Development of Conventional Multiplex PCR: A Rapid Technique for Simultaneous Detection of Soil-Transmitted Helminths.

Vivornpun Sanprasert,Kobpat Phadungsaksawasdi,Siriporn Srirungruang,Surang Nuchprayoon,Sarit Charuchaibovorn,Ruthairat Kerdkaew

doi:10.3390/pathogens8030152

Vivornpun Sanprasert, Kobpat Phadungsaksawasdi + Show 4 more

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https://doi.org/10.3390/pathogens8030152

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Abstract

Soil-transmitted helminths (STHs) are the most common intestinal parasites infecting humans worldwide. STH infections are a major cause of morbidity and disability. Accurate diagnostic tools are pivotal for assessing the exact prevalence of parasitic infections. Microscopic examination and culture techniques have been used to observe the presence of eggs or larvae of parasites in stool samples, but they are time-consuming and have low sensitivity. Therefore, accurate, simple, and inexpensive diagnostic techniques are still required for simultaneous detection of STH infections. Although molecular-based techniques, such as real-time PCR and multiplex real-time PCR, have been developed, they are not suitable for routine diagnosis due to the requirement for expensive reagents and instruments. In this study, we established a conventional multiplex PCR for simultaneous rapid detection of Ascaris lumbricoides, Necator americanus, and Strongyloides stercoralis in stool samples. Our results show that the multiplex PCR could detect the DNA of STHs at a very low target gene concentrations (lower than 1 pg) with no cross-amplification. Multiplex PCR had five times higher sensitivity than the formalin–ethyl acetate concentration technique (FECT) in the detection of multiple infections, and two times higher for detection of S. stercoralis. However, multiplex PCR was comparable to FECT in the detection of A. lumbricoides and N. americanus. In conclusion, this method could be used as an alternative method for the detection of STHs, especially for S. stercoralis.

Highlights

Soil-transmitted helminths (STHs) or geohelminths pose a major public health threat to at least2 billion people each year around the world
STH infections are associated with malnutrition, cognitive impairment, and health problems in children, as suggested by the World Health Organization (WHO)
We developed a conventional multiplex polymerase chain reaction (PCR) for simultaneous infections of A. lumbricoides, N. americanus, and S. stercoralis, providing an efficient and convenient technique for epidemiological and clinical studies

Summary

Introduction

Soil-transmitted helminths (STHs) or geohelminths pose a major public health threat to at least2 billion people each year around the world. STH infections are associated with malnutrition, cognitive impairment, and health problems (e.g., intestinal obstruction and anemia) in children, as suggested by the World Health Organization (WHO). These parasites are typically found in tropical and sub-tropical countries where sanitation and hygiene are lacking, especially in low- and middle-income countries [1]. These parasites are transmitted and co-infected via the fecal–oral route (food or water containing infective eggs or infective larvae) or skin surface contact (hookworms and S. stercoralis) [1,3,4]. This technique has poor sensitivity for S. stercoralis infections [8,9,10]

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