Loading...
![Thumbnail Image](assets/images/Collection.png)
Theses
Browse
7 results
Search Results
ThesisItem Open Access Tick and tick-borne parasitic diseases of dog prevalent in and around Guwahati, Assam(College of Veterinary Science, Assam Agricultural University, Khanapara Campus, 2022) Devi, Pallavi; Islam, SaidulThe study was conducted to record the prevalence of ticks on dog and haemoparasites borne by them. Molecular identification and characterization of the prevalent tick species was also done in order to know their taxonomic status and evolutionary relationship. The study was conducted from March, 2021 to February, 2022. 1440 dogs were examined for the presence of ticks on their body. Out of 1440 dogs, 986 (68.47%) were found to be infested with ticks. 504 (35.00%) ticks were identified as Rhipicephalus sanguineus, 260 (18.06%) were identified as Haemaphysalis bispinosa and 222 (15.42%) were found to be mixed infestation. The highest prevalence of R. sanguineus ticks during August (61%) and the lowest during February and December (25%). Highest prevalence of H. bispinosa was recorded during August (26.67%) and the lowest during February and January (10.00%). Mixed infestation was highest during August (35.0%) and lowest during December (5.0%). The month-wise prevalence was found to be non-significant by statistical analysis (P value: 0.964948). The highest prevalence of ticks was recorded during the monsoon (95.42%) followed by the post-monsoon and the lowest during winter (37.5%). Statistically season has no significant influence on the prevalence of ticks on dogs (P value: 0.6541). The highest percentage of dogs infested with Rhipicephalus and Hemaphysalis tick at the age of 2-3 years (53.49%) and 6 month to 1 year (18.24%) respectively and lowest in 9-10 years (0%). The prevalence of ticks was significantly influenced by the age of dogs (P: 2.28695E-08). Labrador breed of dogs (98.23%) were found to be most infested by ticks followed by Local/mongrel (91.30%) and the least in St. Bernard (7.14%). The breeds of dog were found to be significantly influence the prevalence of ticks (P: 2.1666E-17). The female dogs showed higher prevalence of ticks (52.54%) than their male counterparts (47.46%). The most preferred site of attachment of the ticks on the dog‟s body was ears (28.39%) followed by head (18.56%) and paws/toes (17.64%). Morphological study of both the prevalent tick species were done and characteristic diagnostic features were figured out with photograph. Molecular identification and characterization of prevalent tick species were done by amplification of the 16S rRNA gene by PCR. Phylogenetic analyses by neighbor joining method were performed and the pair wise distance of 16S rRNA were done with MEGA X software. The pair wise distance study revealed that the R. sanguineus strain As/AAU/RS-01 was found to be 100% identical to the isolates of tropical lineages and is 6.4-7.5% divergent with the isolates of temperate lineages. H. bispinosa strain AS/AAU/HB-01 formed the clade with other isolates of H. bispinosa. The closely related isolates of AS/AAU/HB-01 were observed to be Chipahikhola isolate, Itanagar isolate and Arunachal pradesh isolates. The pair wise distance of AS/AAU/HB-01 showed divergence of 0-3.6% with other isolates of H. bispinosa and with other species of Haemaphysalis was between 9.3-14%. 4119 numbers of Giemsa‟s stained thin blood smears were examined under the light microscope for presence of tick-borne hemoparasites. The prevalent of five tick borne haemoparasites in dogs were recorded were Babesia gibsoni (10.34%), Babesia canis (0.92%), Hepatozoon canis (0.39%), Anaplasma (Ehrlichia) platys (0.12%) and Ehrlichia canis (0.22%). Out of 526 positive blood samples, 100 randomly selected positive samples were confirmed with PCR by amplification of 18S rRNA gene for (B. gibsoni, B. canis, H.canis) and 16S rRNA gene for (A. platys and E. canis) and found that PCR shows more sensitivity than light microscopy. Overall prevalence of tick borne haemoparasites, irrespective of season, breed, age and sex is recorded to be highest in the month of July (16.92%) followed by September (16.19%) and lowest in the month of February (4.18%).The highest prevalence of Babesia gibsoni was noticed in Labrador breed (12.93%) followed by local/mongrel dogs (11.53%) while Babesia canis was mostly encountered in Lhasa apso dogs (2.63%) followed by Boxer breed (2.38%) of dogs. Similarly, Pomeranian breed was found to be mostly infected by Hepatozoon canis (1.96%) followed by Doberman (1.55%) dogs. On the other hand, the rickettsial organism Anaplasma platys affect mostly Shitzu (3.85%) dogs followed by Dalmatian breed of dog (2.56%) and