1.
In-Vetro And In-Vivo Anti-Theilerial Activity Of Medicianal Plants
by Mukhtar Ahmad | Prof. Dr. Muhammad Ashraf | Prof. Dr. Muhammad Sarwar Khan.
Material type: ; Format:
print
; Literary form:
drama
Publisher: 2013Dissertation note: In vitro study was carried out in order to estimate the anti-theileria effect of Calotropis procera and Peganum harmala. Water and chloroform extracts of each plant were used in-vitro study along with standard drug Buparvaqoune (Butalex®). For this four concentrations i.e.4, 8, 12 and 16mg/ tested solutions of each extracts of each plant were applied on cultured lymphocytes exposed to theileria parasite infection. ELISA reader findings showed that the mean OD were found less in treated theileria infected lymphocytes cell culture as compared to untreated culture wells mean OD values. The highest cell reduction (94.36%) with C. procera chloroform extract treatment was observed at a concentration of 16 mg/ml solution. Lowest concentration (1mg/ml solution) of C. procera chloroform extract reduced non-significant (P>0.05) lymphocytes cell proliferation (40.97%) as compare to control negative group. P. harmala water extract was effective against the theileria parasite as significantly lower (P<0.05) mean OD value (1.802 ±0.341) was measured at a concentration of 4mg/ml solution and maximum inhibitory effect (92.20) was seen at a concentration of 16 mg/ml solution. ELISA reader findings showed that P. harmala chloroform extract treatment failed to inhibit lymphocytes cells propagation even at highest concentration. The highest inhibitory effect (85.33%) against theileria infected lymphocytes propagation was seen at a concentration of 16 mg/ml solution.
Plant extract was evaluated in respect of feed intake in rabbits. It showed that when administered extracts of C. procera in rabbits at dose of 3 mg and 5 mg/kg body weight, did not affect on feed intake in rabbits. However the chloroform and water extracts of both plant i.e. C. procera and P. harmala when were administered in rabbits parentally at dose 10 of mg/kg body. It showed that the feed intake of rabbits was non-significantly reduced as compared to other treatments groups. Hematological parameters such as WBC X103 count, RBC X 106 count and Hb g/dl values were measured at various days. Findings showed that significantly lesser RBC X 106 count was in group A3 and D3 than control at day 30 of experiment. A non-significant difference (P>0.05) was seen in RBC X 106 count and Hb g/dl measurements in all treatments groups. Kidney and liver functions were evaluated by measuring biochemical parameters, uric acid, creatinine and ALT at 0 days, 9 days and 30 days. Findings showed that serum creatinine and urea enzyme levels were significantly higher (P<0.05) in group A3 as compare to control group at day 30 of experiment. Serum level of urea was also significantly higher (P<0.05) in group B3 and D3 at day 30 of experiment. A non-significant difference (P>0.05) was seen in ALT in all treatment groups at day 30 than control. Post-mortem was performed at day 30 of experiment. Gross lesions consisting of hemorrhages, congestion, and lung emphysema were seen in rabbits treated with high dose i.e. 10 mg/kg of both extract of C. procera. Rabbits treated with P. harmala chloroform extract at dose 10 mg/kg showed moderate gross lesions. Histopathology of organs such as lungs, kidney, liver and heart was performed. Toxicity lesions were seen in rabbits treated with high dose i.e. 10 mg/kg of both extract of C. procera. Rabbits treated with P. harmala chloroform extract at dose 10 mg/kg showed histopathological lesions in lungs, liver and kidney.
Theileria infection was studied in vivo by developing through theileria infected Hayalomma ticks in crossbred calves (n=30) through. At day 15 of infection maximum increase in mean rectal temperature (105.24 ± 0.46F) was observed, twenty four calves had pyrexia (104.1- 105.6 F) and six claves were showing pyrexia > 105.6 F. A significant increase (P<0.05) in pre-scapular lymph node enlargement score of challenged calves was seen by day 7 of infection and maximum lymph node score (grossly enlarged size) was noticed in twenty calves (Table 4.14 , Plate 4.16) with peak mean score (2.73±0.44) on day 13 of infection. The piroplasm peak score (3.80±0.83) was observed in challenged calves at 22 day of infection and remained significantly higher (p<0.05) (2.60±0.54) in untreated calves until the 36 day of infection (Fig.4.29 and Table 4.16). A significant increase (P<0.05) in mean schizonts was observed in pre-scapular lymph node biopsy smear from day 7 of infection to onwards. Blood samples of challenged calves (n =30) were confirmed theileria positive through PCR test. The amplification of Theileria species were amplified at 1098 bp (Plate 4.20 and Theileria annulata was amplified at 721 bp (Plate 4.21).
