Spatial and temporal variations of currently used pesticides (CUPs) concentrations in ambient air in Wallonia, Belgium.

Pesticides are among the most widely used chemicals thus contributing to a global contamination of the environment. Studies in North America and Europe have reported ambient air concentrations of Currently Used Pesticides in rural and urban locations as well as in remote areas. Monitoring pesticides in air is required for a better understanding of human exposure through inhalation and to assess potential health effects related to this exposure pathway. In this study, 46 pesticides were analyzed in ambient air in sampling stations distributed over Wallonia during a year, from May 2015 to May 2016. Different typologies were defined for sampling sites (remote areas, urban sites, agricultural sites, livestock area, and sites with other professional uses). Ambient air was sampled for 14 days with an active air sampler at a flow rate of 4 m³/h. Quartz filters and PUF/XAD-2/PUF cartridges were used to sample both gas and particulate phase pesticides. On the 46 pesticides studied, 6 insecticides, 18 herbicides and 18 fungicides were detected. Herbicides were measured in 68.3% of samples throughout the year, whereas fungicides and insecticides were measured in 62.6% and 13.2% of the samples, respectively. The highest mean concentrations for all pesticides were measured in spring-summer, whereas few pesticides were measured at low concentrations in winter. Six pesticides were measured in the remote sampling station at lower concentrations than in all other sites highlighting volatility of these pesticides. The highest number of different pesticides and the highest concentrations were measured in agricultural stations, where uses of plant protection products are higher. Finally, less volatile pesticides were only detected near application areas and at low concentrations. Together, these results provide better insight on the spatial and temporal variations of pesticides concentrations in ambient air, which were related to pesticides uses as well to atmospheric volatility and persistence.

PermaNet Dual, a new deltamethrin-chlorfenapyr mixture net, shows improved efficacy against pyrethroid-resistant Anopheles gambiae sensu lato in southern Benin.

Pyrethroid-chlorfenapyr nets have demonstrated improved entomological and epidemiological impact in trials across Africa. This is driving increased demand for this novel net class in malaria-endemic countries. PermaNet Dual is a new deltamethrin-chlorfenapyr net developed by Vestergaard Sàrl to provide more options to malaria control programmes. We performed an experimental hut trial to evaluate the efficacy of PermaNet Dual against wild, free-flying pyrethroid-resistant Anopheles gambiae sensu lato in Covè, Benin. PermaNet Dual induced superior levels of mosquito mortality compared to a pyrethroid-only net and a pyrethroid-piperonyl butoxide net both when unwashed (77% with PermaNet Dual vs. 23% with PermaNet 2.0 and 56% with PermaNet 3.0, p < 0.001) and after 20 standardised washes (75% with PermaNet Dual vs. 14% with PermaNet 2.0 and 30% with PermaNet 3.0, p < 0.001). Using a provisional non-inferiority margin defined by the World Health Organisation, PermaNet Dual was also non-inferior to a pyrethroid-chlorfenapyr net that has demonstrated improved public health value (Interceptor G2), for vector mortality (79% vs. 76%, OR = 0.878, 95% CIs 0.719-1.073) but not for blood-feeding protection (35% vs. 26%, OR = 1.424, 95% CIs 1.177-1.723). PermaNet Dual presents an additional option of this highly effective net class for improved control of malaria transmitted by pyrethroid-resistant mosquitoes.

Comparative functional survival and equivalent annual cost of 3 long-lasting insecticidal net (LLIN) products in Tanzania: A randomised trial with 3-year follow up.

BACKGROUND: Two billion long-lasting insecticidal nets (LLINs) have been procured for malaria control. A functional LLIN is one that is present, is in good physical condition, and remains insecticidal, thereby providing protection against vector-borne diseases through preventing bites and killing disease vectors. The World Health Organization (WHO) prequalifies LLINs that remain adequately insecticidal 3 years after deployment. Therefore, institutional buyers often assume that prequalified LLINs are functionally identical with a 3-year lifespan. We measured the lifespans of 3 LLIN products, and calculated their cost per year of functional life, to demonstrate the economic and public health importance of procuring the most cost-effective LLIN product based on its lifespan. METHODS AND FINDINGS: A randomised double-blinded trial of 3 pyrethroid LLIN products (10,571 nets in total) was conducted at 3 follow-up points: 10 months (August-October 2014), 22 months (August-October 2015), and 36 months (October-December 2016) among 3,393 households in Tanzania using WHO-recommended methods. Primary outcome was LLIN functional survival (LLIN present and in serviceable condition). Secondary outcomes were (1) bioefficacy and chemical content (residual insecticidal activity) and (2) protective efficacy for volunteers sleeping under the LLINs (bite reduction and mosquitoes killed). Median LLIN functional survival was significantly different between the 3 net products (p = 0.001): 2.0 years (95% CI 1.7-2.3) for Olyset, 2.5 years (95% CI 2.2-2.8) for PermaNet 2.0 (hazard ratio [HR] 0.73 [95% CI 0.64-0.85], p = 0.001), and 2.6 years (95% CI 2.3-2.8) for NetProtect (HR = 0.70 [95% CI 0.62-0.77], p < 0.001). Functional survival was affected by accumulation of holes, leading to users discarding nets. Protective efficacy also significantly differed between products as they aged. Equivalent annual cost varied between US$1.2 (95% CI $1.1-$1.4) and US$1.5 (95% CI $1.3-$1.7), assuming that each net was priced identically at US$3. The 2 longer-lived nets (PermaNet and NetProtect) were 20% cheaper than the shorter-lived product (Olyset). The trial was limited to only the most widely sold LLINs in Tanzania. Functional survival varies by country, so the single country setting is a limitation. CONCLUSIONS: These results suggest that LLIN functional survival is less than 3 years and differs substantially between products, and these differences strongly influence LLIN value for money. LLIN tendering processes should consider local expectations of cost per year of functional life and not unit price. As new LLIN products come on the market, especially those with new insecticides, it will be imperative to monitor their comparative durability to ensure that the most cost-effective products are procured for malaria control.

