Vertical stratification and development aspects of phlebotomine sand flies (Diptera: Psychodidae) in an area of Atlantic Forest tree species in a metropolitan region in northeastern Brazil.

In the state of Rio Grande do Norte in northeast Brazil, cases of visceral leishmaniasis (VL) occur mainly in the periurban areas of the city of Natal. Lutzomyia longipalpis Lutz & Neiva 1912 (Diptera: Psychodidae), a vector of Leishmania chagasi (Protozoa: Trypanosomatidae) to humans, is found throughout the state. Flora and fauna influence the distribution of sand fly species, whose horizontal or vertical stratification can be used as a parameter for identifying potential vectors, considering the presence of vertebrate hosts in the area. The purpose of this study was to obtain information about the vertical stratification of phlebotomine sand flies in an endemic area of leishmaniasis in Rio Grande do Norte, and associate it with the presence of other animals in the peridomiciliary environment as well as to analyze, under laboratory conditions, aspects of L. longipalpis reproduction in wild females. The sand flies were captured with light traps hung at different heights in species of Atlantic Forest trees and in a peridomiciliary environment in animal shelters. The traps were placed between 17:30 and 6:00 of the following day, in a peridomiciliary and extradomiciliary area of a forest fragment in both dry and rainy months. In the extradomiciliary environment, the traps were installed at 1, 3 and 5 m above the ground. The biological cycle of L. longipalpis was followed from the eggs of 200 wild females. Specimens of L. lenti, L. walkeri, and L. migonei were captured. The comparison and statistical analysis showed that L. longipalpis is more abundant at a height of 3 m and L. evandroi at 1 m. In the animal shelters (chickens, horses, and armadillos), we captured mainly specimens of L. longipalpis and L. evandroi. The duration of the biological cycle of L. longipalpis was approximately 38 days at a temperature of 28 degrees C.     

Deltamethrin-impregnated dog collars have a potent anti-feeding and insecticidal effect on Lutzomyia longipalpis and Lutzomyia migonei

Deltamethrin-impregnated PVC dog collars were tested to assess if they were effective in protecting dogs from sand fly bites of Lutzomyia longipalpis and Lu. migonei. A protective effect against Old World species Phlebotomus perniciosus was demonstrated before. Four dogs wearing deltamethrin collars and three dogs wearing untreated collars (not impregnated with deltamethrin) were kept in separate kennels for over eight months in a village on the outskirts of Fortaleza in Ceará, Brazil. Periodically, a dog from each group was sedated, placed in a net cage for 2 h in which 150 female sand flies had been released 10-15 min before. Lu. longipalpis were used 4, 8, 12, 16, 22, 27, and 35 weeks after the attachment of the collars. Lu. migonei were used 3, 7, 11, 15, 22, 26, and 36 weeks after attachment. During 35 weeks, only 4.1% (81 of 2,022) Lu. longipalpis recovered from the nets with the deltamethrin collared dogs were engorged, an anti-feeding effect of 96%. Mortality initially was over 90% and at 35 weeks was 35% with half of the sand flies dying in the first 2 h. In contrast, 83% of the 2,094 Lu. longipalpis recovered from the nets containing the untreated collared dogs were engorged and the mortality ranged from zero to 18.8% on one occasion with 1.1% dying in the first 2 h. Similar findings were found with Lu. migonei: of 2,034 sand flies recovered over this period, only 70 were engorged, an anti-feeding effect of 96.5%, and mortality ranged from 91% initially to 46% at 36 weeks. In contrast, engorgement of controls ranged from 91 to71% and a mortality ranged from 3.5 to 29.8%. These studies show that deltamethrin impregnated collars can protect dogs against Brazilian sand flies for up to eight months. Thus, they should be useful in a program to control human and canine visceral leishmaniasis.     

Gene silencing in phlebotomine sand flies: Xanthine dehydrogenase knock down by dsRNA microinjections

