A Novel Synthetic Odorant Blend for Trapping of Ma

Authors: Wolfgang R Mukabana Collins K Mweresa Bruno Otieno Philemon Omusula Renate C Smallegange Joop J A van Loon Willem Takken

Publish Date: 2012/03/18

Volume: 38, Issue: 3, Pages: 235-244

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Abstract

Estimating the biting fraction of mosquitoes is of critical importance for risk assessment of malaria transmission Here we present a novel odorbased tool that has been rigorously assessed in semifield assays and traditional African villages for estimating the number of mosquitoes that enter houses in search of a blood meal A standard synthetic blend SB consisting of ammonia Slactic acid tetradecanoic acid and carbon dioxide was complemented with isovaleric acid 45 dimethylthiazole 2methyl1butanol and 3methyl1butanol in various combinations and concentrations and tested for attractiveness to the malaria mosquito Anopheles gambiae Compounds were released through low density polyethylene LDPE material or from nylon strips nylon Studies were done in a semifield facility and two traditional villages in western Kenya The alcohol 3methyl1butanol significantly increased the attraction of SB The other compounds proved less effective or inhibitory Tested in a village 3methyl1butanol released from LDPE increased the attraction of SB Further studies showed a significantly enhanced attraction of adding 3methyl1butanol to SB compared to previouslypublished attractive blends both under semifield and village conditions Other mosquito species with relevance for public health were collected with this blend in significantly higher numbers as well These results demonstrate the advent of a novel reliable odorbased sampling tool for the collection of malaria and other mosquitoes The advantage of this odorbased tool over existing mosquito sampling tools is its reproducibility objectiveness and relatively low cost compared to current standards of CDC light traps or the human landing catchBloodquesting mosquitoes mainly rely on olfactory cues to locate their hosts Takken 1991 Costantini et al 1996 Takken and Knols 1999 For the principal African malaria vectors Anopheles gambiae Giles sensu stricto An arabiensis Patton and An funestus Giles these cues are strongly reminiscent of those released by humans ie the principal source of the mosquitoes’ blood meals Gillies and Coetzee 1987 Odorants from human skin and carbon dioxide from breath are particularly important Costantini et al 1996 Mukabana et al 2004 Spitzen et al 2008 The anthropophilic malaria mosquito Anopheles gambiae sensu stricto hereafter termed An gambiae primarily takes blood meals on humans whereas its close sibling An arabiensis is more opportunistic feeding on humans and animals This difference in host preference is expressed clearly in odorguided behavior where An arabiensis responds significantly stronger to carbon dioxide from breath than does An gambiae Costantini et al 1996 These behavioral differences also are reflected in the role of these species as malaria vectors An gambiae being the principal species even though An arabiensis is equally susceptible to the Plasmodium parasite but due to its different feeding behavior is of less importance as a vector Host preference and hence the selection of hosts based on their odorants is of principal importance to understand the role of these mosquitoes in malaria epidemiologyGiven the strong association of An gambiae with humans unraveling the odor cues that mediate this behavior is of scientific as well as practical importance For An gambiae the principal olfactory cues of humans originate from the feet De Jong and Knols 1995 and recent work has demonstrated that these cues are partially produced by the microbial flora present on the feet Verhulst et al 2010 From these studies several chemical compounds have been identified that play a critical role in the odormediated behavior of An gambiae Smallegange et al 2005 2009 Verhulst et al 2011a Detailed information on the role of these compounds allows for the development of synthetic blends that can be used to better understand the hostseeking behavior of this mosquito Such blends also have the potential to be used for mosquito surveillance or intervention through mass trappingStudies on the development of kairomones for malaria vectors have demonstrated strong behavioral responses to synthetic blends of human odorant compounds Smallegange et al 2010a In laboratory and semifield studies these blends attract a large proportion of hostseeking mosquitoes but when tested against a natural human host or against natural human odorants released from a nylon matrix these blends demonstrate poor competitive characters compared to the natural odorants This suggests that either the concentration of the odorants in the blend was insufficient or that one or more compounds