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The specificity of the double-stranded RNA (dsRNA) used in the RNA interference (RNAi) technique is crucial for the success of sequence-specific gene silencing. Currently, RNAi-mediated insect control is a trending research topic. However, the off-target effects of the dsRNA in RNAi are a major concern. In this study, the dsHvβ´COPI (coat protein complex I, β´ subunit)-treated and untreated transcriptomes of the 28-spotted potato lady beetle (Henosepilachna vigintioctopunctata) were compared to understand its off-target gene silencing effects. The RNA-seq results revealed that 63 and 44 differentially expressed genes (DEGs) were upregulated and downregulated, respectively, in the dsHvβ´COPI treated group as compared with the control. Validation of the differential expressions of some selected DEGs via reverse transcription-quantitative PCR (RT-qPCR) analysis confirmed the reliability of the transcriptome analysis results. Further downstream analysis revealed that there were no genes homologous with Hvβ´COPI in H. vigintioctopunctata. Additionally, no genes with a >11 bp continuous match with dsHvβ´COPI were found in the H. vigintioctopunctata transcriptome. Six genes (Hvcitron, Hvhelicase, Hvtransposase, Hvserine, Hvdynein, and HvE3 ubiquitin) were selected to examine the off-target activity of dsHvβ´COPI based on their potential involvement in various H. vigintioctopunctata metabolic pathways. The severity of silencing these six off-target genes was evaluated by employing RNAi. The RNAi results confirmed the downregulation of the expression of all six genes, although there was no significant lethality. The findings of this study will be helpful in the risk analysis of future RNAi-mediated pest control experiments.

Abstract Invasive genotypes may be associated with their ability to access the invasion habitat. The whitefly, Bemisia tabaci Q, has been an important agricultural pest in China since 2008. In order to identify the invasion routes and to provide insight into its invasion success in China, we analyzed the composition, distribution, and genetic diversity of mitochondrial haplotypes of B. tabaci Q. Samples were obtained from 23 provincial level administrative units in 2011, and analyses conducted based on the mtCOI. Our results revealed five haplotypes (abbreviated as Q1H1-Q1H5) were present in the Q1 subclade based on 773-bp mtCOI fragment analysis. The diversity of haplotypes indicated the B. tabaci Q populations were derived from multiple invasion sources originating from the western Mediterranean region. Among the haplotypes, Q1H1 was dominant, followed by Q1H2. The whitefly populations were generally characterized by low levels of genetic diversity based on the 773-bp mtCOI fragment. Similar results were obtained when the 657-bp fragment was analyzed using the procedure in a previous report. Potential mechanisms contributing to the dominance of the Q1H1 in China are also discussed. These results will be helpful in revealing the mechanisms that enabled the successful invasion of B. tabaci Q into the country.

Tamarixia radiata (Waterston) is a predominant parasitoid of the Asian citrus psyllid (ACP), a destructive citrus pest and vector of huanglongbing (HLB) disease in the fields of southern China. To explore the functioning of target genes in T. radiata, the screening of specific reference genes is critical for carrying out the reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) under different experimental conditions. However, no reference gene(s) for T. radiata has yet been reported. Here, we selected seven housekeeping genes of T. radiata and evaluated their stability under the six conditions (developmental stage, sex, tissue, population, temperature, diet) by using RefFinder software, which contains four different programs (geNorm, ΔCt, BestKeeper, and NormFinder). Pairwise variation was analyzed by geNorm software to determine the optimal number of reference genes during the RT-qPCR analysis. The results reveal better reference genes for differing research foci: 18S and EF1A for the developmental stage; PRS18 and EF1A for sex, PRS18 and RPL13 for different tissues (head, thorax, abdomen); EF1A and ArgK between two populations; β-tubulin and EF1A for different temperatures (5, 15, 25, 35 °C); and ArgK and PRS18 for different feeding diets. Furthermore, when the two optimal and two most inappropriate reference genes were chosen in different temperatures and tissue treatments, respectively, the corresponding expression patterns of HSP70 (as the reporter gene) differed substantially. Our study provides, for the first time, a more comprehensive list of optimal reference genes from T. radiata for use in RT-qPCR analysis, which should prove beneficial for subsequent functional investigations of target gene(s) in this natural enemy of ACP.

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.

