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Environmental and Clinical Strains of Vibrio cholerae Non-O1, Non-O139 From Germany Possess Similar Virulence Gene Profiles.

ABSTRACT: Vibrio cholerae is a natural inhabitant of aquatic ecosystems globally. Strains of the serogroups O1 and O139 cause the epidemic diarrheal disease cholera. In Northern European waters, V. cholerae bacteria belonging to other serogroups (designated non-O1, non-O139) are present, of which some strains have been associated with gastrointestinal infections or extraintestinal infections, like wound infections or otitis. For this study, environmental strains from the German coastal waters of the North Sea and the Baltic Sea were selected (100 strains) and compared to clinical strains (10 isolates) that were from patients who contracted the infections in the same geographical region. The strains were characterized by MLST and examined by PCR for the presence of virulence genes encoding the cholera toxin, the toxin-coregulated pilus (TCP), and other virulence-associated accessory factors. The latter group comprised hemolysins, RTX toxins, cholix toxin, pandemic islands, and type III secretion system (TTSS). Phenotypic assays for hemolytic activity against human and sheep erythrocytes were also performed. The results of the MLST analysis revealed a considerable heterogeneity of sequence types (in total 74 STs). The presence of virulence genes was also variable and 30 profiles were obtained by PCR. One profile was found in 38 environmental strains and six clinical strains. Whole genome sequencing (WGS) was performed on 15 environmental and 7 clinical strains that were ST locus variants in one, two, or three alleles. Comparison of WGS results revealed that a set of virulence genes found in some clinical strains is also present in most environmental strains irrespective of the ST. In few strains, more virulence factors are acquired through horizontal gene transfer (i.e., TTSS, genomic islands). A distinction between clinical and environmental strains based on virulence gene profiles is not possible for our strains. Probably, many virulence traits of V. cholerae evolved in response to biotic and abiotic pressure and serve adaptation purposes in the natural aquatic environment, but provide a prerequisite for infection of susceptible human hosts. These findings indicate the need for surveillance of Vibrio spp. in Germany, as due to global warming abundance of Vibrio will rise and infections are predicted to increase.

SUBMITTER: Schwartz K

PROVIDER: S-EPMC6474259 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Environmental and Clinical Strains of Vibrio cholerae Non-O1, Non-O139 From Germany Possess Similar Virulence Gene Profiles.

Schwartz Keike K Hammerl Jens Andre JA Göllner Cornelia C Strauch Eckhard E

Frontiers in microbiology 20190412

Vibrio cholerae is a natural inhabitant of aquatic ecosystems globally. Strains of the serogroups O1 and O139 cause the epidemic diarrheal disease cholera. In Northern European waters, V. cholerae bacteria belonging to other serogroups (designated non-O1, non-O139) are present, of which some strains have been associated with gastrointestinal infections or extraintestinal infections, like wound infections or otitis. For this study, environmental strains from the German coastal water ...[more]

PMID: 31031724

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Project description:Cholera caused by pathogenic Vibrio cholerae is still considered one of the major health problems in developing countries including those in Asia and Africa. Australia is known to have unique V. cholerae strains in Queensland waterways, resulting in sporadic cholera-like disease being reported in Queensland each year. We conducted virulence and antimicrobial genetic characterization of O1 and non-O1, non-O139 V. cholerae (NOVC) strains (1983 to 2020) from Queensland with clinical significance and compared these to environmental strains that were collected as part of a V. cholerae monitoring project in 2012 of Queensland waterways. In this study, 87 V. cholerae strains were analyzed where O1 (n = 5) and NOVC (n = 54) strains from Queensland and international travel-associated NOVC (n = 2) (61 in total) strains were sequenced, characterized, and compared with seven previously sequenced O1 strains and 18 other publicly available NOVC strains from Australia and overseas to visualize the genetic context among them. Of the 61 strains, three clinical and environmental NOVC serogroup strains had cholera toxin-producing genes, namely, the CTX phage (identified in previous outbreaks) and the complete Vibrio pathogenicity island 1. Phylogenetic analysis based on core genome analysis showed more than 10 distinct clusters and interrelatedness between clinical and environmental V. cholerae strains from Australia. Moreover, 30 (55%) NOVC strains had the cholix toxin gene (chxA) while only 11 (20%) strains had the mshA gene. In addition, 18 (34%) NOVC strains from Australia had the type three secretion system and discrete expression of type six secretion system genes. Interestingly, four NOVC strains from Australia and one NOVC strain from Indonesia had intSXT, a mobile genetic element. Several strains were found to have beta-lactamase (blaCARB-9) and chloramphenicol acetyltransferase (catB9) genes. Our study suggests that Queensland waterways can harbor highly divergent V. cholerae strains and serve as a reservoir for various V. cholerae-associated virulence genes which could be shared among O1 and NOVC V. cholerae strains via mobile genetic elements or horizontal gene transfer. IMPORTANCE Australia has its own V. cholerae strains, both toxigenic and nontoxigenic, that are associated with cholera disease. This study aimed to characterize a collection of clinical and environmental NOVC strains from Australia to understand their virulence and antimicrobial resistance profile and to place strains from Australia in the genetic context of international strains. The findings from this study suggest the toxigenic V. cholerae strains in the Queensland River water system are of public health concern. Therefore, ongoing monitoring and genomic characterization of V. cholerae strains from the Queensland environment are important and would assist public health departments to track the source of cholera infection early and implement prevention strategies for future outbreaks. Understanding the genomics of V. cholerae could also inform the natural ecology and evolution of this bacterium in natural environments.

