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The function of food volatiles: insect behaviour and pest control

Christian Olsson

Tryck:Reprocentralen

Lunds UniversitetLund 2001

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Contents

1. INTRODUCTION 51.1 Problems associated with stored product insects 51.2 Target species 5

1.2.1 Pyralidae (Lepidoptera) 51.2.2 Tenebrionidae (Coleoptera) 6

2. FOOD AND FEEDING OF STORED PRODUCT INSECTS 72.1 Infested food types 71.2 Diet requirements 7

3. FOOD VOLATILES 103.1. Identification of food volatiles 10

3.1.1 Sampling 113.1.2 Recording antennal activity 113.1.3 Chemical identification 133.1.4 Behavioural confirmation 13

3.2 Behaviours induced by food volatiles 143.2.1 Ephestia cautella 143.2.2 Ephestia kuehniella 143.2.3 Plodia interpunctella 153.2.4 Tribolium castaneum 153.2.5 Tribolium confusum 17

4. NON-VOLATILE FOOD COMPOUNDS 174.1 Definition 174.2 Behaviours induced by non-volatile food compounds 17

4.2.1 Pyralidae 174.2.2 Tenebrionidae 18

5. FOOD VOLATILES IN INTEGRATED PEST MANAGEMENT 195.1 General use of food volatiles in IPM 195.2 Effects of food volatiles in combination with pheromones 21

5.2.1 Usefulness of combining food attractants and pheromones 215.2.2 General insect examples 215.2.3 Stored product insect examples 24

6. CONCLUSIONS AND FUTURE PERSPECTIVES 267. ACKNOWLEDGEMENTS 278. REFERENCES 27

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1. INTRODUCTION

1.1 Problems associated with stored product insectsDue to great economical losses, caused by stored product insects, control of pests

infesting warehouses, factories, ships and mills is of main interest for a lot of foodmanufacturers and distributors (Levinson & Levinson, 1978). It is estimated that 35%

of crops all over the world are destroyed by insect pests (Shani, 2000). The use of

pesticides has been the major method of pest control, but extensive use has madestrains of the target insects resistant to, e.g. malathion (Schaafsma, 1990),

methylbromide (Taylor, 1994) and phosphine (Zettler & Arthur, 1997), and it has led

to environmental pollution and health hazards to non-target organisms, includinghumans. The demand for pest control without, or at least with less, pesticides is great

and biorational pest management methods have become abundant (Phillips, 1997).This concept of controlling pest populations includes use of natural enemies of the

pest, such as parasitoids, pathogens and predators (Brower, 1988; McGaughey &

Berman, 1988; Johnson et al., 1998); temperature regulation (Lhaloui et al., 1988;Lewhtwaite et al., 1998; Na & Ryoo, 2000), protective packing of the stored products

and pheromone-based methods. Pheromones are known of over 35 species of storedproduct insects (listed in Burkholder & Ma, 1985 and Phillips, 1994) and they are used

for both monitoring and direct control in the field. Since the majority of the synthetic

pheromone compounds used, however, is originally produced by females with thepurpose of attracting males for mating, the pheromone-based methods mainly control

the males of a pest population (Trematerra & Battaini, 1987) and the unaffected malescan still maintain a high population density (Foster & Harris, 1997). To efficiently

reduce the population density it is important to find a control method that targets the

females of a population, or perhaps even better, a control method affecting both sexessimultaneously.

1.2 Target species1.2.1 Pyralidae (Lepidoptera)

The almond moth (Ephestia [Cadra] cautella, Figure 1), the Mediterranean flour moth(E. kuehniella, Figure 2) and the Indian meal moth (Plodia interpunctella, Figure 3)

belong to the subfamily Phycitinae in the Pyralidae family and are well-known

cosmopolitan indoor pests active during night. Lepidopteran pests are a very diverse

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group found all over the world and in a variety of

artificial habitats (Table 1). The original habitats of the

moths are unknown since the moths are rarely foundoutdoors. Only a few outdoor trapping experiments have

been reported and the captures are always in the vicinityof production or storage facilities and usually directly

after insecticide fumigation (Doud & Phillips, 2000).

The moths feed as larvae, whereas all other stages arenon-feeding. Hence, most damage associated with

stored product moths is due to the larvae of the moths.

Larvae cause direct damage to the accumulation ofgrains by feeding and producing faeces and debris. Also

webbing causes problems by forming clumps of grainthat clog machinery (Wool et al., 1987). Laboratory

experiments with moth larvae feeding on wheat have

shown that at least 60% weight is lost and that no seedsgerminate after seven days of feeding (Madrid & Sinha,

1982). They also cause indirect damage by making theproducts less attractive for the consumers since pest

infestation is associated with bad hygien.

1.2.2 Tenebrionidae (Coleoptera)

Flour beetles belong to the Tenebrionidae family withinthe order Coleoptera and are the most common group of

stored product insects all over the world. The two most

abundant flour beetles are the rust-red flour beetle(Tribolium castaneum, Figure 4) and the confused flour

beetle (T. confusum, Figure 5). Tribolium beetles areconsidered secondary pests, meaning that they only

infest already damaged and deteriorated grains

(Trematerra et al., 2000). The fact that they feed both aslarvae and as adults make them a more complicated

problem than the moths and methods must be developed

to control both stages.

