Contrasting Responses of Southern House Spiders and Raccoons to Blister Beetle Prey

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CONTRASTING RESPONSES OF SOUTHERN HOUSE SPIDERS AND RACCOONS TO BLISTER BEETLE PREY

JAMES E. CARREL

Division of Biological Sciences University of Missouri-Columbia Columbia, Missouri 65211-7400

(Received May 28, 1998; accepted February 4, 1999)

Abstract—Female southern house spiders, Kukulcania hibernalis, readily consumed blister beetles, Lytta polita, regardless of cantharidin content when field-tested late in January and again three weeks later in February 1997. In contrast, free-ranging raccoons, Procyon lotor, initially ate many L. polita, particularly female beetles that contained only one third as much cantharidin as males, but when retested the raccoons ate only a few meloids. These results suggest that raccoons, unlike southern house spiders, quickly form an aversion to blister beetle prey, which is induced by cantharidin. Chemical analyses revealed that southern house spiders ingested 99% of the cantharidin contained within their prey.

Key Words—Cantharidin, chemical defense, Coleoptera, Meloidae, Araneae, Filistatidae, Mammalia, Procyonidae, feeding aversion.

INTRODUCTION

Blister beetles (Coleoptera, Meloidae) are notorious because they contain cantharidin, a noxious substance reputed to possess aphrodisiac properties in prepa- rations called Spanish fly (Keh, 1985; McCormick and Carrel, 1987). Can- tharidin is highly toxic to humans and other mammals, causing blistering of skin and similar damage to internal tissues when administered systemically (Till and Majmudar, 1981; Schmitz, 1989; Wang, 1989). Surprisingly little was known about the natural function of this compound until relatively recently, when it was demonstrated that the substance is a potent feeding deterrent to ants, cara- bid beetles, and some spiders (Carrel and Eisner, 1974; Smedley et al., 1996). Previously we raised the possibility that many avian and mammalian predators might possess oral sensitivity or behavioral aversion to cantharidin, but because 1295

0098-0331/99/0600-1295$16.00/0 C 1999 Plenum Publishing Corporation 1296 CARREL of its toxicity we refrained from performing the appropriate experiments to test our ideas (Eisner et al., 1990; Yosef et al., 1996). I here present data, based on experiments in which I offered live meloid beetles to free-ranging spiders and raccoons, showing that the spiders routinely ingest meloids whereas the raccoons evidently learn to reject meloids. To my knowledge, this is the first demonstration of an induced aversion to eating meloids in a vertebrate predator. I also show that the spiders consume virtually all of the cantharidin present within the meloid prey.

