Entomological Studies in Itaúna, Brazil, an Area With Visceral Leishmaniasis Transmission: Fauna Survey, Natural Leishmania Infection, and Molecular Characterization of the Species Circulating in Phlebotomine Sand Flies (Diptera: Psychodidae).

Among neglected tropical diseases, visceral leishmaniasis (VL) shows great relevance in global terms and is a serious public health concern due to the possibility of severe and lethal forms in humans. In this study, we evaluate entomological factors such as diversity and abundance of phlebotomine sand flies (Diptera:Psychodidae) and the Leishmania species circulating in these species in possible association with VL transmission in the Brazilian town Itaúna. The entomological collections were performed during three consecutive nights, always in the third week of each month, within a period of 12 mo. A total of 1,786 sand fly specimens were collected, from which 20% were collected inside houses. The influence of three local climatic variables (temperature, rainfall, relative humidity) on the population sizes of these insects was evaluated. Temperature was the most influential factor, with a significant positive correlation with the local population size of phlebotomine sand flies collected per month. Lutzomyia longipalpis (Lutz & Neiva, 1912) was the predominant species in the study area. Leishmania DNA was detected in nine out of 133 pools of sand fly females, using nested/PCR, which resulted in a minimal natural infection rate of 2.91%. DNA from Leishmania infantum Nicolle, 1908 (Kinetoplastida: Trypanosomatida), was detected in Evandromyia cortelezzii (Bréthes, 1923), Ev. evandroi (Costa, Lima & Antunes, 1936), Ev. lenti (Mangabeira, 1938), and Ev. termitophila (Martins, Falcão & Silva, 1964), besides Lu. longipalpis. Our study indicates favorable conditions for VL spreading in Itaúna due to the presence of Lu. longipalpis and Le. infantum-infected phlebotomine sand flies.

Detection of Leishmania DNA in phlebotomines captured in Campo Grande, Mato Grosso do Sul, Brazil.

Over the past years, leishmaniases have become a public health issue in the Brazilian state of Mato Grosso do Sul, particularly in Campo Grande, the state capital. The purpose of this study was to detect the presence of Leishmania DNA in the population of phlebotomine sandflies using DNA amplification by PCR. Insect captures were carried out from 4 pm. to 7 am for 4 consecutive days each month from October 2005 to September 2006 in 16 neighborhoods located in 7 urban regions of Campo Grande. Traps were placed indoors and in the vicinity of households. As many as 971 males and 203 females were collected. One hundred and five naturally fed females were identified and grouped as 1- to 4-specimen pools. DNA extraction was carried out using whole insects. Lutzomyia longipalpis predominated, accounting for 99.15% of the phlebotomines captured. Also found was Nyssomyia whitmani, the vector of tegumentary leishmaniasis. Abundance was greatest in the vicinity of households (69.8% of the phlebotomines captured). As revealed by PCR, parasites were present in 1.9% of the Leishmania spp. specimens investigated and confirmed for visceral leishmaniasis.

Are light traps baited with kairomones effective in the capture of Lutzomyia longipalpis and Lutzomyia intermedia? An evaluation of synthetic human odor as an attractant for phlebotomine sand flies (Diptera: Psychodidae: Phlebotominae).

Phlebotomine sand flies are often captured with human bait and/or light traps, either with or without an animal bait. More recently, synthetic attractants have been used as bait in traps to improve the capture of phlebotomine sand flies as well as other insects of medical and veterinary importance. The aim of the present study was to evaluate the effects of the kairomone 1-octen-3-ol (octenol) and the synthetic human odor BG-Mesh Lure (BGML--lactic acid, caproic acid and ammonia) baited in modified CDC light traps on the capture of phlebotomine sand flies. The experiments followed the 5x5 Latin square design. Among the species caught, Lutzomyia intermedia apparently presented a dose-dependent response to octenol. The response obtained with the BGML, alone or in combination with octenol (5 mg/h), indicated some degree of attractiveness of these baits to different phlebotomine sand fly species. Octenol seems to be more attractive to L. intermedia than to Lutzomyia longipalpis, while the BGML presented a higher success in capturing L. longipalpis. When the components of the BGML were used separately, there was no increase in catching the female of L. intermedia. Apparently, there was no synergistic effect between the octenol and the BGML. In conclusion, the octenol and the BGML were demonstrated to be possible baits to attract some phlebotomine sand fly species.

