3.
Pathobiological Investigations Of Peste Des Petits Ruminants (Ppr) Virus With Reference To Antiviral Activity Of Nigella Sativa (Black Seed)
by Kiran Aqil (2008-VA-456) | Dr. Muti Ur Rehman Khan | Prof. Dr. Asim Aslam | Dr. Aqeel Javeed.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Peste des Petits Ruminants (PPR) is a highly contagious, infectious, acute or sub-acute transboundary viral disease of domestic and wild small ruminants. It is an economically important viral disease of sheep and goats causing varying degree of morbidity and mortality in susceptible animals which may be as high as 100 and 90 per cent, respectively. PPR is responsible for serious socioeconomic problems. There is no data available regarding pathogenesis and field virus characterization to compare it with vaccinal strain for any difference. Nigella sativa(Black Seed) has antiviral activity against many viruses. Therefore present studywas undertaken to investigate the antiviral effect of Black Seed in vivo and in vitro against PPR virus. Further more time course detection of virus is still needed to be studied.
Nigella sativa (Black seed) has antiviral activity against PPR virus.
Pathogenesis can better be studied through histopathology, necropcy findings and morphometric changes.
A total of 250 clinically positive samples suspected for PPR virus were included in the study. Samples were consisted of nasal, ocular and anal swabs; whole blood in EDTA were collected from suspected animals. In case of mortality morbid material included lungs, liver, spleen and mysenteric lymph nodes were included in the study. Samples were subjected to immune capture Elisa for detection of viral antigen in suspected samples. Samples which found positive foe IC – Elisa were then subjected to RT-PCR for confirmation of virus. After confirmation of virus through IC – Elisa and RT-PCR the positive samples were subjected to virus isolation on vero cell. After isolation of virus, the TCID 50 of the virus was calculated for preparation of inoculum for further use. In this experiment mesenteric lymph nodes and spleen found to be major organ for isolation of PPRV.RT-PCR found to be most reliable and confirmatory diagnostic test for PPRV. Field Virus adaptation on vero cells found to be difficult to optimize.
In this experiment antiviral activity of black seed was checked on vero cells infected with PPRV. Three extracts of N. Sativa were prepared to check the in vitro antiviral activity of black seed. In this study poly saccharides extracted from black seed found to be more effective against PPRV. Adaptation of field virus was done on Vero cell line. Antiviral activity of Black Seed extract was determined in vitro on Vero cell on bases of CPE (Cytopathic effect). The ethanolic and aqueous extract were found to be more toxic to consistency of monolayer of vero cells. The TCID50 of virus was calculated after treating cells with different extracts. In this study poly saccharides extract exhibit lower TCID50‘s as compared to ethanolic and aqueous extract which showed higher TCID50’s.So less cytopethic effect was observed in vero cells treated with black seed extracts. Antiviral activity was determined on base of CPE.
Pathogenesis of virus in natural host was studied through time course detection of virus in body secretions, blood, organs. Histopathological changes were studied.20 goats were procured from market divided into four groups (n=5) A,B,C and D. In animals of group A prophylactic effect of N.Sativa was studied. In group B complete pathogenesis of PPR virus was studied without any prophylactic or therapeutic measure. In group C therapeutic effect of N. Sativa was studied after onset of clinical picture of disease. At the end of this experiment, clinical picture, gross pathology, histopathology, and morphometric changes revealed that N. Sativa has noticeable prophylactic effect on PPR infected goats. It can be used as a therapeutic agent in PPR infected goats but it can’t control pathological effect of virus after onset of infection.
SUMMARY
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Data collected were statistically analyzed by using Microsoft Excel (Microsoft Excel, 2007) and SPSS (for Windows, Version 16.0). The data were put the descriptive analysis and Chi square test was employed to test the significance and test of hypotheses
It was concluded that Black Seed therapy possessed marvelous prophylective effect against PPR virus and RT-PCR was the most efficient methodology to confirm the virus. Availability: Items available for loan: UVAS Library [Call number: 2890-T] (1).
4.
Comparative Efficacy Of Mycotoxin Binders And Effects Of Aflatoxin B1 On Health Status Of Lactating Beetal Goats
by Haq Aman Ullah (2005-VA-180) | Prof. Dr. Aneela Zameer Durrani | Dr. Muhammad Ijaz | Dr. Aqeel Javeed.
Material type: ; Literary form:
not fiction
Publisher: 2017Dissertation note: Aflatoxins are the secondary metabolites produced by moulds particularly by certain strains of Aspergillus flavus and Aspergillus parasiticus. The most toxic is the aflatoxin B1 (AFB1) and is considered as Hepatocarcinogenic. In present study aflatoxin B1 contamination status of different concentrate feeds (cotton seed cake, wanda, wheat bran and homemade concentrate mixture) of dairy goats was investigated in district Lahore of Pakistan. Twenty goat farms were randomly selected and 40 feed samples (two from each farm) were collected. The samples were analysed for the estimation of AFB1 using reverse phase High Performance Liquid Chromatography (HPLC) in Quality Operations Laboratory, UVAS, Lahore. AFB1 was detected in 33 feed samples out of 40 thus with a percent contamination rate of 83%. The quantity of aflatoxin B1 ranged from 0-225.736 ppb (μg/kg). The maximum level of AFB1 was detected in cotton seed cake (mean level 137.059 ± 22.293 ppb) and minimum level was detected in wheat bran (mean level 5.676 ± 1.047 ppb). Amongst the positive concentrate feed samples, 28 samples (85% of the positive) were having the concentration of AFB1 higher than the permissible level recommended by European Communities which is 5 ppb for concentrates. Thus the milk from goats consuming AFB1 contaminated concentrate feed can be a potential hazard for public health. This part of the study fulfills the first objective of the research project, which is to determine the aflatoxin B1 contamination status of concentrate feeds of dairy goats in District Lahore.
