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Systems to record the frequency of animal health events in Pakistan are limited. A participatory approach was used to address gaps in farmers' knowledge and understanding of bovine health and production issues in five agroecological zones (AEZs) of Pakistan. Participatory tools, including simple ranking, pairwise ranking, constraint impact scoring, and constraint profiling were used in group discussions with farmers and animal health professionals (AHPs) in six districts of two provinces, Punjab and Sindh. The results of the ranking activities showed that foot-and-mouth disease (FMD), clinical mastitis, ticks, hemorrhagic septicemia, reproductive disorders, blackleg, and endoparasites were the most important bovine health and production constraints for small-scale dairy farmers. Constraint impact scoring showed that the participants perceived that: (1) milk production was severely affected by FMD and mastitis; (2) blackleg and parasitism led to poor growth rates and reduced meat production; (3) reproductive disorders and mastitis caused major economic losses (due to the high cost of treatment); and (4) blackleg and hemorrhagic septicemia were the leading causes of mortality in cattle and buffaloes. Although there was strong agreement in responses and constraint impact scores between farmers and AHPs, farmers were more concerned about health issues that cause high mortalities, whereas AHPs emphasized the importance of disorders with a high economic impact. Despite socioeconomic differences among AEZs, farmers' knowledge about bovine health and production constraints was similar. The findings from this study revealed that farmers had limited understanding of the risk factors and routes of transmission of various infectious diseases of bovines, which emphasizes the need to develop and implement tailored extension programs in Pakistan to control contagious diseases of animals and to improve the profitability of small-scale dairy farmers., (Copyright © 2020 Ghafar, McGill, Stevenson, Badar, Kumbher, Warriach, Gasser and Jabbar.)

A cross-sectional study was conducted to estimate the seroprevalence of bovine brucellosis in smallholder farms in seven regions in Pakistan, identify herd and individual level risk factors for seropositivity and assess the level of engagement of farmers with risk factors. In total, 1063 cattle and buffalo belonging to 420 herds in seven districts were sampled. The Rose Bengal test (RBT), indirect enzyme-linked immunosorbent assay (I-ELISA) and competitive enzyme-linked immunosorbent assay (C-ELISA) were used for the serological diagnosis of bovine brucellosis on all the serum samples. The associations between herd-and animal-level risk factors and seropositivity were investigated using logistic regression analyses. In addition, herd management practice scores, created to quantify the number of management practices undertaken that pose a risk for Brucella transmission, were calculated and compared between seropositive and negative herds within each district. Overall herd and animal prevalence were estimated to be 16.2% (95% CI, 13-20%) and 8.7% (95% CI, 7.2-10.6%), respectively, across all districts sampled. Herds with a history of last trimester abortion were found to be more likely to be positive than herds without such history (OR = 2.06, 95% CI, 1.09-3.89), providing validation of our findings and identifying that clinical disease is occurring in this region. It was also identified that herds with five to eight buffalo (OR = 3.80, 95% CI, 1.69-8.49), and those with more than eight buffalo (OR = 3.81, 95% CI, 1.51-9.58) were more likely to be positive for Brucella than those with less (one to two and three to four) buffaloes present in the herd. The presence of other domestic animals at the farm and purchasing animals in last year were found to have no association with seropositivity. The findings of this study support the need for the development of targeted intervention strategies specific to the disease status of each district., (© 2019 The Authors. Veterinary Medicine and Science Published by John Wiley & Sons Ltd.)

The present study aimed to assess the extent of knowledge and understanding of brucellosis in smallholder dairy farmers and identify practices at the farm and household level that might pose a risk for humans contracting brucellosis. Between February and June 2015 a cross-sectional study was conducted among smallholder farms (n = 420) in five districts of Punjab and two districts of Sindh province. Farmers were interviewed using a questionnaire to obtain information on farmers' knowledge about brucellosis and the potential risks for contracting the disease that are present for dairy farmers and their families. Logistic regression and ordinal logistic models were used to investigate potential predictors for risky behaviours. The results show almost all farmers (97%) were not aware of the modes of transmission of brucellosis. Relating to risk, the majority (66%) of the farmers' families were reported to consume raw milk and its products, live in shared housing with animals (49%) and not cover hand cuts during contact with animals (74%). All farmers performed at least one risky practice on a regular basis for brucellosis transmission from animal to human. A multivariable analysis highlighted that the respondents with no formal education and those who had not heard of the disease displayed greater risky behaviour. Poor understanding of the disease, presence of multiple risky practices on farm and at the household, and incorrect perception supports the need for an educational awareness program in order to ensure uptake of improved practices.

