Journal of Chemical Ecology, Vol. 22, No. 3, 1996

ROLE OF P L A N T V O L A T I L E S IN T H E S E A R C H FOR A H O S T BY P A R A S I T O I D Diglyphus isaea (HYMENOPTERA: EULOPHIDAE)

VALI~R1E F I N I D O R I - L O G L I ,

ANNE-GENEVII~VE

JEAN-LUC

B A G N I ~ R E S , * and

CLI~MENT

C, N. R.S., l_ztboratoire de Neurob#~h~gie

Communication Chimique 31. Chemin Joseph-Aiguier, B.P. 71 13402 Marseilh, Cedex 9. France ¢Received July 25, 1994: accepted November 9. 1995)

Abstracl--Diglyphus isueu Walker is a larval ectoparasitoid used in biological pest control againsl the American serpentine leaf miner Liriomyza trifolii Burgess. We studied the parasitoid's host searching behavior, using olfactometric methods. Our data show that the parasitoids locate host larvae (a leafmining dipteran) on the basis of volatile signals released by the plant-host complex. Female D, isaea are strongly attracted to the odors arising from damaged bean plants, whereas they show practically no response to intact plants, The results of our chemical analyses showed that about 15 components were present, two of which, cis-3-hexen-I-ol and 4-hydroxy-4-methyl-2-pentunone, were present in significantly larger quantities in the leaf extracts from mined or damaged bean plants than in those from healthy plants. The damage inflicted by the host larvae on these plants triggers the release of larger amounts of these substances, which probably lead the parasites to their hosts. The compounds thus act as synomones. Key Words--Parasites, volatile signals, synomone, host detection, Hymenoptem, Eulophidae, Diglyphus isaea.

INTRODUCTION

Chemical signals play an important role in the interactions between plants, phytophagous insects, and the latter's natural enemies. When searching for their hosts, numerous entomophagous parasitoids rely on volatile chemical stimuli *To whom correspondence should be addressed. 541 0098~0331/96/t)3{X)-1)541 ~

5{)/0 ,c, 1~6 Plenum PuNishing Coq~r~alion

542

FINIDORI-LOGLI~ BAGNERES, AND CL~.MENT

arising from their host's natural habitat (Vinson, 1975, 1981; Weseloh, 1981; Van Alphen and Vet, 1986). Once the host's habitat has been located, the parasitoids have to find the hosts themselves. For this to be possible, the parasitoids have to be able to detect their hosts from a distance and to orient their movements in response to stimuli produced or induced by the host larvae or by their activity on the plant (Weseloh, 1981). The natural enemies of several phytophagous insects are strongly attracted by volatile compounds released by the host plants when they are damaged (Drost et al., t988; Dicke et al., 1990; Turlings et al., 1990, 1991). The components of the plant-host complex may originate from either the plant, the host insect, or from the latter's feces or saliva (Vinson, 1976; Kennedy, 1984; Mudd et al., 1984; Takabayashi and Takahashi, 1989). In order to understand insect behavior involving chemical communications, it is necessary to determine the nature of the signals arising from the insect's natural environment, that play an important role in their intra- and interspecies relationships. Chemical signals are known to play a role in Dacnusa sibirica (Dicke and Minkenberg, 1991) and in Opius dissitus (Petitt et al.+ 1992). In the present study, we analyzed the chemical stimuli used by the entomophagous parasitoid, Digfyphus isaea Walker (family Eulophidae). D. isaea is used in biological control of agromyzid leaf miners in greenhouses (Lyon, 1986; Nedstare et al., 1987; Van Lenteren and Woets, 1988). The ectoparasitoid uses several leaf miners as its hosts, especially Liriornyza tlifolii, a pest that inflicts considerable damage on greenhouse vegetable and flower crops. Female parasitoids oviposit in the last larval stages of L. trifi)lii (Minkenberg and Van Lenteren, 1987). More specifically, the aim of the study was to investigate whether D. isaea uses volatile chemicals to locate its hosts, and if so, to determine which of the chemical signals emitted by the plants, the agromyzid larva, and larval secretions (saliva, feces) influence the behavior of the parasitoids. METHODS AND MATERIALS

