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Profiling of milk miRNAs associated with the innate immune system and pathway analysis in dairy cows with S. aureus-infected subclinical mastitis

Published online by Cambridge University Press:  06 March 2025

Özge Özmen Open the ORCID record for Özge Özmen [Opens in a new window] ,
Berna Kaya ,
Kardelen Karaman  and
Fatma Seda Bilir
Özge Özmen*
Affiliation:
Faculty of Veterinary Medicine, Department of Genetics, Ankara University, Ankara, Türkiye
Berna Kaya
Affiliation:
Faculty of Veterinary Medicine, Department of Genetics, Ankara University, Ankara, Türkiye
Kardelen Karaman
Affiliation:
Faculty of Veterinary Medicine, Department of Animal Breeding, Kırıkkale University, Kırıkkale, Türkiye
Fatma Seda Bilir
Affiliation:
Faculty of Veterinary Medicine, Department of Food Hygiene and Technology, Ankara University, Ankara, Türkiye
*
Corresponding author: Özge Özmen; Email: [email protected]
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Abstract

We address the hypothesis that bioinformatics analysis can effectively identify miRNAs and target genes associated with the innate immune response to Staphylococcus aureus-induced mastitis in cows. Gram-positive bacteria, such as S. aureus, are some of the mastitis pathogens that cause subclinical infection and pose one of the most severe threats to dairy cattle due to their contagiousness and incidence. Therefore, newer molecular markers must be identified to predict bovine mastitis non-invasively with low error and high specificity. To address this, we conducted in-silico analysis to identify hub genes related to the innate immune system specific to the S. aureus pathogen in subclinical mastitis infections in cows, and experimental validation of the miRNAs of these determined hub genes was completed. As a result of bioinformatic analysis, we identified five hub genes (TLR2, MYD88, CASP4, NOD2 and TBK1) related to the innate immune system that are specific to the S. aureus pathogen. We investigated the expression levels of miRNAs associated with these genes (bta-miR-15a, bta-miR-16b, bta-miR-23a, bta-miR-27a-3p, bta-miR-103, bta-miR-146b, and bta-mir-374b) using real-time quantitative PCR. Except for bta-miR-15a and bta-miR-23a, all miRNAs studied varied in expression between healthy cows and cows with subclinical mastitis infected with S. aureus. This is the first study to bioinformatically determine hub genes specific to the innate immune system and the S. aureus pathogen in subclinical mastitis infections and then validate the determined miRNAs in milk between healthy and subclinical mastitis infected cows. The findings of our study expand our understanding of the roles of these miRNAs in cows with S. aureus-infected subclinical mastitis.

Keywords

Bioinformaticsinnate immunitymilk miRNAsS. aureussubclinical mastitis
Type
Research Article
Information
Journal of Dairy Research , First View , pp. 1 - 6
Copyright
Copyright © The Author(s), 2025. Published by Cambridge University Press on behalf of Hannah Dairy Research Foundation

