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IntroductionEight microsatellite loci were used to define genetic diversity among five native water buffalo breeds in Pakistan.Material and MethodsBlood samples (10 mL) from 25 buffaloes of each of the Nili, Ravi, Nili-Ravi, Kundhi, and Azi-Kheli breeds were collected aseptically from the jugular vein into 50 ml Falcon tubes containing 200 µl of 0.5 M EDTA. The phenol-chloroform method was used to extract DNA and the regions were amplified for microsatellite analysis. The eight microsatellite markers ETH10, INRA005, ILSTS029, ILSTS033, ILSTS049, ILSTS052, ETH225, and CSSM66 were analysed.ResultsThe effective number of alleles across all loci was as usual lower than the observed values with a mean value of 2.52 alleles per locus. The overall allele frequency varied from 0.0041 for alleles B, I, and J over respective loci ILSTS052, INRA005, and ILSTS029 to 0.80 for allele H over locus ILSTS029. The average observed and expected heterozygosity values across all polymorphic loci in all studied buffalo breeds were 0.43 and 0.53, respectively. The overall value for polymorphic information content of considered microsatellite markers was 0.53, suggesting their appropriateness for genetic diversity analysis in buffalo. The mean Fis value was 0.13 and all loci except ILSTS049 were found significantly deviated from HWE, most likely due to non-random breeding. The five buffalo populations were genetically less diverse as indicated by a small mean Fst value (0.07). The average gene flow (Nm) indicative for population migration was calculated as 3.31. Nei’s original measures of genetic distance (Ds) revealed ancient divergence of the Nili and Azi-Kheli breeds (Ds = 0.1747) and recent divergence of the Nili and Ravi breeds (Ds = 0.0374).ConclusionThese estimates of genetic diversity were seen to coincide with phenotypic differentiation among the studied buffalo breeds. The present study reports the first microsatellite marker-based genetic diversity analysis in Pakistani buffalo breeds, and might facilitate similar studies in other livestock breeds of Pakistan.

The present study has been designed to devise a pen-side hematological formula for estimation of hemoglobin (Hb) from Packed Cell Volume (PCV) in Cholistani breed of cattle being reared under pastoralism in Cholistan desert, Pakistan. It also aims to validate the soundness of rule of calculating Hb concentration as one-third of PCV and vice versa as being used in human medicine. Cholistani cattle (n = 364) were bled for PCV determination through microhematocrit method and Hb estimation through validated human hematology analyzer (HbD) as well as through calculation being one-third of PCV (HbC). Means (± SE) and 95% CI were computed using prescribed formulae. Difference between HbC and HbD and between HbD and corrected Hb (CHb) for all groups was computed through independent t-test.Polynomial regression analyses were carried out, scatter-plots were drawn and regression prediction equations were accordingly computed. Attaining a highest adjusted r-square value from these equations, corrected Hb (CHb) was calculated. Signi cantly (P ≤ 0.01) higher positive correlation coe cient was noticed for female adult stock (r = 0.893; adjusted r-square = 0.79) between HbD and PCV, and between HbD and HbC. The regression equation hence attained i.e. Hb = 0.27 (PCV) + 1.9 is considered valid for deducing Hb from PCV in female adult stock. For other groups, regression prediction equation [− 0.673 + 0.6433 (PCV) − 0.008136 (PCV 2 ) + 0.00031 (PCV 3 )] attained from the cubic regression having highest adjusted r-square value (0.59) is recommended for computing Hb concentration from PCV. The present study may assist to give information that can be applicable for the eldwork for the clinical diagnosis of anemia in livestock in general, and in Cholistani cattle in speci c.

The levels of expression of surface molecules and release of cytokines and chemokines of human monocyte-derived dendritic cells were determined after their exposure to adult H. contortus excretory/secretory (ES) products or a combination of ES products and bacterial lipopolysaccharide (LPS). Worm products provoked a weak response and only partial maturation of the dendritic cells, consistent with the hyporesponsiveness and more tolerogenic immune environment present in parasitized animals and humans. Co-stimulation with LPS demonstrated that H. contortus secretions, like those of other helminths, contain immunomodulators capable of reducing some aspects of the strong T(H)1/T(H)2 response evoked by bacterial LPS. There were significant reductions in the release of some cytokine/chemokines by LPS-stimulated mdDCs and a trend (although not significant at P < 0.05) for reduced expression levels of CD40, CD80 and HLA-DR. A prominent feature was the variability in responses of dendritic cells from the four donors, even on different days in repeat experiments, suggesting that generalized conclusions may be difficult to make, except in genetically related animals. Such observations may therefore be applicable only to restricted populations. In addition, previous exposure to parasites in a target population for immunomodulatory therapy may be an important factor in assessing the likelihood of adverse reactions or failures in the treatment to worm therapy.

Since the first report of infectious bursal disease in Pakistan in 1987, outbreaks have been common even in vaccinated flocks. Despite appropriate administration of vaccines, concerns arise if the circulating strains are different from the ones used in the vaccine. Here, we sequenced the hypervariable region (HVR) of the VP2 gene of circulating strains of infectious bursal disease virus (IBDV) originating from outbreaks (n = 4) in broiler flocks in Pakistan. Nucleotide sequencing followed by phylogeny and deduced amino acid sequence analysis showed the circulating strains to be very virulent (vv) and identified characteristic residues at position 222 (A), 242 (I), 256 (I), 294 (I) and 299 (S). In addition, a substitution at positions 221 (Q→H) was found to be exclusive to Pakistani strains in our analysis, although a larger dataset is required to confirm this finding. Compared to vaccine strains that are commonly used in Pakistan, substitution mutations were found at key amino acid positions in VP2 that may be responsible for potential changes in neutralization epitopes and vaccine failure.