Insects. The parasitoids used in these experiments were from a permanent rearing station set up by the INRA at Antibes, using the "alternate open field rearing and caged ovipositing" method (Lyon, 1976). Plants. Bean plants (Phaseolus vulgaris) were grown in a nursery garden and were harvested at the cotyledon stage. Behavioral Analyses. Behavioral observations were conducted with 2-dayold female D. isaea in their rearing cages (50 x 50 x 60 cm)+ For each experiment, two bean plants, each in its own pot, were provided; one plant contained only two healthy leaves and the other, two leaves mined by stage 2

543

PLANT V O L A T I L E S IN H O S T S E A R C H I N G

(L2) larvae. Five female parasitoids were used in each test and 15 replications were conducted. Parasitoid visits to each of the two kinds of leaves were counted during a period of 20 min. Tests were carried out at 25°C and 50% relative humidity. The Olfactometer. The linear olfactometer was a combination between the olfactometer designed by Leconte and Thibout (1981) and that described by Zagatti (1985). It was 30 cm in length, 2 cm in diameter, and was divided into compartments numbered 0-5 through which pure (control) air and air containing the odor to be tested were pushed alternately for 4 min. The device was placed between two neon-shelving (upper tube: 60 W, lower tube: 2 x 60 W). The parasitoids were classified as weakly attracted, moderately attracted, or strongly attracted depending on where they were located in the olfactometer. Five 2-dayold mated female parasitoids were placed in compartment 0 at the beginning of each test. All the tests were carried out at 25°C with 50% relative humidity. Circulating air (10 cm/sec) was filtered through an active charcoal disk and then through a tube containing fibreglass wool. The odor streams were delivered as follows: for each test, 2 p.l of the extract was placed on a 2-cm 2 filter paper dish and when the solvent had completely evaporated, the filter paper was placed in the olfactometer and swept by a stream of humidified air. The control airstreams were passed over a similar Whatman paper filter treated only with solvent. Linear Olfactometer Data Processing. The number of females in each compartment was recorded every 30 sec, and 16 replications were conducted. These data were used to calculate the relative attraction indices as described by Zagatti (1985). 5 i=0

5

init- ~ inio 5

i=0

× t00

(5 - i)nio i=0

where i is the number of compartments in the olfactometer (from 0 to 5), hi, is the number of insects in the compartment i at time t, and nio is the number of insects in the compartment i at t = 0. The results were analyzed statistically with the Mann-Whitney test. Collecting and Analyzing Volatile Substances. A cold-trapping method was used to collect the odors arising from healthy beans, beans infested by stage two larvae, and beans that had been mechanically damaged by cutting the leaves with a razor blade. A steady flow (500 ml/min) of purified air was circulated through a closed 2-liters experimental chamber containing 25 leaves during 20 rain. The molecules emitted were trapped on the walls of a capillary tube (10 c m x 0.2 mm) that was embedded in dry ice at - 8 0 ° C . The trapped odors

