In this paper, we developed a rapid and accurate method for the detection of Vibrio parahaemolyticus strains, using multiplex PCR and DNA--DNA hybridization. Multiplex PCR was used to simultaneously amplify three diag...In this paper, we developed a rapid and accurate method for the detection of Vibrio parahaemolyticus strains, using multiplex PCR and DNA--DNA hybridization. Multiplex PCR was used to simultaneously amplify three diagnostic genes （tlh, tdh andfla） that serve as molecular markers of V. parahaemolyticus. Biotinylated PCR products were hybridized to primers immobilized on a microarray, and detected by chemiluminesce with avidin-conjugated alkaline phosphatase. With this method, forty-five samples were tested. Eight known virulent strains （tlh＋/tdh＋/fla＋） and four known avirulent strains （tlh＋/tdh /fla＋） of the V. parahaemolyticus were successfully detected, and no non-specific hybridization and cross-hybridization reaction were found from fifteen closely-related strains （tlh-/tdh-/fla＋） of the Vibrio spp. In addition, all the other eighteen strains of non-Vibrio bacteria （tlh-/tdh /fla-） gave negative results. The DNA microarray successfully distinguished V. parahaemolyticus from other Vibrio spp. The results demonstrated that this was an efficient and robust method for identifying virulent strains of V. parahaemolyticus.展开更多
Objective To establish an ultra-sensitive,ultra-fast,visible detection method for Vibrio parahaemolyticus(VP).Methods We established a new method for detecting the tdh and trh genes of VP using clustered regularly int...Objective To establish an ultra-sensitive,ultra-fast,visible detection method for Vibrio parahaemolyticus(VP).Methods We established a new method for detecting the tdh and trh genes of VP using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 12a(CRISPR/Cas12a)combined with recombinase polymerase amplification and visual detection(CRISPR/Cas12a-VD).Results CRISPR/Cas12a-VD accurately detected target DNA at concentrations as low as 10^(-18)M(single molecule detection)within 30 min without cross-reactivity against other bacteria.When detecting pure cultures of VP,the consistency of results reached 100%compared with real-time PCR.The method accurately analysed pure cultures and spiked shrimp samples at concentrations as low as 10^(2)CFU/g.Conclusion The novel CRISPR/Cas12a-VD method for detecting VP performed better than traditional detection methods,such as real-time PCR,and has great potential for preventing the spread of pathogens.展开更多
Objective:To identify a potential bacterium which produces antimicrobial peptide(vibriocin),and its purification,characterization and production optimization.The bacteria subjected in the study were isolated from a hi...Objective:To identify a potential bacterium which produces antimicrobial peptide(vibriocin),and its purification,characterization and production optimization.The bacteria subjected in the study were isolated from a highly competitive ecological niche of mangrove ecosystem.Methods:The bacterium was characterized by phenotype besides 16S rRNA gene sequence analysis.The antibacterial activity was recognised by using agar well diffusion method.The vibriocin was purified using ammonium sulphate precipitation,butanol extraction,gel filtration chromatography,ion-exchange chromatography and subsequently,by HPLC.Molecular weight of the substance identified in SDS-PAGE.Production optimization performed according to Taguchi's mathematical model using 6 different nutritional parameters as variables.Results:The objective bacterium was identified as Vibrio parahaemolyticm.The vibriocin showed 18 KDa of molecular mass with mono peptide in nature and highest activity against pathogenic Vibrio harveyi.The peptide act stable in a wide range of pH,temperature.UV radiation,solvents and chemicals utilized.An overall^20%of vibriocin production was improved,and was noticed that NaCl and agitation speed played a vital role in secretion of vibriocin.Conclusion:The vibriocin identified here would he an effective alternative for chemically synthesized drugs for the management of Vibrio infections in mariculture industry.展开更多
