Effect of indoor residual spraying on sandfly abundance and incidence of visceral leishmaniasis in India, 2016-22: an interrupted time-series analysis and modelling study.

BACKGROUND: Efforts to eliminate visceral leishmaniasis in India mainly consist of early detection and treatment of cases and indoor residual spraying with insecticides to kill the phlebotomine sandfly Phlebotomus argentipes that transmits the causative Leishmania protozoa. In this modelling study, we aimed to estimate the effect of indoor residual spraying (IRS) on vector abundance and transmission of visceral leishmaniasis in India. METHODS: In this time-series analysis and modelling study, we assessed the effect of IRS on vector abundance by using indoor vector-abundance data (from 2016 to 2022) and IRS quality-assurance data (from 2017-20) from 50 villages in eight endemic blocks in India where IRS was implemented programmatically. To assess a potential dose-response relation between insecticide concentrations and changes in sandfly abundance, we examined the correlation between site-level insecticide concentrations and the site-level data for monthly sandfly abundances. We used mathematical modelling to link vector data to visceral leishmaniasis case numbers from the national Kala-Azar Management Information System registry (2013-21), and to predict the effect of IRS on numbers of averted cases and deaths. FINDINGS: IRS was estimated to reduce indoor sandfly abundance by 27% (95% CI 20-34). Concentrations of insecticides on walls were significantly-but weakly-associated with the degree of reduction in vector abundance, with a reduction of -0·0023 (95% CI -0·0040 to -0·0007) sandflies per mg/m2 insecticide (p=0·0057). Reported case numbers of visceral leishmaniasis were well explained by trends in vector abundance. Village-wide IRS in response to a newly detected case of visceral leishmaniasis was predicted to reduce disease incidence by 6-40% depending on the presumed reduction in vector abundance modelled. INTERPRETATION: Indoor residual spraying has substantially reduced sandfly abundance in India, which has contributed to reductions in visceral leishmaniasis and related deaths. To prevent the re-emergence of visceral leishmaniasis as a public health problem, surveillance of transmission and sandfly abundance is warranted. FUNDING: Bill & Melinda Gates Foundation. TRANSLATION: For the Hindi translation of the abstract see Supplementary Materials section.

Toxicity of Trixis vauthieri DC essential oil on Lutzomyia longipalpis (Diptera, Psychodidae, Phlebotominae), a proven vector of visceral leishmaniasis in Brazil.

BACKGROUND OBJECTIVES: In Brazil, one of the visceral leishmaniasis control measures in urban environments is the elimination of Lutzomyia longipalpis, which occurs through the application of pyrethroid insecticides with residual action in homes and outbuildings. Due to the loss of sensitivity of this vector to these insecticides, the search for more efficient insecticide compounds against L. longipalpis has been intensified. The objective of this work was to evaluate the toxicity of Trixis vauthieri essential oil on adult sandflies of the species L. longipalpis, and identify the phytochemical composition of these essential oils. METHODS: Essential oils from leaves collected from T. vauthieri at different times were obtained at concentrations of 5, 10 and 20 mg/mL. Twenty sandflies were exposed to the essential oils and the mortality was evaluated after 1, 2, 4, 16, 24, 48 and 72 h. The chemical constituents of the essential oil were also identified. RESULTS: The essential oils of T. vauthieri at a concentration of 20 mg/mL were the most toxic to sandflies, reaching a mortality rate of 98.33% and 95%, respectively, after 72 h of exposure. The analysis of chemical constituents revealed the presence of triterpenes and/or steroids, tannins, flavonoids, alkaloids, saponins and coumarins. INTERPRETATION CONCLUSION: The results obtained suggest that T. vauthieri essential oil is fairly promising as an insecticidal potential against L. longipalpis. A more detailed analysis of the oil's phytochemical composition is necessary to identify active and pure compounds that can be used in vector control of visceral leishmaniasis.

Chemical control and insecticide resistance status of sand fly vectors worldwide.

