Uterine, vaginal and placental blood flows increase with dynamic changes in serum metabolic parameters and oxidative stress across gestation in buffaloes.

The aims of the present study were to determine uterine, vaginal and placental blood flows by Doppler ultrasound cross-buffalo gestation and to evaluate the relationships among reproductive Doppler parameters and serum metabolic parameters as well as oxidative stress. Uterine (UA) and vaginal (VA) arteries were scanned every month, and placentome was scanned from month 4 till 8 in gestation. Time-averaged maximum velocity (TAMV), pulsatility index (PI), resistance index (RI), systolic/diastolic ratio (SD) and arterial diameter (AD) were used for accessing UA and VA hemodynamics. Time-averaged maximum velocity positively correlated with and AD, and both negatively correlated with their PI, RI and SD in UA and VA. TAMV and AD increased constantly in pregnancy, with maximum increase in months 4 and 9. Pulsatility index, RI and AD of UA decreased between months 4 and 9, while PI, RI and AD of VA decreased between months 5 and 9 and then increased in month 10 in pregnancy. Time-averaged maximum velocity of placentome blood flow increased exponentially from months 4 to 8, but decreased at the last two months in pregnancy. Serum lipids were significantly higher in the first month compared to all other months, while glucose was significantly lower in months 9 and 10. Malondialdehyde increased from month 3 till term, but peaked in month 5 and 10. Glutathione and catalase were highest in the first month and remained after. Time-averaged maximum velocity and AD for both UA and VA negatively correlated with serum lipids, glucose, catalase and glutathione, while positively correlated with malondialdehyde and total protein. Thus, increases in uterine blood flow (UtBF), vaginal blood flow (VaBF) and placental blood flow (PaBF) are associated with increased metabolism and oxidative stress in buffalo pregnancy.

Differential expression of miRNAs and related mRNAs during heat stress in buffalo heifers.

The present experiment was aimed to study differential expression of miRNAs and related mRNAs during heat stress (HS) in buffalo heifers. Twelve Murrah buffalo heifers aged between 1.5 and 2.0 years, weighting between 250 and 300 Kg were randomly assigned into two equal groups. The animals were kept in the psychrometric chamber under Thermo-neutral (TN; THI = 72) and HS (THI = 87-90) conditions for 6 h every day between 1000 and 1600 h for 21 days. The blood sampling was done at 1500 h on 15th day of the experiment and physiological parameters viz. pulse rate (PR), respiratory rate (RR) and rectal temperature (RT) were recorded at 1500 h on day -5, -3, -1, 0, +1, +3, +5 with respect to blood sampling. PBMCs were used for extraction of miRNAs and total RNA; and first strand cDNA was synthesized. qPCR was performed for relative gene expression studies. Physiological, hematological (erythrocytic indices), biochemical (triglycerides, urea, ALT, AST, LDH), redox (SOD, ROS) and endocrine parameters (T4) altered significantly (P < 0.05) during HS as compared to TN. Out of eight targeted miRNAs only four were expressed in buffalo heifers. The relative expression of bta-mir-142, bta-mir-1248 and bta-mir-2332 was significantly (P < 0.05) up-regulated whereas expression of bta-mir-2478 was significantly (P < 0.05) down-regulated during HS as compared to TN. The relative expression of the predicted target genes i.e. HSF1, HSP60, HSP70, HSPA8 and HSP90 were significantly (P < 0.05) up-regulated whereas HSF4 expression was significantly (P < 0.05) down-regulated during HS as compared to TN. It can be concluded that a THI of 87-90 could lead to a moderate HS in buffalo heifers. Differential expression studies of miRNAs and related mRNAs in present study deciphers the role of miRNAs in the heat tolerance in buffalo heifers.

Mineral availability in soil and fodders affecting blood profiles in Nili-Ravi dairy buffaloes.