The present study was conducted to observe the seasonal variation in aflatoxin M1 and nutritional quality of milk along informal marketing chains. Milk samples (485) were collected from three different chains over a period of one year. The average concentrations of aflatoxin M1 during the autumn and monsoon seasons (2.60 and 2.59 ppb) were found to be significantly higher (standard error of the difference, SED = 0.21: p = 0.003) than in the summer (1.93 ppb). The percentage of added water in milk was significantly lower (SED = 1.54: p < 0.001) in summer (18.59%) than in the monsoon season (26.39%). There was a significantly different (SED = 2.38: p < 0.001) mean percentage of water added by farmers (6.23%), small collectors (14.97%), large collectors (27.96%) and retailers (34.52%). This was reflected in changes in milk quality along the marketing chain. There was no difference ( p = 0.178) in concentration of aflatoxin M1 in milk collected from the farmers (2.12 ppb), small collectors (2.23 ppb), large collectors (2.36 ppb) and retailers (2.58 ppb). The high levels of contamination found in this study, which exceed the standards set by European Union (0.05 ppb) and USFDA (0.5 ppb), demand radical intervention by regulatory authorities and mass awareness of the consequences for consumer health and safety., Competing Interests: The authors declare no conflict of interest.

The buffalo is an important livestock resource in several countries of South Asia and the Mediterranean regions. However, reproductive efficiency is compromised due to known problems of biological and management origins, such as lack of animal selection and poor nutrition. Under optimal conditions puberty is attained at 15 to 18 months in river buffalo, 21 to 24 months in swamp buffalo and is influenced by genotype, nutrition, management and climate. However, under field conditions these values deteriorate up to a significant extant. To improve reproductive efficiency, several protocols of oestrus and ovulation synchronization have been adopted from their use in commercial cattle production. These protocols yield encouraging pregnancy rates of (30% to 50%), which are comparable to those achieved in buffaloes bred at natural oestrus. The use of sexed semen in buffalo heifers also showed promising pregnancy rates (50%) when compared with conventional non-sexed semen. Assisted reproductive technologies have been transferred and adapted to buffalo but the efficiency of these technologies are low. However, these latest technologies offer the opportunity to accelerate the genetic gain in the buffalo industry after improving the technology and reducing its cost. Most buffaloes are kept under the small holder farming system in developing countries. Hence, future research should focus on simple, adoptable and impact- oriented approaches which identify the factors determining low fertility and oestrus behaviour in this species. Furthermore, role of kisspeptin needs to be explored in buffalo.

Breeding records of 57 Arabian and 66 Thoroughbred mares were analysed to assess their reproductive performance under the subtropical conditions of Pakistan. The Arabian mares showed significantly higher conception rates (p<0.05) in second mated oestrus and foal heat mated oestrus compared to Thoroughbred mares. However, conception rates for first lifetime mated oestrus were similar in both breeds of mares. Age at first mating (1,301±40 vs 1,500±32 days) was significantly (p<0.05) less in Arabian compared to Thoroughbred mares. Both breeds of mares showed significantly (p<0.05) higher frequencies of oestrous cycles and conception rates during the winter (October to March) compared to summer (June to August) months. Age of mares affected the conception rates, as mares at ages 3 to 7 and 8 to 12 years of ages had significantly higher conception rates (p<0.05) than those ≥18 years old in both breeds. This study demonstrates that i) reproductive performance in Arabians is better than Thoroughbred mares under the subtropical conditions of Pakistan, ii) mares remain cyclic throughout the year and iii) conception rates were higher in mares bred during winter compared to summer months.