BACKGROUND: Phlebotomine sand flies are prominent vectors of Leishmania parasites that cause leishmaniasis, which comes second to malaria in terms of parasitic causative fatalities globally. In the absence of human vaccines, sand fly chemical-based vector control is a key component of leishmaniasis control efforts. METHODS AND FINDINGS: We performed a literature review on the current interventions, primarily, insecticide-based used for sand fly control, as well as the global insecticide resistance (IR) status of the main sand fly vector species. Indoor insecticidal interventions, such as residual spraying and treated bed nets are the most widely deployed, while several alternative control strategies are also used in certain settings and/or are under evaluation. IR has been sporadically detected in sand flies in India and other regions, using non-standardized diagnostic bioassays. Molecular studies are limited to monitoring of known pyrethroid resistance mutations (kdr), which are present at high frequencies in certain regions. CONCLUSIONS: As the leishmaniasis burden remains a major problem at a global scale, evidence-based rational use of insecticidal interventions is required to meet public health demands. Standardized bioassays and molecular markers are a prerequisite for this task, albeit are lagging behind. Experiences from other disease vectors underscore the need for the implementation of appropriate IR management (IRM) programs, in the framework of integrated vector management (IVM). The implementation of alternative strategies seems context- and case-specific, with key eco-epidemiological parameters yet to be investigated. New biotechnology-based control approaches might also come into play in the near future to further reinforce sand fly/leishmaniasis control efforts.

Impact of IRS: Four-years of entomological surveillance of the Indian Visceral Leishmaniases elimination programme.

BACKGROUND: In 2005, Bangladesh, India and Nepal agreed to eliminate visceral leishmaniasis (VL) as a public health problem. The approach to this was through improved case detection and treatment, and controlling transmission by the sand fly vector Phlebotomus argentipes, with indoor residual spraying (IRS) of insecticide. Initially, India applied DDT with stirrup pumps for IRS, however, this did not reduce transmission. After 2015 onwards, the pyrethroid alpha-cypermethrin was applied with compression pumps, and entomological surveillance was initiated in 2016. METHODS: Eight sentinel sites were established in the Indian states of Bihar, Jharkhand and West Bengal. IRS coverage was monitored by household survey, quality of insecticide application was measured by HPLC, presence and abundance of the VL vector was monitored by CDC light traps, insecticide resistance was measured with WHO diagnostic assays and case incidence was determined from the VL case register KAMIS. RESULTS: Complete treatment of houses with IRS increased across all sites from 57% in 2016 to 70% of houses in 2019, rising to >80% if partial house IRS coverage is included (except West Bengal). The quality of insecticide application has improved compared to previous studies, average doses of insecticide on filters papers ranged from 1.52 times the target dose of 25mg/m2 alpha-cypermethrin in 2019 to 1.67 times in 2018. Resistance to DDT has continued to increase, but the vector was not resistant to carbamates, organophosphates or pyrethroids. The annual and seasonal abundance of P. argentipes declined between 2016 to 2019 with an overall infection rate of 0.03%. This was associated with a decline in VL incidence for the blocks represented by the sentinel sites from 1.16 per 10,000 population in 2016 to 0.51 per 10,000 in 2019. CONCLUSION: Through effective case detection and management reducing the infection reservoirs for P. argentipes in the human population combined with IRS keeping P. argentipes abundance and infectivity low has reduced VL transmission. This combination of effective case management and vector control has now brought India within reach of the VL elimination targets.

Insecticide susceptibility of Phlebotomus argentipes sandflies, vectors of visceral leishmaniasis in India.

OBJECTIVES: Indoor residual spraying (IRS) with insecticides is the main vector control intervention for the elimination of visceral leishmaniasis in India. After a change in IRS policy in 2015 due to widespread resistance of Phlebotomus argentipes to DDT, IRS with DDT was replaced with alpha-cypermethrin IRS in 2016. The objective of the present study was to evaluate the susceptibility of P. argentipes to DDT and its alternatives, namely malathion and pirimiphos-methyl (organophosphates); alpha-cypermethrin, deltamethrin, lambda-cyhalothrin and permethrin (pyrethroids), and bendiocarb and propoxur (carbamates), in support of visceral leishmaniasis elimination in India. METHODS: Phlebotomus argentipes sandflies were collected from the visceral-leishmaniasis endemic states of Bihar, Jharkhand and West Bengal. In the WHO tube tests, the phenotypic susceptibility of F1, 2-day old, non-blood fed females were determined against filter papers impregnated with DDT 4%, malathion 5%, pirimiphos-methyl 0.25%, alpha-cypermethrin 0.05%, deltamethrin 0.05%, lambda-cyhalothrin 0.05%, permethrin 0.75%, bendiocarb 0.1% and propoxur 0.1%, which were sourced from Universiti Sains Malaysia. The knockdown of sandflies after 1-h exposure and mortality at 24 h after the 1-h exposure period were scored. RESULTS: Mean mortality of P. argentipes 24 h after exposure in tube tests was 22.6% for DDT and ≥ 98% for other insecticide-impregnated papers tested. CONCLUSION: Phlebotomus argentipes continues to be highly resistant to DDT with no reversal of resistance after DDT's withdrawal from IRS. P. argentipes was fully susceptible to pyrethroid, organophosphate and carbamate insecticides tested. Regular monitoring is warranted for insecticide resistance management in sandfly vectors.