Figure 2. Mediterranean flourmoth, Ephestia kuehniella

Figure 1. Almond moth, Ephestia(Cadra) cautella

Figure 3. Indian meal moth,Plodia interpunctella

Figure 4. Rust-red flour beetle,Tribolium castaneum

Figure 5. Confused flour beetle,Tribolium confusum

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2. FOOD AND FEEDING OF STORED PRODUCT INSECTS

2.1 Infested food typesStored products are susceptible to insect infestation, since accumulations of potential

food have a high attractivity to animals. Infested products range from newly harvestedraw products; e.g. fruits, nuts, seeds and spices; to semimanufactured products; e.g.

milled cereals and dried products, and finally to end products; e.g. crackers, oils and

pasta (Table 1).Larvae of both E. cautella and P. interpunctella feed on a variety of diets with

diverse origins. E. cautella feeds on grains, cocoa, peanuts, tree nuts, dried fruit and

meal (Wool et al., 1987). P. interpunctella feeds on cereal products, nuts, almonds,cocoa beans, chocolate, leguminous seeds, dried fruits, plant tissue and tobacco

(Levinson & Levinson, 1978), sunflower seeds, birdseeds and dog food (Arbogast etal., 2000). These two moths, together with the closely related Ephestia elutella, are

usually included in the group ”chocolate moths” because they cause serious problem in

the chocolate industry. E. kuehniella is more restricted in its food preference and rarelyinfests other stored products than flour (Benson, 1973; Levinson & Levinson, 1978).

Both T. castaneum and T. confusum are extremely polyphagous but showgreatest preference for cereal products and seeds since these groups contain the diet

requirements of the beetles (Levinson & Levinson, 1978). The beetles can also feed

on, e.g. meat meal, yeast, dried figs and fish meal and still maintain a high growth andreproductive rate (Fraenkel & Blewett, 1943).

2.2 Diet requirementsStored product insects have developed specific requirements concerning the

constituents of their diets. Due to low water content of dried food products theseinsects have characteristically adapted to require less water in their diets (Levinson &

Levinson, 1978). Water is nevertheless important for stored product insects. For E.

cautella, but not for E. kuhniella, drinking water had a pronounced effect on fecundity

of the females, which was increased with at least 100% when water was provided to

the moths (Norris, 1934). The water drinking also increased the longevity, for bothfemales and males, with more than 100%. In full-scale trapping experiments with E.

cautella and P. interpunctella in a storage house in Taiwan, traps baited with water and

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Table 1. A selection of common coleopteran and lepidopteran stored product insects and their foodpreferences.Scientific name Common name Main food preference ReferenceColeoptera

AnobiidaeLasioderma serricorne Cigarrette, tobacco

beetleTobacco; wheat, yeast,locust bean,groundnuts

Munro, 1966;Ashworth, 1993;Chuman et al., 1995

AnthribidaeAraecerus fasciculatus Coffee bean weevil Cassava chips Kumar et al., 1996BostrichidaeDinoderus minutus Bamboo powderost

beetleCassava chips,harvested bamboo

Åkerlund, 1991

Prostephanus truncatus Larger (greater)grain borer

Corn Houseman & Thie,1993

Rhyzoperta dominica Lesser grain borer Cassava chips, cerealgrains

Tata-Hangy &Lutete, 1997

ChrysomelidaeCallosobruchuschinensis

Pulse beetle Dried peas Munro, 1966

Callosobruchusmaculatus

Cowpea weevil Dried peas,leguminous seeds(larvae),pollen, nectar (adults)

Munro, 1966Levinson &Levinson, 1978

CucujidaeCryptolestes pusillus Flat (Rust-red) grain

beetleWheat, yeast, rolledoat

Munro, 1966

Oryzaephilus mercator Merchant grainbeetle

Cereal seeds andproducts, oilseeds,dried fruit

Levinson &Levinson, 1978

Oryzaephilussurinamensis

Sawtoothed grainbeetle

Cereal seeds andproducts, dried fruit,animal tissues

Levinson &Levinson, 1978

CurculionidaeSitophilus granarius Grain, Granary

weevilGrains, crackers Åkerlund, 1991

Sitophilus oryzae Rice weevil Whole wheat kernels,cereals, pasta, peas

Phillips et al., 1993;Trematerra &Girgenti, 1989;Trematerra et al.,1999

Sitophilus zeamais Maize weevil Corn Houseman & Thie,1993

DermestidaeTrogoderma granarium Khapra beetle Barley, beans, yeast Munro, 1966Trogoderma variabile Warehouse beetle Barley, locust beans,

yeastMunro, 1966

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NitidulidaeCarpophilus hemipterus Driedfruit beetle Ripening and dried

fruitsBartelt et al., 1990

TenebrionidaeTenebrio molitor Yellow mealworm Cereal products Åkerlund, 1991Tribolium castaneum (Rust-) Red flour

beetleCereals, dried fruits,spices, millet,damaged wheatkernels

Sokoloff, 1972Seifelsnasr et al.,1982Phillips et al., 1993

Tribolium confusum Confused flourbeetle

Cereals, dried fruits,spices

Sokoloff, 1972

Tribolium destructor Dark flour beetle Cereal products, driedproducts, dead insects

Åkerlund, 1991

TrogositidaeTenebroidesmauritanicus

Cadelle Cereal seeds andproducts, animaltissues

Levinson &Levinson, 1978

LepidopteraGelechiidaeSitotroga cerealella Angoumois grain

mothWheat, corn, nuts,almonds, leguminousseeds

Munro, 1966;Levinson&Levinson, 1978; Ge& Weston, 1995

PyralidaeAmyelois transitella Navel orangeworm Walnut Johnson et al., 1998Corcyra cephalonica Rice moth Rice Hall et al., 1987Ephestia cautella Almond (Tropical

warehouse) mothGrains, nuts, driedfruit, peanuts, tree nuts

Benson, 1973;Wool et al., 1987;Gothilf et al., 1993

Ephestia elutella Tobacco, Cocoa,Warehouse moth

Grain, cocoa, driedfruit, nuts, tobacco

Benson, 1973

Ephestia figulilella Raisin moth Dried fruits Benson, 1973Ephestia kuehniella Mediterranean flour

mothFlour Benson, 1973

Plodia interpunctella Indian meal moth Walnut, wheat, cocoabeans, chocolate, driedfruits, plant tissues,

