DOI: 10.5061/dryad.600nd
title: Data from: Cholera outbreaks (2012) in three districts of Nepal reveal clonal transmission of multi-drug resistant Vibrio cholerae O1

description: Background: Although endemic cholera causes significant morbidity and
 mortality each year in Nepal, lack of information about the causal
 bacterium often hinders cholera intervention and prevention. In 2012,
 diarrheal outbreaks affected three districts of Nepal with confirmed cases
 of mortality. This study was designed to understand the drug response
 patterns, source, and transmission of Vibrio cholerae associated with 2012
 cholera outbreaks in Nepal. Methods: V. cholerae (n = 28) isolated from
 2012 diarrhea outbreaks {n = 22; Kathmandu (n = 12), Doti (n = 9), Bajhang
 (n = 1)}, and surface water (n = 6; Kathmandu) were tested for
 antimicrobial response. Virulence properties and DNA fingerprinting of the
 strains were determined by multi-locus genetic screening employing
 polymerase chain reaction, DNA sequencing, and pulsed-field gel
 electrophoresis (PFGE). Results: All V. cholerae strains isolated from
 patients and surface water were confirmed to be toxigenic, belonging to
 serogroup O1, Ogawa serotype, biotype El Tor, and possessed classical
 biotype cholera toxin (CTX). Double-mismatch amplification mutation assay
 (DMAMA)-PCR revealed the V. cholerae strains to possess the B-7 allele of
 ctx subunit B. DNA sequencing of tcpA revealed a point mutation at amino
 acid position 64 (N [rightwards arrow] S) while the ctxAB promoter
 revealed four copies of the tandem heptamer repeat sequence
 5[prime]-TTTTGAT-3[prime]. V. cholerae possessed all the ORFs of the
 Vibrio seventh pandemic island (VSP)-I but lacked the ORFs 498-511 of
 VSP-II. All strains were multidrug resistant with resistance to
 trimethoprim-sulfamethoxazole (SXT), nalidixic acid (NA), and streptomycin
 (S); all carried the SXT genetic element. DNA sequencing and deduced amino
 acid sequence of gyrA and parC of the NAR strains (n = 4) revealed point
 mutations at amino acid positions 83 (S [rightwards arrow] I), and 85 (S
 [rightwards arrow] L), respectively. Similar PFGE (NotI) pattern revealed
 the Nepalese V. cholerae to be clonal, and related closely with V.
 cholerae associated with cholera in Bangladesh and Haiti. Conclusions: In
 2012, diarrhea outbreaks in three districts of Nepal were due to
 transmission of multidrug resistant V. cholerae El Tor possessing cholera
 toxin (ctx) B-7 allele, which is clonal and related closely with V.
 cholerae associated with cholera in Bangladesh and Haiti.
description: Dendrogram showing
 genomic fingerprints of V. cholerae O1 strains isolated in Nepal in
 2012.The fingerprint pattern
 in the gel was analyzed using a computer software package, Bionumeric
 (Applied Maths, Belgium). After background subtraction and gel
 normalization, the fingerprint patterns were typed according to banding
 similarity and dissimilarity as recommended by the manufacturer. , using
 the Dice similarity coefficient and unweighted-pair group method employing
 average linkage (UPGMA) clustering, by using pulsed-field gel
 electrophoresis (PFGE) images of NotI- restriction digested genomic DNA of
 the tested Nepalese V. cholerae strains; PFGE images of contemporary V.
 cholerae O1 El Tor strains from Haiti and Bangladesh, including El Tor
 reference strain N16961 were also included. The scale bar at the top
 (left) indicates similarity coefficient. The banding patterns and the
 similarity coefficient revealed 100% similarity for all the tested
 Nepalese, Haitian, and Bangladeshi V. cholerae strains included in the
 dendrogram, suggesting high level of clonal
 relatedness.FIg
 2.pptx