Comparison of two multilocus sequence typing schemes for Mycoplasma bovis and revision of the pubMLST reference method

Authors: Register, Karen; LYSNYANSKY, INNA - Kimron Veterinary Institute; JELINSKI, MURRAY - University Of Saskatchewan; Boatwright, Jr, William; WALDNER, MATTHEW - University Of Saskatchewan; Bayles, Darrell; PILO, PAOLA - University Of Bern; Alt, David

Submitted to: Journal of Clinical Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/7/2020
Publication Date: 5/26/2020
Citation: Register, K.B., Lysnyansky, I., Jelinski, M., Boatwright Jr, W.D., Waldner, M., Bayles, D.O., Pilo, P., Alt, D.P. 2020. Comparison of two multilocus sequence typing schemes for Mycoplasma bovis and revision of the pubMLST reference method. Journal of Clinical Microbiology. 58(6):e00283-20. https://doi.org/10.1128/JCM.00283-20.
DOI: https://doi.org/10.1128/JCM.00283-20

Interpretive Summary: Mycoplasma bovis causes pneumonia, pharyngitis, otitis, arthritis, mastitis and reproductive disorders in cattle and bison. Two different schemes for genetic typing of isolates have been developed for M. bovis, one of which has been adopted as a universal reference method, but no comparison of the schemes has been undertaken. Although the reference scheme has proven to be highly informative, modification is necessary due to the recent discovery of isolates missing one of the genes that it uses to classify isolates, specifically a gene named adh-1. Such isolates can be only partially typed, which is unacceptable for a reference method. The goal of our study was to compare the performance of the two typing schemes and identify a new reference scheme capable of fully typing all isolates. We evaluated 448 isolates arising from diverse geographic and anatomic sites that collectively represent cattle, bison, deer and a goat. The current reference scheme outperformed the alternative scheme, but use of the adh-1 gene must be discontinued if it is to be retained as a reference method. A comprehensive analysis of the genome sequences available for 436 of the isolates used here identified a gene named dnaA as the optimal replacement for adh-1. The revised scheme will be implemented as the new universal reference method and related data will be gathered and archived in a publicly available, web-based database (https://pubmlst.org/mbovis/).

Technical Abstract: Mycoplasma bovis causes pneumonia, pharyngitis, otitis, arthritis, mastitis and reproductive disorders in cattle and bison. Two multilocus sequence typing (MLST) schemes have been developed for M. bovis, one of which serves as the PubMLST reference method, but no comparison of the schemes has been undertaken. Although the PubMLST scheme has proven to be highly discriminatory and informative, modification is necessary due to the recent discovery of isolates missing one of the typing loci, adh-1. The goal of our study was to compare the performance of the two MLST schemes and identify a new reference scheme capable of fully typing all isolates. We evaluated 448 isolates arising from diverse geographic and anatomic sites that collectively represent cattle, bison, deer and a goat. The discrimination index (DI) for the PubMLST and alternative scheme is 0.909 (91 STs) and 0.842 (77 STs), respectively. Although the PubMLST scheme outperformed the alternative scheme, the adh-1 locus must be retired from the PubMLST scheme if it is to be retained as a reference method. The DI obtained using the six remaining PubMLST loci (0.897, 79 STs) fails to reach the benchmark recommended for a reference method (0.900), mandating the addition of a seventh locus. Comparative analysis of genome sequences from the isolates used here identified the dnaA locus from the alternative scheme as the optimal replacement for adh-1. This revised scheme, which will be implemented as the new PubMLST reference method, has a DI of 0.914 and distinguishes 88 STs from the 448 isolates evaluated.