RNAi-based silencing of proteasome 20S subunit alpha 2 affected the survival and development of Henosepilachna vigintioctopunctata.

Henosepilachna vigintioctopunctata is a notorious pest of solanaceous plants in Asia, which is mainly managed by chemical pesticides. RNA interference (RNAi) technique is considered to be a promising and effective alternative for pest control. In this study, we selected the proteasome 20S subunit alpha 2 (Prosα2) gene, a cellular protein involved in many proteins regulatory processes, to explore the RNAi efficiency in H. vigintioctopunctata. The obtained results confirmed the significant lethal effects of HvProsα2 silencing on the H. vigintioctopunctata 1st instar larvae at concentrations of 100, 50, and 5 ng/µL. Ingestion of the bacterially expressed dsHvProsα2 caused high mortality in both larvae and adults. Moreover, silencing of HvProsα2 resulted in feeding disorders, growth delay, and abnormal intestinal development of the larvae. Overall, HvProsα2 acts as an important regulator for the growth and development of H. vigintioctopunctata, and can serve as a candidate target gene for the RNAi-based control of H. vigintioctopunctata.

RNAi assays in the striped flea beetle (Phyllotreta striolata) suggest Psγ-COPI and PsArf1COPI as potential molecular targets for pest control.

Phyllotreta striolata (Fabricius), commonly known as the striped flea beetle (SFB), is a notorious insect pest that attacks Brassicaceae plants worldwide, leading to tremendous economic losses. RNA interference (RNAi) has been proposed as a promising strategy for sustainable and eco-friendly pest control. In this study, a total of nine housekeeping genes including PsVATPA, PsHSP90, PsEF1A, PsRPL6, PsRPS24, PsActin, PsTUBA, PsRPS18, and PsRPL4 were evaluated under four different conditions (organization, population, sex, and RNAi). PsEF1A and PsVATPA were identified as the best reference genes for RNAi bioassay. Furthermore, a total of 24 target genes were selected to investigate their RNAi effects in SFB adults with double-stranded RNAs (dsRNAs), five of them showed significant mortality (28.00% to 70.00%), namely Psα-COPI, Psß-COPI, PsRPS18, Psγ-COPI, and PsArf1COPI. We found that gene transcript levels of the two most lethal genes, Psγ-COPI and PsArf1COPI, were significantly decreased after treated with the target dsRNAs either by feeding or injection method. The findings from this study demonstrated that the introduction of dsRNAs via oral feedings or injection induces the RNAi-mediated silencing of target genes and can lead to insect mortality. Overall, the identified target genes can be explored in developing RNAi-based insecticides for SFB control.

RNAi suppression of the nuclear receptor FTZ-F1 impaired ecdysis, pupation, and reproduction in the 28-spotted potato ladybeetle, Henosepilachna vigintioctopunctata.

Fushi-tarazu factor 1 (FTZF1) is an ecdysone-inducible transcription factor that plays a vital role during the metamorphosis in insects. In this study, we functionally characterized HvFTZ-F1 in H. vigintioctopunctata, a dreadful solanaceous crop pest, by using a dietary RNA interference technique. The HvFTZ-F1 expression levels were elevated in the 1st and 2nd-instars before molting and declined immediately after ecdysis. The HvFTZ-F1 silencing led to high mortality in the 1st instars, while the expression of the osmosis-regulative gene, HvAQPAn.G, was significantly increased in the 1st instars. HvFTZ-F1 silencing downregulated the Halloween and 20E-related genes, decreased the ecdysteroids titer, suppressed the expression of pigmentation-related genes, and reduced the catecholamines titer. In the 4th instars, HvFTZ-F1 silencing caused 100% mortality by arresting the development at the prepupal stage and preventing new abdominal cuticle formation. In the female adults, HvFTZ-F1 silencing caused an evident decrease in fecundity, prolonged the pre-oviposition period, reduced the number of eggs and hatching rate, severely atrophied the ovaries. Moreover, the 20E-related genes and the dopamine synthesis genes were suppressed in the dsHvFTZ-F1-treated females. Overall, our results revealed that HvFTZ-F1 regulates ecdysis, pupation, and reproduction in H. vigintioctopunctata, thereby could be a promising molecular target for the development of RNAi-based biopesticides to control H. vigintioctopunctata.

