Research Article

Received: 24 June 2022 Revised: 25 December 2022 Accepted article published: 11 January 2023 Published online in Wiley Online Library: 24 January 2023

(wileyonlinelibrary.com) DOI 10.1002/ps.7355

Cytochrome P450s genes CYP321A9 and

CYP9A58 contribute to host plant adaptation
in the fall armyworm Spodoptera frugiperda
Li He,a Yang Shi,a Wenbing Ding,b,c Hong Huang,d Hualiang He,a Jin Xue,a
Qiao Gao,a Zhixiang Zhang,e Youzhi Lia,b and Lin Qiua*

Abstract
BACKGROUND: The fall armyworm, Spodoptera frugiperda is one of the most destructive agricultural pests, which can complete
their entire life cycle on various plants. At present, some detoxification genes have been proved to be involved in the adaptabil-
ity to plants in insects. However, the genetics behind insect pest responses to host switches, and their ability to adapt to new
host plants, remain poorly understood. This study was conducted to evaluate the adaptation of S. frugiperda to host plant and
determine the roles of CYP321A9 and CYP9A58 in the detoxification metabolism of the fall armyworm.
RESULTS: The results revealed that feeding on maize was more suitable for S. frugiperda to develop compared with rice. In addi-
tion, knocking down of SfCYP321A9 and SfCYP9A58 resulted in a prolonged developmental time of S. frugiperda larvae that fed
on rice. Meanwhile, RNAi knockdown of SfCYP321A9 resulted in significantly higher mortality of S. frugiperda larvae when
exposed to the rice allelochemicals, ferulic acid, gramine and tricin. Furthermore, overexpression of SfCYP321A9 significantly
reduced mortality in Drosophila melanogaster when exposed to gramine and tricin.
CONCLUSION: Our results suggest that CYP321A9 and CYP9A58 genes play a key role in host plant adaptation in S. frugiperda,
which contribute to a greater understanding of the molecular basis of host plant adaptation and provide the means to develop
effective management tools for S. frugiperda resistance.
© 2023 Society of Chemical Industry.

Supporting information may be found in the online version of this article.

Keywords: Spodoptera frugiperda; host adaptation; CYP321A9; CYP9A58; detoxification metabolism; plant allelochemicals

1 INTRODUCTION composition.8 In fact, insects could be threatened by allelochem-

The fall armyworm (Spodoptera frugiperda, Lepidoptera: Noctui-
dae) is one of the most destructive agricultural pests due to its
strong migratory behavior and extreme polyphagy, attacking
large numbers of cultivated plants, thereby causing significant eco-
nomic losses.1 While native to the Americas,2,3 it has recently
invaded Africa and Asia (https://www.cabi.org/isc/fallarmyworm),
most recently spreading rapidly in China.4 S. frugiperda has two
icals from host plants resulting in the changes of the growth and
* Correspondence to: L Qiu, Hunan Provincial Key Laboratory for Biology and
Control of Plant Diseases and Insect Pests, College of Plant Protection,
Hunan Agricultural University, Changsha, 410128, China, China. E-mail:
qiulin@hunau.edu.cn

sympatric strains, the ‘corn strain’ (C strain), which prefer to feed
on maize and other large grasses, and the ‘rice strain’ (R strain) that
preferentially feeds on rice and various pasture grasses,5–7 yet
which can all complete their entire life cycle on various plants own-
ing to its high degree of polyphagy and adaptability. Consequently,
understanding how insects adapt to their host plants is essential to
develop efficient resistance management tools.
Host plant adaptation and host shift have been previously
investigated in S. frugiperda. For example, the performance of
S. frugiperda when it fed on forage species (e.g., Arachis, Axonopus,
and Cynodon) compared to maize, indicated that bermudagrass
was the most suitable alternative host for its development. Fur-
thermore, a multivariate correlation analysis suggested that the
L. He and Y. Shi contributed equally to this work.
a Hunan Provincial Key Laboratory for Biology and Control of Plant Diseases and
Insect Pests, College of Plant Protection, Hunan Agricultural University, Chang-
sha, China
b National Research Center of Engineering & Technology for Utilization of
Botanical Functional Ingredients, Hunan Agricultural University, Changsha,
China
c Hunan Provincial Engineering & Technology Research Center for Biopesticide
and Formulation Processing, Changsha, China
d Hunan Institute of Plant Protection, Changsha, China
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e College of Plant Protection, South China Agricultural University, Guangzhou,

