Slot blot immunoassay as a tool for plasmid-encoded toxin detection in enteroaggregative Escherichia coli culture supernatants

Diagnostic Microbiology and Infectious Disease
Andréa Bernardes Vilhena-CostaMarina B Martinez

Abstract

Plasmid-encoded toxin (Pet) is a heat-labile enterotoxin encoded in the enteroaggregative Escherichia coli (EAEC) virulence plasmid. Several evidence support the role of this 108-kDa secreted protein in the pathogenesis of EAEC diarrhea. In this study, we standardized a slot blot immunoassay for Pet detection. EAEC culture supernatants were applied onto a polyvinylidene difluoride membrane, and, using rabbit polyclonal Pet antisera, the expression of the toxin by slot blot immunoassay was observed in 9.5% of the isolates studied. In addition, no negative control reacted with Pet antiserum in this assay. This assay is a rapid, specific, reproducible, and low-cost methodology, therefore demonstrating its potential in diagnosing Pet expression. Moreover, we describe for the first time that expression of Pet can be directly detected from EAEC culture supernatants and may be used in clinical laboratorial routine instead of polymerase chain reaction detection of the pet gene, especially in developing countries where the EAEC pathotype has been considered an emerging pathogen.

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Citations

Jun 9, 2009·Journal of Bacteriology·William LainhartGerald B Koudelka
Feb 4, 2011·BMC Research Notes·Márcio A MenezesRoxane Mf Piazza
Jun 21, 2014·BioMed Research International·Rita C RuizRoxane M F Piazza
May 4, 2011·Memórias do Instituto Oswaldo Cruz·Caroline A MagalhãesRoxane M F Piazza
Nov 22, 2016·Brazilian Journal of Microbiology : [publication of the Brazilian Society for Microbiology]·Tânia A T GomesMarina B Martinez

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