544

FINIDORt-LOGL[, BAGNERES, AND CLI~MENT

were eluted with 300 ml of dichloromethane. The extract was then concentrated to obtain 50/~1 and used in behavioral tests in the olfactometer and for chemical analyses. Samples were analyzed with a gas chromatograph, a Delsi 330 apparatus, equipped with a flame ionization detector and a Supelcowax 10 polar capillary column (30 m x 0.32 mm, 0.25-p.m film thickness). The split/splitless injector was kept in the splitless mode for 15 sec after the injection. The carrier gas was helium at a pressure of 1 bar. The temperature program was isothermal at 30°C for t0 min and then increased in 5°C/rain steps to 220°C. The data were captured and processed by a Chromatopac C-R4AX Shimadzu integrator. The results were quantified by introducing a solution containing two internal standards: 375 ng of n-dodecane and 400 ng of octanol. Leaf extracts were obtained by soaking about 50 previously weighed leaves in 200 ml of dichloromethane for 24 hr. The extract was then filtered and concentrated in a nitrogen atmosphere. GC analyses confirmed that these samples contained the same substances as those obtained with the air-entrainment trapping method, but in higher concentrations. For identification of components, it was decided, therefore, to soak the leaves and to use gas chromatography coupled to mass spectrometry (GC-MS). The equipment consisted of a HewlettPackard 5890 Series lI GC equipped with a single Supelcowax 10 polar column (30 m x 0.32 mm, with 0.25-tzm film thickness), connected to a HewlettPackard 5989A MS Engine (EI, 70 eV). The apparatus was controlled by an MS Chemsystem HPUX multitask data processor using the Unix system. Helium at a pressure of 2 bars was used as the carrier gas. The GC oven temperature was maintained at 30°C for 5 min before being increased in 5°C/min steps to 210°C, and maintained for 10 rain. The temperature was 240°C at the source, 250°C at the interface, and 100°C at the quadrupole. The masses were swept at 35-300 mass units at a rate of one scan per second. A standard range of n-alkanes from n-Ci2 to tl-C2o was coinjected, and comparisons with the Wiley 138 library of reference spectra aided in identification of the main compounds present. RESULTS

Behavioral Data. In the experimental cage, D. isaea made more frequent visits (Table t) to mined leaves (43 visits) than to intact leaves (15 visits, Z = 1.3 x 10 -4, P < 0.001). The parasitoid therefore appeared capable of distinguishing between the two categories of leaves. The mined leaves had yellowish tunnels that were clearly visible at the surface, each containing one larva. It is possible that the parasitoids located the mines through visual cues. In any case, the results of this test show that D. isaea are able to distinguish between healthy leaves and those containing a host larva.

545

PLANT VOLATILES IN HOST SEARCHING

TABLE 1, BEHAVIOR OF

Diglyphus isaea

RELEASED IN CAGE CONTAINING UNDAMAGED

AND DAMAGED BEANSa

Number of visits/20 rain trial Mined leaves Healthy leaves

4 1 3 3 3 3 1 2 2 2 3 4 4 4 4 =43 1 2 0 1 0 2 l 1 1 0 1 3 1 0 1 =15

"The results from 15 replications were compared using the Mann-Whitney test. (Z = -3.82; P < 0.001).

The results of the linear olfactometer tests on female parasitoids exposed to trapped odors o f healthy plant leaves are given in Figure 1. During the first 3 rain, the females reacted very little to the trapped volatile substances originating from a healthy bean plant. The relative attraction indices (RAI) were low (5.6, 6.9, 7.4, 9.9, 9.5). F e m a l e parasitoids nevertheless s h o w e d a characteristic b e h a v i o r in which they positioned t h e m s e l v e s facing the o n c o m i n g flow o f odor-

]

30

20'

ae~my leaf

J

10

|

I'30

2'30

3'30

Time (mla)

FIG. 1. Bioassay of healthy bean plant leaves and the control (pure air) stimulus for attraction of parasitoid females. There were 16 replications involving five females in each replication. Statistical analysis: Mann-Whitney test, *P < 0.05.

546

FINIDORI-LOGLI, BAGNI~RES, AND CLI~MENT 80

60

1-7-"

Mined

leaves

l

{D

i

40

20

i

I'30

2'30

3'30

"I'Ve (rain.)

FIG. 2. Movement of five parasitoid females (16 replications) towards extracts of mined bean plant leaves compared with the control stimulus (pure air). Statistical analysis: Mann-Whitney test, *P < 0,05; ***P < 0.001.

