Altitude and hillside orientation shapes the population structure of the Leishmania infantum vector Phlebotomus ariasi.

Despite their role in Leishmania transmission, little is known about the organization of sand fly populations in their environment. Here, we used 11 previously described microsatellite markers to investigate the population genetic structure of Phlebotomus ariasi, the main vector of Leishmania infantum in the region of Montpellier (South of France). From May to October 2011, we captured 1,253 Ph. ariasi specimens using sticky traps in 17 sites in the North of Montpellier along a 14-km transect, and recorded the relevant environmental data (e.g., altitude and hillside). Among the selected microsatellite markers, we removed five loci because of stutter artifacts, absence of polymorphism, or non-neutral evolution. Multiple regression analyses showed the influence of altitude and hillside (51% and 15%, respectively), and the absence of influence of geographic distance on the genetic data. The observed significant isolation by elevation suggested a population structure of Ph. ariasi organized in altitudinal ecotypes with substantial rates of migration and positive assortative mating. This organization has implications on sand fly ecology and pathogen transmission. Indeed, this structure might favor the global temporal and spatial stability of sand fly populations and the spread and increase of L. infantum cases in France. Our results highlight the necessity to consider sand fly populations at small scales to study their ecology and their impact on pathogens they transmit.

Insecticide susceptibility of Phlebotomus argentipes & assessment of vector control in two districts of West Bengal, India.

BACKGROUND & OBJECTIVES: Kala-azar or visceral leishmanisis (VL) is known to be endemic in several States of India including West Bengal (WB). Only meager information is available on the vector dynamics of its vector species, Phlebotomus argentipes particularly in relation to control measure from this State. Hence, a pilot study was undertaken to assess the control strategy and its impact on vector in two endemic districts of WB, India. METHODS: Two villages each from the two districts, Maldah and Burdwan, were selected for the study. Seasonal variation of sandflies was observed during pre-monsoon, post-monsoon and winter seasons. Susceptibility test of P. argentipes against DDT and bioassay on DDT sprayed wall and on long lasting insecticide nets (LN) Perma Net [®] 2.0 were conducted as per the WHO standard methods. RESULTS: P. argentipes density was high during March to October. Susceptibility status of P. argentipes ranged from 40 to 61.54 per cent. Bioassay test showed 57.89 per cent mortality against LN PermaNet [®] -2.0. and 50 per cent against DDT on wall within 30 min of exposure. INTERPRETATION & CONCLUSIONS: Despite the integrated vector management approach, the sandfly population was high in the study area. The reason could be development of resistance in P. argentipes against DDT and low effectiveness of LN PermaNet [®] -2.0. The more pragmatic step will be to conduct large studies to monitor the susceptibility level in P. argentipes against DDT.

Estudo populacional de flebotomíneos (Diptera: Psychodidae) no Município de Belo Horizonte e no Parque Estadual do Sumidouro – Minas Gerais - Brasil

O município de Belo Horizonte e sua região metropolitana ilustram o mais sério exemplo de expansão da leishmaniose visceral no sudeste do Brasil. O município e o parque Estadual do Sumidouro localizam-se no bioma cerrado, segundo maior bioma terrestre da América do Sul. Os objetivos deste projeto foram:estudar a variação sazonal da fauna flebotomínica no DS Venda Nova (Belo Horizonte) e no PES; descrever os padrões de riqueza e diversidade fauna flebotomínica nas formações vegetais do PES e no DS Venda Nova; determinar a taxa de infecção natural de fêmeas capturadas; estudar a variação morfológica das populações de Lu. longipalpis coletadas nos diferentes ambientes; avaliar os impactos das ações de controle realizadas no DS Venda Nova nas taxas de ocorrência sazonal e densidade das populações de flebotomíneos. Foram realizadas coletadas mensais no período de agosto de 2011 a agosto de 2013. Os insetos foram identificados de acordo com Galati 2003. As fêmeas coletadas não ingurgitadas foram submetidas à extração de DNA e reações de PCR e de PCR-RFLP para a averiguação da infecção natural por espécies de Leishmania. Para comparação das localidades de estudo foi utilizada análise descritiva e índices ecológicos. Para a comparação morfológica trinta casais (quinze proveniente do DS Venda Nova e quinze do PES) da espécie Lu. longipalpis foram medidos, e foram comparados tantos as médias como as variâncias dos caracteres entre os grupos.