The durability of long-lasting insecticidal nets distributed to the households between 2009 and 2013 in Nepal.

BACKGROUND: Understanding and improving the durability of long-lasting insecticidal nets (LLINs) in the field are critical for planning future implementation strategies including behavioral change for care and maintenance. LLIN distribution at high coverage is considered to be one of the adjunctive transmission reduction strategies in Nepal's Malaria Strategic Plan 2014-2025. The main objective of this study was to assess the durability through assessment of community usage, physical integrity, residual bio-efficacy, and chemical retention in LLINs: Interceptor®, Yorkool®, and PermaNet ®2.0 which were used in Nepal during 2009 through 2013. METHODS: Assessments were conducted on random samples (n = 440) of LLINs from the eleven districts representing four ecological zones: Terai plain region (Kailali and Kanchanpur districts), outer Terai fluvial ecosystem (Surkhet, Dang, and Rupandhei districts), inner Terai forest ecosystem (Mahhothari, Dhanusa, and Illam districts), and Hills and river valley (Kavrepalanchock and Sindhupalchok districts). For each LLIN, fabric integrity in terms of proportionate hole index (pHI) and residual bio-efficacy were assessed. However, for chemical retention, a representative sample of 44 nets (15 Yorkool®, 10 Permanet®2.0, and 19 Interceptor®) was evaluated. Data were analyzed using descriptive statistics stratified by LLINs brand, districts, and duration of exposure. RESULTS: On average, duration of use of LLINs was shortest for the Yorkool® samples, followed by PermaNet® 2.0 and Interceptor® with median ages of 8.9 (IQR = 0.4), 23.8 (IQR = 3.2), and 50.1 (IQR = 3.2) months, respectively. Over 80% of field distributed Yorkool® and PermaNet® 2.0 nets were in good condition (pHI< 25) compared to Interceptor® (66%). Bio-efficacy analysis showed that average mortality rates of Interceptor and Yorkool were below World Health Organization (WHO) optimal effectiveness of ≥ 80% compared to 2-year-old PermaNet 2.0 which attained 80%. Chemical retention analysis was consistent with bio-efficacy results. CONCLUSION: This study shows that distribution of LLINs is effective for malaria control; however, serviceable life of LLINs should be considered in terms of waning residual bio-efficacy that warrants replacement. As an adjunctive malaria control tool, National Malaria Control Program of Nepal can benefit by renewing the distribution of LLINs in an appropriate time frame in addition to utilizing durable and effective LLINs.

Monitoring the physical and insecticidal durability of the long-lasting insecticidal net DawaPlus® 2.0 in three States in Nigeria.

