Oecologia (2001) 129:571–576 DOI 10.1007/s004420100763
Jürg Friedli · Sven Bacher
Mutualistic interaction between a weevil and a rust fungus, two parasites of the weed Cirsium arvense Received: 19 March 2001 / Accepted: 18 June 2001 / Published online: 2 August 2001 © Springer-Verlag 2001
Abstract We present a mutualism between a stem-boring weevil, Apion onopordi Kirby (Coleoptera: Apionidae), and a rust fungus, Puccinia punctiformis (Str.) Röhl. (Uredinales), both parasites of the creeping thistle, Cirsium arvense (L.) Scop. (Asteraceae). Females, but not males, of A. onopordi induced systemic rust infections of thistle shoots in the season after they were attacked by the weevil, indicating that insect oviposition is a crucial stage in pathogen transmission. Adult weevils emerged from systemically infected thistle shoots were heavier than weevils from healthy C. arvense shoots. Heavier females had a higher fecundity and laid larger eggs. The weevil preferred to deposit eggs in systemically rust-infected over healthy thistle shoots, which seemed to be a sub-optimal host. This is to our knowledge the first report of a mutualistic interaction between an herbivorous insect and a biotrophic plant pathogen. The mechanism responsible for the advantage of rust-infected shoots for A. onopordi causes a different outcome in other thistle herbivores, and therefore can not be explained by a general enhancement of nutritional quality in rust-infected tissue. This mutualism likely has evolved from a competitive relationship. Unlike other thistle herbivores A. onopordi seems to be better suited as mutualist for P. punctiformis because of its small impact on the host plant and its feeding niche on plant parts not directly associated with pathogen reproduction. Keywords Apion onopordi · Herbivory · Multitrophic interactions · Phytopathogen · Puccinia punctiformis
Introduction Obligate biotrophic plant pathogens depend on living plant tissue during all growth stages of their life cycle. J. Friedli · S. Bacher (✉) Zoologisches Institut, Universität Bern, 3012 Bern, Switzerland e-mail:
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Other parasites of their host plants feed on the same resource and should therefore have a competitive (–,–) relationship with the pathogen (e.g. Karban et al. 1987). The interaction may approach neutrality (0,–) if the impact of the competitor on the plant tissues infected by the pathogen is small. Most herbivores do not kill the plant individuals on which they feed. Therefore, herbivores are often considered as parasites rather than predators, with a negative or neutral impact on the performance of their host plants. The extent of herbivore impact on plant performance is mainly determined by their intensity of attack (Crawley 1997). In the case of small herbivores such as insects, attack intensity often depends on herbivore density (e.g. Bacher and Schwab 2000). It follows that herbivores that do not kill their host plant should have a negative impact on biotrophic plant pathogens when feeding on infected plants. Therefore, plant pathogens should avoid herbivory on infected tissues, for example by lowering nutritional quality of plants or single organs or producing repellent or toxic substances. In a recent review, Hatcher (1995) showed that insect herbivores are indeed negatively affected by feeding on host plants infected by biotrophic plant pathogens. However, this pattern seems to be valid only for strict herbivores and obligate biotrophic pathogens; omnivores and herbivores feeding on necrotrophic fungi may well profit (Hatcher 1995). It is well known that herbivorous insects serve as vectors for plant pathogens (Agrios 1980). If the indirect benefits gained by the pathogen exceed the direct costs due to herbivory, such relationships are advantageous for the pathogen (+,–). To our knowledge, however, no example is known of a herbivore receiving a reciprocal benefit for distributing a biotrophic plant pathogen. In this paper we present a mutualism between a stemboring weevil, Apion onopordi Kirby (Coleoptera: Apionidae), and a rust fungus, Puccinia punctiformis (Str.) Röhl. (Uredinales), both parasites of the creeping thistle, Cirsium arvense (L.) Scop. (Asteraceae). We discuss likely conditions under which such a relationship may evolve.
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Materials and methods Biology of the species involved The creeping thistle, C. arvense, is a deep-rooted herbaceous perennial that reproduces vegetatively as well as from seeds. The species produces creeping roots at depths from 10 to more than 30 cm below ground, on which buds develop to new aerial shoots in spring. Shoots die back to the root system in winter (Donald 1994). Creeping thistle is considered as one of the world's worst weeds (Holm et al. 1977) and the third most important weed in Europe (Schröder et al. 1993). A. onopordi is a small oligophagous apionid weevil that lives on several genera in the sub-tribe Carduinae (Zwölfer 1965; Freese 1995). In spring females drill out egg chambers in stems of C. arvense close to the soil surface, and lay a single egg inside each chamber. Larvae mine above- and below-ground and pupate in the stem. In autumn, adults emerge from the plant and, after a short feeding period, overwinter in the leaf litter (Scherf 1964; Wanat 1995). The autoecious rust fungus P. punctiformis infects exclusively C. arvense. As is common in rust fungi, it has a complex life cycle which is still not fully understood. In spring, thistle shoots systemically infected by the rust grow from the overwintering root system of C. arvense. On leaves and stem form yellow-orange pycnia, pustules that produce nectar and fragrance volatiles which presumably attract insects for cross-fertilization of the monokaryotic mycelia (Buller 1950). After fertilization aroma production ceases, and shoots develop sori which release short-lived dikaryotic urediniospores. Systemically infected shoots usually die before flowering (Watson and Keogh 1980). Urediniospores spread to healthy thistle shoots and cause localized, slow-growing infections, which again produce urediniospores. Later in the season, local infection by urediniospores leads to the formation of sori which release teliospores. Teliospores survive the winter and are assumed to be the stage that induces systemic infections in springemerging shoots via infection of root buds (Van den Ende et al. 