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Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples

Objective: To compare the efficiency of routine diagnostic PCR assays in detecting pathogenic Leptospira isolated from water and soils. Methods: Seven routine assays targeting six genes (lipL32, flaB, gyrB, lfb1, secY and ligB) were evaluated and compared on the cultures of two groups of pathogenic...

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Main Authors: Yap, May Ling, Sekawi, Zamberi, Chee, Hui Yee, Ong, Alan Han Kiat, Vasantha Kumari, Neela
Format: Article
Language:English
Published: Wolters Kluwer - Medknow 2019
Online Access:http://psasir.upm.edu.my/id/eprint/80964/1/LEPTOSPIRA.pdf
http://psasir.upm.edu.my/id/eprint/80964/
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spelling my.upm.eprints.809642020-10-14T20:41:52Z http://psasir.upm.edu.my/id/eprint/80964/ Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples Yap, May Ling Sekawi, Zamberi Chee, Hui Yee Ong, Alan Han Kiat Vasantha Kumari, Neela Objective: To compare the efficiency of routine diagnostic PCR assays in detecting pathogenic Leptospira isolated from water and soils. Methods: Seven routine assays targeting six genes (lipL32, flaB, gyrB, lfb1, secY and ligB) were evaluated and compared on the cultures of two groups of pathogenic Leptospira from different sources. One group included 19 described reference strains recovered from infected human or animals, and another group included 22 environmental isolates from recreational and residential sites in Malaysia. The latter have been confirmed for presence of pathogenic Leptospira DNA. PCR positivity or detection sensitivity of each assay was determined and compared between the two groups. Results: Validation on reference strains showed 100.0% PCR sensitivity for all assays except ligB-PCR (95.0%) that failed to amplify Leptospira interrogans serovar Pomona. In marked contrast, there was a notable decline in sensitivity in the environmental isolates (lipL32-PCR, 95.5%; flaB-PCR, 90.9%; gyrB-PCR, 77.3%; lfb1-PCR, 59.1%; secY-PCRs, 40.9% G1/G2-PCR, 36.4%; ligB-PCR, 13.6%), implying a large genetic distance between the two groups, as well as nucleotide polymorphism among environmental isolates. Conclusions: High proportion of false-negative PCR results suggests a need of prudent selection of primers in detecting environmental pathogenic Leptospira. These findings offer valuable insights on the extensive biodiversity of genus Leptospira and its impact on the efficacy and development of molecular detection tool. Wolters Kluwer - Medknow 2019 Article PeerReviewed text en http://psasir.upm.edu.my/id/eprint/80964/1/LEPTOSPIRA.pdf Yap, May Ling and Sekawi, Zamberi and Chee, Hui Yee and Ong, Alan Han Kiat and Vasantha Kumari, Neela (2019) Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples. Asian Pacific Journal of Tropical Medicine, 12 (10). pp. 472-478. ISSN 1995-764 10.4103/1995-7645.269908
institution Universiti Putra Malaysia
building UPM Library
collection Institutional Repository
continent Asia
country Malaysia
content_provider Universiti Putra Malaysia
content_source UPM Institutional Repository
url_provider http://psasir.upm.edu.my/
language English
description Objective: To compare the efficiency of routine diagnostic PCR assays in detecting pathogenic Leptospira isolated from water and soils. Methods: Seven routine assays targeting six genes (lipL32, flaB, gyrB, lfb1, secY and ligB) were evaluated and compared on the cultures of two groups of pathogenic Leptospira from different sources. One group included 19 described reference strains recovered from infected human or animals, and another group included 22 environmental isolates from recreational and residential sites in Malaysia. The latter have been confirmed for presence of pathogenic Leptospira DNA. PCR positivity or detection sensitivity of each assay was determined and compared between the two groups. Results: Validation on reference strains showed 100.0% PCR sensitivity for all assays except ligB-PCR (95.0%) that failed to amplify Leptospira interrogans serovar Pomona. In marked contrast, there was a notable decline in sensitivity in the environmental isolates (lipL32-PCR, 95.5%; flaB-PCR, 90.9%; gyrB-PCR, 77.3%; lfb1-PCR, 59.1%; secY-PCRs, 40.9% G1/G2-PCR, 36.4%; ligB-PCR, 13.6%), implying a large genetic distance between the two groups, as well as nucleotide polymorphism among environmental isolates. Conclusions: High proportion of false-negative PCR results suggests a need of prudent selection of primers in detecting environmental pathogenic Leptospira. These findings offer valuable insights on the extensive biodiversity of genus Leptospira and its impact on the efficacy and development of molecular detection tool.
format Article
author Yap, May Ling
Sekawi, Zamberi
Chee, Hui Yee
Ong, Alan Han Kiat
Vasantha Kumari, Neela
spellingShingle Yap, May Ling
Sekawi, Zamberi
Chee, Hui Yee
Ong, Alan Han Kiat
Vasantha Kumari, Neela
Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples
author_facet Yap, May Ling
Sekawi, Zamberi
Chee, Hui Yee
Ong, Alan Han Kiat
Vasantha Kumari, Neela
author_sort Yap, May Ling
title Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples
title_short Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples
title_full Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples
title_fullStr Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples
title_full_unstemmed Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples
title_sort comparative analysis of current diagnostic pcr assays in detecting pathogenic leptospira isolates from environmental samples
publisher Wolters Kluwer - Medknow
publishDate 2019
url http://psasir.upm.edu.my/id/eprint/80964/1/LEPTOSPIRA.pdf
http://psasir.upm.edu.my/id/eprint/80964/
_version_ 1681490835420479488
score 13.211869

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