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.

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.

The Potential of Systemic Control of Sand Flies Using Fipronil in the Novel Leishmania major (Kinetoplastida: Trypanosomatidae) Reservoirs Meriones tristrami (Rodentia: Muridea) and Meriones crassus (Rodentia: Muridea).

Leishmania major (Yakimoff & Schokhor, 1914), an important causative agent of Old World Cutaneous Leishmaniasis (CL), is transmitted by sand flies among a limited number of gerbilline reservoir-species. We can take advantage of this strong dependency to break the pathogen transmission cycle by using systemic insecticides that render the host toxic to the blood-feeding vector. We evaluated the potential of this approach with two novel reservoir species, incriminated for CL expansion in several sites in the Middle East. Specifically, we evaluated: 1) the residuality of the systemic insecticide fipronil in Meriones tristrami (Thomas, 1892) fed on fipronil-treated baits and 2) the treatments' adulticide effect on sand flies that blood fed on treated and untreated M. tristrami and M. crassus (Sundevall, 1842). We fed M. tristrami with food pellets containing 0.1 g/kg fipronil and used gas chromatograph-mass spectrometery analysis and bioassays to examine its residual toxicity to blood-feeding female sand flies. In M. tristrami, fipronil was rapidly metabolized to fipronil sulfone, found in the blood, urine, and feces for ≥31 d after fipronil admission. The survival of sand flies that blood fed on fipronil-treated M. tristrami and M. crassus was significantly reduced for at least 15 and 9 d respectively, after fipronil admission. These results hold promise for the potential contribution of systemic control approaches to CL integrated management strategies against novel CL (due to L. major) outbreaks in Israel and elsewhere.

Fluralaner (Bravecto®) induces long-term mortality of Lutzomyia longipalpis after a blood meal in treated dogs.

BACKGROUND: Leishmania infantum is the etiological agent of visceral leishmaniasis (VL) in the New World, where the sand fly Lutzomyia longipalpis and domestic dogs are considered the main vector and host reservoirs, respectively. Systemic insecticides have been studied as an alternative to control vector-borne diseases, including VL. Fluralaner, an isoxazoline class compound, is a systemic insecticide used in dogs, with proven efficiency against different species of phlebotomine sand flies. However, to date no studies have demonstrated the efficacy of fluralaner on Lu. longipalpis. The aim of this study was to evaluate the insecticidal effect of fluralaner (Bravecto®) on the sand fly Lu. longipalpis after blood meal in treated dogs. METHODS: Healthy mongrel dogs (n = 8) were recruited from the Zoonoses Control Center in the city of Natal, Rio Grande do Norte, Brazil, and randomized into two groups: fluralaner treated (n = 4) and non-treated control (n = 4). Colony-reared female specimens of Lu. longipalpis (n = 20) were allowed to feed on all dogs for 40 min before treatment (for fluralaner-treated dogs), at day 1 after treatment and then monthly until 1 year post-treatment. RESULTS: In the treatment group, there was 100% mortality of Lu. longipalpis for up to 5 months after treatment initiation, decreasing to 72.5% at 6 months post-treatment initiation. The efficacy of fluralaner ranged from 100% at day 1 (P = 0.0002) to 68% ( P = 0.0015) at 6 months, decreasing to 1.4% at 1 year post-treatment. Sand fly mortality carried out blood meal in non-treated control dogs remained constant at ≤ 15%. CONCLUSIONS: Taken together, our results suggest that fluralaner may be used as a control strategy for VL in dogs in VL endemic areas.

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

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.

Susceptibility of phlebotomine sandflies (Diptera: Psychodidae) collected in the field, to alpha-cypermethrin in four municipalities endemic to leishmaniasis.

