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Open Access
Studies on the protective effects of medicinal plant extract against Escherichia coli in poultry
(CSK HPKV, Palampur, 2022-12-21) Choudhary, Sahil; Asrani, R.K.
The present experimental study was planned to identify the most potent plant extract through in vitro studies and then test the most effective plant extract against experimental E. coli (O101) organism intraperitoneally in Delham Red (DR) chicks to evaluate the effects on clinical signs, mortality, body weight, serum biochemical parameters and pathological changes. The 70% aqua-ethanolic extract of Artemisia annua (A. annua) showed maximum antibacterial effect against experimental E. coli (O101) organism in vitro. The pilot experiment was conducted to determine the LD50 dose which was found 3.6 x 107 cfuml-1 . In the experiment, a total of 300, day-old Delham Red (DR) birds were randomly divided into 6 groups. The group CX acted as control group, group EX was given E. coli infection only, groups EA, EB and EC were administered with both E. coli and 70% aqua-ethanolic extract of A. annua at the dose rate of 0.5 g, 1.0 g and 2.0 g per litre of water, respectively. The group AX was provided with 70% aqua-ethanolic extract of A. annua only at the dose rate of 2 g/l of water. E. coli (O101) infection was given to the birds intraperitoneally on 7th day of age. The 70% aqua-ethanolic extract of A. annua was given in drinking water to birds from 0 day to day 14. The birds from each group were sacrificed at 1, 3, 5, 7, 10 and 14 day post infection (DPI). The findings of the present study revealed that group EX (E. coli infection only) exhibited clinical signs of reduced feed and water intake, dullness, depression, huddling, ruffling of feathers, reluctance to move and whitish watery diarrhoea. No clinical signs were observed in group CX and AX. The values of biochemical parameters such as ALT, AST and creatinine were increased, whereas concentrations of total protein and albumin were decreased in group EX (E. coli infection only) as compared to the group CX (control group). However, the clinical signs and serum biochemical values in the groups EA, EB and EC were significantly lower in dose-dependent manner as compared to group EX (E. coli infection only). Mortality was highest in group EX (E. coli infection only) in comparison to treatment groups i.e EA, EB and EC. The body weight was higher in group EC and in comparsion to that in group EX. The gross pathology comprised of fibrinous perihepatitis, fibrinous pericarditis, airsacculitis, splenomegaly and peritonitis with higher severity in group EX and there was significant reduction in the gross lesions in groups EA, EB and EC in dose-dependent manner. Microscopically, the liver and heart of group EX showed severe perihepatitis, pericarditis, vacuolar changes, leukocytic infiltration, degenerative changes and enhanced cytoplasmic granularity. Similarly in group EX, the microscopic lesions in the spleen were characterized by reticuloendothelial cell hyperplasia and an increase in eosinophilic coagulum material. Airsacs exhibited varying degree of fibrinous airsacculitis.The microscopic lesions in the liver, heart, spleen and airsacs were of less severity in group EA, EB and EC which were attributed to antibacterial effect of plant extract use in the present study.
Open Access
Studies on pathology of Escherichia coli infection in bovine calves and exploration of in vitro antimicrobial potential of certain plant extracts
(CSK HPKV, Palampur, 2022-12-17) Priyanka; Patil, R.D.
