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Prevention of canine Leishmania infantum infection is critical to management of visceral leishmaniasis in people living in endemic areas of Brazil. A bill (PL 1738/11), currently under consideration, proposes to establish a national vaccination policy against canine leishmaniasis in Brazil. However, there is no solid scientific evidence supporting the idea that this could reduce transmission from infected vaccinated dogs to sand flies to a level that would significantly reduce the risk of L. infantum infection or visceral leishmaniasis in humans. Thus, we advocate that insecticide-impregnated collars should the first line protective measure for public health purposes and that vaccines are applied on a case-by-case, optional basis for individual dog protection.

Objective:\ud Failure to control domestic Triatoma infestans in the Chaco is attributed to vulnerable adobe construction which provides vector refuges and diminishes insecticide contact. We conducted a pilot to test impact of housing improvement plus indoor residual spraying (IRS) on house infestation and vector abundance in a rural community in the Bolivian Chaco.\ud \ud Methods:\ud The intervention included three arms: housing improvement + IRS [HI], assisted IRS [AS] in which the team helped to clear the house pre-IRS, and routine IRS [RS]. HI used locally available materials, traditional construction techniques, and community participation. Vector parameters were assessed by Timed Manual Capture for 2 person-hours per house at baseline and medians of 114, 173, 314, 389, and 445 days post-IRS-1. A second IRS round was applied at a median of 314 days post-IRS-1.\ud \ud Results:\ud Post-intervention infestation indices and abundance fell in all three arms. The mean odds of infestation was 0.29 (95% CL 0.124, 0.684) in the HI relative to the RS arm. No difference was observed between AS and RS. Vector abundance was reduced by a mean 44% (24.8, 58.0) in HI compared to RS, with no difference between AS and RS. Median delivered insecticide concentrations per house were lower than the target of 50mg/m2 in >90% of houses in all arms.\ud \ud Conclusion:\ud Housing improvement using local materials and community participation is a promising strategy to improve IRS effectiveness in the Bolivian Chaco. A larger trial is needed to quantify the impact on re-infestation over time.

This repository contains datasets used to parametrised attraction profiles of Lu. longipalpis.Each dataset is provided as a tabulated text file, with each row corresponding to the outcome of a single experiment. Data were collated from the following sources: Dataset A:Bray, D.P.; Carter, V.; Alves, G.B.; Brazil, R.P.; Bandi, K.K.; Hamilton, J.G.C. Synthetic Sex Pheromone in a Long-Lasting Lure Attracts the Visceral Leishmaniasis Vector, Lutzomyia longipalpis, for up to 12 Weeks in Brazil. PLoS Negl. Trop. Dis. 2014, 8, e2723, doi:10.1371/journal.pntd.0002723. Dataset B:González, M.A.; Bell, M.; Souza, C.F.; Maciel-De-Freitas, R.; Brazil, R.P.; Courtenay, O.; Hamilton, J.G.C. Synthetic sex-aggregation pheromone of Lutzomyia longipalpis, the South American sand fly vector of Leishmania infantum, attracts males and females over long-distance. PLoS Negl. Trop. Dis. 2020, 14, e0008798, doi:10.1371/journal.pntd.0008798. Dataset C:Dilger, E. The Effects of Host-Vector Relationships and Density Dependence on the Epidemiology of Visceral Leishmaniasis. Ph.D. Thesis, University of Warwick, 2013. Dataset D:Dye, C.; Davies, C.R.; Lainson, R. Communication among phlebotomine sandflies: A field study of domesticated Lutzomyia longipalpis populations in Amazonian Brazil. Anim. Behav. 1991, 42, 183–192, doi:10.1016/S0003-3472(05)80549-4.

Background In Brazil, members of the sand fly species complex Lutzomyia longipalpis transmit Leishmania infantum, a protist parasite that causes visceral leishmaniasis. Male Lu. longipalpis produce a sex pheromone that is attractive to both females and males. During a cluster randomised trial, to determine the combined effect of synthetic sex-aggregation pheromone and insecticide on Le. infantum transmission Lu. longipalpis had been continuously exposed to insecticide for 30 months. The objective of this study was to determine the effect of continuous exposure to the insecticides used in the trial on the susceptibility of Lu. longipalpis population. Methods During the trial the sand flies had been exposed to either lambda-cyhalothrin [pheromone + residual insecticide spray (PI)], deltamethrin [dog collars (DC)] or no insecticide [control (C)], for 30 months (November 2012 to April 2015). The insecticide treatment regime was kept in place for an additional 12 months (May 2015-April 2016) during this susceptibility study. Sand flies collected from the field were exposed to WHO insecticide-impregnated papers cyhalothrin (0.05%), deltamethrin (0.5%) and control (silicone oil) in a modified WHO insecticide exposure trial to determine their susceptibility. Results We collected 788 Lu. longipalpis using CDC-light traps in 31 municipalities across the three trial arms. Probit analysis showed that the knockdown times (KDTs) of Lu. longipalpis collected from the lambda-cyhalothrin exposed PI-arm [KDT50: 31.1 min, confidence interval (CI): 29.6–32.6 and KDT90: 44.2 min, CI: 42.1–46.7] were longer than the KDTs from the non-insecticide-treated C-arm (KDT50: 26.3 min, CI: 25.1–27.6 and KDT90: 38.2, CI: 36.5–40.2) (no-overlapping 95% CIs). KDTs of Lu. longipalpis collected from the deltamethrin exposed DC-arm had similar values (KDT50: 13.7 min, CI: 10.1–16.2 and KDT90: 26.7 min, CI: 21.8–30.6) to those for the C-arm (KDT50: 13.5 min; CI: 12.2–14.8 and KDT90: 23.2 min, CI: 21.4–25.4) (overlapping CIs). The wild-caught unexposed Lu. longipalpis (C-arm), took approximately twice as long to knock down as laboratory-colonised specimens for both insecticides. Conclusions Our study reveals slight changes in KDT, in sand flies after prolonged exposure to lambda-cyhalothrin in the presence of pheromone. These changes are not considered to have reached the reference levels indicative of resistance in sand flies suggesting that pheromone and insecticide treatment at the level indicated in this study do not constitute a significant risk of increased insecticide resistance. Prolonged exposure to deltamethrin in dog collars did not result in changes to KDT. Electronic supplementary material The online version of this article (10.1186/s13071-019-3364-4) contains supplementary material, which is available to authorized users.

BackgroundVisceral leishmaniasis (VL) is a potentially fatal protozoan parasitic disease of humans and dogs. In the Americas, dogs are the reservoir and Lutzomyia longipalpis is the sand fly vector. A synthetic version of the vector’s sex-aggregation pheromone attracts conspecifics to co-located lethal insecticide, reducing reservoir infection and vector abundance. Mathematical models of spatially deployed interventions are lacking, thus best practise using this novel lure-and-kill vector control approach to reduce infection incidence has not been fully explored.MethodsWe developed a predictive mathematical model of vector host-seeking behaviour combined with spatially explicit transmission models to evaluate changes in human and canine spatial infection incidence under variable pheromone implementation scenarios and demographic conditions.ResultsThe risk of human infection increased exponentially with canine incidence, but at different rates between rural and urban settings with spatial clustering of high forces of human infection related to their proximity to canine infected households. A predicted 70% household coverage using a cost-effective amount (50mg) of pheromone, plus insecticide, reduced rural and urban setting infection incidence by approximately 44% and 50% in dogs, and by 64% and 68% in humans, within 1-2 years. Near or complete transmission elimination in dogs and humans was achieved after 10 years intervention using 50mg of pheromone under 70% household coverage in urban settings when newly acquired (immigrant) dogs had no pre-existing infections, but in rural settings required 90% coverage using 500mg. The willingness to pay (WTP) price per 10mg unit of pheromone is likely to be ConclusionsIntegrated stochastic and spatial models capturing vector host-seeking behaviour, is a useful mathematical framework to evaluate spatially dependent intervention methods, fine-scale transmission dynamics, and to identify best practise.

