On the taxonomic status of Phlebotomus breviductus Barretto, 1950 (Diptera: Psychodidae: Phlebotominae).

The taxonomic status of Phlebotomus breviductus Barretto, 1950 was evaluated based on the morphological and morphometric analysis of the male holotype described from Fazenda Jerusalém, located in the municipality of Alegre, Espírito Santo state, Brazil. After measuring, drawing and photographing the holotype, we concluded that the head and wing of this specimen belong to Trichopygomyia sp., but that the thorax and abdomen belong to a different species, Nyssomyia umbratilis, with an anomalous gonostylus having five spines. The holotype slide consists of parts of specimens representing two different species, thus P. breviductus is not a valid species.

Diversity and temporal distribution of sand flies in an endemic area of cutaneous leishmaniasis in Centre-West Colombia.

The community structure of sand flies indicates the level of adaptation of vector species in a region, and in the context of vector management and control, this information allows for identifying the potential risks of pathogen transmission. This study aimed to analyze sand fly diversity and spatial-temporal distribution in an endemic area of cutaneous leishmaniasis. The study was carried out in the Carrizales hamlet (Caldas), between September 2019 and October 2021. The monthly distribution of sand fly species was evaluated through collections with CDC traps. Shannon and evenness indices were calculated and used to compare species frequencies at each house. The association between climatic variables and the frequency of sand flies was evaluated using Spearman's correlation. A total of 6,265 females and 1,958 males belonging to 23 species were found. Low diversity and evenness were observed, with the dominance of Nyssomyia yuilli yuilli (Young & Porter). Ecological and diversity indices did not reveal differences between the houses. The sand fly community was composed of 3 dominant species, Ny. yuilli yuilli, Psychodopygus ayrozai (Barretto & Coutinho), and Ps. panamensis (Shannon), representing 75.8% of the total catches. No statistical association was found between the absolute frequency of sand flies, rainfall, and temperature. The results show one dominant species, this fact has epidemiological relevance since density influences parasite-vector contact. The high densities of sand flies recorded in peri- and intradomiciliary areas highlight the necessity of periodic monitoring of vector populations and control activities to reduce the risk of Leishmania transmission in this endemic area.

Molecular phylogeny of Psychodopygina (Diptera, Psychodidae) supporting morphological systematics of this group of vectors of New World tegumentary leishmaniasis.

New World sandflies are vectors of leishmaniasis, bartonellosis, and some arboviruses. A classification based on 88 morphological characters was proposed 27 years ago when the New World phlebotomines were organized into two tribes Hertigiini and Phlebotomini. The latter was structured into four subtribes (Brumptomyiina, Sergentomyiina, Lutzomyiina, and Psychodopygina) and 20 genera. The subtribe Psychodopygina, including most of the American vectors of tegumentary Leishmania comprises seven genera from which no molecular work has been produced to support this classification. Here, we carried out a molecular phylogeny based on combined sequences (1,334 bp) of two genes: partial 28S rDNA and mtDNA cytochrome b from 47 taxa belonging to the Psychodopygina. The Bayesian phylogenetic reconstruction agreed with the classification based on morphological characters, supporting the monophyly of the genera Psychodopygus and Psathyromyia, whereas Nyssomyia and Trichophoromyia seemed to be paraphyletic. The paraphylies of the two latter groups were exclusively caused by the doubtful position of the species Ny. richardwardi. Our molecular analysis provides additional support to adopt the morphologic classification of Psychodopygina.

Title: La phylogénie moléculaire des Psychodopygina (Diptera, Psychodidae) soutient la systématique morphologique de ce groupe de vecteurs de leishmaniose cutanée du Nouveau Monde. Abstract: Les phlébotomes du nouveau monde sont des vecteurs de leishmaniose, de bartonellose et de certains arbovirus. Une classification basée sur 88 caractères morphologiques a été proposée il y a 27 ans lorsque les phlébotomes du nouveau monde ont été organisés en deux tribus Hertigiini et Phlebotomini. Cette dernière était structurée en quatre sous-tribus (Brumptomyiina, Sergentomyiina, Lutzomyiina et Psychodopygina) et 20 genres. La sous-tribu des Psychodopygina, qui inclut la plupart des vecteurs américains de la leishmaniose cutanée, comprend sept genres mais aucun travail moléculaire n'a été produit pour soutenir cette classification. Dans cet article, nous avons réalisé une phylogénie moléculaire basée sur des séquences combinées (1334 pb) de deux gènes : ADNr 28S partiel et cytochrome b (ADNmt) chez 47 taxons appartenant aux Psychodopygina. La reconstruction phylogénique bayésienne est en accord avec la classification basée sur les caractères morphologiques, soutenant la monophylie des genres Psychodopygus et Psathyromyia, alors que Nyssomyia et Trichophoromyia semblent être paraphylétiques. Les paraphylies des deux derniers groupes sont exclusivement causées par la position douteuse de la seule espèce Ny. richardwardi. Notre analyse moléculaire fournit une raison supplémentaire pour adopter la classification morphologique des Psychodopygina.