METHODS AND MATERIALS

Except for the chemical analyses, which were done at my Columbia laboratory, the experiments were carried out at night at the Archbold Biological Station, Highlands County, Florida, where the animals are nocturnally active. The meloid beetles, Lytta polita, are large, pine-pollen feeding insects (Car- rel et al., 1990) and were obtained at night by attracting them with two ultraviolet lights. One "black" light was positioned horizontally beside a conventional 1.2- m-long fluorescent light in a fixture above a white polyester sheet attached on the side of a 1.7-m-tall wooden shelf standing on the cement balcony along the back of the main station building. The other ultraviolet light was placed above a conventional bucket-style insect trap far away from the building, surrounded by typical Florida scrub (Abrahamson et al., 1984). Starting on the evening of January 27, 1997, for four nights in a row I checked the lights hourly between 6 and 11 PM to collect the blister beetles. Meloids (N = 190) were individually put in plastic vials and taken to the laboratory where they were sexed, weighed, and placed in screened cages isolated by sex. Care was taken to prevent reflex bleeding when manipulating the beetles. No beetle was used in more than one trial. Feeding Trails with Meloid Beetles. Meloids were fed to adult female southern house spiders, Kukulcania hibernalis (Araneae, Filistatidae) (N = 20), whose relatively permanent webs were located under the eaves of other station buildings, out of sight of the black lights. These spiders were selected because corpses of L. polita are sometimes visible in their webs in winter (Edwards, 1983; Carrel, unpublished). Each spider was presented with either a male or female meloid selected at random. A beetle was flipped into the web from a vial in which it was confined after removal from the cage. Ensuing events were monitored, and the fate of a beetle was recorded. Early season tests were performed at 8–10 PM on January 28–30. They were repeated in the same fashion with the same spiders on February 17-18. Intact carcasses of most eaten beetles, recovered after one to three days when spiders cut the prey from their web and dropped them to the ground, were cleaned of silk, dried for a week at room tem- RESPONSES TO BLISTER BEETLE PREY 1297 perature, weighed, and individually stored at -20°C in clean vials for chemical analysis. Meloids were also fed to a family of three raccoons, Procyon lotor (Mam- malia, Procyonidae), who routinely foraged between 8 and 11 PM at the main station building. Having seen the mother and her two offspring eat assorted insects attracted to the illuminated sheet, I decided to determine whether they would attack and eat blister beetles. On one night early in the season (January 29) I immobilized 24 male and 24 female beetles by chilling and then placed them at ~9 PM in a random alignment at 0.1-m intervals on the cement balcony beneath the light. I retreated to monitor the events from a distance of 15 m. I recorded whether the mother or her cubs visibly ate meloids. After the raccoons ambled off, I retrieved the remaining meloids and took them back to the laboratory to check on injury and the sex of surviving individuals. The tests using the same spiders and raccoons were repeated in a similar fashion three weeks later (February 17 or 18) to ascertain whether their predation behavior had changed. My hypothesis was that the spiders likely would continue to eat meloids, but the raccoons might exhibit a feeding aversion to meloids. Although I could not individually recognize the raccoons, a leading authority on Florida mammals assured me that I almost certainly employed the same raccoons in all of my tests. His reasoning was that it would be virtually impossible for another family unit of the same size to invade this location during my short-term study (J. N. Layne, Senior Research Biologist Emeritus, Archbold Biological Station, personal communication). Cantharidin Analyses. Cantharidin content of L. polita beetles was quan- titatively determined by capillary gas chromatography according to a published technique (Carrel et al., 1985). Two types of samples were analyzed: (1) whole, air-dried meloid bodies (15 individuals of each sex), and (2) remains of air-dried meloids after they had been eaten by southern house spiders (15 of each sex).

RESULTS

Predation on Meloids by Spiders The blister beetles proved very acceptable to southern house spiders (Table 1). In the two feeding trials, only four of 40 meloids given to spiders were not killed and eaten. The spiders showed equal preference to male and female beetles both on the first trial in January and when retested three weeks later in February. Statistical tests revealed no significant effect from the sex of the prey or the time of testing (chi square = 1.11, df= 1, P > 0.1). The attack strategy of female K. hibernalis was very distinctive. Spiders stealthily moved from their retreats and approached the beetles as they struggled in the cribellate web. After contacting the beetles, or silk near the entrapped 1298 CARREL

TABLE 1. MELOID BEETLES (Lytta polita) EATEN OR UNEATEN BY 20 SOUTHERN HOUSE SPIDERS (Kukulcania hibernalis)a

Eaten (N) Uneaten (N)

