Pei Yang

Zong-Bo Li

Da-Rong Yang

Yan-Qiong Peng

Finn Kjellberg

Comparison of the antennal sensilla of females of four fig-wasps associated with Ficus auriculata

Keywords: Host-parasitoid community, Ficus auriculata, Foundress, Antennal sensilla

A comparison was performed of the antennal sensilla of females of four chalcid wasp species Ceratosolen emarginatus Mayr, 1906, Sycophaga sp., Philotrypesis longicaudata Mayr, 1906, and Sycoscapter roxburghi Joseph, 1957, which are specific and obligatory associated with Ficus auriculata (Lour, 1790). The four species exhibit different oviposition strategies in the fig ovules where their offspring hatch and develop. Antennal sensilla morphology was evaluated using scanning electron microscopy. Females of the four species present 11 morphologically similar types of sensilla: trichoid sensilla, sensilla obscura, chaetica sensilla 1 and 2, which all have mechanosensory functions; uniporous basiconic sensilla, which are presumably contact chemosensilla; basiconic capitate peg sensilla, coeloconic sensilla 1, multiporous basiconic and placoid sensilla, which may be regarded as olfactory sensilla, and coeloconic sensilla 2 and 3, which are presumed to be proprioreceptors or pressure receptors. The four species have significant differences in the abundance and arrangement of trichoid sensilla and chaetica sensilla 1 on the flagellum. The coeloconic sensilla and sensilla obscura only occur on the antennae of C. emarginatus that enter figs. The chemosensilla which are presumably involved in host discrimination, i.e., basiconic sensilla, multiporous placoid sensilla and basiconic capitate peg sensilla, are similar in shape and configuration, although they present some differences in abundance. These findings provide practical information on the adaptations of fig wasps and the relationship between multisensory antennae and functions in fig wasp behaviour.

Introduction

 [START_REF] Cook | Mutualists with attitude: coevolving fig wasps and figs[END_REF][START_REF] Weiblen | How to be a fig wasp[END_REF]. They locate their host using stage-specific chemical cues emitted by the figs of their host species [START_REF] Hossaert-Mckey | Floral scents: their roles in nursery pollination mutualisms[END_REF][START_REF] Proffit | Chemical mediation and niche partitioning in non-pollinating fig-wasp communities[END_REF][START_REF] Proffit | Signalling receptivity: comparison of the emission of volatile compounds by figs of Ficus hispida before, during and after the phase of receptivity to pollinators[END_REF][START_REF] Wang | The mechanism of pollinator specificity between two sympatric fig varieties: a combination of olfactory signals and contact cues[END_REF]. In insects, the antenna is an organ capable of detecting odours. It is a multisensory detector carrying various receptors that sense a series of signals, including smell, but also taste, temperature, sound, gravity, wind speed, and even the geomagnetic field [START_REF] De Oliveira | Ant antennae: are they sites for magnetoreception?[END_REF][START_REF] Isidoro | Morphology of antennal gustatory sensilla and glands in some parasitoids Hymenoptera with hypothesis on their role in sex and host recognition[END_REF][START_REF] Meyhöfer | Mechano-and chemoreceptors and their possible role in host location behavior of Sympiesis sericeicornis (Hymenoptera: Eulophidae)[END_REF][START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF][START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF][START_REF] Staudacher | Antennal movements and mechanoreception: Neurobiology of active tactile sensors[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF][START_REF] Zacharuk | Antennae and sensilla[END_REF]. The extent to which variation in the structure of the antennae as well as the presence and distribution of different types of sensory receptors among wasp species can be explained by differences in behaviour and oviposition strategies are poorly known (but see [START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF]. For fig wasps, the few studies of the sensilla on the antenna have almost exclusively focused on pollinators. These wasps enter figs to oviposit and the host figs are specialized to attract them [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF][START_REF] Ware | Repeated evolution of elongate multiporous plate sensilla in female fig wasps(Hymenoptera: Agaonidae: Agaoninae)[END_REF]. Little is known about the sensory organs on the antennae of non-pollinating fig wasps (NWFWs), which oviposit from outside the fig without providing any known benefits to the mutualism [START_REF] Cook | Mutualists with attitude: coevolving fig wasps and figs[END_REF][START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF].

In this study we contrast the antennal ultrastructure of four fig-wasp species associated with one host species, Ficus auriculata: a pollinator (Ceratosolen emarginatus Mayr, 1906), belonging to the Agaonidae, and three NWFW species, one belonging to the Sycophaginae (Sycophaga sp.) and two belonging to the Sycoryctinae (Philotrypesis longicaudata Mayr, 1906, Sycoscapter roxburghi Joseph, 1957). The aim of the study was to investigate similarities among species but also differences that may be associated with how the antennae contact the fig surface and with the fig-entering behaviour of the pollinating wasps. Our working hypothesis is that several traits of the first three segments of pollinator antennae are associated with the figentering behaviour, while the distal segments of the antennae would show many more similarities with NPFW antennae. A second hypothesis is that wasps that contact the fig surface with the whole length of the flagellomere will have different sensilla from wasps that only tap the surface of the fig with the distal segment of the antennae. A third hypothesis is that since pollinators only live a few hours as adults while NPFW adults are longer lived, they may present a more specialized, less diversified, detection system.

We compared here the morphology, number and distribution of the different types of sensilla on the antennae of four fig wasps associated with the host F. auriculata using SEM imagery. We discuss the likely roles of different antennal sensilla according to their morphology and interpret their presence according to global functioning of the antennae in connection with fig wasp behaviour.

Materials and methods

The study species

Ficus auriculata Lour., 1790, is a widespread species ranging from Pakistan to Thailand and Indochina through south China [START_REF] Berg | Moraceae-Ficus[END_REF]. The tree is pollinated by the species complex of C. emarginatus and it is host to 4 non-pollinating fig wasps: Sycophaga sp1., Sycophaga sp2., P. longicaudata and S. roxburghi [START_REF] Peng | The population dynamics of a non-pollinating fig wasp on Ficus auriculata at Xishuangbanna, China[END_REF]; also see figweb (http://www.figweb.org)). These wasps exhibit strong sexual dimorphism: the females are winged, as in most chalcid wasps, and effect host plant choice while the males are wingless and usually do not leave the cavity of their natal fig. Sycophaga sp2. was present at low frequencies, while the other species were more abundant and may occur together in a fig [START_REF] Peng | The population dynamics of a non-pollinating fig wasp on Ficus auriculata at Xishuangbanna, China[END_REF]; personal observation). Ficus auriculata produces large figs, and the different wasp species oviposit sequentially at different fig development stages (Fig. 2). Therefore, the different wasp species are probably exposed to different stage-specific chemicals or mixes of chemicals. Further, the pollinating wasp C. emarginatus, enters the fig to oviposit, losing part of the flagellomere in the process while the other species oviposit from outside the fig.

