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20194
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Subject: Veterinary Microbiology × End date: 2020 × Start date: 2019 × Type: Thesis × Thesis Type: M.V.Sc. ×
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    ThesisItemOpen Access
    DETECTION AND IDENTIFICATION OF EHRLICHIA SPECIES AMONG DOGS, INCONTACT HUMANS AND TICK VECTORS IN HIMACHAL PRADESH
    (CSHHPKV Palampur, 2019-09-30) ANGARIA, SHIVANI; Chahota, Rajesh
    Vector-borne diseases (VBDs) in animals and humans have gained worldwide importance in the climate change scenario. Ehrlichiosis is one of such rickettsial disease. Ehrlichia is an obligate, intra-cytoplasmic, gram-negative bacteria that cause ehrlichiosis in dogs and other vertebrates, transmitted by tick-vectors and occur mainly in the humid and warm regions. These bacteria multiply within the cytoplasm of White Blood Cells (WBCs) of vertebrate hosts and affect various organs. Canine ehrlichiosis is an important tick-borne disease of dogs worldwide and may lead to zoonosis. Till date, not much work has been done on ehrlichiosis, molecular identification and genetic characterization of native strains/species of Ehrlichia prevalent in Himachal Pradesh, therefore, this study was planned to detect the prevalence of Ehrlichia in dogs, tick vectors and humans in different locations of the state. A total of 215 samples were collected including blood samples from dogs (n = 177), blood samples from in-contact humans (n = 24) and ticks collected from the body of the dogs (n = 24). Samples were collected from 105 locations covering 38 tehsils of 9 districts of Himachal Pradesh. All the samples were tested using Ehrlichia genus and three Ehrlichia species (E. canis, E. ewingii and E. chaffeensis) specific PCR tests. 11 (6.2%) of blood samples from dogs were found positive for ehrlichiosis by examination of buffy-coat smears. By PCR tests, out of 177 blood samples of dogs, 107 (60.4%) were found positive for one or more Ehrlichia species. 61 (34.4%) samples were found harboring multiple Ehrlichia species. All the ticks were identified as Rhiphicephalus sanguineus. From ticks, 4.2 per cent samples were found positive, while no human blood sample was found positive. Phylogenetic positions of all the detected Ehrlichia species/strains were determined which were genetically characterized after nucleotide sequencing of partial 16S rRNA gene. It was found that all E. canis, E. platys species/strains were making clusters with earlier reported species/strains from different parts of India and abroad.
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    ThesisItemOpen Access
    DETECTION OF ENTERIC DNA VIRUSES FROM DIARRHEIC DOGS OF HIMACHAL PRADESH
    (CSKHPKV. Palampur, 2019-07-20) THAKUR, DEEPIKA; Dhar, Prasenjit
    Diarrhea is among the most common clinical conditions seen in dogs leading to high morbidity and mortality. It can have various causes, including infectious agents, like viruses, bacteria and parasites. The aim of the study was to isolate enteric DNA viruses from diarrheic dogs of Himachal Pradesh and characterize them at molecular level. A total of 238 fecal/other samples were collected from diarrheic / apparently healthy dogs along with brief clinical history of the animal. The faecal samples were subjected to DNA extraction and PCR for the detection of enteric DNA viruses viz. CPV, CAV-1 and Dog CV with primers targeting the VP2, pⅦ and replicase gene of CPV, CAV-1 and Dog CV respectively of each virus were used in the study. 109 (45.79 %) samples were found positive for CPV-2b and 4 (1.68%) samples were found positive for CAV-1. No samples were found positive for Dog CV. HA along with HI and DotELISA were standardized for virus detection from clinical samples and infected cell culture fluids using hyper immune sera raised in rabbits against CPV and CAV-1. Dot-ELISA was found to be almost similar to in sensitivity HA-HI for detection of viruses. Growth studies of CPV and CAV- 1 were done on MDCK, A-72 and Vero cells. CPV could be adapted to MDCK and A-72 cells but not to Vero cells while adaptation of CAV-1 was not possible on any cell culture. Physicochemical characterization of CPV was done by checking the effect of temperature, pH, chloroform and formalin on the virus. The VP2 (CPV-2) and pⅦ (CAV-1) gene fragment was sequenced directly which revealed close genetic similarity to Indian and Italian isolates of CPV and CAV, respectively. Microbiological studies on fecal samples revealed 37 bacterial isolates that were resistanct to penicillin and streptomycin in antibiogram and were found concurrently associated with clinical cases of viral diarrhea. In conclusion, diarrhea in dogs in H.P are mostly caused by CPV type 2b strain and occasionally by CAV-1 and is frequently associated with other bacterial species.
