttata tcgtgccgaa gactgctcta gaaacttggt aaacgtggtg gcaacgcagt 47640 gatgcacttc gtcaacaata acggtgccga attctttgc caaagcggct ccgtgtttgt 47700 taactgtctg tatattacta ataacgatag gagaatcgat attaaatttg ccagacccaa 47760 taactcctgg ctcaatccca aagaacttgc gtacctcctt ctcccacata gctcgaatgg 47820 tagtgttggt acatataact aacgttttct gcccaagtttt atgggcgata gcaagagcaa 47880 ggattgtttt accgaaccca ggcttaccat tataataca cgtatcatta cagtcttcat 47940 atatagggag ctgatcccct ggtcgcaggg tgaacgatgg ctttggaata tccataggaa 48000 ccagagtacg tttatctacg atctctgtca cttttgccc gaaagattct agaagatcta 48060 atctggttac gggaaaccac ttgatctt tacctaccga tcctgagttc tggaacatta 48120 gaggatactt tgccccaggt ttaagatct ggtagaagt gtgttttaat aagtattccc 48180 acagttcgtt gctgggttta caatagattt tattagatat actactctc atatcttaac 48240 tctaagtcta ggagttctg gctcctctc gtgcgtgtca aaaataactg gtgaaccgcg 48300 aaccaacgcg tacgaataat agttggcagg ttcactaagt aagaaggat atggaacgtt 48360 cttgacataa cactggtact ttccgttgaa gattctagtt gatcctctaa cgcgttccgt 48420 gataatgttg taataagtcg taggttcca tgttactatc ttacctgtgg agtcgatgaa 48480 atgctttagt ccagagttta ttatctgaga taagtacttg aactgttttct ttaaggata 48540 gagcttataa ggtaaatttt cccgctcacc aaagaaaa agccttcgct gagagaatgt 48600 cccaggaagg cttggattgt ccagcacata ccttgtaaat ctcgtctgga taacaatgta 48660 ctccccctct tggaagta catcataagc ccttaaagca tatactggta agctaagtt 48720 aagcaccaag aaccctgcga acgttgacaa tatctttgct gatttccggc aaaaacttaa 48780 tattggcgta cttatcgtga tcagggtggt tttcatcgtt agccgcaata gcagcgtagt 48840 cgaagtcctc cataccgata atactacgta ctttctcctc aaactccttg tcctcaatgc 48900 aagcaacaga aggacgctgt ttcttgatct taccacagga gtaatcacga gaaccgccag 48960 cctcggagtc agagtcgata ccaataggac agcccggaat actaatacca cggtctacct 49020 gaatgttacg gattagcatc tcgttatact gatcaactaa atcctcacga acaatggcaa 49080 ccacggagtc gtgaaccagc atgataatct tcatttcgtc ctgcagaccc agagagcgaa 49140 tctcattatc ggtatcgata gcgcctagca ggagactatc agaagatgca gactgaatga 49200 tggcgttgaa accagaacgc agttcttcac cctgtacacc acggtcttcg gagttaatgt 49260 tgtgcagacg acgcttacga ccaaagtggc tgtaaatgaa gccgttagtt tggatctggg 49320 catgagattt atcgatccat ttcttaagct gagggaatcg gccaaagtag gtttcgatgt 49380 actctttcgc atcaccagta gtacattcag tgtacggttt accggtcttc atgtgctctt 49440 ctaggagagc ttcgtttact gacgctgcta ctttagccgg gccagaaccg taaagaatac 49500 cgaatgaaat agcttttgca gcctgtcgta gtgcaggata cagtttttta acctcagtcg 49560 gtttacacgg cagagcaaat accatgtgtg caattgaacc gtggaagtct gagtagtttt 49620 ccgggtcgtt ctgcatgttg ataaacactt gctgcatgtt aatgtcacca gacagtacag 49680 cggcgtagta aatctcagca gttgttaagt cccatgcgat aatacgataa cctacagggg 49740 caacaataca acctttaata acagactcat cacgaggtaa ctgctgaagg ttcagcttac 49800 cagacgaact cagacgaccg gatgtagtca tatggatatg gaatccagta cgaatgcaac 49860 cgtcagcatc aatactgatc agcatcttct cgatatacgt cgagagcagc ttggagatct 49920 tacgaatctc tagcagagtc ttagctatcg gatgctggtc cgaaagctct ttcagggcat 49980 ctgcgccggt ggaatctgca cccgtatctg tcatgatacc cgtcggggtt aagccaacat 50040 aatcgaacag gagcttacga agctgtacaa cagaggctgc gttaaatact gagccctgat 50100 cttcctctag cttacgaact tctggatact catacagctt agcttttgca agctgtagcg 50160 cagtcattaa ctgaacctga gcctctttca gacgatcttt ggagatcggt acaccacggt 50220 cttccatacg ctgtaggaat acacatcccg gcatcagtac atcgtaatac agagacttca 50280 gtcgagggtt agcttcgact tttggcagga agaaattgga tagacgcagt gttgcatcgg 50340 tatccttcgc tgcgtaaggc cacatgatat caaacggaat gagatcatac gagaaatcct 50400 ctttcttgat cttatgcgtc ttgcaatacg tctccttgaa ctggtcgagt tcaaaatcgt 50460 agtcccccat atcggtatac ttcatcgcca aagatttcaa accgtgtgta ccacgtcgtt 50520 catctagggc gtagtgcatc agcatggtat cgtggaggcg tttctcttct gctgccttat 50580 caaaggatag acctaagtga tagcagtaga agtgcatatc aaacttcaag ttatggaaca 50640 caacgccatg ctccgggcta tcgaacagtt tctgtaagta atagacggta ttttctgtga 50700 tgacgtctgc atcaatatat acgccctgat actcttggtg ggagattgag atccccagca 50760 tgtaaccatc tcggcagtat agcgcactgg tttcggagtc atatgcgatc atgccaggac 50820 acatggtata caccatcttc acatacgctt cggcttcgtc cgggcactga atcgggcggt 50880 aatcccctgc tttagagcgt ttttcacgac cattgagaat ggcatgaatg ctctctactg 50940 tggcttcaaa tacaggcttc atttccggtt taaagtggag ctgggccgga ctgatacttg 51000 caatccagtt agcataccca tcatgctcaa cacgtttacc tgagtaatcg gaaatgccct 51060 ttttagctgc aaacttcaag aacgggtctg cacctaccag aataacgtaa tcaaaatctt 51120 ccggattgaa cgggttctcc ggtgttccga tagtgatatg tttcttgagc agtcgcccgg 51180 ttactttctc actagccatg aagaaggtct caacctcatg gtcaaagagt tcgaagtgtt 51240 tctgatagcg gacgttattc ggtgatttgt ctacaactgc gattttcatt gatttctcct 51300 attaggtaag caagtaaaac gtcagtgttt cttctcaaac ttacccaatt attatatcaa 51360 atcttgacat gttcagcaat caagatttct acgtttctga tgagttggtc gacttcttca 51420 gcatctaggt ctccagggtc tttgccctct ggaagcaaga agttacctat aattggtata 51480 attctagtct tatcacggat taatttggcc aacttcttag ccgcctcatt accggactta 51540 tcattgtcca atatgatgac tacgtgagta gtaccggcga tttggaacgg catgaattta 51600 tctgcaatgt tatctagtga gaactggtga gtaccaaaac aacatgaaac gttatggcaa 51660 cctttatctt ctagattcaa catgtcgaat ataccttcga ctaggattag ggctgggggtg 51720 ttatagcgaa cagggaagat tggggggag accttcttag gtctaatcat atatttcgga 51780 ggggcagagc tatccatatt acggcctaag aagaataagt ttctccctac ggcatccgta 51840 ataggaaaca ctacacgacc ttcccagtcc tctgtttgtt ggaatgcaaa gtatttcttg 51900 aaagtctctg ctttgatacc acgaaagtct tgatcgaata tcatggctga ttctggaatt 51960 tctagactgc gaccctcagt tcggaggtca ctaatcattc tgcgaactct caggagtcta 52020 ggcgacgtcc tgtactgttc ttcattgaag taatgaaaga tgcttggtat accacgacca 52080 aatccgcagc ttaggcagtg gaaaacccg gaatcggggt caacacgcaa actaggatga 52140 gcatcctcat gatcaggatt gagacagcgg atgaggatgt ccccacccgt atctctgtat 52200 tcaatccctt tgagatctaa aagctcggtt attctactca taagtctctt gaggattctc 52260 cggtgggatt agtatcctca tccgaagcct ttttcctttt tggtttgtct ttcggtttat 52320 cgctttcaag tgggataaca aactctgctt ccatctgacg aatgtcctct agcttaaggt 52380 cagaagtctg atccatacgc agtgagttcc agttgatctt cggcatgaat gtcataccct 52440 cggagttacg agtttttacg aagtcgaact tgattgcacc gtcatgcccc tcgttctgtt 52500 tagcagcatt taggtttgca gccatatcac aggaatcgag aatacctctt gccatacggg 52560 cgttaccctg ttggtcaatc tggtaagggg ctacaccagc aacgttatgc ttttggcacg 52620 tagacttgaa tgtagaggag acaaccatct gctgcttcca atcgtacata tctagtgtct 52680 tcgtatccgg caaacgtacc tggttgatgt agtctagcag ggcgattgtt accttatctc 52740 catacttgga ggttagctta gttagttcaa catcgatggt tgctgtacta agttccggat 52800 catacacaat gatcatcgga gtgtgcaggg gtctttcctg cattagcata gtctccatgt 52860 catggaaatc actcatctcg ttgagcgtat aacgtgatac aaaatcgtta tatacttcca 52920 gaccgccttc aaacattttt gcacgagtct tcgcaagttt gttgagttca ataccttgta 52980 aggagtcatt gcgcatagcc ttagcggaaa caccggccat aatggctagg ttacgtctaa 53040 agacctcatt ctctggcatt tcgatggaga aatatggggc gatatccccg ttgtaatatt 53100 gggctacttg aatgttagaa cagatgattg atttacctgt accacgccat ccaccgagta 53160 gcagtgtttc ttttctggca attccgccaa gttgagcatc aaattcgttt gagatgccga 53220 gagagataag atttaacatg gagtcttctt ttctcttgaa aacacgcata gtatcggcgt 53280 tgaagacctt accagtcgtt gttactcgtt cctctagctt catgtgcaga gccgctactc 53340 gatcaagtaa ttctccctga tccagtatgg taatatcctg caacacatca gtttctagca 53400 gattcaggaa tagatcctgt gtgtattccg actctaggac atgaatcgca tgcgatatgt 53460 caacgtctgg aatctgagtg tttgctaaaa cagttagtgc ttgagacaac cttgcgtttc 53520 tgttagactc tagcattaga gcatctaacg aaggcattcc gttatgcttt ttgtaataat 53580 tctggacagc ttggtagatc gcggtaaatg cgtcattgaa gtgatctttg cgcagtctag 53640 agaatgtctc tagggccacc tgcttttgat cggaggctag gagcattttc aaaactactg 53700 cttggacgtt atacataaga tactctctac gccgcgttag cgtctactta aacgaaaaag 53760 gggagaggaa caaaattcct ctcccctcag gtcaaatcaa ttacttggcg gatttagcgt 53820 caagtttagc gcgcttagca acaccatcgt gatccttagc agagatacca cggcggctca 53880 catagactt aacaccacgc tcggatttac cagttgcttc tgcgatctca caaacggtca 53940 tatcggcgat attcagacct tccagaacgt cttcgcgtgc cttagcatta gagacttcct 54000 ggaccggcat agcttcgata cggccttcac cgagcaggct cagagcttta ccacggatct 54060 gcttaacggt acgttcgaac tgagcagcga ttgcttcaac ggacgcgcca ctggcaacgg 54120 cattaatgaa agcagcttcc tgatccgggc taaagctacg tacagcagcg gccttttcag 54180 tcggcttaac agccgcagtc agttccaggc tcaggatttt accctgtact tgtttagcgg 54240 tataagcacc accaactaca gcagcagcga tttcagcgta ggtatactga cccggatgat 54300 cgttcaggaa tgcaaccagt tcatcttcct gacccggagt ccacggagat ttggttacgt 54360 cggatgcttt ctgtacttcg aagccttctt tacgcagttt agaaccaacc gaacgggcgg 54420 taacttcttt accagtttca gcagccagtt cagcagcgat gttagcaacg gcttcctgag 54480 agataacggt cgcgttcaga gcgttagctt tagcggtcag ggatgcagtg atttcttcgt 54540 tccaggttaa tttagccatt tttatttatt ctccagtaga attttaatcg tcaagacttc 54600 gatgcccagc gattgtgctt ttttgtatga agaagaaccg atcttggttt catcttcgca 54660 gattaagtag tttacagcct tagtcacgga ggttttcacc gtatacccaa gactttccag 54720 atatttggta gcgtccgcac gatttttgaa gtcgttgagc ttaccagtaa tgcaaagaac 54780 aactccgttg gattcagtcg ccggggtaac ttgagcttcc cgcgccccac tgaatttcca 54840 tggtagatta attacgttct gtcctgccgg tgtattcttc cacgaagcta agttttctcc 54900 agctttacct tccgctttca cagattggaa atcgttgtaa agctgggata atttcttagc 54960 agcaacttca ccgataagag ggatactaag agtccaaga accgaacca agtcaacatc ttgtctcagc ttagtttcca actctagctc aagttttcgtg gcgactttat cacctacagc 55080. tctgaccaa tcttctttag tcaggaaaaa gagttctga actttcgtga tttccagttt agcaatcgtc tgtgggccaa agccctttaa cttcatcttt agcagaagt tctcgattaa tttactgctt tgagctggac acatagactt gttacggcag aacaactgat cgttaactaa gtccagtttc gaaccacaag agtggcaatt cagtggaatt tcaatttca ttgattttct tcctcatca tttatataaa tattatagca agaaaatcag gactaagca tcgaaatttt tcagaatta gctcagtctc tcaccaaata tttctttccc tcaactgaat gataatagta 55500. 55500. 55500. 55500. 55500. 55500. 55500. 55500. 55500 55560. ttactcgtaa acacgttcta cgatgcgagg gatgacccca ccactacgaa ttacgcgaat ctggcaaccg atttctaggt caagggcagt gatatacca acgttattca gggtagcttt tgaaatgaca gcgtcgtcga tgactaccgg ggtgaagtat ccgacaggag 55680 taactttacc ggaagcccca acttgccatt ctacccgttc gagcgtagta acctcgcctt 55740 catcatcttc tttgatggcg tacgcaccac gcgggaactt gttagtccaa cctgctgcgt 55800 taaattggtt gttgccatcc atacgaacca ccacgccgtc cgttgggaac caatctactc 55860 gactggagat atccagacag gtaacaaaac catcaccctg gaggatatgc atatctccac 55920 agaaagtagc ggtcaggcca actttgccgg tttcacactg aatactatat gccgtgaaca 55980 tcaggccacc ttccgcaata cgggagagaa actctccaga atctttcagg ttaatagcac 56040 cagaggcgaa gttacgcata ttttcaactt ctttggtgat gtgtacttca ccagtaatct 56100 gcaccggagt cttttgggag atccgtttag ggatgttcag taacttcaca ttatgagtta 56160 catcattacc taagacgccg ttaccacggg ttaacgcaga aacgaacttt ccgtcaatat 56220 acagaagtga aatagcacag ccatcgagct tcggcgtttc aatgccctgg aagggtactt 56280 cttcgccgcg gcctgggtac actttctgta gggagaacat acggaatagg tgaggtacat 56340 cacctttagg gccaatttct tcctctagcg gaaaacgacg gattaaggca tcatactctt 56400 catcagagat aatcgacatg cctttgtagt acgcgtcttg gcacagctta ataaattctt 56460 taacgtgttg cattatttgg cctcgtagta atcgaacagc atctgagtct tcccttccca 56520 gtaaaagttt tcgcataccg cctggaaggt ggacattacc ctacgcttag attttggact 56580 gtacaattcg catactagta cgttcttctc ataccacatg cctagcatct ggaagcctga 56640 cattgtccgt actacaggtt tgcccttctc tttctctaac caggttagga ctagctcctg 56700 accctgaagt ggcgggctgg agtatgcgtt aagcgttatc ttttgcattc tttaatgcct 56760 cgttaacaat tctagacatg tgtgctctca gctgttgagt gtgagcatac gtcttagcag 56820 cttcggtcag cttattaact aattgagcga tttcatgatc tttcataaag atctccttta 56880 aactatgaga atattatatc aaggtttaaa acactaagca attgagattt taaagaaaaa 56940 gccagcaact tagtcgtcac tggcttgggt cttttgagat tcacggtaaa cttcatgaag 57000 aacctctgaa ttgcttagta ttttagtaaa tgcacggaat agcgtgctgg tcatttctaa 57060 cgtgtacggg aacgagaagc ctgatttagt agggaaccag ttatcttcaa tatctagtaa 57120 ccataatcgg aaaccaaagt ataaattgcc tctaaactcc gagacagtca tacgaatctg 57180 tcgcccttca tcttcaaaga taacgtggga ctgatcatca atatgtcctt cgtaatcctg 57240 acaaatgttt tgattcattt ttattgttct cagataaaca aaaagccccc gaaggggctg 57300 attgcgtctt aacgacctac tggagtagct cgatcaagtt cttggttgag gctggtgatg 57360 cgcttaatat tcgcaatcgg aatatagcgg aatttatcgc tagagcggct aaagaccagg 57420 atatgatctt tgtcagcggg ttccaggcct tcacggacga ttcgctcggc cagatattta 57480 tcttgcttag gatcgaactc agtcgtaccg tagaggtaag ttaccccctt ggtacgcagt 57540 ttggtgtatt gcatgcaaaa ctcaccgtgt ttctcacaga tagctacgac ttgtgcttta 57600 ttcattgcaa taccctctta tttagtgatt ttacggatag cttcagccag gtgagaggca 57660 gctttaccgg tcagtttgtc aacgatcgat tcgtccagga agccatcttc cagtccagcg 57720 tcgttaaatg cagcgcgag ttcagcctga gcatcagact tagaagtagg gccaccagct 57780 ttaggggcat caccaccagt tttagcggta gacttaccag cttctttctt gacgtatacg 57840 ccagctttgg acagcttcat acgaaggccg ttcgggggaaa caccgtgctc tttggcgatc 57900 tcagctacga tctccatgga aacgccagga cgctcatctt ccgggaagag ttcgattttg 57960 gcgacgtagg cagaagtcag ttcagcttcg agttcaggag accattgagt tggagtagac 58020 atatttatta cttccttat tattgttaaa aagagttttt aagttttgtt tcagactatg 58080 agaatattat aacaagttac tgagctgcaa agcaaacaga attttctagg atttggttaa 58140 cagctaccaa accattaggg gacaaacgat acatcttcag caaaccacga tcgtatttgg 58200 taagccagcc attatcttct gcttcgtcca tgggaagtgc atagtgcaga tcccgtag 58260 ctgtagcacc ctcggaatag atctgaaata agttcaggag ttgatattta ctcaccttta 58320 cctctctttt ctagtttcag ggctgctttt acagcgttat taaccagttc agctacgtct 58380 tctttacccc acttgaaccc cagggacttg atatcaacac caagtgctac cagatgttcg 58440 acagacgcca gatcccaata catgtagtta acatactgct ggcgggcttc ggacagcagg 58500 taaacacgat acgcgcctac cggattatcc agagctttt taacttcgcc aatacactga 58560 tagccaggaa cccaaacgtg ttcgcctact tcgaatacct ctttcattga ttcttctgga 58620 atcgcaggtg gaaagagcgg gtcaacctcg ccattaacac gaagcagagc accgtgagca 58680 gacagaacca ttctgatcat gtttgcggaa cggtagaaac gctcggcaat tgcctcaaaa 58740 ctatcacctg acaggtacga atcgataacg ttggccagtt cagccccttc gattttagta 58800 ccacgtttct tttacgcat ttcagcggaa acacgctggt tatctttcca ttcctcgata 58860 aggcgttcca tcgtggagtt agacgctacg ccgaggattt cgcaagcgcc tttcttggtt 58920 ttgccttcct ctagccactg aatcgcttcc ttaaactttt cgtccgggat gctattagca 58980 tgaagttttt tgcggattgc catgttcttt atcctctctc gaatttatga aactattata 59040 atacagattt agcttctaag caatcgcgat tttcatagaa cgtagggtcg aaagtaacgt 59100 gcatgaattc agatagactc ttgtacccca tgttaagttc catgttgatg cgtatagaaa 59160 ggcgtgtctt tcgctgagcc tctagccaat catagatatt tggcatctgt gctaagcgaa 59220 taggtattac tctacacaat tctatctctt tagctttccg tgctttcgta tcgcgtttgg 59280 gaaacttcct tgtttcccag cgggcatcac gcttgctcat tacgtactcc aagttcgttt 59340 tgcgcttcat cggccagcgc cttctcctcg tccgtcataa actcatacgg tacagagata 59400 ccctcgatag aacagtacgt gcgataccag tctacgatct gctccgtctc catttcctta 59460 ttaataccca tttgggttaa gtacatacgg gcatatttag ggttggcaga ctgtcccgtc 59520 ctaataaaga agtccttctt gtgtttcttc agggcttcaa taaaggggct aatcgtcacg 59580 ttagaacact tcttaatatc cttccagaat atatcttgaa gtttatggaa gaactccgcc 59640 cgcttgtgtg gtttcttact aatccacatc tccgcatgtt cctgtgtgat actcttgata 59700 gagtccaggt ggaagtactt gaccaacatt agcttggctt cgtatgcgtc cgcatgacgt 59760 ggtgcacacc aatccggggt actactcaaa atagcagaga ctcgttcagc actactctct gcggccgagc tatcaatctt