Purpose: To evaluate the applicability of spiral plating method and multiplex real-time quantitative PCR (qPCR) to quantify six non-O157 E. coli serogroups in cattle feces.
Methods: Cattle fecal samples (n = 1,152) collected from eight commercial feedlots were suspended in E. coli broth. Diluted fecal suspensions were spiral plated onto selective chromogenic medium. Concentration (CFU/g) was determined by counting chromogenic colonies using a counting grid and testing ten randomly picked colonies individually by a multiplex conventional PCR targeting six serogroups and four virulence genes. Concentration of each serogroup was determined based on the proportion of colonies positive for the serogroup. DNA extracted from fecal suspensions was subjected to two qPCR assays: Assay 1 - O26, O103, and O111; Assay 2 - O45, O121, and O145. Fecal suspensions incubated at 40°C for 6 h were subjected to a culture method for the detection of six non-O157 E. coli serogroups.
Results: A total of 787 (68.3%) samples were positive by culture method for at least one of the six non-O157 E. coli serogroups, and of those 139 (17.7%) were quantifiable (> log 2.9) by spiral plating method and 493 (42.8%; > log 4) by qPCR assays. E. coli O103 was the predominant serogroup quantified by spiral plating method (8.7%) and qPCR (25.5%).
Significance: Spiral plating method can be used to determine the concentration of non-O157 E. coli serogroups and those that carry Shiga Toxin genes in cattle feces. The qPCR assays are more sensitive but allow quantification only at the serogroup level.