Thumbnail Image

Theses

Browse

20223
Reset filters
Subject: Veterinary Microbiology × End date: 2022 × Start date: 2020 × Type: Thesis × Thesis Type: M.V.Sc. ×
Reset filters

Search Results

Now showing 1 - 3 of 3
  • Thumbnail Image
    ThesisItemOpen Access
    Physicochemical properties of live attenuated duck plague vaccine and evaluation of stabilizer efficacy for lyophilization
    (College of Veterinary Science, Assam Agricultural University, Khanapara Campus, 2022) Barua, Jonmoni; Das, Sutopa
    Duck plague (DP) or Duck Viral Enteritis (DVE) is an acute contagious herpesvirus infection of ducks and waterfowl of the family Anatidae of the order Anseriformes. Anatid Herpesvirus-1 (AHV-1) or duck enteritis virus (DEV)of the family Herpesviridae is the responsible agent for DP or DVE which is a member. The disease is known to have a global distribution and is associated with significant economic losses worldwide. The only method for preventing and controlling the disease is vaccination. Also, an active decontamination process for an effective vaccination programme in field conditions is important. So, in the present study emphasis has been laid to understand the physicochemical properties of a DPV vaccine strain along with evaluation of thermostability of freeze-dried vaccine with different combinations of stabilizers. In the present study, a vaccine strain of DPV available in the DBT-ADMaCDepartment of Veterinary Microbiology, College of Veterinary Science, AAU, Khanapara was revived in CEF and selected for study on the basis of identity with DPV by observing CPE, PCR and molecular characterization. Characteristic CPE like vacuolation, rounding, syncytium formation and ultimately detachment of cells were observed, in case of PCR band was observed at 1510 bp which proves similar identity with DPV. Molecular characterization revealed homology with DPV isolates from India (Kerala and Assam) and China. Quantitation was done at each step to find out the titre by TCID50/ml after every evaluation right from initial titre, loss of titre during lyophilization, loss of titre during the evaluation of physicochemical treatment, stability evaluation of the freeze-dried vaccine vial, as well as reconstituted vials. The initial titre was found to be 6.9±0.17. The vaccine virus was found to be sensitive to temperatures exceeding 56ºC and above, pH 3 and below, and pH 11 and above. It was also found sensitive to ether and trypsin. On sterility test, no growth was found on the culture. Lyophilization was carried out with 3 combinations of stabilizers namely LS, PTI and LHT. On quality evaluation, PTI and LHT showed uniform cake formation along with minimal loss of titre due to lyophilization. To check the thermostability of freeze-dried vaccines and reconstituted vaccines, vials were exposed at different temperatures. Among the freeze-dried vaccine, LHT could keep the highest titre when exposed to different temperatures and sampled at different time intervals. Although, LS and PTI too could keep with the infectivity titre with minimal loss of titre. In case of the reconstituted vaccine, NSS showed better stability at different temperatures than PBS, though the differences were minimum between the two. Finally, it can be concluded that LHT is one of the better stabilizers for DPV freeze-dried vaccine production. Alternatively, LS and PTI can be used by utilizing a suitable freeze-drying protocol. PBS and NSS both can be used as a diluent for the lyophilized DPV vaccine although in this study NSS was found to be superior. Hence, stabilizer LHT with diluent NSS was found to be superior for the DPV vaccine strain under this study.
  • Thumbnail Image
    ThesisItemOpen Access
    Neutralization efficacy of classical swine fever c-strain specific antibody to different genotypes circulating in North Eastern states, India
    (College of Veterinary Science, Assam Agricultural University, Khanapara Campus, 2022-09) Sarma, Jayashree; Barman, N N
    Classical swine fever (CSF) or Hog cholera is a highly contagious viral diseases affecting domestic and wild pigs. It has been a big threat to the piggery sector globally, causing negative impact on the economic background. The disease is highly endemic in India including NER. Assam too records highest CSF outbreaks. The recent outbreaks recorded occurrence of genotype 2.2, 2.1 and 1.2 besides wide prevalence of the historical genotypes 1.1. Outbreaks in vaccinated herds and shift in genogroup 1 to 2 globally, has raised the concern over the antigenic variation, protective immune response and neutralizing capacity of C-strain vaccine antibodies. Thus the present study was undertaken to explore the cross- protection efficacy of C-strain vaccine antibodies to the different genotypes or the need for potential vaccine candidate. Tissue samples and lyophilized isolates were selected for the study from CSFV repository, Department of Veterinary Microbiology. Sandwich ELISA and nested RT-PCR was done to determine the presence of the virus. Out of total 49 samples, overall positivity in SELISA was 36.0% (18) and in nested RT-PCR was 30.0% (15). The recovery rate of tissue samples was lower (35.0%) in comparison to lyophilized isolate 40.0%. Molecular characterization of the samples found positive in screening test was done based on E2 full length gene. Five isolates representing each state from north-east was successfully amplified at 1119bp for full length amplification of E2 gene. Genogrouping and phylogentic analysis revealed, genogroups 1.1 and 2.2 circulating in NER showing 98% and 84% nucleotide identity, respectively with the reference Alfort/187 strain. Whereas 99% nucleotide identity within the genogroup. The five isolates with known genogroups were propagated in PK-15 cell line upto 5th passage level and confirmed by S-ELISA and nested RT-PCR. Four isolates were isolated successfully except the isolate from Assam. The OD value at different passage level ranged from 0.589 to 1.763, showing an increase in titre with each subsequent passage. CSFV_AAU_Mg01 showed highest