A mineral assessment study for dairy buffaloes, Nili-Ravi breed, was carried out seasonally at silvopasture farm at Sargodha, Pakistan. Sampling for soil, forage, and buffalo serum was done seasonally for a period of 1 year for mineral (P, Mg, Na, K, Ca) evaluation. In the study, experiments on forages showed significant differences in mean concentrations of Mg and P with changing seasons, whereas the values for Ca, Mg, Na, and K did not vary significantly. Three groups of buffaloes were checked for mineral concentration in blood serum, viz., calves, lactating and non-lactating. Ca mean concentration varied significantly for non-lactating buffaloes and calves between two seasons. Na mean level showed a significant difference for two sampling seasons among calves and lactating buffaloes. Mean level for Mg only differed significantly in serum of non-lactating buffaloes between the two seasons. Mean levels for Ca and P in soil and forage remained higher than the optimal level, whereas soil Na was below the critical level. K concentrations in soil and blood serum were above the critical level, while Mg levels remained below the optimal range in soil, forage, and blood serum. K concentration in forage remained below the optimal level. All buffalo categories displayed lower levels of Ca and P in serum. Na concentration in forage and serum exceeded the critical level. Furthermore, Ca yielded significant and positive correlation between soil and serum level, whereas K had significant but negative correlation for soil-forage and forage-serum. Overall, it was determined that P deficiency existed among buffaloes, which could be improved via supplementation. Similarly, K deficiency and Mg deficiency were exhibited for forage and soil respectively which could be countered through addition of fertilizers rich for these minerals. In brief, the mineral utilization of buffaloes is affected by reduced availability of essential nutrients and may result in lower productivity. It is suggested to add mineral supplementation in addition to natural diet of buffaloes to enhance the productivity of these animals.

Comparative biochemical profiles, utero-ovarian function, and fertility of the postpartum buffalo with and without subclinical endometritis.

In postpartum buffaloes, the process of uterine involution and changes in blood metabolic profile has not been studied in relation to development of subclinical endometritis (SCE). In this study, buffaloes (n = 100) approaching calving were identified. Weekly blood samples were collected on the day of calving up to 6 weeks post-calving. The diameter of uterine horns and onset of ovarian cyclicity (corpus luteum) were recorded through ultrasonography. On the basis of polymorphonuclear cell (PMN) cell count in endometrial cytology at days 45-50 postpartum, buffaloes were divided into two groups, viz., with SCE (> 5% PMN; n = 38) and without SCE (≤ 5% PMN; n = 62). Buffaloes with SCE took longer (P < 0.05) time to complete uterine involution and had larger (P < 0.05) uterine horn diameter between 3rd and 6th weeks postpartum and lower prostaglandin F2α metabolite (PGFM) concentration on the day of calving (P < 0.05) and 1 week (P < 0.001) post-calving than without SCE group. Buffaloes with SCE had lower (P < 0.001) concentration of glucose at weeks 2 and 3, higher (P < 0.001) ß-hydroxybutyric acid (BHBA) at week 3, and lower serum albumin concentration throughout the sampling period (P < 0.05 to 0.001) except at 1 week post-calving as compared to without SCE group. The urea concentration was significantly lower (P < 0.05 to 0.001) in buffaloes with SCE from 4 weeks post-calving onwards than without SCE group. The calcium concentration was lower in buffaloes with SCE at weeks 5 (P < 0.001) and 6 (P < 0.05) postpartum, whereas the concentration of magnesium and phosphorus was uniform between the two groups. No significant (P > 0.05) difference in onset of ovarian cyclicity between the 2 groups was observed, whereas buffaloes with SCE had longer (P = 0.001) median days open (141 days) than their counterpart (117 days). The first service conception rate, cumulative pregnancy rate, and pregnancy rate at 150 days postpartum were lower (P < 0.05) in buffaloes with SCE than without SCE group. In summary, higher BHBA and lower serum concentrations of glucose, albumin, urea, and calcium control onset of subclinical endometritis which in turn has negative impact on fertility of buffaloes.

