Skip to main content
Download PDF

Expression profiles analysis and roles in immunity of transient receptor potential (TRP) channel genes in Spodoptera frugiperda

BMC Genomics volume 26, Article number: 401 (2025) Cite this article

Abstract

Background

Transient receptor potential (TRP) ion channels play crucial roles in mediating responses to environmental stimuli, as well as regulating homeostasis and developmental processes in insects. Several members of the TRP superfamily are potential molecular targets for insecticides or repellents, indicating their research value in pest control. This study focuses on Spodoptera frugiperda, an important invasive pest in China known for its wide host range and strong reproductive capacity. Currently, there is a lack of molecular research on the TRP channels of the invasive pest S. frugiperda.

Results

In this study, we identified 15 TRP family genes in S. frugiperda, which were classified into six subfamilies. The TRPP subfamily gene was not identified, whereas the TRPA subfamily contained the highest number of members in this insect. Real-time quantitative polymerase chain reaction (RT-qPCR) experiments revealed widespread expression of TRP channel genes across various developmental stages of S. frugiperda. However, TRPM and TRPML were highly expressed only in eggs. Transcripts of TRP channel genes were detected in the sensory organs of mature adults, including the mouthparts, antennae, compound eyes, legs, wings, harpagones, and ovipositors, as well as in tissues of 5th instar larvae (hemocytes, central nervous system, midgut, fat body, and Malpighian tubules). To explore the potential role of TRP channels in immunity, we detected their levels in larvae 24 h after infection with Serratia marcescens. The expression levels of TRPML, TRPL, and the Pain genes were significantly up-regulated, suggesting their important roles in immune responses to S. marcescens.

Conclusions

The results of this study extend our knowledge of these critical sensory channels in S. frugiperda. This knowledge provides a basis for the future development of insecticides that target these channels, thereby promoting the safe and effective control of this key pest.

Peer Review reports

Background

Transient receptor potential (TRP) channels are key proteins in insects, essential for sensing changes in the external environment and regulating physiological processes such as vision [1], olfaction [2], auditory sensation [3], taste [4], temperature sensing [5], and mechanical sensation [6, 7]. Based on primary amino acid sequence homology, the TRP family is divided into seven subfamilies listed as follows: TRP-Canonical (TRPC), TRP-Ankyrin (TRPA), TRP-No mechanoreceptor potential C (TRPN), TRP-Vanilloid (TRPV), TRP-Melastatin (TRPM), TRP-Mucolipin (TRPML), and TRP-Polycystin (TRPP). Moreover, TRP channel families can be classified into two groups according to their differences in sequences and topologies: Group 1 contains the TRPC, TRPA, TRPN, TRPV and TRPM subfamilies, while Group 2 consists of the remaining TRPP and TRPML subfamilies [8].

In recent studies, a number of TRP channel members have been identified as molecular targets for insecticides or repellents. For example, TRPV subfamily members, including Nanchung (Nan) and Inactive (Iav) channels, are activated by pymetrozine, pyrifluquinazon, and afidopyropen, causing coordination dysfunction in adult insects, ultimately leading to death [9,10,11]. TRPA subfamily members, including the Water with (Wtrw) channel, can be activated by afidopyropen, pymetrozine, and the endogenous agonist, nicotinamide [12]. Furthermore, silencing TRPM subfamily channels significantly reduced the repellent activity of L-menthol and menthoxypropanediol (MPD) against Tribolium castaneum [13]. Additionally, TRP1, a homolog of the TRPC subfamily channel, TRPgamma (TRPγ), can be directly activated by natural repellents such as, citronellal, citronellol, and camphor in Mesobuthus martensii [14]. Thus, it is important to determine the sequences and understand the properties of TRP channels for effective pest control strategies.

Although numerous TRP channels have been identified and characterized in various insects [15,16,17,18,19], the functional roles of TRP channels in the agricultural pest Spodoptera frugiperda remains limited. This insect, also known as the fall armyworm, is a common polyphagous agricultural pest [20, 21], that can affect approximately 353 plant species across 76 different families [22, 23]. The wide host range, strong fertility, rapid migration, and adaptability to diverse habitats of S. frugiperda contribute to its characteristic large-scale, concentrated outbreaks over short periods of time, making prevention and control challenging. The damage caused by S. frugiperda is mainly due to its larvae feeding on crop leaves, which affects plant photosynthesis, ultimately reducing crop yield and causing economic losses [24]. To reduce production losses, chemical control is widely used in agricultural as the simplest and most convenient method for large-scale application. However, S. frugiperda has developed resistance to various insecticides, including lambda-cyhalothrin [25], organic phosphate esters [26], chlorpyrifos [27], chlorantraniliprole [28], lufenuron [29], and spinosad [30], due to the long-term overuse of chemical pesticides. Therefore, it is necessary to develop new prevention and control technologies, such as strategies based on biological control.

The use of organisms and microorganisms to manage pests is an essential component of integrated pest management. Among bacteria suitable for pest management, Serratia marcescens (Enterobacterales: Enterobacteriaceae) is a Gram-negative bacterium widely distributed in natural environments [31]. S. marcescens produces a variety of exoenzymes, including chitinase, which hydrolyze and destroy the surface and periplasmic structures of insects, eventually causing death. This trait renders the bacterium highly pathogenic to numerous agricultural and forestry pests [32,33,34,35]. When S. marcescens infects the fall armyworm, the larvae show a strong immune response [36], which reduces their survival rate, development rate and adult emergence rate [37, 38]. S. marcescens infection in other insects leads to upregulation of immune-related genes in Spodoptera exigua [39]. Additionally, it affects growth, development, and reproduction in Mythimna separata (Walker) and Spodoptera litura (Fab.) [40, 41].

In this study, we identified 15 TRP channels in S. frugiperda and investigated their expression patterns across different developmental stages and in multiple tissues of both larvae and adult S. frugiperda. In addition, we analyzed the immune response of S. frugiperda TRP channels following injection of the conditionally pathogenic bacterium, S. marcescens. Our characterization of TRP channels in S. frugiperda lays the groundwork for future functional studies and may contribute to the development of innovative pest control methods.