Although the chemical control against leishmaniasis began in 1953 in Brazil, little information is available on how this strategy has affected populations of phlebotomine sandflies in the field. The objective of this study was to analyze the susceptibility profile of four populations of phlebotomine sandflies to the insecticide alpha-cypermethrin. Sandflies collected in field in four Brazilian municipalities were evaluated using CDC bottles in different concentrations of alpha-cypermethrin. A total of 1,186 phlebotomine sandflies were used in the bioassays. The LD50 ranged from 1.48 to 2.57 ug/mL in the field populations. For a dose of 5 ug/mL of alpha-cypermethrin, the LT50 and LT95 ranged from 17.9 to 27.5 minutes, and LT95 from 39.7 to 61.5 minutes, respectively. All the populations studied were highly susceptible to the insecticide alpha-cypermethrin. Routine studies are needed to detect changes in sandflies susceptibility to insecticides.

Galactosamine reduces sandfly gut protease activity through TOR downregulation and increases Lutzomyia susceptibility to Leishmania.

In sandflies, males and females feed on carbohydrates but females must get a blood meal for egg maturation. Using artificial blood meals, this study aimed to understand how galactosamine interferes with sandfly digestive physiology. We also used galactosamine to manipulate the digestive physiology of Lutzomyia longipalpis to investigate its influence on sandfly digestion and Leishmania development within their insect vectors. Galactosamine was capable to reduce Lu. longipalpis trypsinolytic activity in a dose-dependent manner. This effect was specific to galactosamine as other similar sugars were not able to affect sandfly trypsin production. An excess of amino acids supplemented with the blood meal and 15 mM galactosamine was able to abrogate the reduction of the trypsinolytic activity caused by galactosamine, suggesting this phenomenon may be related to an impairment of amino acid detection by sandfly enterocytes. The TOR inhibitor rapamycin reduces trypsin activity in the L. longipalpis midgut. Galactosamine reduces the phosphorylation of the TOR pathway repressor 4EBP, downregulating TOR activity in the gut of L. longipalpis. Galactosamine reduces sandfly oviposition, causes an impact on sandfly longevity and specifically reduces sandfly gut proteases whereas increasing α-glycosidase activity. The administration of 15 and 30 mM galactosamine increased the number of promastigote forms of Le. mexicana and Le. infantum in galactosamine-treated L. longipalpis. Our results showed that galactosamine influences amino acid sensing, reduces sandfly gut protease activity through TOR downregulation, and benefits Leishmania growth within the Lu. longipalpis gut.

Vaccination against canine leishmaniasis in Brazil.

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.

Visceral leishmaniasis on the Indian Subcontinent: Efficacy of fipronil-based cattle treatment in controlling sand fly populations is dependent on specific aspects of sand fly ecology.

BACKGROUND: Visceral leishmaniasis (VL) is a deadly disease transmitted by the sand fly Phlebotomus argentipes on the Indian subcontinent, with a promising means of vector control being orally treating cattle with fipronil-based drugs. While prior research investigating the dynamic relationship between timing of fipronil-based control schemes and the seasonality of sand flies provides insights into potential of treatment on a large scale, ecological uncertainties remain. We investigated how uncertainties associated with sand fly ecology might affect our ability to assess efficacy of fipronil-based control schemes. To do this, we used a previously-described, individual-based, stochastic sand fly model to quantify how uncertainties associated with 1) the percentage of female sand flies taking blood meals from cattle, and 2) the percentage of female sand flies ovipositing in organic matter containing feces from treated cattle might impact the efficacy of fipronil-based sand fly control schemes. PRINCIPAL FINDINGS: Assuming no prior knowledge of sand fly blood meal and oviposition sites, the probabilities of achieving effective sand fly population reduction with treatments performed 3, 6 and 12 times per year were ≈5-22%, ≈27-36%, and ≈46-54%, respectively. Assuming ≥50% of sand flies feed on cattle, probabilities of achieving efficacious control increased to ≈8-31%, ≈15-42%, and ≈52-65%. Assuming also that ≥50% of sand flies oviposit in cattle feces, the above probabilities increased further to ≈14-53%, ≈31-81%, and ≈89-97%. CONCLUSIONS: Our assessments of the efficacy of fipronil-based cattle treatments in controlling sand fly populations depend on our assumptions regarding key aspects of sand fly ecology. Assessments are most sensitive to assumptions concerning the percentage of sand flies ovipositing in feces of treated cattle, thus emphasizing the importance of identifying sand fly oviposition sites. Our results place the evaluation of fipronil-based cattle treatment within a broader ecological context, which could aid in the planning and execution of a largescale field trial.

