Genotype of MTNR1A gene regulates the conception rate following melatonin treatment in water buffalo.

Buffaloes have tendency to show seasonal reproduction and remain in anestrus due to limited ovarian activity during summer. The seasonal reproductive behavior is ascribed the effect of melatonin related to photoperiod. Treating animals with melatonin could be a possible strategy to overcome the problem. The role of MTNR1A gene has not been fully explained in the buffalo. Therefore, we conducted a study on 114 buffalo heifers to detect the polymorphic site in MTNR1A gene and further treated them with melatonin implants to investigate the role of most frequent genotype following melatonin treatment on pregnancy. The present investigation is the first to investigate the association between melatonin treated different MTNR1A genotype buffalo and pregnancy. We confirmed SNP at position 72 in 812 bp fragment exon II of MTNR1A gene. RFLP of PCR products with Hpa I enzyme resulted in three genotypes: TT (812bp), CT (812, 743, 69bp) and CC (743, 69bp). Next, buffaloes of each genotype (TT, CC, CT; n = 28 for each) were treated with melatonin implants to compare the conception rate with their corresponding untreated control (n = 10 for each genotype). Melatonin concentrations were higher (P < 0.05) for the treatment groups of all genotypes compared to their respective untreated control from day 1-28. The pregnancy rate was significantly associated with the MTNR1A genotype. The conception rate was higher (P < 0.05) for TT genotype than for the other genotypes of buffaloes treated with melatonin. Furthermore, buffaloes of TT genotype treated with melatonin started exhibiting estrus activity soon from second week of melatonin treatment (14.1 ±â€¯2.1; range: 10-17 days) and were found to be 7.8 times more likely to become pregnant compared to other genotypes following melatonin treatment. In conclusion, TT genotype of MTNR1A gene is more sensitive to melatonin treatment that favours pregnancy in buffaloes during summer.

Insecticide resistance status of three vectors of Japanese encephalitis in east central India.

Japanese encephalitis (JE) has been reported in different districts of Odisha state (east central India) since 1992. During 2016, a major outbreak of JE and acute encephalitis syndrome (AES) occurred in the Malkangiri district of Odisha, causing 103 deaths in children, of which 37 were caused by JE and 66 by AES. Information on insecticide resistance in JE vectors is important for the selection of appropriate insecticides for use in vector control. The present study was designed to determine the resistance status of three important vectors of JE, Culex vishnui, Culex tritaeniorhynchus and Culex bitaeniorhynchus (Diptera: Culicidae), against dichlorodiphenyltrichloroethane (DDT), malathion and deltamethrin in three districts of Odisha state affected by JE. Female adult mosquitoes were collected using mouth aspirators both indoors and outdoors in JE-affected villages and used in susceptibility bioassays following World Health Organization guidelines. Knock-downs were recorded every 10 min up to 1 h and mortality rates were recorded at 24 h post-exposure. Culex vishnui and Cx. tritaeniorhynchus showed resistance to DDT, malathion and deltamethrin, whereas Cx. bitaeniorhynchus was susceptible in all study districts. The information generated by this study will be highly useful in the planning and implementing of appropriate vector control operations for the prevention and control of JE in east central India.

Bionomics of Anopheles fluviatilis and Anopheles culicifacies (Diptera: Culicidae) in Relation to Malaria Transmission in East-Central India.

The southern districts of Odisha State in east-central India have been highly endemic for falciparum malaria for many decades. However, there is no adequate information on the abundance of the vector species or their bionomics in relation to space and time in these districts. Therefore, a study was carried out on the entomological aspects of malaria transmission to generate such information. Collections of mosquitoes were made once during each of the three seasons in 128 villages selected from eight districts. Villages within the foot-hill ecotype had a significantly greater abundance of Anopheles fluviatilis James s. l., whereas the abundance of Anopheles culicifacies Giles s. l. was significantly greater in the plain ecotype. The abundance of An. fluviatilis was maximum during the cold season, whereas An. culicifacies abundance was highest during summer and rainy seasons. The maximum likelihood estimation of the malaria infection rate in An. fluviatilis was 1.78%, 6.05%, and 2.6% in Ganjam, Kalahandi, and Rayagada districts, respectively. The infection rate of An. culicifacies was 1.39% only in Kandhamal district; infected females were not detected elsewhere. Concurrently, the annual malaria parasite incidence (MPI) was significantly higher in hill-top (17.6) and foot-hill (14.4) villages compared to plain villages (4.1). The districts with more villages in hill-top and foot-hill ecotypes also had a greater abundance of An. fluviatilis, the major malaria vector, and exhibited a higher incidence of malaria than villages within the plain ecotype, where An. culicifacies was the most abundant vector.