Lutzomyia longipalpis are vectors of medically important visceral leishmaniasis in South America. Blood-fed adult females digest large amounts of protein, and xanthine dehydrogenase is thought to be a key enzyme involved in protein catabolism through the production of urate. Large amounts of heme are also released during digestion with potentially damaging consequences, as heme can generate oxygen radicals that damage lipids, proteins and nucleic acids. However, urate is an antioxidant that may prevent such oxidative damage produced by heme. We investigated xanthine dehydrogenase by developing the RNAi technique for sand flies and used this technique to knock down the Lu. longipalpis xanthine dehydrogenase gene to evaluate its role in survival of adult females after blood feeding. The gene sequence of Lu. longipalpis xanthine dehydrogenase is described together with expression in different life cycle stages and RNAi knock down. Semi-quantitative RT-PCR of xanthine dehydrogenase expression showed a significant increase in expression after bloodmeal ingestion. Microinjection of dsRNA via the thorax of 1-day-old adult female sand flies resulted in approximately 40% reduction of xanthine dehydrogenase gene expression in comparison to flies injected with a control dsRNA. A significant reduction of urate in the whole body and excretions of Lu. longipalpis was observed after dsRNA xanthine dehydrogenase microinjection and feeding 96h later on rabbit blood. Sand flies injected with XDH dsRNA also exhibit significantly reduced life span in comparison with the mock-injected group when fed on sucrose or when rabbit blood fed, showing that urate could be indeed an important free radical scavenger in Lu. longipalpis. The demonstration of xanthine dehydrogenase knock down by dsRNA microinjection, low mortality of microinjected insects and the successful bloodfeeding of injected insects demonstrated the utility of RNAi as a tool for functional analysis of genes in phlebotomine sand flies.     

Phlebotomine sand flies (Diptera: Psychodidae) in the municipality of Várzea Grande: an area of transmission of visceral leishmaniasis in the state of Mato Grosso, Brazil

Visceral leishmaniasis (VL) has been naturally transmitted in periurban areas due to the emergence and reemergence of its vectors in such areas. Aimed to further knowledge on ecological aspects affecting the occurrence of phlebotomine sand flies in VL transmission areas in the municipality of Várzea Grande, state of Mato Grosso (MT), Brazil, sand fly captures were carried out. Monthly collections of sand flies were undertaken with CDC light-traps, which were left in both intradomiciliary and peridomiciliary areas of ten residences during four consecutive days between January 2004 and June 2006. Twenty-two species of genus Lutzomyia and one of Brumptomyia were captured. The most abundant species was Lutzomyia longipalpis (65.23%), followed by L. evandroi (16.26%), L. lenti (7.69%), L. whitmani (4.92%), L. sallesi (2.34%) and L. termitophila (1.32%). The highest density of the main VL vector, L. longipalpis, was found in peridomiciliary areas, mostly males. No significant correlation was found between environment (temperature, air relative humidity and rain fall) and phlebotomine density; although a slight increase in sand fly density has been observed in the period following rainfalls, particularly L. longipalpis. No correlation was observed between distribution and density of L. longipalpis, prevalence of human VL cases and the presence of serologically positive dogs. The presence of infected dogs, increased vector density, susceptibility rate and interruption of epidemiological surveillance may raise the risk of VL transmission to man in Várzea Grande.     

Sand flies (Diptera: Psychodidae) in a Cerrado area of the Maranhão state, Brazil

The present paper aims to increase the knowledge on the sand fly fauna in the cerrado areas of Maranh?o state in urban, rural and forest environments. The research was carried out from October 2007 to September 2008, between 18:00h and 06:00h, in the municipality of Chapadinha, northeast Maranh?o. For insect sampling, CDC light traps were set up in peridomicile and domicile areas of urban and rural zones as well as in Cerrado and Gallery forests. The total of 1,401 specimens belonging to 17 species were sampled, all within the genus Lutzomyia. Lutzomyia longipalpis (Lutz & Neiva) (52.5%), Lu. evandroi (Costa Lima & Antunes) (18.3%), Lu. whitmani (Antunes & Coutinho) (12.1%), Lu. lenti (Mangabeira) (4.7%) and Lu. termitophila (Martins, Falc?o & Silva) (4.0%) were the most frequently collected. From an epidemiological viewpoint, five from all of the collected species are vectors of leishmaniasis: Lu. longipalpis, Lu. whitmani, Lu. flaviscutellata (Mangabeira), Lu. gomezi (Nitzulescu) and Lu. chagasi (Costa Lima). Lutzomyia chagasi was registered for the first time in Maranh?o state and Lu. saulensis (Floch & Abonnenc), Lu. monstruosa (Floch & Abonnenc) and Lu. gomezi were found for the first time in the eastern part of the state, since they had been reported only in the Amazonian region of Maranh?o. Regarding to the studied environments, the urban chicken house had the highest number of specimens collected (801), while the Gallery Forest was the most diverse (15 species). This study demonstrates that the northeast cerrado exhibits a mixed sand fly fauna characterized by an extremely important species vectors assortment involved in the epidemiological cycle of leishmaniasis in Maranh?o state.     

