A Novel High-resolving Method for Genomic PCR-fingerprinting of Enterobacteria

Abstract

We developed a novel PCR–fingerprinting system for differentiation of enterobacterial strains using a single oligonucleotide primer IS1tr that matches the inverted terminal repeats of the IS1 insertion element. Compared to widely used BOX–PCR and ribotyping methods, our system features higher resolution allowing differentiation of closely related isolates that appear identical in BOX–PCR and ribotyping but differ in their phage sensitivity. The IS1–profiling system is less sensitive to the quality of the material and equipment used. At the same time, BOX–PCR is more universal and suitable for bacterial strain grouping and reconstruction of the low–distance phylogeny. Thus, our system represents an important supplement to the existing set of tools for bacterial strain differentiation; it is particularly valuable for a detailed investigation of highly divergent and rapidly evolving natural bacterial populations and for studies on coliphage ecology. However, some isolates could not be reliably differentiated by IS1–PCR, because of the low number of bands in their patterns. For improvement of IS1–fingerprinting characteristics, we offer to modify the system by introducing the second primer TR8834 hybridizing to the sequence of a transposase gene that is widely spread in enterobacterial genomes.

Phages associated with horses provide new insights into the dominance of lateral gene transfer in virulent bacteriophages evolution in natural systems

“…E. coli is a minor but permanently present component of the horse gut ecosystem. In our previous work we demonstrated that E. coli diversity at the strain level in the horse feces is extremely high, reaching several hundreds of lineages per sample (Golomidova et al 2007;Isaeva et al 2010). At the same time, all the coliphages cultured by us from E. coli feces to date appear to be virulent (Golomidova et al 2007;Kulikov et al 2014).…”

The ecogenomics of dsDNA bacteriophages in feces of stabled and feral horses

“…Our metagenomic data does not provide any direct estimates of bacterial diversity and/or phage-host relationships at the strain level. However, high intraspecies diversity of E. coli within horse feces associated with high diversity of co-occurring coliphages, having relatively narrow host ranges, was previously demonstrated using culture-based approaches [42] , [43] . Additional culture-based evaluation of the strain-level diversity of a more prevalent species than E. coli combined with characterization of its co-occurring phages, may help to shed more light over the pattern of the phage-host relationships in the horse gut ecosystem.…”

“…The E. coli host population was found to be highly divergent, and represented by hundreds of strains simultaneously present in the same sample. The overlap of the sensitivity of these strains to co-occuring bacteriophages was limited [42] , [43] with ~1–5% of the total E. coli counts being suitable hosts for any particular phage isolate. The data of Golomidova et al [42] , [44] and the results of the longitudinal study of G7C-related bacteriophages persistence and evolution within the ecosystem of a horse stable [45] indicated the flow of the coliphage genotypes between the animals.…”

Function of bacteriophage G7C esterase tailspike in host cell adsorption

“…A number of N4-like phages and their hosts have been isolated from complex multispecies environments recently, including phage G7C and its O22-like E. coli strain 4s host (Golomidova et al, 2007;Isaeva et al, 2010;Knirel et al, 2015). Bioinformatic analysis shows that all known virion proteins of G7C and N4 are similar except for those responsible for host cell recognition and attachment.…”