Visceral Leishmaniasis (VL) due to Leishmania donovani is a neglected protozoan parasitic disease in humans, which is usually fatal if untreated. Phlebotomus orientalis, the predominant VL vector in East Africa, is a highly exophilic/exophagic species that poses a major challenge to current Integrated Vector Management (IVM). Here we report results of pilot studies conducted in rural villages in Gedarif state, Sudan, to evaluate outdoor residual spraying of 20mg active ingredient (a.i.) /m2 deltamethrin insecticide applied to the characteristic household compound boundary reed fence and to the outside of household buildings (Outdoor Residual Insecticide Spraying, ODRS), and as an alternative, spraying restricted to the boundary fence only (Restricted Outdoor Residual Insecticide Spraying, RODRS). Four to six clusters of 20 households were assigned to insecticide treatments or control in three experiments. Changes in sand fly numbers were monitored over 2,033 trap-nights over 43–76 days follow-up in four sentinel houses per cluster relative to unsprayed control clusters. Sand fly numbers were monitored by sticky traps placed on the ground on the inside (“outdoor”) and the outside (“peridomestic”) of the boundary fence, and by CDC light traps suspended outdoors in the household compound. The effects of ODRS on sand fly numbers inside sleeping huts were monitored by insecticide knockdown. After a single application, ODRS reduced P. orientalis abundance by 83%-99% in outdoor and peridomestic trap locations. ODRS also reduced numbers of P. orientalis found resting inside sleeping huts. RODRS reduced outdoor and peridomestic P. orientalis by 60%-88%. By direct comparison, RODRS was 58%-100% as effective as ODRS depending on the trapping method. These impacts were immediate on intervention and persisted during follow-up, representing a large fraction of the P. orientalis activity season. Relative costs of ODRS and RODRS delivery were $5.76 and $3.48 per household, respectively. The study demonstrates the feasibility and high entomological efficacy of ODRS and RODRS, and the expected low costs relative to current IVM practises. These methods represent novel sand fly vector control tools against predominantly exophilic/exophagic sand fly vectors, aimed to lower VL burdens in Sudan, with potential application in other endemic regions in East Africa., Author summary Phlebotomus orientalis is the predominant vector of visceral leishmaniasis (VL, kala azar) in Sudan and other countries of East Africa, where the disease causes high morbidity and mortality. This sylvatic sand fly species is abundant in wild habitats characterized by presence of black cotton soil and vegetation dominated by Balanites aegyptiaca and/or Acacia seyal trees. In villages, the vector bites people in the household yard and in nearby peri-domestic locations, exhibiting limited indoor resting behaviour. The marked exophagic and exophilic behaviours of P. orientalis represent a profound challenge for VL control by excluding indoor residual spraying of insecticides (IRS) and compromising the efficacy of insecticide-impregnated bednets (ITNs). In this study, we evaluated the entomological efficacy of residual pyrethroid applied outdoors to household boundary fences and the exterior walls of household huts (outdoor residual insecticide spraying, ODRS), to reduce the abundance of P. orientalis inside and outside houses. We also evaluated the entomological impact of a restricted outdoor residual insecticide spraying (RODRS), whereby insecticide was applied only to the boundary fence. The study was carried out in June 2016-June 2017 in Jebel-Algana and Umsalala villages, Gedarif state, eastern Sudan, which are highly endemic for VL. The results showed that a single ODRS application of 20mg a.i. /m2 2.8% deltamethrin provided average reductions of 83%-99% in outdoor and peridomestic P. orientalis sand fly numbers relative to unsprayed control clusters. RODRS reduced outdoor and peridomestic P. orientalis by 60%-88%. The average cost of ODRS and RODRS per household were $5.76 and $3.48, respectively. The costs of these community-based control measures were substantially lower than the costs of LLINs, which is the only evidence-based tool used to protect against VL in the area. Future studies should evaluate the impact of ODRS/RODRS transmission of VL incidence in endemic villages and in seasonal agricultural farms.

Background 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. Methods 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. Results 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). Conclusions 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., Author summary The predominant sand fly vector of the intracellular parasite Leishmania infantum, that causes human and canine visceral leishmaniasis in the Americas, is Lutzomyia longipalpis. Vector control tools to reduce transmission are needed. A sex-aggregation pheromone released by male Lu. longipalpis attracts female conspecifics which facilitates blood-feeding and transmission. This study, conducted in Brazil, quantifies the effects of community deployment of a synthetic version of the sex-aggregation pheromone, in a controlled-release dispenser, and co-located with lethal insecticide applied to a small area of the household compound or outbuilding wall. 50mg synthetic pheromone dispensers were used since they attracted substantially more vectors than 10mg dispensers. Deploying this novel lure-and-kill method to houses in eight replicate study blocks in two suburban endemic areas, demonstrated that it reduces vector numbers at both pheromone + insecticide treated houses, and neighbouring untreated houses. The presence of chickens (a known additional attraction to blood-seeking female Lu. longipalpis) increased the beneficial effects of the intervention. The method used only 1–2% of the quantity of insecticide necessary for IRS for an average sized house. The results demonstrate the efficacy and potential cost-effectiveness of this novel lure-and-kill control method.

Vector-borne diseases (VBDs) such as malaria, dengue, and leishmaniasis exert a huge burden of morbidity and mortality worldwide, particularly affecting the poorest of the poor. The principal method by which these diseases are controlled is through vector control, which has a long and distinguished history. Vector control, to a greater extent than drugs or vaccines, has been responsible for shrinking the map of many VBDs. Here, we describe the history of vector control programmes worldwide from the late 1800s to date. Pre 1940, vector control relied on a thorough understanding of vector ecology and epidemiology, and implementation of environmental management tailored to the ecology and behaviour of local vector species. This complex understanding was replaced by a simplified dependency on a handful of insecticide-based tools, particularly for malaria control, without an adequate understanding of entomology and epidemiology and without proper monitoring and evaluation. With the rising threat from insecticide-resistant vectors, global environmental change, and the need to incorporate more vector control interventions to eliminate these diseases, we advocate for continued investment in evidence-based vector control. There is a need to return to vector control approaches based on a thorough knowledge of the determinants of pathogen transmission, which utilise a range of insecticide and non–insecticide-based approaches in a locally tailored manner for more effective and sustainable vector control., Author summary Vector-borne diseases (VBDs) such as dengue, Chagas disease, human African trypanosomiasis (HAT), leishmaniasis, and malaria exert a huge burden of morbidity and mortality worldwide. The principal method by which these diseases are controlled is through vector control. The authors chart the history of vector control through time from elucidation of the transmission route of VBDs to the present day. Pre-1940 vector control relied heavily on environmental management and larval control based on a thorough understanding of pathogen transmission but was replaced by insecticide-based vector control, often deployed as a monotherapy. The authors call for increased political will and investment in vector control and a return to locally tailored vector control that draws on the entire toolbox of interventions available.

Background: Screening for Trypanosoma cruzi infection was performed amongst children in a rural community in the Bolivian Chaco, an area known for high prevalence. The force of infection (FOI) was estimated. Methods: A total of 423 children attending the local school were screened using the Chagas Detect Plus (CDP) rapid test (InBios International, Inc.). CDP-positive specimens were further tested by indirect haemagglutination assay (IHA) and Wiener Recombinante v3.0 ELISA. A catalytic model was used to estimate FOI. Results: Confirmed seroprevalence was 0.22, rising steeply with age. The mean age of seropositive individuals was 13 years. The calculated specificity of the rapid test was 91.9%. The annual incidence estimated from the FOI was 0.021. Conclusions: This study demonstrates persistent transmission and continuing high levels of T. cruzi infection in the Bolivian Chaco, and highlights the practicality of school-based screening. Keywords: Chagas disease, Trypanosoma cruzi, T. cruzi, Bolivian Chaco, Screening study

This study was designed to investigate the prevalence of members of the Mycobacterium tuberculosis complex (MTBC) in the environment of pastoralists and villagers in the Iringa district, adjacent to the Ruaha National Park in Tanzania. Utilising specific qPCR assays, bothMycobacterium bovisandMycobacterium tuberculosiswere detected in cattle faeces, boma soil, water and household dust.M. boviswas also found in goat faeces and goat boma soil. This is the first report of faecal shedding ofM. bovisin goats and the first molecular survey of faecal shedding in cattle. The prevalence of both bacterial species varied by village, area, season and sample type. Geographical and temporal correlations across sample types were suggestive of cross species transmission. This non-invasive test has previously been rigorously validated for screening other mammals; in this study it has successfully been applied to detectM. bovisandM. tuberculosisin livestock faeces and the environment.