Effect of plastic foreign body impaction on rumen function and heavy metal concentrations in various body fluids and tissues of buffaloes.

Rumen impaction by plastic waste (PW) is a common condition in the developing countries where ruminants are reared under extensive grazing. As heavy metals (HM) are used in the manufacture of plastic, we hypothesized that buffaloes with PW impaction would have suboptimal rumen function and higher content of HM in the rumen fluid, sera and body tissues. Buffaloes at an abattoir were categorized into impacted with PW or not (Group C and B, respectively; n = 30/group). From the animals of group C and B, rumen fluid and sera were collected ante-mortem, while muscle, liver and kidney were collected after exsanguination. Blood and rumen fluid of stall-fed buffaloes (Group A; n = 15) served as known negative control. Rumen function was assessed by protozoal density, motility, pH, methylene blue reduction time (MBRT) and sedimentation activity time (SAT). Concentration of HM such as mercury (Hg), lead (Pb), cadmium (Cd), chromium (Cr) and copper (Cu) in the body fluids, tissues and impacted PW was estimated by atomic absorption spectrometry (AAS). A significant decrease in the rumen protozoal density, motility and increase (P < 0.05) in the rumen fluid pH, MBRT and SAT were observed in group C as compared to group A and B. Concentration of Pb, Hg and Cd showed a moderate to strong significant negative correlation with rumen function indicators. Quantitatively, Cu content (ppm) was highest in PW with 0.9 ± 0.04. The concentration of HM in the body fluids and tissues was significantly (P < 0.05) elevated in group C, but was below the toxic levels. In stall-fed buffaloes, Hg, Pb and Cd were undetectable in body fluids. Collectively, the results indicate that long term exposure of buffaloes to HM from PW affect the health, productivity and increase the risk of entering the food chain.

Serum kisspeptin: New possible biomarker for sexual behaviour and sperm concentration in buffalo bulls.

The study was designed to decipher the inter-relationship between peripheral hormones (kisspeptin and testosterone), sexual behaviour and seminal variables of Murrah buffalo bulls (n = 134). In this study, we recorded that 13%, 37%, 40%, 6% and 4% Murrah buffalo bulls had reaction time of <30, 31-60, 61-180, 181-300 and >300 s, respectively. Further, it was observed that 4%, 85% and 10% buffalo bulls were sexually aggressive, active and dull, respectively, during semen collection. The courtship behaviour was not found to be desirable for the bulls used for the semen collection. Mean of ejaculate volume, sperm concentration and mass motility (0-5 scale) were 3.57 ml, 977.11 million/ml, 2.7, respectively. Correlation studies revealed that the reaction time was positively correlated with courtship behaviour and body weight, and negatively correlated with sexual aggressiveness and sperm concentration. Serum kisspeptin in buffalo bulls, measured for the first time, was found to 3.8 ± 0.7 ng/ml. Serum kisspeptin and testosterone level are negatively correlated to each other and kisspeptin level influenced the sexual behaviour (reaction time, sexual aggressiveness and penile erection) of study bulls. Serum kisspeptin was higher in the buffalo bulls with higher sperm concentration indicating its role in spermatogenesis. In conclusion, for the first time basic information related to sexual behaviour of Murrah buffalo bulls in large population along with its inter-relationship with peripheral hormones (kisspeptin and testosterone) has been documented.

Serum proteomics analysis reveals the thermal fitness of crossbred dairy buffalo to chronic heat stress.

