Female Lu Research Articles

Species diversity, barcode, detection of pathogens and blood meal pattern in Phlebotominae (Diptera: Psychodidae) from northeastern Mexico

More than 90 species of phlebotomines are vectors of parasites, bacteria, and viruses, which cause disease in animals and humans. Therefore, their study is necessary to establish prevention and control strategies. Mexico is an endemic country for leishmaniasis, mostly in the center and southern regions of the country, yet only few studies have been conducted in the northern part of the country. The present study aims to: (a) assess the alpha diversity of Phlebotominae in an annual cycle, (b) to correlate climatic variables with abundance, (c) to generate barcodes of these insects as part of the integrative taxonomy, and (d) to detect Leishmania, Wolbachia and blood sources in an area close to where a case of autochthonous leishmaniasis has been detected in Nuevo Leon, Mexico. A systematic sampling was conducted during three consecutive nights from 17:00 to 22:00 h., placing Shannon traps, CDC traps with incandescent light, and BG Sentinel 2 + BG Lure traps. A total catch effort of 660 nights/traps/hours was achieved, in which a total number of 707 phlebotomines (58% female and 42% male) of six species were collected and identified. The most abundant species were Psathyromyia cratifer (57%) and Psathyromyia shannoni sensu stricto (26%). The highest abundance (72%; 507/707) was collected during March, April and May 2021. Barcodes were generated for four species of phlebotomines, which represent new records for Mexico. For the molecular detection of microorganisms, 302 specimens were analyzed, although no specimens were positive for Leishmania spp. Wolbachia strains were detected in phlebotomines with an infection rate of 1.32% (4/302) and found in Pa. cratifer and Lu. cruciata. Likewise, human DNA was identified in female Lu. cruciata and Pa. cratifer phlebotomines. These findings indicate the presence of potential vector species of the parasite Leishmania spp. This result shows the need for further entomological surveillance to elucidate the transmission mechanisms in these northern areas of the country.

Odour of domestic dogs infected with Leishmania infantum is attractive to female but not male sand flies: Evidence for parasite manipulation.

Globally visceral leishmaniasis (VL) causes thousands of human deaths every year. In South America, the etiologic agent, Leishmania infantum, is transmitted from an infected canine reservoir to human hosts by the bite of the sand fly vector; predominantly Lutzomyia longipalpis. Previous evidence from model rodent systems have suggested that the odour of infected hosts is altered by the parasite making them more attractive to the vector leading to an increased biting rate and improved transmission prospects for the pathogen. However, there has been no assessment of the effect of Le infantum infection on the attractiveness of dogs, which are the natural reservoirs for human infection. Hair collected from infected and uninfected dogs residing in a VL endemic city in Brazil was entrained to collect the volatile chemical odours present in the headspace. Female and male Lu. longipalpis sand flies were offered a choice of odour entrained from infected and uninfected dogs in a series of behavioural experiments. Odour of uninfected dogs was equally attractive to male or female Lu. longipalpis when compared to a solvent control. Female Lu. longipalpis were significantly more attracted to infected dog odour than uninfected dog odour in all 15 experimental replicates (average 45.7±0.87 females attracted to infected odour; 23.9±0.82 to uninfected odour; paired T-test, P = 0.000). Male Lu. longipalpis did not significantly prefer either infected or uninfected odour (average 36.1±0.4 males to infected odour; 35.7±0.6 to uninfected odour; paired T-test, P = 0.722). A significantly greater proportion of females chose the infected dog odour compared to the males (paired T-test, P = 0.000). The results showed that the odour of dogs infected with Le. infantum was significantly more attractive to blood-seeking female sand flies than it was to male sand flies. This is strong evidence for parasite manipulation of the host odour in a natural transmission system and indicates that infected dogs may have a disproportionate significance in maintaining infection in the canine and human population.

Modelling Sand Fly Lutzomyia longipalpis Attraction to Host Odour: Synthetic Sex-Aggregation Pheromone Dominates the Response.