As ações de controle da LV no DS Venda Nova realizadas nos período de janeiro de 2011 a dezembro de 2013 foram analisadas descritivamente e relacionadas à curva sazonal de Lu. longipalpis. A riqueza, a diversidade e a equitabilidade da fauna de flebotomíneos foram marcadamente diferentes entre o DS Venda Nova e o PES. Na área urbana foram coletados 2.247 espécimes pertencentes a cinco gêneros e oito espécies, a curva de acumulação de espécies atingiu a saturação na 19º amostragem. No DS Venda Nova a curva de variação sazonal foi delineada principalmente por Lu. longipalpis e 95,3% dos espécimes pertenciam a esta espécie.

Lu. longipalpis apresentou taxas de infecção natural de 1,01% para Leishmaniainfantum e 1,77% para Leishmaniabraziliensis. No PES foram coletados 4.675 espécimes pertencentes a cinco gêneros e 25 espécies e a curva de acumulação atingiu a saturação na 16º amostragem. No PES não houve uma única espécie que apresentasse padrão tão pronunciado de dominância. Lu. longipalpis correspondeu a 5,35% dos espécimes. Diversas espécies com envolvimento (suspeito ou comprovado) nos ciclos de LTA foram registradas. Uma fêmea do complexo cortellezzi apresentou detecção positiva para Le. braziliensis. A comparação morfométrica e de composição de feromônios indicou grande similaridade entre as duas populações de Lu. longipalpis avaliadas. A análise realizada para correlação das ações de controle com a variação sazonal de Lu. longipalpis indicam a necessidade de estudos pormenorizados para inferências mais robustas. Os resultados evidenciam a dominância da espécie Lu. longipalpis no ambiente urbano, e indicam que as mudanças antrópicas no cerrado podem alterar a composição das populações de flebotomineos e os ciclos de transmissão de patógenos aos humanos.

Isolation of Chandipura virus (Vesiculovirus: Rhabdoviridae) from Sergentomyia species of sandflies from Nagpur, Maharashtra, India.

BACKGROUND & OBJECTIVES: An outbreak of acute encephalitis syndrome was reported from Vidarbha region of Maharashtra s0 tate, India, during July 2012. Anti-IgM antibodies against Chandipura virus (CHPV) were detected in clinical samples. Sandfly collections were done to determine their role in CHPV transmission. METHODS: Twenty nine pools of Sergentomyia spp. comprising 625 specimens were processed for virus isolation in Vero E6 cell line. Diagnostic RT-PCR targeting N-gene was carried out with the sample that showed cytopathic effects (CPE). The PCR product was sequenced, analysed and the sequences were deposited in Genbank database. RESULTS: CPE in Vero E6 cell line infected with three pools was detected at 48 h post infection. However, virus could be isolated only from one pool. RT-PCR studies demonstrated 527 nucleotide product that confirmed the agent as CHPV. Sequence analysis of the new isolate showed difference in 10-12 nucleotides in comparison to earlier isolates. INTERPRETATION & CONCLUSIONS: This is perhaps the first isolation of CHPV from Sergentomyia spp. in India and virus isolation during transmission season suggests their probable role in CHPV transmission. Further studies need to be done to confirm the precise role of Sargentomyia spp. in CHPV transmission.

Transmission of visceral leishmaniasis in a previously non-endemic region of Tunisia: detection of Leishmania DNA in Phlebotomus perniciosus.

Visceral leishmaniasis (VL) has been endemic in northern Tunisia and has occurred sporadically in the center of Tunisia. Recently, there have been several cases from areas known to be free of VL. We report in this work all human and canine cases of VL recorded between 2003 and 2011 and an entomological study of phlebotomine fauna in a previously non-endemic region. Sixty-three cases of VL were diagnosed and identified as L. infantum using several different methods. Eight species of 179 sand flies were caught and identified by both morphological and molecular methods. Two genera were present, Phlebotomus and Sergentomya, with an abundance of the subgenus Phlebotomus (Larrousius) spp., a classic vector of VL in Tunisia. Moreover, Leishmania DNA was detected in seven unfed Phlebotomus pernicousus and L. infantum was identified in three of them. This result confirms the establishment of a transmission cycle of VL in the studied region by the coexistence of infected vectors with infected hosts.

First microscopical and molecular-based characterization of Leishmania major within naturally infected Phlebotomus salehi (Diptera; Psychodidae) in Fars province, southern Iran.

Zoonotoc cutaneous leishmaniasis is endemic in several parts of Iran. Jahrom district is one of the most important endemic foci of leishmaniasis located in Fars province, southern Iran. To identify the vectors of leishmaniasis in this area, a total of 349 sandflies were collected during May to August 2009. They were caught from outdoors in five regions of Jahrom district including villages of Mousavieh, Ghotb-Abad, Heydar-Abad, Fath-Abad and Jahrom County. Eleven species of Phlebotomine (three Phlebotomus spp. and eight Sergentomyia spp.) were detected. To determine the sandflies naturally infected by Leishmania spp., 122 female sandflies were dissected and evaluated microscopically using Giemsa-stained slides. Natural infection of 2 out of 38 (5.26%) P. papatasi and 1 out of 8 (12.5%) P. salehi to Leishmania major was confirmed in the region. Sequencing and nested polymerase chain reaction-based detection of Leishmania were carried out to confirm the microscopic findings. Five (13.16%) P. papatasi and two (25%) P. salehi were positive in nested polymerase chain reaction assay. All positive samples were shown 72-76% similarity with L. major Friedlin. On the basis of our knowledge, this is the first molecular detection of L. major within naturally infected P. salehi in this region in southern Iran.

