RJ WEST,^ M KENIS,' GW Bun, and SM BENNETT Abstract

PARASITOID COMPLEX OF ZEZZUPHERA CANADENSZS (LEPIDOPTERA: TORTRICIDAE) AND EVALUATION OF TYCHERUS OSCULATOR (HYMENOPTERA: ICHNEUMONIDAE) AS A BIOLOGICAL CONTROL AGENT

RJ WEST,^ M KENIS,' GW Bun, and SM BENNETT Natural Resources Canada, P.O. Box 960, Comer Brook, Newfoundland, Canada A2H 653

Abstract The Canadian Entomologist 131: 465 - 474 (1999) A survey of larval and pupal populations of the spruce bud moth, Zeiraphera canadensis Mutuura and Freeman (Lepidoptera: Tortricidae), conducted from 1994 to 1996 in eastern Newfoundland, disclosed that the incidence of endemic parasitism by Hymenoptera was up to 50% for Tycherus osculator (Thiinberg) (Ichneu- monidae), up to 15% for Earinus zeirapherae (Walley), under 3% for Ascogaster (Wesmael 1835) sp. and Clinocentrus (Haliday 1833) sp. (Braconidae), and under 1% for Lamachus (Foerster 1868) sp. and Triclistus (Foerster 1868) sp. (Ichneu- monidae). Tycherus osculator, E. zeirapherae, Ascogaster sp., and Clinocentrus sp. represent new distributional range extensions to Newfoundland, and to the nearctic region in the case of Z osculator. The biology of European populations of Z osculator was studied on a natural host, Zeiraphera diniana (GuenCe). Only females overwintered and ovarian maturation did not occur until after several months of exposure to near-freezing temperatures. Tycherus osculator successfully parasitized prepupae and pupae of Z. diniana of all ages but, in the laboratory, appeared to prefer pupae. Host feeding by Z osculator was common but not necessary for ovarian maturation. Tycherus osculator imported from Europe attacked and successfully developed within the spruce bud moth host in laboratory screenings. Morphological comparisons indicated that Z osculator reared from Z. canadensis were smaller than those reared from Z. diniana. Tycherus osculator obtained from either Newfound- land or Europe may have potential as a biological control of Z. canadensis in mainland Canada, where it is presently absent.

West RJ, Kenis M, Butt GW, Bennett SM. 1999. Complexe parasitoide de Zeiraphera canadensis (Lepidoptera: Tortricidae) et Bvaluation de Tycherus osculator (Hymenoptera: Ichneumonidae) titre de biopesticide. The Canadian Entomologist 131: 465-474.

Une Ctude des populations larvaires et nymphales de la tordeuse de I'Cpinette, Zei- raphera canadensis Mutuura et Freeman (Lepidoptera: Tortricidae), effectuke de 1994 a 1996 B I'est de Terre-Neuve a montrC que le taux de parasitisme par les Hy- mCnoptbres atteignait 50% pour Tycherus osculator (Thiinberg) (Ichneumonidae), 15% pour Earinus zeirapherae (Walley), moins de 3% pour Ascogaster (Wesmael 1835) sp. et Clinocentrus (Haliday 1833) sp. (Braconidae), et moins de 1% pour La- machus (Foerster 1868) sp. et Triclistus (Foerster 1868) sp. (Ichneumonidae). Ty- cherus osculator, E. zeirapherae, Ascogaster sp. et Clinocentrus sp. sont pour la premibre fois mentionnCs Terre-Neuve alors que T. osculator n'avait encore jamais CtC trouvC dans la rCgion nCarctique. La biologie de populations europkennes de Z osculator a Ctt CtudiCe sur un h8te naturel, Zeiraphera diniana (GuenCe). Seules les femelles ont hivernC et la maturation ovarienne n'a CtC observCe qu'aprks une pCriode hivernale de plusieurs mois B basse tempkrature. Tycherus osculator a para- sit6 avec succbs les prC-nymphes et nymphes de tous Bges mais, au laboratoire, il a prCfCrC les nymphes. Les femelles ont souvent CtC observkes s'alimentant sur I'hhte, mais cette activitC n'Ctait pas nCcCssaire a la maturation ovarienne. Une population