Zoontic visceral leishmaniasis (ZVL) due to Leishmania infantum is a potentially fatal protozoan parasitic disease of humans and dogs. In the Americas, dogs are the reservoir and the sand fly, Lutzomyia longipalpis, the principal vector. A synthetic version of the male sand fly produced sex-aggregation pheromone attracts both female and male conspecifics to co-located insecticide, reducing both reservoir infection and vector abundance. However the effect of the synthetic pheromone on the vector’s “choice“ of host (human, animal reservoir, or dead-end host) for blood feeding in the presence of the pheromone is less well understood. In this study, we developed a modelling framework to allow us to predict the relative attractiveness of the synthetic pheromone and potential alterations in host choice. Our analysis indicates that the synthetic pheromone can attract 53% (95% CIs: 39%–86%) of host-seeking female Lu. longipalpis and thus it out-competes competing host odours. Importantly, the results suggest that the synthetic pheromone can lure vectors away from humans and dogs, such that when co-located with insecticide, it provides protection against transmission leading to human and canine ZVL.

Community deployment of a synthetic pheromone of the sand fly Lutzomyia longipalpis co-located with insecticide reduces vector abundance in treated and neighbouring untreated houses: Implications for control of Leishmania infantum.

The rising incidence of visceral leishmaniasis due to Leishmania infantum requires novel methods to control transmission by the sand fly vector. Indoor residual spraying of insecticide (IRS) against these largely exophilic / exophagic vectors may not be the most effective method. A synthetic copy of the male sex-aggregation pheromone of the key vector species Lutzomyia longipalpis in the Americas, was co-located with residual pyrethroid insecticide, and tested for its effects on vector abundance, hence potential transmission, in a Brazilian community study. Houses within eight defined semi-urban blocks in an endemic municipality in Brazil were randomised to synthetic pheromone + insecticide or to placebo treatments. A similar number of houses located >100m from each block were placebo treated and considered as "True Controls" (thus, analysed as three trial arms). Insecticide was sprayed on a 2.6m2 surface area of the property boundary or outbuilding wall, co-located within one metre of 50mg synthetic pheromone in controlled-release dispensers. Vector numbers captured in nearby CDC light traps were recorded at monthly intervals over 3 months post intervention. Recruited sentinel houses under True Control and pheromone + insecticide treatments were similarly monitored at 7-9 day intervals. The intervention effects were estimated by mixed effects negative binomial models compared to the True Control group. Dose-response field assays using 50mg of the synthetic pheromone captured a mean 4.8 (95% C.L.: 3.91, 5.80) to 6.3 (95% C.L.: 3.24, 12.11) times more vectors (female Lu. longipalpis) than using 10mg of synthetic pheromone. The intervention reduced household female vector abundance by 59% (C.L.: 48.7, 66.7%) (IRR = 0.41) estimated by the cross-sectional community study, and by 70% (C.L.: 56.7%, 78.8%) estimated by the longitudinal sentinel study. Similar reductions in male Lu. longipalpis were observed. Beneficial spill-over intervention effects were also observed at nearby untreated households with a mean reduction of 24% (95% C.L.: 0.050%, 39.8%) in female vectors. The spill-over effect in untreated houses was 44% (95% C.L.: 29.7%, 56.1%) as effective as the intervention in pheromone-treated houses. Ownership of chickens increased the intervention effects in both treated and untreated houses, attributed to the suspected synergistic attraction of the synthetic pheromone and chicken kairomones. The variation in IRR between study blocks was not associated with inter-household distances, household densities, or coverage (proportion of total households treated). The study confirms the entomological efficacy of the lure-and-kill method to reduce the abundance of this important sand fly vector in treated and untreated homesteads. The outcomes were achieved by low coverage and using only 1-2% of the quantity of insecticide as normally required for IRS, indicating the potential cost-effectiveness of this method. Implications for programmatic deployment of this vector control method are discussed.

