# Analysis of Phlebotomine sandflies in Laos from 2014–2024: Inventory, description of a new species, screening for Leishmania and detection of Trypanosoma

## TL;DR

This study explores sandfly diversity and potential pathogens in Laos, discovering a new species and detecting unknown Trypanosoma in sandflies.

## Contribution

The discovery of a new sandfly species and the detection of unknown Trypanosoma in sandflies from Laos.

## Key findings

- A total of 25 sandfly species were identified, with the highest diversity in karstic cave environments.
- A new sandfly species, Sergentomyia sutherlandi, was described for the first time.
- Unknown Trypanosoma species were detected in two sandfly species using a Leishmania-targeted PCR.

## Abstract

Phlebotomine sandflies are the principal vectors of leishmaniasis. Laos is a landlocked country bordering Thailand, where autochthonous cases of leishmaniasis have been reported. However, the status of leishmaniases in Laos is unknown due to a lack of testing. In the past decades, very few studies on sandflies and sandfly-borne pathogens have been conducted in Laos. Therefore, the knowledge of sandfly diversity, distribution, and their related pathogens is lacking. We hypothesized that both known and putative sandfly-borne pathogens may silently circulate in Laos but remain undetected. Herein, we aimed to report species diversity data and Leishmania detection among sandflies collected from seven provinces of Laos.

Sandflies were collected using CDC light traps from various habitats, including caves and peri-caves in karstic areas, domestic animal sheds, forests, and plantations, across seven provinces in northern Laos. Sandfly collections were conducted between 2014 and 2024 from different seasons. Sandflies were identified by morphological and molecular techniques. PCR targeting the ITS1 region was used to screen for Leishmania in the sandfly samples. Though a specific Trypanosoma PCR was not used, this PCR is also able to detect the ITS1 region in some Trypanosoma spp.

A total of 3,857 sandflies from 25 species belonging to five genera were collected and examined: Chinius eunicegalatiae, Idiophlebotomus longiforceps, Phlebotomus barguesae, Ph. breyi, Ph. (Adlerius) sp., Ph. kiangsuensis, Ph. mascomai, Ph. seowpohi, Ph. shadenae, Ph. sinxayarami, Ph. stantoni, Sergentomyia anodontis, Se. bailyi, Se. barraudi group, Se. brevicaulis, Se. dvoraki, Se. hivernus, Se. khawi, Se. perturbans, Se. phasukae, Se. sutherlandi n. sp., Se. sylvatica, Se. tambori, Se. gemmea-like (Se. sp 1) and Grassomyia indica. The highest diversity of sandflies was found in karstic areas where sandflies were collected from cave and peri-cave areas. One new sandfly species, Se. sutherlandi n. sp. is described. We also report for the first time in Southeast Asia a Phlebotomus female belonging to the subgenus Adlerius and we also discuss about the taxonomy of Sergentomyia brevicaulis. Although no Leishmania DNA was detected from screened sandflies, unknown Trypanosoma species were detected from 24 individual sandflies: Chinius eunicegalatiae (n = 22) and Idiophlebotomus longiforceps (n = 2) using a PCR assay that was primarily meant to screen for Leishmania.

This study reveals a notable diversity of sandfly species across seven provinces in Laos, with the highest species richness observed in karstic cave environments. While no Leishmania DNA was detected, the unexpected identification of unknown Trypanosoma species in Chinius eunicegalatiae and Idiophlebotomus longiforceps suggests the potential presence of unrecognized trypanosomatid in the region. These findings underscore the need for broader geographic surveillance and increased sampling efforts to improve our understanding of sandfly ecology and the pathogens they may carry in Laos.

Phlebotomine sandflies are best known as insect vectors of arboviruses and leishmaniasis, a parasitic disease of global health concern. While neighboring Thailand has reported locally acquired cases, the status of leishmaniasis in Laos has remained uncertain due to limited surveillance. In this study, sandflies from seven provinces in Laos were surveyed and tested for the presence of Leishmania parasites. The survey revealed 25 sandfly species, with the greatest diversity found in karstic cave environments, and led to the discovery of a new species, Sergentomyia sutherlandi. Although Leishmania parasites were not detected, DNA from an unknown Trypanosoma species was unexpectedly found in two sandfly species using a molecular test originally designed to detect Leishmania. These findings highlight both the rich, previously undocumented sandfly biodiversity in Laos and the potential presence of other parasitic pathogens. This work expands the foundation for future research on vector-borne diseases in Southeast Asia and underscores the need for broader entomological and pathogen surveillance in underexplored regions.