Genetic and Phenotypic Virulence Potential of Non-O1/Non-O139 Vibrio cholerae Isolated from German Retail Seafood

Abstract

Non-O1 and non-O139 Vibrio cholerae (NOVC) can cause gastrointestinal infections in humans. Contaminated food, especially seafood, is an important source of human infections. In this study, the virulence potential of 63 NOVC strains isolated from retail seafood were characterized at the genotypic and phenotypic levels. Although no strain encoded the cholera toxin (CTX) and the toxin-coregulated pilus (TCP), several virulence factors, including the HlyA hemolysin, the cholix toxin ChxA, the heat-stable enterotoxin Stn, and genes coding for the type 3 and type 6 secretion systems, were detected. All strains showed hemolytic activity against human and sheep erythrocytes: 90% (n = 57) formed a strong biofilm, 52% (n = 33) were highly motile at 37 °C, and only 8% (n = 5) and 14% (n = 9) could resist ≥60% and ≥40% human serum, respectively. Biofilm formation and toxin regulation genes were also detected. cgMLST analysis demonstrated that NOVC strains from seafood cluster with clinical NOVC strains. Antimicrobial susceptibility testing (AST) results in the identification of five strains that developed non-wildtype phenotypes (medium and resistant) against the substances of the classes of beta-lactams (including penicillin, carbapenem, and cephalosporin), polymyxins, and sulphonamides. The phenotypic resistance pattern could be partially attributed to the acquired resistance determinants identified via in silico analysis. Our results showed differences in the virulence potential of the analyzed NOVC isolated from retail seafood products, which may be considered for further pathogenicity evaluation and the risk assessment of NOVC isolates in future seafood monitoring.

Non-O1/Non-O139 Vibrio cholerae—An Underestimated Foodborne Pathogen? An Overview of Its Virulence Genes and Regulatory Systems Involved in Pathogenesis

“…We were able to show that genes encoding these virulence factors are also present in NOVC isolated from seafood and the environment in previous studies [30,31]. Further investigations showed that other virulence genes are also present in the NOVC genomes which could also play a role in the infection process, such as hapA for hemagglutinin protease; mshA for mannose sensitive hemagglutinin; and frhA, gbpA, and mam7 for non-specific adhesins [30].…”

“…The combined virulence factors identified in the genomes of NOVC isolated from the above-mentioned infection cases and local outbreaks are the Vibrio pathogenicity islands VSP-2 and VPI-2, genomic islands (GI) encoding type III (T3SS) and type VI secretion systems (T6SS), enterotoxins (RtxA and Stn), and the hemolysin HlyA. We were able to show that genes encoding these virulence factors are also present in NOVC isolated from seafood and the environment in previous studies [30,31]. Further investigations showed that other virulence genes are also present in the NOVC genomes which could also play a role in the infection process, such as hapA for hemagglutinin protease; mshA for mannose sensitive hemagglutinin; and frhA, gbpA, and mam7 for non-specific adhesins [30].…”

“…The intracellular concentration of c-di-GMP can be increased in the presence of bile [65]. The presence of vpsR in NOVC was confirmed by Dua et al (2018) [66], and variations in VpsR between O1/O139 V. cholerae and NOVC were identified in our previous studies, including point mutation and gene fragment deletions [30]. However, 99% of NOVCs could form stable biofilms in our previous studies, and all the biofilm-relevant genes were present in NOVCs [30,31].…”

“…In V. cholerae, the cadABC operon first described in Escherichia (E.) coli is important for protecting the bacteria from acid hydrolysis [40]. The genetic presence of the cadABC operon was identified in 90% of NOVC isolated from seafood and the environment in our previous studies, with genetic identities over 90% compared to the V. cholerae O1 El Tor biotype, suggesting a fully functional cadABC operon [30,31]. In particular, the cadA gene encodes a lysine decarboxylase that binds protons through the production of cadaverine and carbon dioxide.…”

“…In the stomach, nitrite from food and saliva that is exposed to the acidic milieu results in acidified nitrite, which can be reduced by reactive nitrogen species (RNS) to antimicrobially active nitric oxide. By the detoxification of RNS, the expression of the genes nnrS and hmpA plays an important role in V. cholerae, and these genes were also identified in NOVCs from seafood and the environment [30,31]. Both genes encode enzymes that are capable of destroying nitric oxide.…”