Field performance of malaria rapid diagnostic test for the detection of Plasmodium falciparum infection in Odisha State, India.

BACKGROUND & OBJECTIVES: Rapid diagnostic tests (RDTs) have become an essential surveillance tool in the malaria control programme in India. The current study aimed to assess the performance of ParaHIT-f, a rapid test in diagnosis of Plasmodium falciparum infection through detecting its specific antigen, histidine rich protein 2 (PfHRP-2), in Odisha State, India. METHODS: The study was undertaken in eight falciparum malaria endemic southern districts of Odisha State. Febrile patients included through active case detection, were diagnosed by Accredited Social Health Activists (ASHAs) for P. falciparum infection using the RDT, ParaHIT-f. The performance of ParaHIT-f was evaluated using microscopy as the gold standard. RESULTS: A total of 1030 febrile patients were screened by both microscopy and the RDT for P. falciparum infection. The sensitivity of ParaHIT-f was 63.6% (95% CI: 56.0-70.6) and specificity was 98.9% (95% CI: 97.9-99.5), with positive and negative predictive values (PPV and NPV) of 92.6% (95% CI: 86.0-96.3) and 93.0% (95% CI: 91.0-94.5), respectively. When related to parasitaemia, the RDT sensitivity was 47.8% at the low parasitaemia of 4 to 40 parasites/µl of blood. INTERPRETATION & CONCLUSIONS: The results showed that the performance of the RDT, ParaHIT-f, was not as sensitive as microscopy in detecting true falciparum infections; a high specificity presented a low frequency of false-positive RDT results. t0 he sensitivity of ParaHIT-f was around 60 per cent. It is, therefore, essential to improve the efficiency (sensitivity) of the kit so that the true falciparum infections will not be missed especially in areas where P. falciparum has been the predominant species causing cerebral malaria.

Triple insecticide resistance in Anopheles culicifacies: A practical impediment for malaria control in Odisha State, India.

BACKGROUND & OBJECTIVES: In Odisha State, the control of malaria vectors has become dependent on synthetic pyrethroids, which are used for treatment of all approved long-lasting insecticidal nets (LLINs). The vast use of just one class of insecticide has led to the problem of resistance to insecticides in malaria vectors. One of the major malaria vectors in Odisha State is Anopheles culicifacies Giles. The aim of this study was to determine the resistance status of An. culicifacies to deltamethrin, a synthetic pyrethroid and other common insecticides used by the National Vector Borne Diseases Control Programme (NVBDCP) for indoor residual spraying in Odisha State. METHODS: Mosquitoes were collected during April 2014 - June 2014 from 15 randomly selected villages in five p0 lasmodium falciparum endemic southern districts of Odisha State. The blood-fed wild caught females were exposed to the diagnostic dosage of DDT (4.0%), malathion (5.0%) and deltamethrin (0.05%) for one hour. Mortality was recorded at 24 h after the exposure. RESULTS: Results indicated that An. culicifacies was resistant to all the three insecticides used in the malaria control programme in the five districts of Odisha State. INTERPRETATION & CONCLUSIONS: Resistance management strategy by appropriate rotation of different groups of insecticides including carbamates and incorporating a synergist with synthetic pyrethroids for treating mosquito nets should be considered for the control of malaria vectors in the area, especially where An. culicifacies is predominant. Periodical monitoring of susceptibility/resistance status of An. culicifacies to different insecticides is warranted.

Usage pattern, physical integrity & insecticidal efficacy of long-lasting insecticidal nets in Odisha State, India.