[ Objective ] The dynamic change of heterobacteria and vibrios in larvae industrialized culture system was studied to provide scientific reference for healthy cultivation of shrimp. [ Method ] The heterobacteria, vibr...[ Objective ] The dynamic change of heterobacteria and vibrios in larvae industrialized culture system was studied to provide scientific reference for healthy cultivation of shrimp. [ Method ] The heterobacteria, vibrios and pathogenic vibrio parahaemolyticus were monitored in larvae industrialized culture system. [ Result] The heterobacteria, vibrios and pathogenic vibrio parahaemolyticus were the most in fertilized eggs of shrimp but the least in nauplius, then their number would increase with growth. During whole rearing period, both boterobacteria in larvae, vibrios in water would increase by one order of magnitude, while both vibrios in larvae and heterobacteria in water would increase by two orders of magnitude. There were many heterobacteria and vibrios but few vibrio parahaemolyticus in living bait. The correlation coefficients between larvae and heterobacteria and vibrios in water were 0. 704 and 0. 840 in culture system respectively, while the correlation among heterobacteria, vibrios in living bait and larvae, water were weak or negative. [ Conclusion ] There was a dynamic relation between water and larvae in rearing period, and restrictly control of culture condition would restrain the occurrence of disease caused by vibrio parahaemolyticus, besides that bacteria number in bait was not obviously correlated with bacteria nubmer in culture system.展开更多
[ Objective] To investigate the in vitro bacteriostatic activity of Chinese herbal medicines （ rhubarb, Baikal skullcap root, honeysuckle, eucommia bark and Chinese date） and their compounds against two common patho...[ Objective] To investigate the in vitro bacteriostatic activity of Chinese herbal medicines （ rhubarb, Baikal skullcap root, honeysuckle, eucommia bark and Chinese date） and their compounds against two common pathogens of Scylla serrata （ Vibrio parahaemolyticus and Aeromonas hydrophila）. [Method] Drug sensitivity of the bacteria was determined by paper method. Minimum inhibitory concentration （MIC） and minimum bactericidal concentration （MBC） were determined by dilution method and plate method, respectively. [Result] The bacteriostatic order of single medicines on V. parahaemolyticus were Baikal skullcap root 〉 honeysuckle 〉 rhubarb 〉 Chinese date 〉 eucommia bark. The bacteriostatic order of the compounds were A =B = C 〉 D. Among these medicines, the Baikal skullcap root and the compounds A, B, C and D had better bactericidal effects on V. parahaemolyticus. The bacteriostatic order of single medicines on A. hydrophila was rhubarb 〉 Baikal skullcap root 〉 honeysuckle 〉 Chinese date 〉 eucommia bark. The bacteriostatic order of the compounds were A 〉 B 〉 C = D, while the rhubarb and the compound C had better bactericidal effects on A. hydrophila. [ Conclusion] The Chinese herbal medicines including rhubarb, Baikal skullcap root, honeysuckle, eucommia bark and Chinese date have good bacteriostatic effects on V. parahaemolyticus and A. hydrophila, and they can be used to treat bacterial infection in Scylla serrata.展开更多
Vibrio parahaemolyticus, the leading cause of seafood-borne gastroenteritis, has the ability to form biofilms on biotic and abiotic surfaces. Biofilm formation is a complicated process involving many specific structur...Vibrio parahaemolyticus, the leading cause of seafood-borne gastroenteritis, has the ability to form biofilms on biotic and abiotic surfaces. Biofilm formation is a complicated process involving many specific structures and regulatory processes. The most significant of the structures and processes include polar and lateral flagella, mannose-sensitive hemagglutinin typeⅣpili, chitin-regulated pili,capsular polysaccharide (CPS), exopolysaccharide展开更多
Summary What is already known on this topic?Vibrio parahaemolyticus(V.parahaemolyticus)is frequently resistant to common antimicrobials such as ampicillin and generally highly susceptible to most clinically used antim...Summary What is already known on this topic?Vibrio parahaemolyticus(V.parahaemolyticus)is frequently resistant to common antimicrobials such as ampicillin and generally highly susceptible to most clinically used antimicrobials.What is added by this report?V.parahaemolyticus were highly resistant to cefazolin and ampicillin:94.4% and 37.0%,respectively.展开更多
