Long-lasting control of by a single spray application of micro … · 2017-04-05 · RESEARCH Open...

Long-lasting control of Anopheles arabiensis by asingle spray application of micro-encapsulatedpirimiphos-methyl (Actellicreg 300 CS)Oxborough et al

Oxborough et al Malaria Journal 2014 1337httpwwwmalariajournalcomcontent13137


RESEARCH Open Access

Long-lasting control of Anopheles arabiensis by asingle spray application of micro-encapsulatedpirimiphos-methyl (Actellicreg 300 CS)Richard M Oxborough123 Jovin Kitau23 Rebecca Jones1 Emmanuel Feston2 Johnson Matowo23Franklin W Mosha23 and Mark W Rowland13

Abstract

Background Pyrethroid-resistant mosquitoes are an increasing threat to malaria vector control The Global Plan forInsecticide Resistance Management (GPIRM) recommends rotation of non-pyrethroid insecticides for indoor residualspraying (IRS) The options from other classes are limited The carbamate bendiocarb and the organophosphatepirimiphos-methyl (p-methyl) emulsifiable concentrate (EC) have a short residual duration of action resulting inincreased costs due to multiple spray cycles and user fatigue Encapsulation (CS) technology was used to extendthe residual performance of p-methyl

Methods Two novel p-methyl CS formulations were evaluated alongside the existing EC in laboratory bioassaysand experimental hut trials in Tanzania between 2008-2010 Bioassays were carried out monthly on sprayedsubstrates of mud concrete plywood and palm thatch to assess residual activity Experimental huts were used toassess efficacy against wild free-flying Anopheles arabiensis in terms of insecticide-induced mortality andblood-feeding inhibition

Results In laboratory bioassays of An arabiensis and Culex quinquefasciatus both CS formulations produced highrates of mortality for significantly longer than the EC formulation on all substrates On mud the best performing CSkilled gt80 of An arabiensis for five months and gt50 for eight months compared with one and two monthsrespectively for the EC In monthly bioassays of experimental hut walls the EC was ineffective shortly after sprayingwhile the best CS formulation killed more than 80 of An arabiensis for five months on mud and seven monthson concrete In experimental huts both CS and EC formulations killed high proportions of free-flying wild An arabiensisfor up to 12 months after spraying There was no significant difference between treatments All treatmentsprovided considerable personal protection with blood-feeding inhibition ranging from 9-49 over time

Conclusions The long residual performance of p-methyl CS was consistent in bioassays and experimental hutsThe CS outperformed the EC in laboratory and hut bioassays but the EC longevity in huts was unexpectedLong-lasting p-methyl CS formulations should be more effective than both p-methyl EC and bendiocarbconsidering a single spray could be sufficient for annual malaria control IRS with p-methyl 300 CS is a timelyaddition to the limited portfolio of long-lasting residual insecticides

Keywords IRS Pirimiphos-methyl Actellic Anopheles arabiensis Vector control Malaria Tanzania

Correspondence oxandbullhotmailcom1Department of Disease Control London School of Hygiene and TropicalMedicine (LSHTM) London UK2Department of Entomology and Parasitology Kilimanjaro Christian MedicalUniversity College (KCMUCo) of Tumaini University Moshi KilimanjaroTanzaniaFull list of author information is available at the end of the article

copy 2014 Oxborough et al licensee BioMed Central Ltd This is an Open Access article distributed under the terms of theCreative Commons Attribution License (httpcreativecommonsorglicensesby20) which permits unrestricted usedistribution and reproduction in any medium provided the original work is properly credited The Creative Commons PublicDomain Dedication waiver (httpcreativecommonsorgpublicdomainzero10) applies to the data made available in thisarticle unless otherwise stated

Oxborough et al Malaria Journal 2014 1337 Page 2 of 15httpwwwmalariajournalcomcontent13137

BackgroundIndoor residual spraying (IRS) has produced profoundchanges in malaria burden in a range of settings withseveral different insecticide classes [1] Interruption ofmalaria transmission in the USA was achieved partlythrough DDT house-spraying and led to the initiationof the World Health Organization (WHO)-led GlobalMalaria Eradication Scheme (1955-1969) [2] Malariawas subsequently eradicated from Europe parts of theSoviet Union Israel Lebanon Syria Japan and ChineseTaiwan Despite numerous positive outcomes the bene-fits were not on the global scale that was anticipatedThere were about 20 pilot IRS projects in sub-SaharanAfrica between the mid 1950s and early 1960s [3] thatdemonstrated IRS significantly reduced malaria trans-mission even in highly endemic (intense transmission)areas [4] Despite this Africa was largely sidelined foreradication due to the high malaria burden while else-where dramatic reversals were seen once IRS sprayingwas prematurely reduced in countries such as India andSri Lanka [56] As a result interest in IRS subsequentlywaned and was not taken to scale in most sub-Saharanmalaria-endemic countries as part of the global eradicationcampaign [47]Southern Africa was the exception IRS programmes

using DDT began in the 1960s and were supported forseveral decades with later introduction of pyrethroidsand carbamates Countries with sustained IRS activitiesin Africa including South Africa Zambia NamibiaSwaziland Zimbabwe and Botswana achieved sizeablereductions in malaria vector populations and malariaincidence [7] Focal IRS in the southern Africa regionhas remained important in areas of higher malaria bur-den and at risk of epidemics In 2007 about 14 millionpeople in southern Africa were protected by IRS [47]In 2006 WHO reaffirmed the importance of IRS as a

primary intervention for reducing or interrupting mal-aria transmission [89] In recent years an unprecedentedlevel of funding has initiated new IRS campaigns acrosssub-Saharan Africa often in parallel with long-lastinginsecticide-treated bed nets (LLIN) distribution In 2012Presidentrsquos Malaria Initiative (PMI) supported IRS in15 African countries covering seven million structures[10] The implementation of new IRS programmestogether with sustained IRS programmes in SouthernAfrica has elevated the importance of IRS as a primaryintervention for malaria control in Africa Greater em-phasis has been placed on ensuring that IRS in Africacan be sustained [11]Pyrethroids are the only group of insecticides approved

by WHO Pesticide Evaluation Scheme (WHOPES) forLLINs [12] Pyrethroid insecticides have also been pre-ferred for IRS in Africa in recent years due to low costlongevity of three to six months and low mammalian and

non-target toxicity [13] Subsequently pyrethroid resist-ance has become widespread in malaria vectors acrossAfrica [14] Reduced efficacy of insecticide interventionsin areas of pyrethroid resistant malaria vectors has beendemonstrated in several settings A notable example is inSouth Africa where four years after the introduction ofdeltamethrin IRS a four-fold increase in malaria cases wasrecorded in Kwa-Zulu Natal coinciding with re-invasionof pyrethroid resistant Anopheles funestus ss This trendwas reversed after re-introduction of IRS with DDT in2000 and new introduction of artemisinin-based com-bination therapy in 2001 with an accompanied declinein malaria cases by 91 [15] In Bioko Island EquatorialGuinea a single spray round with pyrethroid failed to re-duce the population density of pyrethroid-resistantAnopheles gambiae ss Subsequent spraying of a carba-mate significantly reduced the number of An gambiaess caught exiting in window traps thus demonstratingthe utility of non-pyrethroid IRS [16]The residual lifespan of alternative IRS insecticides is

of key importance Based on WHOPES recommenda-tion DDT is the longest lasting IRS with a duration ofeffective action greater than six months [17] Howeverthe Stockholm Convention on Persistent Organic Pol-lutants stipulates that lsquocountries using DDT are encour-aged to reduce and eliminate the use of DDT over timeand switch to alternative insecticidesrsquo [18] Carbamatesare a commonly used alternative to DDT and pyre-throids and were sprayed in ten African countries in2012 through PMI funding Based on WHOPES recom-mendation bendiocarb has a short residual action ofonly two to six months [17] In areas of intense year-round (perennial) transmission multiple spray roundsof short lasting insecticides are expensive logisticallydemanding and inconvenient to householders [8] Des-pite added impetus for the development of new publichealth insecticides notably from Innovative Vector Con-trol Consortium (IVCC) alternative classes of insecticidefor public health use are emerging slowly [11] For im-proved cost-effectiveness of IRS programmes it is import-ant to develop new long-lasting formulations of currentlyavailable insecticides [19]Encapsulation technology can extend the residual per-

formance of established insecticides Pirimiphos-methyl(p-methyl) is an organophosphate insecticide most com-monly and intensively used in the protection of cerealgrain [20] Several small and medium scale IRS trialsconducted since the 1970s showed high toxicity toanopheline mosquitoes [21] leading to WHOPESrsquo rec-ommendation According to WHOPES p-methyl ECformulation has a relatively short residual IRS activity oftwo to three months but was used successfully for IRSin Malawi and Zambia in 2012 [22] The overall aim ofthis study was to evaluate longevity of two capsule


suspension (CS) formulations in comparison with emul-sifiable concentrate (EC)