BACKGROUND & OBJECTIVES: One of the major strategies being pursued for malaria control by the National Vector Borne Disease Control Programme is the distribution of long lasting insecticidal nets (LLINs) in endemic areas since 2009. Information on durability of insecticidal efficacy and physical integrity of LLINs and community usage at different time intervals of their use is essential to plan net replacements to maintain universal coverage for an effective and sustainable malaria control strategy. Therefore, a study was undertaken to assess these parameters in two malaria endemic districts of the Odisha State. METHODS: A total of 309 households were selected in 15 villages of two community health centres (CHCs) (Borigumma and Laxmipur) from Koraput district and one (Khairput) from Malkangiri district. Data on net usage were collected during March to July 2014 using semi-structured questionnaires. PermaNet 2.0 were sampled from all households, replaced with new ones, and bioassays were carried out to determine the insecticidal efficacy of LLINs after four and half, four and two years of field use following the standard procedure of World Health Organization. RESULTS: LLIN use rate varied from 57.9 to 90.2 per cent in the study CHCs. The annual washing rate per net in Borigumma, Khairput and Laxmipur was 6.6, 3.2 and 4.8, respectively. The LLINs used two years in the field caused 100 per cent mortality and four to four and half years caused below 80 per cent mortality, except one net. INTERPRETATION & CONCLUSIONS: Nearly 20 per cent of the people were out of net coverage and hence the Programme to ensure 100 per cent coverage. The community should adequately be educated so as to increase the net use rate and avoid incorrect washing practices.

Effect of house spraying with lambdacyhalothrin 10 per cent capsule suspension (CS) formulation in comparison with 10 per cent wettable powder (WP) against malaria vector in Malkangiri district, Odisha, India.

BACKGROUND & OBJECTIVES: Selection of an insecticide and its appropriate formulation is a prerequisite of formulating any chemical control strategy against vectors. A hut scale field trial was carried out to study the effectiveness of house spraying with capsule suspension (CS) formulation of lambdacyhalothrin in comparison with its wettable powder (WP) formulation on mortality, density and behaviour of malaria vector in Malkangiri district, Odisha, India. METHODS: The two formulations were tested at 20 and 30 mg (a.i.)/m 2 for their effectiveness in terms of deterrence, excito-repellency, blood-feeding success, mortality and residual activity against Anopheles fluviatilis, the major malaria vector, in experimental huts in Malkangiri district, Odisha State, India. RESULTS: Both CS and WP formulations prevented the entry of An. fluviatilis in to the sprayed huts by >90 per cent for >6 months, the entire peak malaria transmission season in the area. The exit rate increased (90-99%) with different treatments and the feeding rate was reduced (91-97%). There was no significant difference between WP 30, CS 20 and CS 30 mg/m 2 in these respects. However, WP 20 mg/m 2 caused a lesser effect than the other three groups. The formulations or the dosages differed only in causing vector mortalities. Overall, the total mortality rate of An. fluviatilis was higher in the huts sprayed with CS 30 (58%) than the huts sprayed with CS 20 (48%), WP 20 (37%) and WP 30 mg/m 2 (47%). INTERPRETATION & CONCLUSIONS: Considering the duration of residual action and the effect on entry, exit, feeding and mortality of An. fluviatilis, the dosage 20 mg/m 2 of CS formulation of lambdacyhalothrin could be considered for further use.

Environmental management through sluice gated bed-dam: a revived strategy for the control of Anopheles fluviatilis breeding in streams.

BACKGROUND & OBJECTIVES: Integrated vector management (IVM) emphasizes sustainable eco-friendly methods and minimal use of chemicals. In this context, the present study highlights the environmental control of breeding of Anopheles fluviatilis, the primary malaria vector, through water management in a natural stream in Koraput district, Odisha, India. METHODS: The District Rural Development Agency (DRDA), Koraput, constructed two bed-dams across streams, one in Barigaon and the other in Pipalapodar village. The bed-dam in the former village was fitted with two sluice gates whereas the bed dam constructed in the latter village was without the sluice gate. the sluice gates were opened once in a week on a fixed day to flush out the water from the dam. Anopheles immatures were sampled systematically in the streams using a dipper for density measurement and species composition. RESULTS: There was a reduction of 84.9 per cent in the proportion of positive dips for anopheles larvae/pupae and a reduction of 98.4 per cent in immature density (number/dip) of An. fluviatilis in the experimental downstream compared to the control following opening of the sluice gates. INTERPRETATION & CONCLUSIONS: Our findins showed that opening of sluice gates of the bed-dam regularly once in a week resulted in the control of vector breeding in the downstream due to the flushing effect of the water released with a high flow from the bed-dam that stagnated water in the upstream. The outcome of the study encourages upscaling this measure to other areas, wherever feasible.