Susceptibility of laboratory-reared female Lutzomyia longipalpis (Lutz & Neiva, 1912) to infection by different species and strains of Leishmania Ross, 1903

A study was undertaken to compare the susceptibility of laboratory-reared female Lutzomyia longipalpis to infection by different species or strains of New World Leishmania. The sand flies proved to be highly susceptible to infection by a strain of Le. guyanensis, with flagellates developing in all (18/18) of the specimens examined. A lower infection rate of 37% (10/27) was recorded in flies exposed to infection by a strain of Le. amazonensis. Flagellates developed in 13% (6/46) of the sand flies that blood fed on dogs in the early stage of experimental infection with an old laboratory strain of Le. chagasi. In contrast, promastigotes did not develop in sand flies that blood fed on dogs with naturally acquired Le. chagasi. The naturally infected dogs were in an advanced stage of disease. Flagellates developed in 9% (3/32) of the sand flies that blood fed on lesions of hamsters infected with a strain of Le. braziliensis and in 9% (3/34) of those that fed on hamsters with lesions due to a parasite of the mexicana complex (strain MHOM/BR/73/BH121). Sand flies did not develop flagellate infections after blood feeding on hamsters bearing lesions induced by strain MHOM/BR/71/BR49. Factors influencing the susceptibility of Lu. longipalpis to infection by New World species of Leishmania are discussed.     

Detection of Leishmania infantum in naturally infected Lutzomyia longipalpis (Diptera: Psychodidae: Phlebotominae) and Canis familiaris in Misiones, Argentina: the first report of a PCR-RFLP and sequencing-based confirmation assay

In this study, a genotypification of Leishmania was performed using polimerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and sequencing techniques to identify species of Leishmania parasites in phlebotomine sand flies and dogs naturally infected. Between January-February of 2009, CDC light traps were used to collect insect samples from 13 capture sites in the municipality of Posadas, which is located in the province of Misiones of Argentina. Sand flies identified as Lutzomyia longipalpis were grouped into 28 separate pools for molecular biological analysis. Canine samples were taken from lymph node aspirates of two symptomatic stray animals that had been positively diagnosed with canine visceral leishmaniasis. One vector pool of 10 sand flies (1 out of the 28 pools tested) and both of the canine samples tested positively for Leishmania infantum by PCR and RFLP analysis. PCR products were confirmed by sequencing and showed a maximum identity with L. infantum. Given that infection was detected in one out of the 28 pools and that at least one infected insect was infected, it was possible to infer an infection rate at least of 0.47% for Lu. longipalpis among the analyzed samples. These results contribute to incriminate Lu. longipalpis as the vector of L. infantum in the municipality of Posadas, where cases of the disease in humans and dogs have been reported since 2005.     

Sand fly vectors (Diptera, Psychodidae) of American visceral leishmaniasis areas in the Atlantic Forest, State of Espírito Santo, southeastern Brazil

The aim of this study was to evaluate the sand fly fauna of American visceral leishmaniasis (AVL) endemic areas within the Central Atlantic Forest Biodiversity Corridor, State of Espírito Santo, southeastern Brazil. The sand fly captures were performed between January, 1989 and December, 2003 in localities where autochthonous cases of AVL were recorded, as well as in their boundary areas. Sand flies were collected from surrounding houses and domestic animal shelters using two to five CDC automatic light traps, and manual captures were also performed using mouth aspirators in one illuminated Shannon trap during the first four hours of the night. We used cladistic analysis to determine the geographic relationships among the collected sand fly species as well as the index species for the occurrence of other sand flies. A total of 62,469 sand flies belonging to 17 species and eight genera was collected in 164 localities from nine municipalities with AVL records. The richness (S=17) and diversity (H=0.971) of sand flies were lower than in conservation areas and similar to modified environments in the Atlantic Forest of Espírito Santo. Lutzomyia longipalpis was identified in 79 localities. The cladistic analysis identified Evandromyia lenti as the index species for Lutzomyia longipalpis. The latter seems to be the main vector of AVL in the Central Atlantic Forest Biodiversity Corridor due to its high abundance and distribution matching the disease occurrence. Therefore, Evandromyia lenti may be used as an index species for the occurrence of Lutzomyia longipalpis.     

External morphology of sensory structures of fourth instar larvae of neotropical species of phlebotomine sand flies (Diptera: Psychodidae) under scanning electron microscopy.

