The Circadian Clock Gene PHYTOCLOCK1 Mediates the Diurnal Emission of the Anti-Insect Volatile Benzyl Nitrile from Damaged Tea (Camellia sinensis) Plants.

Benzyl nitrile from tea plants attacked by various pests displays a diurnal pattern, which may be closely regulated by the endogenous circadian clock. However, the molecular mechanism by the circadian clock of tea plants that regulates the biosynthesis and release of volatiles remains unclear. In this study, the circadian clock gene CsPCL1 can activate both the expression of the benzyl nitrile biosynthesis-related gene CsCYP79 and the jasmonic acid signaling-related transcription factor CsMYC2 involved in upregulating CsCYP79 gene, thereby resulting in the accumulation and release of benzyl nitrile. Therefore, the anti-insect function of benzyl nitrile was explored in the laboratory. The application of slow-release beads of benzyl nitrile in tea plantations significantly reduced the number of tea geometrids and had positive effects on the yield of fresh tea leaves. These findings reveal the potential utility of herbivore-induced plant volatiles for the green control of pests in tea plantations.

Time series prediction of insect pests in tea gardens.

BACKGROUND: Tea-garden pest control is crucial to ensure tea quality. In this context, the time-series prediction of insect pests in tea gardens is very important. Deep-learning-based time-series prediction techniques are advancing rapidly but research into their use in tea-garden pest prediction is limited. The current study investigates the time-series prediction of whitefly populations in the Tea Expo Garden, Jurong City, Jiangsu Province, China, employing three deep-learning algorithms, namely Informer, the Long Short-Term Memory (LSTM) network, and LSTM-Attention. RESULTS: The comparative analysis of the three deep-learning algorithms revealed optimal results for LSTM-Attention, with an average root mean square error (RMSE) of 2.84 and average mean absolute error (MAE) of 2.52 for 7 days' prediction length, respectively. For a prediction length of 3 days, LSTM achieved the best performance, with an average RMSE of 2.60 and an average MAE of 2.24. CONCLUSION: These findings suggest that different prediction lengths influence model performance in tea garden pest time series prediction. Deep learning could be applied satisfactorily to predict time series of insect pests in tea gardens based on LSTM-Attention. Thus, this study provides a theoretical basis for the research on the time series of pest and disease infestations in tea plants. © 2024 Society of Chemical Industry.

Comparative transcriptomic analysis of resistant and susceptible tea cultivars in response to Empoasca onukii (Matsuda) damage.

MAIN CONCLUSION: Transcriptomic studies in resistant and susceptible tea cultivars have been performed to reveal the different defense molecular mechanisms of tea after E. onukii feeding. The molecular mechanism by which tea plants respond to small green leafhopper Empoasca onukii (Matsuda) damage is unclear. Using the resistant tea plant cultivar Juyan (JY) and the susceptible tea plant cultivar Enbiao (EB) as materials, this study performed RNA-seq on tea leaf samples collected at three time points (6 h, 12 h, 24 h) during exposure of the plants to leafhopper to reveal the molecular mechanisms that are activated in susceptible and resistant tea plant cultivars in response to leafhopper damage. The numbers of DEGs in the susceptible tea cultivar during early (6 h) and late (24 h) stages of leafhopper induction were higher than those in the resistant cultivar at the same time points. The stress responses to leafhopper were most intense at 12 h in both tea cultivars. Pathway enrichment analysis showed that most up-regulated DEGs and their related metabolic pathways were similar in the two tea cultivars. However, during the early stage of leafhopper induction (6 h), jasmonic acid (JA)-related genes were significantly up-regulated in the resistant cultivar. The terpenoid biosynthetic pathway and the α-linolenic acid metabolic pathway were activated earlier in the resistant cultivar and remained activated until the late stage of leafhopper damage. Our results confirmed that after leafhopper damage, the resistant tea cultivar activated its defense responses earlier than the susceptible cultivar, and these defense responses were mainly related to terpenoid metabolism and JA biosynthetic pathway. The results provide important clues for further studies on resistance strategy of tea plants to pest.

Proteomic analysis of "Oriental Beauty" oolong tea leaves with different degrees of leafhopper infestation.