Comparison of efficacy of five types of long-lasting insecticidal nets against Anopheles fluviatilis, the primary malaria vector in east-central India.

Five types of long-lasting insecticidal nets (LNs), namely, Olyset, Netprotect, PermaNet, DuraNet, and Interceptor, were tested after 20 washes for efficacy in terms of mortality, deterrence effect, blood-feeding inhibition, and induced exophily of the malaria vector Anopheles fluviatilis in experimental huts in Malkangiri district of Odisha State, India. Efficacy of the three synthetic pyrethroids (SPs) used in the LNs was also analyzed. Use of LNs reduced the entry of An. fluviatilis into the huts by 73.3-83.2%, and the five LNs were comparable in terms of deterrence. The exit rate of An. fluviatilis from the huts with untreated net was 56.3%, and relative to this, Olyset followed by DuraNet induced significantly a higher exophily. In contrast, the exit rate was significantly lower with Interceptor. Among the three SPs, permethrin induced significantly greater exophily relative to the untreated control, and as a result of this, permethrin-treated Olyset produced a lower mortality. Blood-feeding rate of An. fluviatilis was significantly lower with all the five LNs than the control. Similarly, all the three SPs significantly inhibited blood feeding compared with the control. Interceptor and DuraNet, both alphacypermethrin-treated LNs, caused relatively a higher mortality of An. fluviatilis than the other LNs. The five brands of LNs and three SPs tested in the current study were equally effective in terms of deterrence and blood-feeding inhibition; only exiting and killing effect differed among them. Permethrin-treated LNs induced greater exophily, while, overall, alphacypermethrin-treated LNs killed more An. fluviatilis that entered the huts. Advantage of deterrence, excito-repellent, and killing effects of LNs and appropriate selection of SP for net treatment are discussed in this paper.

Response of malaria vectors to conventional insecticides in the southern districts of Odisha State, India.

BACKGROUND & OBJECTIVES: Updating information on response (susceptible / resistant status) of vectors to the insecticides in use is essential to formulate and introduce appropriate resistance management strategy. Therefore, a study was undertaken in the 10 southern districts of Odisha State, which are endemic for Plasmodium falciparum malaria, to determine the insecticide susceptibility/ resistance status of Anopheles fluviatilis and An. culicifacies, the vectors of malaria. METHODS: Mosquitoes were collected during September 2010 - February 2012 from 60 randomly selected villages in the 10 districts and blood-fed females were exposed to the diagnostic dosage of DDT (4.0%), malathion (5.0%) and deltamethrin (0.05%) for one hour. Mortality was recorded at 24 h after the exposure. The test mortality was corrected to the control mortality. RESULTS: An. fluviatilis was susceptible to the three insecticides tested while, An. culicifacies was resistant to DDT and malathion in all the 10 districts except in two, where its response against malathion was under 'verification required' category. Against deltamethrin, An. culicifacies was susceptible in two districts; while in the other eight districts its response was under 'verification required' category. INTERPRETATION & CONCLUSIONS: Since An. fluviatilis the vector species primarily associated with transmission of malaria, was still susceptible to DDT, indoor residual spraying with DDT could be continued in the 10 districts. Also, in view of the large scale implementation of long lasting insecticidal nets and the signs of development of resistance in An. culicifacies to deltamethrin, response of the vectors to synthetic pyrethroids needs to be periodically monitored.

Estimation of vectorial capacity of Anopheles minimus Theobald & An. fluviatilis James (Diptera: Culicidae) in a malaria endemic area of Odisha State, India.