Insecticide-impregnated netting: A surface treatment for killing Lutzomyia longipalpis (Diptera: Psychodidae), the vector of Leishmania infantum

The sand fly Lutzomyia longipalpis is the main vector of Leishmania infantum in Brazil. Synthetic male-produced sex/aggregation pheromone co-located with micro-encapsulated λ-cyhalothrin in chicken sheds can significantly reduce canine infection and sand fly densities in a lure-and-kill strategy. In this study, we determined if insecticide-impregnated netting (IN) could replace insecticide residual spraying (IRS). We compared numbers of Lu. longipalpis attracted and killed in experimental and real chicken sheds baited with pheromone and treated with a 1 m2 area of either insecticide spray or netting. First, we compared both treatments in experimental sheds to control mortality established from light trap captures. We then compared the long-term killing effect of insecticide spray and netting, without renewal, in experimental sheds over a period of 16 weeks. Finally, a longitudinal intervention study in real chicken sheds compared the numbers and proportions of Lu. longipalpis collected and killed before and after application of both treatments. In Experiment 1, a higher proportion of males and females captured in IRS- and IN-treated sheds were dead at 24 h compared to controls (P < 0.05). No difference was found in the proportion of females killed in sheds treated with IN or IRS (P = 0.15). A slightly higher proportion of males were killed by IRS (100%) compared to IN (98.6%; P < 0.05). In Experiment 2, IN- and IRS-treated traps were equally effective at killing females (P = 0.21) and males (P = 0.08). However, IRS killed a significantly higher proportion of females and males after 8 (P < 0.05) and 16 (P < 0.05) weeks. In Experiment 3, there was no significant difference between treatments in the proportion of females killed before (P = 0.88) or after (P = 0.29) or males killed before (P = 0.76) or after (P = 0.73) intervention. Overall, initially the IN was as effective as IRS at killing female and male Lu. longipalpis in both experimental and real chicken sheds. However, the relative lethal effect of the IN deteriorated over time when stored under prevailing environmental conditions.

Synthetic sex-aggregation pheromone of Lutzomyia longipalpis, the South American sand fly vector of Leishmania infantum, attracts males and females over long-distance.

BackgroundIn South America the sand fly Lutzomyia longipalpis is the predominant vector of Leishmania infantum, the parasite that causes canine and human visceral leishmaniasis. Co-location of synthetic male sex-aggregation pheromone with an insecticide provided protection against canine seroconversion, parasite infection, reduced tissue parasite loads, and female sand fly densities at households. Optimising the sex-aggregation pheromone + insecticide intervention requires information on the distance over which female and male Lu. longipalpis would be attracted to the synthetic pheromone in the field.Methodology/Principal findingsWild Lu. longipalpis were collected at two peridomestic study sites in Governador Valadares (Minas Gerais, Brazil). Sand flies were marked with coloured fluorescent powder using an improved protocol and then released into an existing domestic chicken shed at two independent sites, followed by recapture at synthetic-pheromone host-odour baited traps placed up to 30 metres distant from the release point.In total 1704 wild-caught Lu. longipalpis were released into the two chicken sheds. Overall 4.3% of the marked flies were recaptured in the pheromone baited experimental chicken sheds compared to no marked flies recaptured in the control sheds. At the first site, 14 specimens (10.4% of the marked and released specimens) were recaptured at 10m, 36 (14.8%) at 20m, and 15 (3.4%) at 30m. At the second site, lower recapture rates were recorded; 8 marked specimens (1.3%) were recaptured at 5 and 10m and no marked specimens were recaptured at 15m. Approximately 7x more marked males than females were recaptured although males were only 2x as common as females in the released population. 52% of the marked Lu. longipalpis were collected during the first night of sampling, 32% on the second night, and 16% on the third night.Conclusions/SignificanceThe study established that both male and female sand flies can be attracted to the synthetic sex-aggregation pheromone in the presence of host odour over distances up to at least 30m in the field depending on local environmental and meterological conditions.

Behavioral Aspects of Phlebotomine Sand Flies Associated with a Case of Cutaneous Leishmaniasis in Atlántico, Northern Colombia.

