Phlebotomine Fauna (Diptera: Psychodidae) and Infection by Leishmania spp. in Forest Fragments of a University Campus, Western Amazon.

Forest fragments consist of important ecosystems for the maintenance of sand fly populations and Leishmania hosts. This study sought to identify the phlebotomine fauna and its infection by Leishmania spp. in forest fragments on the campus of the Federal University of Acre (UFAC), Western Amazon. Monthly collections with CDC traps were carried out from March 2020 to June 2021, in four forest fragments of UFAC. Male and female insects were processed and identified at species level. A sample of females was subjected to polymerase chain reaction (PCR) analysis to verify the presence Leishmania DNA. In total, 465 specimens were collected, of which 238 were males and 227 were females. The most frequent species were Nyssomyia antunesi (Coutinho, 1939) (47.3%), Trichophoromyia sp. (Mangabeira, 1942) (18.70%), and Ny. whitmani (Antunes & Coutinho, 1939) (8.81%). Molecular analysis detected the presence of Leishmania (Ross, 1903) DNA in a specimen of Ny. antunesi, and another one of Evandromyia walker (Newstead, 1914). The forest fragments of the university campus harbor a diverse sand fly fauna with the presence of Leishmania DNA in these insects, in addition to the presence of other species considered incriminated vectors of Leishmania parasites.

Remarkable diversity, new records and Leishmania detection in the sand fly fauna of an area of high endemicity for cutaneous leishmaniasis in Acre state, Brazilian Amazonian Forest.

The species richness of Amazonian phlebotomines is considered to be one of the highest in the world. In the present study, we investigated the richness and diversity of phlebotomine fauna in Xapuri city, Acre state, Western Brazilian Amazonia, which is an area that is highly endemic for cutaneous leishmaniasis. Sand fly collections were performed monthly from August 2013 to July 2015 (288 h total of sampling effort) in intradomiciliary, peridomiciliary, and forested environments of two localities. Collected females were dissected, microscopically examined for flagellates in their guts, and preserved in ethanol. A total of 21,197 specimens comprising 14 genera and 57 species were collected, and the majority of these were Nyssomyia, Psychodopygus, and Trichophoromyia genera. Three new records of phlebotomine species for Acre are presented here, including Brumptomyia brumpti, Psathyromyia pradobarrientosi, and for the first time in Brazil, Th. omagua. In Xapuri, the phlebotomine fauna of different ecotopes was varied in regard to abundance, diversity, and frequency, and they included proven and permissive vectors of Leishmania spp. The fauna discovered in the forested areas (57 species) was richer and more diverse than was that (33 species) identified in the peri­ and intra-domiciles. The identification of Leishmania subgenera that were present in sand fly guts according to SSU rRNA sequences revealed ten and three species harboring Leishmania of subgenera Viannia and Leishmania (most likely Leishmania amazonensis), respectively. The presence of Leishmania (Leishmania) in sand flies are reported here for the first time in Acre. The presence of L. (Viannia) spp. in Brumptomyia sp. and Lutzomyia sherlocki. and the occurrence of mixed infections with Leishmania of both subgenera in Ps. lainsoni have been reported for the first time in Brazil. Taken together, data from previous studies and from the present study highlight the remarkable complexity of phlebotomine fauna that is possibly due to the well-preserved Xapuri forested areas sustaining vital economic activities of plant extraction and ecological tourism. Our findings also provide new insights into the ongoing adaptation of Trichophoromyia and Psychodopygus species to human habitats.

First report of an Onchocercidae worm infecting Psychodopygus carrerai carrerai sandfly, a putative vector of Leishmania braziliensis in the Amazon.

Sandflies are insects of public health interest due to their role as vectors of parasites of the genus Leishmania, as well as other pathogens. Psychodopygus carrerai carrerai is considered an important sylvatic vector of Leishmania (Viannia) braziliensis in Amazonia. In this study, sandflies were collected in a forested area in the Xapuri municipality, in the State of Acre (Northern Brazil). Two Ps. carrerai carrerai females were found parasitized with a larval form of a filarial worm, one in the labium of the proboscis, the other after the head was squashed, suggesting they were infective larvae. Sandflies were identified through morphological characters as well as amplification and sequencing of the cytochrome oxidase gene (COI). This was the first sequence obtained for Ps. carrerai carrerai for this marker. The obtained nematodes were also characterized through direct sequencing of a fragment of COI and 12S genes, both mitochondrial, and ITS1, a nuclear marker. Phylogenetic analyses revealed that the filarial nematodes belong to a species without sequences for these markers in the database, part of family Onchocercidade and closely related to genus Onchocerca (12S tree). Although sandfly infection with nematodes including members of the Onchocercidae has been reported in the Old World, this is the first report of sandfly infection by a member of the Onchocercidae family in the New World, to the best of our knowledge. Considering that the phylogenetic relationships and location in the insect, it can be expected that this is a parasite of mammals and the transmission cycle should be clarified.