Oviposition and behavioural events of fig wasps

Ficus auriculata is a gynodioecious fig species that produces a globose-shaped inflorescence called the syconium or fig. The development of the fig is separated into 5 developmental stages (A-E) [START_REF] Berg | Moraceae-Ficus[END_REF]. The four most abundant wasp species associated with F. auriculata figs oviposit at different stages of fig development [START_REF] Peng | The population dynamics of a non-pollinating fig wasp on Ficus auriculata at Xishuangbanna, China[END_REF] Personal observation; Fig. 1). Sycophaga sp. is an as yet undescribed galler that oviposits early in fig development before female flower receptivity, from outside the fig into ovules in male and seed figs, using its long ovipositor [START_REF] Peng | The population dynamics of a non-pollinating fig wasp on Ficus auriculata at Xishuangbanna, China[END_REF] Figs from which wasps were soon to emerge were placed in a nylon bag so that females could be collected as soon as they emerged through the exit hole chewed by male pollinators. SEM samples were prepared for observation using two techniques. First, foundresses were fixed in 2% glutaraldehyde and the number and distribution of sensilla on the antennae of each species were examined. Second, typical characteristics of the sensillar surface, including the cuticular grooves, pores, and basal sockets, were examined in detail. The female wasps were cleaned, stored in 3.5% glutaraldehyde, and then dehydrated in a graded series of alcohol as well as pure isoamyl acetate [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF]. After dehydration, the samples were criticalpoint dried with a Quorum K850 drier (Quorum Technologies, Deben, UK), gold-coated in a Quorum Q150R sputter device (Quorum Technologies, Deben, UK), and observed with a Zeiss EVO-LS10 (Zeiss, Oberkochen, Germany) at 10 kV. The antennae were viewed from the ventral, dorsal, lateral and top sides. The antennae of more than 12 female wasps were examined for each species. (B), P. longicaudata (C), and Sycoscpater roxburghi (D) and depicts the Radicula (Rd), Scape (Sc), Pedicel (Pe), and Flagellum (Fl), which is subdivided into the Anellus (An), Funicle (Fu) and Clava (Cl). Important morphological differences among the four species are indicated with a solid arrow (flagellomere one (F1) base with a spine-like expansion), a dashed loop (sensilla obscura on the dorsal side of the pedicel), an asterisk (deep socket), and a dashed arrow (radicula). Scale bar = 100 μm.

Sensilla number and analysis

The sensilla on the different sides of the antennae of each species were identified, recorded, and measured from the printed SEM pictures. For the irregular antennal segments, such as the scape on the antennae of C. emarginatus, sensilla numbers were directly counted on the ventral and dorsal sides. When counting sensilla on the cylindrical segments, such as the flagellomere (except for the terminal segment, which is conical and examined from the top side, see Fig. 4B), the total numbers were calculated from 2/3 sides of the total viewing surface area [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Ware | Repeated evolution of elongate multiporous plate sensilla in female fig wasps(Hymenoptera: Agaonidae: Agaoninae)[END_REF]. Each sensillary type was measured in at least 20 individuals from each species. To test for significance of variation among species, Kruskal-Wallis tests were used to analyse the distribution and density of multiporous placoid sensilla, which are a synapomorphy of chalcid wasps and were the most conspicuous type of sensillum and occurred on the flagella of females of the four wasp species.

Results

Antennae

The female antennae of the four species studied here are geniculate (angular in presentation) and consist of three functional segments: the scape with radicula (the area that is located proximally on the scape, is limited distally by a constriction and bears the basal knob proximally), the pedicel (the antennal segment that is connected proximally to the scape and distally to the flagellum) and the flagellum (Fig. 3 A-D). The mean length of the different segments are shown in Table 1. The radicula (Rd) is separated from the scape by a constriction, particularly in the two Sycoecine species, P. longicaudata and S. roxburghi (Fig. 3 C,D). It is relatively short for chalcid wasps [START_REF] Heraty | A phylogenetic analysis of the megadiverse Chalcidoidea (Hymenoptera)[END_REF][START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF]. It has a particular type of sensillum (Figs. 3C and5G). The radicula fits into the antennal socket, where it connects with the head through a ball-and-socket joint that allows movement in all directions. The scape (Sc) in C. emarginatus is irregular in shape and has two deeper grooves on the ventral side (Fig. 3A) that may serve to hold the basal part of the flagellum when the pollinators pass through the bracts. On the other hand, in NPFWs, the scape is cylindrical and is approximately 5-times longer than wide (Fig. 3 B-D). The pedicel (Pe) in the four species is a short, pearshaped segment that connects to the scape by a hinge-joint articulation, thus restricting the movement of the antennae to one plane. The elongate flagellum consists of eight, eleven, eleven and nine distinct flagellomeres in C. emarginatus, Sycophaga sp., P. longicaudata and S. roxburghi, respectively; thus, the entire antenna is composed of varying segment numbers. We did not determine which segments were fused in the context of the basic organisation of chalcid wasp antennae, so we did not number the segments of the flagellum. The flagellum is subdivided into a basal ring-like sub-segment, the anellus (An), the median funicle (Fu) and the distal clubshaped clava (Cl) (Fig. 3). In this study, flagellomere one is only described in detail for C. emarginatus because its base is irregular and produces a pointed spine-like expansion, traditionally called a tooth, which is clearly visible on the antenna (Figs. 3A and4D). The surfaces of the antennae of the four species present a similar aspect, although the type, number and distribution of the antennal sensilla on the four species differ. These morphologically different sensilla have been given various names in the literature. We will use the terms trichoid sensilla, chaetica sensilla 1 and 2, uniporous and multiporous basiconic sensilla, multiporous placoid sensilla, basiconic capitate peg sensilla, sensilla obscura and coeloconic sensilla 1 to 3. These eleven different types of sensilla were never observed to occur together in a single species (Table 2). Trichoid sensilla are present on all segments where they are abundant except for the flagellum of S. roxburghi, where they are few (Table 2; Fig. 6). These sensilla are inserted at a sharp angle, inclined in the direction of the axis of the antennae, and their tips are usually curved and point toward the distal end of the antennae (Fig. 4A-D; Fig. 5A andB). In the four species, TS vary in length between 4.13 μm and 49.54 μm depending on their location on the antennae. They present 8-11 longitudinal grooves reaching up to the tip. The base is inserted in a small asymmetric socket with a slightly raised ridge, and we did not detect any pore (Fig. 5 E,E1). Trichoid sensilla were most abundant in C. emarginatus and Sycophaga sp., slightly less so in P. longicaudata and were distinctly fewer in S. roxburghi (Table 2).

Chaetica sensilla (ChS)

We classified these sensilla into two subtypes: Chaetica sensilla type 1 (ChS-1) and type 2 (ChS-2). These subtypes present differences in morphology, size, location and cuticular attachment (Fig. 5D,F). ChS-1 is elongate and was only observed on the flagellum of P. longicaudata and S. roxburghi (Table 2; Fig. 6E-G), where they are intermixed with trichoid sensilla and partly compensate for the lower number of TS in these two species when compared to C. emarginatus and Sycophaga sp. (Fig. 5A). These sensilla are straight or medially bent bristles that arise directly from the cuticle. Their diameter decreases progressively finishing in a pointed apex. In these two species, the length of the ChS-1 Table 1 varies noticeably in different locations on the antennae, ranging from 0.89 μm to 8.35 μm. ChS-1 are the most numerous sensilla on S. roxburghi antennae, representing 40% of the total number of sensilla, while they are less abundant on P. longicaudata antennae (Table 2). ChS-2 is only observed on the base of the radicula and at the scape-pedicel joint in the antennae of all species (Table 2; Fig 4C; Fig. 6 A,C,D,G,H). There are about 20 per antenna. This type of sensillum is constituted by a short, tapered bristle with a bulbous base that projects perpendicularly to the antennal axis from a wide flexible socket (Fig. 5 F). ChS-2 has a constant length of 4.01 ± 0.24 μm and a basal diameter of 1.11 ± 0.26 μm. Both types of ChS are smooth and do not have apparent pores (Fig. 5 D,F).

