Background/Objectives: Subclinical mastitis is a common and economically significant infection in dairy cows. This study aimed to assess the prevalence and antimicrobial susceptibility of bacteria causing subclinical mastitis in dairy cows in the Upper Cheliff Region, Northern Algeria, and to investigate the effects of subclinical mastitis on milk production and reproductive performance. Methods: A total of 263 cows from 23 farms were screened for subclinical mastitis using the California Mastitis Test (CMT) and sampled for isolation and identification of bacteria by MALDI-TOF MS. Antimicrobial susceptibility testing (AST) was performed using the Vitek 2 system and disk diffusion method. Methicillin resistance in staphylococci and mammaliicocci was confirmed by mecA/mecC detection. Results: The results revealed a prevalence of subclinical mastitis of 58.9% at the cow-level and 31.1% at the quarter-level. The most prevalent microorganisms identified were Enterococcus faecium (E. faecium) (24.4%) and Enterococcus faecalis (E. faecalis) (20.5%), followed by non-aureus staphylococci (NAS) (16.9%), Escherichia coli (E. coli) (7.9%) and Staphylococcus aureus (S. aureus) (7.08%). Risk factors significantly associated with the prevalence of subclinical mastitis included parity, lower milk production (<12 L/day), poor animal cleanliness and a history of previous mastitis. Reproductive performance was significantly impaired in cows with subclinical mastitis, cows with longer calving-to-first-service intervals (130 vs. 102.7 days; p < 0.0001), more services per conception (2.5 vs. 1.9; p < 0.0001) and a lower pregnancy rate at first service (24.5% vs. 48.1%; p < 0.0001). Furthermore, 43.2% of cows with subclinical mastitis required three or more inseminations to achieve pregnancy. AST showed a low resistance rate for the antimicrobial agents most commonly used in human and veterinary medicine. Coagulase-negative staphylococci (CoNS) showed remarkable rates of methicillin-resistance (22.2%), as well as resistance towards fosfomycin (37.8%) and tetracycline (31.1%). A substantial proportion of E. coli isolates exhibited resistance to piperacillin (40%) and ciprofloxacin (15%). All S. aureus isolates were classified as MSSA without detection of mecA and mecC genes. Conclusions: The results of this study demonstrated that subclinical mastitis is prevalent in the Upper Cheliff region and is associated with reduced milk production and reproductive performance. The detection of pathogenic and resistant microorganisms in milk is alarming and requires effective management strategies to control subclinical mastitis and improve dairy farm productivity.

Staphylococcus aureus colonizes the skin and nasal orifices of both humans and animals. Biofilm-related infections and acquired antimicrobial resistance render this bacterium a major health concern. This study examined the biofilm-forming capacity of eight S. aureus isolates originating from humans, sheep, and food matrices in northern Algeria. The efficiency of sanitizers (povidone–iodine [PVI], peracetic acid [PAA], quaternary ammonium compounds [QACs] and chlorhexidine [CHX]) in inhibiting and detaching biofilms was also assessed. Biofilm-production ability was assessed using a tissue culture plate. The minimal inhibitory concentration (MIC) and minimal bactericidal concentration were measured for each sanitizer, and antibiotic susceptibility profiles were determined by performing the Kirby-Bauer disk diffusion assay. All S. aureus isolates demonstrated the ability to produce biofilms, and among these, food and sheep mastitis isolates exhibited a strong capacity for biofilm formation. CHX exhibited the highest efficacy in bacterial biofilm inhibition and was considered more potent than the other sanitizers (P < 0.0001). Further, it exhibited lower MIC interval values (0.98-1.9 µg/g). However, MIC interval values for PVI, PAA, and QACs were 3125-25000, 39.06-156.25, and 0.98-3.9 mg/L, respectively. PVI did not affect biofilm detachment at different concentrations. All isolates were sensitive to methicillin (with mecA/mecC genes not detected), gentamicin, sulfamethoxazole–trimethoprim, vancomycin, and amikacin; however, two isolates were resistant to erythromycin and clindamycin. These findings demonstrated the biofilm-forming potential of S. aureus isolates and confirmed the effectiveness of sanitizers against S. aureus isolated from different sources.