ObjectiveTo evaluate the efficacy of a synthetic sex-aggregation pheromone of the sand fly vector Lu. longipalpis, co-located with residual insecticide, to reduce the infection incidence of Leishmania infantum in the canine reservoir, and to reduce sand fly vector abundance. To compare the outcomes to those resulting from fitting deltamethrin-impregnated collars to the canine reservoir.MethodsA stratified cluster-randomised trial was designed to detect a 50% reduction in canine incident infection after 24 months in 42 recruited clusters, randomly assigned to one of three intervention arms (14 cluster each): pheromone + insecticide, insecticide-impregnated dog collars, or placebo control. Infection incidence was measured by seroconversion to anti-Leishmania antibody, Leishmania parasite detection and canine tissue parasite loads. Changes in relative Lu. longipalpis abundance within households were measured by setting three CDC light traps per household.ResultsA total 1,454 seronegative dogs were follow-up for a median 15.2 (95% C.I.s: 14.6, 16.2) months per cluster. The pheromone + insecticide intervention provided 13% (95% C.I. 0%, 44.0%) protection against anti-Leishmania antibody seroconversion, 52% (95% C.I. 6.2%, 74·9%) against parasite infection, reduced tissue parasite loads by 53% (95% C.I. 5.4%, 76.7%), and reduced household female sand fly abundance by 49% (95% C.I. 8.2%, 71.3%). Variation in the efficacy against seroconversion varied between trial strata. Equivalent protection attributed to the impregnated-collars were 36% (95% C.I. 14.4%, 51.8%), 23% (95% C.I. 0%, 57·5%), 48% (95% C.I. 0%, 73.4%) and 43% (95% C.I. 0%, 67.9%), respectively. Comparison of the outcomes of the two interventions showed no statistically consistent differences in their efficacies; however, the errors were broad for all outcomes. Reductions in sand fly numbers were predominant where insecticide was located (chicken and dog sleeping sites), with no evidence of insecticide-induced repellency onto humans or dogs.ConclusionThe synthetic pheromone lure-and-kill approach provides protection particularly against L. infantum parasite transmission and sand fly vector abundance. The effect estimates are not dissimilar to those of the insecticide-impregnated collars, which are documented to reduce canine infection incidence, and human infection and clinical VL disease incidence, in different global regions. As a low-cost alternative or complimentary vector control tool, optimisation of best community deployment of the pheromone + insecticide are now underway.Author summaryThe sand fly vector of the intracellular parasite Leishmania infantum causing human and canine visceral leishmaniasis in the Americas is Lutzomyia longipalpis. Dogs are the proven reservoir. Vector control tools to reduce transmission suited to this predominantly exophilic vector are lacking. Insecticide-impregnated dog collars protect dogs against infectious bites from sand fly vectors, resulting in reductions of new infections in both dogs and humans. However, collars are costly particularly for endemic communities, and alternative approaches are needed. Recent bulk synthesis of a sex-aggregation pheromone produced by male Lu. longipalpis was shown to attract large numbers of conspecific females to lethal pyrethroid insecticides. This study, conducted in Brazil, evaluated the efficacy of this novel lure-and-kill approach to reduce seroconversion and infection incidence with L. infantum in the canine reservoir, in addition to measuring its impact on household abundance of Lu. longipalpis. Deployed in 14 stratified clusters, the outcomes were compared to those resulting attributed to the collars fitted to dogs in another 14 clusters; each intervention was compared to the 14 clusters that received placebo treatments. The beneficial effects of the lure-and-kill method were most noticeable on confirmed infection incidence and clinical parasite loads, and in reducing sand fly abundance. The overall effect of the two interventions were not statistically dissimilar, though the confidence intervals were broad. We conclude that the novel low-cost lure-and-kill approach should be added to the vector control toolbox against visceral leishmaniasis in the Americas.

BackgroundThe parasite Leishmania infantum causes zoonotic visceral leishmaniasis (VL), a potentially fatal vector-borne disease of canids and humans. Zoonotic VL poses a significant risk to public health, with regions of Latin America being particularly afflicted by the disease.Leishmania infantum parasites are transmitted between hosts during blood feeding by infected female phlebotomine sand flies. With a principal reservoir host of L. infantum being domestic dogs, limiting prevalence in this reservoir may result in a reduced risk of infection for the human population. To this end, a primary focus of research efforts has been to understand disease transmission dynamics among dogs. One way this can be achieved is through the use of mathematical models.MethodsWe have developed a stochastic, spatial, individual-based mechanistic model of L. infantum transmission in domestic dogs. The model framework was applied to a rural Brazilian village setting with parameter values informed by fieldwork and laboratory data. To ensure household and sand fly populations were realistic, we statistically fit distributions for these entities to existing survey data. To identify the model parameters of highest importance, we performed a stochastic parameter sensitivity analysis of the prevalence of infection among dogs to the model parameters.ResultsWe computed parametric distributions for the number of humans and animals per household and a non-parametric temporal profile for sand fly abundance. The stochastic parameter sensitivity analysis determined prevalence of L. infantum infection in dogs to be most strongly affected by the sand fly associated parameters and the proportion of immigrant dogs already infected with L. infantum parasites.ConclusionsEstablishing the model parameters with the highest sensitivity of average L. infantum infection prevalence in dogs to their variation helps motivate future data collection efforts focusing on these elements. Moreover, the proposed mechanistic modelling framework provides a foundation that can be expanded to explore spatial patterns of zoonotic VL in humans and to assess spatially targeted interventions.

Made available in DSpace on 2018-12-11T17:06:15Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-09-15 Intensity of peripheral parasite infection has an important role in the transmission of Leishmania spp. from one host to another. As parasite load quantification is still an expensive procedure to be used routinely in epidemiological surveillance, the use of surrogate predictors may be an important asset in the identification of dogs with high transmitting ability. The present study examined whether common clinical and laboratory alterations can serve as predictors of peripheral parasitism in dogs naturally infected with Leishmania spp. Thirty-seven dogs were examined in order to establish correlations between parasite load (PL) in multiple peripheral tissues and common clinical and laboratory findings in canine visceral leishmaniasis (CVL). Quantitative polymerase chain reaction was employed to determine PL in conjunctival swabs, ear skin, peripheral blood and buffy coat. Additionally, a series of hematological, biochemical and oxidative stress markers were quantified. Correlations between net peripheral infection and severity of clinical alterations and variation in laboratory parameters were assessed through a new analytical approach, namely Compressed Parasite Load Data (CPLD), which uses dimension reduction techniques from multivariate statistics to summarize PL across tissues into a single variable. The analysis revealed that elevation in PL is positively correlated with severity of clinical sings commonly observed in CVL, such as skin lesions, ophthalmic alterations, onycogriphosis, popliteal lymphadenomegaly and low body mass. Furthermore, increase in PL was found to be followed by intensification of non-regenerative anemia, neutrophilia, eosinopenia, hepatic injury and oxidative imbalance. These results suggest that routinely used clinical and laboratory exams can be predictive of intensity of peripheral parasite infection, which has an important implication in the identification of dogs with high transmitting ability. Departamento de Clínica Cirurgia e Reprodução Animal Faculdade de Medicina Veterinária de Araçatuba UNESP- Univ Estadual Paulista, Rua Clóvis Pestana 793, Dona Amélia Departamento de Medicina Veterinária Preventiva e Reprodução Animal Faculdade de Ciências Agrárias e Veterinárias UNESP-Univ Estadual Paulista, Via de Acesso Prof. Paulo Donato Castellane s/n Departamento de Apoio Produção e Saúde Animal Faculdade de Medicina Veterinária de Araçatuba UNESP- Univ Estadual Paulista, Rua Clóvis Pestana 793, Dona Amélia School of Life Sciences and Warwick Infectious Disease and Epidemiology Research Group (WIDER), Gibbett Hill Road Departamento de Clínica Cirurgia e Reprodução Animal Faculdade de Medicina Veterinária de Araçatuba UNESP- Univ Estadual Paulista, Rua Clóvis Pestana 793, Dona Amélia Departamento de Medicina Veterinária Preventiva e Reprodução Animal Faculdade de Ciências Agrárias e Veterinárias UNESP-Univ Estadual Paulista, Via de Acesso Prof. Paulo Donato Castellane s/n Departamento de Apoio Produção e Saúde Animal Faculdade de Medicina Veterinária de Araçatuba UNESP- Univ Estadual Paulista, Rua Clóvis Pestana 793, Dona Amélia

Background In 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 findings Wild 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/Significance The 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., Author summary Visceral leishmaniasis (VL) is a disease caused by an insect transmitted protist parasite. In South America and Brazil in particular, it causes significant morbidity and mortality, with thousands of human cases and deaths reported every year. Domestic dogs are the most important source of human infection. Controlling the sand flies that transmit the parasite is one way to reduce the number of VL cases and recent research has shown the potential for a new pheromone-based approach to vector control. In a recent cluster-randomised control trial, co-location of a synthetic copy of the male sand fly sex-aggregation pheromone with pyrethroid insecticide reduced numbers of sand flies in households and provided protection for dogs against leishmaniasis infection incidence. The current study was carried out to determine the distance over which the synthetic sex-aggregation pheromone could attract Lu. longipalpis in a peridomestic environment in a Brazilian city. Male and female Lu. longipalpis were attracted up to 30m in one night towards a source of the pheromone. This information will help to inform the optimisation of placement of sex pheromone/insecticide intervention in Brazil.