First report on natural infection of Phlebotomus sergenti with Leishmania promastigotes in the cutaneous leishmaniasis focus in southeastern Tunisia.

During September 2010, 133 female sand flies were caught inside houses of patients with cutaneous leishmaniasis in the focus for this disease in southeastern Tunisia and subsequently dissected. One specimen was positive for Leishmania protozoa. This sand fly species was identified as Phlebotomus sergenti, and the parasite was identified as L. tropica. This is the first report of P. sergenti involvement in transmission of L. tropica in Tunisia.

Incidence of visceral leishmaniasis in the Vaishali district of Bihar, India: spatial patterns and role of inland water bodies.

The role of the distribution of inland water bodies with respect to the transmission of visceral leishmaniasis (VL) and its dominant vector, Phlebotomous argentipes, has been studied at the regional scale in Bihar, eastern India. The Landsat TM sensor multispectral scanning radiometer, with a spatial resolution of 30 m in the visible, reflective-infrared and shortwave-infrared (SWIR) bands, was used to identify water bodies using the normalized differential pond index (NDPI) calculated as follows: (Green - SWIR I)/(Green + SWIR I). Nearest neighbour and grid square statistics were used to delineate spatial patterns and distribution of the sandfly vector and the disease it transmits. The female P. argentipes sandfly was found to be associated with the distance from open water and particularly abundant near non-perennial river banks (68.4%; P <0.001), while its association with rivers was focused further away from the water source (χ(2) = 26.3; P <0.001). The results also reveal that the distribution of VL is clustered around non-perennial riverbanks, while the pattern is slightly random around the perennial river banks. The grid square technique illustrate that the spatial distribution of the disease has a much stronger correlation with lower density of open waters surfaces as well as with sandfly densities (χ(2) = 26.0; P <0.001). The results of our study suggest that inland water presence poses a risk for VL by offering suitable breeding sites for P. argentipes, a fact that should be taken into account when attempting to control disease transmission.

Differences in the salivary effects of wild-caught versus colonized Phlebotomus papatasi (Diptera: Psychodidae) on the development of zoonotic cutaneous leishmaniasis in BALB/c mice.

Preimmunization of mice with salivary gland homogenate (SGH) of long-term colonized (F29) female Phlebotomus papatasi Scopoli (Diptera: Psychodidae) induced protection against Leishmania major Yakimoff & Schokhor (Kinetoplastida: Trypanosomatidae) co-inoculated with the same type of SGH. In contrast, preimmunization of mice with SGH of wild-caught female P. papatasi did not confer protection against L. major co-inoculated with the same type of SGH. Similarly, SGH from recently colonized (F1) female P. papatasi did not protect mice against L. major. These results suggest that when developing a sand fly saliva-based vaccine, the natural vector populations should be considered.

Host preference of Phlebotomus argentipes and Phlebotomus papatasi in different biotopes of West Bengal, India.

Host preference Phlebotomus argentipes and Phlebotomus papatasi in different biotopes was investigated in two highly endemic Kala-azar districts of West Bengal, India for a better understanding of the transmission dynamics. Blood meals of 304 P. argentipes and 206 P. papatasi, collected from different biotopes from two Kala-azar affected districts in West Bengal, were tested against seven different antisera by modified Ouchterlony gel diffusion techniques. It appeared that host preference of P. argentipes varied widely in different biotopes, which is mainly zoophilic (62.80%), preferring to feed on man as the second choice (24.92%); however it is also a "chance feeder" according to biotopes. Multiple blood meals are also prevalent in P. argentipes at a much higher rate than that of P. papatasi. The implication in relation to epidemiological significance has been discussed.

Spatial and temporal patterns of phlebotomine sand flies (Diptera: Psychodidae) in a cutaneous leishmaniasis focus in northern Argentina.

