Biology and host specificity of Apion miniatum (Coleoptera: Apionidae) from Israel, a potential biological control agent for Emex australis and Emex spinosa (Polygonaceae) in Australia

Biological Control 33 (2005) 20–31 www.elsevier.com/locate/ybcon

Biology and host speciWcity of Apion miniatum (Coleoptera: Apionidae) from Israel, a potential biological control agent for Emex australis and Emex spinosa (Polygonaceae) in Australia John K. Scott ¤, Paul B. Yeoh CSIRO Entomology, Private Bag 5, PO Wembley, WA 6913, Australia Received 13 June 2004; accepted 13 January 2005

Abstract The annual southern African plant, Emex australis (Polygonaceae), is a major weed of crops and pastures in Australia. The only other member of the genus is the Mediterranean species Emex spinosa, which is a comparatively minor weed. Both species are targets for biological control in Australia. Past biological control eVorts using two weevil species from South Africa and Morocco have not been successful and this has been attributed to the hot dry summers of the Australian regions infested by the weed. An apionid weevil, Apion miniatum, is known from Europe and West Asia and was discovered on E. spinosa in Israel in hot, dry climates similar to E. australis infested regions of Australia. The larvae tunnel in stems, crowns, and roots and the adults feed on leaves and lay eggs into stems and petioles of E. spinosa. Apion miniatum from Israel was imported into Australia where it was reared on E. australis and host speciWcity tested on 81 plant species in quarantine. Its host range is restricted to Emex species and some Rumex species including, Rumex conglomeratus, R. crispus, R. pulcher, and R. obtusifolius, species that are also declared targets for biological control in Australia. It was concluded that A. miniatum does not present a risk to economic crops or native Australian Xora. Based on the Wndings summarized within this paper, the Australian regulatory bodies approved the insect’s release within Australia and the inaugural release onto E. australis Weld populations began in 1998. The insect appears not to have established in Australia despite being selected from a region with extreme summer conditions. The biology of A. miniatum is discussed within this context.  2005 Elsevier Inc. All rights reserved. Keywords: Apion miniatum; Australia; Biological control; Emex australis; Emex spinosa; Host speciWcity; Israel

1. Introduction Emex australis Steinh. (Polygonaceae) is an annual weed of cropping systems across southern Australia. It originates in southern Africa and has been established in Australia since the 1830s (Gilbey and Weiss, 1980). In Western Australia E. australis is especially serious, infesting about 500,000 ha of cereal crop and signiWcantly reducing wheat yields (Gilbey and Weiss, 1980). The plant has a long-lived seed bank and germination is from autumn to early winter (March to June). ¤

Corresponding author. Fax: +61 8 9333 6646. E-mail address: [email protected] (J.K. Scott).

1049-9644/$ - see front matter  2005 Elsevier Inc. All rights reserved. doi:10.1016/j.biocontrol.2005.01.009

The rosette develops a strong taproot. Stems have indeterminate growth, producing spiny achenes (dry, indehiscent fruit containing a single seed) at stem nodes. The plants senesce in October to November when the rains cease. Cultivation may kill seedlings, but larger rosette plants with their well developed taproots can survive. The disturbed soil also favors subsequent germination of the weed. Chemical control is successful in cereal crops. However, in pastures, herbicides are diYcult to use because of the damage caused to the annual legumes. Sheep will graze plants when little other food is available, but the spiny achenes can cause lameness in lambs (Gilbey and Weiss, 1980). The spiny fruits also cause severe contamination of lupins, peas, and dried fruits.

J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

Perapion antiquum (Gyllenhal) (Coleoptera: Apionidae) was collected from E. australis in South Africa in 1956 and successfully used as a biological control agent in Hawaii, but in Australia it failed to establish at most sites and where it did establish; it did not control the weed (Julien, 1981, 1992). This species was unable to survive the summer period at most of the release locations within Australia (Julien, 1981) and climatic models constructed by Scott (1992) predicted that this was because the summer periods at these sites were dryer and hotter than that experienced by the insects in South Africa. Coastal areas in southern Australia were predicted to be climatically favorable to P. antiquum, however these were not typically areas where E. australis is a major problem. Extensive searches for other biological control agents in southern Africa during the 1980s concluded that there were few prospects for introducing into Australia the natural enemies that have co-evolved with E. australis because most were not suYciently host speciWc (Scott and Way, 1990). The most promising of all agents was the southern African fungus, Phomopsis emicis Shivas (Sphaeropsidales). However, it was subsequently discovered to be already widespread in Australia where it causes up to 30% reduction in seed survival (Scott and Way, 1990; Shivas et al., 1994; Shivas and Scott, 1994). The second species in the genus, E. spinosa (L.) Campd., is a minor weed in Australia. It originates from the Mediterranean basin and has a similar biology to E. australis. Lixus cribricollis Boheman (Coleoptera: Curculionidae) was collected from E. spinosa in Morocco and released against Emex and Rumex species (Polygonaceae) in the early 1980s, but appears not to have survived (Julien et al., 1982; Julien, 1992). Surveys in Israel in 1988 (Scott unpublished) showed that the apionid weevil Apion miniatum Germar ( D Erythrapion miniatum or Apion (Erythrapion) miniatum) (Coleoptera: Apionidae) was damaging on E. spinosa. This insect is also known from Europe, Syria, and the Caucasus (Wagner,

21

1910). It was considered a potential biological control agent because it occurs in locations with a Mediterranean climate in which the summer period is as hot, and as dry, as many of the places where E. australis is a problem in Australia (Yeoh et al., 2002). In addition, it appeared to have a limited host range. The published host records of A. miniatum and the seven related species are mostly from the Polygonaceae, and in particular, Rumex species (Table 1, Alonso-Zarazaga, 1990). The recorded hosts of A. miniatum include two of the four Rumex subgenera (Rumex and Acetosa) (Table 1). Samedov (1963 in Ter-Minassian, 1972) records A. miniatum in Azerbaijan “on” Rumex acetosa L., Rumex pulcher L., Rheum, and Ribes. The life stage or degree of attack by the insect, if any, is not mentioned. In this paper, we report on the biology and host speciWcity of A. miniatum collected from Israel. Based on these Wndings, the Australian regulatory bodies approved this insect’s release within Australia and the inaugural release onto E. australis Weld populations began in 1998. Despite speciWcally selecting A. miniatum because of its ability to endure extreme summer conditions, it again seems to have been a potential biological agent that was unsuccessful in Australia. Aspects of the insect’s biology are discussed with respect to its apparent inability to establish in Australia so that future biological control agents can be selected or rejected more eYciently.