Systemic insecticides in dogs have been suggested as a public health intervention to prevent human cases of Zoonotic Visceral Leishmaniasis (ZVL). But, currently there are no systemic insecticides for dogs registered against zoo-anthropophilic pool blood feeding phlebotomine flies. We predict the impact of community-wide use of systemic insecticide in dog populations as a public health measure to control transmission of Leishmania infantum to humans using a mathematical model. We developed a Susceptible-Exposed-Infected (SEI) compartmental model to describe L. infantum transmission dynamics in dogs, with a vectorial capacity term to represent transmission between L. infantum-hosting dogs via phlebotomine flies. For Infected (I) dogs two levels of infectiousness were modelled, high infectiousness and low infectiousness. Human incidence was estimated through its relationship to infection in the dog population. We evaluated outcomes from a wide range of scenarios comprising different combinations of initial insecticide efficacy, duration of insecticide efficacy over time, and proportion of the dog population treated (60%, 70% & 80%). The same reduction in human infection incidence can be achieved via different combinations of insecticide efficacy, duration and dog coverage. For example, a systemic insecticide with an initial efficacy of 80% and 6 months above 65% efficacy would require treating at least 70% of the dogs to reduce the human infection incidence by 50%. Sensitivity analysis showed that the model outcome was most sensitive to baseline values of phlebotomine fly daily survival rate and insecticide coverage. Community-wide use of systemic insecticides applied to the “L. infantum canine reservoir” can significantly reduce human incidence of L. infantum infection. The results of this mathematical model can help defining the insecticide target product profile and how the insecticide should be applied to maximise effectiveness., Author summary Zoonotic visceral leishmaniasis (ZVL) is a potentially deadly disease in humans caused by Leishmania infantum. This leishmania species can be delivered by pool blood feeding zoo-anthropophilic phlebotomine flies to several mammals, the dog population being recognized as the main reservoir. Transmission from infected dogs to humans is through the bite of female phlebotomine sand flies. The disease is endemic in several countries and Brazil has a high prevalence of cases with over 3000 ZVL cases reported per year. The main, inefficient and highly controversial, control measure in Brazil has been culling sero-positive dogs. The community-wide use of systemic insecticides in dogs could be an alternative to control L. infantum transmission from phlebotomine flies to humans. The rationale is that phlebotomine flies which sampled their blood meals from dogs treated with systemic insecticides would die reducing the risk of L. infantum transmission. To reduce the number of ZVL cases, a large proportion of dogs in the community should be treated and the systemic insecticide used should be effective in killing phlebotomine flies acting as vectors of L. infantum parasites for a significant amount of time. We used a mathematical model mimicking L. infantum transmission to show that this novel vector control strategy could be effective. We identified the combination of different key parameters (e.g. insecticide efficacy, duration and proportion of dogs treated) that could lead to a significant reduction of the risk of L. infantum infection in humans.

Visceral leishmaniasis (VL) is a life-threatening outcome of Leishmania infantum or Leishmania donovani infection. Dogs are the primary domestic reservoir of L. infantum parasites, and ownership of infected dogs increases the risk of human VL. Controlling infection within dog populations is regarded as critical to VL management in endemic countries, both preventing progression of canine disease and limiting parasite transmission to humans and dogs. Here we discuss various strategies that are used to diagnose canine visceral leishmaniasis (CVL) and the possibilities of adapting these for use within population screening and control programs. In addition, given the variable transmissibility of L. infantum to the sand fly vector, we outline some possibilities for the preferential identification of ‘super-spreader’ dogs among the overall infected population.\ud \ud

Quantitation of the nonlinear heterogeneities in Leishmania parasites, sand fly vectors, and mammalian host relationships provides insights to better understand leishmanial transmission epidemiology towards improving its control. The parasite manipulates the sand fly via production of promastigote secretory gel (PSG), leading to the “blocked sand fly” phenotype, persistent feeding attempts, and feeding on multiple hosts. PSG is injected into the mammalian host with the parasite and promotes the establishment of infection. Animal models demonstrate that sand flies with the highest parasite loads and percent metacyclic promastigotes transmit more parasites with greater frequency, resulting in higher load infections that are more likely to be both symptomatic and efficient reservoirs. The existence of mammalian and sand fly “super-spreaders” provides a biological basis for the spatial and temporal clustering of clinical leishmanial disease. Sand fly blood-feeding behavior will determine the efficacies of indoor residual spraying, topical insecticides, and bed nets. Interventions need to have sufficient coverage to include transmission hot spots, especially in the absence of field tools to assess infectiousness. Interventions that reduce sand fly densities in the absence of elimination could have negative consequences, for example, by interfering with partial immunity conferred by exposure to sand fly saliva. A deeper understanding of both sand fly and host biology and behavior is essential to ensuring effectiveness of vector interventions., Author summary We review recent research that sheds light on the quantitative biology of leishmanial transmission between sand flies and mammalian hosts and use these insights to better understand transmission, the observed epidemiology of the disease, and their implications in choice of control strategy. Using animal models, we show how the parasite-induced processes manipulate sand fly blood-feeding behavior and the infectious metacyclic dose to promote host infection and to differentially regulate the onward transmission potential of individual vectors and hosts. The existence of subpopulations of mammalian and sand fly “super-spreaders” provides a biological basis for the spatial and temporal clustering of clinical leishmanial disease. While tools are unavailable to distinguish these individuals in mixed populations, blanket interventions will be necessary to ensure inclusion of transmission hot spots. Interventions that reduce sand fly densities without elimination could interfere with vector—host dynamics and conferred partial immunity to host populations.

Visceral leishmaniosis is a zoonotic disease that is transmitted by Lutzomyia longipalpis sandflies. Dogs are the main peri-urban reservoir of the disease, and progression of canine leishmaniosis is dependent on the type of immune response elaborated against the parasite. Type 1 immunity is characterized by effective cellular response, with production of pro-inflammatory cytokines such as tumour necrosis factor alpha (TNF-α). In contrast, Type 2 immunity is predominantly humoral, associated with progression of the disease and mediated by anti-inflammatory cytokines such as interleukin 10 (IL-10). Although seemly important in the dynamics of leishmaniosis, other gene products such as toll-like receptor 2 (TRL-2) and inducible nitric oxide synthase (iNOS) exert unclear roles in the determination of the type of immune response. Given that the dog skin serves as a micro-environment for the multiplication of Leishmania spp., we investigated the parasite load and the expression of TLR-2, iNOS, IL-10 and TNF-α in the skin of 29 infected and 8 control dogs. We found that increased parasite load leads to upregulation of TLR-2, IL-10 and TNF-α, indicating that abundance of these transcripts is associated with infection. We also performed a xenodiagnosis to demonstrate that increased parasitism is a risk factor for infectiousness to sandflies.

The leishmaniases comprise a complex of diseases characterized by clinical outcomes that range from self-limiting to chronic, and disfiguring and stigmatizing to life threatening. Diagnostic methods, treatments, and vector and reservoir control options exist, but deciding the most effective interventions requires a quantitative understanding of the population level infection and disease dynamics. The effectiveness of any set of interventions has to be determined within the context of operational conditions, including economic and political commitment. Mathematical models are the best available tools for studying quantitative systems crossing disciplinary spheres (biology, medicine, economics) within environmental and societal constraints. In 2005, the World Health Assembly and government health ministers of India, Nepal, and Bangladesh signed a Memorandum of Understanding to eliminate the life threatening form of leishmaniasis, visceral leishmaniasis (VL), on the Indian subcontinent by 2015 through a combination of early case detection, improved treatments, and vector control. The elimination target is

Visceral Leishmaniasis (VL) is a neglected vector-borne disease. In India, it is transmitted to humans by Leishmania donovani-infected Phlebotomus argentipes sand flies. In 2005, VL was targeted for elimination by the governments of India, Nepal and Bangladesh by 2015. The elimination strategy consists of rapid case detection, treatment of VL cases and vector control using indoor residual spraying (IRS). However, to achieve sustained elimination of VL, an appropriate post elimination surveillance programme should be designed, and crucial knowledge gaps in vector bionomics, human infection and transmission need to be addressed. This review examines the outstanding knowledge gaps, specifically in the context of Bihar State, India.\ud \ud The knowledge gaps in vector bionomics that will be of immediate benefit to current control operations include better estimates of human biting rates and natural infection rates of P. argentipes, with L. donovani, and how these vary spatially, temporally and in response to IRS. The relative importance of indoor and outdoor transmission, and how P. argentipes disperse, are also unknown. With respect to human transmission it is important to use a range of diagnostic tools to distinguish individuals in endemic communities into those who: 1) are to going to progress to clinical VL, 2) are immune/refractory to infection and 3) have had past exposure to sand flies.\ud \ud It is crucial to keep in mind that close to elimination, and post-elimination, VL cases will become infrequent, so it is vital to define what the surveillance programme should target and how it should be designed to prevent resurgence. Therefore, a better understanding of the transmission dynamics of VL, in particular of how rates of infection in humans and sand flies vary as functions of each other, is required to guide VL elimination efforts and ensure sustained elimination in the Indian subcontinent. By collecting contemporary entomological and human data in the same geographical locations, more precise epidemiological models can be produced. The suite of data collected can also be used to inform the national programme if supplementary vector control tools, in addition to IRS, are required to address the issues of people sleeping outside.

