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ThesisItem Open Access EPIDEMIOLOGICAL AND MOLECULAR STUDIES ON CULICOIDES MIDGES IN NORTHERN TELANGANA REGION(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2022) DASMABAIBANOTHU; K. J. ANANDAA study was conducted to know the prevalence and abundance of Culicoides biting midges in Northern part of Telangana of rural and urban districts by both morphological and molecular methods of identification. A total of 28,800 Culicoides midges were collected of which 25,800 (89.58 %) were females and 3,000 (10.41 %) were males. The female fly population was significantly more when compared to with male fly population. The sex ratio was statistically significant. Among the total 28,800 Culicoides midges collected, 12,200 (42.36 %) were C. oxystoma, 9,135 (31.71 %) were C. imicola, 5,950 (20.66 %) were C. peregrinus, 555 (1.92 %) were C. similis; 500 (1.73 %) were C. shivasi; 130 (0.45 %) were C. anopheles; 120 (0.41%) were C. brevipalpis; 110(0.38%) were C. actoni and 100 (0.34%) were C. palpifer. Among all these C. oxystoma and C. imicola were found to be the most predominant species followed by C. peregrinus. and C. similis. In rainy season a total of 15,326 (53.21 %), a total of 10,032 (34.83 %) in winter season and in summer season a total of 3,442 (11.95 %) midges were collected. The highest number of Culicoides was recorded in rainy season followed by winter and it was least during summer. Six species of Culicoides viz., C. oxystoma, C. imicola,C.peregrinus, C. similis , C. anophelis, C. shivasi were confirmed using ITS l region specific primers PanCulF/PanCulR. All the six species got amplified at the range of 400 bp amplicon confirming that all the flies belonged to the genus Culicoides. CO1gene of Culicoides was amplified by universal primers CO1F/CO1R. C. oxystoma and C. peregrinus, C. imicola which yielded specific amplicon size of 658bp. Whereas C. anophelis,C.shivasiand C. similis yielded a specific amplicon at 648 bp. All CO1 sequences were edited and submitted to Genbank and accession numbers were obtained. Finally, barcode was generated using BoldSystems v3, thereby morphologically identified Culicoides species were confirmed by molecular method. Phylogenetic tree was constructed for Culicoide soxystoma, C. peregrinus, C. imicola, C. similis, C. shivasi and C. anophelis spp. The in vitro trials of Metarhizium anisopliae and Bauveria bassiana showed highest mortalities of 82 % and 80 % respectively against larvae. Also, the adult mortality was 66% for Beauveria bassiana and 70 % for (Metarhizium anisopliae). The efficacy range of plant extracts of Annona squamosa and Eucalyptus tereticornis against larvae was from 6.66 % to 40% and 10-50% respectively. The bioassay using Bacillus thuringiensis indicated that the efficacy of Bacillus thuringiensis against Culicoides larvae was 10 % & 26.6 %.when used at 10 ppm and 160 ppm concentration.ThesisItem Open Access STUDIES ON ANTHELMINTIC RESISTANCE TO COMMONLY OCCURRING STRONGYLES IN SHEEP IN HASSAN DISTRICT(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR-585401, 2021-01-01) VEENA M.; G. C. PUTTALAKSHMAMMAThe present research work was carried out to investigate the present status of anthelmintic resistance in Hassan district. A total of 70 randomly selected semi organized sheep farms of Hassan district were included in the study. Questionnaire survey analysis revealed that the farmers have used the anthelmintics irrationally. The anthelmintic resistance to commonly occurring strongyles in sheep was detected by FECRT and EHA. By FECRT, resistance was detected in 15 farms to fenbendazole and six farms to ivermectin. The EHA detected 18 (69%) farms to be resistant to fenbendazole and was more sensitive than FECRT (58%). In the study, Arasikere taluk had more number of resistant farms. Morphological and molecular characterization of predominant larvae in post treatment coproculture confirmed Haemonchus contortus as the predominant gastrointesetinal nematode. The molecular genotyping of β-tubulin