Molecular surveillance of insecticide resistance in Phlebotomus argentipes targeted by indoor residual spraying for visceral leishmaniasis elimination in India.

Molecular surveillance of resistance is an increasingly important part of vector borne disease control programmes that utilise insecticides. The visceral leishmaniasis (VL) elimination programme in India uses indoor residual spraying (IRS) with the pyrethroid, alpha-cypermethrin to control Phlebotomus argentipes the vector of Leishmania donovani, the causative agent of VL. Prior long-term use of DDT may have selected for knockdown resistance (kdr) mutants (1014F and S) at the shared DDT and pyrethroid target site, which are common in India and can also cause pyrethroid cross-resistance. We monitored the frequency of these marker mutations over five years from 2017-2021 in sentinel sites in eight districts of north-eastern India covered by IRS. Frequencies varied markedly among the districts, though finer scale variation, among villages within districts, was limited. A pronounced and highly significant increase in resistance-associated genotypes occurred between 2017 and 2018, but with relative stability thereafter, and some reversion toward more susceptible genotypes in 2021. Analyses linked IRS with mutant frequencies suggesting an advantage to more resistant genotypes, especially when pyrethroid was under-sprayed in IRS. However, this advantage did not translate into sustained allele frequency changes over the study period, potentially because of a relatively greater net advantage under field conditions for a wild-type/mutant genotype than projected from laboratory studies and/or high costs of the most resistant genotype. Further work is required to improve calibration of each 1014 genotype with resistance, preferably using operationally relevant measures. The lack of change in resistance mechanism over the span of the study period, coupled with available bioassay data suggesting susceptibility, suggests that resistance has yet to emerge despite intensive IRS. Nevertheless, the advantage of resistance-associated genotypes with IRS and under spraying, suggest that measures to continue monitoring and improvement of spray quality are vital, and consideration of future alternatives to pyrethroids for IRS would be advisable.

Plant Secondary Metabolites as Defense Tools against Herbivores for Sustainable Crop Protection.

Plants have evolved several adaptive strategies through physiological changes in response to herbivore attacks. Plant secondary metabolites (PSMs) are synthesized to provide defensive functions and regulate defense signaling pathways to safeguard plants against herbivores. Herbivore injury initiates complex reactions which ultimately lead to synthesis and accumulation of PSMs. The biosynthesis of these metabolites is regulated by the interplay of signaling molecules comprising phytohormones. Plant volatile metabolites are released upon herbivore attack and are capable of directly inducing or priming hormonal defense signaling pathways. Secondary metabolites enable plants to quickly detect herbivore attacks and respond in a timely way in a rapidly changing scenario of pest and environment. Several studies have suggested that the potential for adaptation and/or resistance by insect herbivores to secondary metabolites is limited. These metabolites cause direct toxicity to insect pests, stimulate antixenosis mechanisms in plants to insect herbivores, and, by recruiting herbivore natural enemies, indirectly protect the plants. Herbivores adapt to secondary metabolites by the up/down regulation of sensory genes, and sequestration or detoxification of toxic metabolites. PSMs modulate multi-trophic interactions involving host plants, herbivores, natural enemies and pollinators. Although the role of secondary metabolites in plant-pollinator interplay has been little explored, several reports suggest that both plants and pollinators are mutually benefited. Molecular insights into the regulatory proteins and genes involved in the biosynthesis of secondary metabolites will pave the way for the metabolic engineering of biosynthetic pathway intermediates for improving plant tolerance to herbivores. This review throws light on the role of PSMs in modulating multi-trophic interactions, contributing to the knowledge of plant-herbivore interactions to enable their management in an eco-friendly and sustainable manner.

Harris hawks optimization algorithm for model order reduction of interconnected wind turbines.

This paper illustrates the investigation of higher-scale interconnected wind turbine to realize its analytical design and simulation performance in terms of reduced order model. The present research is aimed to visualize the efficacy of the propounded harris hawks optimization (HHO) approach for different reduced order transfer functions with reference to model order reduction studies. Compared to the higher-scale model, the applied methodology aims to obtain better results in terms of steadiness and computational effort. Moreover, to realize a higher-dimensional power system, the implementation of the HHO may proffer more effectiveness and robustness as opposed to the state-of-the-art optimization techniques. The applied methodology aims to obtain better results in terms of stability and computational effort. Furthermore, to realize a higher dimensional system, implementation of the HHO may offer more effectiveness and robustness as opposed to the state-of-the-art optimization techniques.