Project description:Global warming has caused an increase in the emergence of Vibrio species in marine and estuarine environments as well as fresh water bodies. Over the past decades, antimicrobial resistance (AMR) has evolved among Vibrio species toward various antibiotics commonly used for the treatment of Vibrio infections. In this study, we assessed virulence and resistance patterns of Vibrio cholerae non-O1/non-O139 strains derived from Germany and other European countries. A total of 63 clinical and 24 environmental Vibrio cholerae non-O1/non-O139 strains, collected between 2011 and 2021, were analyzed. In silico antibiotic resistances were compared with resistance phenotypes according to EUCAST breakpoints. Additionally, genetic relatedness between isolates was assessed by two cgMLST schemes (SeqSphere +, pubMLST). Both cgMLST schemes yielded similar results, indicating high genetic diversity among V. cholerae non-O1/non-O139 isolates. Some isolates were found to be genetically closely related (allelic distance < 20), which suggests an epidemiological link. Thirty-seven virulence genes (VGs) were identified among 87 V. cholerae non-O1/non-O139 isolates, which resulted in 38 virulence profiles (VPs). VPs were similar between clinical and environmental isolates, with the exception of one clinical isolate that displayed a higher abundance of VGs. Also, a cluster of 11 environmental isolates was identified to have the lowest number of VGs. Among all strains, the predominant virulence factors were quorum sensing protein (luxS), repeats-in-toxins (rtxC/rtxD), hemolysin (hlyA) and different type VI secretion systems (T6SS) genes. The genotypic profiles revealed antibiotic resistance genes (ARGs) associated with resistance to beta-lactams, quinolones, macrolides, tetracycline, antifolate, aminoglycosides, fosfomycin, phenicols and sulfonamide. Carbapenemase gene VCC-1 was detected in 10 meropenem-resistant V. cholerae non-O1/non-O139 isolates derived from surface water in Germany. The proportion of resistance among V. cholerae non-O1/non-O139 species isolates against first line treatment (3rd generation cephalosporin, tetracycline and fluoroquinolone) was low. Empirical treatment would likely have been effective for all of the clinical V. cholerae non-O1/non-O139 isolates examined. Nevertheless, carbapenem-resistant isolates have been present in fresh water in Germany and might represent a reservoir for ARGs. Monitoring antimicrobial resistance is crucial for public health authorities to minimize the risks for the human population.