BACKGROUND: Following guidance from the US President's Malaria Initiative, durability monitoring of DawaPlus® 2.0 brand of long-lasting insecticidal net (LLIN) distributed during the 2015/16 mass campaign was set up in three ecologically different states: Zamfara, Ebonyi and Oyo. METHODS: This was a prospective cohort study of representative samples of households from each location, recruited at baseline, 1 to 6 months after the mass campaign. All campaign nets in the households were labelled and followed up over a period of 36 months in Zamfara and Ebonyi and 24 months in Oyo. Primary outcome was the "proportion of nets surviving in serviceable condition" based on attrition and integrity measures and the median survival in years. The outcome for insecticidal durability was determined by bio-assay from sub-samples of campaign nets. RESULTS: A total of 439 households (98% of target) and 1096 campaign nets (106%) were included in the study. Definite outcomes could be determined for 92% of the cohort nets in Zamfara, 88% in Ebonyi and 75% in Oyo. All-cause attrition was highest in Oyo with 47% no longer present after 24 months, 53% in Ebonyi and 28% in Zamfara after 36 months. Overall only 1% of all campaign nets were used for other purposes. Estimated survival in serviceable condition of the campaign nets was 80% in Zamfara, 55% in Ebonyi (36 months follow-up) and 75% in Oyo (24 months follow-up) corresponding to median survival of 5.3, 3.3, 3.2 years, respectively. Factors associated with better survival were exposure to social messaging combined with a positive net-care attitude and only adult users. Failing to fold the net when hanging and having children under 5 years of age in the household negatively impacted net survival. Insecticidal effectiveness testing at final survey showed knock-down rates of 50-69%, but 24-h mortality above 95% resulting in 100% optimal performance in Ebonyi and Oyo and 97% in Zamfara. CONCLUSIONS: Results confirm the strong influence of net-use environment and behavioural factors in the physical survival of the same LLIN brand, which can increase the time until 50% of nets are no longer serviceable by up to 2 years.

Small-scale field testing of alpha-cypermethrin water-dispersible granules in comparison with the recommended wettable powder formulation for indoor residual spraying against malaria vectors in Benin.

BACKGROUND: Pyrethroids are the most common class of insecticide used worldwide for indoor residual spraying (IRS) against malaria vectors. Water-dispersible granules (WG) are a pyrethroid formulation to be applied after disintegration and dispersion in water with less risks of inhalation than using the usual wettable powder (WP) formulation. The objective of this small-scale field study was to evaluate efficacy and duration of insecticidal action of a new alpha-cypermethrin WG (250 g a.i./kg) against susceptible Anopheles gambiae in comparison with the WHO reference product (alpha-cypermethrin WP, 50 g a.i./kg) on the most common indoor surfaces in Benin. METHODS: Both formulations were applied at two target-dose concentrations in houses made of mud and cement in the Tokoli village in southern Benin. We measured the applied dose of insecticide by chemical analysis of filter paper samples collected from the sprayed inner walls. We recorded An. gambiae mortality and knock-down rates every 15 days during 6 months using standard WHO bioassays. RESULTS: The alpha-cypermethrin WG formulation did not last as long as the WP formulation on both surfaces. The difference is higher with the 30 mg/m2 concentration for which the WP formulation reached the 80% mortality threshold during 2 months on the mud-plastered walls (3 months on cement) whereas the WG formulation last only one month (2 months on cement). CONCLUSIONS: The new WG formulation has a shorter efficacy than the WHO recommended WP formulation. In this trial, both the WG and WP formulations had low durations of efficacy that would need at least two rounds of spray to cover the entire transmission season.

Author Correction: Contributions of cuticle permeability and enzyme detoxification to pyrethroid resistance in the major malaria vector Anopheles gambiae.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Contributions of cuticle permeability and enzyme detoxification to pyrethroid resistance in the major malaria vector Anopheles gambiae.

To tackle the problem of insecticide resistance, all resistance mechanisms need to be studied. This study investigated the involvement of the cuticle in pyrethroid resistance in a strain of Anopheles gambiae, MRS, free of kdr mutations. Bioassays revealed MRS to be resistant to pyrethroids and DDT, indicated by increasing knockdown times and resistance ratios. Moreover, biochemical analysis indicated that metabolic resistance based on enhanced CYP450 activity may also play a role. Insecticide penetration assays showed that there were significantly lower amounts of insecticide in the MRS strain than in the susceptible control. Analysis of the levels of the selected transcripts by qPCR showed that CYP6M2, a major pyrethroid metaboliser, CYP4G16, a gene implicated in resistance via its contribution to the biosynthesis of elevated epicuticular hydrocarbons that delay insecticide uptake, and the cuticle genes CPAP3-E and CPLCX1 were upregulated after insecticide exposure. Other metabolic (CYP6P3, GSTe2) and cuticle (CPLCG3, CPRs) genes were also constitutively upregulated. Microscopic analysis showed that the cuticle layers of the MRS strain were significantly thicker than those of the susceptible strain. This study allowed us to assess the contribution made by the cuticle and metabolic mechanisms to pyrethroid resistance in Anopheles gambiae without target-site mutations.

An experimental hut evaluation of Olyset Plus, a long-lasting insecticidal net treated with a mixture of permethrin and piperonyl butoxide, against Anopheles fluviatilis in Odisha State, India.