In murine models of leishmaniasis, IgG subclass expression is a proxy measure for Th1/Th2 cellular immune response bias. However, in dogs, the reservoir of zoonotic visceral leishmaniasis, no consistent association has been described between IgG subclass ratios and disease resistance. Inconsistent results may reflect lack of specificity of commonly used commercial antibodies. Our aim was to measure IgG1 and IgG2 responses to crude Leishmania antigen using commercial polyclonal antibodies for comparison with a panel of commercially unavailable monoclonal antibodies, in a cohort of 60 naturally infected dogs, and to compare associations between subclass responses and clinical or parasitological outcomes. IgG1 and IgG2, measured by both antibodies, were higher in clinically symptomatic than in asymptomatic dogs (P

Leishmania infantum is the causative agent of infantile visceral leishmaniasis (IVL) in the Mediterranean Basin and, based on isoenzyme typing of a few isolates from patients and domestic dogs, this parasite was considered to predominate in the Kaleybar focus of IVL in northwest Iran. However, in the current investigation only one out of five sandfly infections was found to be L. infantum, based on PCR detection and sequencing of parasite internal transcribed spacer (ITS) rDNA infecting Phlebotomus perfiliewi transcaucasicus. The four other infections were of haplotypes of L. tropica, the causative agent of anthroponotic cutaneous leishmaniasis in the Middle East and a parasite occasionally detected in the viscera of dogs and patients in Iran and elsewhere. The widespread distribution of L. tropica in the Kaleybar focus suggests that this parasite is not a transient introduction. Kaleybar has been used for a deltamethrin dog collar intervention to reduce the biting rates of the vectors of L. infantum and this has significantly reduced the incidence of Leishmania infections both in children and the domestic dog, the usual reservoir host of IVL. The implications of finding L. tropica widespread in the heart of the intervention area are discussed. Extensive and intensive typing of natural Leishmania infections is a characteristic of epidemiological investigations in the Neotropics and the current report indicates that this will also be necessary in some regions of the Old World.

Synanthropic flies have been shown to be important mechanical vectors of Chlamydia trachomatis, which causes trachoma. However entomological studies have not been forthcoming in Latin America. This study assesses the relationship between household dipteran fly densities and active childhood trachoma in a village on Marajó Island, Pará state, Brazil. For 78 households, members were examined for signs of trachoma, relative abundance of potential trachoma vectors (Diptera, Chloropidae and Diptera, Muscidae) was quantified by trap counts, and additional measures of household hygiene associated with C. trachomatis transmission were assessed. Active childhood trachoma prevalence was 24.1% (45/187), present in 46.2% of sampled households with evidence of case clustering. Childhood prevalence was positively associated with increased fly densities, whereas indirect measures of sanitary conditions (latrine ownership and perceived importance of flies) showed a protective effect. This study indicates that C. trachomatis can be transmitted by synanthropic flies in this region of Latin America. Insetos sinantrópicos são importantes vetores mecânicos de Chlamydia trachomatis, causadora de tracoma, contudo, estudos entomológicos não são freqüentes na América Latina. Esse estudo determina a relação entre densidade de dípteros domésticos e tracoma ativo na infância em uma vila na Ilha do Marajó, Estado do Pará, Brasil. Moradores de 78 residências foram examinados para sinais de tracoma e a relativa abundância de potenciais vetores de tracoma (Diptera, Chloropidae e Diptera, Muscidae) foi quantificada junto com medidas adicionais de higiene doméstica associada com a transmissão de C. trachomatis. A prevalência de tracoma ativo na infância foi 24,1% (45/187), presente em 46,3% das residências amostradas com evidência de aglomeração de casos. A prevalência na infância foi positivamente associada com o aumento das densidades de insetos, enquanto medidas indiretas de condições sanitárias (possuir latrina e perceber a importância dos insetos) foram protetoras. Esse estudo indica que C. trachomatis pode ser transmitida por insetos sinantrópicos nessa região da América Latina.

Background Visceral leishmaniasis has been targeted for elimination as a public health problem (less than 1 case per 10,000 people per year) in the Indian sub-continent by 2017. However, there is still a high degree of uncertainty about the natural history of the disease, in particular about the duration of asymptomatic infection and the proportion of asymptomatically infected individuals that develop clinical visceral leishmaniasis. Quantifying these aspects of the disease is key for guiding efforts to eliminate visceral leishmaniasis and maintaining elimination once it is reached. Methods Data from a detailed epidemiological study in Bangladesh in 2002–2004 was analysed to estimate key epidemiological parameters. The role of diagnostics in determining the probability and rate of progression to clinical disease was estimated by fitting Cox proportional hazards models. A multi-state Markov model of the natural history of visceral leishmaniasis was fitted to the data to estimate the asymptomatic infection period and the proportion of asymptomatic individuals going on to develop clinical symptoms. Results At the time of the study, individuals were taking several months to be diagnosed with visceral leishmaniasis, leading to many opportunities for ongoing transmission. The probability of progression to clinical disease was strongly associated with initial seropositivity and even more strongly with seroconversion, with most clinical symptoms developing within a year. The estimated average durations of asymptomatic infection and symptomatic infection for our model of the natural history are 147 days (95 % CI 130–166) and 140 days (95 % CI 123–160), respectively, and are significantly longer than previously reported estimates. We estimate from the data that 14.7 % (95 % CI 12.6-20.0 %) of asymptomatic individuals develop clinical symptoms—a greater proportion than previously estimated. Conclusions Extended periods of asymptomatic infection could be important for visceral leishmaniasis transmission, but this depends critically on the relative infectivity of asymptomatic and symptomatic individuals to sandflies. These estimates could be informed by similar analysis of other datasets. Our results highlight the importance of reducing times from onset of symptoms to diagnosis and treatment to reduce opportunities for transmission. Electronic supplementary material The online version of this article (doi:10.1186/s13071-015-1136-3) contains supplementary material, which is available to authorized users.

Visceral leishmaniasis (VL), caused by Leishmania infantum, is an important disease of domestic dogs. Here, we present data on the IgG subclass antibody response to crude L. infantum antigen in a cohort of naturally infected Brazilian dogs. Specific IgG1-IgG4 responses could be detected in 98, 58, 70 and 82%, respectively of 57 dogs that were seropositive for specific IgG. Levels of all IgG subclasses were strongly inter-correlated. Levels of all IgG subclasses increased at the time of seroconversion, before reaching a plateau after several months. Levels of all IgG subclasses were higher in sick dogs than healthy dogs, and levels of all except IgG2 were higher in parasite-positive (by PCR) than parasite-negative dogs. However, levels of IgG2 relative to IgG1 were lower in sick or parasite-positive dogs compared to healthy or parasite-negative infected dogs. In contrast to previous studies, the results suggest that canine VL is associated with upregulation of specific antibody of all IgG subclasses, particularly IgG1, IgG3 and IgG4.

