Phlebotomine Fauna Research Articles

Phlebotomine Fauna (Diptera: Psychodidae) of Species of Medical Importance from Caves in the States of Minas Gerais, Rio De Janeiro and Sao Paulo, Brazil

This study aimed to update the phlebotomine fauna in caves and compare the species of medical importance with those documented in previous studies conducted in the southeastern region, specifically in Minas Gerais, Rio de Janeiro, and São Paulo. Notably, there is no recorded research on cave fauna in the state of Espírito Santo. This research was conducted based on the review of the Brazilian cave-dwelling phlebotomine fauna published in 2022.

Identification of phlebotomine sand fly (Diptera: Psychodidae) in Atlantic forest fragments and their dispersal to urban area.

The geographical distribution of sand flies in Brazil has been the subject of some studies, yet there is no information about the phlebotomine fauna in João Pessoa, State of Paraíba, Brazil. The aim of this work is to evaluate the occurrence and distribution of sand flies in the Atlantic forest fragments and to evaluate a possible dispersion in 06 nearby districts. Light traps were used during three consecutive nights, supplemented by an aspirator during the dry period and rainy season. A total of 222 sand flies were found, 143 (130 males and 13 females) in the Atlantic forest, and 79 in urban areas. During the entire dry season, three species of phlebotomine sand flies were recorded in 11 forest fragments, Lutzomyia longipalpis, Lu. migonei and Lu. whitmani. During the rainy season, only Lu. longipalpis was found. This was the only species identified in the studied neighborhoods during both seasons. The differences in diversity of sand flies encountered between natural habitats and urban areas may thus be correlated mostly with adaptations to particular habitats and availability of food. One species (Lu. longipalpis) appears to be rapidly adapting to urban areas because of deforestation.

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.

Biodiversity assessment of Phlebotomine (Diptera: Psychodidae) in an environmental impacted area in Rio de Janeiro State, Brazil.

ABSTRACTFrom October 1994 to September 1996, in the municipality of Mangaratiba, Rio de Janeiro State, phlebotomines were captured on the walls of the houses by means of a manual suction tube and light traps, in the household, the peridomicile and in the forest. As it is an area undergoing a real expansion and an increment in the local tourism, with the purpose of assessing changes in the phlebotomine fauna caused by environmental changes in the region, new captures were made in the same location from October 2015 to September 2016, this time using only light traps. In the two phases of the study, a total of 6,681 phlebotomines were captured. The results indicated that Ny. intermedia and Mg. migonei are fully adapted to this anthropic environment, while Pi. fischeri was more abundant and eclectic, and was probably attracted to exercise hematophagy. Nyssomyia intermedia can be suggested as the main vector of the etiological agent of the American Tegumentary Leishmaniasis in the studied areas. Pintomyia fischeri can also transmit Leishmania braziliensis, both in the environment altered by human action, and in the wild environment. Phlebotomines were captured in greater numbers between 6 and 8 pm in the peridomicile and between 9 and 11 pm in the household. The highest densities of phlebotomines were recorded in December, January and February. Despite almost 20 years between the two studies in the municipality of Mangaratiba, there was no change in the profile of the phlebotomine fauna in general; however, there was a greater density of species of medical importance in areas that suffered environmental impacts.

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.

Phlebotomine sand flies (Diptera, Psychodidae) from iron ore caves in the State of Pará, Brazil

The present study aimed to evaluate the distribution of sand fly species in iron ore caves in the State of Pará, Brazil and to associate the richness and abundance of these insects with the capacity of leishmaniasis transmission. Entomological captures were carried out in the years 2010, 2013, 2014 and 2015, throughout active samples with brushes, along the entire caves’ extension, in dry and rainy periods. A total of 9,807 sand flies were counted during the 532 samplings events, being 4,340 in the dry period and 5,467 in the rainy period. A random sample of 802 morphologically identified specimens consisted of 8 genera and 17 species, being 369 males (46%) and 433 females (54%). The predominant species was Sciopemyia sordellii with 60.6% of the total of sand flies collected. Differences in composition and richness were observed between caves located inside of forest and anthropized areas. The mean richness and abundance were different between the wet and rainy periods, with a greater abundance of these insects in the rainy period. The phlebotomine fauna proved to be rich and abundant in the sampled caves, however, environmental degradation seems to be the main factor determining changes in the composition and richness, reinforces the importance of these places as a shelter for sand flies in degraded areas.

