Therapeutic compositions and methods

Abstract

The present invention relates to the reduction of methane production in humans, in particular humans suffering from irritable bowel syndrome. In particular, the invention relates to viruses capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human, for use in a method of reducing the production of methane in a human subject. The invention also relates to pharmaceutical compositions comprising a virus capable of reducing the formation of methane in vivo in the human gastrointestinal tract, and the use of these pharmaceutical compositions in methods of reducing the production of methane in humans. The invention further relates to methods of reducing the production of methane in humans, comprising administering to the human a virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract.

Description

[0001] THERAPEUTIC COMPOSITIONS AND METHODS

[0002] FIELD OF INVENTION

[0003] The present invention relates to the reduction of methane production in humans, in particular humans suffering from irritable bowel syndrome. In particular, the invention relates to viruses capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human, for use in a method of reducing the production of methane in a human subject. The invention also relates to pharmaceutical compositions comprising a virus capable of reducing the formation of methane in vivo in the human gastrointestinal tract, and the use of these pharmaceutical compositions in methods of reducing the production of methane in humans. The invention further relates to methods of reducing the production of methane in humans, comprising administering to the human a virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract.

[0004] BACKGROUND

[0005] Irritable bowel syndrome (IBS) is a functional gastrointestinal disorder affecting the human alimentary tract with a worldwide prevalence estimated at 12%. The disease affects the female population approximately twice as much as the male population and is most common in people younger than 45 years of age.

[0006] Bowel movements vary considerably in their nature among IBS patients and this gives rise to categorisation into three main subtypes: diarrhoea-predominant IBS (D-IBS), constipation- predominant IBS (C-IBS), and mixed IBS (M-IBS). Faecal microbial composition differs among IBS patients depending on their stool quality and IBS categorisation. Stool firmness strongly correlates with faecal microflora diversity, there being a multispecies richness in people with firm stools which decreases across the spectrum of stool consistency reaching a minimum for diarrhoea afflicted individuals.

[0007] Methanogenic archaea (commonly referred to as methanogens) are the predominant biological sources of methane in nature and Methanobrevibacter smithii is the predominant methanogen in the human intestine. These methanogens may contribute 10% of the gut microbiome and are found in the colon but also can be demonstrated to reside in the small bowel, especially when small intestine bacterial overgrowth exists. An increased abundance of these methanogens is found in harder stools of C-IBS patients compared with diarrhoea of D-IBS patients and elevated production of methane is associated with constipation. Indeed, methane is a putative neurotransmitter acting on the cholinergic pathway of the enteric nervous system causing delayed intestinal transit time of ingesta and promoting segmental (non-propagating) peristaltic contractions.

[0008] IBS is an important health care concern as it greatly affects patients’ quality of life. In particular, C-IBS may negatively impact general wellbeing through abdominal pain, bloating and altered bowel habits. Given that C-IBS patients experience increased methane production due to an increased abundance of methanogens, a reduction of methanogenesis in these patients is one therapeutic strategy to achieve symptomatic relief. Previously, this has been performed using pharmaceuticals such as the prodrug lactone form of lovastatin, or by antibiotics such as rifaximin and neomycin. However, these drugs have drawbacks in that they disrupt the enteric microbiome through indiscriminate effects. Thus, there is a need for more targeted strategies for reducing the production of methane in patients suffering from IBS, particularly those with C-IBS, because elevated production of methane is associated with adverse symptoms including discomfort due to bloating, and constipation, so a reduction in methane production in these patients would improve their quality of life. The present invention addresses this need.

[0009] WO2024 / 013394 has a priority date before the priority date of the present application and a publication date after the priority date of the present application. It discloses solid and semisolid compositions for administration to a ruminant which comprise a substance capable of inhibiting the production of methane in vivo in the ruminant, such as a virus capable of inhibiting archaea that produce methane in vivo in a ruminant. Such compositions are disclaimed herein.

[0010] SUMMARY OF INVENTION

[0011] In a first aspect of the invention, there is provided a virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human for use in a method of reducing the production of methane in a human subject, the method comprising administering the virus to the subject.

[0012] In a second aspect of the invention, there is provided a pharmaceutical composition comprising a virus capable of inhibiting the formation of methane in vivo in the human gastrointestinal tract. The pharmaceutical composition according to this aspect may be for use in a method of reducing the production of methane in a human subject, the method comprising administering the composition to the subject. In a third aspect of the invention, there is provided a method of reducing the production of methane in a human subject, comprising administering to the subject a virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract.

[0013] The inventive viruses for use, pharmaceutical compositions and methods provide an effective means to reduce methane production in humans, particularly in patients with C-IBS who exhibit elevated levels of methane production. The reduction of methane production in these patients reduces symptoms associated with C-IBS, including constipation.

[0014] DEFINITIONS

[0015] The term “comprising”, “including” and “includes” as used herein means that at least all of the listed elements must be present, but other elements that are not mentioned may also be present.

[0016] The term “consisting of” as used herein means that only the listed elements must be present, such that other elements that are not mentioned are not present.

[0017] The terms “constipation-predominant irritable bowel syndrome” and “C-IBS” as used interchangeably herein define the subtype of irritable bowel syndrome which is defined as abdominal pain with hard or lumpy stools characterising more than 25% of bowel movements and loose or watery stools constituting less than 25% of bowel movements for an average of three days per month over a span of last three months using the Rome IV criteria for irritable bowel syndrome.

[0018] The term "virus" according to the present invention includes double-stranded or singlestranded RNA or DNA viruses, which infect cells of bacteria, archaea, plants and / or animals. The term “archaea” as used herein refers to any single-celled prokaryotic organism.

[0019] The term "substrate" according to the present invention is understood to mean any solid phase material to which a virus may be immobilised.

[0020] The terms “virus capable of inhibiting the production of methane” and “virus” are used interchangeably herein to refer to a virus capable of inhibiting the production of methane in vivo in a human.

[0021] As used herein, the term “about,” as applied to one or more values of interest, refers to a value that is similar to a stated reference value. In some embodiments, the term “about” refers to a range of values that fall within 25% of the reference value in either direction (greater than or less than the reference value). In some embodiments, the term “about” refers to values that fall within 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less of the stated reference value (except where such number would exceed 100% of a possible value).

[0022] The term “coated” as used herein with respect to a particular pharmaceutical composition refers to the presence of a coating on at least 10% of the external surface of the pharmaceutical composition. In some embodiments, the coating is present on at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 95% of the external surface.

[0023] The term “encapsulated” as used herein with respect to coating of pharmaceutical compositions means that the pharmaceutical composition is enclosed within the coating (in other words, the pharmaceutical composition is contained entirely within the coating).

[0024] As used herein, “location-controlled release” means that the virus is released at a particular location within the human digestive tract after it is administered.

[0025] DETAILED DESCRIPTION

[0026] The present invention relates to a virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human, for use in a method of reducing the production of methane in a human.

[0027] The present invention further relates to a pharmaceutical composition comprising a virus capable of reducing the formation of methane in vivo in the human gastrointestinal tract. The pharmaceutical composition may be in the form of a liquid, syrup, capsule, tablet, bead, granule, pellet, lozenge, suppository or pill, preferably a tablet. Even more preferably, the pharmaceutical composition may be in the form of tablet which is coated with an enteric coating. The pharmaceutical composition may be for use in a method of reducing the production of methane in a human subject, the method comprising administering the composition to the subject. The pharmaceutical composition preferably also comprises a pharmaceutically acceptable carrier.

[0028] The present invention also relates to methods for reducing methane production in a human subject, comprising administering to the subject a virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract. In preferred embodiments, the human has IBS. In particularly preferred embodiments, the human has C-IBS. This is advantageous because methane production is particularly elevated in patients with C-IBS, causing adverse effects such as constipation. Thus, the reduction of methane production in these patients alleviates these adverse effects.

[0029] The administration of the virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract is preferably oral administration or rectal administration, most preferably oral administration. In this way, the virus is directly introduced into the gastrointestinal tract and can reduce methane production there.

[0030] The virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract may be a virus which interrupts the metabolic pathway of methane production in the gastrointestinal tract, and / or may be a virus capable of disrupting the enteric microbiome. In preferred embodiments, the virus is an inhibitor of methanogens. In particularly preferred embodiments, the virus is an inhibitor of methanogens in the Methanobrevibacter genus. In particularly preferred embodiments, the virus is an inhibitor of Methanobrevibacter smithii. These viruses are particularly effective in reducing methane production in humans because Methanobrevibacter smithii is the predominant enteric methanogen, so administration of a virus which inhibits it will be particularly effective in reducing methane production in a human.

[0031] In preferred embodiments, the virus capable of inhibiting methane production in vivo in a human is a stabilised virus. For example, the virus is stabilised as described in EP 1496919 B1, by immobilisation to a substrate. This is advantageous because it ensures that the virus retains its ability to inhibit the production of methane while also improving its stability relative to a free virus.

[0032] In some embodiments, as described in EP 1496919 B1, the virus is immobilised to the substrate via covalent bonds formed between the substrate and the virus.

[0033] In preferred embodiments, the virus is immobilised via its head, leaving the tail free. This ensures that the virus retains its activity, because the tail of the virus is involved in its ability to recognise and infect specific bacteria and / or archaea. In some embodiments, the virus may be a bacteriophage.

[0034] Advantageously, given that methane production in the gastrointestinal tract is caused by specific prokaryotic methanogens, the use of viruses offers a particularly effective means to provide targeted disruption of the methane production process. Unlike other methane- inhibiting agents, which provide a less targeted approach and can cause toxicity to the human, viruses are specific to their target bacteria and / or archaea and thus do not have problematic side effects for the human. Thus, they provide an effective and targeted means for reducing methane production in humans and offer a promising method for alleviating symptoms associated with elevated methane production, such as those suffered by patients with C-IBS.

[0035] In a preferred embodiment, the virus is a bacteriophage which targets bacteria in the enteric microbiome. This is advantageous because the inhibition of these bacteria will, by extension, negatively impact the production of methane in the gastrointestinal tract. In another preferred embodiment, the virus is a virus which targets archaea which are involved in the production of methane in the gastrointestinal tract of humans. In a particularly preferred embodiment, the virus is a virus which targets metabolic processes leading methanogenesis.

[0036] It will be appreciated that viruses capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human can be isolated by the skilled person. For example, to isolate a virus which targets archaea, archaea cultures can be prepared in suitable growth media such as 119 Broth. Then, liquid can be harvested from a suitable source of viruses, for example, bovine rumen fluid, rumen faecal material, municipal sewage systems, or environmental samples from sources that provide habitats for Archaea, such as anaerobic soils including peat and peat bogs, thermal springs deep ocean thermal vents. Viruses can then be separated from the liquid, concentrated, and purified, for example by PEG precipitation and ultracentrifugation. The archaea cultures can then be inoculated with the resulting virus suspension and the cultures allowed to grow to allow viruses with specificity for the archaea to initiate a cycle of infection and amplification. These viruses can be filtered from the cultures and passaged repeatedly (i.e. , repeating the previous steps with fresh archaea) to obtain pure active viruses. Similar procedures can be followed to isolate other viruses.

[0037] Following a procedure as described above, a novel virus with activity against Methanobrevibacter smithii was isolated, which had a sequence as set forth in SEQ ID NO:1. The isolation method is described in more detail in the Examples and in WQ2024 / 013394 A1 , which is incorporated by reference in its entirety. In some embodiments, the nucleic acid of the capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human comprises at least 50% identity to SEQ ID NO:1. In some embodiments, the nucleic acid of the virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human comprises at least 60% identity to SEQ ID N0:1. In some embodiments, the nucleic acid of the virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human comprises at least 70% identity to SEQ ID NO:1. In some embodiments, the nucleic acid of the virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human comprises at least 80% identity to SEQ ID NO:1. In some embodiments, the virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human has a nucleic acid comprising at least 90% identity to SEQ ID NO:1. In some embodiments, the virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human has a nucleic acid comprising at least 95% identity to SEQ ID NO:1. In particularly preferred embodiments, the virus has a sequence as set forth in SEQ ID NO:1.

