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Home > Clinical and Phylogenetic Analysis of Hepatitis B Virus Infection and Associations of its Genotypes With Antiviral Therapy in Pakistani Patients.

Clinical and Phylogenetic Analysis of Hepatitis B Virus Infection and Associations of its Genotypes With Antiviral Therapy in Pakistani Patients.

Thesis Info

Access Option

External Link

Author

Mahmood, Majid

Program

PhD

Institute

Pir Mehr Ali Shah Arid Agriculture University

City

Rawalpindi

Province

Punjab

Country

Pakistan

Thesis Completing Year

2014

Thesis Completion Status

Completed

Subject

Zoology

Language

English

Link

http://prr.hec.gov.pk/jspui/bitstream/123456789/10516/1/Majid%20Mahmood_Zoology_2014_Arid_IndegScholar_PMAS_Rwp.pdf

Added

2021-02-17 19:49:13

Modified

2024-03-24 20:25:49

ARI ID

1676725731816

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Genotypes of hepatitis B virus and their role in disease severity and treatment response has not been studied from all over the Pakistan. This research was aimed to study HBV genotype prevalence in all parts of Pakistan, relationship of HBV genotypes with viral and biochemical factors, influence of different treatments and pretreatment factors on patients'' response, and the mutations responsible for resistance. A total of 840 samples was collected from all over the Pakistan and genotyped by type specific primer PCR method. Viral load, ALT and HBsAg tests were performed for 154 treatment naive patients and the values were compared by ANOVA. One hundred and sixty two chronic HBV patients were compared for three treatments viz. Tenofovir, Entecavir and Peg-Interferon. Patients were monitored for virological response, combined response, HBsAg clearance and HBeAg clearance. The data was analyzed by logistic regression and Chi square tests. Viral reverse transcriptase domain of 20 non-responder patients was sequenced and mutations were detected by aligning the sequences with wild type strains. Genotype D was found to be the most prevalent genotype from all parts of the country as it was present in 71.2% of the samples. The second major sample population (17.3%) was found to be infected with a mixture of different combinations of genotypes, with the dominant combination of A+D (13.7%) while genotype A was the third more prevalent genotype (7.7%). Genotypes B, C and E were found from less than 1% of the samples. Significantly higher values of viral load were found in patients with genotype A+D while no relation of genotype with ALT and HBsAg levels was found. Correlation, however was found between the levels of HBsAg and the HBV DNA quantities of the patients having HBeAg negative infection. Combine response of peginterferon treated patients was higher than entecavir and tenofovir treated patients which have higher rate of virological response than peginterferon treated patients. Genotype A and low baseline viral load were associated to better treatment response. HBV genotype, baseline viral load, HBeAg and ALT were found to be significant factors in at least two of the treatment arms. In tenofovir group, only HBeAg and ALT had a significant influence on response rate, where high base line ALT and negative HBeAg were related with better virologic response. In both entecavir and peginterferon arms, genotype A, low viral load, low ALT and negative HBeAg were associated with better virologic response of the patients. Universally accepted resistant mutations were identified from 8 samples while some new mutations were also found which are still not known for their role in resistance. Known mutations rtL180M/I and rtA181V were found from 6 (30%) samples while rtM204V and rtV173L were found from 5 (25%) samples each. New mutations, rtY135S and rtN248H were found from 13 (65%) and 12 (60%) samples respectively. The study concludes that HBV/D is the most prevalent HBV genotype in Pakistan followed by A+D and A genotypes. Mix infection with A+D is associated with high HBV DNA quantities and low rate of treatment response. Genotype A or D alone, low viral load and negative HBeAg, are significant predictors of high response rate. Treatment with tenofovir and entecavir have better virologic response rate as compared to peginterferon treatment.
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(۱)

ایہناں متراں نیں ہر جا اُتے دھوکہ دتا ہئے ۔ہن حالت ایہہ ہو گئی اے کہ چاروں پاسے دھوکہ نظر آندا اے ۔

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(۲)

رب نے انسان کوں رزق وکھو وکھ ڈھنگ راہیں عطا کیتا اے ۔کتے فقیری دی صورت وچ تے کتے فاکہ کشی توںبعد۔

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Seroepidemiology Zoonotic Potential & Chemotherapy of Neosporosis in Dogs & Cattle.