2. Materials and methods 2.1. Source and rearing of insects Apion miniatum (533 adults) were collected in Israel in 1993 and 1994. They and their oVspring were maintained in cages of Terylene voile sleeves over potted plants of E. australis, E. spinosa, or occasionally on Rumex crispus L. Egg-laying adults were moved to new cages after 2

Table 1 Published host range of A. miniatum and closely related species as deWned in Alonso-Zarazaga (1990) Apion species

Host rangea

Reference

Apion species A. cruentatum Walton A. frumentarium (Paykull)

Atraphaxis, Calligonum, Polygonum, Rumex Rumex acetosa L. R. acetosa, R. acetosella L., R. angiocarpus Murb., R. bucephalophorus L., R. patientia L., R. thyrsoides Desf. Calligonum caput-medusae Schrenk R. tingitanus L. R. conglomeratus Murray, R. hydrolapathum Hudson, R. obtusifolius L., R. sanguineus L., R. thyrsoides R. crispus L. R. acetosa, R. pulcher L., Rheum and Ribes (Grossulariaceae) R. acetosella R. acetosella R. teucriumb, Oenanthe sp. (Apiaceae)

Alonso-Zarazaga (1990) Dieckmann (1973) HoVmann (1958)

A. gallicola Ter-Minassian A. henoni Ab. A. miniatum Germar

A. rubens Walton A. sanguineum (DeGeer) a b

Ter-Minassian (1971) PeyerimhoV (1926) HoVmann (1958) Dieckmann (1973) Samedov (1963 in Ter-Minassian, 1972) HoVmann (1958) HoVmann (1958) Ter-Minassian (1972)

All the hosts are from the Polygonaceae (mostly tribe Rumiceae), except two cases (the family is indicated in parentheses). Authority unknown.

22

J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

weeks and larvae were allowed to complete development. All observations and experiments were carried out in a controlled temperature quarantine glasshouse set to a 10–15/25 °C night/day regime. The insects used in this study were identiWed as A. miniatum at the British Museum (Natural History) and compared with identiWed specimens in the Israeli National Insect Collection in Tel Aviv. Specimens have been lodged with the Australian National Insect Collection and the Israeli National Insect Collection. 2.2. Host-speciWcity tests The test plant list of 81 species was selected following the centrifugal phylogenetic sequence outlined by Wapshere (1974). The list has many similarities with previous host-speciWcity test lists prepared for other biological control agents against Emex (Harley and Kassulke, 1975; Harley et al., 1979; Julien et al., 1982; Scott and Way, 1989a,b; Shepherd, 1989; Shivas et al., 1994), but includes a re-examination of some criteria, especially the selection of economic plants to be included because A. miniatum was collected from the Eastern Mediterranean region, whereas the past source areas for biological control agents were the Western Mediterranean and southern Africa. The test list included cultivated and native Australian Polygonaceae species that have an overlapping distribution with Emex in Australia. Species tested included three of the four subgenera in the genus Rumex; Acetosa, Acetosella, and Rumex. The remaining subgenus, Platypodium, was not found in Australia during this study (cf. Rechinger, 1984) and was examined in the Weld in Israel. The selection of orders and families related to the Polygonaceae was based on Cronquist (1981) excluding Basellaceae and Molluginaceae which do not have economic species in Australia. A species of Drosera was added because of indications from analyses of nucleotide sequences are that Droseraceae may have aYnities with Polygonaceae (Chase et al., 1993). Oxalic acid is suspected of providing a feeding stimulus for insects which specialize on plants of the Polygonaceae (Matsuda and Matsumoto (1975) in Matsuda, 1976), therefore representatives of two plant genera (Begonia and Oxalis) also known to contain high levels of oxalic acid were included in the tests. Representatives of non-Polygonaceae plant families recorded in the literature as hosts of Apion (as deWned in Alonso-Zarazaga, 1990) were included if present in Australia (Table 1). Plant names follow the usage of Mabberley (1987) and Hnatiuk (1990). 2.3. Host-speciWcity test methods Host choice is entirely determined by adult A. miniatum because larval development is completed within the

plant chosen by the female. Three types of host-speciWcity tests were undertaken with adult A. miniatum, short term with no choice, long term with no choice, and short-term choice tests. The aim of the short term with no-choice tests was to determine the degree to which adults would feed and lay eggs in test plants and also to examine the survival of early instar larvae on plants where eggs were laid. The aim of the long-term tests was to determine if the plant was suitable for A. miniatum to complete its larval development; only plants likely to be hosts were included, either based on the literature or based on the results of the short-term tests. In the choice tests, adult A. miniatum were able to choose between the target and test plant species for adult feeding and oviposition. In all tests, E. australis, the main target weed, was used as the control. 2.4. Short-term, no-choice tests Polygonaceae test and target (control) plants were caged individually using a white Terylene voile sleeve secured to their pot and supported by a wire frame. NonPolygonaceae test plants were similarly caged, however rather than caging and testing individual plants, a single plant from each of approximately Wve diVerent test species (average D 4.5, SE D 0.88, n D 11) was placed simultaneously within a single plastic tray (32 £ 40 £ 15 cm) and then a Terylene voile sleeve was secured to the tray to enclose all the plants together. This “group testing” was done to conserve insects and to speed up the testing procedure. It was based on the assumption that there was only a small probability of any of the non-Polygonaceae species being susceptible and therefore the insects were being oVered no suitable choice (rather than no choice in the strict sense). To control for the possibility of the insects acquiring adequate resources (food/oviposition sites) from the simultaneous attack of numerous species, species were to be re-run individually if the total level of attack within the cage (all species combined) approached that observed within the control plants (>20% of control values in the same block). No tests needed to be re-run. Each test plant was replicated six times and six E. australis plants were set up as controls with every batch of test plants. At least one mating pair per cage, but normally two or three pairs of A. miniatum, was used (average 2.4 males, 2.3 females per cage, total D 217). All caged test and target plants within a particular experimental batch were randomly assigned positions on benches within the same section of the glasshouse. Insects were usually removed after 14 days however a shorter duration (minimum 7 days) was sometimes necessary to prevent excessive egg-laying on the control plants as this caused the plants to die prematurely. Insects set up in experiments at the start of their breeding season were left on the plants for a longer duration (25 days) to compensate for individual variation in the onset of sexual activity and egg-laying.