In order to estimate the pattern of disease severity, severity score was measured by summation of mean score of piroplasms, schizonts, lymph node swelling and rectal temperature. From day 7, mild response (3-5 score) was seen in infected calves (n=10).
With increase in the severity of disease a significant decrease (P<0.05) was observed in mean values of the Hb g/dL amount, WBC and RBC count, Hct (%) concentration and lymphocytes percentage from day15 of infection onward to 36 day of infection. A non-significant decrease (P>0.05) in the mean values of MCH pg was seen throughout the experiment. A significant decrease (P<0.05) in mean values of MCHC g/dL along with significant increase (P<0.05) in the mean measurement of MCV fL (64.14±3.53) values was seen at day 36 of infection as compare to day 0 values, indicating macrocytic hypochromic anemia in challenged calves. These findings showed a significant increase (P<0.05) in excretory products (uric acid and creatinine) from day 15 of infection and onward as compared to day 0 values, indicating damaged kidney in infected calves. Biochemical analysis showed the significant increase (P<0.05) in liver enzymes (ALT, AST) from day 15 infection and onward.
Anti-Theileria activities of drugs were estimated by evaluating clinical manifestation of the disease and parasitological findings. Beside this treatment effect on hematological and biochemical reactions of liver and kidney functions was determined. A significant difference (P<0.05) in rectal temperature of calves groups (B and E) was observed than control positive (group F) at day 21 of post-treatment. On other hand calves treated with treatments A, C and D had a non-significant difference (P>0.05) in rectal temperature compared with untreated calves (group F). It was found that calves (n=5) dosed with C. procera chloroform extract (group A) had rectal temperature in normal range by the day 7 of post-treatment. Similarly calves (n=5) treated with Butalex were found with normal rectal temperature from the day 7 of pos-treatment. On other hand, at day 21 of treatment 40%, 20%, 40% and 80% calves were found with pyrexia in treatments groups B, C, D and F, respectively (Table 4. 46).
By the day 14 of treatment, calves of treatment groups B and E showed no parasitemia (piroplams ?1). Disease severity was estimated on accumulative score of rectal temperature, lymph node swelling and parasitological findings (piroplasms and schizonts score). It was found a significant decrease (P<0.05) occurred in the disease severity of score of disease in calves of groups B and E as compare to A, C and F at day 3 of post-treatment. At day 21 of treatment all treated calves were recovered from anemia.
Availability: Items available for loan: UVAS Library [Call number: 1580,T] (1).
2.
Pathology Of Experimental Enterotoxemia In Sheep And Goats
by Azam Ali Nasir | Prof. Dr. M. Yonus Rana | Dr. Muhammad | Dr. Muti-ur Rehman Khan.
Material type: ; Format:
print
; Literary form:
drama
Publisher: 2012Dissertation note: During the present study intestinal scrapings were collected from sheep suspected for enterotoxaemia. Samples were subjected for the isolation by repeated culturing in Reinforced Clostridium medium. Growth on blood agar revealed characteristic colonies of Clostridium perfringens after 18-24 hours. Biochemical and mice inoculation tests were performed. Isolated organism was identified by indirect ELISA. The pure growth was subcultured in RCM in bulk quantity and calculated the dose for experimental infection.
In the second part of the study, sheep and goats were procured, dewormed and kept in the experimental house of UVAS, Lahore. The experimental infection comprised of the whole culture of C. perfringens type D was inoculated intraduodenally via para-mid line between animals of group A and B while in animals of group C and D inoculated only starch solution to achieve the objectives. Accumulative clinical score in sheep was found to be 9 to 16, 13 to 22 and 15 to 23 at 10, 20 and 30 hours PI respectively while in goats the accumulative scores varied from 5 to 15, 9 to 16 and 14 to 21 at above mentioned time intervals. The highest mean score for clinical findings in sheep was anorexia, frothing followed by dehydration while in goats, the highest mean score was recorded for diarrhea dehydration, and anorexia. No significant clinical findings were noted in control groups.
