Characterization of Novel Alleles of Toxin Co-Regulated Pilus A Gene (tcpA) from Environmental Isolates of Vibrio cholerae

Abstract

Vibrio cholerae is causative agent of life threatening diarrheal disease, cholera. The toxin co-regulated pilus (TCP) is a critical colonization factor of V. cholerae and it also serves as receptor for CTXФ. In this study, we describe nucleotide sequence of four novel alleles of tcpA gene from toxigenic and non-toxigenic V. cholerae isolated from environmental sources. The phylogenetic analysis of tcpA revealed that it is related to tcpA of newly emerged O1 strain and unrelated to tcpA of wild type (classical and El Tor strains). All strains showed variant tcpA and also harbored intact Vibrio Pathogenicity Island (VPI). The expression of all variant alleles was demonstrated by RT-PCR.

Occurrence in Mexico, 1998–2008, of Vibrio cholerae CTX + El Tor carrying an additional truncated CTX prophage

“…S1). The Mexican and Chinese strains with analogous tcpA sequences formed a separate cluster to the 10 clusters for tcpA alleles reported to date (11). Overall, results of the analysis clearly indicate that tcpA of Mex-2058 and Mex-3065 have a lineage distinct from the seventh pandemic ET.…”

“…Like CTXΦ, TCP is biotype-specific, with two distinct alleles of tcpA encoding the major protein "pilin" (10,11). In Mexico, the TCP variant of V. cholerae O1 strains carries a tcpA sequence that is different from tcpA types reported to date, including CL and ET (10,11), although an analog of the Mexican type tcpA deposited in GenBank is from a nontoxigenic O1, reportedly isolated in 2006 from China. V. cholerae O1 strains from Mexico carrying the variant tcpA are nontoxigenic.…”

“…Phylogenetic Analysis of tcpA. To determine the genetic relatedness of tcpA in the Mexican V. cholerae O1 strains, tcpA of representative ET, CL, and TCP variants was sequenced, using primers and PCR conditions as described previously (11). Multiple sequence alignment of trimmed tcpA (600 bp, encoding mature protein) and phylogenetic analysis of tcpA alleles were done using the ClustalW multiple alignment program and the Molecular Evolutionary Genetic Analysis program, version 5.05 (MEGA5), respectively.…”

“…Multiple sequence alignment of trimmed tcpA (600 bp, encoding mature protein) and phylogenetic analysis of tcpA alleles were done using the ClustalW multiple alignment program and the Molecular Evolutionary Genetic Analysis program, version 5.05 (MEGA5), respectively. TcpA sequences of two Mexican strains (Mex-2058 and Mex-3065) were used for construction of a phylogenetic tree with 33 tcpA sequences, of which 31 were of serogroup O1 and non-O1/non-O139 from a previous study (11), and two of nontoxigenic O1 strains from China (ZJ59 and LN93097, accession nos. EU622531 and AF512425, respectively, in GenBank).…”

Serendipitous Isolation of Non-VibrioBacterial Strains Carrying the Cholera Toxin Gene from Environmental Waters in Indonesia

“…However, we cannot verify the absence of tcpA and tolQRA genes by PCR alone. As of 2011, Kumar et al [14] reported that there were several variants of the tcpA gene which could be classified into 10 genetically distinct clusters in V. cholerae and V. mimicus. Therefore, it remains to be determined whether these isolates carry tcpA and tolQRA by not PCR but southern blotting analysis.…”