Milk microbiota of Holstein Friesian cattle reared in Lahore: Association with mastitis.

The dairy industry is reshaping itself and becoming commercialized in Pakistan due to the increased demand for milk to overcome the shortage. Exotic breeds such as Holstein Friesian, a high milk producing breed have started being reared more on farms in Pakistan. Along with other issues, mastitis does affects the milk production of this breed. The objective of this study was to evaluate the milk composition in terms of bacterial communities in Holstein Friesian reared in Punjab, Pakistan and alteration in microbial composition with healthy and mastitic udder. Milk samples (n = 36) from farms rearing Holstein Friesian were collected. Among these samples, 05 samples from each three groups, HHF(healthy), CHF (clinical mastitis) and SHF (subclinical mastitis), based on their udder health condition, were processed using the 16 S r=RNA gene based technique. Diversity assessment as carried out by alpha diversity indices showed that milk samples from the udder infected with clinical mastitis were the least diverse and those from the healthy udder were more diverse. Beta diversity across samples showed a scattered pattern suggesting overlap amongst bacterial communities across different groups samples as depicted by PCA plots of beta diversity indices. The taxonomic profile revealed that Proteobacteria Firmicutes, Bacteroidota and Actinobacteriota were the major phyla detected across all groups. Proteobacteria dominated the HHF and SHF group while abundance of Firmicutes was higher in CHF group. Differences at other levels including order, genus and species were also recorded. The overall picture concludes that diverse microbiota is associated with different udder health conditions.

Investigation of milk microbiota of healthy and mastitic Sahiwal cattle.

BACKGROUND: Sahiwal cattle is an indigenous cattle breed of Pakistan and mastitis is one of the major problems faced by Sahiwal cattle which hinders its production potential. The study was designed to investigate the milk microbiota of healthy and mastitic Sahiwal cattle as part of a multistep project to develop probiotics for the mitigation and control of mastitis. Milk samples of Sahiwal cattle (healthy clinical mastitis and subclinical mastitis) reared under similar husbandry and management practices were processed for 16S rRNA gene base metagenomics analysis. RESULTS: Results revealed that Proteobacteria were dominant in the healthy group and subclinical mastitis group (56.48% and 48.77%, respectively) as compared to the clinical mastitis group (2.68%). In contrast, Firmicutes were abundant in the clinical mastitis group (64%) as compared to the healthy and subclinical mastitis groups (15.87% and 38.98%, respectively). Dominant species assigned in the healthy group were Ignavibacterium album, Novosphingobium capsulatum, Akkermansia muciniphila and Lactobacillus fermentum.The clinical mastitis group was dominated by Streptococcus dysgalactiae and Corynebacterium bovis, while subclinical mastitis group included Lactobacillus fermentum and uncultured acidobacteriales and Akkermansia muciniphila as dominant species. Alpha diversity indices showed higher microbial diversity in the healthy group compared to the clinical and sub-clinical mastitis groups. CONCLUSION: It is concluded that the milk microbiota of healthy sahiwal cattle has higher diversity and dominant taxa in the different groups may be used as signature microbes for mastitis susceptibility. Akkermansia muciniphila is one of candidate specie that was identified and may be used for development of probiotics.

Exploring the Milk Microbiota of Healthy and Mastitic Nili Ravi Buffalo Using 16S rRNA Gene Base Metagenomic Analysis.

The Nili Ravi, a buffalo breed from Pakistan, significantly contributes to the dairy industry. Mastitis is one of the major challenges affecting milk production in this breed. The objective of the current study was to identify the bacterial communities and diversity in healthy and mastitic milk of this breed. Milk samples (n = 14) were collected from Nili Ravi buffaloes with different udder health statuses, i.e., healthy (5), subclinical mastitis (4), and clinical mastitis (5). The DNAs were extracted, subjected to partial amplification of 16S rDNA (V3 and V4 regions), and sequenced using the Illumina platform. The results revealed variations in the bacterial communities in the milk of animals with different udder health statuses. Proteobacteria was the predominant phylum in the healthy group, while clinical and subclinical mastitis milk had a higher abundance of Firmicutes. Dominant bacterial genera in the healthy group were Streptococcus (11.60%), Herbaspirillum (7.65%), and Staphylococcus (4.70%), whereas the clinical mastitis group was dominated by Streptococcus (33.96%), Staphylococcus (7.87%), and Corynebacterium (2.68%), and the subclinical mastitis group was dominated by Bacillus (15.70%), Corynebacterium (6.70%), and Staphylococcus (6.58%). Assignment of operational taxonomic units at the species level resulted in most species being assigned to uncultured or unknown bacteria or remaining unassigned. Alpha diversity indices indicated lower microbial diversity in the clinical mastitis group, while beta diversity indices showed a scattered pattern of sample clustering in PCA plots among different groups. It is concluded that bacterial diversity in the milk of Nili Ravi buffaloes suffering from clinical mastitis is lower compared to healthy and subclinical mastitis cases. It is concluded that the variations in the microbiota of healthy and mastitic milk may be further investigated and exploited as signature microbes associated with the udder health status of Nili Ravi buffalo.