Ehrlichia canis was found to affect mostly Lhasa apso (2.63%) followed by Boxer dogs (2.38%). It has been noticed that out of all 18 breeds of dog in the present study, Labrador breed of dog is found to infect most by different haemoparasites followed by local/ mongrel dogs. Female dogs were recorded to be more affected than the male dogs. Statistical analysis revealed that sex of dog has significant influence on the prevalence of tick-borne hemoparasites (P: 0.040134). The highest prevalence of hemoparasites was seen in 1-2 years of age (17.70%) followed by <1 year (16.32%) and 2-3 years (14.88%) of age. The lowest prevalence was observed in 11-12 years of age (2.86%) and mixed infection of Babesia canis and Babesia gibsoni is found to be highest of all other combination of infection.ThesisItem Open Access Efficacy of selected herbal preparations against gastrointestinal nematodes with special reference to haemonchus contortus in goats(College of Veterinary Science, Assam Agricultural University, Khanapara Campus, 2022) Deka, Neelakshi; Phukan, Sulekha ChoudhuryThe present study was carried out to ascertain the prevalence of gastro-intestinal nematode parasites of goats in and around the undivided Kamrup district of Assam and to estimate the anthelminthic efficacy of the three selected indigenous herbal plants viz., Butea frondosa (Polakh), Carica papaya (Omita) and Corchorus fascicularis (Morapat) by in-vitro and in-vivo evaluation in goats naturally infected with gastro-intestinal nematodes. The study was conducted for a period of one-year w.e.f. October 2020 to September 2021 in Kamrup Rural and Metro districts of Assam. A total of 576 faecal samples were collected for the study. Faecal examination by Flotation Technique and Modified McMaster Method was carried out and overall prevalence of gastro-intestinal nematodes was recorded to be 63.89 %. Month-wise prevalence of gastro-intestinal nematode was also recorded where July month recorded the highest prevalence of 91.84 percent and February recorded the lowest prevalence of 31.81 percent. Monsoon season recorded the highest prevalence of 84.28 percent and winter season recorded the lowest prevalence of 35.06 percent. Among the three breeds of goat examined during the present study, Assam Hill goats recorded highest prevalence of 69.80 percent. The 6 months to 1 year age-group animals showed highest prevalence of 76.42 percent and female animals (doe) recorded the highest prevalence of 76.34 percent. The positive faecal samples were put in faecal culture for development of nematode larvae which revealed Haemonchus sp., Trichostrongylus sp., Oesophagostomum sp. and Strongyloides sp. larva in the study area. Polymerase chain reaction (PCR) was employed for molecular identification of Haemonchus contortus which showed a distinct band at 265 bp. Three types of leaf extract viz., ethanolic, hydroethanolic and aqueous extracts of each of three plants were prepared. The percent yield of ethanolic extract of B. frondosa, C. papaya and C. fascicularis was found to be 11.02, 7.81 and 13.72 % (w/w) respectively. For hydroethanolic extract the percent yield was found to be 12.81, 15.19 and 19.60 % (w/w) respectively and the percent yield of aqueous extract was found to be 16.02, 35.28 and 32.87 % (w/w) respectively. Phytochemical analysis of ethanolic, hydroethanolic and aqueous extract of leaves of B. frondosa, C. papaya and C. fascicularis revealed the presence of flavonoids, glycosides and triterpenes. Acute toxicity studies with ethanolic, hydroethanolic and aqueous extracts of the leaves of B. frondosa, C. papaya and C. fascicularis did not show any behavioural change or gross abnormality within 48 hours @ 2000 mg/kg body weight in mice. The extracts were considered to be safe up to a maximum dose of 2000 mg/kg. Three different concentrations viz., 50, 100 and 200 mg/ml was used for all the nine leaf extracts prepared from the three plants. Among the three plants studied in in-vitro trial, B. frondosa was observed as the best plant having in-vitro anthelminthic efficacy. Based on the findings of the in-vitro studies, the hydroethanolic extract of B. frondosa was selected to undergo in-vivo evaluation in goats naturally infected with Haemonchus sp., Trichostrongylus sp., Oesophagostomum sp. and Strongyloides sp. using two doses, 250 and 500 mg/kg body weight. At 250 mg/kg body weight, the percent efficacy was 80.18 % and 72.79 % on Day 7 and 14 respectively, and at 500 mg/kg body weight the percent efficacy was 69.46 % and 73.12 % on Day 7 and 14 post-treatment respectively. The pulverized leaves of B. frondosa was also incorporated in urea-molasses-block (MUMB, Herbal Anthelminthic) and fed to goats naturally