The objective of the study was to assess the post-weaning growth response of Sahiwal calves reared on four different pre-weaning dietary regimens. The four diets were: (a) whole cow's milk, starter ration (SR; CP = 20%, total digestible nutrients (TDN) = 72%) and Berseem hay (H; Egyptian clover; CP = 21%, TDN = 63%); (b) whole cow's milk + H; (c) milk replacer (MR; reconstituted to supplier specification; Sprayfo®) + SR + H; and (d) MR + H. The protein and fat percentages of reconstituted MR were 2.22 and 1.84, respectively. Milk or MR were fed at the rate of 10% of the calves' body weight (BW) until 56 days of age, and then withdrawn gradually until weaned completely by 84 days of age. The average initial BW of calves in groups A, B, C and D were 56.3 ± 1.0, 47.5 ± 1.0, 40.4 ± 1.0 and 30.3 ± 1.0 kg, respectively. Initially, there were 12 calves in each group with six of each sex; however, one male calf died from each of groups B and C and were not replaced. During the post-weaning period, 13 to 24 weeks, the calves were fed a single total mixed ration ad libitum based on maize, canola meal, wheat straw and molasses containing 16% CP and 70% TDN. Daily feed intake and weekly BW gains were recorded. The data were analyzed by MIXED model analysis procedures using the statistical program SAS. The intake of calves as percent of their BW, feed conversion ratio and cost per kg of BW gain were not different (P > 0.05) across treatments. The daily gain at 24 weeks of age for the pre-weaning treatments A, B, C and D were 746 ± 33, 660 ± 33, 654 ± 33 and 527 ± 33 g/day and the final liveweights of calves were 119 ± 4.2, 102 ± 4.2, 95 ± 4.2 and 75 ± 4.2 kg, respectively. Gains were influenced significantly (P < 0.05) by pre-weaning treatments. The calves fed MR and H only during the pre-weaning period were unable to catch up post weaning with calves on other dietary treatments. The calves fed whole milk from birth at the rate of 10% of liveweight together with concentrates had higher weaning weight and superior growth rate post weaning as well. Thus, pre-weaning feeding was important for higher weaning weights and superior growth rates post weaning.

The objective was to study the growth potential of Sahiwal calves given milk or milk replacer with or without concentrates. For this purpose, forty-eight Sahiwal calves were divided into four groups of 12 animals each with equal sex ratio. In each group, the calves were offered either milk or a milk replacer (MR) at a rate of 10% of their body weight adjusted weekly. In addition to this, calves were fed either a starter ration plus Egyptian clover hay (SR + H) or hay only (H) until the end of trial. The milk or MR was withdrawn gradually from day 56 until animals were weaned completely by day 84. Calves offered milk grew faster than those offered MR (357 ± 9 vs. 162 ± 9 g/day; p < 0.05) and displayed higher weaning weights (51.6 ± 0.8 vs. 35.2 ± 0.8 kg; p < 0.05). The calves offered SR + H grew faster (311 ± 9 vs. 208 ± 9 g/day; p < 0.05) and displayed higher weaning weights (48.7 ± 0.8 vs. 38.1 ± 0.8 kg; p < 0.05) than those fed H alone. Calves offered milk plus SR + H showed the highest growth rate and weaning weights (401 ± 13 g/day and 56.3 ± 1 kg, respectively). The lowest growth rate and weaning weights were observed in calves offered MR and H only (115 ± 13 g/day and 30.3 ± 1 kg, respectively). Calves offered the MR had higher number of scour days than those offered milk (13.5 vs. 3.3). The feeding of whole milk in combination with the starter ration and hay resulted in superior growth rates, higher weaning weights, and healthier calves than the other feeding regimens.

