37 results about "Edwardsiella ictaluri" patented technology

Live-attenuated vaccines against Edwardsiella ictaluri or against Pasteurella piscicida are disclosed. Both vaccines are incapable of reversion to virulence, because both are made by deletion mutations in the aroA gene, the purA gene, or both. These vaccines may be used not only to vaccinate fish against Edwardsiella ictaluri or Pasteurela piscicida, but also to serve as vectors to present antigens from other pathogens to the fish, thereby serving as vaccines against other pathogens as well, with no risk of infection by reversion to the virulent form of the pathogen in which the antigen occurs naturally.

A rapid and sensitive PCR-based assay is provided to facilitate early detection of Edwardsiella ictaluri in channel catfish. This bacteria is the causative agent in the disease, Enteric septicemia of catfish (ESC). Also provided is a method of selecting breeding stock for use in selective breeding programs to improve disease resistance in channel catfish. Also provided is a method of determining the efficacy of vaccines produced against ESC.

The invention provides a gene chip for detecting various seawater cultured animal pathogenic bacteria and its application. The pathogenic bacteria include: Vibrio anguillarum, Vibrio alginolyticus, Vibrio parahaemolyticus, Vibrio harveyi, Vibrio fischeri, Vibrio splendidus, Vibrio riverina, Vibrio vulnificus, Vibrio cholerae non-O1, Vibrio mimicus, Vibrio salmonicida, Aeromonas hydrophila, Aeromonas sobria, Aeromonas salmonicida, Edward tarda Bacillus, Edwardsiella catfish, Renalobacter salmonidum, Mycobacterium marinum, Pseudomonas, Streptococcus.

The invention provides a breeding method of excellent strains of hybrid pelteobagrus fulvidraco resistant to Edwardsiella ictaluri.The method comprises the following steps: collecting the pelteobagrusfulvidraco and pelteobagrus vachelli, cultivating till mature, and then reproducing to obtain the fry of purebred pelteobagrus fulvidraco and the pelteobagrus vachelli with different populations; soaking all the fingerlings of the pelteobagrus fulvidraco and the pelteobagrus vachelli with a soaking method according to determined medial lethal concentration of the Edwardsiella ictaluri; performingbreeding on the population resistant to the Edwardsiella ictaluri twice, screening the excellent disease-resistant strains by the survival rate of mixed-cultured fry, respectively cultivating the strains of the pelteobagrus fulvidraco resistant to the Edwardsiella ictaluri, and the pelteobagrus vachelli to sexual maturity, and then producing the hybrid pelteobagrus fulvidraco according to a universal artificial reproduction method of fish. According to the breeding method of the excellent strains of the hybrid pelteobagrus fulvidraco resistant to the Edwardsiella ictaluri, hole-in-head-disease-resisitant parents of the pelteobagrus fulvidraco and the pelteobagrus vachelli can be cultivated, the incidence and mortality of hole-in-head disease in the seedling and culture process are improved, and the survival rate of the culture is improved.

The invention discloses a pelteobagrus fulvidraco beta defensin gene, and beta defensin antibacterial peptide and application thereof. An in vitro recombinant expression technology is used for successfully expressing a pelteobagrus fulvidraco beta defensin recombinant protein PET-32a-Pf_BD, and antibacterial activity identification results show that the pelteobagrus fulvidraco beta defensin gene has a certain bacteriostasis for staphylococcus aureus in gram-positive bacterium, escherichia coli in gram-negative bacterium, aeromonas hydrophila, edwardsiella ictaluri and flavobacterium columnare. Therefore, a foundation is laid for application of the defensin recombinant protein, used as a broad-spectrum antibacterial drug, in aquaculture.