J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

After the adults were removed, the number of feeding holes was counted and an estimate of the number of eggs laid was made by examination under a binocular microscope. The test continued for a further 2 weeks after which the plants were dissected under a binocular microscope, the number of eggs, larvae, and tunnels was counted and the head-capsule width of larvae was measured. Some larvae, and in particular the Wrst instars, were diYcult to measure because they were destroyed by other larvae, damaged during dissection, decomposed or just not found. Their Wnal developmental stage was estimated from the diameter of their tunnels. A second estimate of the number of eggs laid was made when the plants were dissected and the larger of the two estimates reported (no consistent diVerence found; Wrst method slightly higher with diVerence D 0.8 eggs per plant, SE D 1.31, n D 74). The duration of the short-term tests was suYcient for larvae to have reached the third instar in Emex species. 2.5. Long-term, no-choice tests Rumex species were suspected of being suitable hosts and they were included in long-term tests without going through the short-term test phase. Only one test species, Rheum rhabarbarum L. (Polygonaceae), had live larvae at the conclusion of short-term tests, thus it was also included in the long-term tests. Long-term tests were conducted on individually caged plants as described above in the short-term test section (average 2.1 males, 2.0 females per cage, n D 128). The adults were removed and the feeding holes were counted after 9–16 days. One month after the last emergence of any adult, which was usually 4–5 months after the experiment was initially set up, the plants were dissected under a binocular microscope and the fate of any egg was determined. In the Wrst of these experiments, the number of eggs laid into each plant was not estimated when the adults were removed. After 4–5 months, at the conclusion of the experiment, it was impossible to determine how many eggs and dead larvae were present initially. As a consequence, for all subsequent experiments the number of eggs laid was estimated by the same methods as described in the short-term no-choice tests. Based upon these egg counts, the long duration of these experiments generally resulted in many of the dead larvae being unaccounted for (average diVerence D 4.3 individuals per plant, SE D 0.78; paired t test; t D 5.49, 83 df, P < 0.01).

23

experiments were run, one using four pairs, the other using six pairs of A. miniatum per replicate. Adult A. miniatum were introduced via a hole in the top of the cage. They were then removed after 2 weeks at which time the number of feeding holes was counted and an estimate of the number of eggs laid was made. The plants were dissected after a further 2 weeks and live and dead larvae were counted separately. 2.7. Analysis of results To enable comparison between and within tests, results were scaled to the number of feeding holes per adult per fortnight or the number of eggs and larvae per female per fortnight (time during which adults were egglaying in the tests). All results are presented as means § standard error. The data in the choice tests were analyzed as two-way ANOVA with cages treated as “blocks” to remove any bias caused by cage to cage variation due to insect fecundity, cage position and/or time of dissection. Data were either log10 + 1 or arcsine transformed and tested for homogeneity of variance. 2.8. Field observations of host speciWcity Polygonaceae species were examined in the Weld in Israel when they were found growing with E. spinosa containing A. miniatum. At least 30 plants or 30 stems per species were dissected for evidence of tunneling or Apion larvae.

3. Results 3.1. IdentiWcation of Apion miniatum Two observations indicate that the insects collected from E. spinosa and identiWed as A. miniatum may belong to a diVerent biotype to the same species found on Rumex species in Europe. Adult males from Rumex have a release behavior (tapping of the abdomen) when mounted by other males. Adult males from E. spinosa appeared to recognize the sexes and the tapping behavior was not displayed. Second, the adult Israeli biotype is tan colored when newly emerged, becoming red with maturity, whereas the color range of the European biotype is blood red or orange (HoVmann, 1958). 3.2. Biology of the Israeli biotype

2.6. Choice tests Test plant species and E. australis were randomly placed together on single plastic trays (32 £ 40 £ 15 cm) and caged by using a white Terylene voile sleeve secured to each tray and supported by a wire frame. Two choice

Apion miniatum adults are 3.3–4.3 mm long. Sexually active males within a group can be readily identiWed by their mounting behavior, however any adult can be sexed by counting the number of sternites visible on the ventral side of the abdomen. Males have Wve whereas

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J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

females only have four visible sternites. The translucent eggs (length 0.6 mm and width 0.35 mm) were laid singly in holes excavated by the rostrum of the female in the petiole or stem of E. spinosa. Egg-laying occurred from winter through to the senescence of the plant at the start of summer. The extruding portion of the egg (approx. 0.2 mm diameter) was covered with black frass. The eggs hatched after 5–7 days. The larvae in the petiole tunneled towards the stem and those in the stem tunneled towards the crown of the plant. Later instars were found in the stem, crown, and upper taproot. The most extensive tunneling occurred in the crown of the plant. Instar size, as measured by head-capsule width, was the same for larvae reared from either species of Emex (ANOVA, between Emex species F1,139 D 0.07, P D 0.79; between instars F1,139 D 1241, P < 0.0001; interaction term F2,139 D 2.46, P D 0.089). The width of the head capsules (mean § SE) for individuals reared under glasshouse conditions on either species of Emex was as follows: Wrst instars 0.26 § 0.005 mm (n D 17, range 0.22–0.30 mm), second instars 0.43 § 0.004 mm (n D 71, range 0.36– 0.50 mm), and third or Wnal instars 0.70 § 0.005 mm (n D 57, range 0.60–0.78 mm). Pupation occurred inside the stem, crown, or upper taproot. The newly emerged adult was tan and changed to red after feeding for about 1 week. The adults fed by making small (approx. 1.6 mm2) holes in leaves of E. spinosa. The site of adult aestivation in the Weld is unknown, but in cages they hid among dead leaves on the plant or moved to the top of the cage and rested in crevasses and similar sheltered positions. If placed upon the trunk of a rough-barked tree, they will force their way into the crevices until they are out of sight. Aestivation lasted 4–6 months. The exact clues for the resumption of feeding and development of eggs are unknown, but can be stimulated by shortening day length (approx. 10 h 30 min). 3.3. Host-speciWcity tests Feeding by adult A. miniatum was insigniWcant, but was greatest on plants from the tribe Rumiceae (Emex, Rumex, and Rheum) with negligible or no feeding on other species (Tables 2 and 3). The adult’s mouth parts are small and make small “shot holes” in the leaf surface, with a mean of 1.6 mm diameter, representing 2.0 mm2 consumed. A very low number of feeding holes was recorded on two non-Polygonaceae species, begonia and raspberry (Table 3). In contrast, species from the tribe Rumiceae were consumed at a rate of 50 mm2 per individual per fortnight. However, on other tribes there were only low levels of attack and less than 6 mm2 of leaf per individual per fortnight was consumed. The maximum observed average feeding rate (from all species tested) of only 109 mm2 per individual per fortnight indicates that, by itself, adult feeding is unlikely to cause any signiWcant

