# Comparative quantification of Leishmania infantum in experimental phlebotomine sand fly infections using kDNA and single-copy Meta-1 gene qPCR assays

## TL;DR

This study compares two qPCR methods to measure Leishmania infantum in sand flies, showing that combining them improves accuracy and sensitivity in tracking parasite transmission.

## Contribution

The novel contribution is demonstrating that combining kDNA and Meta-1 qPCR assays enhances quantification of Leishmania in sand fly vectors.

## Key findings

- Combining kDNA and Meta-1 qPCR assays improves sensitivity and accuracy in quantifying Leishmania infections.
- Significant differences in parasite proliferation were observed among sand fly species and Leishmania strains.
- Phlebotomus orientalis is confirmed as a highly competent vector for Leishmania donovani complex.

## Abstract

Leishmaniasis is a parasitic disease transmitted by female sand flies and caused by protozoan parasites of the genus Leishmania. Accurate quantification of parasite load within vectors is essential for understanding transmission dynamics and vector competence. This study compares two quantitative polymerase chain reaction (qPCR) methods for detecting and quantifying Leishmania infantum in three experimentally infected sand fly species (Phlebotomus perniciosus, Phlebotomus argentipes, and Phlebotomus orientalis).

One method targets kinetoplast minicircle DNA, which offers high sensitivity but limited quantitative precision, while the other targets the single-copy Meta-1 gene, providing more precise quantification but reduced sensitivity in low-level infections.

A positive correlation between the two molecular markers supports a combined approach to maximize both sensitivity and accuracy in surveillance and transmission studies. Following this methodological comparison, significant differences were observed in parasite proliferation among sand fly species and L. infantum strains, with Ph. orientalis confirmed as a highly competent vector for Leishmania donovani complex.

Together, these findings highlight that combining high-sensitivity (kinetoplast DNA [kDNA]) and single-copy (Meta-1) targets enables both accurate and sensitive quantification of Leishmania infections in sand flies, improving the assessment of parasite–vector interactions.

The online version contains supplementary material available at 10.1186/s13071-025-07231-x.