MOLECULAR CHARACTERIZATION, VIRULENCE, AND ANTIMICROBIAL SUSCEPTIBILITY OF MYCOPLASMA BOVIS ASSOCIATED WITH CHRONIC MASTITIS IN DAIRY COWS
Authors: Gioushy Magdy , Abdelaziz Soliman Eid Elsaid , Elkenany Rasha M. , El-Alfy El-Sayed , Abd Elaal Ahmed , Abd-El Razik Khaled Abd-El Hamid , El-Khodery Sabry 
Published: December 2024

Mycoplasma bovis (M. bovis) is the most common Mycoplasma species, which has a growing impact on the dairy industry. The purpose of this study was to investigate the molecular characterization, virulence, and antibiotic susceptibility of M. bovis isolates from dairy cows with chronic mastitis in Egypt. Eighty-four composite milk samples from mastitic cows were aseptically collected from dairy farms in Egypt. Based on microbiological examination and the polymerase chain reaction (PCR) technique targeting the 16S rRNA, 14 (16.7%) of all milk samples were positive for Mycoplasma spp. PCR targeting M. bovis-specific gene identified six (7.14%) of the 14 mycoplasma isolates as M. bovis. PCR assays for different virulence genes showed that all M. bovis isolates exhibited the presence of vspA gene, while other virulence genes (uvrC, gap, and p40 pseudogenes) were determined in only two M. bovis isolates (2/6). The 16S rRNA gene phylogenetic analysis demonstrated 100% homology with reference strains of M. bovis isolated from different species and locations. When compared to other isolates in the GenBank, the amino acid sequence alignment of our isolate vspA-like protein indicated a distinct mutation. The in vitro antimicrobial susceptibility testing of the six M. bovis isolates in this study to seven antimicrobial drugs revealed that tilmycosin and tylosin had the lowest minimum inhibitory concentrations (MIC) values (≤1 μg/mL), while danofloxacin, streptomycin and florfenicol had the highest MIC values (<4 μg/mL). Nonetheless, this study showed the virulence of M. bovis in dairy cows in Egypt, and macrolides were found to be the most potent compounds in vitro against all tested isolates, but further studies in nationwide surveys are needed.
Molekularna karakterizacija, virulenca in protimikrobna občutljivost z mikoplazmo M. bovis povezanega kroničnega mastitisa pri kravah molznicah
Izvleček: Mycoplasma bovis (M. bovis) je najpogostejša vrsta mikoplazme, ki ima vse večji vpliv na mlečno industrijo. Namen te študije je bil molekularno karakterizirati in raziskati virulenco in občutljivost izolatov M. bovis za antibiotike, in sicer iz krav molznic s kroničnim mastitisom v Egiptu. Štiriinosemdeset mešanih vzorcev mleka krav z mastitisom je bilo aseptično zbranih na mlečnih farmah v Egiptu. Na podlagi mikrobiološkega pregleda in analize izražanja 16S rRNA z verižno reakcijo s polimerazo (PCR) je bilo 14 (16,7 %) od vseh vzorcev mleka pozitivnih na Mycoplasma spp. Z metodo PCR smo šest (7,14 %) od 14 izolatov mikoplazme identificirali kot M. bovis. Analize PCR za različne gene virulence so pokazale, da je bil pri vseh izolatih M. bovis prisoten gen vspA, medtem ko so bili drugi geni virulence (uvrC, gap in psevdogeni p40) izraženi le pri dveh izolatih M. bovis (2/6). Filogenetska analiza gena 16S rRNA je pokazala 100-odstotno homolognost z referenčnimi sevi M. bovis, izoliranimi iz različnih vrst in lokacij. V primerjavi z drugimi izolati podatkovne zbirke GenBank je poravnava aminokislinskega zaporedja proteina, podobnega VspA, pokazala na značilno mutacijo. In vitro testiranje protimikrobne občutljivosti šestih izolatov M. bovis za sedem protimikrobnih zdravil je pokazalo, da sta imela tilmikozin in tilozin najnižje vrednosti minimalnih inhibitornih koncentracij (MIC) (≤1 μg/ml), medtem ko so imeli danofloksacin, streptomicin in florfenikol najvišje vrednosti MIC (<4 μg/ml). Kljub temu da je ta študija pokazala virulenco M. bovis pri kravah molznicah v Egiptu, makrolidi pa so se izkazali za najmočnejše spojine proti vsem in vitro testiranim izolatom, so potrebne nadaljnje študije v nacionalnih raziskavah.
Ključne besede: antimikrobna občutljivost; kronični mastitis; Mycoplasma bovis; analiza zaporedja; geni virulence