Ecological aspects and molecular detection of Leishmania DNA (Kinetoplastida: Trypanosomatidae) in phlebotomine sand flies (Diptera: Psychodidae) from a rural settlement in the Eastern Amazon, Brazil

ABSTRACT Notifications concerning American tegumentary leishmaniasis (ATL) are increasing in the northern areas of Brazil, particularly due to the ongoing increase in human settlements inside the Amazon Forest. Notwithstanding the economical and sanitary importance of the ATL, the ecological aspects of its potential vectors, the Phlebotomine sand flies (Diptera: Psychodidae), remains largely neglected. This study aimed to investigate the diversity of the phlebotomine fauna as well as the detection of Leishmania DNA in these insects in the rural settlement Perimetral Norte, in the state of Amapa, eastern region of the Amazon, endemic region for tegumentary leishmaniasis. Sand flies were collected bimonthly from February 2018 to February 2019, using CDC light traps exposed for three consecutive nights in 10 houses of settlement residents. DNA extraction and multiplex PCR were performed to detect trypanosomatids. A total of 3,946 sandflies belonging to 37 species were collected. The most abundant species were Ty. trichopyga (Floch & Abonnenc, 1945) (21.9%) and Mi. rorotaensis (Floch & Abonnenc, 1944) (16.9%). A total of 36 positive pools were found with Leishmania DNA (Kinetoplastida: Trypanosomatidae), representing 17.9% of the total pools tested (201), those with the highest Minimum infection rates were Mg. migonei (Franca, 1920) and Lu. gomezi (Nitzulescu, 1931), Leishmania DNA was also detected in Bi. flaviscutellata (Mangabeira, 1942), Ny. umbratilis (Ward & Fraiha, 1977), Ny. anduzei (Roseboom, 1942) and Ny. antunesi (Coutinho, 1939). These data confirm the probable participation of these four species in the tegumentary leishmaniasis cycle in the eastern Amazon.

Characterization of the sand fly fauna in Barbalha, one of the municipalities with the highest leishmaniasis rates in Brazil.

Leishmaniases are a complex of sand fly-borne diseases that are considered a public health issue in several countries. Brazil presents high leishmaniases rates. The South of Ceará State, known as Cariri region, shows worrying statistics mainly on American tegumentary leishmaniasis. In Barbalha, which is one of the municipalities in this region, there is still a lack of studies regarding the local phlebotomine (Diptera: Psychodidae) fauna in order to help clarify the high rates. This study aimed to characterize such fauna by capturing sand flies with light traps during a four-year period. A total of 3730 sand flies were captured, of which 37.8% were females. Fourteen species were found: 13 of the Lutzomyia genus and one of the Brumptomyia genus. Of the Lutzomyia species, four were proven and five had potential involvement in leishmaniasis transmission. Lutzomyia longipalpis was the most common species (66.97%). This predominance, especially in the urban area, indicates its epidemiological importance and adaptation to environmental conditions modified by human activity. In fact, further studies are still required to accurately determine the behavioral features of these vectors in order to guide public health measurements towards its control and prevention.

An Inventory of the Sand Flies (Diptera: Psychodidae) of Rudbar County, a New Focus of Leishmaniasis in Northern Iran, with a Taxonomic Note on the Subgenus Larroussius.

