Diwakar Singh Dinesh

Vector control by insecticide-treated nets in the fight against visceral leishmaniasis in the Indian subcontinent, what is the evidence?

Visceral leishmaniasis (VL) is a deadly vector‐borne disease that causes an estimated 500 000 new cases a year. In India, Nepal and Bangladesh, VL is caused by Leishmania donovani, which is transmitted from man to man by the sandfly Phlebotomus argentipes. In 2005, these three countries signed a memorandum of understanding to eliminate VL from the region. Integrated vector management is one of the pillars of this elimination strategy, alongside early case detection and treatment. We reviewed the evidence of effectiveness of different vector control methods, to examine the potential role of insecticide treated bednets (ITNs). Indoor residual spraying has shown poor impact for various reasons and resistance to DDT is emerging in Bihar. Environmental management performed poorly compared to insecticide based methods. ITNs could give individual protection but this still needs to be proven in randomized trials. Given the constraints of indoor residual spraying, it is worthwhile to further explore the use of ITNs, in particular long lasting ITNs, as an additional tool in the VL elimination initiative.

Effect of village-wide use of long-lasting insecticidal nets on visceral leishmaniasis vectors in India and Nepal: a cluster randomized trial.

Background Visceral leishmaniasis (VL) control in the Indian subcontinent is currently based on case detection and treatment, and on vector control using indoor residual spraying (IRS). The use of long-lasting insecticidal nets (LN) has been postulated as an alternative or complement to IRS. Here we tested the impact of comprehensive distribution of LN on the density of Phlebotomus argentipes in VL-endemic villages. Methods A cluster-randomized controlled trial with household P. argentipes density as outcome was designed. Twelve clusters from an ongoing LN clinical trial—three intervention and three control clusters in both India and Nepal—were selected on the basis of accessibility and VL incidence. Ten houses per cluster selected on the basis of high pre-intervention P. argentipes density were monitored monthly for 12 months after distribution of LN using CDC light traps (LT) and mouth aspiration methods. Ten cattle sheds per cluster were also monitored by aspiration. Findings A random effect linear regression model showed that the cluster-wide distribution of LNs significantly reduced the P. argentipes density/house by 24.9% (95% CI 1.80%–42.5%) as measured by means of LTs. Interpretation The ongoing clinical trial, designed to measure the impact of LNs on VL incidence, will confirm whether LNs should be adopted as a control strategy in the regional VL elimination programs. The entomological evidence described here provides some evidence that LNs could be usefully deployed as part of the VL control program. Trial registration ClinicalTrials.gov CT-2005-015374

Insecticide susceptibility of Phlebotomus argentipes in visceral leishmaniasis endemic districts in India and Nepal.

Objectives To investigate the DDT and deltamethrin susceptibility of Phlebotomus argentipes, the vector of Leishmania donovani, responsible for visceral leishmaniasis (VL), in two countries (India and Nepal) with different histories of insecticide exposure. Methods Standard WHO testing procedures were applied using 4% DDT and 0.05% deltamethrin impregnated papers. The effect of the physiological status (fed and unfed) of females on the outcome of the bioassays was assessed and the optimal time of exposure for deltamethrin was evaluated on a colony population. Field populations from both countries were tested. Results Fed and unfed females responded in a similar way. For exposure time on field samples 60 min was adopted for both DDT and deltamethrin. In Bihar, knockdown and mortality with DDT was respectively 20 and 43%. In Nepal almost all sand flies were killed, except at the border with Bihar (mortality 62%). With 0.05% deltamethrin, between 96 and 100% of the sand flies were killed in both regions. Conclusions Based on literature and present data 4% DDT and 0.05% deltamethrin seem to be acceptable discriminating concentrations to separate resistant from susceptible populations. Resistance to DDT was confirmed in Bihar and in a border village of Nepal, but the sand flies were still susceptible in villages more inside Nepal where only synthetic pyrethroids are used for indoor spraying. The low effectiveness of indoor spraying with DDT in Bihar to control VL can be partially explained by this resistance hence other classes of insecticides should be tested. In both countries P. argentipes sand flies were susceptible to deltamethrin.

Measurement of recent exposure to Phlebotomus argentipes, the vector of Indian visceral Leishmaniasis, by using human antibody responses to sand fly saliva.

Antibody (IgG) responses to the saliva of Phlebotomus argentipes were investigated using serum samples from regions of India endemic and non-endemic for visceral leishmaniasis (VL). By pre-adsorbing the sera against the saliva of the competing human-biting but non-VL vector P. papatasi, we significantly improved the specificity of a P. argentipes saliva enzyme-linked immunosorbent assay. Using this method, we observed a statistically significant correlation between antibodies to P. argenitpes saliva and the average indoor density of female sand flies. Additionally, the method was able to detect recent changes in vector exposure when sera from VL patients were assayed before, during, and after hospitalization and protected from sand fly bites under untreated bed nets. Collectively, these results highlight the utility of antibodies to P. argentipes saliva as an important tool to evaluate VL vector control programs.

Long-lasting insecticidal nets fail at household level to reduce abundance of sandfly vector Phlebotomus argentipes in treated houses in Bihar (India)

Objective  To determine whether the use of long‐lasting insecticidal nets (LLINS) at household level are effective in reducing the abundance of Phlebotomus argentipes, vector of anthroponotic visceral leishmaniasis in India, Nepal and Bangladesh.

