Population dynamics of phlebotomine sand flies (Diptera: Psychodidae) in cutaneous leishmaniasis endemic areas of Kurunegala District, Sri Lanka.

Sand flies are the primary vectors of leishmaniasis. Disease management with effective vector control depends on the knowledge of vector population dynamics. In Sri Lanka, despite few isolated entomological collections, long-term studies are not reported to date. In the reported study, monthly entomological surveillance was conducted from May 2017 to December 2018 in Polpithigama, Maho, and Galgamuwa Medical Officer of Health (MOH) areas using standard entomological techniques. Climatic data were collected from the Department of Meteorology, Sri Lanka. Patient records were collected from each MOH office. A total of 38,339 sand flies were collected. The majority were Phlebotomus argentipes (99.50%, n = 38,147), while the rest was Sergentomyia punjabensis (0.50%, n = 192). Wind speed reduces sand fly abundance (r = -0.519, P < 0.05) evidently by limiting the movement. Rainfall increases sand fly abundance (r = 0.842, P <0.05, lag = 6 months) probably by improving favorable conditions in breeding sites. The actual effect of humidity is not conclusive without further research. The observed higher prevalence of leishmaniasis in the study sites could be due to the higher sand fly density in these areas. The number of patients shows a strong positive correlation (r = 0.516, P <0.05) to the sand fly abundance with a lag of 7 months. Systematic surveillance of sand flies as a part of general healthcare services is strongly recommended to identify and prevent possible outbreaks through timely planned vector control measures.

Diversity of midgut bacteria in larvae and females of Aedes aegypti and Aedes albopictus from Gampaha District, Sri Lanka.

BACKGROUND: The midgut microbiota of mosquitoes maintain basal immune activity and immune priming. In recent years, scientists have focused on the use of microbial communities for vector control interventions. In the present study, the midgut bacteria of larvae and adults of Aedes aegypti and Ae. albopictus were assessed using both field-collected and laboratory-reared mosquitoes from Sri Lanka. METHODS: Adults and larvae of Ae. aegypti and Ae. albopictus were collected from three selected areas in Gampaha Medical Officer of Health area, Gampaha District, Western Province, Sri Lanka. Bacterial colonies isolated from mosquito midgut dissections were identified by PCR amplification and sequencing of partial 16S rRNA gene fragments. RESULTS: Adults and larvae of Ae. aegypti and Ae. albopictus harbored 25 bacterial species. Bacillus endophyticus and Pantoea dispersa were found more frequently in field-collected Ae. aegypti and Ae. albopictus adults, respectively. The midgut bacteria of Ae. aegypti and Ae. albopictus adults (X2 = 556.167, df = 72, P < 0.001) and larvae (X2 = 633.11, df = 66, P < 0.001) were significantly different. There was a significant difference among the bacterial communities between field-collected adults (X2 = 48.974, df = 10, P < 0.001) and larvae (X2 = 84.981, df = 10, P < 0.001). Lysinibacillus sphaericus was a common species in adults and larvae of laboratory-reared Ae. aegypti. Only P. dispersa occurred in the field-collected adults of Ae. aegypti and Ae. albopictus. Species belonging to genera Terribacillus, Lysinibacillus, Agromyces and Kocuria were recorded from Aedes mosquitoes, in accordance with previously reported results. CONCLUSIONS: This study generated a comprehensive database on the culturable bacterial community found in the midgut of field-collected (Ae. aegypti and Ae. albopictus) and laboratory-reared (Ae. aegypti) mosquito larvae and adults from Sri Lanka. Data confirm that the midgut bacterial diversity in the studied mosquitoes varies according to species, developmental stage and strain (field vs laboratory).

Molecular Characterization of Culturable Aerobic Bacteria in the Midgut of Field-Caught Culex tritaeniorhynchus, Culex gelidus, and Mansonia annulifera Mosquitoes in the Gampaha District of Sri Lanka.

BACKGROUND: Larval and adult mosquito stages harbor different extracellular microbes exhibiting various functions in their digestive tract including host-parasite interactions. Midgut symbiotic bacteria can be genetically exploited to express molecules within the vectors, altering vector competency and potential for disease transmission. Therefore, identification of mosquito gut inhabiting microbiota is of ample importance before developing novel vector control strategies that involve modification of vectors. METHOD: Adult mosquitoes of Culex tritaeniorhynchus, Culex gelidus, and Mansonia annulifera were collected from selected Medical Officer of Health (MOH) areas in the Gampaha district of Sri Lanka. Midgut lysates of the field-caught non-blood-fed female mosquitoes were cultured in Plate Count Agar medium, and Prokaryotic 16S ribosomal RNA partial genes of the isolated bacteria colonies were amplified followed by DNA sequencing. Diversity indices were used to assess the diversity and richness of the bacterial isolates in three mosquito species. The distribution pattern of bacterial isolates between different mosquito species was assessed by Distance-Based Redundancy Analysis (dbRDA). RESULTS: A total of 20 bacterial species (Staphylococcus pasteuri, Bacillus megaterium, Staphylococcus cohnii, Pantoea dispersa, Staphylococcus chromogenes, Bacillus aquimaris, Staphylococcus arlettae, Staphylococcus sciuri, Staphylococcus warneri, Moraxella osloensis, Enterobacter sp., Klebsiella michiganensis, Staphylococcus hominis, Staphylococcus saprophyticus, Streptomyces sp., Bacillus niacin, Cedecea neteri, Micrococcus luteus, Lysinibacillus sphaericus, and Bacillus licheniformis) were identified. All of these species belonged to three phyla, Proteobacteria, Firmicutes, and Actinobacteria, out of which phylum Firmicutes (71.1%) was the most prominent. The least number of species was recorded from Actinobacteria. The relative distribution of midgut microbes in different mosquito species differed significantly among mosquito species (Chi-square, χ 2 = 486.091; df = 36; P ≤ 0.001). Midgut microbiota of Cx. tritaeniorhynchus and Cx. gelidus indicated a similarity of 21.51%, while Ma. annulifera shared a similarity of 6.92% with the cluster of above two species. The gut microbiota of Cx. tritaeniorhynchus was also significantly more diverse and more evenly distributed compared to Ma. annulifera. Simpson's diversity, Margalef's diversity, and Menhinick's diversity indices were higher in Cx. gelidus. Of the recorded species, P. dispersa and strains of nonpathogenic species in Bacillaceae family (B. megaterium, B. niacini, B. licheniformis, and L. sphaericus) can be recommended as potential candidates for paratransgenesis. CONCLUSION: The relative distribution of midgut microbes in different mosquito species differed significantly among the three studied adult mosquito species. The present data strongly encourage further investigations to explore the potential usage of these microbes through paratransgenic approach for novel eco-friendly vector control strategies.

