216 results about "Vibrio parahaemolyticus" patented technology

The invention discloses an antibacterial peptide and application thereof, and belongs to the technical field of biology. The antibacterial peptide with an inhibition effect on various clinically common pathogenic microorganisms is obtained, and the antibacterial peptide comprises gram-negative bacteria such as escherichia coli, klebsiella pneumoniae, pseudomonas aeruginosa, acinetobacter baumannii, vibrio parahaemolyticus, salmonella gallinarum, salmonella typhimurium and enterobacter sakazakii, and gram-positive bacteria. The antibacterial peptide provided by the invention can be used as an antibacterial peptide, such as staphylococcus aureus, listeria monocytogenes and bacillus cereus, and the antibacterial peptide provided by the invention is novel in structure and easy to prepare.

The invention discloses a fermentation method of recombinant bacillus subtilis for expressing vibrio parahaemolyticus outer membrane protein, which comprises the following steps: inoculating a recombinant bacillus subtilis strain into a liquid culture medium for activation and amplification to obtain a seed solution; transferring the seed solution into a fermentation tank according to the inoculum size of 5%-10%, controlling the temperature to be 35-37 DEG C and the pH to be 6.8-7.2, introducing sterile air and stirring, and monitoring the thallus concentration OD600 value in real time; when the OD600 value reaches 35-38, 10% of an inducer is added for pre-induction; when the OD600 value reaches 40-45, the remaining 90% of the inducer is added for main induction, and the real-time dissolved oxygen value is collected; when the real-time dissolved oxygen value is continuously higher than a first preset threshold value, the flow acceleration rate of the inducer is increased, the fermentation temperature is increased at the same time or step by step, and the increase amplitude of the dissolved oxygen value is in positive correlation with the flow acceleration rate and the temperature increase amplitude; when the OD600 value is stabilized in a preset interval, it is judged that fermentation is completed, and the recombinant bacillus subtilis thalli are obtained. The yield and quality of the vibrio parahaemolyticus outer membrane protein can be effectively improved.

The invention discloses a lytic bacteriophage vBVpaSR40Z and application of the lytic bacteriophage vBVpaSR40Z to prevention and control of vibrio parahaemolyticus in an aquaculture environment. The preservation number of the vibrio parahaemolyticus phage vBVpaSR40Z is as follows: GDMCC No: 67440-B1, and the vibrio parahaemolyticus phage vBVpaSR40Z is The R40Z can be used for cracking multiple strains of vibrio parahaemolyticus and vibrio alginolyticus, has the capability of adapting to an aquaculture environment, and keeps relatively high activity within the range of 4-25 DEG C and pH of 4-10; no virulence gene or drug-resistant gene is detected through genome analysis, and phylogenetic and whole genome comparison shows that the similarity between R40Z and a related phage is lower than a species level threshold value, so that R40Z is a new phage species; the R40Z can obviously inhibit the growth of a host under various infection complex numbers. The R40Z has the advantages of strong splitting capacity, good environmental stability, high genome safety, unique evolutionary characteristics and the like, and is an aquatic vibriosis biological prevention and control bacteriophage with application potential.

The invention belongs to the technical field of gene engineering, and particularly relates to ALFPm3 modified antibacterial peptide, a molecular design and construction method and application of the ALFPm3 modified antibacterial peptide, and the amino acid sequence of the ALFPm3 modified antibacterial peptide is selected from any one of SEQ ID NO: 1-SEQ ID NO: 7. Analysis and design are carried out by combining the structure and action site of vibrio parahaemolyticus and the molecular structure and function relationship of the antibacterial peptide ALFPm3 as a basic template, the antibacterial activity of the antibacterial peptide ALFPm3 is improved, and other biological activities such as hemolysis are reduced.