F'resent address: Box 515, Portugal Cove, Newfoundland, Canada, AOA 3K0 F'resent address: CABI-Bioscience, 1, chemin des Grillons, DelCmont CH-2800, Switzerland 465 466 TH@CANADIAN ENTOMOLOGIST JulyIAugust 1999

europtenne de T. osculator a parasit6 avec succbs la tordeuse de 1'Cpinette au laboratoire. Des comparaisons morphologiques ont montrt que les T. osculator euro- pCens obtenus de Z. canadensis Ctaient plus petits que ceux obtenus de Z. diniana. Tycherus osculator, de Terre-Neuve ou d'Europe, est consider6 cornme un agent de lutte biologique potentiel contre Z. canadensis sur le continent nord-amCricain, ou il est apparemment absent.

Introduction

The spruce bud moth, Zeiraphera canadensis Mutuura and Freeman (Lepidoptera: Tortricidae), is native to North America and attacks white spruce, Picea glauca (Moench) Voss (Pinaceae), throughout Canada and the northeastern United States (Turgeon 1992). Damage by spruce bud moth may cause severe crown deformation, multiple leaders, and reduction in height and volume growth (Carroll et al. 1993). The spruce bud moth is expected to become a pest in western and central New- foundland when spruce plantations are established in areas traditionally covered by balsam fir, Abies balsamea (L.) Miller (Pinaceae), forest, but little is known of parasitoid species attacking this insect in Newfoundland. An understanding of the endemic parasitoid complex is necessary to identify vacant niches that could be exploited by exotic parasitoids imported as biocontrol agents and indicate occupied niches where introduced parasitoid species would be obliged to compete. White spruce trees growing on the Avalon Peninsula near St. John's are severely attacked by spruce bud moth and provide excellent sources of material to determine endemic parasitoid levels. The native complex of natural enemies of the spruce bud moth in Canada was summarized by Turgeon (1992) from records made in Quebec and New Brunswick. Trichogramma rninutum Riley and Trichogramma sp. (Hymenoptera: Trichogrammatidae) have eclosed from spruce bud moth eggs at incidences over 50% (see also Ostaff and Quiring 1994; Ostaff 1995). Twenty-four species of Hymenoptera (nine Braconidae, three Pteromalidae, three Eulophidae, and nine Ichneumonidae) have been recorded as attacking larvae or pupae. Rates of apparent larval parasitism recorded in Quebec were under 13% (Pilon 1965). However, Earinus zeirapherae (Walley) (Ichneumonidae) accounted for over 50% parasitism of larvae in New Brunswick (Turgeon 1992). Mills (1993) and Schonberg (1993) reported the results of surveys of parasitism of European populations of Zeiraphera ratzeburgiana (Ratzeburg), Zeiraphera diniana (GuenCe), and Zeiraphera rufimitrana (Herrich-Schaffer), three species with a life his- tory similar to that of the spruce bud moth. Mills identified several Ichneumonidae, Phytodietus (Gravenhorst 1829) spp., Chorianaeus cristator (Gravenhorst), and Ty- cherus osculator (Thiinberg) as potential candidates for the control of spruce bud moth. Schonberg (1993) recommended that the Ichneumonidae 7: osculator and Tranosema carbonellurn (Thomson) be studied further as potential candidates for introduction on the basis of their specificity to Zeiraphera species from Europe and compatibility with the life cycle of the spruce bud moth. We present a summary of larval and pupal parasitism of the spruce bud moth populations from eastern Newfoundland and results of screening 7: osculator reared from Z. diniana against larvae and prepupae of spruce bud moth in the laboratory. Additional data from laboratory studies are presented on the behaviour and development of T osculator exposed to Z. diniana. We studied 7: osculator because it is the most important natural enemy of Z. ratzeburgiana (Schonberg 1993), the European species of Zeiraphera most similar to 2. canadensis. We worked with Z. diniana because Z. ratzeburgiana populations were too low to allow collection of sufficient numbers of pupae during the period of study. Volume 131 THE CANADIAN ENTOMOLOGIST 467