Levinson &Levinson, 1978;Madrid & Sinha,1982

Pyralis farinalis Meal moth Wheat, plant tissues Levinson &Levinson, 1978;Madrid & Sinha,1982

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a detergent caught females, of which 99% were fertilized, leading to the conclusion

that water is an attractive resource for gravid females (Chow et al., 1977). This

conclusion can be questioned since the experiments did not show if the attractivity wasdue to the water or the detergent. Female moths need water not only for fecundity and

longevity (Norris, 1934) but also for egg maturation (Chow et al., 1977). Willis andRoth (1950) showed that flour beetles were dependent on the water content of the

food, which additionally emphasizes the importance of water for stored product

insects. Both moths and beetles need sterols and vitamins, especially of the B-group,both for reproduction and survival. Moths also require carbohydrates (Fraenkel &

Blewett, 1943) and lipids in their diets for succesful development of wing scales

(Levinson & Levinson, 1978) and pyridoxine, mainly found in the bran (pericarp) ofcereals, for complete growth and development (Madrid & Sinha, 1982). Beetles breed

well on diets without carbohydrates, but better on diets with carbohydrates (Fraenkel& Blewett, 1943).

In a study on larval development, semolina (a wheat-derived baking product)

and soybean flour were both excellent diets for P. interpunctella, resulting in fasterdevelopment than when wheat alone was used as a diet (Locatelli & Biglia, 1995).

Semolina and soybean flour contain the necessary fatty acids, steroids and vitamins.E. kuehniella moths do not have the same restricted vitamin requirement and were able

to develop successfully on semolina, wheat meal, soybean flour and two basic wheat

flour types. Locatelli and Biglia (1995) also tested different semimanufacturedproducts, containing different mixtures of the individual ingredients. These products

seemed equally attractive to both P. interpunctella and E. kuehniella, and theyconcluded that the infestation risk is higher for semimanufactured products since they

provide all nutrients required for successful larval development, probably for several

insect pests.

3. FOOD VOLATILES

3.1 Identification of food volatilesSubstances or mixtures of substances that emanate from food and disperse easily

through air, due to high volatility, are commonly denoted food volatiles. These

compounds consist of a chain of 5-20 carbon atoms and a variety of functional groups,

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e.g. ketones, aldehydes, esters and alcohols. The volatile compounds are working in

long-distance attraction due to the high and fast dispersing ability, and can be

differentiated by their effect on the recieving insect; attractants, causing orientedmovement towards the source of the chemical, and repellents, causing oriented

movement away from the source (Dethier et al., 1960).

3.1.1 Sampling

There are two well-established ways to obtain the chemicals/volatiles from a putativeodour source (Golub & Weatherston, 1984 and references therein). The first is direct

extraction by mixing the sample, e.g. a pheromone gland or an aliquot of food, with a

solvent and then filter and distill the mixture to get a concentrated distillate with thevolatiles (e.g. Loschiavo, 1965, Heydanek & McGorrin, 1981). The major

disadvantage with this method is that a large fraction of the extracted chemicals maynot be interesting, from an olfactory point of view, since they are nonvolatiles, not

functioning as olfactory cues. This problem is avoided in the other sampling

alternative, where the approach is to capture the volatiles emanating from the odoursource on an odour trap, e.g. an adsorptive filter, and then extract the filter with a

solvent. This method, called headspace sampling, has been employd both inpheromone and food volatile identification (Hougen, 1971; Maga, 1978; Shani, 1990).

3.1.2 Recording antennal activityIn order to find behaviourally active compounds the electroantennogram (EAG) assay

is a widely used method, especially in pheromone research (Roelofs, 1984). Bystimulating an antenna with a volatile compound the olfactory receptor potential, as a

result of receptor membrane depolarization, can be measured. Compounds eliciting a

potential larger than the spontaneous antennal activity, are considered to beelectrophysiologically active. The EAG method is conclusive in that sense that a

compound not eliciting an electrophysiological response, can be excluded as anolfactory cue, provided that olfaction is mediated by receptors on the antenna only. In

olfactory research the EAG technique has been mostly used for lepidopteran species

but also for coleopteran species of stored product insects. The pheromoneidentification of Corcyra cephalonica, a pest infesting rice and rice products (Hall et

al,. 1987) and the identification of host plant volatiles of Ostrinia nubilalis, a corn pest

attacking plants in the field (Marion-Poll & Thiéry, 1996) are two examples where the

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EAG technique has been successful for lepidopterans. For flour beetles it has been

reported that the R,R-isomer of 4,8-dimethyldecanal was the main component of the

aggregation pheromone for both T. castaneum and T. confusum (Levinson & Mori,1983), which was concluded from electrophysiological and confirmed by behavioural

experiments. The EAG technique has been used for identification of volatiles fromstored products as well, e.g. for the stored product insect Oryzaephilus surinamensis

(Coleoptera, table 1) EAG has been used to identify active volatiles from oat (White et

al. 1989).To be able to identify specific components in e.g. extracts, the GC-EAD (gas

chromatograph combined with an electroantennographic detector) technique is used,

where the responses from the GC and the antenna are recorded simultaneously. Theability to distinguish between active and non-active peaks in a GC-trace is essential in

the work with food volatiles, since 100-200 peaks are not unusual when analysingextracts of cereal products (Figure 6). The technique was developed by Moorhouse et

al. (1969) while working on the red bollworm, Diparsopsis castanea. From GC-EAD

runs with female gland extracts the authors concluded that the male moths could detect

five different substances in the female extract. Another good example of simultaneous

A

B

Figure 6. Gas chromatograms of extracts from oat (A) and wheat (B). The GCs evidentlyshow the high number of volatiles emanating from stored products. From Heydanek &McGorrin, 1981 and Nara et al., 1981, respectively.