Susceptibility of wild-caught Lutzomyia longipalpis (Diptera: Psychodidae) sand flies to insecticide after an extended period of exposure in western São Paulo, Brazil.

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.

Attraction of Lutzomyia longipalpis to synthetic sex-aggregation pheromone: Effect of release rate and proximity of adjacent pheromone sources.

In South America, the Protist parasite that causes visceral leishmaniasis, a potentially fatal human disease, is transmitted by blood-feeding female Lutzomyia longipalpis sand flies. A synthetic copy of the male produced sex-aggregation pheromone offers new opportunities for vector control applications. We have previously shown that the pheromone placed in plastic sachets (lures) can attract both females and males to insecticide treated sites for up to 3 months. To use the pheromone lure in a control program we need to understand how the application of lures in the field can be optimised. In this study we investigated the effect of increasing the number of lures and their proximity to each other on their ability to attract Lu. longipalpis. Also for the first time we applied a Bayesian log-linear model rather than a classic simple (deterministic) log-linear model to fully exploit the field-collected data. We found that sand fly response to pheromone is significantly related to the quantity of pheromone and is not influenced by the proximity of other pheromone sources. Thus sand flies are attracted to the pheromone source at a non-linear rate determined by the amount of pheromone being released. This rate is independent of the proximity of other pheromone releasing traps and indicates the role of the pheromone in aggregation formation. These results have important implications for optimisation of the pheromone as a vector control tool and indicate that multiple lures placed in relatively close proximity to each other (5 m apart) are unlikely to interfere with one another.

Indoor residual spraying for kala-azar vector control in Bangladesh: A continuing challenge.

BACKGROUND: Visceral leishmaniasis (VL) in the Indian subcontinent is a fatal disease if left untreated. Between 1994 to 2013, the Ministry of Health of Bangladesh reported 1,09,266 cases of VL and 329 VL related deaths in 37 endemic districts. Indoor residual spraying (IRS) using dichlorodiphenyltrichloroethane (DDT) was used by the national programme in the 1960s to control malaria. Despite findings of research trials demonstrating that the synthetic pyrethroid deltamethrin 5 WP was very effective at reducing vector densities, no national VL vector control operations took place in Bangladesh between 1999 to early 2012. In 2012, IRS using deltamethrin 5 WP was re-introduced by the national programme, which consisted of pre-monsoon spraying in eight highly endemic sub-districts (upazilas). The present study aims to evaluate the effectiveness of IRS on VL vectors, as well as the process and performance of the spraying activities by national programme staff. METHODS: Five highly endemic upazilas of Mymensingh district were purposively selected (Fulbaria, Trishal, Mukthagacha, Gaforgaon and Bhaluka) to conduct the present study using the WHO/TDR monitoring and evaluation tool kit. IRS operations, conducted by 136 squads/teams, and 544 spraymen, were observed using check lists and questionnaires included in the WHO/TDR monitoring and evaluation tool kit. A household (HH) acceptability survey of IRS was conducted in all study areas using a structured questionnaire in 600 HHs. To measure the efficacy of IRS, pre-IRS (two weeks prior) and post-IRS (at one and five months after), vector density was measured using CDC light traps for two consecutive nights. Bioassays, using the WHO cone-method, were carried out in 80 HHs (40 sprayed and 40 unsprayed) to measure the effectiveness of the insecticide on sprayed surfaces. RESULTS: Of the 544 spraymen interviewed pre-IRS, 60%, 3% and 37% had received training for one, two and three days respectively. During spraying activities, 64% of the spraying squads had a supervisor in 4 upazilas but only one upazila (Mukthagacha) achieved 100% supervision of squads. Overall, 72.8% of the spraying squads in the study upazilas had informed HHs members to prepare their houses prior to spraying. The required personal protective equipment was not provided by the national programme during our observations and the spraying techniques used by all sprayers were sub-standard compared to the standard procedure mentioned in the M&E toolkit. In the HH interviews, 94.8% of the 600 respondents said that all their living rooms and cattle sheds had been sprayed. Regarding the effectiveness measurements (i.e. reduction of vector densities), a total of 4132 sand flies were trapped in three intervals, of which 3310 (80.1%) were P. argentipes; 46.5% (1540) males and 53.5% (1770) females. At one month post-IRS, P. argentipes densities were reduced by 22.5% but the 5 months post-IRS reduction was only 6.4% for both male and female. The bioassay tests showed a mean corrected mortality of P. argentipes sand flies at one month post-IRS of 87.3% which dropped to 74.5% at 4 months post-IRS in three upazilas, which is below the WHO threshold level (80%). CONCLUSION: The national programme should conduct monitoring and evaluation activities to ensure high quality of IRS operations as a pre-condition for achieving a fast and sustained reduction in vector densities. This will continue to be important during the maintenance phase of VL elimination on the Indian subcontinent. Further research is needed to determine other suitable vector control option(s) when the case numbers are very low.

