Serovar prevalence of Leptospira in semirural India and the development of an IgM-based indirect ELISA.

Sara Chandy,Krishnan Venkataraman,Priya Hemavathy,Lokeshwaran Kirubanandhan,Anees Mohammad Khadeeja,Kristine Mørch,Dilip Mathai,Anand Manoharan,Siby Jacob Kurian

doi:10.3855/jidc.8067

Abstract

Leptospirosis is a major public health problem in India. However, it has been underreported and under-diagnosed due to a lack of awareness of the disease, a functional surveillance system, and appropriate laboratory diagnostic facilities. This multicenter study aimed to understand the Leptospira serovars causing leptospirosis in seven secondary-level hospitals in six states in India. Since early and accurate diagnosis of leptospirosis is one of the challenges faced by clinicians in India due to the poor specificity and sensitivity of commercially available diagnostic systems, an in-house indirect enzyme-linked immunosorbent assay (ELISA) was developed. Genomic DNA from L. interrogans serovar Canicola was used for polymerase chain reaction amplification, cloning, and expression of the lipL32 gene in E. coli to amplify, clone, and express the lipL32 gene. Australis was the common serovar seen at all the study centers. Serovar Icterohaemorrhagiae was seen in samples from Tamil Nadu and Assam. In-house ELISA was standardized using the purified recombinant LipL32 polypeptide and was used to evaluate serum. Subsequently, acute serum samples from leptospirosis patients (n = 60) were screened. Compared to the gold standard, the microscopic agglutination test, sensitivity and specificity of the in-house ELISA was 95% and 90%, respectively. Understanding Leptospira serovars circulating in leptospirosis-endemic areas will help to formulate better vaccines. LipL32-based ELISA may serve as a valuable tool for early diagnosis of leptospirosis.

Highlights

Leptospirosis is a major public health problem in India
All patients > 5 years of age admitted with acute undifferentiated fever (AUF) for 2–14 days were recruited from 7 secondary hospitals located at Ambur (Bethesda Hospital) and Oddanchatram (Christian Fellowship Hospital) in Tamil Nadu, Anantapur (Rural Development Trust) in Andhra Pradesh, Ratnagiri (BKL Walawalkar Hospital) in Maharashtra, Mungeli (Christian Hospital) in Chattisgarh, Raxaul
Screening enzyme-linked immunosorbent assay (ELISA) positive (n = 185), discrepant (n = 7), equivocal (n = 3), and negative (n = 51) isolates were tested by the microscopic agglutination test (MAT) (n = 246) and included samples from Ambur (n = 47), Oddanchatram (n = 16), Ratnagiri (n = 54), Tezpur (n = 80), Anantpur (n = 22), Raxual (n = 17), and Mungeli (n = 10) (Table 1)

Summary

Introduction

Leptospirosis is a major public health problem in India. It has been underreported and under-diagnosed due to a lack of awareness of the disease, a functional surveillance system, and appropriate laboratory diagnostic facilities. Compared to the gold standard, the microscopic agglutination test, sensitivity and specificity of the in-house ELISA was 95% and 90%, respectively. Despite being a treatable disease, human leptospirosis is a significant public health problem and is severely neglected in its endemic hotspots of southern and western India [1]. The old phenotypic classification system based on a cross-agglutination absorption test (CAAT) identified approximately 250 serovars among the Leptospira species and serogroups [2]. Presence of serovars varies depending on local animal species and adaptability of serovars to new hosts [5]. Isolation of leptospires is time consuming and has Chandy et al – Novel diagnostic tools for leptospirosis

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Discussion

Conclusion