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recording was the experiment with pheromone detection in Lobesia botrana, a wine

pest, where a peak of an active component was found in extracts of females (Arn et al.,

1975). GC-EAD has since been extensively used also for pheromone identification instored product insects, e.g. with Plodia interpunctella, whose pheromone blend

consists of at least four components (Zhu et al., 1999).

3.1.3 Chemical identification

Once the electrophysiologically active peaks of a gas chromatogram are determinedthey can be identified with coupled gas chromatograph-mass spectrometry (GC-MS)

(Heath & Tumlinson, 1984). Gas chromatography is an excellent method for

separating and quantifying components of an extract and it is possible by the means ofmass spectrometry to identify the separated components, making the combination a

very powerful tool (Rose, 1990). Identification work with GC-MS has been carriedout for some volatiles of the stored products susceptible to pest infestation, e.g. for

maize (Hougen et al., 1971), oat (Heydanek & McGorrin, 1981), sesame seeds

(Shimoda et al., 1996), Argentinean peanuts (Burroni et al., 1997), and earth-almonds(Cantalejo, 1997). All these studies were pure identification studies with no connection

to the biological relevance of the volatiles. Some general identification studiesincluding induced insect behaviours have been done on oat with O. surinamensis

(White et al., 1989) and on wheat germ with Trogoderma glabrum (Nara et al., 1981).

In both cases did the volatiles cause an aggregation of the tested animals.

3.1.4 Behavioural confirmationThe chemical identity and electrophysiological activity of a compound are of little

value if no behavioural response can be associated with it. Hence, the concluding step

in the identification process is to test the identified and synthesized substance, toconfirm its impact on the investigated species. As mentioned above, in the

electrophysiological work with the different isomers of 4,8-dimethyldecanal, theaggregation pheromone of flour beetles, the achieved results were confirmed in a

behavioural bioassay (Levinson & Mori, 1983). In another coleopteran example the

identified oat volatiles 1-octen-3-ol, 3-octanone and nonanal were tested behaviourallyin dose-response experiments, and synthetic 1-octen-3-ol and nonanal attracted both

sexes of the Oryzaephilus surinamensis beetles (White et al., 1989). In a subsequent

study it was confirmed that nonanal was attractive to both O. surinamensis and O.

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mercator, a closely related species (Pierce et al., 1990). In their study also several

other oat volatiles, such as aldehydes and fatty acids, attracted the beetles. A mixture

of hexanal, octanol and nonanal elicited the highest response. More than 100components have been detected in oat, and aldehydes and fatty acids constitute large

groups of the naturally occuring chemicals in oat and are common products of lipidoxidation (Mikolajczak et al., 1984). Pierce et al. (1990) concluded that these two

cucujid beetles use oat volatiles for host-finding and that the three-compound aldehyde

mixture would be a good alternative to the pheromone as a trap bait.

3.2 Behaviours induced by food volatiles3.2.1 Ephestia cautella

Odours from 2 g medium used for rearing, including whole wheat, rolled oats, wheat

bran and dried yeast, influence the choice of oviposition site of gravid E. cautella

females (Barrer, 1977). Barrer & Jay (1980) showed that gravid females of E. cautella

both orientate and oviposit in response to wheat odour. When testing attraction for

female E. cautella in a two-choice bioassay, wheat was significantly superior inattraction to a variety of alternative dried fruits (Gothilf et al., 1993). Wheat was the

major ingredient in their lab rearing diet and the high behavioural responses could be aresult of food adaptation for several generations in the lab or of wheat being an

extraordinary suitable food source for E. cautella providing all necessary diet

requirements. The only dried fruit type that was more attractive than wheat was date,which was not really explained, but it was hypothesized that date had earlier been a

natural host in the wild for E. cautella. Other stored products, e.g. almond, peanut,walnut and corn, were less attractive than wheat, although not significantly so.

3.2.2 Ephestia kuehniella

A variety of studies have shown how some components of fitness of the moths, e.g.

larval development, reproduction and survival, are related to their food preference andhabits (Fraenkel & Blewitt, 1943; Benson, 1973; Levinson & Levinson, 1978;

Locatelli & Biglia, 1995). However, the importance of food odours responsible for

induced behaviours of E. kuehniella has not been of main interest in stored productresearch. Compared to other closely related moth species, E. kuehniella is a

”specialist” and therefore its olfactory system may be tuned to very specific odours

associated with only a few suitable types of food. Hence, the work with identification

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of food volatiles can perhaps focus on fewer sources of behaviourally active odours,

making research less time consuming.

3.2.3 Plodia interpunctella

Gravid females are attracted for oviposition by extracts of roasted almonds andhazelnuts but not by extracts of roasted cocoa beans (Hoppe, 1981). In addition nut-

containing chocolate products were more attractive than mere chocolate, indicating

that there are some attractive substances in almonds and nuts alone. Volatiles of nutsand almonds, responsible for the induced oviposition behaviour, have not yet been

identified. Also the development of the moths is greatly affected by the presence or

absence of nuts in the diet, resulting in no development at all in chocolate productswithout nuts.

Four-day-old gravid females of Plodia interpunctella were shown to lay eggsin a rearing diet based on corn meal (Phillips & Strand, 1994). They tested rearing diet

(corn meal, chick starter mash and glycerol), both in a two-choice bioassay for

oviposition, using patches of cheesecloth contaminated with rearing diet and forupwind flight in a wind tunnel, using 10 g of rearing diet as bait. The rearing diet

stimulated oviposition and induced upwind flight, indicating attractive cues in the foododour, mediated by olfaction, the only host-finding modality tested for this species

(Ramaswamy, 1988).