.After the first autochthonous case of cutaneous leishmaniasis was reported in the Atlántico department in the Caribbean region of Colombia, entomological sampling was conducted in the specific areas where the infection might have occurred. CDC traps were installed inside and outside dwellings in the peri-urban and rural areas of a settlement in the municipality of Luruaco. Sampling was performed during the night with protected human bait, and phlebotomine sand flies were actively sampled from potential diurnal resting sites within dwellings. Ten species of the genus Lutzomyia were identified; Lutzomyia evansi was the dominant species (78%) in the rural and peri-urban areas as well as in the different sampled habitats, followed by Lutzomyia panamensis and Lutzomyia gomezi. There was a 100% household infestation by Lu. evansi, and its indoor mean abundance was 13.3 sand flies/CDC trap/night. The indoor mean abundance of Lu. panamensis and Lu. gomezi was only 0.9 and 0.8 sand flies/CDC trap/night, respectively. Female Lu. evansi were collected with protected human bait, mostly in the peridomestic area, with sustained activity during the night and a slight increase in the activity from 19:00 to 23:00 hours. Of the total sand flies captured in the diurnal resting sites, 73.1% were collected from the walls of bedrooms and corresponded to Lu. evansi, Lutzomyia cayennensis cayennensis, and Lutzomyia trinidadensis. Owing to their vectorial importance, the species on which entomological surveillance should be focused are Lu. evansi, Lu. panamensis, and Lu. gomezi. The biting and resting behavior reported in this study will help guide vector prevention and the control of leishmaniasis within the study area.

Natural infection by Leishmania infantum in the Lutzomyia longipalpis population of an endemic coastal area to visceral leishmaniasis in Brazil is not associated with bioclimatic factors.

Visceral leishmaniasis (VL) is a zoonosis caused by the protozoan Leishmania infantum and in Brazil is transmitted mainly by the bite of Lutzomuyia longipalpis sand flies. Data about the presence, distribution, natural infection rate, seasonal and monthly dynamics of the vector population are important for optimizing the measures to control VL in endemic areas. This study aimed to identify sand fly fauna in an endemic area for VL to detect the prevalence of L. infantum infection in the Lu. longipalpis population and to elucidate the influence of bioclimatic factors on the monthly fluctuations of this vector. HP light traps were monthly set in the intradomicile and peridomicile of residences located in the central and beachfront areas of Camaçari, a VL endemic area. The sand fly collection was conducted in two periods: i) period 1—between December 2011 and November 2012 and ii) period 2—August 2014 and July 2015. Sand fly species were identified and detection of L. infantum infection by qPCR was performed in pools of female Lu. longipalpis. For the first time, the parasite load of positive pools was correlated with the number of Lu. longipalpis captured per month in both periods. Correlation analyses between the monthly fluctuation of the sand fly population and bioclimatic indices of the municipality in both collection periods were also performed. In both evaluated periods, more than 98% of the collected sand flies were Lu. longipalpis, confirming the predominance of this species in the region. It was captured mostly in the beachfront area in all months evaluated (99%). For the period 1, Leishmania DNA was detected in 81% of tested pools representing a minimal infection rate of 9.6%. In the period 2, 40% of the pools were positive with a minimal infection rate of 10.2%. Infected sand flies were only detected in the beachfront area in both periods. The parasite load was low and did not vary in the evaluated months despite the number of collected sand flies. No correlation was observed for climatic factors in both areas of Camaçari. These findings emphasize the high risk of Leishmania transmission in Camaçari regardless of the season and that other factors, aside from bioclimatic elements, are influencing the sand fly population monthly fluctuation.

Attraction of the sandfly Lutzomyia longipalpis to possible biomarker compounds from dogs infected with Leishmania infantum.

Lutzomyia longipalpis (Diptera: Psychodidae) is the primary vector of Leishmania infantum (Kinetoplastida: Trypanosomatidae) in the Americas. Studies have been carried out to identify new alternatives for monitoring and controlling this sandfly species, particularly with the use of chemical baits. The attractiveness of odours emitted by foxes and alcohols found in some plants has already been demonstrated in laboratory tests with Lu. longipalpis. However, no studies have evaluated the responses of these insects to volatile organic compounds (VOCs) emitted by dogs. The present study was carried out to investigate the effects on Lu. longipalpis of individual and blends of VOCs identified in hair from dogs infected with L. infantum. Effects in male and female Lu. longipalpis were assessed using wind tunnel methodology. Individual compounds including octanal, nonanal, decanal and heptadecane showed capacity for activating and/or attracting male Lu. longipalpis. Only decanal and nonanal showed effects on females. The combination of octanal, decanal and heptadecane increased activation and attraction behaviour in male sandflies, as did the blend of octanal and decanal. These findings indicate that VOCs emitted by dogs may be an interesting source of new attractants of sandflies.

