Development of PCR assay for targeting partial lipL21 and lipL41 gene of leptospira

Chandan, S.; Umesha, S.; Bhure, S. K.; Haraprasad, N.; Chandrashekar, S.
January 2011
Nepal Journal of Biotechnology;2011, Vol. 1 Issue 1, p22
Academic Journal
Leptospirosis is a bacterial zoonotic disease caused by spirochetes of the genus Leptospira that affects human and a wide range of animals. The direct method of diagnosis of leptospirosis, has been so far by culture isolation but it is time consuming and potentially biohazardous. Another traditional method is the detection of antibodies (Serological tests) which is also a time consuming method and fails to identify the infecting serovar. To overcome these limitations associated with the cultivation and serology, we developed PCR assay targeting partial lipL21 gene and lipL41 gene of Leptospires using in-house designed P28/29 and P30/31 primers, with a product size of 385bp and 427bp. The amplicons were subjected to restriction enzyme digestion using RsaI, Pvu II and HindIII for product of P28/29 and ClaI, TaqI and RsaI were used for product of P30/31. The protocols were standardized and the assay targeting the partial lipL21 and lipL41 gene was found to be specific for eight pathogenic Leptospires out of nine leptospires tested. The products were then cloned in pGEMT Easy vector and sequenced to facilitate further studies. PCR could detect the target bacterial gene without any ambiguity and showed good efficiency in detection of targeted species in the sample. This simple, rapid and cost-effective method can be applicable in a prediction system to prevent disease outbreak by these Leptospira species and can be considered as an effective tool for disease diagnosis of Leptospira species.


Related Articles

  • Leptospirosis in Febrile Patients: Diagnosis by Serology and Polymerase Chain Reaction. Perwez, Khalid; Suman, V. B.; Kumar, Nayanatara Arun; Hussain, Mohammad Asif // Journal of Biology & Life Science (JBLS);Aug2011, Vol. 2 Issue 1, p1 

    Introduction: Leptospirosis is a zoonotic disease found in all tropical and temperate areas of the world. Humans contract leptospira through mucosal or percutaneous exposure to environments contaminated by the urine of chronically infected animal sources mainly rodents, dogs, pigs and cattle....

  • INVESTIGAÇÃO DO CONHECIMENTO DE PESSOAS INFECTADAS POR Leptospira spp. E DA SOROPREVALÊNCIA, LEPTOSPIRÚRIA E LEPTOSPIREMIA DE SEUS CÃES CONTACTANTES. Gaspareto dos Santos, Leonardo; Trochmann Cordeiro, Carolina; da Costa Vieira, Rafael Felipe; Wischral Jayme Vieira, Thállitha Samih; Oliveira Stedile, Simone Tostes // Archives of Veterinary Science;2019, Vol. 24 Issue 4, p83 

    Leptospirosis is a zoonotic disease, endemic in Brazil, who affects several mammals, including man. Asymptomatic dogs can transmit and it is considered that the number of animals with this status is underestimated. Although most of studies being performed by serology, the serology doesn't...

  • Validation of reference genes for RT-qPCR normalization in common bean during biotic and abiotic stresses. Borges, Aline; Tsai, Siu; Caldas, Danielle // Plant Cell Reports;May2012, Vol. 31 Issue 5, p827 

    Selection of reference genes is an essential consideration to increase the precision and quality of relative expression analysis by the quantitative RT-PCR method. The stability of eight expressed sequence tags was evaluated to define potential reference genes to study the differential...

  • Development of Real-Time PCR for in wood-detection of Ceratocystis platani, the agent of canker stain of Platanus spp. Pilotti, Massimo; Lumia, Valentina; Lernia, Giovanni; Brunetti, Angela // European Journal of Plant Pathology;Sep2012, Vol. 134 Issue 1, p61 

    Ceratocystis platani is a quarantine fungal pathogen agent of canker stain, a destructive disease affecting Platanus. Despite its diagnosis being critical for disease control, there is still no effective diagnostic tool as all known mycological and biological detection assays are problematical....

  • Serological and Molecular Detection of Brucella abortus from Cattle by RBPT, STAT and PCR and Sample Suitability of Whole Blood for PCR. Karthik, K.; Rathore, R.; Thomas, P.; Elamurugan, A.; Arun, T. R.; Dhama, K. // Asian Journal of Animal & Veterinary Advances;2014, Vol. 9 Issue 4, p262 

    Brucella abortus, one of the major pathogen causing abortions in cattle worldwide and a zoonotic agent, need to be detected earlier in order to prevent its spread among animals. The present study aimed at to know the prevalence of B, abortus in cattle population of three states (Uttar Pradesh,...

  • SEROLOGICAL EVIDENCE OF LEPTOSPIRA SPP. IN THE LAMI TUCO-TUCO RODENTS (CTENOMYS LAMI). Tostes Oliveira, Simone; Guiomara Stein, Gisele; Pires Santos, Andrea; Welker Biondo, Alexander; Messick, Joanne Belle; Diaz Gonzalez, Felix Hilario // Archives of Veterinary Science;2014, Vol. 19 Issue 3, p69 

    Although rodents are reportedly the major reservoirs of Leptospira spp. in the wildlife of Brazil, the role of the widely distributed native tuco-tuco rodent (Ctenomys lami) has yet to be determined. Accordingly, a total of 40 serum and eight urine samples from wild C. lami were collected from...

  • Expanding the Diagnostic Use of PCR in Leptospirosis: Improved Method for DNA Extraction from Blood Cultures. Villumsen, Steen; Pedersen, Rebecca; Krogfelt, Karen Angeliki; Jensen, Jørgen Skov // PLoS ONE;2010, Vol. 5 Issue 8, p1 

    Background: Leptospirosis is a neglected zoonosis of ubiquitous distribution. Symptoms are often non-specific and may range from flu-like symptoms to multi-organ failure. Diagnosis can only be made by specific diagnostic tests like serology and PCR. In non-endemic countries, leptospirosis is...

  • Quantitative PCR-based detection of pathogenic Leptospira in Hawai'ian coastal streams. Viau, Emily J.; Boehm, Alexandria B. // Journal of Water & Health;Dec2011, Vol. 9 Issue 4, p637 

    Pathogenic Leptospira, the causative agents of leptospirosis, are mainly associated with tropical freshwaters, but little is known about their fate in the environment. This study analyzed the distribution of pathogenic Leptospira genomes in 22 tropical coastal streams using quantitative...

  • Molecular identification of the ompL1 gene within Leptospira interrogans standard serovars. Dezhbord, Mehrangiz; Esmaelizad, Majid; Khaki, Pejvak; Fotohi, Fariba; Moghaddam, Athena Zarehparvar // Journal of Infection in Developing Countries;Jun2014, Vol. 8 Issue 6, p688 

    Introduction: Leptospirosis, caused by infection with pathogenic Leptospira species, is one of the most prevalent zoonotic diseases in the world. Current leptospiral vaccines are mainly multivalent dead whole-cell mixtures made of several local dominant serovars. Therefore, design and...


Read the Article


Sorry, but this item is not currently available from your library.

Try another library?
Sign out of this library

Other Topics