Corrigendum: Effects of lng Mutations on LngA Expression, Processing, and CS21 Assembly in Enterotoxigenic Escherichia coli E9034A.

[This corrects the article on p. 1201 in vol. 7, PMID: 27536289.].

Effects of lng Mutations on LngA Expression, Processing, and CS21 Assembly in Enterotoxigenic Escherichia coli E9034A.

Enterotoxigenic Escherichia coli (ETEC) is a major cause of morbidity in children under 5 years of age in low- and middle-income countries and a leading cause of traveler's diarrhea worldwide. The ability of ETEC to colonize the intestinal epithelium is mediated by fimbrial adhesins, such as CS21 (Longus). This adhesin is a type IVb pilus involved in adherence to intestinal cells in vitro and bacterial self-aggregation. Fourteen open reading frames have been proposed to be involved in CS21 assembly, hitherto only the lngA and lngB genes, coding for the major (LngA) and minor (LngB) structural subunit, have been characterized. In this study, we investigated the role of the LngA, LngB, LngC, LngD, LngH, and LngP proteins in the assembly of CS21 in ETEC strain E9034A. The deletion of the lngA, lngB, lngC, lngD, lngH, or lngP genes, abolished CS21 assembly in ETEC strain E9034A and the adherence to HT-29 cells was reduced 90%, compared to wild-type strain. Subcellular localization prediction of CS21 proteins was similar to other well-known type IV pili homologs. We showed that LngP is the prepilin peptidase of LngA, and that ETEC strain E9034A has another peptidase capable of processing LngA, although with less efficiency. Additionally, we present immuno-electron microscopy images to show that the LngB protein could be localized at the tip of CS21. In conclusion, our results demonstrate that the LngA, LngB, LngC, LngD, LngH, and LngP proteins are essential for CS21 assembly, as well as for bacterial aggregation and adherence to HT-29 cells.

CS21 positive multidrug-resistant ETEC clinical isolates from children with diarrhea are associated with self-aggregation, and adherence.

BACKGROUND: Enterotoxigenic Escherichia coli (ETEC) colonize the human intestinal mucosa using pili and non-pili colonization factors (CFs). CS21 (also designated Longus) is one of the most prevalent CFs encoded by a 14 kb lng DNA cluster located in a virulence plasmid of ETEC; yet limited information is available on the prevalence of CS21 positive ETEC isolates in different countries. The aim of this study was to evaluate the prevalence of CS21 among ETEC clinical isolates from Mexican and Bangladeshi children under 5 years old with diarrhea and to determine the phenotypic and genotypic features of these isolates. METHODS: ETEC clinical isolates positive to lngA gene were characterized by genotype, multidrug-resistance, self-aggregation, biofilm formation, and adherence to HT-29 cell line. RESULTS: A collection of 303 E. coli clinical isolates were analyzed, the 81.51% (247/303) were identified as ETEC, 30.76% (76/247) were st (+)/lt (+), and 25.10% (62/247) were positive for the lngA gene. Among the lngA (+) ETECs identified, 50% of isolates (31/62) were positive for LngA protein. The most frequent serotype was O128ac:H12 found in 19.35% (12/62) of lngA (+) ETEC studied. Multidrug-resistance (MDR) lngA (+) ETEC isolates was identified in 65% (39/60), self-aggregation in 48.38% (30/62), and biofilm formation in 83.87% (52/62). ETEC lngA (+) isolates were able to adhere to HT-29 cells at different levels. Two lngA isogenic mutants were constructed in the ETEC E9034A and ETEC73332 clinical isolate, showing a 77% and 98% reduction in adherence, respectively with respect to the wild type. CONCLUSION: ETEC isolates that have the lngA gene showed features associated with self-aggregation, and adherence to HT-29 cells, important characteristics in the human gut colonization process and pathogenesis.