Ecological interactions of sand flies, hosts, and Leishmania panamensis in an endemic area of cutaneous leishmaniasis in Colombia.

BACKGROUND: The transmission dynamics of leishmaniasis are complex. There is also a lack of information about the ecological relationships between the vector/host/parasite at a more local and specific level. The Andean region concentrates more than 50% of Colombia's cutaneous leishmaniasis (CL) cases. The study of the ecological interactions of sand flies through the identification of blood sources has provided information on the female's opportunistic behavior, feeding on various hosts. Therefore, this study aimed to determine sand flies' ecological interactions with Leishmania parasites and their blood sources in an endemic area of CL. RESULTS: A total of 4,621 sand flies were collected, comprising 20 species, in which the most abundant were Nyssomyia yuilli yuilli (55.4%), Psychodopygus ayrozai (14.5%) and Ps. panamensis (13.4%). Sequences of 12S gene fragment were analyzed using the BLASTn search tool. Blood-meal source identification was successfully performed for 47 sand flies, detecting seven vertebrate species, human and armadillo being the most frequent. Leishmania DNA was amplified in four female pools, constituted by Ny. yuilli yuilli and Ps. ayrozai, and the identification through RFLP detected Leishmania (Viannia) panamensis in the positive pools. CONCLUSIONS: The interactions between the sand fly species, local mammalian fauna and the Leishmania parasite in this active focus of CL, provide evidence of the potential role of two different species in the maintenance of the parasite transmission, important information for the understanding of the ecoepidemiology and transmission dynamics of the disease in Andean endemic areas. However its necessary further evaluations of the vector and host competence in the transmission and maintenance of Leishmania spp, in these complex and diverse areas.

Detection of Leishmania (Leishmania) infantum RNA in fleas and ticks collected from naturally infected dogs.

The occurrence of the insect vector (sand flies) with low rates of Leishmania infection, as well as autochthonous transmission in the absence of the natural vector in dogs, have been reported. These unexpected data suggest a hypothesis of other arthropods as a possible way of Leishmania transmission. The prevalence of Leishmania (Leishmania) infantum in fleas and ticks collected from dogs with canine visceral leishmaniasis (CVL), as well as parasite viability, were evaluated herein. The presence of L. (L.) infantum was assayed by PCR and ELISA in ectoparasites and biological samples from 73 dogs living in a Brazilian endemic area. As the occurrence of Leishmania DNA in ticks and fleas is expected given their blood-feeding habits, we next investigated whether parasites can remain viable inside ticks. PCR and ELISA confirmed that 83% of the dogs had CVL. Fleas and ticks (nymphs, male and female adults) were collected in 55% and 63% of the 73 dogs, respectively. Out of the 60 dogs with CVL, 80% harbored ectoparasites infected with L. (L.) infantum. The infection rates of the ectoparasites were 23% and 50% for fleas and ticks, respectively. The RNA analysis of the extract from ticks left in laboratory conditions during 7 to 10 days after removal from CVL dogs showed that parasites were alive. In addition, live parasites were also detected inside adult ticks recently molted in laboratory conditions. These findings indicate a higher infection rate of L. (L.) infantum in ticks and fleas, but they do not conclusively demonstrate whether these ticks can act as vectors of CVL, despite the fact that their rates were higher than those previously described in Lutzomyia longipalpis. The presence of viable L. (L.) infantum in ticks suggests the possible importance of dog ectoparasites in CVL dissemination.

Comparison of BG-Lure and BG-Sweetscents attractants for field sampling of phlebotomine sand flies.

