Detection of Wuchereria bancrofti DNA in paired serum and urine samples using polymerase chain reaction-based systems.

Camila Ximenes,Paulo Araujo,Tamisa Rego,Fabio L Melo,Rafael Medeiros,Luiz Carvalho,Eduardo Brandão,Paula Oliveira,João Ferraz,Christian Reis,Ana Aguiar-Santos,Abraham Rocha

doi:10.1590/0074-0276140155

Abstract

The Global Program for the Elimination of Lymphatic Filariasis (GPELF) aims to eliminate this disease by the year 2020. However, the development of more specific and sensitive tests is important for the success of the GPELF. The present study aimed to standardise polymerase chain reaction (PCR)-based systems for the diagnosis of filariasis in serum and urine. Twenty paired biological urine and serum samples from individuals already known to be positive for Wuchereria bancrofti were collected during the day. Conventional PCR and semi-nested PCR assays were optimised. The detection limit of the technique for purified W. bancrofti DNA extracted from adult worms was 10 fg for the internal systems (WbF/Wb2) and 0.1 fg by using semi-nested PCR. The specificity of the primers was confirmed experimentally by amplification of 1 ng of purified genomic DNA from other species of parasites. Evaluation of the paired urine and serum samples by the semi-nested PCR technique indicated only two of the 20 tested individuals were positive, whereas the simple internal PCR system (WbF/Wb2), which has highly promising performance, revealed that all the patients were positive using both samples. This study successfully demonstrated the possibility of using the PCR technique on urine for the diagnosis of W. bancrofti infection.

Highlights

Lymphatic filariasis (LF) has the Wuchereria bancrofti nematode as its main etiological agent and is transmitted by Culicidae, which live in the tropical and subtropical regions of 83 countries and infect approximately 120 million individuals around the world, placing 1.2 billion people at risk of being infected by this parasite (Chandy et al 2011, WHO 2011)
This study successfully demonstrated the possibility of using the polymerase chain reaction (PCR) technique on urine for the diagnosis of W. bancrofti infection
The development of more specific and sensitive tests are important for the GPELF, allowing to (i) suggest which areas should be involved in mass drug administration (MDA), (ii) measure the efficacy of the intervention, (iii) help to decide when to stop MDA and (iv) suggest how to monitor populations after the ending of MDA, thereby preventing the re-occurrence of transmission of the parasite (Weil & Ramzy 2006, WHO 2008)

Summary

Introduction

Lymphatic filariasis (LF) has the Wuchereria bancrofti nematode as its main etiological agent and is transmitted by Culicidae, which live in the tropical and subtropical regions of 83 countries and infect approximately 120 million individuals around the world, placing 1.2 billion people at risk of being infected by this parasite (Chandy et al 2011, WHO 2011). The use of DNA investigation by polymerase chain reaction (PCR) for diagnosis of W. bancrofti infection has been presented by various authors (Zhong et al 1996, Rocha et al 2002). The aim of the present study was to standardise PCRbased systems for the diagnosis of bancroftian filariasis in serum and urine samples and as a potential assessment of interventions proposed by the GPELF. Target population and ethical considerations - All the individuals came from Recife, metropolitan region in the Brazilian state of Pernambuco and they were at-

Objectives

Results

Conclusion