MethodsInsecticide formulationsTwo capsule suspension (CS) formulation variants ofActellic 300CS containing 300 gL p-methyl and codedas CS lsquoBrsquo and CS lsquoBMrsquo (Syngenta Basel Switzerland)were evaluated alongside the existing EC formulation(Actellic 50ECreg Syngenta Basel Switzerland) in labora-tory bioassays and experimental hut trials at 1 gm2Lambdacyhalothrin CS (003 gm2) (Icon CSreg SyngentaBasel Switzerland) is a WHOPES recommended formula-tion that was sprayed in Tanzania as part of the nationalmalaria control programme (NMCP) from 2007-2012[23] and was included in laboratory bioassays as a posi-tive control but was not sprayed in experimental huts(due to availability of huts)

Laboratory assessment of residual performanceCone bioassays to assess insecticidal duration on sprayedmud concrete and plywood substrates were conductedevery month based on WHO guidelines [9] Substrateswere stored at ambient temperature and humidity (~20-28degC 40-80 RH) For each formulation three blocks weresprayed and ~ nine replicates of ~ ten female Anophelesarabiensis were tested (ie three replicates per block) foran exposure of 60 minutes This is longer than the 30 mi-nutes standard exposure time as specified by WHO forIRS cone bioassay regardless of the insecticide [9] Testmosquitoes were transferred to 150 ml paper cups with10 glucose solution provided ad libitum and mortalityrecorded after 24 hours Substrates were sprayed at an ap-plication rate of 40 mlsq m using a Potter Tower PrecisionSprayer (Burkard Scientific Uxbridge UK) Resistancestatus of insectary-reared female test mosquitoes Anarabiensis Dondotha Culex quinquefasciatus TPRI andCx quinquefascaistus Muheza was determined in WHOsusceptibility tests (Table 1)

Indoor residual spraying experimental hut trialsAn experimental hut trial was conducted at KilimanjaroChristian Medical University College (KCMUCo) FieldStation in Lower Moshi Rice Irrigation Zone (3deg22rsquoS 37deg

Table 1 Resistance status of insectary-reared mosquitoes to p

Species Strain

Anopheles arabiensis Dondotha

Culex quinquefasciatus TPRI

Culex quinquefasciatus Muheza

Results of susceptibility testing with insectary strains exposed for one hour using W

19rsquoE) nightly for 12 months between December 2008 andDecember 2009 The walls and ceiling of the p-methyl EChut were covered with untreated plastic sheeting for 1month in January 2010 to investigate the possibility ofmosquito movement between huts To determine the rela-tive contribution of the sprayed mud and concrete walls tomortality of An arabiensis the palm thatch ceiling wascovered with unsprayed plastic sheeting every secondweek for 2 months from March-April 2010 in all hutsFurther description of the supplementary experimentalhut tests is included in the results section Anopheles ara-biensis densities were heavily dependent on rice croppingcycles with flooded rice fields adjacent to the Field Stationbeing the main breeding site In 2009 wild An arabiensiswere tested in WHO cylinder bioassays and were found tobe susceptible to organophosphates including p-methyland resistant to permethrin (Table 2)Verandah experimental huts were constructed to a de-

sign described by WHO [9] The working principle ofthese huts has been described previously [24] The inter-ior walls of experimental huts were plastered with eithermud or concrete A palm thatched mat typical of organicfibres used in some rural housing [25] was affixed to thewooden ceiling before sprayingThe walls and ceiling were sprayed at an application

rate of 40 mlsq m with a Hudson X-pert sprayer (H DHudson Manufacturing Company Chicago Ill USA)with flat fan 8002E nozzle [26] A constant flow valve(CFV) was not used but compression was maintainedat 55 psi by repressurizing after each swath Flow ratewas 840 mlminute A guidance pole was used to ensurea consistent vertical swath 71 cm wide and swathboundaries were marked out with chalk on walls andceiling to improve spray accuracy High performanceliquid chromatography (HPLC) was not done to con-firm the accuracy of the spray concentration Verandahswere protected during spraying by blocking the openeaves with a double layer of plastic and Hessian sack-cloth IRS treatments were randomly assigned to hutsRotation of IRS treatments was not feasible as the mudand concrete substrates were permanent Hut positionis known to bias the number of mosquitoes entering ahut but is unlikely to affect the primary proportionaloutcomes per cent mortality and per cent blood-fed of

yrethroid and organophosphate insecticides

Mortality (n)

Lambdacyhalothrin 005 Malathion 5

100 (100) 100 (100)

97 (208) 99 (200)

35 (105) 100 (200)

HO diagnostic dosages in cylinder bioassays

Table 2 Resistance status of wild Anopheles arabiensis topyrethroid and organophosphate insecticides

Insecticide Dosage () Number tested Mortality ()

P-methyl 0025 40 98

P-methyl 005 40 100

P-methyl 025 40 100

Malathion 5 201 100

Permethrin 075 111 90

Two- to five-day old sugar-fed offspring (F1) of Anopheles arabiensis collectedfrom cattle-sheds in Lower Moshi were exposed for one hour in WHO cylinderslined with papers treated with diagnostic dosages of malathion and permethrinand a range of dosages of p-methyl


those entering the huts The following treatments weresprayed in a total of six experimental huts

Pirimiphos methyl CS lsquoBrsquo 1 gsq m (one mud andone concrete walled hut)

Pirimiphos methyl CS lsquoBMrsquo 1 gsq m (one mud andone concrete walled hut)

Pirimiphos methyl EC 1 gsq m (one mud walledhut)

Unsprayed (one mud walled hut)

The trial protocols were based on WHOPES proce-dures for small-scale field trials for IRS [9] Adult trialparticipants gave informed consent and were offered freemedical services during the trial and up to three weeksafter the end of participation An adult volunteer slept ineach hut nightly from 2030-0630 Sleepers were rotatedbetween huts on successive nights to reduce any biasdue to differences in individual attractiveness to mosqui-toes Each morning mosquitoes were collected from theverandahs and window traps of huts and recorded asblood-fed or unfed and dead or alive Live mosquitoes inthe sprayed room were not collected in order to allowfor natural resting times on treated surfaces and wereonly collected after exiting to verandahs or window traps10 glucose pads were placed in the window traps and ve-randahs to prevent death by starvation Live mosquitoeswere transferred to 150 ml paper cups and providedwith 10 glucose solution before scoring delayed mor-tality after 24 hours All members of the An gambiaespecies complex identified by morphological characteris-tics were assumed to be An arabiensis based on recentPCR identification [27]

Experimental hut bioassaysCone bioassays of the sprayed walls and ceiling wereconducted monthly using sugar-fed two to five day-oldfemale An arabiensis dondotha for an exposure of 60minutes In each experimental hut four to eight repli-cates of ten female mosquitoes were tested on the walls

and ceiling surfaces Cones were positioned randomlyfor each test

Fumigant activityThe possibility of fumigant activity of the treatmentswas determined using insectary-reared wild female F1An arabiensis (no tarsal contact) [9] Wire cages meas-uring 15 cm times 10 cm times 10 cm covered with netting werehung in the corner of the room ~5 cm from the walland 25 mosquitoes exposed overnight Testing was donemonthly in for all treatments until mortality decreasedto low levels

Analysis of laboratory assessment of residualperformanceTreatments were compared according to the time inter-val since spray application for mortality to fall to 80(based on WHOPES criteria) and 50 [9] Mixed effectlogistic regression models were used to fit mortality tra-jectories over time separately for each strain of mos-quito (An arabiensis Dondotha Cx quinquefasciatus TPRIand Cx quinquefasciatus Muheza) treatment (P-methylEC CS lsquoBrsquo and CS lsquoBMrsquo and lambdacyhalothrin CS) andsubstrate (mud concrete and plywood) All statistical mod-elling was performed on the log odds scale at the individualmosquito level and results back transformed to the propor-tion scale Linear quadratic and cubic terms in time werespecified as predictors in the models to allow for potentialdrops and then levelling off in mortality rates over time Arandom effect was specified in all models to account forsimilarities in mosquitoes tested at the same time point andfor potential behavioural clustering within the same testbatch The cubic equations given by the estimates from thepolynomial models were solved to obtain estimates of thetime points at which mortality fell to 80 and 50 Ninety-five per cent confidence intervals (CI) were estimated usingthe bias corrected bootstrap method with 2000 replica-tions Differences between treatments in estimated time formortality to fall to 80 and 50 were calculated and statisti-cally significant differences inferred from the bootstrap 95CI (p = 005)