infected with Haemonchus sp., Trichostrongylus sp., Oesophagostomum sp. and Strongyloides sp. The anthelminthic efficacy upon feeding of medicated urea molasses block (MUMB, Herbal Anthelminthic) incorporated with B. frondosa was found to have 88.16 % and ii 86.34 % efficacy on Day 7 and 14 post-treatment respectively. Ivermectin was used as the standard anthelminthic drug which gave 96.90 % and 93.79 % efficacy on Day 7 and 14 post-treatment respectively. Haemato-biochemical studies of treated goats under the in-vivo trial showed significant improvement in haemoglobin, packed cell volume, total erythrocyte count and total leukocyte count in groups fed with hydroethanolic extract of leaves of B. frondosa and MUMB (incorporated with B. frondosa) from Day 0 pre-treatment to Day 28 post-treatment. Total protein and albumin concentration improved significantly in groups treated with hydroethanolic extract of B. frondosa. Aspartate transaminase and alanine transaminase was found to improve significantly in groups treated with hydroethanolic extract of leaves of B. frondosa and MUMB (incorporated with B. frondosa) from Day 0 pre-treatment to Day 28 post-treatment. B. frondosa leaf extract and MUMB was found to possess significant in-vitro and in-vivo anthelminthic activity and improved the haemato-biochemical parameters substantially during the present study. This indicates that B. frondosa has the potential to be used as herbal anthelminthic. MUMB with herbal incorporation can be recommended to be used for the control of gastro-intestinal nematodosis in goats. This indicates that the extract of leaves of B. frondosa is a potent and effective herbal anthelminthic. Incorporation of the herbal leaves as powder form in medicated urea molasses block (MUMB, Herbal Anthelminthic) can be used for the control of gastro-intestinal nematodes in goats. The conventional use of synthetic anthelminthics has led to the development of anthelminthic resistance in goat farms not only in Assam but also India and the world as a whole. Therefore, newer anthelminthics need to be developed to overcome the problem of anthelminthic resistance and control of gastro-intestinal nematode parasites of goat.ThesisItem Open Access Epidemiology and molecular identification of trematode parasites of duck with special reference to echinostome(College of Veterinary Science, Assam Agricultural University, Khanapara Campus, 2022-03) Borah, Nanswita; Phukan, Sulekha ChoudhuryA survey was conducted in five different districts of Assam for a period of one year from September‟ 2020 to August‟2021 to investigate on the epidemiology and molecular identification of trematode parasites of duck along with the pathological changes caused by the latter and furthermore, identification of the snail harboring the echinostome cercariae and molecular identification of echinostome cercariae. Out of 1411 fresh faecal samples examined, 411 samples were found positive for helminths with 29.13 per cent prevalence. Infectivity percentage was highest in Kamrup (rural) district and lowest in Udalguri district. Trematodes evinced highest prevalence of 18.00 per cent followed by nematodes with prevalence of 17.51 per cent and finally cestodes with lowest prevalence of 2.62 per cent. Study revealed eggs of Notocotylus sp., Prosthogonimus sp., Echinostoma sp., segments of cestodes, larvae of Strongyloides avium, porrocaecum sp., Tetrameres sp., Heterakis sp. in faecal samples of infected ducks. 303 faecal samples were positive for single infection with prevalence of 21.47 per cent and 108 samples were positive for mixed infection with prevalence of 7.65 per cent. The seasonal prevalence of trematodes on faecal examination of ducks from different districts of Assam depicted highest in monsoon with 28.93 per cent and lowest in winter with 7.82 per cent. The prevalence of trematodes on faecal examination of duck raised under free-range was higher (19.91%) than semi-intensive system of rearing (16.10%). Out of 2786 ducks examined on post-mortem, 2092 ducks were found positive for helminths with 75.09 per cent prevalence. Infectivity percentage was highest in Kamrup (rural) district and lowest in Udalguri district, respectively. Cestodes showed overall highest prevalence of 50.90 per cent followed by trematodes with prevalence of 44.29 per cent and finally nematodes with lowest prevalence of 17.26 per cent. 1146 ducks with an overall prevalence of 41.13 per cent were positive for single infection and 946 ducks with prevalence of 33.96 per cent for mixed infection. On the basis of morphological studies, 9 different species of trematodes, 7 different species of cestodes and 4 different species of nematodes were identified and