Background:Different forms of leishmaniasis are significant infectious diseases in Iran. While, Rudbar County of Guilan Province has been introduced as a new cutaneous leishmaniasis focus, there are few published data about the phlebotomine sand flies (Diptera: Psychodidae) of the province.Methods:To study the phlebotomine fauna of Rudbar County, the sampling was performed in 12 collection sites by light traps, sticky traps and manual aspirators throughout August–December 2015. Sand flies were removed from the sticky traps, rinsed in acetone and stored in 80% ethanol along with the collections of light traps and hand catches.Results:In total, 2186 sand flies were collected and ten species representing two genera were morphologically identified: Phlebotomus (Adlerius) halepensis (0.27%), Ph. (Larroussius) kandeladii (0.10%), Ph. (Lar.) neglectus (0.91%), Ph. (Lar.) perfiliewi (53.88%), Ph. (Lar.) tobbi (43.45%), Ph. (Paraphlebotomus) sergenti (0.82%), Ph. (Phlebotomus) papatasi (0.10%), Sergentomyia (Parrotomyia) baghdadis (0.27%), Se. (Sintonius) clydei (0.05%) and Se. (Sin.) tiberiadis (0.10%). The species Ph. halepensis, Ph. neglectus, Ph. perfiliewi, Se. baghdadis, Se. clydei and Se. tiberiadis were reported for the first time in Guilan Province. This study also verified the presence of Ph. neglectus (Ph. major krimensis as a synonym and morphotype) in Iran. Moreover, the taxonomy of the subgenus Larroussius of the province was discussed.Conclusion:The prevalence of suspected or proven cutaneous and visceral leishmaniasis vectors is noteworthy. The study of ecology of sand flies and detecting the exact vectors of leishmaniasis and phlebotomine fever by molecular specific tests in Guilan Province are recommended.

Correction to ‘Complex ecological interactions across a focus of cutaneous leishmaniasis in Eastern Colombia: novel description of Leishmania species, hosts and phlebotomine fauna’

[This corrects the article DOI: 10.1098/rsos.200266.].

Complex ecological interactions across a focus of cutaneous leishmaniasis in Eastern Colombia: novel description of Leishmania species, hosts and phlebotomine fauna

This study aimed to analyse the patterns of diversity, blood sources and Leishmania species of phlebotomines in a focus of cutaneous leishmaniasis in Arboledas, Eastern Colombia. In total, 1729 phlebotomines were captured in two localities (62.3% Siravita and 37.7% Cinera) and five environments of Norte de Santander. We identified 18 species of phlebotomines: Pintomyia ovallesi (29.8%), Psychodopygus davisi (20.3%), Pi. spinicrassa (18.5%) and Lutzomyia gomezi (15.8%) showed the highest abundance. Species diversities were compared between Cinera (15.00) and Siravita (20.00) and among five microenvironments: forest remnants (19.49), coffee plantations (12.5), grassland (12.99), cane plantations (11.66) and citrus plantations (12.22). Leishmania DNA was detected in 5.8% (80/1380) of females, corresponding mainly to Pi. ovallesi (22/80; 27.2%), Lu. gomezi (17/80; 21.3%) and Pi. spinicrassa (11/80; 13.8%). Leishmania species were 63.1% L. braziliensis, 18.5% L. panamensis, 13.2% L. infantum and 6.1% L. amazonensis. The most frequent feeding sources were Homo sapiens (50%), Bos taurus (13.8%) and Canis lupus familiaris (10.3%). This focus of cutaneous leishmaniasis has a high diversity of Leishmania-carrying phlebotomines that feed on domestic animals. The transmission of leishmaniasis to human hosts was mainly associated with Lu. gomezi, Pi. ovallesi and L. braziliensis.

Morphological Keys for Identification of Sandflies from Morocco (Diptera: Psychodidae: Phlebotominae)

The Moroccan phlebotomine fauna is composed of 24 species whose morphological determination may be difficult for non-taxonomists. The identification keys, presented here, allow the identification of sandflies genera and species in Morocco and will facilitate future studies on these insects of medical importance, Leishmaniasis vectors.

