The cabbage armyworm, Mamestra brassicae L. (Lepidoptera: Noctuidae), is a highly polyphagous insect. It is mainly recognized as a pest of cabbage (Brassica oleracea L.), but it has also been reported from a wide range of other plant species (Balachowsky 1972). Its known food plants include >70 species of 22 families, of which Brassicaceae and Chenopodiaceae are among the most preferred (Popova 1993). Several aspects of M. brassicae biology have been studied extensively, such as courtship and mating behavior (Birch et al. 1989, Noldus and Potting 1990, Poppy and Birch 1994) and oviposition behavior (Rojas et al. 2001). To characterize the chemical orientation of M. brassicae toward its host products and mates, it is necessary to know the structure of the olfactory system of the moth.

Insect antennae play an important role in detection of various external stimuli involved in finding suitable habitat and locating mates (Baker et al. 2004, Pophof et al. 2005, Sun et al. 2011). In insects, the antenna is suggested to be the principal organ carrying the olfactory receptors (Callahan 1975, Lavoie-Dornik and McNeil 1987, Schneider 1964).

Numerous studies have been conducted to characterize antennal sensilla of various insects (Bleeker et al. 2004, Faucheux et al. 2006, Keil and Steinbrecht 1984, Liu et al. 2013, Städler 1984, Zacharuk 1980), but very few of such studies have been focused on Noctuidae (Chang et al. 2015, Hu et al. 2021, Koh et al. 1995, Malo et al. 2004, Mochizuki et al. 1992, Seada 2015). The antennal sensilla of M. brassicae have not been studied despite the important role they play in the life of the insect. In this study, as part of our ongoing research on mate and host location mechanisms of M. brassicae, we used scanning electron microscopy (SEM) techniques to characterize and determine the abundance and distribution of the antennal sensilla of this cabbage moth. This work is a prerequisite for future electrophysiological studies of the antennal sensory system involved in intraspecific chemical communication.

Materials and Methods

Results

Types of sensilla

Based on their morphological features, we identified sensilla chaetica (SCH; subtypes 1 and 2), sensilla trichodea (ST; subtypes 1, 2, and 3), sensilla basiconica (SB; subtypes 1 and 2), sensilla styloconica (STY), Böhm bristles (BB), sensilla coeloconica (SCO), and sensilla squamiformia (SSQ). The morphological features and lengths (mean ± SD) of each antennal sensillum type are listed in Table 1.

Table 1. Length, basal diameter, and morphological characteristics of different types of antennal sensilla of M. brassicae (n = 10 per sex).

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SCH1

Each sensillum was straight, wide at the basal part, and slightly curved at the distal part where the articulation was countersunk into a central socket enclosed by a bulge (Fig. 1A). SCH1 had circumferential grooves on their surfaces (Fig. 1B). Lengths were 86.54 ± 8.47 μm (female) and 83.41 ± 8. 68 μm (male), and basal diameters were 3.35 ± 0.41 μm (female) and 3.78 ± 0.43 μm (male).

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ST1

ST1 were hairs with slightly sharp tips and a helical wall and were nearly parallel to the antenna surface or slightly curved toward the antennal shaft (Fig. 2A). They became progressively more pointed from the base onward. Lengths were 44.69 ± 9.77 μm (male), and basal diameters were 1.78 ± 0.45 μm (male).

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ST3

High magnification images of these sensory hairs revealed that they were curved along the longitudinal axis, with a helical surface texture and a blunt tip (Figs. 2D, 3A). Lengths were 28.54 ± 4.79 μm (female) and 31.69 ± 6.05 μm (male), and basal diameters were 1.59 ± 0.27 μm (female) and 1.57 ± 0.27 μm (male).

