Whole genome sequencing of Moraxella bovis strains from North America reveals two genotypes with different genetic determinants

Authors: Wynn, Emily; HILLE, MATTHEW - University Of Nebraska; LOY, JOHN - University Of Nebraska; Schuller, Genevieve - Gennie; Kuhn, Kristen; Dickey, Aaron; Bono, James - Jim; Clawson, Michael - Mike

Submitted to: BMC Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/17/2022
Publication Date: 10/21/2022
Citation: Wynn, E.L., Hille, M.M., Loy, J.D., Schuller, G., Kuhn, K.L., Dickey, A.M., Bono, J.L., Clawson, M.L. 2022. Whole genome sequencing of Moraxella bovis strains from North America reveals two genotypes with different genetic determinants. BMC Microbiology. 22(1). Article 258. https://doi.org/10.1186/s12866-022-02670-3.
DOI: https://doi.org/10.1186/s12866-022-02670-3

Interpretive Summary: Infectious bovine keratoconjunctivitis (IBK), also known as pinkeye, is the most common eye disease affecting cattle worldwide, and is estimated to cost the United States hundreds of millions of dollars each year. IBK is a complex disease, with multiple bacterial species implicated in having a role in its development, however, only one, Moraxella bovis, has been experimentally shown to cause IBK. M. bovis has a number of virulence determinants such as; 1) a toxin commonly known as hemolysin that can lyse cells in the eye, 2) pilin that are important for M. bovis attachment to cells in the eye, and 3) filamentous haemagglutinin-like proteins that are encoded by plasmids (extrachromosomal DNA elements that contain genes), and are also thought to be important for M. bovis attachment to host cells. Prior to this study, only one completely assembled M. bovis genome (bacterial chromosome and plasmids) was publicly available for a single strain, and that genome was produced by members of our group several years ago. In this study, we sequenced and completely assembled the chromosomes and plasmids of a diverse collection of 36 M. bovis strains from cattle across North America. We compared their gene content with each other, and with Moraxella bovoculi, which is closely related to M. bovis. We found there are two major strain types, or genotypes of M. bovis that differ in their genome content, and that they encode different versions of hemolysin and pilin. We further found that the two genotypes had different types of plasmids, and that only one of the two genotypes co-occurred with a plasmid encoding filamentous haemagglutinin-like proteins. The results from this study provide a means to study M. bovis pathogenesis and virulence from the context of two genotypes that may not associate equally with IBK, and to develop immunological reagents against one or both of them.

Technical Abstract: Background: Moraxella bovis and Moraxella bovoculi both associate with infectious bovine keratoconjunctivitis (IBK), an economically significant and painful ocular disease that affects cattle worldwide. There are two genotypes of M. bovoculi (genotypes 1 and 2) that differ in their gene content and potential virulence factors, although neither have been experimentally shown to cause IBK. M. bovis is a causative IBK agent, however, not all strains carry a complete assortment of known virulence factors. The goals of this study were to determine the population structure and depth of M. bovis genomic diversity, and to compare core and accessory genes and predicted outer membrane protein profiles both within and between M. bovis and M. bovoculi. Results: Phylogenetic trees and bioinformatic analyses of 36 M. bovis chromosomes sequenced in this study and additional available chromosomes of M. bovis and both genotype 1 and 2 M. bovoculi, showed there are two genotypes (1 and 2) of M. bovis. The two M. bovis genotypes share a core of 2015 genes, with 121 and 186 genes specific to genotype 1 and 2, respectively. The two genotypes differ by their chromosome size and prophage content, encoded protein variants of the virulence factor hemolysin, and by their affiliation with different plasmids. Eight plasmid types were identified in this study, with types 1 and 6 observed in 88 and 56% of genotype 2 strains, respectively, and absent from genotype 1 strains. Only type 1 plasmids contained one or two gene copies encoding filamentous haemagglutinin-like proteins potentially involved with adhesion. A core of 1403 genes was shared between the genotype 1 and 2 strains of both M. bovis and M. bovoculi, which encoded a total of nine predicted outer membrane proteins. Conclusions: There are two genotypes of M. bovis that differ in both chromosome content and plasmid profiles and thus may not equally associate with IBK. Immunological reagents specifically targeting select genotypes of M. bovis, or all genotypes of M. bovis and M. bovoculi together could be designed from the outer membrane proteins identified in this study.