Optimization of In-House Indirect-ELISA & EITB Assays Employing Cysticercus cellulosae Antigens for Serological Detection and PCR Assays for Molecular Detection of Porcine Cysticercosis.

Background: Porcine cysticercosis, caused by metacestodes of Taenia solium is an important neglected zoonosis. We evaluated the presence of anti-cysticercal antibodies and T. solium specific DNA in pig sera and blood samples respectively collected from Maharashtra, India. Methods: A total of three antigens (Scolex Antigen (SA), Membrane Body Antigen (MBA) and Excretory-Secretory Antigen (ESA)) were prepared from metacestodes of T. solium and employed in an in-house developed indirect-IgG ELISA for serological screening of 1000 porcine sera samples at Department of Veterinary Public Health, Nagpur Veterinary College, Maharashtra, India. The ELISA positive sera samples were subjected to EITB Assay for detection of immunodominant peptides. An effort has been made for molecular detection of porcine cysticercosis by PCR assay targeting large subunit rRNA gene of T. solium from blood samples of the corresponding ELISA-positive pigs. Results: The overall seroprevalence of porcine cysticercosis employing SA, MBA and ESA was 12.6%, 8.7% and 12.5% respectively. The lower and medium molecular weight peptides were the most frequently recognised in EITB assay. The numbers of bands recognised in EITB assay were observed to be proportionate with the corresponding ELISA O.D. values. An amplification product of 286 bp was observed in 22.98% (20/87), 30.35% (30/99) and 17.14% (12/70) of the sero-positive samples against SA, ESA and MBA respectively. Conclusion: EITB still remains the gold standard serodiagnosis test for cysticercosis. The inclusion of a greater number of positive samples and purification of antigens may improve the diagnostic efficacy of the tests.

Characterization of crude extract prepared from Indian curd and its potential as a biopreservative.

The adverse effects of chemical preservatives used to prevent food spoilage have led to the search for various biopreservatives. Considering this, a study was undertaken to prepare crude extract (CE) from Indian curd (a fermented dairy product) and characterized it in terms of antioxidant and antimicrobial activities against some common food-borne bacteria. The CE exhibited well pronounced antimicrobial activity against Bacillus cereus and Salmonella typhimurium. The minimum inhibitory concentration (MIC) of CE was recorded for 2-fold concentrated solution prepared from a 10-fold stock. The CE exhibited a significantly higher (p < 0.05) antioxidant and antimicrobial activities compared to its fractions. The CE was found to be heat stable (up to 100 ℃ for 30 min) and exhibited a significant (p < 0.05) increase in activity at pH 2-7 and in combination with 2% citric acid solution. Trypsin treatment suggested it to be of proteinaceous in nature. The antibacterial activity of CE remained intact at 4 ℃ for seven days, whereas non-significant (p > 0.05) changes in its activity were noted during storage at -20 ℃ for 30 days. The curd sample used for preparation of CE, when tested for bacteriocin production and subsequent antimicrobial activity, did not show inhibition against S. typhimurium. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis analysis of CE and its fractions revealed multi-banding pattern. By virtue of its bioactivities observed, CE can be explored as a promising food biopreservative.

Pathological and molecular identification of porcine cysticercosis in Maharashtra, India.

Porcine cysticercosis, caused by metacestodes of Taenia solium is an important emerging zoonotic disease with public health and economic significance. Pigs acquire the disease through consumption of Taenia solium eggs excreted by human tapeworm carriers. The present study was conducted to investigate the prevalence of porcine cysticercosis in Nagpur and Mumbai region of Maharashtra, India by P/M examination of carcasses followed by histopathology of affected organs in infected animals and molecular identification of cysts for confirmation. Out of 1000 pigs examined during slaughter, three pigs were found to be heavily affected with T. solium cysts giving a prevalence of 0.3%. Histological section of brain in infected animals revealed marked vascular congestion of meninges, mild neuronal degeneration, perivascular cuffing and gliosis while the liver showed the infiltration of mononuclear cell, predominantly eosinophils throughout the parenchyma. Some degree of calcification was observed in the cysts lodged in liver while calcification was not evident in case of cysts lodged in brain, tongue, diaphragm and skeletal muscle. Molecular identification by PCR using two sets of oligonucleotide primers against LSU rRNA gene and Mt-Cox1 gene of T. solium confirms the cysts to be that of T. solium. The molecular diagnostics methods have been considered for validation in conjunction with P/M inspections, parasitological and histopathological examinations. The study confirms the presence of porcine cysticercosis in the two regions and demands proper sanitary measures to minimize the risk of infection from zoonoses and food safety point of view.