The present study was conducted to determine the pathology caused by Escherichia coli infection in bovine calves in Himachal Pradesh. A total of 222 fecal samples were collected from dead, clinically sick and apparently healthy calves upto one year of age. Necropsy examination of 30 calves upto one year of age including 24 males and 6 females was done during the time period of 1 June, 2021 to 30 June, 2022. Grossly, enteritis was present in 26 (86.67%) calves, ulcerative abomasitis in 5 cases (16.67%), diptheretic enteritis in 1 case (3.33%), milk rumenitis in 1 case (3.33%), nodular abomasitis in 1 case (3.33%) and haemorrhagic typhlitis in 1 (3.33%) case. Histopathologically, vascular changes (congestion, edema, haemorrhages etc.), villous denudation and sloughing, cryptal necrosis, hyperplastic activity of enterocytes, increased goblet cell activity and infiltration of inflammatory cells were the consistent findings. Along with this, macroshizonts and microschizonts of coccidia in tips of villi, bacterial colonies, depletion of lymphocytic element from Peyer’s patches and proliferation of fibrous connective tissue in Peyer’s patches were also found. E. coli was found to be the major cause for gastrointestinal pathologies (55.78%) and responsible for deaths among calves. The prevalence of E. coli in Himachal Pradesh was found to be 47.75% out of which pathogenic E. coli contributed to 40.1%. Molecular pathotyping revealed that enteropathogenic E. coli (EPEC) was present in 57/222 (25.67%) samples, enterohaemorrhagic E. coli (EHEC) was 31/222 (13.96%), enterotoxigenic E. coli (ETEC) was 28/222 (12.61%) and enteroaggregative E. coli (EAEC) was present in 80/222 (36.04%) fecal samples. 25 different serotypes of all the 106 E. coli isolates were identified with serotype O119 (12.26%) being most common followed by O98 (10.38%) and O126 (9.43%). CST for 106 isolates of E. coli against 16 commonly used antibiotics revealed that highest resistance was shown by Cloxacillin (58.49%) followed by Furazolidone (53.77%). 21 E. coli isolates were found to be resistant against one antibiotic group, 17 were resistant against two antibiotic groups, 26 were found to show resistance against three antibiotic groups and rest 42 samples were resistant to more than three antibiotic groups. Upon screening of 15 plant extracts (50% aqua- alcoholic) for their antimicrobial potential against 6 serotypes of multidrug resistant (MDR)-E.coli, five plant extracts i.e. Artemisia nilagirica (C.B. Clarke) Pamp., Asparagus adscendens Roxb., Phyllanthus urinaria, Physalis peruviana Linn. and Tinospora cordifolia were found to have zones of inhibition of varying diameters at different concentrations. In conclusion, the present study revealed that E. coli infection leading to enteritis was the major cause of death in bovine calves upto 1 year of age in Himachal Pradesh. The plant extracts seem to be future alternative to counteract the antimicrobial resistance in animals and humans.
Open Access
Studies on the protective effects of medicinal plant extract against Salmonella infection in poultry
(CSK HPKV, Palampur, 2022-12-13) Soni, Mridul; Asrani, R.K.
The present experimental study was designed with an aim to identify the most potent plant extract through in vitro studies and then subjecting the most potent plant extract against experimental Salmonella enterica serovar Typhimurium 4, 12:i:1,2 infection in Delham Red birds to observe the effects on clinical signs, mortality, growth response and pathological changes. The 80% methanolic root extract of Saussurea costus showing maximum antibacterial potential was used in the experimental trial. LD50 dose of Salmonella enterica serovar Typhimurium was found to be 2x 106 cfu ml-1 during pilot experiment using intraperitoneal route. Day old three hundred Delham Red birds were procured and were randomly divided into six groups : Group (CX) - plain control; Group (DX) - Salmonella Typhimurium infection only; Group (SA) - Salmonella Typhimurium infection + 80% methanolic root extract of Saussurea costus @ 0.5g/l; Group (SB) - Salmonella Typhimurium infection + 80% methanolic root extract of Saussurea costus @ 1g/l; Group (SC) - Salmonella Typhimurium infection + 80% methanolic root extract of Saussurea costus @ 2g/l; Group (SD) -80% methanolic root extract of Saussurea costus @ 2g/l only. The methanolic root extract of Saussurea costus was provided in the drinking water as early as 4th day of age followed by infection at 7th day of age. Sacrifice of the birds in different groups was done at regular intervals i.e.1, 3, 5, 7, 10, 14 & 21 DPI was done. The clinical signs recorded in the plain infection group included huddling near the light source, dullness, loss of appetite, soiling of the vent and drooping of head and wings. There were no clinical signs in the plain control group and the intensity of signs among infected groups was least in the group treated with highest dose of extract as compared to other groups. Mortality rate was found to be highest in group DX in comparison to other groups SA, SB & SC and no mortality was recorded in the groups CX & SD. Body weight gain was minimal in the group DX and highest in the group SC treated with the highest dose of plant extract. The liver and kidney parameters (ALT, AST, total protein, albumin and creatinine) among different treatment groups SA, SB & SC were significantly generally lowered as compared to the group DX. The gross pathology comprised of hepatomegaly, necrotic foci, liver paleness, splenomegaly and fibrinous pericarditis were reduced in Saussurea costus treated group as compared with group DX. Microscopically the liver in group DX comprised of severe multifocal areas of necrosis, infiltration of heterophils and lymphocytes, sinusoidal dilation, degenerative changes and increased cellular granularity. Severe homogenous eosinophilic areas of necrosis, reticuloendothelial cell hyperplasia, depletion of lymphoid follicles and blood-filled sinuses were present in the spleen. The microscopic lesion score intensity was highest in group DX on 5, 7 & 10 DPI. The similar lesions were observed in the treatment groups SA, SB & SC but a significant reduction in the gravity of the lesions was observed among these groups. Preservation of the normal structure of spleen and liver was observed among treatment groups suggesting the antimicrobial and hepatoprotective effect of Saussurea costus against the pathology produced by Salmonella enterica serovar Typhimurium in experimental bird model.