BACKGROUND: Fast development of pyrethroid resistance in malaria vectors prompted the development of new vector control tools including combination of insecticides with different modes of action as part of resistance management strategies. Olyset Plus® is a new long-lasting insecticidal net, in which, permethrin and a synergist, piperonyl butoxide (PBO), are incorporated into filaments. Mixture nets such as this may have application against resistant mosquitoes, particularly those whose resistance is based on oxidative metabolism. There may also be enhanced activity against susceptible mosquitoes since mixed function oxidases are involved in a many metabolic activities including activation to form bioactive compounds. METHODS: Bio-efficacy of Olyset Plus was evaluated against susceptible malaria vector, Anopheles fluviatilis in experimental huts. Deterrence, blood feeding inhibition, induced exophily and killing effect were measured to assess the bio-efficacy. The results were compared with Olyset Net®, a polyethylene permethrin-incorporated LLIN and a conventionally treated polyester net (with permethrin) washed to just before exhaustion. RESULTS: Results showed significant reduction in entry (treatment: 0.4-0.8; control: 4.2 per trap-night) and increase in exit (56.3-82.9 % and 44.2 %) rates of Anopheles fluviatilis in the treatment arms compared to control (P < 0.05). While blood feeding rates declined in treatment arms (18.8-30.6 %), it increased in control (77.6 %) (P < 0.05). This was further evident from the blood-feeding inhibition rates in treatment arms (60.6-90.6 %). Total mortality was significantly higher in all treatment arms (96.3-100 %) compared to control arm (2 %) (P < 0.05). Chemical analysis for active ingredient (AI) showed retention of 75 and 88 % in Olyset plus and Olyset net respectively after 20 washes. Performance of Olyset Plus washed 20 times was equal to the CTN and Olyset Net against the susceptible malaria vector An. fluviatilis, fulfilling the WHO efficacy criteria of Phase II evaluation for LLIN. However, the benefit of incorporating PBO and permethrin together in a long-lasting treatment could not be demonstrated in the current study as the target vector species was fully susceptible to pyrethroids. CONCLUSION: Olyset Plus, with its intrinsic bio-efficacy could be an effective vector control tool to prevent transmission of malaria by susceptible vectors like An. fluviatilis. However, the results of the current study need to be further supported by testing the net at village level (Phase III) for community acceptability. Before taking the net to village level, it needs to be verified whether the net is better than pyrethroid nets in terms of bio-efficacy against resistant An. culicifacies, another malaria vector that has developed resistance to synthetic pyrethroids in India.

Interceptor® long-lasting insecticidal net: phase III evaluation over three years of household use and calibration with Phase II experimental hut outcomes.

BACKGROUND: Long-lasting insecticidal nets (LN) are an effective tool for malaria prevention. The World Health Organization Pesticide Evaluation Scheme has established evaluation criteria to facilitate registration for public use. A household randomised trial was conducted in Tanzania according to WHOPES Phase III procedures to evaluate the alpha-cypermethrin coated Interceptor® LN (BASF) over three years' use. Outcomes were calibrated against results of Phase II experimental hut trials. METHODS: Interceptor LN (200 mg/m(2) alpha-cypermethrin) and conventionally treated nets CTN (40 mg/m(2) alpha-cypermethrin) were randomly distributed to 934 households. At 6-monthly intervals, household surveys recorded net use, durability, adverse effects, user acceptance and washing practices. Concurrently, 30 nets of each type were collected and tested for knock-down and kill of Anopheles gambiae mosquitoes in cone and tunnel bioassays. Alpha-cypermethrin content of nets was assessed annually. RESULTS: At 12 months 97% of Interceptor LN met the efficacy criteria by cone or tunnel test; this pass rate declined to 90% at 24 months and 87% at 36 months. In contrast only 63% of CTN met the efficacy criteria at 12 months, 14 % at 24 months and 0% at 36 months. The alpha-cypermethrin content at 36 months on Interceptor LN was 20% (42 ± 13 mg/m(2)) of the initial content but on CTNs only 4% (1.3 ± 1.6 mg/m(2)) remained. Interceptor LN was reported to be used year-round and washed 4.3 times/year. A few recalled facial tingling during the first days of use but this did not deter usage. The average number of holes at 36 months was 18, hole area per net was 229 cm(2) and hole index was 332. Insecticide content and cone bioefficacy of LN and CTN after 36 months' use were similar to that of LN and CTN used in earlier Phase II hut trials, but while the 20 times washed LN tested in experimental huts gave adequate personal protection the 20 times washed CTN did not. CONCLUSIONS: More than 80% Interceptor LN fulfilled the WHOPES Phase III criteria at 36 months and thus the LLIN was granted full WHO recommendation. Phase III outcomes at 36 months were anticipated by Phase II outcomes after 20 standardized washes.

Durability of Olyset campaign nets distributed between 2009 and 2011 in eight districts of Tanzania.