There was a significant increase in blood glucose, urea and serum creatinine in infected group of sheep and mean values reached up to 141 mg/dl, 92 mg/dl and 7.5 mg/dl respectively at 30 hours PI while in goats a similar pattern was observed with the mean values raised to 142 mg/dl, 111 mg/dl and 10.2 mg/dl for blood glucose, urea and serum creatinine respectively. There was no significant change found in RBC and platelet count of both species but there was an early increase in the mean WBC count of sheep 19.7x103/µl at 10 hours but then decreased to 14.7x103/µl at 30 hours PI while in goats it was 23.6x103/µl and then decreased to 15.3x103/µl. The mean PCV % age increased in animals of both infected groups but more in goats and reached to 52% in 30 hours.
During the third part of the present study, the animals were slaughtered and PME performed. The accumulative score for gross lesions were recorded and it was found between 14 to 24 in animals of group A with the highest score for congestion and edema of different organs whereas in goats it was between 12 to 22 with congestion and hemorrhages of intestine having highest scores. The samples were kept in formalin for histopathological examination and accumulative lesions score was noted in different organs. The highest mean score in sheep was recorded in kidneys and lungs and in goat intestine, lung and kidneys were the major organs affected.
A polymerase chain reaction was optimized under our own laboratory condition for the detection of alpha and epsilon toxins of Clostridium perfringens type D from different tissues. Alpha gene was amplified at annealing temperature 52.2oC with amplicon size 247bp and ETX gene at the annealing temperature.
50.2oC with amplicon size 665bp.
The erythrocytes of different species were used to know the sensitivity against culture supernatants of C. perfringens type D. It was observed maximum hemolysis occurred in human erythrocytes (68%) followed by mice (57%) at 37oC. It was also recorded that a significant increase was found at 37oC as compared to25oC except for dog and rabbits where no significant difference was observed.
Availability: Items available for loan: UVAS Library [Call number: 1596,T] (1).
3.
Phenotypic And Genetic Aspects Of Some Performance Traits Of Buchi Sheep In Pakistan
by Maqsood Akhtar | Prof. Dr. Khalid Javed | Prof. Dr. Muhammad Abdullah.
Material type: ; Format:
print
; Literary form:
drama
Publisher: 2013Dissertation note: Abstract Availability: Items available for loan: UVAS Library [Call number: 1598,T] (1).
4.
Chemical Characterizaton And Toxicological Screening Of Auto-Rickshaw Emissions Particulate
by Khaleeq Anwar | Prof. Dr. Muhammad Ashraf | Dr. Aftab | Dr. Aqeel Javeed.
Material type: ; Format:
print
; Literary form:
drama
Publisher: 2013Dissertation note: Vehicular air pollution is a mounting health issue of the modern age, particularly in urban populations of the developing nations. Auto rickshaws are not considered eco-friendly as to their inefficient engines producing large amount of particulate matter (PM), which poses a significant environmental threat.
Major transformations in the environmental composition are principally attributable to the combustion of fuels by automobiles. Motorized gasoline powered two-stroke auto-rickshaws (TSA) and CNG powered four-stroke auto-rickshaws (FSA)are major sources of air pollution in south Asia and produce toxic amount of PM to the environment. In this study, during the first phase, the PM of TSA and FSA was characterized by using proton induced x-ray emission (PIXE) analysis. The observations of the existing investigation recognized significant increase in Al (P < 0.05), P (P < 0.01), and Zn (P < 0.01) from the PM samples of FSA. In addition, the concentrations of Cu, Fe, K, Mn, Mg, Na, S and Si were also observed exceeding the recommended NIES limits. On the contrary, increased concentration of Sr and V were observed in the PM samples from TSA. It is generally believed that FSA generates smaller amount of PM but the data obtained from this study clearly shows that emissions from FSA are comprised of potentially more toxic substances than TSA. The current research is specific to the metropolitan population and has evidently revealed an inconsistent burden of exposure to air pollutants engendered by FSA in urban communities, which could lead to disruption of several biological activities and may cause severe damage to entire ecological system.
The second phase of this study was conducted to ascertain toxic effects on angiogenesis, embryo development, embryonic movement and phytotoxicity of the PM from TSA and CNG powered FSA. Based on high amounts of aluminum quantified during PIXE analysis of PM from TSA and FSA, different concentrations of aluminum sulfate were also tested to determine its eco-toxicological potential. The PM solution from FSA, TSA and Aluminum sulfate exhibited anti-angiogenic potential with reduction in total area of CAM. Morphological evaluation of embryos exhibited varying degrees of hemorrhages in different groups. In case of phytotoxicity screening using Zea mays, the results demonstrated that all three tested materials were equally phytotoxic at higher concentrations in seed germination(p<0.001). Aluminum sulfate proved to be a highly phytotoxic agent even at the lowest concentration examined.