Phlebotomines are important vectors of bacteria, viruses and protozoan parasites. Protozoans of the genus Leishmania which cause visceral and cutaneous leishmaniases, are among the most important etiologic agents transmitted by sand flies. Different blends of human and animal volatiles have been evaluated for use in surveillance and/or control of hematophagous insects. With regard to phlebotomine sand flies there are few records of attraction under laboratory and field conditions. This study was carried out at two collecting sites located in a Brazilian town with a high prevalence of cutaneous leishmaniasis. The aim of the present study was to evaluate the effect of adding two kinds of commercial bait which mimic human odors to light traps to attract sand flies and compare the results with those of light traps without any additional bait. The commercial baits, BG-Lure® and BG-Sweetscent®, were developed to catch anthropophilic mosquitoes. Three treatments were evaluated: 1 - HP light trap with BG-Lure® added; 2 - HP light trap with BG-Sweetscent® added and 3- HP light trap without any attractant. A total of 3,682 sand flies were collected during the study. Constrained correspondence analysis was applied to perform ordination of the captured community of sand fly species that could be explained by attractant, sampling site and sex. The most abundant species: Pintomyia nevesi, Nyssomyia whitmani and Nyssomyia antunesi were further investigated by fitting generalized linear mixed models. Only Ny. whitmani showed a slight indication of an increase in catches with BG-Lure®.

Dispersal and survival of Nyssomyia intermedia and Nyssomyia neivai (Diptera: Psychodidae: Phlebotominae) in a cutaneous leishmaniasis endemic area of the speleological province of the Ribeira Valley, state of São Paulo, Brazil.

The dispersal and survival of the phlebotomines Nyssomyia intermedia and Nyssomyia neivai (both implicated as vectors of the cutaneous leishmaniasis agent) in an endemic area was investigated using a capture-mark-release technique in five experiments from August-December 2003 in municipality of Iporanga, state of São Paulo, Brazil. A total of 1,749 males and 1,262 females of Ny. intermedia and 915 males and 411 females of Ny. neivai were marked and released during the five experiments. Recapture attempts were made using automatic light traps, aspiration in natural resting places and domestic animal shelters and Shannon traps. A total of 153 specimens (3.48%) were recaptured: 2.59% (78/3,011) for Ny. intermedia and 5.35% (71/1,326) for Ny. neivai. Both species were recaptured up to 144 h post-release, with the larger part of them recaptured within 48 h. The median dispersion distances for Ny. intermedia and Ny. neivai, respectively, were 109 m and 100 m. The greatest dispersal range of Ny. intermedia was 180 m, while for Ny. neivai one female was recaptured in a pasture at 250 m and another in a pigsty at 520 m, showing a tendency to disperse to more open areas. The daily survival rates calculated based on regressions of the numbers of marked insects recaptured on the six successive days after release were 0.746 for males and 0.575 for females of Ny. intermedia and 0.649 for both sexes of Ny. neivai. The size of the populations in the five months ranged from 8,332-725,085 for Ny. intermedia males, 2,193-104,490 for Ny. intermedia females, 1,687-350,122 for Ny. neivai males and 254-49,705 for Ny. neivai females.

Considerations on the Species Complex of the Cortelezzii series (Diptera: Psychodidae) and Description of Evandromyia chacuensis sp. nov., a New Phlebotomine Species of the Chaco Region, Argentina.

A new species of phlebotomine sand fly is described and illustrated based on male and female specimens collected in rural and semi-urban areas of the Chaco Province, Argentina. A separation of the Cortelezzii series in two species complexes is proposed, as is a species key for its identification. The morphological characters of the new species permit its inclusion in the Cortelezzii series of genus Evandromyia, subgenus Barrettomyia (Diptera: Psychodidae). The species is named Evandromyia chacuensis sp. nov. Szelag, Rosa, Galati, Andrade Fhilo & Salomón (Diptera: Psychodidae).

Entomological Survey of the Mosquitoes in an Area of Ecological TOurism in the Brazilian Amazon Basin.