Methods

Local Parasitoid Complex. Study sites and plot layout. The study was conducted during the summers of 1994-1996 near St. John's, Newfoundland, Canada. Field sites containing open-grown, white spruce trees infested by Z. canadensis were surveyed to determine its endemic parasitoid complex and to assess levels of parasitism. The first site (Arterial, 47"3ON, 52"47W), located 1 km west of St. John's, was 3 ha in size, dominated by 2-3 m tall white spruce and containing a mixture of balsam fir; white birch, Betula papyrifera Marshall (Betulaceae); and pin cherry, Prunus pensylvanica L. (Rosaceae). The second site (Torbay, 47"39'N, 52"43W), located next to the ocean and 6 km north of St. John's, was a 1- to 2-ha stand of 2- to 5-m high white spruce. Ten trees, about 2 m tall and showing evidence of moderate to severe defoliation by the spruce bud moth, were sampled at each location each year. Larvae. Branches were sampled once a week starting at the time of spruce bud moth egg hatch, usually at the end of May. On each sampling date, two midcrown branch tips, about 25 cm in length, were arbitrarily chosen and pruned from each tree, placed in labeled paper bags, transported in coolers, stored at 5°C upon arrival at the laboratory, and processed within 24 h. Newly burst buds and elongating shoots were dissected to obtain spruce bud moth larvae. Seventy-five larvae from each collection were arbitrarily chosen and reared in groups of five in 25-mL cups containing 10- 15 mL of spruce budworm artificial diet (Bio-Serv Inc., Frenchtown, New Jersey) until pupation, parasitoid emergence, or death. The diet was modified for 1995 and 1996, when one or two fresh buds from an uninfested site were added to each cup containing the artificial diet in an attempt to increase the survival of first- and second-instar larvae. Diet was changed weekly. Throughout the rearing process, larvae were monitored for survival, mortality, and parasitism. Collections continued until the end of the larval period and lasted approximately 6 weeks. Pupae. Fourth-instar larvae dropping from the foliage to pupate in the litter were collected in 1 x 1 m wooden trays filled to a depth of approximately 3 cm with vermiculite and placed beneath each sample tree for 1 week in early July. The upper edge of the trays was coated with ~an~lefoot~to prevent removal of pupae or larvae by insect predators. After 1 week the vermiculite was placed in large plastic bags and transported to the laboratory where pupae and small clumps of vermiculite containing pupal cells created by the trapped larvae were extracted by sifting. One hundred spruce bud moth per site were subsequently reared individually in 60-mL vials. Foam corks soaked in distilled water were used to prevent desiccation within the vial. Pupal cells that did not yield adult bud moths or endemic parasitoids after 1 month were kept in a rearing chamber at 22°C and 60% RH for an additional 3 months, then held for 12 weeks at 4°C to simulate overwintering conditions. These were then returned to 22°C and 60% RH for 1 month to obtain parasitoids that normally overwinter inside the host. Overwintering pupae that did not yield parasitoids were dissected to determine the incidence of undeveloped and unemerged parasitoids. Larval and pupal parasitoids reared to the adult stage were pinned and sent for identification to specialists at the Eastern Cereal and Oilseed Research Centre in Ottawa and the Zoologische Staatssammlung in Munich.

Biology of Tycherus osculator on Zeiraphera diniuna. Laboratory studies were conducted to determine various aspects of the biology of T osculator on Z. diniana, including ovarian maturation, the preoviposition period (i.e., the elapsed time from transfer to room temperature following overwintering in cold storage and the occurrence of the first oviposition), the stage and age of host preferred for oviposition, and whether host- feeding was required before oviposition. Tycherus osculator adults were reared from 468 THE CANADIAN ENTOMOLOGIST J~lylA~g~st1999