The objective of this study was to determine the effect of oestrous synchronization methods on oestrous behaviour, timing of ovulation and pregnancy rate during the breeding and low breeding seasons in Nili-Ravi buffaloes. In Experiment 1, oestrous behaviour and timing of ovulation were determined from (n=34) oestruses. The mean (+/- S.E.M.) time of ovulation after the onset of standing oestrus was greater (P<0.05) in PGF(2alpha)-induced luteolysis (30.6+/-1.5h) compared to Ovsynch buffaloes (15.0+/-0.8h). In Experiment 2, pregnancy rates were compared between two methods of synchronization (detected oestrus and Ovsynch protocol) during the breeding and low breeding seasons. Pregnancy rates of buffaloes bred at detected oestrus (62.5%) or by the Ovsynch protocol (36.3%) during the breeding season did not differ significantly (P>0.05) from those which were inseminated during the low breeding season (55.5%) and (30.4%), respectively. This study demonstrates clearly that (1) timing of ovulation in Nili-Ravi buffalo is about 30h after the onset of standing oestrus and (2) buffaloes can be successfully synchronized with optimum fertility using either PGF(2alpha) alone (detected oestrus) or using (Ovsynch protocol) during low breeding season, to calve during the period when milk availability is short.

The objective of this study was to characterize the follicular waves and associated ovarian events during spontaneous and PGF(2alpha)-induced oestrous cycles in Nili-Ravi buffaloes. In Exp. 1, (n=13 oestrous cycles) follicular and luteal development was monitored by ultrasonography and jugular blood samples were collected simultaneously on alternate days. Of 12 oestrous cycles, 9 (75%) had two waves of follicular activity and only 3 (25%) had three waves. The mean (+/-S.E.M.) length of the oestrous cycle was shorter (P<0.05) in buffaloes with two waves than in those with three waves (21.2+/-0.1 days versus 22.8+/-0.1 days). In Exp. 2, follicular dynamics were compared in buffaloes undergoing spontaneous (n=12 oestrous cycles) and PGF(2alpha)-induced (n=9) regression of the corpus luteum (CL). The dynamics of ovulatory follicular growth during the 3 days before oestrus were similar (P>0.05) in buffaloes undergoing spontaneous and PGF(2alpha)-induced luteolysis. These results show that (1) the majority of buffaloes had a two wave pattern of follicular growth and emergence of a third wave was associated with a longer luteal phase, and (2) follicular dynamics during the 3 days before oestrus were similar in buffaloes undergoing spontaneous and PGF(2alpha)-induced luteolysis.

This study evaluated the meiotic competence of buffalo oocytes with different layers of cumulus cells. A total of 588 oocytes were collected from 775 ovaries averaging 0.78 oocytes per ovary. Oocytes with homogenous cytoplasm (n = 441) were selected for in vitro maturation (IVM) and divided into four groups based on their cumulus morphology: a) oocytes with > or = = 3 layers of cumulus cells, b) 1-2 layers of cumulus cells and oocytes with partial remnants or no cumulus cells to be cocultured c) with or d) without cumulus cells. Oocytes in all four groups were matured in 100 microL drop of TCM-199 supplemented with 10 microg/mL follicle stimulating hormone (FSH), 10 microg/mL luteinizing hormone (LH), 1.5 microg/mL estradiol, 75 microg/mL streptomycin, 100 IU/mL penicillin, 10 mM Hepes and 10% FBS at 39 degrees C and 5% CO2 for 24 hours. After IVM, cumulus cells were removed from oocytes using 3 mg/mL hyaluronidase, fixed in 3% glutaraldehyde, stained with DAPI and evaluated for meiotic competence. The oocytes with > or = 3 layers of cumulus cells showed higher maturation rates (p<0.05: 64.5%) than oocytes with partial or no cumulus cells (8.6%) and oocytes co-cultured with cumulus cells (34.5%) but did not differ from oocytes having 1-2 layers of cumulus cells (51.4%). The degeneration rates were higher (p<0.05) for oocytes with partial or no cumulus cells (51%) than rest of the groups (range: 13.8% to 17.4%). These results suggest that buffalo oocytes with intact layers of cumulus cells show better IVM rates than oocytes without cumulus cells and the co-culture of poor quality oocytes with cumulus cells improves their meiotic competence.