adaptation index positively correlated with the complexity of host China

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development of insects, but in response to the plant stress, the
activity of enzymes in insects was changed to reduce the harm
of allelochemicals to the organizes.9 Current understanding of
the mechanisms involved in the ability of S. frugiperda to adapt
to primary (corn) and alternate (rice) host plants suggest that total
protease, class-specific trypsin, and chymotrypsin-like protease
activities differ significantly when it feeds on the two hosts. The
knockdown of these genes results in increased mortality, suggest-
ing that digestive enzymes and related genes play a crucial role in
the host plant adaptation of S. frugiperda.10 Additionally, func-
tional characterization of unbiased transcriptome revealed that
detoxification genes exhibit a high level of plasticity, indicating
an increased ability to respond rapidly to novel host plants.11
Although the significance of insect adaptation to host plants has
long been recognized,12 the underlying molecular mechanisms
involved in response to their host plant defenses remain unclear.
Previous studies have shown that phenolic acids can affect
insect feeding and development.13,14 Ferulic acid is a methoxy-
hydroxylated derivative of cinnamic acid belonging to the
phenolic acid group which accumulates in rice in response to Nila-
parvata lugens.15 High levels of ferulic acids and p-coumaric are
associated with maize resistance to Sesamia nonagrioides.16 Fur-
ther studies have shown that caffeic, ferulic, and p-coumaric acids
reduce grain aphid populations by 35–45%.13 At the same time,
insects have evolved defense mechanisms in response to pheno-
lic acids. For example, GST activity was induced in Sitobion avenae
by higher concentrations of phenolics in resistant host plants,17
indicating that the application of phenolic acids may induce up-
regulation of aphid GST activity.18 Collectively, these findings sug-
gest that ferulic acid is an antibiotic against many herbivores,
which in turn, induces the expression of the detoxification
enzymes to play a protective role in these insects. Additionally,
it has also been reported that the feeding behavior of pests could
be inhibited by gramine and tricin which were widely found in
rice, resulting in the reduction of their survival rates and reproduc-
tion rates.19,20 Meanwhile, the activities of two important detoxifi-
cation enzymes, CarE and GST were induced to increase when
Sitobion avenae fed on an artificial diet containing gramine.21
However, the mechanism of detoxification enzymes in herbivores
which are regulated in response to rice allelochemicals is poorly
understood.
Increasing studies have shown that the ability to metabolize
and detoxify allelochemicals is considered as a significant evolu-
tionary adaptation in the defense mechanisms used by
herbivores,22 and the genes involved with detoxification, diges-
tion, and transport are regarded as the key factors for host adap-
tation.23 Meanwhile, it is well known that cytochrome P450s can
be regulated by secondary plant metabolites which play a key role
in plant-herbivore interactions, and they fulfill many important
tasks in insects through the building of detoxification defenses
and basal metabolism adaptations (for example, hormone balance,
rate of development and reproduction).24,25 P450s have been com-
posed of CYP2, CYP3, CYP4 and the mitochondrial CYP clade.26
Interestingly, it has been investigated that the CYP3 clan (CYP321A
and CYP9 genes belong to CYP3 clan) was associated with
detoxification.27–29 For example, for CYP321As, the CYP321A2/
CYP321A1 paralogs in Helicoverpa zea that are induced by the plant
allelochemicals flavone and coumarin, as well as the flavone-
responsive regulatory element XRE-Fla, were detected in the pro-
moter region of the above genes.30 The CYP321A9 gene has been
proved to be induced by xanthotoxin in S. frugiperda.31 Further-
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more, for CYP9s, RNA-seq analysis showed the complex expression
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L He et al.
profile of the CYP9A clustered genes exposed to xanthotoxin and
ricin in S. litura.32 RNAi knockdown of S. litura CYP9A40 resulted in
increased susceptibility to quercetin, cinnamic acid, delta-methrin
and methoxyfenozide.33 The overexpression of CYP9A59 and a
CYP9A-like gene were responsible for deltamethrin and chlorpyrifos
resistant S. frugiperda.34 The above observations show that
CYP321A9 and CYP9A58 belong to CYP3 clan may be also involved
in the defense mechanisms of herbivores. However, it is unclear
that whether CYP321A9 and CYP9A58 are involved in the detoxifica-
tion of plant allelochemicals (ferulic acid, gramine and tricin), and
the underlying mechanism for these two genes regulating allelo-
chemicals remains unclear.
The purpose of this study was to understand the adaptation of
S. frugiperda to host plants and to evaluate the genes involved
in allelochemical detoxification. In this study, we found that