The invention discloses vibrio parahaemolyticus deoxyribozyme screening and label-free biosensor development. The vibrio parahaemolyticus deoxyribozyme screening and label-free biosensor development comprises the following steps: (1) obtaining a vibrio parahaemolyticus deoxyribozyme sequence; (2) optimizing reaction conditions of the vibrio parahaemolyticus label-free biosensor; and (3) detecting the vibrio parahaemolyticus. The detection principle of the sensor is as follows: when vibrio parahaemolyticus exists, deoxyribozyme is triggered to be self-cleaved, a generated cleaved short chain can be used as a primer for starting rolling circle amplification (RCA), a large number of long-chain DNA products with repeated series G-quadruplex sequences are generated by amplifying a designed RCA annular template, G-quadruplex and ThT can form a G-quadruplex / ThT compound, and the long-chain DNA products can be used for detecting the vibrio parahaemolyticus. The fluorescence is generated under the excitation wavelength of 439 nm, and the higher the concentration of the vibrio parahaemolyticus is, the higher the fluorescence intensity of the system is, so that the simple, convenient, rapid, low-cost and high-sensitivity detection of the vibrio parahaemolyticus is realized.

The present application belongs to the field of aquaculture technology, and mainly relates to a traditional Chinese medicine composition for preventing and treating acute hepatopancreas necrosis disease of prawn, a preparation method and application. The traditional Chinese medicine composition comprises the following raw materials in parts by weight: 28-32 parts of Radix Sanguisorbae, 28-32 parts of Radix Scutellariae, 18-22 parts of Phellodendri Cortex and 8-12 parts of Lonicerae Japonicae Flos. The traditional Chinese medicine composition can significantly inhibit the activity of the main pathogen of the disease, i.e. Vibrio parahaemolyticus, improve the immune function and disease resistance of prawn, and further repair the hepatopancreas and intestinal tissues of the diseased prawn, thereby achieving the effect of effectively preventing and treating the acute hepatopancreas necrosis disease of prawn.

The invention relates to the field of microbiological detection, in particular to a method for detecting vibrio parahaemolyticus by combining RPA-CRISPR / Cas12a with a microfluidic biosensor. According to the method, the preloaded freeze-drying reagent is combined with centrifugally driven liquid operation, so that rapid and closed system detection is realized, and the pollution risk is effectively reduced. According to the invention, quantitative detection of vibrio parahaemolyticus is realized in a centrifugal micro-fluidic chip by systematically optimizing an RPA amplification primer and crRNA. The detection method shows high specificity and high sensitivity to non-target vibrio and other food-borne bacteria, and the lower detection limit is 6.08 copies / mu L. In a word, the invention provides a rapid, portable and sensitive platform, can realize quantitative detection in a wide linear range, provides a practical tool for real-time monitoring of pathogens, and has application prospects in monitoring of aquaculture biosafety and marine product safety.

The invention belongs to the technical field of biology, and particularly discloses a high-temperature-resistant antibacterial protein Cystatin B sourced from marine fish as well as a preparation method and application of the high-temperature-resistant antibacterial protein Cystatin B. The high-temperature-resistant antibacterial protein Cystatin B is derived from pomfret, and the amino acid sequence of the high-temperature-resistant antibacterial protein Cystatin B is as shown in SEQ ID NO. 1; the nucleotide sequence of the gene for coding the high-temperature-resistant antibacterial protein Cystatin B is as shown in SEQ ID NO.2, and the sequences of primers adopted for cloning the gene are as shown in SEQ ID NO.3-4. The high-temperature-resistant antibacterial protein Cystatin B has a remarkable antibacterial effect on gram-negative bacteria such as escherichia coli, vibrio parahaemolyticus and edwardsiella tarda as well as gram-positive bacteria such as staphylococcus aureus; and after being treated at the temperature of 100 DEG C, the antibacterial agent still can keep relatively high antibacterial activity. The antibacterial protein is simple in preparation method, can replace traditional antibiotics, and can quickly and efficiently prevent and treat various common bacterial diseases in aquaculture.

