OBJECTIVETo understand the phenotypic diversity of toxigenic Vibrio cholerae O1 El Tor strains isolated from different provinces in China during the last 50 years.

METHODSTraditional biotyping testings including susceptibility to polymyxin B, sensitivity to group IV phage, Voges-Proskauer test and haemolysis of sheep erythrocytes were conducted.

RESULTSData from Biotype-specific phenotype analysis revealed that only 133 isolates carried the typical El Tor phenotypes while the other 251 isolates displayed atypical El Tor phenotypes. Combined with ctxB, rstR genotypes and phenotypic characteristics, 64 isolates were identified as typical El Tor biotype, 21 were El Tor variants that showing the typical El Tor biotype-specific phenotype but with ctxB(class). 280 isolates were defined as the hybrid groups with traits of both classical and El Tor biotypes that could be further classified into 45 groups, based on the combination of genotypes of ctxB, rstR and phenotypic characteristics.

CONCLUSIONToxigenic Vibrio cholerae O1 El Tor strains that isolated from different provinces in China displayed high phenotypic diversity. The traditional biotype traits could not be used to correctly distinguish the two different biotypes.

China ; Genotype ; Phenotype ; Polymorphism, Genetic ; Vibrio cholerae O1 ; genetics

OBJECTIVETo investigate the serologic type, phage-biotype and toxic factor of Vibrio cholerae isolated from different sea products, analyze the relation between the Vibrio cholerae in sea products and cholera epidemiology, and provide references for forecasting cholera epidemic situation and drawing out a preventing plan.

METHODThe biotype of strains isolated was analyzed by using type and phage-biotype serological methods. The toxic gene was detected by PCR.

RESULTSThe constituent ratio of V. cholerae O139, Ogawa and Inaba were, respectively, 48.44%, 20.31% and 31.25% in 64 strains of V. cholerae. The result of phage-biotype showed that the 26 strains of V. cholerae O1 were all non-epidemic strains. The result of toxic gene detecting showed that positive rate of V. cholerae O139 was higher than those of Ogawa and Inaba.

CONCLUSIONThe positive rate of toxic gene in V. cholerae O139 was high and the V. cholerae O139 was mainly in turtle, breed aquatics water and crustacean, so these sea products were the important sectors in cholera prevention and control.

Animals ; Bacteriophage Typing ; DNA, Bacterial ; genetics ; Seafood ; microbiology ; Serotyping ; Vibrio cholerae ; classification ; genetics ; isolation & purification ; Vibrio cholerae O1 ; isolation & purification ; Vibrio cholerae O139 ; isolation & purification

OBJECTIVEThrough systematic monitoring of the number and strain types of O1 and O139 Vibrio cholerae in the Pearl River estuary waters to analyze it's relevance with the temperature of environment, and the relevance between strains in water and isolates during outbreaks and epidemics as well as to estimate the methods used for environmental water detection and the potential role in cholera surveillance program.

METHODSTwenty-four stations along the Pearl River were selected and the water samples were collected monthly from March 2006 to February 2007. V. cholerae O1 and O139 strains were isolated from the samples. Real-time PCR established in our laboratory was used to detect V. cholerae O1 and O139. Air temperature and water temperature were collected during sampling. Pulsed field gel electrophoresis (PFGE) was applied in molecular typing of the isolates.

RESULTS862 water samples were collected during the study period. A total number of 77 O1 and O139 V. cholerae were isolated in 67 water samples and the positive rates were 7.77% for isolation and 26.33% for real-time PCR. Seasonal trend of positive rates by month were approximately coincident with the change of water temperature. The positive rates in the stations in urban area were higher than those in other areas. Toxigenic O139 strains were found in one station located in downstream of a marine market. Most of the O1 and O139 isolates were non-toxigenic. No trend of seasonal variation of the strains was noticed. Within these 75 isolates, 49 PFGE patterns were identified and the patterns differed widely with the similarity of 57.4% - 100%.

CONCLUSIONV. cholerae existed as the natural habitat in estuary water of the Pearl River and showed obvious genetic diversity. Data from monitoring waters might show the separation of strains with certain seasonal association. But the crowd did not show the relationship between the infections. Results from water surveillance program might provide indicators on the appearance of cholera pathogen which might be used in assessing the environmental risk of cholera epidemics as well as the alert of cholera.

Electrophoresis, Gel, Pulsed-Field ; Environmental Monitoring ; Phylogeny ; Polymerase Chain Reaction ; Seasons ; Temperature ; Vibrio cholerae O1 ; classification ; genetics ; isolation & purification ; Vibrio cholerae O139 ; classification ; genetics ; isolation & purification

OBJECTIVETo establish a duplex nested PCR assay system which is capable for detecting O1 and O139 groups of Vibrio cholerae simultaneously, and is applicable to environmental specimens from routine cholera surveillance.