S. frugiperda larvae was more adapted to corn plants compared
with rice plants. Additionally, CYP321A9 and CYP9A58 were eventu-
ally chosen to complete the next experiments according to the lit-
erature review. Then RNAi knockdown of cytochrome P450
(SfCYP321A9 and SfCYP9A58) resulted in increased mortality of lar-
vae feeding on rice plants compared to control larvae. Meanwhile,
we performed allelochemical bioassays using RNAi knockdown and
overexpression of the SfCYP321A9 gene in S. frugiperda and
D. melanogaster, respectively. Our results provide an insight into
the molecular basis of host plant adaptation, and the means to
develop efficient resistance management tools for S. frugiperda.
2 MATERIALS AND METHODS
2.1 Insects
S. frugiperda larvae were originally collected from maize fields in
Chenzhou City, Hunan province, China, in May 2019, belonging
to the ‘corn strain’. The larvae were reared separately on an artifi-
cial diet, maize leaves and rice leaves under laboratory conditions
and were maintained in a light incubator (at 25 ± 1 °C, relative
humidity (RH) of 70 ± 10% under a 14 h light, 10 h dark photope-
riod). Adults were fed 10% sucrose.
2.2 Determining the host plant adaptation of
S. frugiperda
To evaluate the fitness of S. frugiperda fed with maize and rice,
newly hatched larvae were reared on corn and rice leaves, respec-
tively. The leaves were washed with distilled water, then dried at
room temperature before being fed to the larvae. To prevent can-
nibalism, once the larvae reached their third instar they were
transferred to small dishes with an 8.5 cm diameter and a height
of 4 cm. The dishes were cleaned daily to remove feces and leaf
residues, and fresh leaves were added. The developmental time
of larvae was recorded daily, with three replications carried out
for each treatment. Larval and pupal weights were also recorded.
A total of 45 larvae were used with three replicates in each
treatment.
2.3 Production of SfCYP321A9 and SfCYP9A58 double-
stranded RNA
The method of dsRNA expression35 was used to express
CYP321A9 or CYP9A58 dsRNA derived from the CYP321A9 and
CYP9A58 genes with specific primers (Table S1). We con-
structed two fragments into the pET-2p plasmid and
sequenced them. We then transformed the recombinant plas-
mid into Escherichia coli HT115 (DE3) competent cells and inoc-
ulated the individual colonies in a Luria broth medium
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supplemented with 50 mg μL−1 kanamycin. Positive clones
were induced to express dsRNA by adding 0.1 mmol L−1 IPTG
(isopropyl ⊎-D-1-thiogalactopyranoside) until the growing cul-
tures reached an OD600 value of 1.0, after which the bacteria
were cultured for an additional 4 h. The quality of the dsRNA
was verified by electrophoresis on 1% agarose gel. Bacteria
were centrifuged at 5000 g for 10 min and resuspended in
0.05 M phosphate buffered saline (PBS, pH 7.4) at a ratio of
20:1 (concentration of ×10) to obtain the bacterial suspensions
containing dsRNA.
2.4 RNA interference knockdown of SfCYP321A9 and
SfCYP9A58
Two independent bioassays were carried out. Firstly, newly
hatched larvae that were fed on rice leaves, were treated with
an E. coli suspension containing either CYP321A9, CYP9A58, or
EGFP dsRNA. Ten larvae were transferred to a dish as a repeat,
and each treatment was replicated four times. A total of four
dishes were used. To extract total RNA for qPCR, four replicates
were collected after continuous feeding for 24, 48, 72, and
96 h, with 10 larvae were used in each replication. The remaining
larvae were used to observe growth and development. Sec-
ondly, newly hatched larvae were fed an artificial diet immersed
in a bacterial suspension containing dsRNA. After 48 h of feed-
ing, 15 individual larvae were transferred to a Petri dish
containing an artificial diet supplemented with ferulic acid, gra-
mine, or tricin. The larvae were placed in an incubator at 26 °C,
and larval mortality was recorded.
2.5 Construction of transgenic Drosophila melanogaster
and bioassays in vivo
The SfCYP321A9 gene was selected as a research representa-
tive to verify the involvement of the P450 genes in deto-
xifying allelochemicals in transgenic D. melanogaster.
Specifically, the ORF of the SfCYP321A9 gene was amplified
from S. frugiperda cDNA using Phanta® Max Super-Fidelity
DNA Polymerase (Fermentas). The purified PCR products were
individually cloned into the pJFRC-MUH vector between the
XbaI and NotI (Fermentas) sites to prepare the UAS-
SfCYP321A9 construct. Using the PhiC31 system, the recombi-
nant plasmid was injected into the germline of the w1118
strain, which carries the attP40 docking site on chromosome
2. The transgene CYP321A9 was expressed by crossing it with
the Act5C-GAL4 strain. PCR analysis was used to confirm the
expressions of the SfCYP321A9 gene in transgenic
D. melanogaster using gene-specific primers (Table S1). For
bioassays, 15 flies (2–5 days after eclosion) were added to a
vial which contained 4 mL of a corn meal medium supplemen-
ted with 30 mg mL −1 gramine, or 0.5 mg mL −1 tricin and
placed in an incubator at 25 °C under a 12:12 (Light: Dark)

