Urinary Cell-Free miR-99a-5p as a Potential Biomarker for Estrus Detection in Buffalo

Accurate estrus detection method is the need of the hour to improve reproductive efficiency of buffaloes in dairy industry, as the currently available estrus detection methods/tools lack high sensitivity and specificity. Recently, circulating miRNAs have been shown as non-invasive biomarkers by various studies. Hence, in order to evaluate their potential as estrus biomarkers, the objective of this study was to identify and compare the levels of 10 hormone-responsive miRNAs in the urine collected at proestrus (PE), estrus (E), and diestrus (DE) phases of buffaloes (n = 3) pertaining to a discovery sample. Among 10 urinary miRNAs, the levels of bta-mir-99a-5p (E/PE 0.5-fold, P < 0.05; DE/PE 1.9-fold), bta-miR-125b (E/PE 0.5-fold; DE/PE 0.7-fold), bta-mir-145 (E/PE 1.5-fold; DE/PE 0.7-fold), bta-mir-210 (E/PE 1.2-fold, DE/PE 0.7-fold), mir-21 (E/PE 1.5-fold, DE/PE 2-fold), and bta-mir-191 (E/PE 1.3-fold; DE/PE 0.8-fold) were found to be altered during different phases of buffalo estrous cycle. In contrast, bta-mir-126-3p, bta-let-7f, bta-mir-16b, and bta-mir-378 were undetected in buffalo urine. Furthermore, a validation study in an independent group of 25 buffalo heifers showed the increased levels of urinary bta-mir-99a-5p during the DE (3.92-fold; P < 0.0001) phase as compared to the E phase. Receiver operating characteristic curve analyses also revealed the ability of urinary miR-99a-5p in distinguishing the E from the DE phase (area under the curve of 0.6464; P < 0.08). In silico analysis further showed an enrichment of miR-99a-5p putative targets in various ovarian signaling pathways, including androgen/estrogen/progesterone biosynthesis and apoptosis signaling, implicating the role of miR-99a-5p in ovarian physiology. In conclusion, significantly lower levels of bta-mir-99a-5p at the E phase than the DE phase in buffalo urine indicate its biomarker potential, which needs to be further explored in a large cohort in the future studies.

An Alternative Buffalo Urine-based Non-invasive Early Estrus Test using Wheat and Mung Bean Seed Germination

Background: The silent estrus problem in buffaloes is one of the major bottlenecks in buffalo management. Here, we present for the first time a simple and urine-based non-invasive seed germination inhibition test to detect the early estrus in buffaloes.Methods: The urine samples were collected from buffaloes on 0, 6, 10, 14, 18, 21 and 28 days after detection of signs of heat. The mungbean and wheat seeds (n=15 each) were treated with diluted (1:2 and 1:4) and undiluted urine samples. The results were analyzed in terms of germination inhibition percentage (GI%) and shoot length (SL) measurements. Control was established using distilled water in place of urine.Conclusion: In 1:4 dilution urine samples, a significant (P ≤ 0.05) decrease in GI% and SL were observed after 48 hours and 5 days, respectively in both the seeds, which could be useful as an estrus test. No significant results were observed for GI% and SL with undiluted and 1:2 diluted samples in both seeds. Our findings demonstrate that GI% and SL were inhibited around peri-estrus events, increased from 10th-14th days and then gradually decreased with 1:4 dilution of urine. This estrus test may assist the farmers in timing AI.

Evaluation of Physico-Chemical and Antioxidant Properties of Dairy Cow, Goat and Buffalo Urine in Two Different Seasons in A Sub-Tropical Region of India

The present study was conducted to evaluate the physicochemical and anti-oxidative properties of cow, buffalo and goat urine samples in two different successive seasons (summer and winter). For the physicochemical study, pH, electrical conductivity (EC), total dissolved solids (TDS) was measured and for the antioxidant properties, FRAP, DPPH and ABTS level was evaluated by standard methods. The data set were executed through the independent ‘t’ test by using IBM SPSS software version 22.0. The results showed that the EC level in all the (except cow) was significantly pandlt;0.5 lower in the winter season than in the summer season. TDS level in buffalo was significantly pandlt;0.5 lower in the winter season than the summer season but significantly higher in the winter season of the goat urine sample. In the case of antioxidant markers, FRAP and DPPH levels were significantly pandlt;0.5 higher in the summer season in all the animals, but there were no significant changes in the case of ABTS in all the animal’s urine samples. From these results, it is concluded that in the study area, the stress in the summer season as heat stress prevails in dairy cows, buffalo and goats. From the overall findings, it may be concluded that urine could be a good biomarker to evaluate the status of livestock health.

The bacteriological prevalence of leptospiral infection in cattle and buffaloes in West Malaysia

SUMMARYA cross-sectional bacteriological survey of cattle in West Malaysia revealed 14·4% (32/222) had leptospiral infection. Isolates were obtained from all except one herd with prevalence of infection in herds ranging from 0–44·8%. A small number of buffalo urine samples were examined and all of them were found to be negative. A leptospiral isolate obtained from a bovine kidney proved to be n new scrovar ofLeplospira interrogansand the nameunipertamawas assigned to it. Six other leptospiral serovars were isolated, namelycanicola, australis, javanica,ballum, pomonaandhardjo. All six serovars were isolated for the first time in cattle in Malaysia.Cattle in Malaysia appear to be the maintenance host for serovarhardjo. The presence of the other serovars in cattle was probably duo to contact with the maintenance hosts, pigs for serovarpomonaand rodents for the other three serovars. It appears that the epidemiology of leptospiral infection in cattle in Malaysia is similar to that reported overseas.

Control of flea beetle, Phyllotreta nemorum L. (Coleoptera: Chrysomelidae) using locally available natural resources

Aqueous extracts of six different plants (Acorus calamus, Ageratum conyzoides, Azadirachta indica, Duranta repens, Spilanthes acmella and Urtica dioca) and diluted animal urine (buffalo and cow) were tested for mortality rate of flea beetle (Phyllotreta nemorum) in the laboratory. Results were compared with the effects of commercial neem product (neem azal) on flea beetle mortality. The host plant taken for the study was radish (Rhaphanus sativus). Three concentrations of aqueous plant extracts (1kg/5 l, 1kg/10 l and 1kg/20 l of water), three concentrations of animal urine (20%, 15% and 10%) and two concentrations of neem azal (0.1% and 0.01%) were tested in three replications. Observations on the beetle mortality were made at 24 hrs and thereafter on alternate days for a week (168 hrs). All tested concentrations of S. acmella, buffalo urine and cow urine were effective in flea beetle control; A. calamus, A. indica and U. dioca were significantly better in controlling flea beetle (P