Even though introductions of exotic species provide ready-made experiments of rapid evolution, few studies have examined the genetic structure of an exotic species shortly after its initial introduction and subsequent spread. To determine the genetic structure of its populations during the initial introduction, we investigated the invasive sweet potato whitefly (Bemisia tabaci Q, commonly known as B. tabaci biotype Q) in China, which was introduced in approximately 2003. A total of 619 B. tabaci Q individuals in 20 provinces throughout China were collected and analyzed using five microsatellite loci.The introduced populations of B. tabaci Q in China represent eight genetic clusters with different geographic distributions. The populations in Yunnan Province, where B. tabaci Q was first detected, are genetically different from the other populations in China.The introduced populations of B. tabaci Q in China have high spatial genetic heterogeneity. Additional research is required to determine whether the heterogeneity results from multiple introductions, rapid evolution following one or few introductions, or some combination of multiple introductions and rapid evolution. The heterogeneity, however, is inconsistent with a single introduction at Yunnan Province, where B. tabaci Q was first detected, followed by spread.

BACKGROUND: Bemisia tabaci (Gennadius) is a phloem-feeding insect poised to become one of the major insect pests in open field and greenhouse production systems throughout the world. The high level of resistance to insecticides is a main factor that hinders continued use of insecticides for suppression of B. tabaci. Despite its prevalence, little is known about B. tabaci at the genome level. To fill this gap, an invasive B. tabaci B biotype was subjected to pyrosequencing-based transcriptome analysis to identify genes and gene networks putatively involved in various physiological and toxicological processes. METHODOLOGY AND PRINCIPAL FINDINGS: Using Roche 454 pyrosequencing, 857,205 reads containing approximately 340 megabases were obtained from the B. tabaci transcriptome. De novo assembly generated 178,669 unigenes including 30,980 from insects, 17,881 from bacteria, and 129,808 from the nohit. A total of 50,835 (28.45%) unigenes showed similarity to the non-redundant database in GenBank with a cut-off E-value of 10-5. Among them, 40,611 unigenes were assigned to one or more GO terms and 6,917 unigenes were assigned to 288 known pathways. De novo metatranscriptome analysis revealed highly diverse bacterial symbionts in B. tabaci, and demonstrated the host-symbiont cooperation in amino acid production. In-depth transcriptome analysis indentified putative molecular markers, and genes potentially involved in insecticide resistance and nutrient digestion. The utility of this transcriptome was validated by a thiamethoxam resistance study, in which annotated cytochrome P450 genes were significantly overexpressed in the resistant B. tabaci in comparison to its susceptible counterparts. CONCLUSIONS: This transcriptome/metatranscriptome analysis sheds light on the molecular understanding of symbiosis and insecticide resistance in an agriculturally important phloem-feeding insect pest, and lays the foundation for future functional genomics research of the B. tabaci complex. Moreover, current pyrosequencing effort greatly enriched the existing whitefly EST database, and makes RNAseq a viable option for future genomic analysis.

The impact of symbionts on their hosts depends on their infection density. In the current study, we investigated the effects of host plant and insecticide resistance on the relative amount of symbionts PorHera, Hamiltonella, Rickettsia, and Cardiniwm in the whitefly Bemisia tabaci (Gennadius) B biotype. The relative amount of symbionts in three host plant-adapted subpopulations (cucumber, Cucumis sativus L.; cabbage, Brassica oleracea L.; and cotton, Gossypium herbaceum L.) with the same genetic background and insecticide (thiamethoxam)-resistant and -susceptible sub-populations with the same genetic background were measured by quantitative polymerase chain reaction. The results showed that the cucumber population harbored more Portiera than the cabbage and cotton populations, the cabbage population harbored more Hamiltonella than the cucumber population, Hamiltonella amount did not statistically differ between the cotton and cucumber or the cotton and cabbage populations, and the cabbage population harbored more Rickettsia and Cardinium than the cucumber and cotton populations. In addition, the thiamethoxam-susceptible population harbored more Portiera and Hamiltonella than the thiamethoxam-resistant population, whereas the thiamethoxam-resistant population harbored more Rickettsia than the thiamethoxam-susceptible population. These results indicated that relative amounts of symbionts were affected significantly by host plant-adaption and insecticide resistance, and the response to host plant and insecticide differed among the symbionts. [ABSTRACT FROM AUTHOR]

The article focuses on a study which examined the gene expression in thiamethoxam resistant and susceptible B-biotype sweetpotato whitefly Bemisia tabaci. The study utilized suppression subtractive hybridization (SSH) library approach, and included 72 and 52 up- and down-regulated genes. It found that expressed sequence tags (ESTs) belong to various functional categories based on their gene ontology annotation.