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.

Determinants for progression from asymptomatic infection to symptomatic visceral leishmaniasis: A cohort study.

BACKGROUND: Asymptomatic Leishmania donovani infections outnumber clinical presentations, however the predictors for development of active disease are not well known. We aimed to identify serological, immunological and genetic markers for progression from L. donovani infection to clinical Visceral Leishmaniasis (VL). METHODS: We enrolled all residents >2 years of age in 27 VL endemic villages in Bihar (India). Blood samples collected on filter paper on two occasions 6-12 months apart, were tested for antibodies against L. donovani with rK39-ELISA and DAT. Sero converters, (negative for both tests in the first round but positive on either of the two during the second round) and controls (negative on both tests on both occasions) were followed for three years. At the start of follow-up venous blood was collected for the following tests: DAT, rK39- ELISA, Quantiferon assay, SNP/HLA genotyping and L.donovani specific quantitative PCR. RESULTS: Among 1,606 subjects enrolled,17 (8/476 seroconverters and 9/1,130 controls) developed VL (OR 3.1; 95% CI 1.1-8.3). High DAT and rK39 ELISA antibody titers as well as positive qPCR were strongly and significantly associated with progression from seroconversion to VL with odds ratios of 19.1, 30.3 and 20.9 respectively. Most VL cases arose early (median 5 months) during follow-up. CONCLUSION: We confirmed the strong association between high DAT and/or rK39 titers and progression to disease among asymptomatic subjects and identified qPCR as an additional predictor. Low predictive values do not warrant prophylactic treatment but as most progressed to VL early during follow-up, careful oberservation of these subjects for at least 6 months is indicated.

Phytosome complexed with chitosan for gingerol delivery in the treatment of respiratory infection: In vitro and in vivo evaluation.

Respiratory infection is a viral spreading disease and a common issue, particularly in kids. The treatments are available but have so many limitations because the drawback of this disease is more morbidity and mortality in the severely immune compromised. Even, the phyto-constituent antibacterial drug Gingerol was selected to treat respiratory infection but it exhibits low bioavailability profile, less aqueous-solubility issue and most important is rapidly eliminated from the body. To overcome these problems, novel drug delivery (nanoparticle) based phytosome complexed with chitosan approach was implemented. In this research work, the phytosome (GP) was prepared by blending of gingerol with soya lecithin in organic solvent using anti-solvent precipitation technique and it was further loaded in the aqueous solution of chitosan to formulate the phytosome complexed with chitosan (GLPC). To optimize the formulations of gingerol, it was characterized for percentage yield, percentage entrapment efficiency, drug loading and particle size, physical compatibility studies etc. which demonstrated the confirmation of complex of GLPC with soya lecithin and chitosan. The % entrapment efficiency and % drug loading of GLPC was found (86.02 ±â€¯0.18%, 08.26 ±â€¯0.72%) and of GP (84.36 ±â€¯0.42%, 08.05 ±â€¯0.03%), respectively. The average particle size and zeta potential of GLPC and GP were 254.01 ±â€¯0.05 nm (-13.11 mV), and 431.21 ±â€¯0.90 nm (-17.53 mV), respectively which confirm the inhibition of particle aggregation by using chitosan in complex. The in vitro release rate of GP (86.03 ±â€¯0.06%) was slower than GLPC (88.93 ±â€¯0.33%) in pH 7.4 phosphate buffer up to 24 h by diffusion process (Korsmeyer Peppas model). The optimized GLPC and GP were shown irregular particle shapes & spherical and oval structures with smooth surface by SEM analysis. Furthermore, GLPC has shown the potent in vitro antioxidant activity, susceptible antibacterial activity and effective anti-inflammatory activity as compared to GP against stress, microbial infection and inflammation which were causable reason for the respiratory infections. GLPC has improved the significant bioavailability and also correlated the hematological values on rabbit blood against the incubation of microorganisms. Thus, the prepared nanoparticle based approach to deliver the gingerol, has the combined effect of chitosan and phytosome which shown better sustained-release profile and also prolonging the oral absorption rate of gingerol with effective antibacterial activity to treat respiratory infection.

Knockdown resistance mutations predict DDT resistance and pyrethroid tolerance in the visceral leishmaniasis vector Phlebotomus argentipes.

