Comparison of antimicrobial resistance and pan-genome of clinical and non-clinical Enterococcus cecorum from poultry using whole-genome sequencing

Authors: SHARMA, POONAM - Orise Fellow; GUPTA, SUSHIM - Orise Fellow; Barrett, John; Woodley, Tiffanie; Hiott, Lari; KARIYAWASAM, SUBHASHINIE - Pennsylvania State University; Jackson, Charlene; Frye, Jonathan

Submitted to: Foods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/24/2020
Publication Date: 5/26/2020
Citation: Sharma, P., Gupta, S., Barrett, J.B., Woodley, T.A., Hiott, L.M., Kariyawasam, S., Jackson, C.R., Frye, J.G. 2020. Comparison of antimicrobial resistance and pan-genome of clinical and non-clinical Enterococcus cecorum from poultry using whole-genome sequencing. Foods. https://doi.org/10.3390/foods9060686.
DOI: https://doi.org/10.3390/foods9060686

Interpretive Summary: Enterococcus cecorum is an emerging avian pathogen, particularly in chickens, but has been detected in both diseased (clinical) and healthy (non-clinical) poultry. Differences between the two groups of isolates have not been defined. In this study, antimicrobial resistance genes and the genome of clinical and non-clinical E. cecorum from chickens were analyzed using whole-genome sequencing to better define their differences. The comparison confirmed that non-clinical isolates contained more resistance genes than clinical isolates. Resistance genes were both shared and exclusive to each group indicating varying genetic characteristics among E. cecorum isolates. The genome analysis revealed that the non-clinical E. cecorum genomes were comparatively diverse due to acquisition of additional genes while genome reduction in the conserved clinical genomes suggested better host adaptability. This data is important for human health due to clinical disease potential in humans and antimicrobial resistance in the bacterium that may be passed to humans through food. Identification of unknown virulence determinants is important for animal health for determination of the pathogenesis of E. cecorum infections in poultry and for production of a vaccine candidate for protection of poultry flocks. This research is useful for food safety scientists as well as veterinarians who treat poultry for infections caused by E. cecorum.

Technical Abstract: Enterococcus cecorum is an emerging avian pathogen, particularly in chickens, but can be found in both diseased (clinical) and healthy (non-clinical) poultry. To better define differences between E. cecorum from the two groups, antimicrobial resistance genes as well as the pan-genome among the isolates was analyzed using whole-genome sequencing (WGS). Eighteen strains selected from our previous study were subjected to WGS using Illumina MiSeq and comparatively analyzed. Assembled contigs were analyzed for resistance genes using ARG-ANNOT(Antibiotic Resistance Gene-ANNOTation). Resistance to erythromycin was mediated by ermB, ermG, and mefA, in clinical isolates and ermB and mefA, in non-clinical isolates. Lincomycin resistance genes were identified as linB, lnuB, lnuC, and lnuD with lnuD found only in non-clinical E. cecorum; however, lnuB and linB were found in only one clinical isolate. For both groups of isolates, kanamycin resistance was mediated by aph3-III while tetracycline resistance was conferred by tetM, tetO, and tetL. No mutations or known resistance genes were found for isolates resistant to either linezolid or chloramphenicol suggesting possible new mechanisms of resistance to these drugs. Comparison of WGS results confirmed that non-clinical isolates contained more resistance genes than clinical isolates. The pan-genome of clinical and non-clinical isolates resulted in 3651 and 4950 gene families, respectively, whereas the core gene sets were comprised of 1559 and 1534 gene families in clinical and non-clinical isolates, respectively. Unique genes were found more frequently in non-clinical isolates than clinical. Phylogenetic analysis of the isolates and all the available complete and draft genomes showed no co-relation between healthy and diseased poultry. Additional genomic comparison is required to elucidate genetic factors in E. cecorum that contribute to disease in poultry.