In the present study, some morphological structures of antennae, maxillary palps and caudal setae of fourth instar larvae of laboratory-reared phlebotomine sand flies (Lutzomyia longipalpis, L. migonei, L. evandroi, L. lenti, L. sericea, L. whitmani and L. intermedia) of the State of Ceará, Brazil, were examined under scanning electron microscopy. The antennal structures exhibited considerable variation in the morphology and position. A prominent digitiform distal segment has been observed only on the antenna of species of the subgenus Nyssomyia. The taxonomic relevance of this and other antennal structure is discussed. The papiliform structures found in the maxillae and the porous structures of the caudal setae of all species examined may have chemosensory function. Further studies with transmission electron microscopy are needed to better understand the physiological function of these external structures.     

Reactive oxygen species scavenging by catalase is important for female Lutzomyia longipalpis fecundity and mortality

The phlebotomine sand fly Lutzomyia longipalpis is the most important vector of American visceral leishmaniasis (AVL), the disseminated and most serious form of the disease in Central and South America. In the natural environment, most female L. longipalpis are thought to survive for less than 10 days and will feed on blood only once or twice during their lifetime. Successful transmission of parasites occurs when a Leishmania-infected female sand fly feeds on a new host. Knowledge of factors affecting sand fly longevity that lead to a reduction in lifespan could result in a decrease in parasite transmission. Catalase has been found to play a major role in survival and fecundity in many insect species. It is a strong antioxidant enzyme that breaks down toxic reactive oxygen species (ROS). Ovarian catalase was found to accumulate in the developing sand fly oocyte from 12 to 48 hours after blood feeding. Catalase expression in ovaries as well as oocyte numbers was found to decrease with age. This reduction was not found in flies when fed on the antioxidant ascorbic acid in the sugar meal, a condition that increased mortality and activation of the prophenoloxidase cascade. RNA interference was used to silence catalase gene expression in female Lu. longipalpis. Depletion of catalase led to a significant increase of mortality and a reduction in the number of developing oocytes produced after blood feeding. These results demonstrate the central role that catalase and ROS play in the longevity and fecundity of phlebotomine sand flies.     

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.     

The molecular detection of different Leishmania species within sand flies from a cutaneous and visceral leishmaniasis sympatric area in Southeastern Brazil

Over the last 20 years, there has been an increase in the number of leishmaniasis cases in Brazil. Belo Horizonte (BH) is one of the most highly populated Brazilian cities that is affected by visceral leishmaniasis (VL). The health services in BH are coordinated by a central nucleus that is subdivided into nine sanitary districts. Historically, the highest level of human VL cases was found in the northeast sanitary district (NSD). The objective of our study was to detect Leishmania infection in the phlebotomine sand flies collected in the NSD by dissection and molecular approaches. Following the occurrence of human VL cases in 2005, entomological captures were performed from July 2006-June 2007. Out of the 245 sand flies dissected, only three Lutzomyia longipalpis spp contained flagellates. The female sand flies were grouped into 120 pools according to date, collection site and species, with approximately 10 individual sand flies in each pool. Subsquently, the DNA was extracted and Leishmania spp and other parasites were detected and identified by polymerase chain reaction (PCR) and PCR-restriction fragment length polymorfism. Leishmania infantum was present in at least 19% of the Lu. longipalpis collected, in 3.8% of the Nyssomiya whitmani collected, in 33.3% of the Evandromiya termitophila collected and in 14.3% of the Nyssomiya intermedia collected. When the females of the cortelezzii complex were compared with each other, 3.2% of the females were infected with Leishmania braziliensis, whereas 3.2% of the females were infected with trypanosomatids.     

Fixed Differences in the paralytic Gene Define Two Lineages within the Lutzomyia longipalpis Complex Producing Different Types of Courtship Songs

The sand fly Lutzomyia longipalpis (Diptera: Psychodidae: Phlebotominae), the most important vector of American visceral leishmaniasis, is widely distributed in Latin America. There is currently a consensus that it represents a species complex, however, the number and distribution of the different siblings is still uncertain. Previous analyses have indicated that Brazilian populations of this vector can be divided into two main groups according to the type of courtship song (Burst vs. Pulse) males produce during copulation. Nevertheless, no diagnostic differences have been observed between these two groups with most molecular markers used to date. We analyzed the molecular divergence in a fragment of the paralytic (para) gene, a locus involved in the control of courtship songs in Drosophila, among a number of Lu. longipalpis populations from Brazil producing Burst and Pulse-type songs. Our results revealed a very high level of divergence and fixed differences between populations producing the two types of songs. We also compared Lu. longipalpis with a very closely related species, Lutzomyia cruzi, which produces Burst-type songs. The results indicated a higher number of fixed differences between Lu. cruzi and the Pulse-type populations of Lu. longipalpis than with those producing Burst-type songs. The data confirmed our previous assumptions that the presence of different sibling species of the Lu. longipalpis complex in Brazil can be divided into two main groups, one representing a single species and a second more heterogeneous group that probably represents a number of incipient species. We hypothesize that para might be one of the genes directly involved in the control of the courtship song differences between these two groups or that it is linked to other loci associated with reproductive isolation of the Brazilian species.     