accccagctt tcactgaaga tatcgatagt tgctccctga tccagcagat aataccgcag agtactgata aagttacggg actctagttc acgacgttta acatacagct gctgagctaa ttctagccaa ccctcatcga tgttatactg ctccgaaccc gcagagaatc cgctggtgga cttatcgatg aaatagaaga tattcttagc tccccggtcg cgcctgattg cctggaagcg catgtttggc gcctggttag cggtacgagt grandfather60120 acgttgtcga father gtcaacacca ctcgttactg acggactgca cagtagacaa tcaatctgct gttcaattaa ctcattcgtt gtataatcca agatacgacg aatatctata tcagatgtag agtttgagtg aatctcttta acaatcgcac ctgtattgtt tcttagcgtc aggcctttct cgtttaactc atccggccca after cgtttaactc tttctctccc 60360. atctctaggg acgtttggag ggcaacccaa atctggatt catcaggga ctcatacgcc tgagcatccg agagcatctt acgatggtgt father agcggctt gtcaaaatct atgagagatc catatgcctc aatggtttct gcgctgatat ccccgtcaga caggataacg acttttgcag atagcagaat ttcacgaaga acttttgcag actcacgacg ttgtttaaca actggagca ataacaggtc attcataact gcgtcgcact catcgataaa throwcatcg atctgcccaa caaagttttt aaacttgtgg atctgcccaa catacgatca atggcaccgc gcttgaagtt taacatatct acggacttat catactgtcc ggcttcaaac ttcttagcgt taggaac cagggcacgg gtgttagtaa ctgcaagga attacctttc agctgatcac gttctagcca cttcgtcact gcggtggttt tacccgtacc tagagaagcc ttgataaacg ttaaatgccc ttccggtggt ggacggttga tcttcaaata attcatcccg tctgggctat cagaggtcaa cttgtgtaca ggaacgcctt taacattcga ctcaggaata tctcgcatcg agttgttcac aaacgctttt aatgcctgct tacgaccgtt attgaagtaa tcctgaagac tgcgggaatt atccttcgtg ttgatataat cggacaaagc tgaggtaatt tctttctcaa gccaggcgaa atcaacaccg tcctccagag ctctgtggta cagtttagga 61200 ataatacgca gataaacacc atcctctgct tcttccagct ctgcaatggt ttgttccact 61260 ttctcagagg ctacgcgctt gcctttaatc tgattaagta gggatagaaa ctcttcttta 61320 aattccccac gagtggcttc ataatctggc atagtattgg gcagttcaat cctagcaccg 61380 ttaactttaa ccagacgtgg cttaccctcc gctttaaacg gatcggtaaa ctgatcccgg 61440 aaaatcgggt cagcgaaata gtgtagctgt acagatgaat aataagctaa gtcggcaata 61500 tcaaaaccat acttctgccg agagctatca ttgagggacg taaatagaaa ctttaactga 61560 ccctgagtaa cgggaatatt actctctaat agcatgtgca tacgaatccc cggcttaatt 61620 ccagccgaag acgacgcatg ggcaataaac cctgcgttaa gcgggaacat gtcctcgctg 61680 atgctattta ataacttaat gacatgacgt cccataccca caatgtcaaa cctgtcgcat 61740 cctccggtat cagagattcc atcgacgtcc atagcaataa tattgctctt atggtcgatt 61800 ttgaagttag tacgtttacg acgcacggcc ttctctgcaa ctaagcatgt acctcgtact 61860 gctacgagat gtggatcttg agtgagctga accatcagct catacgcttc attaagattt 61920 ttggggtcaa ctgtatccac tacgttaaat ttgtagggca ttgaggctgg cttaccttct 61980 ggatgctcgt tagaaaactc cttcgcaaaa aggtaatcag ttgtcttcac ttccttccag 62040 tttccggaag cgggatcgcg ggaaaaaccc gcgtgtcctt ctagaataga aaacattacg 62100 ttttctccta tgttgaagaa taaagcgatc tcgaactatc aaggatagaa cagcttaaaa 62160 gtccactgac tcaacaaatt tccttataga tggaggggtg gtcacttcct cattgctaga 62220 actggccgct tcgcacaact cccacacagc atgtaatcgc cgctaagctc catgtggttg 62280 acccctattc gtcgctattg cgcagaggta cgttgctact tacagcttca cttcttggtc 62340 atttcagacc gcggtgctat ttgctccaca tgactaacga acaaatatct gagagcctcg 62400 cgatgccaac ctctcttacc cgcattctag atcaattatc tataattccg ttcggaatca 62460 atttcccaat aagccaaacc caaccaagca tgtagcagcc ggactgttct gtatgctcgg 62520 aagtgaatga gaggaattat gtcagttttg taactcacat catttctctc aacctatgaa 62580 tatattatat aaaaatcgtg gcgagattgc aagtaaaata tttaaatacg tggtaggtaa 62640 ctgggtttaa gcatagaaaa gaccagagga taatcccctg gtctaagcgt tcaactaggc 62700 attccaaccc samaaattaa ttgagttaga agaggcattt tgtttgccct cacgaagaac 62760 ctctggagca tacgtccaag gctcatcagc taccaagaac cccggagaaa tacctgcccc 62820 cggttcccgg aatgctgcaa aattagaggg gtggaacaca acgttgttca ccagaaccat 62880 tagcccaga tccgagaggt catcccaatc aaagaggatc tcataatcct gggccaaact 62940 acctttggac aaaccaacac acacgtcatg ctggatgatg aagcaaagct cctcgtccct 63000 caacgcggct cgcagcagat tctctgctac atcaatcaat tccttgggga tatccgccca 63060 ggagatacgt tccatttag gaatactcat ttagttaacc cttcaacttg gaaaccgaag 63120 cgacgcagca cctggatcca accgttaata ttttccggag cataataatc ttcattacgt 63180 ccgacccaaa tacctgctga aacacctgta accccaaccg ggatatagga ctcacgcagc 63240 agacacacgt agcctagtgc aaacaccagc cccatctcag tcagatcgcc ccagtctagg 63300 acttcatctt gatcttcgtc cataggaacc catagcccca ggtgcatgtt gtcataatcg 63360 aagtagacct ccatattatc gcaccacaca cgaagatggt ccagcagaga ctcttttacg 63420 tccagaggaa ttcagccca gggaactagc ccaacccctt gattaacaac cttttcattg 63480 accagcatat tcttccccca tttggatgaa ttcgtgccac tgctcagagt ccgctcgtaa 63540 aacctgatag tcgtccgata ggtgactcat atcttgtaca tcactgcgat aacaccaact 63600 accgttgggc cataacagga tatcatcggg atggtacata aatttctcc tccagccaag 63660 cctcaacttg acctatgaaa tgtttgttgc taatacatt ggcagcagta tcataccaaa 63720 cggggatccc gcgttcttcg tacaaactct gctccaggta gttaataccg ttgttcacta 63780 cccacacctt agtacctcga ctatgggctg ggaatagttc ccaattaaag tcgattactt 63840 ggttactagc accaacaacg attaccagat cctgggttgt cagggtatca aacaggttga 63900 tctgccccat ataccacgga gcggtttccc cgaagaacgt aattgcaggc ttaacccact 63960 catacttacg gtaatcaacc gcagtgtacc cgatatcttc cacgatcaca tccttacctc 64020 ggcgatacat tatctccgta aggtagccat gagcatggat cacatcatca tggggaatgc 64080 cagcacgctc tagtagatca tccacgttag tagtaaagtt aacgacttga ccagggaagc 64140 gctcatacca ctcagcaatt ctcaagtgag caatgttagg ctccaccgta cccagctcct 64200 gtcgccgcat gttgtagaac tcatgggtta gctcgtacag gttgcgacta ggctcatcat 64260 cagatccctc agaaagggct tctaacgggg gcatagggtc gttcagatag tgaaacccct 64320 tatcgaacgc ctgaatattg catacttcgg ttagtttgta tttatcccac aggggagtga 64380 tggcaccctt acggaacgtt ggcactcaat ccagcaccac ttacgataat caatctacgc 64440 attacaggaa ctccatatag tttctcattt cttctttccg caactgctca gccttttcgg 64500 acaagcactc agaggttctg ggatcgactt tatctttcca caagtggtta cggcatcgca 64560 gaagttttac cacgtcttct cgggaaatag atccgtatgc tccagagtcc atcataaagt 64620 agatacgctt ttcatcagtt ttatggattt tcataaatcc tccgtttctc aattacaact 64680 atcttatcaa aaacggaggt atttagcaag tgatttattt caaattcatg tgtttgaccc 64740 tttgcattac ttcctgttgc ttaccagggt taaagggtct actaccagga ctacctaagt 64800 acccacaaac gcgacgggta accgagatat tccccgaccc acattgagga catacaaagc 64860 cgtgctctcc ggatacagac tctcctagat atccgcactc ttcacattca tcaactggaa 64920 tgttaacccc tatgtagtga gatttactta atccataatc aacaacccac tctagtgcag 64980 ggataaactt acgcatctcc ggaagttcca caaaggatat attaccccca ttggcgatgg 65040 tggtaaaatt agcctcatag tcaaacttca cgttaggggc aacttttgta cggacatcca 65100 ggtggtggct attagtcagg tatcccttat ccgttagcca gtcgtactca gggtactgtt 65160 cggcaatttt agtattgaac ctattgcaca gagactcact aggggtagca taaaggctga 65220 atccaagatt agttcctca gctttttgtgt tacaacggtc tttcatgtgg ttgagaacct 65280 gtacagcaaa ttcaatcgca ggtggggaca tggggtcggt atcctcaaac attacttcaa 65340 ccagctcgtt aataccgata tatcccaaag acactgatgc cctgccttca aagatgggcc 65400 atacaagatc attggctttc aaccgtaccc caaaagctcc gtgcatatag agaattggtg 65460 cctgtttagc acgcacacgc ttcaaccgct caatagccca atcgtgggca gccatcgctt 65520 tatcaatgta ttcgtccaac aacttccaga acctatcgaa gtatccctca gactctacag 65580 caactagcgg caggttaatg gatacaaccc ctaggttatt gcgccccgca gtctccccgg 65640 actcaatagc tgaaaggaaa gaccgacaac ccatagggaa cttaaaatca cctgtaacag 65700 ccgttactct ttcataactc acgtaatccg ggtacatcct cttagaagtg caggtgagcg 65760 ccagctgttt gatatcgtag ttgggatcgc tgggagattt attaagcccc tctttaactg 65820 cgaaaataag tttaggaaat acagcagtat gccctgaagc acccaggcct cttatacgaa 65880 cttccagcat agctttctgc aacatgcgtg cctcccagga ttctcctaga ccaaacccaa 65940 atgtcacaaa tggctgctga ccattagagt tgaacaacgt attaacttcg tactctagcc 66000 cctggcacgc atcatatact tctttttctg tcatttcagt agcatacact gctgccttct 66060 tactatcgtg aagccaacgc tggccaatag ctaagtgctt atcataggac ttacggacat 66120 acggggcaaa gacctcgtca atacgatcga tagacgtacc cccatactga cacgagctta 66180 cctgggcaat aacttgggca gttactgcag ctgcagtaga gatagattttt ggagtttcaa 66240 tctctgctcc accaatttta gttccattct taaacatccc cgctaaatct accaggcagc 66300 agttagtata cccctgcgct cggtagtcca tatcgtgaat gtgtatctct cctctattat 66360 gagccgccaa taggtatgct ggaagctcct gtgctaccac atacttagag acttccccag 66420 ctatcatgtc cctctgtgta gggaattgct cgcttgcctt attggcatta ttaaacataa 66480 ggtcttcttc accctcgtta tcaaggatac tatagatatt atcaatcagg tctaattcgt 66540 tcatatattc ctccagttag gcatatatta taccaaatag acacaactat gtcaacctta 66600 ttttattata atcaacagta acttgatctg gatcatcaaa acaataaaaa ctggaaaatt 66660 ttatttgctt attcacccta ttttatatta ttcgtgtgcg cccgcgcatc gcgcgaataa 66720 aggaaaggat ccgtcctcca tatcatccta gaaaatttca gttgcaatcc acgccaaaac 66780 ttggtatact ggtattctaa attgataaac atggtcgaaa tttgaccctc tgtagacctc 66840 taggagatc gcaatgagaa aagcagcacg tcgtaaggag tcgcgccgta acggtagcgc 66900 aaaacgtgaa cgtcacgaaa acgtcatccc cgttgatttt gaagcacggg aacgttttca 66960 accaaccgca aaagaactca agccgaagaa cgctgagcaa aagcactata ttagcaccat 67020 ccgtaacttt acggtgacag ttggtattgg agaagcaggt acgggtaaaa cgtttatccc 67080 atctgttctc gctgctcaag agttagcaac gcctggttct gtgtacgaga agttcatcct 67140 agtccgtccg aacgaacctc ttggtaaatc cctagggatg ttgccaggag atctgaacga 67200 gaagatggca ccgtggttag aaccaatcgc tgacggcttc aagtgggcct taggggaacg 67260 ttcttatcag gggctggttg aacgcaaggc aatccaatac cttgctatcg aacatgcacg 67320 cggtcgtacc ttcaacaatt cttatgtaat tgttgacgaa gctcaaaata tctccgtaga 67380 agcaatgaag tgtattctaa cccgcgtagg tcaggattgc aaattagtaa tctgtggcga 67440 cgtagcgcag aaagacatta aatccgactc tggtctgcaa ctgatcatgg atatctacga 67500 tcagtacgaa catgtgcctt tctccttagt agagttgcat gataacgtgc gttccgctga 67560 atccaaagca ttccaggcaa tctttaacga tatgggaatc taatatggtt actgaactga 67620 tcattggtta tggtgagggt attacctctg aagaaaactg gggctttgta ggcttcggtg 67680 aaggtatcac ttctcacgac gaacgtcctg acctctaact ctaggagcgc gtatgaacaa 67740 cgtatttaca ctgaacaact tccgcactcg taaaacgaaa gtgcatccag tatccctggc 67800 aacagtcaat aaatacaatg ctaactatcc tgaggatgag cggcgacacc atgccgcttt 67860 caagatagca aacgaatttc ctaaccaacc cctcggtact aaagagttag ttagtcgaat 67920 gaaaaaatta cacttttatt aaagtagacc gacacacagg cagcctcgaa aggggttgcc 67980 tgttttgtc tgtaaaatct catttgcaaa atggccgaaa acaaagtaga atatctttta 68040 aatcctgaca acttccaaaa ggagttacca atgacccaac gtattgaata cgttatcaag 68100 cgtgacggta caaaagaacc gttcatggcc cagaagctga atgactgggc aaaatacatc 68160 ggaatccgaa gcgatgttcc gtggtctccg gtagcggtag ccgcagttaa gaacctgccg 68220 aaaggtgatg tacactcaga cgatctgcaa acaatgctga ttaagtctgc cgaatcaatg 68280 atcgagcgtg atcatcgtta cgaccgattc gccctagagc tgcgtttagc ccagctccgt 68340 aagaacctgt ttgattctta caccccgccg tctctgcgtt tcttccacga ccacatggta 68400 gagctgggag cgtgggaaga tatgagcggc tggatctctg acgaccagtt cgaagctcta 68460 aacgaagtta tcgaccacag ccgtgacgaa ctattcacta atgctggtct gaagcagttc 68520 atggataaat actctagacg taatatctat acggaagaga tctatgaaac tccgcagttt 68580 gcctacatgg gtatggcaat ggcaatgctt tctcaacctc actggagtat gctcgatgca 68640 atcgaccttt ataacgcgct gtcgctacag aaaatcaacg tcccaactcc gccgctcgtg 68700 ggccttcgga gtgctgacag gggcttcgct tcttgctgtc ttgttgatgg tactgacacc 68760 ctggactcca ttgacgcagc agagcacgtc gtcttcaaaa tggtcgctgc ccgagcaggg 68820 atcggctatc atcttgagag tagatcaatt gctgatccgg tgcgaaaagg cgcgttccca 68880 cactctggaa aactgccata ttatcgacac atcgaccgct ccgtaaaagc taacactcag 68940 cagtcccgtg gcggttctgc aaccgtttac actgcgttct tcgatccgga aatcattcag 69000 gtaatggaag ctaagagcaa ccgttcccct gatgaaaaga aaatcgataa gatggactac 69060 aacctgaagt ttaacagtat tctcctgaaa cgcttcctgc gtaaagagaa catcacactg 69120 atgagcttcc tgtacgctcc agaagtatac gcagcattcg actctggtga tgtagcagaa 69180 tttgaacgcc tgtacatcgc agccgaaaaa cgtctagcgg gcgttaccaa acgtggtttc 69240 aagggtgaag ttctccctgt agccccagtg attcctgccg cagaactgat tgagttctgg 69300 aaaactgtac gtatggaaac tggccgcctg tacactatgg acgctggtga agtgaaccgc 69360 aacagccgtt acaaagatcc ggtgcgtatg tccaaccttt gtgttgagat tgtacagcca 69420 acgttcccga tccctcacgt acgttccg acgaagaact ggataaaatg gatgtttcag agtatgggga agtatctctg tgcaacttag gcgggttcgc cctaggtcgt 69540 atcaaaactc tggaagagtg ggagaagatc tcttacatcc tgttgaagtt cgttgatacc atcatcgaga ttcagcacta tccgttccca gctatgaagt acacggcatt acgccgccgt aacgtaggta ttgggctgat gaacgccgca ggagcgatgg ccgctgaagg cctggcattc gaggcgag aagcccgtaa ctggattcac cgtgaagcag agaaggcatc attcttcctt 69780 cataaagcgt ctgtacgtct ggcaaaagag atcggcccat gtgaatggtt ccatcgtact cacacttctg atggtacact gttaatcgat acttacaaga agactgtgga tgacctggtt 69900 tccgtaggtc tggaatgga ctggggagagc ctccgtgaag agatcaaaac tcacggtatg cgcaactctg ttctgacggc aagtatgccg ggagagtcca gctctgttct gattggtgtt accaacgcag tagagcctcc tcgcagtgcg gtaactatca agacttcggg cgttaacaaa gtaattactg tagctccggg tctagacgat tgggacacca tgcagtccta tagtatgcg ttcgatattg accgtactga gcacattaag tggttagcag ttctgcagaa gttcacggat 70200 caggctatta gtgccaacct gtactacgac tttaacaaat atcctggagg tattattccg 70260 ggtactgaga ttatcaaaga tctccttaac tccactaaat atgggattaa gaacctctac 70320 tacgcaaact ttgacgtaga taccggaggc tctgcagcag agcagggttg ttccagtggc 70380 ggttgtactc tgtaattaag ctcgatccaa ccgaaagcaa aattttattt gcaagtaggt 70440 tggatttttt gttataatat tcgtatatta aatgagagag gaaattaaac aatggcaaca 70500 gtatttaacc gcgaatggga tcacactgag tctaaactat tcctgggcca agacctggga 70560 atcgccgact atgtaaacgt tcgctatcct cgtttagagg aacttgcatt actgcaacgc 70620 tcgcagttct gggttgagac tgagatctcc ctggaagccg ataagaaaca atggcctaat 70680 ctgccacaac atattaaaaa caaaaccctg ctgaacttag cctggcaaac ccaagccgac 70740 tccattatca cccgtgctcc ggaagatgcc attctgaaac tggtatctcg ccctgaactg 70800 gaaggtatgc ttattcagtg gagctacttt gagaatattc atagccgtgc gtattccaac 70860 attattcgta acgttctccc taatccgggt gagttcattg caaccgtaca ggctaacgac 70920 gaagcattcg cacgtctagc actgccggtt tctgttattg atgagttggc cgaaattgcc 70980 gacatttggc tggacgctag agccaatctg gaaatcgctg agaaggaagg tactctggaa 71040 tacaccgaag aggcagactt cttagcactt actgaacaag tacagcagaa gattctggag 71100 ttctactacg ctgtatacgc tctagaagcc attatgttct acgcgtcgtt tgcgtgcacc 71160 tttgctctgg cagaaaacga tatcctaacc ggtattgcaa agaacttaca gctgatcgct 71220 aaagatgagg cactccatac cgttatggct atggaagttc tccgtattct tcaaaatggg 71280 gaaattcccc cacatgtagt ggcagcagct caggctaacg ccccgaagat cctgcgcagt 71340 attctggaga cagaaattaa ctgggcgcat tatattttcc cggaaggcga agatattcca 71400 ggcctgaacg ccgatttgct ggtagagtac ctgtactaca acgcacgcct ggcatttatg 71460 gcgatcaaca ttccgtggcc ggaagatctc ccggtaatta tggaagaccc gattggctgg 71520 atgaaaggtt ggctgaacac caagaaccag caagtagctc cgcaagaagc acagattacc 71580 aactatcgtg ttggtgcaac ctctcaggct aacccggacg atctgtctga tgaatttgga 71640 gagttcctgt aatgattaca gctttatacg caatgcgagt tgacgccgcc ttcgggattt 71700 ttaatccggc tacaatggat gcctacgggg aacttccctg gggctccatt cccgaggagc 71760 tagaacagtt ctacagaatc ctggatacct atcaggttgt tatagtgggc cataatacct 71820 acgaaacagc ccctccacga ctgaaaaagg cactggagaa gaaatccatg gtgtacgtag 71880 taggttctaa ggctccagtt ctgataaaaa accctccccg taatgtgcgg tttattaccc 71940 acttgggttc caaaattcgg gacttctgta atgaagttga ggtagtctgc ataggaggga 72000 aagctctgct agaaacccta gcaactatgg gctgtctgga tgcgatttat cgctccacca 72060 tttatcctaa ggccggtaca gtaccaagcc tagaccacat catgtatctg gaacacccga 72120 tacttacgtc cactcctccg gatgctgtag taactcatat agcttctggg gaaaatgagc 72180 gctatcgttt tgttatggaa