Basiconic sensilla (BS)

Two types of basiconic sensilla were observed. Uniporous basiconic sensilla (UBS) are erect, and they were only present at the distal end of the terminal flagellomere of the antennae in all species. Because of their length (limited variation among species; 24.75 ± 5.93 μm in length, 2.33 ± 0.39 μm in basal diameter) and orientation, the tips of these sensilla project further than the other sensilla TS/ChS-1 (Fig. 4A andB; Fig. 6A,B,F). UBS are inserted in flexible basal sockets, present a longitudinal groove on their shafts, and their tips are blunt, suggesting the presence of a single apical pore (Fig. 5A). Multiporous basiconic sensilla (MBS) are short, upright, straight or medially slightly bent bristles distributed on the lateral-ventral side of the flagellomere, on the funicle and clava of all species (Fig. 4A andB; Fig. 6A,B,E,F). They are however much more abundant in C. emarginatus and in Sycophaga sp. than in the two other wasp species. These MBS present limited variation and are 21.41 ± 4.09 μm long and their basal diameter measures 2.02 ± 0.25 μm. The shaft arises from a slight depression in the cuticle Mean length of the female antennal segments of the four wasps associated with F. auriculata.

BCPS 10 ± 1 12 ± 1 8 ± 1 8 ± 1 ChS-1 - - 63 ± 3 102 ± 10 ChS-2 24 ± 2 22 ± 1 22 ± 2 18 ± 2 SO 27 ± 1 - - - CoS-1 1 - - - CoS-2 24 ± 2 - - - CoS-3 34 ± 4 - - - Total
564 ± 14 497 ± 23 410 ± 18 256 ± 11 Values are the mean ± S.E of different types of antennal sensilla on each antennae per species (n = 12). The letter abbreviations state the type of sensilla described in text. and5B). The MBS wall is relatively smooth and presents apparent pores on high resolution images (Fig. 5B1).

Multiporous placoid sensilla (MPS)

Multiporous placoid sensilla are numerous and conspicuous sensory structures on the antennae of all species (Fig. 3 A-D; Fig. 6 A,B,E,F). This type of sensillum rises directly from the antennal cuticle to form a sausage-shaped plate, and they cover each funicular and claval flagellomere, in a ring-like arrangement (Fig. 4 A-B, D). Multiporous placoid sensilla lie parallel to the antennal axis, present a pointed end and are always located between rows of TS/ChS-1 (Fig. 4A,D). The MPS plate is smooth and presents two pores per 400 nm at median length (Fig. 5C). The pore density of MPS surface per 1 μm 2 is 27.86 ± 3.44 in C. emarginatus, 27.75 ± 5.94 in Sycophaga sp., 26.00 ± 4.92 in P. longicaudata, and 26.13 ± 6.10 in S. roxburghi (n = 8), without significant difference among the four species (χ 2 = 0.579, P = 0.901).

3.2.5. Basiconic capitate peg sensilla (BCPS) Basiconic capitate peg sensilla are found on the lateral-ventral side of the apical end of each flagellomere on the funicle and clava on the antennae of all species (Fig 4B; Fig. 6E andF). BCPS are nipple-like structures with an expanded terminal part and a short stalk that sits within a distinct cuticular depression. In the four species, the BCPS shapes are slightly variable and present major differences in the length of the stalk and the diameter of the nipple. There are usually 1, sometimes 2, and very occasionally 4 BCPS per flagellomere (Fig. 4B). Their nipples contain 11 finger-like projections jointing at the tip of the stalk. The grooves between projections are relatively deep and may indicate the existence of pores (Fig. 5H). Coeloconic sensilla are recessed in a pit and only distributed on segments one and three of the antennae of the pollinator C. emarginatus (Table 2; Fig. 6 A,C,D). Based on the peg exterior appearance, location and cuticular attachment, three different types of CoS were identified (Fig. 5I-K). Type 1 CoS (CoS-1) is ear-shaped, rare (only 1 per antennae), and always close to the crossed groove on the tooth-like expansion. This sensillum sits in a shallow oval socket with a central opening, and its cuticle has 4-5 deep longitudinal grooves reaching the terminal tip (Fig. 5I). Type 2 CoS (CoS-2) is spiral, bud-shaped protrusion that covers the tooth-like expansion and the top dorsal side of the scape, and it mixes with type 1 (CoS-1) and 3 CoS (CoS-3). The shaft is inserted into a deep, circular socket and has a diameter of 0.78 μm that tapers gradually (Fig. 5J). CoS-3 is a small-coned protrusion that is obviously lower than the depth of the wide circular socket (Fig. 5K). CoS-3 always mixes with CoS-2 but never with CoS-1. Both CoS-2 and CoS-3 have smooth walls without pores (Fig. 5J-K).

Sensilla obscura

The name sensilla obscura (SO) was originally proposed by [START_REF] Cockerill | External Morphology of Antennal Sensilla in Relation to the Host Searching Behaviour of Liporrhopalum Tentacularis Grandi (Hymenoptera: Agaonidae)[END_REF] and has been used in subsequent work on fig pollinating wasps [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF]. They are highly distinctive sensilla grouped dorsally on the pedicel of the pollinator C. emarginatus (Table 2 6A,C). Each SO is flattened and awl-like in shape and strongly tapered to an elongate and slender tip parallel with the antennal surface (Fig. 5G). These sensilla measure 12.24 ± 1.12 μm in length and 2.41 ± 0.40 μm in basal diameter. Between 5 and 6 widely longitudinal grooves line the shaft, which is inserted into an oval socket that is almost equal to the length of the shaft. There are approximately 27 SOs, all of which point toward the head (Fig. 4C).

Abundance and distribution of female antennal sensilla in the four wasp species

The four species differ significantly in the distribution and abundance of some sensillar types on the antennae (Table 2; Fig. 6). TS occur on the flagellum of the four species but are significantly more abundant in C. emarginatus and Sycophaga sp. than in P. longicaudata and S. roxburghi (Table 2;Fig 1;Fig 7). Similarly, MBS are more abundant in C. emarginatus and Sycophaga sp. than in P. longicaudata and S. roxburghi. MBS are restricted to the lateral-ventral side of each flagellomere on the funicle and clava. In contrast, the long, rigid ChS-1 present on the flagellum are numerous on P. longicaudata and S. roxburghi, and none are found on C. emarginatus and Sycophaga sp. (Table 2; Fig. 7). The short and strong ChS-2 only occur on the lateral-ventral side of the radicula and ventral surface of the pedicel at the scape-pedicel joint, and no major differences in distribution and number were found among the four species. UBS are always located at the apex of the last flagellomere, and a constant number (approximately 6) is found on the antennae of each species (Table 2). MPS are present in all four species. MPS lengths vary according to species, while their width does not vary (Fig. 8). Their size and number are negatively correlated among the four species (r = -0.96, P = 0.02; Pearson correlation). Smaller MPS sizes, such as lengths of 32.44 ± 3.52 μm on the antennae of Sycophaga sp., are associated with higher numbers of MPS (χ 2 = 27.809, P < 0.001), thus leading to more ring-like arrangements on the antennae and larger total surface for the smaller MPS (χ 2 = 27.152, P < 0.001; Table 2;