Aeromonas hydrophila is a reemerging pathogen of channel catfish (Ictalurus punctatus); recent outbreaks from 2009 to 2014 have caused the loss of more than 12 million pounds of market size catfish in Alabama and Mississippi. Genome sequencing revealed a clonal group of A. hydrophila isolates with unique genetic and phenotypic features that is highly pathogenic in channel catfish. Comparison of the genome sequence of a representative catfish isolate (ML09-119) from this virulent clonal group with lower virulence A. hydrophila isolates revealed four fimbrial proteins unique to strain ML09-119. In this work, we expressed and purified four A. hydrophila fimbrial proteins (FimA, Fim, MrfG, and FimOM) and assessed their ability to protect and stimulate protective immunity in channel catfish fingerlings against A. hydrophila ML09-119 infection for vaccine development. Our results showed catfish immunized with FimA, Fim, FimMrfG, and FimOM exhibited 59.83%, 95.41%, 85.72%, and 75.01% relative percent survival, respectively, after challenge with A. hydrophila strain ML09-119. Bacterial concentrations in liver, spleen, and anterior kidney were significantly (p<0.05) lower in vaccinated fish compared to the non-vaccinated sham groups at 48 h post-infection. However, only the Fim immunized group showed a significantly higher antibody titer in comparison to the non-vaccinated treatment group (p<0.05) at 21 days post-vaccination. Altogether, Fim and FimMrfG recombinant proteins have potential for vaccine development against virulent A. hydrophila infection. Genomic subtraction revealed three outer membrane proteins present in strain ML09-119 but not in the low virulence reference A. hydrophila strain; the major outer membrane protein OmpAI (OmpA1), TonB-dependent receptor (TonB-DR), and transferrin-binding protein A (TbpA). Here, the genes encoding OmpAI, tonB-DR, and tbpA were cloned from A. hydrophila ML09-119 and were expressed into Escherichia coli. The purified recombinant OmpA, TonB-DR, and TbpA proteins had estimated molecular weights of 37.26, 78.55, and 41.67 kDa, respectively. Catfish fingerlings vaccinated with OmpA1, TonB-DR, and TbpA emulsified with non-mineral oil adjuvant were protected against the subsequent A. hydrophila ML09-119 infection with 98.59%, 95.59%, and 47.89% relative percent survival (RPS), respectively. Furthermore, the mean liver, spleen, and anterior kidney bacterial loads were significantly lower in catfish vaccinated with the OmpA1 and TonB-DR than the non-vaccinated control group. ELISA demonstrated that catfish immunized with OmpA1, TonB-DR, and TbpA produce significant antibody response by 21 days post-immunization. Therefore, data generated during the study suggest that OmpAI and TonB-DR proteins could be used as potential candidates for vaccine development against A. hydrophila epidemic strain infection. However, TbpA protein failed to provide such strong protection. Recombinant ATPase from A. hydrophila also showed promise as a vaccine antigen. A live attenuated vaccine was prepared that combined the advantages of a live attenuated vaccine (ESC-NDKL1 (ΔgcvPΔsdhCΔfrdA) mutant of Edwardsiella ictaluri) against enteric septicemia of catfish (ESC) and three immunogenic recombinant proteins (Fim, FimMrfg, and ATPase) against A. hydrophila infection. Our results showed channel catfish fingerlings immersion-vaccinated with ESC-NDKL1::pETfim, ESC-NDKL1::pETmrfG, and ESC-NDKL1::pETATPase exhibited 100%, 91.67%, and 100% percent survival after challenge with the A. hydrophila ML09-119, which was significantly less than non-vaccinated group (88.89% mortality). In a second study, Catfish immunized with NDKL1::pETfim, ESC-NDKL1::pETmrfG, ESC-NDKL1::pETATPase had significantly (p<0.05) lower mortalities than sham-vaccinated group.

The invention provides a kit for detecting fish pathogenic bacteria; the kit comprises a primer with a nucleotide sequence as shown in SEQ ID NO. 1-2, and the primer is used to amplify genes of edwardsiella ictaluri. The invention also provides a method for detecting fish pathogenic bacteria, and an application of the primer shown in SEQ ID NO. 1-2. The kit provided by the invention can rapidly and accurately detect a pathogenic bacterium of ictalurus punctatus - edwardsiella ictaluri, and thus realizes pertinent control and detection of ictalurus punctatus enteric septicemia, and the invention has wide application prospects.