Toxicity of fluralaner against vegetable pests and its sublethal impact on a biocontrol predatory ladybeetle.

Fluralaner, a systemic pesticide, was originally registered with the US Food and Drug Administration in 2014 under the trade name Bravecto for flea treatment for pets. As a GABA antagonist, the footprint of fluralaner has expended beyond medical and veterinary pests in recent years. In this study, we examined the acute toxicity of fluralaner against three pests of Henosepilachna vigintioctopunctata, Megalurothrips usitatus, and Phyllotreta striolata in the Solanaceae, Fabaceae, and Cruciferae families, respectively, and the sublethal impact of fluralaner on Propylaea japonica, a widely distributed predatory ladybeetle. Based on LC50, fluralaner was effective against H. vigintioctopunctata (0.098 mg a.i. L-1 for the second instar larvae), M. usitatus (0.134 mg a.i. L-1 for adult females), and P. striolata (0.595 mg a.i. L-1 for adults). For P. japonica, however, fluralaner was substantially less effective (1.177 mg a.i. L-1 for the third instar larvae). Furthermore, the LC10 and LC30 of P. japonica were also consistently higher than the LC50 of the three pests. In addition, we did not observe any significant impacts of fluralaner at LC10 and LC30 on the life history traits, including body weight, developmental time, pre-oviposition period, and fecundity of P. japonica. Based on our results from acute toxicities and sublethal impacts, fluralaner is effective against vegetable pests, while potentially friendly to P. japonica when employed as a biological control agent.

Double-stranded RNA targeting vATPase B reveals a potential target for pest management of Henosepilachna vigintioctopunctata.

The development of genetic based techniques, specifically RNA interference (RNAi), has emerged as a powerful tool in novel pest management strategies for pestiferous coleoptera. The 28-spotted ladybird beetle, Henosepilachna vigintioctopunctata, is a dynamic foliar pest of solenaceous plants, primarily potato plants, and has quickly become one of the most important pests attacking many crops in Asian countries. In this study, we demonstrate the efficacy of dietary RNAi targeting vATPase B, which led to significant gene silencing. Downstream effects of vATPase B silencing appeared to be both time- and partial dose-dependent. Our results indicate that silencing of vATPase B caused a significant decrease in survival rate, as well as reduced the food stuffs consumption and inhibited the overall development of H. vigintioctopunctata. Furthermore, results demonstrate expression of insect melanism related genes, TH and DDC, was significantly up regulated under the dsvATPase B (RNAi molecule designed against vATPase B) treatment. The impact of oral dsvATPase B delivery on the survival of 1st, 3rd instars, and adults was investigated through bacterially expressed dsRNA. The effectiveness of RNAi-based gene silencing in H. vigintioctopunctata provides a powerful reverse genetic tool for the functional annotation of its genes. This study demonstrates that vATPase B may represent a candidate gene for RNAi-based control of H. vigintioctopunctata.

De Novo Assembly of the Asian Citrus Psyllid Diaphorina citri (Hemiptera: Psyllidae) Transcriptome across Developmental Stages.

Asian citrus psyllid Diaphorina citri Kuwayama is an important economic pest of citrus, as it transmits Candidatus Liberibacter asiaticus, the causative agent of huanglongbing. In this study, we used RNA-seq to identify novel genes and provide the first high-resolution view of the of D. citri transcriptome throughout development. The transcriptomes of D. citri during eight developmental stages, including the egg, five instars, and male and female adults were sequenced. In total, 115 million clean reads were obtained and assembled into 354,726 unigenes with an average length of 925.65 bp and an N50 length of 1733 bp. Clusters of Orthologous Groups, Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes analyses were conducted to functionally annotate the genes. Differential expression analysis highlighted developmental stage-specific expression patterns. Furthermore, two trehalase genes were characterized with lower expression in adults compared to that in the other stages. The RNA interference (RNAi)-mediated suppression of the two trehalase genes resulted in significantly high D. citri mortality. This study enriched the genomic information regarding D. citri. Importantly, these data represent the most comprehensive transcriptomic resource currently available for D. citri and will facilitate functional genomics studies of this notorious pest.