ggagtctatc tgtgattcac tatattaatg aaggcaaacg 72240 cattcttgaa gaaggtgtat ggctggagaa ccctcgaacg ggggttcgtt gtttaaccgt 72300 aatagggtca aactttgaat atgatgtact cggaaagaaa ttcccactta tcacaactcg 72360 taaagcgtat gctcttcagg ccatcatgga gcttatcgga tatctgcgag gatatgattc 72420 cgcagagcaa ttccgtgcta tcggctgcaa tacgtggaac gctaacgcca acgaaaacga 72480 ggcgtggttg gtcaacccta acagaaaggg aaccgacgat atgggtagag tctatggagt 72540 acagggcaga acgtggcttc gaccggatgg ttccccacttt gaccaactct ataagatcta 72600 tgagaattta cgacgaggca ttgatgaccg cggagagatt ctcaccttct ggaaccctgg 72660 cgaatttgat caaggatgct tacgaccgtg catgcatacc caccagttct cgctgctcaa 72720 tggaaatctg tatctcgact ccttccagag atccaatgac ttcctattgg gacaagcctt 72780 taacatggtc caatgctaca cgtttcttgc gcttatggcc cagatcacag ggaaccgggc 72840 aatcagagca aaccaacgta tagtaaatat gcacatctac gagaatcagt acaaggttct 72900 catggaacat gggcaatttg accgcaaacc ttcccggct cctcgtctag agatcaaccc 72960 ggagataaaa accctagagg acgtactcac ctgggttttct aaggacgact ttaaaatcgt 73020 ggggtataag tctcatgacc ctattgcata tccgttcact gcgtgaggac ttatgggatt 73080 attack cgccccaaaa taacttctc cgaagggag gagtctcagc tgaagttctct 73140 ggttcaatcc tcgggattac acatagatgt tatcctaggg atggtaagt acagggcat 73200 ggacgcacta atgcgtcagt ttgcccctaa accacctaaa gaaaaccctc cagccaagcg 73260 ggactataac agtaatttac tggttcctcc agctaatta ctataactaa ggtggccttc 73320 gggccactta tattaaaaat tcatttgcaa accacgtgaa tttctaatat atagattca 73380 taaatttgag agaggaata aatgtctt ttactgatgc aaagcaatg gcagctaag 73440 ccaaaaggtc taacgatatg gcagttatag cagctcgcag atctata tcaatattg 73500 atggatcagc ttctcaggt aaaacagaag tagattctta cgcattgaat ggcctgccta 73560 tcgccgcacg gtctcagatt atggaagacc taaaggacgc gggctatgaa gtaaaagtaa 73620 atcatccgtt tgaccaacgc gatactgaat caattaccat ttcctgggg cacgcataat 73680 gtttcatgtt tatactgatg gtggttgccg cggtaatacc cgtggggtag acaacgttgg 73740 tgcttgggct atggtagttt acaactccag cgaagagcaa atcggcacca agtctgctcc 73800 caaacgcaac acaactaaca acgagatgga gcttcaagca gtcctagagg ctctgctatg 73860 gtctaacaaa aaccctggcc gaccaatgac aatctatctg gactcaacct acgttaagaa 73920 tggttgtgag tcctgggttt ggggctggga acgtaaaggc tggagaagg cagatggtga 73980 tacacctctg aacttagacc agtggaagtg gattatcgac gaactcaaga agtatcgtct 74040 aaaccacaat gagatcccaa ctttcgttaa ggttaagggc cactctggtg tagaaggtaa 74100 tgaggccgca gataacctgc taaatgttcg aatgaccgaa ctggaaatgg aggatatgtg 74160 atgttggaga atttacgccg cctagttagt gaaatgaaat atgaagtgct cctgatggag 74220 cctggtgtgg atcgagtagt aatgaaattg cgaatcgcac gtatggaagc ccaaatcttt 74280 gaagcggagt ggaagctct gaggggtgga gatgaattat aatggcacca gatttgagggg 74340 atttgttccc aaacgtacca cagtaccagc tcgatctcta cgccgcgtttt ctggaagcgt 74400 caaatcggg gaaccctcta cgcgtctacc gccagaccg caggcacggg aaatcctgga 74460 tcctgagatg gctgaaagg aacgagccgc tcttaagaa attgagcgaa agaactctg 74520 tacagcaccg gcatacaca aaggtgggta ccagtacatc agctcagaag agcaggcaaa 74580 acatacggg cggtaacaga tacgagttca tcatcttcga tgatcttgta gatgaaaatg 74640 aaaaaacgca gttgctaaac gctaaaaatt aacgtataat agattcataa attackaa 74700 ccaatcaaaa aaggaat aaatcatggc taagcagacc tctaaaaag cagtagaaac 74760 caaagttgca acttttccca aaactgaagc gatcgcaa gcacgtctgg agcgtcatct 74820 gcgtaaacat ccggctgata ctcaagcagc atcagctgta ggcaaccgg ccccgcgtcg 74880 taagaaaccg gtaactagg gtttacctc cggctacgtc tccagatcg tcggttggag 74940 tacgccggat aaggcagaca ccaaagaagt cctgcgcaag acccagggcc gtttcggtag 75000 cgtcaagccg aatatcttcg gttgcgaata cagccgtgag aatgttcgcg ctctgtgcta 75060 cggcgtaggt attaaattta cgggcaaggc aataagccc cgtaatcaaa aacgcaaacc 75120 75180 cataagtcgg ccctagatta ttctagggtc aactcccggg aactgatgga ttcgtgctac 75240 gaagaactcg aagattgggc ggctgactgg ccgtcgatgg aagaaaactg ggatgtgtcc 75300 gaagatatga ctaagccacc accggaggtg gcttcttaaat gtgataacac atctagaagc 75360 aactttaata aacaaaa aatatgcaa aacttacaag accgctggat ctctgtttgt 75420 gatatcgaat ccctgggaac tccgggagat tgcaagagca cctttatcgc tatgccgttt 75480 ttcgctttcg tactgatgaa agatctatcg ttagatccgt atatcgttct aggcactcct 75540 aacgttgccc agcagttggc cctcggtgct aaagtctccg ccggaactat tgcattctgg 75600 atgaatgagg ctcgtgctgg tagtgctccg tcgctgtcca ttattgaagc tctgaacgct 75660 aaagacggtg agtctactgt tctgtctgc aatccgactc atgaatcacc gtatccaag 75720 catacgttca tggatctgat ctgcccgttc gtagaggcaa aacaggtaat cgaaggcatt 75780 atcgatgagc aaggtatcga cactcgttcc ctgcgccact acggtaatgg cccgcagttc 75840 gatatgtcta tttacgaac agtggcggct caggctaatg tctctctcc gtctgatcct 75900 gcaatcgttc cgtggaaatt ctgggatatc tccagcgccc gtaacccgcg tgactatttc 75960 gaggctctcg gaggagactg gaagcattg gtacgttgtg ctgaaatcta cgcacacgac 76020 gtaatcgagc gttacaacct gattcctgag ggggtctatc cgtcgaaaca tgatccggtg 76080 tttgacgccc tggtagaagc gtactgtatt aaaactacg aatcgaaatt gaaatttga 76140 tttgactaaa tggctgattt cttgatatac tattattct gatcaaga gtaaggaga 76200 aaatgaaa gcagccattc taatgatctc catcctcact agcttccatg cgcaggcaaa 76260 gattgatgcg catgagatag agtgcatagc taaaaatgcg tactttgaag ctagagggga 76320 aggagttaaa ggaatgaccg cagttgcaca agtaacgaaa atcgtgtta actatggaaa 76380 attcccgtcc acatattgta aagtagtcta tcaccaggg cagttcagct gggttggtaa 76440 gagaaacat aactcgatc gtaagacga agagtggaa caagctaag agatagctag 76500 actagtttac tatatggatc ttccagtaga tccgacgaag ggagctttgt actttcacag 76560 window aaaccttact ggaaaaaga aaggactc aaaagaaaa gtaaaattgg 76620 CAccatgtg ttcttaaat taaaatctca gttgcctaat gcttaaaag ttgatataat 76680 agtttcataa attagagaga acagcgggcc gattacgctg aacttgtttt aatgtaagac 76740 tcgtcgagca ttaagattcg ttgaatagaa tcggtgagtt aagagtgatc acctgacaaa 76800 gttcgtgcat ccagtccgga taccggccag ggagcgtatt ggacgggcgt aggttagaga 76860 tctcttgtaa aactcgggag ctgtccccgga gtctcgtaaa ctctctccct gatttataga 76920 aaatggtgga atctcacttt aacctgttcg accagaatga agcatgtgc gtggcctgcc 76980 ggagaggtgg ggagctgagc ctgagaaact cgtatagctc ccacactgac ctgcctaggc 77040 cgcaaattgg agacatgtgt atagcgtctg gtgctagtta ttgccaccat agtggggtgt 77100 cgaaatcact tatgtagcaa cactgccgtc ctaagttaat tggtagactt ctgcacgcgg 77160 gggagctatc cccagatgcg gaaagtgcag gttcgaatcc tgcggtcgga aagagttaga 77220 taactgatgg ttctattggc gaaccactcg ctgctggagt acctcctccc ctcccaggga 77280 aagacttggg agactcgcgg gaacgaaaca gctaaaccac gaaggactag gatgctgatt 77340 tggctaccct tacctgatat atgcccaaat caaaagaaga actttaggcg gtagcctaga 77400 tagtgaagtt tttcctccac gaggagtcgt ttttcaagct atcaaaactt cacatcgaaa 77460 attcggaggt gtgattttaa cagaccgcgt ggaggatggc tctccagaag tgccaaagtt 77520 agtccttaaa tacggcatga tgatcgtatg aactttggcg aggtgggatc tcgcctgatt 77580 ctaagtggga aagagtattc ggatagccgt tcgtacaaac ggtggctccg agacctaagt 77640 gggacacggt tgagtataca gctccgcgga agctcgtata ccctgtgtaa gcatgtgaca 77700 atggagtcat gaccggagtc accccgagcg aaagcataac cactatttat agcaccctta 77760 cacaggctag aagtgcccct gtttggagtg tgccctcgcc tagaccgcat tagccgacgg 77820 atatccgttg gccgggatct ggcaaggtat attagtcacc ttgactataa ctggaagata 77880 cgtctgacag aagggccgca tagcttaaac aggggaaggc ccctacctac tggcggtatc 77940 tttaagagct ttcacgagag ttcttaaaga tactaattga cgaggtctat atgaacgatt 78000 tagtatgtt aactaagata cgctctgata ttgagtctat ggtctctcgt cgtagtgagc 78060 tgactaaggc taaacagatc atcagcggtg gtacacagaa acgcttcaca ttgcaagccg 78120 gggatattaa gtttgaccta tgtggcagcc agactcgaga ttacactttc gaaatgaaac 78180 cgtgttacga tatggtgaag ctggggctta tcaaagctct agacaaacaa atagatcagt 78240 gtacggacgc aatcaaaacc ctaaacgtcc agttcgccgc tgaatgcgat cgtctcaaaa 78300 actctatcaa ggtataataa tggtggcatc tgtgcatact cctccgtatg aacgtccagc 78360 acctaatctg acacctgaac agaaacagct aatcgccagg cgcactctag agtttaaaga 78420 gtcgctgcat aagagcgttg gccggtattc tgaacaggtt catgatctgg ttgtcaaac 78480 acttaaactt tattaatgct tccttagctc agaggataga gcaacggtct tctaaaccgt 78540 gggtcacagg ttcgaatcct gtaggta ccactttaat cgctgcccac acgtaatatt 78600 cagtgggcag cggcagaaag cccgcaggggc gcagattaag attgattcta caaattttt 78660 cttgcaattt tgtcagaaat ttgtataat agttctttaa atcaaagaga aagtagagga 78720 taaaatggct tacaaattg atattaa gaaaggggtt ctcactgagc tggttatcga 78780 tgcgaacatg gctcgcaacg agggcactaa atccgtattc tataaagacg gtagcgtagc 78840 tcgtatgatc ataccgaag acatcagga tctgtacgtg atctccgacg aagaagcagg 78900 tttcgtaaaa gatcctgagc cggctgaaga tactccaact gagacactc cggtagcgga 78960 taccactact gagaacctc cggtggaagg cactccggaa gatgagcag cagtataat 79020 aactaagggc tagaatagc ccttctcctt gggccggtag cttaaagtta aagcagtggc 79080 ctcataagcc aacgagtggg agttagagtc tccctggcc caccaatttc atacaggag 79140 actaaaatgt ctactaaaaa cgcaatcgta tccttcgtgg atgacagtgg tatcgtccta 79200 gagtccacag taacagat cagtcctaaa cgtctgctcc atcgtgg ggatatccctg 79260 gaaatcctta ataacggtc tgagactatg ctggtcattc ctgtaaatcg tcttctctcg 79320 attaaaatcg tgtgggagga ctaatggacg ctcaattaca aacccagtac tatatgcttt 79380 taggcatgtt agaggatgct ggcccgacag tcagagggca ctatgagcgt cacaagcag 79440 cttttgaagc cctactaaaa gaagttaatg agaacgaagg tggtaaggc tctgactcct 79500 atgccgcatt cattatcgcg ctacaagtttt tcctaatca tcaaccaag tataggcta 79560 atatgctga atattaacg taaaggtttt ttctacaagt ggcttaattt ctcttccgcg 79620 tctttcacct accggctgaa cgacaacaga gtcaccttgt gtagcttt ttggcactca 79680 gtgtgtatt tcctgctaca gattgcgta actgctattg ctgtactctt ctccttgggg 79740 atgggaagta tcttatctac gttctttggtt ctcaccttg agctgggaat tactccctgg 79800 tacatgcttg tagggttaag tctagcagga ctatctacga taatagcgat tctgctggcg 79860 atagctggga tcggctgggc ctgcgctaag ataggggatc ggatccagga atggaacgcg 79920 agtaaatcct ttgaaagggc acagaaagag tataatgctc gcgatgaaga gctccgcttc 79980 ggtaatatct accagaagat gcgaatctat aagaaggaca aactttgtcc gctcattcgt 80040 gtagaccacg gcgagtagtt tgcatagtga cctgcgttgc cgggtcacta ccaaaatact 80100 tgaaaaattc aattgctaaa tgctctagaa tttgagataa tatttatatt gaaagggaat 80160 agccaagtgg ttacggcatc ggcctttgac tccgagatcg gtaggttcaa ctcctcttc 80220 ccttgccaaa tttaaaatgc tcctgtcgtc taagctggtt aggacaccac tctttcacag 80280 tgggaacacg ggttcgaacc ccgttgggag taccaaattt actgaaaaat tcagttgcta 80340 aatgcttaac aaattgatat aatatttata tagtttgtta aggaattaat ctgatactga 80400 gccggaacca aaaaacacgc ttgcagcttg ctcggtcatt gtactgaggg ttcgcaagaa 80460 tagcggagtg gctctccgaa acgcctggta agcgtaggt gtatcgtatt tgtgaagagc 80520 gggtatcctc gcatggctaa ttgcttaggg tattacgcgc accggattaa ttgccttaac aaattattgg gggatgggtc tgctggggagt ggacaccgca cttgcaatgc gggaatcaga acggttcaaa tccgttatcc tccaccaaac aaatggggat gtggcgaat tggcagccgc gctagattta ggttctagtg gtgaaatatc cgtgtgggtt cgaccccctc catccctacc 80760 aaataacggg agttcgtgat gaacatagaa atcatgcagt tggatcgtaa gaaaaatgag ttccgtaagg tccatacctt tccgagtaaa gaagctctag agtttcatat tagtgtatg ggactggtgc taccggaag cgagatcttc gacttagcat gtgctaacgg agttctgtac gtctgggaa tcacgtatca tgctgatcct gatgaactcc gtaaagaagt agagcaata ctaactggag agtagtgctc atgggagcaa gctgacttga aatcagtcgc catcggaaac 81120. ggtgagggtt cgattccttt attctccgcc aaacaacagg aaagctggtg aaatggtagc cacgcatcac tgctaatgat gagtccgcaa gggcatgaag gttcaagtcc ttcgctttcc gccaaacaat ggcccgacga gtaacggttg cccctttcta gagaattttc tgggcgaaaa 81240 tccagaagta atcaactagt tgtaagtggg aagagtttct tagggccgcc aaatttgagt 81300 atcattggta ttgtaggagt cgtacgctgg gactacgtag tacactgagt gtcctatgcc 81360 cagtacgcgg tctaaccaaa tacggtgtgc gatgtgcatt ttatttgtgg aggtctgatc 81420 agccctcgat ggtactcaaa tttgggatat tatcataact ggataatgac ctcgattgtg 81480 gatcgagtct atcttggttc gaatccaaga tatccctcca gattactgca ccattagttt 81540 aatggataga atatagagct acgaactcta tggttgaggt tcgattcctc gatggtgtac 81600 cataaaatct ctgctgagag tgacgaggtt agccctctgg gtcacatccc tcttctagcc 81660 gcctcccata aatgcaagca tttatgttct ggccctagac acaacgttgt actcagcaga 81720 gtgcgcataa ttggggtata gctcagtagg tagagcggag gtctctgaag ccttaggtca 81780 caagttcgat tcttgttgcc cctgccaatt gcaccgtaga ggagaggccg tcctcgccag 81840 tctcataagc tggagatcgc aagttcgat cttgccggag catccaattc taggaaga 81960. tgatgagaat ctctttcaca gaacgagtac taggtactgg agtaatgcta atcacttcct gggatggaga tagctggtgt aacgtgacag gcttacgtaa gtcagaacaa acacccgaga atatcgctaa aatcaagaaa cgaatggcag aagctgctag tcgtcctgga gcacctcgta atggtaaacg ttgaggtaac tatgactcgc tatcaaggta tgctaattaa tacccacaca aaagagattg tattcttggc accggctttt cacgacacct acaatgaagc cgaggagac gctaggatcg ctaaataca cccggacgag gaaatctgcg tccgtcagca agaacaataa tgcttcaata gctcagttgg taggcaaac gaccgataat cgttaggtca ctggttcgag tccagttcgg agtaccaatt tttgccccct tccagttcggat atcctccagc gatggtacgt aggravate ttacctgtta agcctagaca aagttacttt gtgggcaaaa 82440 tag tagtag tagtagtagt aagattcttg aggcaaattc tttaggtcag 82500 ttggcagaga tggtttatgc actcgcttca tacgtgagac tacagtggtt cgagtccact 82560 attgaccacc aaatacttgc ttagctcaat cgggagagca tcgtctttac acggcgaggg 82620 tagctggttc gaaaccagca gcaagtacca atttacagtg agacttcggt cagggccgtc 82680 aaacgtccga gaagctgtaa tttctgatgg ggatgatccc agattcgtaa gaatacccca 82740 tccttacctt cacaggtttt agcgcaatgc tgagtaaaag tctagggaat cgactccatg 82800 ccatccagga gtataaatgg cgaatctagg gacttagacg gtcattgccc tagcgcgttg 82860 atgccgtgcc ggaatcggta tacggggctt cctaaagggc gtaagcagaa catccactct 82920 gcataagata cccgaaaacg aggctgagat aaatggtcta tcttgtgaag gttcgatccc 82980 ttccggggtt agcgtgtagt ggggctaatc aatccccacg ttaaacgttc aagttaagat 83040 ttgttagttg cagaagcggg gaggctatca accttcagtc cctgaaagct ctcagcgctt 83100 ggaactcttt cgtccgtaag gaatagatcc cctttcgctt tgactcgcaa acttcaaacc 83160 aagtgtaact aacagctgca agcctggatt accgctctcc ggagcggata attcaaaaat 83220 ttatttgcta aacgctctta tttactgtat atagttaca taattaatg agaggagtta 83280 gcaaatggaa aagattactg caacggtat tgaatctgca ctggtggttg attggggccgg 83340 atgggacgga gatcacgaat ggatggtctt ttatagctgt acgcttcagc cagaactatg 83400 gactaggctt accgatgagc atgctatgcc ctacggtatc atagatgtag aaattgagat 83460 taacaaactg gttggcacca ttatgtgca tcgagccgaa ggcgatcaca aggaaatctt 83520 ccgtaaagc attaactgg tagttctac cggtgacttt atttaacaat taactgagga 83580 atactatgt accacgtcc tactaatgga aattctgctg tagtccgcct gatgattgtt 83640 caggacaacc tgtccaacaa catcgagtct ctagaccgtc gtattgagga gtacgcact 83700 gagatgctgt ctctgatgcg cgaacgtgag gccagatcg aagagcagct agaggtttgc 83760 gaagctatcg accgcctggt tgacggacc gcagtattta tggcggaagc tcccgcagag 83820 cctaccttca ccccggtagc accagctgac atgcagtatg ctattctgcc tttccatctg 83880 gaagaagaag atggcgaagg cccgtcgctg gaagacgttg tcgctcct gcttgcctcc 83940 ggtttcccga acggaggtcg ttaatggatt ttatcgttgt ttgcggagcg aatactgact 84000 gcttcgagct gttaaacgac gccctagaca aggttgatga acacatgcag gaaggcagaa 84060 ctcctacatt cattgacctg tctcaaggta aaacttactt ctacccgtct cttgacgtag 84120 agcctacagt tctacctatc ttcatgcact ccctctcgtg ggacgaagaa gacgattaat 84180 gaaaaaattc atttgcctag tactctgaaa tttagtataa tatttatatt gaaacgaaag 84240 aggaaaagca aatggaagat gaagttaaat tgtacatttg ttcagattgc ggtgagttaa 84300 gtgaatgtca ttgggagtgt ccgacttgtg agtcagacga cttaatggaa tacgaaccgg 84360 aataacaaat taagcggcta tggtgttcag cggtcaacat accggcctgt cacgtcggag 84420 ccacgggttc gaatcccgtt agccgcgcca aattctctct gggctaagag atatgaccgt 84480 ctagaggtat cctcggacac aagggtcagc cctagacaca ataagttagc aaatatctcg 84540 tttcgacgag gtatttgaag aggttcttac gagagcttct tcaaatacgg gggaatattc 84600 tgtaagtggt agcagagcgg tctgtaaaat cgttgccatt