Results

Identification and sequence analysis of TRP channels in S. frugiperda

Fifteen TRP channel genes were identified in S. frugiperda that exhibited sequence homology to known Drosophila TRP channel sequences (Fig. 1 and Table 1). Phylogenetic analysis categorized these channels into subfamilies: three TRPC, seven TRPA, one TRPN, two TRPV, one TRPM, and one TRPML subfamily members. Interestingly, no TRPP subfamily members were identified in S. frugiperda (Fig. 1 and Table 1). These 15 S. frugiperda TRP channel genes were classified into six subfamilies and divided into two groups based on structural specificity and homology (Fig. 1 and Table 1). Group 1 included five subfamilies: TRPC (SfruruTRP, SfruTRPL, SfruTRPγ), TRPA (SfruTRPA1, SfruTRPA5, SfruPain, SfruPyx1, SfruPyx2, SfruWtrw1, SfruWtrw2), TRPN (SfruNompC), TRPV (SfruIav, SfruNan), and TRPM (SfruTRPM). Group 2 comprised only SfruTRPML of the TRPML subfamily. Sequence analysis identified six transmembrane domains in all S. frugiperda TRP channels (Table 1). Most group 1 TRP proteins had 1 to 29 N-terminal ankyrin repeat (AR) domains, except for the TRPM subfamily. Conversely, no AR domains were detected at the N-terminus of the group 2 TRPML subfamily protein (Table 1). BLASTP analyses of protein sequence alignments showed that all S. frugiperda TRP channels exhibited a high level of sequence identity (> 62%) with their counterparts in Drosophila melanogaster (Table 1).

Fig. 1
figure 1

Phylogenetic analysis of TRP channels in Spodoptera frugiperda and other insects. The tree was constructed using the software MEGA 6.06 with 1000 bootstrap replicates based on the Maximum Likelihood method. The numbers on branch nodes denote levels of bootstrap support. Species abbreviations are Sfru, Spodoptera frugiperda, Bmor, Bombyx mori, Dmel, Drosophila melanogaster, Tcas, Tribolium castaneum

Full size image
Table 1 TRP channels identified from S. frugiperda
Full size table

TRP channel transcript levels in different S. frugiperda developmental stages

Next, we performed RT-qPCR analyses to detect TRP channel expression patterns at different stages of S. frugiperda development, including eggs, larvae, pupae, and adults (Fig. 2). The results showed that SfruTRP was highly expressed in 1-day-old, with lower expression in 3-day-old adult males (Fig. S1 A). SfruTRPL was primarily expressed in 6 th instar larvae and pupae, while its expression was low at other stages (Fig. S1B). SfruTRPγ transcripts were generally detected at all stages tested, with the highest expression levels observed in pupae and 1-day-old adults (Fig. S1 C). SfruTRPA1 was abundantly expressed in 1 st instar larvae, with significantly higher levels compared toother stages (Fig. S1D). SfruTRPA5, which is specific to the Lepidoptera, was detected in pupae, 1-day-old adults, and 3-day-old males (Fig. S1E). SfruPain was more highly expressed in adults compared to other developmental stages (Fig. S1 F). Both SfruPyx splice forms were abundantly expressed in 3-day-old males (Fig. S1G and H). SfruPyx1 was also expressed in pupae, while SfruPyx2 showed additional expression in 1-day-old adult males. Both transcripts encoding Wtrw molecules were highly expressed in 1-day-old and 3-day-old adult males (Fig. S1I and J). In contrast to SfruWtrw1, SfruWtrw2 was also expressed in 1 st to 4 th instar larvae (Fig. S1 J). Similar to SfruTRP, high SfruNompC transcripts levels were detected in 1 st instar larvae, as well as 1- day-old adults and 3-day-old males (Fig. S1 K). SfruIav was widely expressed across all tested stages, particularly at the pupal stage (Fig. S1L). In contrast, SfruNan was mainly highly expressed in 1 st and 2nd instar larvae (Fig. S1M). Unlike most other TRP channels, which are predominantly expressed in adult stage insects, SfruTRPM and SfruTRPML were almost exclusively highly expressed in eggs (Fig. S1 N and O).

Fig. 2
figure 2

Relative expression level of TRP channels in different developmental stages of Spodoptera frugiperda, including eggs (1 to 2 h), larvae (1 st, 2nd, 3rd, 4 th, and 5.th instar), pupae, immature adult male and female insects (1-day-old), and mature adult male and female insects (3-day-old)

Full size image

Tissue distribution of TRP channels in S. frugiperda adults

To explore the possible roles of TRP channels in S. frugiperda, RT-qPCR analyses were performed to determine their relative transcription levels in various external sensory organs of adults, including mouthparts, antennae, compound eyes, legs, wings, harpagones, and ovipositors (Fig. 3). Both SfruTRP and SfruTRPL were highly expressed in compound eyes (Fig. S2 A and B), whereas SfruTRPγ exhibited high expression in all tissues except the wings and ovipositors (Fig. S2 C). SfruTRPA1 was mainly expressed in mouthparts, antenna, and compound eyes (Fig. S2D). Meanwhile, SfruTRPA5 expression levels were high in most tissues, except for the compound eyes and ovipositors (Fig. S2E). Similar to SfruTRPA1, transcripts of SfruPain were abundantly detected in the mouthparts, antennae, and compound eyes (Fig. S2 F). SfruPyx1 expression levels were highest in the compound eyes (Fig. S2G), while SfruPyx2 showed peak expression in the mouthparts (Fig. S2H). SfruWtrw1 was highly expressed in the antennae (Fig. S2I), whereas SfruWtrw2 was hardly detectable in this tissue (Fig. S2 J). The antennae displayed the highest levels of SfruNompC, SfruIav, SfruNan, SfruTRPM, and SfruTRPML mRNAs (Fig. S2 K-O). In addition, SfruNompC and SfruTRPM showed high transcription levels in the mouthparts (Fig. S2 K and N), while SfruIav was expressed at higher levels in the compound eyes (Fig. S2L). Beyond the antennae, SfruTRPML mRNA expression levels were also increased in the mouthparts, legs, harpagones, and ovipositors, but were lower in the compound eyes and wings (Fig. S2O).