In Pakistan, Livestock is considered as a social security for poor villager as it can be a source of cash at the time of need. Degnala disease reduces the production of these animals directly. Along with other side issues related to Degnala disease, this study was done to diagnose the actual cause of Degnala disease by applying different latest scientific techniques. Prevalence along with risk factors was calculated in the rice growing areas of Punjab, Pakistan. Fungal isolation (n=40) was performed from the rice straw feedings of the Degnala disease affected animals through the technique of spot culture on SDA. Then these fungal isolates were identified through comparing their microscopic and macroscopic characters. Then toxigenic potential was checked for all these isolates through the application of TLC and HPLC. After that, from those isolates which were positive for mycotoxin production potential, most cytotoxic isolate was checked with the application of MTT assay. Then the most cytotoxic isolate was inoculated on non-contaminated rice straw and fed to the experimental animals to see a similarity of natural cases of Degnala disease. Finally treatment was conducted to see a proper combination of various drugs against this disease. Toxigenic potential of different candidate fungi, isolated from rice straw feeding of Degnala disease affected bovines was analysed along with Species, age, gender and season wise prevalence. Out of 1536, 104 (6.77%) cases showed positive signs for this disease with a significant association (p<0.05) between rice straw feeding in buffaloes, winter season and bovines having an age of more than one year. Complete Blood Count showed marked increase in erythrocyte sedimentation rate and all white blood cells numbers, except lymphocytes in positive cases. There was a significant increase (p<0.05) in Alanine amino transferase, Aspartate amino transferase and Alkaline phosphatase noticed in Liver Function Test. At the same time, increased value of Creatinine was noticed in Renal Function Test. For isolation and screening of toxigenic fungi, rice straw samples (n=40) being fed to the positive cases were processed further, out of which there were 85 fungal isolates mainly of Aspergillus (57), Penicillium (10), Fusarium (04), Zygomycetes (03), Curvularia (01) and unidentified (10). All isolated fungi were subjected for mycotoxin production and only 11 showed mycotoxin producing capability (including Aspergillus, Penicillium and Fusarium isolates) analysed by Thin Layer Chromatography and quantified through High Performance Liquid Chromatography. It is concluded that all the fungi, contaminating rice straw feeding of Degnala affected animals are not toxigenic. This work will help in establishing major mycotoxin producing fungi leading to the probable cause of Degnala disease in bovine. With the help of MTT assay on vero cell line, most cytotoxic fungus was identified. After an incubation with vero cells, OD values of all the candidate fungi were compared through one way ANOVA. Results of this analysis showed that Fusarium was at the highest ranking and then was the A. flavus with a significant value of 0.006 and 0.039. Finally it was concluded through these systematic steps of converging the diagnosis that, out of all the 85 suspected fungi, Fusarium (isolate number S 8.1) was the most cytotoxic isolate obtained from the rice straw feedings of Degnala affected animals in our study. For molecular diagnosis of the most cytotoxic isolate of Fusarium, PCR was conducted and the results showed that ultimately the final PCR product was successfully amplified against the mentioned primer of ITS conserved region for Fusarium genera and the DNA product was with a length of 570 base pairs. Experimental feeding trials were conducted by inoculating Fusarium (the most cytotoxic isolate) and A. flavus (second most cytotoxic one after Fusarium) separately and in combination compared with the negative control group, all groups were of eight animals each. It was concluded that alone Fusarium was able to produce Degnala disease, while its combination with A. flavus was more lethal. Ultimately the treatment trials proceeded with penta-sulphate, oxytetracycline and antiseptic topical application as therapeutic treatment were shown to be very effective against Degnala cases. While in all the affected animals feeding of affected rice straw was ceased. Only withdrawal of affected rice straw from the feedings of Degnala affected animals was not effective unless proper treatment as mentioned here was not conducted. Analysed. The expected results of the study shall be helpful to make exact diagnosis and treatment of infected buffaloes and cattle that is further helpful for timely prophylaxis and control of the Degnala disease in the rice growing areas of Pakistan and South Asia.