J Insect Behav DOI 10.1007/s10905-014-9470-4
Prey Preference of Aphidoletes Aphidimyza on Acyrthosiphon Pisum: Effect of Prey Color and Size Farhad Farhoudi & Hossein Allahyari & Seyed Mohammad Tabadkani & Majid Gholizadeh
Revised: 22 September 2014 / Accepted: 24 September 2014 # Springer Science+Business Media New York 2014
Abstract Color polymorphism in insects as well as factors contributing to its occurrence and maintenance have been of a great interest. Pea aphid (A. pisum) exhibits a noteworthy color polymorphism which occurs as red and green. The preference of the predatory gall midge Aphidoletes aphidimyza (Rondani) for the two color morphs of pea aphid at two life stages (adult and second instar) was investigated. Red adults, red nymphs, green adults, and green nymphs were offered to the larvae of the predator in different sets. The larvae attacked red aphids more than green ones of a same size. But whenever adults were offered along with nymphs, the nymphs experienced significantly more attacks. Although visual cues could result in more predation on red aphids, our findings showed that the size of aphids was the more important factor. The results showed that previous findings, suggesting more predation on the red morph, are valid when the same size of aphids is used. The ovipositing females exhibited no differences in oviposition choice between stalks infested with either of two colors. Keywords Pea aphid . aphididae . predatory gall midge . cecidomyiidae . color polymorphism . life stage
Introduction Polymorphism among individuals of a particular species, especially in insects, is a widespread phenomenon that has been recognized for almost two centuries (Losey et al. 1997). Some aphid species represent a noteworthy color polymorphism. Color morphs may vary in life parameters, host range, defensive behavior, susceptibility to natural enemies, and some other features (Ahsaei et al. 2013; Braendle and Weisser 2001; Caillaud and Losey 2010; Lambers 1966; Losey et al. 1997; Markkula 1963; Tahmasebi et al. 2012; Tomanovic et al. 1996; Tsuchida et al. 2010; Weisser and F. Farhoudi (*) : H. Allahyari : S. M. Tabadkani : M. Gholizadeh Department of plant protection, Faculty of Agriculture Karaj, College of Horticulture and Plant Protection, Tehran, Iran e-mail:
[email protected]
J Insect Behav
Braendle 2001). Pea aphid, Acyrthosiphon pisum (Harris) (Hemiptera: Aphididae) is a good example of an obvious color polymorphism which occurs as red and green. Body color of pea aphid is genetically determined, with red being dominant over green (International Aphid Genomics Consortium 2010; Caillaud and Losey 2010). Different populations of this aphid have different proportions of color morphs (Balog and Schmitz 2013; Lowe and Taylor 1964). Various biotic and abiotic elements may influence the frequency of these color morphs in a population; some studies suggest that different patterns of parasitism and predation affect prey polymorphism (Balog and Schmitz 2013; Henter and Via 1995; Libbrecht et al. 2007; Losey et al. 1997; Tomanovic et al. 1996). For instance, it has been shown that the predatory ladybird Coccinella septempunctata L. (Coleoptera: Coccinellidae) attacks red aphids more frequently when offered with two color morphs of pea aphid (Losey et al. 1997). It has been suggested that the mechanism behind this preference is visual; red aphids which are more detectable on the green surface of a leaf are more easily found and preyed upon (Nakamuta 1984). The evidence for parasitic wasps is weak and contradictory. Some researchers have found heavier parasitism on red aphids (Henter and Via 1995), while other studies in field and laboratory conditions showed that green aphids are more susceptible to parasitism (Michaud and Mackauer 1994; Losey et al. 1997; Libbrecht et al. 2007). Also previous encounters, are suggested to alter preferences of a parasitoid wasp (Langley et al. 2006). Various factors affect the prey preference of natural enemies, which could in turn affect the frequency of color morphs. Differences in chemicals and odors of various species/morphs can cause differential attractiveness to their predators and parasitoids. Different defensive behaviors can also cause discrimination among different color morphs (Braendle and Weisser 2001; Losey et al. 1997). Lowe and Taylor (1964) reported that red pea aphid employed more dropping off the plant as a consequence of disturbance by natural enemies. Braendle and Weisser (2001) also reported more escape behavior by red aphids in response to artificial stimuli. In spite of various studies regarding prey preference of generalist natural enemies, there is no study on the effect of a specialist furtive predator on color morphs of a given aphid species. The predatory gall midge, Aphidoletes aphidimyza (Rondani) (Diptera: Cecidomyiidae), is a specialist predator of aphids used as an effective biological control agent in greenhouse for more than four decades (Harris 1973; Markkula 1963). The larvae, i.e. the predatory stage, crawl along a leaf and suck out the aphids’ body fluid, after biting and paralyzing them. Using olfactory and visual cues, neonate larvae can find close prey (~3 mm) and will die of starvation if they are more than 63 mm away from food (Lucas and Brodeur 2001; Wilbert 1973). Plant species, plant varieties, artificial substrates, and plants infested or not infested by aphids affect females’ ovipositing decision (Mansour 1975). Jandricic et al. (2013) reported the aphids’ patch density as the main factor affecting oviposition decision of A. aphidimyza under controlled conditions, but under more natural conditions, prey species also play an important role. The density-dependent predation is the main advantage of, A. aphidimyza over other aphidophages. Although A. aphidimyza can survive to adulthood on only few aphids (e.g. as few as seven peach aphids) (Uygun 1971), in high populated patches, these predatory larvae kill many more than they can consume (Harris 1973).