damage to any plant attacked (based upon the very low densities of insects observed in the Weld whilst collecting the initial stocks from Israel). 3.4. Short-term, no-choice tests Larvae were only able to feed and develop in plants of the Polygonaceae tribe Rumiceae (Emex, Rumex, and Rheum species). Egg-laying adults were exposed to 34 non-Polygonaceae families including 49 species of plants. The assumption of minimal attack rates and therefore the validity of group testing the non-Polygonaceae held true with the total number of eggs laid and feeding holes produced per cage (all species combined) averaging 0.5% (SE D 0.54%, n D 11) and 2.7% (SE D 1.0%, n D 11), respectively, of the corresponding control plants. Of all the non-Polygonaceae species, slight feeding damage was recorded only for Rubus idaeus L. (Rosaceae) and Begonia sp. (Begoniaceae) (4.7 and 5.3%, respectively, of the damage recorded in control plants from the same batch). Eggs were only laid on Begonia sp. (Begoniaceae). These were in low numbers (6% of the number observed in the control E. australis plants) and none of them developed past Wrst instar (Table 3). Eggs were laid on 6 of the 15 Polygonaceae species tested in the tribes Coccolobeae, Persicarieae, and Polygoneae (0.5–11.0% of the number observed in the controls from the same block), but no development occurred past Wrst-instar larvae. No eggs were laid in buckwheat, Fagopyrum esculentum Moench, the only crop plant in this group (Table 3). Eggs were laid on all species in the tribe Rumiceae (Emex, Rumex, and Rheum). A higher level of oviposition, but not adult feeding, was noted on E. spinosa when compared to the E. australis controls in the same batch, with only 11.4 § 3.43 eggs/female/fortnight laid into E. australis (t D 3.13, 10 df, P < 0.05) but 37.3 § 6.77 feed holes per insect per fortnight recorded (t D 0.26, 10 df, ns). Two sets of experiments were run for rhubarb (Rheum rhabarbarum), the Wrst using Weld-collected A. miniatum which had developed on E. spinosa and the second using laboratory-reared insects which had developed on E. australis. For both experiments, the petioles of rhubarb received approximately 50% of the eggs observed on the corresponding control E. australis plants. In contrast to plants of Emex spp., rhubarb plants produced copious amounts of thick mucilage at the sites of A. miniatum damage. Eggs and larvae were sometimes observed trapped in beads of mucilage extruding from the oviposition site. Most larvae within petioles of rhubarb were found dead, trapped within their tunnels Wlled with mucilage. In the Wrst set of experiments, no live larvae were found in rhubarb when they were dissected and in the second set of experiments, a single larva developed to second instar and the

J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

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Table 2 Families, species, and common names of plants included in host-speciWcity tests of A. miniatum with an average score of <1 and maximum scores of <2 feed holes per adult per fortnight and on which no eggs were laid Family

Species

Common name, (A) Australian native

Actinidaceae Aizoaceae Amaranthaceae Anacardiaceae

Actinidia chinensis Planchon Carpobrotus edulis (L.) N.E.Br. Amaranthus tricolor L. Anacardium occidentale L. Mangifera indica L. Anethum graveolens L. Apium graveolens L. Pastinaca sativa L. Ananas comosus (L.) Merr. Opuntia Wcus-indica (L.) Miller Carica papaya L. Dianthus caryophyllus L. Beta vulgaris L. spp. cicla (L.) Kock Cistus sp. Arctotheca calendula (L.) Levyns Leucanthemum vulgare Lam. Sedum sp. Sempervivum sp. Brassica rapa L. Brassica rapa L. var. sylvestris (Lam.) Briggs Drosera capensis L. Erica holosericea Salisb. Vaccinium corymbosum L. Phalaris aquatica L. x Triticosecale sp. Ribes nigrum L. Ribes rubrum L. Acacia cyclops G.Don Cytisus proliferus L.f. Glycine max (L.) Merr. Medicago truncatula Gaertn. Trifolium subterraneum L. Vigna radiata (L.) R.Wilczek Linum ustatissimum L. Gossypium hirsutum L. Humulus lupulus L. Eucalyptus diversicolor F. Muell. Bougainvillea spectabilis Willd. Oxalis deppei Loddiges PassiXora edulis Sims Phytolacca octandra L. Limonium sinuatum (L.) Miller Muehlenbeckia c.f. cunninghamii (Meissner) F. Muell. Muehlenbeckia horrida Gross Persicaria odorata (Lour.) Soják Persicaria prostrata (R.Br.) Soják Polygonum salicifolium Willd. Portulaca grandiXora Hook. Banskia caleyi R.Br. Dryandra polycephala Benth. Macadamia integrifolia Maiden & Betcke Litchi chinensis Sonn. Zingiber oYcinale Roscoe

Kiwifruit Pig-face Amaranthus Cashew nut Mango Dill Celery Parsnip Pineapple Prickly pear Pawpaw Carnation Silverbeet Rock rose Capeweed Ox-eye daisy Stonecrop Houseleek Turnip Rapeseed Drosera Erica Blueberry Phalaris Triticale Blackcurrant Redcurrant Wattle (A) Tree lucerne Soybean Barrel medic Sub clover Mung bean Linseed Upland cotton Hops Karri (A) Bougainvillea Wood sorrel Passion fruit Pokeweed Statice Lignum (W.A.) (A) (A) Vietnamese Mint Creeping knotweed (A) Slender knotweed (A) Portulacca Banksia (A) Dryandra (A) Macadamia (A) Litchi Ginger

Apiaceae

Bromeliaceae Cactaceae Caricaceae Caryophyllaceae Chenopodiaceae Cistaceae Compositae Crassulaceae Cruciferae Droseraceae Ericaceae Gramineae Grossulariaceae Leguminosae

Linaceae Malvaceae Moraceae Myrtaceae Nyctaginaceae Oxalidaceae PassiXoraceae Phytolaccaceae Plumbaginaceae Polygonaceae

Portulacaeae Proteaceae

Sapindaceae Zingiberaceae

rest were dead. During the same period, 33 and 36% (respectively) of the eggs laid in the control E. australis plants had successfully developed to second or third instar. Because of the presence of a live larva in the second set of experiments, rhubarb was re-run in a longterm test.