Figure 1. The fitness of S. frugiperda larvae that fed on rice and maize. (a) The larval developmental period of S. frugiperda that fed on corn and rice. Each
color represents the developmental time of insects that fed on rice and maize. 1st to 6th indicate the larval instars. (b) Pupal developmental periods.
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(c) Larval weight. Instars are abbreviated (L1 to L6). (d) Pupal weight of S. frugiperda that fed on corn and rice. Asterisks indicate significant difference
between the two groups (*P < 0.05, ***P < 0.001, Student's t test).

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Figure 2. Oral RNA interference knockdown of SfCYP321A9 and SfCYP9A58 transcripts in S. frugiperda larvae and associated developmental time after
feeding on rice leaves. (a) Relative estimates of SfCYP321A9 transcripts from qPCR of S. frugiperda larvae that fed on rice treated with either SfCYP321A9
or EGFP dsRNA for 24, 48, 72, and 96 h. Student's t tests, compared to that of the EGFP dsRNA group. n.s., no significant difference (*P < 0.05, ***P < 0.001).
(b) Relative estimates of SfCYP9A58 transcripts from qPCR of S. frugiperda larvae that fed on rice treated with either SfCYP9A58 or EGFP dsRNA for 24, 48,
72 and 96 h. Student's t test compared to that of EGFP dsRNA group. n.s. = no significant difference (*P < 0.05, **P < 0.01). (c) The developmental time of
larvae after continuous feeding on SfCYP321A9 and SfCYP9A58 dsRNA. Data are shown as means ± SE derived from three or five biological replicates.