Fast Population Growth in Physogastry Reproduction of Luciaphorus perniciosus (Acari: Pygmephoridae) at Different Temperatures.

Luciaphorus perniciosus Rack is one of the most serious pests of several cultivated mushroom species including Ganoderma lucidum (Fr.), Flammulina velutipes Karst., Auricularia polytricha (Mont.) Saac., Lentinus polychrous Lev., and Lentinus squarrosulus (Mont.) Singer in Thailand. Adult female Lu. perniciosus produce offspring inside their physogastric hysterosomas, with all embryos developing through to the adult stage while remaining in the abdomen. Once the abdomen ruptures, the female parent dies and the offspring consisting of mostly fertilized female adults along with a few male adults continue to emerge from the cadaver of the mother for a period of several days. This peculiar type of reproduction after the death of the mother is a special case for life table analysis and has not been discussed previously in demographic analyses. In this study, the life table data of this mite fed on Le. squarrosulus were collected at 25, 30, and 35 °C and analyzed by using the age-stage, two-sex life table. The standard errors of population parameters were estimated by using the bootstrap technique (200,000 bootstraps). At 25, 30, and 35 °C, females started reproduction at ages 9, 5, and 3 d, respectively; the net reproductive rates (R0) were 192.27, 253.81, and 234.11 offspring. Due to their rapid development and high fecundity, the r values were as high as 0.4189, 0.8653, and 1.0892 d-1 at 25, 30, and 35 °C, respectively. Computer projection indicated that the mushroom mites Lu. perniciosus is capable of a threefold daily increase at 35 °C.

Lundep, a Sand Fly Salivary Endonuclease Increases Leishmania Parasite Survival in Neutrophils and Inhibits XIIa Contact Activation in Human Plasma

Neutrophils are the host's first line of defense against infections, and their extracellular traps (NET) were recently shown to kill Leishmania parasites. Here we report a NET-destroying molecule (Lundep) from the salivary glands of Lutzomyia longipalpis. Previous analysis of the sialotranscriptome of Lu. longipalpis showed the potential presence of an endonuclease. Indeed, not only was the cloned cDNA (Lundep) shown to encode a highly active ss- and dsDNAse, but also the same activity was demonstrated to be secreted by salivary glands of female Lu. longipalpis. Lundep hydrolyzes both ss- and dsDNA with little sequence specificity with a calculated DNase activity of 300000 Kunitz units per mg of protein. Disruption of PMA (phorbol 12 myristate 13 acetate)- or parasite-induced NETs by treatment with recombinant Lundep or salivary gland homogenates increases parasite survival in neutrophils. Furthermore, co-injection of recombinant Lundep with metacyclic promastigotes significantly exacerbates Leishmania infection in mice when compared with PBS alone or inactive (mutagenized) Lundep. We hypothesize that Lundep helps the parasite to establish an infection by allowing it to escape from the leishmanicidal activity of NETs early after inoculation. Lundep may also assist blood meal intake by lowering the local viscosity caused by the release of host DNA and as an anticoagulant by inhibiting the intrinsic pathway of coagulation.

First comparative transcriptomic analysis of wild adult male and female Lutzomyia longipalpis, vector of visceral leishmaniasis.

Leishmaniasis is a vector-borne disease with a complex epidemiology and ecology. Visceral leishmaniasis (VL) is its most severe clinical form as it results in death if not treated. In Latin America VL is caused by the protist parasite Leishmania infantum (syn. chagasi) and transmitted by Lutzomyia longipalpis. This phlebotomine sand fly is only found in the New World, from Mexico to Argentina. However, due to deforestation, migration and urbanisation, among others, VL in Latin America is undergoing an evident geographic expansion as well as dramatic changes in its transmission patterns. In this context, the first VL outbreak was recently reported in Argentina, which has already caused 7 deaths and 83 reported cases.Insect vector transcriptomic analyses enable the identification of molecules involved in the insect's biology and vector-parasite interaction. Previous studies on laboratory reared Lu. longipalpis have provided a descriptive repertoire of gene expression in the whole insect, midgut, salivary gland and male reproductive organs. Nevertheless, the study of wild specimens would contribute a unique insight into the development of novel bioinsecticides. Given the recent VL outbreak in Argentina and the compelling need to develop appropriate control strategies, this study focused on wild male and female Lu. longipalpis from an Argentine endemic (Posadas, Misiones) and a Brazilian non-endemic (Lapinha Cave, Minas Gerais) VL location. In this study, total RNA was extracted from the sand flies, submitted to sequence independent amplification and high-throughput pyrosequencing. This is the first time an unbiased and comprehensive transcriptomic approach has been used to analyse an infectious disease vector in its natural environment. Transcripts identified in the sand flies showed characteristic profiles which correlated with the environment of origin and with taxa previously identified in these same specimens. Among these, various genes represented putative targets for vector control via RNA interference (RNAi).