Phlebotomines (Diptera: Psychodidae) from a Urban Park of Belém, Pará State, Northern Brazil and Potential Implications in the Transmission of American Cutaneous Leishmaniasis.

In urban ecotourism parks, the life cycle of American cutaneous leishmaniasis (ACL) agents can remain established, where phlebotomines may comprise potential risks for visitors. The present study aimed to survey the phlebotomine fauna of a forest park ‘Bosque Rodriques Alves-Jardim Botânico da Amazônia’ (BRAJBA), in the urban area of Belém, Brazil. The park was monthly surveyed in 2018 using CDC light traps placed in ground and canopy strata. Leishmania spp. isolated from dissected females were characterized by polymerase chain reaction-restriction fragment length polymorphism analysis (PCR-RFLP) analysis. Fluctuations in specimen capture were correlated with rainfall. Nyssomyia antunesi (Coutinho, 1939) was predominant for all surveyed ecotopes and capture methods in both areas. Females of Ny. antunesi resting on tree bases were observed attempting to bite researchers during early morning. One Bichromomyia flaviscutellata (Mangabeira, 1942) and one Trichophoromyia brachipyga (Mangabeira, 1942) were found naturally infected by flagellates. Only the strain from Th. brachipyga was isolated and characterized as Leishmania (Viannia) lainsoni Silveira, Shaw, Braga and Ishikawa, 1987. Monthly fluctuations of the three most abundant species, Ny. antunesi, Trichophoromyia ubiquitalis (Mangabeira, 1942) and Th. brachypiga, had statistically significant negative correlations with rainfall. The present study provided further information to better understand ACL ecology in the Belém urban area, where the urban parks surveyed appeared to offer potential risk of contracting the disease, thus requiring environmental management. These observations highlighted the need for including Ny. antunesi, Bi. flaviscutellata, Th. ubiquitalis, and Th. brachypiga in the priority list for continuous entomological surveillance.

Phlebotomine (Diptera: Psychodidae) fauna in a cavern containing cave paintings and its surrounding environment, Central-West Brazil

The interior of caves usually presents climate, light, soil, flora and fauna that distinguish it from the external environment. Among the sandfly species found in caves, several may act as vectors of Leishmania spp. and others are a nuisance to humans due to their painful bites. This study sought to identify the phlebotomine fauna in a cavern with cave paintings (“Gruta do Pitoco”) which attract tourists in Alcinopólis municipality, Mato Grosso do Sul state, Brazil. The sandflies were collected from March 2014 to September 2015, with automatic light traps, installed monthly, in the entrance and interior of the cave and outside it. A total of 7645 specimens belonging to 27 species were collected. Lutzomyia dispar, Martinsmyia oliverai and those of the Lu. longipalpis complex (Lu. cruzi and Lu. longipalpis) were the predominant species. Among the three environments sampled, no great difference between species richness was observed. Comparing these environments, the interior of the cave presented the lowest diversity index due to the high frequency of Lu. dispar (46%). For the first time, specimens (only females) of the genus Deanemyia were collected in Mato Grosso do Sul state. Moreover, the study revealed the sympatric occurrence of Lu. cruzi and Lu. longipalpis, both proven vectors of L. infantum, and also Lu. dispar, a highly anthropophilic species, which often causes annoyance to those who visit the cave.

Surveillance of phlebotomine fauna and Didelphis marsupialis (Didelphimorphia: Didelphidae) infection in an area highly endemic for visceral leishmaniasis in Colombia.