to make the blend sufficiently competitive were missing Smallegange et al 2005 2010a Verhulst et al 2011a Recent progress however has demonstrated the development of odor blends that approach the attractiveness of natural human skin odorants Okumu et al 2010bIn addition there is sometimes a mismatch between laboratory behavioral results and field trials which can be caused by differences in concentration or spatial effects Verhulst et al 2009 2011a Assessment of existing synthetic attractant compounds under semifield and field conditions provides a potential for the development of technologies that can be used for sampling and control of malaria mosquitoes Kline 2006 Jawara et al 2009 Okumu et al 2010b The current study was designed to evaluate the attractiveness of selected synthetic blends and human hosts to hostseeking mosquitoes in western Kenya with emphasis on the malaria vectors An gambiae sensustricto and An arabiensis Thus the comparative trapping efficacy of the attractant blends and the physiological status of the mosquitoes trapped were investigatedBehavioral responses of mosquitoes to synthetic attractants were evaluated under field and semifield conditions The semifield experiments utilized a laboratory colony of the Mbita strain of An gambiae Aquatic stages of the mosquitoes were reared under ambient atmospheric conditions in screenwalled greenhouses at the Thomas Risley Odhiambo TRO campus of the International Centre of Insect Physiology and Ecology icipe located near Mbita Point township in western Kenya Mosquito eggs were placed in plastic trays containing filtered water from Lake Victoria All larval instars were fed on Tetramin® baby fish food supplied three times a day Pupae were collected daily transferred to adult holding rooms and placed in meshcovered cages 30 × 30 × 30 cm prior to adult emergence Adult mosquitoes were fed on 6 glucose solution through wicks made from adsorbent tissue paper Mosquitoes used for semifield experiments were placed in mosquitogauze covered plastic cups and starved for 8 hr They had no prior access to a blood meal Only water availed on wet cotton towels placed on top of mosquito holding cups was provided during starvation All semifield experiments were carried out at night 2030–0630 h inside the screenhouses Verhulst et al 2011b Two hundred 8hstarved mosquitoes were used each experimental nightAll chemicals used to constitute the synthetic attractant blends in this study with the exception of carbon dioxide water sugar and yeast were purchased from SigmaAldrich Chemie GmbH Germany The chemicals included propionic acid 996 butanoic acid ≥999 pentanoic acid 99 heptanoic acid 98 octanoic acid ≥99 tetradecanoic acid ≥99 ammonia solution purity 25 Slactic acid 85 isovaleric acid 998 45dimethylthiazole 97 2methyl1butanol 99 a racemic mixture of the R and S isomers of unknown ratio and 3methyl1butanol purity ≥ 985 Carbon dioxide was produced by mixing 250 g sucrose Sony sugar company Limited Kenya 175 g dry yeast Angel Yeast company limited China and water 2 L as described in Smallegange et al 2010bField studies were carried out in Lwanda and Kigoche villages of Homa Bay and Kisumu counties of western Kenya respectively Lwanda village is located on the southern shore of the Winam Gulf of Lake Victoria 00°28′28″S 34°17′22″E at an altitude of 1169 m above sea level Verhulst et al 2011a Average rainfall and relative humidity are 1200 mm and 65 respectively The mean temperatures vary between 18°C and 34°C Hoof prints of cattle and nightgrazing hippopotami provide excellent mosquito breeding sites in Lwanda Fishing and livestock keeping are the main occupation of the local inhabitants Kigoche village lies adjacent to the Ahero rice irrigation scheme 00°08′19″S 34°55′50″E at an altitude of 1160 m above sea level Kigoche has an average annual rainfall of 1000–1800 mm and an average relative humidity of 65 Mean annual temperatures in the area vary between 17°C and 32°C Rice cultivation is the main occupation of the inhabitants Most houses in the two villages are mudwalled with open eaves have corrugated ironsheet roofs have no ceiling and are either single or doubleroomed Eaves about one foot wide increase ventilation in the houses and form the predominant entry points for mosquitoes Snow 1987 Lindsay and Snow 1988 Malaria caused by Plasmodium falciparum is endemic in the two villages The villages experience two rainy seasons between April–June and September–October During these periods mosquito breeding grounds proliferate and mosquito populations rapidly increase in size Cattle goats chicken dogs cats and a few sheep constitute the domestic animal population with cattle being most abundant Maize millet and sorghum are cultivated at subsistence level in Lwanda whereas rice is a main cash crop in Kigoche

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