Analysis of experimental hut bioassaysAnalysis of hut bioassays was similar to that describedfor laboratory bioassays For wall assays separate modelswere fitted for each hut For ceiling assays data fromhuts treated with the same insecticide (but with differentwall materials) were combined There was little evidenceof a departure from a linear decrease in the log odds ofmortality over time for either the wall or ceiling assaysso a linear term in time was specified as the only predictorin all models


Analysis of experimental hut trialThe number of mosquitoes collected from the two closedverandahs was multiplied by two to adjust for the unre-corded escapes through the two open verandahs whichwere left unscreened to allow routes for entry of wild mos-quitoes via the gaps under the eaves [924] The data wereanalysed to show the effect of each treatment in terms of

Overall mortality = Total proportion of mosquitoesdead on the morning of collection plus delayedmortality after holding for a total of 24 hoursBlood feeding inhibition = Percentage of blood-fedmosquitoes from a treated hut relative to percentagefrom the unsprayed negative controlMortality-feeding index = The null hypothesis was thatmortality and blood-feeding are independent so thatmosquitoes surviving or killed by the treatment have anequal probability of having fed or not Deviation fromthe null hypothesis shows whether there is associationbetween feeding and mortality and may indicate thesequence of events experienced by individual mosquitoesafter entering in the hut The mortality-feeding indexis calculated as follows

Mortalityminusfeeding index frac14total bloodminusfed dead=total bloodminusfedeth THORN

ndash total unfed dead=total unfedeth THORNInterpretationof mortalityminusfeeding index

0 frac14 equal chance of unfed and blood

minusfed mosquitoes being killed

0 tominus1 frac14 deviation towards unfed mosquitoes

being killed

0 to 1 frac14 deviation towards bloodminusfed mosquitoes

being killed

Separate mixed effect logistic regression models werefitted to the mortality and blood-feeding data The mainpredictors in each model were treatment one or moretime parameters and interactions between treatmentand each of the time terms There was little evidence ofa departure from a linear decrease in the log odds ofmortality over time since spraying so only linear termsin time were specified in the statistical model for mor-tality A model with linear quadratic and cubic terms intime provided the best fit to the blood-feeding data Arandom effect was specified in both models to accountfor similarities among mosquitoes entering huts on thesame day and potential behavioural clustering Both modelscontrolled for sleeper Predicted trajectories were plottedover the duration of the 12 months for mortality alongsideactual results

ResultsLaboratory residual bioassayThe duration of residual activity of the p-methyl formu-lations on mud concrete and plywood are presented inTable 3 and the differences in residual activity are shownin Table 4 Using gt80 mortality and gt50 mortality asthe duration of residual efficacy there was evidence thatthe two CS formulations showed significantly longer activ-ity than the EC on mud and concrete substrates for bothAn arabiensis and for two strains of Cx quinquefasciatusbut differences between the two CS formulations werenon-significant in most instances There was no evidencethat treatment performance differed between species orstrainsWhen sprayed on mud the EC had a particularly short

residual action against An arabiensis and killed gt80for only one month (95 CI 07-18) CS lsquoBrsquo and CSlsquoBMrsquo showed substantial improvement over the EC withmortality gt80 for 49 months (95 CI 44-55) and44 months (95 CI 38-51) respectively (P lt 005) Theresidual times for 50 mortality to be reached (RT 50)were 75 months (95 CI 57 to dagger) for CS lsquoBrsquo 62 months(95 CI 54-70) for CS lsquoBMrsquo and 19 months (95 CI12-42) for EC (Table 3 Figure 1) On concrete CS lsquoBrsquoproduced gt80 mortality for 41 months (95 CI 36-47) longer than the EC against An arabiensis (P lt 005)(Table 4) Based on observed data on plywood both CSlsquoBrsquo and CS lsquoBMrsquo killed gt80 An arabiensis for 12 monthsThe EC killed gt80 for eight months followed by a rapiddecline to lt30 after nine months (Figure 2)

Residual activity of formulations in experimental hutsOne-hour cone bioassays of An arabiensis were con-ducted on walls and ceilings at monthly intervals Both CSformulations showed improvement over the EC on mudconcrete and palm thatch Mortality was 100 one weekafter spraying the CS lsquoBrsquo and CS lsquoBMrsquo formulations onmud and concrete walls (Figure 3) Mortality was gt80for CS lsquoBrsquo for 48 months (95 CI 19-69) on mud and70 months (95 CI 54-83) on concrete compared with09 months (95 CI 0-44) and 66 months (95 CI 30-90) for CS lsquoBMrsquo respectively (Table 5) The EC was inef-fective on mud and killed a small proportion one weekafter sprayingActellic CS on palm thatch ceiling was highly effective

with close to 100 mortality recorded for both CS formula-tions after six months (Figure 4) and gt80 for 84 monthsfor CS lsquoBrsquo (95 CI 74-94) and 108 months for CS lsquoBMrsquo(95 CI 99-117) (Table 5) Mortality remained high forthe CS formulations and was gt50 up to 12 months (95CI 112-127) and 144 (137-152) months after sprayingfor CS lsquoBrsquo and lsquoBMrsquo respectively The EC initially killed afairly high proportion of An arabiensis but showed a

Table 3 Estimated time (months) for mortality to decrease to 80 and 50 for Anopheles arabiensis Culexquinquefasciatus TPRI and Muheza strains tested on laboratory sprayed substrates

Substrate Insecticide Estimated time to 80 mortality Estimated time to 50 mortality

Time (months) 95 CI Time (months) 95 CI

Anopheles arabiensis dondotha

Mud P-methyl EC 10 (07 to 18) 19 (12 to 42)

P-methyl CS B 49 (44 to 55) 75 (57 to dagger)

P-methyl CS BM 44 (38 to 51) 62 (54 to 70)

Concrete P-methyl EC 23 (18 to 27) 31 (27 to 33)

P-methyl CS B 64 (61 to 68) 72 (69 to 75)




Lambda CS 29 (27 to 33) 37 (34 to 40)

P-methyl CS B 62 (53 to 76) dagger dagger



Lambda CS 50 (47 to 53) 59 (57 to 61)

P-methyl CS B 82 (75 to 93) 97 (89 to 107)




Lambda CS dagger dagger 09 (05 to 14)

P-methyl CS B 40 (35 to 46) 71 (55 to 110)



Lambda CS 11 (08 to 16) 18 (15 to 22)

P-methyl CS B 49 (42 to 56) 65 (58 to 74)


daggerIndicates that statistical models produced estimates outside the study period for Culex quinquefasciatus TPRI estimated mortality for Actellic CS-B on mud washigher than 50 throughout the entire study period for Culex quinquefasciatus Muheza estimated mortality for Lambda CS was lower than 80 throughout


marked reduction to lt50 24 months (95 CI 0-61)after spraying

Twelve-months experimental hut trial against wildfree-flying Anopheles arabiensisAll formulations of p-methyl (CS lsquoBrsquo CS lsquoBMrsquo and EC)were highly effective against free-flying wild An arabiensisshortly after spray application (Figure 5) Mortalitygradually decreased over time for all formulations up tofive months after spraying followed by a small increasebetween months five to seven possibly due to climaticchanges Subsequently between months seven to 12there was a gradual decrease in mortality (Figure 5)Overall mortality rates remained high for both CS treat-ments up to12 months after spraying regardless of wallsubstrate P-methyl EC performed equally well as CS lsquoBrsquoand CS lsquoBMrsquo after 12 months based on 95 CIs from

estimated curves Twelve months after spraying predictedmortality was 628 (95 CI 544-712) for EC 720(95 CI 645-796) for CS lsquoBrsquo (mud) and 695 (95 CI620-770) for CS lsquoBMrsquo (mud) (Table 6)Blood-feeding was high in the unsprayed hut through-

out the study but did show considerable variation overtime and ranged from 40 (after nine months) to 90(five and 12 months) (Figure 6) The two periods of lowestpercentage blood feeding in the unsprayed hut one andnine months after spraying coincided with the period ofhighest mosquito density during rice transplantation cy-cles (Figure 6) For the first month after spraying treatedhuts provided no protection from being bitten by host-seeking An arabiensis Between two and 12 months afterspraying all treatments provided some degree of personalprotection (Figure 6) Blood-feeding inhibition was rela-tively high after six and nine months across all treatments

Table 4 Between treatment differences in estimated time for mortality to fall to 80 and 50 for mosquitoes tested oninsecticide-treated substrates

Substrate Treatmentcomparison

Difference in estimated time to 80 mortality Difference in estimated time to 50 mortality

