Combined PCR and MAT improves the early diagnosis of the biphasic illness leptospirosis

Abstract

The diagnosis of leptospirosis remains a challenge due to its non-specific symptoms and the biphasic nature of the illness. A comprehensive diagnosis that includes both molecular (polymerase chain reaction (PCR)) and serology is vital for early detection of leptospirosis and to avoid misdiagnosis. However, not all samples could be subjected to both tests (serology and molecular) due to budget limitation, infrastructure, and technical expertise at least in resource-limited countries. We evaluated the usefulness of testing the clinically suspected leptospirosis cases with both techniques on all samples collected from the patients on the day of admission. Among the 165 patient's blood/serum samples tested (from three hospitals in Central Malaysia), 43 (26%) showed positivity by microscopic agglutination test (MAT), 63 (38%) by PCR, while 14 (8%) were positive by both MAT and PCR. For PCR, we tested two molecular targets (lipL32 by qPCR and 16S rDNA or rrs by nested PCR) and detected lipL32 in 47 (29%) and rrs gene in 63 (38%) patients. The use of more than one target gene for PCR increased the detection rates. Hence, a highly sensitive multiplex PCR targeting more than one diagnostic marker is recommended for the early detection of Leptospira in suspected patients. When the frequencies for positivity detected either by MAT or PCR combined, leptospirosis was diagnosed in a total of 92 (56%) patients, a higher frequency compared to when samples were only tested by a single method (MAT or PCR). The results from this study suggest the inclusion of both serology and molecular methods for every first sample irrespective of the days post-onset of symptoms (DPO) collected from patients for early diagnosis of leptospirosis.

Leptospirosis as a cause of fever associated with jaundice in the Democratic Republic of the Congo

“…Several authors have recently recommended combining serological tests with PCR (Polymerase Chain Reaction). PCR is a complementary test, especially when no specific antibodies were detected by serological methods and allows to detect more cases than a serological test would have missed [41][42][43]. We believe that this also contributed to the underestimation of the results reported in this study.…”

Predictors of severe leptospirosis: a multicentre observational study from Central Malaysia

“…A prospective multicenter observational study was performed on patients with clinically suspected leptospirosis. Among the 165 suspected patients, 92 were confirmed leptospirosis [ 1 , 19 ]. The laboratory confirmation of these patients is defined as positive by polymerase chain reaction (PCR) or microscopic agglutination test (MAT) (first sample > 1:400; seroconversion or fourfold increase in the paired samples) [ 16 ].…”

“…The samples positive by 16S rDNA were subjected to sequencing (MyTACG Bioscience Enterprise, Malaysia) to identify the infecting Leptospira species [ 1 ]. The serovars used for MAT assay comprised of Australis, Autumnalis, Batavia, Canicola, Celledoni, Grippotyphosa, Hardjoprajitno, Icterohaemorrhagiae, Javanica, Pyrogenes, Tarrasovi, Djasiman, Patoc, Pomona (international serovars were obtained from World Health Organization (WHO)) and IMR LEP 1; saprophyte, IMR LEP 115; saprophyte, IMR LEP 175; saprophyte, IMR LEP 803/11-Copenhageni, IMR LEP 27-Hardjobovis, IMR LEP 22-Lai (local serovars were obtained from the Institute for Medical Research (IMR), Federal Territory of Kuala Lumpur, Malaysia [ 19 , 20 ]. Among the 92 confirmed leptospirosis, a total of 83 patients with complete clinical admission data were recruited for classification of mild and severe (severe included dead patients) categories.…”

Predictors of Severe Leptospirosis: A Multicentre Observational Study From Central Malaysia

“…The samples positive by 16S rDNA were subjected to sequencing (MyTACG Bioscience Enterprise, Malaysia) to identify the infecting Leptospira species [1]. The serovars used for MAT assay comprised Australis, Autumnalis, Batavia, Canicola, Celledoni, Grippotyphosa, Hardjoprajitno, Icterohaemorrhagiae, Javanica, Pyrogenes, Tarrasovi, Djasiman, Patoc, Pomona (international serovars were obtained from World Health Organization, WHO) and IMR LEP 1; saprophyte, IMR LEP 115; saprophyte, IMR LEP 175; saprophyte, IMR LEP 803/11-Copenhageni, IMR LEP 27-Hardjobovis, IMR LEP 22-Lai (local serovars were obtained from the Institute for Medical Research (IMR), Federal Territory of Kuala Lumpur, Malaysia [19,20].…”