During the last phase, of the study, the MTT assay demonstrated a significant (p<0.001) dose dependent cytotoxic effect for TSA, FSA and aluminum sulfate on the BHK-21 cell line, establishing that the PM from FSA is a highly cytotoxic material. Mutagenicity was assessed by fluctuation Salmonella reverse mutation assay adopting TA100 and TA98 mutant strains with (+S9) and without (-S9) metabolic activation. Despite the fact that different concentrations of PM from both sources i.e. TSA and FSA were highly mutagenic (p<0.001) even at lower concentrations, the mutagenic index was higher in TSA. The chronic toxicity study revealed that chronic exposure to PM emitted from FSA and TSA resulted in peribrochiolitis, emphesema and infilteration of leukocytes in lung tissues. On the other hand liver, cardiac and kidney tissues exhibited degeneration and necrosis. The data shows that all tested materials are equally ecotoxicand if the existing trend of atmospheric pollution by auto-rickshaws is continued, air-borne metals/heavy metals will seriously affect the normal growth of local inhabitants and increased contamination of agricultural products, which will amplify the dietary intake of toxic element and could result in genetic mutation or long-term health implications.
Availability: Items available for loan: UVAS Library [Call number: 1795,T] (1).
5.
Production, Purification & Characterization Of Recombinant Thermostable Phytase And Its Biological Evaluation In Broiler Chicks
by Furqan Sabir (2007-VA-524) | Dr. Muhammad Tayyab | Dr. Abu Saeed Hashmi | Dr. Ali Raza Awan.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Phytate is the principle storage form of phosphorus in plants particularly in cereal grains and legumes. Mono-gastric animals doesn’t have ability to utilize phytate as phosphorus source. The animals release the undigested phytate from body with manure that cause environmental pollution. Phytases are responsible for the hydrolysis of phytate, resulting in availability of free phosphorus for the animal. The present study deals with the production and characterization of recombinant thermostable phytase and its biological evaluation in the broiler chicks. The PCR resulted in the amplification of 1.8 kb phytase gene using the genomic DNA of Thermotoga naphthophila as template. The purified PCR product was ligated in pTZ57R/T and the ligated material was utilized for the transformation of E.coli DH5α cells. The positive clones were selected on the basis of blue white screening. The restriction digestion of plasmid DNA from positive clones using NdeI and Hind III resulted in the release insert from the vector. The purified phytase gene after restriction digestion was ligated into pET21a already restricted with the same restriction enzymes and the expression was analyzed using E.coli BL21 CodonPlus (DEL) cells. SDS-PAGE demonstrated the intra-cellular production of recombinant phytase. The conditions were optimized for the optimal production of recombinant phytase (PHYTN). The maximal production of PHYTN was recorded when the BL21 CodonPlus cells having recombinant pET21a having phytase gene were induced with 1.4 mM IPTG and 6 hours post induction incubation period. The recombinant protein was purified using various chromatographic techniques and the purified protein was utilized for characterization. PHYTN showed optimal activity at 80 °C and pH 6 in sodium acetate buffer. The enzyme was found metal dependent and presence of Fe3+ or Cu2+ showed enhancing effect on PHYTN activity. Thermostability studies demonstrated that PHYTN retains 90% residual
SUMMARY
71
activity when the protein was incubated at 80 °C for 1h in the presence of 1.5 mM Fe3+. The kinetic studies of PHYTN demonstrated km and Vmax values of 50 mM and 2500 μmole/min respectively when sodium phytate was used as substrate. The characterized PHYTN was used for poultry trials to check the efficacy of the enzyme in poultry birds.
The results depicted that PHYTN put significant effect on the bird weight gain, feed intake and feed efficiency ratio. Presence of 1000 IU/kg of PHYTN resulted in the weight gain in 3rd, 4th and 5th week of trials from 504.766 to 533.535 g, 767.933 to 823.733 g and 999.833 to 1120.277 g respectively when compared with the control. The study demonstrated that this recombinant thermostable phytase is suitable for poultry feed industry and its domestic production will contribute the economic availability of PHYTN for the poultry feed industry. Availability: Items available for loan: UVAS Library [Call number: 2870-T] (1).