recorded. Trematodes were identified as Echinostoma revolutum (7.11%), Echinostoma paraulum (18.23%), Echinostoma friedi (0.29%), Hypoderaeum conoideum (18.52%), Psilochasmus oxyurus (0.04%), Tracheophilus cymbius (0.04%), Prosthogonimus ovatus (0.04%), Notocotylus attenuates (0.04%). Cestodes were identified as Hymenolepis collaris (14.28%), Hymenolepis lanceolata (11.59%), Fimbriaria fasciolaris (7.93%), Raillietina tetragona (4.74%), Raillietina echinobothridia (5.03%), Raillietina cesticillus (3.30%), and Amoebotaenia cuneata (4.02%). Nematodes were identified as Porrocaecum sp. (0.68%), Heterakis gallinarum (5.53%), Heterakis dispar (4.02%), and Tetrameres sp (7.04%). Notocotylus attenuates and Amoebotaenia cuneata was first time reported from Assam in duck. The prevalence of trematodes on post-mortem examination of ducks on the basis of sex depicted higher prevalence in females (46.00%) than males (42.62%). The prevalence of trematodes in post-mortem examination of ducks on the basis of age showed higher in adults (48.93%) than young ducks (38.95%). The prevalence of trematodes on post-mortem examination of ducks on the basis of breed showed highest in Pati ducks (49.24%), followed by Khaki Campbell ducks (45.76%), Chara-chambelli ducks (42.14%) and lowest in White pekin ducks (39.16%). Seasonal prevalence of trematodes on post-mortem examination of ducks depicted highest in monsoon with 49.62 per cent and lowest in winter with 38.25 per cent. Prevalence of trematodes on ii post-mortem examination of ducks on the basis of rearing duck showed higher in free-range (47.44%) than semi-intensive system of rearing (40.91%). For molecular identification of trematodes namely, Echinostoma revolutum, Echinostoma paraulum, Hypoderaeum conoideum and Echinostoma friedi (morpholocigally identified) were subjected to PCR (COX -1 gene of Echinostoma revolutum, Echinostoma paraulum, Hypoderaeum conoideum and ITS2 gene of Echinostoma friedi). The trematode, Echinostoma revolutum collected from different districts on the basis of COX-1 sequence identity showed 97.80-99.63% identity with Echinostoma revolutum, Echinostoma paraulum collected from different districts on the basis of COX-1 sequence identity showed 99.07-99.54% identity with Echinostoma robustum, Hypoderaeum conoideum collected from different districts on the basis of COX-1 sequence identity showed 99.08%-99.74% identity with Hypoderaeum conoideum. Echinostoma friedi on the basis of ITS2 sequence identity from Kamrup (rural) and Kamrup metropolitan districts showed 100% identity with Echinostoma friedi. DNA extracted from adult trematode parasites yielded positive results for COX-I genes of E. revolutum, E. paraulum and Hypoderaeum conoideum by PCR analysis, yielded the amplicons size of 818 bp, 216 bp, and 765 bp, respectively and 1000bp for E. friedi for ITS2 gene. On post-mortem examination, intestine of ducks infected with the species under the genus Echinostoma and Hypoderaeum showed slight gross lesions except catarrhal enteritis and congestion in the mucosal surface, mainly in the jejunum, ileum and caecum. Psilochasmus oxyurus showed catarrhal enteritis and congestion in the mucosal surface of the small intestine. Trachea infected with Tracheophilus cymbius exhibited bloody mucosal exudates in the site of attachment. No observable gross pathological lesions could be seen in oviduct and caecum affected with Prosthogonimus ovatus and Notocotylus attenuates, respectively. This may be due to low level of infection encountered in the present investigation. Microscopic changes associated with the species under the genus Echinostoma, Hypoderaeum, and Psilochasmus showed marked thickening with infiltration of inflammatory cells including lymphocytes, eosinophil, plasma cells and macrophages in the intestine. Some of the villi were degenerated and necrosed. Desquamated epithelial lining of intestinal villi were found to be deposited in the lumen of the intestine. Haemorrhagic patches could be seen in the tip of some villi. Hypertrophy of goblet cells was seen in the villi. The glandular epithelium showed vasculation and distortion which have further undergone necrosis. Most of the lymphoid follicles in ileum were found to be enlarged and degeneration of lymphoid cells was also observed. Muscle cells showed necrosis and hyalinization. Blood vessels showed congestion and dilation along with haemorrhages in subserosal layer of intestine. Histopathology, of trachea infected with Tracheophilus cymbius showed heavy infiltration of leucocytes in the mucosa and degeneration with necrosis in submucosal layer. Snail, Indoplanorbis exustus (0.55%) and Lymnaea luteola f. ovalis (0.37%) were recorded as the intermediate