Development of a PCR test for Staphylococcus chromogenes detection in caprine milk

Subclinical mastitis, often caused by Staphylococcus chromogenes, is related to milk quality and productivity loss in goats. The correct identification of the pathogen causing the infection allows a more effective treatment strategy. The objective of this study was to develop a specific PCR test for S. chromogenes identification, aiming to diagnose subclinical mastitis in goats. For this, the OrthoVenn3 platform was used to identify core genes exclusive to S. chromogenes and primers were designed using the Primer3Plus tool to amplify an exclusive gene (QDW81589.1). Primer specificity was evaluated in silico using BLAST and ehu.es and. Additionally, an in vitro validationusing DNAs from different Staphylococcus species was performed. Sensitivity was assessed using goat milk samples contaminated with S. chromogenes at different concentrations, which were subsequently evaluated by lactocultures and by the PCR developed in this present study. After this, the procedure developed in this study was used to detect S. chromogenes in mastitic goat milk. The results indicated that the primers developed specifically amplified a 200 bp fragment in DNA obtained from Staphylococcus chromogenes cultures and did not amplify the fragment in DNA obtained from other Staphylococcus species, confirming the test specificity. Also, it was observed the amplification in milk samples containing bacterial concentrations higher than 2.82 × 10⁶ CFU/ml and in mastitic goat milk. The PCR developed in the study allowed the Staphylococcus chromogenes identification It was also possible to detect the pathogen in goat milk samples, demonstrating the possibility of using the test in the diagnosis of mastitis.