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C 2006) Journal of Insect Behavior, Vol. 19, No. 1, January 2006 ( DOI: 10.1007/s10905-005-9001-4

Host Location and Ovipositional Behavior of Platygaster demades Walker (Hymenoptera: Platygastridae), an Egg Parasitoid of Apple and Pear Leaf Curling Midges W. R. M. Sandanayaka1,2 and J. G. Charles1 Published online: February 9, 2006

The foraging responses of 1–2-day-old na¨ıve female Platygaster demades to odors of apple and pear foliage and host insect eggs were measured. The host origin of P. demades had no effect on the parasitoids’ longevity, host preference, or foraging behavior. Four distinct behaviors related to oviposition were identified. In choice experiments, more female parasitoids responded to apple foliage with no midge eggs than to midge eggs alone. In a Y-tube olfactometer, parasitoids preferred the plant cues to clean air, and responded equally to both apple and pear odors. The results indicate that P. demades utilizes plant cues to locate the habitat of its host and then searches for host eggs to parasitize. KEY WORDS: Platygaster demades; Dasineura mali; Dasineura pyri; Host finding behavior.

INTRODUCTION The apple leaf curling midge (ALCM), Dasineura mali Kieffer (Diptera: Cecidomyiidae) and pear leaf curling midge (PLCM), Dasineura pyri ´ are two closely related but morphologically distinct (Gagne and (Bouche) Harris, 1998) pests of apples and pears respectively. Both ALCM and 1 The

Horticulture and Food Research Institute of New Zealand, Private Bag 92169, Auckland, New Zealand. 2 To whom correspondence should be addressed at HortResearch, Mount Albert Research Centre, Private Bag 92169, Auckland, New Zealand; e-mail: msandanayaka@hortresearch. co.nz. 99 C 2006 Springer Science+Business Media, Inc. 0892-7553/06/0100-0099/1

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PLCM cause similar leaf damage. Females lay eggs on new shoots and flower buds, and larvae hatch after a few days. Larval feeding distorts leaf growth and prevents young leaves from unfurling. Mature larvae exit the ‘leaf roll’ and drop to the ground, where they spin a cocoon and pupate. In New Zealand they may become a contaminant of export fruit when larvae fall into the stem end or crawl to the calyx of the fruit and pupate. Platygaster demades Walker (Hymenoptera: Platygastridae), an egg parasitoid of ALCM and PLCM, was introduced to New Zealand in 1925 to control PLCM and was first observed parasitising ALCM in 1954 (Todd, 1956). P. demades females oviposit in ALCM and PLCM eggs, but larvae complete their development in the larvae and early pupae of both species before also pupating in the soil. Since its establishment, P. demades has often achieved high levels of parasitism of ALCM and PLCM populations despite considerable variation within a season. Maximum levels of ALCM parasitism by P. demades in 1955–1956, 1956–1957, and 1957–1958 were 95.6%, 82.4%, and 60.5% respectively (Todd, 1959). However, despite high levels of parasitism, both ALCM and PLCM infestations can cause extensive foliage damage. This appears to be because of a lack of synchrony in the second generation (Berry and Walker, 1989; Shaw and Wallis, 2004). The life cycle of P. demades is more or less well synchronized with the first generation of its hosts. Todd (1956) reported that P. demades emerged 2–3 days after the start of ALCM emergence, while Dumbleton (1935) found that P. demades emerged 3 weeks after PLCM. Tomkins et al. (2000) observed that P. demades emergence was earlier than ALCM in the early 1998–1999 season. ALCM populations increased considerably during the 1980s (Tomkins et al., 1994), and some of the insecticides applied to control ALCM and other orchard pests significantly reduced parasitism (Shaw et al., 2003). While field studies have focused on the host/parasitoid responses at a population level, very little is known about the behavior of individual adult female parasitoids. In this study, we aimed to understand the searching and oviposition behaviors of P. demades; determine whether the parasitoid origin (midge species and host plant) affects adult longevity; and determine if the original insect–host plant combination influences host finding behavior. MATERIALS AND METHODS Host-Egg Production Wild ALCM and PLCM larvae were collected on apple and pear leaves respectively, from orchards in Auckland, Nelson and Hawke’s Bay. In the