BACKGROUND: Long-lasting insecticidal nets (LLINs) are the first line choice for malaria vector control in sub-Saharan Africa, with most countries adopting universal coverage campaigns. However, there is only limited information on LLIN durability under user conditions. Therefore, this study aimed to assess the durability of Olyset(®) LLINs distributed during campaigns between 2009 and 2011 in Tanzania. METHODS: A retrospective field survey was conducted in eight districts in Tanzania mainland to assess the durability of Olyset campaign nets. Household questionnaires were used to assess attrition, i.e. net loss. All nets remaining in households were collected. A sub-sample of 198 Olyset campaign nets was examined for bio-efficacy against Anopheles gambiae s.s. mosquitoes, permethrin content and physical integrity following standard World Health Organization (WHO) methods. RESULTS: Of 6067 campaign nets reported to have been received between 2009 and 2011, 35% (2145 nets) were no longer present. Most of those nets had been discarded (84%) mainly because they were too torn (94%). Of the 198 sub-sampled Olyset LLINs, 61% were still in serviceable physical condition sufficient to provide personal protection while 39% were in unserviceable physical condition according to WHO proportionate Hole Index (pHI). More than 96% (116/120) of nets in serviceable condition passed WHO bioefficacy criteria while all nets in unserviceable condition passed WHO bioefficacy criteria. Overall mean permethrin content was 16.5 g/kg (95% CI 16.2-16.9) with 78% of the sub-sampled nets retaining recommended permethrin content regardless of their age or physical condition. Nets aged 4 years and above had a mean permethrin content of 14 g/kg (95% CI 12.0-16.0). The only statistically significant predictor of reduced physical net integrity was rats in the house. CONCLUSIONS: Two-to-four years after a mass campaign, only 39% of distributed nets remain both present and in serviceable physical condition, a functional survival considerably below WHO assumptions of 50% survival of a 'three-year' net. However, the majority of nets still retained substantial levels of permethrin and could still be bio-chemically useful against mosquitoes if their holes were repaired, adding evidence to the value of net care and repair campaigns.

A phase III trial to evaluate the efficacy, fabric integrity and community acceptance of Netprotect using a recommended long-lasting insecticidal net as positive control.

BACKGROUND: The evaluation of new long-lasting insecticidal bed nets (LLINs) is coordinated by the WHO Pesticide Evaluation Scheme (WHOPES). In 2007, Netprotect® was granted WHOPES interim recommendation after Phase I and II evaluations. Present study evaluates Netprotect in a Phase III trial in rural Cambodia. METHODS: A randomized, prospective longitudinal study design was used to assess the performance of Netprotect over a period of three years, using conventionally-treated nets and a WHOPES recommended LLIN (PermaNet 2.0) as positive controls. The primary outcomes were the physical integrity, insecticide content and cone bioassay performance using. RESULTS: The baseline deltamethrin concentration of 43% of Netprotect nets were below the tolerance limit while 27% of PermaNet 2.0 nets were above the target dose limits. By 36 months Netprotect retained 35% while PermaNet 2.0 retained 49% of baseline insecticide dose. Moreover the proportion of the inactive deltamethrin R-alpha isomer in the Netprotect nets was 33% at the baseline and increased to 69% after three years while it was low and almost constant for PermaNet® 2.0 (3-7%). Only 71% of Netprotect met the WHO criteria for bio-efficacy after three years while at least 80% is required. Moreover Netprotect nets failed for the WHOPES criteria after 12 and 24 months. The reference LLIN met the WHOPES criteria throughout the study. Over the entire three years the reference LLIN did obtain significant higher mosquito mortality than Netprotect. The physical integrity was based on the proportionate hole index and after three years, 25% of Netprotect and 30% of PermaNet 2.0 were in a mediocre or poor state. CONCLUSION: Netprotect did not meet the minimum WHO criteria for bio-efficacy after 12, 24 and 36 months. The use of a reference LLIN as positive control was helpful for data interpretation. However, for future three-year studies, it is essential that before initiating any study nets should be checked for their specifications and this for both the candidate LLIN as well as for the reference LLIN. Moreover, to improve the accuracy of the success rate of the candidate LLIN more nets should be tested for their bio-efficacy at the end of the trial.

Comparison of the laboratory standard washing using CIPAC washing agent and the domestic washing on three recommended types of long-lasting insecticidal mosquito nets.