gcggctcggg tggttcgact 84660 ccatcttccc ccaccaattt tgacaatcaa agagatccgg gttcgaatcc cgcgtacgca 84720 acaggcggca taaagcctt gtagcttaa tggacaaag cgctttggtt gttaaatcta 84780 tttgggag aagcagtaag tggtataggc ggtcgcctgt taagcgaatg acagtgagtt 84840 cgaatctcac ctctcccgcc aattactg aaaggttat ttattacc taggcacgcg 84900 taggaatact gccaccgcaa tgaagagatg tggcccggtt tggaggta tgccgctaca 84960 cgctacggtg ctaactccgt ctaggtgaaa atcctagtca gtaatacagt attaggtaa 85020 gaacagcgac ttaggttgca gtgaagcgtt agcaatcggt ggaactccgg tgcgccctaa 85080 ttaaacaatg catcattggc cgagtgacta ggcagaggct tgcaaccct cgaagcatgg 85140 ttaaaatcca tgatggtgct ccaattacca gaggtactca tgattaagta taaggcgtttt 85200 gtaacaagag agtcccaac aggattca agtattaat tcgaggtac aacgttacat 85260 gatacattg aagccgcttt aacagaggcg gaaacccata tagtatcgaa aagctgctat 85320 gctcatgtat gggaagtaaa cactatttta gatcgctagc tcaatggtta gagcactcgc 85380 cttttaagcg ataggttccg ggttcgagtc ccggcggtc taccatta tgggacttca 85440 gctaatcggc taagcatcat agatagcggg gatcccggt ctgtgagggg agtacacgt 85500 agtctcccaa gttccaccaa acatcagagc agcgtgatgt actccaaga cccaagacc 85560 ggggtggact tcttgattca aaggagcagg cacgaattac gcaactgtag ctcagcgagg 85620 tgagagcact gtttgaag tccaggggtc gttcgttcaa atcgaccgg ttgcaccaaa 85680 ttaatagtat caatagctta tggagctgaa tagtcctaa gctcacggta aagcagctgg 85740 ccgttagtga gcagacttgg aaacgtctgg tgatctctg gttagccggt gcgttggttc 85800 gatccaact gatacttcca aatcccaagt agcccgttac aggctctg attackactca 85860 gabaagcagg taatgggtga caggataaa taagccactg atttccggcc gtttgacgtg 85920 gtaaagtcct ccagtcaaga agaatttctg cgtgcagaaa tcactgaaag caatggggca 85980 gacaacttga gatagattgc ctataacgtc tcaagtagtg gcattgactt agggtggcat 86040 tttcccgaaa gttcgaaaaa ttcatttgct aaacgctctt attataata tatatttat 86100 ataattaat gagagggctt acctaatgaa tcaaaaaatt cttatgcgat ataatccacg 86160 agctttatgg ttccgctggg aagtgattgt atcgtaccag atacgagtgc gtaacggtga 86220 tcctgagaac aacattatcg tgctggagac attctctaat agagatgcgg cagttaagtt 86280 cctgaacacc atcgacaata ctttaatcaa ggtatattaa taatggatat ctttactact 86340 cctgctatca acttggtcgg tgtcggccta ttccaggcaa cggtctatcg tattgatgat 86400 agcactgacg ttgtaacgtt cattgtaccg gagttcttcc ttgagaagtt ctttgaagag 86460 tttgagcaat tccgtgaaga gcatgatgct tactccaata tggaagatct agcagcgatg 86520 ttcccaactg tatacggcta catctttgaa ggcaatgatc tgcttttgga taagtcagag 86580 ctggtggaac tcaactgggg cattagcttt gaagtgggct ctccgttccc gcggtatttc 86640 caaggtctgg agattcgata atgggcggct actctaattt catcgagaat tacattaatt 86700 ccgtagattc ctggaatcag gaaactctag tggtagtgct taaagagaga ttcaatatct 86760 ctactctgga agccctagag gctatagaag cctacttgga taacgattaa cacccacttg 86820 gtccaatctg gtagaggcat gaggcttaag acttcagggt tcccggttcg agtccgggag 86880 tgggtaccaa attatggggc acgactgatg gggaagtcgg cctggctagg gttgagatac 86940 ggttcgattc cgtaggcaaa tggtctcgta attccgcaca ggtgatggaa ggttcgagtc 87000 cttctcccat aactatacaa attcccctta gctcagtctg gcagagcggg cgctttggga 87060 gcgtcaggtc aagtgttcaa atcacttagg ggagaccaat cttgcctcaa tagctcagcc 87120 gggagagcaa ccgccttgta agcggtaggt cgtgggttcg attcctactt ggggcaccat 87180 tttcgctggt tatggttaca gtgtgattaa gtttacattg tcattaaatt ccggttcaat 87240 tccggaagcc agcaacccta aaaggtaagc atatgaggat tctttccata gaagatgtta 87300 tgtgtgacag atgctacggc tccatattct ctggtggctg tagttgtaag taatctatct 87360 aatgaccttc tgtagatgta aaataaatat gaagggtcga tgctcgcggc taccgacaga 87420 tcgtaatgat caagtagtag tctgaggtga agagactagc ccattaattc cgcgactaac 87480 tgtacgggtt acagcgtccg ccttccaagc ggtactgagt ggggttcgat tcccctagt 87540 cgctccaaat tcggtaaaat gaccgtcccc tgacgttacg gggaattaaa caaggaactg 87600 ggagccggta aggccatttt atcgatacta cttgagaggt acaacaatga aagcctttga 87660 tgcagaacta gtgttctcac tcttagctga gatggaagcc tgcgtagatc gtgtacgtgc 87720 gctgcgtctt agtatgttta gctcttaaat tatttgctgt tctagagcca atgtagtccg 87780 gggtgccgac gtctgcctgg aactcgactc taggatagtc aggacgtcac ttaacagcca 87840 gagatggcaa actataggag aagcaaatgt taacagttaa ggtaatgtca cctaacggtg 87900 gcgaagaaat tcacgatggt tcaagcgttg ggtttaatcc taagcagaag agcatctcta 87960 ttgctggtct tgatcagcat atcttcctga aagaagatga ggtagcctac gttatgaatc 88020 agaatggtaa gacagtatcc gtctaccacg gtagctaatt ttattccccg agtgttactg 88080 gacagcacgc cggtctccaa aaccgtgcag taggagttcg agtctcctgg ggtttgccag 88140 tttaaaggag tgatcatgat cacttatagc accaatttta tggggccagt ctccaacaat 88200 tggtacatcc gtatgggtat tccgtatacg gagtaaccg agccgaatcg ttttgcagat 88260 ggcgggcaac taactcgaaa agtatttgcc aaacgatatg ccggtggtcg cattgacgtt 88320 cgtggtactg atgactattt cggccaggaa atcggtgttc cgattatgga ggccgagtct 88380 tggaatgaac tacagcagtt tctctggaca ttctcgtccg ataaggttct gacattggag 88440 caaattgtac aggctctaga ggacgagaca ggtttcgca ttgtttggtt taggagcca 88500 gcatgtacat agatcgtaac caactgttca agttcctaga gcttgacctt cgttggcctc 88560 tatcagtcaa tccgggacga gctactggca agacgttcga ggctattaat acagcctatg 88620 agtttgcagt atttaaaggt attcaagcag tatacgtagc atccggtgtc cgtgaaatgg 88680 cacgcctaga gaaggatt aacgagcttc aacctcatgt taaaattaca acgtatagatca 88740 tgttagaacc ctaccgtata ggtcgtcgat tcagctgcat catgtttgac gaacctagct 88800 tggctattaa gtacggcgta aacgcttacg tggttcgtat agccagagaa aaccagtgtc 88860 ctgtaattat ttttggag taaagtggaa caaatttc aagtctcgt agacgccctct 88920 aagagagttt tattcataca agccaccgat gaggtcatg gcctgcaact ttccttcgac 88980 tctctggagc aaataacca gattgttctg agggcacaa atcattaga aaagaacact 89040 gaggcacctc ctgacctcta aaaaagcc ccttgctct tgagcgagg ggctttttct 89100 tttaaattc acttgcaaaa atgccaattt tgccgtataa tagattatta aattgatgag 89160 agagatttcc aagatgaag taacaatgga aacacagaa gagtttattg ctactctgtac 89220 tgcgtagca gatacgcttc cacctgaggg tatggacgac catacaatgc agttagtggc 89280 ggatatctac agattagcag agttagcta agacacac aacaggctgg tgtacgtca 89340 agaacgccctc gaatgatgg aataggagta atcaatgac gactcaatg aactgaatga 89400 attgcattac gctgaacgtg ctattgacga gctggattc gctggtggtt attatacccg 89460 acacgtcaac gcaatgaccg ccgaagggct taatagtaag tctgcgattg ctgcggagtt 89520 agcagttcgg gactttgtta ttgatagtct ccagaagact attagcaact tgagcgaaaa 89580 caacaaagct gctctggaag ccctggataa gctgtccaac catcttctcg cattggggat 89640 caaataatga ataagcaatc tctacgtggt atccgagtat ttcgctctag ccttgttgat 89700 tccttctata tctggggcaa ggcaactcga cgcactgttg agcaagcact agacggtacg 89760 ttctatgatt ggcgagaaaa agagcgaaac cccgtattca gccgcccagg gctgtatcac 89820 gatcgtgtat ctaagacagc gtggtacgaa attgaagtaa ctcctggcgt tatcagagcg 89880 ttctatacgg attgggagca cgaaaagtgg gtttgggaca atcagattgc tcccggtgat 89940 cgtattatga actatgccga atataaagaa atgaagcgca tgttcgagct atacgatgtt 90000 cctaggctgt ctcgcccggc tatcttcatt gctagccaag agtactggca tactgttcgt 90060 atgaagcgtg attttaacaa gcatcacctg cgctatgaga aagaacacgg cacactgcgg 90120 gaacgtgtgg ccaagagaaa agccgaactg cgtgaaaaac gcctggagaa gaaatatggt 90180 gaaagctagt ctcttacgct tcactcctgg agttggcctc cagtataaaa ttgttggggg 90240 ccacaagttc caatacttca ccccgggtaa gctctacttc gtcgagctac acgatagtag 90300 agcaggctac aaattacgaa gcgacgcaaa tgagggcatt tgggtgagct tcacacaagt 90360 taaacgctgg tttacagttg aaggatataa tgattatgag taaagttgta tatttcgtaa 90420 aatgctgccg tgacacctta gagttggtcg accacaaaat gcagacctgt aaatgtggtg 90480 cttcctccat agatggtgta gccggagcct acgtccgttt tctaggcgac aaaagtaact 90540 tcatgcgtct gaatgagttc cagctcgaag ttgaaaagaa tcgaccggct ctagaggcag 90600 aggctgagcg tctcaaagat ttcgatggga acatcgtagc atataacatg gtaagcggta 90660 aagacttctg gtctgaactg gctgaaaaac ttaacatgcc gcggcaggcg gctaaggctt 90720 tatatcatgg cttcaactac tcgccacgtt ggaattaagt tttccaacag taacagaact 90780 tacgtttacc gagtacctag ttattggaaa tactccccgg agataggtga tgtagtagtc 90840 ataccgggga atgttatgtt taataatcct cgtcgcgcca aagtagtaga agtgcatggt 90900 atgtatgta agccagagta t'aagaaaga aagaacatct cttatgtaga gctgcatgat 90960 tacttaccga agggaacg taatggacga taaaattgct cgggaagca tagaacttgt 91020 tcgtaaacga ctggaagaac gtaatgtcga ggtgccgaaa atatgctta ttggctacgg 91080 cggcatcggc ggatttccca gcttcactga cctagaaaga atggaacgag agtcccaagc 91140 cagctttcta gagttagaat cctacgctcg ggaatggaa cacgaacatc cgattggtaa 91200 taactttatg ccacgtagct cccgccagga ggttactcac ggcaagacta attcctggcc 91260 cactccgaaa cgcagaggta gaaaatgatt actacaggat ttggatattc tcacgaagaa 91320 ttgtgcaaaa tggttgaaag cgctccgtttt attagaagt tagtcgaaga gcaacgaccg 91380 gtgtgtttac atgctgcctg taccaagtgt cacggtactg gtgttgataa aaacggcaag 91440 atgtgcgtac atgcactgtc ctgcccgtgt cctaaatgta gctggagctg ttaatatgga 91500 tctaggatat tgtgtagtac atgaattcat ggagcagggg ttgccggatc gtatttgcgt 91560 tgtaacctct cgtaatctgg aagcagccca atcactagtg gagcgactct ctggctacta 91620 tcgtgaccat gaacgttacc agcagaaagt atttgacctg acaaaactgt accatcaaaa 91680 agctctggat atgccaactc cacagctgga cgatctcaag cagtttagtc ctgaggcctg 91740 gtatagcgtc aaagacgcca gtccggtaga ctataccgta caggtattct cgcattatgg 91800 tactcgtcac tggataaacc gtaaatggga ttccatggtt gactacctta attcggaact 91860 cgaaaagggg gctgccgaat acaaaagaaa gcgtgccgaa gccaaagtat tgaaaaattc 91920 agttgcattg taggcgattt ttcgatataa tagttacata aattgatagg gagagcttca 91980 tgagcaaact aagtattgaa agcattatcc gcccgttaat gcacgggtat gtacaaggtt 92040 cctgtgtaag cgaaactgaa gctctcaatg ttatcgaaga agagctggtg gctaacgggt 92100 ataatcttca cgaaggtgtt atcgaggatc tcttctggca gaccgcggaa gatatggaga 92160 tctttcggtg cgtgaattgc gggtggtggt gtcctgcttt tgaaagggcg gagaaccaaa 92220 tagaggaaat ctgccgggac tgcgagccag acctcgaagg tgaagtggat gaacaagata 92280 acgaaggtga agactatgag taagcgttca attgcagcaa tcatcgcatt cagtatgatg 92340 tactctggag tatctctggc cgcagagcgt aaaagtag agatctctga taacgggcgg 92400 gttcgtgtaa caacgaatgg gattactaaa ggagctggta agttcgtaa atctgaaacc 92460 cgttttgggg aaactaagat ctacacgaac aaaacctatg gcaagcccgc tgtgacactg 92520 gaccgctatg gtcgtcaggt agaagacgag gatgatagcg atgagtaatc ttcaccccaa 92580 acttcaagag accctagact ggattaatga agagtgtgct tttgaggaag ctccttactg 92640 tgtctgggca agagctggtg cagccccctc ggagtggtgt actgtgtttg ataatcgcta 92700 ccggataaca gttgaactaa gcctaaaaga ggacaaagta tacgctaaag cctctatgac 92760 cgcgctagga ttacgggt tcgtggagat gcaagagctc tgtatgccta acactcacct 92820 tcgggttcag atcgagcagt tggcaacaat tcgtttgatg ttgccggaag ataacatcaa 92880 tgaccatttc cataaggtta ttgaaaatga atacaaacta cggcggcaac gtcgcaaagc acgacggga gtagagaaga ctcggatgat gtgtaacatg aatccacacg tataacggta gggccgttat aggagtcacc ttcctgttcc tatccggagt cgtgcccgcg atgaacgaag 93060 tggtgagttg gtcgccgacc actaactat tcaaagacaa actgaggaag caaaatgttt actttattta tactggcagt atcggcgtgg atggcagttg gtatcaatca tggtctggac tctgctaagc tgctgtcagc taaagccttc gagttcctgg ctaagtttgc cacccgtaaa gatatcgagg ccatcatcgc aaaaggcggt gccaaagatg catcgagtgt cctgaagtcc 93300 ttcgataaga tcctagagct tcgtaacggt aagcatgccg cagagctacg ctgtatgagt cgcaagacca taggcagact gtgtaaggcc atcttcatcg tacagggggc gttgaaaggc ccattcgcta father ggatagtatc aagcgagcca agatctttaa cgattactgt gtggaacacc acccgttaaa tcgctaacta cccaagccca gcatcgaaag gtgttgggct tttctttttt aaaattact tgcgctctcg cttaaaatgt tgtatatag ttttataaat gagaggagaa ctaacatgcg tattatctct aaattaaagg acgtatatga tctacaaggt 93660 actatgtatg atgcagaaag agcatggtat cgcgaagaag tgaaagaggt tgtaaacgta 93720 tccgctgatt tcgaacaaat tatttctac gctgaaatct tgcgtaaccg cacttcctct 93780 ggatatgggg gtgttaatat gggaactctg gaagttcgtc cagtattgat ttgtggtaca 93840 ctacgctggc tgtacggtta tcacactggc ctaggagcag atgctgtgca tatccagact 93900 ttcgatccag ttaaggtaaa agaggtactg gaagaacaag gatactacct gcggatgggt 93960 tgggacatga atacgattga gaagatcgac gctcatgttc gtaacgcaac cgccacggct 94020 tctgcgttcc tggagacttt caacaagccc attgcgatgg catgggatgc cgcaaagtct 94080 aagacagacc caacggttaa tattaccgtt aaaactgatt tcaacttcca tgcggaagat 94140 ttccccgtggc aggaaattga tccaaacctg tatcgctggc accagacctt agaatcatat 94200 atcttcggtg tgctgggtca aggggaacca aagaccgagt ctacgtctga tcgtgatcgg 94260 ttattgcta agggcttcga tgctaaagtt tctttcagga atatggagcg gtaagactgg agcaattctg tttctggcaa ttgcagcagg caccttcggg ggtgcctact acattaccaa tagctaca gatatgtcca gttctttaca gtccctatca aaccgtaatg aacaactaga aaagacagtt ggtaatctcc aaacagagat tcgaatcga gatcgcaata ccactaccta cataaccaac ctagctaaga accaggaaga tctggatggg cgtattaata aactggatgc agctagagcc aaagagggcg tggttgccgc taacctaaa cttgctacca aagtggcaaa agacaaagtt aatgagttcc aggagagact ttcatgcgta actggaaata tggactcctg ctctcggctg caattatcgc atccgggtgt gcagaacggc cagacccaag tagcacagta acggggggttg agccacagca cctaccgtgg ccagcaagcc tacagacttg cccgtttaat tttgagttca taacgaaga agggaaagtg tatgttcgca taccttacca agggaagtc acgatgggca agtgtaatga gcaggtctac acctacattg ccaatctgac tgcattaacg tgtacctatc gcgtgtcact 94980. tgtacctatc gcgtgtcact tgaaatacgt tttaggatta attggtgatg ccggcgcagg caaagacacc tttgctgaca 95040 tggctaaggt ttgggcctgg gaagtcctcg gtccggagta ttctatcagt aaatttagtt 95100 ttgcagctcc agtttacgaa ctcgcggctg obeyccttgg agtcacgcca gaaaagctgg 95160 cagagcgcag gacaaaagag attaagcagt ggttttgggt gactcaggaa gcccttgagc 95220 gtactgctaa cgtctggaag cgttttgga tcgacaagta cgccgatttc tcttacgttt 95280 ggcctcagtt tgaagcgtca gcactatatc cgttgattgc taagactgcc ccagactttt 95340 atcaaggtcg tgagaccccg ttgtacccat tatacacgtc ccctcgtaaa atgctagaat 95400 ttgttggcac ggagttagga cgcgctctgg tggatgagaa cttgtggttg aatattgtgg 95460 tagatcgtat cactgctacc aaagctgaca ttagtattat ttctgatgtt cgttttgata 95520 atgaagccgc gctagttcgg aacttccccg gtgcgcagaa ctctagtatt cttaaagttc 95580 acgcccctaa caacatccac gctatccaat ctactcatgc ctccgctaga ggtgttgcac 95640 ctgagtttat tgatgtg gtaacaaa actttgacgg actggaaaat ttccgtaaaa 95700 atgtaaacgc attttgcgac gagaggattc tctttatata aggtaacaat atgggacaa 95760 ttaataacgt agagcagaaa ggtggcaata agactcctaa ttactcgcc tcctagtgg 95820 cgactaaggc agagtataat atttaccact atcatctcga tgggccgata gtcgatgtag 95880 actactaccg cgacctatca gtcactctgg caactatgca agagggac actacttaatc 95940 tttatattaa tagtcccggc gggtacgttg atacagcagt tcagttgtgc aaccttatca 96000 tgaactgcca gggaaccgta attgggcacc tggttgggcc atcagcatca gctgcctgct 96060 caatcttcct agcatgtcat gttggctgg tgcacctta cgttatgttg atgggtcata 96120 cctatcgtgg tgcgcactat gctagggca aaaatgagat ccagcactat gctgatcagt 96180 ttaactcgtt cttcgaggat atgatgctag acctgtacta cccgttctc tcttagaag 96240 agatcactga gatgatagaa gtggtagg atatttggct gacctccaaa gaatcaacg 96300 aacgtgtaga tcgtatggct gcgcaccgag aacaagaagc tcgtaggct gcgggtcaat 96360 96420. aaaactaagt aggccagggc tttttagctc tggcctttttt ctttatctaa attctacttg acatttgtct caaagtatga tccgagcaat aactataat ttttattgca acttcctcca father father father tggttagagg father tggataatt fathercagttg atatcgctgt tactacaaga agcgaaggat ccagcttctc ttcttaaacg tttgctaacc ttactggttg gtctagttat ctacctcttt atagctaata cgagtgaagt catgtcgtac ttaaagactt tctctacttc ggcggtccta caagatgtta aagtacaacg cacactagag tttccaaatg tggcacgaga gaaggcaatg atcctattct cacagactcg agctgatgcc gtcttcgtgg father gcctgaggct father accused catagcttgg gaaagcaacg ttcagttgga taaatccgat gtatccgaca aagcagttga caagacgtct atgctctacc gagcgcattt agacggtctg aactttgcaa ttgatgcaag ggaaaaacga ggtttatcga agtggtctgg aacgggtttg ccaccgttca aggtgcaaa tttcgagtat gtgtacacgt gcccatattt taaccttaat aacatctact ctggatatgt tgccgtcgcg 97080 