[0038] SEQ ID NO:1:

[0039] GTCCTGGTGTTTCTCCAAGGTCTACTTTTATGCTTTCATTTTTCTCTAACCATTTTGTGT

[0040] TATCTCCCGAACCGTTTAATTTTTGCCAGCACCCAATCAATACTTCTTCTACTGATTGTT

[0041] CGCTAGTTTCAAATGGTTCTAATTTTGGATTACAATCACATACTGTGAGATAATTGAAAT

[0042] CTACAAGTGTTGGTAAAGCATGAATATATAATTCTCCTCTTGGTTTTACATAGGGGAATC

[0043] T T GAAAAT T C AAAAAGT T T T GT T GT AT GAG AT GT T AAT GAT AC AAAAT C AC AT CCT T T T A

[0044] CTCCTACTTTTTCTTTGACTGTTTTTGTTGCAGTGGTTGTTGTTTCTGCTTCTCCAGGAG

[0045] TTAATTTAACTTTTTTACATTTACCATGACCCCATTTGTCTCCTCCACAGTATCCGCAGT

[0046] AATCTGCGTCACATCCTCCTTTTTTCATGCTGCAAGTTATTTCGCCTTCGTACACTTTTT TAGGATTGTCGGTTAATGTTCCAGTTTTGCCACAGTTTGGGCAGTAGTTTTTCCAGGTTT TTGTGTATCTTTTATATGCTGTTCCTGCACAACAGCCACATGATGGCATCATGTTTACGG TTATTGTGTTGGGATCTCCTGTTGTGACATTTTCAGTTACCTGTTTTTCTACTTCTTTTT CAACATATTTCATGAATTTTATTGACTTATTTTTGTAGTCAATAGTTATTTCGCTTTCAT CCTGAATTAGATTCGGTTGTTGGAAATATATCATTGAAGTATTGGGATCATCATCTGCTA TTCCCTGATTAGTTATTTCAATTCCTAGTGTGGATGCTCCTTTTATTTTTTTAATTTTGA GAATAGGGTATGATTCGCTGATATATTTTTCAACATCTAAAATAGTACATCCTGGGATTT GAC GT GGT GT AAT GGT T GGGAT T T T AT T T GAAGAGGT T GT AT T T T GT GT GT AC AT GAAT G GCATGTATTCGTAGTTTTCATAATCTTTTATATAGTGAATATTTTCTTGTACAAATGTTA AAGAAAC AGT GT CT AT T AT T T T T AGT T C AT T AGAAT T AAAGAAAAC AT CTTTTTTTGTTG TTAAATATTTTGCAAGAACATATATCAAATATTGGAAGCTTAGTCTTCCACCTGTAATTG ATAAGCTGTTTGGTGCTTTTGTTTGAGTTATGCAGTACGATTTGATTATTCTTGCTAATT GTATTATTTCATCCTGTTTTATGATGCAGTCAATATGGCTGTGTGATGGTAAATCTGCAC AAT C TAG AAT T T T AAGT GAAAT GT T T T C AGAGT T AT T GT CT T C AGT GT T GAT TAT T GGT T CTGCAAGTAGAAGTATCATTTGTCCCATGCCTATTTTAACATTGTAGACTTTGTTTTTTG ATGAATCTTCATATATTTCACTATGTGATATTTCAAACTTGTTTGGGAAGTATTTTTGAA CATCAAGAAGTACTAATAGTTTTTCAGCACAATCAATAATGTCTTGTATGTAAAAATCAT ATTCAAATAATGTTGGTGGAATATTATTGTTTACGTCAAGATTGTAAAATTGTCTAAGTA TTTTTGAATAATCTACTTCATCAGTATTTTCTGAAGAAATTACTGGCATTATACCTAATG CATTTTTGGTTTCATCACTACTATATGATAATTCATTAGTATTTTCTCCAATTACAACAC TTTCTACAATTTTATTGTGTGTTACTCCATAATTTGCAGGTTTTAGTAATTGTATTTCTT TTATAATGTTATTATTTTTTAAAGAATAATTTGTTTTAAATACTAATCCTGATTGTTTTT C AAT AGCT T T T AAT AAGT T GT AT T T T GT T AT T GAGCC T T GAAC AT T GAT T AAT CT T T T AC TAT AAT C AAT AC T GT C AAT AT CT GT T T T T GT AAC AT T GAAAAAT C CT T T C AGT AAT T T GT TTAGGAATAATTTACTGATAATGATTGTATTTCCATTTACATGTGAAGCATAATTGGGAT CTCGGATATAGAATGGTTTACAATTGTTTAATTCTACAATTATTTCTTCTGCATCTACAT CAAT AGAT TT GT CT AT AT AATCAAAGGT GCAT TCACT ATT AAT AACAAACAAACAAT TAT CAACAAAGATTTTGTTTCCTTGTTTGAATAATTGCTTGTCTCTTTTAACATTTGTAAGTT

[0047] TATGTGATAGTTTTAATGTTTCATATCCTTGATATTCATCAGTAATATGTATTTCTGATA

[0048] ATTCAGTATCTAAAAATTGAAGAATATTTTCTTGATTATCTAAAACAACAACCATACTTT

[0049] TTATCCTCCTTTTTCATAGTAAATTACATCAGTCACTTTACAATTGATACTGTTTTCAAA

[0050] CTTGAACCATTCACTATATTTTTCATAGAATACATTTCTATTTTCACAATCAATTTTAAT

[0051] CTTTGTTGTTTCAGGTAAGTTTGTTATAAACTCTCCTTCAAGTTTCATTAGTTGTCCAGA

[0052] T T CT GAT T C AAT T AT T T GT AT GCT T GT T GAT T C AT CT T C AC T T AAT T T AT AC AAAAT AAT

[0053] GTCTGGTTTTACTTTTCCAATTGTGGTTATTGTTCCTGCGAAACTTTTTCTTAATGGGTT

[0054] AATATTTCTACTTAAACCACTAGGGATTATTAAGTTCACTTCACAATCATAACCTCCAAC

[0055] C AT T GC T T C T GC AT C AAT T G AAT C T T C T AT AT AGT AAT C AAAAC AT T C T T C AG GT GC GAA AAAGAAACTAATACTTTTTAGAATTGGTGAATCTAATGTGTCTCTTTCAGGATACAGGTA

[0056] GTCACTTACATATTTCATGATTTCTGTTGATTCTTCAAAATCATCTCCGTAAACTCTGAA

[0057] TTTTAATTTGATTTTTTTATCATCAACATTGACTCTTATAGGGTATTTCCCATCTGCTCC

[0058] TTCAATTTTATATTTTTCTGTATCAAAATTAGCTCCACGAGGTATTTCTGTTTCAGGACT

[0059] CATACTTATAAGGAAATATTTGCAACTAACACCATTAATATAGAAATCGCAGTCTTCTCC

[0060] ACTATTTTTTGAATAATATACTGTGATTTGCACATTTTTCATTTCGACTTGGACTTGAGT

[0061] GAC AT T GT C AAAT GC AT C AT C AAT T T GT AAGAAAAAC C T T AAAT CAT T T AAT AT T GT GT T GAT T T CT GGAAAT GGAAT GC C GAAAT TAT GGAAT T T T CC AC C GAAT T T AAAGT T AGT GT C

[0062] ATTTCTAGTGATGTTTATGCTGTCTAAATAGTAATCTAATTCATCATCATCTAATGTTCC

[0063] AACACCACAAATTAGATTAATTGATTCTTCAACACTTATGTCTCCAGTTATTTCTATTCC

[0064] ATGAATCAGTACATTATTGTTTTCTAATCCTTGCCAGTTGAAACCTTCAAAGAAATGTTT

[0065] TGTAGATAAAGCTAACTCTGGAGGTTCAAGTAATAAATTTGCAGTGCTGCCTACATCACC

[0066] TAACAAATATCTTTTTGGAGATAAAGCCAATACTGGATATTCATAACCTGCATATCTTTC

[0067] TTTAAGTATTAAAGATAAATTACCAAATTCTACAGTTCCATAGTCAAAATCTATGAAATT

[0068] CCCATAAAAAATTAGATAGATTGGTTTACTTGCATCATATGTAAACTCACAGGATAACTC

[0069] TTCCCATATGTCCTTAACACTGATTTGATTGGTGTATGAATTTGTTGTTCCATTTACAAC

[0070] ACAAACTCTTAAGTTTTTCAAACCTTTTTCAATTTTAGAAGCATTTAACAGTCTTCCTAA

[0071] TGCTGAGAAAACATAAGTTTCACCATGTTCAAGGTTTGATAATGTGAATTCTGATAAATT

[0072] CAATATTGAATAATAGGATTTATATGAATCAGGAATGTTTACTTTCACTTTATCTGTGAA

[0073] AACACCTAATAATAATGGACTGGTCATTTCAATATTATGCTCTCCACTACTTTTAGAGTA

[0074] T AAAGGAAT AT T T GT AT C AAAAAC AAT GT T T C C ACT AT CAT CT T T T T GC CAT T GT GAT T T TGAAATTTGTAAAATATTATTTGATAATCCTGGAACAGGTTCTTTTACAGTTAATCCTCC

[0075] AAAATTCAGATTTAGTGTTAAATCTAATGGAGCATCAGTTTTTAAAGTTATGTTTAAGTA

[0076] TGTTAAATCTGTAACTGTATCTGTACTAATTTCAATTTCTTTGTTATTTGCATAGAAATC

[0077] CCATACTTCCAATTGGGGTTTTTCTACTACAAATGTTGTTTTTGTTGAATGGCCACTATT

[0078] GT T T T CAT C AAGT GT GAT T GT AAAC AAT CCT T C AGAT GT T GGAGT T AC AT AGAAT AT T AA

[0079] TTCTGCTTTACCATTTGTTGTTGCTTGTCTCCATATGCCTGTGGTCCAGTCATAATATGG

[0080] TGAATCATATGTGATTTCACAGTCTCCATTTGTGATTATCTTTATGTTTGCCGATATTTC

[0081] TTCTGAAGGTTTTGTTCCACTGGTTTCAACCATTGTGCATTTTACAGCTACTTTTTCACC

[0082] GTAACTAACAATATTTGGTTCAATGCTAATGCTTGGAATATAATTTCCAGTTGTATACAC

[0083] TACAATTACTCTTAAGTATTTCATTTCAATGTAACATGGGTTTGTAGATGTATTTTGTGG

[0084] TAAATCAAATTTAACATCAAAATTTGGAGTGTTAATGTCTTTTCCTTTTAGATTCAAATT

[0085] ATTAAATTGAACTTCATATTCTCCAAAATTATTTCTAGGTGGGGCATTACCTGTTTTTGA

[0086] CAGGTTTAAACTACTTAATATGATTTCTGGATTTGAAAAACTTCCATGTGCAGTTTGACT

[0087] TATCTGTGCAGAGTATGCTAATTTTCTATATGAATATTGTACTATTATACTCGTTACTTT

[0088] AGCT GAAT CT GGAAT T AAT C CAT CAT AT T T AAAAT CAT AT C C AC GT AT GGT T GAT GGAT G TTTTCTTGAACCATTTCTTCCAGCAATCGCATTTTTATATTTTGCTCCAGGATCTGCATA

[0089] ACCC C AAT GT CC C AC AGT AT T T AAAT CAT T T GT AAT AT T T T C AAT AT GAT C AC AT GAAC G

[0090] ATAAGGTATGTTGTTTCCTTCTATATTTTGAACTTTACTAGGATATCTTAAAATACTAGG

[0091] CAATTTAATAATCCCTCCATCATTTTTTTTAATTTTTAATAAAACCATCATACTCTGAAT

[0092] TAAACAATAAACAGTTTAATTGAAAGCTAATGTAATCTCCTTTACTTACCCCATCAGGAA AAGC AAAT T C AAC ACT C AC AT AT T T AGC ACC AGC AGGT AT T T T T GAT GT T T T AAT AGC AC