host of echinostome cercariae. No cercarial infection was recorded from Lymnaea acuminata f. rufescens, and Bellamya bengalensis f. typica. District-wise Monthly prevalence of echinostome cercaria in Indoplanorbis exustus depicted maximum shedding from the month of June to September from the different districts of Assam. District-wise Monthly prevalence of echinostome cercaria in iii Lymnaea luteola f. ovalis depicted maximum shedding from the month of June to October from the different districts of Assam. Morphological identified echinostome cercariae (4 representative samples) collected from different districts of Assam were subjected for molecular identification where ITS2 and COX -1 gene of the parasite was subjected to PCR. Cercariae of Hypoderaeum conoideum and Echinostoma revolutum (molecularly identified) are commonly prevalent in the snails of Assam.ThesisItem Open Access DIGESTIVE TRACT PROTOZOAN PARASITISM IN DOMESTIC BIRDS WITH SPECIAL REFERENCE TO Trichomonas gallinae IN ASSAM(College of Veterinary Science, Assam Agricultural University, Khanapara, Guwahati, 2019-07) SAIKIA, MUNMI; Bhattacharjee, KantaThe present study was undertaken to ascertain the prevalence of protozoan parasites inhabiting the digestive tract of domestic birds which included pigeon (Columba livia domestica), chicken (Gallus gallus domesticus), duck (Anas platyrhynchos domesticus) and quail (Coturnix coturnix japonica) in the state of Assam, India. The study was conducted for a period of two years w.e.f. April 2017 to March 2019 in 8 districts of Assam viz. Kamrup (Rural and Metro), Dhubri, Barpeta, Nalbari, Darrang, Baksa, Lakhimpur and Dhemaji. A total of 1278 pooled faecal samples and 1207 throat swabs were collected for study. Faecal examination by floatation method for the presence or absence of oocyst of coccidia and by Modified Ziehl- Neelsen staining and Kinyoun’s staining method for detection of Cryptosporidium infection was carried out in domestic birds and overall prevalence of digestive tract protozoa was recorded as 34.66%. Identified species were Eimeria labbeana (26.35%), E. columbarum (8.69%) and E. columbae (3.53%) from pigeon; Eimeria tenella (19.13%), E. acervulina (9.46%), E. necatrix (6.88%), E. maxima (2.36%) and Cryptosporidium baileyi (3.01%) from chicken; Eimeria battakhi (19.86%) from duck and Eimeria tsunodai (13.29%), E. bateri (5.69%), E. uzura (3.16%) and Cryptosporidium meleagridis (4.43%) from quail. Season wise, highest prevalence was recorded from pigeon in pre monsoon (58.33%) and lowest in monsoon (27.17%); in chicken highest in monsoon (57.00%) and lowest in pre monsoon (28.69%); in duck highest in winter (52.45%) and lowest in post monsoon (17.28%); in quail highest prevalence was seen during monsoon (61.11%) and lowest in winter season (14.28%). District wise, highest prevalence (80%) was recorded from Kamrup (rural and metro) and lowest from Dhemaji (22.22%) in pigeon; in chicken highest (82.60%) from Dhubri and lowest from Lakhimpur (28.16%); in case of duck and quail highest prevalence was recorded from Dhubri (51.72%) in duck and (45%) in quail and lowest percentage was recorded from Baksa (22.22%) and (17.85%) from Darrang respectively. Observation on the prevalence of T. gallinae was done by Giemsa staining and culture and overall prevalence was recorded as 28.91%. In pigeon, the prevalence was recorded as high as 71.12% and in chicken it was 6.25% but T. gallinae was not recorded from both duck and quail in natural condition. In pigeon, prevalence was found in squab as 79.47% which was the highest. In young bird, it was 61.11% and in adult, prevalence was 70.00%. Female birds showed a prevalence rate of 75.51% while in male, it was 66.36%. In chicken prevalence rate was 6.73% in females and 6.00% in males. Season wise, highest number of cases (87.12%) in pigeon was recorded in winter and lowest in monsoon (60.58%). In chicken, T. gallinae infection was recorded only in two seasons; post monsoon showed slightly higher prevalence (15.49%) than winter (13.79%). District wise, maximum number of positive samples (78.65%) was recorded from Kamrup (rural and metro) in pigeon and in chicken, highest prevalence of Trichomonas infection was reported from Baksa district (34.78%). Comparative evaluation of direct smear (Giemsa staining) and culture methods (Wet mount) for detection of T. gallinae revealed culture method to be sensitive and superior to direct smear method. Five media, viz. modified Diamond’s media, Medium199, Minimum Essential Medium (MEM), RPMI 1640 and Nutrient broth were used for culture and maintenance 3 of T. gallinae parasite. Medium 199 showed the highest growth of organism upto 144-168 hours with motile trophozoites