BACKGROUND: One of the best ways to prevent malaria is the use of insecticide-treated bed nets. Manufacturers pursue easier, safer and more efficient nets. Hence, many studies on the efficacy and wash resistance using World Health Organization standards have been reported. The commonly used detergent is "Savon de Marseille", because it closely resembles actually used soaps. At the 54(th) Collaborative International Pesticides Analytical Council (CIPAC) Technical Meeting in 2010, it was suggested to replace it by a standardized "CIPAC washing agent". The aim of this study was to investigate the difference between a laboratory hand washing simulation using the CIPAC washing agent (method-1) and a domestic washing (method-2) on different bed nets, as well as the effect of the drying process on the release of active ingredient. METHODS: Interceptor®, Permanet®2.0 and Netprotect® nets were used in three treatments, each repeated 20 times. The first treatment included method-1 washing and indoor drying. The second treatment included method-2 washing and indoor drying. The third treatment used method-2 washing and UV-drying. The residual insecticide contents were determined using gas chromatography. RESULTS: The washing procedure and the number of washes have a significant effect on the release of active ingredient. Statistically, the two washing methods have the same effect on removing the active ingredient from the Interceptor® and Permanet®2.0 net, but a significantly different influence on the Netprotect® nets. The drying process has no significant effect on the insecticide. CONCLUSION: Both washing procedures affected the amount of insecticide remaining on nets independently of the impregnation technology. The active ingredient decreases with the number of washing cycles following an exponential or logarithmic model for coated nets. The laboratory hand washing simulation had more impact on the decrease of active ingredient content of the Netprotect® nets. All net types seemed to be effectively protected against UV-light.

Efficacy of Olyset® Plus, a new long-lasting insecticidal net incorporating permethrin and piperonyl-butoxide against multi-resistant malaria vectors [corrected].

Due to the rapid extension of pyrethroid resistance in malaria vectors worldwide, manufacturers are developing new vector control tools including insecticide mixtures containing at least two active ingredients with different mode of action as part of insecticide resistance management. Olyset® Plus is a new long-lasting insecticidal net (LLIN) incorporating permethrin and a synergist, piperonyl butoxide (PBO), into its fibres in order to counteract metabolic-based pyrethroid resistance of mosquitoes. In this study, we evaluated the efficacy of Olyset® Plus both in laboratory and field against susceptible and multi-resistant malaria vectors and compared with Olyset Net, which is a permethrin incorporated into polyethylene net. In laboratory, Olyset® Plus performed better than Olyset® Net against susceptible Anopheles gambiae strain with a 2-day regeneration time owing to an improved permethrin bleeding rate with the new incorporation technology. It also performed better than Olyset® Net against multiple resistant populations of An. gambiae in experimental hut trials in West Africa. Moreover, the present study showed evidence for a benefit of incorporating a synergist, PBO, with a pyrethroid insecticide into mosquito netting. These results need to be further validated in a large-scale field trial to assess the durability and acceptability of this new tool for malaria vector control.

Validation of a multi-residue method to determine deltamethrin and alpha-cypermethrin in mosquito nets by gas chromatography with electron capture detection (GC-µECD).

BACKGROUND: Nowadays long-lasting insecticidal mosquito nets (LNs) are frequently used around the world to protect people against malaria vectors. As they contain insecticide, laboratory control is needed to check whether the content of the active ingredient follows the conditions of the manufacturer and also if the active ingredient is still present after some time of use. For this purpose, an analytical method had to be developed. The fact that LNs include a range of polymers for the yarn and use coated or incorporated technologies for the active ingredient, it is a challenge to find only one analytical method determining the active ingredient in LNs, which takes into account both impregnation technologies. Some methods are provided by international organizations but are limited by the determination of only one pesticide per method. The aim of this study was to optimize a short time extraction method for deltamethrin and alpha-cypermethrin from coated and incorporated mosquito nets and also to detect both insecticides in one analytical run, using gas chromatography with electron capture detection (GC-µECD). METHODS: Based on the literature, the most suitable solvent and the adequate extraction process for the insecticides used for net making were identified and adapted for the new multi-residue method. RESULTS: The validation data of the multi-residue method to determine deltamethrin and alpha-cypermethrin in mosquito nets by GC-µECD are given. Depending on the concentration of the active ingredient spiked on the nets, the mean recovery for alpha-cypermethrin ranged between 86% and 107% with a relative standard deviation below 3.5%. For deltamethrin it ranged between 90% and 108% with a relative standard deviation also below 3.5%. The limit of detection is 0.009 g.a.i/kg of net (0.3 mg a.i./m2 of net) both for alpha-cypermethrin and deltamethrin. CONCLUSIONS: Data obtained are excellent. A 30 minutes reflux extraction method with xylene was developed to determine alpha-cypermethrin and deltamethrin in long-lasting insecticidal mosquito nets (LNs) by gas chromatography with electron capture detection (GC-µECD). The method can be easily extended to others pyrethroid used for mosquito net treatment. This paper also presents an overview of the studies dealing with pesticide determination in mosquito nets.

Evidence for a useful life of more than three years for a polyester-based long-lasting insecticidal mosquito net in Western Uganda.