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SB2

The SB2 inserted into a shallow socket in the integument and were obliquely oriented to the antennal surface (Fig. 3D). Oblique and longitudinal ridges covered the surface of the sensillum, forming almost vertical rows near the base and developed into a herringbone pattern over the distal half. Pores lined the interstices between the ridges (Fig. 4A). Lengths were 16.29 ± 3.50 μm (female) and 17.42 ± 2.57 μm (male), and basal diameters were 1.35 ± 0.38 (female) and 1.30 ± 0.23 μm (male).

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SCO

SCO were small sensilla, commonly called pit pegs. A sensillum was projected from the center of a shallow cuticular pit (Fig. 4D). A fringe of 14–16 teeth could be seen along the edge of the pit. SCO were clumped at the ventral side of the flagellum (Fig. 5A).

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Discussion

The general shape and structure of the antennae of male and female M. brassicae are similar to those reported for other species of Lepidoptera (Bawin et al. 2017, Castrejón-Gómez et al. 1999, Koh et al. 1995, Zheng et al. 2014, Zheng et al. 2023). The antennal flagellomeres of males and females are cylindrical and most of the sensilla are located on the ventral surface, whereas the dorsal surface possesses numerous overlapping scales (Fig. 5D). Functions of the antennal scales have been described as protecting the antenna (Koh et al. 1995), contributing to the detection of stimulus direction (Van der Pers et al. 1980), and trapping and concentrating odorous molecules (Wall 1978); however, these hypotheses have not been tested. Based on morphological criteria and according to the published scientific reports, we inferred the function of the sensilla in M. brassicae described here.

SCH1 are similar in shape to those reported for Spodoptera exigua Hübner (Jefferson et al. 1970), Noctua pronuba L. (Faucheux 1990), and Catocala remissa Staudinger (Zheng et al. 2014) but differ in length. SCH2 are similar in morphology and location to those reported on Spodoptera frugiperda (J.E. Smith) (Malo et al. 2004), Synanthedon scitula (Harris) (Frank et al. 2010), and Nyctemera annulata Boisduval (Mark et al. 2018). Several researchers have suggested that SCH have both a contact chemo- and mechanoreceptive function because they arise from a socket and possess a terminal pore (Altner and Prillinger 1980, Van der Pers et al. 1980). Jørgensen et al. (2007) noted that SCH of Heliothis virescens (F.) possess one mechanosensory neuron and four gustatory receptor neurons that aid in the detection and discrimination of noxious and edible food sources. A similar function seems likely in M. brassicae, particularly because SCH1 and SCH2 tend to extend beyond the antennal surface, thus allowing increased contact with the substrate. In studies of sesiid species, the presence of a terminal pore has not been documented, and the primary function of these sensilla was considered mechanoreceptive (Karalius 1994, Sellier 1977). Although the presence of a terminal pore on SCH2 of the M. brassicae could not be confirmed, the behavior of females suggests that the role of these sensilla may involve both mechano- and contact chemoreception.

ST1 are longer than the other ST subtypes and are present on only male antennae in M. brassicae. This sensillum is similar in structure to those reported for other moths (Castrejón-Gómez et al. 2003, Frank et al. 2010, Ndomo-Moualeu et al. 2014). In codling moths, the long ST houses highly sensitive pheromone-detecting olfactory receptor neurons (Ebbinghaus et al. 1997). In several moth species, the long ST on male antennae responds to female sex pheromones (Baker et al. 2004, Cossé et al. 1998, Faucheux et al. 2006, Hansson 1995). Thus, the ST in M. brassicae also might be responsible for the reception of sex pheromones. The response of ST1 to conspecific female sex pheromones in M. brassicae could be studied via single sensillum recording.