tgggagaagt atcccctaca agacgaagac atgggtatgt ttaatgacta tatggcaaaa 97140 atctgcgcat cgccgcagag atcattaggg agatcaatat gagtttcaga tttggaaacc 97200 gtagtctcca gcagctcgat actgtagatc ctaaactcaa ggctctagct attcgtgctc 97260 tggaactctc accgcacgat tttaccatca ttcagggcaa acgtaccgta caacagagtg 97320 cccagaacat tgctaatggt acttcatttt tgaaagaccc tagcaagtct aaacacgtta 97380 cgggcaaagc cattgacttc gccccatata tcaatggtaa gattgactgg aatgacttag 97440 aagcattctg ggctatcgtc ggtgcattta aaaaggccgc aaatgagatg aacattgccg 97500 ttcgtttcgg agcggattgg aataactctg gcgactaccg tgatgaaatc caacgtggta 97560 cttatgacgg cggacacgta gagttactgt aacattaagg ggagctttta ggctcccctt 97620 ctttgtatgg agcaagcaaa taggaggtgg ttatgtttgc aagtttagta actataatgc 97680 tactcaagat atttcaattc ggtatagtat tcttttccgc aacatggatt ttaatacgcg 97740 gtgccattta ttttcgcaga acacgcttag ctagggttgc tagatggctt ctacacttcg 97800 tatttggtat tttcggtgtg ttccgctgcc catgtgagaa aaagaggggga acagggttct 97860 gctggctctg gttaggtcta gagaattgct tctcattgtc cttagcgcta ctatttggta 97920 tgatagtcct agctataacg ctagcattta taccactaat gtacgcaggt ggggtgacgt 97980 ctatcctaac ccttacttct cctgtgttga tgtactcagt ataccccatt acactatttt 98040 tcgtaagacg aggcaagcac tgcaattaat gaaaaattca attgactttc cgctcgattt 98100 agcgtataat attttataa attcgagaga ggagatacaa aatgtcagac agattctaca 98160 ctcaaatgtg cgagcatttt aaagtatctc cctacgagtt aaatatagct ttgtgggatc 98220 gtgaatcacc ggagtttaag aaaattgcta agaaatcgga gggtgttatg tccaacggta 98280 agaaaatgac ccgaattgac cttaacaatg cgctaaccaa actgctcggc gttaacattg 98340 agggccagaa gctctctatg ccaactttga ctactatttt ggagaaagtc aaggctggag 98400 atgttaaaaa agtagcagtg cctgagggtc gtctgaaaaa gccgtatcag gaggctataa 98460 ttgaggcttt cggagaaaag ctagacctgg ataccgcaac cgtaaaaaca atgaaagcac 98520 tactggagag tattaataat gtctaagaaa attgtattcc ttaaaggttc tagctgcgtt 98580 ccttgcaagc aatttgaacc agttttgat aaactcaccg ctgagttcaa cctgcccgtt 98640 gaaaaacgca cggacgacgt agattcccta cgtaagttcg gccttcgcac tgtacccgca 98700 gtagtcctgg tggatgtgga gaacggacgt gaagaagcac atcacattct tagtggtgcc 98760 acgcttcgct ctgcagtagt tagtaaagct atccaagact ttatcgacta cgtagaagaa 98820 tatacctaa ccccggctca atacgagacc ggggtttttt attatttgct aaacgcttaa 98880 aatttgata tatatttat attgaattga gagaggatta cacaatgact aagcaagcat 98940 atcttatcct gaataatggt tttgcagttg gtactacctt cgttgatctg gggtatacaa 99000 aagaagagtg gcaagctctc gatgctgccc agaagaatca gcttgttaat gaagccgcct 99060 gggagtatgc agaggcttac gtggaggcgg tagatgacga gttagttatt gtcgtttccc 99120 ttggtgccgt aggttgtgat gctcatgtac atacagactt ccagagcgaa gaagagtggg atgagttaga tctaacccac caaaatgctc tgattaacga agcgttctgg gaagttgtag actgctacgt tgcgttctgt aaggacgatg atgaagctaa cacctgcact aactatggct atgaacacga cgatgtggag tgtgcataat gcaaaaattc tctagagact ggtcttctga tatggcccgt aaaaatcgag ctgctgccta ctataataag aaaatacagc tcgataagtt aataaagggt atcacctaca atgtaggag gggcttttct ggtataaaag taggctag gagcctggac tactcgtgca tactgtgggc tagcagaat gggtacgcgt ttaaacgaat aggcaatgag atactgattg cctgggaacc cgaaggtcta gtgcaatacg fathercga cccttatcga ggggaatatg taagagatca caggcagcag cccacagact tctcgctgcc aaaacgttac tatttagaga cgaggtatta atgaacgtac atgaaacagt taccgtgccc gacaatgcta atatcttctt cattggggat attcatggcg agtacgacat gatgatgggt gccctgaaac ttgcagggta cgaggaaggt cgtgactacg tgttctgtgt aggggatctt 99840 atcgaccgtg gcccgaaaaa cttacaggtt ctagcgaagt tcctgtacaa cccgaaattc 99900 cgctctgttc gcgggaatca tgacgaattc atgatccaag acgactacgc taactggatg 99960 tataatggcg gtagctggac tatcacggaa ggtttcgata cggataccat gaaaggtatc 100020 gcggaagaca tggacagcaa gatgccatac attatgaccg ttgaacaccg tggcaagcgt 100080 tacggggtag ttcatgctgg tatccctctg cgctaccagg ctcaaggtat gggtgtaact 100140 gtaccggtgt gggacgacat tgtgcacgaa cacgaatcta caccagattt acgtcgtttg 100200 ggtgtactgc tgtgggatcg tgatgtaatc caagaagttg ggtttaatct gtatcgtagt 100260 ggggagaagc atccgtactt cgatcgctac gccagcttct ccgaggagtg cgcagtcgat 100320 gtgcctgaaa tcgtgggagt agactacacg tttcatggcc atactggtgt cccgttcccc 100380 attcgctgga agaaccgtgt ctatttggac acaggtggca cctttaacgg tcgtatgacc 100440 gtggcgtcgc cggttctagg tcaattatac acattcacca cagatcgtga tgatccttgt 100500 ggttccgctg acattattta ataggtgaaa catgaaactt ttcaaagatc tggagaagaagg 100560 tgaagtattc gtagttgctg gtggcttcga gttgcagaaa tgtgtagcaa tgctggacaa 100620 cggaaactcc gtgtttacag acgacgcgaa tatctcggtt actattgccc cggacactga 100680 aacctggaaa cccaaagaat tctgggagat ccataaagat cgtccattgg acgacctact 100740 ggacgacatt ctattcacag cctgaggtaa atcatgaaac tcgaatctcg ttatattgta 100800 tttaagcagt cagatgccgc taggatatcta accagcactg ctattcggga gataaacgac 100860 agcttatccc tgatctataa aggccgagag gctgacggca aggtaggatt cccgaactat 100920 atcgttctag aagaggattg gccggagtat cctatcgcta aagaagctct agaaggacgt 100980 atcgtgctgg aagagtttaa taaaagggcg gagagaaac gtggggctaa ggctgcagaa 101040 gatcactata agcagcacca gtctacggaa ctactccggg gtatttctcc attctgtgca 101100 gggtggaacg actacatgag aagactggta atcgaggagt aaccatgtat actggaatgg 101160 gaaatgatat ggccaagatg tttattggcc tcttaatcct ggctgcgtta gtcggtgcag 101220 caattgtagg cggaatctgg gcgctcgtag catttgtatt ttaaattcag ttgcttccga 101280 gccaattttt ctgtataata gctgtataaa ttgatgagga agcaaatatg aaaaacgcaa 101340 tgatcgaact gaatgcaaat atggaatccc ttcgccacgc agttaacacc gcccgtgctt 101400 cttttaacct gctgatgcgc gatgagagca taccgctgtc cgctcgtgtc aaggcatttg 101460 aggagttcgc cgatgagctt cttcatatgg gagactatct cagcgactcc ccgtttaatg 101520 aagatcgccg ggattatcaa cacgcctact gcaatcgtgg ggaaatagtt tacctgaccg 101580 atgttctgga gagtgtgctg gagtatgcga actctttcat gcgaactcct gacgaatggg 101640 aagacgcttc gaacgactat gttctagacg agattcaaaa gaactggcca gaaattaaga 101700 aactggtaga agaacacatt cattctgaag tttacgctta ccggatcgat tggtaaaaag 101760 tttaaaaatt cacttgctaa gtagttagaa tttcggtata atatacgcat aaattaacga 101820 aacggacgga aatcattatg gctaagaaca ctatctctta caccactggt aaaactgctg 101880 agaacaagc taacaccctg accaaagacg aaatggttgc ggtactggta atcctgctgg 101940 atatgagtgg gttcgaaggt cagcttgcta aactgtctct cccggcactt cgtgctctgt 102000 acgaaggcac taaaaaaat gctgcggcct acaacctcgc taaaaatgaa gcgcgctggg 102060 ccaaggaaca ccagcaggtt gctgaacgtc gtgctgagag tttcgaacgt gacctgaaac 102120 gtgaaaaggc gaaaaaag taatggtact agaaattctc tccgggctgt taatagcagc 102180 cctagtaact ggaacgggtc tggcggtttg ggtatcgata ctgcgggaaa ataaccggag 102240 aatgagatta acaacaacg ggctacacga gaagttgatg gatcaagtac aagatgctga 102300 tgaattctct gctgcggctg agcgactttt agttcgcctg gcgaagattg aagagattat 102360 cggccaagat tccacaatgt cgaagacaac caaaatgcgg ttgatcacgg agattaagaa 102420 atgatctcac caattgtagc ggcactttac ttagtagttg ttggctacct gtccgggaag 102480 taccacggct tcggtttaaa aggtactatt aaggcagcaa tgctggttcc tctgtatcct 102540 ctggcaattc tgttaaccgg gtactacgcc tgtgttttaa gaatctttgg ccggggcaaa 102600 gttaactacg ataactgcac agcgttgctg gacgatattg aaaacaccat taagaaggaa 102660 gaaaaatgaa ccttaactcc aaagaacgcc aagtgttagt agatgctctg cgtcaggtag 102720 tagaccacga tttgctttgt gatgaagata tcgttgagtc aattgcccta atcgccaaga 102780 ttgagcttgc tgagaaggac tcatggcgtc ctttgagtga gcttctctcct ctaggcttgg 102840 caatcgtagt acaacgtgct gatggcgctc cgtttaacac tgtgatggtt cgcagagatc 102900 tggcaaagtc ctactccccg gatatcatca cgttgcacac taagatcacc aacgaacctt 102960 tcgagtttaa tactcgtcac tattactgga gactgaccaa tgcttaatca gcttcgaatc 103020 tataatttcc tggattgcaa cttccaggtc tggcgagaac tccccagccg tttcctaggc 103080 tgggcattat tcctctccat ggtggtttca gtcttccata acgtcccagc gacgttctat atggaagcaa ttcaaccagt cactgtattc atttacgaga tgtacctcgt agcaattac gggtggaag atggccgcat caactgaagt tttaaatcaa tattttaatc gtgagcacca agagttcagc gatctgttca tccagatgtt cgtcaacgct aataacgctc tggactatcg tttcttcaac gagttccatg agaccacgtt cagccatcag gatattaact ccgctctgaa agactcatt ggctctaaag tgataccatt ccgacagact gcaaatgcag agacactaga gcttagtgtc gtttggggct tgttcaaga agcatatgag tttggcaagt atcagaacgc acgtcactgg atttacgagg tgtatctaaa cactgaagtc attctacctc gccaaatgat gttgggctgg attgcaaaac aacgtcctga aacgcaatgca aaatcattcg crack cgacgggac ttataccatg caagtgaaaa atttgatgct ccaaaatccg tggcttgatc gcctggcgca agtaggaag atgcttctgg ttgaaggtct gaccgaaga gacatttgtc tgaactcctt ctacaactgc aaaactcatg taatgcaggc gctagacgaa gaacgtgttg 103800 aactgagtaa gtttatggtg aacatcgcca cagctcaagt tcactggca acccaggggct 103860 tatccgccga cgatatcctg aaacataccc tgaacgctat tgcagagtat gggaaagctc 103920 gtggtgaagc tctcttggct tcaaagaag agtttgataa atcggagtca atgctgaaaa 103980 tggcaatcga tatccatatg gaaggcatcg acggtactat tcactaagag ggtgacaaat 104040 ggctaagaaa cgcgttgtgg taaacttcct agaggagat tctggggact gcgagtacgg 104100 ttgctggaat actggctacg gtgttgaggt tattgtggat ggcaatgcg tccaccgtca 104160 ggaagcctgg gcaagctgca ctaacaacag cagcgtagac ttgacgtat tggcccatgt 104220 tcttcagggc atcagacga aagaaggcta cccggtcaa gcagatcaca tcgactttgg 104280 agatccttcc gattatccag aggagttct ggatctgttc acctacaaa gaggccaggg 104340 ctaattgctc tggcctttcg tggttttagg atattaatg aaatctatag acaactattt 104400 gcgcgggga aaccctgtag atcaagcagc agttactgtt gaaaaggtca ggaaagagtg ttttatactt actcaacgtg gaggtggtaa tcggcctaat cgagtgtatc ttaattggac tcagtccaaa gacctgtatg aggregate aggregate aggregate aggregate cagagata atcaagagct agtatggga gctaatcct tccccatta tcaattcctt aaacgtgacg gttcgatcaa ccataatag ataaagcaac ttaggccaga tttcaggcaa gatgctaaga acctcatctt tattaaccgt gctctagacg ctcatggagt attcttcgga 104820. 104820. 104820. 104820. 104820. 104820. 104820. 104820. 104820. 104820. 104820. 104820 104880. caagattact ttatgcaaac tgctagaagc caccttgagg gtaacggttg ccaaaaatgc aggcaccgca tggttaccag agttacggat tatggaagtt acaccgtgcc agcatactat cacaaatttt caattgacgg agactccatt atttggtata ataactccag taattgata aaacctttgg aggttaaga tgaaatacga ttccctgaat aatccgagca ccaactatct gactgaccag tcagtttctg agatcaagtt ccatccgaac tactccccgg actctagcaa 105120 gccgagtgta gcagctatct ccttccgctt ccgcaatctg cgctttacgt tcgttggaga 105180 ggaagacaag atgatctcta taattgacaa ggttaaagca gtaagcgagc tgtccggtag 105240 cgataccgtt aagttcgaag cattaacctc gttgctgctg actagtgggg ctaccgttgg 105300 taagttcgaa cttattcagc cgcatgtttc tgcactgacc aataccagaa acttctggga 105360 tcaagccaat gtcgagagtc tcataaaatg ggatagtgct actgagttct acaacaagta 105420 agagggctag gatatgttat tctgtacagt tgattttgaa gaagctaacg aaacgtatat 105480 tgtctacggt atgtctgaaa gcaaagtacg tatcctctgg aatcagttcc aattagaagt 105540 accggacgac atatccaaga ctccgaaaga cttctttcat ttaatagata ttaaggcagt 105600 aaaagctcgt aagaagctaa ctccttatgt gttcccaggt gcagtattcg tccatgagtt 105660 aacagcgtat actaacgtgg ttctcaaaaa gtctcgacag cacccaggct atctcacgat 105720 gttgacgtac aaagttggag ctattcacga cggtgagctg gtggttcggg tagatgcacg 105780 cttgcagcaa gaagtcgaag aaatgattag gcagtgccaa aataagcag aattaaaca aagagcacgc ctattcgaca tggcagcacc tagcgaggca gttgccgcat atcacggctt ttataaaga atagctgaat ccgatgaaga tttctttatg tag acacaatgaa cgagaaatat gaagtatgga ctccggttgg ggagaattgt agctatcttc ttcgtaccct gtgtactcgg gaagatggca catctttctc agaatacttg agtgaatgcc acgctaaggc ccagcaggac aacccgctat ttaagatcag aggagaggat atcctcaaag ttaacggcgt gccgtacacg ccagtggaca gtttcgccgc tcttcaggtg tttaaggaac acagagagcg ggagcaccgg aggatgattg aacgcttgac aggtaggag ccgttttcac atcctaggtg 106260 106320. 106320. 106320. cctggttaag aaagctgacc aaaagaact ctcagagctg agtgaagaag agtatcaagc agtcctcttc tgttgttcgg caatgccagc gaagctcgat ggagtactgg caaagtcaga catccacaac 106440 ggtaaggaaa caactttcca gccgccgtat aaatggctag cgtccaacat ccagcaaatg 106500 gtgggtaaag ttacgggatt ctcaaataga aagacgccta acatctttat tgacattact 106560 cctagaaccc ctgagtttac gaaggactgg agagacgcat tggattcatt tccgtcatgg 106620 aaagtctttt ataaacctga cgatgaaacg tatgcacatc ttcccttttt gaagcaccca 106680 ggctacacag ttgaagaccc aagttctggg gttaatttca aagacttcaa gtgtactgac 106740 gagaacattg cttacggact catgagaaca tccgtgagga ttgcaatgga tcatgaacta 106800 gacaaacaag atctagcagt cattgcactt tgtaaggatc gttatattaa agttaagagg 106860 atcgcagaga aactttccgt actttcttgc tttgaaacaa tccgagattg tgaaccagag 106920 ggtgaatatc ctaaaggttc cctctactgg aaagatgtta aacatctagg actatcagaa 106980 gaagccgtat tcctaggctt agtagttacg ggaagattcc tgaggctaca ggagaaataa 107040 tttgtacttc gtctatattt tcgctaatga agatcagcgg cttatcccta ggcggaccaa 107100 ggtgggttac tcgcgtgacc cattttatcg tctaaattcg cttcaacttc acagactacc 107160 tcatcgtgga ttacaggata tcgttattca ttctgtctat ataatagatg acgccagcga 107220 gttctcagca aagttgctag agaaggccgc gcataaaaag tttaagccta tgcgagttaa 107280 tttcggtgac aagtttgatg ggcatacaga atggtttgat gttgaacctc acgttattga 107340 gaagttcttc ctgtcagttg gggcaaaaca agtacctatt gacaaactca tagcccaaga 107400 acaaaagatt cgtaaatcca cgaaaaagta gttgaaaaaa aataaaacgc atgttataat 107460 aatctcatag attacataca atttgaaaaa tttatgaagc taggcggatt atgaaccaga 107520 atcctggact cggggcgacc aagcctacga tgtcacagag actttctggt ctcacataat 107580 ccgcatgcct agcgcttcgt aatctcacat gaacgatgga gaacctataa ctataataaa 107640 actaaggaga aaatagggaa acgtgggaat gaactcacga ttactcacgt ttaaaagtag 107700 gtaatcctta cacaccatcg tttcgcggct ctagggttga actctagtca tctttctata 107760 cagggcggtt ctttctacac acctaagggg agggcggtta ttttccaacc ctaaaaatcc 107820 ataacattt tctggaattg cacaaatttt cacaccgtct alatccc acgaagccct 107880 cacacatctc acgcaatccc tctagtacgc ccaagttcta agcgtcactc cacaaagtcc 107940 tcacgaagtc cccacacaaa cccacgtcat tcacccaagt tctaagcgtc aaacattatc 108000 ggatttgcac attgctccta gatattaaag aagccaagca tttctagatt tctagcaatt 108060 gtcggatttg handicapgacaa aacgattgtc ggaaatgcac attaactctg aatccaaaaa 108120 cacggtcgaa tttgcacata taaaaactac tatcggattt gcacatactc aaacgcttgt 108180 cggaaatgca cattattact gaagtacggt agctgtggat caaaatcttc gattctgcga 108240 atataaaata tctcctgaaa tttcaagcct ttttacacgg tgtgtcaccg tgagttcgct 108300 tgttttttag agccacagga attttttctg tgtcaacccc tttaccccca tttaactcat 108360 aatattttcc cttatctgcc ccctgccccc atcatccccg aattgcccca attaaccccc 108420 tttatccctc ccccaaaaat acccatcgga tttgcacata ctcacccaaa ctcgcgccca 108480 ccgctgcgct gcagcaaact gcgcgccttc ccctaaaaa ttattatcgg ttttgcacat 108540 attagtatcg gatttgcaca tgcccctagg gcggtaaaaa taaatttca tcggatttgc 108600 acatacccta gcagcctcac gtaatatttt gcttgacaca ttgtcggatt cgcacataag 108660 cgcaggaagg gccgggggtc ggaaatgcaa atacgaatga gagtgattcg catttgagaa 108720 gttgaatgcg aatgataatt attctcgttt gataagctga atacgaatga gaatgattcg 108780 cattccggaa aggaagtgag aaacactatc atttaagtta tcaccagagt tatacaaaat 108840 tgtaaattag cttgacgctt gagaaggtcg cagacgagtt atccacagac ttattaacag 108900 gttatcctac tggttattta tacagtagtt cccggcaaca ctttctgcta atgtcgattt 108960 taagcgccct aggaagcgtt ctaagcgatt taaatttcgt gggggtattt tagtaagggg 109020 tcacgcggtg gaggaggggcg gcgacactct tacatatcgc cgtggaaaca ctcaggccaa 109080 aagtgattta caactaac tattagcagg gtaatgattg cgctaatac tatcatccc 109140 aaaaaacatta ggccgctcat tattggcagc tctgcaaaa gactagggtc atataataa 109200 ccatgatgac acgaaaaccc cactcaccgc cgccaccgtc accgccttc ttctgaccgt 109260 cacgaataac gttcaatttgg taagtattgt ggctacggct tttgggacta ccaccgcgta 109320 aagttcgcat taaaaagccct ccttattaaa caagtatca taggaggcat agccgcaaaa 109380 agcgataaaa cccaaaacca