Fig. 3
figure 3

TRP channels relative expression levels in mature adult Spodoptera frugiperda various tissues, including mouthparts, antennae, compound eyes, legs, harpagones, ovipositors

Full size image

Tissue distribution of TRP channels in S. frugiperda larvae

The internal tissues of insect larvae, including the hemocytes, central nervous system (CNS), midgut, fat body, and Malpighian tubules (MT), play crucial roles in maintaining homeostasis. To investigate the role of TRP channels in larval internal tissues, we examined their relative transcription levels, revealing distinct expression patterns (Fig. 4). SfruTRP was highly expressed in the CNS, with low expression levels detected in other tissues (Fig. S3 A). Transcription levels of SfruTRPL were higher in the MT than in other tissues (Fig. S3B). Abundant transcripts of SfruTRP, SfruTRPA1, SfruPain, SfruWtrw1, SfruNompC, and SfruIav were detected in the CNS. However, their expression levels were low in other tissues (Fig. S3 C, D, F, H, J, K). High levels of SfruTRPA5 and SfruPyx2 transcripts were detected in hemocytes, while their expression was minimal in other tissues (Fig. S3E, H). Moreover, SfruIav transcripts were highly expressed in the CNS, hemocytes, and fat body (Fig. S3 K). Similar to SfruWtrw, SfruTRPML transcripts were also highly expressed in the MT (Fig. S3M).

Fig. 4
figure 4

TRP channels relative expression levels in larval Spodoptera frugiperda various tissues, including hemocytes, central nervous system (CNS), midgut, fat body, and Malpighian tubules (MT)

Full size image

Effect of S. marcescens infection on the expression of TRP channels

To investigate the response of TRP channels to S. marcescens infection, we monitored the expression of the 15 TRP genes in insect hemocytes. The bacteria were injected into the mid-region of 5 th instar S. frugiperda larvae, and gene expression was analyzed 24 h post-injection. Only seven TRP genes were expressed 24 h after inoculation with S. marcescens, while the levels of the other genes were barely detectable. SfruTRPML gene expression increased 2.64-fold 24 h post-infection (Fig. 5A), while SfruTRPL levels were significantly up-regulated by 2.68-fold (Fig. 5B). In addition, SfruPain expression increased 1.48-fold compared to wild-type (WT) controls (Fig. 5C). Since SfruTRP5 and SfruPyx2 were highly expressed in hemocytes, we speculated that they might play a role in immune responses. However, after injecting the bacteria, no significant differences in the expression levels of these genes were observed compared to WT controls (Fig. 5D). Furthermore, SfruPyx1 and SfruTRPM expression levels showed no significant changes following S. marcescens injection (Fig. 5E–G).

Fig. 5
figure 5

The relative expression levels of TRP channels in the hemocytes larval Spodoptera frugiperda 24 h after injection with Serratia marcescens. Error bars represent standard error. “ns” indicates no significant difference, while asterisks indicate values significantly different from control values determined from unpaired t -test (*p < 0.05, ** p < 0.01, *** p < 0.001)

Full size image

Discussion

In this study, we identified 15 TRP channels in S. frugiperda, which were classified into six subfamilies, TRPC, TRPA, TRPN, TRPV, TRPM, and TRPML, based on structural and phylogenetic analyses. However, TRPP, considered the oldest subfamily, was not found in S. frugiperda, suggesting functional compensation by other TRP channels or alternative pathways in this insect. Among all TRP channels subfamilies, TRPA exhibits the highest diversity in arthropods [42], consistent with our finding that the TRPA subfamily in S. frugiperda has more members than other TRP subfamilies. In addition, the gene encoding TRPA5 is widely found in Lepidoptera, Isoptera, and Blattodea [17], although its function has not been reported yet. In this study, the gene encoding TRPA5 was detected in S. frugiperda, along with two Wtrw genes (SfruWtrw1 and SfruWtrw2). Multiple Wtrw transcripts are widely present only in the Lepidoptera, which may be related to their specific habitats and life histories [17].

Except for SfruTRPM and SfruTRPML, which were almost exclusively expressed in egg, all TRP channels in this study were detected across all developmental stages of S. frugiperda. These results indicate that different TRP channels may be involved in distinct physiological processes, offering opportunities for the development of insecticides targeting specific life cycle stages. S. frugiperda TRPM was only highly expressed during the egg stage and was almost undetectable in other tissues. This situation also occurs in the Drosophila and B. dorsalis, where this gene is crucial for Mg2+and Zn2+ homeostasis [43, 44]. Unlike other TRPML genes, which are highly expressed in the gut and MT [16, 17], S. frugiperda TRPML was not detected in these tissues, but like TRPM, it was highly expressed during the egg stage. Hence, this gene may be necessary for the early development of S. frugiperda.