The study of the interaction between the parasite, the vector and the mammalian hosts, including man, allows to understand the behavior of the leishmaniases. To determine the presence of Lutzomyia species and to detect the Leishmania infection in Didelphis marsupialis in an endemic area for visceral leishmaniasis. Phlebotomine fauna and individuals of D. marsupialis were collected with CDC and Tomahawk™ traps, respectively. The species of Lutzomyia were identified using the Young and Duncan key (1994). Ear and tail biopsies and blood samples from D. marsupialis were taken to identify the Leishmania species by amplifying a fragment of the gene associated with the 70 kD heat shock protein. Seven Lutzomyia species were identified: Lu. evansi, Lu. gomezi, Lu. panamensis, Lu. dubitans, Lu. cayennensis cayennensis, Lu. rangeliana and Lu. trinidadensis. The first three species have epidemiological importance in Colombia because of their implications in the transmission of the Leishmania parasite. Sixty-five tissue samples from 19 D. marsupialis individuals were negative for Leishmania spp. The presence of the Lutzomyia species that have been identified as vectors for Leishmania inside and around houses in the village of El Bledo, in El Carmen de Bolívar represents a risk of infection. Furthermore, the presence of Lu. panamensis is reported for first time in El Carmen de Bolívar in Colombia. Although the lack of detection of Leishmania spp. in D. marsupialis samples may suggest that D. marsupialis does not play an important role in the transmission cycle of Leishmania in this region, it is necessary to carry out further longitudinal studies to confirm this hypothesis.

Ecology of phlebotomine sand flies in an area of leishmaniasis occurrence in the Xakriabá Indigenous Reserve, Minas Gerais, Brazil.

Leishmaniasis is a complex vector-borne infectious diseases caused by protozoan parasites in the genus Leishmania and spread by hematophagous phlebotomine sand flies (Diptera: Psychodidae, Phlebotominae). The aim of this study was to investigate the phlebotomine fauna, endophily and exophily of the species found, and possible influence of climatic factors on their populations. The study was conducted in the Xakriabá Indigenous Reserve (XIR) in the municipality of São João das Missões in northern Minas Gerais state, Brazil. Insects were collected over three consecutive nights in the last week of each month for 12 months from July 2015 to May 2016 from four houses in four different villages. Two traps were set up in each house: one in the intra-domicile and another in the peri-domicile. A total of 2,012 phlebotomine sand fly specimens representing 23 species and belonging to 10 different genera were captured and identified. Among the studied villages, Riacho do Brejo showed the highest density and diversity of phlebotomine sand flies. The species Lutzomyia longipalpis (80.3%) and Nyssomyia intermedia (7.3%), which are major vectors of visceral and cutaneous leishmaniasis, respectively, had the highest population densities, both in the intra- and peri-domicile. No correlation was observed between climatic factors and the density of phlebotomine sand flies. The results of the present study may contribute to a better understanding and targeting of the measures for preventing and controlling leishmaniasis by the authorities responsible for indigenous health.

Three new records of the genus Lutzomyia of the subgenus Helcocyrtomyia (Diptera: Psychodidae: Phlebotominae) from Southwestern Brazilian Amazonia

The sand fly phlebotomine fauna of the Brazilian municipality of Assis Brasil, Acre was sampled between April 2013 and March 2014 using modified CDC light traps. A total of 22,334 sand flies were collected, 41 of them belonging to three species: Lutzomyia gonzaloi, Lu. kirigetiensis and Lu. munaypata, of the subgenus Helcocyrtomyia, which have previously only been recorded in Peru. This paper presents their first reported collection in Brazil.

Disney Trap Attractiveness for the Cortelezzii Series (Diptera: Psychodidae, Phlebotominae) in a Fishing Tourism Area, Bonito Municipality, Mato Grosso do Sul State, Brazil.

Sand fly fauna is frequently sampled using automatic light or Shannon traps, yet few studies have been devoted to investigating the effectiveness of Disney traps baited with live animals as an attractant. This study sought to identify the phlebotomine fauna attracted to Disney traps having hamsters (Mesocricetus auratus) as bait. A hamster-baited Disney trap was installed in a gallery forest located in Águas do Miranda District, Bonito Municipality, Mato Grosso do Sul state, Brazil, from November 2011 to October 2012. A total of 717 phlebotomines were collected, in which male (251) and female (463) specimens of the Cortelezzii series (Diptera: Psychodidae) predominated (99%). Males were attracted from January to July, whereas the females were predominantly attracted from January to May. No significant correlation was observed between the monthly climatic averages of temperature, rains and humidity, and the numbers of insects collected. Although these findings showed the attractiveness of both sexes of the species of the Cortelezzii series to golden hamsters, further studies are needed to investigate the blood meal preferences of these females to other rodents. As rodents have been reported as reservoirs of Leishmania spp. (Kinetoplastida: Trypanosomatidae), research using live-baited traps can reveal feeding preferences of sand flies and the importance of various rodent species in the zoonotic cycle.

