Evaluation of immunotherapeutic and antioxidative effects of some herbs in bovine subclinical mastitis

Name : Dhiraj Kumar Gupta I.D. No. : 23313 Year of Admission : Semester I, 2004-05 Degree : Ph.D. Major : Veterinary Medicine Minor : Veterinary Pharmacology Department : Clinical Veterinary Medicine, Ethics & Jurisprudence Thesis Title : Evaluation of immuno-therapeutic and antioxidative effects of some herbs in bovine subclinical mastitis Advisor : Dr. V.S. Rajora, Professor & Head, Deptt. of Clinical Veterinary Medicine In the present study in vitro antibacterial activity of hydro-alcoholic extracts of Withania somnifera roots, Ocimum sanctum leaves, Azadirachta indica bark and Tinospora cordifolia stem individually and in combination against the mastitis pathogens viz., Staphylococcus aureus, Streptococcus agalactiae and Escherichia coli was studied and the activity of best extract mix was compared with that of standard antibiotic. The best extract mix was tried in vivo in subclinical mastitic cows to assess its immuno-therapeutic and antioxidative effects. The per cent yield of extracts of W. somnifera roots, O. sanctum leaves, A. indica bark and T. cardifolia stem was 14.6%, 9.5%, 6.1% and 5.9%, respectively. The extract mix of W. somnifera and O. sanctum was observed to be the best combination against all tested bacteria with the efficacy of 66.25%, 74.81% and 75.56% for Staphylococcus aureus, E. coli and Streptococcus agalactiae, respectively in comparison to enrofloxacin. The MIC of hydro-alcoholic extract of W. somnifera roots and A. indica bark was observed to be 125 mg/ml and 500 mg/ml, respectively for both Staphylococcus aureus and E. coli and 500 mg/ml and 1000 mg/ml, respectively for Streptococcus agalactiae. Whereas, the MIC of hydro-alcoholic extract of O. sanctum leaves was recorded to be 125 mg/ml for Staphylococcus aureus and 250 mg/ml for both Streptococcus agalactiae and E. coli. Overall prevalence of SCM was 32.98% and 57.72% cow wise and quarter wise, respectively with higher involvement of right hindquarters, between 2 nd to 4 th parity and between 31-60 days of lactation. Staphylococcus spp. was present in maximum (67.95%) cases followed by Streptococcus spp. (23.72%). The mean CMT point score, SCC and TBC in SCM cows were recorded to be 2.15±0.06, 6.15±0.14 x 10 5 and 4.93 x10 3 /ml, respectively. No statistical difference in CMT score, pH, EC, fat, protein, lactose, SNF, SCC and TBC values on day 3, 7 and 15 was observed in Gr I and Gr II during the present course of investigation. In Gr III cows (enrofloxacin treated) significant decrease in CMT score, SCC, TBC and neutrophil % was recorded on day 3. The mean phagocytic neutrophils in this group increased significantly on day 3 and 7 in comparison to day 0 post treatment. In Gr IV (extract mix treated) and Gr VI (extract and powder mix treated) CMT score, SCC, neutrophil% and lymphocyte% rose significantly on day 3 and decline subsequently on day 7 and 15 post treatment. The TBC declined gradually in this group during the period of study. Mean phagocytic neutrophils and phagocytic index significantly increased on day 3 and 7 post treatment. A significant rise in lactose and SNF values was observed in this group. Administration of PBS (Gr V) did not cause any recovery. Standard antioxidant therapy im (Gr VII) showed a significant decrease in CMT score, EC and neutrophil percentage on day 3, 7 and 15 as compared to day 0. However, the TBC varied significantly on day 7 and 15 as compared to day 0 post treatment. There was no significant alteration in the values of SCC, fat, protein, lactose and SNF and lymphocyte % in milk of cows of group VII throughout the period of study. The phagocytic neutrophil percentage in Gr VII increased significantly on day 7 as compared to day 0 post treatment. The phagocytic index did not vary significantly on day 3 and 7 as compared to day 0. No complete cure of SCM quarter in Gr VII was observed. In Gr VIII (powder mix treated) a significant decrease in CMT score, EC, SCC, TBC, lactose and SNF, neutrophil % was observed on day 15 in comparison to day 0 post treatment. However, lymphocyte % rose significantly on day 3 as compared to day 0 but afterwards declined on day 7 and 15 post treatment. Gr VIII experienced a significant increase in phagocytic neutrophils and phagocytic index on day 3 and 7 in comparison to day 0 post treatment. The activities of oxidative stress related enzymes varied significantly after treatment in Gr III, Gr IV, Gr VI, Gr VII and Gr VIII. The recovery was 91.67%, 88.33%, 75.00% and 50.00% in Gr III, Gr VI, Gr IV and Gr VIII, respectively. No adverse reaction and milk losses were observed in herbs treated groups during the period of treatment. Enrofloxacin was graded as superior treatment for subclinical mastitic cows followed by extract and powder mix of W. somnifera and O. sanctum, extract of W. somnifera and O. sanctum and powder mix. The total cost of treatment of one quarter with intramammary alone, intramammary and oral use and oral administration of herbal preparations was computed as Rs. 94.99, Rs.107. 60 and Rs. 25.20, respectively as against Rs. 201.00 on intramammary infusion of enrofloxacin. The intramammary infusion of extract of W. somnifera and O. sanctum and oral administration of powder of W. somnifera and O. sanctum mix gave very encouraging results in bovine subclinical mastitis. Therefore, in future the use of combination of these herbs may be explored in clinical mastitis and the active ingredients of these herbs may be isolated and tried for their efficacy in subclinical mastitis as well as in clinical mastitis. (V.S. Rajora) Advisor (Dhiraj Kumar Gupta) Author