Studies in Drosophila have shown that TRP and TRPL channels are involved in phototransduction [1, 45]. SfruTRP, SfruTRPL, and SfruTRPγ were expressed at high levels in the compound eyes, implying that they are important for photoreception. In addition to compound eyes, we detected SfruTRPγ in the mouthparts, antennae, and legs of S. frugiperda. In other insects, TRPγ is also highly expressed in these tissues, and participates in coordinated locomotion, with insects lacking DmTRPγ unable to properly coordinate leg locomotion [17, 46]. The proprioceptive neurons are distributed in the appendage joints of Drosophila, including the legs and wings [47]. TRPA1 is involved in temperature sensation and avoidance of noxious heat, aversive odorants and tastants, non-volatile irritants, strong lights, and mechanical stimuli [48]. The presence of olfactory, auditory, and gravitational sensory organs in insect antennae [48], along with the high expression of TRPA1 in S. frugiperda antennae, suggests that TRPA1 may contribute to these functions. Similar to Pieris rapae [17], the Pain channel is widely distributed in S. frugiperda, which may be important for the sensation of gravity, as well as the avoidance of noxious heat, mechanical stimulation, and dry environments [48]. Pyx is involved in the gravity sensing in Drosophila [6], while Wtrw detects dry air and mosquito repellent [12, 49]. The widespread distribution of taste receptor neurons in the labellum, wings, legs, and ovipositors of female Drosophila contributes to food selection and aversion behaviors [50], which is consistent with our findings of relatively high levels of SfruPyx2 and SfruNompC expression in the mouthparts of S. frugiperda. The dominant expression of Nan and Iav in antennae has been previously reported [11]. We also observed higher expression of these genes in S. frugiperda gut samples. Nan and Iav may play a critical role in gravity, sound sensation, and feeding in S. frugiperda. Pymetrozine and pyrifluquinazon disrupt mechanosensation and chordotonal organ functions by activating the Nan-Iav channel complex, thereby impairing gravity perception, sound perception, and feeding behavior in insects [11, 48]. Previous studies have shown that TRPM channels mediate the repellent responses of Tribolium castaneum to l-menthol and MPD. These findings suggest that these TRP channels may play a key role in insect perception and response to insecticides [13]. The distribution of SfruTRPM in S. frugiperda antennae and mouthparts suggests a similar role in this insect. Our data show high expression of SfruTRPM in S. frugiperda MT, suggesting a functional similarity to D. melanogaster. Drosophila TRPM impacts noxious cold sensation and gentle touch mechanical sensation [51], and we found that it is highly expressed in the mouthparts and compound eyes of S. frugiperda. These genes play a role in temperature perception and avoidance of harmful stimuli. Therefore, inhibitors targeting these channels can affect the response capability of the S. frugiperda to environmental changes, thereby reducing its adaptability.

Regarding larval tissues, our data showed that several TRP channels (SfruTRP, SfruTPRγ, SfruTRPA1, SfruPain, SfruWtrw1, SfruNompC, and SfruIav) were detected in the CNS. We hypothesize that these genes play significant roles in sensory perception, motor control, behavior regulation, endocrine modulation, as well as learning and memory in S. frugiperda larvae. TRPM is essential for Mg2+ and Zn2+ homeostasis, and it has been reported that knocking out this channel in D. melanogaster leads to shortened MT and results in larval growth arrest during the larval stage [44]. Furthermore, TRPML is involved in locomotion, autophagy, and apoptotic cell clearance [48]. SfruTPRL, SfruWtrw2, SfruTRPM and SfruTRPML are highly expressed in MT, suggesting functional roles in this tissue. Although SfruTPRA5 and SfruPyx2 are highly expressed in blood cells, their functions have been rarely studied.

In our study, transcript levels of SfruTRPML, SfruTRPL, and SfruPain were found to be significantly higher in S. frugiperda after being infected by the Gram-negative bacterium, S. marcescens, suggesting their involvement in immune response functions. Similar immune-related expression of SfruTRPML has also been observed in Drosophila, suggesting that expression of TRPML in hemocytes is essential for antibacterial immune responses [52]. TRPL belongs to the TRPC subfamily and is mainly involved in photoreception transduction in insects, which has rarely been reported in the context of insect immunity. However, TRPC subfamily molecules contribute to Ca2+ signaling in immune cells of other organisms [53]. Other TRP channel subfamilies have a wide range of functional roles in inflammation and immunity. TRPM2 channels are widely expressed in immune cells [54], where they are essential for regulating cation balance in inflammatory environments [55]. TRPV4, a member of the vanilloid (TRPV) subfamily of TRP channels, forms a widely expressed mechanosensitive channel which can also be found in many (innate) immune cells. It plays a regulatory role in increasing proinflammatory cytokine expression induced by bacterial lipopolysaccharides [56, 57]. Collectively, this study identifies three TRP genes activated by S. marcescens, providing preliminary evidence for their potential as target genes for insecticides. However, other members did not exhibit a response regarding immune function; these genes may still play significant roles in additional important physiological processes requiring further investigation. Therefore, they remain viable research targets for developing insecticides.

Conclusions

In summary, the gene expression results suggest that TRP channels may be involved in immune responses, warranting further investigation into their specific roles and mechanisms. Our study expands knowledge of these functions, as well as key sensory pathways in S. frugiperda, laying the foundation for new strategies to control these important pest insects.

Materials and methods

Insects

S. frugiperda larvae used in this study were collected from Qilin North Farm, South China Agricultural University, and raised in an incubator. The insects had been cultured for more than 10 generations. Individual S. frugiperda were reared at 26 °C ± 1℃ and 75% ± 1% relative humidity, with a photoperiod cycle of 16-h light and 8-h dark. Collected eggs were soaked in a 5% formaldehyde solution for 20 min, rinsed with water for 5 min, dried, and placed in a 90 mm petri dishes sealed with film. Hatched larvae were fed an artificial diet described by Li et al. [58]. Then, 3rd to 5 th instar larvae were transferred to a new 150 mm petri dish, with approximately 10 larvae per dish. Fresh artificial feed, composed of bean powder 100 g, wheat bran 80 g, yeast powder 26 g, casein 8 g, ascorbic acid 8 g, distilled water 500 mL, agar 26 g, choline chloride 1 g, sorbic acid 1 g, inositol 0.2 g, streptomycin 0.1 g, penicillin sodium 0.1 g, and propylparaben 2 g, was replaced daily, and feces and dead insects in the culture dishes were cleaned. After pupation, pupae were housed in homemade paper tubes (diameter: 85 mm; height: 150 mm) until adults emerged. Adults were provided with honey water (approximately 5%). Egg mass collection began 2 days after successful adult mating.