Time months 95 CI p Time months 95 CI p


Mud CS B vs EC 39 (31 to 46) lt005 56 (30 to 129) lt005

CS BM vs EC 35 (26 to 43) lt005 42 (20 to 54) lt005

CS B vs CS BM 04 (-04 to 13) ns 13 (-07 to 117) ns

Concrete CS B vs EC 41 (36 to 47) lt005 41 (37 to 46) lt005


CS B vs CS BM 15 (08 to 22) lt005 07 (01 to 13) lt005


Mud CS B vs EC 44 (34 to 58) lt005 dagger dagger dagger


Lambda vs EC 12 (06 to 17) lt005 16 (10 to 21) lt005

CS B vs Lambda 32 (22 to 46) lt005 dagger dagger dagger

CS BM vs Lambda 44 (38 to 52) lt005 60 (49 to 74) lt005

CS B vs CS BM -12 (-24 to 04) ns dagger dagger dagger


CS BM vs EC 60 (-02 to 64) ns 72 (67 to 78) lt005


CS B vs Lambda 32 (24 to 43) lt005 38 (29 to 48) lt005

CS BM vs Lambda 18 (-44 to 24) ns 27 (21 to 33) lt005





Lambda vs EC dagger dagger dagger -03 (-09 to 03) ns

CS B vs Lambda dagger dagger dagger 61 (42 to 102) lt005

CS BM vs Lambda dagger dagger dagger 55 (46 to 66) lt005




Lambda vs EC 01 (-03 to 05) ns 04 (-01 to 09) ns



CS B vs CS BM 06 (-02 to 14) ns 08 (00 to 19) ns

daggerIndicates that statistical models produced estimates outside the study period for one or more of the treatments or their 95 CI and treatment differences cannottherefore be estimated


ranging between 39-49 for CS formulations and 36-43 for EC (Table 7) Blood-feeding inhibition wassimilar for both CS and EC formulations over the trialThe mortality-feeding index (total blood-fed deadtotalblood-fed) ndash (total unfed deadtotal unfed) was 008and 005 for CS lsquoBrsquo and 008 and 003 for CS lsquoBMrsquo onconcrete and mud walled huts compared with 007 forEC and 015 for the unsprayed hut (mud walls) For alltreatments the mortality-feeding index was close to 0

indicating mosquitoes had an equal chance of survivingwhether fed or unfedFumigant activity tested in small cages resulted in

100 mortality of An arabiensis F1 one week and twomonths after spraying for CS lsquoBrsquo lsquoBMrsquo and EC formula-tions A large decrease to 42 fumigant mortality wasrecorded after three months for CS lsquoBMrsquo (concrete)with fumigant mortality less than 10 for all othertreatments

025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12

Time since spraying (months)

Actellic CS-B Actellic CS-BMActellic EC

025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12



Observed Trajectories Predicted Trajectories

Figure 1 Mortality of Anopheles arabiensis dondotha on mud blocks after one-hour bioassays Mud blocks were sprayed with p-methyl CSlsquoBrsquo CS lsquoBMrsquo and EC and tested at monthly intervals Mortality for unsprayed blocks was lt15 for all bioassays


Supplementary explanatory bioassays in experimentalhutsThe walls and ceiling of the p-methyl EC hut were cov-ered with untreated plastic sheeting between months12-13 This was done to investigate the possibility of mos-quito movement between huts picking up a lethal dosage

025

5075

100

Mor

talit

y (

)

0 2 4

Time since s

Actellic CS-BActellic EC

Figure 2 Mortality of Anopheles arabiensis dondotha on plywood blocp-methyl CS lsquoBrsquo CS lsquoBMrsquo and EC and tested at monthly intervals Mortality f

of p-methyl CS before exiting flying into the EC hut anddying All other huts were left uncovered Mortality forthe covered EC hut was 29 which was greater than theunsprayed hut 1 (P = 0001) but less than huts sprayedwith CS lsquoBrsquo 65 78 and CS lsquoBMrsquo 67 74 with con-crete and mud walls respectively (P = 0001) (Table 8) The

6 8 10 12

praying (months)

Actellic CS-BM

ks after one-hour bioassays Plywood blocks were sprayed withor unsprayed blocks was lt15 for all bioassays

025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21


Untreated control - mud Actellic CS-B - concreteActellic CS-B - mud Actellic CS-BM - concreteActellic EC - mud Actellic CS-BM - mud

025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21



Figure 3 Mortality of Anopheles arabiensis dondotha after one-hour bioassay on experimental hut walls Time after spraying is shown inmonths Mortality for unsprayed walls was lt15 for all bioassays


proportion of An arabiensis that blood-fed was signifi-cantly higher in the covered EC hut (63) than for CSformulations (19-38 P lt 005) but was less than the un-sprayed hut 94 (P = 0001)To determine the relative contribution of the sprayed

mud and concrete walls to mortality of An arabiensisthe palm thatch ceiling was covered with unsprayed plasticsheeting every second week between months 15-16 Asthe palm thatch ceiling remained highly insecticidal overthe duration of the study (Figure 4) the hypothesis wasthat it masked any differences in efficacy between the con-crete and mud walls (Figure 3) The covering of the ceilinghad little impact on overall mortality trends for the EChut (mud) with 43 mortality when uncovered and 46

Table 5 Estimated time (months) for mortality to decrease to(pyrethroid susceptible) tested on sprayed experimental hut

Insecticide Substrate Estimated time to 80

Time (months)

Hut walls

P-methyl EC Mud dagger

P-methyl CS B Concrete 70

Mud 48

P-methyl CS BM Concrete 66

Mud 09

Hut ceilings

P-methyl EC Thatch dagger

P-methyl CS B Thatch 84

P-methyl CS BM Thatch 108

daggerIndicates that statistical models produced estimates outside the study period in arespectively) throughout the entire study period

covered (P = 0255) (Table 8) For both CS lsquoBrsquo and CS lsquoBMrsquoany differences in mortality after covering the ceiling weresmall for both mud and concrete hutsExtended cone bioassays of up to 12 hours were

undertaken as may occur when mosquitoes enter ahouse early in the evening to blood-feed and subse-quently rest on treated surfaces until the followingmorning before exiting With one-hour exposure fourmonths after spraying the CS lsquoBrsquo and CS lsquoBMrsquo killed a fargreater proportion (P = 0001) of An arabiensis than ECwith mortality 18 for EC compared with 57 and 79for CS lsquoBrsquo and CS lsquoBMrsquo (Figure 7) With longer exposureof two hours the EC killed 88 of An arabiensis com-pared with 100 for CS formulations A similar trend

80 and 50 for Anopheles arabiensis dondothawalls (concrete and mud) and ceiling (thatch)

mortality Estimated time to 50 mortality

95 CI Time (months) 95 CI

dagger dagger dagger

(54 to 83) 113 (102 to 124)

(19 to 69) 114 (99 to 130)

(30 to 90) 160 (135 to 206)

(dagger to 44) 90 (64 to 110)

dagger 24 (dagger to 61)

(74 to 94) 120 (112 to 127)

(99 to 117) 144 (137 to 152)

ll cases estimates were lower than the specified mortality (50 or 80

Figure 4 Mortality of Anopheles arabiensis after one-hour bioassay on experimental hut ceiling One-hour cone bioassay of insectary-reared Anopheles arabiensis dondotha on palm thatch ceiling over time (months) after spray application

Oxborough et al Malaria Journal 2014 1337 0 of 15httpwwwmalariajournalcomcontent13137

was observed after ten months as the EC killed 15 withone-hour exposure but killed 73 with a four-hour ex-posure compared with 80 for CS lsquoBMrsquo (P = 0401) and97 for CS lsquoBrsquo (P = 0014) After 17 months mortalitywas low for both CS lsquoBrsquo (20) and EC (20) with one-hour exposure but increased to 52 for EC 72 CS lsquoBrsquoand 98 for CS lsquoBMrsquo with 12-hour exposure

DiscussionLaboratory bioassays showed that p-methyl CS lsquoBrsquo and CSlsquoBMrsquo formulations were effective at killing high proportions

025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12



Figure 5 Mortality of wild Anopheles arabiensis freely entering experimosquitoes recorded on a daily basis were variable Graphs of observed m

(gt80) of An arabiensis and Cx quinquefasciatus for sig-nificantly longer than the EC formulation on mud concreteand plywood substrates The most important improvementwas observed on mud The EC was ineffective on mud andkilled gt80 of An arabiensis and Cx quinquefasciatus forone month or less In contrast the best performing CSformulation killed gt80 of An arabiensis for five monthsand sustained control above 50 for longer than sevenmonths Similar longevity was observed in The Gambiawhere p-methyl CS sprayed in village houses persisted forat least five months (when testing was ended) on mud and

025

5075

100

Pre

dict

ed m

orta

lity

()

0 2 4 6 8 10 12


Actellic CS-B - concrete Actellic CS-B - mudActellic CS-BM - concrete Actellic EC - mudActellic CS-BM - mud

mental huts over 12 months after spraying Data on wildortality over time plot data pooled for each month since spraying

Table 6 Estimated mortality () three six nine and twelve months after spraying for wild mosquitoes collected ininsecticide treated huts

Mortality (95 CI)

