Klebsiella pneumoniae infection causes mitochondrial damage and dysfunction in bovine mammary epithelial cells

Abstract

Klebsiella pneumoniae, an important cause of bovine mastitis worldwide, is strongly pathogenic to bovine mammary epithelial cells (bMECs). Our objective was to determine the role of mitochondrial damage in the pathogenicity of K. pneumoniae on bMECs, by assessing several classical indicators of mitochondrial dysfunction, as well as differentially expressed genes (DEGs). Two K. pneumoniae strains (HLJ-D2 and HB-AF5), isolated from cows with clinical mastitis (CM), were used to infect bMECs (MAC-T line) cultured in vitro. In whole-transcriptome analysis of bMECs at 6 h post-infection (hpi), there were 3453 up-regulated and 3470 down-regulated genes for HLJ-D2, whereas for HB-AF5, there were 2891 up-regulated and 3278 down-regulated genes (P < 0.05). Based on GO term enrichment of differentially expressed genes (DEGs), relative to the controls, the primary categories altered in K. pneumoniae-infected bMECs included cellular macromolecule metabolism, metabolic process, binding, molecular function, etc. Infections increased (P < 0.05) malondialdehyde concentrations and formation of reactive oxygen species in bMECs. Additionally, both bacterial strains decreased (P < 0.05) total antioxidant capacity in bMECs at 6 and 12 hpi. Furthermore, infections decreased (P < 0.05) mitochondrial membrane potential and increased (P < 0.01) mitochondrial calcium concentrations. Finally, severe mitochondrial swelling and vacuolation, as well as mitochondrial rupture and cristae degeneration, were detected in infected bMECs. In conclusion, K. pneumoniae infections induced profound mitochondrial damage and dysfunction in bMECs; we inferred that this caused cellular damage and contributes to the pathogenesis of K. pneumoniae-induced CM in dairy cows.

Suitability of milk lactate dehydrogenase and serum albumin for pathogen-specific mastitis detection in automatic milking systems

“…The reason for investigating SA in this in vivo study is that SA appears earlier than LDH (2 vs. 5 h) in experimentally induced mastitis as reported in other studies (Wall et al, 2016;Wellnitz et al, 2016). In addition to the transfer via the blood-milk barrier, both SA and LDH may originate from mammary epithelial cell production that may cause an increase of SA (Shamay et al, 2005), and mammary cell damage may contribute to the increased concentration of LDH in milk (Wellnitz et al, 2016;Cheng et al, 2021). The potential contribution of the 2 sources of SA and LDH may support the possibility to distinguish between IMI-causing pathogen types.…”

Correlation between Polymerase Chain Reaction Identification of Iron Acquisition Genes and an Iron-Deficient Incubation Test for Klebsiella pneumoniae Isolates from Bovine Mastitis

“…K. pneumoniae strains with multiple iron-acquisition systems account for >90% of pathogenic K. pneumoniae strains in humans [17]. Virulence genes encoding enterobactin, aerobactin, and yersiniabactin have also frequently been detected in PCR analyses of K. pneumoniae isolates from environmental samples collected from dairy farms, as well as mastitis milk samples [2,11,20]. Coliform bacteria, including K. pneumoniae, can profitably utilize several iron chelators when invading and subsequently surviving within the mammary glands due to the selective and effective diversion of these acquired substances for their survival in the dairy environment [2].…”

“…The iron concentration in the iron-deficient medium described previously and used in this study is less than 0.5 µM [3,25]. Our study using iron-deficient incubation tests can be developed by including other laboratory tests, such as bovine mammary epithelial cell culture to identify bacterial adhesion function, for the further investigation of the virulence genes facilitating K. pneumoniae-associated bovine mastitis [20].…”

“…Additionally, an iron-deficient incubation test has previously been used to evaluate the growth rates of K. pneumoniae with or without iron-acquisition genes. Bovine mammary epithelial cell cultures have previously been used to identify the adhesion activity associated with fimbriae genes for the in vitro assessment of coliform isolates from bovine mastitis samples [20]. However, mastitis-associated coliform bacteria have not been incubated in an iron-deficient medium previously.…”

Emerging connections between gut microbiome bioenergetics and chronic metabolic diseases

“…Interestingly, PTSs are widespread in Enterobacteriaceae but found only within selected genera of Firmicutes (e.g., Clostridium, Enterococcus, Streptococcus, Staphylococcus). Microbial metabolites from these bacteria have been reported to cause mitochondrial swelling and cristae degeneration to occur via multiple signaling pathways in mammalian epithelial cells (Cheng et al, 2021), suggesting a direct potential to disrupt host metabolism (Cheng et al, 2021). While antibiotic-resistant isolates from foodborne (Lawal et al, 2021) and hospital-acquired origins (Zhang et al, 2013) are commonly enriched in PTSs, commensal strains from healthy individuals usually lack PTS-encoding genes.…”