Polymorphic Amplified Typing Sequences and Pulsed-Field Gel Electrophoresis Yield Comparable Results in the Strain Typing of a Diverse Set of BovineEscherichia coliO157:H7 Isolates

Abstract

Polymorphic amplified typing sequences (PATS), a PCR-based Escherichia coli O157:H7 (O157) strain typing system, targets insertions-deletions and single nucleotide polymorphisms at XbaI and AvrII restriction enzyme sites, respectively, and the virulence genes (stx1, stx2, eae, hlyA) in the O157 genome. In this study, the ability of PATS to discriminate O157 isolates associated with cattle was evaluated. An in-depth comparison of 25 bovine O157 isolates, from different geographic locations across Northwest United States, showed that about 85% of these isolates shared the same dendogram clade by PATS and pulsed-field gel electrophoresis (PFGE), irrespective of the restriction enzyme sites targeted. The Pearson's correlation coefficient, r, calculated at about 0.4, 0.3, and 0.4 for XbaI-based, AvrII-based and combined-enzymes PATS and PFGE similarities, respectively, indicating that these profiles shared a good but not high correlation, an expected inference given that the two techniques discriminate differently. Isolates that grouped differently were better matched to their locations using PATS. Overall, PATS discriminated the bovine O157 isolates without interpretive biases or sophisticated analytical software, and effectively complemented while not duplicating PFGE. With its quick turnaround time, PATS has excellent potential as a convenient tool for early epidemiological or food safety investigations, enabling rapid notification/implementation of quarantine measures.

Evaluation of clonal relatedness among different sources of E. coli isolates in Iranian children with urinary tract infection (UTI) and age-matched healthy people.

“…To summarize, we can conclude that our results suggest that the presence of certain features such as PAI ICFT073, IICFT073, PAI I536, PAI IV536, PAI II J96, PAI II536, gafD In addition, failure to recognize relatedness concepts and similarity coefficients may have facilitated acceptance of dendrograms from single enzyme PFGE as precise relatedness measures. BioNumerics software produces banding patterns analysis from large sets of isolates, but it cannot be trusted to consider the presence or lack of bands without visual perception of the gel picture" [14,21,36]. The present study findings allow us to conclude epidemiologic relationships among isolates from PFGE data: however, more restriction enzymes would be required to make a reasonable approximation of genetic relatedness.…”

Shiga toxin-producing Escherichia coli (STEC) O22:H8 isolated from cattle reduces E. coli O157:H7 adherence in vitro and in vivo

“…Although the in vitro study was small, the statistic procedure applied was robust and suggests that infection of calves with the isolated strains of O22:H8 inhibits E. coli O157:H7 colonization. Since a epithelial cell line like HEp-2 seems to carry different adhesin receptors than RAJ cell types (Asper et al, 2007;Kudva et al, 2012a), a mixture of the three STEC O22:H8 strains was used to challenge the animals to account for the differences observed in vitro.…”

Cost-effective procedure for Streptococcus pyogenes immobilized DNA preparation and miniPFGE subtyping

“…However, in all cases, the patterns showed less intense bands than did the ones resolved from the miniplugs subjected to the standard enzymatic treatment (Figure 2A). R mean values did not differ between the patterns obtained from samples incubated with lysozyme for different times (F [3,13] =1.06, p =0.399).…”

“…It has been successfully applied to a broad range of different Gram-negative and Gram-positive bacteria [4][5][6][7][8][9][10][11]. Bacterial subtyping by PFGE relies on the determination of the relationship between different isolates by comparing their DNA macrorestriction patterns [12][13][14][15].…”