Identification of TRP channels

The methods used to identify TRP channels in various species were similar to those reported in our previous study [16]. To search exhaustively all TRP genes in each species, we screened several types of databases including assembled genomes, reference sequence (RefSeq) database from National Center for Biotechnology Information (NCBI) (https://www.ncbi.nlm.nih.gov/refseq/) and transcriptomic data acquired from NCBI Sequence Read Archive (SRA) Databases (https://www.ncbi.nlm.nih.gov/genbank/tsa/). We obtained the genome data of Spodoptera frugiperda, Bombyx mori, Tribolium castaneum from Silkworm Genome database (http://silkworm.genomics.org.cn/), and Beetlebase (http://www.beetlebase.org/) respectively. Firstly, candidate S. frugiperda TRP genes were identified by TBLASTN searches against genome and transcriptomes with an E-value cutoff of 1e−5, using known TRP protein sequences of D. melanogaster. Then, candidate genes were further verified using BLASTP versus non-redundant NCBI protein sequences without species limits and with a cut-off e-value of 1e−5. The same procedure was used to identify TRP genes of B. mori, T. castaneum by a homology-based approach. Transmembrane segments were predicted using TMHMM 2.0 (http://www.cbs.dtu.dk/services/TMHMM-2.0/) and SMART (http://smart.embl-heidelberg.de/). Multiple alignments of complete amino acid sequences were performed with ClustalW2 (http://www.ebi.ac.uk/Tools/msa/clustalw2/), and the results were displayed using BioEdit (https://bioedit.software.informer.com/). Phylogenetic trees and molecular evolutionary analyses were performed using the maximum likelihood method in MEGA 6.06 software with 1000 bootstrap replicates.

Real-time quantitative PCR (RT-qPCR) analysis

S. frugiperda is a holometabolous insect, and its developmental stages include the egg, larva, pupa, and adult phases. To study the spatiotemporal distribution of TRP channels in S. frugiperda, samples were collected from various developmental stages, including eggs (1 to 2 h), larvae (1 st, 2nd, 3rd, 4 th, and 5 th instar), pupae (male: female ratio: 1:1), immature adult male and female insects (1-day-old), and mature adult male and female insects (3-day-old). In addition, we selected the important tissues required for insects to perceive environmental stimuli and maintain internal balance. Tissues, including mouthparts, antenna, compound eyes, legs, harpagones, ovipositors, hemocytes, central nervous system (CNS), midgut, fat body, and Malpighian tubules (MT) were dissected from equal numbers of male and female 3-day-old adults. Temporal distribution of gene expression was analyzed using pooled samples of eggs (n = 100 per pool); 1 st (n = 20), 2nd (n = 10), 3rd (n = 5), 4 th (n = 3), 5 th (n = 3), and 6 th (n = 3) instar larvae; pupae (n = 3); and single 1-day-old (immature adult) and 3-day-old (mature adult) male and female adult insects. Tissue distribution analysis was performed with 20 adult tissue samples included in each pool. At least three biological replicates were carried out for each experiment.

RNA was extracted using TRIzol reagent (Invitrogen, Carlsbad, CA, USA) and RNA quantities measured using a Nanodrop 2000 spectrophotometer (Thermo Scientific Inc., Bremen, Germany). Reverse transcription was then performed with 1 μg RNA samples using TransScript one-step gDNA removal and cDNA Synthesis SuperMix (TransGen Biotech, China). Synthesized cDNA served as a template for RT-qPCR, which was performed on a Stratagene Mx3000P thermal cycler (Agilent Technologies, Wilmington, DE). Each reaction mixture contained 5 μL TB Green Premix Ex Taq II (Tli RNase H Plus) (TaKaRa Bio, Otsu, Japan), 0.4 μL of each primer (0.2 μM), and 0.8 μL of template cDNA, with sterile distilled water added to a final volume of 10 μL. Thermal cycling conditions were: 30 s at 95 °C, then 40 cycles of 95 °C for 5 s and 60 °C for 34 s. Three sample replicates were performed for each group, and no-template negative controls were included in each run, to detect possible contamination or carryover. A series of gene-specific primers were designed for RT-qPCR using the software, Primer 3 (http://bioinfo.ut.ee/primer3-0.4.0/) (Additional file 1: Table S1). Primers were used to investigate the relative expression levels of selected samples and melting curve analysis was performed between 60 °C to 95 °C for all reactions, to ensure the specificity and consistency of generated products (Fig S4). The specificity of all RT-qPCR reaction products was further established via electrophoresis on 1.0% agarose gel prior to sequencing. All experiments were performed independently at least twice, to ensure their reliability and reproducibility. Transcript levels of different genes were quantified using the 2−ΔΔCT method [59], with SfruRPS3 A and SfruL17, commonly used reference genes in the S. marcescens, serving as normalization genes.

Microbial infection by injection

We selected the well-recognized model pathogen, S. marcescens, which is widely used in the study of insect immune responses. Fifth instar larvae were dehydrated for 24 h without food and then injected with S. marcescens cultured in LB medium at 37 °C with shaking (200 rpm). Bacterial cultures were harvested at an optical density at 600 nm (OD600) of 1.0, corresponding to approximately 5 × 108 colony-forming units per mL. Bacterial cultures (250 mL) were pelleted by centrifugation (10 min, 4000 g) and washed twice with phosphate buffered saline (PBS). For larval systemic infection, bacterial pellets were resuspended in PBS and adjusted to OD600 = 0.025 and 1 µL bacterial solution injected into the middle part of larvae using a syringe, while larvae injected with PBS served as controls. Hemocytes were collected from larvae 24 h after injection. Pools of ten larvae were used, with three biological replicates and two independent experimental replicates conducted.

Statistical analysis

Statistical analysis of data was performed using Prism 8.0 (GraphPad Software). The t-test was used for unpaired comparisons between two groups of data, and one-way analysis of variance (ANOVA) followed by Tukey’s multiple pairwise comparison test was applied for comparisons of three or more groups. Significance was set at p < 0.05. Data are presented as mean ± standard error of the mean (SEM). The heatmap analysis was performed using OmicShare tools (https://www.omicshare.com/tools/).