The invention relates to an extraction method and application of hippophae rhamnoides leaf polyphenol, and belongs to the technical field of food functional component extraction. According to the method, the hippophae rhamnoides leaf polyphenol is extracted by innovatively adopting a wall-breaking-step extraction system combining enzymatic pretreatment with ultrasonic induction, and the method is specifically characterized in that hippophae rhamnoides leaf powder is used as a substrate, and normal-temperature enzymolysis wall-breaking treatment is performed. Furthermore, a sound field is combined to promote non-thermal dissociation of secondary metabolites in the hippophae rhamnoides leaf powder, and a continuous process of'normal-temperature enzymolysis of cellulase-ultrasonic extraction 'is constructed. Then, adding an ethanol solution, and placing in a water bath for reaction extraction; and centrifugally freeze-drying to obtain the crude polyphenol of the sea buckthorn leaves. An antibacterial experiment shows that the hippophae rhamnoides leaf polyphenol crude extract has a remarkable antibacterial effect on food-borne pathogenic bacteria such as vibrio parahaemolyticus and aeromonas hydrophila, the technical scheme effectively keeps the biological activity of polyphenol, meanwhile, the solvent consumption and the oxidative degradation rate of heat-sensitive components are lower than those of a traditional process, and the method is suitable for development of natural preservatives and functional foods.

The invention belongs to the technical field of microbial antibacterial preparations, and particularly discloses a novel strong vibrio parahaemolyticus bacteriophage isolated strain vBVpSCC6 and application of the novel strong vibrio parahaemolyticus bacteriophage isolated strain vBVpSCC6. The bacteriophage vBVpSCC6 is preserved in the Guangdong Microbial Culture Collection Center on September 14, 2020, and the preservation number is GDMCC (China General Microbiological Culture Collection Center) NO: 61189-B1. The bacteriophage has good tolerance to environments such as temperature, pH, ultraviolet rays and the like, and has a good prevention and control effect on aquatic product pollution caused by vibrio parahaemolyticus. The bacteriophage has a lysis effect on multi-drug-resistant vibrio parahaemolyticus, can replace antibiotics to play a bacteriostatic role, effectively solves the problem of drug resistance faced by current antibiotic treatment, and has relatively good utilization value and application prospect. The bacteriophage can also be prepared into a composition with other bacteriophages to play a synergistic effect, and the bacteriophage can be applied to wider-spectrum antibacterial or bacteriostatic application.

The invention discloses a multiplex fluorescent quantitative PCR (polymerase chain reaction) detection kit for prawn culture pathogenic bacteria, which is characterized in that the kit contains primers and probes for detecting the prawn culture pathogenic bacteria, wherein the primers and the probes are respectively used for detecting TDH-related hemolsin genes of vibrio parahaemolyticus, ORF107 genes of white spot syndrome viruses, small subunit ribosomal genes of shrimp enterocytozoon hepatopenaei, 37 kDa coat protein genes of infectious subcutaneous and hematopoietic necrosis viruses, DNA (deoxyribonucleic acid) primer genes of full-eye iridovirus 1 and PirA [pi] genes of pathogens of bacterial prawns acute hepatopancreas necrosis; the method has the advantage that the detection of eight prawn culture pathogens can be realized at the same time. The method comprises the following steps of: obtaining a tcdB gene of a high-pathogenicity vibrio and a hemolsin gene of a vibrio harveyi;

The invention discloses a primer probe composition for detecting prawn disease related pathogens, a kit and application of the primer probe composition. The primer probe composition comprises a first primer probe group used for detecting vibrio parahaemolyticus, a second primer probe group used for detecting a tetrapod iridovirus 1, a third primer probe group used for detecting vibrio vitriocultatus and a fourth primer probe group used for detecting enterocytozoon hepatopenaei. The primer probe composition has the characteristics of high sensitivity, good repeatability and strong specificity, and can be used for specifically detecting vibrio parahaemolyticus, tetrapod iridovirus 1, vibrio vitrioculi and enterocytozoon hepatopenaei.

The invention discloses a nano antibody for detecting vibrio parahaemolyticus as well as a preparation method and application thereof, and aims to provide a nano antibody for detecting vibrio parahaemolyticus, which is high in detection accuracy, strong in specificity and simple in obtaining method. The amino acid sequence of the nano antibody is as shown in SEQ ID NO. 1, SEQ ID NO. 2, SEQ ID NO. 3, SEQ ID NO. 4 or SEQ ID NO. 5. Belongs to the technical field of biological detection.