in comparison with other four media in the present study. Polymerase chain reaction (PCR) employed for amplification of Fe hydrogenase gene of Trichomonas gallinae from positive cultured materials of pigeon and chicken showed clear 290 bp band fragment. Molecular characterization of T. gallinae from pigeon and chicken isolate of Assam in the present study showed 100% similarity with isolates of Iran and Austria. To determine the virulent nature of the trichomonads and its transmissibility to different hosts, experiment was performed with or without immunosuppressive drugs in chicken, duck, quails and mice taking pigeon as its natural host. Pigeon strain of T. gallinae orally inoculated at concentration of 4x104 trophozoites in birds and intra peritoneal inoculation in mice revealed presence of parasites in all bird species while mice developed abscess which is an indicator of T. gallinae infection. Pathological lesions like yellowish to whitish masses of caseous necrotic materials were seen in the beak, oral cavity, oeosophagus, crop and proventriculus of pigeon and mild gross alterations were observed in other experimental host. Histopathological alterations were also found more in pigeon than other infected birds. In the present study, five different drugs used for in vitro and in vivo efficacy against T. gallinae were Flagyl 400 (Metronidazole), Ornida (Ornidazole), Tiniba 300 (Tinidazole), Sulcoprim (sulphadiazine and trimethoprim) and Vetfur-TL (Metronidazole, Furazolidone and Loperamide). Concentration of drugs @10, 20 and 30 mg/ml for in vitro study and 20 and 30 mg/kg for in vivo study revealed 100% efficacy of the drugs Metronidazole, Ornidazole and combination of Metronidazole, Furazolidone and Loperamide at 30mg/ml in vitro and 30mg/kg in in vivo condition.ThesisItem Open Access STUDIES ON Dirofilaria immitis IN DOGS AND ITS ASSOCIATION WITH Wolbachia SPECIES(College of Veterinary Science, Assam Agricultural University, Khanapara, Guwahati-781022, 2013-06) Borthakur, Sonjoy Kumar; Deka, Dilip Kr.Dirofilaira immitis is an important canine filarial nematode. An epidemiological study was carried out to record the prevalence of D. immitis in dogs in two different geographical locations viz., Guwahati, Assam and Aizawl, Mizoram of North Eastern Region in India, from February, 2011 to July, 2012. The study also included to evaluate the persistence of Wolbachia endosymbiont with D. immitis. In the present study, dogs were grouped into three categories, i.e., stray, pet and working dogs, their respective numbers being 413, 266 and 103 irrespective of the study regions. Three different methods were used for the study, i.e., microscopy (wet film and KCT), immunological (Ag ELISA by SNAP®4Dx kit) and molecular techniques (PCR). The study revealed overall heartworm prevalence in Guwahati to be higher (18.23%) than in Aizawl (17.68%) irrespective of categories of dogs. Sex-wise, the infection was higher in male (18.12%) than in female (17.90%), though the difference was statistically non-significant. The overall efficacy percentage for detection of heartworm by wet film, KCT, Ag ELISA and PCR test revealed 6.26, 11.38, 18.03 and 13.93 percent, respectively. Ag ELISA test was found to be the best amongst the three types of tests compared. Using molecular tools, prevalence of D. immitis in dogs was 13.52 percent in Guwahati and in Aizawl was 14.62 percent. With PCR, 4 cases of D. repens could be diagnosed in stray dogs from Guwahati. The study revealed overall 22.69 percent occult infection, of which, highest cases were recorded in working dogs (60%). Occult infection was calculated by finding the difference between heartworm prevalence based on Ag ELISA and PCR test. Dot ELISA test using monoclonal antibody of D. immitis for detection of heartworm antigen in dog blood samples was standardized. The test revealed 72% specificity against known positive D. immitis blood samples at SNAP®4Dx commercial kit. Molecular technique using PCR was standardized to detect D. immitis using published primers with slight modification of thermal condition. Two different primers were used viz., specific primers for D. immitis only and another, pan filarial primers for detecting six different canine filariids. Both the primers resulted desired amplification product size against different filarial parasites. Molecular cloning and characterization of D. immitis for ITS-2 region of Guwahati isolates were conducted. The results showed the Guwahati isolates had a close relationship with that of South Asian isolates of D. immitis. Pair-wise homology analysis revealed 98.6 - 98.9% similarity with a few sequences available at NCBI GenBank. Similarly, phylogenetic analysis of