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Meta-Analysis
. 2019 Aug 1;8(8):CD011871.
doi: 10.1002/14651858.CD011871.pub2.

Nucleic acid and antigen detection tests for leptospirosis

Bada Yang  1 , Sophia G de Vries, Ahmed Ahmed, Benjamin J Visser, Ingeborg M Nagel, René Spijker, Martin P Grobusch, Rudy A Hartskeerl, Marga Ga Goris, Mariska Mg Leeflang
Affiliations
Meta-Analysis

Nucleic acid and antigen detection tests for leptospirosis

Bada Yang et al. Cochrane Database Syst Rev. .

Abstract

Background: Early diagnosis of leptospirosis may contribute to the effectiveness of antimicrobial therapy and early outbreak recognition. Nucleic acid and antigen detection tests have the potential for early diagnosis of leptospirosis. With this systematic review, we assessed the sensitivity and specificity of nucleic acid and antigen detection tests.

Objectives: To determine the diagnostic test accuracy of nucleic acid and antigen detection tests for the diagnosis of human symptomatic leptospirosis.

Search methods: We searched electronic databases including MEDLINE, Embase, the Cochrane Library, and regional databases from inception to 6 July 2018. We did not apply restrictions to language or time of publication.

Selection criteria: We included diagnostic cross-sectional studies and case-control studies of tests that made use of nucleic acid and antigen detection methods in people suspected of systemic leptospirosis. As reference standards, we considered the microscopic agglutination test alone (which detects antibodies against leptospirosis) or in a composite reference standard with culturing or other serological tests. Studies were excluded when the controls were healthy individuals or when there were insufficient data to calculate sensitivity and specificity.

Data collection and analysis: At least two review authors independently extracted data from each study. We used the revised Quality Assessment of Diagnostic Accuracy Studies tool (QUADAS-2) to assess risk of bias. We calculated study-specific values for sensitivity and specificity with 95% confidence intervals (CI) and pooled the results in a meta-analysis when appropriate. We used the bivariate model for index tests with one positivity threshold, and we used the hierarchical summary receiver operating characteristic model for index tests with multiple positivity thresholds. As possible sources of heterogeneity, we explored: timing of index test, disease prevalence, blood sample type, primers or target genes, and the real-time polymerase chain reaction (PCR) visualisation method. These were added as covariates to the meta-regression models.

Main results: We included 41 studies evaluating nine index tests (conventional PCR (in short: PCR), real-time PCR, nested PCR, PCR performed twice, loop-mediated isothermal amplification, enzyme-linked immunosorbent assay (ELISA), dot-ELISA, immunochromatography-based lateral flow assay, and dipstick assay) with 5981 participants (1834 with and 4147 without leptospirosis). Methodological quality criteria were often not reported, and the risk of bias of the reference standard was generally considered high. The applicability of findings was limited by the frequent use of frozen samples. We conducted meta-analyses for the PCR and the real-time PCR on blood products.The pooled sensitivity of the PCR was 70% (95% CI 37% to 90%) and the pooled specificity was 95% (95% CI 75% to 99%). When studies with a high risk of bias in the reference standard domain were excluded, the pooled sensitivity was 87% (95% CI 44% to 98%) and the pooled specificity was 97% (95% CI 60% to 100%). For the real-time PCR, we estimated a summary receiver operating characteristic curve. To illustrate, a point on the curve with 85% specificity had a sensitivity of 49% (95% CI 30% to 68%). Likewise, at 90% specificity, sensitivity was 40% (95% CI 24% to 59%) and at 95% specificity, sensitivity was 29% (95% CI 15% to 49%). The median specificity of real-time PCR on blood products was 92%. We did not formally compare the diagnostic test accuracy of PCR and real-time PCR, as direct comparison studies were lacking. Three of 15 studies analysing PCR on blood products reported the timing of sample collection in the studies included in the meta-analyses (range 1 to 7 days postonset of symptoms), and nine out of 16 studies analysing real-time PCR on blood products (range 1 to 19 days postonset of symptoms). In PCR studies, specificity was lower in settings with high leptospirosis prevalence. Other investigations of heterogeneity did not identify statistically significant associations. Two studies suggested that PCR and real-time PCR may be more sensitive on blood samples collected early in the disease stage. Results of other index tests were described narratively.