Beetal is the best dairy goats’ breed of Pakistan. In second part of the study, lactating beetal goats were used. In this experiment, effects of dietary AFB1 on health status of lactating beetal goats and its transfer from feed into milk as metabolite aflatoxin M1 (AFM1) were investigated. Thirty two lactating beetal goats of 3-4yr old, 6-8 wks. lactation period and weighing 40.91 ± 0.285 Kg, were selected from Small Ruminant Farm, Pattoki, University of veterinary and Animal
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Sciences Lahore, and were randomly divided into four groups A, B, C and D, each having 8 animals. Group A was kept as negative control, while groups B, C and D were fed with aflatoxin B1 through naturally contaminated cotton seed cake. The cotton seed cake was having high level of AFB1 which was 150 μg/kg. After 7 days of adaptation period, each animal of groups B, C and D was individually fed with 30μg, 40μg and 50μg of aflatoxin B1 per day respectively through naturally contaminated cotton seed cake for a 10 days period., thus each animal of groups B, C, and D was fed with 200 g, 266.66 g, and 333.33 g of cotton seed cake per day respectively, to provide the required amount of AFB1 to each experimental animal. Milk samples were collected 24hr before the first AFB1 feeding and on 3rd, 7th and 10th day post aflatoxin B1 feeding. The samples were tested for AFM1 through HPLC, somatic cell count (SCC) and total viable count (TVC). Blood samples were analyzed for aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase (ALP) levels before and after aflatoxin B1 administration. AFM1 was detected in all milk samples of the Groups B, C and D, with concentration beyond permissible level which is 0.05ppb (European communities).The amount of AFM1 excreted in milk was positively correlated with the amount of aflatoxin B1 ingested. The AFB1 was excreted in the milk as AFM1 at 1.35-1.59%. The mean SCC of groups A, B, C and D was 1.44×106 ± 1.75×105, 4.13×106 ± 4.09×105, 3.63×106 ± 4.60×105, 1.38×106 ± 1.56×105 cells/ml respectively. TVC obtained was 2.4×104 ± 1.9×103, 4.3×104 ± 1.6×103, 4.8×104 ± 3.3×103, 3.6×104 ± 3.3×103 cfu/ml of milk for group A, B, C and D respectively. The mean ALP levels were 62 ± 7, 223 ± 22, 147 ± 14, 175 ± 16 U/L for group A, B, C and D respectively. Mean levels of AST of group A, B, C and D were 79 ± 2.214, 110 ± 5.386, 104 ± 2.015, 126 ± 9.456 respectively. Mean levels of ALT of group A, B, C and D were 14 ± 0.326, 20 ± 1.118, 21 ± 1.106, 28 ± 1.250 U/L respectively. The ingestion of AFB1 even at low dose of 30μg in goats resulted in excretion
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of AFM1 in milk beyond the permissible level. SCC, TVC, AST and ALT levels increased with ingestion of dietary AFB1. This part of the study fulfills the 2nd objective of the research project which is to investigate the effects of dietary aflatoxin B1 on health status of lactating beetal goats and its transfer from feed into milk as AFB1.
In the third part of the study, the efficacy of two different mycotoxin binders Toxfin® and Elitox® was determined, in terms of reduction in milk AFM1 excretion, improvement in milk quality and liver health. Groups A and B were kept negative and positive controls respectively. Experimental animals, cotton seed cake and number of groups remained the same as used in part 2 of the experiment. After initial 10 days of the experiment, each animal of groups B, C and D was daily fed with 266.66 g of cotton seed cake to provide 40 μg of AFB1 per day to each animal, for next 7 days. In the meantime, group C was fed with mycotoxin binder Toxfin® (Kemin industries, Inc. USA), at a dose rate of 3 g per Kg of cotton seed cake, while group D was fed with mycotoxin binder Elitox® (Impextraco Belgium ) at a dose rate of 1 g per Kg of cotton seed cake. Toxfin® and Elitox® both significantly decreased the excretion of AFM1 in goats’ milk, detected through high performance liquid chromatography (HPLC). Toxfin® and Elitox® reduced the excretion of AFM1 in milk by 56 % and 48 % respectively, thus the detoxifying ability of Toxfin® in feed was significantly higher than Elitox®. The SCC and TVC remained statistically unchanged after the administration of mycotoxin binders. The enzyme activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were not significantly affected by the mycotoxin binders. The alkaline phosphatase (ALP) level significantly increased in animals receiving either Toxfin® or Elitox®. The mycotoxin binders used in this study were not able to significantly improve milk quality and liver’s health in AFB1 treated goats, during the experimental period. This part of the study fulfills the 3rd objective of the research project, which was to determine the comparative efficacy of two different mycotoxin binders in lactating goats fed with mold contaminated diet. Availability: Items available for loan: UVAS Library [Call number: 2860-T] (1).