Chronic heat stress (CHS) reduces the production efficiency of the buffalo dairy industry. Relatively low-abundance proteins with particular functions in biological processes are changed by CHS. The present study aimed to quantify the differences in low-abundance proteins of crossbred dairy buffaloes under CHS and thermal-neutral (TN) conditions. With label-free quantification, 344 low-abundance proteins were identified in serum. Of these, 17 differentially expressed low-abundance proteins with known functions were detected, and six of the differentially expressed proteins related to heat stress were validated with parallel reaction monitoring. Lipase (LPL), glutathione peroxidase 3 (GPX3), cathelicidin-2 (CATHL2), ceruloplasmin (CP), and hemoglobin subunit alpha 1 (HBA1) cooperatively played roles in the thermal fitness of dairy buffalo by decreasing heat production and increasing blood oxygen delivery. Also, dairy buffaloes may adapt to CHS and hypoxia with high levels of RBCs, HBA1 and CP to increase blood oxygen delivery capacity.

Proteomic analysis of Fasciola gigantica excretory and secretory products (FgESPs) interacting with buffalo serum of different infection periods by shotgun LC-MS/MS.

Fasciolosis, caused by Fasciola hepatica and Fasciola gigantica, is an important zoonotic disease in the world. It affects livestock, especially for sheep and cattle, causing major economic loss due to morbidity and mortality. Although the excretory and secretory products (ESPs) of F. hepatica have been relatively well studied, little is known about the interaction between the ESP and host, and the mechanism of the key proteins involved in interaction. In this study, buffaloes were infected by Fasciola gigantica, and infection serum was collected at three different periods (42dpi, 70dpi, and 98dpi). The interaction proteins were pulled down with three different period serum by Co-IP assay, respectively, and then identified by LC-MS/MS analysis. A number of proteins were identified; some of them related to the biological function of the parasite, while most of them the functions were unknown. For the annotated proteins, 13, 5, and 7 proteins were pulled down by the infected serum in 42dpi, 70dpi, and 98dpi, respectively, and 18 proteins could be detected in all three periods. Among them, 13 belong to the cathepsin family, 4 proteins related to glutathione S-transferase, and 3 proteins are calcium-binding protein; other proteins related to catalytic activity and cellular process. This study could provide new insights into the central role played by ESPs in the protection of F. gigantica from the host immune response. At the same time, our research provided material for further studies about the interaction between F. gigantica and host.

Comparative proteomic analysis of high- and low-fertile buffalo bull spermatozoa for identification of fertility-associated proteins.

The present study identified few potential proteins in the spermatozoa of buffalo bulls that can be used as an aid in fertility determination through comparative proteomics. The sperm proteome of high-fertile buffalo bulls was compared with that of low-fertile buffalo bulls using two-dimensional difference gel electrophoresis (2D-DIGE), and the differentially expressed proteins were identified through mass spectrometric method. The protein interaction network and the functional bioinformatics analysis of differentially expressed proteins were also carried out. In the spermatozoa of high-fertile bulls, 10 proteins were found overexpressed and 15 proteins were underexpressed at the level of twofold or more (p ≤ 0.05). The proteins overexpressed in high-fertile spermatozoa were PDZD8, GTF2F2, ZNF397, KIZ, LOH12CR1, ACRBP, PRSS37, CYP11B2, F13A1 and SPO11, whereas those overexpressed in low-fertile spermatozoa were MT1A, ATP5F1, CS, TCRB, PRODH2, HARS, IDH3A, SRPK3, Uncharacterized protein C9orf9 homolog isoform X4, TUBB2B, GPR4, PMP2, CTSL1, TPPP2 and EGFL6. The differential expression ranged from 2.0- to 6.1-fold between the two groups, where CYP11B2 was high abundant in high-fertile spermatozoa and MT1A was highly abundant in low-fertile spermatozoa. Most of the proteins overexpressed in low-fertile spermatozoa were related to energy metabolism and capacitation factors, pointing out the possible role of pre-mature capacitation and cryo-damages in reducing the fertility of cryopreserved buffalo spermatozoa.

Serum haptoglobin and protein electrophoretic fraction modifications in buffaloes (Bubalus bubalis) around calving and during early lactation.

