The Role of Antennae in Mate Recognition in Phoracantha semipunctata (Coleoptera: Cerambycidae)

Journal of Insect Behavior, Vol. 18, No. 2, March 2005 ( C© 2005)DOI: 10.1007/s10905-005-0478-7

The Role of Antennae in Mate Recognitionin Phoracantha semipunctata (Coleoptera:Cerambycidae)

Orlando Lopes,1 Paulo C. Marques,1,2 and Jorge Araujo1

Accepted August 31, 2004; revised September 30, 2004

The mating behavior of the eucalyptus longhorned borer Phoracanthasemipunctata was studied to understand its mate recognition system. Bioas-says were conducted to determine the existence of a cuticular chemical on itsbody surface and how individuals perceived it. Males walked oriented to andattempted copulation with live conspecifics only upon antennal contact withtheir bodies. They showed similar responses to intact dead females and males,but failed to respond to washed bodies. Dummies carrying male extracts weremore likely to elicit copulation attempts than control dummies. This consti-tutes behavioral evidence that unidentified chemical(s) on the body surfaceplay a major role in mate recognition, and can only be perceived after an-tennal contact. Sensilla trichodea on the antennal flagellum are candidatesfor this contact chemoreception. They are distributed throughout the entireflagellum, especially along its margins and at the tip of the distal flagellom-ere, and share structural features with contact chemosensory sensilla of otherinsects.

KEY WORDS: mate recognition; bioassay; antennal sensilla; mechano-contact chemorecep-tors; eucalyptus longhorned borer; Phoracantha semipunctata.

1Universidade de Evora, Unidade Ecologia Quımica, Centro Ecologia e Ambiente, RuaRomao Ramalho 59, 7000-671 Evora, Portugal.

2Correspondence should be addressed at Universidade de Evora, Departamento de Biologia,Laboratorio Biologia, Colegio Luis Verney, Rua Romao Ramalho 59, 7000-671 Evora,Portugal. E-mail: [email protected].

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0892-7553/05/0300-0243/0 C© 2005 Springer Science+Business Media, Inc.

244 Lopes, Marques, and Araujo

INTRODUCTION

In cerambycids, mate location can be achieved by a combination of amale long-range pheromone and a female short-range pheromone (e.g.,Xylotrechus pyrrhoderus) (Iwabuchi, 1982, 1985), a male short-rangepheromone (e.g. Monochamus alternatus) (Kim et al., 1992) or a femaleshort-range pheromone (e.g. Paraglenea fortunei) (Wang et al., 1990),(Semanotus japonicus) (Kim et al., 1993).

The eucalyptus longhorned borer Phoracantha semipunctata(Coleoptera: Cerambycidae) is a species native to Australia and a se-rious pest on eucalyptus plantations in Portugal since its accidentalintroduction. Evidence gathered to the present day show that the sexesrely mainly on the host plant odors to meet (Barata, 1997; Hanks, 1999), astrategy called the stressed host strategy (Hanks, 1999).

Although no evidence has yet fully proved nor dismissed the existenceof a short-range pheromone in this species, the males unequivocally dependon antennal contact in order to recognize females (Marques, 1992; Hankset al., 1996). It is plausible that they respond to a contact sex pheromone onthe cuticule of females in a way similar to that described for other ceram-bycids (Akutsu and Kuboki, 1983a, b; Fukaya and Honda, 1992; Kim et al.,1992, 1993; Wang, 1998; Fukaya et al., 1996, 1999; Akino et al., 2001; Ginzelet al., 2003; Ginzel and Hanks, 2003). This hypothesis, which had never beeninvestigated before, required that behavioral and chemical evidence of itsexistence was made available, and that a contact chemoreception systemallowing the perception of this putative contact pheromone through anten-nal contact be present in the antennae of this species. In fact, five types ofsensilla had been known to exist on the antennal flagella of P. semipunc-tata: chaetica, trichodea, grooved, and basiconica I and II (Lopes, 2000), andbased on the peripheral and/or distal location, size and external appearanceof the sensilla trichodea. Lopes (2000) had considered them as likely con-tact chemoreceptors on the antennae of this species. Ultrastructural dataon their functional morphology, which would allow us to infer on their pos-sible physiological function was however still missing and thus needed to beinvestigated as well.