3.5. Long-term, no-choice tests No eggs were laid in 8% of the cages whereas plants of the same species in other cages were suitable for oviposition (Table 4). The reason for this is unknown but presumably this is due to females remaining unmated 9–

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J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

Table 3 Families, species, and common names of plants in short- and long-term host-speciWcity tests of A. miniatum where feeding or egg-laying were observed Family

Common name, (A) Australian native

N

Begonia sp. Rubus idaeus L.

Begonia Raspberry

6 6

1.2 § 0.62 1.6 § 0.97

Antigonon leptopus Hook. & Arn. Homalocladium platycladum (F.Muell.) L.Bailey Muehlenbeckia adpressa (Labill.) Meissner Muehlenbeckia complexa (A.Cunn.) Meissner Muehlenbeckia cunninghamii (Meissner) F.Muell. Muehlenbeckia polybotrya Meissner Persicaria capitata (D.Don) Gross Persicaria hydropiper (L.) Spach Fagopyrum esculentum Moench cv Mancan Polygonum aviculare L. Polygonum bistorta L. Emex australis Steinh. Emex spinosa (L.) Campd. Rheum rhabarbarum L. cv Sydney Crimson Rumex acetosa L. Rumex acetosella L. Rumex alcockii K.H.Rech. Rumex brownii Campd. Rumex conglomeratus Murray Rumex crispus L. Rumex drummondii Meissner Rumex dumosus Meissner Rumex obtusifolius L. Rumex pulcher L. Rumex tenax K.H.Rech. Rumex vesicarius L.

Coral vine Tapeworm plant Climbing lignum (A)

6 6 6 6 6 6 6 6 12 6 5 91 6 16 6 6 6 6 6 6 5 6 6 5 6 6

2.6 § 0.99 1.5 § 0.49 2.1 § 0.25 3.2 § 1.64 13.4 § 1.51 1.2 § 0.74 6.9 § 2.30 0.3 § 0.33 2.2 § 0.97 0.8 § 0.59 0.8 § 0.56 20.9 § 1.36 39.4 § 4.25 10.7 § 1.68 7.0 § 2.71 6.0 § 2.07 54.3 § 6.50 20.7 § 5.04 26.2 § 3.38 32.4 § 1.93 27.4 § 2.07 10.9 § 1.98 43.6 § 5.16 32.9 § 8.76 37.6 § 7.39 4.2 § 0.81

Tribe Begoniaceae Rosaceae Polygonaceae Coccolobeae

Persicarieae Polygoneae

Rumiceae

No. of feed holesa (mean § SE)

Species

Lignum (A) (A) Knotweed (A) Water pepper (A) Buckwheat Wireweed Snakeroot Doublegee Lesser jack Rhubarb French sorrel Sorrel (A) Swamp dock (A) Clustered dock Curled dock (A) (A) Broad dock Fiddle dock (A) Rosy dock

No. of eggsa (mean § SE) 0.4 § 0.44b 0

1.2 § 0.80b 0.1 § 0.05b 0 0 0.1 § 0.13b 0 0.2 § 0.17b 0 0 0.3 § 0.29b 0.4 § 0.34b 8.6 § 0.79 31.3 § 5.35 5.0 § 1.10 0.4 § 0.42b 0.5 § 0.32b 13.5 § 3.14 5.6 § 1.58 4.6 § 0.90 4.3 § 1.19 3.1 § 0.83 2.8 § 1.01 c c

8.6 § 2.88 2.6 § 0.99

a

Feeding is expressed as the number of feeding holes in leaves observed per adult per fortnight. The number of eggs is per female per fortnight. Not one larvae (in any plant) was found to survive past second instar stage. c In this early experiment, the high number of eggs laid lead to the target being killed by the developing larvae. The number of eggs was also not counted on removal of the adult A. miniatum. b

16 days after set up. The overall insect mortality rate during this period was 8.8% in cages with no eggs versus 7.0% in cages with eggs. In Experiment 1, the level of attack on E. australis, R. obtusifolius L., and R. pulcher was higher than they could sustain with all individual plants dying before any A. miniatum larvae could complete development (Table 4). The exposure to egg-laying adults was reduced for all subsequent experiments resulting in only two E. australis being killed. Apion miniatum laid eggs into all Rumex spp. Survival of these eggs was however only favored in the species that were not Australian natives. An exception to this was R. vesicarius L. for which insect development appeared to be progressing normally with tunneling larvae within the stem and crown region until all plants (attacked or unattacked) completed seed production, and senesced as is normal for this short-lived annual species. The highest rate of survival within the Australian native species was 3.2% for R. dumosus Meissner (Table 4).

In the native Australian Rumex species (all Rumex subgenus Rumex), eggs were laid in the petioles where larval development commenced, but larvae were unable to penetrate the stems, except for the distal end when soft. In R. drummondii Meissner and R. tenax K.H.Rech., no adults developed although one dead pupa was found in the stem of R. drummondii. In contrast, 30% of eggs on the E. australis control became adults. In R. brownii Campd., one adult emerged alive but two others were trapped inside the stem, which appeared to be too ligniWed for them to escape. The success rate on R. brownii was only 2% of that in the corresponding E. australis controls. The crown of R. brownii was not attacked and the plants regrew. In R. dumosus and R. alcockii K.H.Rech., larvae that successfully completed development did so entirely within large petioles. In one case, R. alcockii, the larvae tunneled from the petiole into the center leaf vein where development was completed. In most cases, attacked petioles collapsed and the larvae died. Six percent of eggs on R. dumosus and 1% of eggs

J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

27

Table 4 Number of eggs (mean § SE) laid per plant during long-term no-choice experiments and the corresponding number of oVspring that emerged Experiment

Species

N (set up)

Eggs laid per plant

Adults emerged per plant

Plants with eggs

% survival (egg to adult)a

Plants killedb

1 1 1 2 2 2 3 3 3 4 4 4 5 5 6 6 6

E. australis R. obtusifolius R. pulcher E. australis R. crispus R. dumosus E. australis R. conglomeratus R. drummondii E. australis R. brownii R. vesicarius E. australis R. rhabarbarum E. australis R. alcockii R. tenax