The elimination of seropositive dogs in Brazil has been used to control zoonotic visceral leishmaniasis but with little success. To elucidate the reasons for this, the infectiousness of 50 sentinel dogs exposed to natural Leishmania chagasi infection was assessed through time by xenodiagnosis with the sandfly vector, Lutzomyia longipalpis. Eighteen (43%) of 42 infected dogs became infectious after a median of 333 days in the field (105 days after seroconversion). Seven highly infectious dogs (17%) accounted for >80% of sandfly infections. There were positive correlations between infectiousness and anti-Leishmania immunoglobulin G, parasite detection by polymerase chain reaction, and clinical disease (logistic regression, r2=0.08-0.18). The sensitivity of enzyme-linked immunosorbent assay to detect currently infectious dogs was high (96%) but lower in the latent period (

The Mycobacterium genus comprises over one-hundred-and-fifty recognised species, the majority of which reside in the environment and many of which can be pathogenic to mammals. Some species of environmental mycobacteria may interfere with BCG vaccination efficacy and in tuberculin test interpretation. Examining biogeographic trends in the distribution of members of the mycobacteria across a number of physicochemical and spatial gradients in soil and water environments across Ethiopia using oligotyping identified differential distributions of pathogenic and significant species. The tuberculosis complex was identified in more than 90% of water samples and taxonomic groups implicated in lower BCG vaccine efficiency were core in both soil and water Mycobacterium communities. A reservoir of Mycobacterium bovis was identified in water, with up to 7.3×102 genome equivalents per ml. Elevation, temperature, habitat and vegetation type were important predictors of both soil and water Mycobacterium communities. These results represent the first step in understanding the potential risk of exposure to environmental mycobacteria that may undermine efforts to reduce disease incidence.\ud \ud

Evaluating the risk of disease spill-over from domestic dogs to wildlife depends on knowledge of inter-specific contact rates and/or exposure to aetiological agents in dog environments. Here, contact rates of crab-eating foxes (Cerdocyon thous) with sympatric domestic dog populations were measured over 25months in Amazon Brazil. Foxes and dogs were serologically and clinically monitored for exposure to canine parvovirus (CPV-2) and canine distemper virus (CDV), pathogens known to have caused wildlife population declines elsewhere. Twenty-two of 24 (92%) tagged foxes visited one or more houses in a median 2 (range 1-3) villages per night where dog densities ranged from 7.2 to 15.4 per km(2) (mean 9.5 per km(2)). Foxes spent an average 6.4% (0-40.3%) of their 10h nocturnal activity period in villages, the equivalent of 38m (range 0-242) per night. The rate of potential exposure to disease agents was thus high, though varied by 3 orders of magnitude for individual foxes. Overall, 46% of the fox population was responsible for 80% of all contacts. None of the 37 monitored foxes however showed serological or clinical evidence of infection with CPV-2 or CDV. Seroprevalences for CPV-2 and CDV antibodies in the local domestic dog population were 13% (3/23) and 9% (2/23), respectively, and 89% of 97 monitored pups born during the study presented clinical signs consistent with active CPV-2 infection (haemorrhagic diarrhoea, vomiting, rapid morbidity and emaciation). Although there was no evidence for infection with either virus in foxes, the high level of contact of foxes with peridomestic habitats suggests that the probability of potential spill-over infections from dogs to foxes is high.

Mycobacterium bovis is the causative agent of bovine tuberculosis (bTB) in cattle and wildlife. Direct aerosol contact is thought to be the primary route of infection between conspecifics, whereas indirect transmission via an environmental reservoir of M. bovis is generally perceived not to be a significant source for infection. Here, we report on the application of molecular technology (PCR) to quantify the prevalence of M. bovis in the environment and to explore its epidemiological significance. We show that the detectability of viable M. bovis at badger setts and latrines is strongly linked to the frequency of M. bovis excretion by infected badgers, and that putative M. bovis in the environment is prevalent on a large proportion of endemic cattle farms in Britain. These results raise important questions about the role of an environmental reservoir in bTB persistence.

Background There is a need for sensitive and specific rapid diagnostic tests (RDT) for canine visceral leishmaniasis. The aims of this study were to evaluate the diagnostic performance of immunochromatographic dipstick RDTs using rK39 antigen for canine visceral leishmaniasis by (i) investigating the sensitivity of RDTs to detect infection, disease and infectiousness in a longitudinal cohort study of natural infection in Brazil, and (ii) using meta-analysis to estimate the sensitivity and specificity of RDTs from published studies. Methodology We used a rK39 RDT (Kalazar Detect Canine Rapid Test; Inbios) to test sera collected from 54 sentinel dogs exposed to natural infection in an endemic area of Brazil. Dogs were sampled bimonthly for up to 27 months, and rK39 results compared to those of crude antigen ELISA, PCR, clinical status and infectiousness to sandflies. We then searched MEDLINE and Web of Knowledge (1993–2011) for original studies evaluating the performance of rK39 RDTs in dogs. Meta-analysis of sensitivity and specificity was performed using bivariate mixed effects models. Principal Findings The sensitivity of the rK39 RDT in Brazil to detect infection, disease and infectiousness was 46%, 77% and 78% respectively. Sensitivity increased with time since infection, antibody titre, parasite load, clinical score and infectiousness. Sixteen studies met the inclusion criteria for meta-analysis. The combined sensitivity of rK39 RDTs was 86.7% (95% CI: 76.9–92.8%) to detect clinical disease and 59.3% (37.9–77.6%) to detect infection. Combined specificity was 98.7% (89.5–99.9%). Both sensitivity and specificity varied considerably between studies. Conclusion The diagnostic performance of rK39 RDTs is reasonable for confirmation of infection in suspected clinical cases, but the sensitivity to detect infected dogs is too low for large-scale epidemiological studies and operational control programmes., Author Summary Canine visceral leishmaniasis is a vector-borne disease caused by the intracellular parasite Leishmania infantum. It is an important veterinary disease, and dogs are also the main animal reservoir for human infection. The disease is widespread in the Mediterranean area, and parts of Asia and South and Central America, and is potentially fatal in both dogs and humans unless treated. Diagnosis of canine infections requires serological or molecular tests. Detection of infection in dogs is important prior to treatment, and in epidemiological studies and control programmes, and a sensitive and specific rapid diagnostic test would be very useful. Rapid diagnostic tests (RDTs) have been developed, but their diagnostic performance has been reported to be variable. We evaluated the sensitivity of a RDT based on serological detection of the rK39 antigen in a cohort of naturally infected Brazilian dogs. The sensitivity of the test to detect infection was relatively low, but increased with time since infection and the severity of infection. We then carried out a meta-analysis of published studies of rK39 RDTs, evaluating the sensitivity to detect disease and infection. The results suggest that rK39 RDTs may be useful in a veterinary clinical setting, but the sensitivity to detect infection is too low for operational control programmes.

SUMMARYZoonotic visceral leishmaniasis (ZVL) caused byLeishmania infantumis an important disease of humans and dogs. Here we review aspects of the transmission and control of ZVL. Whilst there is clear evidence that ZVL is maintained by sandfly transmission, transmission may also occur by non-sandfly routes, such as congenital and sexual transmission. Dogs are the only confirmed primary reservoir of infection. Meta-analysis of dog studies confirms that infectiousness is higher in symptomatic infection; infectiousness is also higher in European than South American studies. A high prevalence of infection has been reported from an increasing number of domestic and wild mammals; updated host ranges are provided. The crab-eating foxCerdocyon thous, opossumsDidelphisspp., domestic catFelis cattus, black ratRattus rattusand humans can infect sandflies, but confirmation of these hosts as primary or secondary reservoirs requires further xenodiagnosis studies at the population level. Thus the putative sylvatic reservoir(s) of ZVL remains unknown. Review of intervention studies examining the effectiveness of current control methods highlights the lack of randomized controlled trials of both dog culling and residual insecticide spraying. Topical insecticides (deltamethrin-impregnated collars and pour-ons) have been shown to provide a high level of individual protection to treated dogs, but further community-level studies are needed.