Host Location and Ovipositional Behavior of P. demades

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laboratory in Auckland, ALCM and PLCM colonies were held in separate rooms with the same controlled environmental conditions of 24 ± 1◦ C, 65 ± 10% relative humidity and a 16:8 photoperiod. Midge-infested leaves were placed on top of a 3–4 cm deep layer of potting mix held in plastic boxes (20 cm × 12 cm × 8 cm) for mature larvae to crawl into the potting mix and pupate. The boxes were sprayed with water once a week to maintain humidity. When midges started to emerge, the boxes were moved to nylon gauze covered cages (50 cm × 50 cm×50 cm). Emerging adult ALCM and PLCM were collected into vials and then released into separate oviposition cages with uninfested apple shoots and pear shoots respectively, obtained from Royal Gala apple and Doyenne de Comice pear trees maintained in a shade-house. The cut stems of the shoots were submerged in water during experiments. Young leaves were examined daily under a binocular microscope for midge eggs, which were harvested and used in experiments with P. demades within 3 days of oviposition.

Parasitoid Production Emerging adult parasitoids were also collected from the field-collected leaves, individually into glass vials (7.5 cm × 2.5 cm diameter) with gauze-covered stoppers. Parasitoids emerging from the different hosts were labeled and kept separately. All parasitoids were fed with a honeyagar diet (Doug Allan, HortResearch, personal communication) except those tested for longevity in the absence of food. Na¨ıve parasitoids were tested in every experiment to avoid conditions that favored learning of host-associated cues.

P. demades Longevity The longevity of adult P. demades reared from ALCM and PLCM and fed on honey agar, or water, or in the absence of both was measured within the glass collection vials. Forty-four females and 30 males reared from each host midge were held on honey agar (1 cm diameter droplet/vial). Thirty males and females from each host midge were held with water and no food. Filtered water was provided within 3 cm × 1 cm diameter glass vial closed with a plug of absorbent cotton wool and placed in the parasitoid collection glass vial. This test was carried out in 24 ± 1◦ C, 65 ± 10% relative humidity and a 16:8 photoperiod. Longevity (in days) was assessed daily at midday, starting at 0.5 days for those parasitoids that died on the day of their emergence.

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Oviposition and Host Location Behavior All direct visual observations were carried out in the laboratory at 23 ± 2◦ C in daylight supplemented by fluorescent light. Oviposition Behavior Ten female parasitoids fed on honey agar and 10 without food or water were individually observed at the same time of day under a binocular microscope (10 × 0.63) for 1 h daily from emergence to death in the presence of ca. 200 ALCM eggs. The ALCM eggs had been collected from infested apple shoots using a fine brush and placed onto a wet, white filter paper (5.5 cm diameter) in a 5.5 cm diameter Petri dish. One parasitoid ( 0.1) (Table II).

Response (%) TRT∗

Choice 3

7.59 ± 1.27b (n = 7/40) 17.5

5.85 ± 0.55a (n = 42/50) 1.66 ± 0.11a (n = 42/50) 84 4.91 ± 0.86a (n = 19/50) 27.5 —

PLCM eggs on pear

2.19 ± 0.38a (n = 7/40) 17.5

5.49 ± 0.62a (n = 35/50) 2.50 ± 0.36b (n = 35/50) 70 3.27 ± 0.64a (n = 24/50) 37.5 —

ALCM eggs on apple

2.35 ± 0.68a (n = 7/40) 17.5

—

—

—

—

Clean pear shoot

5.49 ± 0.75a (n = 26/50) 52 2.32 ± 0.45a (n = 9/40) 22.5

—

—

—

Clean apple shoot

7.99 ± 1.8a (n = 6/50) 12 —

—

—

—

ALCM eggs on filter paper

Note. time to reach target. TFP∗ : time to first probe. n: number reached the target/total number tested. Means followed by different letters within rows are significantly different (binomial GLM test). Figures are shown as mean ± SEM.

Response (%)

Response (%) TRT∗

Choice 2

TRT∗ :

Response (%) TRT∗

TFP∗ after TRT

TRT∗

Activity

Treatments

Direct Visual Observations of Host Location Behavior: Time Taken to Reach the Target and the Percentage Response of P. demades, in No-Choice and Choice Tests in a Square Perspex Box

Choice 1

No-choice

Test type

Table II.