Trial Male Female Male Female

First 9 8 1 2 Second 10 9 0 1 aEach spider was given one beetle of the same sex in each trial. The intertrial interval was three weeks. insects, briefly with palps and the front legs (Figure 1A), spiders bit only the tips of the nearest legs or antennae with their small chelicers (Figure 1B), and retreated while their venom took effect. Within 80-300 sec of being bitten, the meloids were immobilized by the spiders' venom. Thereafter the spiders returned, swathed the beetles in silk, and then they began to feed at the head or thorax of the beetles (Figure 1C). None of the attacked meloids reflex bled and there was no visible cleansing of the mouthparts by spiders. Four beetles managed on their own to struggle free and fall from the webs before being attacked by spiders. This process was slow, ranging from 140 to 320 sec, primarily because the long tarsal claws of meloids repeatedly became entangled in fine loops of silk. Predation on Meloids by Raccoons. The results with raccoons were in sharp contrast to those with spiders. Of a total of 96 meloids presented in the two trials, only 32 were eaten by the three raccoons. In the first trial (January 29), the raccoons ate 29 living meloids (Table 2). However, in this initial trial they showed a distinct preference for female meloids over males when both were presented in equal numbers (chi square with Yates' correction = 12.5, df= 1, P < 0.001). When retested three weeks later in February, the raccoons ate only three beetles, all of which were females, and they rejected the remaining 45 animals regardless of sex. The decline in raccoon predation on meloids as a function of time was highly significant (chi square with Yates' correction = 36.8, df= 1, P< 0.001). In the first trial, all three raccoons were observed to inspect and sniff meloids on the balcony before eating them. Because it was early in the season and the evenings were cool, few large insects other than meloids were to be found at the lights on the balcony. In several instances after eating blister beetles, raccoons wiped their mouths and then shook their heads from side to side. In the second trial, all three raccoons ate many other large insects that had been attracted to the lights, but only the two cubs approached and contacted blister beetles. The mother raccoon climbed onto the wooden shelf, picked an uniden- tified katydid off the sheet with her forepaws, and put it in her mouth before RESPONSES TO BLISTER BEETLE PREY 1299

FIG. 1. Three consecutive stages in the attack of a southern house spider (Kukulcania hibernalis) on a meloid beetle (Lytta polita). (A) The spider uses its pedipalps and first pair of legs to gently contact the beetle struggling in its web. (B) The spider bites only the tarsus on the beetle's left hindleg and injects its paralytic venom before retreating from its prey. (C) Soon after immobilization is complete, the spider returns to feed in the cephalic region of the beetle. 1300 CARREL

TABLE 2. MELOID BEETLES (Lytta polita) EATEN OR UNEATEN BY FAMILY OF THREE RACCOONS (Procyon lotor).a

Eaten (N) Uneaten (N)

Trial Male Female Male Female

First 8 21 16 3 Second 0 3 24 21 aRaccoons were given 24 beetles of each sex in each trial. The intertrial interval was three weeks. she resumed ingesting crunchy scarab beetles and mole crickets. After the raccoons departed, I found two dead female meloids wet with raccoon saliva and 43 unharmed meloids remaining on the balcony. Cantharidin Analyses. Whole blister beetles contained much cantharidin (Table 3). Male meloids had three times as much cantharidin as females, even though females weighed nearly twice as much as males. Hence, the concentration of cantharidin in L. polita males (about 1.1% of dry weight) was about five times higher than that detected in females (about 0.2% of dry body mass). Remains of blister beetles eaten by southern house spiders weighed con- siderably less than uneaten beetles (Table 3). When cut into fragments in preparation for extraction of cantharidin residues, I noticed that each beetle's remains consisted of an integumental shell largely devoid of fat body, gut, muscle, and reproductive tissues. The spiders had assimilated nearly two thirds of the dry mass of meloid prey, including virtually all of the cantharidin in their bodies.

TABLE 3. MASS AND NET CANTHARIDIN CONTENT OF UNEATEN BLISTER BEETLES (Lytta polita AND REMAINS OF BLISTER BEETLES EATEN BY SPIDER (Kukulcania hibernalis)a

Cantharidin Dry mass (mg) content (ug)

Whole bodies Male (N = 15) 56.4 + 3.3 b 627 + 104 a Female (N = 15) 110.8 ± 11.1 a 204 ± 34 b Remains Male (N = 15) 35.4 ± 3.2 c 1.5 ± 0.6 c Female (N = 15) 68.7 ± 5.6 b 0.6 ± 0.2 c

aValues are given as mean ± SE. Values sharing the same letter in each column are not significantly different (P > 0.05, Tukey HSD test). RESPONSES TO BLISTER BEETLE PREY 1301

