The Characterisation of Vibrio parahaemolyticus isolates from Food Grade Shellfish and Their Susceptibility to Antimicrobials Relevant to Clinical or Farming Purposes
Creators: name: Capindale, Charlie
Publisher: name: University of Lincoln
Year: 2023

description: Litopenaeus vannamei and Penaeus monodon are two of the most widely aquaculture-farmed shrimp species in South East Asia and generate significant economic influx for countries involved in their production. In 2009 a disease emerged among farmed stocks of these species, decimating the industry with over US$1bn losses per annum recorded at its height. The disease, acute hepatopancreatic necrosis disease, or AHPND, continues to ravage SE Asia and has since appeared in several Mexican states. The aetiological agent involved in the disease is the short, pleomorphic, Gram-negative bacterium: Vibrio parahaemolyticus. Through acquisition of a plasmid harbouring a transposon encoding toxin duplex genes pirA and pirB the bacteria infects the shrimp via a faeco-oral route and spreads rapidly throughout the culture pond causing 40-100% mortality of stock. With historically unchecked use of antibiotics for disease treatment and prophylaxis, intensive farming represents one of the most pressing issues contributing to the development, spread, and persistence of antimicrobial resistance genes. As the outbreak of a bacterial epidemic often prompts the industry to turn to antibiotics, it is important that surveillance of antimicrobial resistance in bacteria from globally exported aquacultured species is routinely carried out.The present study aimed to isolate V. parahaemolyticus from several commercially available seafood sources, while focussing heavily on South East Asian farmed shrimp. V. parahaemolyticus was confirmed by selective culturing and the molecular identification of the tlh gene which is conserved across the species. Human toxin genes and blood agar were used to screen the presence of human pathogenicity determinants tdh and trh genes. Isolates were also screened for the presence of pirAB toxins through duplex PCR. To monitor the status of antibiotic susceptibility in the retrieved V. parahaemolyticus these isolates were screened for susceptibility to 9 antibiotics through microbroth dilution determination of minimum inhibitory concentration (MIC).42 isolates were identified as V. parahaemolyticus, none of the isolates were found to possess the genes encoding human pathogenicity toxins, nor did they possess haemolytic activity on Wagatsuma agar. From the dissected shrimp, 8 isolates were identified containing the pirAB toxin genes. Antibiotic resistance was found to vary through different batches of shrimp. There was also a degree of variance in the country of origin for the shrimp tested. For example, cefotaxime had a particularly high minimum inhibitory concentration when tested against the Bangladeshi isolates, relative to the Vietnamese isolates. All isolates tested in the study had explicitly high MIC values for ampicillin and colistin. As a follow up to the colistin data, the resistant isolates were screened for the presence of the mcr-1 gene, a rapidly disseminating plasmid-borne resistance mechanism. Out of all 42 isolates only one isolate, of Bangladeshi origin, produced a positive result for mcr-1 amplification. In this study parahaemolyticus was successfully and repeatedly isolated from shrimp from multiple origins. Although there was a complete absence of human toxigenicity genes, the rate of retrieval of isolates encoding pirAB AHPND (7 out of 42 isolates) toxigenicity far exceeded what was expected. Antimicrobial susceptibility determination against 9 compounds of clinical relevance or historical use in aquaculture returned interesting results for several compounds. The most important results were those of ampicillin and colistin chiefly, with cefotaxime, gentamicin, and kanamycin showing, lesser but still high, levels of resistance. The study highlights patterns of resistance and susceptibility that demonstrates the need for continued global surveillance on the role of antibiotics in intensively farmed SE Asian shrimp. To the best of the author’s knowledge this is the first study to identify AHPND (+) Vp present in UK seafood market circulation.