Khasapane, Ntelekwane G.Myburgh, JacobusNkhebenyane, Sebolelo J.Khumalo, Zamantungwa Thobeka HappinessRamatla, TsepoThekisoe, Oriel2024-08-012024-08-012024-01Khasapane, N.G.; Koos, M.; Nkhebenyane, S.J.; Khumalo, Z.T.H.; Ramatla, T.; Thekisoe, O. Detection of Staphylococcus Isolates and Their Antimicrobial Resistance Profiles and Virulence Genes from Subclinical Mastitis Cattle Milk Using MALDI-TOF MS, PCR and Sequencing in Free State Province, South Africa. Animals 2024, 14, 154. https://doi.org/10.3390/ani14010154.2076-2615 (online)10.3390/ani14010154http://hdl.handle.net/2263/97394This article belongs to the Special Issue titled 'Mastitis in Farm Animals: Pathogenesis, Diagnosis, Control, and Prevention'.DATA AVAILABILITY STATEMENT : The data used to support the findings of this study are available in the present manuscript.SUPPLEMENTARY MATERIAL: TABLE S1: Accession numbers of Staphylococcus isolates.Staphylococcus species are amongst the bacteria that cause bovine mastitis worldwide, whereby they produce a wide range of protein toxins, virulence factors, and antimicrobial-resistant properties which are enhancing the pathogenicity of these organisms. This study aimed to detect Staphylococcus spp. from the milk of cattle with subclinical mastitis using MALDI-TOF MS and 16S rRNA PCR as well as screening for antimicrobial resistance (AMR) and virulence genes. Our results uncovered that from 166 sampled cows, only 33.13% had subclinical mastitis after initial screening, while the quarter-level prevalence was 54%. Of the 50 cultured bacterial isolates, MALDI-TOF MS and 16S rRNA PCR assay and sequencing identified S. aureus as the dominant bacteria by 76%. Furthermore, an AMR susceptibility test showed that 86% of the isolates were resistant to penicillin, followed by ciprofloxacin (80%) and cefoxitin (52%). Antimicrobial resistance and virulence genes showed that 16% of the isolates carried the mecA gene, while 52% of the isolates carried the Lg G-binding region gene, followed by coa (42%), spa (40%), hla (38%), and hlb (38%), whereas sea and bap genes were detected in 10% and 2% of the isolates, respectively. The occurrence of virulence factors and antimicrobial resistance profiles highlights the need for appropriate strategies to control the spread of these pathogens.en© 2024 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).StaphylococcusVirulenceSubclinical mastitisCattleAntimicrobial resistance (AMR)SDG-03: Good health and well-beingSDG-02: Zero hungerArticle