Genetic and phenotypic characterisation of clinical Vibrio cholerae O1 strains from Malaysia / Tan Yen Joo

Tan, Yen Joo (2018) Genetic and phenotypic characterisation of clinical Vibrio cholerae O1 strains from Malaysia / Tan Yen Joo. Masters thesis, University of Malaya.

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Abstract

Vibrio cholerae serogroup O1 and O139 cause cholera outbreaks globally. The O1 serogroup consists of classical and El Tor biotypes, which cause the fifth, sixth and seventh cholera pandemic. More recently, the sporadic Matlab variant of O1 strain that possess both the classical and El Tor biotype characteristics was found to be emerging. The hybrid and variant strains have been reported to cause cholera outbreaks in India, Pakistan, and Bangladesh, then recently in Kelantan, Malaysia. These hybrid and variant biotypes could be misidentified as El Tor biotype due to their combined characteristics of both the El Tor and classical biotypes. Therefore, this study aimed to characterise and re-identify the biotypes of forty-four V. cholerae O1 El Tor clinical isolates from different outbreaks in Sabah(n=27), Sarawak (n=15) and Klang (n=2) from year 2009 to 2012. The isolates were subjected to phenotypic and genotypic tests for determination of their biotype and genetic characteristics. In order to identify the phenotypic characteristics of these clinical V. cholerae O1 isolates, polymyxin B susceptibility test, Voges-Proskauer test, haemolysis test, and chicken blood cell agglutination test were performed. The presence of genes of tcpA, ctxA, ctxB, hlyA, rtxC, and rstC in the isolates were detected using PCR approach. The results showed that 42 out of the 44 (95%) V. cholerae O1 clinical isolates tested in this study possess all the six genes tested. Both of the isolates from Klang lacked the rstC gene. Notably, based on the phenotypic characteristics and ctxB gene sequence, all of the V. cholerae O1 outbreak clinical isolates examined in this study, which were previously identified as El Tor biotype, was re-identified as either belongs to hybrid or variant biotype. Of the 44 isolates examined, 13 (30%) isolates were classified as the hybrid biotype; while 31 (70%) isolates were identified to be the variant biotype. DNA fingerprinting using REP-PCR was also carried out to determine the relationship among the V. cholerae O1 clinical isolates in this study. Based on the results, four REP profiles (REP 1, REP 2, REP 3 and REP 4) were observed based on the dendrogram generated. REP 1 and REP 2 groups were predominant while the two strains of O139 serogroup demonstrated a distinct profile (REP 4). In conclusion, all of the forty-four V. cholerae clinical isolates from cholera outbreaks occurred in Sarawak, Sabah and Klang in year 2009 to 2012 were not of the typical 7th pandemic El Tor biotype but demonstrated the characteristics of the variant or hybrid biotype, which were reported to be emerging. The isolates causing cholera outbreaks in Sarawak and Sabah were found to be genetically close and demonstrated limited genetic variation based on the REP-PCR genotyping.

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