Introduction 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. Methods 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. Results 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 . Conclusion 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.