Phlebotomine sand flies (Diptera: Psychodidae) were captured in an area of Argentina endemic for American cutaneous leishmaniasis (ACL). A total of 44,944 flies were collected during a 130-wk interepidemic period from 1990 through 1993. These sand flies included Lutzomyia neivai (Pinto) (97.8%), Lutzomyia migonei (Franca) (1.2%), Lutzomyia cortelezzii (Brèthes) (0.8%), Lutzomyia shannoni (Dyar) (0.1%), and Lutzomyia punctigeniculata (Floch and Abonnenc) (0.1%). Lutzomyia neivai was more abundant in secondary forests and peridomestic environments associated with human cases than in primary forest or xeric thorn scrub areas. Time series analyses of species densities suggested a bimodal or trimodal annual pattern related to rainfall peaks, a 5-wk reproductive cycle, and peridomestic local populations that were located adjacent to secondary forests. In general, sand fly abundance was correlated with the rainfall of the previous year. Lutzomyia neivai spatial distributions were consistent with ACL incidence patterns during the study and in the recent outbreaks in Argentina. However, Lu. migonei also may be involved in peridomestic transmission. Our results suggest that there is a need for improved, long-term surveillance of sand flies and ACL cases, as well as development of effective intervention strategies.

Sandfly survey in Nainital and Almora districts of Uttaranchal with particular reference to Phlebotomus argentipes, vector of kala-azar.

Kala-azar continues to pose a major public health problem in Bihar, West Bengal and parts of eastern Uttar Pradesh in India causing great deal of morbidity and mortality. During 1998, several kala-azar cases from Sub-Himalayan region were treated in Delhi hospitals. And a suspected focus of kala-azar was subsequently reported from this area. Therefore a preliminary sandfly survey was carried out during October, 1999 in 18 randomly selected villages at different altitudes in Nainital & Almora districts of Kumaon region Uttaranchal. The surveys revealed relative preponderance of vector sandfly Ph. argentipes as 77%; mainly confined to cattlesheds and mixed dwellings in the villages at an altitude ranging from 350-960 metres main sea level. The other sandfly species encountered were: Ph. papatasi 6.9%, Ph. major 2.9% and 13.2% Sergentomyia spp. 17.4% Ph. argentipes reacted positive with human antisera and 82.6% with bovine but none reacted with goat, pig dog and bird antisera indicating that Ph. argentipes in the area is mainly zoophilic. Ph. argentipes was found to be highly susceptible to DDT; mortality ranging from 98-100%.

Photosynthesis modulates the plant feeding of Phlebotomus papatasi (Diptera: Psychodidae).

Phlebotomus papatasi (Scopoli), the vector of Leishmania major (Yakimoff & Schokhor), feeds on plants in desert habitats in the Jordan Valley. At the end of the dry summer, the life span of sand flies is short and the amount of sugars in their guts is small. In this season the plants are under the stress of heat and dehydration. This stress arrests the photosynthesis and decreases the amounts of the main end products, sucrose and starch. We presumed that the paucity of sugars in the sand fly plant tissue diet resulted from the arrest of photosynthesis. To test this assumption, we compared the feeding of sand flies on branches of Capparis spinosa (L.) that had been kept for 24 h in darkness and on branches cut after a normal day of photosynthesis. In darkness, the branches had lost more than half of their sugar content. Afterward they were fed upon overnight by 45.2% of female and 14.3% of male sand flies. A higher proportion of 81.0% females and 38.7% males fed on branches from natural conditions and these fed flies were significantly heavier. Laboratory experiments also showed that plant tissue meals of P. papatasi often include starch grains. Such grains were found also in 50% of field-caught males and females. The nutritive potential of plant tissues was demonstrated by the 33-d median survival of P. papatasi series that had been maintained on fresh C. spinosa branches and water.

Leishmaniasis cutánea (Botón de Oriente) / Cutaneous leishmaniasis

Se presenta un paciente de 65 años de edad, con una lesión nodular de aproximadamente 1,5 cm de diámetro, eritematosa y asintomática, ubicada en tercio superior, cara anterior de pierna derecha. Como antecedente de importancia, refiere haber viajado a Italia donde permaneció hasta 3 meses antes de la aparición de la afección. Los estudios efectuados demostraron que se trataba de una leishmaniasis cutánea (Botón de Oriente). Se analizan las formas clínicas, métodos diagnósticos y terapéuticos, y se efectúa una revisión de la literatura de esta inusual enfermedad en nuestro medio

Man-bait catch of Phlebotomus perniciosus (Diptera, Psycodidae) in a human and canine leishmaniasis focus in central Italy.

Man-bait catches of Phlebotomus perniciosus were carried out in a human and canine leishmaniasis focus in Central Italy (Monte Argentario, Grosseto) to evaluate the man-sandfly contact of this vector. P. perniciosus females were collected biting on man outside, during the first hours of the night, when the local people are at risk. The results of these captures are discussed and the man-sandfly contact is considered in relation to the epidemiological aspects of the disease in the area under study.