The invention discloses a flounder abdominal dropsy dual inactivated vaccine and a preparation method thereof. The flounder cultivating abdominal dropsy dual inactivated vaccine is prepared by the following method: (1) Edwardsiella tarda inactivated vaccine is prepared; (2) Vibro alginolyticus inactivated vaccine is prepared; and (3) the concentrations of the Edwardsiella tarda inactivated vaccine and the Vibro alginolyticus inactivated vaccine are adjusted to between 10<8> and 10<12> CFU / mL by physiological saline or 0.1 M sterile phosphate buffer the pH of which is 7.2 respectively, then the Edwardsiella tarda inactivated vaccine and the Vibro alginolyticus inactivated vaccine are mixed evenly according to the fact that the ratio of Edwardsiella tarda to Vibro alginolyticus is 1 to between 0.1 and 3, thereby preparing the flounder cultivating abdominal dropsy dual inactivated vaccine. The prepared vaccine can effectively prevent the occurrence and spread of the flounder abdominal dropsy, avoids the harm to the human body due to the residue of a large amount of antibiotics in fish bodies, improves the safety of cultivating products, and can not cause environmental pollution at the same time.

The invention discloses a preparation method of astragalus polysaccharide chitosan nanoparticles. The astragalus polysaccharide chitosan nanoparticles prepared by the method have good prevention and treatment effects on Edwardsiella catarrhalis. According to the preparation method, the astragalus polysaccharide chitosan nanoparticles are prepared by applying an ionic crosslinking method for the first time; the degradation of the medicine before reaching an acting part is reduced, the release speed of the medicine is reduced, the release time is prolonged, and the stability and in-vivo availability of astragalus polysaccharide are improved, so that the curative effect is improved, the administration frequency and dosage are reduced, and the product is more conveniently applied to fish culture.

The invention relates to the field of aquaculture, and discloses lactobacillus acidilactici, an aquatic feed additive based on the lactobacillus acidilactici, fish feed and application of the lactobacillus acidilactici, the aquatic feed additive and the fish feed. The lactobacillus acidilactici can promote the growth performance of cultured fishes, improve the immunity of the cultured fishes, especially improve the capacity of resisting Edwardsiella infection of longsnout catfish, well adapt to the digestive tract environment of the cultured fishes and stably play a role.

A live attenuated Edwardsiella ictaluri bacterium lacking a viable gene encoding a functional evpB protein and a method of using the same to protect fish against infection from virulent Edwardsiella ictaluri. The methods and compositions for protecting fish against infection from virulent Edwardsiella ictaluri comprising administering to a fish a therapeutically effective amount of an attenuated Edwardsiella ictaluri bacterium lacking a viable gene encoding a functional EvpB protein. The bacterium may include an insertion and / or deletion mutation in the evpB gene. The fish include catfish, preferably catfish fingerling or a catfish fry. The composition may be delivered via immersion delivery, an injection delivery, an oral delivery, or combinations thereof.

The invention discloses primer pairs shown in SEQ ID NO.1-2 and SEQ ID NO.3-4 and an application thereof, and further, discloses a kit and detection method for detecting vibrio bacteria and edwardsiella ictaluri. The kit and the method can amplify vibrio bacteria and edwardsiella ictaluri gene fragments respectively and simultaneously, achieve one-step detection of two pathogenic bacteria in a same reaction system, have the advantages of strong specificity, high sensitivity, short consuming time, simple steps, small workload, and low cost, are used for rapid detection of culture water bodies, can timely and effectively prevent happening of fish diseases, and have good application prospects.

The invention discloses multiple PCR primer groups and a PCR detection method for synchronously detecting four pathogenic bacteria of a siluriformes fish. The multiple PCR primer groups and the PCR detection method have the advantages that high detection efficiency and accurate results are achieved, and the four pathogenic bacteria of the siluriformes fish can be synchronously detected. The multiple PCR primer groups comprise a catfish edwardsiella ictaluri primer pair, a flavobacterium cloumnare primer pair, an aeromonas hydrophila primer pair and an aeromonas punctata primer pair. The four pairs of PCR primer groups for different pathogenic bacteria are designed; one multiple PCR is established by utilizing the primer groups; the purpose of synchronously and quickly identifying the four pathogenic bacteria is realized; moreover, the sensibility is good, the specificity is high; relative to a conventional detection method, the cost needed by the PCR detection method is lower; the operation is simpler and more convenient; the needed time is shorter; the pertinence is high; the multiple PCR primer groups can be applied to the analysis and the detection of any one bacterial sample in multiple environments containing catfish edwardsiella ictaluri, aeromonas punctata, flavobacterium cloumnare and aeromonas hydrophila.