D. repens encountered in Guwahati isolate was also done. Endosymbiont Wolbachia association with D. immitis worm as well as in heartworm infected blood was revalidated by PCR method. The findings were substantiated with the presence of the organisms in the worm’s lateral cord by fine structural studies conducted through transmission electron microscopy (TEM). Molecular evidence followed by sequence analysis of Wolbachia revealed 99.4 to 99.8% similarities with other sequences available in NCBI GenBank for Wolbachia endosymbiont of D. immitis. Finding of the present studies establish the endemicity of D. immitis in North East India and validates the association of Wolbachia endosymbiont in D. immitis. Record of D. repens warrants further detail studies owing to its zoonotic significance.ThesisItem Open Access EPIDEMIOLOGY OF LEECH INFESTATION AND EVALUATION OF ANTI LEECH EFFECTS OF SOME LOCALLY AVAILABLE PLANTS(Assam Agricultural University, Khanapara, Guwahati, 2016-07) PATHAK, PALLABI; Islam, SaidulAn epidemiological study was conducted to record the prevalence of aquatic and terrestrial leech species in fifteen districts of Assam. Out of 1991 number leeches collected five aquatic e.g., Hirudinaria manillensis (57.86%), H. granulosa (11.85%), H. javanica (2.71%), Whitmania laevis (2.61%) and Hemiclepsis marginata (0.80%) and one terrestrial leech e.g., Haemadipsa sylvestris (24.16%) was found prevalent in the study area. Molecular identification of H. manillensis based on coI sequence was done. Phylogenetic analysis showed 95.5% sequence similarity with P. manillensis (Accession No: KT693108.1) and lowest with H. manillensis (GQ368747.1). The sequence was submitted into the GenBank, and assigned the Accession No. GenBank KX579976. Bionomics of H. sylvestris was studied in laboratory conditions. The incidence of leech infestation in man and animal in different leech infested areas was conducted using interview method by means of questionnaire. Total protein concentration of crude antigen of H. granulosa, H. sylvestris and H. manillensis was found to be 4.14g/dl, 4.05g/dl and 3.6g/dl. Total protein concentration of E/S antigen of H. manillensis, H. granulosa and H. sylvestris was 3.39g/dl, 3.44g/dl and 3.4g/dl, respectively. Discontinuous gel electrophoresis revealed protein profile of crude antigen of H. manillensis, H. granulosa and H. sylvestris being almost same. The number and size of peptides varied from 9-10 and 12.5 to 96 kDa, respectively. The number of peptides of E/S antigen for H. manillensis, H. granulosa and H. sylvestris were 5 and size varied from 14-96 kDa. PAS staining of glycoprotein antigen could not be detected for E/S product for all three species. Two glycoprotein antigens could be detected in crude H. manillensis and H. granulosa and three in H. sylvestris which ranged from12.5-26 kDa. A total of two immune-reactive peptides were detected for H. manillensis and H. granulosa crude antigens and one in E/S product of H. sylvestris. Hirudinicidal effect of aqueous and methanolic extracts of leaves of Nicotiana sp., seeds of Camellia sinensis and stems of Carica papaya was evaluated against H. manillensis and compared with ivermectin and nicotine. The methanolic extract of Nicotiana sp. and C. sinensis; aqueous extract of Nicotiana sp. and C. sinensis possessed effective hirudinicidal properties. While the concentration and extraction procedure was considered, the aqueous extract of Nicotiana sp. at 5000 mg/ml and 2500 mg/ml showed faster killing effect. While the method of extractions and the level of concentrations were considered against time of repellency, it was found that barring the methanolic extract of C. papaya at the concentrations of 50 mg/ml and 100 mg/ml, all the other extracts at different concentrations showed highly significant (p<0.001) leech repellent effects. Expression of HSP was evaluated by Dot-ELISA in stressed organism using a panel of monoclonal antibodies which comprised of HSP-60, HSP-70 and HSP-90. After application of pure compounds (nicotine and ivermectin) and extracted products (methanolic and aqueous) of Nicotiana sp., C. sinensis and C. papaya, HSP-60, HSP-70 and HSP-90 were detected in excretory and secretory (E/S) product of leeches. HSP-60, HSP-70 and HSP-90 were also detected both in crude and excretory and secretory (E/S) products of the cold stressed leeches. Extracts of Nicotiana sp. and C. sinensis were subjected to in vitro cytotoxicity studies on Vero cell lines using MTT assay. Two fold dilution of the extracts were used from 10.00 µg/µl up to 0.0096 µg/µl. Both the aqueous and methanolic extracts of Nicotiana sp. and metahnolic extract of C. sinensis had more per cent viability of Vero cells