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Parasitological Confirmation and Analysis of Leishmania Diversity in Asymptomatic and Subclinical Infection following Resolution of Cutaneous Leishmaniasis

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Rosales-Chilama, Mariana, Gongora, Rafael E, Valderrama, Liliana, Jojoa, Jimena, Alexander, Neal, Rubiano, Luisa C, Cossio, Alexandra, Adams, Emily ORCID: https://orcid.org/0000-0002-0816-2835, Saravia, Nancy G and Gomez, Maria Adelaida (2015) 'Parasitological Confirmation and Analysis of Leishmania Diversity in Asymptomatic and Subclinical Infection following Resolution of Cutaneous Leishmaniasis'. PLoS Neglected Tropical Diseases, Vol 9, Issue 12, e0004273.

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Abstract

Background
The contribution of individuals with subclinical infection to the transmission and endemicity of cutaneous leishmaniasis (CL) is unknown. Immunological evidence of exposure to Leishmania in residents of endemic areas has been the basis for defining the human population with asymptomatic infection. However, parasitological confirmation of subclinical infection is lacking.

Methods
We investigated the presence and viability of Leishmania in blood and non-invasive mucosal tissue samples from individuals with immunological evidence of subclinical infection in endemic areas for CL caused by Leishmania (Viannia) in Colombia. Detection of Leishmania kDNA was conducted by PCR-Southern Blot, and parasite viability was confirmed by amplification of parasite 7SLRNA gene transcripts. A molecular tool for genetic diversity analysis of parasite populations causing persistent subclinical infection based on PCR amplification and sequence analysis of an 82bp region between kDNA conserved blocks 1 and 2 was developed.

Principal Findings
Persistent Leishmania infection was demonstrated in 40% (46 of 114) of leishmanin skin test (LST) positive individuals without active disease; parasite viability was established in 59% of these (27 of 46; 24% of total). Parasite burden quantified from circulating blood monocytes, nasal, conjunctival or tonsil mucosal swab samples was comparable, and ranged between 0.2 to 22 parasites per reaction. kDNA sequences were obtained from samples from 2 individuals with asymptomatic infection and from 26 with history of CL, allowing genetic distance analysis that revealed diversity among sequences and clustering within the L. (Viannia) subgenus.

Conclusions
Our results provide parasitological confirmation of persistent infection among residents of endemic areas of L. (Viannia) transmission who have experienced asymptomatic infection or recovered from CL, revealing a reservoir of infection that potentially contributes to the endemicity and transmission of disease. kDNA genotyping establishes proof-of-principle of the feasibility of genetic diversity analysis in previously inaccessible and unexplored parasite populations in subclinically infected individuals.

Author Summary
A variable and often high proportion of individuals residing in areas where cutaneous leishmaniasis is endemic are exposed to Leishmania parasites, yet do not develop symptoms of disease. The role of this asymptomatic population in the transmission of disease is unknown and could interfere with the effectiveness of community or population-based control measures. Exposure to Leishmania is indirectly assessed by immunological tests; however, immunological evidence does not discriminate between historical exposure to the parasite and actual presence of parasites without causing clinical manifestations. We sought to determine whether viable Leishmania are present in individuals with immunological evidence of asymptomatic infection. Our results showed that at least 24% of individuals having immunological evidence of subclinical or asymptomatic infection harboured live Leishmania. These individuals may be at risk of activation of disease, or could represent an unperceived reservoir of parasites for vector-borne transmission. Characterization of Leishmania causing asymptomatic infection has not been possible due to technical limits of detection of parasites in low grade infections. We developed a molecular method that allows genotypic analysis of parasites involved in subclinical infection and potentially provides a means to assess their involvement in transmission.

Item Type: Article
Subjects: QW Microbiology and Immunology > Immune Responses > QW 700 Infection. Mechanisms of infection and resistance.
QX Parasitology > Protozoa > QX 70 Mastigophora. (e.g., Giardia. Trichomonas. Trypanosoma. Leishmania)
WR Dermatology > Parasitic Skin Diseases > WR 350 Tropical diseases of the skin. Mucocutaneous leishmaniasis. Leishmaniasis
Faculty: Department: Biological Sciences > Department of Tropical Disease Biology
Digital Object Identifer (DOI): https://doi.org/10.1371/journal.pntd.0004273
Depositing User: Jessica Jones
Date Deposited: 29 Jan 2016 16:12
Last Modified: 15 Jun 2018 10:14
URI: https://archive.lstmed.ac.uk/id/eprint/5570

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