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Subject: Veterinary Microbiology × End date: 2006 × Start date: 2006 × Type: Thesis ×
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    ThesisItemOpen Access
    PARTIAL SEQUENCING AND CHARACTERIZATION OF VP2 GENOME OF BLUETONGUE VIRUS SEROTYPE-2
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2006-09) BALUMAHENDIRAN, M; SREENIVASULU, D(MAJOR); SATYANARAYANA CHETTY, M; SRILATHA, Ch.
    ABSTRACT: Occurrence of multiple serotypes and presence of antigenic diversity with in serotypes of BTV is the major constraint for developing a protective vaccine. Hence, there is a need to identify and characterize the circulating BTV serotypes in the endemic region. In this direction, a modest study was taken up to amplify, sequence and analyse the phylogenetic relationship of Indian isolate of BTV serotype-2. A pair of primers specific to BTV-2 was designed using VP2 sequences available in Genbank to ampllfy 1797-2932 bp region. Partial amplification of 1135 bp product of VP2 gene of BTV-2 was obtained by RT-PCR and cloning of the amplified product was carried out using pTZ 57R/T plasmid vector. The positive clones were confirmed by colony lysis, PCR, restriction analysis and sequencing. A sequence of 1074 nucleotides were read from the chromatogram. The sequence analysis using BLAST N search showed that the GC content of the gene was 43 percent. Comparison of nucleotide and amino and sequence over the region 1781-2854bp (1074 bases) revealed that Indian isolate BTV-2 was having 66- 86 and 67-86 percent identity respectively with BTV-2 isolates of different regions. Whereas, 55-62 and 3548 percent identity of nucleotide and amino acid respectively with other BTV serotypes from different regions. The multiple alignment of nucleotide and amino acid sequence over the proposed region of VM revealed that there were conserved bases among BTV-2 isolates at position 2170-2223 and 754-767aa respectively. Like wise conserved bases among different serotypes at position 2828-2849 and 916-933aa respectively. The alignment of 1074 bases of VP2 gene of Indian isolate BTV-2 showed 86 percent homology with BTV Taiwan isolate and indicate its closest relationship. Restriction map analysis over the same proposed region revealed that a single conserved SalI site (at 2380bp) was present in both Indian and Taiwan isolate of BTV-2 and confirms the genetic relationship among both isolates. Phylogenetic study also revealed that Indian isolate of BTV-2 more closely related to Taiwan isolate BTVs. Thus, the study of VM sequence comparison revealed that BTV-2 of the same serotypes but from different geographical region were closely related at the nucleotide and amino acid levels and both isolates might have evolved from a common evolutionary pathway. The present study enlightened further approach of developing a recombinant vaccine by expressing the VP2 gene in suitable expression system, so as to develop a suitable desired recombinant vaccine which is the need of hour to help the sheep rearing farmers.