Data availability

All sequence data that support the findings of this study were listed in Additional file 1: Table S1.

References

  1. Montell C, Rubin GM. Molecular characterization of the Drosophila trp locus: a putative integral membrane protein required for phototransduction. Neuron. 1989;2:1313–23.

    Article  CAS  PubMed  Google Scholar 

  2. Colbert HA, Bargmann CI. Odorant-specific adaptation pathways generate olfactory plasticity in C. elegans - ScienceDirect. Neuron. 1995;14:803.

    Article  CAS  PubMed  Google Scholar 

  3. Matsuura H, Sokabe T, Kohno K, Tominaga M, Kadowaki T. Evolutionary conservation and changes in insect TRP channels. BMC Evol Biol. 2009;9:228.

    Article  PubMed  PubMed Central  Google Scholar 

  4. Zhang YF, Hoon MA, Chandrashekar J, Mueller KL, Cook B, Wu DQ. Coding of sweet, bitter, and umami tastes: Different receptor cells sharing similar signaling pathways. Cell. 2003;112:293–301.

    Article  CAS  PubMed  Google Scholar 

  5. Caterina MJ, Schumacher MA, Tominaga M, Rosen TA, Levine JD, Julius D. The capsaicin receptor: a heat-activated ion channel in the pain pathway. Nature. 1997;389:816–24.

    Article  CAS  PubMed  Google Scholar 

  6. Sun Y, Liu L, Ben-Shahar Y, Jacobs JS, Eberl DF, Welsh MJ. TRPA channels distinguish gravity sensing from hearing in Johnston’s organ. Proc Natl Acad Sci USA. 2009;106:13606–11.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Wang LX, Niu CD, Zhang Y, Jia YL, Zhang YJ, Zhang Y. The NompC channel regulates Nilaparvata lugens proprioception and gentle-touch response. Insect Biochem Mol Biol. 2019;106:55–63.

    Article  CAS  PubMed  Google Scholar 

  8. Venkatachalam K, Montell C. TRP channels. Annu Rev Biochem. 2007;76:387–417.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Nesterov A, Spalthoff C, Kandasamy R, Katana R, Rankl NB, Andres M. TRP channels in insect stretch receptors as insecticide targets. Neuron. 2015;86:665–71.

    Article  CAS  PubMed  Google Scholar 

  10. Kandasamy R, London D, Stam L, Von Deyn W, Zhao X, Salgado VL. Afidopyropen: new and potent modulator of insect transient receptor potential channels. Insect Biochem Mol Biol. 2017;84:32.

    Article  CAS  PubMed  Google Scholar 

  11. Wang LX, Niu CD, Salgado VL, Lelito K, Stam L, Jia YL, Zhang Y, Gao CF, Wu SF. Pymetrozine activates TRPV channels of brown planthopper Nilaparvata lugens. Pestic Biochem Physiol. 2019;153:77–86.

    Article  CAS  PubMed  Google Scholar 

  12. Kandasamy R, Costea PI, Stam L, Nesterov A. TRPV channel nanchung and TRPA channel water witch form insecticide-activated complexes. Insect Biochem Mol Biol. 2022;149:103835.

    Article  CAS  PubMed  Google Scholar 

  13. Shimomura K, Oikawa H, Hasobe M, Suzuki N, Yajima S, Tomizawa M. Contact repellency by l-menthol is mediated by TRPM channels in the red flour beetle Tribolium castaneum. Pest Manag Sci. 2021;77:1422–7.

    Article  CAS  PubMed  Google Scholar 

  14. Tian Q, Wang P, Xie C, Pang P, Zhang Y, Gao Y. Identification of an arthropod molecular target for plant-derived natural repellents. Proc Natl Acad Sci U S A. 2022;119:e2118152119.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Kim HG, Margolies D, Park Y. The roles of thermal transient receptor potential channels in thermotactic behavior and in thermal acclimation in the red flour beetle, Tribolium castaneum. Insect Physiol. 2015;76:47–55.

    Article  CAS  Google Scholar 

  16. Su HA, Bai X, Zeng T, Lu YY, Qi YX. Identification, characterization and expression analysis of transient receptor potential channel genes in the oriental fruit fly, Bactrocera dorsalis. BMC Genomics. 2018;19:674.

    Article  PubMed  PubMed Central  Google Scholar 

  17. Mao F, Lu WJ, Yang Y, Qiao X, Huang J. Identification, characterization and expression analysis of TRP channel genes in the vegetable pest, Pieris rapae. Insects. 2020;11:192.

    Article  PubMed  PubMed Central  Google Scholar 

  18. Wang LX, Niu CD, Wu SF, Gao CF. Molecular characterizations and expression profiles of transient receptor potential channels in the brown planthopper, Nilaparvata lugens. Pestic Biochem Physiol. 2021;173:104780.

    Article  CAS  PubMed  Google Scholar 

  19. Zhang YT, Liu HW, Cao S, Li B, Liu Y, Wang G. Identification of transient receptor potential channel genes and functional characterization of trpa1 in Spodoptera frugiperda. J Integrat Agric. 2024;23:1994–2006.

    Article  CAS  Google Scholar 

  20. Sparks AN. A review of the biology of the fall armyworm. Florida Entomol. 1979;62:82.

    Article  Google Scholar 

  21. Nagoshi RN, Shelby F, Meagher RL, Mirian HR, Ayub K, Gabriela MM. Fall armyworm migration across the lesser antilles and the potential for genetic exchanges between North and South American populations. PLoS One. 2017;12:e0171743.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Montezano DG, Specht A, Sosa-Gómez DR, Roque-Specht VF, Hunt T. Host plants of Spodoptera frugiperda (Lepidoptera: Noctuidae) in the Americas. African Entomol. 2018;26:286–300.

    Article  Google Scholar 

  23. Sun X, Hu C, Jia H, Wu Q, Wu K. Case study on the first immigration of fall armyworm Spodoptera frugiperda invading into China. J Integrat Agric. 2019;18:2–10.