[0093] ATCTTCCTTTATTACCCATACCTTCATTAGCAATATCCATAATTTCCTGATTTTCAATTA

[0094] ATTTACCATCTTTATCGAAAAATACAAACCCAACAAGTATGCTTTCAATAGTGTTTTTTG

[0095] CATCATAATTAGATGAAAACACAATACTGGAAGCATCATCAGGTATGTGTACTCCTTGAT

[0096] T GT T CT GAT GGAT T AAAAGAAAAAAAGGAGAAGT T AAAT C AAC AT T CT C C GAGAGAT AT G T T T C T T T AACT T T C AAAGT AT AAC CAT T T CT AAAAGGAAC AACT T C AAAAAAAT C AC TAT AT GGGT T GAAAT T AAAAT C AAAAT GT T C AC AAAC AAAC AAACT AGAT AAACT AGAT T CC A TACAATATTGAATATCTTCACTAAATCTGTTAGTTAATGAAACATCTTTACTTAATTTAA AT GAAT AT AC AT T GT AAGAT T CCC AT T C AT C AGGT T T TAG AT TAT T T AAT GAAACC AT AC ATTTATGTGATTCAAAATCGTTTTTATCTTCCAGATATCCATGTTTGTTACTTATGATAT TACCGAAGATTTCACATTCTTCAGGATTAATTTTTATAGTTTCCATTATCTTTTAACCTC TCATTCTTTTAATACGATTGTCTCTTTTAATTAAAGCTTCAGCAATGCGGTCAATTAATT TGGAATCAGTTATCACTTCTTTTAAAGTTGTGATTATTGATTTTTCATCAATATCTTCTC CT T C AAC AGT AAC AT T AAGAT T AT GAT C T AT T GT T AAAT GC AC AT C AT C AGAT GT AT AAG ACACTAAATTATCAGCAGTAGTATTATTCCTATTATGTTCAATATTCTGGAAGTATGCAT T T T GAT T T GT T T T CAT T AAAT C AAT AAT AT TAT AT AT T CC AT CT GT AAT GAT T CC GAAT A CATCTTTAGAACTTCCTCCAGGACTTGCTGAACCAAAACTAACTCCAGAACCTCCGCGGA ATATTCCTCCACGTTTTTGGAATTGTGTCATATCATATATTTGACCATTAACCATAGCTG CTACATGGCCTATTCCATTCCATGATCCATGAATCATGTGTCCTGATAATCCCAACATAG AT GC T AT T T GAAT TAT CAT T T CT GC ACC AT C AAAAC AGT T GC AT C T T AC T T GAT CCC AAA CTTGCTGATTACTATATCTTGAATTATAATAAAATTCATAAGTTCCAGGGTTTTGGAATC CTCTTGCAGTTAAGATTTTTCTTAGAAGATTTTCAAAGTTTCCAGGATTTAATTGCGGTT GTTTTCCATCAC GGAAAT CTTCCACAT GAT AAT C T GAAGGT AT T T GAAT T C C AAGGAAC C ATGGGTCTGCTATCCTCCATTTGTATGCAGTATTCATTATTTTATTCACATTAGTATCAG GAATTCCTGCAAAACAAGTATTTGGATTAACACAACCATTAACTCTTGCCAAATCATTAA TTACGCTTGTTGGTAAATTATAATCTAAAAGATTTAACCATGATTTGCTTTTATTGTTTT GAGGATGTTTTACTTTAGAATTAGTGTCAATTACTCTGCCTTTTCCTTGTCCTAATCCTC CAGCAGCATAACTTGCTATTGTGCCTTTTGTGGATTTTCCAAAGTTAATGCTTCCACCAG GTAATGTTCTGCCTCCAATTAATCCTCTTGTTGAATTAGTTGCTGGGCCTGCAGGCAGTC CACTACTGAAATGTGCATTGGCTAATTGATTATAGAAACTTGCAATACTTCTATGAAGGC TGCTGAATTTATTGTATGATTGTGTTTGTATGCTTCCTGCAGCTGAAACAATATTGTCTT TCATTACTCCCCATGCATTAGTCATTTTATTTGTTACATCTATTGTGGAGTTACGTACTT GATTTAAGTTCTGGGTTGTTGTTGATTTAATATTGTTCCATGCTTGAGTATTTGAATTAC TCATGCTTGTTAATGCAGTGGTTACTCCATTACGTGTTGTTTGGAAGCTTGTTACTACTC CTGCAACTCTCATTTGAATAGCTCCTGCATTTAATGCTATTCCTGCAGTTAATGTTGAAA AGCTTGTTCCGATCATATTGTTGCTTGCAGTGATGCTTTCACTAGCTAATGATACTTCTT CAACAAGACCATTATATTGGTCTCCAGTTTCTTCTGTACCTGAATTATTATTGCTTAAAT CAACATTTGCACTATCTGTTTCAACTTCAGTGTTTGTTGTTACTGGAGTTGTGATAGTTG T T GAAAT AT TAT T T AAT GC AT T T T C AAC AT C AGC AGT AT T GAAT C CAT T TAG AAT GT T T T TTCCAACATTTTTACCTGCTTCATATGCTGATTTTCCTTTGTCTTTAACTCTTTGTAGCA TGTTAAAGAATTCTAGTACTACTTTTTCTTGGATTATTCCTGGTGAATGTATTCCCATTG CGGAAAGCATTCCGTCTACAATTTTTTTACCGATTTGCTTTGCATTTTCAACTAATTGTG AACCTGCAGATAGTATTCTTGAGCCTATGTTTAGGAATTCCTGGTATACTTTTTGAGGTA ATTGCTTTATCCAGGACATTACTCCAGATACTACTTGTGATGCTTTTGTTTTAGCATTAC TTACCCATTGTGCTCCTGCAGTTATTACTCTTACAGCAACTGCTAAAAGATACATTTTTA TTTTATCTGGCAATTGTCGGATAAATTGAACTACTCCTGTAAGGAAGTTTCTACCAGTAT T AAC T GC AT T AT T CC AC AT T GACT GAGC CC AACT AAC AAC AC GAT T AAT C AC AT T T GAT A AT AT T GT GCT GAT T GT T GAT T GAAT AGC T GCC C AAGC CC AAGC AAT T GC T T GT T GAAGAC T T AT T T GACC AGT AAT T AAC T GGC T AAAAAT T GT T GC C AGT T GC AT GAAT AAAT T GAAT A AT GGAGTT ACAAT ATC AAGT ACAAGTT GCCAT GCT GGACCAAAT ACACT T AAT AAGAAT G ATGCAATTCCATTGAAAACATCCCATATTATGGTGAGTATGTCTATTGCAAGTTGAACTC TAACTTTGAAACTTTCAAATACTGCTCCTATTGCATCGATAACTGCTCTTGTTCCATCAA CTTTACCTTTTGCACTTTCTGGGAATATTTCATCCCATATTCCTTTCAACCAATCTACAA CT GGT T T GAGAGAT T C AT T C AAAT C AGACC AT GC AT C T T GAAT T GCT T T GAT AGT T GCT T TTACATCAGGATGATTTATGAATGCATCCCATAATCTACCAATATTATTCTTTATGGCTT CGAGCATGGTTCCTACATCTTTCCACCAACCAAAAGCTTTACCTACTTCATATATTGCTA CTGCTATTGCAACTAATGCCACAACAACCCATGTTAATGGATTTGATAAAATTGCTCCTG CAACTGCTAAAAAACTAGCCGCCAAACCCATATTTGCTATTTCAACACCTAATGCTCCAG CAATATAAGTTACAAAACTAGTCGCACTAGCAGCTATTGCGGGAATTACTGAACCAAGAA TTACAATACCCAACCCAACTAAAGCTGTTGAAAGTACACCAATAGTTACTGCTGCTTTAG CCCAATCGGGCATGCTTGACCATGCTGATTGTAATGCTTCAACCATCCAAGTAATCATTT CAACAACTGTTATAATTATGGGAGTTATTGGGACAAGGATAGATTGTATTAATTTGCTTC CAGTTACTGTTAATGCTGCCCAAGCATCATCTAATGTTACAATTTCTTGTGCGGACTGTT CAAAACCCATATCGCTAAGTGTTTTGTTCATTGCTTGGAGTAATCCAGTTTTATTATTTA TATCTCCATCCCATCCATTTTTCATGAGCATGTCTTGAGATATGCCTATTTCCTGCAGTC

[0097] TTCTGAATTGGCCATCCATTGCATCTGATACTGCAAGTATTGCATCTTCTTGTGTTCTTC

[0098] CTTCTTTTACGAAAGCTGAACTCATTACTGCAGTTGTTTTAGTAAGTTCATCCATTGAAG

[0099] CTTTAGGAAGTTTTAATTTCACTCCCATTTCTAATGCGGCTGCACCTACTGCATTCATAT

[0100] CTACTTTTCTGAAGCTATCTTGCATTTTGTCAACAGCCCCATGGAATTGTTGTAGTTCTC

[0101] CTTCAGTCATTCCTAATCTTTGACCGAATCTTTCAAAACTTGCAGCTGCATTTATTGATT

[0102] CTCTTGCTCCTTGAACCATACTGTTTACAAGGTCAAAACCAATCATTCCTACAGTCATAC

[0103] TTGCTGCTGTTCTAAGAAATCCAAGACCTCCACCTGCAGTTTTAGCTCCGTTTTCAAGTC

[0104] CTCCGAAACCAGTTCCTGTTTTTTGTGAACTTGTTCTTAAACCTTCTAAAGCTAATTGTG

[0105] CTTCTCTTGCATCTCTTGTAACATTATCTAAACCAGTAGTATGGAAGTTTAGCAATGAAT

[0106] TTACTTGTTGAAGTATACCAATTAAAACTAATAATATTGCTTTTAGTAGTTCTGCTGATG

[0107] CTGTTGTTCTACTGAATCCCGAACCATCAATTGAATCTAAGGCTGCATCTGTTGAATGTG

[0108] CACTTGTTCTAGCTCGATCTAATGCCATGTCTAATTGTGCTGCACTTGATGAAGTTGCTG

[0109] TTAATGTGTATCCAGTTATACTATTTATTCCACTGCTTAAACTAAGTGCACTGGATGAAG

[0110] CTTGTTTTAAACTTGAAGATAATAATCTGGTACTATTATTTGAACTATTCATCGAGGAAG

[0111] ATAATCTTGTAGCACTAGATGATGCTGTGAGTAAATTACCTGAATTAATTGTGTTGATTG AAGT GT T AAGT C TAT AT GC ACT AT T CAT C GAT GT AT C CAT T GAAGT GGT T AAT AGT T T T G

[0112] CAGAATTAGATAATTGATTAATTGTATTGGGATTGATTGAATTTAATGCTGTTTTTGTAG

[0113] AATTAGTGCGATTTGTAACTTCAGTTAATTCTTTATCTAACTGATTAGCAGAGTTTTTGG

[0114] CTTTATCCATACTTGAGGAATCAAATTGTCGCATTGCATTTTGAGCTTGTTGTGCTGCTT

[0115] TTTGAACTTGCTCCATTTGTGCTTGTATCTCTTTTAATTGAGCTTTAACACGGTTTTCCA

[0116] GTTCAATTATTAACTTTACAATATTATTGCTCATTATTCTATTCCTCTGCTAATTTCTTG T T T T T T T C T T CT GAG AC GGT CT T T T AGAGAAT GT GAAGC AT GAT T T GT ACCT GAAGAT T G

[0117] TTTATCCTGTTTATCACGAACTAAGGGTACTGCATGATTCAAGAATAATTTCTGCCTAAT

[0118] AGTTAAATCTGCTTGGTTTTCTGCTAACATATAACCGATGTCTTGAAGAGTTACTACTTC

[0119] AATAGCTATGTTTTCATTCTTCTTCACGAAATTCTGCTATATTATTATCCATTGATTTAA

[0120] GGTTATCTAAACCACTGAAAATTAATGCTTCATTTACAATTAAGTCTAATGTTCCTGCTT

[0121] TTAATTGTTCAAGGTCTTTTTTGGTGAATTTATCAGGATTGTCTGCATTGTCTAATACTT

[0122] TTAATGCAAGGTTCATTTTTGCATCATATTTCTTTTTTTCCATTGCTGCGAGACTGGTTT

[0123] TGAATTTTGTGGTGTCTTCTGCAAGGCTGGTTTGTTTTGCTTTGATGGTGGAAATCATGT

[0124] CGAAGTTTCCTACATCACGGTATGCTGATGTGAATTGGTTGTATTCAAGGTCGCTTATTT

[0125] CTCTTATTGCTACTTCATTATCGTACATTGGTAGTTCGATTATTTTTGTGTTTTTGATTC

[0126] CGCCGATGATCATTTCTTTTGTAAGCATAAATATCACATTTTTATATAATAATTTTTTTT

[0127] T GAAT T T T AAAT AAAGT AAT T GAAT GAG AAT AAT CATATTCTCATTTCAT T T AAT C T GT G

[0128] TAAACAGTGATTGTTTTTGTTTTACTTGCACCATCAATTAAAATATGTGCTGGTTCTACT

[0129] TTAGTTGCAGTTGGTTCACCAGTCAACTCTATATCATACACTCCATCTTCAGCAGTAAGT GT TACT T C ACC AT TAG T T GAAGAT T C AGO AT C AAC AGT T GT GGAT OCT T T CAT T AAT GAT

[0130] ACTGTGTAATCAGTTGTTAATTTAGCACCTTCCTTATCAACAAGTTTTATTGTTGCATCA

[0131] TAAGTTGGGTTGGGATCACTGTCTGCTTTTCTTGCAAATAATGCTCCAGCTAAATTTTTA

[0132] T AAT GGT C CAT T GT T GAT T GAAGT GT T T T T GAT AAAC AAAC AAT ACT T AAT T CAT AT T TA

[0133] ACTGGGTCTGCATTCATTGTTGCAGATGGTTGGTTGATAATACACATTGGACAATATAAT T C GAAT T T T T T AT GAG CAT T T CT AAT GT AGAAC AT T AAT T C AAAGT AT T CT CC T GC AAGG

[0134] TATCTTCTTGGTCCTTCCTCACTTCCCCAGTACATTTCCAAGTATTTTTCATCAGTATCA

[0135] TCTACTGTTAAGTTTAAACCAATTTCTCTTTTAGCTGCTCTTGGTATTTTACACATAAAC

[0136] CTTGAGCCCATTCCTCTTGCATCATCAGTATTAACATTGTTTTTGATTTCAAGACTTATT

[0137] TTATTGGTTTTACATCTCATTTCAGTCCATTCATCTTCACCAAATCTTCTCATTTTAACA

[0138] CTGTCTATATGGTAAAATGATAATGGTAATTTTCCAAAGTTAAACTCTGATAATTTTTTT

[0139] AATGGTTGTTTACTATTGATTTTTGATTTGATTTCTGCTGATGCAGTTAAGAATTCTGAT

[0140] TCTACTTCAATACTTAAACTGTCCATCACCATACCTAAAATTTCCATCTCGTAGGTTGCA

[0141] CATCCTGCCATCACAGTATAAGAAGGTAAGATACTACTATTTGTACCGTAGATAATATCT

[0142] CCTTCTCTTCTTCCCAATACTGCTTCAAGAAGGTATTCTAATGTTTCCATTTGTAAGTTG

[0143] CAACTGAAACTGTTTTCAGGTACATAGTAAGCAGCTACACCAGATTGATAATCCCTACTT

[0144] ACTACTGAATCATATTTTGTAAATTGCTCTGGTGGAGATACATCCATCTCTGATATTTCA

[0145] ATATCAATAGATTCTCCACTAAGGTCTTTTTTTGCAAATTCTCCTTCTTTTACTAATCTT

[0146] AAATATTTTAGTTCTTCTGCGGCTGTTACCATATTCTTTAGCCTCCATCACATTTTGAAT

[0147] TCTTAATTTTAAATCTTATTTGGAATGTTACACCTGCACTATACACATTTTTCTTTTTAC

[0148] CCAGACCATATGGAACCCATCCGAAATCCTTGGTTTTAATATTGAAAAATTCTAAGTCTC

[0149] CAAGTAATCTTGAACTTATTATTCTACGTTCTGCCTGTGAAACTATACTTGTAGCTTCAT GAATACCTATCTTAGGATTTTTATTGTTAAGCACCTGTGAGGATAATATTATTTCTCCTT