6 6 5 5 6 6 6 6 5 6 6 6 6 6 6 6 6

c

0

4 6 4 5 5 5 6 6 5 6 6 5 5 6 6 6 5

c

4 5 2 1 0 0 0 0 0 0 0 0 1 0 0 0 0

c c

10.4 § 2.86 8.5 § 2.38 5.7 § 2.03 14.8 § 1.83 9.2 § 1.80 6.2 § 1.66 27.5 § 1.69 12.8 § 3.62 5.8 § 2.26 10.2 § 3.54 13.5 § 4.09 9.3 § 1.93 17.3 § 4.04 11.0 § 3.70

1.5 § 1.50 1.4 § 0.98 1.4 § 0.87 2.0 § 1.00 0.3 § 0.21 4.8 § 1.19 1.0 § 0.63 0 15.8 § 4.76 0.2 § 0.17 0 4.2 § 1.49 0.2 § 0.17 1.3 § 0.61 0.2 § 0.17 0

c c

17.3 § 11.85 27.7 § 14.43 6.3 § 4.84 29.6 § 6.90 8.8 § 5.33 0 58.3 § 16.70 0.9 § 0.88 0 43.7 § 15.32 0.8 § 0.76 16.4 § 7.57 0.9 § 0.93 0

a

% survival ignored plants in which no eggs were laid. Number of plants that died prematurely with no A. miniatum completing development. They were presumed to have been killed because they had extensive internal damage caused by the A. miniatum larvae. c In this early experiment, the number of eggs was not counted when the adult A. miniatum were removed. b

on R. alcockii developed into adults versus 17 and 16% in the controls (Table 4). All the native Australian Rumex tested regrew from the stems and/or crown that were untouched by the insects. A single adult developed in rhubarb, where the larva had tunneled from the petiole into the crown. This petiole had dried out and was dying and mucilage production was reduced. The tunnel in the crown was also without mucilage. Apart from damage from the A. miniatum larvae, there were no other apparent reasons for the petiole dying. The larvae within all other petioles of this plant, like those in all other rhubarb plants within this experiment, had died as described for the short-term tests. For the long-term experiment, less than 1% of eggs completed development to adult in rhubarb. The survival rate in the E. australis control was 40 times higher (Table 4).

3.6. Choice tests In the choice experiment, the number of adult feeding holes was similar between the Emex species, but was signiWcantly less on the two Rumex species (Table 5). Fewer eggs were laid on R. drummondii than on the other species in the same experiment, although this was not statistically diVerent. In the second experiment, the highest number of eggs was laid on R. brownii and signiWcantly less were laid on R. crispus. Some eggs failed to hatch on R. dumosus (2.1 § 1.32 unhatched eggs per plant), but unhatched eggs were not observed on the other species of plants. The size of larvae found at the time of dissection was similar in E. australis, E. spinosa, and R. crispus, with the majority of the larvae being live second or third instar (Table 6). In the native Australian Rumex spp., there was

Table 5 Mean § SE (minimum–maximum) number of feeding holes and eggs laid per female per fortnight in plants where adult A. miniatum had a choice for feeding and oviposition Experiment

Species

N

Number of feeding holesa,b

Total eggsa,b

7

E. australis E. spinosa R. drummondii R. dumosus E. australis R. alcockii R. brownii R. crispus R. tenax

6 6 6 6 6 6 6 6 6

5.0 § 1.79 (1–11) 4.1 § 1.05 (2–7) 4.7 § 0.97 (2–9) 5.0 § 1.24 (0–10) 6.4 § 1.21a (2–10) 1.2 § 0.34b (0–2) 3.2 § 1.00ab (0–7) 2.0 § 0.56b (1–4) 6.1 § 1.58a (1–11)

4.2 § 1.21 (0–7) 6.5 § 2.35 (1–14) 2.1 § 0.66 (0–5) 6.9 § 2.42 (0–15) 4.8 § 1.41a (2–11) 4.5 § 0.97a (2–8) 6.7 § 1.49a (3–13) 1.8 § 0.37b (0–3) 5.6 § 1.04a (2–9)

8

a ANOVA on number of feeding holes; Experiment 7: F3,15 D 0.1, ns; Experiment 8: F4,20 D 4.5, P < 0.01. ANOVA on total eggs laid; Experiment 7: F3,15 D 1.2, ns; Experiment 8: F4,20 D 4.5, P < 0.01. b Means marked with the same letter are not signiWcantly diVerent (LSD test at 5%).

28

J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

Table 6 Mean § SE of the number of larvae found alive and dead (and their stage of development) for each plant species within choice experiments Experiment Species

7

8

E. australis E. spinosa R. drummondii R. dumosus E. australis R. alcockii R. brownii R. crispus R. tenax

First instar

Second instar

Third instar

Total larvae

Dead

Live

Dead

Live

Dead

Live

Deadb

Missingc (dead?)

Livea,b,e

0 0 0.4 § 0.15 0.5 § 0.26 0.4 § 0.09 2.4 § 0.67 1.1 § 0.18 0.6 § 0.22 2.1 § 0.63

0.1 § 0.07 0.8 § 0.50 0.2 § 0.11 0.2 § 0.16 0.3 § 0.13 0.1 § 0.06 0.2 § 0.07 0.1 § 0.06 0.2 § 0.14

0.1 § 0.05 0.2 § 0.16 0.3 § 0.14 0.4 § 0.17 0.1 § 0.08 1.0 § 0.28 1.2 § 0.22 0.1 § 0.06 1.2 § 0.56

1.9 § 0.47 1.7 § 0.48 0.4 § 0.26 0.1 § 0.05 1.7 § 0.76 0.6 § 0.26 0.8 § 0.31 0.3 § 0.16 1.0 § 0.31

0 0.1 § 0.05 0 0 0 0.0 § 0.03 0.1 § 0.09 0.1 § 0.06 0

1.4 § 0.69 1.6 § 0.66 0 0.1 § 0.05 1.4 § 0.28 0.1 § 0.09 0.7 § 0.32 0.3 § 0.10 0.3 § 0.15

0.1 § 0.05 0.2 § 0.16 0.7 § 0.24 1.4 § 0.65 0.6 § 0.19 3.4 § 0.90 2.5 § 0.23 0.8 § 0.25 3.3 § 1.16

0.7 § 0.56 2.2 § 1.35 0.7 § 0.46 5.2 § 1.99 0.7 § 0.32 0.3 § 0.18 2.5 § 1.54 0.3 § 0.23 0.8 § 0.37

3.4 § 1.09a 4.1 § 1.19a 0.7 § 0.28b 0.3 § 0.16b 3.5 § 1.02a 0.8 § 0.39b 1.7 § 0.41ab 0.7 § 0.17b 1.5 § 0.50b