J Insect Behav
Locating prey is a complicated task in an environment filled with different plants and animal species. Equipped with specialized sensory nervous systems, predators use a variety of physical and/or chemical cues in order to find and identify target organisms. These cues may be useful for long or short range attraction to prey. In addition, discrimination among different types of prey may have various short-term or longterm effects on population features. In this study we investigated if the larvae and ovipositing females of A. aphidimyza express any preference for the red or green color morphs of pea aphid. Differential defensive behaviors in different life stages of a given aphid species have been also shown to affect prey preference (Chau and Mackauer 1997; Gerling et al. 1990; Losey and Denno 1998; Wyckhuys et al. 2008). Aphids possess a large repertoire of defensive tactics including kicking with rear legs, walking away, dropping off plants, and producing cornicle droplets (Losey and Denno 1998; Nelson and Rosenheim 2005; Roitberg and Myers 1979). Here, we also studied the effect of two life stages of pea aphid, reflected in its body size, on the preference of the larvae of A. aphidimyza. Results of this study will help us understand the extent of the role of another member of the aphid-natural enemy guild in population dynamics of color morphs. Moreover, we need to further understand the prey-preference decisions of a commercially available predator for which prey preference has not been fully characterized.
Materials and Methods Plants Broad bean plants (Vicia fabae), on which aphids were reared and experiments carried out, were planted within pots containing sawdust (we used sawdust as potting media in order to avoid soil-born diseases). Plants were maintained under greenhouse conditions (24±1 °C, 50–55 R.H. and 16:8 L:D photoperiod) at the Department of Plant Protection, College of Agriculture, University of Tehran.. Aphid Colonies Pea aphids (A. pisum) were collected initially from the experimental alfalfa farms of University of Tehran, in Karaj, Iran. Red and green morphs of the aphid were reared on broad bean plants for several generations, in separated growth chambers (24±1 °C, 50–55 R.H. and 16:8 L:D photoperiod). These colonies were refreshed every year by introducing new collected aphids. In order to avoid producing winged morphs in the colonies, several new broad bean plants were added twice a week and old plants were discarded. Predator Colony Aphidoletes aphidimyza were collected in larval stage on cotton plants from Bojnourd (in northeast Iran). By the time that adult midges emerged, they were fed by a solution of sucrose (5 %). 24 h after adults’ emergence, Aphis fabae on broad bean plants were offered to them for oviposition. After egg hatching, larvae were reared on A. fabae. Black bean aphids were used because of the ease of rearing and maintenance. All these procedures were carried out under controlled conditions (24 ± 1 °C, 50–55 R.H. and 16:8 L:D photoperiod).
