AUTHOR=Sheikh Nazish , Kumar Sanjay , Sharma Harsh Kumar , Bhagyawant Sameer S. , Thavaselvam Duraipandian 

TITLE=Development of a Rapid and Sensitive Colorimetric Loop-Mediated Isothermal Amplification Assay: A Novel Technology for the Detection of Coxiella burnetii From Minimally Processed Clinical Samples

JOURNAL=Frontiers in Cellular and Infection Microbiology

VOLUME=Volume 10 - 2020

YEAR=2020

URL=https://www.frontiersin.org/journals/cellular-and-infection-microbiology/articles/10.3389/fcimb.2020.00127

DOI=10.3389/fcimb.2020.00127

ISSN=2235-2988

ABSTRACT=Q fever is an important worldwide zoonosis that is caused by infection with Coxiella burnetii. Because of the low infectious dose, aerosol transmission, resistance to heat, drying, and many common disinfectants, C. burnetii is deemed as a category B agent for bioterrorism. Transmission of infection to humans is largely through the inhalation of contaminated aerosols generated from birth products of animals, but may also occur after ingestion of unpasteurized dairy products. Thus, rapid and accurate detection of C. burnetii in shedders, especially those that are asymptomatic, is important for early warning which allows controlling its spread among animals and animal to human transmission. In the present study, we have developed a colorimetric loop-mediated isothermal amplification (LAMP) assay to detect the presence of the transposase gene insertion element IS1111a of C. burnetii in sheep vaginal swabs. The sensitivity of this LAMP assay is very similar to quantitative PCR (qPCR) method with a detection limit of 3 copies of the gene, which is equivalent to about one cell of C. burnetii. The utility of the colorimetric LAMP assay for use in clinical samples was determined by testing 145 vaginal swab samples collected from the sheep breeding farms with history of still birth and repeated abortions. Compared to qPCR, colorimetric LAMP had a sensitivity of 93.75% (CI: 69.77%-99.84%) and specificity of 100% (CI: 97.20%-100%), with a PPV and NPV of 100 and 99.24%, respectively. There was a very high level of agreement between both colorimetric LAMP and reference qPCR assay. The colorimetric LAMP assay reported here is a rapid and simple test without extensive sample preparation and has a short turn-around time of less than 45 min. To the best of our knowledge this is the very first report describing the use of colorimetric LAMP assay for the detection of C. burnetii in vaginal swab samples with minimal sample processing for DNA extraction.