The invention belongs to the field of prevention and control of bacterial diseases of aquatic animals, and discloses an acid-resistant broad-spectrum vibrio parahaemolyticus bacteriophage LVPP58 and application of the acid-resistant broad-spectrum vibrio parahaemolyticus bacteriophage LVPP58 in vibrio disease prevention and control, and the preservation number of the bacteriophage is CCTCC M 2025322. The bacteriophage is a broad-spectrum vibrio parahaemolyticus bacteriophage, not only can split vibrio parahaemolyticus, but also can split vibrio alginolyticus, the bacteriophage LVPP58 is stable in titer at the pH of 3-11 and the temperature of 30-60 DEG C, the bacteriophage and antibiotics are combined for use, a synergistic effect can be generated, the treatment effect is enhanced, the occurrence rate of drug resistance is reduced, and a new strategy is provided for solving the problem of bacterial drug resistance.

The invention belongs to the field of microbial culture, and particularly relates to an enrichment medium for reducing expression of surface polysaccharide of vibrio parahaemolyticus. According to the formula of the polysaccharide low-expression enrichment culture medium, nutritional ingredients in a specific proportion and specific polysaccharide expression inhibition conditions are selected, so that the expression quantity of surface lipopolysaccharide and capsular polysaccharide after enrichment of vibrio parahaemolyticus is lower than the expression quantity of polysaccharide after enrichment of normally cultured vibrio parahaemolyticus; expression of lipopolysaccharide and capsular polysaccharide on the surface layer can be inhibited to a certain extent, so that outer membrane protein can be better exposed on the surface of a thallus, and immunodetection of vibrio parahaemolyticus is facilitated. The invention provides a technical basis for developing a complete vibrio parahaemolyticus immunodetection method.

The invention relates to the technical field of microbiological detection, and particularly discloses a primer probe combination for detecting six food-borne pathogens and application of the primer probe combination. The detection primer probe combination for the six food-borne pathogens comprises six pairs of specific primer pairs and six probes, the nucleotide sequences of the specific primer pairs are respectively shown as SEQ ID NO.1, SEQ ID NO.4, SEQ ID NO.8, SEQ ID NO.1, SEQ ID NO.13, SEQ ID NO.16, SEQ ID NO.19, SEQ ID NO.22, SEQ ID NO.26, SEQ ID NO.29, SEQ ID NO.32 and SEQ ID NO.35, and the nucleotide sequences of the probes are respectively shown as SEQ ID NO.37-SEQ ID NO.42; the six food-borne pathogens comprise listeria monocytogenes, salmonella, vibrio parahaemolyticus, norovirus, escherichia coli O157: H7 and staphylococcus aureus. According to the present invention, the pathogens of six common food-borne diseases can be detected, the detection method has characteristics of high specificity and high sensitivity, and the pathogens do not interfere with each other and do not inhibit each other during the detection so as to achieve good stability.

The invention relates to an on-site synchronous quantitative determination method for three pathogenic bacteria in marine sediments, and belongs to the technical field of pathogenic bacteria detection.The three pathogenic bacteria are vibrio parahaemolyticus, vibrio alginolyticus and escherichia coli O157, and the method comprises the steps that marine sediment samples are collected and put into a centrifugal tube filled with sterile seawater, and are evenly mixed and centrifuged; taking the supernate as a template for LAMP amplification, respectively adding the supernate into detection tubes of an LAMP reaction system preloaded with the three pathogenic bacteria, and sealing. Placing the detection tube into a detection channel of a portable electronic isothermal gene amplification instrument, and determining a delta C4-t curve, a dv / dt-t curve and Tp; and after the determination is finished, substituting the Tp value into a working curve formula to calculate the content / concentration of the three pathogenic bacteria of the corresponding marine sediment sample in the detection tube. According to the method disclosed by the invention, Bst DNA polymerase and betaine are not needed in the reaction, so that the cost of LAMP analysis is reduced by at least 50%, the operation is simple and convenient, and simultaneous quantitative determination of the three pathogenic bacteria can be completed on site within 1h.