In this study we set two objectives: (1) Observe the species matingbehavior, particularly the male’s mating sequence, and identify the male’smost relevant behaviors associated with mate recognition; test how likelythe stimuli provided by intact dead and dead washed conspecifics, as wellas crude extracts of dead conspecific bodies are in eliciting the exhibitionof these male behaviors; (2) Investigate the fine structure of the antennalsensilla trichodea in order to obtain insight into their functional morphol-ogy, which might indicate a possible contact chemoreceptive function. We

Role of Antennae in Mate Recognition in Phoracantha semipunctata 245

provide evidence that mate recognition is mediated by contact chemore-ception of a yet unidentified contact pheromone, and that the fine structureof the antennal sensilla trichodea is consistent with a contact chemosensoryfunction.

MATERIALS AND METHODS

Insect Source

Specimens of P. semipunctata used for electron microscopy were ob-tained as pupae from larvae infested eucalyptus logs collected in planta-tions at Pedra do Rato (Crato), in southern Portugal. Larvae were rearedon an artificial diet in individual Petri dishes in a controlled environment(25 ± 3◦C; 70 ± 10% RH). Adults used in behavioral experiments were feda honey solution (7 ml honey/100 ml water) since day one of emergence,following Marques’ procedure (1992), and held in a controlled environment(26◦ ± 1◦C, 60–70% RH), under a 16 white light: 8 red light photoperioduntil needed.

Behavioral Experiments

Dyad Observations

Dyads of mature individuals were observed in light floor acrylic arenas(200 mm diam. × 100 mm high), 2 h after the beginning of the red lightperiod, at 26◦ ± 1◦C, 60–70% RH, and recorded using a 8-mm monochro-matic camera, specially adapted for dim light, with a video Kama 0.5X,SWK-21-ccd macro lens (49–52 mm). Dyads were either (a) male–femalepairs (N = 16), or (b) same sex pairs (two females, N = 16, or two males,N = 16). Observations were focused on the displacement of individuals inthe arena prior to encounters and on their subsequent behaviors after that.On such encounters we observed whether the individuals approached eachother, and how they did it, or moved away from one another. In male–maleor male–female dyads we followed the male’s mating sequential behavior,as one male in a dyad attempted to hold, mount and copulate with the otherindividual.

Bioassays

Mate recognition was further studied by testing two groups of males,under the same controlled environmental conditions, in two experimental


sets: Experiment A, and B. We tested the null hypothesis that all treat-ments elicit equal proportions of dashing and genital touch by males. Inboth experiments we used a randomized block experiment design contain-ing values of a dichotomous variable: ‘behavior exhibited,’ or ‘behaviornon-exhibited’; whenever the behavior was exhibited we scored it as “1”;whenever it was not exhibited we scored it as “0.” Repeated observationswere made on the same individuals, each individual being a block. Datafrom both experiments (A and B) was analyzed with Cochran’s Q test(Zar, 1984), and a posteriori paired comparisons were performed using theMarascuilo and McSweeney’s (1967) procedure for data subjected to theCochran Q test.

Experiment A. Individual males were randomly assigned to each of thefollowing four treatments: (1) an intact freshly dead female, (2) an intactfreshly dead male, (3) a dead washed female body, and (4) a dead washedmale body. Freshly dead, intact bodies were obtained by keeping males andfemales separately in containers at −18◦ to −45◦C for 4–5 h. Dead washedbodies were obtained by washing separately male and female dead bodiesin 400 ml of chloroform in a Soxhlet apparatus for 12 h, a method similarto that described by Fukaya and Honda (1992).