Laboratory assessment of the anti-feeding effect for up to 12 months of a slow release deltamethrin collar (Scalibor®) against the sand fly Phlebotomus perniciosus in dogs.

BACKGROUND: Leishmaniosis/leishmaniasis consists of a wide group of diseases, caused by different Leishmania species and having different hosts. Leishmaniosis caused by Leishmania infantum, a disease primarily of dogs and humans, occurs after susceptible hosts are exposed to the feeding behavior of infected sand flies. A one-year laboratory study in dogs was designed to determine the 364-day anti-feeding efficacy of a slow release deltamethrin collar against the sand fly P. perniciosus, a common host of L. infantum in the Mediterranean basin. METHODS: In this assessor-blinded study, 16 Beagle dogs were randomized into two groups using P. perniciosus engorgement rates from a Day -7 challenge. On Day 0, dogs in Group 1 received a placebo collar, while dogs in Group 2 received a deltamethrin collar (Scalibor® Protector Band). All dogs were caged, sedated and then exposed for 1 h to 85 (± 10) female and 15 (± 5) male P. perniciosus on Day 7 and every 28 days through Day 364. All flies, alive and dead, were aspirated from cages and from dogs, immediately counted and then frozen for assessment of blood engorgement. Anti-feeding efficacy was determined by comparing the arithmetic means of engorged female flies (alive, dead and moribund) in the deltamethrin group to the control group means. Insecticidal efficacy at the time flies were retrieved was assessed by comparisons between groups of mean live female fly counts. RESULTS: In the deltamethrin group, relative to the control group, there was a significant reduction in arithmetic mean numbers of engorged P. perniciosus of 94-98% from Day 7 through Day 364. On Day 28, in the treated group relative to the control group, there was a 74% reduction in mean live fly counts, with between-group differences significant from Days 7 through 196, although insecticidal activity remained less than 50% from Day 56. CONCLUSION: Deltamethrin collar application to dogs reduced sand fly feeding by ≥ 94%, relative to unprotected control dogs, for 364 days. Thus, one collar applied to a dog can prevent or reduce the risk of sand fly transmission of Leishmania for one full year.

Experimental hut to study the indoor behaviour and effects of insecticide-treated bednets on phlebotomine sand flies (Diptera: Psychodidae).

INTRODUCTION: Behavioural effects of insecticides on endophagic phlebotomine sand fly vectors of Leishmania are poorly understood mainly because of the lack of an experimental hut (EH) in which to study them. OBJECTIVE: To build an EH to evaluate the effects of long-lasting insecticide-treated nets (LLINs) on Lutzomyia longiflocosa. METHODS: The study had two phases: (1) Laboratory experiments using tunnel tests to select the traps for the EH; and (2) EH construction and evaluation of the effects of deltamethrin and lambda-cyhalothrin LLINs on L. longiflocosa females inside the EH. FINDINGS: Phase 1: The horizontal-slit trap was the best trap. This trap collected the highest percentage of sand flies, and prevented them from escaping. Therefore, this trap was used in the EH. Phase 2: The main effects of LLINs on L. longiflocosa in the EH were: landing inhibition, inhibition from entering the bednet, induced exophily, and high mortality (total and inside exit traps). CONCLUSIONS: The EH was effective for evaluating the effects of LLINs on endophagic sand flies. Although both types of LLINs showed high efficacy, the lambda-cyhalothrin-treated LLIN performed better. This is the first report of induced exophily in sand flies.