METHODSBased on nucleic acid sequences available in GenBank, six sets of primers were designed by PrimerSelect program of DNAStar, targeting the rfb gene that encodes the O antigens of O1 and O139 V. cholerae, respectively. The specificity of several primer combinations was tested. A duplex nested PCR assay system for simultaneously detecting O1 and O139 V. cholerae was established, subsequently, its sensitivity, specificity, reproducibility and field evaluation were tested. The sensitivity of this assay was evaluated by comparing detection limits of nested PCR and conventional PCR. Its reproducibility was tested by 32 positive samples (11 samples positive for O1, 21 samples positive for O139) from environmental surveillance. In addition, the selected amplicons from positive samples were sequenced and analyzed with relevant sequences.

RESULTSThis newly-established duplex nested PCR assay might distinguish O1 V. cholerae from O139 V. cholerae, based on fragment lengths of amplicons, with reliable reproducibility, and no specific amplification was observed as compared with other vibrio species. The sensitivity of this nested PCR was (15 000) higher than conventional PCR, and there was no interference observed with multiple primers and complicated templates in the same vial. In its field evaluation, 32 positive DNA samples were detected and be further confirmed with double or triple tests, implying reliable reproducibility and consistency of this system. These results indicated that this assay had reliable reproducibility. No amplification was observed in all negative specimens and also suggested the acceptable specificity of this assay. Sequence analysis of the selected amplification products revealed 100% homogeneous with relevant genes from V.cholerae, indicating that these amplicons were originated from V. cholerae.

CONCLUSIONThis duplex nested PCR assay system should be rapid, sensitive and especially applicable to small laboratories, and be suitable for dynamic environmental surveillance.

DNA, Bacterial ; Environmental Monitoring ; methods ; Polymerase Chain Reaction ; methods ; Sequence Analysis, DNA ; Vibrio cholerae O1 ; genetics ; isolation & purification ; Vibrio cholerae O139 ; genetics ; isolation & purification

Objective: To analyze the etiological and epidemiological characteristics of Vibrio cholerae in Beijing during 2015-2021 and provide evidence for the prevention and control of cholera. Methods: The V. cholerae strains isolated in Beijing during 2015-2021 were analyzed by serotyping and virulence genes detection. Pulsed field gel electrophoresis (PFGE) was performed for the molecular typing of the strains. Based on the collected epidemiological and clinical data of cholera cases,the epidemiological characteristics of cholera were analyzed by descriptive epidemiology method. Results: A total of 76 Vibrio cholerae O1 strains were isolated in Beijing during 2015-2021, including 61 strains from human, 10 strains from environment and 5 strains from seafood. The 76 strains consisted of 68 Ogawa strains and 8 Inaba strains. Six Ogawa strains isolated from sporadic cases carried ctxAB. After NotⅠ digestion, 76 strains were divided into 33 PFGE patterns. From 2015 to 2021, a total of 38 cholera epidemics were reported in Beijing, most of them were sporadic ones, accounting for 92.11% (35/38). A total of 45 cases were reported, and the cases occurred during June-September accounted for 97.78% (44/45). Cholera cases occurred in 9 districts of Beijing, and the cases reported in Chaoyang district accounted for 42.22% (19/45) and in Changping district accounted for 31.11% (14/45). The age of the cholera cases ranged from 19 to 63 years. Except for one case with unknown clinical symptoms, 44 cases had diarrhea symptoms with 84.09% (37/44) of the cases reporting diarrhea (3-9 times/day), followed by yellow watery stool (95.45%, 42/44), abdominal pain (68.18%, 30/44), nausea and vomiting (40.91%, 18/44) and fever (36.36%, 16/44). Conclusion: Vibrio cholerae strains isolated in Beijing during 2015-2021 were mainly O1 serotype Ogawa,most of which were non-toxigenic. The PFGE of the strains varied. Cholera epidemics occurred in 9 districts of Beijing, but most were sporadic ones with incidence peak during June-September.
Adult ; Beijing/epidemiology* ; Cholera/epidemiology* ; Diarrhea/epidemiology* ; Electrophoresis, Gel, Pulsed-Field ; Humans ; Middle Aged ; Vibrio cholerae O1/genetics* ; Young Adult

OBJECTIVETo study the molecular characteristics and antibiotic resistances of Vibrio cholerae (V. cholerae) O1 isolates in Hangzhou in 2009.

METHODSThe virulence genes ctxA and tcpA of the thirty V. cholerae O1 isolates from 7 counties and districts of Hangzhou were detected by PCR. Pulsed-field gel electrophoresis (PFGE) was performed for molecular typing and similarity analysis. Antibiotic resistances of these isolates were measured by the Kirby-Bauer method.