isotype-1 gene of H. contortus revealed SNPs at E198A and F200Y, whereas SNP at F167Y was not observed and of T. colubriformis did not reveal the mutation at any position. Upon molecular genotyping of Hco-avr-14b amplicon of H.contortus the polymorphism at 239th nucleotide was observed. The real time PCR revealed that the transcription level of Pgp-2 gene was significantly upregulated in the resistant isolates as compared to Pgp-9.2 gene. LDA and LMIA were used for the evaluation of the effect of anthelmintics along with third generation Pgp inhibitors viz., zosuquidar and tariquidar. The use of third generation Pgp inhibitors increased the efficacy of the existing anthelmintics on the resistant larvae in vitro. Tariquidar had a significant effect than ZosuquidarThesisItem Open Access MORPHOLOGICAL AND MOLECULAR IDENTIFICATION OF TABANIDS IN WESTERN GHATS OF KARNATAKA AND EVALUATION OF FLY TRAPS(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2021-01-01) RAJU METRI; G. C. PUTTALAKSHMAMMATabanids are haematophagous flies of medical and veterinary importance. The study was conducted from March 2019 to February 2020 to record the prevalence of flies in Western Ghats of Belagavi division Karnataka by both morphological and molecular methods of identification. A total of 912 flies belong to twenty species were found to be prevalent viz., T. indianus, T. biannularis, T. diversifrons, T. sexcinctus, T. gertrudae, T. triceps, T. dorsilinea, T. rubidus, T. jucundus, T. auristriatus, T. tenebrosus, H. montana, H. javana, H. longipennis, H. brevis, H. albimedia, C. dispar, C. pellucidus, A. virgo and P. taprobanes. T. jucundus was reported for the first time from Karnataka. The season wise prevalence revealed highest number of species were prevalent during rainy season (17 species) followed by summer (9 species) and winter (4 species). The most diverse genus was Tabanus with 11 species followed by Haematopota with five species, Chrysops with two species and Atylotus and Philoliche with one species each. The morphometric characters of 20 species of tabanids with 10 flies of each species was studied and grouped into large, medium and small flies. The DNA barcoding of twenty different flies was carried out targeting COI a 658 bp amplicon. The sequence of these species after BLAST analysis confirmed the species which was identified based on morphology. The sequence of these species submitted to GenBank and barcode was generated. Phylogenetic analysis of 20 species was carried out and highest and lowest interspecific divergence was found between Philoliche taprobanes and Chrysops dispar (18%); and Tabanus tenebrosus and Tabanus indianus (1%), respectively. A total of 9974 tabanid and Stomoxys flies belonging to eight species were caught by Nzi, modified Manning and H-traps. Nzi trap was superior to the modified Manning and H-traps in collecting tabanid flies. Approximately three times more flies caught by traps baited with fermented (aged) cow urine than unbaited traps. The duration of the olfactory attractive property of fermented (aged) cow urine was tested and it was found to be effective for 12 days.ThesisItem Open Access SYSTEMATIC STUDY ON ARTHROPOD INFESTATION IN SHEEP WITH SPECIAL REFERENCE TO TICKS(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2018-08-01) SUDHA RANI, R.; PLACID E. D’SOUZAA systematic study on arthropod infestation in sheep in Karnataka based on season, breed, management system, agroclimatic zone and site preference was undertaken in nine organized and ten unorganised farms. The prevalence of ectoparasite infestations was 20.7% and 43.7% in organised and unorganized farms. Among all the ectoparasites, prevalence of ticks was highest followed by lice, mites, fleas, nasal bots and flies. The ticks were found resistant to deltamethrin, cypermethrin and were susceptible to amitraz using LPT and AIT-DD test. Among the two EPNs assessed, H.indica was found to be more virulent than S.abbasi. Among the phytoacaricides, Carica papaya seed extract was found to be more virulent than Ricinus