Insecticide Substrate 3 months 6 months 9 months 12 months

P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)

P-methyl CS B Concrete 810 768 718 663

(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)

P-methyl CS BM Concrete 825 798 769 738

(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)

Estimates are adjusted for sleeper and account for similarities among mosquitoes entering huts on the same day and potential behavioural clustering


painted walls [28] Mud is a problematic substrate forIRS owing to loss of available insecticide due to sorp-tion Early work in Tanzania in the 1960s characterizedthe performance of organophosphates and carbamateson various types of soil and showed rapid loss of efficacyon several types of mud while on less porous substratessuch as wood high levels of mortality were recordedover several months [2930] In the present study micro-encapsulation substantially improved the surface bioavail-ability of p-methyl on mud Mud or adobe is still acommon wall material in rural low-income areas of

025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12



Figure 6 Percentage blood-fed Anopheles arabiensis collected in expeAnopheles arabiensis caught per treatment over time (right) Data on wblood feeding and number of mosquitoes caught over time plot data poo

Africa In Tanzania in 2010 78 of houses were con-structed from a form of mud the most common typesbeing mud plaster (27) sun-dried mud bricks (28)and burnt mud bricks (23) [25]Both CS formulations showed improved longevity over

EC on concrete and wood substrates in bioassays Thealkaline Ph of concrete can rapidly degrade insecticidescommonly used for IRS particularly pyrethroids resultingin reduced residual efficacy [17] In laboratory bioassayson plywood CS formulations lasted for several monthslonger than the EC and killed gt80 of An arabiensis 12

020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12



rimental huts over time by treatment (left) and number ofild mosquitoes recorded on a daily basis were variable Graphs of

led for each month since spraying

Table 7 Estimated blood feeding () three six nine and twelve months after spraying for wild mosquitoes collectedin insecticide treated huts


Blood fed () BFI () Blood fed () BFI () Blood fed () BFI () Blood fed () BFI ()

(95 CI) (95 CI) (95 CI) (95 CI)

Untreated control Mud 90 - 81 ndash 57 ndash 93 ndash

(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)

P-methyl CS B Concrete 71 22 49 40 32 44 84 9

(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)

P-methyl CS BM Concrete 68 24 48 41 29 49 63 33

(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)

Estimates are adjusted for sleeper and account for similarities among mosquitoes entering huts on the same day and potential behavioural clusteringBFI = blood-feeding inhibition compared to the untreated control


months after spraying Wood is relatively non-porous witha tendency for long residual bioavailability of organophos-phates and pyrethroids [2931]Cone bioassays on mud and concrete experimental hut

walls showed similar findings to laboratory results andshowed that both CS formulations were effective for sig-nificantly longer than the EC For all bioassays in the

Table 8 Supplementary experimental hut results for percentaspraying

Time after spraying Outcomemeasures

Untreated(Mud)

13 Months (EC Walls amp CeilingCovered)

Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -

15-16 Months (Ceiling Uncovered) Total Caught 411

Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -

15-16 Months (Ceiling Covered) Total Caught 303

Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -

During month 13 the walls and ceiling of the hut sprayed with p-methyl EC were cevery hut were covered with plastic sheeting for one out of every two weeks DataBFI = blood-feeding inhibition compared to untreated control

laboratory and experimental huts an exposure time of60 minutes was used rather than the standard WHOPES30 minutes exposure It is likely that the residual durationof action would be shorter if tested using WHOPESguidelinesResults for free-flying wild An arabiensis showed that

huts sprayed with p-methyl CS formulations maintained

ge mortality and blood-feeding 13-16 months after

CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22

overed with plastic sheeting Between months 15 and 16 the treated walls ofare grouped according to whether the walls were covered or uncovered

Figure 7 Results of extended duration bioassays on walls of experimental huts Percentage mortality of insectary-reared Anopheles arabiensisdondotha following cone bioassay with standard exposure time of one hour (light bars) and extended exposure (darker bars) of two hours (fourmonths) four hours (ten months) 12 hours (17 months) on sprayed mud walls Mortality for unsprayed walls was lt20 for all bioassays


high rates of mortality for up to 12 months after sprayingThis finding is comparable to that in Benin where 1 gsq mof p-methyl sprayed in mud and concrete experimentalhuts killed around 75 of wild free-flying An gambiae ssten months after spraying [32]In Tanzania there was an increase in mortality for all

formulations five to seven months after spraying be-tween May-July This was the cool season when meannight-time temperature outdoors dropped to 20degC com-pared with 24degC inside the experimental huts (USBWireless Touchscreen Weather Forecaster Maplin UK)This may have resulted in longer indoor resting timeswhich would explain the increase in mortality during thisthree-month period It has been reported elsewhere thatat higher altitude where differences between indoor andoutdoor temperature are greatest indoor resting is morecommon [3334]An unexpected finding was that the EC formulation

matched the performance of the CS against wild free-flyingAn arabiensis despite being considered by WHOPES tohave an effective duration of only two to three months[1732] Recent studies in Ghana on painted cement andMozambique on several surfaces showed high levels ofmortality for the EC formulation gt four months after spray-ing indicating that the EC can remain effective for a rela-tively long duration [35] In this study the EC maintainedhigh levels of mortality for wild free-flying An arabiensisbut paradoxically showed poor performance in one-hourcone bioassay on hut walls only weeks after sprayingSeveral explanations were postulatedMosquito resting location Mortality in the EC hut may

have been generated by tarsal contact with palm thatchceiling with mud walls providing a small proportion of

overall mortality Covering the ceiling with untreatedplastic did not result in a decrease in mortality indicat-ing that mosquitoes were able to pick up a lethal dosagefrom treated mud wallsMosquito movement between huts It was plausible that

mosquitoes were picking up a lethal dosage of p-methylCS before exiting through open verandahs flying intothe EC hut and falsely being recorded as killed by theEC Covering all sprayed surfaces (walls and ceiling) withuntreated plastic for one month (13 months after spray-ing) in the EC hut should have resulted in low mortalityrates similar to an unsprayed hut if there was no move-ment of mosquitoes between huts When covered mortal-ity was 29 which although slightly higher than theunsprayed hut suggested that few mosquitoes were flyingbetween huts Throughout the trial mortality in the un-sprayed control was lt20 This suggests that mortalitywas generated by insecticidal activity within each individ-ual hut and any movement of mosquitoes between hutshad a limited effect on mortality trendsMosquito resting duration The standard exposure time

as specified by WHO for IRS cone bioassay is 30 minutesregardless of the insecticide [9] This exposure time isprobably suitable for excito-repellent insecticides such aspyrethroids and DDT Resting times of blood-fed Angambiae on a wall sprayed with a non-irritant insecticidesuch as p-methyl may be longer than 30 minutes For thisstudy an exposure of one hour was selected for monthlybioassays with supplementary bioassays of up to 12 hoursIn the EC hut the finding that one-hour bioassays killed asmall proportion of An arabiensis while hut collectionsshowed high levels of mortality may indicate that mosqui-toes either i) rested for a short time and exited before


picking up lethal dosage or ii) rested for several hours Ex-tended cone bioassay of two hours after four months andfour hours after ten months showed high levels of mortal-ity for both EC and CS formulations Anopheles arabiensismay have rested on treated surfaces for several hours over-night and may partially explain why EC mortality wassimilar to that of the CS formulations for wild free-flyingAn arabiensis While this offers some understanding towhy the EC was effective for a longer duration than ex-pected it does not provide a full explanation for this Asnew insecticides are developed for IRS with low excito-repellency WHOPES may have to revisit the standard 30minutes exposure for IRS if this period of exposure doesnot provide an accurate prediction of field performanceThe mortality-feeding index showed that unfed mosqui-

toes were equally likely to be killed by p-methyl as thoseblood-fed The concept of IRS is to kill mosquitoes thatblood-feed and then rest on treated surfaces while pro-cessing the blood meal This finding indicates that someAn arabiensis rested on hut surfaces before attemptingto blood-feed and explains why there was some protect-ive effect of p-methyl IRS [36] There were apparentseasonal changes in percentage blood-feeding in the un-sprayed hut The periods of lowest proportion blood-fedcoincided with peak mosquito densities during ricetransplantation It is likely that a larger proportion ofnewly emerged An arabiensis entered experimentalhuts from adjacent paddies for resting or sugar feedingrather than host-seeking [37]There was a fumigant effect of all formulations that

killed a high proportion of mosquitoes in cage bioassaysduring the first two months after spraying The microcap-sules in the CS would have limited any fumigant effect be-cause the majority of active ingredient is enclosed withinthe capsule membrane however some active ingredient isalso present in external solution Slow release of active in-gredient from microcapsules was sufficient for contactmortality but insufficient for a fumigant effect Question-naires of volunteers sleeping during the hut trial resultedin Actellic EC ranked consistently last in terms of odourappeal with typical comments including ldquoSmells like cab-bage and white spiritrdquo or ldquoNot pleasant and produces irri-tationrdquo The CS formulations ranked better and weregenerally considered to be much milder than the EC withcomments such as ldquoSmells like cow insecticide appealingas not too strongrdquoOf 17 African countries sprayed with PMI-funded IRS

in 2012 only one was classified as having pyrethroidsusceptible anophelines the remainder had confirmed oremerging resistance [10] The Global Plan for InsecticideResistance Management (GPIRM) states that in areas ofpyrethroid resistance IRS rotations should be used withnon-pyrethroid insecticides [38] Despite added impetusfrom the IVCC there have been no new insecticides for