Mosquitoes have a great epidemiological importance, since females of several species transmit etiological agents of human diseases, such as malaria and arboviruses. In Brazil, the state of Acre has been highlighted in recent years for its high incidence of diseases of relevance to public health, such as malaria, dengue, and leishmaniases, whose etiological agents are transmitted by insect vectors, including mosquitoes. Nevertheless, few studies have been carried out on the mosquito fauna in Acre State; this study aims to identify the culicid fauna of an ecological tourism area of the municipality of Xapuri, Acre State, Brazilian Amazon. Mosquito collections were carried out monthly at the Seringal Cachoeira settlement, between April 2014 and March 2015 using 6 automatic Centers for Disease Control and Prevention light traps, one in each ecotope. During the study period, 1,021 specimens belonging to the genera Aedeomyia, Aedes, Anopheles, Coquillettidia, Culex, Mansonia, Orthopodomyia, Psorophora, Trichoprosopon, and Uranotaenia were collected. The genus Culex was predominant, Culex (Culex) quinquefasciatus being the most frequent species, followed by Cx. (Melanoconion) gnomatos. The importance of these findings is discussed.

[Sand flies in the Aguapeí river floodplain, northwest area of State of São Paulo, Brazil]. / Flebotomíneos de várzea do rio Aguapeí, região noroeste do Estado de São Paulo, Brasil.

OBJECTIVE: To investigate the seasonal variation of sand flies regarding their occurrences and densities. METHODS: The study was conducted in the Aguapeí river floodplain in the state of São Paulo, southeastern Brazil. Sand flies were monthly captured with automatic light traps set up between 18:00 and 7:00 hours, over a period of one year (2004-2005), at two sites: veranda of a domicile and in a forest on the banks of Aguapeí River. The Williams' average was used for estimating the seasonal variation and the Chi-square test for comparison. RESULTS: A total of 35,995 specimens were captured: five Brumptomyia avellari, one Psathyromyia (Xiphomyia) hermanlenti and the rest Nyssomyia neivai, which had the highest frequency during the winter. Ps. hermanlenti is first recorded in state of São Paulo. CONCLUSIONS: The high density of Nyssomyia neivai, a suspected vector of cutaneous leishmaniasis, points out to the risk of transmission of this disease in this setting, especially during drier periods of the year.

An illustrated guide for characters and terminology used in descriptions of Phlebotominae (Diptera, Psychodidae).

Phlebotomine (Diptera, Psychodidae, Phlebotominae) taxonomy has been studied extensively, primarily due to the role of these flies as vectors of various parasites, including species of Leishmania, Bartonella and arboviruses that cause diseases in humans and other vertebrates. We present some topics discussed at a round-table on phlebotomine taxonomy held at the Ninth International Symposium on Phlebotomine Sandflies (ISOPS IX) in Reims, France, in June 2016. To date, approximately one thousand phlebotomine species have been described worldwide, although in varying languages and mostly without standardization of characters and terminology. In the interest of standardization, we list the characters that should minimally be considered in the description of new phlebotomine taxa as well as annotated illustrations of several characters. For these characters, multiple illustrations are provided to show some of the variations. The preferred terms for all pertinent characters are listed as well as their synonyms in English, Portuguese, and French. Finally, we offer an updated list of abbreviations to be used for generic and subgeneric names.

Reproduction of Leishmania (Leishmania ) infantum chagasi in conditioned cell culture growth medium.

Leishmanias can be produced by inoculation in conditioned McCoy cell culture growth medium (CGM). Leishmania (Leishmania) infantum chagasi (100 parasites) grown in NNN medium was inoculated in 2.5 mL CGM, kept in plates (24 wells) and its multiplication was observed for five days (120 hours). After day 5, the medium was saturated with the flagellate forms of the parasite (promastigotes). The reproduction of the leishmanias was observed every 24 hours and the number of parasites was calculated by counting the parasites in a drop of 10 microL and photomicrographed. So the number of Leishmanias was adjusted to 1 mL volume. The advantage of the technique by isolation of Leishmania in CGM demonstrated in this study is its low cost and high efficacy even with a small quantity of parasites (10(2) promastigotes) used as inoculum. Additionally, isolation of the leishmania can be obtained together with an increase in their density (180 times) as observed by growth kinetics, within a shorter time. These results justify the use of this low-cost technique for the isolation and investigation of the behavior and multiplication of Leishmania both in vertebrates and invertebrates, besides offering means of obtaining antigens, whether whole antigens (leishmanias) or the soluble antigens produced by the parasites which may be useful for the production of new diagnostic kits.

Kinetics of growth of Leishmania (Leishmania) chagasi cycle in McCoy cell culture.