The efficacy of indoor CDC light traps for collecting the sandfly Phlebotomus argentipes, vector of Leishmania donovani

Abstract The efficacy of light traps for collecting sandflies (Diptera: Psychodidae) varies both inter‐specifically and intra‐specifically (by gender and physiological status) as a result of significant differences in phototropic and other behavioural characteristics. The efficacy of miniature CDC light traps for collecting Phlebotomus argentipes Annandale & Brunetti, a vector of Leishmania donovani Laveran & Mesnil (Kinetoplastida: Trypanosomatidae), was assessed in the Indian state of Bihar. Sandflies were collected during the night from 16 houses in each of three villages over 3 months (four times at fortnightly intervals) using CDC light traps indoors, and by aspirator collection (carried out by one person for 30 min/house) from the walls of the same houses the following morning. Incidence rate ratios (IRRs) between CDC light trap collections and aspirator collections were obtained through a negative binomial regression with household as random effect. CDC light traps were especially effective in catching males (IRR 3.08, 95% confidence interval [CI] 2.12–4.46) and unfed females (IRR 3.50, 95% CI 2.37–5.16) of P. argentipes, and to a lesser extent gravids (IRR 1.77, 95% CI 1.07–2.93). However, only a relatively small proportion of all blood‐fed P. argentipes were collected by light trap (IRR 0.45, 95% CI 0.28–0.73). Despite its limitations in collecting blood‐fed female sandflies, the CDC light trap appears to trap a sufficient proportion of the indoor population of sandflies for sampling purposes, and as this light trap is also more convenient and more easily standardized than the aspirator method, we conclude that it is the most efficient method for monitoring P. argentipes populations in the Indian subcontinent.

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

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

Peridomestic Breeding and Resting Sites of Sandflies (Diptera: Psychodidae) in Bihar, India

ABSTRACT: Bihar has been endemic for Visceral Leishmaniasis for over a century and shares approximately 90 percent cases of India. The Old World sandflies are mostly endophilic and endophagic. However, outdoor breeding sites were found in the peridomestic areas of human dwellings and cattle sheds from one to 100 meters distant. Sandflies were collected using CDC light traps from between abandoned wells covered with wild plants, bushes of bamboo (Bambusa bambos), banana (Musa sapientum) and wild herbs, old stacks of bricks covered with smaller plants making the environment cool and providing food to larvae from organic debris due to litter. The resting sandflies were also collected by aspirator and flash light from the dwellings and cattle sheds. The findings will assist in mapping of peridomestic breeding and resting sites of sandflies which will be of immense help during implementation of control programs for the disease in Bihar, India.

Oviposition behaviour of Phlebotomus argentipes - A laboratory-based study

The breeding habitat of sandflies is a little studied and poorly understood phenomenon. More importantly, oviposition behaviour is a largely neglected aspect of sandfly biology and this knowledge gap further undermines our understanding of the biology of sandflies. Pheromones released by the eggs play an important role in identifying good sites for oviposition by female insects. Several recent studies have examined the oviposition pheromone. The present study provides a preliminary report on the oviposition behaviour of Phlebotomus argentipes, the only vector of kala-azar (or visceral leishmaniasis) on the Indian sub-continent. Sandflies prefer to oviposit their eggs on surfaces that contain organic substances, especially substances with an odour of decaying animal products and the remains of conspecific eggs. The results presented here suggest that the odour released by the organic substances of old sandfly colony remains that contain dead flies, old unhatched eggs, larval food containing vertebrate faeces, frass and other organic matter serves as an attractant for the ovipositing females of P. argentipes and hence greatly increases the number of oviposited eggs compared to eggs deposited in controlled oviposition pots. This result will be helpful in maintaining an efficient colony of P. argentipes and may be a promising tool for monitoring and controlling the target insect as part of a synergistic approach.

Insecticidal effect of plant extracts on Phlebotomus argentipes (Diptera: Psychodidae) in Bihar, India

Background & objectives: Phlebotomus argentipes (Diptera: Psychodidae), the established vector for kala-azar is presently being controlled by indoor residual spray of DDT in kala-azar endemic areas in India. Search for non-hazardous and non-toxic biodegradable active molecules from botanicals may provide cost-effective and eco-friendly alternatives to synthetic insecticides. The present study was aimed at evaluating various plant extracts from endemic and non-endemic areas of Bihar for their insecticidal activity against sandfly to identify the most effective plant extract. Methods: Bio-assay test was conducted with larvae and adult of P. argentipes with different plant extracts collected in distilled water, hexane, ethyl acetate, acetone and methanol. Thin layer chromatography (TLC), column chromatography and high performance liquid chromatography (HPLC) were conducted for detection of active molecules. Results: Adults and larvae of sandflies exposed to the aqueous extract of Nicotiana tabacum resulted in 100 per cent mortality. The hexane extract of Clerodendrum infortunatum was found to kill 77 per cent adults but was ineffective against larvae. Bio-assay test of the ninth fraction (hexane extract-methanol phase) separated by column chromatography was found to be 63 per cent effective. The purple spot on the TLC of this fraction indicated the presence of a diterpenoid. HPLC of this fraction detected nine compounds with two peaks covering 20.44 and 56.52 per cent areas with retention time of 2.439 and 5.182 min, respectively supporting the TLC results. Interpretation & conclusions: The column separated 9th fraction of C. infortunatum extract was found to be effective in killing 63 per cent of adult P. argentipes. Compounds of this fraction need to be evaluated further for identification and characterization of the active molecule by conducting individual bio-assay tests followed by further fractionation and HPLC. Once the structure of the active molecule is identified and validated, it may be synthesized and formulated as a product.