Global Multilocus Sequence Typing Analysis of Mycoplasma bovis Isolates Reveals Two Main Population Clusters

Abstract

Mycoplasma bovis is a major bovine pathogen associated with bovine respiratory disease complex and is responsible for substantial economic losses worldwide. M. bovis is also associated with other clinical presentations in cattle, including mastitis, otitis, arthritis, and reproductive disorders. To gain a better understanding of the genetic diversity of this pathogen, a multilocus sequence typing (MLST) scheme was developed and applied to the characterization of 137 M. bovis isolates from diverse geographical origins, obtained from healthy or clinically infected cattle. After in silico analysis, a final set of 7 housekeeping genes was selected (dnaA, metS, recA, tufA, atpA, rpoD, and tkt). MLST analysis demonstrated the presence of 35 different sequence types (STs) distributed in two main clonal complexes (CCs), defined at the double-locus variant level, namely, CC1, which included most of the British and German isolates, and CC2, which was a more heterogeneous and geographically distant group of isolates, including European, Asian, and Australian samples. Index of association analysis confirmed the clonal nature of the investigated M. bovis population, based on MLST data. This scheme has demonstrated high discriminatory power, with the analysis showing the presence of genetically distant and divergent clusters of isolates predominantly associated with geographical origins.M ycoplasma diseases cause substantial economic losses, particularly in intensively farmed cattle production systems worldwide, as a result of poor growth, morbidity, and deaths, as well as the costs associated with increased control and prophylactic measures. Mycoplasma bovis has increasingly been recognized as one of the main pathogens involved in the bovine respiratory disease complex, on its own or in association with other respiratory pathogens (1). M. bovis can also be found in association with mastitis, in which outbreaks can affect more than 20% of the cows in a herd, regardless of the stage of lactation, and infections are usually refractory to treatment. Arthritis and otitis have also been associated with M. bovis, usually appearing once pneumonia or mastitis is already established in the herd (1, 2). The control of M. bovis infections relies strongly on antimicrobial therapy, which has variable success rates in the field (3). Vaccination has been used in the early stages of cattle development and is mostly based on autogenous vaccines, which limits their use and the potential for widespread control of M. bovis infections (3).Taking into consideration the limited tools available for M. bovis disease management, the development of a dependable molecular typing scheme able to offer robust and reproducible epidemiological information would provide a valuable addition to control measures targeting this pathogen. M. bovis isolates have been characterized previously using multiple molecular typing methods, including amplified fragment length polymorphism (AFLP) analysis (4), random amplified polymorphic DNA (RAPD) analysis (5), pulsed-fiel...

Flow cytometry follow-up analysis of peripheral blood leukocyte subpopulations in calves experimentally infected with field isolates of Mycoplasma bovis

“…MLST analysis showed that isolate 1 was MLST sequence type 36 (ST36) that did not cluster with any of the clonal complexes previously observed (Rosales et al, 2015). Isolates 2 and 3 produced two different STs, ST30 and ST32, respectively, which clustered in the same clonal complex 1.…”

“…Isolate 2 was derived from the lungs of a 5-month-old pneumonic calf, and isolate 3 was obtained from bovine fetal stomach contents, both originating from England. Multilocus sequence typing (MLST) was also performed on these isolates using the method described by Rosales et al (2015).…”

Development of a Multilocus Sequence Typing Scheme for Molecular Typing of Mycoplasma pneumoniae

“…MLST has been used to genotype several species of bacteria, including several Mycoplasma species: Mycoplasma agalactiae, Mycoplasma bovis, and Mycoplasma hyorhinis (30)(31)(32). This study has described the successful development of a novel M. pneumoniae MLST scheme to allow the characterization of clinical isolates.…”

Prevalence of pathogens from Mollicutes class in cattle affected by respiratory diseases and molecular characteristics of Mycoplasma bovis field strains

“…So far, M. bovis isolates have been characterised in some countries using genotyping tools such as RAPD, multilocus variable number tandem repeat (VNTR) analysis, and multilocus sequence typing (MLST) (2,3,14,15,23,29) and it revealed that M. bovis strains are divided into two major clonal complexes. However, MLST results published by Dudek et al (7) revealed that a Polish strain (not included in this study) did not cluster with any previously described clonal complexes.…”

“…Due to the difficulty of effective isolation of mycoplasmas under laboratory conditions, molecular methods are considered to be the most suitable and have recently been generally recommended for diagnosis of mycoplasmal infection (28). The survey of M. bovis spread and studies on its genetic diversity were performed in different parts of the world because M. bovis is well-equipped to generate diversity, and new clones may appear (2,7,23,29).…”