Live as % of total eggsa,d, e

78.8 § 9.93 73.1 § 10.80 39.4 § 15.16 23.8 § 19.17 72.8 § 5.16a 16.5 § 6.35c 27.8 § 5.25bc 40.6 § 3.93b 25.3 § 8.74bc

a ANOVA on total live larvae; Experiment 7: F3,15 D 10.4, P < 0.01; Experiment 8: F4,20 D 4.1, P < 0.05. ANOVA on the % surviving out of the total eggs laid; Experiment 7: F3,13 D 2.1, ns; Experiment 8: F4,20 D 12.8, P < 0.001. b Includes the larvae that were unable to be sized because they were damaged during dissection. c Insect were classed missing if tunnels were present but no insect found. d 1 E. australis and 1 R. dumosus plant excluded because no eggs found on them. e Means marked with the same letter are not signiWcantly diVerent (LSD test at 5%).

a relatively higher early mortality rate and proportionally fewer individuals present as live, late instars. The total number of live larvae per plant was consistently higher in the Emex spp. when compared to the Rumex spp. (signiWcantly higher for all species tested except R. brownii) with at least twice as many live larvae (on average) being present. Over 70% of the eggs initially laid into the Emex spp. were accounted for as live larvae at the time of dissection. Rumex crispus had the highest proportion of surviving progeny (40%) within the Rumex species. 3.7. Field observations of host speciWcity Emex spinosa is the only known host of A. miniatum in Israel. Polygonum equisetiforme Sibth. & Sm., Rumex pictus Forssk. (subgenus Acetosa), and Rumex bucephalophorus L. (subgenus Platypodium, the fourth subgenus in Rumex (Rechinger, 1984)) were examined near to and south of Tel Aviv and no evidence of A. miniatum was found. In addition, Rumex cyprius L. (subgenus Acetosa) was also examined near the Dead Sea in an area without E. spinosa. No Apion spp. were found. Rumex (Rumex) species, e.g., R. crispus, R. conglomeratus Murray, and R. pulcher, are known to occur in Israel (Zohary, 1966), but were not abundant and were not found with E. spinosa during Weld work.

4. Discussion The tests on 81 species from 35 families showed that A. miniatum completed normal development only on Emex species and Rumex (Rumex) species. The results indicate a very low level of development on rhubarb. Rhubarb is extensively grown in Europe without reported attack, which supports the conclusion that

there is a very low probability of this plant being attacked in Australia. The only other economic Polygonaceae crop, buckwheat (Fagopyrum esculentum), was neither attacked in the tests nor is it known to be a host. The literature record of blackcurrants (Ribes rubrum L.) being a host (Samedov, 1963 in Ter-Minassian, 1972) is questioned as both red and black currants (R. rubrum and Ribes nigrum L.) were unattacked in these tests. The observation that organisms feeding on Emex species also develop well on Rumex subgenus Rumex (e.g., R. conglomeratus, R. crispus, R. pulcher, and R. obtusifolius) has been made in speciWcity-testing programs on other Emex associated species. Examples are the weevils, Rhytirrhinus inaequalis (F.) (Coleoptera: Curculionidae) (Scott and Way, 1989a), Lixus cribricollis (Julien et al., 1982), the aphid, Brachycaudus rumexicolens (Patch) (Hemiptera: Aphididae) (Scott and Yeoh, 1998), and the pathogens Phomopsis emicis (Shivas et al., 1994) and Uromyces rumicis (Schum.) Wint. (Morris, 1982). Emex and Rumex species share a number of specialist herbivore species in the Mediterranean region such as Perapion violaceum (Kirby) (Coleoptera: Apionidae) and Pegomyia spp. (Diptera: Anthomyiidae) (Scott and Sagliocco, unpublished). While Emex is probably derived from Rumex or a common ancestor, the most closely related Rumex to Emex has not been identiWed. However, the results of speciWcity tests on a range of organisms and Weld observations indicate that there is a natural similarity, be it chemical or physical, between Emex species and the Rumex (Rumex) species. The last biological control agent to be released against Emex was Lixus cribricollis. At that time (1979), authorization was obtained in Australia for release against both Emex and Rumex. Although A. miniatum demonstrated its ability to attack and even kill Rumex spp. (of European origin) within our laboratory experiments, it was selected primarily for release in Australia against the more

J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

susceptible Emex species. Emex spinosa plants with larvae present were notable in the Weld in Israel due to their shortened internode length and stunted appearance compared to unattacked plants. In contrast, the biotype associated with Rumex in Europe is not abundant and from Weld observations in Europe, Scott (1985) considered it doubtful that it would cause signiWcant damage to Rumex species. When reviewing potential biological control agents for the control of R. obtusifolius and R. crispus in Switzerland, Grossrieder and Keary (2004) also doubted the level of damage would be suYcient to give eVective control. The diVerence in the impact of A. miniatum upon Rumex spp. versus E. spinosa maybe due to the distribution of A. miniatum larvae within the plants. Freese (1995) found that A. miniatum larvae were restricted to the basal area of the stems for samples of R. crispus from Germany. Higher stem positions were occupied by a similar insect, Perapion violaceum. Emex spinosa sampled in Israel did not contain other Apionidae species and the larvae were found mostly at the bottom of stems, but also in the crown and taproots. In our laboratory culture, larvae were found along the length of the stems as well as in the crown and taproots of both E. australis and E. spinosa. Taxonomically, the native Australian Rumex species are placed in separate subsections of Rumex (Rumex) from the introduced species R. conglomeratus, R. crispus, R. obtusifolius, and R. pulcher (Rechinger, 1984). The level of structural damage inXicted upon the native Australian Rumex species by A. miniatum larvae was less than observed upon either the introduced Rumex or Emex species. The stems, crowns and/or taproots of the introduced species were damaged whereas damage to the native Rumex species was largely restricted to the leaves and petioles; apparently due to some physical barrier or ligniWed tissue at the base of stems which prevented the larvae from penetrating the crown and root systems. Although the attacked petioles or leaves of the native Australian Rumex species often died, the plants usually quickly recovered with new leaves being produced. Three Australian Rumex species, R. stenoglottis K.H.Rech., R. bidens R.Br., and R. cystallinus Lange, were not available to the testing program. However, the testing program included Wve other native Australian Rumex species and these all proved to be unsuitable or at best, suboptimal hosts for A. miniatum. We considered the other species to not be threatened by A. miniatum for the following reasons: Rumex stenoglottis is a perennial species found in New South Wales and is morphologically very similar to R. dumosus, from which it was recently separated, and R. drummondii (Rechinger, 1984). We predict it would be subjected to the same level of damage by A. miniatum as R. dumosus, and R. drummondii. Rumex bidens is a semi-aquatic plant found in fresh water swamps in the cooler parts of Australia. The