BACKGROUND: Indoor residual spraying (IRS) with DDT has been the primary strategy for control of the visceral leishmaniasis (VL) vector Phlebotomus argentipes in India but efficacy may be compromised by resistance. Synthetic pyrethroids are now being introduced for IRS, but with a shared target site, the para voltage-gated sodium channel (VGSC), mutations affecting both insecticide classes could provide cross-resistance and represent a threat to sustainable IRS-based disease control. METHODOLOGY/PRINCIPAL FINDINGS: A region of the Vgsc gene was sequenced in P. argentipes from the VL hotspot of Bihar, India. Two knockdown resistance (kdr) mutations were detected at codon 1014 (L1014F and L1014S), each common in mosquitoes, but previously unknown in phlebotomines. Both kdr mutations appear largely recessive, but as homozygotes (especially 1014F/F) or as 1014F/S heterozygotes exert a strong effect on DDT resistance, and significantly predict survivorship to class II pyrethroids in short-duration bioassays. The mutations are present at high frequency in wild P. argentipes populations from Bihar, with 1014F significantly more common in higher VL areas. CONCLUSIONS/SIGNIFICANCE: The Vgsc mutations detected appear to be a primary mechanism underlying DDT resistance in P. argentipes and a contributory factor in reduced pyrethroid susceptibility, suggesting a potential impact if P. argentipes are subjected to suboptimal levels of pyrethroid exposure, or additional resistance mechanisms evolve. The assays to detect kdr frequency changes provide a sensitive, high-throughput monitoring tool to detecting spatial and temporal variation in resistance in P. argentipes.

Development of a Simple Dipstick Assay for Operational Monitoring of DDT.

BACKGROUND: Indoor residual spraying (IRS) of DDT is used to control visceral leishmaniasis (VL) in India. However, the quality of spraying is severely compromised by a lack of affordable field assays to monitor target doses of insecticide. Our aim was to develop a simple DDT insecticide quantification kit (IQK) for monitoring DDT levels in an operational setting. METHODOLOGY/ PRINCIPLE FINDINGS: DDT quantification was based on the stoichiometric release of chloride from DDT by alkaline hydrolysis and detection of the released ion using Quantab chloride detection strips. The assay was specific for insecticidal p,p`-DDT (LoQ = 0.082 g/m2). Bostik discs were effective in post spray wall sampling, extracting 25-70% of active ingredient depending on surface. Residual DDT was sampled from walls in Bihar state in India using Bostik adhesive discs and DDT concentrations (g p,p`-DDT/m2) were determined using IQK and HPLC (n = 1964 field samples). Analysis of 161 Bostik samples (pooled sample pairs) by IQK and HPLC produced excellent correlation (R2 = 0.96; Bland-Altman bias = -0.0038). IQK analysis of the remaining field samples matched HPLC data in identifying households that had been under sprayed, in range or over sprayed. INTERPRETATION: A simple dipstick assay has been developed for monitoring DDT spraying that gives comparable results to HPLC. By making laboratory-based analysis of DDT dosing accessible to field operatives, routine monitoring of DDT levels can be promoted in low- and middle- income countries to maximise the effectiveness of IRS.

DDT-based indoor residual spraying suboptimal for visceral leishmaniasis elimination in India.

Indoor residual spraying (IRS) is used to control visceral leishmaniasis (VL) in India, but it is poorly quality assured. Quality assurance was performed in eight VL endemic districts in Bihar State, India, in 2014. Residual dichlorodiphenyltrichloroethane (DDT) was sampled from walls using Bostik tape discs, and DDT concentrations [grams of active ingredient per square meter (g ai/m(2))] were determined using HPLC. Pre-IRS surveys were performed in three districts, and post-IRS surveys were performed in eight districts. A 20% threshold above and below the target spray of 1.0 g ai/m(2) was defined as "in range." The entomological assessments were made in four districts in IRS and non-IRS villages. Vector densities were measured: pre-IRS and 1 and 3 mo post-IRS. Insecticide susceptibility to 4% DDT and 0.05% deltamethrin WHO-impregnated papers was determined with wild-caught sand flies. The majority (329 of 360, 91.3%) of pre-IRS samples had residual DDT concentrations of <0.1 g ai/m(2). The mean residual concentration of DDT post-IRS was 0.37 g ai/m(2); 84.9% of walls were undersprayed, 7.4% were sprayed in range, and 7.6% were oversprayed. The abundance of sand flies in IRS and non-IRS villages was significantly different at 1 mo post-IRS only. Sand flies were highly resistant to DDT but susceptible to deltamethrin. The Stockholm Convention, ratified by India in 2006, calls for the complete phasing out of DDT as soon as practical, with limited use in the interim where no viable IRS alternatives exist. Given the poor quality of the DDT-based IRS, ready availability of pyrethroids, and susceptibility profile of Indian sand flies, the continued use of DDT in this IRS program is questionable.