[0150] C AAAGGT T T GAG T T AAACC AGT AT C AGT AAT AT T T T T AAAT GGT T CAT C T ACC C AT AAT T CCAGATAAGGTAATTCTGGTGTTCCCAGACTTACACCCATAAGAGATAAATGTTTAATAG TGCCATCTTCAATCATTTCAGATAAGACCAGTTGGAAACCTTCCATTAGTTTATCATAAG CTTCTATGAAATCTTTCATAATCCCACCTCTTTTAATGTTTCAGACACATATTTGTTAAT TTTTGATTCAGTAGTTTTAACTGCATGAGGATAATAATGGTATCCCCTGAATGCTGCTAC TTTTTTAGGTTTAGGGACAAAAACATCTTTACCAGATTTATCCACCCAATGCAGGGCTTT T T T AT T T T T AGC T GAGAAAC T T CC AC GACC AT CAT GAAC AT AT T T T T C AT AAC CT GAAGG TTTTCCTCTTGCTTTTATAACATACATTTCATCTGTTTCCCTTCTTGCAGTTATTGCAGT TAAAAGATTGTAACGATTGTATTTTATCCTACTTTGAAGGTAAGTTTTAGCATCTTTACT TGCTCCATAACTTACCTTTTTACCTAATCTTGGAGGTACCCGTAATATTTTTTCTTGCAG TCTTTCCCATCCCGCATAATCAAAATCAAAATGTATCATAAAAAATCATCAACTCTTATA AAAAAT T AAAAAAAAT GT T T AGAT T GAGAAAAC AT GAAT CT T AGAT T T T T T AAT GT AAGG CTTCATTCTTTTTTCAATGTCTTCTGTGAAAATATTGTTTACTGCCTCATCAAAATCAAA ATTTTCATGATCAGTAATTCCCAAGTCCTGTCTTACAGCATGGCTTCTAATAATGTTACT GGT T AAT T C AAT AAT GAT T T GT AT TAG AT CAT C AGGAAT T T T AT C T GGT AT AT CAT AT T T CTTTTTAACCCATGATTCCATTGCAGTGTAATAATCATTAATATAATTATCTAATTCAGT AGTTTCTAAATTAAATAATTCTTCAGTATTTTTATTAGCTCCACTGTACTGTTTGATTTT TTTTAAGGTGTTTTCATCCAACATAAATTATTCCTATGTTTTAGTCATTGTAATGGTTAA TGTAGTTGAATCATCACCTACAGTATAATTACCTGTGCTTTCAAAGTTTTCATAACCAGT ACATGTTGCACCATATGAATAAACTCCATAAGGAACATTACTTATTGTACAGCCTCCAGC ACTACCAGTACCGTTACAAGTGTAAGTTTTACCATCAGTAATATTTGTTAAAGTTACAAT AGCTCCTTCAACAGGATTTGTTCCATCATCAACACTTACACCAATATTTTTAGTACTAGT T GT T GT GC T GGAGT T T AT GC T T CC T T T AT GT AT GC AAGAGC AGAT GC AT C AGACC AT GCA AACTGAATATCAGCATACATTGTAGAAGCAATATAATACTTGTTAGATTTTAAATCAAAT TCAGATTCAACAATAATATTATCAGGATCCGCCAACCATTGAATGTTTTCTTTATGAGTT AAAATTACTGGTTTTTTAGTGAAACCATTACGAAGAGTACTGAAAGCTGGAATTGGTACT AATGGTACTTCCTCAATAATGATGTCCCCATCTTTAGTAATAACTACATCATTAACAGCA TACTTGTCATGATTATCTGCTACATATCTCCATACTGCTCTTTTAAAACTGTATGGTACA AATGCAGCTACCCCACCATCATTTTTATATTTATCAGGGAATAAATCCAACATTCTACGG AATTCTTTTAACGGATTGGAATCTGTTGCAGTTAAATCTATTGTTTCTTGATCTATATCA GAAT CAT C T T CT AAT T T T T T C AAGAT AC CAT CAT T TACT T TAT AT GAAGT T GC AAC AT TA GATTCAGTACTTGCAGTATTCCCATAGATTAATGTTCTTTCTAATGCTCTACCATTTGCA CTACCAAATTGACCAGTTAAAGTATTCATGAAATTCTTTTGCTCAATACTGTCATATAAT ACAGTTCTGTGAACACCAGTTAAAGCACGATATTCTTCTGCATCAAAAGCCCTATTAGTG AATGTTGGGTCCTGTTCATTGCTTAAAGTTTGTGGTGTACCACTGATTCTTCCAGCTTCC AAT T GAAT AT C AAAAC T CAT CAT GT CC AAT TCTCTTTTAT GAT T GT GAGT T GGAAC AACT TTAGTCTTGTTTAAAAATGCTGTTTCTTCTTGAACAGCTTGCATGTATTTATCTGCTTTT TCCGCTTGAAGAACTCCTTTACCGAGTTTCCCACTTCCTTCTCCAATATCTACAAATTTA AGGATAAAATCTTCATTATGCATGATTTTATCAGCTAACGCTTGTTCTACTTGTTTAATT GTCATTTTATATCAAACCTTCTCATAAAAAATTTAATTATTAAGTATAATATCTTCTTTT TTTCTACCAGGTCATTCCATTTGAATTCCTACCCATTCTCTCAAGGAATGATTTTTCAGA AGCATTGCCTTTTGCAAGGTCAGGATCAATACTTTTACTAACAGTAGCATCTGGATTAAC AGCACCATCAGCAGTAATAGTAGTATCCACTTTAACTTCTCCTCCATCATCATCTTTTGG AGGTTCTGATTTTTTAACAGCTCCAGGAACAGGATCCTCTTTTGGAGGTTCTGGTGGTTT TTCCCCACCTTCAAGTTTAGTTATCCTTGCATCCATAGTATCTAATCTTTTATCGATTTT ACCTAGTGCTTCTAAAATTTTATTTTCATCTTCCACAGGTTCATTATTTTGTGCTGGAGG TTCAGTTGGTTCTTGAGGTTCAGATTTAGCAACTAAACCATTAATTAATTTTTCAAGAAT ACCTTCTGAAACCTGCACATTATTTTTTCCATTGTCTTCTTTTGTCATAACTTCCACCTC TTCTGGTGTGATATATTTTTTCACAAATTCTTCATCATCTTCATATACTTCAAAAACCGC C AT AGGAT GT GAT GGT T GAT C AAC AAT ACT GAT T GT T GAT GGT T T CC AAT CT T T AAT GT C TTTAAATTTCAAAGCCATATTTAACCTCCTAATCCTCTACTCATTTGAGCAACACTTTTA TATGGTGCAGCCAGTATGCTGAAACCATTGTATTCCCCATCTCGGATTGCTTGTTGTATT TCCTCATCAGTTACATCAACAGAAATAAACCAGGATCCTTTGGGGTAAGTGTTTCCCCTG AAAGTGGTTGGTGATTCAAGAATATATGATTCTAAAATTCTTCCTACAGGTTGGAGTGTG TGCTGTACATCAACTCCTAATCCCAGGCGATTGTATATTAATGATGCTTGTCTTATTGTT TCTTCATCCAGTATGTCTCCTGTTGCATCAGGGATTCCTGGTATGCAGACTGCACCTTTC ACAATCATAGTTTTTTTTGACTCCCCATTTTTTTATATTGGAAAAAAATTAGTTTTGTTG GAGT T T T T T T T AGT AAAAAAGAAAGGAGGAT T T T AAAAAAAGAT AAAT GAT GT T AAGT T A

[0151] CTAAAATAAAAATATTTTTTTATCCCCCTTCAATATATAGGAGTGGAAAAATAGAAAAAA

[0152] ACGAAAAGTTAAATAAATTTATTCAACAATTTTGACCATAGGTTTAAATTGAGGATATTT

[0153] ATCAAGAAAATCTAAAGCATATTGTATATGATATTCCCTAGCTTCTTTGTGTTTCCCTTC

[0154] ATATTTTAATCTATGAGTTACAGCCCGTTCATTCTGATCCAAAGATTCAAACTCCGGATG

[0155] T T C AAT AC ACT GAAT T AAT C T T AAT C GAT T AAT AT T AGGAAT AAC T T C AC GT AC AAT ACT

[0156] CATTAATCTATGAGGTAAACCTGAAGGTGTTGTAATGAAATCTCTTAATTGATCCTCAGA

[0157] C C AAT T T T T AAC AAAT T C AT AAT T AGGAAT GAAT TTTTCAACCAT GAT AAT GAAC C C C AG

[0158] TATTTGTTAGTATATTATATATAAAACTTCCTATATAAAAATTACGATATCATTTCCCAA T GAGC AAT AAAC T CT GAT T T T T C AC GT AT T AAT CT T AAAT C AAC AT T T T T AT C AAAC AAA

[0159] ATTTCAAACTCCTGAGGAGTTTCAGTTATTCCTTCAATATACAGAATTTTAATTCCTTTT

[0160] GGAATGACAATATAATTAATATACTCCCCATATTCTTCAGGTTTAACATTTTCGGAAATA

[0161] GAAGT T GAT AAAAAT GAAT C T GAAT GAT AAT TAT CT C CT T C TAG AAAT T T AGT CAT T GGA CTTTTTTGTCTTCTAACTACAACAAGACTTTCTTCTAATATTTTACCTTTCTTAAATACT

