Cholera is still an important public health problem in several countries, including Thailand. In this study, a collection of clinical and environmental V. cholerae serogroup O1, O139, and non-O1/non-O139 strains originating from Thailand (1983 to 2013) was characterized to determine phenotypic and genotypic traits and to investigate the genetic relatedness. Using a combination of conventional methods and whole genome sequencing (WGS), 78 V. cholerae strains were identified. WGS was used to determine the serogroup, biotype, virulence, mobile genetic elements, and antimicrobial resistance genes using online bioinformatics tools. In addition, phenotypic antimicrobial resistance was determined by the minimal inhibitory concentration (MIC) test. The 78 V. cholerae strains belonged to the following serogroups O1: (n = 44), O139 (n = 16) and non-O1/non-O139 (n = 18). Interestingly, we found that the typical El Tor O1 strains were the major cause of clinical cholera during 1983–2000 with two Classical O1 strains detected in 2000. In 2004–2010, the El Tor variant strains revealed genotypes of the Classical biotype possessing either only ctxB or both ctxB and rstR while they harbored tcpA of the El Tor biotype. Thirty O1 and eleven O139 clinical strains carried CTXϕ (Cholera toxin) and tcpA as well four different pathogenic islands (PAIs). Beside non-O1/non-O139, the O1 environmental strains also presented chxA and Type Three Secretion System (TTSS). The in silico MultiLocus Sequence Typing (MLST) discriminated the O1 and O139 clinical strains from other serogroups and environmental strains. ST69 was dominant in the clinical strains belonging to the 7th pandemic clone. Non-O1/non-O139 and environmental strains showed various novel STs indicating genetic variation. Multidrug-resistant (MDR) strains were observed and conferred resistance to ampicillin, azithromycin, nalidixic acid, sulfamethoxazole, tetracycline, and trimethoprim and harboured variants of the SXT elements.For the first time since 1986, the presence of V. cholerae O1 Classical was reported causing cholera outbreaks in Thailand. In addition, we found that V. cholerae O1 El Tor variant and O139 were pre-dominating the pathogenic strains in Thailand. Using WGS and bioinformatic tools to analyze both historical and contemporary V. cholerae circulating in Thailand provided a more detailed understanding of the V. cholerae epidemiology, which ultimately could be applied for control measures and management of cholera in Thailand.

e Vibrio parahaemolyticus is a seafood-borne pathogenic bacterium that is a major cause of gastroenteritis worldwide. We investigated the genetic and evolutionary relationships of 101 V. parahaemolyticus isolates originating from clinical, human carrier, and various environmental and seafood production sources in Thailand using multilocus sequence analysis. The isolates were recovered from clinical samples (n ‫؍‬ 15), healthy human carriers (n ‫؍‬ 18), various types of fresh seafood (n ‫؍‬ 18), frozen shrimp (n ‫؍‬ 16), fresh-farmed shrimp tissue (n ‫؍‬ 18), and shrimp farm water (n ‫؍‬ 16). Phylogenetic analysis revealed a high degree of genetic diversity within the V. parahaemolyticus population, although isolates recovered from clinical samples and from farmed shrimp and water samples represented distinct clusters. The tight clustering of the clinical isolates suggests that disease-causing isolates are not a random sample of the environmental reservoir, although the source of infection remains unclear. Extensive serotypic diversity occurred among isolates representing the same sequence types and recovered from the same source at the same time. These findings suggest that the O-and K-antigen-encoding loci are subject to exceptionally high rates of recombination. There was also strong evidence of interspecies horizontal gene transfer and intragenic recombination involving the recA locus in a large proportion of isolates. As the majority of the intragenic recombinational exchanges involving recA occurred among clinical and carrier isolates, it is possible that the human intestinal tract serves as a potential reservoir of donor and recipient strains that is promoting horizontal DNA transfer, driving evolutionary change, and leading to the emergence of new, potentially pathogenic strains. Vibrio parahaemolyticus is a seafood-borne pathogenic bacterium and is the leading cause of traveler's diarrhea and gastroenteritis worldwide due to the consumption of undercooked contaminated seafood, particularly shellfish. The organism possesses a number of virulence factors, including a thermostable direct hemolysin (TDH) (1), a thermostable direct hemolysinrelated hemolysin (TRH) (2), and type 3 secretion system 1 (TTSS1) and TTSS2 (3). Strains that possess tdh, trh, and TTSS2-encoding genes are generally pathogenic and responsible for the vast majority of clinical cases. While such strains account for only 1 to 2% of isolates sampled from water and different marine species (4-6), they are known to have a widespread global distribution, having been reported from cases of infection in Japan, India, Thailand, Malaysia, China, Indonesia, the United Kingdom, Italy, and the East and Gulf Coasts of the United States (7-19).The public health and commercial burdens associated with V. parahaemolyticus contamination are very high in Thailand due to the wide consumption of seafood. Although undercooked seafood has been identified as a source of V. parahaemolyticus infection (20, 21), the relative likelihood of contamination from differ...

Acute hepatopancreatic necrosis disease (AHPND) is an emerging shrimp disease that causes massive die-offs in farmed shrimps. Recent outbreaks of AHPND in Asia have been causing great losses for shrimp culture and have become a serious socioeconomic problem. The causative agent of AHPND is Vibrio parahaemolyticus, which is typically known to cause food-borne gastroenteritis in humans. However, there have been few reports of the epidemiology of V. parahaemolyticus AHPND strains, and the genetic relationship among AHPND strains is unclear. Here, we report the genetic characterization of V. parahaemolyticus strains isolated from AHPND outbreaks in Thailand. We found eight isolates from AHPND-suspected shrimps and pond water that were positive for AHPND markers AP1 and AP2. PCR analysis confirmed that none of these eight AP-positive AHPND strains possesses the genes for the conventional virulence factors affecting to humans, such as thermostable direct hemolysin (TDH), TDH-related hemolysin (TRH) and type III secretion system 2. Phylogenetic analysis by multilocus sequence typing showed that the AHPND strains are genetically diverse, suggesting that AHPND strains were not derived from a single genetic lineage. Our study represents the first report of molecular epidemiology of AHPND-causing V. parahaemolyticus strains using multilocus sequence typing, and provides an insight into their evolutionary mechanisms.

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