λ-Carrageenan Suppresses Tomato Chlorotic Dwarf Viroid (TCDVd) Replication and Symptom Expression in Tomatoes.

The effect of carrageenans on tomato chlorotic dwarf viroid (TCDVd) replication and symptom expression was studied. Three-week-old tomato plants were spray-treated with iota(É©)-, lambda(λ)-, and kappa(κ)-carrageenan at 1 g·L-1 and inoculated with TCDVd after 48 h. The λ-carrageenan significantly suppressed viroid symptom expression after eight weeks of inoculation, only 28% plants showed distinctive bunchy-top symptoms as compared to the 82% in the control group. Viroid concentration was reduced in the infected shoot cuttings incubated in λ-carrageenan amended growth medium. Proteome analysis revealed that 16 tomato proteins were differentially expressed in the λ-carrageenan treated plants. Jasmonic acid related genes, allene oxide synthase (AOS) and lipoxygenase (LOX), were up-regulated in λ-carrageenan treatment during viroid infection. Taken together, our results suggest that λ-carrageenan induced tomato defense against TCDVd, which was partly jasmonic acid (JA) dependent, and that it could be explored in plant protection against viroid infection.

Health & Demographic Surveillance System profile: the Muzaffarpur-TMRC Health and Demographic Surveillance System.

The Muzaffarpur-TMRC Health and Demographic Surveillance System (HDSS), established in 2007, was developed as an enlargement of the scope of a research collaboration on the project Visceral Leishmaniasis in Bihar, which had been ongoing since 2005. The HDSS is located in a visceral leishmaniasis (VL)-endemic area in the Muzaffarpur district of Bihar state in India. It is the only HDSS conducting research on VL, which is a vector-borne infectious disease transmitted by female phlebotomine sandflies and is fatal if left untreated. Currently the HDSS serves a population of over 105,000 in 66 villages. The HDSS collects data on vital events including pregnancies, births, deaths, migration and marriages, as well as other socio-economic indicators, at regular intervals. Incident VL cases are identified. The HDSS team is experienced in conducting both qualitative and quantitative studies, sample collection and rapid diagnostic tests in the field. In each village, volunteers connect the HDSS team with the community members. The Muzaffarpur-TMRC HDSS provides opportunities for studies on VL and other neglected tropical diseases (NTDs) and their interaction with demographic events such as migration. Queries related to research collaborations and data sharing can be sent to Dr Shyam Sundar at [drshyamsundar@hotmail.com].

Adverse Effects of UV-B Radiation on Plants Growing at Schirmacher Oasis, East Antarctica.

This study aimed to assess the impacts of ultraviolet-B (UV-B) radiation over a 28-day period on the levels of pigments of Umbilicaria aprina and Bryum argenteum growing in field. The depletion of stratospheric ozone is most prominent over Antarctica, which receives more UV-B radiation than most other parts of the planet. Although UV-B radiation adversely affects all flora, Antarctic plants are better equipped to survive the damaging effects of UV-B owing to defenses provided by UV-B absorbing compounds and other screening pigments. The UV-B radiations and daily average ozone values were measured by sun photometer and the photosynthetic pigments were analyzed by the standard spectrophotometric methods of exposed and unexposed selected plants. The daily average atmospheric ozone values were recorded from 5 January to 2 February 2008. The maximum daily average for ozone (310.7 Dobson Units (DU)) was recorded on 10 January 2008. On that day, average UV-B spectral irradiances were 0.016, 0.071, and 0.186 W m(-2) at wavelengths of 305, 312, and 320 nm, respectively. The minimum daily average ozone value (278.6 DU) was recorded on 31 January 2008. On that day, average UV-B spectral irradiances were 0.018, 0.085, and 0.210 W m(-2) at wavelengths of 305, 312, and 320 nm, respectively. Our results concludes that following prolonged UV-B exposure, total chlorophyll levels decreased gradually in both species, whereas levels of UV-B absorbing compounds, phenolics, and carotenoids gradually increased.

Strong association between serological status and probability of progression to clinical visceral leishmaniasis in prospective cohort studies in India and Nepal.