ST2 resemble “sensilla trichodea I” in two tortricid moths, Cydia pomonella (L.) and Cydia succedana (Denis and Schiffermüller) (Roh et al. 2016) and in the defoliating moth, Phauda flammans (Walker) (Liu et al. 2018). ST3 located on the flagellomere of M. brassicae are very similar to the “ridgeless-type trichoidea” in the beet armyworm, S. exigua (Mochizuki et al. 1992) and the “sensilla trichoidea Type III” of C. remissa (Zheng et al. 2014). Sensilla trichodea subtypes II and III were more abundant in females than in males. We do not know at present the functions of ST2 and ST3 on female antenna of M. brassicae. Subtypes of females may be able to detect their own sex pheromones and plant volatile compounds for female-specific behaviors. For example, antennae on females of Spodoptera littoralis (Boisduval) (Ljunberg et al. 1993) and Trichoplusia ni (Hübner) (Birch 1977) responded to conspecific sex pheromones. The short ST of female antennae in S. littoralis and Cactoblastis cactorum (Berg) (Lepidoptera: Pyralidae) detected plant volatiles (Anderson et al. 1995, Pophof et al. 2005). Functions of ST2 and ST3 needed to be studied in regard to responses of the females to their own sex pheromones and host plant volatiles.

Two types of SB were identified on both sexes in this study. SB1 were found on flagellomeres of M. brassicae as in other moths (Faucheux 1999, Faucheux et al. 2006, Razowski and Wojtusiak 2004, Sun et al. 2011). The SB1 were lined up longitudinally in M. brassicae (our study) and in moths of the Oecophoridae (Faucheux 1978), Tortricidae (Cuperus 1985), and Pyralidae (Faucheux 1991). SB2 in M. brassicae appear to be identical to the “sensilla basiconica type 4” in N. annulata (Mark et al. 2018). SB are important olfactory setae that are structurally adapted to the recognition of plant odors in phytophagous insects (Laue et al. 1994, Schneider 1964, Steinbrecht et al. 1995). Scanning and transmission electron microscopy studies of SB in the coleopteran Phoracantha semipunctata (F.) (Cerambycidae) revealed such structure, and single sensory cell recordings revealed responses to plant odors with no difference between males and females (Lopes et al. 2002). In other studies, olfactory neurons housed in SB responded to plant-associated odorants in S. littoralis (Binyameen et al. 2012) and Manduca sexta (L.) (Ghaninia et al. 2014). The surface of SB in this study (Fig. 3C) was multiporous, suggesting that SB are olfactory receptors used to detect volatile plant odors (Pophof et al. 2005).

STY of M. brassicae possess several general features in common with STY of many other lepidopterans, namely, the presence of a stylus and a sensory cone and the distal distribution on the lateral branches (Faucheux 1997, 1999). These sensilla were considered thermo- and hygrosensitive in Bombyx mori (L.) (Steinbrecht 1989) and members of the Pyralidae (Hallberg et al. 1994, Hansson 1995); however, the presence of an apical pore in the Tortricidae suggested a contact chemoreceptive function (Castrejon-Gomez and Carrasco 2008, Wall 1978). Aporous STY have been demonstrated to be thermo- and hygroreceptors containing two antagonistic hygroreceptive cells and one thermoreceptive cell responding to cold in B. mori (Steinbrecht 1989, Steinbrecht and Müller 1991) and Antheraea species (Gödde and Haug 1990, Haug 1985). An apical pore was not found on STY of M. brassicae (Fig. 4B), suggesting that these STY may have a thermo- and hygrosensory function.

BB of M. brassicae antennae are similar in morphology to those on the scape and pedicel of Micropterix calthella L. (Faucheux 1997), Clostera anastomosis L. (Liu et al. 2013) and Plodia interpunctella (Hübner) (Ndomo-Moualeu et al. 2014). BB location is as reported by Schneider (1964): “a typical bristle is always found in areas opposite the intersegmental membrane between head and scape, as well as between scape and pedicel on the scape and pedicel, respectively.” The functions attributed to these sensilla are ambiguous. Schneider (1964) suggested that these hair plates may play a mechanosensitive role. Results of recent studies suggest that they mediate antennal positioning in moths during flights (Krishnan et al. 2012). The BB located in the pedicel respond to changes in location and vibration of the whole antenna, whereas those located near the scales–pedicel boundary transfer information about the position of the antenna (rather than its movement) to the brain (Krishnan et al. 2012). Some studies of these sensilla on Cnaphalocrocis medinalis (Guenée) (Lepidoptera: Pyralidae) revealed sporulated walls, suggesting an olfactory function (Sun et al. 2011). Therefore, further research, especially single sensilla recordings, are needed with the BB in M. brassicae to determine their precise function.