Ecoepidemiological aspects of visceral leishmaniasis in an endemic area in the Steel Valley in Brazil: An ecological approach with spatial analysis.

Leishmaniases are a group of infectious diseases transmitted by phlebotomine sand flies, and their distribution depends on the presence of vectors, parasites, reservoirs and susceptible hosts in the same environment. In the last decades, visceral leishmaniasis (VL) has become urbanized and reached economically important cities in countries within the transmission zone. Our study was conducted in one of those cities-Ipatinga-in the state of Minas Gerais, Brazil, where the first autochthonous case of VL dates back to 2011. Since no data regarding the epidemiological triad of VL (etiological agent/vector/domestic reservoir) were available for this city, we characterized the local entomological fauna, identified the presence of specific Leishmania DNA in the captured phlebotomine sand flies, and assessed the incidence of canine and human VL. For the entomological survey, we set twenty light traps in ten districts of the city with reports of human and canine VL. The insect captures were performed monthly, during one year, starting in March 2015. A total of 1501 specimens of phlebotomine sand flies belonging to 16 distinct species were captured, with predominance (61.9%) of Lutzomyia longipalpis. Leishmania infantum DNA was detected in L. longipalpis and in Evandromyia cortelezzii test samples. A total of 9,136 dogs were examined, 1,355 of which (14.8%) were serologically positive for VL. The cases were georeferenced and the data were plotted in thematic maps, along with human cases of VL registered by the local Department of Health, during the study period. Our results confirm that the VL transmission cycle is active in Ipatinga, with the presence of vectors carrying Leishmania DNA, canine and human cases of the disease. Spatial analysis allowed for the observation of a positive relationship between canine and human cases of VL and the identification of areas with high priority for control actions in the city. The mapping of high-risk areas, together with an epidemiological study in urban areas, is fundamental to improve the efficacy of the Program for Surveillance and Control of VL (PSCVL) in Brazil.

Seroprevalence and molecular characterization of Leishmania in dogs from an endemic area of zoonotic visceral leishmaniasis in Brazil.

Visceral leishmaniasis (VL) can cause large-scale and tenacious epidemics with high fatality rates. Current seroprevalence and circulating Leishmania species were evaluated in dogs domiciled in the municipality of Sabará, a small historic and touristic city in the Brazilian state of Minas Gerais. A total of 3926 dogs domiciled in seven different districts of Sabará were serologically tested for canine visceral leishmaniasis (CVL) by indirect enzyme-linked immunosorbent (ELISA) and immunofluorescence (IFA) assays, in a two-years census survey (2011-2012). The average positivity rate of canine infection was 3.4%. Three additional diagnostic tests - imprint/smear direct parasitological, molecular (LnPCR) and myeloculture - were performed in a random sample of fifty seropositive dogs composed of symptomatic (39) and asymptomatic (eleven) animals. LnPCR showed 100% of positivity for Leishmania DNA in, at least, one among four tissue samples tested (mesenteric lymph node, skin, spleen and bone marrow), independently of the clinical canine group. Higher and statistically equivalent positivity rates (98% and 96%) for Leishmania DNA were found in canine lymph node and spleen. Asymptomatic dogs showed expressive positivity rates in all three additional diagnostic techniques. Leishmania infantum was confirmed as the etiological agent of CVL in Sabará.

Importance of Lutzomyia longipalpis in the dynamics of transmission of canine visceral leishmaniasis in the endemic area of Porteirinha Municipality, Minas Gerais, Brazil.