Open Access
Studies on pathology of respiratory viral diseases in bovine calves
(palampur, 2022-09-24) Singh, Rahul; Patil, R. D
The present investigation was conducted to study the pathology of pneumonia and to identify the possible respiratory viral etiology in bovine calves under one year of age during 1st June 2021 to 30th June 2022. A total of 30 calves were necropsied for studying pathology of pneumonia by help of cytological, gross, histopathological and immunohistochemical examination. To detect viral agents, 228 samples were collected from necropsy (30 tissues), clinically sick calves (130 including 121 nasal and 9 ocular swabs) and apparently healthy calves (68 nasal swabs). In addition, 221 samples were gathered from clinically sick calves (121 nasal swabs), apparently healthy calves (68 nasal swabs) and necropsy (32 including 30 nasal and 2 lung swabs) for isolation and identification of associated bacteria. Pneumonia was found to be the main cause of death in bovine calves under one year of age. Male calves showed maximum mortality i.e., 80%. Maximum mortality was observed in 3-12 months of age group with 24/30 (80%) cases followed by 1-3 months of age group with 4/30 (13.33%) cases and least mortality was observed in <1 month of age with 2/30 (6.67%) cases. Pneumonia present in all of 30 necropsied calves revealed different pattern with bronchointerstitial pneumonia in 14/30 (46.67%), suppurative bronchopneumonia in 4/30 (13.33%), fibrinous bronchopneumonia in 5/30 (16.67%) and interstitial pneumonia 7/30 (23.33%) cases. In bovine respiratory syncytial virus (BRSV) and bovine parainfluenza virus type 3 (BPIV3) positive cases necrotising bronchiolitis and bronchitis, multiple number of syncytial cells in alveolar lumen and infiltration of inflammatory cells mainly lymphocytes were seen. Using AgELISA, the prevalence of each of the bovine herpes virus-1 (BoHV) and BRSV was 56.67%, BPIV3 46.67% and bovine viral diarrhoea virus (BVDV) 26.67% in necropsy cases. Using AgELISA, 5/33 (15.15%) samples tested positive for all the 4 tested viruses, 4/33 (12.12%) tested positive for the concomitant 3 viruses and 11/33 (33.33%) were positive for 2 viruses. The prevalence of BRSV was found to be 50% (15/30) in necropsy cases and 23.73% (47/198) in clinical samples using nested RT-PCR of Nucleoprotein (N) giving a product size of 435 bp after secondary PCR. The maximum positivity in necropsy cases was seen in 3-12 months age group with 13/30 (43.33%) cases and in clinical samples the highest prevalence was observed in 1-3 months age group with 23/198 (11.61%) samples. The prevalence of BPIV3 was found to be 40% (12/30) in necropsy cases with maximum positivity rate in 3-12 months age group having 10/30 (33.33%) cases using RT-PCR of Matrix (M) protein having product size of 127bp. In addition, 13 different bacterial isolates from clinical samples and 9 different bacterial isolates from necropsy samples were isolated and identified. Staphylococcus aureus was the most prevalent bacteria in both clinical (15.87%) as well as necropsy samples (12.50%). Other bacterial isolates including Staphylococcus epidermidis, Streptococcus mutans, Streptococcus pyogenes, Streptococcus uberis, Klebsiella oxytoca, Klebsiella pneumoniae, Citrobacter koseri, Citrobacter freundii, Pasteurella spp., Salmonella spp., Escherichia coli and Proteus spp. were also isolated and identified. Pathogenic S. aureus and S. epidermidis were also identified. 3/5 (60%) isolates of C. freundii were found to be positive for shiga toxin pathotype. Different pathotypes of E. coli such as enteroaggregative, enterotoxigenic, enteropathogenic and enterohaemorrhagic were also detected. This study concludes that correlation of cytological findings and pathological findings coupled with molecular techniques aids in accurate diagnosis of respiratory viral diseases.