The SSQ in M. brassicae also are frequently found in many lepidopterans, where these sensilla are distributed among the lamellar scales (Faucheux 1999). Typically, this sensilla type is characterized by slender lamellar scales (Fig. 5B), although some variation in morphology exists (e.g., Liu et al. 2013, Yang et al. 2009). SSQ observed in M. brassicae are of the more conventional form and bear particular resemblance to those sensory structures seen in male Zamagiria dixolophella Dyar (Castrejón Gómez et al. 2003) and in a number of skipper butterflies (Yuan et al. 2014). SSQ in M. brassicae are similar in morphology and location to those reported on other moth species such as P. interpunctella (Ndomo-Moualeu et al. 2014) and Z. dixolophella (Castrejón Gómez et al. 2003). Sexual dimorphism of SSQ has been described in some species (e.g., Castrejón Gómez et al. 2003, Zheng et al. 2014), although the functional significance of this dimorphism is unknown. No sexual dimorphism was apparent in the morphology of SSQ in M. brassicae; however, because the SSQ density could not be estimated accurately, a difference between males and females cannot be ruled out. SSQ are thought to have a mechanoreceptive function (Faucheux 1999), and their location on the dorsal side of the antennal spindle in M. brassicae suggests that in this species they likely have the same function.

SCO present on the antenna of M. brassicae are similar to those described in members of the Pyralidae such as Hypsipyla grandella (Zeller) (Callahan 1975), Homoeosoma nebulella Denis and Schiffermüller (Faucheux 1991), Ostrinia nubilalis (Hübner) (Hallberg et al. 1994), Ostrinia furnacalis (Guenée) (Jung et al. 1999), and Z. dixolophella (Castrejón-Gómez et al. 2003). Similar results were reported in some members of the Tortricidae such as Argyrotaenia velutiana (Walker) (Callahan 1975), Cydia nigricana (F.) (Wall 1978) and some members of the Noctuidae such as Pseudaletia unipuncta (Haw) (Lavoie-Dornik and McNeil 1987). The function of SCO might be olfactory (Yang et al. 2009). Two types of SCO have been found in some insects (Roux et al. 2005), whereas only one type has been found in others (Gao et al. 2007, Steinbrecht 1984). In the present study, only one type was found in the antennae of both sexes of M. brassicae, and these grooved pegs may function as chemo- or thermoreceptors. SCO that are recessed below the antennal surface and located within cavities may be involved in the perception of humidity, temperature, heat, or CO₂ (Altner et al. 1983, Steinbrecht 1984), may play a role in preventing desiccation (Kristoffersen et al. 2006), and may have an olfactory function (Gao et al. 2007, Keil 1999, Ochieng et al. 2000, Roux et al. 2005, Steinbrecht 1997). Results of a transmission electron microscopic study of the tobacco hornworm, Manduca sexta L. (Lepidoptera: Sphingidae) suggested that olfactory molecules entered through the grooves in SCO (Shields and Hildebrand 2001). In the SCO of M. brassicae, the presence of deep longitudinal grooves on the surface of central and circumferential pegs also suggests their olfactory function (Fig. 5A).

We identified and characterized nine types of antennal sensilla of M. brassicae and discussed their probable functions. These results provide necessary background information for our ongoing study of the chemical ecology of M. brassicae. Future studies on the functional morphology of the antennal sensilla by use of transmission electron microscopy coupled with electrophysiological recordings will likely confirm the functions of the sensilla identified in this study.