Spatial Ecology And Distribution Of Soil Borne Burkholderia Mallei In Punjab, Pakistan
Material type: Book ; Literary form:
Publisher: 2017 Dissertation note: Burkholderia mallei is a causative agent of glanders, the disease of equines. The disease is characterized by pulmonary, nasal and cutaneous forms. B. mallei is excreted through nasal discharge, lacerated skin/wounds and expiration. Diseased animals shed bacteria through the discharges contaminating soil, water, fodder and other susceptible animals in its vicinity. The present study was designed to map and investigate the association of different physical factors and soil chemistry analytes with persistence of B. mallei genome in soil of 10% percent villages (n=456) from eight selected districts of Punjab province, Pakistan.
Eleven (0.48%) out of 2, 280 soil samples were positive for B. mallei genome in varied locations of Punjab. Higher prevalence (2.37%) for genome was detected in Sheikhupura district followed by Chakwal district (2.10%). None of the samples from Gujranwala, Sahiwal, DG Khan, Attock, Faisalabad and Sargodha districts were found positive for B. mallei genome. The genome of B. mallei was distributed in 25% study districts of Punjab, Pakistan. In Chakwal district, the genome of B. mallei was strongly associated with moisture (p=0.008) in all positive samples ranging from 0.80 to 39.20%, Phosphorous (p=0.050) ranging from 1.74 to 21.75 mg/Kg. While, this association in Sheikhupura district soil samples was with Sodium (p=0.018) and moisture (0.026) ranging from 1.90 to 133.59 mg/Kg and 0.80 to 39.20%, respectively. The odds of detecting DNA of B. mallei were recorded higher (1.4, 6.8, 5.0, 2.8 and 10.6 ) when soil sample sites were < 500 meters away from vehicular traffic roads, < one kilometer from animal markets, < 100 meters from canal, animal density < 1,000 animals and human population < 300 houses/village. While the odds of detecting DNA of B. mallei were 0.1, 0.3, 0.4, 0.2 and 0.5 when soil sample sites were > 500 meters from vehicular traffic roads, > one kilometer from animal markets, > 100 meters from canal, animal density > 1000 animals and human population > 300 houses/village, respectively. Soil-borne B. mallei DNA is more likely to be detected in areas closer to roads with vehicular traffic along the interstate routes in Punjab and soil containing low level of moisture.
It was concluded that soil of two districts out of eight selected was positive for B. mallei genome in Punjab province. Odds of less distance from main road to animal farm and high animal density at farm were positively associated with B. mallei DNA persistence in soil. Moisture, sodium and phosphorus were positively associated with persistence of B. mallei DNA in soil.
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Prevalence, Molecular Diagnosis And Chemotherapy Of Degnala Disease In Large Ruminants Of Punjab.
Material type: Book ; Literary form:
Publisher: 2017 Dissertation note: 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.
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Evaluation Of White Sesame Seed Oil As A Functional Food Ingredient And Its Role To Mitigate Hyperglycemia
Material type: Book ; Literary form:
Publisher: 2017 Dissertation note: White sesame seed oil (WSSO) contains appreciable amount of various bioactive components including tocopherols, polyphenols, phytosterols and lignans (sesamin & sesamolin) known to have positive impact against certain diseases. Characterization of white sesame seed oil (PB Til-90) showed the presence of bioactive components make it suitable for human consumption. The comparison of WSSO based functional cookies and vegetable fat (VF) based cookies showed that energy and fat% were significantly higher (P < 0.05) in WSSO than VF cookies. At 60th day, mean moisture, peroxide value, and acidity were higher (P < 0.05) in VF cookies. Over time, protein and fiber% decreased significantly (p < 0.05) in both cookies but remained higher (P < 0.05) in WSSO at 60 days. By the end of the 60 days of storage time, moisture content in SO cookies increased approximately 34% (p < 0.05), while other components decreased significantly (p < 0.05) over time; (protein: -0.2%, fat: -3%, fiber: -5.5%, and ash: -7.9%). At 60 days there were significant (p < 0.05) differences between groups. Moisture was significantly higher in VF verses SO, whereas all other components were significantly (p < 0.05) lower in VF group compared to SO group; (protein: -7.6%, fat: -9%, fiber: -5% and ash: -11 %).
Over time, from baseline to 60 days, peroxide value increased approximately 252% in SO cookies. Additionally, in SO, acidity, nitrogen free extract, and thiobarbituric acid values increased (35%, 3%, 54% respectively), while bioactive components, sesamin and sesamol, decreased significantly (p <0.05) over time (i.e., -0.22% and -1.2% respectively). A similar trend was observed in VF cookies. Over the period from baseline to 60 days, the mean rating on each attribute decreased significantly (p < 0.05) for each cookie type. For SO cookies, colour decreased by about -5.5%, flavour -8%, taste -16%, texture -11.6%, crispness
-8% and overall acceptability by -14%. A similar trend was observed in VF cookies. In VF cookies, the mean rating for colour decreased -9%, flavour decreased by -11%, taste decreased by -11%, texture decreased by -12%, crispness decreased by -7% and overall acceptability decreased by -5.5%. By day 60, there were significant (p < 0.05) differences in the sensory rating between groups.