A study of Lutzomyia longipalpis (Lutz and Neiva, 1912) (Diptera: Psychodidae), the primary vector of American visceral leishmaniasis (AVL), and the canine form of the disease, was carried out in Porteirinha. The city is situated in the northern part of the Brazilian State of Minas Gerais and is an endemic area of AVL. Systematic phlebotomine captures were performed in seven districts with previously reported cases of canine visceral leishmaniasis, during 2 years (January 2000--December 2001). A total of 2328 specimens of L. longipalpis were captured. The association between the local climate variables and the population density of L. longipalpis was evaluated and rainfall was determined to be a major factor, with increased populations during the rainy season (October--March). At the same time period, blood samples from every dog domiciled in the same seven districts, in total 14,077 animals, were analyzed for infection by viscerotropic Leishmania using indirect immunofluorescence assay (IFA). Accumulated incidence rates of canine VL per district varied from 3.40 to 14.34 for the 2-year period. A positive correlation between the population density of L. longipalpis and the canine cases of visceral leishmaniasis in Porteirinha was observed.

[HP: an improved model of suction light trap for the capture of small insects]. / HP: um modelo aprimorado de armadilha luminosa de sucção para a captura de pequenos insetos.

This paper describes a new model of light trap, named HP, based on the widely used CDC trap. Its advantages include high efficiency, easy handling and transport. Moreover, it is almost totally made of national materials, which reduces considerably the final cost.

Assessment of PCR in the detection of Leishmania spp in experimentally infected individual phlebotomine sandflies (Diptera: Psychodidae: Phlebotominae).

DNA amplification by the polymerase chain reaction (PCR) was applied in the investigation of the presence of Leishmania (Kinetoplastida: Trypanosomatidae) parasites in single phlebotomine sandflies. Three phlebotomine/parasite pairs were used: Lutzomyia longipalpis/Leishmania chagasi, Lutzomyia migonei/Leishmania amazonensis and Lutzomyia migonei/Leishmania braziliensis, all of them incriminated in the transmission of visceral or cutaneous leishmaniasis. DNA extraction was performed with whole insects, with no need of previous digestive tract dissection or pooling specimens. The presence of either mouse blood in the digestive tract of the sandflies or the digestive tract itself did not interfere in the PCR. Infection by as few as 10 Leishmania sp. per individual were sufficient for DNA amplification with genus-specific primers. Using primers for L. braziliensis and L. mexicana complexes, respectively, it was possible to discriminate between L. braziliensis and L. amazonensis in experimentally infected vectors (L. migonei).

Association between the prevalence of infestation by Rhipicephalus sanguineus and Ctenocephalides felis felis and the presence of anti-Leishmania antibodies: A case-control study in dogs from a Brazilian endemic area.

The association between the prevalence of infestation by Rhipicephalus sanguineus and Ctenocephalides felis felis and the presence of anti-Leishmania antibodies has been evaluated in dogs located in a city of Brazil endemic for canine visceral leishmaniasis. Blood samples from 5556 domestic dogs domiciliated in the urban area of Belo Horizonte (Minas Gerais state) were submitted to enzyme linked immunosorbent (ELISA) and indirect immunofluorescent antibody (IFAT) assays, and 432 (7.8%) animals tested positive. Seropositive (n=200) and seronegative (n=200) dogs were randomly selected and examined for the presence of ticks and fleas, the results of which were expressed qualitatively as infested or non-infested, irrespective of the intensity of infestation. The prevalence of infestation by R. sanguineus was significantly greater (ρ=0.04) among seropositive dogs (38.5%) compared with their seronegative counterparts (29.0%). Similarly, the prevalence of infestation by C. felis felis was significantly greater (ρ<0.01) within the seropositive group (36.5%) than within the seronegative group (15.0%). Moreover, the probability of seropositivity for Leishmania was 53% higher in tick-infested dogs and 300% higher in flea-infested dogs in comparison with non-infested animals. Our data provide evidence of the vectorial capacity of these ectoparasites in transmitting Leishmania to the canine population, although further studies are needed to confirm or reject this hypothesis.

[Phlebotomine sandflies in Parque Nacional Cavernas do Peruaçu, Minas Gerais state, Brazil]. / Flebotomíneos do Parque Nacional Cavernas do Peruaçu, MG.

We surveyed the phlebotomine fauna in the Parque Nacional Cavernas do Peruaçu, Minas Gerais, Brazil, aiming to associate the presence of vector with the risk of leishmaniasis transmission. Field captures were performed with CDC light traps between February and March 2003. A total of 436 sand flies were captured, belonging to 14 species. The predominant species (28.7%) was Lutzomyia ischnacantha Martins, Souza e Falcão, followed by L. renei (27,06%) and L. cavernicola (13,07%). The finding of L. intermedia, a species that is incriminated as vector of cutaneous leishmaniasis, has to be taken into account.