impregnated air with their antennae extended, and then made a series of antennal movements before advancing against the airstream. A significant difference was found to exist 4 min after the beginning of the experiments (t + 4 min) between the RAI indices obtained with impregnated and pure air at the 5% significance level (Z = 0.25) (RAI = 13.4 vs. RAI = 3.1 under control conditions). In response to the airstream carrying trapped volatile substances extracted from bean plants infested with stage 2 larvae, female parasitoids oriented much more quickly towards the oncoming airstream (Figure 2). The RAI increased much more quickly, from 7.6 after 1 min to 61.7 after 3 min, and reached 75.5 by the last minute of observation. Females actively worked their way up against the odor-laden stream of air, frequently contracting their abdomen. This behavior is typical of females moving towards a mined area. As early as 1 min after the beginning of the test, the differences between the control test and the tests with extracts were significant at the 5 % threshold level (Z = 0.014), and shortly afterwards, they became significant at the 1% level (Z = 1.8 x 10 -6 at the fourth minute of the test). Figure 3 gives the comparative data on movements of the female parasitoid towards healthy versus mined bean plants. These females also responded to the airstream carrying the trapped odor of mechanically damaged leaves (Figure 4).

547

P L A N T V O L A T I L E S IN H O S T S E A R C H I N G

80

60

i

Healthy bean leaf I Mined bean leaf

40

I

j 20

S

'i

I

2

3

Time

(min.)

FiG. 3. Movement of parasitoid females (5 females, 16 replications) towards extracts of healthy bean leaves compared with mined bean leaves. Statistical analysis: Mann-Whitney test, ***P < 0.001.

40

30

,~

Control

2o •

Mechanically

damaged

bean

lO

!

1

2

3

' Time

(rain.)

FIG. 4. Attraction of parasitoid females (5 females, 16 replications) by extracts of mechanically slashed bean plant leaves compared with the control stimulus (pure air). Statistical analysis: Mann-Whitney test, *P < 0.05; ***P < 0.001.

548

FINIDORI-LOGLI, BAGNERES, AND CLEMENT 4o

30 0

[ ----~"-'o

20

C ontrol Larvae

10

i

1

2

3

4

Time (min.)

F1c. 5. Movement of parasitoid females (5 females, 16 replications) towards leaf miner larvae previously fed with bean plant leaves. Statistical analysis: Mann-Whitney test, not significant•

807060 "O

..= ,= 0

50

-----v--- Control L s Fecal extract

40 ¢= -w

30 20 10 0

i

1

2

3

4

Time (min.)

FIG• 6. Movement of parasitoid females (5 females, 16 replications) towards fecal extracts of leaf miner larvae compared with the control stimulus (pure air). Statistical analysis: Mann-Whitney test, *P < 0.05.

P L A N T V O L A T I L E S IN H O S T S E A R C H I N G

549

The attraction indices increased by 8.8 at t + I min, and reached 34.7 at t + 4 min. In comparison with pure air conditions, the RAI differed significantly at the 5% threshold (Z = 0.025) at t + 3 rain, and then at the 1% threshold (Z = 7.76 × 10 -3 a t t + 3.5 minutes, Z = 2.24 × 10 3 a t t + 4rain). Here the parasitoids' behavior was fairly similar to that observed with mined leaves. The movements of D. isaea females towards stage 2 Liriomyza trifiglii larvae did not differ from those elicited by control air (Figure 5). The relative attraction indices followed the same pattern in both cases, ranging between 2.2 and 10 with odor-impregnated air and between 1 and 7.2 with pure air (Z = 0.82 a r t + 1 rain a n d Z = 0.5 at t + 5 rain). These data show that the host larvae alone exerted no attraction for the parasitoids. The odor of the feces of leafminer larvae was attractive to the parasitoid females (Figure 6). The RAI reached 16.8 within t + 4 min, as compared with 6.7 in pure air (Z = 0.06). The differences observed between the indices recorded under control conditions and with the fecal odors were significant within 2 rain at the 5% threshold level (Z = 0.19 at t + 2 min, Z = 0.02 at t + 2.5 rain, Z = 0 . 1 7 a t t + 3rain). Chemical Analysis. Representative chromatograms obtained with trapped volatile extracts from healthy, mined, and mechanically damaged bean plant leaves are shown in Figure 7. Fifteen peaks occurred consistently from one test to another. None of these compounds was detected in the blank trapping extracts used as controls. Figure 8 shows details of a typical reconstructed ion chromatogram obtained with mined leaves that were soaked in dichloromethane. Some of the substances were present only in trace quantities in the healthy leaves (peaks 7, 9, and 10). The composition of all substances present in larger amounts could be clearly determined. Three of the quantified components (peaks 6, 7, and 12) varied with the state of the leaves (whether healthy, mined, or damaged). Table 2 presents the identity of compounds, the diagnostic ions used as the basis for identification, and the quantities of the three main components. Most of the substances were oxygenated molecules and included primary and tertiary alcohols (1-pentanol, cis-3-hexen-l-ol, 3-octanol, 5-octa-5-dien-3-ol), aldehydes (2-butenal, propanal-2-methyl oxime, and 2,4-heptadienal), or ketones (4-hydroxy-4-methyl-2-pentanone, previously identified by Hefetz and Lloyd (1983) in a formicine (ant) anal gland. Some short-chain hydrocarbons were identified also, including n-dodecene (peak 3), n-tridecane (peak 5), 4,8-dimethyl tetradecane (peak 14), and the monoterpene camphene (peak 2). Table 2 and Figures 7 and 8 show that the volatile profiles of healthy, mined, and damaged plants did not differ qualitatively with the exception of 1-octen-3-ol. This compound was present only in mined leaves at the same retention time as that of 2,4-heptadienal.

