Abstract
In South America, various species of Leishmania are endemic and cause New World tegumentary leishmaniasis (NWTL). The correct identification of these species is critical for adequate clinical management and surveillance activities. We developed a real-time polymerase chain reaction (PCR) assay and evaluated its diagnostic performance using 64 archived parasite isolates and 192 prospectively identified samples collected from individuals with suspected leishmaniasis enrolled at two reference clinics in Lima, Peru. The real-time PCR assay was able to detect a single parasite and provided unambiguous melting peaks for five Leishmania species of the Viannia subgenus that are highly prevalent in South America: L. (V.) braziliensis, L. (V.) panamensis, L. (V.) guyanensis, L. (V.) peruviana and L. (V.) lainsoni. Using kinetoplastid DNA-based PCR as a gold standard, the real-time PCR had sensitivity and specificity values of 92% and 77%, respectively, which were significantly higher than those of conventional tests such as microscopy, culture and the leishmanin skin test (LST). In addition, the real-time PCR identified 147 different clinical samples at the species level, providing an overall agreement of 100% when compared to multilocus sequence typing (MLST) data performed on a subset of these samples. Furthermore, the real-time PCR was three times faster and five times less expensive when compared to PCR - MLST for species identification from clinical specimens. In summary, this new assay represents a cost-effective and reliable alternative for the identification of the main species causing NWTL in South America.
Highlights
The leishmaniases are a globally widespread group of vectorborne diseases that are endemic to 88 countries and affect an estimated 12 million people, with approximately 350 million people at risk worldwide [1]
Identification of Leishmania species traditionally relies on the multilocus enzyme electrophoresis (MLEE) assay, but it can only be applied to culture-positive samples and takes at least six weeks of intense laboratory work
This paper describes a novel real-time polymerase chain reaction (PCR) assay for identification of the five main species that cause tegumentary leishmaniasis in the New World
Summary
The leishmaniases are a globally widespread group of vectorborne diseases that are endemic to 88 countries and affect an estimated 12 million people, with approximately 350 million people at risk worldwide [1]. Depending on the parasite species and host genetic background, infection can range from self-healing cutaneous ulcers to disfiguring mucocutaneous forms and lethal visceral disease [2]. In South America, New World tegumentary leishmaniasis (NWTL) is mainly caused by species of the Viannia complex. In Peru, 70% of the country’s region is affected with leishmaniasis, 96% of which are caused by three species: L. Diagnosis of leishmaniasis is based on criteria that consider epidemiological data, clinical features and laboratory test results [2]. Most laboratory methods are based on findings of the etiological agent microscopically or by culture; they have relatively low sensitivity and they do not identify the infecting species [2]. Correct diagnosis of leishmaniasis should be based on clinical suspicion of leishmaniasis with a confirmed laboratory diagnosis, where the physician must take into account the reliability of the given laboratory result
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