Discussion

A wide variety of sensilla have been recorded on the antennae of female parasitoids [START_REF] Basibuyuk | Gross morphology of multiporous plate sensilla in the Hymenoptera (Insecta)[END_REF][START_REF] Isidoro | Morphology of antennal gustatory sensilla and glands in some parasitoids Hymenoptera with hypothesis on their role in sex and host recognition[END_REF][START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF], and their detailed structures and configurations can help interpret sensilla function with regard to sensory-guided behaviours, such as host discrimination and exploitation [START_REF] Das | Abundance of antennal chemosensilla in two parasitoid wasps with different degree of host specificity may explain sexual and species differences in their response to host-related volatiles[END_REF][START_REF] Isidoro | Morphology of antennal gustatory sensilla and glands in some parasitoids Hymenoptera with hypothesis on their role in sex and host recognition[END_REF][START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF][START_REF] Roux | Antennal structure and oviposition behavior of the Plutella xylostella specialist parasitoid: Cotesia plutellae[END_REF][START_REF] Staudacher | Antennal movements and mechanoreception: Neurobiology of active tactile sensors[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF]. There is as yet no standardised naming of sensilla and the same types of antennal sensilla are subdivided and assigned to different sensilla types by different authors despite presenting similar morphologies and distributions. Therefore, are difficult and associations between name and function are fuzzy. In this study, we assigned sensilla to different types based on the presence of distinct differences in external appearance and cuticular attachment following the nomenclature used by [START_REF] Basibuyuk | Gross morphology of multiporous plate sensilla in the Hymenoptera (Insecta)[END_REF], [START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF], [START_REF] Dweck | Antennal sensory receptors of Pteromalus puparum female (Hymenoptera: Pteromalidae), a gregarious pupal endoparasitoid of Pieris rapae[END_REF][START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF], 2014). For example, although hairs had various sizes and were widely distributed on the antennae of the four species (Fig. 4A-D; Fig. 5A-B), they presented similar shafts and basal sockets. Therefore, we used the same name for all of them, i.e. trichoid sensilla, and did not subdivide them again into several subtypes or classify them under other names [START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF]. Chaetica sensilla should reflect the same rigid spines with a flexible basal socket as type 1 and should have a similar appearance and distribution but may vary in size.

Here, we reported 11 different types of female antennal sensilla in four wasp species associated with the figs of F. auriculata. These wasps depend largely on species-specific chemicals to discriminate their host figs from other fig species [START_REF] Hossaert-Mckey | Floral scents: their roles in nursery pollination mutualisms[END_REF][START_REF] Proffit | Chemical mediation and niche partitioning in non-pollinating fig-wasp communities[END_REF][START_REF] Wang | The mechanism of pollinator specificity between two sympatric fig varieties: a combination of olfactory signals and contact cues[END_REF]. These types of antennal sensilla never occur simultaneously on a single species and their types are consistent with those observed on other hymenopteran parasitoids [START_REF] Amornsak | External morphology of antennal sensilla of Trichogramma australicum Girault (Hymenoptera : Trichogrammatidae)[END_REF][START_REF] Basibuyuk | Gross morphology of multiporous plate sensilla in the Hymenoptera (Insecta)[END_REF][START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF][START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF][START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF][START_REF] Steinbrecht | Pore structures in insect olfactory sensilla: a review of data and concepts[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF]. The sensilla observed on C. emarginatus were similar to those found on the closely related fig pollinating wasp C. solmsi marchali [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF]. On the other hand, while the global organisation of sensilla is similar, coeloconic sensilla are absent in the pollinator of F. curtipes, Waterstoniella sp. [START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF]. SO are restricted to fig pollinating wasps and always grouped dorsally on the pedicel [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF][START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF]. They have not been observed in other wasp species (Polidori and NievesAldrey, 2014;[START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF]. Analysing the distribution of the different types of sensilla in connection with their morphology and the behaviour of the wasps may provide practical information and theoretical insights into their functions in these fig wasps.

Putative functions of the different sensilla types

Hair-like TS were abundant on the antennae of the four study species and have been reported on the antennae of nearly all investigated hymenopteran parasitoids [START_REF] Amornsak | External morphology of antennal sensilla of Trichogramma australicum Girault (Hymenoptera : Trichogrammatidae)[END_REF][START_REF] Bleeker | Antennal sensilla of two parasitoid wasps: a comparative scanning electron microscopy study[END_REF][START_REF] Das | Abundance of antennal chemosensilla in two parasitoid wasps with different degree of host specificity may explain sexual and species differences in their response to host-related volatiles[END_REF][START_REF] Gao | Antennal morphology, structure and sensilla distribution in Microplitis pallidipes (Hymenoptera: Braconidae)[END_REF][START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF][START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF][START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF][START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF][START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF][START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF][START_REF] Roux | Antennal structure and oviposition behavior of the Plutella xylostella specialist parasitoid: Cotesia plutellae[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF]. In the four analysed species, the TS occur in various forms and lengths and at different locations, but they always present more longitudinal grooves on the ventral side in the direction of the antennal axis. The sockets of the TS hairs are small, flexible, and asymmetric, thus allowing for the directional movement of the hair; they present the functional morphology of mechanoreceptors [START_REF] Staudacher | Antennal movements and mechanoreception: Neurobiology of active tactile sensors[END_REF].

The absence of apical or peripheral pores and traces of wall pores observed on the TS in the present study further establishes that they are not olfactory receptor [START_REF] Steinbrecht | Pore structures in insect olfactory sensilla: a review of data and concepts[END_REF]. A transmission electron microscopy investigation has further confirmed a mechanosensory function of TS in two pollinating fig wasp species, C. solmsi marchali and Waterstoniella sp. by showing lack of innervation of the TS body [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF].

We found two types of non-porous ChS on the antennae of the four species. ChS-1 that were distributed on the flagellum of both P. longicaudata and S. roxburghi are similar to those described in some braconid Microplitis [START_REF] Gao | Antennal morphology, structure and sensilla distribution in Microplitis pallidipes (Hymenoptera: Braconidae)[END_REF][START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF] and pteromalid Pteromalus species [START_REF] Dweck | Antennal sensory receptors of Pteromalus puparum female (Hymenoptera: Pteromalidae), a gregarious pupal endoparasitoid of Pieris rapae[END_REF][START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF], while they are absent in pollinating fig wasps (Agaonidae) [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF][START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF]. ChS-1 have been assumed to have a mechanosensory functions because of their socket-like insertions, and because they present non-porous walls as revealed from the SEM images [START_REF] Gao | Antennal morphology, structure and sensilla distribution in Microplitis pallidipes (Hymenoptera: Braconidae)[END_REF][START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF][START_REF] Staudacher | Antennal movements and mechanoreception: Neurobiology of active tactile sensors[END_REF]. Although the location and number of ChS-1 do not vary, their sizes vary even on the same flagellomere, such as on the funicle on S. roxburghi. We propose that the various ChS-1 can sense the magnitude of external pressure from the antennal waving frequency and protect the fragile chemoreceptors from damage. ChS-1 are entirely replaced by TS in C. emarginatus and Sycophaga sp., two species in which the whole length of the flagellum may contact the fig surface.

ChS-2 are widely found on various parasitoids, although they have been given different names, such as Böhm's bristles or hair plates [START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF][START_REF] Staudacher | Antennal movements and mechanoreception: Neurobiology of active tactile sensors[END_REF], aporous type 4 sensilla trichodea [START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF], and non-porous sensilla chaetica [START_REF] Dweck | Antennal sensory receptors of Pteromalus puparum female (Hymenoptera: Pteromalidae), a gregarious pupal endoparasitoid of Pieris rapae[END_REF]. These small erect sensilla only occur at the level of the radicula and scape-pedicel joint in all species and may function as proprioreceptive position detectors that keep the wasps informed of the relative position of the antennal segments [START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF].