Experiment B. Males were presented with a 25-mm length solid glassrod dummy covered with a male extract, a female extract, or an equalamount of solvent. Individual males were randomly assigned to: (1) adummy with 3-FE extract (three-female equivalent concentration extract),(2) a dummy with 5-FE extract, (3) a dummy with 3-ME extract (3-equivalent male concentration extract), (4) a dummy with 5-ME extract,and (5) a control (a similar 25-mm length solid glass rod dummy impreg-nated with chloroform). Extracts were obtained dipping five individuals perml of chloroform for 24 h at room temperature, condensed under nitrogento a total volume of 800 µl, and than kept at –18◦ to −45◦C until needed.Extracts were applied on the dummy using a micropipette after condensingthe extract to a 20 µl final volume.

Ultrastructure Analysis

Scanning Electron Microscopy

Adults were dehydrated in ethanol series 70, 80, 95, 100 and twoacetone baths. The antennae were removed from the head, and thenair-dried, gold coated with a sputter coater (SCD 040 Balzer) and ob-served using a scanning electron microscope (JEOL, J.S.M. 6400 F) at10 kV.


Transmission Electron Microscopy

Antennae from CO2 anesthetized males and females of P. semipunc-tata were removed and directly immersed in 2.5% glutaraldehyde fixativein 0.1 M sodium cacodilate buffer (pH 7.2). Antennomeres were then sep-arated and left in the fixative at room temperature overnight. After rinsingin buffer, the samples were post-fixed in 1% osmium tetroxide in the samebuffer for 2 h, rinsed in buffer, dehydrated in a graded ethanol series (70,90, and 100%), transferred to propylene oxide and lastly embedded in Spurrresin. Ultrathin sections were taken using a diamond knife on a Leica SuperNova ultramicrotome, then mounted on formvar coated grids, contrastedwith uranyl acetate and lead citrate and examined in a Philips CM 10 trans-mission electron microscope. The sensillar terminology adopted here wasthat of Dyer and Seabrook (1975), which is based on the size, shape, poros-ity and articulation of the socket of the cuticular processus of the sensillum.

RESULTS

Behavioral Experiments

Dyad Observations

In all three types of dyads we observed the males and females roam-ing about the arena, walking with their antennae oriented forward, slightlyarched and touching the ground, head down, continuously touching theground with their palpi. On no occasion were the individuals in any of thedyad types observed approaching each other in oriented walk, from a dis-tance, prior to antennal contact.

Male–Female Pairs

Females were very active, spending more than two thirds of their timeroaming and interacting with males. Males did the same slightly above50% of their time. During their displacements the males eventually cameinto contact with the female’s elytra and or female’s head through theirstretched, and moving antennae. Upon touching their bodies, the males in-variably approached the females. The majority (81%) did it by ‘jumpingforward,’ often reorienting their bodies, their head up, antennae open andoriented forward, a behavior we called dashing. This male behavior con-trasted with that of females in these dyads, as upon contact the females


never approached the males. Instead females responded to male’s anten-nal contact by changing direction and running away from males, (88%),or stopped and immobilized on contact (12%). Ninety four percent of theapproaching males were able to hold the females dorsally with their pro-thothoracic legs, and 73% of these mounted the females, by aligning theirbodies with the female’s longer axis, and holding onto them with their pro-thothoracic and mesothoracic legs, and attempted copulation by bendingtheir abdominal tip and touching the female’s abdominal tip, a behavior wecalled genital touch, which was followed by intromission. Following mount-ing, and during copulation males were often observed licking the female’spronoto with their palpi.

Same Sex Dyads

In 88% of the male dyads, both males were observed initiating contactvia their antennae and then dashing to the receiving male. In these dyadsthe receiving males were also observed approaching the initiating male inresponse to contact, although the percentage of dyads where that happenedwas somewhat smaller (69%). Males were often observed mounting othermales and attempting copulation with them. Whenever this happened, themounted males adopted a passive posture, while being hit on their headswith the mounting male’s mouthparts. These interactions usually endedwith the male on top dismounting and moving away.