    Google Scholar 

  24. Johnson SJ. Migration and the life history strategy of the fall armyworm, Spodoptera frugiperda in the western hemisphere. Int Trop Insect Sci. 1987;8:543–9.

    Article  Google Scholar 

  25. Ríos-Díez JD. Saldamando-Benjumea CI, Susceptibility of Spodoptera frugiperda (Lepidoptera: Noctuidae) strains from central colombia to two insecticides, methomyl and lambda-cyhalothrin: a study of the genetic basis of resistance. J Econ Entomol. 2011;5:1698–705.

    Article  Google Scholar 

  26. Arias O, Cordeiro E, Corrêa AS, Domingues FA, Guidolin AS, Omoto C. Population genetic structure and demographic history of Spodoptera frugiperda (Lepidoptera: Noctuidae): implications for insect resistance management programs. Pest Manag Sci. 2019;75(11):2948–57.

    Article  CAS  PubMed  Google Scholar 

  27. Renato A. Carvalho, Celso, Omoto, Linda, Investigating the molecular mechanisms of organophosphate and pyrethroid resistance in the fall armyworm Spodoptera frugiperda. PLoS One. 2013;8:e62268.

    Article  Google Scholar 

  28. Bolzan A, Padovez FE, Nascimento AR, Kaiser IS, Lira EC, Amaral FS. Selection and characterization of the inheritance of resistance of Spodoptera frugiperda (Lepidoptera: Noctuidae) to chlorantraniliprole and cross-resistance to other diamide insecticides. Pest Manag Sci. 2019;75:2682–9.

    Article  CAS  PubMed  Google Scholar 

  29. Nascimento ARBd, Farias JR, Bernardi D, Horikoshi RJ, Omoto C. Genetic basis of Spodoptera frugiperda (Lepidoptera: Noctuidae) resistance to the chitin synthesis inhibitor lufenuron. Pest Manag Sci. 2016;72:810–5.

    Article  PubMed  Google Scholar 

  30. Okuma DM, Bernardi D, Horikoshi RJ. Inheritance and fitness costs of Spodoptera frugiperda (Lepidoptera: Noctuidae) resistance to spinosad in Brazil. Pest Manag Sci. 2018;74:1441–8.

    Article  CAS  PubMed  Google Scholar 

  31. Petersen LM, Tisa LS. Friend or foe? A review of the mechanisms that drive Serratia towards diverse lifestyles. Can J Microbiol. 2013;59:627–40.

    Article  CAS  PubMed  Google Scholar 

  32. Zhao LN, Ma Y, Yang X, Iqbal A, Ruan CC, Zang LS. Identification of Serratia marcescens isolated from Antheraea pernyi eggs and determination of bacterial pathogenicity and transmission pathway. J Invertebr Pathol. 2020;169:107297.

    Article  PubMed  Google Scholar 

  33. Wang S, Dos-Santos ALA, Huang W. Driving mosquito refractoriness to Plasmodium falciparum with engineered symbiotic bacteria. Science. 2017;357:1399–402.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Wei H, Fan K, Zhanjun L, Ning Z, Tingtao C. Killing effects of an isolated Serratia marcescens KH-001 on Diaphorina citri via lowering the endosymbiont numbers. Front Microbiol. 2018;9:860.

    Article  Google Scholar 

  35. Arp AP, Martini X, Pelz-Stelinski KS. Innate immune system capabilities of the Asian citrus psyllid, Diaphorina citri. J Invertebr pathol. 2017;148:94–101.

    Article  CAS  PubMed  Google Scholar 

  36. Aggarwal C, Paul S, Nain V, Tripathi V, Khan MA. Comparative response of Spodoptera litura challenged per os with Serratia marcescens strains differing in virulence. J Invertebr Pathol. 2021;183:107562.

    Article  CAS  PubMed  Google Scholar 

  37. Tao A, Wang T, Pang F, Zheng X, Ayra-Pardo C, Huang S. Characterization of a novel chitinolytic Serratia marcescens strain TC-1 with broad insecticidal spectrum. AMB Express. 2022;12:1–13.

    Article  Google Scholar 

  38. Zhang L, Ringbauer JA, Goodman CL, Reall T, Stanley D. Prostaglandin-mediated recovery from bacteremia delays larval development in fall armyworm, Spodoptera frugiperda. Archiv Insect Biochem Physiol. 2018;97:e21444.

    Article  Google Scholar 

  39. De Mandal S, Lin B, Shi M, Li Y, Xu X, Jin F. iTRAQ-based comparative proteomic analysis of larval midgut from the beet armyworm, Spodoptera exigua (Hübner) (Lepidoptera: Noctuidae) challenged with the entomopathogenic bacteria Serratia marcescens. Front Physiol. 2020;11:442.

    Article  PubMed  PubMed Central  Google Scholar 

  40. Lin DJ, Zhou JX, Ali A, Fu HY, Gao SJ, Jin L. Biocontrol efficiency and characterization of insecticidal protein from sugarcane endophytic Serratia marcescens (SM) against oriental armyworm Mythimna separata (Walker). Int Biol Macromol. 2024;262:129978.

    Article  CAS  Google Scholar 

  41. Devi S, Saini HS, Kaur S. Assessing the pathogenicity of gut bacteria associated with tobacco caterpillar Spodoptera litura (Fab.). Sci Rep. 2022;12:8257.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Peng G, Shi X, Kadowaki T. Evolution of TRP channels inferred by their classification in diverse animal species. Mol Phylogenet Evol. 2015;84:145–57.

    Article  CAS  PubMed  Google Scholar 

  43. Georgiev P, Okkenhaug H, Drews A, Wright D, Lambert S, Flick M. TRPM channels mediate zinc homeostasis and cellular growth during Drosophila larval development. Cell Metabol. 2010;12:386–97.