photoperiod. The male progeny were from the cross between
the Act5C-GAL4 and w1118 strain that possessed the attP40
docking site but did not carry the transgene. This strain was
also used as the control. Mortality was recorded after 24, 48,
72, 96, 120, 144 h. Six replicates were used for each
experiment.
2.6 Quantitative real-time PCR
Total RNA (1 μg) was reverse transcribed using the PrimeScript RT
reagent kit with gDNA eraser (TaKaRa). qPCR reactions were car-
ried out on a CFX-96™ PCR Detection System (Bio-Rad) using qPCR
Mastermix plus for SYBR® Premix Ex Taq™ (TaKaRa). The PCR con-
ditions were as follows: 1 cycle at 95 °C for 30 s, 40 cycles at 95 °C
for 10 s, and 59 °C for 30 s. A fivefold dilution series of cDNA was
used to construct a relative standard curve. Melting curve analysis
and standard curves were performed from 55 to 95 °C to assess
primer specificity and PCR efficiencies, respectively. The relative
expression and each target gene were assessed according to
the 2−ΔΔCT method, incorporating PCR efficiency after normaliza-
tion with the housekeeping genes ⊎-Actin and Rpl32.36,37
2.7 Statistical analysis
The statistical significance of differences was analyzed using Stu-
dent's t test for two samples and one-way ANOVA for more than
two samples in Graphpad Prism 7.0. Differences were considered
significant if the P-value was <0.05.
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3 RESULTS
3.1 Effects on the development of S. frugiperda by
feeding on different plants
To investigate how invading S. frugiperda larvae adapt to living on
rice plants (alternative host), larvae were fed maize (preferred
host) and rice for three generations. Larvae fed on rice had a sig-
nificantly longer developmental time compared to those fed on
corn (Fig. 1(a)). The mean developmental time for rice and
maize-fed larvae was 20.26 ± 1.43 days and 18.74 ± 1.34 days,
respectively. Pupal development and weight were also affected
significantly by the host plant. The developmental period of pupa
was longer on rice (9.66 ± 0.75 days) compared to maize (7.98
± 0.99 days) (Fig. 1(b)). The larval weight was also affected signif-
icantly by hosts (Fig. 1(c)). Pupal weight was significantly different
on different hosts; it was heavier on maize (0.19 g) than on rice
(0.16 g) (Fig. 1(d)).
3.2 RNAi knockdown of CYP321A9 and CYP9A58
transcripts
To evaluate the functions of the CYP321A9 and CYP9A58 genes in
the host adaptation of S. frugiperda, we performed RNAi-mediated
knockdown of these two genes. Our results showed that the
expression levels of CYP321A9 and CYP9A58 were significantly
reduced by oral CYP321A9 and CYP9A58 dsRNA after 48, 72, and
96 h, compared to the dsEGFP control (Fig. 2(a)–(b)). Additionally,
developmental time was increased for larvae that were fed rice
leaves treated with CYP321A9 (22.96 ± 1.38 day) or CYP9A58

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Figure 3. RNA interference knockdown of SfCYP321A9 and SfCYP9A58 transcripts in S. frugiperda larvae and subsequent mortality of larvae after feeding
on plant allelochemicals. (a) Mortality of larvae fed on SfCYP321A9 and SfCYP9A58 dsRNA before being exposed to ferulic acid after 48 h. (b) Mortality of
larvae fed on SfCYP321A9 and SfCYP9A58 dsRNA before being exposed to gramine after 48 h. (c) Mortality of larvae fed on SfCYP321A9 and SfCYP9A58
dsRNA before being exposed to tricin after 48 h.

Figure 4. Mortality of D. melanogaster with overexpression of SfCYP321A9, after feeding on plant allelochemicals. (a) The mortality of D. melanogaster at
the 30 mg mL−1 gramine. Student's t test compared to that of the control (*P < 0.05). (b) The mortality of D. melanogaster at 0.5 mg mL−1 tricin. Student's
t test compared to the control (*P < 0.05, **P < 0.01).