RATIONALE: Oriental Beauty, a type of oolong tea native to Taiwan, is highly prized by connoisseurs for its unique fruity aroma and sweet taste. Leaves of Oriental Beauty vary in appearance, aroma, and taste, depending on the degree of tea green leafhopper (Jacobiasca formosana) infestation. In this study, the aim is to investigate the differential expression of proteins in leaves with low, medium, and high degrees of leafhopper infestation. METHODS: Proteomic techniques 2DE (two-dimensional electrophoresis) and nanoscale liquid chromatography/tandem mass spectrometry (LC/MS/MS) were used to investigate the differential expression of proteins in tea leaves with different degrees of leafhopper infestation. RESULTS: A total of 89 proteins were found to exhibit significant differences in expression. In a gene ontology analysis, most of these proteins participated in biosynthesis, carbohydrate metabolism, transport, responses to stress, and amino acid metabolism. CONCLUSIONS: These results indicated that the unique aroma and taste of the leaves might be influenced by their protein expression profiles, as well as related factors such as defensive responses to tea green leafhopper saliva.

Ecologically controlling insect and mite pests of tea plants with microbial pesticides: a review.

Insect and mite pests are damaging stressors that are threatening the cultivation of tea plants, which result in enormous crop loss. Over the years, the effectiveness of synthetic pesticides has allowed for its prominent application as a control strategy. However, the adverse effects of synthetic pesticides in terms of pesticide residue, environmental contamination and insect pest resistance have necessitated the need for alternative strategies. Meanwhile, microbial pesticides have been applied to tackle the damaging activities of the insect and mite pests of tea plants, and their performances were scientifically adjudged appreciable and environmental friendly. Herein, entomopathogenic microbes that were effective against tea geometrid (Ectropis obliqua Prout), tea green leafhopper (Empoasca onukii Matsuda), paraguay tea ampul (Gyropsylla spegazziniana), tea mosquito bug (Helopeltis theivora Waterhouse) and red spider mite (Oligonychus coffea Nietner) have been reviewed. The current findings revealed that microbial pesticides were effective and showed promising performances against these pests. Overall, this review has provided the basic and integrative information on the integrated pest management (IPM) tool(s) that can be utilized towards successful control of the aforementioned insect and mite pests.

Identification and evaluation of reliable reference genes for quantitative real-time PCR analysis in tea plants under differential biotic stresses.

The selection of reliable reference genes (RGs) for normalization under given experimental conditions is necessary to develop an accurate qRT-PCR assay. To the best of our knowledge, only a small number of RGs have been rigorously identified and used in tea plants (Camellia sinensis (L.) O. Kuntze) under abiotic stresses, but no critical RG identification has been performed for tea plants under any biotic stresses till now. In the present study, we measured the mRNA transcriptional levels of ten candidate RGs under five experimental conditions; these genes have been identified as stable RGs in tea plants. By using the ΔCt method, geNorm, NormFinder and BestKeeper, CLATHRIN1 and UBC1, TUA1 and SAND1, or SAND1 and UBC1 were identified as the best combination for normalizing diurnal gene expression in leaves, stems and roots individually; CLATHRIN1 and GAPDH1 were identified as the best combination for jasmonic acid treatment; ACTIN1 and UBC1 were identified as the best combination for Toxoptera aurantii-infested leaves; UBC1 and GAPDH1 were identified as the best combination for Empoasca onukii-infested leaves; and SAND1 and TBP1 were identified as the best combination for Ectropis obliqua regurgitant-treated leaves. Furthermore, our results suggest that if the processing time of the treatment was long, the best RGs for normalization should be recommended according to the stability of the proposed RGs in different time intervals when intragroup differences were compared, which would strongly increase the accuracy and sensitivity of target gene expression in tea plants under biotic stresses. However, when the differences of intergroup were compared, the RGs for normalization should keep consistent across different time points. The results of this study provide a technical guidance for further study of the molecular mechanisms of tea plants under different biotic stresses.

Characterization of Terpene Synthase from Tea Green Leafhopper Being Involved in Formation of Geraniol in Tea (Camellia sinensis) Leaves and Potential Effect of Geraniol on Insect-Derived Endobacteria.