3.2.4 Tribolium castaneum

Adults of both sexes of Tribolium castaneum were reported to be highly attracted tovolatiles from whole wheat flour (Willis & Roth, 1950). Their study was the first to

confirm that Tribolium beetles use olfaction to orient to a food source, since physical

and visual contact with the food were prohibited in their experiments. Seifelnasr et al.(1982) continued the work with food volatiles from wheat and also included millet in

their studies. They found that both sexes of T. castaneum were attracted to differentfractions of both wheat and millet. All fractions of wheat tested; whole kernels, whole

flour, germ, bran and endosperm, elicited response of the beetles. Also diethyl ether

extracts of the wheat kernels were attractive to the beetles. All fractions of millet,whole kernels, whole flour, fermented flour and starch, did also attract the beetles, as

did millet extracts. Strangely enough, virgin females were the best responding group,

which was left unexplained by Seifelnasr et al. (1982). One reason could be that the

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female beetles associate food with suitable partners and orient to the food odours with

the purpose of mating.

An extract from a blend of stored products; cashew nuts, groundnuts, carobs,maize, almonds, raisins, wheat, rye, barley, oats and millet was proven attractive

particularly for T. castaneum when tested on a filter paper disc in a bioassay, causingthe beetles to aggregate (Levinson & Levinson, 1978). One hypothesis is that some of

these stored products contain substances that, when emitted, are responsible for the

behaviour of the beetles and that identification work could determine which substancesare acting on the beetles. In a spin-off experiment it was found that the grain-derived

compounds valeraldehyde, maltol and vanillin, which are all common in several

species of grain, e.g. corn, rye, wheat, barley and rice (Maga, 1978), were attractive tothe rice weevil, Sitophilus oryzae, a species infesting the same habitats as T. castaneum

(Phillips et al, 1993). Unexpectedly, the same compounds were not attractive to T.

castaneum. Instead the T. castaneum beetles were attracted to oils from pressed stored

products, such as rice, soybean, oat, wheat germ and corn, here listed according to

increasing attractivity. These oils did not elicit any response of the S. oryzae weevilsand the difference in attraction is probably because S. oryzae is a primary pest,

infesting whole, undamaged grain, whereas T. castaneum is a secondary pest, infestingalready damaged grains (Trematerra et al., 2000). The odour composition of the grain

is reflected by the degree and type of damage. Trematerra et al. (2000) used 20 beetles

of mixed sex in the bioassay, which can be a drawback since the pheromone attractionbetween sexes and individuals is not considered.

When tested in an two-choice olfactometer, wheat and sorghum were the twocereals with the most attractive odours for adult T. castaneum of both sexes compared

to rice, barley, corn, and millet (Bekon & Fleurat Lassard, 1988). The beetles seem to

develop best when feeding on wheat or sorghum, with high pupal weight andfecundity, explaining the high attraction to volatiles from these two cereal species. In

another study where food preference was coupled to fitness, Bergerson & Wool (1987)concluded that standard medium (wheat flour and Brewer´s yeast) was the most

attractive stimulus when tested in an olfactometer. Even when the beetles were

selected for and cultured on a totally different diet they still preferred standardmedium, probably showing a strong adaptation to the standard diet after 60 years of

breeding in the laboratory. The attractiveness of standard medium was due to flour

odour since standard medium did not attract significantly more beetles than flour

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alone, when presented in a two-choice olfactometer. Again, however, the possible

interactions between the beetles were not considered, since 80-100 beetles of mixed

sexes were released simultaneously in the olfactometer.

3.2.5 Tribolium confusum

Different kinds of extracts from wheat flour elicit both aggregation and feeding

behaviour of both sexes of the confused flour beetle (Loschiavo, 1965). The highest

responses were obtained with a benzen extract of whole flour, ethanol extracts ofwheat bran and an ether extract of wheat germ, indicating that behaviourally active

volatiles are present in these cereals.

4. NON-VOLATILE FOOD COMPOUNDS

4.1 DefinitionNon-volatile compounds are larger molecules with lower vapour pressure, which donot spread easily in air. Therefore, they are acting over short distances and the induced

behaviour is mediated by contact chemoreception (Foster & Harris, 1997). Thesesubstances are divided into stimulants, causing feeding or oviposition, or deterrents,

inhibiting feeding or oviposition (Dethier et al., 1960). A common and general feeding

stimulant is sucrose, which induces feeding in many insect species (Schoonhooven,1982).

4.2 Behaviours induced by non-volatile food compounds4.2.1 Pyralidae

For Amyelois transitella, a pest infesting cultures of different nuts, oil extracts fromboth almond and peanuts acted as attractants for mated females (Phelan et al., 1991).

They concluded that there were some compounds in the oil acting as cues foroviposition, since neither males nor unmated females were attracted. The compounds

mediating the oviposition were identified as oleic acid and linoleic acid, two fatty acids

found in almost all animal and vegetable fats.In an experiment with Plodia interpunctella, with the purpose of studying the

chemicals responsible both for short-range attraction and feeding, extracts of corn,

peanuts and wheat induced both orientation and feeding (Baker & Mabie, 1973). When

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analysing the constituents they found that carbohydrates, mainly sugars, were powerful

feeding stimulants; amino acids had no effect and fatty acids and sterols stimulated

feeding only when presented together with sucrose.

4.2.2 TenebrionidaeFatty acids induce behaviour in T. castaneum beetles. Acids with low carbon atom

numbers (C5-C11) are in general deterrent, whereas larger acids aggregate beetles but

do not induce feeding (Cohen et al., 1974).Wheat contains a lot of chemicals, both volatiles (Hougen et al., 1971 and

Maga, 1978; Figure 6b) and compounds with larger molecular weights, e.g. fatty acids,

proteins and sugars (Loschiavo, 1965). One group of the non-volatile compoundsfound in and isolated from wheat germ is the triglycerides, a lipid consisting of

glycerol esterified with three carboxylic acids and found in both animal fats andvegetable oils. The triglycerides which caused significant aggregation of T. confusum

were identified as 1-palmito-2,3-diolein, 2-linoleo-1,3-dipalmitin and 1-palmito-2-

linoleo-3-olein, all three present in hexane extract of wheat germ (Tamaki et al.,1971a). These triglycerides elicited aggregation both when crude extracts and

synthesized compounds were used in the bioassay. When testing methanol extracts ofwheat germ, probably containing different compounds than hexane extracts, the beetles

responded to the extract only by intensive feeding rather than aggregation, indicating

that the compounds causing feeding and aggregation differ. When testing a largergroup of triglycerides, containing at least one palmitic acid chain often in combination

with the unsaturated linoleic acid, the triglycerides in general seem to attract andaggregate both sexes of adult T. confusum (Tamaki et al., 1971b). The biological

activity of triglycerides is in general affected by the number of double bonds in the

fatty acid moieties. Palmitic acid, stearic acid and oleic acid, common fatty acidmoieties of triglycerides, can induce both aggregation and feeding when being

presented as individual compounds (Loschiavo, 1965). Other feeding stimulants for T.

confusum are maltose and gluten.