OD value at 5th passage. In-situ demonstration of CSFV by IPT revealed reddish brown cytoplasm indicating replication of the virus in the cytoplasm.TCID50 of the passaged viruses were done by FAT showing comaparble titre (4.49-5.16) with that of vaccine strain at 5th passage level. CSFV_AAU_Mg01 showing highest log TCID50 10 5.16 log TCID50 per ml. Hyperimmune sera was raised using purified cell culture adapted lapinised vaccine showing titres of 1:800 and 1:1600 and used for immunological characterization of the isolates by cross neutralization assay. A 50% neutralization titre of the hyperimmune serum ranged from 1/133 to 1/158 when assayed against the different viruses by FAT. Neutralization and cross – neutralization assay with C-strain specific antibody showed 100% neutralization with genotype 1.1, whereas 84% in geno-type 2.2. The study revealed genotypes 1.1 and 2.2 widely circulating in NER with lower neutralization efficacy of vaccine antibodies to heterologous genotypes.
  • Thumbnail Image
    ThesisItemOpen Access
    Seroprevalence and molecular detection of bovine brucellosis and leptospirosis in Assam
    (College of Veterinary Science, Assam Agricultural University, Khanapara Campus, 2022-09) Devi, Bandana; Saikia, G K
    Brucellosis and leptospirosis are neglected zoonotic disease prevalent throughout the world. Bovine brucellosis is predominantly caused by Brucella abortus. Leptospirosis in bovine is mainly caused by Leptospira serovars under the serogroup Sejroe. Both the diseases share some common clinical signs and symptoms and cause severe economic losses. The present study was undertaken to estimate the seroprevalence of bovine brucellosis and leptospirosis and diagnose both the diseases by molecular detection of Brucella and Leptospira organisms in clinically suspected and seropositive cases. The study was carried out in Assam during August 2021 to July, 2022. In this study, a total of 1013 cattle serum samples were collected from 11 districts of Assam viz. Tinsukia, Lakhimpur, Dhemaji, Sonitpur, Nagaon, Kamrup-M, Barpeta, Udalguri, Kokrajhar, Dhubri and Cachar, and screened for Brucella antibodyby Rose Bengal Plate Test (RBPT) and Indirect-enzyme Linked Immunosorbent Assay (i-ELISA) to estimate the seroprevalence of the disease. To detect seroprevalence of leptospirosis, a total of 512 cattle serum samples were collected from 7 districts of Assam viz. Dhemaji, Bishwanath, Nagaon, Kamrup-M, Bongaigaon, Kokrajhar and Dhubri, and tested by i-ELISA for Leptospira antibody. A total of 41 serum samples were found to be positive for Brucella antibody by both the tests with a seroprevalence rate of 4.04% and 19 out of 512 serum samples were found to be positive for Leptospira antibody by i-ELISA with a seroprevalence rate of 3.71%. Higher seroprevalence of brucellosis was recorded in female (4.60%) than in male animals (2.16%). Similarly, higher seroprevalence of leptospirosis was recorded in female (4.53%) than in male animals (1.45%). Age wise seroprevalence of brucellosis was found to be highest in animals of 2.1 to 5 years (1st to 3rd lactation) of age(6.32%) followed by animals of 5.1 years and above (4th lactation onwards) age group (2.90%). In case of leptospirosis, animals of 5.1 years and above (4th lactation onwards) age group showed highest seroprevalence (7.47%) followed by animals of 2.1 to 5 years (1st to 3rd lactation) of age (4.14%). In both brucellosis and leptospirosis, higher seroprevalence rate i.e., 6.54% and 4.33%, respectively was recorded in crossbred than in local cattle (1.07% and 2.97%, respectively). In case of brucellosis, animals reared in organised farms showed higher seroprevalence (8.94%) than the animals reared in semi-organised (3.08%) and backyard farms (1.63%). On the other hand, in case of leptospirosis, animals reared in backyard farms showed higher seroprevalence (7.20%) than the animals reared in semi-organised (2.63%) and organised farms (2.47%). In relation to animal health status, the seroprevalence of both the diseases were found to be highest in clinically ill animals with 40.90% for brucellosis and 8.18% for leptospirosis than apparently healthy animals i.e., 3.22% for brucellosis and 3.06% for leptospirosis. Again, among clinically ill animals, seropositivity for brucellosis was highest in animals with history of abortion (66.66%) followed by animals with retention of placenta (50.0%) and repeat breeding (33.33%). Similarly, in case of leptospirosis, highest seroprevalence was found in animals with history of abortion (33.33%) followed by animals with retention of placenta (25.0%) and repeat breeding (13.33%). In this study, both Brucella and Leptospira antibodies could be detected in 5 out of 512 serum samples screened by i-ELISA specific for both the diseases with a seropositivity rate of 0.976%. For molecular detection of brucellosis, 41 seropositive (32 apparently healthy and 9 clinically ill) and 23 clinically suspected (seronegative) samples (whole blood, aborted foetus, placenta, vaginal swab) were tested by Brucella genus specific PCR. Out of these, 9 clinical samples (39.13%) from seronegative cases and 4 samples (9.75%) from seropositive cases were found to be positive for Brucella genomic DNA in Brucella genus specific bcsp31 PCR. Overall, out of 64 samples examined, Brucella genomic DNA could be detected in 13 number of samples with a positivity rate of 20.31%. All 13 Brucella DNA were confirmed as Brucella abortus by multiplex PCR (AMOS). For molecular detection of leptospirosis, 19 seropositive (15 apparently healthy and 4 clinically ill) and 33 clinically suspected (seronegative) samples (whole blood, aborted foetus, placenta, vaginal swab and urine) were tested by Leptospira lipL32 gene PCR. Out of 33 clinically suspected (seronegative) samples, Leptospira DNA could be detected in 6 number of samples with positivity rate of 18.18%. Leptospira DNA could not be detected from seropositive samples. As a whole, out of 52 samples, Leptospira DNA could be detected in 6 sample with an overall positivity rate of 11.53%.