Metagenomic Analysis of Taxa Associated with Lutzomyia longipalpis, Vector of Visceral Leishmaniasis, Using an Unbiased High-Throughput Approach

BackgroundLeishmaniasis is one of the most diverse and complex of all vector-borne diseases worldwide. It is caused by parasites of the genus Leishmania, obligate intramacrophage protists characterised by diversity and complexity. Its most severe form is visceral leishmaniasis (VL), a systemic disease that is fatal if left untreated. In Latin America VL is caused by Leishmania infantum chagasi and transmitted by Lutzomyia longipalpis. This phlebotomine sandfly is only found in the New World, from Mexico to Argentina. In South America, migration and urbanisation have largely contributed to the increase of VL as a public health problem. Moreover, the first VL outbreak was recently reported in Argentina, which has already caused 7 deaths and 83 reported cases.Methodology/Principal FindingsAn inventory of the microbiota associated with insect vectors, especially of wild specimens, would aid in the development of novel strategies for controlling insect vectors. Given the recent VL outbreak in Argentina and the compelling need to develop appropriate control strategies, this study focused on wild male and female Lu. longipalpis from an Argentine endemic (Posadas, Misiones) and a Brazilian non-endemic (Lapinha Cave, Minas Gerais) VL location. Previous studies on wild and laboratory reared female Lu. longipalpis have described gut bacteria using standard bacteriological methods. In this study, total RNA was extracted from the insects and submitted to high-throughput pyrosequencing. The analysis revealed the presence of sequences from bacteria, fungi, protist parasites, plants and metazoans.Conclusions/SignificanceThis is the first time an unbiased and comprehensive metagenomic approach has been used to survey taxa associated with an infectious disease vector. The identification of gregarines suggested they are a possible efficient control method under natural conditions. Ongoing studies are determining the significance of the associated taxa found in this study in a greater number of adult male and female Lu. longipalpis samples from endemic and non-endemic locations. A particular emphasis is being given to those species involved in the biological control of this vector and to the etiologic agents of animal and plant diseases.

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.

Chicken blood provides a suitable meal for the sand fly Lutzomyia longipalpis and does not inhibit Leishmania development in the gut

BackgroundThe aim of this study was to address the role of chickens as bloodmeal sources for female Lutzomyia longipalpis and to test whether chicken blood is harmful to Leishmania parasite development within the sand flies. Bloodmeal ingestion, excretion of urate, reproduction, fecundity, as well as Leishmania infection and development were compared in sand flies fed on blood from chickens and different mammalian sources.ResultsLarge differences in haemoglobin and protein concentrations in whole blood (dog>human>rabbit> chicken) did not correlate with differences in bloodmeal protein concentrations (dog = chicken>human>rabbit). This indicated that Lu. longipalpis were able to concentrate bloodmeals taken from different hosts using prediuresis and this was confirmed by direct observation. Sand flies fed on chickens or dogs produced significantly more eggs than those fed on human blood. Female Lu. longipalpis retained significantly more urate inside their bodies when fed on chicken blood compared to those fed on rabbit blood. However, when the amounts of urate excreted after feeding were measured, sand flies fed on rabbit blood excreted significantly more than those fed on chicken blood. There was no difference in female longevity after feeding on avian or mammalian blood.Sand flies infected via chicken blood produced Leishmania mexicana infections with a similar developmental pattern but higher overall parasite populations than sand flies infected via rabbit blood.ConclusionsThe results of this study help to define the role that chickens play in the epidemiology of leishmaniasis. The present study using a Lu. longipalpis/L. mexicana model indicates that chickens are suitable hosts to support a Lu. longipalpis population and that chicken blood is likely to support the development of transmissible Leishmania infections in Lu. longipalpis.