基金financial supports from National High Technology Research and Development Program of China(No.2007AA10Z430)National Natural Science Foundation of China(No.30700535)Program for New Century Excellent Talents in Fujian Province University,and Fok Ying Tong Education Foundation(No.111032)
文摘In this paper, we developed a rapid and accurate method for the detection of Vibrio parahaemolyticus strains, using multiplex PCR and DNA--DNA hybridization. Multiplex PCR was used to simultaneously amplify three diagnostic genes （tlh, tdh andfla） that serve as molecular markers of V. parahaemolyticus. Biotinylated PCR products were hybridized to primers immobilized on a microarray, and detected by chemiluminesce with avidin-conjugated alkaline phosphatase. With this method, forty-five samples were tested. Eight known virulent strains （tlh＋/tdh＋/fla＋） and four known avirulent strains （tlh＋/tdh /fla＋） of the V. parahaemolyticus were successfully detected, and no non-specific hybridization and cross-hybridization reaction were found from fifteen closely-related strains （tlh-/tdh-/fla＋） of the Vibrio spp. In addition, all the other eighteen strains of non-Vibrio bacteria （tlh-/tdh /fla-） gave negative results. The DNA microarray successfully distinguished V. parahaemolyticus from other Vibrio spp. The results demonstrated that this was an efficient and robust method for identifying virulent strains of V. parahaemolyticus.
基金supported by the National Key Research and Development Plan of China[2018YFC1602500]the Natural Science Foundation of Tianjin[19JCZDJC39900]
文摘Objective To establish an ultra-sensitive,ultra-fast,visible detection method for Vibrio parahaemolyticus(VP).Methods We established a new method for detecting the tdh and trh genes of VP using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 12a(CRISPR/Cas12a)combined with recombinase polymerase amplification and visual detection(CRISPR/Cas12a-VD).Results CRISPR/Cas12a-VD accurately detected target DNA at concentrations as low as 10^(-18)M(single molecule detection)within 30 min without cross-reactivity against other bacteria.When detecting pure cultures of VP,the consistency of results reached 100%compared with real-time PCR.The method accurately analysed pure cultures and spiked shrimp samples at concentrations as low as 10^(2)CFU/g.Conclusion The novel CRISPR/Cas12a-VD method for detecting VP performed better than traditional detection methods,such as real-time PCR,and has great potential for preventing the spread of pathogens.
基金Supported by the project Drug from Sea,funded by Ministry of Earth Sciences,Govt.of India(Grant#34030020005)
文摘Objective:To identify a potential bacterium which produces antimicrobial peptide(vibriocin),and its purification,characterization and production optimization.The bacteria subjected in the study were isolated from a highly competitive ecological niche of mangrove ecosystem.Methods:The bacterium was characterized by phenotype besides 16S rRNA gene sequence analysis.The antibacterial activity was recognised by using agar well diffusion method.The vibriocin was purified using ammonium sulphate precipitation,butanol extraction,gel filtration chromatography,ion-exchange chromatography and subsequently,by HPLC.Molecular weight of the substance identified in SDS-PAGE.Production optimization performed according to Taguchi's mathematical model using 6 different nutritional parameters as variables.Results:The objective bacterium was identified as Vibrio parahaemolyticm.The vibriocin showed 18 KDa of molecular mass with mono peptide in nature and highest activity against pathogenic Vibrio harveyi.The peptide act stable in a wide range of pH,temperature.UV radiation,solvents and chemicals utilized.An overall^20%of vibriocin production was improved,and was noticed that NaCl and agitation speed played a vital role in secretion of vibriocin.Conclusion:The vibriocin identified here would he an effective alternative for chemically synthesized drugs for the management of Vibrio infections in mariculture industry.
