Validation of Bos taurus SNPs for Milk Productivity of Sahiwal Breed (Bos indicus), Pakistan.

The aim of the present study was the validation of the already reported Bos taurus SNPs in the Sahiwal breed. A total of nine SNPs of the casein gene were studied. Out of nine, seven Bos taurus SNPs of casein protein genes were found to be significantly associated with milk productivity traits. The genomic DNA was extracted from the mammary alveolar endothelial cells of a flock of 80 purebred Sahiwal lactating dams available at Khizrabad Farm near Sargodha. New allele-specific primers were designed from the NCBI annotated sequence database of Bos taurus to obtain 100 nt-long PCR products. Each dam was tested separately for all the SNPs investigated. Animals with genotype GG for the SNPs rs43703010, rs10500451, and 110323127, respectively, exhibited high milk yield. Similarly, animals with genotype AA for the SNPs rs11079521, rs43703016, and rs43703017 showed high milk yield consistently. For the SNP rs43703015, animals with genotype CC showed high milk productivity. These above-mentioned SNPs have previously been reported to significantly up-regulate casein protein contents in Bos taurus. Our results indicated SNPs that significantly affect the milk protein contents may also significantly increase per capita milk yield. These finding suggest that the above-mentioned reported SNPs can also be used as genetic markers of milk productivity in Sahiwal cattle.

Fortification of Extender with Basella rubra Fruit Extract Enhances the Cryosurvival of Ram Semen.

This study aimed to evaluate the impact of Basella rubra fruit extract (BR-FE) on cryopreserved ram sperm's motility, velocity, and membrane integrity. Thirty ejaculates collected from 3 fertile rams (10 from each) were diluted with semen dilution extender (SDE) in a ratio (1:2) and centrifuged to remove 50% supernatant. The remaining sample was mixed with semen cryopreservation extender (SCE) in 1:4 ratio. Then 1.2 mL of SCE diluted sample was divided in four aliquots (0.3 mL each) that were further extended with [(1) control group (0.7 mL of SCE), (2) BR-FE-0.6% group (0.7 mL of SCE supplemented with 0.6% BR-FE), (3) BR-FE-0.8% group (0.7 mL of SCE supplemented with 0.8% BR-FE), and (4) BR-FE-1.6% group (0.7 mL SCE supplemented with 1.6% BR-FE)]. All extended samples were cooled gradually from 25°C to 4°C in half an hour. The 0.1 mL sample from all aliquots was analyzed for precryopreservation sperm parameters and the remaining sample was loaded in 0.5 mL plastic semen straws, cooled gradually to -20°C, and then dipped in liquid nitrogen. After 24 hours of cryopreservation, the straws were thawed for postcryopreservation sperm evaluations. The results (analysis of variance based) showed significantly enhanced percentage of post-thaw sperm membrane integrity, progressive motility, and velocity in BR-FE-0.6% group at both pre- and postcryopreservation stages as compared with all other groups. However, analysis of covariance revealed concentration-dependent cryoprotective effect of BR-FE with maximum percentage of sperm membrane integrity in the 1.6% group. According to these results, BR-FE supplementation adds enormous sperm protective potential to ram sperm cryopreservation medium.

Seasonal Variation and Association of Heavy Metals in the Vital Organs of Edible Fishes from the River Jhelum in Punjab, Pakistan.

The toxic effects of heavy metals are drastic, including accumulation. Fish species are important bio-indicators of heavy metal pollution in aquatic bodies. The current study aimed to assess the seasonal variation of heavy metals in the vital organs of mostly consumed fishes in River Jhelum, Pakistan. Samples of fish, including Wallago attu (Malhi), Rita rita (Khagga), and Mystus seenghala (Singhari), were collected from four different sites, i.e., Khushab, Muhammad Wala (M. Wala), 8.R.D and Rasool barrage during summer and winter seasons. Heavy metals such as iron (Fe), lead (Pb), chromium (Cr), cobalt (Co) and Cadmium (Cd) were estimated through acid digestion and spectrometric analysis. Results showed a significantly higher (P < 0.05) amount of these metals in the liver, followed by the kidneys of fish species. There were seasonal variations in the absorption of these metals as well. Cr (11.71) and Fe (58.66) were detected in higher amounts in Khagga which showed the greatest affinity for certain metals in some cases. In contrast, Singhari showed the greatest affinity to other metals in other cases. Comparative analysis revealed that there was a highly significant (P < 0.05) difference for the accumulation of almost all metals in both seasons and summer had the highest concentration of Cd, Pb, Co, Cr and Fe as compared to winter in all four sampling stations in the case of kidney and liver of all the three fishes. Elevated levels of heavy metals were detected in the summer due to increased temperature. Heavy metals found in the River Jhelum may demonstrate that metals can significantly affect the fish species.