[Four new records of the phlebotominic fauna of the Genus Lutzomyia França (Diptera: Psychodidae, Phlebotominae) of the rural district of Brejo do Mutambal, Varzelândia, State of Minas Gerais, Brazil]. / Quatro novos registros da fauna flebotomínica do gênero Lutzomyia França (Diptera: Psychodidae, Phlebotominae) do distrito rural de Brejo do Mutambal, Varzelândia, MG.

During April and May 2006, experiments were carried out in Brejo do Mutambal, Varzelândia Town, Minas Gerais State, to evaluate the attractiveness of phlebotomine sandflies to CDC light traps, baited with kairomones. Of the 19 species of Lutzomyia already registered for the region, L. lutziana (Costa Lima), L. longipennis (Barreto), L. goiana (Martins, Falcão & Silva) and L. brasiliensis (Costa Lima) were recorded for the first time, thus increasing the diversity of phlebotomine sandflies fauna in this area to 23 species. The new registered species and distribution are shown and discussed herein.

Are light traps baited with kairomones effective in the capture of Lutzomyia longipalpis and Lutzomyia intermedia? An evaluation of synthetic human odor as an attractant for phlebotomine sand flies (Diptera: Psychodidae: Phlebotominae)

Phlebotomine sand flies are often captured with human bait and/or light traps, either with or without an animal bait. More recently, synthetic attractants have been used as bait in traps to improve the capture of phlebotomine sand flies as well as other insects of medical and veterinary importance. The aim of the present study was to evaluate the effects of the kairomone 1-octen-3-ol (octenol) and the synthetic human odor BG-Mesh LureTM (BGML - lactic acid, caproic acid and ammonia) baited in modified CDC light traps on the capture of phlebotomine sand flies. The experiments followed the 5x5 Latin square design. Among the species caught, Lutzomyia intermedia apparently presented a dose-dependent response to octenol. The response obtained with the BGML, alone or in combination with octenol (5 mg/h), indicated some degree of attractiveness of these baits to different phlebotomine sand fly species. Octenol seems to be more attractive to L. intermedia than to Lutzomyia longipalpis, while the BGML presented a higher success in capturing L. longipalpis. When the components of the BGML were used separately, there was no increase in catching the female of L. intermedia. Apparently, there was no synergistic effect between the octenol and the BGML. In conclusion, the octenol and the BGML were demonstrated to be possible baits to attract some phlebotomine sand fly species.

Flebotomíneos do Parque Nacional Cavernas do Peruaçu, MG / Phlebotomine sandflies in Parque Nacional Cavernas do Peruaçu, Minas Gerais state, Brazil

O presente trabalho teve como objetivo conhecer a fauna flebotomínica na Gruta do Caboclo, localizada no Parque Nacional Cavernas do Peruaçu, MG, e associá-la ao risco de transmissão das leishmanioses. As capturas foram realizadas utilizando-se armadilhas luminosas tipo CDC, nos meses de fevereiro e março de 2003. O total de 436 flebotomíneos foi capturado, pertencendo a 14 espécies. A espécie predominante foi Lutzomyia ischnacantha Martins, Souza & Falcão, com 28,7 por cento, seguida por L. renei (27,06 por cento) e L. cavernicola (13,07 por cento). O encontro de L. intermedia, espécie incriminada como vetora de leishmaniose cutânea, deve ser levado em consideração.

We surveyed the phlebotomine fauna in the Parque Nacional Cavernas Do Peruaçu, Minas Gerais, Brazil, aiming to associate the presence of vector with the risk of leishmaniasis transmission. Field captures were performed with CDC light traps between February and March 2003. A total of 436 sand flies were captured, belonging to 14 species. The predominant species (28.7 percent) was Lutzomyia ischnacantha Martins, Souza e Falcão, followed by L. renei (27,06 percent) and L. cavernicola (13,07 percent). The finding of L. intermedia, a species that is incriminated as vector of the cutaneous leishmaniasis, has to be taken into account.