For efficacy study on rats, sixty-three male Sprague-Dawley rats were randomized into standard diet groups (normal control, NCON, n=21) and (diabetic control, DCON, n=21) and a diabetic sesame oil (DSO) (n=21) group which were fed a diet containing 12% WSSO. Blood samples were analyzed at 0, 30 and 60 days. Differences between groups and across days were assessed with two-way repeated measures ANOVA. At baseline, GLU and INS were similar in both diabetic groups (mean 248.4 + 2.8 mg/dl) and (mean 23.4 ± 0.4 μU/mL) respectively. At 60 days, GLU was significantly (p < 0.05) higher in DCON (298.0 ± 2.3 mg/dl) as compared to DSO (202.1 ± 1.0 mg/dl). Activities hepatic antioxidant enzymes increased significantly (p < 0.05) in each variable across time from baseline to 60 days; SOD: (9.7 ± 0.1 to 15.5 ± 0.6 IU/mg), CAT: (6.6 ± 0.1 to 12.5 ± 0.8 IU/mg), GPx (11.1 ± 0.3 to 35.9 ± 3.2 IU/mg), APx (48.7 ± 1.6 to 76.1 ± 1.9 IU/mg) in the DSO group as compared to the DCON and NCON groups. In the DSO group, CK decreased significantly (p < 0.05) from baseline (291.1 ± 0.9 U/L) to 60 days (245.5 ± 7.2 U/L) from both the control groups, while CK-Mb decreased significantly (p < 0.05) from baseline (550.5 ± 3.9 U/L) to 60 days (510.8 ± 6.8 U/L) from NCON group but was not significantly different from DCON group. Among liver function tests, ALP increased over time in both diabetic groups (i.e., in DSO group from baseline to 60 days it raised from 246.7 ± 3.3 U/L to 277.7 ± 2.8 U/L) and at 60 days was significantly higher (p < 0.05) than NCON in both groups but were not significantly different from each other. In contrast, ALT from baseline (81.5 ± 3.7 U/L) to 60 days (67.4 ± 2.7 U/L) and AST from baseline (148.7 ± 3.5 U/L) to 60 days (118.3 ± 1.2 U/L) significantly decreased
(p < 0.05) in the DSO group as compared to DCON or NCON resulting in significantly lower values than both control groups by 60 days. At 60 days, urea in the DSO group decreased from baseline (38.5 ± 2.3 to 30.9 ± 1.1) such that it was significantly lower (p < 0.05) than both control groups. From baseline to 60 days, creatinine significantly increased (p < 0.05) in the two diabetic groups; in DSO group at baseline creatinine was (0.3 ± 0.0 mg/dl) and increased up to (0.4 ± 0.1) after 60 days whereas it remained fairly stable in the NCON group. At 60 days, creatinine was significantly higher in both the diabetic groups as compared to NCON. At 60th day; cholesterol, triglyceride, VLDL, and LDL was significantly lower (p < 0.05) and HDL significantly was significantly higher than DCON, and NCON. The results indicated that there were no significant differences between the DSO or DCON groups in electrolyte balance, minerals, and hematological values.
For efficacy study on humans, forty-six subjects with Type 2 diabetes were recruited and randomly divided into two equal groups (diabetic control, DCON) and diabetic sesame oil (DSO). At baseline, 30, 60, and 90 days, blood samples were drawn and analyzed. Two-way repeated measures ANOVA was used to evaluate the difference between groups and across time. In both groups GLU, INS, and HbA1c were not significantly different at baseline; (mean 187.07 + 5.63 mg/dl), (mean 12.12 ± 1.03 μU/mL), and (mean 7.55 + 0.37 %) respectively. At 90 days, GLU was significantly (p < 0.05) decreased in DSO (137.83 ± 3.16 mg/dl) when compared with DCON (218.13 ± 5.92 mg/dl) while insulin was significantly increased in DSO (23.13 ± 1.15 U/ml) as compared to DCON (7.93 ± 0.38 U/ml). At 90th day HbA1c was significantly lower (p < 0.05) in DSO as compared to DCON. TBARS was significantly lower (p < 0.05) in DSO (1.08 ± 0.05 [MDA] nmol/ml) as compared to DCON (2.26 ± 0.07 [MDA] nmol/ml). In DSO, activities of hepatic antioxidant enzymes (SOD, CAT, and GPx) increased while in DCON these activities decreased significantly (p < 0.05) across time period. Biomarkers of liver, cardiac and renal functions improved significantly in
DSO as compared to DCON. At 90th day; cholesterol, triglyceride, VLDL, and LDL were significantly lower (p < 0.05) and HDL was significantly higher than DCON. There were no significant differences between the DSO or DCON groups in electrolyte balance, minerals, and hematological values.
It was concluded that consumption of white sesame seed oil significantly improved blood glucose regulation, reduced oxidative stress, improved antioxidant activity and biomarkers hepatic, cardiac and liver enzymes in male sprague dawley rats and type 2 diabetic patients.
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