Serum protein distribution and concentration can be affected by different physiological and pathological conditions. The aim of this study was to evaluate the changes in the concentration of serum protein fractions and haptoglobin in clinically healthy dairy buffaloes during late pregnancy and early lactation. Blood and milk samples were collected from 30 buffaloes at around 7 d before expected calving (blood only) and 7, 30 and 50 d after calving. In serum samples, the total protein, haptoglobin, albumin, α1-, α2-, ß1-, ß2-, γ-globulins, and albumin/globulin ratio (A/G) values were evaluated. In milk, fat%, protein%, lactose%, somatic cell score (SCS) were assessed, along with milk yield (MY) and daily milk production (DMP). The peripartum period significantly influenced (P < 0.005) total protein, albumin, haptoglobin, α2-, ß2- and γ-globulins (P < 0.005). Milk yield, DMP and fat% changed significantly throughout the monitoring period (P < 0.005). Milk yield and DMP were positively correlated with total protein, albumin, ß2-globulins and A/G ratio, and negatively correlated with haptoglobin and α2-globulins. These results provide new knowledge about the serum protein electrophoretic pattern in Italian Mediterranean Buffaloes during the last phase of pregnancy and early stages of lactation.

Immune status and haemato-biochemical profile of buffalo calves supplemented with phytogenic feed additives rich in tannins, saponins and essential oils.

The present study was performed to ascertain the synergistic effects of phytogenic feed additives (PFA-7) supplementation on immune status and haemato-biochemical profile of buffalo calves. The PFA-7 is a mixture of neem seed cake (Azadirachta indica), mahua seed cake (Madhuca longifolia), fennel seed (Foeniculum vulgare), harad (Terminalia chebula), fruit pulp of bahera (Terminalia bellirica), fruit pulp of amla (Phyllanthus emblica) and ajwain seed (Trachyspermum ammi) mixed in 2:2:2:1:1:1:1 proportion. Male buffalo calves (n = 21) having similar age and body weight were allotted to three groups in a completely randomised design. The dietary treatments were viz. T1: control (without PFA-7) and T2 and T3: provided with PFA-7 at 2 and 4% of dry matter intake (DMI), respectively, plus sodium sulphate at 0.06% of DMI. The feeding trial was carried out for 4 months, and serum isolation was done on days 0, 60 and 120 post-feeding. The concentrations of total protein, albumin, globulin, aspartate transaminase and alanine transaminase increased, whereas cortisol and glucose decreased in the supplemented groups as compared to the control. The levels of triglycerides, urea, albumin/globulin ratio, calcium, phosphorus and alkaline phosphatase were not affected by the supplementation of PFA-7. Both cell-mediated and humoral immune response increased in the supplemented groups. The results revealed that PFA-7 positively impacted haemato-biochemical profile and both cellular and humoral immunity of the growing calves. The PFA-7 can be used as an alternative for chemical feed additives in the diet of growing calves.

Responses of pro-inflammatory cytokines, acute phase proteins and cytological analysis in serum and cerebrospinal fluid during haemorrhagic septicaemia infection in buffaloes.