Variation in both host defense and prior herbivory can alter plant-vector-virus interactions.

BACKGROUND: While virus-vector-host interactions have been a major focus of both basic and applied ecological research, little is known about how different levels of plant defense interact with prior herbivory to affect these relationships. We used genetically-modified strains of tomato (Solanum lycopersicum) varying in the jasmonic acid (JA) plant defense pathways to explore how plant defense and prior herbivory affects a plant virus (tomato yellow leaf curl virus, 'TYLCV'), its vector (the whitefly Bemisia tabaci MED), and the host. RESULTS: Virus-free MED preferred low-JA over high-JA plants and had lower fitness on high-JA plants. Viruliferous MED preferred low-JA plants but their survival was unaffected by JA levels. While virus-free MED did not lower plant JA levels, viruliferous MED decreased both JA levels and the expression of JA-related genes. Infestation by viruliferous MED reduced plant JA levels. In preference tests, neither virus-free nor viruliferous MED discriminated among JA-varying plants previously exposed to virus-free MED. However, both virus-free and viruliferous MED preferred low-JA plant genotypes when choosing between plants that had both been previously exposed to viruliferous MED. The enhanced preference for low-JA genotypes appears linked to the volatile compound neophytadiene, which was found only in whitefly-infested plants and at concentrations inversely related to plant JA levels. CONCLUSIONS: Our findings illustrate how plant defense can interact with prior herbivory to affect both a plant virus and its whitefly vector, and confirm the induction of neophytadiene by MED. The apparent attraction of MED to neophytadiene may prove useful in pest detection and management.

De Novo Transcriptome Analysis Reveals Abundant Gonad-specific Genes in the Ovary and Testis of Henosepilachna vigintioctopunctata.

Henosepilachna vigintioctopunctata (Coleoptera: Coccinellidae) is a major pest affecting Solanaceae plants in Asian countries. In this study, we sequenced the ovary and testis transcriptomes of H. vigintioctopunctata to identify gonad-related genes. Comparison of the unigene sequences in ovary and testis libraries identified 1,421 and 5,315 ovary- and testis-specific genes, respectively. Among the ovary-specific genes, we selected the RC2-like and PSHS-like genes to investigate the effects of gene silencing on the mortality, percentage infertility, pre-oviposition period, fecundity, daily number of eggs laid, and hatching rate in female adults. Although the percentage mortality and infertility of females did not differ significantly among dsRNA treatments, fecundity was significantly reduced in the dsRC2-like and dsPSHS-like treatment groups. Moreover, the pre-oviposition period was markedly prolonged in response to dsPSHS-like treatment. This is the first reported RNA sequencing of H. vigintioctopunctata. The transcriptome sequences and gene expression profiles of the ovary and testis libraries will provide useful information for the identification of gonad-related genes in H. vigintioctopunctata and facilitate further research on the reproductive biology of this species. Moreover, the gonad-specific genes identified may represent candidate target genes for inhibiting the population growth of H. vigintioctopunctata.

Genome-Wide Analysis of Carboxylesterases (COEs) in the Whitefly, Bemisia tabaci (Gennadius).