The invention discloses enrofloxacin chitosan nanoparticles which are prepared by using the following methods: 1, adding chitosan into an acetic acid solution, performing stirring dissolution, and performing swelling over night so as to obtain a chitosan gel; 2, dissolving sodium tripolyphosphate into deionized water so as to obtain a sodium tripolyphosphate solution; 3, adding enrofloxacin into the chitosan gel, performing uniform stirring, adding NaOH to adjust the pH value to 5.0, continuing stirring, adding dropwise the sodium tripolyphosphate solution, when light blue opalescence is generated when the sodium tripolyphosphate solution is added dropwise, and continuing stirring for 1 hour so as to obtain an enrofloxacin chitosan nanoparticle mixed suspension; and 4, drying the enrofloxacin chitosan nanoparticle mixed suspension, so as to obtain the enrofloxacin chitosan nanoparticles. The nanoparticles are simple in preparation method, free of adverse smell, good in palatability andvery good in killing function on catfish edwardsiella tarda.

The invention relates to the field of aquatic product vaccinology, in particular to a vibrio harveyi, vibrio anguillarum and Edwardsiella tarda triple inactivated vaccine, and preparation and application thereof. The triple inactivated vaccine contains inactivated vibrio harveyi T4, vibrio anguillarum C312 and Edwardsiella tarda TX1. The highest relative percent survival (RPS) of the obtained inactivated vaccine for the vibrio harveyi, the vibrio anguillarum and the Edwardsiella tarda can be respectively 89.29%, 85.71% and 76.67%, and the RPS still can be about 50% in six months after immunization is carried out. The obtained vaccine has a simple preparation process and a low construction cost, does not have potential safety hazards, has a good immunization effect and can prevent and control three kinds of different-species pathogen infection through one-time immune inoculation.

The invention relates to a Chinese herbal medicinal bait for preventing and controlling Ictalurus punctatus Edwardsiellasis, and a preparation method thereof. The Chinese herbal medicinal bait is prepared through processing the following components, by weight, 3-5 parts of a nucleic acid yeast extract product, 0.5-1 part of composite vitamins, 10-15 parts of a Chinese herbal medicine extract paste, 40-60 parts of an enteric polymer, and 5-8 parts of vitamin E acetate; and the Chinese herbal medicine extract paste is obtained through processing the following Chinese herbal medicinal raw materials, by weight, 5-8 parts of Andrographis paniculata, 5-8 parts of Weeping Forsythia, 5-12 parts of radix bupleuri, 8-12 parts of Artemisia argyi, 5-8 parts of Radix Codonopsis, 4-8 parts of Radix Astragali and 7-15 parts of honeysuckle flower. The Chinese herbal medicinal bait has the advantages of reasonable compatibility, balanced nutrition, low processing cost, good palatability and feed attraction property, high Ictalurus punctatus utilization rate, few metabolites, substantial improvement of the immunity of the Ictalurus punctatus, great reduction of the Edwardsiellasis infection probability of the Ictalurus punctatus, no drug residual or drug resistance, and good safety.

The present invention is directed to a novel live attenuated isolate and sub-isolets thereof of a strain of the pathogen Edwardsiella ictaluri for protecting fish from ESC infection or disease, a vaccine composition comprising the isolet, a method of oral delivery of the vaccine and any vaccine, and an apparatus designed to effectively mix the vaccine with fish feed and deliver the vaccine to fish.

The invention discloses a recombinant plasmid pMDMcherry, a construction method of the recombinant plasmid and a method for marking Edwardsiella ictaluri with a red fluorescent protein gene. A base sequence of the recombinant plasmid pMDMcherry is shown as SEQ ID NO:7. The Edwardsiella ictaluri, in which a pMDMcherry recombinant vector is introduced, shows red fluorescence when observed under a fluorescence microscope. A stability test shows that a stability rate of the plasmid can reach 100% when the recombinant bacterium is transferred to the 25th generation. With the introduction of the red fluorescent protein gene, a simple and intuitive relation is provided for researching a relation between the Edwardsiella ictaluri and a host.