communis leaves against ticks by LPT and AIT. By microscopy the prevalence rate of Theileria spp was 16.4% and 64% in organized and unorganized farms respectively. By PCR 24.7 % were positive for T.luwenshuni and 4.7% for T.ovis in organised farms. Also 72 % were positive for T.luwenshuni, 19 % positive for T.ovis and 4 % positive for B.ovis in unorganised farms. Staining of tick tissues with Geimsa revealed 13.5% and 10% of parasitic infection in organized and unorganized farms respectively. Similarly, MGP stain revealed 11.86% and 15.55% infection in organized and unorganized farms. Using PCR, the prevalence of T.luwenshuni in H.Kutchensis was 36% and of T.ovis in H.a.anatolicum was 4% in organised farms. Prevalence of T.luwenshuni and T.ovis was 77% in H.kutchensis and 47 % in H. a. anatolicum in unorganised farms. H.kutchensis ticks was considered as potential vector in disease transmission of T.luwenshuni and T.ovis in sheep in Karnataka. Key words: Arthropods prevalence, sheep farms, acaricides, phytoacaricides, EPNs, vector potentiality.ThesisItem Open Access EPIDEMIOLOGICAL AND MOLECULAR STUDIES ON OVINE THEILERIOSIS FROM SELECTED DISTRICTS OF NORTHERN KARNATAKA(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2019-06-01) SHANTVEER BIRADAR; PLACID E. D’ SOUZAA study was undertaken to observe the prevalence of ovine theileriosis in selected districts of northern Karnataka (South India) belonging to five different agro-climatic zones. The prevalence was found to be 44.95 and 66.66 per cent in sheep by microscopic examination and PCR, respectively. Theileria luwenshuni (389 bp) was found to be the most predominant species with prevalence rate of 84.85 per cent followed by 1.71 per cent of T. ovis (520 bp). The highest rate of infection was observed in Raichur district (73.33%) followed by Kalaburagi (71.42%), Vijayapur (68%), Bagalkot (64%), Yadgir (66.66%) and Belagavi (60.66%) districts and in the age group of 2 to 10 years (68.64%) followed by sheep of 6 months to 2 year age group (58.65%). Females (68.68%) were more commonly infected than the males (64.03%). Bellary breed of sheep had the highest rate of infection with 73.49 per cent followed by 65.62, 65.60, 65.32 and 65.11 per cent in Shahapuri, Deccani, Kenguri and non-descript (ND) breeds, respectively. The statistical analysis revealed non significant (P< 0.05) difference in the prevalence of Theileria spp. between age, gender breed and districts. The level of parasitemia in the present study ranged from 0.8 to 1.2 and 0.2 to 0.4 per cent in clinical and apparently healthy/tick infested animals respectively and the haemogram level ranged from 2.3 to 4.6 g / dl. The coding region of TLSP gene was cloned and expressed in pTZ57R/T vector in E. coli DH5α cells while pRSET-A vector in BL21 (DE3) pLysS cells respectively. The Western blot analysis confirmed the expression of the recombinant protein (35 kDa) using specific Ni-NTA HRPase conjugate, T. luwenshuni positive serum and anti-rTLSP anti-serum raised in rabbit.ThesisItem Open Access DEVELOPMENT OF RECOMBINANT ANTIGEN BASED SERODIAGNOSTIC ASSAY FOR CANINE BABESIOSIS(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2019-06-01) LAVANYA KV; G.C. PUTTALAKSHMAMMAThe present research work was conducted to study the molecular epidemiology of canine babesiosis in Bengaluru and Hassan districts of Karnataka, India and to develop recombinant BgSA3 based I-ELISA for the diagnosis of B. gibsoni infection in dogs. Out of 328 random dog blood samples screened by microscopy, 12.80 % (42/328) found positive for Babesia spp. encompassing 8.53 % (28/328) of B. gibsoni and 4.26 % (14/328) of Babesia vogeli. Cytb and hsp70 gene based PCR assays were developed for detection of B. gibsoni and B. vogeli infection, respectively. The cytb based PCR revealed the prevalence of 10.97 % (36/328) for B. gibsoni and hsp70 based PCR revealed 25 % (32/128) of B. vogeli infection. Mixed infection was found in 3 dogs. The nucleotide sequences of 24 B. gibsoni isolates revealed 98.41 to 98.69 % similarities with the