550

FINIDORI-LOGLt, BAGNERES, AND CLI~MENT

A- Undamaged bean leaves 13

I I.$2

6

LT.,.13 I

B- Miner infested bean leaves

~

5 i I.S2

6

C- Artificially damaged bean leaves

13 5 ~I.S 2

FIG. 7. Gas chromatogram (Supelcowax 10) of volatiles from undamaged bean leaves (A), damaged bean leaves containing feeding Liriomyza trifolli larvae (B), and artificially damaged bean leaves (C). Internal standards (ISI: n-docosane and IS2: octanol) are shown.

PLANT

VOLATILES

IN HOST

SEARCHING

55 1

5

~u'ndmrllll

12+13

§

100~0

9 M

14

15 8OOOO

!

3 4

40000

2OOOO

'

'

ib

'

'

'

tk

i

i

I

i

i

i

i

i

llmt |ndn.)

FtG. 8. Total ion chmmatogram of damaged beans with numbered peaks: (1) 2-butenal, (2) camphene, (3) n-dodecene, (4) l-pentanol. (5) n-tridecane, (6) 4-hydroxy-4-methyl2-pentanone, (7) cis-3-hexen-l-ol, (8) unknown, (9) propanal, 2-methyl oxime, (10) 3-octanol, (11) unknown, (12) l-octen-3-ol, (t3) 2,4-heptadienal, (14) 4.8-dimethyl tetradecane, (15) 5-octa- 1.5-dien-3-ol, (16) unknown.

Determinations were conducted on three of the main substances in a dozen samples of soaked healthy, mined, and damaged leaves. The amounts of all three substances increased whenever the leaves had been either attacked by biological agents or by the activity of the larvae or artificially damaged. Alcohols such as cis-3-hexen-l-ol (peak 7), which could not be quantified in the healthy leaves because it was present at such low levels, increased to as much as 600 ng/g of leaf matter in mined leaves and up to 700 ng/g in artificially damaged leaves. Likewise, 1-octen-3-ol (peak 12), which was not detected in the healthy leaves, reached the level of 1200 ng/g in mined leaves. 1-Octen-3-ol was not detected in extracts from damaged leaves. Lastly, 4-hydroxy-4-methyl-2-pentanone (peak 6) was extracted from the healthy leaves at about 350 ng/g, but it increased to as much as 6800 ng/g in mined leaves and 5700 ng/g in damaged leaves. DISCUSSION D. isaea are attracted by odor signals emitted by plants and by the host insects living on these plants. The results of our experiments using cold-trapped