The invention belongs to the technical field of pathogenic microorganism screening, and particularly relates to vibrio parahaemolyticus capable of causing prawn vibrio vitreovibriosis and application of the vibrio parahaemolyticus, and the pathogenic bacteria can be used for preparing inactivated vaccines or used as the pathogenic bacteria to screen medicine components with the effect of preventing and treating the prawn vibrio vitreovibriosis or a culture method. The preservation number of the vibrio parahaemolyticus provided by the invention is CGMCC (China General Microbiological Culture Collection Center) The pathogenicity of the vibrio parahaemolyticus with high pathogenicity and strong lethality is about 1000 times stronger than that of the vibrio parahaemolyticus which is collected by the world animal hygiene organization and causes acute hepatopancreatic necrosis, and the strain is popular in main prawn culture areas and causes serious harm to the prawn fry culture industry in China. The vibrio parahaemolyticus provided by the invention can be used as a positive control for preparing a diagnosis product for the prawn vibrio vitreovirus disease; or preparing an immune vaccine after inactivation.

The invention relates to the technical field of microorganisms, and provides a vibrio owneri bacteriophage and application thereof, the vibrio owneri bacteriophage is vibrio owneri bacteriophage VOP240901, and the preservation number is CCTCC (China Center For Type Culture Collection) NO: M20242007. The Erwinia vibrio bacteriophage VOP240901 is a broad-spectrum bacteriophage, can split vibrio alginolyticus, vibrio parahaemolyticus and Erwinia vibrio, can stably exist in the environment with the temperature of 4-55 DEG C and the pH value of 3-11, is suitable for killing and preventing aquatic vibrio microorganisms, especially Erwinia vibrio, and has the sterilization rate of 90%.

The invention discloses ultrasensitive detection of vibrio parahaemolyticus by a CRISPR (clustered regularly interspaced short palindromic repeats) electrochemical biosensor based on PCR (polymerase chain reaction) and a hairpin DNA (deoxyribonucleic acid) probe, which comprises the following steps: preparing a gold nanoelectrode, modifying the gold nanoelectrode by using the hairpin DNA probe, performing polymerase chain reaction amplification to obtain a PCR amplification product, adding the PCR amplification product into a CRISPR system, uniformly oscillating, and detecting the vibrio parahaemolyticus by using the CRISPR electrochemical biosensor. Placing in a 37 DEG C constant-temperature metal bath for pre-reaction for 10 minutes, and dropwise adding the obtained reaction system to the surface of the hairpin DNA probe modified gold nanoelectrode. The method disclosed by the invention is high in overall repeatability and stability, the modification density of the surface of the electrode can be reduced, the steric hindrance effect is further reduced, the trans-cutting efficiency is improved, and in actual detection, the method shows remarkable sensitivity, is high in specificity, short in time consumption, low in cost and simple and convenient to operate, and can be applied to field detection.

The invention provides a primer combination of a multiple detection kit for food-borne pathogenic bacteria and the multiple detection kit. The primer combination comprises probes and primers for detecting nine food-borne pathogenic bacteria: vibrio cholerae general type, vibrio cholerae O1 type, vibrio cholerae O139 type, vibrio parahaemolyticus, salmonella, yersinia enterocolitica, campylobacter, listeria monocytogenes and enterobacter sakakaakii, and the sequence is shown as SEQ1-31; the probe and the primer pair are used for detecting nine characteristic genes, namely a shigella universal type: ipaH, a diarrheogenic Escherichia coli universal type: eltA, sta3, sta and aggR, and enterohemorrhagic Escherichia coli: stx1, stx2, escV and rfbE, and the sequences of the probe and the primer pair are shown as SEQ32-64. According to the invention, by combining a digital PCR multiple coding technology, the detection of 12 food-borne pathogenic bacteria can be realized, and the sensitivity, the specificity and the accuracy are relatively good.

The invention belongs to the technical field of gene engineering, and particularly relates to ALFPm3 modified antibacterial peptide, a molecular design and construction method and application of the ALFPm3 modified antibacterial peptide, and the amino acid sequence of the ALFPm3 modified antibacterial peptide is selected from any one of SEQ ID NO: 1-SEQ ID NO: 7. Analysis and design are carried out by combining the structure and action site of vibrio parahaemolyticus and the molecular structure and function relationship of the antibacterial peptide ALFPm3 as a basic template, the antibacterial activity of the antibacterial peptide ALFPm3 is improved, and other biological activities such as hemolysis are reduced.

