Search

The endosymbiont Wolbachia is known for manipulating host reproduction. Wolbachia also can affect host fitness by mediating interactions between plant and herbivores. However, it remains unclear whether saliva proteins are involved in this process.We found that Wolbachia infection decreased the number of deposited eggs but increased the egg hatching rate in the spider mite Tetranychus urticae Koch (Acari: Tetranychidae), a cosmopolitan pest that infects1000 species of plants. Transcriptomic and proteomic analyses revealed that Wolbachia-infected mites upregulated the gene expression levels of many T. urticae salivary proteins including a cluster of Tetranychidae-specific, functionally uncharacterized SHOT1s (secreted host-responsive proteins of Tetranychidae). The SHOT1 genes were expressed more in the feeding stages (nymphs and adults) of mites than in eggs and highly enriched in the proterosomas. RNA interference experiments showed that knockdown of SHOT1s significantly decreased Wolbachia density, increased the number of deposited eggs and decreased the egg hatching rate.Together, these results indicate that SHOT1s are positively correlated with Wolbachia density and account for Wolbachia-mediated phenotypes. Our results provide new evidence that herbivore salivary proteins are related to Wolbachia-mediated manipulations of host performance on plants. © 2022 Society of Chemical Industry.

These are supporting materrilas for the investigation of microbiota in wild spider mite populations

Ras opposite (Rop) is known to play an essential role in regulating vesicle trafficking, including synaptic transmission and general secretion. The fundamental roles of Rop have been confirmed by the observation that null mutations in many organisms generate lethal phenotypes during embryogenesis. However, the effects of Rop during the postembryonic stages, especially in non-model organisms, remain largely unknown. Here, we provide new data that enhance our understanding of Rop's roles in the adults of multiple species of Tetranychus spider mites (Acari: Tetranychidae), a class of notorious agricultural pests. Our in silico and experimental evidence demonstrated that Rop is under purifying selection and is highly conserved in Tetranychus spp. RNA interference experiments showed that Rop is required for maintaining normal fecundity but has no significant effect on survival. We further demonstrate that knockdown of Rop darkens the body color of spider mites and blocks the excretion of fecal pellets, which is likely to be related to an abnormality in the excretion of food waste in the digestive system. Overall, our findings clarify novel functions of a vesicle trafficking-related gene in the adult stage of multiple Tetranychus species and highlight the need to evaluate the roles of essential genes in various organisms.

Herbivore-associated molecular patterns (HAMPs) enable plants to recognize herbivores and may help plants adjust their defense responses. Here, we report on herbivore-induced changes in a protein disulfide isomerase (PDI) widely distributed across arthropods. PDI from the spider mite Tetranychus evansi (TePDI), a mesophyll-feeding agricultural pest worldwide, triggered immunity in multiple Solanaceae plants. TePDI-mediated cell death in Nicotiana benthamiana required the plant signaling proteins SGT1 (suppressor of the G2 allele of skp1) and HSP90 (heat shock protein 90), but was suppressed by spider mite effectors Te28 and Te84. Moreover, PDIs from phylogenetically distinct herbivorous and nonherbivorous arthropods triggered plant immunity. Finally, although PDI-induced plant defenses impaired the performance of spider mites on plants, RNAi experiments revealed that PDI genes are essential for the survival of mites and whiteflies. Our findings indicate that plants recognize evolutionarily conserved HAMPs to activate plant defense and resist pest damage, pointing to opportunities for broad-spectrum pest management.

BACKGROUND Drosophila suzukii (Matsumura, 1931) (spotted wing drosophila), an invasive species, has recently become a significant global pest of soft-skinned fruits such as berries. Unlike other Drosophila species, female D. suzukii have evolved a specialized sharp, serrated ovipositor that pierces and penetrates ripe and ripening fruits, causing them to lose commercial value and preventing their sale. A first step for the development of biological control agents for pest management may be achieved through the identification of microbes infectious for D. suzukii in the wild. RESULTS We first determined that D. suzukii is susceptible to chemicals commonly used to rear Drosophilids in the laboratory and established a diet able to sustain healthy D. suzukii growth. Using this diet, we demonstrated that of 25 species of culturable bacteria and fungi isolated from field-collected D. suzukii, eight microbes decreased host survival when injected. Three of the eight bacteria (Alcaligenes faecalis, Achromobacter spanius and Serratia marcescens) were acutely pathogenic to both D. suzukii and Drosophila melanogaster adults by injection. Feeding of these bacteria resulted in susceptibility only in larvae. CONCLUSION We successfully identified multiple microbes from field-collected D. suzukii that are pathogenic to both larvae and adults through different routes of infection, some of which could be candidates for biocontrol of this species. © 2020 Society of Chemical Industry.