Unknown I-Octen-3ol

2.4-Hepladienal 4,8-Dimelhyl lemadecane 5-Ocla-l.5-dien-3-ol

II 12

13 14

15

10

Unknown Pmpanal 2-methyl ilxynle 3-Octanol

4-Hydmxy-4-melhyl 2-pentanone Hx-3-Hexen-I ol

2-Butenal Camphene D{~decene I -Pentanol n-Tridecane

Compound

g 9

7

6

Peak

CsH,,O

CTHmO C..H u

43. 57. 72. g l . 85 99. I I0 (M-18)' 53. 81. 95. 110 (M ~ ) 7(I/I, 112/3, 1411/1. 182/3. 211 ( M- 151 ' 41. 57. 70. 79. 93. 106 (M-18)'

41. 59. 83. l01. 112 I M - I g ) '

C,H,.O

C.H,aO

41/2. 55, 7(). H6. g7

41. 55. 67. g2 IM-18)'

42. 55. 70 43. 57. 71. 85. 184 (M'} 43. 59. I01 tM-t5) '

39. 41. 56, 70 I M ' ) 79. 93. 121. 1 3 6 ( M '

Diagnoslic El-MS ions

Call.NO

C,,H,_.O

C,,HpO:

C4H,,O C,,,H., CL.H:a C.Ht:O C~ ~H_..

Funnula

124

I10 226

128

130

g7

I()0

110

184

70 136 16H gg

MW

TABI.E 2. II)ENI'IIq('AII()N OF VrH~A'I'ILI(S BASED ON G C - M S

+

+ trace

+

trace

trace

+ 350 ng!g trace

Undamaged

El [~')NIZATI()N

+

trace + 1200 ng/g trace +

+

+

+ 6800 ng/g + 600 ng/g

Damaged

+

+ trace

trace

+

+

+ 5700 ng/g + 700 ng/g

Aniticially damaged

z

r-

z ~J

z

o

>

oo _C.

o '7

PLANT VOLATILES IN HOST SEARCHING

553

bean plant extracts to attract the parasitoids show that they are able to recognize bean plants on the basis of volatile chemical signals that the plants give off. With the cold-trapping technique used in these experiments, natural conditions are better reconstituted by eliminating heavy, less volatile compounds such as triterpenes and phenols that are removed during the soaking or crushing processes (Wolfender et al., 1993). Bean plants infested with leafminer larvae release much higher quantities of volatile molecules from the leaves than do healthy, non-mined plants. Larvae feeding on these plants may be responsible for the enhanced production of these substances. In our experiments, D. isaea females placed in an olfactometer responded to these chemical signals by moving towards the odor source, and it seem likely that they behave in the same way in natural surroundings. When plants are attacked, they seem to release substances that guide the parasitoids to their hosts, The molecules we identified here have been previously mentioned in the literature, cis-3-Hexen-l-ol, for instance, occurs in more than 25 plant families (Visser et al., 1979) and has been reported from healthy leaves of potato plants (Visser et al., 1979), cowpeas (Whitman and Eller, 1990), several cmcifers (Wallbank and Wheatley, 1976), and in strawberry plant leaves (Buttery et al., 1984; Hamilton-Kemp et al., 1988). Saijo and Takeo (1975) established that cis-3-hexen- I-ol is produced by green plants when the leaf tissues are damaged. l-Octen-3-ol has previously been reported from strawberry foliage (Hamilton-Kemp et al., 1988), in the leaves of a shrub endemic to Florida (Jordan et al., 1992), and in corn tassels (Buttery and Ling, 1980). In the present study, this compound was emitted only by mined leaves. The presence of 2,4-heptadienal was observed in both healthy and damaged leaves. It has also been detected in corn (Buttery et al., 1978; Buttery and Ling, 1980). We also identified some quite rare substances, such as oximes, in the trapped volatiles obtained from mined and artificially damaged bean plant leaves. Oximes have been previously identified in apple leaves attacked by Tetranichus urticae (Takabayashi et al., 1991) and in cucumber leaves (Takabayashi et al., 1994). D. isaea do not use only one pest species as a host, and the question arises as to whether there may exist a common set of molecules used by the parasitoid to detect the mined leaves of any of their potential hosts. In our studies the parasitoid seems to recognize volatiles common to several plants. Guerin et al. (1983) suggested that the recognition of a plant's odor may be based on a synergy between molecules that are common to a whole set of plants, plus some more specific molecules, If true, this hypothesis would explain why D. isaea are so effective in attacking several leafminer pests. Although D. isaea responded actively to plant extracts used in our study, it should be remembered that these results were obtained under experimental