published sequences of B. gibsoni Asian genotype and shared 99.16 to 100 % identity. The phylogeny showed all 24 isolates grouped with B. gibsoni isolates from other Asian countries indicating the monophyletic nature. The sequence analysis of B. vogeli isolates revealed 99.8-99.9 % identity and 99.8 to 100 % similarity with the published sequences. Phylogenetically, all the B. vogeli isolates clustered together with Japan and Taiwan isolates with 97 % bootstrap value indicating genetically similar species circulation in Asian content. Recombinant Δ2BgSA3 based I- ELISA was standardized for the diagnosis of B. gibsoni infection in dogs and showed no cross reactivity with B. vogeli, E. canis, H. canis and D. repens indicating high specificity. The diagnostic sensitivity and specificity of I-ELISA was 86.4 and 93.1 % respectively, in comparison with cytb based PCR. Out of 287 serum samples screened, 34 (11.85 %) were positive for B. gibsoni antibodies. In conclusion, cytb and hsp70 genes can be useful molecular diagnostic and genetic markers, which needs further studies with more number of isolates from different regions of India. The indirect ELISA using rΔ2BgSA3 might be used in clinical diagnosis and in large scale sero surveillance of B. gibsoni infection in dogs.ThesisItem Open Access A SYSTEMATIC STUDY ON TICK INFESTATION IN BOVINES IN THREE DISTRICTS OF H-K REGION OF KARNATAKA(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2019-06-01) KRISHNA MURTHY, C. M.; PLACID E. D’ SOUZAA systematic study on tick infestation of bovines in three districts of Hyderabad- Karnataka region of Karnataka was carried out during February 2018 to January 2019. An overall prevalence of 45.10% tick infestation in bovines including Bidar (49.13%), Gulbarga (42.86%) and Yadgir (44.39%) was observed. The prevalence in cattle and buffaloes was 54.72% and 36.94% respectively. The tick infestation was highest in <1year age group. The females and crossbred animals had higher tick infestation and were highest in monsoon season. Five common species of ticks viz; Rhipicephalus microplus, R. annulatus, R. haemaphysaloides, Hyalomma a. anatolicum, and H. marginatum isaaci were recorded with Rhipicephalus microplus as most prevalent species. AIT-DD showed highest mean % resistance for cypermethrin and deltamethrin in Bidar (28.0% & 34.0%), Gulbarga (36.19% & 40.0%) and Yadgir (24.44% & 30.0%) districts. The LPT revealed the highest mean % resistance for cypermethrin and deltamethrin in Bidar (30.53% & 36.80%), Gulbarga (35.76% & 41.14%) and Yadgir (31.33% & 38.78%) districts. The PCR revealed no mutations either in sodium channel genes or carboxyl esterase genes of field isolates of ticks. Sapota leaf extracts was evaluated for acaricidal efficacy. The AIT showed 100% mortality at 10.0% and 20% concentrations in aqueous extract and 5.0%, 10.0% and 20% concentrations in methanol extracts. An in-vivo study of two species of EPN (Heterorhabditis indica and Steinernema carpocapsae) against R. (B.) microplus was conducted. Out of 10 cattle sheds studied, ticks were found dead only in 2 cattle sheds and in remaining 8 farms no mortality of adult ticks was observed. The non-effectiveness of EPN in cattle sheds could be attributed due to the climatic conditions of the H-K region.ThesisItem Open Access Epidemiological And Immunological Studies With Special Reference To Diagnosis Of Trypanasoma Evansi Infection In Bovines(Karnataka Veterinary Animal And Fisheries Sciences University, Bidar, 2007) P. MalakondaiahTrypanosoma Evansi, A Haemoprotozoan Parasite Of Domestic Livestock Causes A Wasting Disease That Is Better Known Historically As Surra .ThesisItem Open Access Studies On Echinococcus Granulosus In Dogs With Special Reference To Immunological And Molecular Diagnosis(Karnataka Veterinary Animal And Fisheries Sciences University, Bidar, 2007) K.J. AnandaEchinococcosis, A Cyclozoonotic Helminthosis Caused By The Dwarf Dogtapeworm Echinococcus Granulosus Is Highly Endemic And Is Considered To Be One Of The Most Important Parasitic Diseases