The bacterial communities of arthropods play important roles in the growth and development of their hosts, such as by providing nutrition or protecting against parasites and pathogens. However, for Tetranychus truncatus, a major crop pest, very little is known about its bacterial community. To understand the effects of the bacterial community in T. truncatus, mites were fed bean leaves that had been soaked in different concentrations of tetracycline hydrochloride (5 mg to 5 g per L). Antibiotic treatment had a negative impact on the fitness of T. truncatus, decreasing its fecundity and longevity. However, we could not exclude the possibility that the decrease in fecundity was due to a toxic effect of the antibiotic. 16S rRNA genes analyses showed that the antibiotic treatment did not significantly affect the diversity of bacterial species, but it did significantly change their relative abundances. Furthermore, predicted functions of microbiota were also changed with antibiotic treatment and the most abundant genes were related to metabolism. Our research revealed that the antibiotic induced change of bacterial communities mostly decreased the fecundity of T. truncatus, although we couldn't totally exclude the direct toxin of antibiotics on spider mites.

Rice planthoppers are notorious plant sap-feeding pests which cause serious damage. While several microbes in rice planthoppers have been broadly characterized, the abundance and diversity of bacteria and fungi in field planthoppers are largely unknown. This study investigated the bacterial and fungal community compositions of Chinese wild rice planthoppers Laodelphax striatellus and Sogatella furcifera using parallel 16S rRNA gene amplicon and internal transcribed space region sequencing. The bacteria varied significantly between the species and were partitioned significantly by sex, tissues and host environments in each species. The majority of bacteria were affiliated with the genera Wolbachia, Cardinium, Rickettsia and Pantoea. The abundance of Wolbachia was negatively correlated with that of Cardinium in both planthopper species. Compared with bacteria, the abundance and diversity of fungi did not differ between sexes but both were enriched in the gut. The bacterial community as a whole showed no significant correlation with the fungal community. The majority of fungi were related to Sarocladium, Alternaria, Malassezia, Aspergillus and Curvularia. A phylogenetic analysis revealed that these fungi were closely related to botanic symbionts or pathogens. Our results provide novel insights into the bacteria and fungi of rice planthoppers.

Mosquito vectors transmit numerous pathogens, but large gaps remain in our understanding of their physiology. To facilitate future explorations of mosquito biology, with specific attention to the major vector Aedes aegypti, we have created Aegypti-Atlas (http://aegyptiatlas.buchonlab.com/), an online resource hosting RNAseq profiles of Ae. Aegypti body parts (head, thorax, abdomen, gut, Malpighian tubules, and ovaries), gut regions (crop, proventriculus, anterior and posterior midgut, and hindgut), and a time course of blood meal digestion in the gut. Using Aegypti-Atlas, we provide new insights into the regionalization of gut function, blood feeding response, and immune defenses. We find that the anterior and posterior regions of the mosquito midgut possess clearly delineated digestive specializations which are preserved in the blood-fed state. Blood feeding initiates the sequential transcriptional induction and repression/depletion of multiple cohorts of peptidases throughout blood meal digestion. With respect to defense, immune signaling components, but not recognition or effector molecules, show enrichment in ovaries. Basal expression of antimicrobial peptides is dominated by two genes, holotricin and gambicin, that are expressed in the carcass and the digestive tissues, respectively, in a near mutually exclusive manner. In the midgut, gambicin and other immune effector genes are almost exclusively expressed in the anterior regions, while the posterior midgut exhibits the hallmarks of immune tolerance. Finally, in a cross-species comparison between the midguts of Ae. aegypti and Anopheles gambiae, we observe that regional digestive and immune specializations are closely conserved, indicating that our data may yield inferences that are broadly relevant to multiple mosquito vector species. We further demonstrate that the expression of orthologous genes is highly correlated, with the exception of a ‘species signature’ comprising a small number of highly/disparately expressed genes. With this work, we show the potential of Aegypti-Atlas to unlock a more complete understanding of mosquito biology.