J Insect Behav
Preference Assay of Larvae The effect of prey color morph and body size on the preference of A. aphidimyza was assessed in experimental arenas in the laboratory. Experimental arenas consisted of single leaves of broad bean of similar area (approximately 10 cm2) resting horizontally on agar gel in petri dishes (9 cm diameter). In a homogenous distribution, two types of aphids differing in color and/or age stage were settled on each leaf. Beforehand, observations on large populations of both morphs showed that having inserted the stylet into the leaf tissue and started feeding, no aphid left the feeding site unless it was bothered. Therefore, in order for the aphids to stay immobile, they were left undisturbed for at least 1 h before the experiment started. Then a single larva of A. aphidimyza was released on the midpoint of the main leaf vein. Consequently, based on pre-test, eight individuals of each type of aphids (total=16) were offered to the predator in the following combinations: green adults vs. red adults, green adults vs. green nymphs, red adults vs. red nymphs, green adults vs. red nymphs, red adults vs. green nymphs, and red nymphs vs. green nymphs. In another experiment, the larva of A. aphidimyza was allowed to choose among all four types of pea aphid, each consisting of five individuals of red adults, green adults, red nymphs, and green nymphs (total=20). Second instars of pea aphid were used as nymphs. Eaten aphids were not removed or replaced during the experiments. All of the experiments were carried out in 15 replicates using 2-day old larvae of A. aphidimyza. The larvae were allowed to feed on the aphids in controlled conditions (25±1 °C and 16:8, L:D). Given that the larvae were allowed to predate for 24 h, using older larvae could result in pupation. Mean duration of larval stage at the given temperature was 3.27 days or 78.48 h (unpublished data). After 24 h, the number of surviving aphids was recorded. Aphids which could not extract their stylet out of the leaf and escape after being nudged by a fine brush were considered consumed, regardless of being eaten completely or not. Attacked aphids also had forward antennae, whereas the antennae of non-attacked aphids are backward (personal observation). This feature also helped us to find if the dead aphids were just results of midges’ predation or aphids’ autogenic mortality or/and handling mortality. Furthermore, based on the result of another experiment assessing life history of these two morphs, they showed no difference in terms of autogenic mortality in the mentioned time length, so no control (experiments without predator) was set. Prey preference of predator was calculated based on Manly’s preference index (Manly 1974) from the number of aphids consumed: log pi βi ¼ X m log p j j¼1
where βi=Manly’s β for prey class i; pi =proportion of prey i remaining at the end of the experiment relative to the original input (i=1, 2, 3, 4, …, m); pj = proportion of all prey species together remaining at the end of the experiment relative to the original input (j=1, 2, 3, 4, …, m); and m=number of prey classes, Manly’s β can vary between zero and unity. In a two-prey combination (m=2), the value 0.5 represents no preference, values larger than 0.5 indicate a preference for prey A and smaller values indicate a preference for prey B. This
J Insect Behav
method takes into account the prey density depletion by predation during experiments (Roger et al. 2001; Sherratt and Harvey 1993). For the experiments, where two types of aphids differing in color and/or age stage were tested in terms of causing prey preference, calculated indexes were subjected to a paired t-test. In the four-choice assay, the difference of calculated indexes of each type were compared to the expected value; 0.25 using t-test. Egg-Laying Site Preference of A. Aphidimyza Twenty five pairs of broad bean stalks, with two broad leaves (almost same age and area) and almost 15 cm height, were used as feeding sites of the aphids. Aphids were reared on same plants prior to the experiments. Broad beans were planted within plastic pots containing sawdust. Fifty newly emerged adults of each color were released on the leaves of each stalk, separately. Every two stalks, each of which were infested with one color, were settled within separate transparent glass boxes (1×0.5 m) containing eight ovipositing 24-h-old females of A. aphidimyza (gravid A. aphidimyza have reddish abdomen). Therefore, eight females in each box had a choice of two color/stalks to oviposit on. There was 40 cm distance between the two aphid-infested stalks. After 12 h, the number of eggs on each stalk was counted and recorded.
Results When adults of red and green pea aphid were exposed to the predator, larva of A. aphidimyza preferred red adults of pea aphid significantly more than green adults (t=3.7, d.f=14, P =0.002) (Fig. 1). Red nymphs were also preferred by the larva when they were presented to the predator together with the green nymphs (t=2.02, d.f=14, P=0.045) (Fig. 1). When green adults and red nymphs were offered, more preference was observed for the red nymphs (t=3.998, d.f=14, P= 0.001) (Fig. 1). Also when red adults along with green nymphs were exposed to the predator larvae, green nymphs underwent more attacks (t=5.1, d.f=14, P