29

plant’s rhizomes are at least temporarily submerged (Rechinger, 1984). These types of areas are not typically where Emex is a problem so it is unlikely that A. miniatum will ever be exposed to R. bidens. However, if exposure does occur, the plant is highly likely to be fully protected as the stems will be underwater during winter, the period of the year that A. miniatum is egg-laying. Rumex drummondii may be similarly protected since, like R. bidens, it is also found in winter-Xooded habitats (Scott and Yeoh, 1995). Rumex crystallinus is the only annual species among Australian Rumex. It is an ephemeral species found in temporarily Xooded depressions within arid/semiarid inland areas (Rechinger, 1984). It too is unlikely to coexist with Emex as in Australia, Emex is typically a weed of agricultural areas. Like all annual ephemeral arid species, R. crystallinus will respond quickly to any rain events. It is likely that the plant’s life cycle, like that of R. vesicarius, an exotic arid/semiarid annual, will be too short for A. miniatum to complete its development before the plant senesces. The uncertainty of annual rainfall events and the Wxed 1-year life cycle of A. miniatum would ensure that any established populations of the insect went to extinction in the Wrst year that rains were insuYcient for the plant’s germination. The Australian Quarantine Inspection Service (AQIS) and the Department of Environment and Heritage jointly regulate the import, testing, and release of biological control agents in Australia. In 1996, they approved the release of A. miniatum. Mass-rearing methods were developed and, in 1998, extensive releases began upon Australian Weld populations of E. australis. Most releases were in autumn and consisted of 60–2000 adults that had just reached sexual maturity before being liberated at each site (Yeoh et al., 2002). The areas from which A. miniatum were collected in Israel have annual rainfalls of 200–600 mm. Yeoh et al. (2002) found that for the release sites located within the Australian graingrowing region (255–510 mm annual rainfall), insects typically reproduced well during the plant’s growing season with an average 38-fold increase in population size occurring during the year of release. Based upon 28 sites within this region, pre-summer A. miniatum populations were estimated to average 16,300 individuals and the largest population thought to have 113,000 individuals. Despite these pre-summer populations and all eVorts to ensure that this agent ecoclimatically matched that of E. australis in Australia’s grain-growing region, no established population has been detected to date at any site of release. In all cases, the Weld populations have disappeared during the Wrst summer period. Determining the reasons as to why a potential biological agent fails allows researchers to more eYciently select or reject future agents. The simplest explanation of the Australian summer being too hot and dry for the insect to survive, as was also thought to be the reason for the fail-

30

J.K. Scott, P.B. Yeoh / Biological Control 33 (2005) 20–31

ure of P. antiquum based upon available knowledge at the time (Scott, 1992), seems unlikely for A. miniatum as they were collected from and around the Negev region. However, as an added control, some releases were also made in the cooler, wetter, coastal areas of Australia (>510 mm annual rainfall). In these locations, A. miniatum in general performed poorly in winter and failed to survive summer (Yeoh et al., 2002). Ants have been attributed to the failure of many biological control projects, but Yeoh et al. (1999) found, ants had no eVect on the reproductive output of A. miniatum reared within Weld cages. Yeoh et al. (2002) postulated that establishment was not occurring in Australia because the insects were dispersing too widely and therefore not Wnding mates however subsequent releases were made with multiple, clustered releases (2000 individuals at each of four sites spaced 1 km apart) without any improvement to establishment (Yeoh et al., 2004). By releasing A. miniatum, an insect from E. spinosa in Israel, upon E. australis, a plant from South Africa, we are attempting to create an evolutionary new exploiter– victim association. Given that A. miniatum is relatively host speciWc, it could be argued that E. australis is not a suitable host plant and that, under conditions of stress, the insect cannot survive due to inadequate resources. Hokkanen and Pimentel (1989) predict that such new associations actually favor the probability of an agent establishing and impacting upon a pest because the later has not been able to speciWcally develop antifeeding responses to the former. A similar situation appears to be the case for A. miniatum as insects reared on E. australis have higher levels of body fat and are able to withstand longer periods of starvation compared to those reared on E. spinosa (Yeoh and Scott, unpublished). The most plausible reason for the failure of A. miniatum to establish in Australia appears to be that although A. miniatum enters a reproductive diapause over summer, it needs to periodically replenish energy reserves by ingesting simple carbohydrates (Yeoh et al., 2004; Yeoh and Woodburn, 2003). The Australian grain-growing region is relatively devoid of summer Xowering trees when compared to Israel, where tamarisk (Tamarix spp.) trees with their profuse, nectar-laden Xowers are common. Yeoh et al. (2004) demonstrated that survival of A. miniatum during the summer could be increased from 1% to 92% by providing tamarisk plants in Xower. Despite some comparative sequencing techniques that now consider grouping together the Tamaricaceae (includes Tamarix spp.) and the Polygonaceae (includes Emex spp.) (Meimberg et al., 2000), A. miniatum did not feed upon the foliage of tamarisk. Alternative sources of simple carbohydrates (e.g., cane sugar or honey) were suitable alternatives to the tamarisk nectar (Yeoh et al., 2004). If the Azerbaijan biotype of A. miniatum behaves similarly to the Israel biotype, then Samedov (1963 in Ter-Minassian, 1972) observations of the insect “on” Ribes may also be related to sugar intake.

ConWrming the dependence of A. miniatum on Tamarix spp. or some substitute summer Xowering tree will be diYcult without additional releases within Australia or ecological studies within Israel. If true, the lack of potential Xowering trees in the areas of Australia where E. australis is the greatest problem diminishes the value of this insect as a biological control agent for this plant. It is however relatively easy to screen new potential agents for dependence on simple carbohydrates over the summer and this should be one of the Wrst steps for any similar potential agent in the future.

Acknowledgments The Western Panel of the Grains Research and Development Corporation and CSIRO Entomology funded this work. Agriculture Western Australia made available its quarantine facilities for the work. We especially thank the late Professor David Rosen for his advice and encouragement. We also thank his colleagues in Israel for their help. J.-L. Sagliocco helped with collections in Israel and T.L. Woodburn and T. Heard kindly read drafts of the manuscript.

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