基金the National High Technology Research and Development Program of China (863 Program) (2006AA10A406)National Key Technology Research and Development Program of China (2006BAD01A13)Public Agriculture Specific Research Program (nyhyzx07-042)~~
文摘[ Objective ] The dynamic change of heterobacteria and vibrios in larvae industrialized culture system was studied to provide scientific reference for healthy cultivation of shrimp. [ Method ] The heterobacteria, vibrios and pathogenic vibrio parahaemolyticus were monitored in larvae industrialized culture system. [ Result] The heterobacteria, vibrios and pathogenic vibrio parahaemolyticus were the most in fertilized eggs of shrimp but the least in nauplius, then their number would increase with growth. During whole rearing period, both boterobacteria in larvae, vibrios in water would increase by one order of magnitude, while both vibrios in larvae and heterobacteria in water would increase by two orders of magnitude. There were many heterobacteria and vibrios but few vibrio parahaemolyticus in living bait. The correlation coefficients between larvae and heterobacteria and vibrios in water were 0. 704 and 0. 840 in culture system respectively, while the correlation among heterobacteria, vibrios in living bait and larvae, water were weak or negative. [ Conclusion ] There was a dynamic relation between water and larvae in rearing period, and restrictly control of culture condition would restrain the occurrence of disease caused by vibrio parahaemolyticus, besides that bacteria number in bait was not obviously correlated with bacteria nubmer in culture system.
基金funded by the Transformation Project of Scientific Achievements in Guangdong Colleges (cgzhzd0812)
文摘[ Objective] To investigate the in vitro bacteriostatic activity of Chinese herbal medicines （ rhubarb, Baikal skullcap root, honeysuckle, eucommia bark and Chinese date） and their compounds against two common pathogens of Scylla serrata （ Vibrio parahaemolyticus and Aeromonas hydrophila）. [Method] Drug sensitivity of the bacteria was determined by paper method. Minimum inhibitory concentration （MIC） and minimum bactericidal concentration （MBC） were determined by dilution method and plate method, respectively. [Result] The bacteriostatic order of single medicines on V. parahaemolyticus were Baikal skullcap root 〉 honeysuckle 〉 rhubarb 〉 Chinese date 〉 eucommia bark. The bacteriostatic order of the compounds were A =B = C 〉 D. Among these medicines, the Baikal skullcap root and the compounds A, B, C and D had better bactericidal effects on V. parahaemolyticus. The bacteriostatic order of single medicines on A. hydrophila was rhubarb 〉 Baikal skullcap root 〉 honeysuckle 〉 Chinese date 〉 eucommia bark. The bacteriostatic order of the compounds were A 〉 B 〉 C = D, while the rhubarb and the compound C had better bactericidal effects on A. hydrophila. [ Conclusion] The Chinese herbal medicines including rhubarb, Baikal skullcap root, honeysuckle, eucommia bark and Chinese date have good bacteriostatic effects on V. parahaemolyticus and A. hydrophila, and they can be used to treat bacterial infection in Scylla serrata.
基金supported by the National Natural Science Foundation of China [81601809]the Natural Science Foundation of Jiangsu Province [BK20160505]
文摘Vibrio parahaemolyticus, the leading cause of seafood-borne gastroenteritis, has the ability to form biofilms on biotic and abiotic surfaces. Biofilm formation is a complicated process involving many specific structures and regulatory processes. The most significant of the structures and processes include polar and lateral flagella, mannose-sensitive hemagglutinin typeⅣpili, chitin-regulated pili,capsular polysaccharide (CPS), exopolysaccharide
基金supported by the National Science and Technology Major Project of China(Grant agreement 2018ZX10733402).
文摘Summary What is already known on this topic?Vibrio parahaemolyticus(V.parahaemolyticus)is frequently resistant to common antimicrobials such as ampicillin and generally highly susceptible to most clinically used antimicrobials.What is added by this report?V.parahaemolyticus were highly resistant to cefazolin and ampicillin:94.4% and 37.0%,respectively.