Female’s response to antennal contact in same sex dyads, contrastedwith that of females in Male–female pairs in that females in these dyads wereobserved approaching their pair after touching or being touched. Their be-havior also contrasted with that of males in same sex dyads. Only in halfof female dyads were both females observed initiating contact and then ap-proaching the receiving female. In addition, the receiving female never ap-proached the female initiating contact in 44% of female dyads, and in theremaining 56% only one female of the pair moved towards the initiating fe-male. Also unlike males, females never dashed towards the individuals theyapproached.

Bioassays: Experiment A

Highly significant differences were found among proportions of thedashing response elicited by intact and washed dead bodies, following an-tennal contact (Cochran’s test Q = 27.667, p � 0.001). This behavior was


Table I. Pair-Wise Comparisons for Male dashing (DS) Towards Intact Dead and DeadWashed Conspecifics After Antennal Contact: 1 – Dead female; 2 – Dead male; 3 – Dead

washed female; 4 – Dead washed male

GroupGroup mean comparison Difference(15 blocks)a (B vs. A) |GB/b-GA/b|b SEc Sd Sαe Conclusions

1 0.67 4 vs. 1 0.67 0.19 3.51 2.80 Reject H0:u4−ul = 02 0.53 4 vs. 2 0.53 0.19 2.81 2.80 Reject H0:u4−u2 = 03 0.00 4 vs. 3 0.00 0.19 0.00 2.80 Accept Ho:u4−u3 = 04 0.00 3 vs. 1 0.67 0.19 3.51 2.80 Reject H0:u3−ul = 0

3 vs. 2 0.53 0.19 2.81 2.80 Reject H0: u3−u2 = 02 vs. 1 0.14 0.19 0.70 2.80 Accept H0: u2−ul = 0

aFive blocks, from the initial 20 blocks, were deleted from the analysis, because ‘0’s occurredfor all treatments.

b|GB/b-GA/b|: (difference between group means).cSE—Standard error.dS—Statistic test.eSa—Critical value (Square root of χ2

0.05,3).

exhibited towards intact dead females and intact dead males by 67, and 53%of males respectively, whereas the washed bodies of both males and femalesfailed to elicit that male response (Table I). A posteriori paired compar-isons revealed that the intact dead bodies of males and females were equallylikely to elicit the dashing response (Table I).

Highly significant differences were found as well among proportions ofthe genital touch behavior, elicited by these treatments following antennalcontact (Cochran’s test Q = 77.11, p � 0.001) as males showed no responseto dead washed bodies. A posteriori paired comparisons revealed that theintact dead bodies of females were equally likely to elicit this behavior (95%of response) as were the intact dead bodies of males (100% of response)(Table II).

Bioassays: Experiment B

All males approached the dummies impregnated with extracts after an-tennal contact, although very few dashed towards them, their response notbeing significantly different from that exhibited to the control (Cochran’stest Q = 6.182, 0.10 < p < 0.20). However significant differences werefound among treatments for the genital touch response, following antennalcontact (Cochran’s test Q = 38.105, p � 0.001). Eighty and ninetyfivepercent of males exhibited this response towards dummies impregnated


Table II. Pair-Wise Comparisons for male genital touch (GT) towards intact dead and deadwashed conspecifics after antennal contact: 1 – Dead female; 2 – Dead male; 3 – Dead washed

female; 4 – Dead washed male

GroupGroup mean comparison Difference(20 blocks) (B vs. A) |GB/b-GA/b|a SEb Sc Sαd Conclusions

1 0.95 4 vs. 1 0.95 0.24 3.93 2.80 Reject H0: u4−ul = 02 1.00 4 vs. 2 1.00 0.24 4.13 2.80 Reject H0: u4−u2 = 03 0.00 4 vs. 3 0.00 0.24 0.00 2.80 Accept Ho: u4−u3 = 04 0.00 3 vs. 1 0.95 0.24 3.93 2.80 Reject H0: u3−ul = 0