BACKGROUND & OBJECTIVES: Anopheles minimus and An. fluviatilis were incriminated as the major malaria vectors in Keonjhar district of Odisha State recently. This study was carried out to elucidate the potential role of these two vector species in transmission of malaria during different seasons, and vectorial capacity of these species was also estimated. METHODS: Three hilly and forested villages of Keonjhar district were randomly selected. Vectorial capacity (C) was calculated using the Macdonald's formula as modified by Garret-Jones. The human landing density of the vector species was obtained from all night human landing collections (bait protected by bed-net). Man feeding habit was estimated by multiplying the human blood index with feeding frequency, which was obtained on daily basis from the duration of gonotrophic cycle. The probability of survival through the extrinsic incubation cycle was calculated from the probability of survival through one day and duration of sporogonic cycle. RESULTS: The estimated vectorial capacity of An. minimus varied between 0.014 and 1.09 for Plasmodium falciparum (Pf) and between 0.1 and 1.46 for P. vivax (Pv). The C of An. minimus for both Pf and Pv was higher during rainy season than the other two seasons. The estimated C of An. fluviatilis varied between 0.04 and 1.28 for Pf and between 0.20 and 1.54 for Pv. INTERPRETATION & CONCLUSIONS: Based on the estimated values of vectorial capacity of the two vector species, the area could be stratified and such stratification would reflect the difference in the intensity of transmission between different strata and accordingly the appropriate control strategy could be adopted for each stratum.

DNA Barcodes indicate members of the Anopheles fluviatilis (Diptera: Culicidae) species complex to be conspecific in India.

Anopheles fluviatilis, a major vector of malaria in India has been described as a complex of three sibling species members, named as S, T and U, based on variations in chromosomal inversions. Also, ribosomal DNA markers (repetitive Internal Transcribed Spacer 2 (ITS2) and 28S D3 region) were described to differentiate these three sibling species members. However, controversies prevail on the genetic isolation status of these cryptic species. Hence, we evaluated this taxonomic incongruence employing DNA barcoding, the well established methodology for species identification, using 60 An. fluviatilis sensu lato specimens, collected from two malaria endemic eastern states of India. These specimens were also subjected to sibling species characterization by ITS2 and D3 DNA markers. The former marker identified 31 specimens among these as An. fluviatilis S and 21 as An. fluviatilis T. Eight specimens amplified DNA fragments specific for both S and T. The D3 marker characterized 39 specimens belonging to species S and 21 to species T. Neither marker identified species U. Neighbor Joining analysis of mitochondrial cytochrome c oxidase gene 1 sequences (the DNA barcode) categorized all the 60 specimens into a single operational taxonomic unit, their Kimura 2 parameter (K2P) genetic variability being only 0.8%. The genetic differentiation (FST ) and gene flow (Nm ) estimates were 0.00799 and 62.07, respectively, indicating these two 'species' (S & T) as genetically con-specific intermixing populations with negligible genetic differentiation. Earlier investigations have refuted the existence of species U. Also, this study demonstrated that An. fluviatilis and the closely related An. minimus could be taxonomically differentiated by the DNA Barcode approach (K2P = 5.0%).

Seasonal prevalence & resting behaviour of Anopheles minimus Theobald & An. fluviatilis James (Diptera: Culicidae) in east-central India.

BACKGROUND & OBJECTIVES: Anopheles minimus has recently been reported to have re-appeared in Keonjhar district of Orissa after a period of about 45 years of launching the malaria eradication programme. An. minimus and An. fluviatilis were the incriminated major malaria vectors in the district, endemic for falciparum malaria. The information on seasonal prevalence and resting behaviour of the vectors is crucial for implementing appropriate malaria control measures. Therefore, a study was undertaken on seasonal prevalence and resting behaviour of An. minimus and An. fluviatilis in this district. METHODS: Seven randomly selected villages of Keonjhar district, Orissa, were studied during August 2005 to November 2007. Daytime resting collections indoors and outdoors were made covering three seasons of the year. The Anopheles mosquitoes obtained from different habitats were identified. Collections were maintained separately according to different sites as well as heights of the walls in human dwellings. RESULTS: Among the indoor collections, the densities of An. minimus and An. fluviatilis were higher in human dwellings than cattle sheds. An. fluviatilis was the predominant (41.5%) species followed by An. minimus (26.3%) in human dwellings. The density of both the vector species in human dwellings peaked during rainy and winter seasons followed by summer. Walls were the most preferred site by these vectors for resting and the maximum number was collected at a height of 3 to 4 ft. INTERPRETATION & CONCLUSIONS: The resting behaviour of the vector species increases their contact with the sprayed walls and therefore, a quality residual spraying of human dwellings focusing indoor walls could interrupt the malaria transmission in this area.