BACKGROUND: Long-lasting insecticidal nets (LLIN) are now standard for the prevention of malaria. However, only products with recommendation for public use from the World Health Organization should be used and this evaluation includes the assessment of net effectiveness after three years of field use. Results for one of the polyester-based products, Interceptor is presented. METHODS: In five villages, 190 LLIN and 90 nets conventionally treated with the insecticide alpha-cypermethrin at 25 mg/m2 were distributed randomly and used by the families. Following a baseline household survey a net survey was carried out every six months to capture use, washing habits and physical condition of the nets. Randomly selected nets were collected after 6, 12, 24, 36 and 42 months and tested for remaining insecticide content and ability to knock-down and kill malaria transmitting mosquitoes. RESULTS: During the three and a half years of observation only 16 nets were lost to follow-up resulting in an estimated attrition rate of 12% after three and 20/% after 3.5 years. Nets were used regularly and washed on average 1.5 times per year. After three and a half years 29% of the nets were still in good condition while 13% were seriously torn with no difference between the LLIN and control nets. The conventionally treated nets quickly lost insecticide and after 24 months only 7% of the original dose remained (1.6 mg/m2). Baseline median concentration of alpha-cypermethrin for LLIN was 194.5 mg/m2 or 97% of the target dose with between and within net variation of 11% and 4% respectively (relative standard deviation). On the LLIN 73.8 mg/m2 alpha-cypermethrin remained after three years of use and 56.2 mg/m2 after three and a half and 94% and 81% of the LLIN still had > 15 mg/m2 left respectively. Optimal effectiveness in bio-assays (≥ 95% 60 minute knock-down or ≥ 80% 24 hour mortality) was found in 83% of the sampled LLIN after three and 71% after three and a half years. CONCLUSIONS: Under conditions in Western Uganda the tested long-lasting insecticidal net Interceptor fulfilled the criteria for phase III of WHO evaluations and, based on preliminary criteria of the useful life, this product is estimated to last on average between three and four years.

An experimental hut evaluation of PermaNet(®) 3.0, a deltamethrin-piperonyl butoxide combination net, against pyrethroid-resistant Anopheles gambiae and Culex quinquefasciatus mosquitoes in southern Benin.

PermaNet 3.0 is a long-lasting combination net with deltamethrin present on the sides and a mixture of deltamethrin and piperonyl butoxide (PBO), an oxidase synergist, on the top panel. An experimental hut trial comparing unwashed and 20 times washed PermaNet 3.0 and PermaNet 2.0, Olyset Net and a conventional deltamethrin-treated net washed three times was conducted in southern Benin. Anopheles gambiae and Culex quinquefasciatus from this area are highly resistant to pyrethroids through kdr and cytochrome P450 mechanisms. The unwashed PermaNet 3.0 killed slightly more A. gambiae (52%) than the unwashed PermaNet 2.0 (44%) (P=0.036), indicating only partial synergism of resistance. After washing there was significant loss of activity to a similar level, with PermaNet 3.0 killing 31%, PermaNet 2.0 killing 29% and the conventional net killing 26%. Blood-feeding rates were partially inhibited for unwashed PermaNet 3.0 and Olyset Net (27% inhibition). Personal protection against A. gambiae derived from PermaNet 3.0 was similar to that from PermaNet 2.0 before washing (50% vs. 47%), and after 20 washes it decreased to 30%. Against C. quinquefasciatus, no treatment killed >24% entering the huts. The synergism from unwashed PermaNet 3.0 was lower than expected, probably due to an unidentified resistance mechanism unaffected by PBO.

Field efficacy of a new mosaic long-lasting mosquito net (PermaNet 3.0) against pyrethroid-resistant malaria vectors: a multi centre study in Western and Central Africa.

BACKGROUND: Due to the spread of pyrethroid-resistance in malaria vectors in Africa, new strategies and tools are urgently needed to better control malaria transmission. The aim of this study was to evaluate the performances of a new mosaic long-lasting insecticidal net (LLIN), i.e. PermaNet 3.0, against wild pyrethroid-resistant Anopheles gambiae s.l. in West and Central Africa. METHODS: A multi centre experimental hut trial was conducted in Malanville (Benin), Vallée du Kou (Burkina Faso) and Pitoa (Cameroon) to investigate the exophily, blood feeding inhibition and mortality induced by PermaNet 3.0 (i.e. a mosaic net containing piperonyl butoxide and deltamethrin on the roof) comparatively to the WHO recommended PermaNet 2.0 (unwashed and washed 20-times) and a conventionally deltamethrin-treated net (CTN). RESULTS: The personal protection and insecticidal activity of PermaNet 3.0 and PermaNet 2.0 were excellent (>80%) in the "pyrethroid-tolerant" area of Malanville. In the pyrethroid-resistance areas of Pitoa (metabolic resistance) and Vallée du Kou (presence of the L1014F kdr mutation), PermaNet 3.0 showed equal or better performances than PermaNet 2.0. It should be noted however that the deltamethrin content on PermaNet 3.0 was up to twice higher than that of PermaNet 2.0. Significant reduction of efficacy of both LLIN was noted after 20 washes although PermaNet 3.0 still fulfilled the WHO requirement for LLIN. CONCLUSION: The use of combination nets for malaria control offers promising prospects. However, further investigations are needed to demonstrate the benefits of using PermaNet 3.0 for the control of pyrethroid resistant mosquito populations in Africa.