The whitefly (Bemisia tabaci), an important invasive pest that causes severe damage to crops worldwide, has developed resistance to a variety of insecticides. Carboxylesterases (COEs) are important multifunctional enzymes involved in the growth, development, and xenobiotic metabolism of insects. However, systematic studies on the COEs of B. tabaci are scarce. Here, 42 putative COEs in different functional categories were identified in the Mediterranean species of B. tabaci (B. tabaci MED) based on a genome database and neighbor-joining phylogeny. The expression patterns of the COEs were affected by the development of B. tabaci. The expression levels of six COEs were positively correlated with the concentration of imidacloprid to which B. tabaci adults were exposed. The mortality of B. tabaci MED adults fed dsBTbe5 (67.5%) and dsBTjhe2 (58.4%) was significantly higher than the adults fed dsEGFP (41.1%) when treated with imidacloprid. Our results provide a basis for functional research on COEs in B. tabaci and provide new insight into the imidacloprid resistance of B. tabaci.

Insecticides promote viral outbreaks by altering herbivore competition.

While the management of biological invasions is often characterized by a series of single-specieg decisions, invasive species exist within larger food webs. These biotic interactions can alter the impact of control/eradication programs and may cause suppression efforts to inadvertently facilitate invasion spread and impact. We document the rapid replacement of the invasive Bemisia Middle East-Asia Minor I (MEAM1) cryptic biotype by the cryptic Mediterranean (MED) biotype throughout China and demonstrate that MED is more tolerant of insecticides and a better vector of tomato yellow leaf curl virus (TYLCV) than MEAMJ. While MEAM1 usually excludes MED under natural conditions, insecticide application reverses the MEAM1-MED competitive hierarchy and allows MED to exclude MEAMI. The insecticide-mediated success of MED has led to TYLCV outbreaks throughout China. Our work strongly supports the hypothesis that insecticide use in China reverses the MEAMl-MED competitive hierarchy and allows MED to displace MEAM1 in managed landscapes. By promoting the dominance of a Bemisia species that is a competent viral vector, insecticides thus increase the spread and impact of TYLCV in heterogeneous agroecosystems.

Multiple forms of vector manipulation by a plant-infecting virus: Bemisia tabaci and tomato yellow leaf curl virus.

For many insect-vectored plant viruses, the relationship between feeding behavior and vector competence may prove integral to an understanding of the epidemiology of the resulting plant disease. While plant-infecting viruses are well known to change host plant physiology in a way that makes them more attractive to vectors, viral manipulation of the vectors themselves has only recently been reported. Previous research suggested that the rapid spread of Tomato yellow leaf curl virus (TYLCV) throughout China has been facilitated by its primary vector, the whitefly Bemisia tabaci. We conducted two experiments testing the impact of TYLCV infection of the host plant (tomato) and vector (B. tabaci biotypes B and Q) on whitefly feeding behavior. Whiteflies of biotypes B and Q both appeared to find TYLCV-infected plants more attractive, probing them more quickly and having a greater number of feeding bouts; this did not, however, alter the total time spent feeding. Viruliferous whiteflies fed more readily than uninfected whiteflies and spent more time salivating into sieve tube elements. Because vector salivation is essential for viral transmission, this virally mediated alteration of behavior should provide TYLCV a direct fitness benefit. This is the first report of such manipulation by a nonpropagative virus that belongs to an exclusively plant-infecting family of viruses (Geminiviridae). In the context of previous research showing that feeding on TYLCV-infected plants harms biotype B but helps biotype Q, the fact that both biotypes were equally affected by TYLCV also suggests that the virus may alter the biotype B-biotype Q competitive interaction in favor of biotype Q.

Three-way interactions between the tomato plant, tomato yellow leaf curl virus, and Bemisia tabaci (Hemiptera: Aleyrodidae) facilitate virus spread.

Plant defense responses can greatly affect plant viruses and their herbivore vectors. The current article reports on plant defense responses involving jasmonic acid (JA), salicylic acid (SA), and proteinase inhibitor (PI) in the three-way interaction between tomato plants, tomato yellow leaf curl virus, and the whitefly Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae). The results showed that feeding by viruliferous B. tabaci increases the longevity and fecundity of nonviruliferous B. tabaci that subsequently feed on the same plant. Feeding by nonviruliferous B. tabaci alone suppressed plant defense responses involving JA and PI but induced responses involving SA. Feeding by viruliferous B. tabaci increased the suppression of plant defenses involving JA and PI but did not increase responses involving SA. These results indicate that the interactive effects of tomato yellow leaf curl virus and B. tabaci on plants increase vector fitness and virus transmission by reducing plant defense.