The kinetics of growth of Leishmania performed in vitro after internalization of the promastigote form in the cell and the occurrence of the transformation of the parasite into the amastigote form have been described by several authors. They used explants of macrophages in hamster spleen cell culture or in a human macrophage lineage cell, the U937. Using microscopy, the description of morphologic inter-relationship and the analysis of the production of specific molecules, it has been possible to define some of the peculiarities of the biology of the parasite. The present study shows the growth cycle of Leishmania chagasi during the observation of kinetic analysis undertaken with a McCoy cell lineage that lasted for a period of 144 hours. During the process, the morphologic transformation was revealed by indirect immunofluorescence (IF) and the molecules liberated in the extra cellular medium were observed by SDS-PAGE at 24-hour intervals during the whole 144-hour period. It was observed that in the first 72 hours the promastigote form of L. chagasi adhered to the cell membranes and assumed a rounded (amastigote-like) form. At 96 hours the infected cells showed morphologic alterations; at 120 hours the cells had liberated soluble fluorescent antigens into the extra cellular medium. At 144 hours, new elongated forms of the parasites, similar to promastigotes, were observed. In the SDS-PAGE, specific molecular weight proteins were observed at each point of the kinetic analysis showing that the McCoy cell imitates the macrophage and may be considered a useful model for the study of the infection of the Leishmania/cell binomial.

Canine-Based Strategies for Prevention and Control of Visceral Leishmaniasis in Brazil.

Visceral leishmaniasis (VL) is a zoonosis found worldwide. Its incidence has increased in Brazil in recent years, representing a serious public and animal health problem. The strategies applied in Brazil are questionable and are not sufficient to control the disease. Thus, we have compared the efficacy of some of the currently available strategies focused on dogs to prevent and control zoonotic VL in endemic areas by optimizing a mathematical model. The simulations showed that the elimination of seropositive dogs, the use of insecticide-impregnated dog collars, and the vaccination of dogs significantly contribute to reducing the prevalence of infection in both canines and humans. The use of insecticide-impregnated collars presented the highest level of efficacy mainly because it directly affected the force of infection and vector-dog contact. In addition, when used at a coverage rate of 90%, insecticide-impregnated collar was able to decrease the prevalence of seropositive dogs and humans to zero; moreover, because of the easy application and acceptance by the targeted population, these collars may be considered the most feasible for inclusion in public policies among the three simulated measures. Vaccination and euthanasia were efficacious, but the latter method is strongly criticized on ethical grounds, and both methods present difficulties for inclusion in public policies. When we compared the use of euthanasia and vaccination at coverages of 70 and 90%, respectively, the proportion of infected populations were similar. However, on evaluating the implications of both of these methods, particularly the negative aspects of culling dogs and the proportion of animals protected by vaccination, the latter measure appears to be the better option if the total cost is not significantly higher. The comparison of complications and advantages of different control strategies allows us to analyze the optimal measure and offer strategies to veterinary and public health authorities for making decisions to prevent and control zoonotic VL. Hence, improvements in both public and animal health can be achieved in regions with scenarios similar to that considered in the present study; such scenarios are characteristically found in some areas of Brazil and other countries.

Correction: Canine-Based Strategies for Prevention and Control of Visceral Leishmaniasis in Brazil.

[This corrects the article DOI: 10.1371/journal.pone.0160058.].

Redescription of Lutzomyia (Lutzomyia) renei Martins, Falcão & Silva, 1957 (Diptera: Psychodidae: Phlebotominae).

The male genitalia of Lutzomyia (Lutzomyia) renei (Martins, Falcão & Silva, 1957) have four bristles, three fine and one semi-foliaceous, inserted basomesally on the gonocoxite. Nonetheless, in the original description and in other taxonomic studies, these bristles have been illustrated and described in varying formats. In order to clarify the morphology of this species, both sexes are here redescribed based on three males and one female from the type series. A lectotype and two paralectotypes are here designated.

Recent advances in phlebotomine sand fly research related to leishmaniasis control.