In ovo exposure of F-ions and organo-fluoride insecticide (Bifenthrin) cause developmental anomalies of eye in chick embryos.

Objective: The developmental abnormalities of the in-ovo exposure of Fluoride ions (F-ions) and Bifenthrin (BF) on the embryonic chick eye were investigated. Materials and methods: 165 fresh fertilized eggs of zero day and 40-50 g weight were divided into three groups (55 eggs each) on the basis of inter-vitelline treatment of eggs on zero day of study: 1) Control group (CG); 0.1 ml of 5 % DMSO aqueous solution 2),3) Fluoride group (FG), and Bifenthrin group (BFG); 0.01 mg/kg F-ions (from NaF) and 0.01 mg/kg BF in 0.1 ml of 5 % DMSO aqueous solution respectively. After incubation for 14 days at 37 ± 0.5 °C embryos were externalized. Eyes of each embryo were removed for micro-anatomical, micrometric and histopathological studies. Results: The histological sections have shown denser and enlarged marginal mitotic region of the developing eye lenses in FG and BFG. In vertical sections of the eye lenses the nuclei of the crystalline cells in FG and BFG show a highly depressed bow shaped arrangement. Moreover, the nuclei of the core crystalline cells of the lens were apparently smaller in FG and BFG than CG. Out of the six anatomical layers of the retina the nuclear and the plexiform layers were highly enlarged in FG and BFG, similarly the three corneal cell layers (endothelial, parenchymal and epithelial) were enlarged in FG and BFG than CG. The morphometric, histometric and micrometric estimations also show significant variations in FG and BFG than CG. Conclusion: The results indicate subtle developmental anomalies of the eyes attributable to the F-ions and BF exposure indicating their developmental neuro-optic disruption potentials. Results further revealed higher toxicity of BF as compared to F-ions.

Evaluation of the anti-inflammatory, analgesic, anti-pyretic and anti-ulcerogenic potentials of synthetic indole derivatives.

A large number of new synthetic compounds are synthesized in the field of heterocyclic chemistry having a variety of biological potentials. In the present study, some synthetic indole derivatives are used to check anti-inflammatory, analgesic, antipyretic and gastroprotective activity in albino mice. Albino mice of either sex of reproductive age were used for each study (n = 5). In anti-inflammatory activity, the negative control (NC) and positive control group animals were treated with normal saline and 10 mg/kg of indomethacin respectively. The treated groups received the twenty four different synthetic chemicals, after 30 min of sub cutaneous injection of carrageenan. In analgesic activity, hot-plate method is used and for each group the latency period was recorded at zero moment of the provision of required dose and after 30, 60, 90, 120 and 180 min. In anti-pyretic activity, Pyrexia was induced by using Brewer's yeast method. Before any treatment and then after duration of 18 h, the rectal temperatures were recorded. Among all the chemicals, only those chemicals which show any potential related to above mentioned activities were selected for gastroprotective activity. The gastroprotective activity was performed to check the gastric ulcers by using 300 mg/kg of single oral dose of indomethacin to animals of all groups except NC group. This study helped to screen out the most potent indole derivatives 3a-II and 4a-II from the 24 synthetic indole derivatives which demonstrated the best biological potential (anti-inflammatory, analgesic, antipyretic, and gastroprotection) as compared to the remaining ones. The micrometric and biochemical results also support the histological findings. Out of the twenty-four novel indole amines tested, 3a-II and 4a-II have shown the effective pharmacological capacity and additionally have not shown any overt and systemic toxicity. Thus these two indole amines need further in-depth pharmacokinetic and pharmacodynamics studies before they are recommended for any pre-clinical trial.

Histo-anatomical mutilations of developing chick brain induced by in-ovo fluoride and bifenthrin exposure.