Sensitivity of Bemisia tabaci (Hemiptera: Aleyrodidae) to several new insecticides in China: effects of insecticide type and whitefly species, strain, and stage.

Whitefly biotypes B and Q are the two most damaging members of the Bemisia tabaci (Hemiptera: Aleyrodidae) species complex. Control of B. tabaci (and especially of Q) has been impaired by resistance to commonly used insecticides. To find new insecticides for B. tabaci management in China, we investigated the sensitivity of eggs, larvae, and adults of laboratory strains of B and Q (named Lab-B and Lab-Q) and field strains of Q to several insecticides. For eggs, larvae, and adults of B. tabaci and for six insecticides (cyantraniliprole, chlorantraniliprole, pyriproxyfen, buprofezin, acetamiprid, and thiamethoxam), LC50 values were higher for Lab-Q than for Lab-B; avermectin LC50 values, however, were low for adults of both Lab-Q and Lab-B. Based on the laboratory results, insecticides were selected to test against eggs, larvae, and adults of four field strains of B. tabaci Q. Although the field strains differed in their sensitivity to the insecticides, the eggs and larvae of all strains were highly sensitive to cyantraniliprole, and the adults of all strains were highly sensitive to avermectin. The eggs, larvae, and adults of B. tabaci Q were generally more resistant than those of B. tabaci B to the tested insecticides. B. tabaci Q eggs and larvae were sensitive to cyantraniliprole and pyriproxyfen, whereas B. tabaci Q adults were sensitive to avermectin. Field trials should be conducted with cyantraniliprole, pyriproxyfen, and avermectin for control of B. tabaci Q and B in China.

Population structure of the greenhouse whitefly, Trialeurodes vaporariorum (Westwood), an invasive species from the Americas, 60 years after invading China.

Though the greenhouse whitefly, Trialeurodes vaporariorum (Westwood) (Hemiptera: Aleyrodidae) was introduced into China more than 60 years ago, the genetic diversity and structure of this exotic insect pest and virus vector have not been studied. To investigate the population genetic characteristics of this invasive species and to identify potential invasion routes, the genetic diversity and population structure of 17 collections of T. vaporariorum from nine provinces in China were analyzed using seven microsatellite loci. The results of the analyses indicated that the genetic diversity for the populations examined from the four provinces: Jilin, Ningxia, Guizhou and Qinghai, was lower than the genetic diversity of populations from the five provinces: Yunnan, Shandong, Shanxi, Liaoning, and Gansu. The T. vaporariorum populations analyzed in this study grouped as two distinct genetic clusters based on the analysis using STRUCTURE, whereas, 8 clusters were identified based on the BAPS analysis. Of the 136 genetic distance (Fst) values, 128 (94%) were associated with a significant exact test. However, there was no significant relationship between Fst and geographical distance. These results demonstrate that populations of T. vaporariorum in China exhibit significant genetic differentiation, indicating the likelihood that multiple introductions of T. vaporariorum into China have occurred. Also, the populations collected from the provinces of Jilin, Ningxia, Guizhou and Qinghai appear to represent secondary introductions originating from other Chinese provinces.

Two cytochrome P450 genes are involved in imidacloprid resistance in field populations of the whitefly, Bemisia tabaci, in China.