[0162] GATTTTGAAGATTTAATTGCTTTATTAAAAGAGATATAATATTTTGAATTAGCTAAATCC TGCTCCCATAATTTAGAATATTTTCTTTTAATACGCCACCTCTTAACTGGGTTGAGCTCC CCTTTTATTTCTCTGAAATAATTATTCAAACTATAATATCCTTCATCAGAATATGCTTTT ATGAATTTATACTCTTCAGAAGTCAAATCTTCTTTTTTAATTCCATATTTATATTCAAAT GCCAATTTATCTTTAGATGATTCACCAACACCATAAACTGTTTTATTACCAATTTTCACT GCTTTATTTTTAGCTTTACTAAATTTAGTGATAGATTCATTTAATTTAAATTTACTGAAA TCTCTTTTTGAATTTTGTTTTGGTTTTGAAGAAATATTAATCATTACTGTTTTTCTTTTA ACATTTTCAGTGATTTTACTTCCTGTATTTTTAATCTTGTTACCTATATTCGATATGGTT CTAATTATCTTATTCTTAATCTTAGAAGTTATCTTCAAAGAATTTTGATTACTGCTACTT TTATTCTTACCATGTAAGTAAGAATTAGAAGGAATATTAAAAGAACTTTTACTTTTAAGA TTACTTGGAGTACTATTAGTATAGCTTAACCAACATCTGCAATTAGCAACATTCTCAGCA CCTCCAGCTAAATCCCCAGGATACATCATCTCAGTACGATAACTACCAGTAATAATAAAA TAATCATCAATAGGAACACTCTCAATAAAGTTAGCACGATGCCAAATTCTATGTTGTCTA GCACGACCATTCATCCAAACCTTATACTTATACCCCTTATCCAAAGCATGTAAAAAACTC ATCCTAGACTGATTAGTATGCAAAGCATCACGAACAATATTCTTAGCATTCTTACGATAC AACTCCCCATACTTCTTTTTAATCTCACTATCAGACAACTTACTTTCAGCTTCAATCTTA GCTTTTAACTTATAAGCATCACTATCAAAACGTGAAGATAAAATCTTAGAATAATCCCCA ACAGTTTGATCAACACTACCTTGAATAATCTTAGAACGCTTTTTCATTAAAGTATTAGCT TGACCTTGCAAAGCAGTATCAATTATGTTCTCTGCAATCTTTTTACTATTCCCAATTTTA T T T C CAT GAAT AAT AT T AT T T AAT AAT T T AT C T T T AT AAGAAT GAAT AC T T T T AT C AGCT TTATTCGTATGTTTAAACAAATGATTAAAACTTATATCTCTTTTAACCACATAAAGCTGA TGCAATCTTTGAGTTAAAAGCAGATACTGAAGTTTCTTTTTAGAAATCATAATAAACCAT TCTTATTAAAATCAGAATTCAAAGCATCAATCATACTATCCAATTTACCTTCAGCATCCA TTGGAGCATCACCATACAAAACCTTATCCAAAGACTGATTATTCATAAACCTACAATTAT AAT AC TCATCATCTTCACTCATTTT C AAAT C AAAAGAT T C AC C AAAAC GAT T AAC AAAC T CCCCTAAAGTCACTGCACCATTTTGCAGTAATTTAATTCCACGATCCAATACTTTATCTT CCTCATCAAAATTAACAGGCAAATACTCAAGTTTATGTGTAAATATTCCAAATTCCTTTT GAATAATAGTCTTATTAATCAAATTAGCCATCCTCTTTTGCAATGTAGCTACTGTAGATT TACTATAATTTTTAAGCAAAGTTTCAGTACGATTACTAGCAATACCTGTAGACTCAGAAT C ACC T AAAC GT T C AC GAGGT AC AC GAT GAAT AC GAG GAAT AC GAT C AGC C AC ACT AC CT G ACAGTTCTAAAAAACTGCCTTCTTTTTTCTCATCAGCAATTTTAGTAACATTAACACTAA CATTATTTTCTTCACTGGGAATAGTGAAAACTAAAGCAGTTCCAGGCTCATTGGACACTT CACGAAACTCATGTTCCAAATCCTCTTCAAAATCATCAAAAGTATAATCTTCTTTTTCCT CAACATTACCTGTAACAGTAATAATATAATTCGGAATACCATGTGCTTTAAAATGCCCTT GCT GAT AC T GAAT AAT AGC AT TAT C AGT T AAAAT AGC AT CC AACT C GGAC AAAT AT T T C G GTCTACCATAAACAAGACTTTCATTACTTTTAAGATTAAACCAGATTAAATCATTTGCAC GATTTTCATCTGTAATATTATCATCCCACAAACCAGTTTCACGATTCAATTGCTTAATAT TGTCAGGATCATATAATTTGAAATAATTTTCTTTATGCCCTATCTTTTGTACAACACGTT TTTTATCACGACACATTCTCAGGTATAAACTGCTTACATGATTAACACTTTTCAACTCCC CTTTTTCCCTAAGTAGTTCTAATCCTGCAAAACCAAAAGCTTCTAAATCTTCTAGGAATG ATTCTATTTCCTCATCAAAATTAAAATCATTTAAAAAATCATCTAATCCTTCAGGTATTT CTTCTCCTTCTTTAGAGATTATTTTTTTACCTGTGAAAATTGCATCTTCACATTTAACAG TAATACAAATATCATGCAAACCACTAATATCACGCAACTTATCTAATTGGAACGGGTCAT ACGCTGGGTCAATTACTTCTGTACCATATGTTAATTCATCTTTACTCATTTCTTGAGATT TAATCTCATATTCATTTAAAACACTTTTAATCACTGAATCTCTTAAAAAGTTACTTTTAA CTATTTTAACCATTATTAATTTCTCCTTCTACGATTTCTTTTTCTTAAAACTGTAGGTTT T GTT T GAGGAT AT AAACCTT CAT GT AAC AAT GAT ACACT AT CT AC ACGGTCAT CAT GT GT TGTTTCATCATCAGCTATTATTTTCTCTGATGGGAATTTCACAGCTTGTTTCATGAAATC TTTCAACCATTTACCACGAACAAATAATATCCGTCCATTATTCATCCCCCGTATAGTTCT ACT T GCT C T T AT T AAT T T T GAT T T AGGAAC AC GAAT AAAAT T AGGAT GAT AGT CT T C GAA TTCGTTTTCCCAGTATCTTTTAACTATTTTACCTGCAGCTGCAGGCTGATACTCAATCCA ATTATCATACTCTGAATGTTCATCCATAATCCTTGTCATGTATTGTTCTAACTTACCTGG CTTTTTTTGTGTGCTTTTCTGATTATGCACTACACCTACTTTTCCTTGTAATACTGTTGA AAAACTACATGCTGTATAATCAGAAGTACTTTTTTCTGTTGCTGCAATATCCCATGTAAT TACTTCTTGAAGAATATCCTTTTCAGTTAATAATTCATTAAACTCTTGTTTGCTAATTGT TGCTGCTTCAATTGTATCATAATCAAATACATCTCCAGCTCTTATTTCATAATCCCAATT AC C AAT C T GAT AC T GAT AAT CTGCTTTT GAT AAT T C AC GT AAAT TAGCTAAGTATTTTTC TTTGTCAATATATGGATTATGCCAGAAATTCATTTCAAAAAAAGGGTATTGTCCTTTAAC AAATTTTTCATTTAAGTATGTTGATCCATCTGCATCAGCAGGATTACTAATATAGTAAAT TGCTAATGGGAAAGTCATTAACTTATCAGTTCCTCTGAGACTACGATTCAGGAATTGCAG GT T T AC T T T T T C AAAT T C T G AAGC T T C AT C AAC AAT G AT T T T AT G AT AAGC AC GAC T T T T GAATTTCTCTTTGTCTTTTTCCAGAAGCATATATGAATAATAAATCCGTGCATCATTTTC ATGGTTAATGAAACATCTTTTACTTTGATTGTGTTCTATGTAATCGAATGGTTCTGTCCA ATGGTCCAGGTAATCTACTATTCCTCCAGTCGCTATTACATTATCATAAGTAGACCGTAG AATTAAGCAGCGGTAGTATGGTACTTCATAATGTTGTAAGGCTAATACTGCTCCAAGCAT ACTTTTACCAGAATATGCGGAACCTCCAATTAGTTTTCTTGTGTGGCGGTCTGCAATTGC ATATAATTGTCGGTCGTATGGTGTTACTGGAATGTATGGGTTTTCAAGTATTGTTCTTCT AATCAACTTTTTTTGTTGTTGATCTAAGTGTATTTTCTTGTAATCTACTCTCATAATTCA TCAGTGAATTTTTCTAAATCATCGTTAATGGTTAGTAATTCTTCTAATTCTTTGTCGTTC ATATGGATTACATTATCATTTTCTTTGGTGGATGTTTCAATACTTCCTTGTATTTGAGTA T T AT T AT C T T T AAT AT CT GT T GGT T GAC CAT GAGCT AGT CT GAAAT T T C T AT AGAT T AT T TCCGCAGATTTATTTAGATTTAGAAAACTATTTGCTCGTGTAGTTGGAGCATTCTCATTA CTATTTTCTTTTAGTTCTTTTTGTATTTCTTTTAGTAATTCAAAATCATCATTGAAGAAT TCTTGAAACTTAGCATTTCCTTGTTTGAATAATTCATAGTTCCACTCTCTTTCTTTTTGA TCCATATGATTATCATATAATCTGCATCTTTCCACCCAATTCCATCTTGAAGAGAGTGTT TTTAGTTGACTTAGTGAAGGTATGTTTTTTTCTTCAGTATGCTTCGATTTCTTCAATTTT ATTATCTGTGGGAATGTTCTTTTGGAACCTAAATCTCTGTATTCTTTGAATAGTGAATAG CTTTTACTTGGTTCTCCTTTTTGTCTTTCCCATGCTTCAGTCATAATCCCCCCATCCTTA TATTTTTTTTGAACATTATTGTAATGTTTATTCTTTTTTGTTGAACACGAAATTTTAAAA T AC GGGAT TTTTTATTTT CAT AGT GAAC AAT AT AAT AT TGTTTAGTTTT GAAT T T T GAAC AAAAAAAAGTTTTATTAGTTTTTTATAATAGTTTTAGGAAATTCACAATAACAAACTGAA ATCCATCAATCAAAAATACAATCAAAGCACCAACAATTGCTATGAATACTCCACTTTTTA TTGTGAATAATCGATTATCATCATTAGCTTGTTCTTCCTGAATAGCCAACTGTTTTGTTT GAATTTCAATACTTTTATCCATTTTCTTTATCAGTTTTTCTAATTGCTTATTTTGAAACT GATCACTTGCTTCCAGTTTACTTATACGAGATTCCTGTTTACAATATTTCTCATGTAAAT CTTGAACTTGATTATGGGTCATTATTGTTTCCTCCATTGTTAGTGGAGGTGTAAGTTCTT GAAATCCATCCAACTATCCCCCCTAATGCTACTGCTGCTAGTTCGTTGTTTCCAATATAG GTGCTTAGAATTCCTATAATTATTATTCCAATTATTGCTAATGTTGTGTTGTTGAAGTTA GT C AT T AT T AT T T T C ACT CC T AAAAAAAAT T T GT T T T T AAAAAAAAC AT T GGGGAAGGGA TTTGAACCCCTGCGATACTAGTGTATCATTAGATTAGCAGTCTAACGCCCTACCAGGCTA GGCTACCCCAACATTGTTGGTGGAGGGAATTGCACCCTCAACTTTTGAGTGGTTACTAAG T T AC AAT AAAAAAT AAGCT AT T AC C AAC AT CAT GAAC AT GT AT T AAAGGAGAT AAAT AT T TATATTATATTTATGGGAAGAATATTTTTTTTTAAGATATGTGATATTAATTATTTAATC TTTGGAGGATACCTATATATTTTAATGAAGAAAAAAATCCAATCATTCTATTATTCATTA AAAATTTGCTACCATTCCCATAATAATTTTTAAGAATAATAATAAATGAAAAGCGAATAA T AAAC GGAT AAGAAT AT T AT T T T T AAAT T GT C AAAT AAT GAT AAT GC GAAGCT AAT T GT A CAATAACATATGATAAAATATTTTTTATAAGTGTTTTATTACAAGACCACATTTTTTACA AACATACTCATCATGCCACTCATCATAGTAAACCTTTTTGCAATCTTTTTTCAGTTTTTC ACAAACAGGACACTCTATTTCTGCATGTTTCATTTTTTAAGGGAAATACACACCCGCAGT TTGTGCAATTATATTCTGCACGAGATTCATCCAAAACAGTTAAAGGGGATTCAGGAAACT TAATATAACAAGATGTCTTATGATTTCTATAATGTTTTTCTTGTTTTTTCCTTTGTTTCC ACTTCCAGTTACAAACAGGACACTCTATTTCTGCATGTTTCATGTTTTTAAAAAATGTAT TAGCATCCAAATAATATTATCCTCCCATATATAATTATAATAATGTCTTTTATTTTTTTT

[0163] ATGCTCACCACGACTGATATGTCCCTTACTTACAATCCATTTAATCATGGGATCCATCAA AC C T T T T T C AT AT T C T T T AAT AAAAT AAT T C AAT GT C C AAAAAGT AAC C T T T T C AT T AAA

[0164] AATTATCCTTTCAAAAAGTTCTTTAGATGTCTTTTTAAATTTTTTTCTATGAAAAATAAT

[0165] ACTGTCAATTTCATCAATAGTTAAATACAATGCAATATCTTCACCAGAAATATAAATACA

[0166] TTCCTTATCAACATCATACCAATAATACTTCTCATTAGGATTTCTTAATAATTTAAGATA

[0167] CTTGATAAAATTCATATTTTTACTCCTATTCTTTTTAATTCTCTTTTCACAGTTCTTTCC

[0168] TCCAAACCAAAATCACTCAAAGCATGCTGTGAAAGAAATGTTGTACCAGGCTCCCTTAAC

[0169] TCATTACTAATTAACTCCCCATCATTAATTAATTTCCTACGTTTTTGTTGATACTTTGCT

[0170] TTCTGCTCACGTACCGCCCAGGTCCTGCAAGCTTCCCTGCAATAACCTGTTTTATTCTCA

[0171] AACTTAATAAATACTCTGCCACAGTATTTGCATTTGCTGATGTAGAATCTTGTTTGTGGA

[0172] TCATTTAATGTTTTCTTCAATAACATCCTATCATCTTTCTAAATTTTTAGATAGTAAGTT

[0173] T T T C T T C ACT T C AAAAAC AAAC AT GT GAAAAGGT AAAAGT T T ACC CT GAT AT T T C AC AT A

[0174] TTCATTAATTACTTGTGATTTTTTTAATGCTTCAGTTATAGTTTCTTGTTTTAATTCTTC

[0175] ATTTATGCATTCAAAATTGGAGACTGTTCTACTTAAACTAATGTATACTCTTTCATATAG

[0176] AT T AAGGT T AT T GT T T T T C AC AT T CT CC ACC AC AAC AAGAAACT GAAT T T T C AGAAGT GT T GAAT TTACCATCT T C AAT C AT T T C AAT C AAC T C AT T AAGAAT AC GAGT AAC AT GC AT C A

