Deciphering the possible role of ctxB7 allele on higher production of cholera toxin by Haitian variant Vibrio cholerae O1

Cholera continues to be an important public health concern in developing countries where proper hygiene and sanitation are compromised. This severe diarrheal disease is caused by the Gram-negative pathogen Vibrio cholerae belonging to serogroups O1 and O139. Cholera toxin (CT) is the prime virulence factor and is directly responsible for the disease manifestation. The ctxB gene encodes cholera toxin B subunit (CTB) whereas the A subunit (CTA) is the product of ctxA gene. Enzymatic action of CT depends on binding of B pentamers to the lipid-based receptor ganglioside GM1. In recent years, emergence of V. cholerae Haitian variant strains with ctxB7 allele and their rapid spread throughout the globe has been linked to various cholera outbreaks in Africa and Asia. These strains produce classical type (WT) CTB except for an additional mutation in the signal sequence region where an asparagine (N) residue replaces a histidine (H) at the 20th amino acid position (H20N) of CTB precursor (pre-CTB). Here we report that Haitian variant V. cholerae O1 strains isolated in Kolkata produced higher amount of CT compared to contemporary O1 El Tor variant strains under in vitro virulence inducing conditions. We observed that the ctxB7 allele, itself plays a pivotal role in higher CT production. Based on our in silico analysis, we hypothesized that higher accumulation of toxin subunits from ctxB7 allele might be attributed to the structural alteration at the CTB signal peptide region of pre-H20N CTB. Overall, this study provides plausible explanation regarding the hypertoxigenic phenotype of the Haitian variant strains which have spread globally, possibly through positive selection for increased pathogenic traits.
Author summary Cholera remains to be a major public health burden in the developing countries. Toxigenic strains of the Gram-negative organism Vibrio cholerae belonging to serogroups O1 and O139 are the etiological agents for this dreadful disease. These strains secrete CT into the extracellular milieu which upon ingestion of contaminated food or water, results in severe loss of water and electrolytes. In the recent past, V. cholerae O1 strains with the signature ctxB7 allele have propagated through many of the cholera endemic regions in Africa and Asia. These strains were also identified as the cause for the devastating cholera epidemic in Haiti that killed around 8000 victims. It was demonstrated that the Haitian isolate had been associated with hypertoxigenic phenotype and increased virulence. However, whether the ctxB7 allele may have any role to play in increased CT production phenotype of the Haitian variants remains obscure. We observed that the ctxB7 allele plays a pivotal role in heightened CT production, although any link between increased CT production and hypervirulence phenotypes remains undefined.