tcatagaacc aactaaccg agcgccaaca ttgaaaaagc 109440 catttttcga tctcctatat gggtgagtca taataatac atactctag cgggaagcaa 109500 ctactatttt tagtagtaga caagcaaatc cgatttccct atagttgat ctcccggcaa 109560 gcatagaaaa atttgcttg ctaaaaagcg gagaatcttg ctataataga ttctcagcca 109620 aaaaaggag aacagaga tgttacagaa atttacccc gttgcaaatt tgcctatggt 109680 tcgcggtggc gctcgcaacc tgttagatgg tagtaaatgc gcctctatcg gtcatatttt 109740 aggcgtttac cgctcgaata tggaaagtag aaccagccgc gcttttgaac atagccgcga 109800 ctacgttcta gccaaccccg gcgctgcgat tgttattttt cacgacgatc aatatttagt 109860 tgacagccag cctattgatc tgatagtatc taccactacg gacgcttatt tgtataaggc 109920 atccgaaggt aagcaagcgt caagacgttt ttgctaccat gaaagcgaac tgctcgcttt 109980 caccgatgcc cgcgcatgga ttaaaaacct gtgcgatcat ctggaactac caccagcacg 110040 aatttctagc gaaatgatga tttttgtgct tgacaaagac ggaagcatcc tgctaccatg 110100 tgacccttac gatattgata tcgaagaagg ggcaaggact ggaaattatc gttatgatgg 110160 cgagctagaa gaagtcgccc cggctgttac tgaaaatgta gttaacccta ataattttga 110220 gactggagca ttacaaatga atactatcaa atctaccgct accgctatcg ttgccgctaa 110280 caaaaacgct gctgtaaacg ctgctaaact ggaagccggt tctatcgtcc tgaaaaaagt 110340 ttccggcatc gccgccagca aagcgccgtt tatggttcgc ggttatgttg acacggcggt 110400 aggtcgtgtg gtaattgcta acctgctgaa ttcgcggtt agccagtatg cgccgaacaa 110460 ccgcaaagcg gtgattgcgg ctgacgctgc aatgcaagcc gccatgttgg aactggtaca 110520 aagtttcaat gtaggcgaaa tgattgacga agtattgaaa ggcgtcaatc tttctagcct 110580 gattgaaagc gacgtagcag aataattata tctagcgcct ctataatggg gcgctataat 110640 aattaccctg ctaactaact ggagacaaa aatggaacgg ctaaccgcta cttcgaagg 110700 cgaaaagatg acgatcgcta atgtatggca gcgcctgcgc cagaatggcg atcgtggcaa 110760 ttttgctatt ttcatcgagc caaaaaactt ggataatctg gctcgccaga ttgaccgccg 110820 ggactgctac ccggatacag atgatatgct gggaatcccc ttgcggatta tcggggtata 110880 tggttacggc tttgatatct gtattggtga tagtagcttt gaaatcgact gcgaatccgg 110940 cgctaccgaa atcgaagtat tcctaattaa tttaggctcg ctaacgtttt tggatacgcc 111000 accagcggaa ccggaaccgg aaaagctgga ggtgaaaacg tccgttattg ttagttcgct 111060 aactatggat gagctggcgg atatcgtgtc aacgtatgac gaaattcacg ccgacgcgat 111120 taaagagcta aacaaccggc ttgatacttt ccgcgataag ctgtaaaatc cctcccggcg 111180 gctaaatctc tagccgcccc cattattgga gaaaaccatg tttcatgtga aatcttgcgt 111240 accgggcatt aattatacgg tagaagcgga agaaggttta tatttagagg gcgggcgaat 111300 tgagtctcag gaagtcgccg ccgtcctcaa atgtgacact aacgtttgcg gtacgtcctg 111360 gactgacctt cattttttag gtcgtggaat tgacgtcgat tccctttcct gggaaaaggc 111420 ttgcgaacat gctgaaagta tgctaaacga ggatgattgg gatgatgacg acagcgacga 111480 aaaatacgcg aatgctgggg tagaagggtc gttttatatg tactggcccg gtcattcctg 111540 caacctcgtt aatggtggtt cgcccctcca ttccgttcta gagcgggcaa tctatttggg 111600 ctatatccag atagttgacg gaaaggcagt cattaatctg cgcgaattga aaacgtttat 111660 ctatatcccg gatgctgaaa ctatccttca tattgaggaa ggtttaaaat ccggttggaa 111720 ggttagcggg gtcgtatacc tatgatttac atctatgtca acaaatattt tctggcgcac 111780 tataaaacga tggaatctgt gatccagtac gttagccgcc agaatgcaag gcatattgat 111840 gaagtggcta ccctcaaaat cggattgcgg ggcgatgcta tcaatattag ctggcctttg 111900 ctaattctga tttgccgtga tcttgtagcg ggtaaacctg tttccgtttc cgcgctgggg 111960 gaatcttacc cgctttccga tgatctggat ctctatgatc tgctaaccaa atataaaact 112020 gaacgcctgt tttatcgtgg cgggtcggta tgctctagcg gcgaaacaat agaaacggta 112080 ttccgctaat agttagctaa ctaatgataa gcctgcttat aatgagcggg ctttttattt 112140 ttctgcgccc ctctagcaat ggcctttaat atggcctcta aatttaaagc gcttagaatg 112200 cgttatagag cgttctagag gcatattaga gtattaccgc tgccggaata tactgtataa 112260 atatacagta ggataacctg tggataagtc tggggatatc tcgttatacc gcttctcagc 112320 ttgtcaagtc aaaacgtgct gtggataact tgcccggaaa acaggagtgc agcataaaat 112380 aaacgcaagc attattttgc gttttcataa aaataatcgt ttgtctagtg gataactaat 112440 taaatttcat ttttgctttt tgtataagtt attaacaggg tctaaaacgc cctagaatcg 112500 attctaagcg gttttagata aaagccgatg aatcataccg ggtatgaggc gatctcctca 112560 cctcgtgtga aatagatttt gtcaacccct tttgaagaaa aatagtaaaa atattcgttg 112620 actttcttta aatattgtgt cacgtgcgcg cgcttcctat ctatcgtggg cctgttttct 112680 ctttacgctt ctttacaatt ctccgcttgc ggctaaacgc cattgtgtta ttatttatct 112740 cgtagggcgg caagacgaaa cggaaacacg aaagcgtttc actagtagtt aggtatctgg 112800 aggcggcaac attgccccga ctaccgcgaa aggaaaggcc gaaacgatat agtctgcccc 112860 cgccctacac tgctctttaa aaatttggga ttctataaaa agcgttatcg gctttcttta 112920 cttaataagg gaaccgaaaa catgttaaaa gaaaacgtaa tgagtagcga gattgtaaac 112980 gagtttacag tagcagacgc cgaacatttt atagagactt atttaaatgt gtacgacgta 113040 gatttagcct tcattcataa agacggccaa atc ​​113073 <210> 2 <211> 45423 <212> DNA <213> See more of Kle_F17 / 19 <400> 2 tgaggtaacg gcggagtttg ccgaacgatt ctcttcagcc ggatagcaca aagcgccctt 60 tacgggtgct ttttttggc ctatgatctt gagtcgataa gaggaggaaa tttatatgac 120 aactaacgtt tcaaaagagt ttgcgaactg cttgcaaaag ctgttccctg gcgagatctt 180 aacgctgatt gacattgacg ccacgaagtt cggcgggcag atctatcgat tccacaacga 240 gaatatcgcc tactcaaccg aagaactttt ggcagcggtt aacggtggaa cgctgacacc 300 gaaggcgatc acgtttcgcg gtgagcaata cggcccgcgc ccgttcgggga taggcgggat 360 tgcaatgtca agtgacggca cggtagaaaa gccaacgctg acggttagca atattgatgc 420 gcaagcgagt gctcttattc gcgcctacaa cggcctcatg caagcgaaag ttacggtatg 480 ggttttggtt aaggatttgc tgaaagaaga tggtagcgta gctgatggcg atttcggcg 540 gttcgtttac tatatcgaaa gacctaaa ggttgacccg cagaaggcca cgtttgagct 600 aacatccgtc ttcgatatgg atggattgat gatcccggcc cgacagactc aaaccgtttg 660 ttattgggcg caacgcggct ggtacaaaac cgggaaggga tgcggctaca acgggcaaaa 720 cggatacttt gacaatag gaaatagggt agatgatccc agccaggatg tttgcggtgg 780 ccttgtttcg tcgtgcagat tgcgattcgg caatgaggct ttatagtttg gcggttgcgc 840 tactgcaacc ttgaaaagcg gacggtaata tgttaactcc aaaaatcaaa atgcagatca 900 tgcagcatgc caaggaagtt taccctcatg agtgcgcagg gctggtgacg cagaaatcac 960 gcgtgcagaa atatcacaga ctcgataacg tttcgccaga tcctgagaac gaatcaatgc 1020 cggacgaac aagtacgcg ctggcgtcac tggagaga gccgatcgcc ttcgttcact 1080 cccacactgg cgacggggca accaccgttc cgagcgccac agatttatgc ttctgtgatg 1140 aatctggctt gtcgtgggtt atcgtgtcga ttccagaagg tgatatgcgg attatcgaac 1200 cgaagcgcag gccgctgatt ggtcgcccct gggctttggg cgcgtatgat tgctacggcc 1260 tggtgatgga ttttcacaag cgccacggcg tcacgctgac tgaccgccgc ctaccgttcg 1320 aatggtggaa gccggaatac aaagaggatc tttaccgcga ctactggcgc gaagatggat 1380 tcatcgaaaa cactggcgat cccgaagtcg gagatatgat tatcttccag cttcaggcgg 1440 aaaagtggaa ccacgctggg atttacgttg gaaacaataa tatccttcac cacgccttcg 1500 gcaagctatc acgccgcgat atctattccg gatggtatga gcagcacaag gttttaattt 1560 gcagacataa ggatcttaaa catggcatca catacaaaga cgattaaact ttccggctcc 1620 cttgggcgtc ggttcggtgt cttccataac ctagcggtcg attcagttgg cgaatgcatc 1680 cgggcgctat cctaccaggt ggaagggttt aaagccttca tgcagagcaa ggttggttct 1740 aacatgcgct ttggtatcgt ggcggacgga aaaccaatca gcacgaatga ctttgcaacg 1800 ttcgccgtag ctaaggagat ccgaatcatc cccatcccga aggccagaaa gaacggcgga 1860 ttgttgcaga tcgttatcgg cgctgcgatt atggttgcag ccttcttcac tggcggtgca 1920 tcactggcgg caatgggggc gttttcatcg gcggcgttta tggctggtgg cgcaatggtc 1980 ttgggtggcg taatgcagat gattgcaccg cagatgggcg gcaacatgcg agcgagcgag tcgccggaa acaaaccatc gtatgctttc ggcgggccta ttaacaccac ggcagcgggt tatccaatcc aattgccata cggttacaga ttggctggag gcgcattgtt cggttcagga tcttacgccg aagacaacaa ctaattaagc gattcgcttt ttggcctggg ggcatagcct ccgggctttt tgtcgtgtac aattgcaaaa ctattaacag gaggctaaac gatgactaat atcaaggccc gcaagggcgg ttccagcaag acgcgaaccc cggtagaaat gcctgataac ctgatctcga aagacaagat tagttattg ttggccgttt ctgacggtga agtagtggat gacttcagcc tgaagcagtt gcatttcgga ggcgttccgg ttcagaacga agacggtagt father agggcgttat tgcagaattt cgacccggca cgcaaacgca gagttacatc cagggctca gtgaatcaag cgctgaattc caggttgctc gcgacgtcac tcacaacacg ccgtacacgc ttaccgtatc gaacaaaaat ctgtctgcta ttcgctttcg cctgttatgg ccgcgcgtgc tgactcaaaa agataacggc gatatggtcg gatcggttgt tgagtacaag atcgagatgg cggtagacgg cgcaagttat cagacctacc taactggcaa gattgacggc 2760 aagaacacga ctggcggcta cgatcgcagc atccgcgtca acttgccgca agacttcacg 2820 tcgcaggttc ttatccgcgt aagccgaatt acgccggacg ctgacggggt taaggtggtt 2880 gatgccttcc aggttcagtc ctacgctgag gtgatcgatg caaaattccg ctatcctctg 2940 acggcaatgc tttacgtcga gtttgatagt gatctgttcc agaaccagat cccgacaatc 3000 tcgctcaaga agaagtggaa aattattcag gttccaagta actacgaccc aattaaccgg 3060 acatactccg gaacgtggaa cggaactttt aaatgggcgt ggagcaacaa cccggcctgg 3120 gttctttacg acctgatcat gaatcagcgt tatgggttag accagcgcga gttaggaatc 3180 ccggtagaca agtggtctct ctatgaggtg tcgcaatact gtgatgagct tgtgccggac 3240 aatcgcggag ggatggagcc gcgttacctg atggatatga tcgtgcagtc gcaggtcgag 3300 gcgttccagt tagtaaggga tgtttgttcc gcgttccgtg gaatgacgtt ctacaacggt 3360 gaaagccttt cgattatcgt cgataagccg cgcgatccgg tgtacctgtt tacggctgat 3420 aacgtcgttg atggcgtgtt cgttcggacg ttcccaagcg aaaaaacgat gtatacatcg 3480 3540 aaccctgacg ctgccatgcg gttcggtcac aacccaacaa gcatcacagc gatcggttgc 3600 accagacgga cagaagctaa ccgccgcggg cgctggattc tgcaaacgaa cctaagcgca 3660 accaccgttt cgttttctac tggtctggaa ggtatgatcc cttcgtgcgg cgatgttatt 3720 tacgttgcag atccgcactg gcaatcagcc ttcaacctgg tgttgtcagg tcgcattatg 3780 gaagtaacgg gaacgcaggt tttcctagct tttcgctgcg acgcgaaggc gggcgatacg 3840 ctgattctga acactgacga cggcaagcca ttacgccgaa ccattgccag cgtatcagca 3900 gacggaaaaa ccctgacgct aaacgtggga tataacttcg acgttgcgcc ggacagcgta 3960 ttcctgatcg aaagtgatca gcttgtagcg gaacagtatg tagtaacccg gattgaaaag 4020 ggtagtgatg acgacgaatt cactttcgcg atcacggcta cgcagtacaa cccgaacaag 4080 tatgacgcga tcgacaacgg agtaattacc gacgaccgcc caacttcggt tgttgaccct 4140 gattcaatgg gcgctccgaa agacttaaca attagttcgt tttctcgcat tgtccagggg atgagcgtag aaacaatggt tattggatgg tctgctgtgc agtatgcaaa actgtacgag gtgcaatggc gtaaggatgg cgggaactgg aacaacgttc cgcgaacggc gacaacgcag gttgatatcg aggggattta cgctggcgag tatcaggccc gcgtaaggtg cattagcggc 4380 gggaatgtag cgtctccgtg gtctgcgttg gcaaccgcca cactgaccgg gaaagtcgga 4440 gcgccaaaag gcccgataa cctttttgcg tcggacgatg agatctttgg cattcgcgtt aagtgggcca tgccggaagg tgcggaagac acggcataca ttgagcttta ccagtctcaa agcggaaccg atcaggatgc aagcctgctt actctgattc cttatccggc ggctgaatac tggcactcaa ttttgcccgc tggctatgtg aactggtaca aggcgagaag tgtagacagg 4680 atcggaaacg tttcaccttg gactgattac gctcgcggca tgtcatctac tgacgttaac gccatcacgg atgtgatcct ggatgagatc ctcgacagcg acgcaatgaa agaacttcag gagagtgcgc aggatagcgc ggcaaaactc aatgaatacg cgaacagtat cattcaaaac gcattagcga atgatggcga tgttagaata atgagaaagg agaatggcaa gaggaaagct 4920 gaaattaaac acgcagaagt tctcatagca atgagacgg aagccagggt gcagcaggtt 4980 aaccagatct cggcagagtt caacgaaat ctcaatgctg gattactca agttaacgag 5040 gcactagcca atgaactga ggctagggtt acgtcggaag agcactttc agcaaggactc 5100 ggagagaact ccgcggcgat agatcagaaa cttgactcat gggctgacgt taatggcgtc 5160 ggttccatgt acacgatgaa gttgggattg aagtacaacg gccaggaata caatccggg 5220 atggccctac agcttaccgc gcaagggaac aacgttgttt cgcaagttct gtttattgct 5280 gatagattcg ctatcatccg aaatgctgag tctggagcgt acacttgcc gtttgttgtg 5340 cagaatgacc agttttcat gaatacgcg cttattcagg atggttcgat taccaacgcg 5400 aagatcggaa acatcattca gtctacaac tacatcgccg ggcagcagg gtggatggtt 5460 aacaagaatg gtagttctga gttcagcgag gtaacagtaa gagggacaat ctacgcaact 5520 gatggttggt ttaagggtac tgtctatgcc gagcacattg aaggtgacgt gatgattgcg 5580 gagtcaaata cgataccatt caaggattat tcgaatattg atagtggtga ttatgagata 5640 cttaaaatca atggtgaaaa cttcgatagg agcatcgaca caaacctgat tgtccaagtt 5700 aactctacat tcaggaacac atttagagtt attatccaga caccaggggaa agaagagatc 5760 gagttctata gtatagatac tggcaacgac ggcggaacaa gatcctatgc actccgaggc 5820 ttcgatgttc cggcagcagg aaaggggcaa cagaacagga tcatagttag agtcacgcag 5880 agtggtaggg gtagcagtgt aaaaacattt actccgtggg ttgagcgaaa taatgggtct 5940 acagggacag aaagttcaga tggaacaata tctaacattg gttttataag agagaggaca 6000 tacatatcag catacagaaa aggaaataga atagtagtct agtgacaaaa gcgcccggtt 6060 gggcgctttc ttttaatatg aataatagat gtaaaaaagg ggccgaatgg ccccctttttt 6120 tctttgttg ccttgatatt cttgatatgt catatcgcaa caatgtcgat tttgttttcg 6180 gtaactacta ttgttttgga ctgcgccaac cgttgcaggc cgtagttggt atgaattttt 6240 gcgtattgca ttatttgatc ccaatgatta gataaagtttt attagcacgc catcgaaatc 6300 gacaactgcg ccatcatacg cgcgcataga aataactgcc gttgccgctc caggaggaac 6360 gatacctctt ataaagctcc cataaacagc ccaatctgca acgttaaccg gaaggttagc 6420 tgtagggcca gcaatctgtg ttccgttttg gctgtagaat gagaggttta tatttcccgc 6480 cgcattccca tttgcgccga tgtttacaac ccttgcccaa aaagaggtca tataatactg 6540 tcctggtttt acgttaaact gctgagttgc aaaaatgctc aacccggcga ttgaagtcat 6600 cctcatgccg tatgacccag ccttcgcata ctccgcagac acaacggcag tttgtgatgc 6660 agaaccagcg ttattaattg accatccggc attattaccc tgctcaaaac ctgaattata 6720 aacaggtgac aaagatcggt gtacaggaat atttccggct cctgatgcta tgtcgctagt 6780 acaagcattg cacattacag atccaggccc ctcaacaaat gcccgaacca tatccgatga 6840 gttctgttcg aatttgtatg aattaccagg aaatttaacc acattaaata ttatgaatgc 6900 gtttattccg acataaatta taggctgtgt ttgtgctaat ggctgattaa tggtcagtgt 6960 gctttcagaa agaataaatc ttgcaccagt gccagtacag gtaacgtaag gataccacac 7020 cgactgccca gggttttcta tgttacccat tccagacatt gatactgtag ccccatttcc 7080 ggtgatctga actcttgtat taagagttga tgtgccgtca aggccaagat tgaaagcgtc 7140 acatgcaatt atgatcggtg cgccgcttgc gtctgcaacc atcccaccat taaacgttat 7200 gctctcgcct gaatctaaaa gaccagccgg agcatagaat atcgaagtta ttcccttagt 7260 aataacgcac tcattgaact tgtacctcca ggtgctgttt gcacaagaaa ctacataatc 7320 gaagtcatgg aatgagcaat gttctatgca gcactgtcca ttataatatt gcgcgccacc 7380 gggcctgccg ataagtaatc catttttacc agcaacttta gcaccaaagc attcaatccc 7440 ggacagcttg ttgactgtgt tgcggtacag tggggctgga tatgttgctg atgaatagac 7500 gttaatggcg taagtggagg ttacaccaga aaaatctatt gacgcgcaac ctacatggca 7560 gactaaggat ataaatccca agtcaatttc cagcatttcc gaaccagtgt atttatactg 7620 tccagcagag ataactaaac ccaaacgttt tggtttggca tatgaaatcg ccttattgat 7680 ggcgttggtt atatcgccat ccgatggaat accgaaccat gcaacgttta ctgcaccatc 7740 aaaaatccgc ttccagcgct ttccaccagc agtgacaatg cagatacctc cgtcatcaac 7800 ggatgtactg tcagtttgat catactcaaa atagccagtt attggcttgt agcctgaggc 7860 atattttatg acttctatct tttggtttgg taatgtcggc tctattgtcc gtagggtcgc 7920 aatgtcacta cagcgcccta tgtttttagc accatttggc tgagccaata caaccagcgt 7980 tgactgttcg ccggaatcat caatgatctg ctgaatttca tcgcgagttg cttcagcgtc 8040 atctgctgcc tgctgacttg atgccgctgc ctgctgactt gctgccgctg cctgctgact 8100 tgctgatgct gactgctggc ttgctgatgc tgactgctgg cttgctgatg ctgactgctg 8160 gcttgcggca gcttcctctt ttgcctgttc taccaattct tgcaaagtgg ataaatcaaa 8220 gtccttaaag aaatctaccg cctcagcaat gacggtctcc tgcgattggt agtaacgcag 8280 agtttccgca acatcctgcg ccaggccgtc aacggtcagg gagtcgctta acaggatcgc 8340 gtaatcactg gacggaacag ccgcgccgtt tgtggagata gcattgattt gcgtatcact 8400 tacaaccttg ttaactaccg ccatttgaat cggggacgat aaaaacagga tcgtagcgcc 8460 tgggcgaatt agcgttagcg ctgattgcca gttagttcca gtcccggtga cgatgccgtt 8520 tgcgtccatc gcggctttgc cttgtctgta tagtgccatt ttaatacctc ttttggttgg 8580 ttgagtaacg aagagataat atcatcaatg aagcaataaa aaaaggagcc ttgcggcccc 8640 tttagttgtc aaatcagaac gggatatcat catcgaaatc catgccatga