RESULTSVirulence gene analysis showed that 80.00% (24/30) of the genotype in V. cholerae isolates was ctxA- and tcpA+, 13.33% (4/30) was ctxA- and tcpA-, and 6.67% (2/30) was ctxA+ and tcpA+. Twenty-seven isolates tested were typed into 11 PFGE patterns (P1-P11). Twenty-three isolates with genotype ctxA- and tcpA+ were clustered into 7 PFGE patterns (P1-P7, termed P1-like cluster) with the similarity to be equal or greater than 91.4%, and 56.52%(13/23) of them belong to P1. 7 isolates with very high similarity (97.6%), belonging to P1 (6 isolates), and P2 (1 isolate), respectively, were collected from one foodborne disease outbreak. The resistant rates of the 24 isolates with genotype ctxA- and tcpA+ to ampicillin, tobramycin and amikacin were 20.83% (5/24), 4.17% (1/24) and 4.17% (1/24), respectively.

CONCLUSIONThe genotype of the epidemic strains of V. cholerae O1 isolates in Hangzhou in 2009 with high similarity was ctxA- and tcpA+; The level of drug resistances of this kind of V. cholerae O1 isolates were not high.

China ; Cholera Toxin ; genetics ; Drug Resistance, Bacterial ; genetics ; Electrophoresis, Gel, Pulsed-Field ; Genes, Bacterial ; Genotype ; Humans ; Molecular Typing ; Vibrio cholerae O1 ; drug effects ; genetics ; isolation & purification

OBJECTIVETo study the distribution of serotype and the positive rate of toxins among vibrio cholerae non-O(1) isolated from environmental waters in Foshan city.

METHODSWater specimens were collected from river and cultured for vibrio cholerae non-O(1). The PCR method was used to detect cholerae enterotoxin (CT) gene; the ELISA method was used to detect heat-stable toxin (ST) and heat-labile toxin (LT).

RESULTS478 vibrio cholerae non-O(1) strains were isolated from 1 644 water specimens, with a positive rate of 29.07%. Serological assay showed that the main serotype of vibrio cholerae non-O(1) in Foshan city is VBO(7). Positive rate of CT, ST and LT were 1.91%, 13.14% and 12.17%, respectively.

CONCLUSIONSA few non-O(1) strains were found to have several virulent factors simultaneously, and the results suggest that vibrio cholerae non-O(1) in environmental waters is potentially pathogenic and may affect people's health. It is necessary to pay attention to the prevention of diarrhoea caused by vibrio cholerae.

China ; Enterotoxins ; genetics ; Environmental Monitoring ; methods ; Enzyme-Linked Immunosorbent Assay ; Polymerase Chain Reaction ; Seasons ; Serotyping ; Vibrio cholerae non-O1 ; classification ; genetics ; isolation & purification ; Water ; analysis ; Water Microbiology

OBJECTIVETo understand the genetic structures and variations of the superintegron (SI) in Vibrio cholerae isolated in the seventh cholera pandemic.

METHODSPolymerase chain reaction scanning and fragment sequencing were used. Sixty toxigenic V. cholerae O1 El Tor strains isolated between 1961 and 2008 were analyzed.

RESULTSSome variations were found, including insertions, replacements, and deletions. Most of the deletions were probably the result of recombination between V. cholerae repeat sequences. The majority of the variations clustered together. The SIs of the strains isolated in the 1960s and 1970s showed more diversity, whereas SI cassette variations in strains isolated in the 1990s and after were lower, with ∼24 kb signature sequence deletion. This indicates the predominant SI in the host during the epidemic in the 1990s and after. The insertion cassettes suggested the mobilization from the SIs of other V. cholerae serogroups and Vibrio mimicus.

CONCLUSIONThe study revealed that structural variations of SIs were obvious in the strains isolated in epidemics in different decades, whereas the divergence was based on syntenic structure of SIs in these El Tor strains. Also, the continuing cassette flows in the SIs of the host strains during the seventh cholera pandemics were displayed.

Cholera ; epidemiology ; microbiology ; Chromosomes, Bacterial ; genetics ; ultrastructure ; Cluster Analysis ; DNA, Bacterial ; genetics ; Electrophoresis, Gel, Pulsed-Field ; Gene Deletion ; Gene Flow ; Genetic Variation ; Humans ; Integrons ; genetics ; Mutagenesis, Insertional ; Open Reading Frames ; genetics ; Polymerase Chain Reaction ; Tandem Repeat Sequences ; Vibrio cholerae O1 ; classification ; genetics ; isolation & purification ; ultrastructure