Thumbnail Image

Theses

Browse

20093
Reset filters
Subject: Veterinary Microbiology × Author: KAU × End date: 2009 × Start date: 2009 × Thesis Type: M.V.Sc. ×
Reset filters

Search Results

Now showing 1 - 3 of 3
  • Thumbnail Image
    ThesisItemOpen Access
    Development and evaluation of whole cell and membrane protein vaccines against mycoplasma gallisepticum
    (Department of Veterinary Microbiology, College of Veterinary and Animal Sciences, Mannuthy, 2009) Surya, Sankar; KAU; Krishnan Nair, G
    A study was carried out to develop and evaluate the efficacy of vaccines against M. gallisepticum infections in Gallus domesticus using whole cell and membrane proteins produced under iron sufficient and restricted conditions, incorporating the adjuvants aluminium hydroxide and saponin either alone or in combination. A total of 50 samples were collected in broth and in addition 25 tracheal swabs were streaked onto PPLO agar plates also, from birds showing respiratory ailments. Mycoplasma genus specific PCR revealed twelve positive samples and on subjecting these samples to species specific PCR, a total of four samples showed positive results. From four species specific PCR samples, two isolates could be obtained, named as MG UPF-1 and MG UPF-2, which were sub cultured and subjected to morphological, biochemical and serological characterization. The SDS-PAGE profile of whole cell proteins of the two isolates revealed similar profile, indicating their homogeneity. Whole cell and membrane proteins were isolated from cultures grown under iron sufficient and iron deficient conditions. Both the whole cell and membrane proteins revealed bands ranging from 24 kDa to 200 kDa on SDS-PAGE. The whole cell proteins had two prominent bands at 75 kDa and 35 kDa and eight faint bands, which the membrane proteins lacked. The whole cell and membrane proteins produced under iron restricted conditions had an additional band of 52 kDa, which was not seen in the respective protein groups isolated under iron sufficient condition. Most of the proteins resolved on SDS-PAGE in the four sets of proteins were found to be antigenic as found out by western blot analysis. The optimum concentration of the proteins used for vaccine preparation was 84.2 µg per dose of the vaccine. Formalin inactivated vaccines were formulated with saponin and aluminium hydroxide alone and in combination at a final concentration of 100 µg/dose and 25 per cent v/v respectively, thereby producing eight different sets of vaccines. The sterility and safety testing of the vaccines were carried before vaccination trials. The infective dose of M. gallisepticum cells for challenge studies was found to be 1.1 x 105. For vaccination studies, 240 chicks of five weeks age were divided into 10 groups and were inoculated with eight different sets of vaccines. One group was inoculated with commercial vaccine and other was kept as control. Potency testing for humoral response was conducted with HI test and CMI response with LMIT and Blastogenic calorimetry. The mean HI titre of groups vaccinated with iron sufficient and restricted saponin adjuvanated vaccines showed an increase in their titre at first week post booster vaccination followed by decline of the titre on subsequent collections on third and fifth week post booster vaccination. The mean HI titres of groups vaccinated with iron sufficient and restricted saponin-aluminium hydroxide adjuvanated vaccines maintained significantly high HI titres during the entire study period. Commercial vaccine group also showed a similar response. The control group showed significantly less titre from other vaccinated groups during the entire time period. Leukocyte migration inhibition test and Blastogenic calorimetry were carried out to assess the CMI response evoked by different sets of vaccines. A significant CMI response was shown by all vaccinated groups except commercial vaccine and control group. To assess the protective response evoked by different vaccines, two challenge studies were conducted two weeks apart. All the vaccinated birds were having significantly lower lesion score when compared to the control group. Mycoplasma gallisepticum was re-isolated from all the control birds with lesions during the first challenge study and from 70 per cent of the birds in the second challenge study. A latex agglutination test was developed using iron restricted M. gallisepticum whole cell proteins. When compared with HI test, LAT had a sensitivity of 95 per cent and specificity of 93 per cent. The test was statistically significant as indicated by a kappa value greater than 0.81. The saponin-aluminium hydroxide adjuvanated whole cell and membrane protein vaccines had elicited similar humoral immune response than vaccines adjuvanated with saponin alone. With regard to CMI response, the experimental vaccines produced a significant CMI response whereas, commercial vaccines failed to evoke a protective CMI response. In brief, saponin-aluminium hydroxide adjuvanated whole cell and membrane protein vaccines, especially those produced under iron restricted conditions, could be an effective tool to afford protection against M. gallisepticum infection, as it would stimulate both humoral and cell mediated immune responses in host Latex agglutination test was found to be an effective tool for flock screening of birds for M. gallisepticum antibodies and the test is simple, rapid, and easy to conduct, with applicability under field conditions.
  • Thumbnail Image
    ThesisItemOpen Access
    Isolation and characterization of bacteria associated with gastroenteritis in weaned Piglets
    (College of Veterinary and Animal Sciences, Mannuthy, 2009) Atulya, M; KAU; Koshy, John