3 vs. 2 1.00 0.24 4.13 2.80 Reject H0: u3−u2 = 02 vs. 1 0.05 0.24 0.21 2.80 Accept H0: u2−ul = 0

a|GB/b-GA/b|: |difference between group means|.bSE—Standard error.cS—Statistic test.dSα—Critical value (square root of χ2

0.05,3).

with 5EM and 3EM, respectively, whereas all males ignored the control(Table III). Forty-five percent of males attempted copulation with dummiesimpregnated with the female extracts (3EF and 5EF), but a posteriori com-parisons showed that these proportions were not significantly different fromthose elicited by the control (Table III).

Table III. Pair-Wise Comparisons for Genital Touch (GT) with Dummies Impregnated withFemale and Male Crude Extracts, and a Control, After Antennal Contact: 1–ConcentrationEquivalent to 3 Females; 2–Concentration Equivalent to 5 Females; 3–Concentration Equiva-lent to 3 Males; 4–Concentration Equivalent to 5 males; 5-Equivalent Concentration of Chlo-

roform Solvent

GroupGroup mean comparison Difference(20 blocks) (B vs. A) |GB/b-GA/b|a SEb Sc Sαd Conclusions

1 0.45 5 vs. 1 0.45 0.17 2.65 3.08 Accept H0: u5−ul = 02 0.45 5 vs. 2 0.45 0.17 2.65 3.08 Accept Ho: u5−u2 = 03 0.95 5 vs. 3 0.95 0.17 5.59 3.08 Reject H0: u5−u3 = 04 0.80 5 vs. 4 0.80 0.17 4.71 3.08 Reject H0:u5−u4 = 05 0.00 4 vs. 1 0.35 0.17 2.06 3.08 Accept HQ: u4−ul = 0

4 vs. 2 0.35 0.17 2.06 3.08 Accept H0: u4−u2 = 04 vs. 3 0.15 0.17 0.88 3.08 Accept H0: u4−u3 = 03 vs. 1 0.50 0.17 2.94 3.08 Accept Ho: u3−ul = 03 vs. 2 0.50 0.17 2.94 3.08 Accept H0: u3−u2 = 02 vs. 1 0.00 0.17 0.00 3.08 Accept H0: u2−ul = 0

a|GB/b-GA/b|: |difference between group means|.bSE—Standard error.cS—Statistic test.dSα—Critical value (square root of χ2

0.05,4).


Ultrastructure Analysis

Sensilla Trichodea

Sensilla (s.) trichodea occur over the entire antennal flagellum ofP. semipunctata, which consists of nine flagellomeres (Lopes et al., 2002).They are mainly distributed along the anterior and posterior borders of theflagellum, and at the distal tip of the apical flagellomere. S. trichodea varygreatly in length (50–200 µm), generally being shortest on the internal re-gion of the flagellomeres and lengthening peripherally. Their diameter is3.5–6 µm at the base, tapering to about 1 µm at the apex. They are stout,straight or slightly curved, blunt-tapered hairs with longitudinal groovesthat gradually become faint towards their tip (Fig. 1a). They stand clearlyabove the flagellar surface, pointing outwardly and distally. S. trichodea areinserted into a well-developed, flexible socket. Their apex bears a single ter-minal pore surrounded by a sponge-like cuticle (Fig. 1b). Cross sections atthe apical part of the hair shaft indicate that tiny, electron-dense channel-like structures can run through its cuticular wall (Fig. 1c, inset).

S. trichodea house five to six bipolar sensory cells (Fig. 2f). Their den-drites are divided into two distinct segments by a ciliary constriction: theinner and outer dendritic segments (Fig. 2e). The outer dendritic segmentsonly contain microtubules (Fig. 2d).