DISCUSSION

Like most predators, spiders exhibit a wide range of sensitivities to blister beetles as prey. For example, recent studies performed in central Florida show that the orb-weaver, Nephila clavipes, rejects L. polita and many other meloids, whereas a sympatric orb-weaver, Argiope florida, readily feeds on meloids (Smedley et al., 1996). Crystalline cantharidin is an effective feeding deterrent to N. clavipes; hence, the unacceptability of meloids to this spider can be attributed, at least in part, to cantharidin. Although I did not assess the deter- rency of pure cantharidin in my tests, the fact that southern house spiders from the same locality ingested several hundred micrograms of cantharidin along with the internal tissues of L. polita strongly suggests that this substance is not a potent antifeedant to them. Systemic absorption of cantharidin from meloid prey may be of mixed adaptive value to spiders. The most obvious risk to the spider is tissue damage caused by cantharidin-induced inhibition of protein dephosphorylation nec- essary for regulation of cellular activities (Eldridge and Casida, 1995; Liu et al., 1995), which could prove fatal, but it is highly likely that some spiders, frogs, and other predators cope with meloid prey by having cantharidin-resistant protein phosphatases in their cells. A secondary risk, as suggested by Smedley et al. (1996), is that cantharidiniphilic insects, such as pyrochroid beetles and cerato- pogonid flies, might be attracted to spiders and bite them in order to obtain this substance for their own use. On the other hand, dietary acquisition and systemic retention of cantharidin might protect spiders from some of their own enemies. For example, one wonders whether the wasp Allochares azureus (Hymenoptera: Pompilidae) would attack cantharidin-laden southern house spiders or if its par- asitoid larvae would be able to grow and develop as they normally do on this spider (Deyrup et al., 1988). Experiments designed to test these hypotheses are in progress. The strategy of biting the tips of appendages on insect prey in its web is an unusual, but highly effective mode of attack for a spider. Not only does the spider deliver its venom into the insect's circulating hemolymph, enabling the spread of paralysis throughout the body, but it also protects the spider from retaliatory bites of its prey. Our studies demonstrate that the southern house spider performs this behavior when given other large insects having long appendages, even prey known to be devoid of chemical defenses (J. E. Carrel, in preparation). Avoidance of L. polita by raccoons seems to be a discrimination based on direct experience with the beetles. Moreover, it is reasonable to attribute the raccoons' aversion to meloids to the presence of cantharidin in them, considering that male beetles having much cantharidin were avoided more often than female beetles, which were morphologically very similar but had relatively little cantharidin. The aversion was specific for meloids rather than a general malaise, for 1302 CARREL the raccoons ate scarab beetles and other insects attracted to the lights, much as one would expect them to do. In fact, when the tests are performed late in winter, raccoons typically are experiencing food scarcity, hunger, and weight loss because the natural supply of nutritious fruits has been depleted and they subsist on a meager diet of beetles and other insects (Johnson, 1970). The mechanism whereby omnivorous raccoons develop an aversion to meloids may be varied and complex. For example, after handling or chewing meloids, within hours a raccoon may develop cantharidin-induced blisters on the skin of the forepaw or mouth, resulting in activation of nociceptive neural pathways (Simone et al., 1993) and associative learning. Alternatively, a naive raccoon may quickly form a conditioned food aversion associated with delayed ill effects following ingestion of cantharidin-laden meloids, much as happens in many vertebrates after they have eaten toxic food (Brower and Fink, 1985; Frutos et al., 1997; Kyriazakis et al., 1997; Garcia and Riley, 1998). Of course social learning of one sort or another may also influence diet selection in raccoons, much as it does in rats and some other animals (Galef, 1988). Although few formal studies of learning in raccoons apparently have been performed (John- son, 1970, and references therein; Conover, 1989), all published results indicate that these mammals possess a remarkable ability to learn and remember, which would explain their adaptability as generalist predators.

Acknowledgments—This study was supported in part by a grant from the University of Mis- souri Research Board. I thank Margaret Janowski-Bell, Klaus Gerhardt, and Rebecca Sherry for assistance, and the staff of the Archbold Biological Station, Lake Placid, Florida, for providing research facilities.

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