Detection and quantification of Leptospira interrogans in hamster and rat kidney samples: immunofluorescent imprints versus real-time PCR

A major limitation in the clinical management and experimental research of leptospirosis is the poor performance of the available methods for the direct detection of leptospires. In this study, we compared real-time PCR (qPCR), targeting the lipL32 gene, with the immunofluorescent imprint method (IM...

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Bibliographic Details
Published in:	PloS one Vol. 7; no. 2; p. e32712
Main Authors:	Chagas-Junior, Adenizar D, da Silva, Caroline L R, Soares, Luciane Marieta, Santos, Cleiton S, Silva, Carlos D C M, Athanazio, Daniel A, dos Reis, Mitermayer G, McBride, Flávia W Cruz, McBride, Alan J A
Format:	Journal Article
Language:	English
Published:	United States Public Library of Science 29-02-2012 Public Library of Science (PLoS)
Subjects:	Animal models Animals Antigens Chronic infection Comparative analysis Computer simulation Cricetinae Deoxyribonucleic acid DNA Experimental research Experiments Female Genes Gram-negative bacteria Hamsters Health aspects Health sciences Immunoglobulins Infection Infections Inoculation Kidney - metabolism Kidneys Laboratory animals Leptospira Leptospira interrogans Leptospira interrogans - metabolism Leptospirosis Leptospirosis - microbiology Leptospirosis - pathology Medicine Mesocricetus Microscopy Microscopy, Fluorescence - methods Proteins Rats Rats, Wistar Rattus norvegicus Real time Real-Time Polymerase Chain Reaction - methods Rodents Sequence Analysis, DNA Statistical analysis Statistical methods Veterinary Science Brazil
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Summary:	A major limitation in the clinical management and experimental research of leptospirosis is the poor performance of the available methods for the direct detection of leptospires. In this study, we compared real-time PCR (qPCR), targeting the lipL32 gene, with the immunofluorescent imprint method (IM) for the detection and quantification of leptospires in kidney samples from the rat and hamster experimental models of leptospirosis. Using a virulent strain of Leptospira interrogans serovar Copenhageni, a chronic infection was established in the rat model, which were euthanized 28 days post-infection, while the hamster model simulated an acute infection and the hamsters were euthanized eight days after inoculation. Leptospires in the kidney samples were detected using culture isolation, qPCR and the IM, and quantified using qPCR and the IM. In both the acute and chronic infection models, the correlation between quantification by qPCR and the IM was found to be positive and statistically significant (P<0.05). Therefore, this study demonstrates that the IM is a viable alternative for not only the detection but also the quantification of leptospires, particularly when the use of qPCR is not feasible.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Conceived and designed the experiments: ADCJ AJAM. Performed the experiments: ADCJ CLRS LMS CSS CDCMS FWCM. Analyzed the data: ADCJ AJAM. Contributed reagents/materials/analysis tools: DAA MGR FWCM. Wrote the paper: ADCJ DAA MGR FWCM AJAM.
ISSN:	1932-6203 1932-6203
DOI:	10.1371/journal.pone.0032712