Principal coordinate analysis showing the clustering of antimicrobial resistance genes by livestock, municipal and low impact environmental samples.

<p>Antimicrobial resistance genes were detected among cattle (n = 12), low impact environment (n = 8), municipal (n = 12) and swine (n = 12) pooled samples. Data points are colored as follows: green = cattle, red = low impact environment, black = municipal and blue = swine.</p

Model adjusted prevalence (%) of <i>E</i>. <i>coli</i>, <i>Salmonella</i> and <i>Enterococcus</i> species from cattle (n = 48), low impact environment (n = 32), municipal (n = 46) and swine (n = 48) samples^a.

<p>^aDifferent superscripts across rows indicate statistically significant (<i>P</i> < 0.05) differences between pair of sample sources. Bonferroni adjusted for multiple comparisons. For prevalence values of 0 or 100% the logistic regression models did not converge. In those instances exact binomial 95% confidence intervals were used for pairwise comparisons.</p><p>^bAbbreviations: 3GC^r = third generation cephalosporin resistant; COT^r = trimethoprim/sulfamethoxazole resistant; NAL^r = nalidixic acid resistant; ERY^r = erythromycin resistant</p><p>Model adjusted prevalence (%) of <i>E</i>. <i>coli</i>, <i>Salmonella</i> and <i>Enterococcus</i> species from cattle (n = 48), low impact environment (n = 32), municipal (n = 46) and swine (n = 48) samples<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132586#t001fn001" target="_blank">^a</a>.</p

Box plot showing median distribution of antimicrobial resistance genes detected per pooled sample among cattle (n = 12), low impact environment (n = 8), municipal (n = 12) and swine (n = 12) samples.

<p>The bold horizontal lines represent the median. The whiskers represent the upper and lower adjacent values. Superscripts have been assigned to the median. Different superscripts indicate statistically significant (P = 0.0001) differences between pairs of sample sources.</p

Venn diagram showing the number of specific genes identified by sample source.

<p>One gene common to cattle and swine samples and one gene common to low impact and municipal samples were not shown on the Venn diagram. The two genes are common to circles that cannot intersect in this diagram.</p

Number of cattle (n = 12), low impact environment (n = 8), municipal (n = 12) and swine (n = 12) pooled samples harboring specific antimicrobial resistance genes.

<p>Number of cattle (n = 12), low impact environment (n = 8), municipal (n = 12) and swine (n = 12) pooled samples harboring specific antimicrobial resistance genes.</p

Characteristics of <i>E</i>. <i>coli</i> O157:H7 isolates.

<p>Characteristics of <i>E</i>. <i>coli</i> O157:H7 isolates.</p

Model adjusted mean log₁₀ count of <i>E</i>. <i>coli</i>, <i>Salmonella</i> and <i>Enterococcus</i> spp from cattle, municipal, and swine samples^a.

<p>^aDifferent superscripts across rows indicate statistically significant (<i>P</i> < 0.05) differences between pair of sample sources. Bonferroni adjusted for multiple comparisons.</p><p>^bAbbreviations: n = number of samples; 3GC^r = third generation cephalosporin resistant; COT^r = trimethoprim/sulfamethoxazole resistant; ERY^r = erythromycin resistant.</p><p>Model adjusted mean log₁₀ count of <i>E</i>. <i>coli</i>, <i>Salmonella</i> and <i>Enterococcus</i> spp from cattle, municipal, and swine samples<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132586#t002fn001" target="_blank">^a</a>.</p

Comparative genomics of two super-shedder isolates of <i>Escherichia coli</i> O157:H7

<div><p>Shiga toxin-producing <i>Escherichia coli</i> O157:H7 (O157) are zoonotic foodborne pathogens and of major public health concern that cause considerable intestinal and extra-intestinal illnesses in humans. O157 colonize the recto-anal junction (RAJ) of asymptomatic cattle who shed the bacterium into the environment through fecal matter. A small subset of cattle, termed super-shedders (SS), excrete O157 at a rate (≥ 10⁴ CFU/g of feces) that is several orders of magnitude greater than other colonized cattle and play a major role in the prevalence and transmission of O157. To better understand microbial factors contributing to super-shedding we have recently sequenced two SS isolates, SS17 (GenBank accession no. CP008805) and SS52 (GenBank accession no. CP010304) and shown that SS isolates display a distinctive strongly adherent phenotype on bovine rectal squamous epithelial cells. Here we present a detailed comparative genomics analysis of SS17 and SS52 with other previously characterized O157 strains (EC4115, EDL933, Sakai, TW14359). The results highlight specific polymorphisms and genomic features shared amongst SS strains, and reveal several SNPs that are shared amongst SS isolates, including in genes involved in motility, adherence, and metabolism. Finally, our analyses reveal distinctive patterns of distribution of phage-associated genes amongst the two SS and other isolates. Together, the results of our comparative genomics studies suggest that while SS17 and SS52 share genomic features with other lineage I/II isolates, they likely have distinct recent evolutionary histories. Future comparative and functional genomic studies are needed to decipher the precise molecular basis for super shedding in O157.</p></div

Comparative analysis of SS strains and reference O157 genomes.

<p>(A) Dendrogram representing cluster analysis of SS isolates based on PFGE patterns as previously described [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0182940#pone.0182940.ref012" target="_blank">12</a>] highlighting the two representative isolates, SS17 and SS52, that have been completely sequenced; (B) Circular genome representations of SS17 and SS52. Larger circles depict chromosomal DNA with smaller circles representing plasmids. The blue circles represent the ORFs and the outer circle (purple) represents the phages in each genome; (C) Cladogram based on whole genome alignment reveals that SS17 and SS52 clusters closely with lineage I/II “spinach” outbreak isolates (EC4115 and TW14359) as compared with lineage I outbreak isolates (Sakai and EDL933) or the bovine lineage II isolates; (D) Whole genome alignments of SS strains with reference O157 strains using progressiveMauve depicting 8 homology blocks of similarity and patterns of divergence among the strains.</p

Phage diversity in SS and reference O157 strains.

<p>(A) MAUVE alignment of SS17 and SS52 showing the ~37 Kb insertion of CP-933O phage in the SS17 genome; (B) BLAST analysis of the region reveals that SS17, TW14359, and EC4115 all contain the same phage, whereas EDL933, Sakai, and SS52 do not; (C) Dot matrix representation of the results of megaBLAST analysis of the 37.5kb CP-933O region reveals its presence in SS17 (left panel) but not SS52 (right panel).</p