19

5. FOOD AND FOOD VOLATILES IN INTEGRATED PEST MANAGEMENT

5.1 General use of food volatiles in IPMFood-baited traps can, to a large extent, be used in the same manner as pheromone-

baited traps, i.e. in detection of pest outbreaks before being visible, monitoring formaking decisions on temporal and spatial distribution of control actions, and direct

control (Pinniger, 1990). When comparing the two different bait types, food and

pheromone, food-baited traps have several advantages over pheromone-baited ones.Food-baited traps can possibly trap both sexes, several species, both adult and larval

insects and they are cheaper (Chambers, 1990; Pinniger, 1990). The advantages of

using pheromone-baited traps are less competition with the background odour andhigher trap catches of the target species (Table 2). The first traps used for sampling of

insects in stored grain were filled with water, but the water was believed to function asa killing agent rather than an attractant for the target insects (Watters & Cox, 1957).

The first example of food used in traps was an attempt to detect infestations by the

Khapra beetle, Trogoderma granarium, a species feeding on cereal products and otherstored products (Strong, 1970). The food bait used contained poultry laying mash,

rolled barley, whole wheat and whole corn, and did not only capture T. granarium butalso over 20 other species associated with stored product infestation, among them

Tribolium beetles and Pyralidae moths, evidently showing the possibilities of

capturing multiple species using one kind of food bait.

Table 2. Comparison between traps baited with food attractants and pheromones. Modified fromPinniger, (1990).Food attractants PheromonesBoth sexes respond Usually only one sex responds

Multi-specific Mainly species-specificLess potent in competition with

surrounding food sources

Often very potent, may be enhanced by

foodMay attract adults and larvae Attract adults only

Simple and cheap chemicals Mixtures of expensive chemicals

May enhance pheromone attractivityin traps

May be inefficient in absense of food

Spin-off from other food technology

20

In food-baited traps the stimulus can be presented in two ways (Rajendran,

1999). The first type of bait is grains of one or several cereal products in cloth, jute or

plastic bags. The bags are placed within the accumulations of grain and the insectstrapped in the bags are counted. Both larvae and walking insects can be caught and are

not able to escape. This system could be used both for detection and monitoring,possibly for multiple species. Food baits from different kinds of stored products have

been tested, e.g. barley, corn, wheat and coffee beans and have attracted and trapped

both beetles and moth larvae. The other type, baiting with oils and distillates fromcereals, has been successful in trapping stored product insects in the laboratory. Oils of

corn, rice, soybean and wheat germ are attractive to adult T. castaneum (Phillips,

1993) and oils of oat, pumpkin and sesame seeds are attractive to larvae of the Khaprabeetle, Trogoderma granarium, and have been tested both in the laboratory and in the

field (Barak, 1989). Food-derived oils, when applied both in pheromone traps andphysical traps, increased the number of captured insects, meaning that both detection

and monitoring could be improved by adding food attractants to the traps.

Odour of stored wheat can increase the trap catches of unbaited traps, for avariety of species, including Tribolium castaneum (Barrer, 1983). By comparing traps

without grain odour to traps with grain odour, he concluded that the trap efficacy for T.

castaneum was increased two-fold when wheat odour was present. The results

confirmed the multispecies function of the food-baited traps and show that the

simplicity of this monitoring system makes it an economically good alternative. Themagnitude of the stimulus presented was equal to the background level, meaning that

an increased source/background-ratio could increase the trap efficacy further more.This could be achieved by identifying the active compounds in wheat and use these

identified compounds in higher doses in traps. Enhancement of trap catching with food

odours was tested and confirmed in a comparative laboratory study with two storedproduct beetles T. castaneum and Cryptolestes ferrugineus (White & Loschiavo,

1986). Although infesting the same stored products, the preferences of the beetlesseemed to differ, trap catches of C. ferrugineus were enhanced only by present

conspecifics, whereas trapping of T. castaneum was increased almost three-fold by

both wheat and insect-baited traps compared to unbaited traps. By combining the twobaits one could possibly expect an additive or synergistic effect, however this was not

tested.

21

5.2 Effects of food volatiles in combination with pheromone5.2.1 Usefulness of combining food attractants and pheromones

Due to one of the major drawbacks of pheromone-baited traps, namely that female-produced sex pheromones only affect males, some trapping experiments have been

done to evaluate the possibility of using pheromone together with food-derivedattractants. In the most successful cases, the addition of food attractants had a

synergistic effect on the trap catches, and both females and males were captured

(Walgenbach et al., 1987; Trematerra & Girgenti, 1989). Synergism in this case meansthat the total trap catch when pheromone and food attractants were presented together

was higher than the sum of catches in the traps where pheromone and food attractants

were presented individually.