Sudden death is usually the main finding in field animals during haemorrhagic septicaemia outbreaks caused by Pasteurella multocida type B:2 that causes acute, fatal and septicaemic disease in cattle and buffaloes. This situation may be due to failure in early detection of the disease where early treatment of antibiotics may improve the prognosis of the animal and other surviving animals. Thus, there is a grey area on the knowledge on the potential usage of pro-inflammatory cytokines and acute phase proteins as early biomarkers in the diagnosis of haemorrhagic septicaemia. In addition, exploration of the cerebrospinal fluid during infection has never been studied before. Therefore, this study was designed to fill up the grey areas in haemorrhagic septicaemia research. Twenty-one buffalo calves were divided into seven treatment groups where group 1 was inoculated orally with 10 mL of sterile phosphate-buffered saline pH 7 which act as a negative control group. Groups 2 and 3 were inoculated orally and subcutaneously with 10 mL of 1012 colony-forming unit of P. multocida type B:2. Group 4 and 5 buffaloes were inoculated orally and intravenously with 10 mL of lipopolysaccharide broth. Groups 6 and 7 were administered orally and subcutaneously with 10 mL of outer membrane protein broth. During the post-infection period of 21 days, blood and cerebrospinal fluid were sampled for the analyses of pro-inflammatory cytokines, acute phase proteins and cytological examination. Buffalo calves infected with P. multocida and its immunogens via different routes of inoculation showed significant changes (p < 0.05) of pro-inflammatory cytokines, acute phase proteins and cytological changes in both the serum and cerebrospinal fluid. Buffalo calves from groups 3 and 7 showed the highest pro-inflammatory cytokines, whereas group 6 had the highest acute phase protein concentration and group 5 revealed the highest value for cytology changes. In summary, results obtained in this study could be used as a profiling study to add novel knowledge to the haemorrhagic septicaemia research as well as the development of biomarkers.

Association of CYP19A1 gene polymorphisms with anoestrus in water buffaloes.

Cytochrome P450 aromatase (encoded by the CYP19A1 gene) regulates oestrogen biosynthesis and so plays an essential role in female fertility. We investigated the genetic association of CYP19A1 with the risk of anoestrus in Egyptian water buffaloes. A total of 651 animals (326 anoestrous and 325 cycling) were used in this case-control study. Using single-strand conformation polymorphisms and sequencing, four single nucleotide polymorphisms (SNPs) were detected; c.-135T>C SNP in the 5'UTR and three non-synonymous SNPs: c.559G>A (p. V187M) in Exon 5, c.1285C>T (p. P429S) and c.1394A>G (p. D465G) in Exon 10. Individual SNP-anoestrus association analyses revealed that genotypes (CC, AA and GG) and alleles (C, A and G) of the -135T>C, c.559G>A and c.1394A>G SNPs respectively were high risk for anoestrus. A further analysis confirmed that these three SNPs were in linkage disequilibrium. Additionally, haplotypes with two (TAG/122 and CAA/221) or three (CAG/222) risk alleles were significantly associated with susceptibility to anoestrus, lower blood levels of both oestradiol and antioxidant enzymes (superoxide dismutase, glutathione peroxidase (GPX) and catalase) and downregulated expression levels of CYP19A1, oestrogen receptor α and Gpx3 in the ovary, as well as increased serum level of malondialdehyde. This suggests the occurrence of a high incidence of oxidative ovarian damage and subsequently ovarian inactivity in buffaloes carrying risk alleles. Therefore, with this study we suggest the selection of buffaloes with protective alleles at these SNPs to improve the reproductive efficiency of the herd.

Correlation of two radioimmunoassay systems for measuring plasma pregnancy-associated glycoproteins concentrations during early pregnancy and postpartum periods in water buffalo.

This is the first time that PAG determination using two different antisera raised against PAG molecules purified from both caprine (RIA-706) and bubaline placentas (RIA-860) is reported in water buffalo. Ninety-eight buffalo cows, belonging to a buffalo herd subjected to a synchronization and artificial insemination (AI) programme, were enrolled in this study. Blood samples were taken on days 0 (AI), 23, 25, 28, 30 and 45. Pregnancy was confirmed by ultrasonography on days 28 and 45. The blood of 20 buffaloes that had calved was tested every five days from the day of calving until day 50 postcalving. Differences in PAG concentrations were observed between pregnant and nonpregnant buffaloes starting from day 23 post AI using both RIA-706 and RIA-860 (p < 0.001). However, estimated mean concentrations of PAG measured by RIA-706 were higher than RIA-860 (p < 0.001) and Bland-Altman analysis showed biases ranged from 0.0 ng/ml at day 23 to 0.79 ng/ml at day 28 post AI. Moreover, RIA-706 showed greater sensitivity and accuracy both at 23 and 25 days of pregnancy. RIA-706 and RIA-860 decreased below 1 ng/ml from 40 and 30 days postpartum, respectively, suggesting that PAG are better recognized by the antisera raised against the caprine PAG in the postpartum period also. This is essential when using PAG as an appropriate marker of early pregnancy after postpartum for detecting new pregnancies. The results of this study show that the ability of RIA systems to recognize early PAG could be improved using antisera raised against PAG molecules isolated from caprine placenta.

