Activation of a Bovine Mammary Epithelial Cell Line by Ruminant-Associated Staphylococcus aureus is Lineage Dependent

Abstract

Bovine mastitis is a costly disease to the dairy industry and intramammary infections (IMI) with Staphylococcus aureus are a major cause of mastitis. Staphylococcus aureus strains responsible for mastitis in cattle predominantly belong to ruminant-associated clonal complexes (CCs). Recognition of pathogens by bovine mammary epithelial cells (bMEC) plays a key role in activation of immune responsiveness during IMI. However, it is still largely unknown to what extent the bMEC response differs according to S. aureus CC. The aim of this study was to determine whether ruminant-associated S. aureus CCs differentially activate bMEC. For this purpose, the immortalized bMEC line PS was stimulated with S. aureus mastitis isolates belonging to four different clonal complexes (CCs; CC133, CC479, CC151 and CC425) and interleukin 8 (IL-8) release was measured as indicator of activation. To validate our bMEC model, we first stimulated PS cells with genetically modified S. aureus strains lacking (protein A, wall teichoic acid (WTA) synthesis) or expressing (capsular polysaccharide (CP) type 5 or type 8) factors expected to affect S. aureus recognition by bMEC. The absence of functional WTA synthesis increased IL-8 release by bMEC in response to bacterial stimulation compared to wildtype. In addition, bMEC released more IL-8 after stimulation with S. aureus expressing CP type 5 compared to CP type 8 or a strain lacking CP expression. Among the S. aureus lineages, isolates belonging to CC133 induced a significantly stronger IL-8 release from bMEC than isolates from the other CCs, and the IL-8 response to CC479 was higher compared to CC151 and CC425. Transcription levels of IL-8, tumor necrosis factor alpha (TNFα), serum amyloid A3 (SAA3), Toll-like receptor (TLR)-2 and nuclear factor κB (NF-κB) in bMEC after bacterial stimulation tended to follow a similar pattern as IL-8 release, but there were no significant differences between the CCs. This study demonstrates a differential activation of bMEC by ruminant-associated CCs of S. aureus, which may have implications for the severity of mastitis during IMI by S. aureus belonging to these lineages.

Genomic Analysis of Staphylococcus aureus Isolates Associated With Peracute Non-gangrenous or Gangrenous Mastitis and Comparison With Other Mastitis-Associated Staphylococcus aureus Isolates

“…In the peracute gangrenous subgroup, 44% of the isolates belonged to CC133. Interestingly, the S. aureus isolates of CC133 induced a significantly stronger interleukin 8 (IL-8) release from bovine mammary epithelial cells in cell culture than isolates from the other CCs ( Hoekstra et al, 2019 ). In the 11 European countries that participated in the study by Hoekstra et al (2020) , the proportion of CC133 in each country varied from 0 to 25%.…”

Protective Roles of Folic Acid in the Responses of Bovine Mammary Epithelial Cells to Different Virulent Staphylococcus aureus Strains

“…First, the low common DEGs and DSGs induced by different S. aureus lineages showed the great heterogeneity of host immune responses to different S. aureus lineages. The challenge of S. aureus strains with different lineages in bovine mammary epithelial cells results in the differential expression of genes related to inflammation [ 9 , 32 ]. Among which, only a few candidate genes were tested by low-throughput qPCR or ELISA.…”

“…In the mastitis cell model, a previous study observed that the transcriptional expression levels of several candidate genes in primary bovine mammary epithelial cells (pbMECs) or Mac-T cells line are significantly different for different S. aureus lineages challenge [ 8 ]. For the host, the mammary epithelial cells in mammary gland establishes the first physical line of defense against the external pathogen, and a healthy mammary gland is vital in mobilizing the early innate immune response [ 9 , 10 ]. In our previous study, we isolated and cultured 191 S. aureus strains, including five MRSA strains, from bovine milk across nine Holstein dairy farms [ 11 , 12 ], and found that S. aureus is widely distributed in cows with different inflammatory responses.…”

The Synergism of PGN, LTA and LPS in Inducing Transcriptome Changes, Inflammatory Responses and a Decrease in Lactation as Well as the Associated Epigenetic Mechanisms in Bovine Mammary Epithelial Cells

“…These pathogenic bacteria include both Gram-positive bacteria, e.g., Staphylococcus aureus (S. aureus), Streptococcus agalactiae, Streptococcus spp., etc., as well as Gram-negative bacteria, such as Escherichia coli (E. coli), Klebsiella spp., Enterobacter spp., etc. [6][7][8][9][10]. E. coli and S. aureus are amongst the most common and important pathogens to induce mastitis [1,11].…”