Field evaluation of a 0.005% fipronil bait, orally administered to Rhombomys opimus, for control of fleas (Siphonaptera: Pulicidae) and phlebotomine sand flies (Diptera: Psychodidae) in the Central Asian Republic of Kazakhstan.

Plague (Yersinia pestis) and zoonotic cutaneous leishmaniasis (Leishmania major) are two rodent-associated diseases which are vectored by fleas and phlebotomine sand flies, respectively. In Central Asia, the great gerbil (Rhombomys opimus) serves as the primary reservoir for both diseases in most natural foci. The systemic insecticide fipronil has been previously shown to be highly effective in controlling fleas and sand flies. However, the impact of a fipronil-based rodent bait, on flea and sand fly abundance, has never been reported in Central Asia. A field trial was conducted in southeastern Kazakhstan to evaluate the efficacy of a 0.005% fipronil bait, applied to gerbil burrows for oral uptake, in reducing Xenopsylla spp. flea and Phlebotomus spp. sand fly abundance. All active gerbil burrows within the treated area were presented with ~120 g of 0.005% fipronil grain bait twice during late spring/early summer (June 16, June 21). In total, 120 occupied and 14 visited gerbil colonies were surveyed and treated, and the resulting application rate was minimal (~0.006 mg fipronil/m2). The bait resulted in 100% reduction in Xenopsylla spp. flea abundance at 80-days post-treatment. Gravid sand flies were reduced ~72% and 100% during treatment and at week-3 post-treatment, respectively. However, noticeable sand fly reduction did not occur after week-3 and results suggest environmental factors also influenced abundance significantly. In conclusion, fipronil bait, applied in southeastern Kazakhstan, has the potential to reduce or potentially eliminate Xenopsylla spp. fleas if applied at least every 80-days, but may need to be applied at higher frequency to significantly reduce the oviposition rate of Phlebotomus spp. sand flies. Fipronil-based bait may provide a means of controlling blood-feeding vectors, subsequently reducing disease risk, in Central Asia and other affected regions globally.

Impact of insecticide-treated bednets and indoor residual spraying in controlling populations of Phlebotomus duboscqi, the vector of Leishmania major in Central Mali.

BACKGROUND: Cutaneous leishmaniasis (CL) is an endemic neglected tropical disease prevalent in several areas where seasonal malaria transmission is active. We assessed the effect of indoor residual spraying (IRS) and the mass distribution of long-lasting insecticide-treated bednets (LLINs) for malaria control on sand fly population diversity and abundance, and its impact on the risk of Leishmania transmission in the district of Baroueli, endemic for CL in Mali. METHODS: Kemena and Sougoula, two villages in the district of Baroueli, were selected for entomology surveys from March to September 2016 to evaluate sand fly species composition and density, and Leishmania infection rates in the vector Phlebotomus duboscqi. The surveys followed an annual indoor residual spraying and mass distribution of long-lasting insecticide-treated bednets (IRS/LLINs) that began in 2011 for malaria vector control. We also carried out a leishmanin skin test (LST) survey in the two villages to determine the incidence of Leishmania infection in humans living in the endemic area. RESULTS: A total of 2936 sand fly specimens, 1013 males and 1923 females, were collected and identified from the two villages throughout the study period. Fourteen species, 2 belonging to the genus Phlebotomus and 12 to the genus Sergentomyia, were documented. The genus Sergentomyia constituted 91% of collected sand flies versus 9% for the genus Phlebotomus (P. duboscqi and P. rodhaini). Of those, P. duboscqi was the most abundant, representing 99.6% of the collected Phlebotomus species. In both villages, P. duboscqi was most abundant during the end of dry season (June). The prevalence of Leishmania infection in individual females of P. duboscqi by PCR was 3.5%. After 5 years of the IRS/LLINs, the incidence of Leishmania infection in the human population as measured by LST was 4.2%. CONCLUSIONS: Compared to historical data collected from 2005-2008, a considerable reduction was observed in both sand fly density and prevalence of human Leishmania infection in the villages of Kemena and Sougoula, Baroueli District, following IRS/LLINs. This suggests that IRS/LLINs used for mosquito control also impacts sand fly vectors reducing the incidence of leishmaniasis. TRIAL REGISTRATION: NCT00344084 . Registered: 23 June 2006.