ttaccaccac 8700 cgctttgcgg cttcggttgt tgttgttgcg gcttaggttg ctgaggctgg ccccacccac 8760 cctgctgatt accgccgccc tgcgctggtt cgcgctggct gaattccagt tgcggcataa 8820 tcatttcatt gtggctgtaa gttgcgccgt tgtgctcgcg gtttacgatc tgaagtgtcc 8880 ggcaggttac gctaataacc ttgtcgacct gcaacgcttc atcgtaccac gcaatcatgc 8940 tttccttcgc gaagaagacg gcgcgatagt tggtgtatac agtttcgtcc tggccgtcgc 9000 gattgcggat cttcattcgc tccgacaggt ctaccgcata catcttccac ggcccgttat 9060 tgttgctgcc ctcttttacg taaggcgctt ttcggattac ccctgttaca acatgcattt 9120 ttattcctat ggggcggttt cccgcccggt taaattagtt gaaagatgag atatcttgtg 9180 cttcaggttc aggctttgat tctacctgtt ccggctcacg tttcgcaacc tcttgcggct 9240 ttccggggtt aaatccgttc gctggagtca cttttagttg cgcctggcgc ttcgtgatat 9300. cgtcttccgt aaccttccat tccgcaggcg tcaaagtctg tttagccagc ttgtaaatct cgcggagaga ttcgaggtct tcgcacgcat caatgcgttt tttgaagtct tgcggcttca 9420. tcttcgcgat ctctgcgtca tcgtcggcct gcttgatgcc cagcgcggcg gctaacgcat 9480 aacggcgagc gtatgaggtg gtcgatccat acgcttgctc aacggttttg ctgatcggca 9540 tgttgaactg aaacgccatc cactcgccag attcgtgcag aaacatcgtt tcaaggtgca taaccttgtc ggtgctggta tccatcattg attgaatgac cataattttg ttcttttcca gcgatggccc gatggcgtca aggatatcgc cgagattggc gtaggtgttc cccaggtggc 9720 tattcttccc gcttttcttc gcggcgacga aaccagactt cgctttgatt aacgcaacgg 9780 cgatatcttt gaaactttca gaagtacgca tgacaaactt tccttttcct gattggtgga gcgcactata tcacaagtgc gccgtagtgt ttagctattt gtgccgtata cttccggga catgtatttc acgaattgcg gcgtgggcaa cttcacttcc gctgcgttcg attcgtatga 9960 cggccaaacg tcatgcttga cgcattcggc gtactggtga atcacgcttt gatactgctt 10020 ccgcccgatc tcaatctgct gatccgttaa cgtgaacgcg agaggcgcga acggtgattt 10080 tttctcctgc gtcagcaggc ggacaacaac cgggcgtttt tcgttgtacg tcttaacgaa 10140 cagatcgcgc tgcaatgcca ttttgagata gtagcccagg ttgaacgcca gccgcccgaa 10200 gtcatccggt ttagaggtct gagtagtttt gtagtcggta atcaccacca cttcgaaaac 10260 ttcatccggg ttaaaccccc actctttgat gagttcggga tccgacacta cgtcaacatg 10320 atcgagtcga acctttaccg ggacgccgaa aatttcgccg aagattgaca actcgcgttg 10380 cgctgtatcc gattcaatgc acgccgcgtg ccgtgggtta gctagcatca cgtttcgcat 10440 ttgcacgacc gcatcgaaat cagaatactt gaccagcttt cggcctgagt tgatggcggt 10500 gctttcgtcg cagagttcga tcgcccacca cacgttaaca tcaatcccgg cgcggtatgc 10560 catttccagc aattcggggt agtccttgct ggacgtccca atcaggccac acgctttcaa 10620 tttcgcagac atgccgact tcgaagtaat cagatcctta acgtcgcccg gcgcggtcgc 10680 tcgcaggtac tcgccattaa attttgacgt ctcaagcatg caggtatggg agcaagtgcc 10740 gatgaaagc gcagctgttt cttctcgctc cttgtatttc cagtggggccg gggatgttgc 10800 gtaaatctcg ccgaggcttg agccgctaac gtatctggcg caccaggagt taggatcgtg 10860 gtaatcatcg ttggatatt ccgcgttggt gtagcctta aaaattgctt cagccattgg 10920 tatcactccg ttgtggtt cgttgcgtta agtataccca tcacgattcc aggtgcaagg 10980 caaaagtgc tattcgcaac tggtcaaaa atgagcgaaa tttacgtaag atttagtaag 11040 atgcatctta cgtgatttc tttatatatt tcatgcggtt aaccaaaatc ggtaagatgg 11100 taagatgcct atagtaaat attcacagga aaattgggcc gaatcgcaa agcaaatac 11160 atatacccgg agaaaatctt acctacacag aataatagag agagtaataa taatatata 11220 tttagatatc atatacttat acatacta tggttcgata ttggttaaaa atgagcgaa 11280 atttacgtaa gatgcatctt acctaatctt acttaaagtg gtcgaccagt tggtaagttg ctgaattttt ggcataaaaa aaggtaagac tgaaattatc aatcttacct aaaatttgtt cacgctttaa tcagagaagt gatgatgcca cgtagagctt tcgttcgaat ccgcctttga agttgtcatt gttcggcgcg atcactttca tatcgcgttc ttcagccgca gccagcatat cccgatcttt cctgctgcat acgactcgca attctcgctt gccttcgccg ccgccctttc 11580 ctttatatcg ataggccacg atctcaacgt tggacgggat gatgcaggcc cacacgtcac acttaaacga gcttgcgata ttgaagtgca tcgcctccac ccaagagcga gccaggtaaa ttggcccgtt accgccttcg ctctgattgg ttacgatcac cgatccgaaa gtcaggtcgc 11760 cagctaacat cttctcccgg ccctcctcat caatgaata gatgttgcag tattcatcgt ccggcccgtc ttcatgcacc agttgcatcg ggagcgcatg aaataactct tgcctgccgt 11880. tctcgtgggt ttttacgcca acccggtatg attcaacgaa ctcattttca atgccctcat aggcgttac agcgcta tcgacggcct ccgttctgtt cattatcgcc acgacgcgat cgtgatctgc catcttgcca tagtcgtacc cgttatcgcg agtaacctgc ttgttgcgct 12060 taactacgta ctcttgcgga actttgccca ggtatcgccc aagaatgttg atgcactcgc 12120 tatacggctc gccgctcaac ttcattaacc atccgatccc cttatcggca ccgcagccgc 12180 tgcaatatgc cccgccgtcg ccgcgattct ctaatttgtc agtccagcgg aagcgatcct 12240 tgccgccgca gttagggcag tcctggtgtt tgccgttgaa atagcgggag tggatgccgc 12300 aaatattttg tagcgcctcg cgccacatcc ccggcatgta tggcaaaacc tctttttcgt 12360 cgtaaaaatc cacgtcataa cctccaaata aaaaacgccc acgcgagaat cataacccgg 12420 taggcgttta gtagttagtc aaattgtgct atcggactac tcgaagcatt tcgcggcggt 12480 cgcagcggcg cgtaatcggc ttgccgttgc tgtcgaatct gagatccggg cggcaaaatg 12540 aagccctgaa gcctttgcag tcattgcggc ggtaactctt atgcaccagg taagcaccat 12600 cggctgaaat catgccgcgc ttaatccact gctgaacaac ctggatgcta acgcccaact 12660 cctttgctgt tttggcgatg ccgccgtatg cctcgatcac caattccatc cgggtagtca 12720 ggcccgccct tacttcatcc ttcaggacgt aatagcctgt cggcttcttg cggggcttct 12780 tatctttccc gcgtgacgtc ccgttatttc cgttcaaagt tctattgtct acctttgcca 12840 tttgttccat aatttaacct cacagcactt tttgttaaac acgataaaat ttgctctgta 12900 ttatacacgc aaccatgcga atgacaaatc aggattaacc atgctgacaa ttgaacaaca 12960 aattgaagcc tacgcagaca agatcccatc tattcaaaag cggttcacgg tcgggaatat 13020 cgttccgtac ccttaccagg ctgtcgcgta catcgagacg gcgaagcgga tcgcgaacta 13080 tgagcatccg ttctacatta aggcgtccgt atccgcaggc aagacaatca tgatcgccat 13140 gctcgcggcg cagtgcaaaa caatgaactt gcctatgatg gttctcgcca ggcaggccga 13200 gatcgtgaag caggattccg aagaaatcag caatcttgac gtgccaaact ccgtttattg 13260 cgccgggtta ggtacaaaag cggcttactt cccgatcgtc gtcggctccg aagggacggt 13320 ggttaatggc ctgtttaaga tgcttggcga ttacgtgcct tcagtcctgg caatcgacga 13380 atgccatcag gttgactggc aagacctcgc agaagcgatc gcgaacgatg aatccttga 13440 gtacatgagc cgcgcaaagg ataagccgta tcgcgtgaat ggtgagatcg tcgatgccga 13500 ttacccgtac aacgaaaagt tcgaaacggt tgaatttggc ggcgggcgca ctcaatacac 13560 catcgtaatc atcgagttaa tgcgccgttg ccttcagaag acggggcgag aacttcgcat 13620 cgttggctac actggttcgg agtttcgcgg cgtcattccg atcttacagg aagaaagag 13680 ccagccgggt ttctggcgcg agcagatcac ggacattaac acaactacc ttgtcgaatt 13740 cggttcggtg gttccgacaa tctttggtga cactgaggcc gacggcctgg gctatgacct 13800 gtcagagttt cacggctcca gtcaggacgg cacgcaggat tttagcgcgg aagacttgcg 13860 caagatggag aagaaaatcc acgattccgg cgaaatgact aagctgatta tgcagaaggt 13920 ggtcgagcgt gcgcagacgc gaaacggcgt gcttacact tgcgcgggcc agcgccactg 13980 taggaggcg gcgagctact taccgccgga cgccacatac gcgatcatca cagagaagac 14040 gaactcgaag aaacgcggtg aaattctgga caaggctaac cggggcgaga ttaagtacat 14100 cttccaggtg atggccctaa cgactggcgt taacgttccg ttttgggatt tttctgtaat 14160 ccttcgcaag atcggttctc tgacgttgct gattcagctt ttggggcgag gaatgcgatt 14220 gctcaaagac tggcagaagg aagcgcccta ttcgtgggtc aaagaagatc accttgtctg 14280 ggattttgcc ggaacaatgg atgacctggg ccagttatat ttcgatccga ttctggagca 14340 ggcgcagtat cagaagcgca agagcagcaa gaacggcccg aaaatttgcc cggtctgcaa 14400 aggcgaaaat agcgagtacg cccgccgatg catccacaag gatagtaacg gcaatcggtg 14460 cgagtatttt tggacgtccc agcgctgcga agaccaaaag gatccgagga cggggaaaat 14520 caaggtcaag gggtgccacg ctgaaaacga cattgtggcg cgccagtgca gatgttgtgg 14580 cgtgcagtta aaggatccta acgacaacct caccgggaag cattacacgc aaaatgactg 14640 gtatcaggtt gtaggctttg atttgggatt gacacgaaac caaagcggga tcatctttaa 14700 ttacgtgttg ctcaaccatg acggcgagcg tttcacggcc agggaaaaat tcttccctga 14760 gtcagaaagt gcgatttgcg gtaagttgtg gcgacaaaag gcagtcttcc agcatgttga 14820 tgatgctgta atgcgtggca agttgggcgg catgaaaaac gcccgcaaga tcctggagta 14880 tgcggaatac ttccgcgcgc cgaagcgcgt aacgcatcgc atcaacggca agaaggagga 14940 cattatttcc cgcaaagact ttggaggcga ggagtgatta cagacaaagg ggattacctc 15000 gaatattacg gcgggcctgt aaaggcttgc ccgcttgaaa aaatcgacca gatgaacagc 15060 gtttcgtggc tgcgttacga atacccggat tatttgttct ggcatacggt caacgagggc 15120 agcaagcaca aggcgagcgc cgttgctgat catcaaatgg gattgcttaa aggcgtaagc 15180 gatatccttg tgctgattgg gcttggcggc aaatacccgt tcgccgccat tgagctaaag 15240 cgccagggta aatcccaggc atcgccagtg agcaaggagc aacgggaatt cctagcagcc 15300 gtccgaagcc gtggcgggtt cgctgccgtg gcctatggat tcgagcagtt taagatcgcg 15360 tttgaagacg caatcaaata gcactttttg ttaaaactac ccggcgagag tcgggtaata 15420 ttctcccatc gaaacgaatc acggagtatt gaagatgaa aagatgctt gctttaattg 15480 ttctgtctct tggtcttatt gttgtagcg aaaaaccaaaacatatatt tgcggtaacg 15540 aagcattga ggttacgagc gattatatga aggtagttag cggcaagc gctggcgttt 15600 taattgatgg cattggtga aaccagtata agctgtttac gccacttgga acggcttttt 15660 atactgttaa caaaaacacc attgatta aagttagcgt ttatcagaac actctactt 15720 gcgaggttaa gtataatgg caaagacac ccaacaaa vakactcacg acgcgttcac 15780 gacgtttgaa cagcttgaac gtgaaacgtt cataggcaac gcgcttgcca ctggcggaca 15840 ctatcaggcc gtcaagcctg aataattcta acggtaacg ggcaccgct acgcaggtag 15900 caagacgccg gatatcgtcc gcgacaagtg gtcaccgat cgcagcctta ttgcttacat 15960 ggaggagcgt tacggccat acgacctgga cgccgccgcc gaggaagca acacgctg 16020 cccgaagttt tacgacgaaa agacagattg cctcaagcgc tggtggggga aaataagca 16080 cgtatggctt aacccgcctt actcgttccc ggatccctc gtgctgaaag ccattgagca 16140 gatggagcat gacaaccaga tcgatatttt gcttcccggc gacaactcta cggcgtggtt 16200 ccgtgacgca cagaaggcag cggctgagat catttggatt gtcgcagacg tcacggagga 16260 tgaagacgga acgcaactaa gccgatccgg tcgcctggcc ttcattaacg gcctaagcgg 16320 gaaaccagtc gacaacaaca acaagggcag cgtgattttc atcatgcgca aactcaagcc 16380 gggcgaggag caaaaaacgt tctacgttcc ggtaagcgaa atctgcccgt cattaactaa 16440 aaagcgcatg cgcaaacgtg ggatctgaaa aatggaacag cttgaatctt tcaccgaata 16500 cctccacatc gtcgttaccc tgctggacaa gtacggcttc atcggtacga atgaagaaaa 16560 gatggccttc gccgatagca tcgacggcac ttatcaagag ttcatggata acggcacgcc 16620 gcttgctgac tggcccgcga ttattgagcg cgaattgctg gattttcggg cgcgtgaagg 16680 tgcggaatat ttcgcgaaac agcactaatt gctaaacaat gcccgccgcg agcgggtatt 16740 attgttccat caaccaatca ggagctaacg ccatgaacac caaaaccatc gccgacacca 16800 tcaagatcgc acaagttaag ccgcacatcg tatcgcgtca cctgattaac ctttctcgcc 16860 tgtgcatggc cgactacgtg gcgaagcctt tagagaatgg ccttgatggt gaaatcggcg 16920 cgatctactt ccgagcagcc cacggcatcg aagaaattca catgtacgag cagatggctg 16980 aatgcttctg catttacggt gacgaataat gattgtcgaa tctggccgcg ctgccgtatg 17040 gcagcatgcc aaagaggctg gcattagtga cgatatcgtg atgatcgcaa agtatttcga 17100 tatcaaggat atctccattg tgttcaatgg gaaatttacc tatcttcatg agcgccccact 17160 gaagcgcacg cgtatagcgg tcgcgacgcg agcggaggcc gacgcgctga agatgttcat 17220 taatgagtcg aagcagacaa aaaaatatta caagtagcgg gagtgttgag aatgcgatat 17280 attgcgatcc tgattacggc ggtggttctc actatcgcaa tctttaacta cgcaattcaa 17340 ttgggataaa ttatgcaacc taaaatcaca gacgaagaat ttttagccgc ccgcgaggaa 17400 ggcaagacct accgcgagat cgccgaagag ttcggtatga acgttcgcag tgtggagcgt 17460 cgcggcgttc gacttgcccg ccaggggcat ttgcacggta acaaccacgt agcgaagcac 17520 atcccggacg ggttcggggt caaaggcaca tcgaccatga ttcgcgcgaa cggtgacgaa 17580 gttgtgcgct gggtcaagtc agaagtagac cgcgaccgca tgatcgcttt gatggagtcg 17640 gcgcaagcat cattctgcga agacctcccg cgtgccgaac cgcagccgaa ggatgaatcg 17700 aagtttttca ttgaggatca gcttgccctg tatccaatct tcgacttgca catcggcgcg 17760 atggcccaca agcacgaatg cggcgagaac tacgacacca gcacagccga aagggttctt 17820 aaccgcttct ttgactattc cgtttcggtt gccccgcagt cgaaaaaggc ggtcttgctg 17880 gtcggcggtg acttccttca cagtgacggc ctggacgcag taaccccggc aagtggtcac 17940 gttctcgacc aggacagccg ttacgcgaaa cttgtctatg ttgctattcg ctccttgcgc 18000 cgcgccgtgt cgctactgct gagcaatcac gcggaagtag aggtgcatgt gatcgagggg 18060 aaccacgatc aggctggtat gatctggttg cgtgcagcgc tcgcagcgtt ctatgaaaat 18120 gagccgcgcg tattcgttga cgtcagcccc gccattctgc atcgtaccgt gtggggcaaa 18180 acgatgctgg gctacactca cggacacacg atgaagaagg cagaaacgcg cctcgccgca 18240 atggctactg acttccgcaa ggagttcggg caatgcgact acatctacac gcattcgggc 18300 cattggcatc accagaccgt aacggagcac tcgttaggga ttgacgaagt acacggccag 18360 ttaggggcca aagacgctta cgccgctcgc ggtggttggc gctcctatcg ccaggcggcg 18420 gttatctgt acagcaaaga atatggcgaa gtaggccgtt ttatctaccg tccgaacatg 18480 taaccacaac ggccccgcga ggggccaaca aggaaaaaccg atgaataaaa atatctgcat 18540 tttcgatctc gatggcacgt tgtccaatgg caaccaccgt ttgcacctgc tgccgaaaaa 18600 agatctccat ctaaccgaaa gctggacggc gttcaacatg gcggctggcg gagacgctcc 18660 cattacggac acgatccggg taatggaggc catgcgcgca gcaggattta tcgtcatcat 18720 cctgaccggg cgaagtgatg aggcacgcga gatcactgaa aaatggctgg tgcaaaatgg 18780 cgcatcgcat tacgatctgc tgattatgcg ctccgcctcc gacaaccgga aagatacggt 18840 tatcaaagag gaaattttac gcaagattgg cattgagcga atcgttgcgg cgtgggatga 18900 tagcccctct gttattgcgc acttccgtag cctgggtatc acaacttacc aggtctgcga 18960. ctacggcga aaccttcacg accatctga atctcatgga gtagacaaat gaaaatgta atcatcctca acggcgcgcc tggcatcgga aaagacacca tcgcggaat catcgcccgc 19080. aaatgggagt atcggatcct tagtttcaaa tcaccgatgt ttgacattgc gcgcgccatc ctgggtgcaa ccgactttgc ccgcttcgcc gtccgttacc atgaccgcag ccggaaaga gtcaagtgcg actttttagg cgaccgctcg ccgcgtgaat tcctgattca cattagcgaa aatttcttaa agccgacgtt aggcaaaaag cagttcggca agttgctttg cgattccgtc gtgaattcgc ctgttagttg cgtcgtcagc gacggcggat ttgacgaaga agtagcac 19380. atcgcggcgc atgatagcgt aaacgtcttt gtcgttcgcc tgcatcgtga cggcatgact 19440 ttcgagggtg atagccgcaa gcatattagc cgcccggatc tgatttgcga aacttatcac 19500. gaattcgatt ttgatatgac gactggcgag ccggaagacg acgcgcaaaa aatccttgat atggtttttg aagggtatta atattctcta cgtaatgcct ttattatcac cgccttaaac 19620 tttgggagcc ttaacaggtt cccttttttt tgttctttat ttggcccaac gcatatatca 19680 tcgcattatc acttaactaa cagaggttgc atatcatgcg ggaattcatc aacgcggcaa 19740 ccaatggtag cggcggtgtt accctcgcag gctctgcgac cgggcaactt atcattgcgg 19800 ccattggttt atttttcatg attctattcg gctccttcgg cgcgtggttg cgctggcgag 19860 attcaaaggc gcttcgtgac gcgctggaag ccggggatat caaaacggcg gtgaagatca 19920 ggagtaaata acatggggat taaaacgcgg gttacattcg cggcggcgat ggcgatcgct 19980 gtcgccttcc tcccgaaagt ggaggacacg aaatacaagg tttataccga tatcgctggc 20040 gtttcgacgg tctgcgaagg catcacaggg ccggacgtaa tcaaggggaa aacctatacc 20100 cgatcagagt gcgatgctct tttggcgaag catatccagg tcgcaaagcg tgtggttgac 20160 agcaaaatca aagtggccgt gcctgacacc ttcagggcgt cgatgtacag cttcactttc 20220 aacgctggcg gcggcgcata ctccggcagc actatgctga agttgacgaa ccaggggcga 20280 ctatatgagg cgtgcgaaca gctttaccgc tggacgtact atcgcaaccc gaaaacgggc 20340 aagatggaaa aatcgcgcgg cctgcataat cgccgggatc aggaattcaa tctgtgcatt 20400 aaggatctaa aatgagtagc ttaaattttc agagagcact agcaatcggc ttcatcgtgt 20460 gggcggttgc actggtttcc ggttgcgctt ccagcgtccc gatcctttcc gatctggttg 20520 gcagcaagcc agatatgacg gcgcaagtcg gcgcggagaa cgtgaaacag gcggttggcg 20580 tgacgaacaa aacggacacg tcgagcaagc aggaaaccac cttcaaagaa tcggcagttg 20640 gcaaagttga cacttccaac aaaaaggcgg tgacgacctc cagcattcac gcgaaccaga 20700 ttacggcgga gaagatcgag atccgaaacg acgatagcgg aagcatgatt ccgtggctga 20760 ttggtgccgt tggggccatc atgttcgcgg tcagtgtttt cggtatgtgg cgggagcgaa 20820 aaaataaagg ggcgtaagcc ccttttttta tatgtaccgt ttgacgtgca gcagtgccac 20880 gccatcttca tcattcaacc catgttcaac cgtgtttgtc gcggccatgc cttgatatag 20940 caaggtgagc gcggcacgca agtaattttc tggtgtaatc tgattcacgc aaacaagcct 21000 gtgaacctcc g...