The sweet potato whitefly, Bemisia tabaci (Gennadius) (Hemiptera:Aleyrodidae), is an invasive and damaging pest of field crops worldwide. The neonicotinoid insecticide imidacloprid has been widely used to control this pest. We assessed the species composition (B vs. Q), imidacloprid resistance, and association between imidacloprid resistance and the expression of five P450 genes for 14-17 B. tabaci populations in 12 provinces in China. Fifteen of 17 populations contained only B. tabaci Q, and two populations contained both B and Q. Seven of 17 populations exhibited moderate to high resistance to imidacloprid, and eight populations exhibited low resistance to imidacloprid, compared with the most susceptible field WHHB population. In a study of 14 of the populations, resistance level was correlated with the expression of the P450 genes CYP6CM1 and CYP4C64 but not with the expression of CYP6CX1, CYP6CX4, or CYP6DZ7. This study indicates that B. tabaci Q has a wider distribution in China than previously reported. Resistance to imidacloprid in field populations of B. tabaci is associated with the increased expression of two cytochrome P450 genes (CYP6CM1 and CYP4C64).

Tomato spotted wilt virus infection reduces the fitness of a nonvector herbivore on pepper.

Plant pathogens and insect herbivores often share hosts under natural conditions. Hence, pathogen-induced changes in a host plant can affect the herbivore and vice versa. Even though plant viruses are ubiquitous in the field, little is known about plant-mediated interactions between viruses and nonvector herbivores. Here we tested whether the performance of the sweet potato whitefly, Bemisia tabaci (Gennadius) biotype Q, was altered when raised on pepper infected with Tomato spotted wilt virus (TSWV). TSWV infection reduced B. tabaci fecundity and longevity and increased B. tabaci developmental time but did not affect the insect's survival or female body lengths. Our results demonstrate that TSWV infection can decrease the fitness of B. tabaci biotype Q on pepper plants.

RNAi-Based Biopesticides Against 28-Spotted Ladybeetle Henosepilachna vigintioctopunctata Does Not Harm the Insect Predator Propylea japonica.

RNA interference (RNAi)-mediated control of the notorious pest Henosepilachna vigintioctopunctata is an emerging environment friendly research area. However, the characterization of key target genes in H. vigintioctopunctata is crucial for this. Additionally, assessing the risk of RNAi to nontarget organisms (NTOs) is necessary for environmental safety. In this study, the potential of RNAi technology in controlling H. vigintioctopunctata infestation has been investigated by the oral delivery of double-stranded RNA (dsRNA). The results revealed that the silencing of six genes, including HvABCH1, HvHel25E, HvProsbeta5, HvProsalpha6, HvProsbeta6, and HvSrp54k, was highly lethal to H. vigintioctopunctata. The LC50 values of the dsRNAs used to silence these six genes were found to be less than 13 ng/µL. Moreover, the use of the bacterially expressed dsRNAs caused high mortality in the lab and field populations of H. vigintioctopunctata. Further, administration of HvHel25E and HvSrp54k dsRNAs in the predatory lady beetle Propylea japonica confirmed no transcriptional or organismal levels effects. This risk-assessment result ensured no off-target RNAi effects on the NTOs. Overall, the findings of the study suggested that HvABCH1, HvHel25E, HvProsbeta5, HvProsalpha6, HvProsbeta6, and HvSrp54k can be novel promising molecular targets with high specificity for H. vigintioctopunctata management with negligible effects on the NTOs.

Heterogeneous distribution of Cardinium in whitefly populations is associated with host nuclear genetic background.

Inherited bacterial symbionts are very common in arthropods, but infection frequency can vary widely among populations. Experiments and interpopulation comparisons suggest that host genetic background might be important in explaining this variation. Our extensive field investigation showed that the infection pattern of the facultative symbiont Cardinium was heterogeneous across geographical populations of the invasive whitefly Bemisia tabaci Mediterranean (MED) in China, with genetic nuclear differences evident in 2 of the populations: 1 with a low infection rate (SD line) and 1 with a high infection rate (HaN line). However, whether the heterogeneous frequency of Cardinium is associated with the host genetic background remains poorly understood. Here, we compared the fitness of the Cardinium-infected and uninfected sublines with similar nuclear genetic backgrounds from SD and HaN lines, respectively, and further determine whether host extranuclear or nuclear genotype influenced the Cardinium-host phenotype by performing 2 new introgression series of 6 generations between SD and HaN lines (i.e., Cardinium-infected females of SD were backcrossed with uninfected males of HaN, and vice versa). The results showed that Cardinium provides marginal fitness benefits in the SD line, whereas Cardinium provides strong fitness benefits in the HaN line. Further, both Cardinium and the Cardinium-host nuclear interaction influence the fecundity and pre-adult survival rate of B. tabaci, whereas the extranuclear genotype does not. In conclusion, our results provide evidence that Cardinium-mediated fitness effects were closely associated with the host genetic background, which provides a fundamental basis for understanding the mechanism underlying the heterogeneous distribution of Cardinium in B. tabaci MED populations across China.