(22.86 ± 1.34 day) dsRNA compared to those fed with rice leaves
treated with EGFP dsRNA (20.40 ± 1.64 day) (Fig. 2(c)).
3.3 SfCYP321A9 and SfCYP9A58 are involved in the
detoxification of plant allelochemcials
S. frugiperda larvae fed artificial diets treated with dsCYP321A9 for
48 h and subsequently supplemented with 0.8% ferulic acid
showed significantly increased levels of mortality by 6%, 14%
24, 48 and 72 h, respectively (Fig. 3(a)). Similarly, the knockdown
of CYP321A9 could significantly increase the mortality of larvae
exposed to 0.05% tricin by 29%, 41% and 40% at 24, 48 and
72 h, respectively (Fig. 3(c)). However, there was no significance
between the mortality of larvae fed a diet treated with 0.8% gra-
mine and the controls at 24 h. But compared with the controls,
the mortality of larvae fed dsCYP321A9 was significantly increased
by 22% and 27% in S. frugiperda exposed to 0.8% gramine at 48 h
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and 23% compared with those treated with dsEGFP at and 72 h, respectively (Fig. 3(b)). In addition, the knockdown of

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CYP9A58 also could significantly increase the mortality of
S. frugiperda exposed to 0.8% ferulic acid or 0.05% tricin at 72 h.
However, there was no significant difference between the mortal-
ity of larvae treated with dsCYP9A58 and the controls under 0.8%
gramine exposure (Fig. 3). These results indicated the possible
roles of the S. frugiperda CYP321A9 or CYP9A58 genes in plant alle-
lochemical detoxification.
3.4 Overexpression of CYP321A9 enhances
detoxification of allelochemicals
To further investigate the role of the SfCYP321A9 gene in allelo-
chemical detoxification, the GAL4/UAS system and Act5C-GAL4
strain was used to express SfCYP321A9 in transgenic
D. melanogaster. The expression of CYP321A9 in the F1 progeny
was confirmed by PCR and qPCR (Fig. S1). Following its feeding
on an artificial diet treated with 30 mg mL−1 gramine, the mortal-
ity of D. melanogaster expressing CYP321A9 was significantly
reduced by 15% and 20% compared to the control group at
24 and 48 h, respectively. However, there was no significant dif-
ference under 30 mg mL−1 gramine exposure at 72, 96, 120 and
144 h (Fig. 4(a)). In addition, overexpression of CYP321A9 could
significantly reduce the mortality of D. melanogaster exposed to
0.5 mg mL−1 tricin at 72 or 96 h. And the mortality of
D. melanogaster expressing CYP321A9 were lower than the con-
trols at other times under 0.5 mg mL−1 tricin exposure but no sig-
nificant difference between them (Fig. 4(b)).
4 DISCUSSION
There is growing evidence which indicates that transcriptional
plasticity in some herbivorous insects, is involved in the detoxifi-
cation of secondary plant metabolites. However, further studies
are required to reveal the truth of genetics behind insect pest
responses to host switches. In our study, we compared the effects
of different host-plants on the growth and development of
S. frugiperda to determine its host-plant adaptation. In addition,
detoxification metabolism was analyzed to provide a greater
understanding of the mechanisms underlying S. frugiperda host
adaptation and host shift, which enables a better assessment of
the risk of the damage caused to rice by the ‘corn’ strain of
S. frugiperda, to develop effective pest management strategies.
The development and population dynamics of S. frugiperda are
affected by differences between hosts.8 In this study, S. frugiperda
fed on rice had a longer larval developmental time compared to
those fed on maize (Fig. 1). This result is consistent to the finding
in the previous study that the larval developmental time on rice
(alternative host) was longer than on corn (primary host) in the
CS and RS strains of S. frugiperda.11 Above findings further prove
that corn is more suitable for S. frugiperda to develop, also leading
us to suspect that there may be some substances in rice that is not
suitable for the growth and development of it. The reason for this
speculation is that the growth of insects has been found to be
affected by some substances which were composed of secondary
metabolites could be released from plants infested by pests.9 For
example, the pupal weight and adult emergence of S. frugiperda
could be significantly affected by the chloroform extract of
S. crotalarioides.38 Therefore, evaluating the relation between