Effect of season and breed on physiological and blood parameters in buffaloes.

In this Research Communication we describe the effect of temperature and humidity index (THI) on various physiological traits, the plasma heat shock protein 70 (HSP70), heat shock protein 90 (HSP90) and cortisol levels and other blood parameters in crossbred buffalo (Nili-Ravi × Murrah) and Mediterranean buffalo to compare their tolerance to heat stress. As expected, crossbred buffalo had a significantly higher rectal temperature (RT), body surface temperature (BT), respiratory rate (RR), HSP70 and HSP90 levels in summer compared to spring and winter. RT and BT were also significantly higher in spring compared to winter. A significant correlation existed between THI and RT (r = 0·81) and RR (r = 0·84). Importantly, in summer the crossbred buffalo had a significantly lower RT, BT and RR and higher HSP70, HSP90 and cortisol levels than the Mediterranean buffalo. In conclusion, higher THI was associated with significant increase in RT, RR, BT, HSP70, HSP90 and cortisol levels, and the crossbred buffalo were more heat tolerant than Mediterranean buffalo.

Efficacy of different methods for detecting the onset of ovarian cyclicity in post-partum Murrah buffaloes.

This study was designed to investigate the sensitivity and specificity of three methods for detecting the onset of cyclicity in post-partum Murrah buffaloes. The methods investigated were visual signs, transrectal ultrasonography, and serum progesterone (P4) assay. For this study, 102 post-partum Murrah buffalo cows were grouped for monitoring their ovarian activity. The first group of buffaloes was between 26 and 35 days post-partum. Thereafter, the buffalo cows that calved were grouped after every 10 days for the study sample. Thus, the study animals were adjudged between 26 and 35, 36-45, 46-55, 56-65, 66-75, 76-85, and 86-95 days post-partum with an average of 30, 40, 50, 60, 70, 80, and 90 days post-partum, respectively. Visual estrus signs were monitored twice daily, and simultaneously, ultrasound examination was carried out at 10 days interval for accessing the presence of corpus luteum (CL). Serum P4 was estimated in the animals which were adjudged cyclic by ultrasound examination, and the assay was repeated after 10 days. The buffalo cows in estrus were inseminated artificially, and pregnancy status was assessed after 30 days post-insemination. In this study, the sensitivity and specificity of visual observation were low (39.37 and 70.73%, respectively) when compared to P4 assay (98.80 and 96.47%) and ultrasound examination (single, 97.59 and 97.59%; double, 100 and 100%), respectively. Furthermore, the sensitivity and specificity of single and double ultrasound examination and P4 assay were comparable. In conclusion, this study reports that single and double ultrasound examination and P4 assay are more efficient than visual observation in detecting the onset of ovarian cyclicity in post-partum Murrah buffaloes.

Effects of increasing dietary protein supplies on milk yield, milk composition, and nitrogen use efficiency in lactating buffalo.