Open Access
STUDIES ON PATHOLOGY OF RESPIRATORY DISEASES IN BOVINE CALVES WITH REFERENCE TO BACTERIAL AND VIRAL INFECTIONS
(Palampur, 2022-01-10) BISHT, ANMOL; Gupta, Vipan.K.
Open Access
EFFECTS OF ROOT EXTRACT OF SAUSSUREA LAPPA ON CHEMICAL INDUCED LIVER DAMAGE IN RATS
(CSKHPKV Palampur, 2020-09-10) ABHISHEK, KUMAR; Asrani, R.K.
Open Access
STUDIES ON PATHOLOGY OF RESPIRATORY DISEASES IN BOVINE CALVES WITH SPECIAL EMPHASIS ON BOVINE RESPIRATORY SYNCYTIAL VIRUS AND PARAINFLUENZA-3 VIRUS INFECTION.
(CSK HPKV, Palampur, 2019-07-23) Sameeksha; Gupta, V.K.
The present study was conducted to determine the prevalence and the factors responsible for respiratory diseases in bovine calves during April, 2018 to May, 2019. A total 210 nasal swabs were collected from the necropsy (18), clinically sick (130) and apparently healthy calves (62). Besides, 24 tracheal and 12 lung swabs from necropsy cases were also collected. 33 calves were examined at necropsy. Pneumonia was recorded in all 33(100%) calves and was considered an important contributor to death. Out of 33 cases, gross lesions suggestive of bronchopneumonia seen in 7/33 (21.2%), interstitial pneumonia in 4/33 (12.1%) and bronchointerstitial pneumonia in 22 (66.7%). Bronchointerstitial pneumonia revealed mixed characteristic features of suppurative bronchopneumonia and interstitial pneumonia. Staphylococcus aureus was detected in 30.3 % of the clinical samples. E. coli comprised 23.6% of the isolates in necropsy. Bacterial isolates seen in nasal tract were different from the bacterial isolates obtained from lungs / trachea. Single bacterial infection was detected in 51.5% (17/33) cases and polymicrobial infection in 45.5% (15/33) cases. Total 41 lung tissues from necropsy and 104 nasal samples from clinical cases were screened for BRSV using RT-PCR with glycoprotein gene (246bp) primers and showed overall prevalence of 50.4%. BRSV infected lung showed consolidation at the cranioventral and fibrinous pleuritis with range of changes in airways histopathologically multinucleated syncytial cells was seen. Total of 145 cases were screened for BPIV-3 using RT-PCR, no sample was found positive for primer having HN gene. In necropsy, most frequent E.coli pathotype categories were found to be enterotoxigenic (80%) & enteroaggregative (80%). In clinically sick cases frequent pathotype categories found were enterotoxigenic (63.6%). Four (33.3%) of Citrobacter freundii isolates were pathogenic with shiga toxin on PCR analysis. 17.9% Staphylococcus aureus and 15.4% Staphylococcus epidermidis were recorded as pathogenic. Male and female calves showed 20/33 (60.7%) and 13/33 (39.4%) mortalities respectively. Maximum mortality was recorded in <1 M (42.2%) old calves. Most number of deaths were recorded during the summer (36.4%) season. Combination of molecular detection (PCR) along with clinical, gross & histopathology helped in better confirmatory diagnosis of BRSV infection in calves. The present study concluded co-infection of bacterial and BRSV in respiratory infection of young calves.
Open Access
STUDIES ON THE PATHOLOGY OF NEOPLASM IN DOMESTIC ANIMALS.
(CSKHPKV, Palampur, 2019-07) Masand, Rupali; Patil, R.D.