    Article  CAS  Google Scholar 

  44. Hofmann T, Chubanov V, Chen X, Dietz AS, Gudermann T, Montell C. Drosophila TRPM channel is essential for the control of extracellular magnesium levels. PLoS One. 2010;5:e10519.

    Article  PubMed  PubMed Central  Google Scholar 

  45. Hardie RC, Reuss H, Lansdell SJ, Millar NS. Functional equivalence of native light-sensitive channels in the Drosophila trp301 mutant and TRPL cation channels expressed in a stably transfected Drosophila cell line. Cell Calcium. 1997;21:431–40.

    Article  CAS  PubMed  Google Scholar 

  46. Akitake B, Ren Q, Boiko N, Ni J, Sokabe T, Stockand JD. Coordination and fine motor control depend on Drosophila TRPγ. Nat Comm. 2015;6:7288.

    Article  CAS  Google Scholar 

  47. Cheng LE, Song W, Looger LL, Jan LY, Jan YN. The role of the TRP channel NompC in Drosophila larval and adult locomotion. Neuron. 2010;67:373–80.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Fowler MA, Montell C. Drosophila TRP channels and animal behavior. Life Sci. 2021;92:394–403.

    Article  Google Scholar 

  49. Liu L, Li Y, Wang R, Yin C, Dong Q, Hing H. Drosophila hygrosensation requires the TRP channels water witch and nanchung. Nature. 2007;450:294–8.

    Article  CAS  PubMed  Google Scholar 

  50. Thorne N, Chromey C, Bray S, Amrein H. Taste perception and coding in Drosophila. Curr Biol. 2004;14:1065–79.

    Article  CAS  PubMed  Google Scholar 

  51. Turner HN, Armengol K, Patel AA, Himmel NJ, Sullivan L, Iyer SC. The TRP channels Pkd2, NompC, and Trpm act in cold-sensing neurons to mediate unique aversive behaviors to noxious cold in Drosophila. Curr Biol. 2016;26:3116–28.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Edwards-Jorquera SS, Bosveld F, Bellache YA, Lennon-Duménil A-M, Glavic L. Trpml controls actomyosin contractility and couples migration to phagocytosis in fly macrophages. J Cell Biol. 2020;219:3.

    Article  Google Scholar 

  53. Freichel M, Almering J, Tsvilovskyy V. The role of TRP proteins in mast cells. Front Immunol. 2012;3:150.

    Article  PubMed  PubMed Central  Google Scholar 

  54. Fliegert R, Riekehr WM, Guse AH. Does cyclic ADP-Ribose (cADPR) activate the non-selective cation channel TRPM2? Front Immunol. 2020;11:2018.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Robledo-Avila FH, Ruiz-Rosado JDD, Brockman KL, Partida-Sánchez S. The TRPM2 ion channel regulates inflammatory functions of neutrophils during Listeria monocytogenes infection. Front Immunol. 2020;11:97.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Michalick L, Kuebler WM. A missing link between mechanosensation and immunity. Front Immunol. 2020;11:413.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Scheraga RG, Southern BD, Grove LM, Olman MA. The role of TRPV4 in regulating innate immune cell function in lung inflammation. Front Immunol. 2020;11:1211.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Li ZY, Dai QX, Kuang ZL, Liang MR, Wang L, Lu YY, Chen KW. Effects of three artificial diets on development and reproduction of the fall armyworm Spodoptera frugiperda (J. E. Smith). J Environ Entomol. 2019;41:8.

    Google Scholar 

  59. Livak KJ, Schmittgen TDL. Analysis of relative gene expression data using real-time quantitative PCR and the 2-DDCt method. Methods. 2001;25:402–8.

    Article  CAS  PubMed  Google Scholar 

Download references

Funding

This research was supported by the Science and Technology Talent and Platform Project of Yunnan (Academician and Expert Workstation) (202405AF140082), Guangdong Special Support Program-Leading Talents in Science and Technology Innovation (2023TX07A017), the Key-Area Research and Development Program of Guangdong (2020B020223004), and the Guangdong Modern Agriculture Innovation Team Project (2024CXTD21, 2024KJ134).

Author information

Author notes

  1. Hongai Su and Qianxuan Dai contributed equally to this work.

Authors and Affiliations

  1. Department of Entomology, College of Plant Protection, South China Agricultural University, No. 483, Wushan Road, Tianhe District, Guangzhou, 510642, China

    Hongai Su, Qianxuan Dai, Feiyue Teng, Ziyuan Li, Yixiang Qi & Yongyue Lu

Authors
  1. Hongai Su
    View author publications

    You can also search for this author inPubMed Google Scholar

  2. Qianxuan Dai
    View author publications

    You can also search for this author inPubMed Google Scholar

  3. Feiyue Teng
    View author publications

    You can also search for this author inPubMed Google Scholar

  4. Ziyuan Li
    View author publications

    You can also search for this author inPubMed Google Scholar

  5. Yixiang Qi
    View author publications

    You can also search for this author inPubMed Google Scholar

  6. Yongyue Lu
    View author publications

    You can also search for this author inPubMed Google Scholar

Contributions

YXQ and YYL conceived and designed the experimental plan. HAS and QXD analyzed, interpreted the experimental data, and drafted the manuscript. FYT and ZYL reared insects and participated in the experiments. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Yixiang Qi or Yongyue Lu.

Ethics declarations

Ethics approval and consent to participate

The material we used in this study is a notorious agricultural pest, the fall armyworm Spodoptera frugiperda. The strain in this study is a wide used laboratory strain for the oriental fruit fly functional researches. Therefore, it does not involve ethical issues and not need relevant permission.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if you modified the licensed material. You do not have permission under this licence to share adapted material derived from this article or parts of it. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by-nc-nd/4.0/.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Su, H., Dai, Q., Teng, F. et al. Expression profiles analysis and roles in immunity of transient receptor potential (TRP) channel genes in Spodoptera frugiperda. BMC Genomics 26, 401 (2025). https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s12864-025-11599-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s12864-025-11599-6

Keywords

BMC Genomics

ISSN: 1471-2164

Contact us