IRS and LLIN since the pyrethroids in the 1980s [11]As a result the majority of African PMI-funded IRS pro-grammes are currently spraying IRS with bendiocarbwhich has a short residual efficacy of only two to sixmonths and is relatively expensive [1017] In Malawiwhere resistance to both pyrethroids and carbamates wasdetected p-methyl EC was sprayed in 2011 but ldquoalthougheffective the high unit cost substantially increased the IRScosts and PMI subsequently suspended direct support dueto increased costsrdquo [39] Long-lasting p-methyl CS formu-lations should be more cost-effective than both p-methylEC and bendiocarb but this estimation is sensitive to boththe duration of efficacy and the relative cost per unit areasprayed Use of p-methyl IRS + pyrethroid LLIN is prefer-ential for resistance management to pyrethroid IRS + pyr-ethroid LLINs as p-methyl and pyrethroids have differentmodes of action which should result in redundant killingof mosquitoes resistant to a single insecticide [40] Cross-resistance of organophosphates and carbamates due to al-tered acetylcholinesterase (AChE) target site is present atlow frequency in limited parts of west and central Africaand may increase in frequency as a result of currentIRS programmes using bendiocarb Nevertheless IRSwith p-methyl CS should prove an effective solutionfor control of pyrethroid resistant An gambiae andhaving received recent recommendation from WHO[41] is a welcome addition to the limited portfolio oflong-lasting IRS

Ethical approvalEthical approval was granted from the review boards ofLSHTM (5256) and Tanzania National Institute of MedicalResearch (NIMRHQR8cVolI24)

Competing interestsThe authors declare that they have no competing interests

Authorsrsquo contributionsRMO participated in the study design data collection and analysis anddrafted the manuscript JK participated in the study design data collectionand helped to draft parts of the manuscript RJ performed the statisticalanalysis and provided critical comments on the manuscript EF and JMparticipated in data collection and was involved in drafting parts of themanuscript FWM participated in study design and critically revised themanuscript MWR participated in study design analysis and multiple revisionsof the manuscript All authors read and approved the final manuscript

AcknowledgementsRMO and MR are members of the Malaria Centre of the London School ofHygiene and Tropical Medicine httpmalarialshtmacuk The authors thankAndy Bywater and Mark Birchmore of Syngenta for supplying samples andfor technical support Dave Malone and Robert Sloss for technicalprogramme support (IVCC Liverpool UK) We are grateful to C Masenga AMtui E Philipo H Temba P Anthony A Sanga R Athuman K Ezekia for fieldand laboratory assistance

Author details1Department of Disease Control London School of Hygiene and TropicalMedicine (LSHTM) London UK 2Department of Entomology andParasitology Kilimanjaro Christian Medical University College (KCMUCo) ofTumaini University Moshi Kilimanjaro Tanzania 3Department of Entomology


Pan-African Malaria Vector Research Consortium (PAMVERC) MoshiKilimanjaro Tanzania

Received 28 November 2013 Accepted 12 January 2014Published 29 January 2014

References1 Pluess B Tanser FC Lengeler C Sharp BL Indoor residual spraying for

preventing malaria Cochrane Database Syst Rev 2010CD0066572 Griffith M The World-Wide Malaria Eradication Program Presented in the

Plenary Symposium on International Programs at the meeting of theEntomological Society of America in New Orleans November 30 19651965

3 Molineaux L Gramiccia G The Garki Project research on the epidemiologyand control of malaria in the Sudan Savanna of West Africa Geneva WorldHealth Organization 1980

4 WHO Implementation of indoor residual spraying of insecticides for malariacontrol in the WHO African Region Report Geneva Switzerland World HealthOrganization 2007

5 Akhtar RL Learmonth A The resurgence of malaria in India 1965-76GeoJournal 1977 1(5)69ndash80

6 Pinikahana J Dixon RA Trends in malaria morbidity and mortality in SriLanka Indian J Malariol 1993 3051ndash55

7 Mabaso ML Sharp B Lengeler C Historical review of malarial control insouthern African with emphasis on the use of indoor residual house-spraying Trop Med Int Health 2004 9846ndash856

8 WHO Indoor Residual Spraying Use of indoor residual spraying for scaling upglobal malaria control and elimination Geneva Switzerland WHOHTMMAL20061112 2006

9 WHO Guidelines for testing mosquito adulticides for indoor residual sprayingand treatment of mosquito nets Geneva Switzerland WHOCDSNTDWHOPESGCDPP20062003 2006

10 PMI PMI Actual and Estimated Use of Insecticides for the Indoor ResidualSpraying Program 2012 [httpwwwpmigovtechnicalirsPMI_IRS_Insecticide_Trends_080112xlsx]

11 Hemingway J Beaty BJ Rowland M Scott TW Sharp BL The innovativevector control consortium improved control of mosquito-bornediseases Trends Parasitol 2006 22308ndash312

12 WHO WHO recommended insecticide products treatment of mosquito nets formalaria vector control Geneva World Health Organization 2007

13 WHOPES Report of the 4th WHOPES Working Group Meeting WHOHQGeneva WHOCDSWHOPES20012002 2000

14 Ranson H NGuessan R Lines J Moiroux N Nkuni Z Corbel V Pyrethroidresistance in African anopheline mosquitoes what are the implicationsfor malaria control Trends Parasitol 2011 2791ndash98

15 Maharaj R Mthembu DJ Sharp BL Impact of DDT re-introduction onmalaria transmission in KwaZulu-Natal S Afr Med J 2005 95871ndash874

16 Sharp BL Ridl FC Govender D Kuklinski J Kleinschmidt I Malaria vectorcontrol by indoor residual insecticide spraying on the tropical island ofBioko Equatorial Guinea Malar J 2007 652

17 WHO WHO recommended insecticides for indoor residual spraying againstmalaria vectors 2013 httpwwwwhointwhopesInsecticides_IRS_Malaria_25_Oct_2013pdfua=1

18 UNEP Ridding the World of POPs A Guide to the Stockholm Convention onPersistent Organic Pollutants 2010

19 Zaim M Guillet P Alternative insecticides an urgent need Trends Parasitol2002 18161ndash163

20 Mabbett T Pirimiphos methyl - long term solution to the problem ofstored product pests Int Pest Contr 2002203ndash204

21 Nasir SM Ahmad N Shah MA Azam CM A large-scale evaluation ofpirimiphos-methyl 25 WP during 1980-1981 for malaria control inPakistan J Trop Med Hyg 1982 85239ndash244

22 Presidentrsquos Malaria Initiative Malawi and Zambia Malaria Operational Plan FY2013 2013

23 PMI Presidentrsquos Malaria Initiative Tanzania Malaria Operational Plan FY 20132013

24 Curtis CF Myamba J Wilkes TJ Comparison of different insecticides andfabrics for anti-mosquito bednets and curtains Med Vet Entomol 1996101ndash11

25 TDHS Tanzania Demographic and Health Survey 2010 National Bureau ofStatistics Dar es Salaam Tanzania Calverton Maryland USA ICF Macro2011

26 WHO Manual for Indoor Residual Spraying Application of Residual Sprays forVector Control Geneva Switzerland WHOCDSNTDWHOPESGCDPP20072003 2007c

27 Kitau J Oxborough RM Tungu PK Matowo J Malima RC Magesa SM BruceJ Mosha FW Rowland MW Species shifts in the Anopheles gambiaecomplex do LLINs successfully control Anopheles arabiensis PLoS One2012 7e31481

28 Tangena JA Adiamoh M DAlessandro U Jarju L Jawara M Jeffries D MalikN Nwakanma D Kaur H Takken W Lindsay SW Pinder M Alternativetreatments for indoor residual spraying for malaria control in a villagewith pyrethroid- and DDT-resistant vectors in The Gambia PLoS One2013 8e74351

29 Hadaway AB Barlow F The toxicity of some organophosphoruscompounds to adult Anopheles stephensi Bull World Health Organ 19632855ndash61

30 Hadaway AB Barlow F The residual action of two organophosphoruscompounds and a carbamate on dried muds Bull World Health Organ1963 2869ndash76