Population dynamics of sand flies (diptera: psychodidae) at two foci of leishmaniasis in Texas.

Sand flies were collected at a focus of leishmaniasis in Medina County, TX, from April through October 1997 and at a focus in Bexar County, TX, from April 1998 through December 1999. Lutzomyia diabolica (Hall) were collected from April through November with peak abundance in July. The male:female ratio of Lu. diabolica was 1:6.2. Almost all female Lu. diabolica in the collections were unfed. One gravid Lu. diabolica contained 49 ova. Female Lu. anthophora (Addis) were active from February through December with three peaks in abundance suggestive of successive generations. Unfed and gravid Lu. anthophora were collected in about equal numbers. Gravid females contained from 1 to 64 ova per female. The male:female ratio was 1:1.8, with male Lu. anthophora collected in all months. One female Lu. anthophora was found infected with Leishmania in July 1999. Lutzomyia texana (Dampf) were collected from April through October with peak abundance in April during 1997. The male:female ratio was 1:1.4, with most females unfed. Two gravid Lu. texana contained 32 and 102 ova. An undescribed species of Lutzomyia was found only at the Medina County site from May through September 1997. Trapping sites four times per month versus two times per month in 1999 did not appear to adversely affect the abundance of Lu. diabolica or Lu. anthophora. There were marked differences in the species composition and relative abundance at the different sites, indicating that the spatial distribution of sand flies is patchy in nature.

Influence of lysates of the salivary glands of Lutzomyia longipalpis on the development of a Leishmania-major -like parasite in the skin of the golden hamster

Twelve years ago, some mice inoculated with Leishmania major were found to develop larger lesions, containing more amastigotes, if the inoculum used to infect them contained a lysate of salivary glands from Lutzomyia longipalpis than if no lysate was included. In the present study, outbred golden hamsters (Mesocricetus auratus) were each inoculated in a footpad with 10(4), 10(5), 10(6) or 10(7) stationary-phase promastigotes of a Leishmania-major-like parasite (MHOM/BR/71/BH49). Some of the inocula used each contained a lysate of the salivary glands from a laboratory-reared, female Lu. longipalpis. Only the hamsters inoculated with 10(7) promastigotes each developed macroscopic cutaneous lesions (all 10 co-inoculated with lysate but only two of the 10 co-inoculated with diluent). Each of the lesions developed into cutaneous nodule affecting the dermis and underlying subcutaneous tissue of the inoculated footpad, with, histologically, an intensive, diffuse and productive, inflammatory reaction. There were no apparent differences between the lesions of hamsters infected with inocula containing salivary-gland lysate and those seen in the animals infected with lysate-free inocula. Future studies will follow the histological changes at the sites of Lu. longipalpis bites.

Cryptic Speciation in Lutzomyia (Nyssomyia) trapidoi (Fairchild &amp; Hertig) (Diptera: Psychodidae) Detected by Multilocus enzyme electrophoresis

Lutzomyia trapidoi is the major vector of cutaneous leishmaniasis in Ecuador. In the framework of an epidemiologic study, female Lu. trapidoi sand flies were captured on human bait in La Tablada and Paraiso Escondido. Some coloration heterogeneity among the specimens caught led us to look for the existence of cryptic species using multilocus enzyme electrophoresis. In 196 specimens studied, five of seven enzyme loci proved to be variable, making it possible to check for departures from panmixia both by Hardy-Weinberg statistics and linkage disequilibrium analysis. Two discrete groups were clearly distinguished, which could be differentiated by the diagnostic locus glycerophosphate dehydrogenase. The two groups occurred in sympatry within each locality. Genetic distances measured between these two groups were consistent with values usually found between distinct species. These results suggest the existence of a least two sibling species in Paraiso Escondido as well as La Tablada. The epidemiologic relevance of these results is discussed.