Aspects on the ecology of phlebotomine sand flies and natural infection by Leishmania hertigi in the Southeastern Amazon Basin of Brazil

The medical and veterinary importance of sand flies (Diptera: Psychodidae) follow as a result of some species ability to transmit the zoonotic protozoa of the genus Leishmania. Of all municipalities in the state of Mato Grosso, Sinop ranks first in reported cases of American Cutaneous Leishmaniasis (ACL). Sinop urban zone encompasses three permanent forest preservation areas (APPs) that provide refuge for insects and other vertebrate hosts. We assessed ecological parameters and investigated the natural infection by Leishmania spp. of the phlebotomine fauna from four ecotypes with different levels of urbanization in the urban area of Sinop. A total of 62,745 sand flies were collected, of which 52.34% female. Out of 37 species in this study, nine were found to be constant. Sand flies frequency and diversity were highest in APPs (96.85%; 33 species). Lutzomyia dasypodogeton was the most frequent species and exhibited the greatest abundance (SISA=0.977). The neighborhoods around APPs and completely urbanized neighborhoods presented noteworthy ecological similarity. Moreover, eight vector sand fly species with medicalwere identified, and one L. antunesi sample pool was found to be naturally infected with Le. hertigi. We observed a high frequency and diversity of sand flies, including some species that are known to be major vectors of ACL. Further studies are needed on the natural rates of infection in humans, domestic animals, and sylvatic hosts to better comprehend the leishmaniases dynamics.

Visceral leishmaniasis in an environmentally protected area in southeastern Brazil: Epidemiological and laboratory cross-sectional investigation of phlebotomine fauna, wild hosts and canine cases.

BackgroundLeishmaniasis is a rapidly expanding zoonosis that shows increasing urbanization. Concern exists regarding the role of wildlife in visceral leishmaniasis (VL) transmission, due to frequent natural or anthropogenic environmental changes that facilitate contact between wildlife, humans and their pets. The municipality of Campinas, in southeastern Brazil, initially recorded VL in 2009, when the first autochthonous case was confirmed in a dog living in an upscale residential condominium, located inside an environmentally protected area (EPA). Since then, disease transmission remains restricted to dogs inhabiting two geographically contiguous condominiums within the EPA.Methodology/Principal findingsWe conducted a cross-sectional study of the VL focus to investigate Leishmania spp. infection in domestic dogs, wild mammals and sand flies using molecular tools and recommended serological techniques. Canine seroprevalences of 1.5% and 1.2% were observed in 2013 and 2015, respectively. Six insect species, confirmed or suspected vectors or potential transmitters of Leishmania, were identified. Two specimens of the main L. (L.) infantum vector in Brazil, Lutzomyia longipalpis, were captured in the EPA. Natural infection by L. (L.) infantum was recorded in one Expapillata firmatoi specimen and two Pintomyia monticola. Natural infection by L. (L.) infantum and Leishmania subgenus Viannia was also detected in two white-eared opossums (Didelphis albiventris), a known reservoir of VL. Geographical coordinates of each sampling of infected animals were plotted on a map of the EPA, demonstrating proximity between these animals, human residences, including the dogs positive for VL, and forest areas.Conclusions/SignificanceThe EPA, which is inhabited by humans, has an active VL focus. The risk of establishing and maintaining disease transmission foci in similar scenarios, i.e. wild areas that undergo environmental modifications, is evident. Moreover, different epidemiological profiles of VL must be included to elaborate prevention and control measures that consider the particularities of each transmission area.