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References

Arenas-Padilla, M, González-Rascón, A, Hernández-Mendoza, A, Calderón de la Barca, AM, Hernández, J and Mata-Haro, V (2022) Immunomodulation by Bifidobacterium animalis subsp. lactis Bb12: integrative analysis of miRNA expression and TLR2 pathway-related target proteins in swine monocytes. Probiotics and Antimicrobial Proteins 14, 510–522.CrossRefGoogle ScholarPubMed
Bai, X, Wang, X, Lin, T, Dong, W, Gao, Y, Ji, P, Zhang, Y, Zhao, X and Zhang, Q (2022) Toll-like receptor 2 is associated with the immune response, apoptosis, and angiogenesis in the mammary glands of dairy cows with clinical mastitis. International Journal of Molecular Sciences 23, 18.CrossRefGoogle ScholarPubMed
Barnum, DA and Newbould, FH (1961) The use of the California mastitis test for the detection of bovine mastitis. Canadian Veterinary Journal 2, 83–90.Google ScholarPubMed
Chen, L, Liu, X, Li, Z, Wang, H, Liu, Y, He, H, Yang, J, Niu, F, Wang, L and Guo, J (2014) Expression differences of miRNAs and genes on NF-κB pathway between the healthy and the mastitis Chinese Holstein cows. Gene 545, 117–125.CrossRefGoogle ScholarPubMed
Dysin, AP, Barkova, OY and Pozovnikova, MV (2021) The role of microRNAs in the mammary gland development, health, and function of cattle, goats, and sheep. Non-Coding RNA 7, 78.CrossRefGoogle ScholarPubMed
FAO (2021) Food Outlook—Biannual Report on Global Food Markets. Rome. Available at https://www.fao.org/documents/card/en/c/cb9427enGoogle Scholar
Günther, J, Petzl, W, Bauer, I, Ponsuksili, S, Zerbe, H, Schuberth, HJ, Brunner, RM and Seyfert, HM (2017) Differentiating Staphylococcus aureus from Escherichia coli mastitis: S. aureus triggers unbalanced immune-dampening and host cell invasion immediately after udder infection. Scientific Reports 7, 4811.CrossRefGoogle ScholarPubMed
Jin, W, Ibeagha-Awemu, EM, Liang, G, Beaudoin, F, Zhao, X and Guan, LL (2014) Transcriptome microRNA profiling of bovine mammary epithelial cells challenged with Escherichia coli or Staphylococcus aureus bacteria reveals pathogen directed microRNA expression profiles. BMC Genomics 15, 181.CrossRefGoogle ScholarPubMed
Jurkin, J, Schichl, YM, Koeffel, R, Bauer, T, Richter, S, Konradi, S, Gesslbauer, B and Strobl, H (2010) miR-146a is differentially expressed by myeloid dendritic cell subsets and desensitizes cells to TLR2-dependent activation. Journal of Immunology 184, 4955–4965.CrossRefGoogle ScholarPubMed
Kawai, T and Akira, S (2010) The role of pattern-recognition receptors in innate immunity: update on Toll-like receptors. Nature Immunology 11, 373–384.CrossRefGoogle ScholarPubMed
Lai, YC, Fujikawa, T, Maemura, T, Ando, T, Kitahara, G, Endo, Y, Yamato, O, Koiwa, M, Kubota, C and Miura, N (2017) Inflammation-related microRNA expression level in the bovine milk is affected by mastitis. PLoS One 12, e0177182.CrossRefGoogle ScholarPubMed
Livak, KJ and Schmittgen, TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) method. Methods (San Diego, Calif.) 25, 402–408.CrossRefGoogle ScholarPubMed
Momen-Heravi, F and Bala, S (2018) miRNA regulation of innate immunity. Journal of Leukocyte Biology 103, 1205–1217.CrossRefGoogle Scholar
Oviedo-Boyso, J, Valdez-Alarcón, JJ, Cajero-Juárez, M, Ochoa-Zarzosa, A, López-Meza, JE, Bravo-Patiño, A and Baizabal-Aguirre, VM (2007) Innate immune response of bovine mammary gland to pathogenic bacteria responsible for mastitis. Journal of Infection 54, 399–409.CrossRefGoogle ScholarPubMed
Pietrocola, G, Nobile, G, Rindi, S and Speziale, P (2017) Staphylococcus aureus manipulates innate immunity through own and host-expressed proteases. Frontiers in Cellular and Infection Microbiology 7, 166.CrossRefGoogle ScholarPubMed
Rainard, P, Foucras, G, Fitzgerald, JR, Watts, JL, Koop, G and Middleton, JR (2018) Knowledge gaps and research priorities in Staphylococcus aureus mastitis control. Transboundary and Emerging Diseases 65, 149–165.CrossRefGoogle ScholarPubMed
Saikh, KU (2021) Myd88 and beyond: a perspective on MyD88-targeted therapeutic approach for modulation of host immunity. Journal of Immunology Research 69, 117–128.CrossRefGoogle ScholarPubMed
Schanzenbach, CI, Kirchner, B, Ulbrich, SE and Pfaffl, MW (2017) Can milk cell or skim milk miRNAs be used as biomarkers for early pregnancy detection in cattle? PLoS One 12, e0172220.CrossRefGoogle ScholarPubMed
Sordillo, LM (2018) Mammary gland immunobiology and resistance to mastitis. Veterinary Clinics of North America: Food Animal Practice 34, 507–523.Google ScholarPubMed
Srikok, S, Patchanee, P, Boonyayatra, S and Chuammitri, P (2020) Potential role of MicroRNA as a diagnostic tool in the detection of bovine mastitis. Preventive Veterinary Medicine 182, 105101.CrossRefGoogle ScholarPubMed
Tzelos, T, Ho, W, Charmana, VI, Lee, S and Donadeu, FX (2022) MiRNAs in milk can be used towards early prediction of mammary gland inflammation in cattle. Scientific Reports 12, 5131.CrossRefGoogle ScholarPubMed
Umesh, A, Guttula, PK and Gupta, MK (2022) Prediction of potential molecular markers of bovine mastitis by meta-analysis of differentially expressed genes using combined p value and robust rank aggregation. Tropical Animal Health and Production 54, 269.CrossRefGoogle ScholarPubMed
Wang, XP, Luoreng, ZM, Zan, LS, Raza, SH, Li, F, Li, N and Liu, S (2016) Expression patterns of miR-146a and miR-146b in mastitis infected dairy cattle. Molecular and Cellular Probes 30, 342–344.CrossRefGoogle ScholarPubMed
Wang, MQ, Zhou, CH, Cong, S, Han, DX, Wang, CJ, Tian, Y, Zhang, JB, Jiang, H and Yuan, B (2021) Lipopolysaccharide inhibits triglyceride synthesis in dairy cow mammary epithelial cells by upregulating miR-27a-3p, which targets the PPARG gene. Journal of Dairy Science 104, 989–1001.CrossRefGoogle ScholarPubMed
Wellnitz, O and Bruckmaier, RM (2012) The innate immune response of the bovine mammary gland to bacterial infection. The Veterinary Journal 192, 148–152.CrossRefGoogle ScholarPubMed
Zaatout, N, Ayachi, A and Kecha, M (2020) Staphylococcus aureus persistence properties associated with bovine mastitis and alternative therapeutic modalities. Journal of Applied Microbiology 129, 1102–1119.CrossRefGoogle ScholarPubMed
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