[0177] AAGTATGAAGATTAATACTATGAATTATAACATCATCATACTCCATGTCTCTCAAACTAC

[0178] CTGTGCATTCTTTATGAATAGTAAAAGATTGTTCTTCAAATTCCCTGCTTCTTGTTTTAA

[0179] T AAAAT AT AGT AAT AAT T T C T CAT C AAT CAT AC AT AT T T AT CCT C CAT GT AT T T T T T AAC

[0180] TTCTTTCATAAAAACTCCTGATTACAATATAACCTATCTTAATCATCAAAAAAATCAGTA

[0181] AGACATAACACTAAACAACAACTAATAATTACACTTACACTAAAAACTACCCAACCATCC

[0182] ATCATACTATATTCTCCCCCCATTATCTCCAACTTGTTATTAACTCATACTCCTCATGAG

[0183] TTAACTTCTGTTTTAACCTTTCATCAAAACTTCTTACATCTCCAGGATTATTCTCATCCC

[0184] AACAAGACTCAATCTTGGCAGTTAATTCATTAATTTCTTCCTTATTCATTTTCAATATCC

[0185] TCCTATTTAATTTTGAAAGGTTGTAATGGTAGTGTTGGTCTTTCAAATATAATCTGAAAC

[0186] AAAACAGTATCCCCACATATATCTGGATAAATACTTTTTAACTCATGTTTCAACAAATCA

[0187] ACATGCCTATAACCTTCAAACCAAGCATGATTATGTTTCAAATCTTTAAATTTAACCTGT

[0188] TCAACTCTCCGTACTACCCGAAGCAGACAAAAACCCGTCCCCTCAAACCAACATTCTACA

[0189] AGGTCACCTTTTTTCAATCCTTTATCAGATTTTCTTATTGTAGACATTTTTTTCCTACTT

[0190] CTTAAAGGTTCAAAGAAATACGGTTTAAACTTCAATGTTTTCATTTTTATCAACTTCCTT

[0191] TTTTACATATCTTTGCTTCTTTTTTATCATCATCCAACACTATATTATGTTCTTTTAGGT

[0192] ATGATTTTCCAAATAATTCTTCAGCATGTTTACATAACAGATAAGAATCAATTACTCCTT

[0193] CAGCAGATATTATTCCTTCTTTTTCTAATTTTTCAAGTATATCATTACAACTCATTTTTG

[0194] GATCTCCTTTTTATTCCGGACTGGTTGTCTTTGACTTGGTTCAGGAACACCACACCTCAT

[0195] ACGTACCTTTTCAGCAAACTTAACAAACTCATCTGCAAATAAACTTATTCTAAACCACAT

[0196] CAACTGCCACCTTAATATAATATGTTCCTCATCTTCTTTCTTCATGATTCCACCTTTTTT

[0197] AACCTAGTTTGGAAGTTATTTCCCCCATAACGCCTAACATGATCTTCACAAATGATTCTT

[0198] TTCACAAAGGAATCTTTTTGATGTCGAGTTAATCCATACCTTTTCTGCAACTCATCAACA

[0199] GAGATATTTAATTCATGTTTATAATCTCTCCTGAATTTCAAATATGTTTCTTCACCTTTA

[0200] GAAACATCATAATCCGTTTCAACAATATTAAAAACTGGTTTATTCATTTTTTATCTCTCC

[0201] GTATACGTACGTACGTATGTGCACGTACGTGGTTTCCTACAGGTTCTTCGGAAAAATGTA

[0202] AACATAAATGTTTACAAATCATTTTATCACCAAACCATAATTCCGAACATCATAACCCAA

[0203] ATCATACACATGATCCAATAATGACTTATACGGAACTTCTTGCTGCTTACTAATATTCTT

[0204] CAGCTGATCTTTACCAATAAAACCAGCATTATCAACAATCCTATTAACTGTACCCATCAC

[0205] CCCATCAAAGACATTTACACTTCTTTCATGTTCTAAACGCTCAGCACGAATCTTACATAA

[0206] TCTTGATTCACGAGCATTAATCTCATTTTGCAACTCAGCTATCTCTTTTCTCAAACTATT

[0207] TTCCTCATCTTCAGATAAATCAATAGCTAATCTTAACTGCTCTTCTAAAAACTCACTTCT

[0208] GGTTGTCCCCAGTTTGTTTTTAGCTTCCCTCCATACTTCAGAATCAACCATGAATGAAAC

[0209] CTGTATTTTTTCACTCATACATCATCACCTAACTCTTTCATGACAGTTTCTAATTGCATT

[0210] TCCTTTGCAATTAAATCTAATTTTAAAGACCTGATTTCCTCTTTTAACTGGTCCTTTTCA

[0211] ATTAACCTTTGATTTTGTTTACTCACATACATATCAATGCAATGTTCCACGCAGAAACGT

[0212] ACAGTTAAACCATACTTTTCCATTAACTTTTTTGCCCTTGTAGTAATCCTACCAGATACC

[0213] ACTTCAGGGAAATTATCCTGAGTATTCATACTCTTACACCTAACATTTTTTCTGCTTTTT

[0214] T AAT AT GT T T GC AT T TAT AAT GTTCTACTTTTT GAT GT C GGT AGAAAT AAT CT T C AC AAC TACACCACCATCCATCTATACTGTCCCAGTTCACAGTATTGGATCCAGTATTTCCTGATG CCTTGAATTGTATGAATATTATTTCTACCTTAGGTTCCATAATTAAACAGCAACCTCTTT TTCTTGATTTTTAAACCAGTTATATGCTTCTTTGTTTTCAGCTTTGGTCAGTTCACCATC TTTGAATAACTTCATTCTATGAGTATTAATCATAGATGGAGTTATTTCCAGTGAACTGTC CTTTTTATGAATATACTCGCGGATTTTCTGCACAGCACCAGTAGCCTCAACTTTCTCAGT TCTTGCAGAACTACCCTTAGCAGTTTTAACATTAATTTTATCAGAATCCACAACATCTTT TTCAACAATTAGAAACATGTTTGCTAATAAATATCTTTTTAAGTAAGTAATGTAACTGCC TTCACTTTGCATTACATTCATACCTTTATTTAAAGCAACTATTTCAGGCATAGGAACACT GGTGATGACAGATTCCCCAGGTTCATTCCAATTTCTGATTTTCAATTGAGCCAATTCATT T GT AAAT GAAAACT C AAT GAAT AACT CC T GT T C AT AAC AT T C AC AAAAGAT T GC AGGC AA AATATCTTCTAATTCATAGTACTTAAATTTCCCAAATTTATTGTACCCACTTTTAGGTAA TTTTTTATGCAGGAGATTCTTTTGAATTCCTGCAATTTTCTGATATATAGTCATAAAAAA ATAATCCCCCTTTTTACATATTCATCGACTTTTTCATCAAATCCCTAATTTCTTTTAACT GT T C AT T T T C AC T T AT T T T T CT AAAC AT GGT GT T C AGAAT AT AT T CAT C GT AAT CT AT AT TTTTTTCAGCACATAACTCTTCCATCAGTGCTTTAACTATTCTAACTTGATTTTTCTGTA C AAT AT GT T T AAAGAT CAT AT C AAAAAAT GAAT T C AAT T CT GC AGGAT T T T TAGCAT CT A CATTATGGTCTAAAAAGTGGATTGCATTAGTTAATTTCTCCTTTAACTCTTTTTTGTTTA TGATTTCTTCAGTAAATAGTTCTTTCTCCATTAAAATTCCTCCTCAATAATCTCCAATAC T T T GT T T T C AT C AAT AT T T T C C AAAGAT GC T T GAGAGT GT G GAAT GAAAC T T T T T GAC AC CCAGTAGTCATGATACTGGATTTCACCACCACACAGTTCCTGTAAAGTATTCATAATATC AC GT GT GT T CT C AT AAGT AC T GT CAT GAAAAAC AAT T AC AGAT AAGAAAAT AC CAT GT AA GTTTCCATGAGCAGAAATACGTCCGCCCATTTTTTCAACAATTCCTAATAATTGATTGTA AAACCTGTTTTGCTGATCATAAGTAATCATACTAAACACCCATTATTATAAGGATCCATA GGCCCTACTACAAGGAATGCAAAAATGAATAATAAAATTATTAAAATTATTCCTTTTGCA AAGATTATTTCAACTTCATACTTTTCTCTCCAGGACACTTTTTTATGAAGTCTAACTGGA GTAGGTTTATTGTTGAAGAGACTCATAATTGAGCCTCCAACACTTTATACTTAACTTCTT TTTTCATACCAGGAGTATTAAGGAAATCTTTCAATTCCCTTAATCTGGTTTGTTCTTCAA CATACTCTGCATACTCCCCGTATAATGTTTCGAACAATTCTCTTTGTTCAGGATATTCCT GT GC T AGAAAT AC GAAC AGT T CAT CT AT T T C AC AGT AAT CT T T GT ACT GT GAC T GGT T T A TAG GAT AT AAC GC AT T AC GAAT CTTGTTTTCATTTTTC C AAAAT T C AGG AGT GAAC AT T C ATTTACCCCCGTAAGCTAGTTCACCTGCACGAGCAAACTCCAACATTTGATCATCAGACA ATTCATAGATTTGAAATTGCTTATATTCATCAGCCAACCTATGATCTATTTCTTCCTGTG TTTCATAATCCTCAGACAATAATATCTGTTCAAGATAATCAAACTCGACCTCATCTTCAG GTGGAGTAATCGTATACATTGAGTTGTTGTATCTGCAACTCATACGCCCACACTCCTGAA TTTTTGGAGTTCTTCTTCATAGACTCTGTTTTGAGCCTGAAGCCTTTTTGCTTTTAAAAA TACATGTTCAATTTCTTCTTTGGAAGTTAACCTTCCAAAAACTTCAATCTTATTATTATC CATATTACCAACTCGTGGATTTTTTAGAAGATCTTCTTCAGAGTTGCCGCTCTGATTTTT TCTTCTTCAAATAACTATTATACTTAATACTATATAAAGGTTATTATATTTACCTTTATT AAAGGTAAATATATATACTGCAAAACAGAATATATTATATAAGAAATATAAAAATGGAGG TATGCCTATTTTAGAATATACAACAAAATTAAGTAAAGGCGGACCTAATTCTATTAGGTC AAT AGT CC CCC AAGAT GT GAT AAAAT TAT T AGAAT TAGAAC T GGGGGAT T CAT TAG AT T G GATT GT AAAT AT T GAT GAAGGAAT AACT GT AT CT ATT GAAAAAGCT GAAAAAT AATC AGT TAATCCTTCACAAATATTTTTAGTAAATATTAAATATCTTTTTTTATAATAAATAGTATT GGTTACTAAAATTTTTATCAACTCGTGGTATCTATTTTTTGTAGCCTCTAGTTTTTGAGA GGCGAGATGTTGCCGCATCACAGACCCCCTCATCAACTAGGAATTACATTTACTTATTTT GCATTTAATTGTTCTTTTTCAACCTTTTTTTGTTCTGATAACATCATAAAATATGTAAAG TATAATCTATTTACAAAACCATATTCCCCACTATTTTCTCTCCCAATAGACTCTATAATC CCAAGGTCTTTAGCTCTCTTTAAAAAATCATTAAAAGCTCTCTTCTCCCCATTTGTCAAA ATATTTTTCACATCTGATTTTTTAAAATCCATTAATCTATTTTCCCCTAATTTAAGAAAA ATATCAATATATGTATCGCTTCTAATTTTATTTAATTTGGATTTTAATTGTTTTTTAGCT AATTCATCTGTTGCATCAGTAATACCAGAAAATACTATAGATTCATTTATTTTTAAATTA TCTTGAGCATTCCAAAAAATTGAATCACCAATTTGTTGCATTATTAAAGGCATACCCCAT GAAAAATAAACCATAGGTTCTATAGAATTATATCCATCTTCAAACTCAATGCCTACATTA TTAAATGAAGTTGTGAAGAAATCTTCAATATCATTATCCTCTAAATTATCAATTTCAATT AAAT T AAAC AT T CT AGAAAAT GAT T CAT T AAT T GAAC AT AAT T T T T C AAAT T C AT GAGGG T AAC T T AT T AAT GT GAAAAC AAC AGGC AT AT GAT AT T C AT T AACC AAT AAT GT T T C AGAT AATCCTTTATACCAATCTGTAAACTCTTCATTATCAGATAAACCATTAAGATCATCAACA AC AAT AAAT AT T CC AT AAT C T T C AGGT AAT T C T T TAG AAGT AGT AAT C AAAAAAT CT GCA AAAT GT T T T T T AAT AT TAT GAAT T AAAT T AGT AT CAT T AT C T T T T AAAGAT AC T CC AGT C CCTGCAACTTTAAATTCATTAATATTATTCAAAATTTTATCAATTACACTTTTACCTAAA TAATCCTTTTTAAATTCTTTTAATAATCCATCAAGTAACTTTTGTATTAATTCATCAATA GTTTTTCCACCACCATTGTTAATATGGATTGGAATCATTTGAAAATCATCTTCTGCTTTT CTTGAAACATAATTTATAAATGATGTTTTACCCATTCCTCTCTTTCCAGTTATGAAAAAA TGTTCGGGAGCTCCTTGATTTTTAACTTTAGGTAAATATTTTATAATTTTGTTGATATCT TCTTTTCTTCCTTTAAAATTATCTGGAGATACTGGTCTCCCTGGTTCAAAAGGACTTTCT GGGAAATCTTGTGACATGATATTTTTACCCCTCAATACTCATTTATTACAATTTACATTT TGTATTTAACTGTCTTATCAAATTTTTATTACTGTATCAGCTAGAAAATCTGGTGTAACA TTCCGTTTCTTTTTTTGATTTAAAATAGTATTGGACCATTTACTGAACTGTGTAGCTTCT TTACTATTGACGCGATAGCCTACGGAAATCATAGCATCTAAATTATAGAGTATGGGTTGA GT AT T T GC T T C AGGAT T AAT T T T AAT AAC GAT T CC AC T AAT AGT GGAAT C GT T T GGAT T A ACAGTTACTTCATCTTTAACAAGTTCTCCACTATCAAAGATATTATTTAAATGTTTACTA ATGTTTTTTGGTGTTTTATTAAATAAATTAGCCATTGTTTTTTGTGTTGCCCATAATGTT TCGTTTTCTTGATCAATGATAACTTTAATTGATACTGGCCCATCTTCACTTTTATATAAA AATGTTTCAATAAGATTACTTTCCACCATGATTTATATAACCTCTACAATTTTACTCTTC AAAAAAGTTTTTTTATTCTTGTTTTTATATTTATGTTATAAGTTAATATAAATGTTTCTA T T C AAAAAAT AAACCT T AAAAAT AC AGT AT AT T C AAT GAAAAT AAT GAAAC AC T AAAC AT T AT T C AGT AT GAAAAAAT GAAAC GC AT T GT GT T C T AAAT T T T GGT GT T T AAT T AGT T T T G TGAACTGTTAGGTTATTGTTCATTTTTATAATGAATATTTTTTATCTTTTTTTGTTATTA AAAT GT AT T GAT AT T T T T AAT T AAAAAAAT AGAAACC T T TAT AT ACT AT GT AGT AC AAT A TAATAATAGAAATACATGGAGGTGAAAAGTTGTCAGAATCAGAGGCAATAAAAATCATAA TATTGTCTTTGAGTATAATCCTAGAAATACTCAAAAGACAAAAACACAAGTAGGAGGTTA ACCCCCTCTTACAATTATTATATTTTTTATTGCTTCTTATATAAATTTTTCTTAAAAAGG AGGTGAAAAACAAAATGACAATAACAAGCATCATAATACTAATATTATTAATCATATTAG CAATATTAATCTACATTAATAGAAACCAAAAATATACTTGGATAAGTTTAATAATATGGT TGGTTATTTTAATAAGTTTCATCTACGAAACATTAATCTAACCCCCCCATATTTTTTTTA T ATT AT AAGCCAAAAAAAAGGAAGAT GAT AAAAAAT GACT GACTACACCAGCATAAGAAT AAAAAAAGAAATAGCTGAAAAAATACAATTAATAAAAATACAAAACAACTGCAAATCACT AAAC GAAAC AC T AGAAC AAC T AAT AC C C C GAAC AGT AAAC G AAAAC T AC GAGT T C AT AAA AGAAC AAC C AAT AT T C AC AAT AAAC AAC AAAC CC AT AAC AT T C AC AGAC CT GAAAAAC AA CAATACAGGCAAAACATGGGGAAATGAAAAACAAAACGCAACAATAGTATTTAAAGATAA ACAAGGAGCATTTATAAGATTCAATGATGAAGATGAAGTATTCTTAGAATACTACCACTT CATCTAAAACCATTCTTTTTTTAATATTAAAAATAAATCCATAAAACCTCAAATTTTTAA AGTTTTAAAAATTTGTTCCATAAGGTACATAGCATCATCCAATTCACCTTCAGAGATATT ATAAAAAGGATCATAATCTGCTTTTTTTCTTAAATTAAATAAATCATATAGTTTATTCCT AAC AGAAT AC T GAT T G AC C T C AAG AAGT T C AT AT T GAAC TTGCCCATGTTCT GT AAT AAA CTCATTTTCACCAAAGTCTTCTTTAGACAATGGTTTAAATTTATAAACTTCCTGAAGCCA CAAACTAACATATAAATAAGCACTGTAATAGCACCTATTGATAATAGTACTAAACACACA ACGATTATCGCAATTTTCAGGGATTAATTCTTCTTTATTTTCATATAACTGCTTAGTGAA TTTATATAATTCATATTGAGGGTATTTTTCAATATTAATGTATAAAACCCCCAATAAATT ATTTAATTAAAATATAGGTATCTTTATAGAAATCAAACATATTATTTTCCTTTGAAAAGA TAACCATATCATTAATAATTTGGTCTAACAGAGAATATTTTTCTTTCCTATCAATATGAA ATGGAACATTAATTGTAAATACTTTTTCAGGATATGATCCATCATACACTGTATCAATAG ATATATTTTTTCCTAATCCTAAATTGTCAGAAATTTTTATAAGTTCATCATAAAATTCAG TGATTTTACTTTCAAAAGACTCTGTGGATGATTTTTTAAAAAACTTATTTTTAAAAAAAT CCCCTACAATATTAGAAGGTTTATTATATCCATAATTTTTATCAACTGAATACACCATAG ACACCACCTATAAATGTTTATTCTAAAGTTTTTCTAATAATATCCAATGTTTCAGGATTA TTTTGAACATCAATAAGAACTGTATTACTTTTTATTTCTTCATCATTCCTATAAGATAAA GTAAGAATATCCTTATTTATTTCCCCATCTTTTAAAACAAAAGCTGCATTTATAGGTATA AC T T C AAG AT T T T CAAAGT T T T C T T C AAGT T T T T T T AAAAAAT C AAC AT TAAAAAC T T C T TTAAGGGAAATCCCCTCTTGAAGCACTCTTAAGCTTCTTGCATCCTTTTTCATTTAACTT CCTCCAAAATAGTTATTTAATAAATATCTTTTATTAAACATATAAACCTTGTCAAAAAAT TTTTAAATTAATAAATCTTCAAAGTCATTTTCTTTGAAATTTTCTTCTTTTTTCTCAATA TTTCGTAATCTTTCATCAATATCATAGATTAATTTTTCATAATCTTTGATTTTTTCATCT TTTTCAACAATTGTATTTTCTAATTTTAAAAATTCTGGAGATTTAACAGTGATCTTTTCA ACTTCTTTACTAATTGATAATGCTGGTAAATGTTGAATATATTCCTGTTTTAAATCAGCA GGATTGGTCATGAAATATACTTCATCAGTACTGTTTTTTGATTTGCCTTGCAGGTCATTT ACTTTGTCCAGACTCATACCATCATTATATAGTGTTGAGGCGTGGAATTTTCTGAGCATG