Claims

1. A pharmaceutical composition comprising any one of the following combinations of purified bacteriophage and a pharmaceutically acceptable carrier: (a) Bacteriophages with the genome sequence SEQ ID NO: 1 and / or their progeny, and bacteriophages with the genome sequence SEQ ID NO: 3 and / or their progeny; or (b) Bacteriophages with the genome sequence of SEQ ID NO: 1 and / or their progeny, bacteriophages with the genome sequence of SEQ ID NO: 2 and / or their progeny, and bacteriophages with the genome sequence of SEQ ID NO: 3 and / or their progeny; The combination of purified bacteriophages described herein has antibacterial activity against one or more strains of Klebsiella pneumoniae.

2. The pharmaceutical composition according to claim 1, comprising purified bacteriophage (b).

3. The pharmaceutical composition according to claim 1, further comprising one or more purified bacteriophages deposited under accession numbers NCIMB 42915 (F99 / 10), NCIMB 42916 (F27 / 12), and NCIMB 42917 (F95 / 13).

4. The pharmaceutical composition according to claim 3, further comprising purified bacteriophages deposited under accession numbers NCIMB 42915 (F99 / 10), NCIMB 42916 (F27 / 12), and NCIMB 42917 (F95 / 13).

5. The pharmaceutical composition according to any one of claims 1 to 4, further comprising one or more other purified bacteriophages having antibacterial activity against Klebsiella pneumoniae.

6. The pharmaceutical composition according to any one of claims 1 to 4, further comprising one or more other purified bacteriophages having antibacterial activity against Pseudomonas aeruginosa.

7. The pharmaceutical composition according to any one of claims 1 to 4, wherein each of the phages is present in an amount such that it provides a multiple of infection (MOI) of 1 to 10 when the composition is administered to a subject in need.

8. The pharmaceutical composition according to any one of claims 1 to 4, wherein each of the bacteriophages is in the form of 10 9 PFU exists.

9. The pharmaceutical composition according to any one of claims 1 to 4, wherein the composition is formulated for administration as an aerosol.

10. Use of the pharmaceutical composition according to any one of claims 1 to 9 in the preparation of a medicament for treating or reducing the occurrence of a bacterial infection in a subject in need, wherein the bacterial infection is caused by a strain of Klebsiella pneumoniae.

11. The use according to claim 10, wherein the bacterial infection is further caused by a strain of Pseudomonas aeruginosa.

12. The use according to claim 10, wherein the strain exhibits resistance to one or more known antibiotics and / or forms a biofilm.

13. The use according to claim 10, wherein the bacterial infection is a respiratory infection.

14. The use according to claim 13, wherein the bacterial infection is hospital-acquired bacterial pneumonia or an infection associated with cystic fibrosis.

15. The use according to any one of claims 10 to 14, wherein the composition is applied to the lungs as an aerosol.

16. The use according to claim 15, wherein the composition is applied again 4-8 hours after the first application of the composition.

17. The use according to claim 10, wherein the bacterial infection relates to the colonization or growth of bacteria on the mucous membranes of the subject.

18. The use according to any one of claims 10 to 14, wherein the subject is a mammal.

19. The use according to claim 18, wherein the subject is a human.

20. Use of the pharmaceutical composition of any one of claims 1 to 9 in the preparation of a preparation of a pathogenic factor for diagnosing a bacterial infection in a subject, wherein the diagnosis comprises: (i) Culturing tissue samples from the subjects; (ii) Contact the culture from step (i) with the pharmaceutical composition; and (iii) Monitor for evidence of growth or lysis of the culture, wherein evidence of lysis of the culture indicates that the culture contains bacterial strains known to be susceptible to the pharmaceutical composition used in step (ii).

21. The use according to claim 20, wherein the tissue sample is a tissue biopsy or swab collected from the respiratory tract of the subject.

22. An in vitro method for reducing or inhibiting bacterial colonization or growth on a surface, comprising contacting the surface with a pharmaceutical composition according to any one of claims 1 to 9, wherein the bacteria is Klebsiella pneumoniae, and wherein the surface is a non-biological surface.

23. The method of claim 22, wherein the surface is the surface of a medical device or a portion of a medical equipment.

24. The method of claim 22, wherein the surface is the surface of a surgical instrument or a portion of a surgical device.

25. The method according to any one of claims 22 to 24, further reducing or inhibiting the colonization or growth of Pseudomonas aeruginosa.

26. A purified phage assembly comprising: (a) Bacteriophages with the genome sequence SEQ ID NO: 1 and / or their progeny, and bacteriophages with the genome sequence SEQ ID NO: 3 and / or their progeny; or (b) The phage with the genome sequence of SEQ ID NO: 1 and / or its progeny, the phage with the genome sequence of SEQ ID NO: 2 and / or its progeny, and the phage with the genome sequence of SEQ ID NO: 3 and / or its progeny.

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