|Schroeder, Steven - Steve|
|Reinhardt, Timothy - Tim|
Submitted to: Biomed Central (BMC) Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/15/2015
Publication Date: 10/16/2015
Citation: Sun, J., Aswath, K., Schroeder, S.G., Lippolis, J.D., Reinhardt, T.A., Sonstegard, T.S. 2015. MicroRNA expression profiles of bovine milk exosomes in response to Staphylococcus aureus infection. Biomed Central (BMC) Genomics. 16:806. Interpretive Summary: Mastitis is an infection of the mammary gland which is estimated to cost the dairy industry about $2 billion US dollars annually. This study was designed to better understand how expression of small, non-coding RNA inhibitors of protein synthesis known as microRNAs are changed within cow’s milk after bacterial infection of the mammary gland. MicroRNAs within milk have the potential to serve as diagnostic markers of udder health in a lactating cow. Using next-generation sequencing, we interrogated the total RNA fraction isolated from milk of healthy cows and cows artificially challenged with bacterial infection. Our results identified at least 14 microRNAs that change in prevalence due to infection. Two of these microRNAs are not present in healthy cows, thus they serve as candidates for biomarker development. Such biomarkers can be used to categorize lactating animals with bacterial infection and characterize their resistance or susceptibility to mastitis.
Technical Abstract: Background: Milk exosomes are a rich source of microRNAs (miRNAs) that are protected from degradation. Ingestion of milk and subsequent absorption of miRNAs into recipient cells by endocytosis may play a role in the regulation of neonatal innate and adaptive immunity. In contrast, the miRNA content of milk exosomes may also be indicative of a lactating animal’s health; whereby, the presence or absence of specific miRNAs could serve as biomarkers for early detection of bacterial infection that can lead to mastitis. In the present study, we therefore analyzed and compared miRNA expression profiles of milk exosomes from four Holstein cows obtained during mid-lactation prior to and after infection (48 h) of the mammary gland with Staphylococcus aureus. Results: Next generation sequencing of eight, unpooled small RNA libraries derived from milk exosomes produced about 60.5 million high-quality, bovine-specific sequence reads for comparison of miRNA expression between treatments. Sequence identity analysis showed the miRNAs make up about 13% of the average RNA content of these exosomes. Although 417 known bovine miRNAs were identified, miRNAs represented the least diverse class of RNA accounting for only 1% of all unique sequences. The 20 most prevalent unique sequences within this class accounted for about 90% of the total miRNA-associated reads across samples. Non-annotated, unique reads provided evidence for another 303 previously unknown bovine miRNAs. Expression analyses found 14 known bovine microRNAs significantly differed in frequency between exosomes from infected and control animals. Conclusions: Our survey of miRNA expression from uninfected milk exosomes and those produced in response to infection provides new and comprehensive information supporting a role for delivery into milk of specific miRNAs involved in immune response. In particular, bta-miR-142-5p, and -223 are potential biomarkers for early detection of bacterial infection of the mammary gland. Additionally, 22 mammary-expressed genes involved in regulation of host immune processes and response to inflammation were identified as potential binding targets of the differentially expressed miRNAs.