The present work was conducted to study the pathomorphological changes of animal neoplasms by using cytological, gross and histopathological, histochemical and immunohistochemical studies. A total of ninety-six (96) tumor cases were collected during September, 2017 to June, 2019. The incidence of neoplasms was highest in canines (71.87%) followed by the bovines (16.67%), rabbits (7.2%), equines (2.08%) and caprine (1.04%). The maximum incidence of neoplasms occurred in the age group of 3-6 years (28.12%) and the incidence was highest during summer (31.25%) followed by winter (25%), monsoon (23.45%) and spring (19%) season. Benign tumors (53.12%) were higher than malignant tumors (46.87%). Location wise highest occurrence of neoplasms was recorded in mammary gland (20.33%) followed by oral region (18.75%), genitalia (11.45%). On basis of cytology, tumors of epithelial origin were 58.04%, mesenchymal origin 32.25% and round cell tumor 9.6%. Canine mammary tumors (18.84%) were found to be most frequently recorded tumors followed by squamous cell carcinoma (13.04%). Histopathological examination of papilloma, squamous cell carcinoma, Fibroma, fibrosarcoma, haemangioma, haemangiosarcoma, mast cell tumor, melanocytoma, malignant melanoma, sebaceous adenoma, mammary adenoma and adenocarcinoma, mixed mammary tumor, solid carcinoma of mammary gland, hepatocellular carcinoma, cholangiocellular carcinoma, acinar cell carcinoma, malignant renal cell carcinoma, Sertoli cell tumor, cystic papillary carcinoma revealed charecteristic benign and malignant features. Histochemical staining such as Masson’s trichrome, Van Gieson’s, Toluidine blue and Schmorl's confirmed the tumors as fibroma or fibrosarcoma, leiomyoma, mast cell tumors and melanocytoma or malignant melanoma, respectively. Silver stained nucleolar organizer region (AgNOR)count showed significantly higher count in malignant than benign tumors. Amongst malignant tumor, the highest AgNOR count was found in cholangiocellular carcinoma (6.89±2.00) followed by fibrosarcoma of mammary gland (6.64±2.47) and least in haemagiosarcoma (2.96±1.25). Immunohistochemical studies in tumor for the expression of proliferating cell nuclear antigen (PCNA), p53 and H-ras found to have diagnostic value and supported the hisopathological observations of tumors in animals. Further, TdT mediated dUTP nick end labeling (TUNEL) assay was found to be useful for determining the role of apoptosis in different animal neoplasms.
Open Access
INVESTIGATION ON PROTECTIVE EFFECT OF MEDICINAL PLANT(S) AGAINST HEPATIC DAMAGE IN RATS
(CSKHPKV, Palampur, 2019-02-08) Sharma, Rakshita; Asrani, R.K.
The present study was planned with a view to observe the In-vitro cytotoxicity of 70% ethanolic extract of plant Saussurea lappa, Phyllanthus urinaria, Rheum webbianum, Artemisia nilagirica and Zingiber crysanthum on A549, KB and SiHA cell lines. Extracts were screened for the presence of various polyphenols and main bioactive compounds of respective plant. The plant extract from 3 plants namely Phyllanthus urinaria, Saussurea lappa and Rheum webbianum was used in combination (1:2:2) to study the growth response, mortality, clinical signs, biochemical and pathological changes against DEN induced hepatic damage in rats. Acute toxicity was studied in 6 rats. For final experimentation, 42 rats were randomly divided into 6 groups, Group-1: plain control, Group-2: N-diethylnitrosamine (DEN) alone. Group-3: DEN with Silymarin @25mg, Group-4: DEN with combination of plant extracts @100mg/kg, Group-5: DEN with combination of plant extract @250mg, Group-6: DEN with combination of plant extract @500mg. The N-diethylnitrosamine (DEN) was given in drinking water to rats throughout the experiment. The route of plant extract and standard drug was oral intubation throughout the period of experimentation of 17 weeks. Results of the present study showed that rats of group-2 (DEN only) exhibited clinical signs of decrease in body weight, hair loss, dullness, and depression. ALT, AST, Creatinine activity was found to increase, whereas total protein concentration decreased in all the DEN treated groups as compared to the control groups. Biochemical parameters in case of combination groups were lower as compared to the DEN alone group. Grossly, group-2 animal showed hepatomegaly, liver paleness, cirrhosis and multiple nodules over the liver surface. However, in the combination groups the gross lesions of less intensity were observed. Microscopically, the liver of the rats of group-II revealed severe distortion of the hepatic cord arrangement, lobular pattern, bile duct proliferation, nuclear changes, fibrous tissue proliferation, inflammatory cell infiltration, Kupffer cell hyperplasia and degenerative changes. Similarly, in this group spleen showed coalescing lymphoid pulp, increase in the red pulp. Similar changes were noticed in the combination groups (group-3, 4, 5 and 6), but the changes were less severe in intensity as compared to the DEN alone group. Histopathologically, liver and spleen showed regenerative changes in plant treated groups. Immunohistochemical studies showed positive result for the presence of apoptotic bodies in DEN treatment groups. The results indicated hepatoprotective effect of plant extract in combination. DEN was found to be carcinogenic and produced significant hepatic nodules in rat model during in vivo experimentation.

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