Glutathione S transferase activity in Indian vectors of malaria: A defense mechanism against DDT.

Glutathione S transferases (GSTs) are multifunctional enzymes involved in detoxification of xenobiotic compounds in majority of the insect groups. Significance of insect GSTs is their elevated level of activity in association with insecticide resistance. This investigation was to explore the metabolic status of GSTs in two Indian DDT-resistant malaria vectors, Anopheles culicifacies and Anopheles annularis, and one DDT-susceptible vector, Anopheles fluviatilis. Malkangiri and Koraput districts of Orissa State, endemic for falciparum malaria and having a long insecticide spraying history, were the study areas. F1 progeny was raised from wild-caught females of the three vectors and used for biochemical assays to detect the GST-mediated DDT resistance mechanism. Results of the enzyme assay showed a significant 3-fold increase in GST activity in DDT-resistant An. annularis compared with its susceptible population. In DDT-resistant An. culicifacies, the median GST activity (71.8 micromol/min/mg) was almost the same as estimated in the DDT-resistant An. annularis (74.6 micromol/ min/mg), suggesting that the GST activity estimated in An. culicifacies could be an elevated level for detoxification of DDT. Furthermore, the GST activity in DDT-resistant An. culicifacies and An. annularis was significantly higher than that in the DDT-susceptible An. fluviatilis, which had a GST activity of 20.0 micromol/min/mg. Also, the GST-mediated DDT detoxification was confirmed by comparing GST activity in wild-caught females with that in their F1 progeny.

Knowledge, attitude and practice on malaria: a study in a tribal belt of Orissa state, India with reference to use of long lasting treated mosquito nets.

Local knowledge and practice related to malaria is important for the implementation of culturally appropriate, sustainable and effective interventions. In this context, to know people's knowledge, attitude and practice on malaria and its prevention, a study was carried out in two districts viz., Malkangiri and Koraput of Orissa state in India, the former with ongoing insecticide treated mosquito nets (ITNs) programme and the latter without such programme (non-ITNs). Both qualitative and quantitative methods were used for data collection. The local concepts used for malaria describe only the biomedical symptoms of the disease although a few by meaning in local language reflect people's misconceptions about the cause of malaria. About 63% of the respondents mentioned mosquito bite as the cause for this disease and 65% considered malaria as a serious problem. Qualitative data showed that people from remote villages seek treatment from traditional healers, Disharis. About 64% of the respondents stated that avoiding mosquito bites could prevent malaria. Majority (99%) of the people reported using personal protection measures to avoid mosquito bites. Although, majority of the people were aware of the cause and prevention of malaria (about 70% stated sleeping under mosquito net prevents malaria), a sizable proportion still had misconceptions and hence appropriate communication strategies should be developed and imparted alongside ITNs/LLINs distribution for a behaviour change to adopt such preventive measures. Since, the tribes are habituated to seek treatment from traditional healers; they could be involved in motivating people to use ITNs/LLINs to protect from mosquito bites and malaria.

Acceptability, willing to purchase and use long lasting insecticide treated mosquito nets in Orissa State, India.

Long lasting insecticide treated nets (LLINs) that require no re-treatment have been advocated as an effective tool against malaria transmission. However, success of this community based intervention measure largely depends on its acceptability and proper usage by the target population, besides assuring access to bed nets. To determine the acceptability of LLIN, its usage and people's willingness to buy the net, a study was conducted in two tribal districts viz., Malkangiri (with ongoing ITN programme) and Koraput (no ITN programme) of Orissa State, India. Both qualitative and quantitative data collection methods were used to collect information regarding the objective of the study. A total of 2457 LLINs (Olyset Nets) were distributed in the selected villages of these districts at free of cost. In the study villages of Malkangiri, 58% of the households had either ITNs (73%) or other types of mosquito nets aside from the LLINs and in the villages of Koraput, only 8% had other nets, as majority (96%) informed that buying nets from market was not affordable to them. Physical verification of the nets during the house visits revealed that 75.4% and 83% (in ITNs and non-ITNs villages, respectively) of the LLINs and 76% of the other nets (including ITNs) were used by the respondents, the night before the survey as nets were in a hanging position at the time of the visit. Majority of the respondents (76-98%) felt that reduction of mosquito bites as the main perceived benefit of using the LLINs. About 55% and 67% of the respondents from non-ITNs and ITNs areas, respectively, expressed their willingness to buy the LLINs. Among them, 76.8% and 94.7% offered to pay INR<100 for a net and also ready to buy it by cash payment. Social marketing of LLINs at a subsidized price or free supply to the deserving sections of people (socially/economically poor and/or under-privileged) and ensuring the availability of nets during harvesting season could encourage people to buy and use LLINs.