The present invention discloses a Bacillus velezensis GY1, which was deposited in the Guangdong Provincial Microbial Culture Collection Center on June 28, 2024, with a deposit number of GDMCC No: 64792. The Bacillus velezensis GY1 provided by the present invention can degrade 100% and 56.4% of nitrogen in M9 culture medium containing 200 mg / L and 1000 mg / L ammonia nitrogen, respectively, and the ammonia reduction rate in the actual production of aquaculture water reaches 53.3%. It also has a strong inhibitory effect on common pathogens in aquatic products: Vibrio alginolyticus, Salmonella and Vibrio parahaemolyticus.

The invention discloses a kit for simultaneously detecting 16 food-borne pathogenic bacteria. Each group of probes is designed based on bacterial classification, virulence gene specificity or serotype marks of the pathogenic bacteria; a group A probe vibrio detection group is used for detecting vibrio cholerae universal type, vibrio cholerae O1 serotype, vibrio cholerae O139 serotype and vibrio parahaemolyticus; the B group of probe invasive enterobacteriaceae pathogen group is used for detecting escherichia coli O157: H7, shigella, salmonella and yersinia small intestine; the C group probe escherichia coli pathotype detection group is used for detecting enterotoxigenic escherichia coli ETEC, enteroinvasive escherichia coli EIEC, enteroaggregative adherent escherichia coli EAEC and pathogenic escherichia coli EPEC; and the D group of probes are used for detecting campylobacter jejuni, campylobacter colons, listeria monocytogenes and enterohemorrhagic escherichia coli EHEC in the campylobacter and toxin type pathogen group.

The invention belongs to the technical field of microorganisms, and discloses a bifidobacterium animalis subsp. Lactis XB02 strain capable of inhibiting pathogenic bacteria and improving inflammation, the preservation number of the bifidobacterium animalis subsp. Lactis XB02 strain is CGMCC (China General Microbiological Culture Collection Center) No.34942, and the preservation date is June 18, 2025. Meanwhile, the probiotic composition adopting the bacterial strain XB02 also comprises a lactobacillus rhamnosus XB01 bacterial strain. The XB02 strain disclosed by the invention has excellent bacteriostatic activity, especially for vibrio parahaemolyticus and gardneria; the atopic dermatitis symptom can be obviously improved; the intestinal mucosa permeability is reduced, and the intestinal barrier function is enhanced; and when the strain is combined with the XB01 strain for use, a synergistic sleep promoting effect is shown. The invention further provides a probiotic composition containing the strain, a preparation and application of the probiotic composition and the preparation in food, medicine or health care products.

The invention discloses bacillus velezensis Bve1910, which is preserved in the China Center for Type Culture Collection (CCTCC) and has the preservation number of CCTCC NO. M 20221965. The strain and metabolites thereof have strong antagonistic effects on more than ten types of marine culture animal pathogenic bacteria such as vibrio harveyi, vibrio erwiniae, vibrio canbayi, vibrio parahaemolyticus, vibrio alginolyticus, vibrio anguillarum, vibrio cholerae, vibrio shewanii, vibrio rotifer, pseudoalteromonas and photobacterium damarae, are strong in antibacterial activity and wide in antibacterial spectrum, and can be used for preparing marine culture animal pathogenic bacteria. According to the preparation method, the traditional Chinese medicine composition is simple in preparation method and rich and stable in metabolite, an ammonium sulfate precipitation extract, a hydrochloric acid precipitation methanol extraction extract, an ethyl acetate extraction extract and a chloroform extraction extract all have a broad-spectrum bacteriostatic effect, functional substances playing the broad-spectrum antagonistic bacteriostatic effect are compounded and diversified, and the directional selection effect on bacteria in the environment and ecology is not generated; and pathogenic bacteria are not easy to generate tolerance, so that the method has good application safety and ecological safety properties.

