Evaluation of a modified Taqman assay detecting pathogenic Leptospira spp. against culture and Leptospira-specific IgM enzyme-linked immunosorbent assay in a clinical environment.
A Taqman assay for the detection of pathogenic Leptospira was modified to suit the LightCycler instrument. The modified assay was found to have an analytical sensitivity of 10 copies/reaction. The assay was then compared to the current gold standard for acute phase detection, culture, and to a commercially available Leptospira-specific IgM enzyme-linked immunosorbent assay (ELISA). Of the 236 samples including serum and ethylenediaminetetraacetic acid anticoagulated blood sample submitted for testing, polymerase chain reaction (PCR) was able to detect Leptospira DNA in 27 of the 28 culture positives and in 1 negative culture. Discrepant results were resolved by using the microscopic agglutination test on convalescent sera. The PCR was found to have a clinical specificity and sensitivity of 99.5% and 96.4%, respectively, when compared to the culture. Comparisons between culture and ELISA showed that the ELISA lack sensitivity (4.2%) and positive predictability (3.6%) for the detection of acute phase Leptospira infections. These results show that the modified LightCycler Taqman assay could be used as a replacement of culture for the detection of pathogenic Leptospira in a clinical setting.
Slack
Andrew
A
WHO/FAO/OIE Collaborating Centre for Reference and Research on Leptospirosis, Western Pacific Region, Centre for Public Health Sciences, Queensland Health Scientific Services, Brisbane 4108, Australia. andrew_slack@health.qld.gov.au
Symonds
Meegan
M
Dohnt
Michael
M
Harris
Christopher
C
Brookes
Dianne
D
Smythe
Lee
L
D000907:Antibodies, Bacterial / Q000097:blood; D001431:Bacteriological Techniques; D003470:Culture Media; D004269:DNA, Bacterial / Q000097:blood / Q000302:isolation & purification; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007075:Immunoglobulin M / Q000097:blood*; D007919:Leptospira / Q000145:classification / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D007922:Leptospirosis / Q000175:diagnosis* / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D011237:Predictive Value of Tests; D012680:Sensitivity and Specificity; D019914:Taq Polymerase
D000907:Antibodies, Bacterial; D003470:Culture Media; D004269:DNA, Bacterial; D007075:Immunoglobulin M; D019914:Taq Polymerase