Z. diniana collected from Ste-Anne-la Condamine (44'27'N, 6"42'E), Myronnes (44"27'N, 6"47'E), and Le Lauzet (45"01%, 6'28%) in the French Alps in July and Au- gust 1996. Mated females were overwintered in 1.5-L unscreened styrofoam boxes in a cold room maintained in darkness at 2'C at >90% RH and outdoors in a hole in the ground to simulate natural temperatures. Both overwintering methods resulted in less than 20% mortality, but the females kept in the ground were transferred to the cold room in March to delay the onset of spring activity brought about by warmer temperatures. Females were moved from the cold in June and July to the laboratory and exposed to larvae, prepupae, and pupae of Z. diniana collected as larvae at Les Haudkres (46"05'N, 7"31'E), Valais, in the Swiss Alps. All the experiments were carried out in the laboratory at 21 + 2°C under natural light from a large north-facing window. Adult i? osculator were kept in ventilated 1.3-L plastic cylinders, fed with honey, and provided with moistened tissue paper as a source of water. Zeiraphera diniana larvae were reared in gauze-covered wooden cages (25 x 25 x 50 cm) or in ventilated styrofoam boxes (15 x 25 x 25 cm) and fed with fresh larch, Larix decidua Miller (Pinaceae), foliage. Attacked pupae and prepupae were kept in a growth chamber at 20 + 0.5'C under a 12L:12D photoperiod until emergence of the parasitoid or host adult. Ovarian maturation and preoviposition period. Ovarian maturation was determined by dissecting three females daily from the day of transfer from the cold (day 0) to day 9. For each female, the largest egg was measured and yolk formation was observed. The preoviposition period was determined by placing a female following transfer from the cold in a screened, 0.5-1.5 L, styrofoam box containing three pupae 12 days old. Exposures began 2 days (n = 3 repetitions) and 5 days (n = 3 repetitions) following transfer from the cold and ended 13 days later. Pupae were changed daily and reared. Host stage-age and parasitization. Hosts of various ages and developmental stages were exposed to gravid 7: osculator females in 1.5-L ventilated styrofoam boxes. Mature larvae, prepupae, and pupae were placed in holes punched into the box bottoms to curtail movement and covered by a thin (1 mm) layer of frass and larch litter. In the first experiment, the various developmental stages were exposed separately to assess their suitability for parasitoid attack and development. Three groups of four females were confined 24 h with a minimum of 10 and a maximum of 24 pupae 2, 4, 6, and 8 days old. Pupae older than 8 days were not tested because, at room temperature, spruce bud moths start to eclose 11 days after pupation. The same experiment was repeated with a minimum of 10 prepupae and 10 mature larvae, but for 8 h only, to re- duce the probability of pupation during the exposure period. To determine whether parasitoid females prefer to oviposit in pupae or prepupae, five females were exposed to 10 pupae (52 days in age) and 10 prepupae in two-choice tests (n = 10 repetitions) lasting 8 h. In an additional test (n = 12 repetitions), single females, confined with three pupae and three prepupae as described above, were observed for I h to record the occurrence of host feeding and oviposition. Sex ratio, development time, and longevity. Tycherus osculator progeny were checked daily and reared until death to determine sex ratios resulting from attacks by females older and younger than an arbitrarily assigned age of 15 days following overwintering, and the period of development from oviposition to adult emergence and longevity of the adults.

Screening of Tycheius osculator Against the Spruce Bud Moth. Source and shipment of Tycheius. Parasitoids were reared from Z. diniana collected as pupae on 31 July 1995 in a stand of larch at Larche (44"27%, 6"51%), France. Twenty female Volume 13 1 THE C4NADIAN ENTUMOl.CKiTST 469