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    ThesisItemOpen Access
    SERO-EPIDEMIOLOGY OF BLUE TONGUE IN DOMESTIC ANIMALS OF ANDHRA PRADESH
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2006-03) JAYAKAR JOHNSON, P; DHANALAKSHMI, K(MAJOR); JANAKIRAMA SARMA, B; NARASA REDDY, G.V
    ABSTRACT: The present study was taken up with the objectives of conducting epidemiological studies on bluetongue in Andhra Pradesh for the years 1995 - 2005, seroprevalence of bluetongue in various species of domestic animals in Andhra Pradesh and Development of an antigen for detection of group specific antibodies to bluetongue. Epidemiological data like morbidity, mortality, case fatality rate due to bluetongue were collected and analyzed. It was observed that outbreaks have been recorded regularly almost on yearly basis with variation in severity. A total of 3853 outbreaks were recorded during the period with an average case fatality rate of 20.93 per cent with a range of 4.47 to 38.67 per cent. Disease was very severe during 1998 (1 132 outbreaks) and 2005 (880 outbreaks) clinically affecting 1,46.861 and 2.72.41 5 sheep respectively. Studies on district wise outbreaks of BT revealed that disease was comparatively more frequent and severe in districts of Krishna, Nellore, Nalgonda and p-. Studies on seasonal incidence of BT indicated that almost all the ou*- (99.58 per cent) were reported during monsoon periods with 64.18 per cent outbreaks during North East monsoon, 35.40 per cent during South wtst monsoon and 0.42 per cent outbreaks during other periods of the year. Attempts were nho made to analyze the &kct of climatological factors like rainfall, relative humidity, temperature and wind speed. Most outbreaks were mrded in September month followed by October and November. No out breaks were recorded during March and May. The reasons were discussed. Results of seroprevalence of bluetongue in Andhra Pradesh on 1058 serum samples indicated that seropositivity was 9.09 per cent in cattle (n=220), 8.25 in buffaloes (n=291), 35.39 in sheep (ns291), 33.47 (~239i)n goats. Seropositivity was very high in sheep and goats compared to cattle and buffaloes. Seroprevalence of BT also revealed that adult sheep above one year had higher scroprcvalence than animals below one year. Further, seroprevalence was higher in Rayalascema and Telangana regions of Andhra Pradesh. An antigen was developed locally to detect group specific antibodies to BT in agar gel precipitation test and compared with a standard kit. Sensitivity and specificity of the antigen developed were 63.26 per cent and 84.37 per cent. The probable reasons for low sensitivity were explained.
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    ThesisItemOpen Access
    EVALUATION OF IMMUNERESPONSES TO ENTEROTOXAEMIA VACCINE IN SHEEP USING ELISA KIT UNDER LABORATORY AND FIELD CONDITIONS
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2006-02) NAGENDRA REDDY, T; SREENIVASULU, D(MAJOR); SATYANARAYANA CHETTY, M; ESWARA PRASAD, P
    ABSTRACT : The present study was undertaken to evaluate the immune responses in sheep vaccinated against Enterotoxaemia using Enzyme Linked Immuno Sorbent Assay (ELISA). The ELISA was standardised (Department of Microbiology, College of Veterinary Science, Tirupati), to detect the protective titers of antiepsilon antibodies in vaccinated sheep using 12 μg/ml of purified epsilon toxin, serum dilution at 1:100 with conjugate dilution of 1:10,000 used in the study. Results were expressed as Per cent Positivity values and Per cent Positivity value 30 is considered as cut off point to determine the protective titers in the serum samples. A total of six sheep maintained under controlled conditions were vaccinated and serum samples were collected periodically, upto 4 months after post vaccination. The mean Percent Positivity values of post vaccinated sera varied from 71.17 to 31.95. The mean Percent Positivity values of the sera collected at one month after vaccination was 71.17 which indicates good protective titer. The titers were gradually reduced. The decrease in the titer from one month to 4 months of post vaccination period was found to be significant (P < 0.05). The mean protective titers were maintained upto 120 days of post vaccination period studied. A total of 139 sera samples from organised farms were collected from the sheep vaccinated against Enterotoxaemia after 2 months of post vaccination and also 247 serum samples from 7 different villages of Kadapa and Chittoor districts of Andhra Pradesh for comparative study. The immune responses were found to be significantly higher in one year age group of animals when compared to other age groups. Variation in immune responses were observed in sheep maintained under different village and farm conditions. Immune responses of sheep maintained under rural conditions were found to be low when compared with sheep maintained under controlled conditions. Duration of immunity in vaccinated sheep was also varied. The protective titers were maintained upto 3 months period in sheep maintained under village conditions. The immune status among the vaccinates usually be measured by mouse neutralisation test which is cumbersome, expensive, time consuming and requires large number of laboratory animals. As an alternative ELISA could be used to determine the immune status and to study the minimum duration of immunity in sheep vaccinated against Enterotoxaemia.