Project description:A non-O1 non-O139 Vibrio cholerae strain, 10259, belonging to the serogroup O53 was shown to harbor genes related to the vibrio pathogenicity island (VPI) and a cholera toxin (CT) genetic element called CTX. While the nucleotide sequence of the strain 10259 tcpA gene differed significantly (26 and 28%) from those of O1 classical and El Tor biotype strains, respectively, partial sequence analysis data of certain other VPI-associated genes (aldA, tagA, tcpP/H, toxT, acfB/C, and int) and intergenic regions (tcpF to toxT and tcpH to tcpA) of the strain showed only minor variations (0.4 to 4.8%) from corresponding sequences in O1 strains. Strain 10259 also contained CTX element-associated toxin genes with sequences almost identical to those of O1 strains. Growth of the organism in Luria broth (LB) under ToxR inducing conditions (30 degrees C and pH 6.5) led to transcriptional activation of tcpP/H, toxR, toxT, and tcpA genes, but not of ctxA, as determined by reverse transcription-PCR (RT-PCR). Subsequent analysis revealed that strain 10259 possessed only two copies (instead of three or more copies found in epidemic-causing O1 or O139 strains) of the heptanucleotide (TTTTGAT) repeats in the intergenic region upstream of ctxAB. Therefore, a strain 10259 mutant was generated by replacement of this region with a homologous region (1.4 kb) derived from a V. cholerae O1 classical biotype strain (O395) that contained seven such repeats. The resultant recombinant strain (10259R) was found to be capable of coordinately regulated expression of toxT, ctxA, and tcpA when grown under the ToxR inducing conditions. Serological studies also demonstrated that the recombinant strain produced TcpA and a significantly ( approximately 1,000-fold) higher level of CT in vitro compared to that of the parent strain. Virulence gene expression in two other non-O1 non-O139 strains (serogroup O37) containing VPI and the CTX element was studied by RT-PCR and serological assay. One strain (S7, which was involved in an epidemic in Sudan in 1968) showed coordinately regulated expression of virulence genes leading to the production of both CT and TcpA in LB medium. However, the other strain, V2, produced RT-PCR-detectable transcripts of toxT, ctxA, or tcpA genes in the early phase (6 h), but not in the late phase (16 h) of growth in LB medium. These results are consistent with the low levels of production of CT and TcpA by the strain that were serologically detectable. The significance of these results is discussed in relation to the role of virulence genes and their expression to the pathogenic potential of V. cholerae strains belonging to non-O1 serogroups.

Project description:Cholera toxin (CT)-producing Vibrio cholerae O1 and O139 cause acute diarrheal disease and are proven etiological agents of cholera epidemics and pandemics. On the other hand, V. cholerae non-O1/non-O139 are designated as non-agglutinable (NAG) vibrios and are not associated with epidemic cholera. The majority of NAG vibrios do not possess the gene for CT (ctx). In this study, we isolated three NAG strains (strains No. 1, 2, and 3) with ctx from pond water in Kolkata, India, and examined their pathogenic properties. The enterotoxicity of the three NAG strains in vivo was examined using the rabbit ileal intestinal loop test. Strain No. 1 induced the accumulation of fluid in the loop, and the volume of fluid was reduced by simultaneous administration of anti-CT antiserum into the loop. The volume of fluid in the loop caused by strains No. 2 and 3 was small and undetectable, respectively. Then, we cultured these three strains in liquid medium in vitro at two temperatures, 25°C and 37°C, and examined the amount of CT accumulated in the culture supernatant. CT was accumulated in the culture supernatant of strain No.1 when the strain was cultured at 25°C, but that was low when cultured at 37°C. The CT amount accumulated in the culture supernatants of the No. 2 and No. 3 strains was extremely low at both temperature under culture conditions examined. In order to clarify the virulence properties of these strains, genome sequences of the three strains were analyzed. The analysis showed that there was no noticeable difference among three isolates both in the genes for virulence factors and regulatory genes of ctx. However, vibrio seventh pandemic island-II (VSP-II) was retained in strain No. 1, but not in strains No. 2 or 3. Furthermore, it was revealed that the genotype of the B subunit of CT in strain No. 1 was type 1 and those of strains No. 2 and 3 were type 8. Histopathological examination showed the disappearance of villi in intestinal tissue exposed to strain No. 1. In addition, fluid accumulated in the loop due to the action of strain No. 1 had hemolytic activity. This indicated that strain No. 1 may possesses virulence factors to induce severe syndrome when the strain infects humans, and that some strains of NAG vibrio inhabiting pond water in Kolkata have already acquired virulence, which can cause illness in humans. There is a possibility that these virulent NAG vibrios, which have acquired genes encoding factors involved in virulence of V. cholerae O1, may emerge in various parts of the world and cause epidemics in the future.

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Environmental and Clinical Strains of Vibrio cholerae Non-O1, Non-O139 From Germany Possess Similar Virulence Gene Profiles.

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Environmental and Clinical Strains of <i>Vibrio cholerae</i> Non-O1, Non-O139 From Germany Possess Similar Virulence Gene Profiles.

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