than the aqueous extract of C. sinensis. In no case the per cent viability was below 70. The efficacy of extracts against H. sylvestris and aquatic leech was evaluated on human volunteers. The methanolic extract of C. sinenesis failed to exhibit minimum 50% leech repellent effect. Aqueous extract of C. sinenesis exhibited inconsistent effect with 45.83% repellent effect at 20% concentration, but 26.19% effect at 30% concentration. Both the methanolic and aqueous extract of Nicotiana sp. exhibited 100% repellent effect at all the three levels of concentrations which was comparable with DEET (12% concentration).ThesisItem Open Access STUDIES ON Indoplanorbis exustus AND ITS ASSOCIATED SCHISTOSOMES(Assam Agricultural University, Khanapara, Guwahati, 2016-07) Bulbul, Kamal Hashan; Das, ManoranjanA study on ecology, biology and bionomics of Indoplanorbis exustus along with its associated schistosomes was undertaken for a period of two years i.e. from March 2014 to February 2016 in Barpeta, Nalbari and Kamrup district of Assam. A total of 12 aquatic macrophyte species were recorded from the study area. The highest frequency percent (f %) relative frequency percent (RF %) of different macrophytes was found to be 86.67% and 19.70% respectively. Population density of I. exustus in terms of f% and RF% and man hour collection per meter square area (MHC/m2) showed an increasing trend from July to September and then gradually declining towards November. While the snails were reared in biologically balanced aquarium at constant temperature of 15, 20, 25, 30 and 35C and room temperature (20.02-31.75C), all the snails died before attainment of sexual maturity at 15oC. The growth rate in terms of shell diameter and body weight attained maximum size 10.63+0.162 mm and 435.83+23.367 mg, respectively at room temperature at 6th week of age. Egg to egg cycle ranged from 50 days at 30C and 125 days at 20C. Room temperature was found to be more conducive for fecundity wherein as many as 22833 numbers of eggs were hatched out from 1033numbers of egg capsules. The temperature, pH, DO, free CO2 and total alkalinity of ambient water had direct bearing on population density of snails. The ANOVA of physicochemical properties of water was highly significant (p<0.01) between months and areas of water bodies. Out of 161500 numbers of snails collected in two years showed incidence rate of cercariae of Schistosoma spindale (0.72%), S. indicum (0.50%) and S. nasale (0.17%). The cercariae were identified on the basis of morphological and molecular method. The RS data indicated that the distribution of cercariae of S. indicum group was more in the northern bank of Brahmaputra compared to the Southern bank. By means of molecular dissection, brevifurcate cercaria was identified as Trichobilharzia sp. and the longifurcate as Alaria alata with 87-88% clonal relationship with Denmark, Lithuania and Germany isolates. A total of 420 cattle, 179 buffaloes and 171 goats slaughtered at local abattoirs revealed visceral schistosomosis in 37.38, 35.20 and 12.28%, respectively. In regards to coprological examination it was 14.52, 12.85 and 4.68% in order of same sequence as above. The incidence rate in terms of adult parasites present and faecal examination was the highest in monsoon season and the lowest in pre-monsoon season. Concentration of eggs was more in liver (45.45%) followed by ileocaecal junction (37.50%), caecum (25.97%) and small-intestine (18.18%). Based on worm count methods, mild types of intensity was found to be highest followed by moderate and heavy intensity in slaughtered animals. The incidence of nasal schistosomosis was highest in July (18.92%) and lowest in December (2.70%) when nasal swab was examined for the presence of eggs in 363 cattle irrespective of sex and age. Like visceral schistosomosis, nasal schistosomosis also had a higher infection rate in monsoon season and in animals above 8 years of age. The second internal transcribed spacer (ITS2) sequence of S. spindale, S. indicum and S. nasale were found to be amplified showing different repeatitive band patterns. While the mitochondrial cytochrome c oxidase subunit 1 (CO1) genes and the ribosomal gene repeat, part of the 28S RNA gene (28S) were amplified on 372 bp and 1225 bp when subjected to PCR. Phylogenetic analysis of Schistosoma indicum group based on nucleotide sequences of COI and 28S genes revealed Assam isolates of S. indicum showed a clonal relationship with Bangladesh isolates and S. spindale and S. nasale with Nepal isolates. RAPD-fingerprinting using different random primers showed specific polymorphic markers for susceptible and non-susceptible I. exustus to Schistosoma infection due to genetic variability.