INTRODUCTION: Asymptomatic persons infected with the parasites causing visceral leishmaniasis (VL) usually outnumber clinically apparent cases by a ratio of 4-10 to 1. We assessed the risk of progression from infection to disease as a function of DAT and rK39 serological titers. METHODS: We used available data on four cohorts from villages in India and Nepal that are highly endemic for Leishmania donovani. In each cohort two serosurveys had been conducted. Based on results of initial surveys, subjects were classified as seronegative, moderately seropositive or strongly seropositive using both DAT and rK39. Based on the combination of first and second survey results we identified seroconvertors for both markers. Seroconvertors were subdivided in high and low titer convertors. Subjects were followed up for at least one year following the second survey. Incident VL cases were recorded and verified. RESULTS: We assessed a total of 32,529 enrolled subjects, for a total follow-up time of 72,169 person years. Altogether 235 incident VL cases were documented. The probability of progression to disease was strongly associated with initial serostatus and with seroconversion; this was particularly the case for those with high titers and most prominently among seroconvertors. For high titer DAT convertors the hazard ratio reached as high as 97.4 when compared to non-convertors. The strengths of the associations varied between cohorts and between markers but similar trends were observed between the four cohorts and the two markers. DISCUSSION: There is a strongly increased risk of progressing to disease among DAT and/or rK39 seropositives with high titers. The options for prophylactic treatment for this group merit further investigation, as it could be of clinical benefit if it prevents progression to disease. Prophylactic treatment might also have a public health benefit if it can be corroborated that these asymptomatically infected individuals are infectious for sand flies.

The discovery and eradication of potato spindle tuber viroid in Canada.

In 1960s, potato spindle tuber was thought to be a viral disease. In 1971, the agent of the disease was characterised as a low-molecular weight infectious ribonucleic acid (RNA), which was named as 'viroid', specifically Potato spindle tuber viroid (PSTVd)). Since then, more than 30 plant diseases in horticultural and ornamental plants have been shown to be caused by different viroids globally. Viroids are single-stranded RNA, covalently closed circular molecule, without any protein coat. They are the smallest known plant pathogen containing RNA genome ranging from 246 nucleotides (Coconut cadang-cadang viroid) to 399 nucleotides (Chrysanthemum chlorotic mottle viroid). Some viroids are located in the plant cell nucleus (pospiviroids) and others in the chloroplast (avsunviroids). With the recognition of pathogenic nature of viroid, specific detection methodologies were developed, which enabled detection of PSTVd in seed-potato tubers prior to their planting in the field, and thus PSTVd was prevented from spreading the disease. As a result, PSTVd was eradicated from Canada in late 1980s. Viroids similar to PSTVd (Pospiviroid) have been discovered and they are detected in symptomless ornamental plants. Although, PSTVd has been eradicated from Canada, there is a strong possibility of viroid introduction from other plants besides potato and tomato and causing PSTVd like diseases.

Seasonal variation in the prevalence of sand flies infected with Leishmania donovani.

BACKGROUND: Visceral Leishmaniasis (VL) is a life threatening neglected infectious disease in the Indian subcontinent, transmitted by the bite of female sand flies. Estimation of the infectivity in the vector population, collected in different seasons, may be useful to better understanding the transmission dynamics of VL as well as to plan vector control measures. METHODOLOGY: We collected sand flies from highly endemic regions of Bihar state, India for one year over three seasons. The species of the sand flies were confirmed by species-specific PCR-RFLP. Leishmania donovani infection was investigated in 1397 female Phlebotomus argentipes using PCR, targeting the Leishmania specific minicircle of the kDNA region. Further, the parasitic load in the infected sand flies was measured using quantitative PCR. CONCLUSION: Though sand flies were most abundant in the rainy season, the highest rate of infection was detected in the winter season with 2.84% sand flies infected followed by the summer and rainy seasons respectively. This study can help in vector elimination programmes and to reduce disease transmission.

Latent infection with Leishmania donovani in highly endemic villages in Bihar, India.