Gene expression profiling in the thiamethoxam resistant and susceptible B-biotype sweetpotato whitefly, Bemisia tabaci.

Thiamethoxam has been used as a major insecticide to control the B-biotype sweetpotato whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae). Due to its excessive use, a high level of resistance to thiamethoxam has developed worldwide over the past several years. To better understand the molecular mechanisms underlying this resistance in B. tabaci, gene profiles between the thiamethoxam-resistant and thiamethoxam-susceptible strains were investigated using the suppression subtractive hybridization (SSH) library approach. A total of 72 and 52 upand down-regulated genes were obtained from the forward and reverse SSH libraries, respectively. These expressed sequence tags (ESTs) belong to several functional categories based on their gene ontology annotation. Some categories such as cell communication, response to abiotic stimulus, lipid particle, and nuclear envelope were identified only in the forward library of thiamethoxam-resistant strains. In contrast, categories such as behavior, cell proliferation, nutrient reservoir activity, sequence-specific DNA binding transcription factor activity, and signal transducer activity were identified solely in the reverse library. To study the validity of the SSH method, 16 differentially expressed genes from both forward and reverse SSH libraries were selected randomly for further analyses using quantitative realtime PCR (qRT-PCR). The qRT-PCR results were fairly consistent with the SSH results; however, only 50% of the genes showed significantly different expression profiles between the thiamethoxam-resistant and thiamethoxam-susceptible whiteflies. Among these genes, a putative NAD-dependent methanol dehydrogenase was substantially over-expressed in the thiamethoxamresistant adults compared to their susceptible counterparts. The distributed profiles show that it was highly expressed during the egg stage, and was most abundant in the abdomen of adult females.

Transcriptome analysis of megalurothrips usitatus (Bagnall) identifies olfactory genes with ligands binding characteristics of MusiOBP1 and MusiCSP1.

The olfactory system is an important component of insect behavior and is vital for survival and reproduction. However, the genomic characterization and molecular basis of the olfactory response of Megalurothrips usitatus remain relatively unknown. RNA sequencing-built developmental transcriptomes of nymphs, pupae, and adult M. usitatus were examined in order to establish the sequence-based background of M. usitatus olfactory responses. A total of 56,669 unigenes were annotated using GO, NR, Pfam, eggNOG, SwissProt, and KEGG. The number of differentially expressed genes between pupae and nymphs, males and nymphs, and females and nymphs were 10,498, 9,235, and 10,964, respectively. One odorant-binding protein (MusiOBP1) and one chemosensory protein (MusiCSP1) were selected from the transcriptome, and their full-length sequences were obtained using RACE PCR. The relative expression of MusiOBP1 was the highest in primordial females, whereas the relative expression of MusiCSP1 was the highest in primordial pupae. The strongest binding ability to the odor-binding protein MusiOBP1 was observed for ß-citronellol. 3-Hydroxy-2-methyl-4-pyrone showed the strongest binding affinity to MusiCSP1. Our analysis suggests that MusiOBP1 and MusiCSP1 may play significant roles in mediating M. usitatus host recognition. This research will improve our knowledge of odorant-binding proteins and chemosensory proteins, which will in turn improve our understanding of insect olfactory systems.