When insects attack plants, insect-derived elicitors and mechanical damage induce the formation and emission of plant volatiles that have important ecological functions and flavor properties. These events have mainly been studied in model plants, rather than crop plants. Our study showed that tea green leafhopper (Empoasca (Matsumurasca) onukii Matsuda), a major pest infesting tea attack significantly induced the emission of geraniol from tea leaves, but did not affect the crude enzyme activity of geraniol synthase in tea leaves. An enzyme extract of E. (M.) onukii specifically produced geraniol from geraniol diphosphate. Furthermore, a terpene synthase (EoTPS) was isolated from E. (M.) onukii. This terpene synthase was able to convert geraniol diphosphate to geraniol in vitro. In addition, geraniol had in vitro ability to inhibit the growth of Acinetobacter johnsonii that is endobacterial isolated from E. (M.) onukii. This information illustrates that elicitors from piercing-sucking insects can induce the formation of volatiles from crop plants and advances our understanding of the roles of plant volatiles in the interaction among crops-insects-microorganisms.

Systemic identification and analyses of genes potentially involved in chemosensory in the devastating tea pest Basilepta melanopus.

Basilepta melanopus is a serious insect pest of tea plantations in southern China. This tea pest poses a great threat to the tea industry in China. No effective and environmentally friendly methods have been established to control this pest at present. Olfactory genes play key roles in insect behaviour, and can potentially be used as targets for developing environmentally-friendly approaches for pest control. In this study, we produced a transcriptome derived from dissected antennae from B. melanopus using high-throughput sequencing. We identified gene families that are potentially involved in odorant reception and detection, including unigenes encoding 63 odorant receptors (ORs), 16 gustatory receptors (GRs), 18 ionotropic receptors (IRs), four sensory neuron membrane proteins (SNMPs), 46 odorant binding proteins (OBPs), and 19 chemosensory proteins (CSPs). Analyses of tissue expression profiles revealed that all 63 OR transcripts, 14 antennal IRs, one SNMP and six OBPs were predominately expressed in antennae. Real-time quantitative PCR assays were also adapted to examine sex-biased expression of selected antenna-predominant genes. Our results provide valuable information for further functional studies of olfactory genes in B. melanopus and potential novel targets for developing new pest control measures.

Effect of Major Tea Insect Attack on Formation of Quality-Related Nonvolatile Specialized Metabolites in Tea ( Camellia sinensis) Leaves.

Insect attack is known to induce a high accumulation of volatile metabolites in tea ( Camellia sinensis). However, little information is available concerning the effect of insect attack on tea quality-related nonvolatile specialized metabolites. This study aimed to investigate the formation of characteristic nonvolatile specialized metabolites in tea leaves in response to attack by major tea insects, namely, tea green leafhoppers and tea geometrids, and determine the possible involvement of phytohormones in metabolite formation resulting from insect attack. Both tea green leafhopper and tea geometrid attacks increased the jasmonic acid and salicylic acid contents. The abscisic acid content was only increased under tea green leafhopper attack, perhaps due to special continuous piercing-sucking wounding. Tea green leafhopper attack induced the formation of theaflavins from catechins under the action of polyphenol oxidase, while tea geometrid attack increased the l-theanine content. Exogenous phytohormone treatments can affect the caffeine and catechin contents. These results will help to determine the influence of major tea pest insects on important tea quality-related metabolites and enhance understanding of the relationship of phytohormones and quality-related nonvolatile metabolite formation in tea exposed to tea pest insect attacks.

Positive effects of the tea catechin (-)-epigallocatechin-3-gallate on gut bacteria and fitness of Ectropis obliqua Prout (Lepidoptera: Geometridae).

Ectropis obliqua Prout (Lepidoptera: Geometridae) is the most devastating insect pest of tea plants in China and infests thousands of hectares of tea plantations in China annually. (-)-Epigallocatechin-3-gallate (EGCG) is a major phenolic compound in tea leaves and has a strong antibacterial function. Here, we show that EGCG can effectively improve the fitness of E. obliqua larvae and present the reason by which EGCG promotes larval fitness. In this study, we compared the fitness difference among Control, Antibiotic and Treatment of larvae. The fitness of larvae treated with EGCG and antibiotic was similar and better than that of control group. We also demonstrated that EGCG treatment could significantly reduce species richness and abundance of gut bacteria in E. obliqua larvae. Hence that we speculate that EGCG promotes larval fitness and is associated with ECGG antimicrobial activity. In short, our study provides evidence of the E. obliqua larvae have adapted to secondary compounds found in tea leaves, and may even benefit from these compounds. Our study also contributes to a greater understanding of the reason involved in plant-insect interactions.

Transcriptomic and Phytochemical Analyses Reveal Root-Mediated Resource-Based Defense Response to Leaf Herbivory by Ectropis oblique in Tea Plant ( Camellia sinensis).