Impact of lambdacyhalothrin capsule suspension treated bed nets on malaria in tribal villages of Malkangiri district, Orissa, India.

BACKGROUND & OBJECTIVE: Insecticide treated mosquito nets are increasingly being used in malaria control programmes. One of the problems with the treatment of bed nets with conventional formulations of insecticides was that regular washing of treated nets diminish insecticidal effect. Lambdacyhalothrin 2.5 capsule suspension (CS) (2.5% a.i., w/v), a new water-based microencapsulated formulation is reported to have wash-resistant property and longer persistence on the netting material than other formulations. We evaluated the impact of the use of nylon bed nets treated with lambdacyhalothrin 2.5 CS at 10 mg (a.i.)/m(2) in comparison to untreated nets and no nets on malaria in tribal villages in Orissa. METHODS: Nine foothill villages, highly endemic for falciparum malaria, from the Primary Health Centre (PHC) areas of Khairput and Kudumulugumma of Malkangiri district, Orissa, were divided into three groups, each with a population of about 500 and allocated randomly for treated (TN) and untreated nets (UN) and no nets (NN). Bed nets were distributed in September 2001 and retreatment was done in June 2002. The impact was assessed based on the changes in vector density, parous rate, malaria incidence and parasite rates. Indoor-resting collections of Anopheles fluviatilis and An. culicifacies were made at fortnightly intervals from fixed human dwellings. Mass blood surveys before and after distribution of nets and fortnightly active surveillance were carried out to assess the change in parasite rates and malaria incidence. Bioassays were conducted at fortnightly intervals on the bed nets supplied to the villagers. RESULTS: The reductions in indoor resting catches of An. fluviatilis and An. culicifacies were 96 and 38 per cent in villages with treated nets and 2.6 and 23 per cent in villages with untreated nets respectively compared to no net villages. For six months following treatment, 100 per cent mortality of An. fluviatilis was observed on the unwashed nets and on the nets washed once or twice. After re-treatment, 100 per cent mortality of An. fluviatilis or An. culicifacies was observed for nine months even after two washes. Usage rates of treated and untreated nets varied seasonally; 58.9 and 46.3 per cent in rainy season, 48.6 and 37.1 per cent in winter season and 38.1 and 31.6 per cent in summer season respectively. Reductions in malaria parasite rates were about 65 per cent in the treated net villages and 39 per cent in the untreated net villages compared to no net villages. About 75 per cent of treated nets and 60 per cent of untreated nets were in usable condition 19 months after distribution. INTERPRETATION & CONCLUSION: The estimated protection factor based on malaria incidence was 86 per cent for the treated nets during both post-treatment and post-retreatment periods and 34 and 51 per cent for untreated nets for the corresponding periods compared to no nets. The results of the study showed that the use of bed nets treated nets with CS formulation of lambdacyhalothrin at 10 mg (a.i.)/m(2) was acceptable to the community and re-treatment of nets at nine-monthly intervals can significantly reduce density and survival of An. fluviatilis and incidence of falciparum malaria.

Identification of Anopheles fauna in a hyperendemic falciparum area of Orissa State, India.