Authors' conclusions: The validity of review findings are limited and should be interpreted with caution. There is a substantial between-study variability in the accuracy of PCR and real-time PCR, as well as a substantial variability in the prevalence of leptospirosis. Consequently, the position of PCR and real-time PCR in the clinical pathway depends on regional considerations such as disease prevalence, factors that are likely to influence accuracy, and downstream consequences of test results. There is insufficient evidence to conclude which of the nucleic acid and antigen detection tests is the most accurate. There is preliminary evidence that PCR and real-time PCR are more sensitive on blood samples collected early in the disease stage, but this needs to be confirmed in future studies.

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Conflict of interest statement

Conflict of interest

The Leptospirosis Reference Center (AMC), formerly part of the Royal Tropical Institute (KIT), has done multiple diagnostic accuracy studies. Three such studies are included in this review (Gravekamp 1993; Ahmed 2009; Denipitiya 2016).

Financial conflicts

BY: none. SGV: none. AA: none. BJV: none. IN: none. RS: none. MPG: none. RH: none. MG: none. ML: none.

Figures

1
1
Algorithm, assisting with interpretations and conclusions on the outcome of laboratory testing (adapted from Goris 2012). Antibody titres shown in this figure are optimised for leptospirosis cases in the Netherlands. DPO: days postonset of symptoms; IgM ELISA: immunoglobulin M enzyme‐linked immunosorbent assay; MAT: microscopic agglutination test; neg: negative; PCR: real‐time polymerase chain reaction; pos: positive.
2
2
Eligible study designs. 1. Cross‐sectional study; 2. single‐gate case‐control study; 3. two‐gate case‐control study.
3
3
Study flow diagram. MAT: microscopic agglutination test; n: number of records; PCR: polymerase chain reaction.
4
4
All conventional polymerase chain reaction (PCR) studies: risk of bias and applicability concerns. Sukmark 2018 and Widiyanti 2013 were not part of the PCR (blood products) meta‐analysis.
5
5
All conventional polymerase chain reaction (PCR) studies: risk of bias and applicability concerns graph.
6
6
All real‐time polymerase chain reaction (PCR) studies: risk of bias and applicability concerns. Villumsen 2012 BC and Villumsen 2012 U were not part of the real‐time PCR (blood products) meta‐analysis.
7
7
All real‐time polymerase chain reaction (PCR) studies: risk of bias and applicability concerns graph.
8
8
Studies of nested polymerase chain reaction (PCR), PCR performed twice (PCR 2×), loop‐mediated isothermal amplification (LAMP), enzyme‐linked immunosorbent assay (ELISA), dot‐ELISA, immunochromatography‐based lateral flow assay (ICG‐based LFA), and dipstick assay: risk of bias and applicability concerns.
9
9
Studies of nested polymerase chain reaction (PCR), PCR performed twice (PCR 2×), loop‐mediated isothermal amplification (LAMP), enzyme‐linked immunosorbent assay (ELISA), dot‐ELISA, immunochromatography‐based lateral flow assay (ICG‐based LFA), and dipstick assay: risk of bias and applicability concerns
10
10
Forest plot of conventional polymerase chain reaction (PCR) on blood products. Ref test RoB: risk of bias for the 'reference standard' domain.
11
11
Forest plot of conventional polymerase chain reaction (PCR) on urine. Ref test RoB: risk of bias for the 'reference standard' domain.
12
12
Summary ROC plot for conventional polymerase chain reaction (PCR) on blood products. Transparent dots indicate the test accuracy of the individual studies included in the analysis; the black dot indicates the pooled test accuracy. The ellipse around the pooled test accuracy is the 95% confidence region. The size of the transparent dots represents the sample size, with the vertical diameter representing the number of cases and horizontal diameter representing the number of non‐cases.
13
13
Forest plot of conventional polymerase chain reaction (PCR) on whole blood versus conventional PCR on serum.
14
14
Forest plot of real‐time polymerase chain reaction (PCR) on blood products. Ref test RoB: risk of bias for the 'reference standard' domain.
15
15
Summary ROC plot for real‐time polymerase chain reaction (PCR) on blood products. Transparent dots indicate the test accuracy of the individual studies included in the analysis. The solid black line (summary ROC curve) is a graph of the values of sensitivity and specificity that are obtained by varying the threshold across all possible values.The size of the transparent dots represents the sample size, with the vertical diameter representing the number of cases and horizontal diameter representing the number of non‐cases.
16
16
Forest plot of nested polymerase chain reaction (PCR) on serum. Ref test RoB: risk of bias for the 'reference standard' domain.
17
17
Forest plot of loop‐mediated isothermal amplification (LAMP), on whole blood, plasma or urine. Ref test RoB: risk of bias for the 'reference standard' domain.
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References to other published versions of this review

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