Leaf herbivory on tea plants ( Camellia sinensis) by tea geometrids ( Ectropis oblique) severely threaten the yield and quality of tea. In previous work, we found that local defense response was induced in damaged leaves by geometrids at the transcriptome level. Here, we investigated the systemic response triggered in undamaged roots and the potential role of roots in response to leaf herbivory. Comparative transcriptome analysis and carbohydrate dynamics indicated that leaf herbivory activated systemic carbon reallocation to enhance resource investment for local secondary metabolism. The crucial role of jasmonic acid and the involvement of other potential hormone signals for local and systemic signaling networks were supported by phytohormone quantification and dynamic expression analysis of phytohormone-related genes. This work represents a deep understanding of the interaction of tea plants and geometrids from the perspective of systems biology and reveals that tea plants have evolved an intricate root-mediated resource-based resistance strategy to cope with geometrid attack.

A Disease Resistance Elicitor Laminarin Enhances Tea Defense against a Piercing Herbivore Empoasca (Matsumurasca) onukii Matsuda.

The tea plant (Camellia sinensis) suffers heavily from a harmful piercing pest, the tea green leafhopper (TLH) Empoasca (Matsumurasca) onukii Matsuda. In the present study, we studied the effect of an efficient elicitor of plant disease resistance, the ß-1,3-glucan laminarin, on the induced defense against TLH in tea plants. Defense responses elicited by laminarin in tea include the activation of mitogen-activated protein kinases and WRKY, the burst of H2O2, salicylic acid, and abscisic acid, and the accumulation of direct-defense chemicals (including chitinase, phenylalanine ammonia lyase, callose, polyphenol oxidase, and flavonol synthase), as well as the production of volatile compounds. The laminarin-treated tea plants reduced the performance of TLH and enhanced the attractiveness to the egg parasitoid wasp of TLH, Stethynium empoascae Subba Rao. In the field experiment, laminarin application effectively reduced the number of TLH by attracting parasitoids. These results suggest that laminarin can induce protection against TLH by regulating signaling pathways in tea plant. Our study also proposes an environment friendly strategy for the integrated management of an economically important piercing pest.

Understanding the biosyntheses and stress response mechanisms of aroma compounds in tea (Camellia sinensis) to safely and effectively improve tea aroma.

Metabolite formation is a biochemical and physiological feature of plants developed as an environmental response during the evolutionary process. These metabolites help defend plants against environmental stresses, but are also important quality components in crops. Utilizing the stress response to improve natural quality components in plants has attracted increasing research interest. Tea, which is processed by the tender shoots or leaves of tea plant (Camellia sinensis (L.) O. Kuntze), is the second most popular beverage worldwide after water. Aroma is an important factor affecting tea character and quality. The defense responses of tea leaves against various stresses during preharvest (tea growth process) and postharvest (tea manufacturing) processing can result in aroma formation. Herein, we summarize recent investigations into the biosyntheses of several characteristic aroma compounds prevalent in teas and derived from volatile fatty acid derivatives, terpenes, and phenylpropanoids/benzenoids. Several key aroma synthetic genes from tea leaves have been isolated, cloned, sequenced, and functionally characterized. Biotic stress (such as tea green leafhopper attack) and abiotic stress (such as light, temperature, and wounding) could enhance the expression of aroma synthetic genes, resulting in the abundant accumulation of characteristic aroma compounds in tea leaves. Understanding the specific relationships between characteristic aroma compounds and stresses is key to improving tea quality safely and effectively.

Benzothiadiazole and B-Aminobutyricacid Induce Resistance to Ectropis Obliqua in Tea Plants (Camellia Sinensis (L.) O. Kuntz).