The invention discloses a lactobacillus acidophilus FMM01 with the effects of improving the contractility of pelvic floor muscles and repairing mucous membranes and application of the lactobacillus acidophilus FMM01, the lactobacillus acidophilus FMM01 is preserved in the China Center for Type Culture Collection, the preservation date is January 9, 2024, and the preservation number is CCTCC NO: M2024052. The lactobacillus acidophilus FMM01 has the advantages of self-adaption, high acid resistance, strong mucous membrane adhesiveness and no irritation to vaginal mucosa. The lyophilized powder is high in activity and excellent in stability; experimental results show that the lactobacillus acidophilus FMM01 can effectively inhibit bacteria, and especially can effectively inhibit enterobacter cloacae, vibrio parahaemolyticus and / or candida albicans. Besides, the lactobacillus acidophilus FMM01 can improve vaginal relaxation, improve pelvic floor muscle contractility and / or recover vaginal mucosa functions, can promote recovery of healthy florae after mixed vaginitis antibiotic / hormone treatment, improves the treatment effect of mixed vaginitis, and remarkably reduces the recurrence rate of mixed vaginitis.

The invention discloses a preparation method of a boiling antigen for vibrio parahaemolyticus serum agglutination. The preparation method comprises the following steps: selecting common serotype vibrio parahaemolyticus in monitoring for resuscitation; scraping a ring of placenta by using 10 mu L of an inoculating loop, dissolving the placenta in 1 mL of normal saline to prepare a bacterial suspension, and preparing 3 tubes of bacterial suspensions from each strain; boiling the bacterial suspension in boiling water for 1 hour, 45 minutes and 30 minutes respectively; and after boiling, centrifuging at 10000 rpm for 10 minutes, discarding the supernate, and reserving the precipitate as a boiling antigen for serum agglutination. The result shows that the agglutination accuracy after boiling for 1 hour is 100%, and the agglutination degree is 2 + or above; and no significant difference exists between 45 minutes of boiling and 1 hour of boiling, and no significant difference exists between 30 minutes of boiling. Compared with a high-pressure method, the method has the advantages that the operation time is shortened by 2-3 hours, special equipment certificates are not needed, and the method is suitable for daily detection and food poisoning traceability.

The application provides a bacillus desiccabilis strain XT23 microbial agent, a preparation method and application thereof, and belongs to the technical field of biological agents. A biological agent comprises a bacillus desiccabilis strain XT23, a fermentation treatment liquid of the bacillus desiccabilis strain XT23, or supernatant separated from the fermentation treatment liquid of the bacillus desiccabilis strain XT23. The fermentation treatment liquid is obtained by acid treatment and heat treatment of the fermentation liquid of the bacillus desiccabilis strain XT23, which effectively promotes spore formation, inactivates miscellaneous bacteria, significantly improves the stability and storage period of effective live bacteria, maximally retains antibacterial active substances in the fermentation liquid, and makes the inhibition effect of the final biological agent on Vibrio parahaemolyticus equivalent to that of fresh fermentation liquid. Meanwhile, the addition of the auxiliary material has a protective and slow-release effect on the active ingredients, and further enhances the convenience and effect persistence of application.

The invention provides a primer combination for a multiple detection kit for marine pathogenic bacteria and the multiple detection kit. The primer combination comprises primer pairs and probes which are used for detecting 13 marine pathogenic bacteria: vibrio alginolyticus, vibrio parahaemolyticus, vibrio cholerae, vibrio vulnificus, vibrio marine fish, vibrio river, vibrio menivibrio, vibrio mimicus, vibrio harveyi, vibrio occultus, vibrio horii, plesiomonas and aeromonas, and the sequence of the primer combination is as shown in SEQ1-SEQ39. According to the present invention, the genomic DNA of 13 marine pathogenic bacteria is identified and detected in one digital PCR reaction through the ultra-multiple digital PCR fluorescence coding technology, and the kit not only covers the common vibrio species such as vibrio cholerae, vibrio vulnificus, vibrio parahaemolyticus and the like, but also can identify the sea area vibrio, vibrio holii, vibrio occultus and the like, and the kit has good sensitivity, specificity and accuracy.