Phlebotomine sand flies are the subject of much research because of the role of their females as the only proven natural vectors of Leishmania species, the parasitic protozoans that are the causative agents of the neglected tropical disease leishmaniasis. Activity in this field was highlighted by the eighth International Symposium on Phlebotomine Sand flies (ISOPS) held in September 2014, which prompted this review focusing on vector control. Topics reviewed include: Taxonomy and phylogenetics, Vector competence, Genetics, genomics and transcriptomics, Eco-epidemiology, and Vector control. Research on sand flies as leishmaniasis vectors has revealed a diverse array of zoonotic and anthroponotic transmission cycles, mostly in subtropical and tropical regions of Africa, Asia and Latin America, but also in Mediterranean Europe. The challenge is to progress beyond descriptive eco-epidemiology, in order to separate vectors of biomedical importance from the sand fly species that are competent vectors but lack the vectorial capacity to cause much human disease. Transmission modelling is required to identify the vectors that are a public health priority, the ones that must be controlled as part of the integrated control of leishmaniasis. Effective modelling of transmission will require the use of entomological indices more precise than those usually reported in the leishmaniasis literature.

Ecological aspects of phlebotomine fauna (Diptera, Psychodidae) of Serra da Cantareira, Greater São Paulo Metropolitan region, state of São Paulo, Brazil.

Human cases of American cutaneous leishmaniasis (ACL) have been recorded in Serra da Cantareira, in the Greater São Paulo Metropolitan Region, where two conservation units are situated, the Parque Estadual da Cantareira and the Parque Estadual Alberto Löfgren. The present study aimed to identify the sandfly fauna and some of its ecological aspects in these two parks and their surrounding area to investigate Leishmania sp. vectors. The captures were undertaken monthly from January to December 2009, from 6:00 p.m. to 6:00 a.m., with automatic light traps installed in forests and peridomicile areas and with modified black/white Shannon traps in the peridomicile. A total of 12 species and 5,436 sandflies were captured: with automatic light traps (141), Shannon traps (5,219) and attempting to bite the researchers while they were conducting the collection in Shannon traps (76). Pintomyia fischeri and Migonemyia migonei were the most abundant species. Pi. fischeri predominated in all three kinds of captures (49%, 88.8% and 65.8%, respectively). Mg. migonei was the second most prevalent in Shannon traps (10.0%) and attempting to bite the researchers (22.4%). Pi. fischeri females were significantly more attracted to black and those of Mg. migonei to white Shannon traps. A positive and significant correlation was observed between the numbers of Pi. fischeri and the mean of minimum relative humidity values on the fifteen days prior to capture, while there was a negative and significant correlation between the relative humidity on the capture day and the two most abundant species. The anthropophilia and high frequencies of Pi. fischeri and Mg. migonei suggest that both species may be transmitting ACL agents in this region.

Description of the female of Pintomyia (Pifanomyia) deorsa (Diptera, Psychodidae, Phlebotominae) / Descripción de la hembra de Pintomyia (Pifanomyia) deorsa (Diptera, Psychodidae, Phlebotominae)

Pintomyia (Pifanomyia) deorsa (Pérez, Ogusuku, Monje & Young, 1991) was described on the basis of a single male; the female is being described here from specimens collected in Ollantaytambo, Cusco, Peru. Diagnoses for the Pintomyia genus, Pifanomyia subgenus, Verrucarum series and both sexes of Pi. deorsa are presented, as well as an identification key to distinguish the females of the Verrucarum series.

Pintomyia (Pifanomyia) deorsa (Pérez, Ogusuku, Monje & Young, 1991) fue descrita en base a un solo espécimen macho; la hembra es descrita aquí a partir de especímenes colectados en Ollantaytambo, Cusco, Perú. El diagnóstico para en género Pintomyia, el subgénero Pifanomyia, la série Verrucarum y ambos sexos de Pi. deorsa son presentados, así como claves para la identificación y separación de las hembras de la serie Verrucarum.

[Occurrence of Nyssomyia intermedia (Lutz & Neiva) (Diptera: Psychodidae) in Paraná State, south of Brazil]. / Ocorrência de Nyssomyia intermedia (Lutz & Neiva) (Diptera: Psychodidae) e fauna associada, no Paraná

The occurrence of Nyssomyia intermedia s.lat. in the state of Paraná, Brazil, has been registered by several authors; however, studies have identified this taxon as belonging, in Paraná, to Nyssomyia neivai (Pinto). During captures with traps in a hen-house and forested areas, from November 2005 to October 2006, in Adrianópolis, Morretes and Pontal do Paraná, situated in the Atlantic forest domain, Paraná state, seven females of Nyssomyia intermedia s.str. (Lutz & Neiva) were collected together with other 14 sand floy species. Thus the occurrence of N. intermedia on the coast and in areas of Atlantic forest in Paraná is confirmed.