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Choice Experiment 2 Only 32 parasitoids (64%) responded to a choice of clean apple shoots or ALCM eggs on filter paper. There were no differences between blocks. Although more parasitoids found the clean apple shoot (52%) than ALCM eggs on filter paper (12%), they took equally long to find each (p > 0.2) (Table II). Choice Experiment 3 Ten parasitoids (25%) did not respond to any of the treatments. There was a block effect in the choices of four treatments (Tukey’s HSD). Variability between the treatments with the block effect removed showed that the time to find PLCM eggs on a pear shoot was significantly longer than in other three treatments (F = 11.64, df = 3, p < 0.001) (Table II). Sticky Trap Experiment More female and male parasitoids were attracted to the sticky trap containing a clean apple shoot (p < 0.05). Equal numbers of female and male parasitoids were attracted to Hebe and to the blank sticky trap (p < 0.05) (Fig. 3). Y-Tube Experiment Female parasitoids responded to both apple and pear volatiles, but showed no significant differences between the two (χ2 = 0.1015, df = 1, p = 0.61). The responses to the ALCM eggs on a wooden stick were lower than responses to the plants alone, and were not different from the control (F = 0.113, df = 1, p = 0.74). Parasitoids reached the apple foliage sooner than they did pear foliage (F = 6.8143, df = 1, p = 0.01) (Table III). DISCUSSION The host origin of P. demades had no effect on the parasitoids’ longevity, host preference, or ovipositional behavior. Both females and males lived longer when fed with honey agar, and females exhibited oviposition behavior and presumably laid eggs throughout their adult life. Direct visual observations disclosed preovipositional and ovipositional

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Fig. 3. Sticky trap experiment; mean number (± SEM) of female and male P. demades trapped on the sticky traps with apple shoot (host), Hebe (non-host) and control (just the trap). Means with different letters are significantly different at p < 0.05 (Tukey–Kramer HSD test).

behaviors of female P. demades. The relatively rare grooming behavior may be more frequent in the wild (when females are exposed to more odors, dust, and other detritus on host leaves), and may have an impact on landing time. Multiple probing of the same host egg was a common behavior that may lead to multiple parasitism (Todd, 1959). Although two to three probes per minute were observed, the number of probes that led to oviposition was Table III. Y-Tube Experiment—Percentage Response and the Time Taken to Reach the Target of Female P. demades to the Odors of Apple, Pear Foliage, and Apple Leaf Curling Midge (ALCM) Eggs Odor Young pear foliage Odorous air Clean air Young apple foliage Odorous air Clean air ALCM eggs Odorous air Control

Response (%)

Time (min) to target (mean ± SEM)

72 28

5.20 ± 0.35

74 26

4.08 ± 0.3

32.5 40

4.58 ± 0.67


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not determined. Further study of post-oviposition events is required to understand the potential fecundity of P. demades, which may be substantial, as the fecundity of some species in this genus is exceptionally high. Ovaries of P. hiemalis females contained an average of 3322 eggs (Clausen, 1940), and ovaries of P. matsutama contained an average of 1569 eggs (Jeon et al., 1985). Although the number of eggs in P. demades ovaries is unknown, unfed females began to oviposit on the day that they emerged, indicating at least partial pro-ovigeny. Many female parasitoids search for their hosts using chemical or visual cues from their phytophagous hosts and the plants on which their hosts feed (Vet et al., 1992; Agelopoulos and Keller, 1994, Powell et al., 1998; Pinto et al., 2004). Foraging is constrained by the ability to detect direct host cues, and the unreliability of indirect cues. Parasitoids reared on ALCM and PLCM did not show any evidence of the pre-emergence conditioning that must have occurred when they were in contact with the plant material and host larval remains. These results contrast with the behavior of some parasitoids, which is highly influenced by several trophic levels. For example, an early adult experience of cues present in the cocoon was necessary to induce the responsiveness of adult Microplitis demolitor Wilkinson (Hymenoptera: Braconidae) to the host–plant complex ´ volatiles (Herard et al., 1988), and Monge and Cortesero (1996) found that different learning mechanisms were involved in Eupelmus vuilleti, Crawford (Hymenoptera: Eupelmidae) and Dinarmus basalis, Rond. (Hymenoptera: Pteromalidae). In our results, since there was no effect of host origin on host preference in the no-choice test, the host origin of P. demades was ignored in the rest of the experiments. The odors from apple and pear shoots attracted inexperienced P. demades females and affected their host finding behavior. Plants continuously release volatile compounds into the surrounding air and adult ALCM utilize apple volatiles to locate oviposition sites (Harris et al., 1999). The sticky trap experiment showed that female P. demades were attracted to the plant cues rather than color cues, while the choice and Y-tube experiments showed that the plant odors were stronger cues than those of the host eggs, and that apple and pear volatiles were equally attractive to P. demades. In all experiments, P. demades took slightly longer to reach pear shoots than apple shoots. Once there however, parasitoids took less time for initial probing PLCM eggs than ALCM eggs. This may indicate some subtle differences in short-distance oviposition cues that might affect effective parasitism in the field. In conclusion, these laboratory experiments indicated that the behavioral responses of P. demades adults to ALCM or PLCM eggs were very