[0215] TGGCTGCGGAATCTGTTGTAGTTTCCTGCTTTTCCTAATCCTAATTCATTGTTTATTTTA

[0216] ATGAATTGTTGGTTTAGGTAGTCTTCATGTATTTTGAATAGTTGGGATTCTGGTGTTAAA

[0217] TTTTGTCTTGATAGTAAGTGATGATTTATAGCTGTGACTGCTTCTGGACTGCAATAGGTA

[0218] ATATAATATTTCTGTGTTTTTTGCCTTAAAATATTAAAAGTAGGGACAACATTATCAATA

[0219] TTATTTAACACATCTATCATTTCCATAATATTATCTGTATTATGGTATTCTTTAGTTGCG

[0220] TTCATATAGTCCATTACAGTTAAATTTAATGTTTCTCTTCTGGCACAACCTGAACTACTC

[0221] ATGAATAATATAATAGCTTTCATGGTAGAAGTACATATATTAACTGCTTCACGAATAATT

[0222] TCTTTATCAGGTAAATCTTTAAAACTGATCGGTTTGGGATTATTATAACTTTTTTTATCA AT T C GT GGC AGAT CAT GT AT T T C AAT T T C AAAAT AT T T AT AT AC AAC AAGAAC T GGT GT G

[0223] AAAGTATTTGATACAGTGTTATAATAATAATTATCCATCAAATACTTGCGAAAATTTAAT

[0224] AATCTTCTTTTTAATGTACGATGTTTCCATCGTATACCCTGTTCTTCTTCTTTTTCTGCT

[0225] T CT T C GAT T AAT T C GGCT AAT GAT AACT T AT T C AAT T C AC AAT AT T T AT T T AT T GAAT GT TTGTAAATATGCTGTGTTGTTTTTTTATGATTTTTAACAGAATGGATCTCATTTAGGATT

[0226] TCTTCATTGGTTCTCATATTTATCAAATCTATTTTTCTCTTTTTATATTGTTTGATGTAG

[0227] AGT GCATT T GAT AAGT AT GT GAT AAT AT GTCT ACT GT CGT GTCT ACAAC GAGT T AAT TAT AACATACTTAAAACATTCTAATCTATTAAAATATTATATCAAAAATAGGTTTTAAATTTT

[0228] ATCCCCCAAAATTAAACCCTTATCCTCGTACTGCGTTTATAAAAAATGGAAGTAATTAAG

[0229] ATAAATTTCTCCGTATTCATCTTCAAATCTGATTAATGCTCCAAGGTTATCTTTATAGAT

[0230] TAATGTTGCTTTTTCCCCATTACTCCATTGTGTGCCTACTTCCGATTGTTTTAATTCATT

[0231] CCATGATACATTTAAAACTGTTTTTTTGTTTATTAGTGTGAATGCTGGTTGTTCTTGTAT

[0232] GTATTCTTCAGGTGTTGATGTTCCATCAGGTAATAAATGTTTTACTGTTTCATTAACTGA

[0233] TTTGTATCCGTTGATTTCCTGTATTTCTTTTATCTTGTTAATTAAATTTACTTCAAGTTT

[0234] TATAGTACTTCTTTCCATTATTTAATTCCTCCTTTAAAAAAATAATAAAAAAATTATAGT

[0235] CTTTATATTAAATGGATTATAATGATGACTATAATATTTATTATAAGCATTATTGCGTTT

[0236] CGTGTCCTGTTTTCTTCATTTTTCATTGAATATAATAATACTACTGCAAGTATTATTAAA

[0237] ATTATTTGAATTAATGTTAATACTGTGGTTATCATAATTCCCCATCCCTTTTATTTAAAT

[0238] ATTAGTAATGACATATAATTTATTATAAATTATTTTAAAGAGGGATTAATTTTAATCCCC

[0239] CATTAGGGATAATATTATTGATATTATCCCCATAACCATATTTATTAGAAAATATATGTC

[0240] ATTTTTATCAATGTTTTTCACCTCCTTGTAGTTCTATTATTATATTGGTTACTATACTAT

[0241] ATAAAGGTTTCTATTTTTTACTACTTATTTTTATAAAAAAATAAAAGAATTGTTGATAAA

[0242] ACAACAATCTTATTGATACAAATCTGAGAAAATCCAAGCTGGATCATACGCTAAAATCGT

[0243] ACCATTCATACACCAATTACTTGAAACACTGTTTCCATCTAAAACTGCTGCAGGGTCACG

[0244] ATAAATCCAAGAACCCCCAGTATACTTAGAATGTTTCAATCTGAGCCTTACATGCCCAGT

[0245] CCCACTTCTACATCGTACATGAATGAATTGTACATTATAACCTAATGCAAGTCCTAAACG

[0246] GTAAAATAATTGACTTGAATCAGTACAATTAACTCCTTGCTTATTATATATCTTATTAAT

[0247] AGTTTCCTGAGTATTATATTTGGAGTTATAATAATATCCATATCCTCTTCCTCGGATTTT

[0248] GTTCAAGCAAGAGTCTATATCTGTTACACCACCAAATTTGTCGGTGAACCTTTTTAATGT

[0249] GATGTCTGTGGTGTCGCTGGTTGTTCCGTTGCCATGTGGGTCTTCAAGATAAACAATAGC

[0250] TGGACTAACACCATTTAATACTTCATAAGCTGATACACGATTAGCCATACTAACATAACA

[0251] AT T C T TAT AAT ACT T T GT T C C AT C AGT AT C AGAC AC AT AAT T T GGGAGGAAAC T T AAAT C T T TAT TAT C AAAT AT T GC AC AT T C AAC AAACT CT GC AAT AAGACT GT CT T GT T GAT AAT T

[0252] CGTACCTCTCTCTTTAATTGCTTTTATAATTGTGCCATAATCAGCTACACGATTCATGTA

[0253] ACTATACACTTGTCTTGTATTAAACTTTTTATTATTATTCACTAACCAATAAGCAATTCT

[0254] AT T CAT C AAAT AAGT GT AT T T T GT AAAACT AAT AT GT T CAT T AAC CAT T T AT T T T CAT CT CCTTTTTAAATTTTAAATCATCTAAATATTCCTTTAAAATACTGCATTTTATTATTGCAT

[0255] CT T C AAAAC AAC AAGGAACC T CT T T AT C AC AAGGAT C AT AAAAT GC AC AT T T T T C AC AT T CTTCACTAATCATTCCAAAATAATCCTCCTATCAATTTTTAATGTACAATCAGTAATCTT

[0256] GTTTGTTCTAAACGAGGATTCTCTCCAGCAGCTCCTCTTTTTACAATACCTTCCAACATA

[0257] TAACTTCCAGGGTCTAATCTAATCGGCAGTATTGCTCTACCATTAGCATCAGTTGTTCTT

[0258] TCATAAGAAACACCATTTACAATAAAATTAATTACCACATTTTCCCCTGTGTTAATTTCA

[0259] GGATTGGAAGAAATAATTTTACCGTAAACAGAATTTCCATAATTTAATTCTAGTTTAACT

[0260] CCATCTTCTCCAGCAAGGTCTGATAAGCGACCCATTACTAATCTAGTGTTTTTTAAATCA

[0261] TCCATTGTGGTTTCTAATATTTCTTGTCTTGCAAATGTTTCAGAATGTTCATCATCCACC

[0262] AAATCTGCATTTAAACCTGAACCTGCTCCATCCACTTCTTTTAATTTATTTAAAATAGTG

[0263] GTGGGTGTTTCAATGTTTCCATCTTTACCATCTTTTCCTGCAGGTCCAGTATCACCTTTA

[0264] CTTCCTTTATCACCTTTAGGACCTGTTGGTCCTGTTGGTCCAGTATTGCCTTTAGGGCCT

[0265] TTGAAATTCCCCAGTAATATTTTTTTAACCATTAAAAAAAACCTCTTCATTTATTTAATT ATACTCCTATTTTCATTTTCACACCAGACACATTATAACTCATACTTGTACCGCTACTCC ATGCACCTGCTTTTGTAATTAATCTCCACCTATTAGGATTAATTTCTGCTCCTCGACTCT T AT T GGCT GT GT T AAT T GT T T GAGAAT C AT C AC AT AAT AC AT AAC T T AACT T AT CAT GT C TTACTTTGAAAATTAAACCAAAATATTTTCCTGCAACATTCCCATATGTGCCCTTTTGGT CGTAAGTTCTATTTGTTCCACCATCACAAACTATATCAACCCATGTACGGCCTTGCTGAG GGGTGCATACTACACTTATTTTTTCGTTTGTTTCTGGGTTCAATAAGATAAATTGTATAT TAGAGTTAGTTCCTAAGCTGTTAGGACTCCATGATAAAAATAAGTCAAAGTTTTCAAGAT TAAAAGTATCTGTACGATACCCTTCTTGAGTGCTCTGCTTGTAGGGTATTTGTATGGTAT CGTGAACATGTGAGTGAGTAATACTTTTATCTGCAATACTGCTTCCTTCAGGCAATATAA CATCAGATAAAAAGAAACTAAAATCTCTTGTATTAATATTGGTTTTACCATCCAAGCTTG ATTGAAGATCAGTAATATTGCTGATACTGTGACTGTGACTATTGTTAGCTTTACCGTTCA AC GC AT T AGT C AC GAT T T T AT T CT GT AC AGGAT T AAT AGAAGAAC T ACT T AAT GAT GAAT C AAC AACT GT T T T AT T AGC ACC AT C AT C AAT AT T T T C C AAT T T T T T GT C AT GAT CAT AAA CAAAACCATTTACACTTGACTGTCTATTAATTCCATATTCATACACCCACATGCCATCAT CACTATCCTGATCAAGGTCTATACTATCACTTGTATGGCTGTGGTTAACATTTGCTTTAT TTGCTAATTGAGTATTCATTTCTGTTTGTGTAACATACCCTGATAAGTCAACTGTTGTTT CAGTTGAACCAATTTTTTCAAATTTATTATTAACCCAAATATACTCATCATAAATATTAC CCTCAACAGAAGCTGAAGTATTTAATTTTAAATACATTACTCCAACTTCCCCTGTAGATG GTAATGAATTTACTATTGTTGCTGTAAAACCAATTAATGAATTTATCTGAGAATTAATAT TATTTACTTTCGCATCTATTTCAGTTTCAGTATAATACCTTTCATCATGATTATGTCCAA CATTTGATTTTCCACCTAATTTTGAATCAATTTCTGTTTTCTTATAATAATCATTTAAAT GTTTAACAGCAATATAATTGCCAACATGTACCTGATAACTATTTGCTCCATCTTCACTAG CTATATTTAAATATAAATCACCAGTAGTTTCATCATGTTCAAATATTGGAGGGTTGGTTT CATCATTACAATAACACCATAGATTTCCTTCAGCATCTCCTGCTAAAGTGAAAAAACCTG GTGGAGGTACTGTAATATTTAATAATGGGTTTGCCCATTTGTCGAACAATTCATCAATGG CTTTACTTACTGTATTGGCATCTGTCTTTAATATGTCGATGTCTAGGAAATCTTCAATGT TTCCTATTTTTGTGCCTAGGTTTGTTTCTCTTAATCTTGCACATGCATTATTTAATGCTT TTCTTTCATTTTCAGTTAGTTTAGTCATATTACACTCTCTCGTTTAACATTACCTATTTT AATCTCTTTGTCACCATTTAGATAAGGATTATAACTTATTTCTGTTATTTGTGCTTTGAA CACCT CT GAAT CT GGGAAT CTTACAT T C

[0266] In preferred embodiments, the pharmaceutical compositions of the invention further comprise at least one additional virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract of a human. Similarly, in preferred embodiments, the methods in which the viruses and pharmaceutical compositions are used comprise the administration of at least one additional virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract of a human. In preferred embodiments, the at least one additional virus is a virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract of a human selected from those disclosed herein. The viruses may be stabilised as discussed above.

[0267] Archaea and bacteria are known to develop resistance against particular active substances over time. It is therefore advantageous for pharmaceutical compositions and methods to involve a cocktail of two or more viruses to minimise the impact of such resistance. The use of two or more viruses in combination is additionally advantageous because, as discussed above, viruses are specific to their target bacteria and / or archaea. Thus, unlike other methane-inhibiting agents, which provide a less targeted approach and can cause toxicity to the human, viruses will not cause problematic off-target side effects for the human. Further, the use of more than one virus reduces the risk that archaea and bacteria have developed or will develop resistance to the active substance.

[0268] In preferred embodiments, the pharmaceutical compositions of the invention further comprise at least one additional active substance which alleviates the symptoms of irritable bowel syndrome. Similarly, in preferred embodiments, the methods in which the viruses and pharmaceutical compositions are used comprise the administration of at least one additional active substance which alleviates the symptoms of irritable bowel syndrome. In particularly preferred embodiments, the additional active substance is an antispasmodic. Preferably, the antispasmodic comprises hyoscine or a pharmaceutically acceptable salt thereof.

