Reduced Oxygen Condition Is Associated with Genome-Wide Expression Changes in Mastitis-Lineage Staphylococcus aureus During In Vitro Invasion into a Mammary Cell Line.

Staphylococcus aureus (S. aureus) bovine mastitis is a significant public health issue. Despite enormous efforts, important gaps remain regarding host-microenvironmental factors. How intramammary reduced oxygen modulates S. aureus transcription in bovine mammary epithelial cells (MECs) remains unclear. We examined oxygen-associated transcriptional changes in a bovine-mammary adapted S. aureus clone following internalization into MECs and identified functional category enrichments under Normal-O2 and Reduced-O2 exposures. Bovine MAC-T monolayers were infected with a dominant bovine mastitis isolate under Normal-O2 or Reduced-O2 conditions. Triplicate infection experiments were performed for each oxygen condition. Each condition included matched non-reacted bacterial controls maintained under the same gas condition but without MAC-T exposure serving as the reference condition for expression calling. RNA was extracted and profiled using a high-throughput qRT-PCR platform covering genome-wide loci. Expression calls were mapped to curated BioQT roles and interpreted descriptively. Results indicated 211 loci were upregulated and 99 were downregulated under Normal-O2 conditions, versus 53 upregulated and 35 downregulated under Reduced-O2 conditions, relative to their non-reacted controls. Under Normal-O2 conditions, regulated loci covered multiple functional roles, including cellular processes, transport/binding proteins, regulatory functions, and energy metabolism with downregulated loci enriched in transport/binding and cell-envelope categories. Under Reduced-O2 conditions, upregulated loci were abundant in cellular process annotations dominated by pathogenesis/toxin-related functions, whereas downregulated loci were enriched in nucleotide biosynthetic and DNA/cell division categories. Thus, this reveals oxygen-associated shifts in the transcriptional response of intramammary S. aureus in MAC-T cells. Normal-O2 conditions were associated with broader category representation, whereas Reduced-O2 conditions yielded a narrower distribution enriched for selected toxin/pathogenesis- and iron/cation-associated annotations. These oxygen-linked transcriptional-shifts highlight candidate pathways for the intramammary adaptation of S. aureus, potential diagnostic markers, anti-virulence strategies, and targeted therapeutics.