From the ciliary region, a thick, cuticular sheath encloses the outerdendritic segments (Fig. 2c). One dendrite ends in a tubular body at thebase of the hair shaft, which is completely enclosed and separated from theremaining ones by the cuticular sheath (Fig. 2b). These dendrites extenddistally into the hair shaft lumen of the sensillum, almost up to its tip, with-out branching, and wrapped by a common cuticular sheath (Figs. 1c and 2a).The cuticular sheath compartmentalizes the lumen of the hair shaft into twolumina: the “dendritic” lumen, containing the dendrites, and the “extra-dendritic” lumen containing a flocculent material (Figs. 1c and 2a). The“dendritic” lumen extends up to the apical pore, while the “extra-dendritic”lumen ends proximally to the tip of the hair shaft.

Three sheath cells are associated with S. trichodea: inner, intermediate,and outer cells (Fig. 2c, d, e).

DISCUSSION

In contrast to what is known of other cerambycid mating systems wherethe meeting of the sexes is mediated by a short-range pheromone, namelythose of Paraglenea fortunei (Wang et al., 1990), Monochamus alternatus


Fig. 1. (A) and (B). Scanning electron micrographs of flagellar sensilla trichodea of Phoracan-tha semipunctata. (A) Dorsal view of part of an area of the 7th antennal flagellum segment ofa male, showing a sensillum trichodeum (arrow) inserted into a socket surrounded by a raisedcuticular ring (arrowhead). Scale bar = 10 µm; (B) High magnification, showing the apicalpore (arrowhead) at the sensillum’s hair shaft tip. The cuticle around the pore has a spongyappearance. Scale bar = 500 ηm; (C) Transmission electron micrograph of a sensillum tri-chodeum. Cross-section through the hair shaft of the sensillum showing the outer dendriticsegments within the “dendritic” lumen and shielded by the cuticular sheath. The cytoplasmof the outer dendritic segments contains microtubules (arrowhead). Asterisk, extra-dendriticlumen. Scale bar = 0.5 µm. Inset: Higher magnification of a sensillum’s hair shaft near the tipshowing tiny channel-like structures (arrowhead) with an electron-dense material, extendingfrom the sensillar lumen into the cuticular wall of the sensillum. Scale bar = 0.2 µm.

Fig. 2. Transmission electron micrographs of flagellar sensilla trichodea of P.semipunctata. (A) Longitudinal section through the socket of the hair shaft (dou-ble arrowhead) showing dendrites (d) entering the hair shaft lumen and en-veloped by the cuticular sheath (arrowhead). Star, “extra-dendritic” lumen (star)Scale bar = 1 µm. (B) Oblique section at the level of insertion of the hair shaft.Note one dendrite with the tubular body (tb) separated from the remaining den-drites (d) by the cuticular sheath (asterisk with arrowhead). Scale bar = 1.5 µm.(C) Cross section below the hair shaft base. The outer dendritic segments (d)can be seen with microtubules and wrapped by a thick electron-dense cuticu-lar sheath (arrowhead). I, inner sheath cells; In, intermediate sheath cell; O,outer sheath cell. Scale bar = 1.5 µm. (D) Cross section of six dendrites atthe level of the ciliary region. I, inner sheath cell; small arrow, microtubules.Scale bar = 0.5 µm. (E) Longitudinal section through the ciliary sinus (asterisk)with one dendrite cut at the junction of the inner (id) and outer (od) dendriticsegments, d, outer dendritic segments; I, inner sheath cell (I); In, intermediatesheath cell (In); O, outer sheath cell. Scale bar = 2 µm. (F) Longitudinal sectionat the level of the perikarya (N) of three neurons. Axons (ax). Scale bar = 3 µm.


(Kim et al., 1992), and Semanotus japonicus (Kim et al., 1993), we havenot found a clear evidence that supports the existence of a short-rangepheromone produced by either sex in P. semipunctata. Females were clearlyvery active, which could be interpreted potentially as a response to a short-range pheromone emitted by the males, but their encounters with malesseemed to have happened by chance. The same seems to be true for themales that approached other males and females only after antennal contact.The dashing behavior exhibited by P. semipunctata males upon antennalcontact with females and males, either live or freshly dead, and their be-havior towards dummies impregnated with crude extracts, constitute how-ever evidence that a contact pheromone present on the body surface of thisspecies is required to trigger the sequence leading to mating, superimposingover visual or tactile stimuli.