Nitrogen efficiency of lactating buffalo can be increased by providing dietary crude protein (CP) precisely to the requirement. Twelve lactating Nili-Ravi buffaloes (6 primiparous and 6 multiparous) at 76 ± 37.5 days in milk (DIM) were used in this study. The treatments were diets providing three levels of CP (% DM basis): (1) low-protein = 11%; (2) medium-protein = 13.1%; (3) high-protein = 14.2% according to a 3 × 3 Latin square design. The period length of each treatment was 21 days and the total duration of experiment was 63 days. The diets were designed to provide similar energy. The nitrogen intake of buffalo increased linearly by increasing CP levels. Dry matter intake showed a tendency toward decrease in quadratic fashion, whereas milk yield decreased linearly in high-protein diet. No effect was observed on milk protein yield and content. Increasing the dietary CP levels increased plasma urea nitrogen, whereas glucose and triacylglycerol levels remain unaffected. Efficiency of dietary nitrogen utilization to milk averaged 21% and showed both linear and quadratic decreases by increasing the protein supply levels. In conclusion, low CP level showed higher milk production with low plasma urea nitrogen and high nitrogen efficiency in this experiment.

Reactive oxygen metabolites in alpha-herpesvirus-seropositive Mediterranean buffaloes (Bubalus bubalis): a preliminary study.

In the present study on Bubalus bubalis of the Campania Region (Italy) the serum levels of derivatives of reactive oxygen metabolites (d-ROMs), anti-ROM and oxidative stress index (Osi) were evaluated. These data were then related to the seropositive status of the animals against alpha-herpesviruses, precisely Bubaline herpesvirus 1 (BuHV-1) and Bovine herpesvirus 1 (BoHV-1). Clinically healthy Mediterranean buffaloes were selected for this study. The serum samples of these animals were taken, and d-ROMs, anti-ROM and Osi were measured using commercially available tests. The preliminary data demonstrated that animals seropositive to both BuHV-1 and BoHV-1 present more oxidative stress than seronegative animals, as revealed by a significant increase in d-ROMs. Our results provide, for the first time, insight into the reac- tive oxygen species (ROS) modulation induced by the herpesvirus in Bubalus bubalis.

HSFY and ZNF280BY show copy number variations within 17 water buffalo populations.

Recent transcriptomic analysis of the bovine Y chromosome revealed abundant presence of multi-copy protein coding gene families on the male-specific region of the Y chromosome (MSY). Copy number variations (CNVs) of several MSY genes are closely related to semen quality and male reproduction in cattle. However, the CNVs of MSY genes in water buffalo are largely unknown. Therefore, this study aimed to investigate the CNVs of HSFY and ZNF280BY of 298 buffaloes from 17 populations distributed in China, Vietnam and Laos using quantitative PCR. Our results revealed that the median copy numbers of the HSFY and ZNF280BY genes were 47 (ranging from 20 to 145) and 269 (ranging from 73 to 974) respectively. In conclusion, this study indicated that HSFY and ZNF280BY showed abundant CNVs within swamp buffalo populations.

Disposition of a long-acting oxytetracycline formulation in Thai swamp buffaloes (Bubalus bubalis).

The present study aimed to characterize the pharmacokinetic profile of oxytetracycline long-acting formulation (OTC-LA) in Thai swamp buffaloes, Bubalus bubalis, following single intramuscular administration at two dosages of 20 and 30 mg/kg body weight (b.w.). Blood samples were collected at assigned times up to 504 h. The plasma concentrations of OTC were measured by high-performance liquid chromatography (HPLC). The concentrations of OTC in the plasma were determined up to 264 h and 432 h after i.m. administration at doses of 20 and 30 mg/kg b.w., respectively. The Cmax values of OTC were 12.11 ± 1.87 µg/mL and 12.27 ± 1.92 µg/mL at doses of 20 and 30 mg/kg, respectively. The AUClast values increased in a dose-dependent fashion. The half-life values were 52.00 ± 14.26 h and 66.80 ± 10.91 h at doses of 20 and 30 mg/kg b.w, respectively. Based on the pharmacokinetic data and PK-PD index (T > MIC), i.m. administration of OTC at a dose of 30 mg/kg b.w once per week might be appropriate for the treatment of susceptible bacterial infection in Thai swamp buffaloes.