INTRODUCTION: Asymptomatic persons infected with the parasites causing visceral leishmaniasis (VL) usually outnumber clinically apparent cases by a ratio of 4-10 to 1. We describe patterns of markers of Leishmania donovani infection and clinical VL in relation to age in Bihar, India. METHODS: We selected eleven villages highly endemic for Leishmania donovani. During a 1-year interval we conducted two house to house surveys during which we collected blood samples on filter paper from all consenting individuals aged 2 years and above. Samples were tested for anti-leishmania serology by Direct Agglutination Test (DAT) and rK39 ELISA. Data collected during the surveys included information on episodes of clinical VL among study participants. RESULTS: We enrolled 13,163 persons; 6.2% were reactive to DAT and 5.9% to rK39. Agreement between the tests was weak (kappa = 0.30). Among those who were negative on both tests at baseline, 3.6% had converted to sero-positive on either of the two tests one year later. Proportions of sero-positives and sero-converters increased steadily with age. Clinical VL occurred mainly among children and young adults (median age 19 years). DISCUSSION: Although infection with L. donovani is assumed to be permanent, serological markers revert to negative. Most VL cases occur at younger ages, yet we observed a steady increase with age in the frequency of sero-positivity and sero-conversion. Our findings can be explained by a boosting effect upon repeated exposure to the parasite or by intermittent release of parasites in infected subjects from safe target cells. A certain proportion of sero-negative subjects could have been infected but below the threshold of antibody abundance for our serologic testing.

Retrospective Quarterly Cohort Monitoring for patients with Visceral Leishmaniasis in the Indian subcontinent: outcomes of a pilot project.

OBJECTIVE: To evaluate a new tool for the monitoring of Visceral Leishmaniasis (VL) treatment outcomes in primary healthcare (PHC) settings, adapted from the standardised Retrospective Quarterly Cohort Monitoring done in tuberculosis control. METHODS: We developed standard case definitions for early and late VL treatment outcomes, a single register allowing for one-line entry per patient as registration tool, and quarterly reporting formats for the clinical outcomes. We pilot-tested these tools in three Indian Primary Health Centres and two Nepalese district hospitals, as well as in a charity VL treatment centre and a university hospital. RESULTS: Data collection for early treatment outcome was easily implemented but information on late treatment outcome was hard to obtain. Effectiveness of Miltefosine under routine care conditions was about 87% at end of treatment, and 76% at 6 months post-treatment related to the high number of patients lost to follow up at the latter end point. CONCLUSION: A retrospective cohort monitoring methodology is conceptually a good framework for monitoring clinical outcomes for chronic conditions as VL. The monitoring of early outcomes of VL treatment is perfectly feasible in Primary Care settings. The completeness of information on late outcomes can be improved by a number of strategies that remain to be field tested. Generally, clinical outcome monitoring should be strengthened in the VL control programmes.

Visceral leishmaniasis in rural bihar, India.

To identify factors associated with incidence of visceral leishmaniasis (VL), we surveyed 13,416 households in Bihar State, India. VL was associated with socioeconomic status, type of housing, and belonging to the Musahar caste. Annual coverage of indoor residual insecticide spraying was 12%. Increasing such spraying can greatly contribute to VL control.

Post-kala-azar dermal leishmaniasis in visceral leishmaniasis-endemic communities in Bihar, India.

We assessed the prevalence of post-kala-azar dermal leishmaniasis (PKDL), a late cutaneous manifestation of visceral leishmaniasis (VL), in 16 VL-endemic communities in Bihar, India. The prevalence of confirmed PKDL cases was 4.4 per 10 000 individuals and 7.8 if probable cases were also considered. The clinical history and treatment of the post-kala-azar dermal leishmaniasis cases are discussed.

Prevalence of sand flies and Leishmania donovani infection in a natural population of female Phlebotomus argentipes in Bihar State, India.

Leishmaniasis is a vector-borne disease, and in the Indian subcontinent the female Phlebotomus argentipes is the vector for Leishmania donovani. However, data on the extent of sand fly infection rates in natural settings using molecular methods have not been extensively reported in India. In this study a PCR technique was applied targeting the 18S rRNA encoding region to determine the prevalence of Leishmania infection in female P. argentipes captured in the field. For this study, sand flies were collected from 897 houses selected from 50 villages endemic for visceral leishmaniasis (VL) in Muzaffarpur district, Bihar state, using CDC miniature light traps and mouth aspirators. A total of 14,585 sand flies were collected of which 449 were female P. argentipes divided into 132 pools. Molecular detection using PCR targeting the 18S rRNA gene was carried out for the identification of P. argentipes and Leishmania. The overall prevalence of infection was 4.90-17.37% for L. donovani in female P. argentipes in endemic regions of Bihar state. In this study no correlation was found between the presence of infected sand flies and the occurrence of clinical VL. This study provides the first report evaluating the prevalence of Leishmania infection in sand flies in a region endemic for VL in India. Sergentomyia species are the most common species of sand fly. Knowledge of the infection rate in female P. argentipes may help in predicting severity of disease and in vector elimination programs.