The uniporous and multiporous basiconic sensilla recorded in C. solmsi marchali and Waterstoniella sp. were also present in the four species investigated here [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF]. The morphology and location of the uniporous basiconic sensilla suggest that they correspond to the same category as sensilla previously described under other names such as the fluted BS [START_REF] Norton | Antennal sensilla of three parasitic Hymenoptera[END_REF], TS with a tip pore [START_REF] Bleeker | Antennal sensilla of two parasitoid wasps: a comparative scanning electron microscopy study[END_REF], uniporous ChS [START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF], and uniporous pitted sensilla [START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF]. These UBS present an apical pore and emerge above the other sensilla, suggesting that they are involved in contact chemoreception when wasps touch a surface with the tip of their antennae, as has been suggested in many other Hymenoptera [START_REF] Dweck | Antennal sensory receptors of Pteromalus puparum female (Hymenoptera: Pteromalidae), a gregarious pupal endoparasitoid of Pieris rapae[END_REF][START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF][START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF][START_REF] Roux | Antennal structure and oviposition behavior of the Plutella xylostella specialist parasitoid: Cotesia plutellae[END_REF]. We noted that after landing on a fig, female wasps of the different species tapped the surface of the fig with the distal part of the antennae bearing UBS, which further suggests a role of these sensilla in contact chemoreception and fig stage-specific discrimination [START_REF] Proffit | Chemical mediation and niche partitioning in non-pollinating fig-wasp communities[END_REF]. The number of UBS, 6, was remarkably constant among species. The presence of UBS located on the terminal antennal segment seems to be generalised in Chalcidoidea.

Multiporous basiconic sensilla have also been described as curved non-fluted BS [START_REF] Norton | Antennal sensilla of three parasitic Hymenoptera[END_REF], sensilla basiconica I [START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF], TS with wall pores [START_REF] Bleeker | Antennal sensilla of two parasitoid wasps: a comparative scanning electron microscopy study[END_REF], and multiporous pitted TS [START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF]. In general, multiple wall pore sensilla have been assumed to function as olfactory structures [START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF][START_REF] Steinbrecht | Pore structures in insect olfactory sensilla: a review of data and concepts[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF]. We found numerous pores on the cuticle of multiporous basiconic sensilla under SEM examination (Fig. 5B1), supporting an olfactory function of this type of sensillum. MBS are confined to the lateral-ventral side of the flagellum in the four species, suggesting they are used to detect fig surface chemicals when females contact the fig surface with their antennae [START_REF] Moore | Habitat-specific signal structure for olfaction: an example from artificial streams[END_REF][START_REF] Wang | The mechanism of pollinator specificity between two sympatric fig varieties: a combination of olfactory signals and contact cues[END_REF]. In agreement with this speculation, MBS are most abundant in C. emarginatus and Sycophaga sp., the two species that consistently touch the surface of the fig with the whole length of the flagellum.

Multiporous placoid sensilla are widely distributed on the antennae of chalcid wasps [START_REF] Barlin | Multiporous plate sensilla in antennae of the Chalcidoidea (Hymenoptera)[END_REF][START_REF] Basibuyuk | Gross morphology of multiporous plate sensilla in the Hymenoptera (Insecta)[END_REF][START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF][START_REF] Ware | Repeated evolution of elongate multiporous plate sensilla in female fig wasps(Hymenoptera: Agaonidae: Agaoninae)[END_REF]. Usually, MPS are elongate and sausage-shaped and present ring-like arrangements around the flagellomere except for the funicle and the basal flagellomere (F1) [START_REF] Heraty | A phylogenetic analysis of the megadiverse Chalcidoidea (Hymenoptera)[END_REF]. Their multiporous surface system indicates that they are olfactory receptors [START_REF] Bleeker | Antennal sensilla of two parasitoid wasps: a comparative scanning electron microscopy study[END_REF][START_REF] Steinbrecht | Pore structures in insect olfactory sensilla: a review of data and concepts[END_REF]. [START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF] reported that MPS are dose-dependent sensors that perceive the volatiles of host plants. The exposed surface areas of these sensilla in female fig wasps, including NPFWs in this study, are larger than that of other chalcid female wasps [START_REF] Barlin | Multiporous plate sensilla in antennae of the Chalcidoidea (Hymenoptera)[END_REF][START_REF] Ware | Repeated evolution of elongate multiporous plate sensilla in female fig wasps(Hymenoptera: Agaonidae: Agaoninae)[END_REF] and suggests that the detection of host figs is particularly challenging and involves detecting speciesspecific chemicals at a distance. However, the specific function and sensitivity of MPS in fig wasp foundresses must be further explored via single-sensillum recordings.

BCPS have been reported in many chalcid wasp species belonging to a diversity of families, and they are referred to as multiporous grooved peg sensilla on the eulophid Tetrastichus hagenowii (Ratzeburg, 1852) [START_REF] Barlin | Multiporous plate sensilla in antennae of the Chalcidoidea (Hymenoptera)[END_REF][START_REF] Ware | Repeated evolution of elongate multiporous plate sensilla in female fig wasps(Hymenoptera: Agaonidae: Agaoninae)[END_REF], type I coeloconic sensillum on the braconid Cotesia species [START_REF] Bleeker | Antennal sensilla of two parasitoid wasps: a comparative scanning electron microscopy study[END_REF], type s4 sensilla on the mymarommatoids [START_REF] Heraty | A phylogenetic analysis of the megadiverse Chalcidoidea (Hymenoptera)[END_REF], sensilla ampullaceum on the trichogrammatids [START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF], and peg-like sensillum on the eulophid Sympiesis sericeicornis (Nees, 1834) [START_REF] Meyhöfer | Mechano-and chemoreceptors and their possible role in host location behavior of Sympiesis sericeicornis (Hymenoptera: Eulophidae)[END_REF]. These sensilla, which occur in low numbers on the antennae of all species, are short bulbous pegs with multiporous furrows as revealed by TEM [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF]. These sensilla can be olfactory receptors [START_REF] Barlin | Multiporous plate sensilla in antennae of the Chalcidoidea (Hymenoptera)[END_REF][START_REF] Bleeker | Antennal sensilla of two parasitoid wasps: a comparative scanning electron microscopy study[END_REF][START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF][START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF][START_REF] Steinbrecht | Pore structures in insect olfactory sensilla: a review of data and concepts[END_REF], although they may also present mixed functions, such as hygro-, thermo-and mechanoreceptor capabilities [START_REF] Meyhöfer | Mechano-and chemoreceptors and their possible role in host location behavior of Sympiesis sericeicornis (Hymenoptera: Eulophidae)[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF]. Their location in fig-wasps suggest that they may have a role as proprioreceptive relative position detectors [START_REF] Dweck | Antennal sensory receptors of Pteromalus puparum female (Hymenoptera: Pteromalidae), a gregarious pupal endoparasitoid of Pieris rapae[END_REF][START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF] for the segments of the flagellum.

In other parasitic wasps, the three types of CoS that occur together in C. emarginatus are rare and have only been observed in another species belonging to genus Ceratosolen [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF].