[0269] The co-administration of additional active substances which alleviate the symptoms of irritable bowel syndrome is advantageous because this allows for a more complete reduction in a patient’s discomfort and symptoms. For example, a patient’s constipation may be reduced by virtue of the administration of the virus of the invention, while their abdominal pain may also be reduced due to co-administration of an antispasmodic. This provides a more complete treatment.

[0270] In some embodiments, the compositions of the invention further comprise a clay, preferably Fuller’s Earth. In some embodiments, the compositions of the invention further comprise silica gel. In some embodiments, the compositions of the invention further comprise fibre. In some embodiments, the compositions of the invention further comprise probiotics. In some embodiments, the compositions of the invention further comprise one or more food additives. In some embodiments, the compositions of the invention further comprise one or more dietary supplements. In some embodiments, the compositions of the invention comprise one or more of a clay, silica gel, fibre, probiotics, food additives and dietary supplements.

[0271] In some embodiments, the pharmaceutical compositions of the invention are configured to provide location-specific release of the virus to the human. This is advantageous because it allows for the targeting of particular parts of the body for treatment. For example, the pharmaceutical composition may be configured to provide location-specific release of the virus when the pharmaceutical composition enters the target release area (such as the small intestine) in a patient’s gastrointestinal tract, thus allowing for targeted reduction in methane in the parts of the body where its production is most problematic.

[0272] In some embodiments, this is achieved when the pharmaceutical composition is in the form of a tablet which is coated with an enteric coating or, more preferably, encapsulated in an enteric coating. The enteric coating protects the active ingredient (i.e., the virus) from the acidic pH environment in a patient’s stomach, preventing it from being released. Once the tablet passes into a patient’s intestines, the coating breaks down to release the virus in a targeted way.

[0273] EXAMPLES

[0274] Example 1

[0275] A virus with activity against Methanobrevibacter smithii was isolated from abattoir specimens of fresh bovine rumen fluid that had been concentrated by filtration and addition of polyethylene glycol (PEG) and sodium chloride to a final concentration of 10% (w / v) and 0.5M, respectively as described below.

[0276] First, virus particles were separated from the rumen fluid, concentrated and purified using PEG precipitation and centrifugation. Archaea cultures in 119 Broth were inoculated with the purified virus suspension and the cultures were allowed to grow for 48 hours to allow any Archaea specific viruses to initiate a cycle of infection and amplification.

[0277] Following incubation, methane production was measured using gas chromatography-mass spectrometry (GC-MS). Cultures inoculated with the virus preparation were compared to those that were not treated with virus, but were otherwise identical. The presence of active virus was confirmed by a measurable reduction in methane production in cultures exposed to the virus preparation. The viruses released during the infection cycle were recovered from the cultures by filtration with a 0.22pm filter that allowed the separation of virus particles from the Archaea in the culture medium.

[0278] The recovered virus suspension was then used to inoculate a fresh culture of host Archaea cells as before. This process of infection, replication, extraction and re-infection, referred to as passaging, was repeated four and six times to obtain a virus with activity against Methanobrevibacter smithii. The virus with activity against Methanobrevibacter smithii was a novel virus which had the sequence set forth in SEQ ID NO:1.

[0279] Fresh human faeces obtained from three healthy donors were used in an established in vitro colon model to investigate the methane mitigation effect of the virus which had been demonstrated to be active against Methanobrevibacter smithii (ATCC 35061). Because the nature of the intrinsic archaea population within each faecal donor sample was not known, a spike of Methanibrevibacter smithii (ATCC 35061) was introduced to the in vitro colon model to provide a sensitive target for the virus.

[0280] The in vitro colon model comprised 100ml volume, acid-washed, sterilised, air-tight, glass vessels with sealed rubber lids which could be manipulated in an anaerobic chamber containing a standard anaerobic gas mixture of 80% nitrogen, 10% carbon dioxide and 10% hydrogen. The rubber lids were self-sealing and remained air-tight after piercing with a fine bore needle. All materials were de-gassed and stored under anaerobic conditions ready for use within this chamber.

[0281] The faeces of each donor were processed separately and, in each case, experiments were set up in duplicate. Each sample was homogenised in sterile, de-gassed PBS buffer to form a faecal slurry. Other materials included sterile, de-gassed 119 DSMZ culture medium, a planktonic growth of Methanibrevibacter smithii in culture medium, and virus in culture medium.

[0282] The in vitro colon model vessels were filled within the confines of the anaerobic chamber with either faecal slurry and culture medium, or faecal slurry, Methanibrevibacter smithii and culture medium, as shown in Table 1. The rubber lids were applied and the vessels were sealed. Thereafter, the seal was pierced with a 25G needle attached at all times to a gas line carrying a mixture of 90% hydrogen and 10% carbon dioxide at 1 bar above atmospheric pressure and gas was delivered to each vessel for a period of 45 seconds before the needle was withdrawn. Thus, each air-tight vessel in the in vitro colon model contained 20.2ml of liquid under a hydrogen-rich, anoxic gas mixture. The vessels were incubated at 37°C for 24 hours after which 1.0ml of gas was withdrawn via a gas-tight Hamilton syringe for detection of methane to ensure all vessels contained viable growths of archaea. Using a syringe and 25G needle inserted through the rubber lid of each sealed vessel, either 1 ,8ml of culture medium or 1.8ml of virus in culture medium were added, as shown in Table 1. The vessels were incubated further for 48 hours at 37°C, after which 1.0ml of gas was withdrawn and transferred into labelled and sealed 10ml Hungate tubes ready for analysis. Table 1

[0283] Table 1: Initial in vitro colon model liquid contents at the starts of the 24 hour and 48 hour incubation periods.

[0284] Using a gas-tight Hamilton syringe, 200pl of gas were taken from each labelled Hungate tube and analysed separately via gas chromatography (GC). By subtracting the GC reading for methane generated in the Faeces Only control for each faecal sample, the contribution made by the Methanibrevibacter smithii spike could be elucidated. Having corrected for methane in the Faeces Only control, the GC readings for Faeces + Methanibrevibacter smithii could be compared with the GC readings for Faeces + Methanibrevibacter smithii + Virus, thereby determining the impact of virus on methane emissions in the in vitro colon model.

[0285] The data in Table 2 show that each faecal sample cultured in the absence of spiked Methanibrevibacter smithii generated detectable levels of methane. This indicates that each of the three faecal samples were endowed with intrinsic populations of methanogenic archaea and, by inference, the human donors of these faeces carry methanogenic archaea in their gastrointestinal tracts. The faeces of Donor 1 and Donor 2 displayed the highest and lowest innate ability to generate methane, respectively. As shown in Table 2, the presence of virus reduced the average GC reading for each of the donor faecal samples. Having corrected for methane emission caused by intrinsic populations of archaea in the faecal samples, it can be seen that the presence of virus caused an average reduction in methane emission of between 12.16% and 26.13% under the conditions of the experiment.

[0286] It will be appreciated that the in vitro colon model used in this experiment is a representation of the anatomical and physiological makeup of the human colon in which ingesta transit times of up to 72 hours are considered normal. In constipation-predominant irritable bowel syndrome (C-IBS), colon transit times of over 100 hours are commonly experienced. It will be further appreciated, therefore, that the benefit imparted by virus on methane emission in vivo in the human colon may be far greater that that demonstrated in the in vitro colon model over an incubation period of only 48 hours. Furthermore, the experiment depicts a scenario in which a naive colon is provided with virus for the first time leading to an initial infection cycle and lysis of the target archaea. In the event of repeated daily dosing with virus, as may be appropriate for a patient with C-IBS, the contribution of infection cycles already established from previous doses would give rise to a more comprehensive methane mitigation effect. Table 2

[0287] Table 2: Gas chromatography readings for methane at the end of the experiment and analysis of the generated data

[0288] NUMBERED EMBODIMENTS OF THE DISCLOSURE

[0289] 1. A virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human for use in a method of reducing the production of methane in a human subject, the method comprising administering the virus to the subject.

[0290] 2. A pharmaceutical composition comprising a virus capable of reducing the formation of methane in vivo in the human gastrointestinal tract.

[0291] 3. The pharmaceutical composition according to embodiment 2, in the form of a liquid, syrup, capsule, tablet, bead, granule, pellet, lozenge, pill or suppository.

[0292] 4. The pharmaceutical composition according to embodiment 3, in the form of a tablet.

[0293] 5. The pharmaceutical composition according to embodiment 4, wherein the tablet is coated with an enteric coating, preferably wherein the tablet is encapsulated with an enteric coating.

[0294] 6. The pharmaceutical composition of any of embodiments 2-5, wherein the pharmaceutical composition is configured to location-specific release the virus to the human.

[0295] 7. The pharmaceutical composition of any of embodiments 2-6 for use in a method of reducing the production of methane in a human subject, the method comprising administering the composition to the subject.

[0296] 8. A method of reducing the production of methane in a human subject, comprising administering to the subject a virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract.

[0297] 9. The substance for use according to embodiment 1 , the pharmaceutical composition for use according to embodiment 7, or the method according to embodiment 8, wherein the human has irritable bowel syndrome.

[0298] 10. The substance for use, pharmaceutical composition for use or method according to embodiment 9, wherein the irritable bowel syndrome is constipation-predominant irritable bowel syndrome. The virus for use, pharmaceutical composition for use or method of any preceding embodiment, wherein the administration is oral administration or rectal administration, preferably oral administration. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method according to any preceding embodiment, wherein the virus inhibits the metabolic pathway of methane production in the human gastrointestinal tract. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method according to any preceding embodiment, wherein the virus is an inhibitor of methanogens, preferably an inhibitor of methanogens in the Methanobrevibacter genus. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method according to any preceding embodiment, wherein the virus is an inhibitor of Methanobrevibacter smithii. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method according to any preceding embodiment, wherein the virus is a virus which targets metabolic processes involved in methanogenesis. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method according to any preceding embodiment, wherein the virus is stabilised. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method for use according to embodiment 16, wherein the virus is stabilised by immobilisation to a substrate. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method according to embodiment 16 or 17, wherein the virus is immobilised via its head. The pharmaceutical composition, or pharmaceutical composition for use according to any preceding embodiment, wherein the composition further comprises at least one additional virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract of a human. The virus for use, pharmaceutical composition for use or method according to any preceding embodiment, wherein the method further comprises administering at least one additional virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract of a human.

[0299] 21. The pharmaceutical composition, or pharmaceutical composition for use according to any preceding embodiment, wherein the composition further comprises at least one additional active substance alleviates the symptoms of irritable bowel syndrome.

[0300] 22. The virus for use, pharmaceutical composition for use or method according to any preceding embodiment, wherein the method further comprises administering at least one additional active substance alleviates the symptoms of irritable bowel syndrome.

[0301] 23. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method for use according to embodiment 21 or 22 wherein the additional active substance is an antispasmodic.

[0302] 24. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method for use according to embodiment 23, wherein the antispasmodic comprises hyoscine or a pharmaceutically acceptable salt thereof.

[0303] 25. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method according to embodiment 24, wherein the viruses are stabilised.

[0304] The foregoing disclosure provides exemplary embodiments of the invention and is not intended to be limiting. It will be appreciated that various other modifications and variations of the invention are also possible.

Claims

CLAIMS1. A virus capable of inhibiting the formation of methane in vivo in the gastrointestinal tract of a human for use in a method of reducing the production of methane in a human subject, the method comprising administering the virus to the subject, wherein the virus is an inhibitor of methanogenic archaea.

2. A method of reducing the production of methane in a human subject, comprising administering to the subject a virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract, wherein the virus is an inhibitor of methanogenic archaea.

3. A pharmaceutical composition comprising a virus capable of reducing the formation of methane in vivo in the human gastrointestinal tract for use in a method of reducing the production of methane in a human subject, the method comprising administering the composition to the subject, wherein the virus is an inhibitor of methanogenic archaea.

4. A pharmaceutical composition comprising a virus capable of reducing the formation of methane in vivo in the human gastrointestinal tract, wherein the virus is an inhibitor of methanogenic archaea and wherein the composition is not a solid or semi-solid composition for administration to a ruminant.

5. The pharmaceutical composition of claim 4, wherein the composition is a liquid, syrup, capsule, lozenge or suppository.

6. The pharmaceutical composition for use according to claim 3 or the composition of claim 4, wherein the composition is in the form of a tablet.

7. The virus for use according to claim 1 , the pharmaceutical composition for use according to claim 3 or 6, or the method according to claim 4, wherein the human has irritable bowel syndrome, optionally wherein the irritable bowel syndrome is constipation-predominant irritable bowel syndrome.

8. The virus for use, pharmaceutical composition for use or method of any preceding claim, wherein the administration is oral administration or rectal administration, preferably oral administration.

9. The virus for use, pharmaceutical composition for use, pharmaceutical composition or method according to any preceding claim, wherein the virus is an inhibitor ofmethanogens in the Methanobrevibacter genus, optionally wherein the virus is an inhibitor of Methanobrevibacter smithii.

10. The virus for use, pharmaceutical composition for use, pharmaceutical composition or method according to any preceding claim, wherein the virus is stabilised.

11. The virus for use, pharmaceutical composition for use, pharmaceutical composition or method for use according to claim 10, wherein the virus is stabilised by immobilisation to a substrate.

12. The pharmaceutical composition for use or the pharmaceutical composition according to any preceding claim, wherein the composition further comprises at least one additional virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract of a human.

13. The virus for use, pharmaceutical composition for use or method according to any preceding claim, wherein the method further comprises administering at least one additional virus capable of inhibiting the production of methane in vivo in the gastrointestinal tract of a human.

14. The pharmaceutical composition for use, pharmaceutical composition, virus for use or method according to any preceding claim, wherein the composition further comprises at least one additional active substance that alleviates the symptoms of irritable bowel syndrome, optionally wherein the additional active substance is an antispasmodic; or wherein the method further comprises administering at least one additional active substance that alleviates the symptoms of irritable bowel syndrome, optionally wherein the additional active substance is an antispasmodic.

15. The virus for use, pharmaceutical composition, pharmaceutical composition for use or method for use according to claim 14, wherein the antispasmodic comprises hyoscine or a pharmaceutically acceptable salt thereof.

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