We assessed the suitability of 9 internal control genes (ICG) in milk somatic cells of lactating cows to find suitable reference genes for use in quantitative PCR (qPCR). Eighteen multiparous lactating Sahiwal cows were used, 6 in each of 3 lactation stages: early (25 ± 5 d in milk), mid (160 ± 15 d in milk), and late (275 ± 25 d in milk) lactation. Nine candidate reference genes [glyceraldehyde 3-phosphate dehydrogenase (GAPDH), protein phosphatase 1 regulatory subunit 11 (PPP1R11), β-actin (ACTB), β-2 microglobulin (B2M), 40S ribosomal protein S15a (RPS15A), ubiquitously expressed transcript (UXT), mitochondrial GTPase 1 (MTG1), 18S rRNA (RN18S1), and ubiquitin (UBC)] were evaluated. Three genes, β-casein (CSN2), lactoferrin (LTF), and cathelicidin (CAMP) were chosen as target genes. Very high amplification was observed in 7 ICG and very low level amplification was observed in 2 ICG (UXT and MTG1). Thus, UXT and MTG1 were excluded from further analysis. The qPCR data were analyzed by 2 software packages, geNorm and NormFinder, to determine suitable reference genes, based on their stability and expression. Overall, PPP1R11, ACTB, UBC, and GAPDH were stably expressed among all candidate reference genes. Therefore, these genes could be used as ICG for normalization of qPCR data in milk somatic cells through lactation.

Aim:The present study was aimed to study the use of cell counts as an early indicator of mammary health.Materials and Methods:Milk and blood cell counts were estimated from 8 healthy, 8 subclinical (SCM), and 8 clinically mastitis (CM) groups of Karan Fries (KF) cows.Results:Total leucocyte counts and neutrophil percent in blood and milk somatic cells and milk neutrophil percent of healthy cows increased significantly (p<0.05) in SCM cows and CM cows. Viability of blood and milk neutrophils was more in healthy cows, but decreased significantly (p<0.05) in SCM and CM cows. Significant (p<0.05) decrease were also observed in both the blood and milk lymphocytes and monocytes of SCM and CM cows. Phagocytic activity (PA) of blood neutrophils also decreased significantly (p<0.05) in SCM cows. There was no difference between the PA of SCM and CM cows. Milk neutrophil percent was more in the SCM and clinically infected milk than in the blood of these cows. About 96-97% of the neutrophils had segmented nucleus in both healthy and subclinical milk, whereas, 2-3% were having band shaped or immature nuclei. There was a significant decrease in the segmented neutrophils, whereas, band neutrophils increase significantly to about 5% in the infected milk of mastitic cows. Viability of the milk neutrophils decreased more in case of subclinical and clinical milk as compared to that of blood. PA was found to be highest in the milk of healthy group of cows, but decreased significantly (p<0.05) in subclinically infected cows. However, there was no difference between the PA of milk neutrophils of SCM and CM cows. PA of milk was also found to be significantly lower in the milk of healthy cows when compared to that of blood neutrophils.Conclusion:This study indicated that percent neutrophils and their type in conjunction with milk somatic cell counts can be used as a more reliable indicator of mammary health in cows.

Thirty-six cross-bred cows were used to study the effect of long-chain (flaxseed) or short-chain (butyric acid) fatty acid supplementation on metabolic status, ovarian function and reproduction performance during transition period. Control cows received a routine feed of transition diet, while the cows in two treatment groups were supplemented with either 750-g crushed flaxseed or 250 g butyric acid per cow per day. Ovarian activity was monitored by transrectal ultrasonography on 10th, 20th and 30th days post-partum. Blood samples were collected by jugular venipuncture into heparinized polystyrene tubes; plasma was prepared and stored under -20 °C until analysis. Results indicated that cows in flaxseed group were in positive energy balance as indicated by lower NEFA and Beta hydroxy Butyrate and higher glucose concentrations. Uterine involution was completed well within 30 days post-partum in all the cows in flaxseed fed group compared to 76.9% in butyric acid supplemented and 61.5% in control groups. The size of dominant follicle and corpus luteum was significantly higher (p < 0.05) for flaxseed group compared to control group, which in turn resulted in higher concentrations of plasma progesterone. Cows fed on diets supplemented with flaxseed exhibited post-partum heat earlier and bred sooner (p < 0.05) than control cows. It has been noticed that supplementation of flaxseed and butyric acid enhanced involution of uterus, early resumption of cyclicity and thereby early breeding. However, in view of the encouraging results obtained for flaxseed supplemented group, its organic nature and easier availability at farmer's gate, we concluded that flaxseed can be safely included in transition diet to modulate reproductive performance of dairy cattle.

Micronutrients when fed around peripartum may reduce the stress induced by cortisol. With this objective, 24 Sahiwal (SW) and 24 Karan Fries (KF) cows and 24 Murrah (Mu) buffaloes were taken and divided into four groups of six each. Vitamin E (VE), zinc (Zn) and copper were supplemented from 30 days pre- to 30 days postcalving in groups 1, 2 and 3. Animals without supplementation served as control. Blood sampling was done on days 30, 15, 7 and 3 precalving; at calving; and on days 3, 7, 15 and 30 postcalving. Plasma cortisol levels were measured by ELISA. Supplementation of VE and Zn significantly (P<0.05) reduced plasma cortisol levels at calving and at 30 days postcalving. KF cows exhibited best results with VE, whereas SW cows and Mu buffaloes responded best to Zn. Copper increased the peripartum cortisol levels. Reduction in cortisol levels may help in keeping the animal healthy and stronger to fight immuno-suppression generally observed around the period of peripartum.

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