31 Tseng LF Chang WC Ferreira MC Wu CH Rampao HS Lien JC Rapidcontrol of malaria by means of indoor residual spraying ofalphacypermethrin in the Democratic Republic of Sao Tome andPrincipe Am J Trop Med Hyg 2008 78248ndash250

32 Rowland M Boko P Odjo A Asidi A Akogbeto M NGuessan R A newlong-lasting indoor residual formulation of the organophosphateinsecticide pirimiphos methyl for prolonged control of pyrethroid-resistant mosquitoes an experimental hut trial in Benin PLoS One 20138e69516

33 Paaijmans KP Thomas MB The influence of mosquito resting behaviourand associated microclimate for malaria risk Malar J 2011 10183

34 Manguin S Biodiversity of malaria in the world English completely updatededn Montrouge Esther [Esher] Surrey John Libbey Eurotext 2008

35 Fuseini G Ebsworth P Jones S Knight D The efficacy of ACTELLIC 50 ECpirimiphos methyl for indoor residual spraying in Ahafo Ghana area ofhigh vector resistance to pyrethroids and organochlorines J MedEntomol 2011 48437ndash440

36 Oxborough RM Kitau J Matowo J Mndeme R Feston E Boko P Odjo AMetonnou CG Irish S NGuessan R Mosha FW Rowland MW Evaluation ofindoor residual spraying with the pyrrole insecticide chlorfenapyragainst pyrethroid-susceptible Anopheles arabiensis and pyrethroid-resistant Culex quinquefasciatus mosquitoes Trans R Soc Trop Med Hyg2010 104639ndash645

37 Foster WA Takken W Nectar-related vs human-related volatiles behaviouralresponse and choice by female and male Anopheles gambiae (DipteraCulicidae) between emergence and first feeding Bull Entomol Res 200494145ndash157

38 WHO The Global Plan for Insecticide Resistance Management in MalariaVectors (GPIRM) 2012

39 Presidentrsquos Malaria Initiative Malawi Malaria Operational Plan FY 2013 201340 Denholm I Rowland MW Tactics for managing pesticide resistance in

arthropods theory and practice Annu Rev Entomol 1992 3791ndash11241 WHOPES Report of the Sixteenth WHOPES Working Group Meeting WHOHQ

Geneva Review of Pirimiphos-methyl 300CS Chlorfenapyr 240 SC Deltamethrin625 SC-PE Duranet LN Netprotect LN Yahe LN Spinosad 833 Monolayer DTSpinosad 25 Extended Release GR 2013

doi1011861475-2875-13-37Cite this article as Oxborough et al Long-lasting control of Anophelesarabiensis by a single spray application of micro-encapsulatedpirimiphos-methyl (Actellicreg 300 CS) Malaria Journal 2014 1337

Abstract

Background

Methods

Results

Conclusions

Background

Methods

Insecticide formulations

Laboratory assessment of residual performance

Indoor residual spraying experimental hut trials

Experimental hut bioassays

Fumigant activity

Analysis of laboratory assessment of residual performance

Analysis of experimental hut bioassays

Analysis of experimental hut trial

Results

Laboratory residual bioassay

Residual activity of formulations in experimental huts

Twelve-months experimental hut trial against wild free-flying Anopheles arabiensis

Supplementary explanatory bioassays in experimental huts

Discussion

Ethical approval

Competing interests

Authorsrsquo contributions

Acknowledgements

Author details

References


RESEARCH Open Access

Long-lasting control of Anopheles arabiensis by asingle spray application of micro-encapsulatedpirimiphos-methyl (Actellicreg 300 CS)Richard M Oxborough123 Jovin Kitau23 Rebecca Jones1 Emmanuel Feston2 Johnson Matowo23Franklin W Mosha23 and Mark W Rowland13

Abstract

Background Pyrethroid-resistant mosquitoes are an increasing threat to malaria vector control The Global Plan forInsecticide Resistance Management (GPIRM) recommends rotation of non-pyrethroid insecticides for indoor residualspraying (IRS) The options from other classes are limited The carbamate bendiocarb and the organophosphatepirimiphos-methyl (p-methyl) emulsifiable concentrate (EC) have a short residual duration of action resulting inincreased costs due to multiple spray cycles and user fatigue Encapsulation (CS) technology was used to extendthe residual performance of p-methyl

Methods Two novel p-methyl CS formulations were evaluated alongside the existing EC in laboratory bioassaysand experimental hut trials in Tanzania between 2008-2010 Bioassays were carried out monthly on sprayedsubstrates of mud concrete plywood and palm thatch to assess residual activity Experimental huts were used toassess efficacy against wild free-flying Anopheles arabiensis in terms of insecticide-induced mortality andblood-feeding inhibition

Results In laboratory bioassays of An arabiensis and Culex quinquefasciatus both CS formulations produced highrates of mortality for significantly longer than the EC formulation on all substrates On mud the best performing CSkilled gt80 of An arabiensis for five months and gt50 for eight months compared with one and two monthsrespectively for the EC In monthly bioassays of experimental hut walls the EC was ineffective shortly after sprayingwhile the best CS formulation killed more than 80 of An arabiensis for five months on mud and seven monthson concrete In experimental huts both CS and EC formulations killed high proportions of free-flying wild An arabiensisfor up to 12 months after spraying There was no significant difference between treatments All treatmentsprovided considerable personal protection with blood-feeding inhibition ranging from 9-49 over time

Conclusions The long residual performance of p-methyl CS was consistent in bioassays and experimental hutsThe CS outperformed the EC in laboratory and hut bioassays but the EC longevity in huts was unexpectedLong-lasting p-methyl CS formulations should be more effective than both p-methyl EC and bendiocarbconsidering a single spray could be sufficient for annual malaria control IRS with p-methyl 300 CS is a timelyaddition to the limited portfolio of long-lasting residual insecticides

Keywords IRS Pirimiphos-methyl Actellic Anopheles arabiensis Vector control Malaria Tanzania

Correspondence oxandbullhotmailcom1Department of Disease Control London School of Hygiene and TropicalMedicine (LSHTM) London UK2Department of Entomology and Parasitology Kilimanjaro Christian MedicalUniversity College (KCMUCo) of Tumaini University Moshi KilimanjaroTanzaniaFull list of author information is available at the end of the article

copy 2014 Oxborough et al licensee BioMed Central Ltd This is an Open Access article distributed under the terms of theCreative Commons Attribution License (httpcreativecommonsorglicensesby20) which permits unrestricted usedistribution and reproduction in any medium provided the original work is properly credited The Creative Commons PublicDomain Dedication waiver (httpcreativecommonsorgpublicdomainzero10) applies to the data made available in thisarticle unless otherwise stated

Oxborough et al Malaria Journal 2014 1337 of 15httpwwwmalariajournalcomcontent13137














Species Strain









Mortality (n)


100 (100) 100 (100)

97 (208) 99 (200)

35 (105) 100 (200)




P-methyl 0025 40 98

P-methyl 005 40 100

P-methyl 025 40 100

Malathion 5 201 100























being killed


being killed














P-methyl CS B 64 (61 to 68) 72 (69 to 75)




Lambda CS 29 (27 to 33) 37 (34 to 40)




Lambda CS 50 (47 to 53) 59 (57 to 61)

P-methyl CS B 82 (75 to 93) 97 (89 to 107)





P-methyl CS B 40 (35 to 46) 71 (55 to 110)



Lambda CS 11 (08 to 16) 18 (15 to 22)

P-methyl CS B 49 (42 to 56) 65 (58 to 74)


















































025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12



025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12







025

5075

100

Mor

talit

y (

)

0 2 4

Time since s




6 8 10 12

praying (months)

Actellic CS-BM


025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21



025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21









Time (months)

Hut walls



Mud 48


Mud 09

Hut ceilings











(54 to 83) 113 (102 to 124)

(19 to 69) 114 (99 to 130)

(30 to 90) 160 (135 to 206)

(dagger to 44) 90 (64 to 110)


(74 to 94) 120 (112 to 127)

(99 to 117) 144 (137 to 152)






025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12





025

5075

100

Pre

dict

ed m

orta

lity

()

0 2 4 6 8 10 12





Mortality (95 CI)


P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)


(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)


(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)




025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12






020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12








(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References















Species Strain









Mortality (n)


100 (100) 100 (100)

97 (208) 99 (200)

35 (105) 100 (200)




P-methyl 0025 40 98

P-methyl 005 40 100

P-methyl 025 40 100

Malathion 5 201 100























being killed


being killed














P-methyl CS B 64 (61 to 68) 72 (69 to 75)




Lambda CS 29 (27 to 33) 37 (34 to 40)




Lambda CS 50 (47 to 53) 59 (57 to 61)

P-methyl CS B 82 (75 to 93) 97 (89 to 107)





P-methyl CS B 40 (35 to 46) 71 (55 to 110)



Lambda CS 11 (08 to 16) 18 (15 to 22)