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    ThesisItemOpen Access
    DETECTION OF ENTERIC DNA VIRUSES FROM DIARRHEIC DOGS OF HIMACHAL PRADESH.
    (CSKHPKV, Palampur, 2019-07) Thakur, Deepika; Dhar, Prasenjit
    Diarrhea is among the most common clinical conditions seen in dogs leading to high morbidity and mortality. It can have various causes, including infectious agents, like viruses, bacteria and parasites. The aim of the study was to isolate enteric DNA viruses from diarrheic dogs of Himachal Pradesh and characterize them at molecular level. A total of 238 fecal/other samples were collected from diarrheic / apparently healthy dogs along with brief clinical history of the animal. The faecal samples were subjected to DNA extraction and PCR for the detection of enteric DNA viruses viz. CPV, CAV-1 and Dog CV with primers targeting the VP2, pⅦ and replicase gene of CPV, CAV-1 and Dog CV respectively of each virus were used in the study. 109 (45.79 %) samples were found positive for CPV-2b and 4 (1.68%) samples were found positive for CAV-1. No samples were found positive for Dog CV. HA along with HI and Dot- ELISA were standardized for virus detection from clinical samples and infected cell culture fluids using hyper immune sera raised in rabbits against CPV and CAV-1. Dot-ELISA was found to be almost similar to in sensitivity HA-HI for detection of viruses. Growth studies of CPV and CAV- 1 were done on MDCK, A-72 and Vero cells. CPV could be adapted to MDCK and A-72 cells but not to Vero cells while adaptation of CAV-1 was not possible on any cell culture. Physicochemical characterization of CPV was done by checking the effect of temperature, pH, chloroform and formalin on the virus. The VP2 (CPV-2) and pⅦ (CAV-1) gene fragment was sequenced directly which revealed close genetic similarity to Indian and Italian isolates of CPV and CAV, respectively. Microbiological studies on fecal samples revealed 37 bacterial isolates that were resistanct to penicillin and streptomycin in antibiogram and were found concurrently associated with clinical cases of viral diarrhea. In conclusion, diarrhea in dogs in H.P are mostly caused by CPV type 2b strain and occasionally by CAV-1 and is frequently associated with other bacterial species.
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    ThesisItemOpen Access
    MICROBIOLOGICAL INVESTIGATIONS ON RESPIRATORY INFECTIONS OF SHEEP AND GOATS WITH PARTICULAR EMPHASIS ON MYCOPLASMA
    (CSKHPKV, Palampur, 2019-07) Sunaina; Verma, Subhash
    In this study, a total of 163 of 198 morbid samples (nasal and tracheal swabs, lung tissues and pleural fluid) from sheep and goats showing respiratory disease signs were positive for bacteria belonging to seven genera and ten species. Staphylococcus aureus (34.94%) was major bacteria followed by E. coli (21.50%). Other bacteria included Bacillus spp., Pseudomonas aeruginosa, S. epidermidis etc. Nine (6.2%) samples also yielded growth resembling mollicutes which were confirmed by PCR. Out of 37, 83.8% S. aureus carried at least one virulence gene. The mecA was detected in 64.51% S. aureus followed by coa and lukPV in 14 (45.16%) and arcA in 12 (38.50%) isolates. Most S. aureus were sensitive to chloramphenicol, ceftriaxone and tetracycline. Penicillin was at least effective antibiotic. Out of 40, 55% isolates of E. coli carried at least one virulence gene. The eaeA, stx2 and UAL were detected in six, eleven and four E. coli isolates. Most E. coli were susceptible to chloramphenicol and ciprofloxacin. MAR index > 0.2 was recorded for S. aureus (75.6%) and E. coli (10%). Trueperella pyogenes was detected in 10 and Histophilus somni in one nasal swab out 50 nasal swabs of goats. In total, 64 of 198 samples were positive for Mycoplasma spp. using generic-specific primers targeting 16S rRNA gene. Also, 12.5% samples detected positive both for M. mycoides cluster and M. capricolum subsp. capripneumoniae, all of which emanated from an outbreak CCPP having case fatality of 45.07%. The 16S rRNA sequencing identified S. maltophilia, S. entericus and Enterococcus spp. These findings suggest that S. aureus is a major opportunistic pathogen of respiratory tract of sheep and goats harbouring many virulent and AMR genes with high MAR indices and MDR phenotype. This calls for rational usage of antibiotics in veterinary practice. Species-specific PCR should be first choice for diagnosis of CCPP, since MCCP is difficult to culture. Finally, the role of novel bacterial sequences recovered from respiratory tract of goats should be investigated in health and disease.