The aims of the study were to determine the blood feeding preferences of sandflies and to identify species of Leishmania that infected phlebotomines in Caxias, Maranhão, Brazil, an area that is highly endemic for leishmaniasis. Sandflies were captured in light traps located in the peridomiciliary environments of randomly selected houses in urban and rural settings between 1800 and 0600 hours on new moon days between March 2013 and February 2015. DNA extracts from 982 engorged female sandflies were submitted to fragment length polymorphism analysis to identify infecting species of Leishmania, and blood sources were identified for 778 of these specimens. Infection by Leishmania infantum was detected in Lutzomyia longipalpis, Lu. whitmani and Lu. termitophila; L. infantum/L. braziliensis in Lu. longipalpis, Lu. whitmani and Lu. trinidadensis; L. shawi in Lu. longipalpis; L. mexicana in Lu. longipalpis; L. braziliensis in Lu. longipalpis and Lu. whitmani; L. guyanensis in Lu. longipalpis and Lu. termitophila; L. amazonensis in Lu. longipalpis and L. lainsoni or L. naiffi in Lu. longipalpis, while Lu. longipalpis and Lu. trinidadensis were infected with unidentified Leishmania sp. Blood sources were identified in 573 individual phlebotomines and the preferred hosts were, in decreasing order, chicken, dog, rodent and human with lower preferences for pig, horse, opossum and cattle. Lu. longipalpis and Lu. whitmani performed mixed feeding on man, dog and rodent, while Lu. longipalpis was the most opportunistic species, feeding on the blood of all hosts surveyed, but preferably on dog/chicken, dog/rodent and rodent/chicken. Our findings reveal the concomitant circulation of Leishmania species that cause visceral leishmaniasis and tegumentary leishmaniasis in the study area, and explain the occurrence of autochthonous human cases of both clinical forms of leishmaniasis in Caxias, Maranhão. The results support our hypothesis that, in the municipality of Caxias, transmission of Leishmania occurs in close proximity to humans.

Leishmaniasis is a serious public health concern in the Northeastern region of Brazil, where the sand fly fauna is well studied, although few species have been identified as competent vectors. The detection of Leishmania spp. parasites in wild-caught sand flies could help sanitary authorities draw strategies to avoid the transmission of the parasites and, therefore, the incidence of leishmaniases. We detected Leishmania DNA in wild-caught sand flies and correlated that data with aspects of sand fly ecology in the Caxias municipality, Maranhao State, Brazil. The sand flies were sampled in the peridomicile (open areas in the vicinity of human residences) and intradomicile (inside the residences) from July/2019 to March/2020. Leishmania DNA was detected in females, targeting a fragment of the Internal Transcribed Spacer (ITS1) from ribosomal DNA. Among the fourteen species of sand flies identified, five ( Lutzomyia longipalpis, Nyssomyia whitmani, Evandromyia evandroi, Micropygomyia trinidadensis, and Micropygomyia quinquefer ) harbored DNA of Leishmania (Leishmania) amazonensis. The most abundant species in rural ( Ny. whitmani : 35.2% and Ev. evandroi : 32.4%) and urban areas ( Lu. longipalpis : 89.8%) are the permissive vectors of L. (L.) amazonensis , especially Ny. whitmani , a known vector of causative agents of cutaneous leishmaniasis. Although Lu. longipalpis is the vector of L. (L.) infantum, which was not detected in this study, its permissiveness for the transmission of L. (L.) amazonensis has been reported. We suspect that visceral leishmaniasis and cutaneous leishmaniasis are caused by L. (L.) amazonensis, and the transmission may be occurring through Lu. longipalpis, at least in the urban area.

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Studies on experimental sand fly infection require the availability of colonies and laboratory conditions. In Brazil, Lutzomyia longipalpis (Diptera: Psychodidae) (Lutz and Neiva 1912) is responsible for the highest infection rates by Leishmania spp. and this species is one of the most suitable species for laboratory colonization. In this study, we describe a method for growing Lu. longipalpis in laboratory conditions (10 generations) from natural populations sampled from a region of high endemicity for visceral leishmaniasis in the state of Maranhão, Northeastern Brazil. Using two methods (individualized or grouped females), the colony’s highest productivity occurred in the first four generations, where all stages presented with high frequency. Nonviable eggs represented more than 50% of the total eggs produced by engorged females, while pupae were more resistant to fungal contamination, with a mortality rate of only 2%. In both methods, there was a predominance of female emergence; however, the ratio between males and females did not show significant differences, IF (P = 0.8023) and GF (P = 0.1831). Using the method of individualized females, the F4 generation took the longest to appear (234 d; 64 ± 57 d); by grouped females, F3 took the longest to appear (102 d; 47 ± 20 d). This method provides sufficient numbers of insects to perform vector competence tests for Leishmania spp. that cause the cutaneous form of leishmaniasis, usually found in Lu. longipalpis sampled from the study location.