Quatro novos registros da fauna flebotomínica do gênero Lutzomyia França (Diptera: Psychodidae, Phlebotominae) do distrito rural de Brejo do Mutambal, Varzelândia, MG / Four new records of the phlebotominic fauna of the Genus Lutzomyia França (Diptera: Psychodidae, Phlebotominae) of the rural district of Brejo do Mutambal, Varzelândia, State of Minas Gerais, Brazil

Durante os meses de abril e maio de 2006 foram realizados experimentos em Brejo do Mutambal, município de Varzelândia, MG para avaliar a atratividade de flebotomíneos a cairomônios iscados em armadilhas luminosas do tipo CDC. Das 19 espécies do gênero Lutzomyia já registradas para a região, L. lutziana (Costa Lima), L. longipennis (Barreto), L. goiana (Martins, Falcão & Silva) e L. brasiliensis (Costa Lima) foram registradas pela primeira vez, aumentando a diversidade da fauna de flebotomíneos do município para 23 espécies. As espécies relatadas como primeiros registros e sua distribuição são citadas e discutidas.

During April and May 2006, experiments were carried out in Brejo do Mutambal, Varzelândia Town, Minas Gerais State, to evaluate the attractiveness of phlebotomine sandflies to CDC light traps, baited with kairomones. Of the 19 species of Lutzomyia already registered for the region, L. lutziana (Costa Lima), L. longipennis (Barreto), L. goiana (Martins, Falcão & Silva) and L. brasiliensis (Costa Lima) were recorded for the first time, thus increasing the diversity of phlebotomine sandflies fauna in this area to 23 species. The new registered species and distribution are shown and discussed herein.

HP: um modelo aprimorado de armadilha luminosa de sucção para a captura de pequenos insetos / HP: an improved model of sucction light trap for the capture of small insects

Descreve-se um novo tipo de armadilha, resultado do aprimoramento de um modelo muito utilizado, conhecido como CDC. A nova armadilha, denominada HP, apresenta como vantagens alta eficiência, facilidade no manuseio e transporte, além de ser confeccionada quase que exclusivamente com material nacional, o que reduz consideravelmente seu custo final.

Random amplified polymorphic DNA (RAPD) analysis of Lutzomyia longipalpis laboratory populations

O flebotomineo Lutzomyia longipalpis tem sido incriminado como vetor da leishmaniose visceral americana, causada pelo protozoario Leishmania chagasi. Entretanto, tem-se acumulado evidências que sugerem a existência de um complexo e näo apenas uma espécie de L. longipalpis na natureza. Nosso trabalho teve como objetivo comparar, ao nível molecular, quatro populaçöes de L. longipalpis de referência, utilizando especimens criados em laboratório, provenientes de regiöes geograficamente distintas, através de RAPD-PCR (reaçäo de polimerase em cadeia com amplificaçäo por iniciadores ao acaso). Para isso, o DNA genomico de grupos de flebotomineos foi amplificado com iniciadores decamericos unicos com sequencia de nucleotideos arbitraria, na tentativa de se detectar sitios polimorficos...

Assessment of PCR in the detection of Leishmania spp in experimentally infected individual phlebotomine sandflies (Diptera: Psychodidae: Phlebotominae)

DNA amplification by the polymerase chain reaction (PCR) was applied in the investigation of the presence of Leishmania (Kinetoplastida: Trypanosomatidae) parasites in single phlebotomine sandflies. Three phlebotomine/parasite pairs were used: Lutzomyia longipalpis/Leishmania chagasi, Lutzomyia migonei/Leishmania amazonensis and Lutzomyia migonei/Leishmania braziliensis, all of them incriminated in the transmission of visceral or cutaneous leishmaniasis. DNA extraction was performed with whole insects, with no need of previous digestive tract dissection or pooling specimens. The presence of either mouse blood in the digestive tract of the sandflies or the digestive tract itself did not interfere in the PCR. Infection by as few as 10 Leishmania sp. per individual were sufficient for DNA amplification with genus-specific primers. Using primers for L. braziliensis and L. mexicana complexes, respectively, it was possible to discriminate between L. braziliensis and L. amazonensis in experimentally infected vectors (L. migonei)