BACKGROUND & OBJECTIVE: Keonjhar district of Orissa State has been hyperendemic for falciparum malaria since many years with alarming deaths due to cerebral malaria. Therefore an entomological investigation to know more about the relative prevalence of Anopheles species was done. METHODS: Daytime indoor resting and outdoor resting, light trap and double bed net collections were made. Surveys were also made to collect Anopheles immature in streams and paddy fields. The Anopheles mosquitoes obtained by different catching methods were identified and the known vector species were subjected to gut and salivary gland dissection for vector incrimination. The infected specimens of An. fluviatilis and An. minimus were subjected to polymerase chain reaction assay for identification of sibling species. RESULTS: Of the anophelines collected, the most abundant was An. splendidus (18.2%) and An. fluviatilis (17.7%), followed by An. maculatus (14.0%) and An. minimus (9.0%). The sporozoite rate of An. fluviatilis and An. minimus was 0.9 and 1.4 respectively. The infected specimens have been identified as sibling species S of the An. fluviatilis complex and A of the An. minimus complex. INTERPRETATION & CONCLUSION: An. fluviatilis and An. minimus are the major two species in the transmission of malaria in Keonjhar district in Orissa.

Insecticide treated mosquito nets for malaria control in India-experience from a tribal area on operational feasibility and uptake

The study assessed the operational feasibility and acceptability of insecticide-treated mosquito nets (ITNs) in one Primary Health Centre (PHC) in a falciparum malaria endemic district in the state of Orissa, India, where 74 percent of the people are tribes and DDT indoor residual spraying had been withdrawn and ITNs introduced by the National Vector Borne Disease Control Programme. To a population of 63,920, 24,442 ITNs were distributed free of charge through 101 treatment centers during July-August 2002. Interview of 1,130, 1,012 and 126 respondents showed that the net use rates were 80 percent, 74 percent and 55 percent in the cold, rainy and summer seasons, respectively. Since using ITNs, 74.5-76.6 percent of the respondents observed reduction of mosquito bites and 7.2-32.1 percent reduction of malaria incidence; 37 percent expressed willingness to buy ITNs if the cost was lower and they were affordable. Up to ten months post-treatment, almost 100 percent mortality of vector mosquitoes was recorded on unwashed and washed nets (once or twice). Health workers re-treated the nets at the treatment centers eight months after distribution on a cost-recovery basis. The coverage reported by the PHC was only 4.2 percent, mainly because of unwillingness of the people to pay for re-treatment and to go to the treatment centers from their villages. When the re-treatment was continued at the villages involving personnel from several departments, the coverage improved to about 90 percent.Interview of 126 respondents showed that among those who got their nets re-treated, 81.4 percent paid cash for the re-treatment and the remainder were reluctant to pay. Majority of those who paid said that they did so due to the fear that if they did not do so they would lose benefits from other government welfare schemes. The 2nd re-treatment was therefore carried out free of charge nine months after the 1st re-treatment and thus achieved coverage of 70.4 percent. The study showed community...

Utility of the rDNA-ITS2-PCR assay in detecting the life stages of species S of Anopheles fluviatilis complex.

BACKGROUND & OBJECTIVE: Anopheles fluviatilis, which ranks second among the major malarial vectors in India occurs as a complex of three morphologically identical species (species S, T and U) of which only species S is a vector. Hence, it becomes pertinent to have a method for the detection of this vector species under field conditions to map the distribution of this vector. An rDNA-ITS2-PCR assay has been developed earlier for species S of this complex using female adult specimens. In order to widen the range of samples on which this technique can be employed, the utility of this PCR assay in detecting different life stages/gender/parts of the vector species was studied. Also, its reliability in detecting a single species S in pools of species T was studied. METHODS: Mosquitoes were collected from Malkangiri and Koraput districts of Orissa State where species S and T of this complex are reported. The wild caught fed females, after egg laying were subjected to PCR assay for species identification. The F1 progeny of a few PCR identified specimens was raised and samples at larval, pupal and adult stages were used for PCR assay. Single adult specimen of species S was added to pools containing different numbers of adults of species T and the pools were subjected to DNA extraction and PCR assay. RESULTS: The PCR assay could detect species S from pure DNA extracts of the immature stages and crude DNA extracts of parts of adult/whole adult mosquito of either gender. Crude DNA extracts of pools of mosquitoes had to be diluted and used in order to obtain the species diagnostic fragment. INTERPRETATION & CONCLUSION: The rDNA-ITS2-PCR assay producing an amplicon of 350 bp. diagnostic for species S, could detect all stages/gender. Any part of the adult can be used for species identification. Further, a single adult of species S in pools of as many as 99 adults of species T could be detected. Application of this PCR assay will be useful in mapping the distribution of species S, an important malarial vector.