In order to investigate the effect of benzothiadiazole (BTH) and ß-aminobutyric acid (BABA) on the resistance of tea plants (Camellia sinensis) to tea geometrid (Ectropis obliqua), three levels each of benzothiadiazole (BTH) and ß-aminobutyric acid (BABA) were sprayed on 10-year-old tea plants. Generally PPO and PAL activities increased with low concentrations of BTH and BABA treatments. Quantitative RT-PCR revealed a 1.43 and 2.72-fold increase in PPO gene expression, and 3.26 and 3.99-fold increase in PAL gene expression with 75 mg/L BTH and 400 mg/L BABA respectively. Analysis of hydrolysis of synthetic substrates also revealed that chymotrypsin-like enzyme activity present in larval midgut extracts was not significantly inhibited by BTH and BABA. However, proteinase activity was found to be inversely proportional to the age of tea geometrid. Larvae pupation rate decreased by 8.10, 10.81 and 21.62% when tea geometrid were fed with leaves treated with 25, 50 and 75 mg/L BTH solutions, while 100, 200 and 400 mg/L BABA solutions decreased same by 8.10, 16.21 and 13.51% respectively. Also, larvae development period delayed to 23.33 and 26.33 days with 75 mg/L BTH and 400 mg/L BABA treatments respectively. The results in this study; therefore, suggest that benzothiadiazole (BTH) and ß-aminobutyric acid (BABA) play a role in inducing resistance in tea plants to tea geometrid, with the optimal effect achieved at BTH-3 (75 mg/L) and BABA-3 (400 mg/L), respectively.

Functional Characteristics, Electrophysiological and Antennal Immunolocalization of General Odorant-Binding Protein 2 in Tea Geometrid, Ectropis obliqua.

As one of the main lepidopteran pests in Chinese tea plantations, Ectropisobliqua Warren (tea geometrids) can severely decrease yields of tea products. The olfactory system of the adult tea geometrid plays a significant role in seeking behaviors, influencing their search for food, mating partners, and even spawning grounds. In this study, a general odorant-binding protein (OBP) gene, EoblGOBP2, was identified in the antennae of E. obliqua using reverse transcription quantification PCR (RT-qPCR). Results showed that EoblGOBP2 was more highly expressed in the antennae of males than in females relative to other tissues. The recombinant EoblGOBP2 protein was prepared in Escherichia coli and then purified through affinity chromatography. Ligand-binding assays showed that EoblGOBP2 had a strong binding affinity for some carbonyl-containing tea leaf volatiles (e.g., (E)-2-hexenal, methyl salicylate, and acetophenone). Electrophysiological tests confirmed that the male moths were more sensitive to these candidate tea plant volatiles than the female moths. Immunolocalization results indicated that EoblGOBP2 was regionally confined to the sensilla trichoid type-II in the male antennae. These results indicate that EoblGOBP2 may be primarily involved in the olfactory activity of male E. obliqua moths, influencing their ability to sense tea leaf volatiles. This study provides a new perspective of insect GOBPs and implies that olfactory function can be used to prevent and control the tea geometrid.

Gene Identification of Pheromone Gland Genes Involved in Type II Sex Pheromone Biosynthesis and Transportation in Female Tea Pest Ectropis grisescens.

Moths can biosynthesize sex pheromones in the female sex pheromone glands (PGs) and can distinguish species-specific sex pheromones using their antennae. However, the biosynthesis and transportation mechanism for Type II sex pheromone components has rarely been documented in moths. In this study, we constructed a massive PG transcriptome database (14.72 Gb) from a moth species, Ectropis grisescens, which uses type II sex pheromones and is a major tea pest in China. We further identified putative sex pheromone biosynthesis and transportation-related unigenes: 111 cytochrome P450 monooxygenases (CYPs), 25 odorant-binding proteins (OBPs), and 20 chemosensory proteins (CSPs). Tissue expression and phylogenetic tree analyses showed that one CYP (EgriCYP341-fragment3), one OBP (EgriOBP4), and one CSP (EgriCSP10) gene displayed an enriched expression in the PGs, and that EgriOBP2, 3, and 25 are clustered in the moth pheromone-binding protein clade. We considered these our candidate genes. Our results yielded large-scale PG sequence information for further functional studies.

Genome-wide identification of microRNAs responsive to Ectropis oblique feeding in tea plant (Camellia sinensis L.).

The tea plant (Camellia sinensis L.) is vulnerable to the geometrid Ectropis oblique; although microRNAs (miRNAs) are important for plant growth, development and stress response, the function of miRNAs in the response of C. sinensis to stress from E. oblique is unclear. To identify E. oblique stress-responsive miRNAs and their target genes in tea plant, three small RNA libraries were constructed from leaves subjected to mechanical wounding (MW), geometrid attack (GA) and from healthy control (CK) leaves. Using high-throughput sequencing, 130 known miRNAs and 512 novel miRNAs were identified; of these, differential expression under GA stress was observed for 36 known and 139 novel miRNAs. Furthermore, 169 GA-responsive and 173 MW-responsive miRNAs were detected by miRNA microarray. The expression patterns of six GA-responsive miRNAs were validated by qRT-PCR. Several target genes for these miRNAs encode various transcription factors, including ethylene-responsive transcription factors and squamosa promoter-binding-like proteins, which suggests that these miRNAs may regulate stress-responsive transcriptional processes in tea plant. The present findings provide novel insights into miRNA-mediated regulatory mechanisms underlying the response to GA stress, and also offer valuable information for development of pest resistance using RNA interference-based strategies in tea plants.