    Gastroenteritis is a leading cause of morbidity and mortality in piglets. A number of factors are involved in post weaning diarrhoea in piglets. A comprehensive study was performed to examine the incidence, bacterial etiology, drug sensitivity, plasmid profile and pathogenicity of the bacteria isolated from weaned piglets with gastroenteritis prevalent in and around Kerala Agricultural university. Samples were taken only from piglets with diarrhoea that had not been previously treated with antibiotics. Rectal swabs were collected from live diarrhoeic piglets and intestinal contents, pieces of jejunum, ileum, colon, mesenteric lymph nodes, liver, spleen and stomach were collected during post mortem examination after taking all sterile precautions. Isolation of causative bacteria was made by culturing on Brain Heart Infusion Agar, Mac Conkey agar, Mannitol Salt agar, Blood agar, Brucella agar and Cooked Meat medium. The identification of isolates was carried out as per standard protocols. All the procedures of biochemical testing were followed as described by Barrow and Feltham (1993). For classification of Staphylococcus isolates, a system suggested by Baird Parker (1965) was considered. A total of 53 bacterial isolates were identified to species level from 82 samples tested for pathogens. Six different microorganisms were encountered in this study, with Escherichia coli being dominant, followed by Pseudomonas aeruginosa, Staphylococcus subgroups BPIII and BPV, Serratia fonticola, Salmonella typhimurium and Aeromonas hydrophila. Thirteen different serotypes of E. coli were encountered, with O69 being dominant and the others were O38, O5, O84, O132, O80, O79, O56, O41, O25, O109 and O103 and two were untypable. Majority of the isolates exhibited multidrug resistance. Plasmid profile of the Gram negative isolates were determined and 80.43 per cent were found to bear plasmids. Pathogenicity of the isolates was determined by performing in vivo mice pathogenicity test.
  • Thumbnail Image
    ThesisItemOpen Access
    Antioxidant potential of malphigia glabra(acerola) berries in rats
    (College of Veterinary and Animal Sciences , Mannuthy, 2009) Arul Mary Luveena, A; KAU; Karthiayini, K
    The present study was designed to assess the effect of aqueous extract of mature fruits of Malphigia glabra (Acerola) berries on paracetamol induced hepatotoxicity in rats. Forty-two adult male Wistar albino rats were used for the experiment. The rats were randomly divided into five groups with eight animals each in G1, G2 and G5 and nine animals in G3 and G4. Group G1 served as normal control rats. The G2 (untreated vehicle alone) rats were administered with 40 % sucrose syrup p.o. @ 10 ml/kg b.w. on day one and two and then at three days interval for 21 days. The G3 group of rats were administered with paracetamol @ 2 g/kg b.w. p.o., on day one and two and at every three days interval upto day 18. The G4 (curative group) rats were administered with paracetamol p.o. @ 2 g/kg b.w. on day one and two and at every three days interval upto day 18 and Acerola berry extract (20 ml/ kg b.w., p.o.) for 21 days. The G5 (protective group) rats were given Acerola berry extract p.o. @ 20 ml/ kg b.w. for 17 days and paracetamol p.o. @ 2 g/kg b.w. on day 18 and 19. Body weight was recorded at weekly intervals. Blood samples were collected from eight animals in each group on day zero, four, 10, and 21. In G5 group, an additional blood collection was made on 18th day of the experiment before administration of paracetamol. The haematological parameters such as total erythrocyte count (TEC), haemoglobin (Hb) concentration, total leukocyte count (TLC) and differential leukocyte count (DLC) and serum biochemical parameters such as alanine amino transferase (ALT), aspartate amino transferase (AST), alkaline phosphatase (ALP), total cholesterol, total protein, albumin, globulin, albumin:globulin, blood urea nitrogen (BUN), total bilirubin, direct bilirubin, indirect bilirubin, reduced glutathione were analysed. One animal each from G3 and G4 was euthanized on day four and rest of the rats were euthanized on day 21. Levels of liver reduced glutathione, lipid peroxides and superoxide dismutase (SOD) on 21st day were estimated. Representative samples of liver and kidney tissues collected on day four and 21 were subjected to histopathological examination. Administration of paracetamol in G3 group caused a significant (P 0.05) increase in the levels of serum ALT, AST, ALP, total cholesterol, BUN and total, direct and indirect bilirubin while reduced glutathione content was significantly reduced. The activities of liver reduced glutathione and SOD were also decreased significantly whereas the liver lipid peroxide content was significantly increased. Haematological analysis showed significantly decreased TEC and Hb concentration and a significantly increased TLC with monocytosis. No significant (P> 0.05) variation was observed in body weight, and in the levels of serum total protein, albumin, globulin and albumin:globulin. Histopathology indicated necrosis of hepatic cells, diffused haemorrhage, central venous congestion and focal coagulation in liver; while tubular dilatation and congestion was observed in kidney. Administration of Acerola berry extract along with paracetamol in G4 (curative group) rats effectively reversed the levels of serum ALT, AST, ALP, total cholesterol, BUN, total bilirubin (direct and indirect), reduced glutathione (in liver and serum), liver SOD and lipid peroxides in liver to normalcy signifying the antioxidant and hepatocurative effect of the extract. However, the active antioxidant components of Malphigia glabra such as vitamin C and polyphenols has a short half life, so that the berry extract could not produce any prophylactic effect against paracetamol induced toxicity in G5 (protective group) rats, evidenced by toxic range of values.