T. osculator, overwintered outdoors as described above, were shipped from the CABI- Bioscience European Station, DelCmont, Switzerland, on 29 April 1996 and arrived 2 days later at the Canadian Forest Service laboratory in St. John's, Newfoundland. One T. osculator was dead on arrival but the others were healthy and lived as long as 4 mo. Screenings against prepupae and pupae of spruce bud moth were initiated on 3 May 1996 in a quarantine laboratory. The females were held in groups of four or five, in 21 x 12 x 7 cm Frig.O.SealB clear plastic containers ventilated on one side with 0.5- mm screening and provided with fresh water (cotton batting saturated with distilled water) and honey. Containers were cleaned and replaced with fresh water and honey every 2-3 days. Bioassays were conducted during daylight hours at 20-25"C, approximately 60% RH, and under a natural photoperiod provided by a west-facing window. Bioassay I. Two 7: osculator females were confined for 5-10 min in a 1.5 x 9.0 cm Petri dish with two spruce bud moths in the prepupal and (or) pupal stages. Each dish contained a thin layer of humus collected from the field sites monitored for endemic parasitoids, fresh white spruce buds-shoots, and spruce bud moth frass. At the end of each 5- to 10-min exposure the hosts were removed and replaced with unex- posed hosts. Five exposures were run before females were replaced and no female was used for more than five assays per day; however, the same females were used in assays on following days. Sixty-nine bioassays representing 345 exposures were run and a total of 19 females were used. Attacked hosts were removed and reared individually in 60-mL vials until adult emergence, death, or parasitoid emergence. Hosts were reared in a controlled-environment (ConvironTM) chamber with conditions similar to those in the quarantine room. During the bioassays the duration of exposure, number of encounters, number of attacks, and duration of attack were noted. An encounter between a parasitoid and a bud moth host was recorded if the host was touched by the parasitoid's an- tenna. Bioassay 11. Five female parasitoids were confined for 24 h with 10 spruce bud moth prepupae or pupae in a plastic container as described above containing a 1-cm layer of humus, freshly cut buds and shoots of white spruce, and spruce bud moth frass (n = 6 replicates). These containers also contained water and were streaked with honey to provide a food source for the parasitoids. Following exposure to the parasitoids, all hosts were removed, placed in a new vial, and reared until the adult stage, death, or parasitoid emergence. Voucher specimens. The imported 7: osculator and progeny emerging from exposed pupae were retained in the insect collection at the National Resources Canada (Newfoundland) laboratory in St. John's. Body lengths and widths of the head and thorax were measured for eight imported 7: osculator and eight female progeny from spruce bud moth parasitized by 7: osculator. Statistical analysis. Data distributions were tested for normality (Shapiro and Wilk 1965); means were compared with Student's t test (Sokal and Rohlf 1965) or, if nonparametric, distributions were compared with the Mann-Whitney Rank Sum Test (T) (Sokal and Rohlf 1965) (Sigmastat Statistical Software 1994).

Results

Local Parasitoid Complex Larval parasitoids. Lamachus (Foerster 1868) sp. and Triclistus (Foerster 1868) sp. (Hymenoptera: Ichneumonidae) were reared at incidences less than 6% from spruce bud moth collected as larvae from the Arterial site (Table 1). There was a low incidence of other parasitoid species at all sites (Table 1) but these failed to emerge from their cocoons. The cocoons of these unknown species resembled but could not be confirmed as being those of E. zeirapherae. TABLE1. Incidence (%) of endemic parasitoids in Zeiraphera canadensis populations at two sites near St. John's, Newfoundland, 199&1996

Larval collections* Pupal collectionsr Parasitoid species 1994 1995 1996 1994 1995 1996 Torbay Ascogaster sp. Clinocentrus sp. Earinus zeirapherae Lamachus sp. Triclistus sp. Tycherus osculator Unknown Total no. of hosts

Arterial Ascogaster sp. Clinocentrus sp. Earinus zeirapherae Lamachus sp. Triclistus sp. Tycherus osculator Unknown Total no. of hosts

*Obtained from 25-cm branch tips. +Obtained from 1-m2 drop trays trapping late-stage larvae and pupae. %even percent of pupae collected in 1996 from the Torbay site and 30% of pupae from the Arterial site were desiccated and mouldy following the overwintering period and they were not in a condition to determine whether or not parasitism had occurred. The incidence of Earinus zeirapherae, therefore, could not be determined because this species only emerged from overwintered pupae. TIncludes parasitoids obtained by dissection and from overwintering pupae.

Late-stage larval-pupal parasitoids. Hymenopteran parasitoids reared from drop-tray collections of late-stage larvae and pupae of spruce bud moth included three species of Braconidae, E. zeirapherae, Ascogaster (Wesmael 1835) sp., and Clino- centrus (Haliday 1833) sp., and the Ichneumonidae 7: osculator (Table 1). Tycherus osculator and E. zeirapherae were the most frequently encountered species throughout the survey. In 1996, parasitism by 7: osculator accounted for 30% and 50% of pupal mortality at the Arterial and Torbay sites, respectively, but occurred at a much lower incidence in 1994 and 1995 (Table 1). Earinus zeirapherae was recovered at incidences between 6% and 16%, whereas Ascogaster sp. and Clinocentrus sp. were rarely encountered (Table 1). Less than 8% of the material reared from the drop trays contained parasitoids that failed to complete development and thus could not be identified.