The addition of resveratrol in tris citric acid extender ameliorates post-thaw quality parameters, antioxidant enzymes levels, and fertilizing capability of buffalo (Bubalus bubalis) bull spermatozoa.

Resveratrol is a natural grape-derived polyphenol with potent antioxidant properties that protect spermatozoa against lipid peroxidation (LPO) by eradicating free radicals. The objectives of this study were to 1) appraise the effects of resveratrol in extender on post-thaw quality parameters, antioxidant enzymes, adenosine triphosphate (ATP), DNA fragmentation, LPO and 2) fertilizing capability of buffalo bull spermatozoa. Semen was collected from four fertility proven bulls with artificial vagina thrice, evaluated initially, and diluted in five different extenders containing resveratrol (T4 = 100 µM, T3 = 50 µM, T2 = 20 µM, T1 = 10 µM), and control (no resveratrol), and evaluated after post-dilution and post-thawing stage of cryopreservation. Analysis of variance revealed that, there was no difference (P > 0.05) in any parameters due to treatments at post-dilution. However, at post-thawing, sperm progressive motility (%), plasma membrane integrity (%), mitochondrial membrane potential (%) and ATP levels (nmol/106) were found higher in semen samples cryopreserved in T3 and 4 than control. Sperm supravital plasma membrane integrity (%) and viable/acrosome integrity were found higher in semen samples cryopreserved in T4 than control and T1. Furthermore, sperm catalase (U/mL), glutathione peroxidase (µM) and superoxide dismutase (U/mL) concentrations were found significantly higher in resveratrol treated groups as compared to the control. Conversely, DNA fragmentation (%) and LPO (µM/mL) were significantly (P > 0.05) decreased in semen samples cryopreserved in T4 in comparison to the control. Fertilizing capability was found higher in semen samples cryopreserved in T4 as compared to the control (%, 77.33 vs. 57.41, P < 0.05). It is concluded that the addition of resveratrol in extender ameliorates quality parameters, antioxidant enzymes levels and fertilizing capability, and alleviate DNA fragmentation and LPO in buffalo spermatozoa during cryopreservation.

Stimulating effects of Quercetin (QUE) in tris citric acid extender on post thaw quality and in vivo fertility of buffalo (Bubalus bubalis) bull spermatozoa.

Aim of this study was to evaluate the stimulating effects of quercetin (QUE) on post thaw quality and in vivo fertility of buffalo (Bubalus bubalis) bull spermatozoa. QUE is a well-known flavonoid having antioxidant properties to scavenge free radicals. Semen was collected from three buffalo bull through artificial vagina (42 °C) and were evaluated initially for volume, motility, concentration, followed by dilution in five extenders (n = 5 aliquots, control, C = no QUE; treatment 1, T1 = 50 µM QUE; treatment 2, T2 = 100 µM QUE; treatment 3, T3 = 150 µM QUE and treatment 4, T4 = 200 µM QUE supplementation). The experiment was repeated thrice (n = 3 replicates). At post dilution, sperm progressive motility (PM, %) plasma membrane integrity (PMI, %), supra vital plasma membrane integrity (SVPMI, %) and acrosome integrity (ACR-1, %) of buffalo bull were significantly higher (P < 0.05) in extenders supplemented with QUE than C. At thawing, sperm PM of buffalo bull was higher in T3 and T4 than C, T1 and T2. Sperm SVPMI was significantly higher in T2, T3 and T4 than C and T1. Sperm ACR-I was higher in T3 and T4 than C, T1 and T2. Sperm DNA integrity was higher in T4 than C. QUE supplementation increased in vitro semen quality during 30 and 60 min of incubation at 37 °C than C. The in vivo fertility was higher in T3 and T4 than C (%, 61.82 vs. 65.22 vs. 46.90). It is concluded that QUE @ 150 and 200 µM improved post thaw semen quality and in vivo fertility of buffalo bull.

Prevalence of Hepatitis C genotypes in District Swabi, Khyber Pakhtunkhwa.

The present study was conducted to determine the prevalence and genotype of Hepatitis C in District Swabi. Blood samples were collected from 100 seropositive patients from various hospitals of District Swabi, out of which, 93 samples were found positive by qualitative analysis through PCR. Positive samples were genotyped using Nested PCR. It was observed from the analysis of positive samples that 1a, 1b, 2a, 3a, 3b along with mixed genotypes are prevalent in Swabi. The most prevalent genotype was 3a with the rate of 73.13% followed by 3b with 11.82% rate and 1b to be the least common genotype at rate of 1.10%. From the present study it was concluded that HCV genotypes 1a, 1b, 3a and 3b are distributed in various parts of Swabi and genotype 3a is the most frequent genotype.