5.2.2 General insect examplesOlive fruit fly, Bactrocera oleae

In attempts to control the olive fruit fly, Bactrocera (Dacus) oleae (Diptera:

Tephritidae), the major pest of olives in the Mediterranean region, the combined use offood attractants and pheromone has yielded high catches in mass trapping. (Haniotakis

et al., 1991). Mazomenos & Haniotakis (1981) combined ammonium bicarbonate(food attractant) with a pheromone blend and trapping efficiency, at least for females,

was increased compared to traps baited with food alone. The mass trapping based on

food attractants was also superior to traditional control with insecticides, thus fulfillingthe applied goal of reduced insecticide use. In a subsequent study the combined bait

did catch both females and males (Broumas & Haniotakis, 1994). These authors, aftertesting several different known food chemicals, e.g. ammonium bicarbonate,

ammonium carbonate, ammonium sulphate and hexanodiol, concluded that olive fruit

flies has a preference for more general food odour and that trapping efficiency wassimilar for the different food attractants. The report is partly inconclusive, since the

trapping efficiency of pheromone alone was not tested.

The dried fruit beetle, Carpophilus hemipterus

The dried fruit beetle, Carpophilus hemipterus, (Coleoptera: Nitidulidae) is acosmopolitan pest attacking fruits and grains, both before and after harvest. The beetle

causes damage both by feeding on ripening products and as a vector transmitting

phytopathogenic fungi, responsible for souring of fruits. Field tests showed that several

22

Carpophilus species could be captured in fig orchards with chemicals from fermented

fig, however C. hemipterus was by far the most abundant species and further control

was determined to be most valuable for this species (Smilanick et al., 1978). Bothindividual volatiles and mixtures of volatiles were tested as baits in field traps. A 1:1:1

mixture of acetaldehyde, ethyl alcohol and ethyl acetate was the most attractive bait,with compounds acting synergistically and capturing significantly more beetles than

traps baited with the individual components, other combinations of fig volatiles or,

rather surprisingly, crude fig paste. It was hypothesized that fig paste could containcomponents with masking or repelling effects or that a concentration effect could

explain the unexpected results. In laboratory experiments with C. hemipterus the 3-

component mixture acts as a pheromone synergist (Bartelt et al., 1990a). The maleproduced aggregation pheromone was identified and consisted of two tetraene

hydrocarbons, a major component (2E,4E,6E,8E)-3,5,7-trimethyl-2,4,6,8-decatetraeneand a minor component (2E,4E,6E,8E)-3,5,7-trimethyl-2,4,6,8-undecatetraene. By

adding both the mixture and the two components individually to the food-derived 3-

component blend the captured number of beetles was increased at least 4-fold, but upto 10 times higher trap catches were reported, evidently showing a synergistic effect.

The fig-derived 3-component mixture with acetaldehyde, ethyl alcohol andethyl acetate was questioned by Phelan & Lin (1991) since the release rates of the lures

required to catch any beetles, were much higher than the expected natural release rate

of a fig plants. Therefore, they tested the relative attractiveness of banana, yeast(Saccharomyces cerevisae) and banana inoculated with yeast. In a two-choice

experiment the inoculated banana induced upwind flight for significantly more beetlesthan any other stimuli. By analysing the volatile profile of inoculated banana they were

able to create a synthetic mixture which mimicked the natural substrate both

behaviourally and chemically. A subtractive bioassay led to the conclusion that thechemicals responsive for the upwind flight was ethyl acetate, acetaldehyde, 2-pentanol

and 3-methylbutanol, where acetaldehyde can be replaced by the mixture of 2-pentanon and 3-hydroxy-2-butanone and 3-methylbutanol can be replaced by 2-

methylpropanol or butanol. The main difference from the earlier mixture of food

volatiles, except for some alcohols added, is that ethyl alcohol is not required forinducing upwind flight.

A study by Dowd & Bartelt (1991) is contradictory to the previous in the

sense that the preference of C. hemipterus seems to be more general than earlier

23

expected. They tested 60 different food volatiles and found that nearly all attracted

beetles in the wind tunnel and that even more volatiles acted synergistically together

with the aggregation pheromone, 43 out of 60 individual volatiles had a significantsynergistic effect on beetle attraction. The most effective volatiles used alone were

methanol, methyl butyrate, methyl propinate and propanal, whereas ethyl acetate andpropyl propanoate were the most effective synergists. The authors concluded

”synergism in C. hemipterus is a more generally widespread phenomenon” than earlier

appreciated. They also suggested that the combination of pheromone and host volatilescould be useful in the control of C. hemipterus as well as other nitidulids. The

combination of pheromone and food volatiles has also been tested in the field (Bartelt

et al., 1992). They used a 4-component aggregation pheromone (Bartelt et al., 1990b)and added three types of food attractants, fermented fig juice, whole-wheat bread

dough and semisynthetic volatile mixture (SVM). SVM was a mixture of apple cidervinegar, methanol, methyl butyrate and propanal and the effect was again evident, all

food attractants acted synergistically meaning a higher yield of trap catches. For long

term effects the bread dough and the SVM were more suitable and the SVM mixturewas in turn superior over bread dough since it was more consistent and required less

maintenance. Trapping studies in a fruit orchard in Australia also showed a strongattraction to pheromone/food volatiles in combination (James et al., 1994). The 4-

component aggregation pheromone combined with fermented bread dough was

compared with pheromone and dough individually, and the combined bait did alwayscatch significantly more C. hemipterus beetles. Since there was a synergistic effect

working both in the field and in the laboratory, the hypothesis of creating a good,applicable control method by combining pheromone and food attractants was

supported. The sex ratio of the attracted beetles and a comparison with the natural

release rate of hosts were not considered or discussed in the studies, and both thesefactors can possibly have an impact on future work with trapping of Carpophilus

species in the field.