To our knowledge, this study provides the first record of CoS-1, which does not resemble any CoS on other insects [START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF][START_REF] Zacharuk | Antennae and sensilla[END_REF]. The CoS protrusion is strongly grooved along its length, which may be an indicator of surface pores that allow odour molecules to reach receptors [START_REF] Ameismeier | Embryonic development and molting of the antennal coeloconic no poreand double-walled wall pore sensilla in Locusta migratoria (Insecta, Orthopteroidea)[END_REF][START_REF] Steinbrecht | Pore structures in insect olfactory sensilla: a review of data and concepts[END_REF], suggesting that these sensilla may be involved in volatile perception. However, the low numbers of CoS-1 may restrict the sensory ability (Table 1). The non-porous CoS-2 and CoS-3 are always mixed together. CoS-2 is similar to the auriculate sensillum in the female sphinx moth Manduca sexta (Linnaeus, 1763) [START_REF] Shields | Fine structure of antennal sensilla of the female sphinx moth, Manduca sexta (Lepidoptera: Sphingidae). II. Auriculate, coeloconic, and styliform complex sensilla[END_REF], the sensilla coeloconica in the mushroom fly Coboldia fuscipes (Meigen, 1830) [START_REF] Zhang | Morphology, distribution and abundance of antennal sensilla of the oyster mushroom fly, Coboldia fuscipes (Meigen) (Diptera: Scatopsidae)[END_REF], and the CoS-2 in the pollinating fig wasp C. solmsi marchali [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF], whereas CoS-3 resembles the single-walled poreless CoS in some Lepidoptera [START_REF] Faucheux | The antennae of neopseustid moths: morphology and phylogenetic implications, with special reference to the sensilla (Insecta, Lepidoptera, Neopseustidae)[END_REF] and CoS-3 in C. solmsi marchali [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF]. To enter into the fig cavity, foundresses always use the spine-like expansion as a tool for lifting the outer ostiolar bracts (Weiblen, 2002, Fig. 2 A4). Therefore, these surface sensilla could intervene as detectors during ostiolar entry. CoS-2 are likely to be involved in proprioreceptive functions [START_REF] Meyhöfer | Mechano-and chemoreceptors and their possible role in host location behavior of Sympiesis sericeicornis (Hymenoptera: Eulophidae)[END_REF][START_REF] Staudacher | Antennal movements and mechanoreception: Neurobiology of active tactile sensors[END_REF], whereas CoS-3 may sense the pressure of fig bracts [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Pophof | Olfactory responses recorded from sensilla coeloconica of the silkmoth Bombyx mori[END_REF].

No equivalent of SO has been reported from non-agaonid wasps [START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF][START_REF] Zacharuk | Antennae and sensilla[END_REF], and they could be generalised in pollinating fig wasps [START_REF] Cockerill | External Morphology of Antennal Sensilla in Relation to the Host Searching Behaviour of Liporrhopalum Tentacularis Grandi (Hymenoptera: Agaonidae)[END_REF][START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF][START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF]. The shape of SO varies among species. They are leaf-shaped in C. solmsi marchali [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF], teardrop-shaped in Liporrhopalum tentacularis (Grandi, 1926) [START_REF] Cockerill | External Morphology of Antennal Sensilla in Relation to the Host Searching Behaviour of Liporrhopalum Tentacularis Grandi (Hymenoptera: Agaonidae)[END_REF], and are present as trichoid hairs with a bifurcated apex in Waterstoniella sp. [START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF]. Their internal structure is a thick and non-porous wall [START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF]. SO are distributed dorsally in a patch on the pedicel and always point toward the wasp's head. Their functions could be to provide a grip on the ostiolar scales similar to the function of the teeth on the mandibular appendages that help the wasps get through the ostiole [START_REF] Weiblen | How to be a fig wasp[END_REF].

We may summarise an interpretation of the functional organisation of the female antennae of the study species in the following way. The wasps need proprioreceptors enabling them to monitor antenna position. This role is filled by ChS-2 for the basal parts of the antennae and could be filled by BCPS for the flagellum. The principal distance olfactory detection function of the antennae is probably filled by MPS which represent the majority of the receptors that are distributed all along the flagellum. Localised contact chemical stimulation when wasps are tapping the surface of a fig with the tip of their antennae is probably fulfilled by UBS. All along the flagellum, interspersed between the MPS, longer sensilla may deflect obstacles thus protecting BCPS, and allowing the wasps to sense the strength of contact with a surface or an obstacle and allow contact chemical stimulation. These roles are filled by ChS-1, TS and MBS. The well-developed articulated sockets of TS suggest that these sensilla provide finer information on contact than ChS-1. In agreement with this proposition, in C. emarginatus and Sycophaga sp., the two species that contact the surface of the fig with the whole length of the antennae, there are abundant TS and a total lack of ChS-1. The lateral-ventral disposition of MBS suggest that they are involved in contact chemo-stimulation and they are also much more abundant in C. emarginatus and Sycophaga sp. than in P. longicaudata and S. roxburghi. Finally, a whole set of sensilla are restricted to the first three antennal segments of the pollinating wasps suggesting that they are associated with various functional aspects of fig-entering behaviour.

Within this general framework, variation is observed. For instance, Kradibia tentacularis (Grandi, 1926) pollinating F. montana presents exceptionally long antennae and BCPS are present in places that can hardly be linked with a proprioreceptor function [START_REF] Cockerill | External Morphology of Antennal Sensilla in Relation to the Host Searching Behaviour of Liporrhopalum Tentacularis Grandi (Hymenoptera: Agaonidae)[END_REF]. Data on Waterstoniella sp. associated with F. curtipes shows lack of CoS-1 [START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF], a feature that could correlate with the simpler constitution of the tooth-like expansion of the third antennal segment in Waterstoniella comparatively to Ceratosolen. We observed large variation among species in numbers of sensilla. For instance, in fig-pollinators numbers vary from 738 sensilla including 406 TS in C. solmsi marchali [START_REF] Mancini | Chemical composition and phytotoxic effects of essential oils of Salvia hierosolymitana Boiss. and Salvia multicaulis Vahl. var. simplicifolia Boiss. growing wild in Lebanon[END_REF], through 564 sensilla including 346 TS in C. emarginatus, and down to 209 sensilla including 77 TS in Waterstoniella sp. [START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF].

Morphological adaptations and differences among four agaonid species

For female pollinating fig wasps, the modified first three antennal segments and especially the tooth on the third antennal segment and the multiple sensillum types on the surface represent conspicuous antennal adaptations that facilitate pollinator passage through the fig ostiole. The challenge is probably largely mechanical, but could involve sensing ostiolar exudates [START_REF] Machado | Secretory structures at syconia and flowers of Ficus enormis (Moraceae): a specialization at ostiolar bracts and the first report of inflorescence colleters[END_REF], ostiolar firmness [START_REF] Chen | Regulation of fig wasps entry and egress: the role of ostiole of Ficus microcarpa L[END_REF], and diverse odours released by glands located in the ostiolar scales [START_REF] Souza | Diversity of fig glands is associated with nursery mutualism in fig trees[END_REF]. SO that are grouped dorsally on the pedicel and directed toward the head serve a mechanical function that prevents the wasps from slipping backward when foundresses pass through the ostiolar bracts and may have a function similar to that of the teeth on the mandibular appendage, which the wasps use to pull themselves through the ostiole using similar opening and closing mechanisms [START_REF] Van Noort | Convergent evolution of agaonine and sycoecine (Agaonidae, Chalcidoidea) head shape in response to the constraints of host fig morphology[END_REF]. Interestingly, mandibular appendages and their teeth often present large differences among species, suggesting adaptation to variation in ostiole shape. Similarly, SO present large variation among species. If our hypothesis is true then we may predict limited development of SO in fig pollinating wasps, such as the extant species of genus Tetrapus, that loose neither their antennae nor their wings on entry into a fig, because of an easily penetrated ostiole.