To develop long-lasting, topical pour-on insecticides for dogs to control zoonotic visceral leishmaniasis, two deltamethrin-based formulations (emulsifiable concentrate [EC] and suspension concentrate [SC]) were tested for their efficacy against the phlebotomine sandfly Lutzomyia longipalpis Lutz & Neiva (Diptera: Psychodidae), vector of Leishmania infantum Nicolle (Kinetoplastida: Trypanosomatidae). The entomological outcomes tested were anti-feeding effect (proportion of female sandflies unfed), lethal effect (24-h female sandfly mortality) and these two effects combined, and the insecticide persistence time at 50% (residual activity, RA(50)) and 80% (RA(80)) efficacy. On initial application, the proportions of female flies that demonstrated anti-feeding activity or were killed were similar for both formulations, at 0.93 (95% confidence interval [CI] 0.856-0.977) vs. 0.81 (95% CI 0.763-0.858) (anti-feeding) and 0.86 (95% CI 0.787-0.920) vs. 0.76 (95% CI 0.698-0.817) (24-h mortality) for EC and SC, respectively. The RA50 rates for anti-feeding and mortality caused by the EC formulation were 4.7 months (95% CI 4.18-5.84) and 2.5 months (95% CI 2.25-2.90), respectively, compared with 1.1 months (95% CI 0.96-1.15) and 0.6 months (95% CI 0.50-0.61), respectively, for the SC formulation. The RA50 for the combined anti-feeding and mortality effects of EC was 5.2 months (95% CI 4.73-5.96), compared with only 0.9 months (95% CI 0.85-1.00) for the SC formulation. The four-to six-fold superior residual activity of the EC formulation was attributed to the addition of a solvent-soluble resin in the formulation which improved fur adhesion and acted as a reservoir for the slow release of the active ingredient. These results identify the potential of such a low-cost formulation to reduce the inter-intervention interval to 5-6 months, similar to that recommended for deltamethrin-impregnated dog collars or for re-impregnation of conventional bednets, both of which are currently used to combat Leishmania transmission. Finally, a novel bioassay was developed in which sandflies were exposed to fur from treated dogs, revealing no detectable tolerance (24-h mortality) in wild- caught sandflies to the insecticide formulations up to 8 months after the initiation of communitywide application of the insecticides to dogs.

To determine whether the incidence of canine leishmaniasis has increased on Crete, Greece, we fitted infection models to serodiagnostic records of 8,848 dog samples for 1990–2006. Models predicted that seroprevalence has increased 2.4% (95% confidence interval 1.61%–3.51%) per year and that incidence has increased 2.2- to 3.8-fold over this 17-year period.

In response to the increasing need for field trials of experimental DNA vaccines against zoonotic visceral leishmaniasis in dogs, our aim was to validate the use of ELISA protocols which will be suitable for detection of natural infection in vaccinated dogs. We have previously demonstrated that DNA/modified vaccinia virus Ankara (MVA) vaccine expressing tryparedoxin peroxidase (TRYP) induced high titres of TRYP antigen-specific IgG in immunized dogs. Here we report our findings that seroconversion to an unrelated diagnostic antigen rK39 did not occur in vaccinated dogs, and that responses to crude Leishmania infantum promastigote antigen (CLA) were weak and short-lived. This is in contrast to strong responses to both antigens shown in naturally infected dogs. To select an appropriate serological test for measurement of infection incidence, we also tested longitudinal samples from an immunologically well-characterized cohort of naturally infected dogs. The sensitivity of CLA ELISA was superior to that of rK39 in early stage infection (from 2 months before, to 2 months after the first detection of infection by PCR or parasitological culture), and more sensitive than rK39 in cross-sectional sampling (81.0% vs 61.9%). We conclude that CLA ELISA will provide sensitive estimates of L. infantum infection incidence in DNA/MVA vaccinated dogs, though optimal testing would include rK39, or a similar recombinant antigen, to improve overall specificity.

The entomological efficacy of using 25% deltamethrin EC insecticide-treated bednets (ITNs) was evaluated against the sandfly Lutzomyia longipalpis Lutz and Neiva (Diptera: Psychodidae), the principal vector of zoonotic visceral leishmaniasis (ZVL) in Latin America. A crossover field study in Amazon Brazil (Marajo Island, Para State) demonstrated that, compared with untreated nets, the insecticide increased the barrier effect of nets by 39% (95% confidence interval [CI] 34-44%), reduced human landing rates by 80% (95% CI 62-90%) and increased the 24-h mortality rate from 0% to 98% (95% CI 93-99%) inside ITNs. The presence of an ITN also reduced the human landing rate on unprotected persons outside the net in the same room by 56% (95% CI 52-59%), and increased 24-h mortality to 68% (95% CI 62-73%) compared to 0.4% (0.1-2.0%) in untreated houses. The reduction in human landing rates in ITN rooms was associated with a doubling in the proportion of sandflies alighting on walls compared with that in untreated rooms, which was attributed to insecticide-induced excito-repellency. There was no evidence that sandflies were diverted onto unprotected hosts. Human landing catches inside houses peaked between 19.00 hours and 23.00 hours and declined steadily to zero at 02.00 hours and thereafter. House-to-house questionnaires established that only 34% of households owned at least one net (median two, range 1-8), only 20% of the population slept under a net (33% of 0-5-year-old children), and the majority (73%) of the population slept in hammocks. Combined data pertaining to sleeping times for children and sandfly activity period indicate that > 50% of sandfly bites inside houses, and substantially more outside houses, were taken before a third of children were potentially protected by a net. This study demonstrates the clear entomological efficacy of ITNs against Lu. longipalpis in this endemic region. The effectiveness of ITNs at preventing ZVL infection and disease has still to be evaluated.

Several studies have suggested that the PCR could be used in epidemiological mass-screening surveys to detect Leishmania ( Viannia ) spp. infection in human and animal hosts. Dogs from an area of Leishmania braziliensis and Leishmania peruviana endemicity were screened for American cutaneous leishmaniasis (ACL) infection by established PCR-based and enzyme-linked immunosorbent antibody test (ELISA) protocols. PCR detected Leishmania ( Viannia ) infection in a total of 90 of 1,066 (8.4%) dogs: 32 of 368 (8.7%), 65 of 769 (8.5%), and 7 of 42 (16.7%) dogs were PCR positive by testing of whole blood, buffy coat, and bone marrow aspirates, respectively. ELISA detected infection in 221 of 1,059 (20.9%) tested dogs. The high prevalence of Leishmania ( Viannia ) detected by PCR and ELISA in both asymptomatic (7.5 and 19.2%, respectively) and symptomatic (32 and 62.5%, respectively) dogs is further circumstantial evidence for their suspected role as reservoir hosts of ACL. However, the low sensitivity of PCR (31%) compared to ELISA (81%) indicates that PCR cannot be used for mass screening of samples in ACL epidemiological studies. Unless more-sensitive PCR protocols were to be developed, its use should be restricted to the diagnosis of active (canine and human) cases and to the parasitological monitoring of patients after chemotherapy.

The epidemiological role of the crab-eating fox Cerdocyon thous in the transmission of Leishmania infantum is assessed in a longitudinal study in Amazon Brazil. A total of 37 wild-caught foxes were immunologically and clinically monitored, and 26 foxes exposed to laboratory colonies of the sandfly vector Lutzomyia longipalpis, over a 15-month period. In total 78% (29/37) of foxes were seropositive for anti-Leishmania IgG on at least 1 occasion, and 38% (8/37) had infections confirmed by PCR and/or by culture. Point prevalences were 74% (serology), 15% (PCR), and 26% (culture). No signs of progressive disease were observed. None of the foxes were infectious to the 1469 sandflies dissected from 44 feeds. A conservative estimate of the possible contribution of foxes to transmission was 9% compared to 91% by sympatric domestic dogs. These results show that crab-eating fox populations do not maintain a transmission cycle independently of domestic dogs. The implication is that they are unlikely to introduce the parasite into Leishmania-free dog populations.

University of Leeds. School of Biology. Leeds, UK. University of Liverpool. Veterinary Clinical Sciences. Mammalian Immunogenetics Research Group. Liverpool, UK / Unversity of Manchester. Centre for Integrated Genomic Medical Research. Manchester, UK. University of Liverpool. Veterinary Clinical Sciences. Mammalian Immunogenetics Research Group. Liverpool, UK. University of Warwick. Department of Biological Sciences. Ecology and Epidemiology. Coventry, UK. World Health Organization. Communicable Diseases. Geneva, Switzerland. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Bel?m, PA, Brasil University of Leeds. School of Biology. Leeds, UK. University of Liverpool. Veterinary Clinical Sciences. Mammalian Immunogenetics Research Group. Liverpool, UK. University of Liverpool. Veterinary Clinical Sciences. Mammalian Immunogenetics Research Group. Liverpool, UK. University of Liverpool. Veterinary Clinical Sciences. Mammalian Immunogenetics Research Group. Liverpool, UK. Zoonotic visceral leishmaniasis (VL) is a disease of dogs, humans and other animals caused by the intracellular macrophage parasite Leishmania infantum. We examined the relationship between DLA class II alleles (DRB1, DQA1, DQB1) and the course of infection in a cohort of Brazilian mongrel dogs exposed to natural L. infantum infection. DLA alleles were typed by sequence-based typing. DLA-DRB1 genotype was significantly associated with levels of anti-Leishmania IgG and parasite status assessed by PCR. Dogs with DLADRB1*01502 had higher levels of specific IgG and an increased risk of being parasite positive compared with dogs without this allele, controlling for other alleles and significant variables. No significant associations were seen for DLA-DQA1 or DLA-DQB1 alleles. These results suggest that the DLA-DRB1 locus plays a role in determining susceptibility to canine VL. As the domestic dog is the main reservoir for human infection, the identification of genetic factors influencing canine resistance or susceptibility to VL may provide insights into the immunology and potential control through vaccination of VL.