A lipophosphoglycan-independent development of Leishmania in permissive sand flies

Leishmaniases are serious parasitic diseases the etiological organisms of which are transmitted by insect vectors, phlebotominae sand flies. Two sand fly species, Phlebotomus papatasi and P. sergenti, display remarkable specificity for Leishmania parasites they transmit in nature, but many others are broadly permissive to the development of different Leishmania species. Previous studies have suggested that in 'specific' vectors the successful parasite development is mediated by parasite surface glycoconjugates and sand fly lectins, however we show here that interactions involving 'permissive' sand flies utilize another molecules. We did find that the abundant surface glycoconjugate lipophosphoglycan, essential for attachment of Leishmania major in the specific vector P. papatasi, was not required for parasite adherence or survival in the permissive vectors P. arabicus and Lutzomyia longipalpis. Attachment in several permissive sand fly species instead correlated with the presence of midgut glycoproteins bearing terminal N-acetyl-galactosamine and with the occurrence of a lectin-like activity on Leishmania surface. This new binding modality has important implications for parasite transmission and evolution. It may contribute to the successful spreading of Leishmania due to their adaptation into new vectors, namely transmission of L. infantum by Lutzomyia longipalpis; this event led to the establishment of L. infantum/chagasi in Latin America.     

Performance of light‐emitting diode traps for collecting sand flies in entomological surveys in Argentina

The performance of two light‐emitting diode traps with white and black light for capturing phlebotomine sand flies, developed by the Argentinean Leishmaniasis Research Network (REDILA‐WL and REDILA‐BL traps), were compared with the traditional CDC incandescent light trap. Entomological data were obtained from six sand fly surveys conducted in Argentina in different environments. Data analyses were conducted for the presence and the abundance of Lutzomyia longipalpis, Migonemyia migonei, and Nyssomyia whitmani (106 sites). No differences were found in presence/absence among the three types of traps for all sand fly species (p>0.05). The collection mean of Lu. longipalpis from the REDILA‐BL didn´t differ from the CDC trap means, nor were differences seen between the REDILA‐WL and the CDC trap collection means (p>0.05), but collections were larger from the REDILA‐BL trap compared to the REDILA‐WL trap (p<0.05). For Mg. migonei and Ny. whitmani, no differences were found among the three types of traps in the number of individuals captured (p>0.05). These results suggest that both REDILA traps could be used as an alternative capture tool to the original CDC trap for surveillance of these species, and that the REDILA‐BL will also allow a comparable estimation of the abundance of these flies to the CDC light trap captures. In addition, the REDILA‐BL has better performance than the REDILA‐WL, at least for Lu. longipalpis.     

Horizontal stratification of the sand fly fauna (Diptera: Psychodidae) in a transitional vegetation between caatinga and tropical rain forest, state of Bahia, Brazil

A study about the horizontal stratification of the sand fly fauna in two distinct ecosystems, caatinga area, endemic for visceral leishmaniasis, and the tropical rain forest area, endemic for cutaneous leishmaniasis, was performed in the state of Bahia, Brazil. Lutzomyia longipalpis was predominant in the caatinga, and following it came the species L. capixaba and L. oswaldoi. In the tropical rain forest other species were found, such as L. intermedia, L. migonei, L. whitmani, L. yuilli, L.fischeri, L. damascenoi, L. evandroi, L. monticola, and L. lenti. It was found that the geographical limits of the vector species of visceral and cutaneous leishmaniasis are clearly defined by the biological and phytogeographic characteristics.     

