| In order to survey and analyze the antibiotic resistance and micro-evolution of V.parahaemolyticus isolated from clinical,acute hepatopancreatic necrosis disease infecting shrimps,and aquatic production,a total 61 V.parahaemolyticus isolates were studied for antibiotic susceptibility pattern using modified Kirby-Bauer disk diffusion method in 25 kinds of antibiotics.All V.parahaemolyticus isolates were also evaluated for the presence of potential virulence factors(tlh,tdh,trh)through PCR and genetic diversity through multilocus sequence analysis(MLST).The results showed that 61 isolates developed different degrees of resistance to the selected 25 antibiotics,of which at least 4 were resistant and up to 14 were resistant;At the same time,it was found that there may be a correlation between the pathogenic gene types and multidrug resistance The V.parahaemolyticus with the virulence gene of tdh-/trh+/tlh+ seems to develop resistance to more antibiotics.The MLST results revealed the genetic variation and correlation between the isolates,and found that there is a correlation between the strain's virulence genotype and microevolution,among the four different virulence genotypes,the isolates with the virulence genotype of tdh-/trh+/tlh+ are more dispersed in the phylogenetic tree than the isolates with genotype tdh+/trh-/tlh+.The low degree of clustering indicates that the kinship is relatively far.Combined with the heat map analysis,the antibiotic resistance spectrum of the isolates from aquatic products,clinical and diseased shrimps is different to some extent.The antibiotic resistance of aquatic products and disease shrimp isolates are stronger,and they are resistant to more kinds of antibiotics It is speculated that human activities and environmental selection pressure have an impact on the transformation of virulence from non-virulence or weak virulence to strong virulence;And it is important to take measures to strengthen monitor the antibacterial resistance of V.parahaemolyticus in the contaminated aquatic productsFood safety has always been a hot issue of global concern.However,foodborne pathogens,as the most important hazard,cause foodborne diseases and affect food safety In China,pathogenic microorganisms are the primary cause of food-borne diseases,and food-borne pathogens are the most common.At present,diseases caused by foodborne pathogens are mainly treated with antibiotics,however,the problem of resistance due to antibiotic abuse is becoming another focus of attention.In addition,China is a large import and export country of aquatic products in the world.According to the current production structure of aquatic products in China,most aquatic products come from aquaculture,while only less than 30%of aquatic products come from fishing.Vibrio parahaemolyticus is not only a common food-borne pathogen that can cause food poisoning and cause a series of food-borne diarrhea,but also the specific strain lead to a disease called early mortality syndrome(EMS)or more technically known as acute hepatopancreatic necrosis disease(AHPND).Once the disease outbreak will cause persistent large-scale death of shrimp in the infected area,causing huge economic losses to the shrimp farming industry.In order to explore the potential relationship between the antibiotic resistance with micro-evolution of V.parahaemolyticus,so as to provide data support for the control of foodborne diseases and aquatic products diseases caused by V.parahaemolyticus The following review and research work were done:(1)Genitic diversity and microevolution of V.parahaemolyticus isolated from clinical,acute hepatopancreatic necrosis disease infecting shrimps,and aquatic production;(2)Antibiotic resistance and microevolution of V.parahaemolyticus isolated from acute hepatopancreatic necrosis disease infecting shrimps and aquatic production.The main research content and specific results of this research are as follows:1.Genitic diversity and microevolution of V.parahaemolyticus isolated from clinical,acute hepatopancreatic necrosis disease infecting shrimps,and aquatic production To investigate the genetic and evolutionary relationships of V.parahaemolyticus in Shanghai and Guangdong,a total of 184 isolates from clinical samples(VPC,n=40),AHPND infecting shrimps(VPE,n=10),and various aquatic production sources(VPF,n=134)were collected and evaluated through multilocus sequence analysis(MLST).All V.parahaemolyticus isolates were also evaluated for the presence of potential virulence factors(tlh,tdh,trh)through Polymerase chain reaction(PCR)and single nucleotide polymorphisms(SNPs)through genomic sequencing.The analyses of virulence factors showed that the great majority of VPC(97.5%)possess tdh and/or trh,while most of VPF(83.58%)do not encode the hemolysin genes.We may thus hypothesize that the environment serves as a potential reservoir that promotes horizontal DNA transfer,driving evolutionary change,and in turn,lead to the emergence of novel,potentially pathogenic strains.The results of phylogenetic analyses show that VPF-112 is a non-pathogenic maternal strain isolated from aquatic products,with a relatively high evolutionary status.Besides,all the VPE strains and few VPC strains could be grouped into several small subgroups and evenly distributed on phylogenetic trees,which means that there was intraspecific or interspecific gene exchange between strains from different sources.It is possible that anthropogenic activities and environmental selective pressure may be the important determinant of the process of transforming strains from non-pathogenic bacteria to pathogenic bacteria.2.Antibiotic resistance and microevolution of V.parahaemolyticus isolated from acute hepatopancreatic necrosis disease infecting shrimps and aquatic production.In order to survey and analyze the antibiotic resistance and micro-evolution of V.parahaemolyticus isolated from clinical,acute hepatopancreatic necrosis disease infecting shrimps,and aquatic production,a total 61 V.parahaemolyticus isolates were studied for antibiotic susceptibility pattern using modified Kirby-Bauer disk diffusion method in 25 kinds of antibiotics.All V.parahaemolyticus isolates were also evaluated for the presence of potential virulence factors(tlh,tdh,trh)through PCR and genetic diversity through multilocus sequence analysis(MLST).The results showed that 61 isolates developed different degrees of resistance to the selected 25 antibiotics,of which at least 4 were resistant and up to 14 were resistant;At the same time,it was found that there may be a correlation between the pathogenic gene types and multidrug resistance The V.parahaemolyticus with the virulence gene of tdh-/trh+/tlh+ seems to develop resistance to more antibiotics.The MLST results revealed the genetic variation and correlation between the isolates,and found that there is a correlation between the strain's virulence genotype and microevolution,among the four different virulence genotypes,the isolates with the virulence genotype of tdh-/trh+/tlh+ are more dispersed in the phylogenetic tree than the isolates with genotype tdh+/trh-/tlh+.The low degree of clustering indicates that the kinship is relatively far.Combined with the heat map analysis,the antibiotic resistance spectrum of the isolates from aquatic products and diseased shrimps is different to some extent.The antibiotic resistance of disease shrimp isolates is stronger and resistant to more kinds of antibiotics.It is speculated that human activities and environmental selection pressure have an impact on the transformation of virulence from non-virulence or weak virulence to strong virulence;And it is important to take measures to strengthen monitor the antibacterial resistance of V.parahaemolyticus in the contaminated aquatic products. |
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