Biology of Tycherus osculator on Zeiraphera diniana. Ovarian maturation and pre- ovipositional period. Tycherus osculator adults mated immediately after emergence. Eggs began to turn yellow 1 day after incubation, but the size of the largest egg increased until 5 days after incubation (Fig. 1). There were three ovarioles per ovary, with each of the ovarioles containing a maximum of two mature eggs and about 12 immature eggs. Two females started to oviposit 3 and 4 days after incubation, respectively, but the third female never oviposited. When the experiment was repeated, starting 5 days after incubation, two of the three females oviposited on the first day of exposure to Z. diniana. THE CANADIAN ENTOMOLOGIST

Days after incubation at 21 "C FIGURE1. Mean (+SE) length of largest egg dissected from Tycherus osculator females 0-9 days after incubation at 21°C, following overwintering at 2°C. Three females were dissected daily and none were exposed to hosts prior to dissection. The largest egg for each of the three females dissected on day 9 was 0.72 mm.

Host stage-age and parasitization. Tycherus osculator adults eclosed from host pupae of all age groups and only at an incidence of one adult per host pupa. Even the 8- day-old pupae stung by 7: osculator were successfully parasitized. At least three parasitoid adults emerged from pupae attacked as prepupae. In contrast, larvae were not accepted as hosts. Of the 27 attacks observed, nine were under 1 min in duration and did not result in progeny, whereas 18 lasted over 1 min (max. 11 min) and resulted in eight progeny. When females were given the choice between pupae and prepupae, a total of seven pupae (7%)and one prepupa (1%) were successfully parasitized in the 8-h choice tests and a total of only four pupae (11%) and no prepupae were attacked in the 1-h choice tests. In view of these low rates of parasitism, more tests are needed to determine host stage preference. Females usually fed on their hosts before or after oviposition, sometimes up to 10 min, but this activity was not obligatory for ovarian maturation because eggs ma- tured in females that were not exposed to hosts (Fig. 1). No host adult or 7: osculator emerged from the 12 host pupae on which feeding was observed, and this may suggest that feeding may have lethal effects. Sex ratio, development time, and longevity. A total of 33 males and 33 females were obtained from our laboratory experiments. For mothers younger than 15 days (n = 20) following overwintering, the sex ratio of the progeny was 12 males for 31 females, whereas for mothers 15 days and older (n = 20) following overwintering, 21 males and two females were obtained, perhaps because sperm quality decreases with age. The mean (+SE) development time for males (20.9 + 0.2 days, n = 20, range = 20- 23) was shorter (t48= 4.58, P < 0.00001) than that for the females (22.6 + 0.3 days, n = 30, range = 21-27). Females were long-lived and survived up to 13 months following eclosion and up to 120 days following overwintering. Screening of Tycherus osculator against the spruce bud moth. Bioassay I. Tycherus osculator attacked and parasitized exposed spruce bud moth pupae, and its offspring successfully developed within the pupae. Tycherus osculator spent a considerable amount of time searching the litter layer. An average of 27.2 host-parasitoid encounters per assay were recorded. Tycherus osculator occasionally fed from oviposition wounds on attacked host prepupae. Thirteen attacks, lasting approximately 3 min, on three prepupae and four pupae were observed. One adult Tycherus emerged from one of the attacked prepupae. The attack in this case lasted 55 s. No parasitoid progeny emerged from the other attacked hosts, but many of these were desiccated when the growth chamber overheated. Bioassay 11. Tycherus osculator progeny successfully developed and emerged from 16 of 600 prepupae and pupae exposed in 60 bioassays. The developmental time of T. osculator from the time of oviposition until emergence varied from 14 to 19 days. Progeny including the single individual reared in bioassay I had a male to female sex ratio of 1:16. Exposed Z. canadensis prepupae which failed to pupate showed no sign of parasitoid development. Tycherus oscuiator females reared from spruce bud moth were smaller than those reared from 2. diniana for each parameter examined. Mean (+SE) body lengths and widths of the head and thorax were 6.31 * 0.08, 1.24 + 0.02, and 1.31 * 0.03 mm, respectively, for 7: osculator ex Z. diniana and 4.96 * 0.18 (tI4= 6.75, P < 0.00001), 1.07 + 0.04 (T = 94.0, P = 0.00466), and 1.07 + 0.03 mm (t,, = 4.87, P = 0.00020), respectively, for 7: osculator ex spruce bud moth.