Standardization of the method proposed by the World Health Organization for determining the susceptibility levels of leishmaniasis vectors to insecticides

The WHO method for determining insecticide resistance was standardized for several species of Lutzomyia sand flies under laboratory and field conditions. The biological assays were applied solely to optimize the conditions for the control, i.e., without insecticide, and to estimate mortality due to handling or other unfavorable conditions. Adult female flies from 3 laboratory colonies and one field strain were tested: two laboratory strains of Lutzomyia longipalpis, one laboratory strain of Lutzomyia serrana and one field-collected strain of Lutzomyia quasitownsendi. The WHO method was compared with one modified in which, during the post-exposure period, the recommended plain tube apparatus was replaced with a plastic container layered with damp plaster of Paris. Three paper substrate types were compared under each condition: olive oil additive, silicon oil additive and plain paper. The measured variable was percent mortality in 24 h. For the WHO protocol, the L. longipalpis strains indicated a 0-10% mortality, L. serrana 20-80% and L. quasitownsendi 10-50%. With the modified WHO apparatus, the average mortality was < 4% for all species. No significant differences were observed among the paper treatments. These results indicate a strong species-specific effect of post-exposure conditions on sand flies. To establish baseline levels of insecticide resistance in Lutzomyia sand flies, the WHO method is recommended only for L. longipalpis, and the modified method for L. serrana, L. quasitownsendi and closely related species.     

Sand fly specificity of saliva-mediated protective immunity in Leishmania amazonensis-BALB/c mouse model

Immune response of BALB/c mice to the salivary antigens of sand flies was found to vary with different species used, i.e. Phlebotomus papatasi, Phlebotomus sergenti and Lutzomyia longipalpis. Exposure of mice to bites of these sand flies elicits production of antibodies, which are largely specific to different saliva antigens previously identified as unique to the respective fly species. When immunized intradermally (i.d.) with salivary gland lysates (SGL) of L. longipalpis, BALB/c mice developed partial protective immunity against challenges in the contralateral ears with Leishmania amazonensis plus the gland lysates. Preimmunization of these mice with the lysates from the other two species was ineffective, further indicative of the specificity of saliva-mediated immune response. The partial protective immunity observed is significant, although it is not as dramatic as reported previously in a different sand fly-mouse model. There is a correlation of this immunity with a lower number of mononuclear and polymorphonuclear phagocytes at the site of parasite inoculation. Vector species-specificity of this immunity implies its elicitation by unique saliva antigen-an issue which requires attention when designing saliva-based vaccines against leishmaniasis.     

Leishmania chitinase facilitates colonization of sand fly vectors and enhances transmission to mice

Chitinases of trypanosomatid parasites have been proposed to fulfil various roles in their blood-feeding arthropod vectors but so far none have been directly tested using a molecular approach. We characterized the ability of Leishmania mexicana episomally transfected with LmexCht1 (the L. mexicana chitinase gene) to survive and grow within the permissive sand fly vector, Lutzomyia longipalpis. Compared with control plasmid transfectants, the overexpression of chitinase was found to increase the average number of parasites per sand fly and accelerate the escape of parasites from the peritrophic matrix-enclosed blood meal as revealed by earlier arrival at the stomodeal valve. Such flies also exhibited increased damage to the structure of the stomodeal valve, which may facilitate transmission by regurgitation. When exposed individually to BALB/c mice, those flies with chitinase-overexpressing parasites spent on average 2.4-2.5 times longer in contact with their host during feeding, compared with flies with control infections. Furthermore, the lesions that resulted from these single fly bite infections were both significantly larger and with higher final parasite burdens than controls. These data show that chitinase is a multifunctional virulence factor for L. mexicana which assists its survival in Lu. longipalpis. Specifically, this enzyme enables the parasites to colonize the anterior midgut of the sand fly more quickly, modify the sand fly stomodeal valve and affect its blood feeding, all of which combine to enhance transmission.     

Host preferences of the phlebotomine sandfly Lutzomyia longipalpis in Amazonian Brazil

Experiments were undertaken to determine the relative attractiveness of humans, dogs and chickens to Lutzomyia longipalpis, the principal vector of Leishmania chagasi causing American visceral leishmaniasis. Field experiments in two villages on Marajó Island, Pará State, Brazil, showed that one boy attracted significantly more flies than one dog or chicken, and slightly fewer flies than a group of six chickens. Experiments with laboratory-bred female flies showed that a significantly greater number of flies engorged on a single human than on either a single dog or chicken, and man-biting catches demonstrated the willingness of flies to bite in the field. It appears that Lu.longipalpis has catholic feeding habits, the attractiveness of different hosts being largely a function of their relative sizes. These results are discussed with reference to the epidemiology of visceral leishmaniasis in Brazil.