Comparative cell free salivary miRNA profile in Bubalus bubalis between diestrus and estrus stages

Despite buffaloes being primary farm animals, their reproductive performance remains poor mainly due to inaccurate estrus detection methods that ultimately has an economic impact on dairy industry as well as farmers. Recently, numerous studies showed potential of miRNAs as estrus biomarker. However, a miRNA profile of buffalo cell free saliva, a non-invasive fluid, at estrus and diestrus stages is missing. Hence, the present study was planned to identify differential levels of salivary cell free miRNAs in estrus as compared to the diestrus phase of buffalo oestrous cycle (n=3) in order to discover a possible estrus specific miRNAs as biomarkers. miRNA-Seq data analysis showed that in total 10 miRNAs i.e bta-miR-375, bta-miR-200c, bta-miR-30d, bta-let-7f, bta-miR-200a, bta-miR-12034, bta-let-7b, bta-miR-142-5p, bta-miR-2467-3p, bta-miR-30a-5p are significantly altered (log2foldchange >3 and p<0.05) during estrus in comparison to the diestrus phase in buffaloes, suggesting their estrus biomarker potential. Overall, 8 miRNAs i.e bta-miR-375 (6.87 Fold; p-value 0.003), bta-miR-200c (5.98 Fold; p-value 0.003), bta-miR-30d (4.17 Fold; p-value 0.015), bta-let-7f (3.34 Fold; p-value 0.022), bta-miR-200a (4.92 Fold; p-value 0.024), bta-miR-12034 (3.58 Fold; p-value 0.0025), bta-let-7b (3.06 Fold; p-value 0.031), bta-miR-30a-5p (4.7 Fold; p-value 0.036) were upregulated, whereas bta-miR-142-5p (-3.4 Fold; p-value 0.032) and bta-miR-2467-3p (-5.24 Fold; p-value 0.035) were downregulated during estrus. However, further validation study using qPCR is required in a large sample size in order to determine their estrus biomarker potential. In summary, our results revealed differential salivary cell free miRNAs profile during the oestrous cycle that may lead to the development of estrus specific miRNAs based point-of-care test applicable for the reproductive management of buffaloes in the field condition in the near future. Saliva samples were collected from three healthy murrah buffaloes at estrus and diestrus phase of an oestrous cycle.