Comparative developing brain histo-anatomical pathologies of Fluoride ions and Bifenthrin in-ovo exposures were explored in the golden black variety of domestic chick. Three exposure groups were -the Vehicle control group (Vg); Fluoride (F) group and the Bifenthrin (Bn) group each with forty fertilized eggs and received their respective group treatment at zero day of incubation. Embryos were extracted, dissected from head region and the embryonic whole brains were recovered after 14 days of incubation. The embryonic brains were preserved in bouin fixative for 24 h for further studies. The morphological results show the atrophied and hypertrophied embryonic brain in F and Bn groups respectively as compared to Vg group. The toxicological signs of encephalic anatomy and histology of F and Bn exposure were the enlarged third ventricles, optocoeles and arachnoid mater, encephalic spongiosis and decreased neuroglial density. The morphometric data showed significant decrease (p ≤ 0.05) in mean weight and density of whole brain in F and Bn groups compared Vg. The mean length and width of whole brain in F were significantly lower than that of the Bn and Vg. whereas, the mean breadth of third ventricle in Bn remained significantly lower than F and Vg groups. On the other hand, the mean breadth of optocoele and fourth ventricle in F and Bn groups remained significantly higher than Vg. Conversely the mean optic lobe wall thickness in F remained significantly lower than Bn and Vg. Additionally, the mean neuronal density in diencephalon, optic lobe and cerebellum in F group and Bn group remained significantly (p ≤ 0.05) lower than Vg. Results show that low dose in-ovo fluoride or bifenthrin exposure may cause neuro-developmental abnormalities in the developing chick embryos indicating that the Fluoride-ions and Bifenthrin harbor strong developmental neurotoxic capacity.

Mitigating properties of vitamin E and olive oil on the teratomorphogenic impacts of lambda-cyhalothrin in developing chick embryos.

Embryonic toxicity of lambda-cyhalothrin (LCH) and the mitigating properties of extra virgin olive oil (EVO) and vitamin E were investigated in golden black variety of chick embryos. Fifty fertilized eggs distributed in 5 groups received their respective group treatments at zero days of incubation were recovered after 14 days of incubation and fixed in fixative for 48 h for further studies. Results showed that treatment with LCH caused embryonic death, growth retardation and developmental abnormalities such as limbic developments, reduced muscular growth, and embryonic cataract. Post treatment with vitamin E and EVO alone significantly improved all these developmental defects. Morphometric readings also showed that embryos treated with combined dose of vitamin E and EVO had more resemblance towards the control group embryo. These findings suggested that the use of vitamin E and EVO together during pregnancy have the potential to curtail these accidental, environmental and work place exposures of LCH.

Basella alba, Grewia asiatica, Solanum nigrum and Ficus carica fruit extracts ameliorate the testicular histopathology induced by CCl4 exposure in albino mice: An experimental study.

Background: Carbon tetrachloride (CCl 4 ) is extensively used in various industries and induces oxidative stress in humans. Unfortunately, it is one of the neglected potent male reproductive toxicants. Objective: The present research reports the testicular histopathology of CCl 4 and ameliorations by four medicinal fruit pulp extracts (FPEs) in mice. Materials and Methods: Sixty male albino mice were divided into six groups (10/group) as per the following: 1. Vehicle control (Vc); 2. CCl 4 (C); 3-6. CCl 4 +Basella alba (CBa), CCl 4 +Solanum nigrum (CSn), CCl 4 +Ficus carica (CFc) and CCl 4 +Grewia asiatica (CGa). Except for the control group, CCl 4 (0.1 mL of 0.2 mL kg - 1 ) was given to the animals in corn oil. The four plant extracts (0.1 mL each) were respectively given to the relevant FPE group animals for the next five consecutive days, while the animals in the Vc and CCl 4 groups received water instead of FPE. Results: The CCl 4 exposure led to various histometric and histological alterations (loss of interstitial tissue and various dislodged tailless spermatids with enlarged heads) that were recovered in all except Solanum nigrum FPE mice post-treatment. The micrometric data of testicular sections also indicated significant decline in the number of spermatogonia, while the cross-sectional area of the sperm heads remained significantly higher in the CSn and C groups. Conclusion: Except for Solanum nigrum,the three FPEs, especially Ficus carica,showed rehabilitative properties against CCl 4 exposure-related modifications in testicular histopathologies.

Compound heterozygous ASPM mutations in Pakistani MCPH families.

Autosomal recessive primary microcephaly (MCPH) is characterized by reduced head circumference (50% of all reported families. In spite of the high frequency of MCPH in Pakistan only one case of compound heterozygosity for mutations in ASPM has been reported yet. In this large MCPH study we ascertained 37 families including 319 persons (140 patients). Haplotype analysis of eight STS markers suggested linkage by homozygosity in 20 families, and re-analysis of single sib ships in the remaining families demonstrated possible compound heterozygosity in two families. Direct sequencing indeed confirmed compound heterozygosity in two and homozygous mutations in 20 families, respectively, showing that up to 10% of families with MCPH caused by ASPM are compound heterozygous. In total we identified 16 different nonsense or frameshift mutations of which 12 were novel thereby increasing the number of mutations in ASPM significantly from 35 to 47. We found no correlation between the severity of the condition and the site of truncation. We suggest that the high frequency of compound heterozygosity observed in this study is taken into consideration as part of future genetic testing and counseling in Pakistani MCPH families.