Ecology, feeding and natural infection by Leishmania spp. of phlebotomine sand flies in an area of high incidence of American tegumentary leishmaniasis in the municipality of Rio Branco, Acre, Brazil.

BACKGROUND: Phlebotomine sand flies (Diptera: Psychodidae) are insects of medical importance due to their involvement in the zoonotic transmission of Leishmania spp. to vertebrates. The aim of this work was to study the ecology of the sand fly fauna of two types of environments, a rural environment (the Transacreana Road) and an urban park (Horto Florestal Park), both located in the municipality of Rio Branco in the state of Acre, Brazil. Additionally, this study intended to investigate Leishmania infection and blood meal sources of these sand flies using molecular techniques. METHODS: The sand fly fauna was studied in different environments (i.e. forest and peridomestic environments in a rural area, and an urban forest) using Shannon traps and HP light traps to collect sand fly specimens over 13 consecutive months (December 2014 to January 2016). For investigating natural infection by Leishmania and the source of sand fly blood meals, DNA samples were extracted from female sand flies and subjected to polymerase chain reaction targeting ITS1 and cytb genes. DNA sequencing was subsequently used to identify species of Leishmania and the source of blood meals. RESULTS: A total of 2515 individual sand flies of 43 species were collected and identified, Trichophoromyia auraensis (839; 33.35%), Trichophoromyia spp. (537; 21.35%) and Evandromyia saulensis (187; 7.43%) were more abundant in the rural area (S = 41 species) than in the urban forest. No significant differences were found in species richness between forest and peridomestic environments in the rural area (H = 0.04; P > 0.05), but a larger number of species was found in the forest. Leishmania DNA was sequenced in 13 samples, confirming the presence of L. (V.) braziliensis in Th. auraensis (n = 1), Ev. saulensis (n = 2), Ev. walkeri (n = 1), Ps. llanosmartinsi (n = 1), Pi. nevesi (n = 2), Ps. davisi (n = 1), Ps. ayrozai (n = 1), Pa. aragaoi (n = 1), Ny. antunesi (n = 1) and Ev. infraspinosa (n = 1). Only Ps. ayrozai possessed a sequence similar to that of L. (V.) guyanensis (99%). Through microscopic analysis, five specimens of Ev. saulensis were found to possess flagellate forms in the hindgut, with an infection rate of 2.4%. Samples from 33 fed females were submitted to cytb gene amplification, for which sequencing determined that all were similar to the sequence deposited on GenBank for Gallus gallus (domestic chicken). CONCLUSIONS: The high abundance of Trichophoromyia auraensis and Ev. saulensis, and the detection of L. (V.) braziliensis DNA, suggests that both species may be vectors of American tegumentary leishmaniasis. Psychodopygus ayrozai was found to be infected by L. (V) braziliesnsis and L. (V.) guyanensis, and although collected in low abundance, it may be a potential vector in the region. The sand fly fauna was found to be rich and diverse with predominance of the genus Psychodopygus. Identification of food sources of fed females showed that 100% amplified a gene region compatible with the domestic chicken, which although considered refractory in the disease transmission cycle, may have an influence on the population dynamics of sand flies.

Attractiveness of black and white modified Shannon traps to phlebotomine sandflies (Diptera, Psychodidae) in the Brazilian Amazon Basin, an area of intense transmission of American cutaneous leishmaniasis.

In the Amazon region the phlebotomine fauna is considered one of the most diverse in the world. The use of Shannon traps may provide information on the anthropophily of the species and improve the traps' performance in terms of diversity and quantity of insects collected when white and black colored traps are used together. This study sought to verify the attractiveness of the traps to the phlebotomine species of the Brazilian Amazon basin using Shannon traps under these conditions. The insects were collected using two Shannon traps installed side by side, one white and the other black, in a primary forest area of the municipality of Xapuri, Acre, Brazil. Samples were collected once a month during the period August 2013 to July 2015. A sample of females was dissected to test for natural infection by flagellates. A total of 6,309 (864 males and 5,445 females) specimens (36 species) were collected. Psychodopygus carrerai carrerai (42%), Nyssomyia shawi (36%), and Psychodopygus davisi (13%), together represented 90% of the insects collected. Nyssomyia shawi and Psychodopygus davisi were more attracted by the white color. Specimens of Nyssomyia shawi, Nyssomyia whitmani, and Psychodopygus hirsutus hirsutus were found naturally infected by flagellates in the mid and hindgut. This is the first study in Acre state using and comparing both black and white Shannon traps, demonstrating the richness, diversity, and anthropophilic behavior of the phlebotomine species and identifying proven and putative vectors of the etiological agents of leishmaniasis.