European corn borer, Ostrinia nubilalis

The major drawback of pheromone trapping, that traps in most cases capture malesonly, was again shown in an extensive field study on the European corn borer, Ostrinia

nubilalis (Lepidoptera: Pyralidae) (Klun & cooperators, 1976). O. nubilalis infests

beans, pepper, potato and especially corn (Calvin et al., 1988). Traps baited with the

24

sex pheromone of O. nubilalis have yielded high catches of males, but control of

females still remained an unsolved problem (Calvin et al., 1988; Webster, 1988)

It was hypothesized that chemicals stimulating oviposition could be used toattract female cornborers, and the common plant volatile phenylacetaldehyde (PAA)

was tested as a trap bait for female O. nubilalis. The results, however, werediscouraging (Cantelo & Jacobson, 1979). Traps baited with PAA were not superior to

unbaited traps and PAA was excluded as a possible attractant to be used in IPM for O.

nubilalis. Many of the corn volatiles are in fact derivatives of the terpenoidbiosynthetic pathway (Binder et al., 1995; Binder & Robbins, 1997). From an

ovipositional bioassay Binder et al. (1995) concluded that farnesene, a chemical found

in apple coating, ants and aphids, stimulated oviposition of gravid cornborer females.The possibility of using farnesene in control was confirmed by the results, but the high

chemical impurity and the isomeric composition of farnesene could definitely be asource of error in this study (Binder et al., 1995). Binder & Robbins (1997) tested

several monoterpenes and sesquiterpenes, with higher purities, and their results pointed

at a more general response pattern of O. nubilalis, since almost all cyclic terpenesstimulated oviposition and acyclic sesquiterpenes deterred oviposition. My suggestion

is that some of the compounds tested can be used in combination with sex pheromoneto create an environmentally safe control method for O. nubilalis.

5.2.3 Stored product insect examplesIn a laboratory trapping experiment with the maize weevil, Sitophilus zeamais, 4S,5R-

sitophinone (aggregation pheromone, 4S , 5R-5-hydroxy-4-methyl-3-heptanone)together with cracked wheat caught a mean of 36 insects/trap, 4S,5R-sitophinone

alone and cracked wheat alone caught 0.9 and 14.8 insects/trap, respectively,

indicating a strong synergistic effect (Walgenbach et al., 1987). Trapping studies withthe closely related species, Sitophilus oryzae, in which 4S,5R-sitophinone also acts as

an aggregation pheromone, also showed a strong synergistic effect when foodattracants were added to the trap (Trematerra & Girgenti, 1989). Addition of rice

kernels made the trap three times more efficient compared to pheromone or food-

baited traps, and addition of cracker corn further enhanced the trapping, approximatelysix times.

25

Phillips et al. (1993) continued the work on S. oryzae with the identified

odours valeraldehyde (1-pentanal), maltol (3-methoxy-2-methyl-4-pyrone) and vanillin

(3-methoxy-4-hydroxy-benzaldehyde), which are all common in several species ofgrain, e.g. corn, rye, wheat, barley and rice (Maga, 1978). The food attractants were

presented together with 4,5-sitophinone in a bioassay, which resulted in a weak butevident synergistic effect. In the same study, T. castaneum was tested against a

stimulus containing 8:2 mixture of 4R , 8R - and 4R,8S-dimethyldecanal and a

commercial food product consisting of soybean oil, wheat germ, sugar, milk protein,soy protein, natural and artificial flavors and colours. The effect was, if not synergistic,

at least obvious and implied that trap catches of flour beetles can be greatly enhanced

by the addition of food volatiles (Figure 7).

Addition of rearing diet to larval-contaminated cheesecloth patchesresulted in a significantly higher attraction of mated female Plodia interpunctella

(Figure 8) (Phillips & Strand, 1994). The authors reported that larval contamination

elicited strong close-range oviposition responses, but that rearing medium wasrequired to induce upwind flight in the windtunnel. Food and larval-contaminated

patches combined, had a synergistic effect both in close-range and long-range

orientation. Phillips & Strand (1994) concluded that food odours obviously indicatepresence of food, and that larval odours indicate a suitable oviposition site supporting

larval growth.

Mean number responding (+/-SE)

0 5 10 15 20

Control

Food

DMD

DMD+Food

Figure 7. Responses of adult Tribolium castaneum to synthetic pheromone (DMD) and agrain-based food individually or combined in an olfactometer. Bars with different lettersare significantly different. From Phillips et al., 1993.

26

6. CONCLUSIONS AND FUTURE PERSPECTIVES

A number of studies have shown that female insects use food volatiles as olfactory

cues for locating suitable oviposition sites. Also males are attracted to food sources,since the probability of finding a suitable mating partner is higher where food is

available. Finally, larvae, due to their high food demands, are also attracted to foodaccumulations. As mentioned above, identification studies have been done to find the

chemicals responsible for the odour of the cereals. In addition, several studies have

shown what chemicals from stored products can elicit behaviours in the stored productinsects, e.g. aggregation, feeding or oviposition. A new approach in the field of food

volatiles and stored product research would be to combine the identification of cerealvolatiles and the behavioural results, mainly to see which chemical that is responsible

for a particular behaviour. The practical implication is that food volatiles can be used

in traps. The traps can be suitable for multispecies captures, since some volatiles areattractive to several species. Trapping of females can be greatly enhanced, meaning a

more efficient pest control, and food volatiles in combination with pheromones can

make simultaneous trapping of both sexes possible. Finally, by using the foodvolatiles, even larvae can be targets in pest control. The use of food volatiles in

combination with development of trap design opens up a possibility for a new field inpest management; direct control of the larvae, the most damaging life stage of stored

product insects.

.

0

10

2030

4050

60

7080

Larvae Food Larvae+Food

Figure 8. Responses of Plodia interpunctella females in a windtunnel to larval or foododours, either individually or in combination. Differences between treatments weresignificant. From Phillips & Strand, 1994.

% la

ndin

g on

tre

atm

ent

(+/-

SE)

27

7. ACKNOWLEDGEMENTS

I would like to thank my supervisors Olle Anderbrant and Christer Löfstedt for their

helpful comments on earlier drafts. I would also like to thank MISTRA for financialsupport.

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