New Approaches for Escherichia coli Genotyping

Abstract

Easy-to-perform, fast, and inexpensive methods of differentiation of Escherichia coli strains beyond the species level are highly required. Herein two new, original tools for genotyping of E. coli isolates are proposed. The first of the developed method, a PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) test uses a highly variable fliC gene, encoding the H antigen as a molecular target. The designing of universal pair of primers and selection of the optimal restriction enzyme RsaI was preceded by in silico comparative analysis of the sequences of the genes coding for 53 different serotypes of H-antigen (E. coli flagellin). The target fragments of E. coli genomes for MLST method were selected on the basis of bioinformatics analysis of complete sequences of 16 genomes of E. coli. Initially, seven molecular targets were proposed (seven pairs of primers) and five of them were found useful for effective genotyping of E. coli strains. Both developed methods revealed high differentiation power, and a high genetic diversity of the strains tested was observed. Within the group of 71 strains tested, 29 and 47 clusters were revealed with fliC RFLP-PCR and MLST methods, respectively. Differentiation of the strains with the reference BOX-PCR method revealed 31 different genotypes. The in silico analysis revealed that the discriminatory power of the new MLST method is comparable to the Pasteur and Achtman schemes and is higher than the discriminatory power of the method developed by Clermont. From the epidemiology point of view, the outcomes of our investigation revealed that in most cases, the patients were infected with unique strains, probably from environmental sources. However, some strains isolated from different patients of the wards of pediatrics, internal medicine, and neurology were classified to the same genotype when the results of all three methods were taken into account. It could suggest that they were transferred between the patients.

Antibiotic Resistance in Bacteria—A Review

“…IPEC bacteria are associated with infections of the gastrointestinal tract. Several pathotypes can be distinguished among IPEC strains: enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enterohaemorrhagic E. coli (EHEC), enteroinvasive E. coli (EIEC), enteroaggregative E. coli (EAEC), adherent invasive E. coli (AIEC), and diffusely adherent E. coli (DAEC) [ 96 ]. ExPEC strains cause infections in extraintestinal anatomic sites, and in this group include: uropathogenic E. coli (UPEC) associated with urinary tract infection in human and animals, neonatal meningitis-associated E. coli (NMEC), septicaemic E. coli (SePEC) causing systemic infection in human and animals, avian pathogenic E. coli (APEC) that cause avian colibacillosis, and a potentially emerging ExPEC lineage named endometrial pathogenic E. coli (EnPEC) [ 97 , 98 ].…”

Antibiotic Resistance and Virulence Profiles of Escherichia coli Strains Isolated from Wild Birds in Poland

“…Intestinal pathogenic E. coli (IPEC) bacteria are associated with infections of the gastrointestinal tract, while extraintestinal pathogenic E. coli (ExPEC) strains cause infections in extraintestinal anatomic sites. Several pathotypes can be distinguished among IPEC strains: enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC), enterohaemorrhagic E. coli (EHEC), enteroinvasive E. coli (EIEC), enteroaggregative E. coli (EAEC), adherent invasive E. coli (AIEC), and diffusely adherent E. coli (DAEC) [ 1 ]. ExPEC strains include avian pathogenic E. coli (APEC), uropathogenic E. coli (UPEC), neonatal meningitis E. coli (NMEC), and sepsis-associated E. coli (SEPEC) [ 1 , 2 ].…”

“…Several pathotypes can be distinguished among IPEC strains: enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC), enterohaemorrhagic E. coli (EHEC), enteroinvasive E. coli (EIEC), enteroaggregative E. coli (EAEC), adherent invasive E. coli (AIEC), and diffusely adherent E. coli (DAEC) [ 1 ]. ExPEC strains include avian pathogenic E. coli (APEC), uropathogenic E. coli (UPEC), neonatal meningitis E. coli (NMEC), and sepsis-associated E. coli (SEPEC) [ 1 , 2 ]. Characteristic virulence factors, which play a decisive role in pathogenesis, have been defined for each of these pathotypes [ 3 ].…”

Genetic variability of the 16S rRNA gene of Nocardia brasiliensis, the most common causative agent of actinomycetoma in Latin America and the Caribbean

“…Previously, in silico enzymatic restriction studies have demonstrated the intraspecies genetic variability in other bacteria, such as identifying clusters and subclusters in the streptokinase gene of Streptococcus 26 , the identification of E. coli genotypes, which also suggested updating the existing classifications 30 . Its usefulness was also proven in constructing new classifications in the study species, with a highly reliable discrimination power, such as the one performed using the cpn60 gene of phytoplasmas 28 .…”