The abundance and distribution of the mechanoreceptors such as ST and ChS-1 on the flagellum could also be related to fig surface properties. As the fig develops and grows in size, the surface also shifts from a soft to a hard surface with long surface hair. Hence, an alternative explanation to the higher number of ST present on early-arriving wasp species could be the association with softer figs than for the late-arriving wasp species associated with harder figs. Conversely, ChS-1 are regularly observed in the females of NPFWs, which lay eggs in figs that are more advanced in development (Fig. 6; Table 2). This result is consistent with what is observed for ST on the antennae of herb-and wood-galling cynipids [START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF], or female fig wasps in F. hispida [START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF], which may indicate that the rigid ChS-1 provide better protection to chemoreceptors below. But this explanation is only applicable to parasites that feed on plant tissue on the outside of the host or have a similar lifehistory trait [START_REF] Li | Antennal structure, sensilla and their ecological implication of three internally ovipositing female parasites in Ficus curtipes[END_REF][START_REF] Meng | Antennal sensilla of fig wasps (Hymenoptera: Agaonidae): function-driven elaboration in females and degeneration in males[END_REF][START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF].

The variation in the surface coverage area of MPS in the females of the four wasp species could be the result of variation in size and in behaviour. [START_REF] Ware | Repeated evolution of elongate multiporous plate sensilla in female fig wasps(Hymenoptera: Agaonidae: Agaoninae)[END_REF] reported that there are two distinct adaptative strategies that may increase receptor surface. The first strategy is MPS elongation which increases the receptor-exposed surface area, which in turn enhances olfactory sensitivity. MPS in the late-arriving foundresses, including P. longicaudata and S. roxburghi, are typically more elongate than those of other females belonging to the same family of wasps [START_REF] Barlin | Multiporous plate sensilla in antennae of the Chalcidoidea (Hymenoptera)[END_REF][START_REF] Dweck | Antennal sensory receptors of Pteromalus puparum female (Hymenoptera: Pteromalidae), a gregarious pupal endoparasitoid of Pieris rapae[END_REF][START_REF] Onagbola | Scanning electron microscopy studies of antennal sensilla of Pteromalus cerealellae (Hymenoptera: Pteromalidae)[END_REF]. The second strategy is increasing the number of sensilla and as a result increasing the exposed area by lengthening the antennae and thickening antennal sub-segments. In our case, Sycophaga sp. has a larger antenna than the other inverstigated wasps and presents more rings of MPS with a larger ring arrangement (Fig 3;Fig 8). This feature can be explained by the larger body size of Sycophaga sp. compared to the three other species which also translates into larger antennae (Table 1). However, because young figs produce limited quantities of odours [START_REF] Proffit | Signalling receptivity: comparison of the emission of volatile compounds by figs of Ficus hispida before, during and after the phase of receptivity to pollinators[END_REF], the associated larger MPS surface area of Sycophaga sp. may help detect small amounts of fig chemicals. On the other hand among the three similarly-sized wasps, and despite receptive figs producing more odours than other fig stages [START_REF] Proffit | Signalling receptivity: comparison of the emission of volatile compounds by figs of Ficus hispida before, during and after the phase of receptivity to pollinators[END_REF], the pollinator presented a 1.5 times larger surface area of MPS than the two other NPFWs. We may suggest that the reason is that NPFWs have longer life spans and thus can spend a greater amount of time searching for stage-specific figs in the natural environment [START_REF] Compton | African fig wasp parasitoid communities[END_REF]. However, olfactory receptors are extremely sensitive and can be triggered by less than six molecules of host plant volatiles [START_REF] Angioy | Extreme sensitivity in an olfactory system[END_REF], indicating that further study using single-sensillum recordings may provide insights that can explain this adaptation and sensory sensitivity.

In conclusion, females of four fig wasp species associated with figs of F. auriculata may use mechanoreceptors and chemoreceptors to locate and discriminate stage-specific figs. In addition to identifying the spine-like expansion and SO related to ostiole entry, this study also revealed that three types of CoS are likely involved in sensing the interior properties of figs. Two mechanosensory sensilla, including TS and ChS-1, occurred on the flagellum, and inconsistent results were observed for species laying their eggs on early soft figs and late hard figs. The chemosensilla involved in host discrimination, i.e., basiconic sensilla, multiporous placoid sensilla and basiconic capitate peg sensilla, present similar morphologies and configurations and provide a sensory infrastructure for our ongoing studies of host discrimination among these four species, including behavioural interpretations and electrophysiological studies. Further studies on the functional morphology of the antennal sensilla of the four foundresses using

Fig

  Fig wasps (Hymenoptera: Chalcidoidea) are chalcid wasps that oviposit almost exclusively in fig ovules. Many of these wasp species develop on a single Ficus species[START_REF] Cook | Mutualists with attitude: coevolving fig wasps and figs[END_REF][START_REF] Weiblen | How to be a fig wasp[END_REF]. They locate their host using stage-specific chemical cues emitted by the figs of their host species[START_REF] Hossaert-Mckey | Floral scents: their roles in nursery pollination mutualisms[END_REF][START_REF] Proffit | Chemical mediation and niche partitioning in non-pollinating fig-wasp communities[END_REF][START_REF] Proffit | Signalling receptivity: comparison of the emission of volatile compounds by figs of Ficus hispida before, during and after the phase of receptivity to pollinators[END_REF][START_REF] Wang | The mechanism of pollinator specificity between two sympatric fig varieties: a combination of olfactory signals and contact cues[END_REF]. In insects, the antenna is an organ capable of detecting odours. It is a multisensory detector carrying various receptors that sense a series of signals, including smell, but also taste, temperature, sound, gravity, wind speed, and even the geomagnetic field[START_REF] De Oliveira | Ant antennae: are they sites for magnetoreception?[END_REF][START_REF] Isidoro | Morphology of antennal gustatory sensilla and glands in some parasitoids Hymenoptera with hypothesis on their role in sex and host recognition[END_REF][START_REF] Meyhöfer | Mechano-and chemoreceptors and their possible role in host location behavior of Sympiesis sericeicornis (Hymenoptera: Eulophidae)[END_REF][START_REF] Ochieng | Functional morphology of antennal chemoreceptors of the parasitoid Microplitis croceipes (Hymenoptera: Braconidae)[END_REF][START_REF] Romani | Antennal structures used in communication by egg parasitoids[END_REF][START_REF] Staudacher | Antennal movements and mechanoreception: Neurobiology of active tactile sensors[END_REF][START_REF] Van Baaren | Antennal sensilla of hymenopteran parasitic wasps: variations linked to host exploitation behavior[END_REF][START_REF] Zacharuk | Antennae and sensilla[END_REF]. The extent to which variation in the structure of the antennae as well as the presence and distribution of different types of sensory receptors among wasp species can be explained by differences in behaviour and oviposition strategies are poorly known (but see[START_REF] Polidori | Diverse filters to sense: Great variability of antennal morphology and sensillar equipment in gall-wasps (Hymenoptera: Cynipidae)[END_REF]. For fig wasps, the few studies of the sensilla on the antenna have almost exclusively focused on pollinators. These wasps enter figs to oviposit and the host figs are specialized to attract them[START_REF] Li | Ultrastructure of antennal sensilla of female Ceratosolen Solmsi Marchali (Hymenoptera: Chalcidoidea: Agaonidae: Agaoninae)[END_REF][START_REF] Li | Ultrastructure and distribution of sensilla on the antennae of female fig wasp Eupristina sp. (Hymenoptera: Agaonidae)[END_REF][START_REF] Ware | Repeated evolution of elongate multiporous plate sensilla in female fig wasps(Hymenoptera: Agaonidae: Agaoninae)[END_REF]. Little is known about the sensory organs on

  Fig. 1. Oviposition sequence of four fig wasp species associated with F. auriculata figs along the different development stages (A-E) of figs. The dashed lines show the first and the last arrival of wasps landing on the fig surface. The solid lines show the range of oviposition time.

