ABSTRACT
Objective: To identity the variation of sand flies in the Gampaha and Kurunegala districts of Sri Lanka and to assess DNA barcoding as a complementing method for morphological identification. Methods: A total of 38 441 sand flies were collected from selected localities in Gampaha and Kurunegala districts using standard entomological techniques from May 2017 to December 2018. Specimens were identified using morphological features and compared with mitochondrial cytochrome C oxidase subunit I gene- based DNA barcoding as an alternative tool. Results: Morphological and molecular identification confirmed the presence of four species under two genera (Phlebotomus and Sergentomyia). Phlebotomus argentipes was the predominant species, followed by Sergentomyia (S.) punjabensis, S. babu insularis, and an unidentified Sergentomyia sp. Phlebotomus argentipes showed a clear genetic differentiation from other species. S. babu insularis and S. punjabensis showed a higher genetic affinity to each other than the unidentified species. The unidentified Sergentomyia species is morphologically similar to S. zeylanica, but differs only in clavate gonostyle. Conclusions: DNA barcoding is an effective technique for the identification of sand flies. Further studies using molecular techniques will improve the knowledge of the cryptic diversity of Sri Lankan sand fly fauna. Establishing a reliable and standardized identification system for sand fly species in Sri Lanka is recommended.
ABSTRACT
Objective: To compare the DNA sequences of Leishmania (L.)donovani isolated from individuals in two districts of the Northern Province with other parts of Sri Lanka and neighboring countries. Methods: Samples were collected from military personnel at the Army Hospital, Narahenpita, Sri Lanka from November 2018 to March 2020. A portion of the samples was fixed, stained with Giemsa and observed under the light microscope. The genomic The DNA was extracted from the remaining portion of the samples using DNEasy blood tissue kit (Qiagen, Germany) and amplified using Leishmania genus-specific primers for molecular diagnosis initially. DNA was amplified using L. donovani species-specific primers by PCR and the amplified product was sequenced for comparison of nucleotide sequences. Results: Out of 76 suspected patients, at least one biological sample of 45 (59.2%) was positive for L. amastigotes upon microscopy. Overall, 33 (43.4%) were positive in Leishmania genus-specific PCR, but only 23 (30.3%) were positive in L. donovani specific PCR. The dendrogram indicates that the current sequences clustered together with those from Nepal and Gampaha districts (Western Province), Sri Lanka, while the Indian and Eastern African sequences clustered separately. Conclusions: The genetic diversity was low among the isolates, indicating a single and possibly a local point of origin. However, the similarity of Sri Lankan and Nepal strains indicate a possibility of a shared point of origin, which needs more extensive evidence to confirm.