Evaluation of PermaNet 3.0 a deltamethrin-PBO combination net against Anopheles gambiae and pyrethroid resistant Culex quinquefasciatus mosquitoes: an experimental hut trial in Tanzania.

BACKGROUND: Combination mosquito nets incorporating two unrelated insecticides or insecticide plus synergist are designed to control insecticide resistant mosquitoes. PermaNet 3.0 is a long-lasting combination net incorporating deltamethrin on the side panels and a mixture of deltamethrin and synergist piperonyl butoxide (PBO) on the top panel. PBO is an inhibitor of mixed function oxidases implicated in pyrethroid resistance. METHOD: An experimental hut trial comparing PermaNet 3.0, PermaNet 2.0 and a conventional deltamethrin-treated net was conducted in NE Tanzania using standard WHOPES procedures. The PermaNet arms included unwashed nets and nets washed 20 times. PermaNet 2.0 is a long-lasting insecticidal net incorporating deltamethrin as a single active. RESULTS: Against pyrethroid susceptible Anopheles gambiae the unwashed PermaNet 3.0 showed no difference to unwashed PermaNet 2.0 in terms of mortality (95% killed), but showed differences in blood-feeding rate (3% blood-fed with PermaNet 3.0 versus 10% with PermaNet 2.0). After 20 washes the two products showed no difference in feeding rate (10% with 3.0 and 9% with 2.0) but showed small differences in mortality (95% with 3.0 and 87% with 2.0). Against pyrethroid resistant Culex quinquefasciatus, mediated by elevated oxidase and kdr mechanisms, the unwashed PermaNet 3.0 killed 48% and PermaNet 2.0 killed 32% but after 20 washes there was no significant difference in mortality between the two products (32% killed by 3.0 and 30% by 2.0). For protecting against Culex PermaNet 3.0 showed no difference to PermaNet 2.0 when either unwashed or after 20 washes; both products were highly protective against biting. Laboratory tunnel bioassays confirmed the loss of biological activity of the PBO/deltamethrin-treated panel after washing. CONCLUSION: Both PermaNet products were highly effective against susceptible Anopheles gambiae. As a long-lasting net to control or protect against pyrethroid resistant mosquitoes PermaNet 3.0 showed limited improvement over PermaNet 2.0 against Culex quinquefasciatus.

Median knock-down time as a new method for evaluating insecticide-treated textiles for mosquito control.

BACKGROUND: Insecticide treated bed nets are major tools for the Roll Back Malaria campaign. There are two types of Long-Lasting Insecticide-treated Nets (LNs) on the market: coated nets and insecticide-incorporated nets. Nets provided to this market need a recommendation from the World Health Organization to be purchased by donors and NGOs. During laboratory study (phase I), the first step consists in evaluating the wash resistance of a new LN product. When insecticide-incorporated nets are washed, it takes time to regenerate the insecticidal activity, i.e. insecticide must migrate to the net surface to be accessible to mosquitoes. The interval of time required for regeneration must be carefully determined to ensure the accuracy of further results. WHOPES procedures currently recommend the determination of the regeneration time by using mortality data. However, as mortality cannot exceed 100%, a LN that regenerates a surface concentration exceeding the dosage for 100% mortality, will have its regeneration time underestimated. METHODS: The Median Knock Down Time (MKDT) was determined as function of insecticide dosage on an inert surface, glass, and on polyester nettings using an acetone solution or a simple emulsion. Dosage response was also established for mortality data. The same method was then applied to a commercially polyethylene netting, currently under WHOPES evaluation, to determine the dynamics of regeneration as function of repeated washings. The deltamethrin content of these nets was estimated by Capillary Gas Chromatography (GC-ECD). RESULTS: MKDT was a linear function of log insecticide dosage on glass as on nettings. Mortality data were either 0 or 100% for most concentrations except for a narrow range. MKDT was log linear function of total deltamethrin content in a commercial polyethylene net exposed to washings. The regeneration time of this net increased with the number of washes and MKDT became higher. A new, easy and rapid method to determine MKDT is suggested. DISCUSSION: The MKDT is linearly correlated to log dosage on a given substrate and shows no saturation as mortality data do. It is suited to determine regeneration time of a product that is exposed to a stress, like washing or heating, where the process impacts on the bio-availability of the insecticide. Mortality data are useful for measuring product efficacy, whereas MKDT are better to measure dynamics of surface concentration like regeneration after a stressing process. Change in MKDT can be used to illustrate the loss of insecticide due to washing, but the slope of the curve is product and surface-dependent.