The invention provides application of Edwardsiella ictaluri to the preparation of fish vaccines and a fish vaccine. The fish vaccine comprises effective dose of immunogenic inactivated Edwardsiella ictaluri and pharmaceutically acceptable auxiliary materials or carriers. The vaccine or combined vaccine of the invention can pertinently prevent and treat red head disease of Pseudobagrus fulvidraco with good effect and is a low-cost and environmental-friendly prevention method.

The invention belongs to the technical field of biology, and particularly relates to a pelteobagrus fulvidraco red head disease resistance character related SNP marker, a screening method and an application. A CypA gene is analyzed based on pelteobagrus fulvidraco genome information and a mixed pool sequencing technology to obtain a potential SNP locus, and primer amplification is designed for verification. Association analysis is carried out on the potential SNP loci and the red head disease of the pelteobagrus fulvidraco to find that one SNP has significant difference, and allele mutation (T / C) exists on the 528th nucleotide of the CypA genome sequence of the SNP. The SNP 528T / C also has significant difference in the pelteobagrus fulvidraco of another culture group. Besides, a transgeniczebra fish line expressing the yellow pelteobagrus fulvidraco CypA is constructed, and toxicity attacking experiments prove that the transgenic zebra fish has better resistance to Edwardsiella ictaluri, and further illustrates the true effectiveness of the correlation between the SNP site of the pelteobagrus fulvidraco CypA gene and the pelteobagrus fulvidraco ''red head disease'' resistance.

The invention provides a kit for detecting fish pathogenic bacteria; the kit comprises a primer with a nucleotide sequence as shown in SEQ ID NO. 1-2, and the primer is used to amplify genes of edwardsiella ictaluri. The invention also provides a method for detecting fish pathogenic bacteria, and an application of the primer shown in SEQ ID NO. 1-2. The kit provided by the invention can rapidly and accurately detect a pathogenic bacterium of ictalurus punctatus - edwardsiella ictaluri, and thus realizes pertinent control and detection of ictalurus punctatus enteric septicemia, and the invention has wide application prospects.

The invention discloses nucleic acid molecule, an expression vector and an application thereof, an antibacterial drug and a method thereof, which belong to the technical field of biology. A nucleotidesequence of the nucleic acid molecule disclosed in the present invention is shown as SEQ ID NO. 1. The study of the present invention finds that the nucleic acid molecule shown in SEQ ID NO. 1 is overexpressed in fish body, which can significantly inhibit bacterial proliferation and increase the survival rate of the fish body during bacterial infection; for example, the nucleic acid molecule shown as SEQ ID NO. 1 is overexpressed in zebrafish, which can significantly inhibit the proliferation of edwardsiella tarda in zebrafish and significantly improve the survival rate of zebrafish during infection with edwardsiella tarda; the nucleic acid molecule shown as SEQ ID NO. 1 has broad application prospects in the field of aquaculture, can be used for preparing antibacterial drugs or disease-resistant biological products, and improves the antibacterial ability of fish body.

The invention belongs to the technical field of antibacterial agents for water environment, and discloses a compound plant extract for Edwardsiella tarda in water environment. The compound plant extract is mainly made of the following components in mass percentage: Extract 55%-65%, Portulaca oleracea extract 26%-35%, Jinbuchang stem extract 12%-18%. Compared with the safety of traditional GMP disinfectants, the compound plant extract of the present invention belongs to environment-friendly substances, and has a large safe concentration range for scaleless fish. When splashed with water, it has obvious inhibitory effect on Edwardsiella tarda, And it will not be disturbed by organic matter in the water body. When the dosage is ≤5mg / L, it can effectively inhibit the abnormal reproduction of Edwardsiella tarda in the upper, middle and bottom layers of the aquaculture water body, and it is effective against the yellow catfish, channel catfish and channel catfish. Security is good.

The callicarpa nudiflora is used as the adjuvant of the fish vaccine, the effective component of the callicarpa nudiflora is callicarpa nudiflora, the application of the adjuvant can improve the specific protection rate after vaccination, and the adjuvant is used by injection after being mixed with an edwardsiella tarda inactivated vaccine. The callicarpa nudiflora is used as the fish vaccine adjuvant, and has the advantages of low price, low residue, no drug resistance of fish and the like.