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similar, regardless of their host origin. The experiments indicated that there may be some subtle differences in oviposition behavioral responses to host cues over short distances, but they provided no evidence as to how these may influence effective parasitism in the field. Hence, the experiments indicate that the lack of parasitism in the second field generation may be due to asynchrony of host and parasitoid populations at this time (Todd, 1959; Shaw and Wallis, 2004), and environmental factors, rather than constrained the behavior of individual parasitoids. ACKNOWLEDGMENTS Authors wish to thank Mr Patrick Connolly for statistical analysis, Mr Peter Shaw and Dr Louise Malone for their valuable comments on the manuscript and Meulasi Sandanayaka for laboratory assistance. This work was funded by the New Zealand Foundation for Research, Science and Technology. REFERENCES Agelopoulos, N. G., and Keller, M. A. (1994). Plant-natural enemy association in the tritrophic system, Cotesia rubecula-Pieris rapae-brassiceae (Cruciferae): I. sources of infochemicals. J. Chem. Ecol. 20(7): 1725–1748. Berry, J. A., and Walker, J. T. S. (1989). Dasineura pyri (Bouch´e), pear leafcurling midge and Dasineura mali (Kieffer), apple leafcurling midge (Diptera: Cecidomyiidae). In Cameron, P. J., Hill, R. L., Bain, J., and Thomas, W. P. (eds.), A Review of Biological Control of Invertebrate Pests and Weeds in New Zealand 1874 to 1987. Technical communication, CAB International Institute of Biological Control 10, CAB International, Wallingford, UK, pp. 171–175, 424. Clausen, C. P. (1940). Entomophagous Insects Statement of Responsibility, McGraw-Hill, New York, p. 688. Dumbleton, L. J. (1935). Note on pear midge parasite. N.Z. J. Sci. Tech. 16: 163–164. Gagne, R. J., and Harris, M. O. (1998). The distinction between Dasineura spp. (Diptera: Cecidomyiidae) from apple and pear. Proc. Entomol. Soc. Washington 100(3): 445–448. Harris, M. O., Galanihe, L. D., and Sandnayake, M. (1999). Adult emergence and reproductive behavior of the leafcurling midge Dasineura mali (Diptera: Cecidomyiidae). Ann. Entomol. Soc. Am. 92(5): 748–757. ´ Herard, F., Keller, M. A., Lewis, W. J., and Tumlinson, J. M. (1988). Influence of host diet on host-oriented flight chamber responses of Microplitis demolitor Wilkinson. J. Chem. Ecol. 14: 1597–1606. Jeon, M., Lee, B., and Ko, J. (1985). Ecology of Platygaster matsutama and Inostemma seoulis (Hymenoptera: Platygastridae), egg-larval parasites of the pine needle gall midge, Thecodiplosis japonensis (Diptera, Cecidomyiidae). Esakia 23: 131–143. Lo Pinto, M., Wajnberg, E., Colazza, S., Curty, C., and Fauvergue, X. (2004). Olfactory response of two aphid parasitoids, Lysiphlebus testaceipes and Aphidius colemani, to aphidinfested plants from a distance. Entomol. Exp. Appl. 110: 159–164. Monge, J. P., and Cortesero, A. M. (1996). Tritrophic interactions among larval parasitoids, bruchids and Leguminosae seeds; influence of pre- and post-emergence learning on parasitoids’ response to host and host-plant cues. Entomol. Exp. Appl. 80: 293–296.


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