Fig. 2 .

 2 Fig. 2. Females of the four wasp species landing or ovipositing from outside the figs of F. auriculata. (A) Fixed sequence of behavioural events in the pollinator C. emarginatus, including holding the antennae upward (A1), lowering the antennae (A2), tapping the fig surface with the tip of the final flagellomere by curving the antenna downwards (A3), and lifting the ostiolar bracts with the modified spine-like expansion of the third antennal segment (A4). The bottom pictures show aggregative oviposition by the three nonpollinators, Sycophaga sp. (B), P. longicaudata (C), S. roxburghi (D).

Fig. 3 .

 3 Fig. 3. Female antennae of four fig wasps associated with F. auriculata. The figure shows SEM graphs of excised geniculate antennae of females of the four fig wasp species C. emarginatus (A), Sycophaga sp.(B), P. longicaudata (C), and Sycoscpater roxburghi (D) and depicts the Radicula (Rd), Scape (Sc), Pedicel (Pe), and Flagellum (Fl), which is subdivided into the Anellus (An), Funicle (Fu) and Clava (Cl). Important morphological differences among the four species are indicated with a solid arrow (flagellomere one (F1) base with a spine-like expansion), a dashed loop (sensilla obscura on the dorsal side of the pedicel), an asterisk (deep socket), and a dashed arrow (radicula). Scale bar = 100 μm.

Fig. 4 .

 4 Fig. 4. Different types of antennal sensilla recorded on four fig wasp species associated with F. auriculata. (A) Micrograph of the tip flagellomere of P. longicaudata showing the trichoid sensilla (TS), chaetica sensilla type 1 (ChS-1), uniporous (UBS) and multiporous basiconic sensilla (MBS), and multiporous placoid sensilla (MPS) from the dorsal view. The UBS tips are well above the other sensilla (the arc). (B) Distribution of different sensilla, including basiconic capitate peg sensilla (BCPS), occurring on the tip of the clava of C. emarginatus. (C) Basal segment of antennae of C. emarginatus shows the specific sensory organs, including the sensilla obscura (SO) and chaetica sensilla type 2 (ChS-2). The dashed loop shows the mixed distribution of coeloconic sensilla 2 and 3. (D) Spine-like expansion detached from the base of first flagellomere (F1); the dashed arrow shows the enlarged image of the spinelike expansion with coeloconic sensilla 2 and 3 (pits on the surface, D1). Scale bars = 10 μm

Fig. 5 .

 5 Fig. 5. Fine structure of different female antennal sensilla of the four fig wasp species associated with F. auriculata. (A) Uniporous basiconic sensilla (UBS) with a blunt tip (Po) occurred on the terminal part of the clava of the antennae of Sycophaga sp. Other sensilla are also labelled in the figures. (B) Multiporous basiconic sensilla (MBS) with many surface pores (B1) on the terminal claval segment of P. longicaudata. (C) Multiporous placoid sensilla (MPS) on C. emarginatus. (D) Chaetica sensilla type 1 (ChS-1) on P. longicaudata. (E) Trichoid sensilla (TS) with longitudinal grooves insert into a flexible socket (E1) on Sycophaga sp. (F) Chaetica sensilla type 2 (ChS-2) on Sycophaga sp. (G) Sensilla obscura (SO) with longitudinal grooves inserts into an oval socket. (H) Basiconic capitate peg sensilla (BCPS) on P. longicaudata. (I) Coeloconic sensilla type 1 (CoS-1). (J) Coeloconic sensilla type 2 (CoS-2). (K) Coeloconic sensilla type 3 (CoS-3). Scale bar = 2 μm

Fig. 6 .

 6 Fig. 6. Schematic diagrams of the distribution of 11 types of sensilla on the antennae of the four fig wasp species associated with F. auriculata. (A) Dorsal-ventral side of the whole antennae of C. emarginatus. (A1) Dorsal side of the clava on C. emarginatus. (A2) Dorsal/ventral (A3) side of the scape and pedicel, as well as one flagellomere shape on C. emarginatus. (B-D) Lateralventral side of the whole antennae of Sycophaga sp. (B), P. longicaudata (C) and S. roxburghi. (D), respectively. (C1) Ventral side of the second claval flagellomere on P. longicaudata. (D1) Enlarged drawing of the scape and pedicel on S. roxburghi

  Fig 3C; Fig 8). The abundance and distribution of BCPS are not significantly different among the four species. TS/ChS-1, MPS, and BCPS always occur concurrently on the same flagellomere, whereas both CoS and SO only occur on the antennae of the pollinator C. emarginatus that enters the receptive figs (Fig. 6). The abundance and distribution as well the configuration of ST, ChS-1, MBS, MPS, SO, and CoS may indicate the occurrence of morphological adaptations and sensory differentiation among the four species.

Fig. 7 .

 7 Fig. 7. Number of the three major types of sensilla on the flagellum of females of the four fig wasps associated with F. auriculata. TS: trichoid sensilla; MPS: multiporous placoid sensilla; ChS: chaetica sensilla.

Fig. 8 .

 8 Fig. 8. Measurements and total surface area of multiporous placoid sensilla on the flagellum of females of four fig wasp species associated with F. auriculata.

  transmission electron microscopy coupled with electrophysiological techniques are likely to yield a deeper understanding of the differences in chemical utilization and olfactory sensitivity in the fig wasp community. Comparative studies of antennal structure variation among fig hosts within genera of NPFW and within pollinating figs wasp genera could probably help identify causal ecological factors for inter-specific differences.Author contributions PY, ZBL and YQP conceived and designed the study. PY and ZBL performed the experiments and the data analyses. PY and ZBL wrote the first draft of the manuscript after having discussed the results with YQP, DRY and FK. All author contributed to revisions and approved the final manuscript.

Acknowledgements

We gratefully acknowledge Prof. Jean-Yves Rasplus (Centre de Biologie pour la Gestion des Populations, INRA, France) for identifying fig wasps. We also thank Dr Li Wang (Xishuangbanna Tropical Botany Garden, CAS, China) for technical assistance and use of scanning electron microscopy. We also appreciate the support of the Central Laboratory of Xishuangbanna Tropical Botanical Garden. This research was supported by the National Natural Foundation of China (Nos. 31200292, 31100279, 31760107, 31611130108), the West Light Foundation of the Chinese Academy of Sciences, and the Natural Science Foundation of Yunnan Province (No. 2014FB183), and Program for Innovative Research Team (in Science and Technology) in the University of Yunnan Province (No. 51600507).