P. semipunctata males showed no differences in response toward deadconspecifics (males and females) (Bioassay A) although some failed to re-spond to some of the dummies impregnated with female crude extracts(Bioassay B). Their responses thus seem to point to the existence of anon-differentiated chemical or mixture of chemicals between sexes simi-lar to what is known for Monochamus. alternatus (Kim et al., 1992) wherea specific substance present on the cuticular surface of individuals does notidentify the sex of the specific individual. This result contrasts with those ofother cerambycids where the sexes differ in the nature (e.g., Psacothea hi-laris (Fukaya et al., 1996); Anoplophora chinensis (Wang, 1998); A. malasi-aca (Fukaya et al., 1999; Akino et al., 2001)) or proportion of the chemi-cal components present on the cuticle (e.g. S. japonicus (Kim et al., 1993);Anoplophora glabripennis (Zhang et al., 2003)).

In M. alternatus the lack of discrimination is counterbalanced by thefact that females are attracted to the male short-range pheromone (Kimet al., 1992). Given that our results point to the absence of a short-rangepheromone and the existence of a non-differentiated contact pheromone,it is possible that males cue both on the females’ runaway response to an-tennal contact and on their cuticle odor to successfully identify a prospec-tive mate. In fact in cerambycids, female movement has been describedas a relevant behavior used as a cue for mate recognition in Xylotrechuspyrrhoderus (Iwabuchi, 1982), and another female behavior conveying im-portant information for mate recognition has also been reported for otherspecies outside this group, e.g. the Curculionidae Macromerus bicinctuswhere males rely on female-posture to recognize prospective mates (Wcisloand Eberhard, 1989).

Both the presence of a yet unidentified contact pheromone on the cu-ticle of P. semipunctata and its detection through antennal contact are in-dicative of an antennal contact chemoreceptive mechanism in this species.


Information about the contact pheromone might be mediated by S. tri-chodea. Our morphological study clearly showed that the S. trichodea fulfilthe morphological criteria known for other gustatory chemoreceptive sen-silla of insects (Zacharuk, 1980). They bear an apical pore surrounded bya sponge-like cuticle, cuticular channel-like structures crossing the sensillawall at the apical region of the hair shaft and unbranched dendritic seg-ments within the sensillar lumen. Chemical stimuli might access the sensil-lar lumen through the apical pore, and probably the channel-like structures,and reach the dendritic membrane. The distribution pattern, size and ori-entation of S. trichodea are consistent with a contact chemoreceptive role,allowing them to contact with substrata or objects easily.

In conclusion, we consider the characterization of the contactpheromone of P. semipunctata, through the isolation and identification ofactive fractions a requirement to clarify whether this species has a distinc-tive female contact pheromone, or not, which may press for a more con-clusive study on the potential existence of a male short-range pheromonein this species. We also see the male’s antennae playing an important rolein the mating system of this species, as a structure adapted to perceive acontact pheromone, based on the account of the anatomical data on theS. trichodea, but we consider necessary to further investigate these sensilla(e.g., by single-cell recording) so that we may functionally link them withtheir suggested role in antennal contact chemoreception. This will likely bedone as soon as we have identified the contact pheromone.

ACKNOWLEDGMENTS

We thank Dr Moura Nunes (Electron Microscopy Laboratory, CancerPortuguese Institute—Lisbon) who kindly provided us access to the TEMand helped in the film processing; Mrs Teresa Varejao for photographicwork. This research was supported by The Center of Applied Ecology,University of Evora, and by JNICT (PRAXIS XXI/BM/38/94).

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The Role of Antennae in Mate Recognition in Phoracantha semipunctata (Coleoptera: Cerambycidae)

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