plant secondary substances and the growth and development
of S. frugiperda would be a meaningful topic for future research,
which is beneficial to develop phytochemical defense strategy
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and its application in agriculture.
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L He et al.
In this study, the developmental time of larvae which fed rice
leaves treated CYP321A9 or CYP9A58 dsRNA was increased com-
pared with the controls, which further revealed that sfCYP321A9
and sfCYP9A58 might be responsible for S. frugiperda adaptation
to rice. Additionally, our results also further revealed that the
knockdown of sfCYP321A9 or sfCYP9A58 significantly increased
the mortality of S. frugiperda exposed to ferulic acid, gramine
and tricin. This finding was consistent with previous study show-
ing that RNAi of CYP9AQ1 increased the mortality of Locusta
migratoria exposed to fluvalinate.39 Similarly, the knockdown of
CYP321B1 resulted in significantly increased mortality in S. litura
exposed to hlorpyrifos and ⊎-cypermethrin.40 These findings
prove that sfCYP321A9 and sfCYP9A58 may be involved in the
detoxification of plant allelochemicals, the role of sfCYP321A9 of
which were also supported by our results that overexpression of
sfCYP321A9 could enhance the detoxification of allelochemical
in transgenic D. melanogaster expressing sfCYP321A9. Taken
together, CYP321A9 and CYP9A58 may play an important role in
the detoxification of ferulic acid, gramine and tricin in
S. frugiperda, providing a basis to further understand the molecu-
lar mechanisms of host plant adaptation.
Furthermore, it has been reported that insect susceptibility to
insecticides is closely related to plant allelochemicals at the gene
level.41 In our study, CYP321A9 and CYP9A58 were involved in the
metabolism of ferulic acid, gramine and tricin. Interestingly, these
two genes also have been found to be significantly upregulated
in S. frugiperda exposed to chlorantranilipole, an insecticide.42 Sim-
ilarly, CYP6B2 and CYP6B7 were involved in the detoxification
metabolism of permethrin in Helibthis armigera, which also could
be upregulated by monoterpenoids.43,44 The above findings
revealed that insecticide resistance of insect could be affected by
insect defense response to plant allelochemicals. Additionally, sev-
eral studies have shown that allelochemicals can compete with
insecticides for action sites in insects to increase the toxicity of
insecticides.45 However, it is unknown whether CYP321A9 and
CYP9A58 can be served as common target sites of rice allelochem-
icals (ferulic acid, gramine and tricin) and chlorantranilipole for pest
management. Therefore, more research is required to further verify
the role of CYP321A9 and CYP9A58 to determine whether these alle-
lochemicals can be used as synergists for insecticides, which is con-
ductive to develop a more effective pest control strategy.
5 CONCLUSION
Numerous complex interactions occur between plants and herbi-
vores. During their co-evolution, herbivores have evolved various
strategies for metabolizing and detoxifying plant toxins.
S. frugiperda is a global pest that has already caused immense eco-
nomic losses worldwide. In this study, we discovered that
S. frugiperda fed on maize that had a better development com-
pared with those fed on rice. Meanwhile, combining RNAi tech-
nologies with tests in vitro, SfCYP321A9 and SfCYP9A58 were
found to be involved in the metabolism of rice allelochemicals
in S. frugiperda. These findings provide further understanding of
the molecular mechanisms underlying host plant adaptation
and the development of effective tools for managing
S. frugiperda resistance.
ACKNOWLEDGEMENTS
This research was supported by the key research and develop-
ment program of Hunan Province (China) (2020NK2034), the
Pest Manag Sci 2023; 79: 1783–1790

P450 mediate host adaptation of S. frugiperda
science and technology innovation Program of Hunan Province
(2022RC1025) and the Double first-class construction project of
Hunan Agricultural University (SYL2019029).
CONFLICT OF INTEREST
The authors declare no competing interests.
DATA AVAILABILITY STATEMENT
The data that supports the findings of this study are available in
the supplementary material of this article
SUPPORTING INFORMATION
Supporting information may be found in the online version of this
article.
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