Molecular identification of spiders preying on Empoasca vitis in a tea plantation.

Biological control using predators of key pest species is an attractive option in integrated pest management (IPM). Molecular gut analysis can provide an estimation of predator efficiency on a given prey. Here we use a combination of various experimental approaches, both in field and lab, to identify a potential biological control species of the common pest of commercially grown tea, Empoasca vitis (Göthe) (Hemiptera), in a Chinese plantation. We collected 2655 spiders from plantations and established relative abundances of spider species and their temporal overlap with the pest species in tea canopy. We analyzed DNA from 1363 individuals of the most common spider species using targeted RQ-PCR to quantify the potential efficiency of spiders as a predator on E. vitis. The results showed that, in the field, the jumping spider Evarcha albaria was the most abundant, had the closest temporal overlap with the pest, and frequently fed on it. Therefore, this spider may play a key role in pest suppression. The present study demonstrates the potential of our experimental approach to study predator-prey relationships in taxa that do not lend themselves to morphological identification of gut contents, such as spiders.

Mechanism of Fenpropathrin Resistance in Red Spider Mite, Oligonychus coffeae (Acarina: Tetranychidae), Infesting Tea [Camellia sinensis L. (O. Kuntze)].

Red spider mite (RSM), Oligonychus coffeae (Nietner) (Acarina: Tetranychidae), has gained special attention in view of their widespread occurrence as a pest on tea [Camellia sinensis L. (O. Kuntze)]. The development of acaricide (fenpropathrin) resistance has been screened in field populations (FPs) of RSMs from different tea-growing regions of south India and compared with a laboratory-susceptible population (SP) based on toxicity bioassay, detoxifying enzyme activities, analysis of acetylcholine esterase gene (AChE, 2064 bp), and their expression pattern using semiquantitative RT-PCR. The increased resistance ratio (RR, 1.39 to 2.13) in LC50 of fenpropathrin observed in field populations of RSM provides a baseline for screening the development of resistance to fenpropathrin. This resistance developed due to hyperexpression of detoxifying enzymes, i.e., esterase (RR of 1.43 to 2.53) and glutathione S-transferase (RR of 1.11 to 1.86), and overexpression of AChE gene at 1.4 to 2.7-fold. These results necessitate molecular studies and warrant the continuous monitoring of acaricide susceptibility and resistance pattern in order to analyze the usefulness of AChE gene as target for developing alternate pest control strategies and management of pesticide resistance in tea ecosystem.

Midgut-based resistance to oral infection by a nucleopolyhedrovirus in the laboratory-selected strain of the smaller tea tortrix, Adoxophyes honmai (Lepidoptera: Tortricidae).

A strain of Adoxophyes honmai resistant to Adoxophyes honmai nucleopolyhedrovirus (AdhoNPV) was established from a field-collected colony by repeated selection. Fifth-instar larvae of this resistant strain (R-strain) had over 66 666-fold greater resistance in terms of 50 % lethal concentration values to oral infection of AdhoNPV than non-selected strain larvae (susceptible for AdhoNPV; S2-strain). In this study, the mechanism of resistance to AdhoNPV was determined in R-strain larvae. An assessment of viral genome replication in AdhoNPV-infected S2- and R-strain larvae by quantitative PCR showed no viral genome replication occurring in R-strain larvae. Transcription of AdhoNPV ie-1, vp39 and polyhedrin genes was also not detected in R-strain midgut cells. Besides, a fluorescent brightener had no effect on AdhoNPV infection in either S2- or R-strain. However, binding and fusion of occlusion-derived virus with R-strain were significantly lower than those of S2-strain. These findings suggest that R-strain Adoxophyeshonmai larvae possess a midgut-based resistance to oral infection by AdhoNPV in which midgut epithelial cells are infected less efficiently.