The Diversity of Midgut Bacteria among Wild-Caught Phlebotomus argentipes (Psychodidae: Phlebotominae), the Vector of Leishmaniasis in Sri Lanka.

Phlebotomus argentipes is the main suspected vector for leishmaniasis in Sri Lanka. Investigations on the presence of aerobic bacteria in the gut of sand flies which evidence a potential approach to control leishmaniasis transmission through a paratransgenic strategy are still not available for the local sand fly populations. Field-caught unfed female sand flies collected from three selected Medical Officer of Health (MOH) areas (Polpithigama, Maho, and Galgamuwa) in Kurunegala District, Sri Lanka from August to December 2018 were used. Prokaryotic 16S ribosomal RNA partial gene was amplified and sequenced. Morphological identification revealed the presence of only one sand fly species, P. argentipes (n = 1,969). A total of 20 organisms belonging to two phyla (Proteobactericea and Furmicutes) were detected within the gut microbial community of the studied sand fly specimens. This study documents the first-ever observation of Rhizobium sp. in the midgut of P. argentipes. The presence of Bacillus megaterium, which is considered as a nonpathogenic bacterium with potential use for paratransgenic manipulation of P. argentipes suggest that it may be used as a delivery vehicle to block the vectorial transmission of Leishmania parasites. In addition, Serratia marcescens may be used as a potential candidate to block the parasite development in sand fly vectors since it has evidenced antileishmanial activities in previous investigations. Hence, further studies are required to gain full insight into the potential use of this bacterium in the control of Leishmania parasites through paratransgenesis.

Socioeconomic, demographic and landscape factors associated with cutaneous leishmaniasis in Kurunegala District, Sri Lanka.

BACKGROUND: Leishmaniasis is a neglected tropical disease that affects countries in the developing world. In Sri Lanka, cutaneous leishmaniasis is the most common form of the disease. It is prevalent in dry and intermediate zones, mostly associated with rural settings. Understanding basic risk factors is critical in the management of the disease with effective interventions. This study is focused on assessing the demographic, socioeconomic and landscape factors associated with leishmaniasis in Kurunegala District, Sri Lanka. METHODS: A descriptive cross-sectional study was conducted. Households of the past patients and randomly selected households, which had no history of leishmaniasis cases were interviewed. The clinical, socioeconomic, demographic, landscape and awareness-related data were obtained using a pre-tested, interviewer-administered questionnaire. RESULTS: A total of 101 patients and a similar number of controls were included in the study. All the patients had the cutaneous form of the disease. Housewives and personnel with monthly incomes less than Rs. 10,000 (56.76 USD) were 3.9- and 9.5-times more prone to the disease, respectively, according to multivariate analysis. Presence of decaying garbage, termite hills, unclear areas, wet soil and gardening areas were always associated with the increased odds of acquiring the disease. CONCLUSIONS: Demographic factors do not play a pivotal role in the prevalence of leishmaniasis in the area. Housewives, inhabitants with low incomes and individuals who live in areas with conditions suitable for sand fly breeding and resting are major groups with a higher risk of infection. Special attention must be given in raising awareness and environmental management in control activities.

Potential Challenges of Controlling Leishmaniasis in Sri Lanka at a Disease Outbreak.

The present works reviewed the existing information on leishmaniasis in Sri Lanka and in other countries, focusing on challenges of controlling leishmaniasis in the country, in an outbreak. Evidence from recent studies suggests that there is a possibility of a leishmaniasis outbreak in Sri Lanka in the near future. Difficulty of early diagnosis due to lack of awareness and unavailability or inadequacy of sensitive tests are two of the main challenges for effective case management. Furthermore, the absence of a proper drug for treatment and lack of knowledge about vector biology, distribution, taxonomy and bionomics, and reservoir hosts make the problem serious. The evident potential for visceralization in the cutaneous variant of L. donovani in Sri Lanka may also complicate the issue. Lack of knowledge among local communities also reduces the effectiveness of vector and reservoir host control programs. Immediate actions need to be taken in order to increase scientific knowledge about the disease and a higher effectiveness of the patient management and control programs must be achieved through increased awareness about the disease among general public and active participation of local community in control activities.