Discussion

The low rates of larval parasitism are consistent with those reported by Pilon (1965) in Quebec and Turgeon (1992) in New Brunswick. The species of Larnachus and Triclistus were the only parasitoids reared from spruce bud moth larvae in the present study. Triclistus podagricus (Gravenhorst) was commonly observed by Pilon (1965), but Larnachus sp. has not been recorded previously as a parasitoid of 2. canadensis. The only other parasitoid species reared from larvae of spruce bud moth in Newfound- land are Macrocentrus rnarginator Nees (Braconidae) and Diadegrna sp. (Ichneu- monidae) (Bowers and Pardy 1997). Additional Hymenopteran parasitoid species, present in New Brunswick and recorded at incidences under lo%, include Enytus rnontana (Ashmead) (Chalcidae), Glypta sp. (Ichneumonidae), and Apanteles sp. (Braconidae) (Ostaff 1995). The likelihood of available niches for introduced larval parasitoids (e.g., 7: carbonellurn) appears high. Parasitoids emerging from late-stage larvae and pupae of the spruce bud moth were identified as 7: osculator, E. zeirapherae, Ascogaster sp., and Cbinmentrus sp. These species represent new distributional range extensions to Newfoundland, and to the nearctic region in the case of 7: osculator. Earinus zeirapherae was recorded as attacking and killing spruce bud moth in the larval stage in New Brunswick (Turgeon 1992) and it is likely that it emerges also from larvae in Newfoundland because cocoons resembling those of E. zeirapherae were associated with larvae killed in their fourth instar. We can only confirm that E. zeirapherae emerged from host pupae in the present study, accounting for up to 16% parasitism, and this is unusual because Hy- menopteran parasitoids of the subfamily Agathidinae are only known to kill larvae and prepupae (Shaw and Huddleston 1991). Our results also confirm unpublished observations by DP Ostaff (Canadian Forest Service, personal communication) that E. zeirapherae emerges from host pupae. In 1994, up to 11% of spruce bud moth were parasitized by 7: osculatol: Nearly absent from survey data in 1995, 7: osculator Volume 131 MECANADlAN ENTOMOLOGIST 47 3 reappeared during the summer of 1996, at incidences of up to 50%. The unusually mild winter of 1996 may have contributed to increased survival of T. osculator females overwintering from 1995. Incidences up to 50% are comparable to incidences of about 60% parasitism of Z. rufimitrana and Z. ratzeburgiana in Europe recorded by Schon- berg (1993). Up to 30% parasitism of Z. diniana by T. osculator has been observed (M Kenis, unpublished data). The remaining parasitoids, Ascogaster sp. and Clinocentrus sp., appear to be inci- dental species and most probably kill late-stage larvae or prepupae. The subfamily Cheloninae, which includes species of the genus Ascogaster, has exclusively egg-larval or egg-prepupal parasitoids (Shaw and Huddleston 1991). The species of Clinocentrus are also known to kill their host in the prepupal stage (Shaw and Huddleston 1991). Tycherus osculator ex Z. diniana attacked, parasitized, and successfully developed within Z. canadensis, although their progeny had significantly smaller body lengths, head widths, and thorax widths than i? osculator reared from Z. diniana. Similarly, specimens emerging from the larger Z. diniana are larger than those emerging from the smaller Z. ratzeburgiana and Z. rufimitrana in Europe (M Kenis, unpublished data). The size of the host in which the parasitoid develops is a factor known to influence parasitoid size (Bai et al. 1995). Important aspects of the biology of i? osculator were determined and these results will be useful to programs evaluating potential releases of 7: osculator, obtained from either Europe or Newfoundland, in mainland Canada where this parasitoid is apparently absent (e.g., Pilon 1965; Turgeon 1992; Ostaff 1995). Tycherus osculator from the St. John's area would be a convenient source of material for biological control introduc- tions to central Newfoundland should Z. canadensis become a pest there.

Acknowledgements We thank A Carroll, D Ostaff, and two anonymous reviewers for reviewing earlier drafts of the manuscript and A Van Averbeke, S Monnet, S Peddle, and C Lopez- Vaamonde for technical assistance. We are grateful to Erich Diller (Zoologische Staat- sammlung, Munich) and the late John Barron (Eastern Cereal and Oilseed Research Centre, Ottawa) for examining and identifying specimens of Tycherus osculator. Addi- tional identification of parasitoid specimens was kindly given by Michael Sharkey (University of Kentucky).

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