Identification of unconventional Shiga toxin-producing (STEC) and enteropathogenic (EPEC) Escherichia coli from diarrheic calves.

Introduction
Escherichia coli producing the Shiga toxins (STEC) and/or the attaching-effacing (AE) lesion (AE-STEC) cause enteritis and (bloody) diarrhoea in young calves and in humans, and are also present in the intestines of healthy cattle. AE-STEC in humans can belong to scores of O serogroups with 7 most frequent and pathogenic ones (O157, O26, O103, O111, O121, O145 and O165). Conversely the majority of AE-STEC in young calves belong to a narrower range of serogroups (O26, O111, O118, O5). Besides AE-STEC, E. coli producing only the AE lesion (EPEC) are also frequently associated with diarrhoea in young calves. Half of them belong to the O26 and O80 serogroups,, but the serotypes of the second half of isolates remain unidentified (Fakih et al. 2017; Thiry et al., 2018).
Materials and Methods
Different unconventional serogroups (O123/186, O156, O177, O182, O183) were identified by PCR in 5 STEC or EPEC that tested negative for the pre-cited 12 serogroups (Iguchi et al., 2015). The aim of this study was therefore to identify those 5 serogroups and the O146 serogroup amongst 76 AE-STEC, STEC and EPEC isolated between 2008 and 2015 from diarrheic calves at ARSIA (Fakih et al., 2017). Two triplex PCRs were applied either for the O146_O182_O183 serogroups or for the O123/186_O156_O177 serogroups.
Discussion
So far the first triplex PCR identified 4 O182-positive and 2 O183-positive STEC and EPEC. These 6 PCR-positive isolates were confirmed by either uniplex PCR. Two O182-positive isolates were AE-STEC and 2 were EPEC whereas the two O183-positive isolate were STEC. The second triplex PCR identified 10 O123-186-positive, 2 O156-positive and 13 O177-positive STEC and EPEC. These 25 PCR-positive isolates were confirmed by the three uniplex PCR. One O123/186-positive isolate was AE-STEC and 9 were EPEC. One O156-positive isolate was AE-STEC and one was STEC and finally, 12 O177-positive isolates were EPEC and 1 was STEC.
The further steps of the work are: (i) the identification of the same 6 serogroups amongst a collection of (AE-)STEC and EPEC isolated from healthy cattle at slaughterhouse; (ii) the identification of still other serogroups amongst the remaining untyped calf (AE-)STEC and EPEC; (iii) the comparison of the calf-positive STEC and EPEC with human (AE-)STEC and EPEC belonging to the same serogroups.
References
Fakih, I., Thiry, D., Duprez, J. N., Saulmont, M., Iguchi, A., Piérard, D., … Mainil, J. G. (2017). Identification of Shiga toxin-producing (STEC) and enteropathogenic (EPEC) Escherichia coli in diarrhoeic calves and comparative genomics of O5 bovine and human STEC. Veterinary Microbiology, 202, 16–22.

Iguchi, A., Iyoda, S., Seto, K., Morita-Ishihara, T., Scheutz, F., & Ohnishi, M. (2015). Escherichia coli O-genotyping PCR: A comprehensive and practical platform for molecular O serogrouping. Journal of Clinical Microbiology, 53(8), 2427–2432. https://doi.org/10.1128/JCM.00321-15

Thiry, D., De Rauw, K., Takaki, S., Duprez, J. N., Iguchi, A., Piérard, D., … Mainil, J. G. (2018). Low prevalence of the “gang of seven” and absence of the O80:H2 serotypes among Shigatoxigenic and enteropathogenic Escherichia coli (STEC and EPEC) in intestinal contents of healthy cattle at two slaughterhouses in Belgium in 2014. Journal of Applied Microbiology, 124(3), 867–873.