554

FINIDORI-LOGLI, BAGN~RES, AND CLI~.MENT

conditions. Each odor was presented separately in the olfactometer, whereas in natural surroundings, olfactory stimuli are more complex. In our study the volatile chemical signals emitted by the D. isaea-L trifotii-Phaseotus complex mediated communications between the plants, the plant-eating insects, and the insect-eaters. Green plants such as tomatoes, potatoes, and bean plants release volatile compounds that give cut leaves a characteristic " g r e e n " odor. These volatiles are mainly alcohols with six carbon atoms, such as cis-3-hexen-l-ol, cis-2hexen-l-ol, 1-hexanol, and aldehydes such as trans-2-hexenal and their derivatives (Visser et at., 1979; Hamilton-Kemp et al., 1988, 1989; Hemandez et al., 1989). In the present study, the proportions of two of the molecules released by bean plant leaves, cis-3-hexen-l-ol and 4-hydroxy-4-methyl-2-pentanone, increased considerably in plant leaves attacked by the agromyzid larvae and in those that had been mechanically lacerated. The release of these compounds, or possibly just one of them, may inform parasitoids that they are in the vicinity of damaged plants, and the plants subsequently benefit from the messages it has sent out. In the chemical interactions described here, communications seem to occur between the first and third trophic levels. The emitter and the receiver of the chemical messages both benefited from the exchange (Whittaker and Feeny, 1971). Here the tern1 "allomones" does not apply in the strictest sense since the interactions were reciprocal. Nordlund and Lewis (1976) proposed the term synomone, which seems to describe more appropriately the type of signals occurring in our model. Several authors (Whitman, 1988; Dicke and Sabelis, 1988, 1989; Dicke et al., 1990) have explained communication of this kind in terms of the plants calling for help when they are attacked by larvae. Elzen et al. (1983), Nadel and Van Alphen (1987), Dicke and Sabelis (1988), and Turlings et al. (1991) have established that herbivore-damaged plants are more attractive to parasitoids than healthy ones. The idea that plants may have developed a defense mechanism that elicits participation of insect parasitoids seems quite plausible. This hypothesis presupposes that there exists a long-standing association between the plants, the phytophagous insects, and the parasitoids. Plants may be induced to emit attractants utilized by insect parasitoids by either the host saliva (Turlings et al., 1990), its excretory droppings (Ramachandran et al., 1991), and/or its cocoon (Bekkaoui and Thibout, 1993). In our experiments with the olfactometer, D. isaea showed some sensitivity to feces of leafminer larvae feeding on bean plant leaves. We were not able to extract the substance responsible for this attraction. In locating a host, D. isaea wasps have to search for various potential hosts living on different plants growing in diverse habitats. In such a complex chemical

PLANT VOLATILES IN HOST SEARCHING

555

environment, simply searching for a single kairomonal chemical may not suffice for the parasitoid to be able to make the most of all the available resources. Long-range signals probably will convey only information about the host's habitat, while more short-range semiochemical substances emanating from infested plants or from the host's feces or other secretions will provide more direct and more reliable information about the availability of hosts and the sites where they are to be found. A parasitoid searching for a host probably uses a continuous, dynamic flow of information to locate hosts. Some parasitoids appear to be able to make use of what they have previously experienced and learned in order to take advantage of this complex system of interactions (Vinson et al., 1977; Vet and Groenwold, 1990). Aekmm'iedgments--We are grateful to the Conseil Regional Provence-Alpes-C6te-d'Azur and Duclos Agrobiolech Society tot lheir financial support. We would like to thank G. Dusticier and E. Tabone for their cooperation, Jessica Blanc |kJr the English Inmslation, and Arlette Poveda l~'~r helpful secretarial assistance.

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