Dispersal of phlebotomine sand flies (Diptera: Psychodidae) in a Colombian focus of Leishmania (Viannia) braziliensis.

Dispersal of five species of phlebotomine sand flies was studied in a coffee plantation near Arboledas, Colombia, by mark-release-recapture studies using fluorescent powders. The estimated recapture rate for males of Lutzomyia shannoni marked and released during the day was 28.1%, significantly higher than that for all other species (p < 0.05). Recapture rate of female Lu. shannoni was 9.5%, and no females of the other four species were recovered. This suggests either that Lu. shannoni is a more sedentary species than the others, or that the large trees on which these insects were captured and recaptured function as foci of lekking behaviour in males. The high recapture rates of females of this species may indicate that oviposition occurs in close proximity to the bases of these trees. Although most marked sand flies were recaptured within 200 m of their release point, a single female Lu. gomezi was recovered 960 m away 36 h after release. This suggests that the dispersal capacity of Lutzomyia species may be greater than has been thought, an important consideration in future control programs directed against these insects in Leishmania-endemic areas.

Phlebotomine sandflies and leishmaniasis in British honduras (Belize)

In British Honduras (Belize), Leishmania mexicana is mainly a parasite of small forest rodents, in which the infection is usually apparent as a non-ulcerated, discrete lesion near the base of the tail. All infected rats have been captured in forest. The rat most frequently infected with L. mexicana is Ototylomys phyllotis, the most abundant and most successful rodent in forest in British Honduras. It is a strictly forest-dwelling animal with scansorial habits. L. mexicana also occurs in Heteromys desmarestianus, a ground-dwelling forest rat that occasionally extends into savanna, farmland and houses. Accidental rodent hosts of L. mexicana are Sigmodon hispidus, a rat of savanna that sometimes invades forest, and Nyctomys sumichrasti, a forest-dweller with arboreal habits. Man is an accidental host for L. mexicana. A survey revealed that 10·8% of the populations of certain rural communities in British Honduras gave positive responses to Montenegro antigen. Positive reactions were more frequent in men (18·1%) than in women (3·7%). The incidence of human leishmaniasis is highest in the central part of British Honduras, occurring among persons with past or present association with the forest extending eastwards from El Peten Department of Guatemala; L. mexicana infections are rare or absent from the north and south of British Honduras. An account is given of the development of L. mexicana lesions on man, and selected cases of human leishmaniasis are described briefly. Evidence of the role of phlebotomine sandflies in the dissemination of American cutaneous leishmaniasis is reviewed. Details are given of phlebotomine sandflies caught in British Honduras between 1963 and 1969; 3 species of Brumptomyia and 22 species of Lutzomyia were collected. Notes are given of the species of phlebotomines taken on human bait, with rat-baited oil traps, with C.D.C. miniature light traps and from resting-places such as the buttressed roots of trees, beneath decaying leaves on the forest floors, and the walls of caves. Geographical distribution and abundance, vertical zonation and biting rhythms of phlebotomines in British Honduras are discussed and observations on the distribution of sandfly bites on the human body are reported. A description is given of the techniques used to examine phlebotomines for flagellate infections and of methods used to identify such infections. Trypomastigote or epimastigote infections or both were found in specimens of Lu. cruciata, Lu. steatopyga, Lu. shannoni and Lu. trinidadensis. Promastigote infections were discovered in Lu. cruciata, Lu. ovallesi, Lu. olmeca, Lu. panamensis and Lu. permira. Only promastigotes isolated from Lu. olmeca gave rise to characteristic L. mexicana lesions when inoculated into hamsters. Transmission of leishmaniasis to rodents by Lu. olmeca is discussed. A theory is presented to explain how Lu. olmeca, a species not attracted to take blood meals on man, could be responsible for the transmission of L. mexicana to man. It is suggested that leishmaniasis in British Honduras is acquired when the daytime resting places of Lu. olmeca (decaying leaves on the forest floor) are disturbed during hours of daylight. Evidence is offered to show that when man disturbs leaves on the forest floor in the hour or so after dawn, he runs the same risks as a susceptible rat of being bitten by a female Lu. olmeca infected with L. mexicana.