P-methyl CS B 49 (42 to 56) 65 (58 to 74)


















































025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12



025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12







025

5075

100

Mor

talit

y (

)

0 2 4

Time since s




6 8 10 12

praying (months)

Actellic CS-BM


025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21



025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21









Time (months)

Hut walls



Mud 48


Mud 09

Hut ceilings











(54 to 83) 113 (102 to 124)

(19 to 69) 114 (99 to 130)

(30 to 90) 160 (135 to 206)

(dagger to 44) 90 (64 to 110)


(74 to 94) 120 (112 to 127)

(99 to 117) 144 (137 to 152)






025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12





025

5075

100

Pre

dict

ed m

orta

lity

()

0 2 4 6 8 10 12





Mortality (95 CI)


P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)


(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)


(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)




025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12






020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12








(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References



P-methyl 0025 40 98

P-methyl 005 40 100

P-methyl 025 40 100

Malathion 5 201 100























being killed


being killed














P-methyl CS B 64 (61 to 68) 72 (69 to 75)




Lambda CS 29 (27 to 33) 37 (34 to 40)




Lambda CS 50 (47 to 53) 59 (57 to 61)

P-methyl CS B 82 (75 to 93) 97 (89 to 107)





P-methyl CS B 40 (35 to 46) 71 (55 to 110)



Lambda CS 11 (08 to 16) 18 (15 to 22)

P-methyl CS B 49 (42 to 56) 65 (58 to 74)


















































025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12



025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12







025

5075

100

Mor

talit

y (

)

0 2 4

Time since s




6 8 10 12

praying (months)

Actellic CS-BM


025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21



025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21









Time (months)

Hut walls



Mud 48


Mud 09

Hut ceilings











(54 to 83) 113 (102 to 124)

(19 to 69) 114 (99 to 130)

(30 to 90) 160 (135 to 206)

(dagger to 44) 90 (64 to 110)


(74 to 94) 120 (112 to 127)

(99 to 117) 144 (137 to 152)






025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12





025

5075

100

Pre

dict

ed m

orta

lity

()

0 2 4 6 8 10 12





Mortality (95 CI)


P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)


(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)


(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)




025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12






020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12








(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References









being killed


being killed














P-methyl CS B 64 (61 to 68) 72 (69 to 75)




Lambda CS 29 (27 to 33) 37 (34 to 40)




Lambda CS 50 (47 to 53) 59 (57 to 61)

P-methyl CS B 82 (75 to 93) 97 (89 to 107)





P-methyl CS B 40 (35 to 46) 71 (55 to 110)



Lambda CS 11 (08 to 16) 18 (15 to 22)

P-methyl CS B 49 (42 to 56) 65 (58 to 74)


















































025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12



025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12







025

5075

100

Mor

talit

y (

)

0 2 4

Time since s




6 8 10 12

praying (months)

Actellic CS-BM


025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21



025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21









Time (months)

Hut walls



Mud 48


Mud 09

Hut ceilings











(54 to 83) 113 (102 to 124)

(19 to 69) 114 (99 to 130)

(30 to 90) 160 (135 to 206)

(dagger to 44) 90 (64 to 110)


(74 to 94) 120 (112 to 127)

(99 to 117) 144 (137 to 152)






025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12





025

5075

100

Pre

dict

ed m

orta

lity

()

0 2 4 6 8 10 12





Mortality (95 CI)


P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)


(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)


(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)




025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12






020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12








(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References









P-methyl CS B 64 (61 to 68) 72 (69 to 75)




Lambda CS 29 (27 to 33) 37 (34 to 40)




Lambda CS 50 (47 to 53) 59 (57 to 61)

P-methyl CS B 82 (75 to 93) 97 (89 to 107)





P-methyl CS B 40 (35 to 46) 71 (55 to 110)



Lambda CS 11 (08 to 16) 18 (15 to 22)

P-methyl CS B 49 (42 to 56) 65 (58 to 74)


















































025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12



025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12







025

5075

100

Mor

talit

y (

)

0 2 4

Time since s




6 8 10 12

praying (months)

Actellic CS-BM


025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21



025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21









Time (months)

Hut walls



Mud 48


Mud 09

Hut ceilings











(54 to 83) 113 (102 to 124)

(19 to 69) 114 (99 to 130)

(30 to 90) 160 (135 to 206)

(dagger to 44) 90 (64 to 110)


(74 to 94) 120 (112 to 127)

(99 to 117) 144 (137 to 152)






025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12





025

5075

100

Pre

dict

ed m

orta

lity

()

0 2 4 6 8 10 12





Mortality (95 CI)


P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)


(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)


(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)




025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12






020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12








(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References











































025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12



025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12







025

5075

100

Mor

talit

y (

)

0 2 4

Time since s




6 8 10 12

praying (months)

Actellic CS-BM


025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21



025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21









Time (months)

Hut walls



Mud 48


Mud 09

Hut ceilings











(54 to 83) 113 (102 to 124)

(19 to 69) 114 (99 to 130)

(30 to 90) 160 (135 to 206)

(dagger to 44) 90 (64 to 110)


(74 to 94) 120 (112 to 127)

(99 to 117) 144 (137 to 152)






025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12





025

5075

100

Pre

dict

ed m

orta

lity

()

0 2 4 6 8 10 12





Mortality (95 CI)


P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)


(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)


(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)




025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12






020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12








(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References

025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21



025

5075

100

Mor

talit

y (

)

0 3 6 9 12 15 18 21









Time (months)

Hut walls



Mud 48


Mud 09

Hut ceilings











(54 to 83) 113 (102 to 124)

(19 to 69) 114 (99 to 130)

(30 to 90) 160 (135 to 206)

(dagger to 44) 90 (64 to 110)


(74 to 94) 120 (112 to 127)

(99 to 117) 144 (137 to 152)






025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12





025

5075

100

Pre

dict

ed m

orta

lity

()

0 2 4 6 8 10 12





Mortality (95 CI)


P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)


(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)


(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)




025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12






020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12








(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References





025

5075

100

Mor

talit

y (

)

0 2 4 6 8 10 12





025

5075

100

Pre

dict

ed m

orta

lity

()

0 2 4 6 8 10 12





Mortality (95 CI)


P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)


(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)


(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)




025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12






020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12








(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References


Mortality (95 CI)


P-methyl EC Mud 866 805 725 628

(839 to 894) (778 to 833) (679 to 772) (544 to 712)


(777 to 844) (737 to 798) (671 to 766) (583 to 743)

Mud 896 853 794 720

(873 to 920) (829 to 876) (754 to 834) (645 to 796)


(793 to 856) (771 to 826) (729 to 810) (670 to 805)

Mud 839 798 750 695

(809 to 869) (771 to 826) (706 to 794) (620 to 770)




025

5075

100

Blo

od fe

edin

g (

)

0 2 4 6 8 10 12






020

040

060

080

010

0012

00

Mos

quito

s ca

ught

(n)

0 2 4 6 8 10 12








(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References




(95 CI) (95 CI) (95 CI) (95 CI)


(87 to 93) (77 to 85) (52 to 63) (86 to 100)

P-methyl EC Mud 69 24 52 36 33 43 67 29

(64 to 74) (47 to 57) (28 to 37) (49 to 84)


(66 to 76) (44 to 54) (28 to 37) (73 to 96)

Mud 66 26 50 39 31 46 47 49

(61 to 72) (44 to 55) (26 to 35) (26 to 69)


(63 to 73) (43 to 53) (25 to 33) (45 to 81)

Mud 67 26 49 39 31 46 63 32

(61 to 72) (44 to 54) (27 to 35) (44 to 82)







Untreated(Mud)


Total Caught 92

Mortality 1

95 CI (1 to 6)

Blood-fed 94

BFI -


Mortality 5

95 CI (2-12)

Blood-fed 59

BFI -


Mortality 7

95 CI (3-15)

Blood-fed 69

BFI -





CS-B(Concrete)

CS-BM(Concrete)

CS-B(Mud)

CS-BM(Mud)

EC(Mud)

181 204 143 170 115

65 67 78 74 29

(51 to 77) (45-83) (63-88) (61-83) (13-51)

32 30 19 38 63

66 68 80 60 33

592 870 576 685 629

34 42 48 63 43

(27-42) (33-51) (36-59) (46-77) (31-55)

48 53 51 42 52

19 10 14 29 12

557 455 390 498 580

48 49 49 53 46

(41-55) (38-60) (38-59) (41-64) (37-55)

47 46 51 45 54

32 33 26 35 22




































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References



































































Abstract

Background

Methods

Results

Conclusions

Background

Methods





Fumigant activity




Results





Discussion

Ethical approval

Competing interests


Acknowledgements

Author details

References

Long-lasting control of by a single spray application of micro … · 2017-04-05 · RESEARCH Open...

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