1987; Frantzen 1994). In the laboratory, root buds of C. arvense infected by teliospores indeed grew systemically infected shoots (Van den Ende et al. 1987; French and Lightfield 1990). However, attempts to artificially induce systemic infections in naturally growing plants by application of teliospores have so far failed (Frantzen and Scheepens 1993). Only recently it was discovered that systemic P. punctiformis infections can be induced in creeping thistle clones by A. onopordi in the year after thistles have been attacked by the weevil (Friedli and Bacher 2001). Transmission of P. punctiformis by A. onopordi Adults of A. onopordi were collected in the Swiss Valais, in spring 1999. Weevils were kept singly and were allowed contact with a leaf containing urediniospores of P. punctiformis. Roots of C. arvense were collected from a rust-free population near Bern. Root pieces of 10–13 cm length were planted at a depth of 10–12 cm in the center of clay pots (3 l volume). To determine the effectiveness of induction of systemic rust infection by weevils compared with spores alone, we randomly assigned potted C. arvense plants to one of four treatments. On 20 pots, single females of A. onopordi were confined for 72 h to the largest C. arvense shoot in each pot by means of a transparent plastic tube (15 cm diameter, height depending on the shoot length) sealed on top with gauze mesh. On 11 pots each, (1) male instead of female A. onopordi were used as described above, (2) urediniospores of P. punctiformis were applied with a brush to the largest shoot or (3) no treatment was applied; these served as controls. In spring 2000, the pots were checked for systemically infected shoots of C. arvense after a growing period of 5 weeks in early May. Differences in the incidence of systemic infections between treatments were analysed with the G-test (Sokal and Rohlf 1995). Throughout the text means are presented ±SD.
Weight of newly emerged A. onopordi To determine differences in the development of A. onopordi in healthy and rust-infected shoots, 68 healthy C. arvense shoots (including the below-ground shoot parts) and 43 shoots systemically infected with P. punctiformis were collected in late June from a site in the Swiss Valais in 1999. The lower shoot parts, where the pupal cases of A. onopordi are located, were kept singly in plastic boxes (16×12×6 cm) on moist filter paper to prevent desiccation. The emerged A. onopordi adults were weighed (Mettler Toledo AG 204; accuracy ±0.1 mg). The same procedure was repeated in 2000, with 580 healthy thistle shoots and 276 shoots systemically infected with P. punctiformis collected in early June at the same site in the Swiss Valais. In addition, 258 healthy C. arvense shoots and 15 shoots of C. vulgare were collected from a site in the German Rhine Valley, and were treated as described above. Weight differences of newly emerged A. onopordi adults were established using the Fisher Pitman randomization test for small sample sizes (1999 collections) or the Mann-Whitney U-test for larger samples (2000 collections; Zar 1996). Offspring production To evaluate the effect of adult weight on egg laying rate, egg size and hatching success, 23 field-collected females of A. onopordi were weighted and kept singly for 12 days at 15°C and a light/dark regime of 12/12 h in transparent plastic tubes (5 cm in diameter, height 10 cm) on healthy C. arvense leaves. Leaves were exchanged daily and afterwards immediately dissected for eggs. Egg length and width were measured under a stereo microscope. Eggs closely resembled ellipsoids. Therefore, we calculated the egg volume ve using the formula where l is length and w is width of the egg. Eggs were transferred onto moist filter paper in petri dishes and incubated at 15°C and 12/12 h light/dark regime. Larval emergence was checked daily. Eggs were assigned to one of three size classes, 0.38 mm3, and the hatching rate was determined for each size class. The relationship between adult size and egg laying rate and egg size was established by regression analysis (Zar 1996). Differences in hatching success were analysed with the chi-square test (Zar 1996). Oviposition preference The oviposition preference of A. onopordi was determined in dual choice tests. Female A. onopordi were field-collected at a site with healthy and rust-infected thistles. Weevils were fed healthy thistles until the beginning of the experiment. For the experiment, females were kept for 72 h in transparent plastic tubes (5 cm in diameter, height 10 cm) on two C. arvense shoot cuttings (test and control) of about 7 cm length, each harboring one leaf. A healthy shoot cutting served as control in all experiments and was tested simultaneously against one of the following test treatments: (1) a C. arvense shoot cutting systemically infected with P. punctiformis harboring pycnias with the typical flower-like aroma, (2) a C. arvense shoot cutting systemically infected with P. punctiformis that lost its fragrance but instead harbored urediniospores and (3) a healthy C. arvense shoot cutting on which urediniospores were brushed on. Females were randomly assigned to one treatment. After 72 h, shoot cuttings were dissected and the number of shoots with feeding holes and the number of eggs laid in each shoot were recorded. Results were analysed with the chi-square test or Wilcoxon signed-rank test, respectively. In order to compare the effect of rust infection on egg-laying behavior of A. onopordi with another stem-borer, we also investigated the oviposition preference of the weevil Lixus algirus (L.) (Coleoptera, Curculionidae). In spring 1998, 44 shoots of C. arvense systemically infected with P. punctiformis and 52 healthy shoots from a site in the Valais were dissected under a stereo microscope and searched for eggs and early instar larvae of L. algirus. Data were analysed with the chi-square test.
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Results Transmission of P. punctiformis by A. onopordi In autumn 1999, local infections of P. punctiformis were observed on creeping thistle in all treatments except the control, indicating that the urediniospores used in the experiment were viable. In spring 2000, C. arvense shoots developed in all pots. Systemically infected shoots emerged in 40% of the pots where female A. onopordi were confined to thistles, while in the other treatments only healthy thistle shoots were found (Table 1; G=18.1, df=3, P0.30