University of Leeds. School of Biology. Leed, UK / London School of Hygiene and Tropical Medicine. Department of Infectious and Tropical Diseases. London. London School of Hygiene and Tropical Medicine. Department of Infectious and Tropical Diseases. London. University of Leeds. School of Biology. Leed, UK. Minist?rio da Sa?de. Funda??o Nacional de Sa?de. Instituto Evandro Chagas. Bel?m, PA, Brasil. Universty of Cambridge. Molteno Isnstitute of Parasithology. Department of Pathology. Cambridge. Minist?rio da Sa?de. Funda??o Nacional de Sa?de. Instituto Evandro Chagas. Bel?m, PA, Brasil. Minist?rio da Sa?de. Funda??o Nacional de Sa?de. Instituto Evandro Chagas. Bel?m, PA, Brasil / Universidade de S?o Paulo. Instituto de Ci?ncias Biom?dicas. S?o Paulo, SP, Brasil. University of Leeds. School of Biology. Leed, UK. London School of Hygiene and Tropical Medicine. Department of Infectious and Tropical Diseases. London. The sensitivity and specificity of PCR, serology (ELISA) and lymphoproliferative response to Leishmania antigen for the detection of Leishmania infantum infection were evaluated in a cohort of 126 dogs exposed to natural infection in Brazil. For PCR, Leishmania DNA from bone-marrow was amplified with both minicircle and ribosomal primers. The infection status and time of infection of each dog were estimated from longitudinal data. The sensitivity of PCR in parasite-positive samples was 98%. However, the overall sensitivity of PCR in post-infection samples, from dogs with confirmed infection, was only 68%. The sensitivity of PCR varied during the course of infection, being highest (78?88%) 0?135 days postinfection and declining to around 50% after 300 days. The sensitivity of PCR also varied between dogs, and was highest in sick dogs. The sensitivity of serology was similar in parasite-positive (84%), PCR-positive (86%) and post-infection (88%) samples. The sensitivity of serology varied during the course of infection, being lowest at the time of infection and high (93?100%) thereafter. Problems in determining the specificity of serology are discussed. The sensitivity and specificity of cellular responsiveness were low. These data suggest that PCR is most useful in detecting active or symptomatic infection, and that serology can be a more sensitive technique for the detection of all infected dogs.

University of Leeds. School of Biology. Leeds, United Kingdom. London School of Hygiene and Tropical Medicine. Department of Infectious and Tropical Diseases. London. UK University of Leeds. School of Biology. Leeds, United Kingdom. University of Bristol. Department of Pathologyand Microbiology. Bristol, United Kingdom. Minist?rio da Sa?de. Funda??o Nacional de Sa?de. Instituto Evandro Chagas. Bel?m, PA, Brasil. World Health Organization. Prevention and Eradication. Communicable Diseases Control. Geneva, Switzerland. London School of Hygiene and Tropical Medicine. Department of Infectious and Tropical Diseases. London. UK. To elucidate the local tissue cytokine response of dogs infected withLeishmania chagasi,cytokine mRNA levels were measured in bone marrow aspirates from 27 naturally infecteddogs from Brazil and were compared with those from 5 uninfected control animals. Interferon-gmRNA accumulation was enhanced in infected dogs and was positively correlated withhumoral (IgG1) but not with lymphoproliferative responses toLeishmaniaantigen in infecteddogs. Increased accumulation of mRNA for interleukin (IL)?4, IL-10, and IL-18 was notobserved in infected dogs, and mRNA for these cytokines did not correlate with antibody orproliferative responses. However, infected dogs with detectable IL-4 mRNA had significantlymore severe symptoms. IL-13 mRNA was not detectable in either control or infected dogs.These data suggest that clinical symptoms are not due to a deficiency in interferon-gpro-duction. However, in contrast to its role in human visceral leishmaniasis, IL-10 may not playa key immunosuppressive role in dogs.

SummaryThe paper describes a longitudinal field study of canine leishmaniasis in sympatric domestic dog and crab-eating fox (Cerdocyon thous) populations. Dogs were studied in house-to-house surveys, and foxes were studied by live-trapping and radio-telemetry. Because serological tests (IFAT in this case) for leishmaniasis are often of uncertain sensitivity and specificity, we draw conclusions comparatively. Both cross-sectional (age–prevalence) and longitudinal analyses indicate that incidence in dogs was highest in the dry season. Seasonal changes in the age–prevalence relationship for dogs suggest that serological conversion and recovery rates decline with prior exposure to infection, where ‘recovery’ may be due to loss of a positive antibody titre or death from leishmaniasis. The mean incidence in dogs was higher in the rural than in the urban population and higher in hunting dogs than pet dogs. Cross-sectional and longitudinal analyses of fox serological data suggest that foxes remain positive for longer than dogs on average, either because detectable antibody is more persistent or because they experience a lower mortality rate due to leishmaniasis.

Background Studies performed over the past decade have identified fairly consistent epidemiological patterns of risk factors for visceral leishmaniasis (VL) in the Indian subcontinent. Methods and Principal Findings To inform the current regional VL elimination effort and identify key gaps in knowledge, we performed a systematic review of the literature, with a special emphasis on data regarding the role of cattle because primary risk factor studies have yielded apparently contradictory results. Because humans form the sole infection reservoir, clustering of kala-azar cases is a prominent epidemiological feature, both at the household level and on a larger scale. Subclinical infection also tends to show clustering around kala-azar cases. Within villages, areas become saturated over a period of several years; kala-azar incidence then decreases while neighboring areas see increases. More recently, post kala-azar dermal leishmaniasis (PKDL) cases have followed kala-azar peaks. Mud walls, palpable dampness in houses, and peri-domestic vegetation may increase infection risk through enhanced density and prolonged survival of the sand fly vector. Bed net use, sleeping on a cot and indoor residual spraying are generally associated with decreased risk. Poor micronutrient status increases the risk of progression to kala-azar. The presence of cattle is associated with increased risk in some studies and decreased risk in others, reflecting the complexity of the effect of bovines on sand fly abundance, aggregation, feeding behavior and leishmanial infection rates. Poverty is an overarching theme, interacting with individual risk factors on multiple levels. Conclusions Carefully designed demonstration projects, taking into account the complex web of interconnected risk factors, are needed to provide direct proof of principle for elimination and to identify the most effective maintenance activities to prevent a rapid resurgence when interventions are scaled back. More effective, short-course treatment regimens for PKDL are urgently needed to enable the elimination initiative to succeed., Author Summary Visceral leishmaniasis (VL) is a potentially deadly parasitic disease that affects 200,000 to 300,000 people per year in the Indian subcontinent, where an effort is currently underway to eliminate the disease. Studies have identified fairly consistent patterns of risk factors for VL. This information can help to inform the elimination effort. Because humans form the sole source of infection, clustering of VL cases is a prominent feature, both at the household level and on a larger scale. Mud walls, dampness in houses, and peri-domestic vegetation increase infection risk by promoting the survival of the insect vector. Bed nets, sleeping on a cot and indoor residual spraying decrease risk. The presence of cattle is associated with increased risk in some studies and decreased risk in others, reflecting a complex effect on vector abundance and vector infection rates. Poverty is a major underlying factor increasing risk of VL infection and disease. A well-designed demonstration project is needed to provide direct proof of principle for elimination and to identify the most effective maintenance activities to prevent a rapid return of disease when interventions are scaled back.

Minist?rio da Sa?de. Funda??o Servi?os de Sa?de P?blica. Instituto Evandro Chagas. Bel?m, PA, Brasil. The London School of Hygiene & Tropical Medicine. Department of Medical Parasitology. Keppel Street, London. The London School of Hygiene & Tropical Medicine. Department of Medical Parasitology. Keppel Street, London. University of Oxford. Department of Zoology. Wildlife Conservation Research Unit. Oxford, England. University of Oxford. Department of Zoology. Wildlife Conservation Research Unit. Oxford, England. Minist?rio da Sa?de. Funda??o Servi?os de Sa?de P?blica. Instituto Evandro Chagas. Bel?m, PA, Brasil. Minist?rio da Sa?de. Funda??o Servi?os de Sa?de P?blica. Instituto Evandro Chagas. Bel?m, PA, Brasil.