Mosquitoes transmit numerous pathogens, but large gaps remain in our understanding of their physiology. To facilitate explorations of mosquito biology, we have created Aegypti-Atlas (http://aegyptiatlas.buchonlab.com/), an online resource hosting RNAseq profiles of Ae. aegypti body parts (head, thorax, abdomen, gut, Malpighian tubules, ovaries), gut regions (crop, proventriculus, anterior and posterior midgut, hindgut), and a gut time course of blood meal digestion. Using Aegypti-Atlas, we provide insights into regionalization of gut function, blood feeding response, and immune defenses. We find that the anterior and posterior midgut possess digestive specializations which are preserved in the blood-fed state. Blood feeding initiates the sequential induction and repression/depletion of multiple cohorts of peptidases. With respect to defense, immune signaling components, but not recognition or effector molecules, show enrichment in ovaries. Basal expression of antimicrobial peptides is dominated by holotricin and gambicin, which are expressed in carcass and digestive tissues, respectively, in a mutually exclusive manner. In the midgut, gambicin and other effectors are almost exclusively expressed in the anterior regions, while the posterior midgut exhibits hallmarks of immune tolerance. Finally, in a cross-species comparison between Ae. aegypti and Anopheles gambiae midguts, we observe that regional digestive and immune specializations are conserved, indicating that our dataset may be broadly relevant to multiple mosquito species. We demonstrate that the expression of orthologous genes is highly correlated, with the exception of a 'species signature' comprising a few highly/disparately expressed genes. With this work, we show the potential of Aegypti-Atlas to unlock a more complete understanding of mosquito biology.

Symbiont-mediated nutritional mutualisms can contribute to the host fitness of insects, especially for those that feed exclusively on nutritionally unbalanced diets. Here, we elucidate the importance of B group vitamins in the association of endosymbiotic bacteria Wolbachia with two plant-sap feeding insects, the small brown planthopper, Laodelphax striatellus (Fallén), and the brown planthopper, Nilaparvata lugens (Stål). Infected planthoppers of both species laid more eggs than uninfected planthoppers, while the experimental transfer of Wolbachia into uninfected lines of one planthopper species rescued this fecundity deficit. The genomic analysis showed that Wolbachia strains from the two planthopper species encoded complete biosynthesis operons for biotin and riboflavin, while a metabolic analysis revealed that Wolbachia-infected planthoppers of both species had higher titers of biotin and riboflavin. Furthermore, experimental supplementation of food with a mixture of biotin and riboflavin recovered the fecundity deficit of Wolbachia-uninfected planthoppers. In addition, comparative genomic analysis suggested that the riboflavin synthesis genes are conserved among Wolbachia supergroups. Biotin operons are rare in Wolbachia, and those described share a recent ancestor that may have been horizontally transferred from Cardinium bacteria. Our research demonstrates a type of mutualism that involves a facultative interaction between Wolbachia and plant-sap feeding insects involving vitamin Bs.

The endosymbiont Wolbachia is known for manipulating host reproduction in selfish ways. However, the molecular mechanisms have not yet been investigated in embryos. Here, we found that Wolbachia had no effect on the number of deposited eggs in Tetranychus urticae Koch (Acari: Tetranychidae) but caused two types of reproductive manipulation: killing uninfected female embryos via cytoplasmic incompatibility (CI) and increasing the hatching ratio of infected female embryos. RNA sequencing analyses showed that 145 genes were differentially expressed between Wolbachia-infected (WI) and Wolbachia-uninfected (WU) embryos. Wolbachia infection down-regulated messenger RNA (mRNA) expression of glutathione S-transferase that could buffer oxidative stress. In addition, 1613 and 294 genes were identified as CI-specific up-/down-regulated genes. Compared to WU and WI embryos, embryos of CI cross strongly expressed genes involved in transcription, translation, tissue morphogenesis, DNA damage and mRNA surveillance. In contrast, most of the genes associated with energy production and metabolism were down-regulated in the CI embryos compared to the WU and WI embryos, which provides some clues as to the cause of death of CI embryos. These results identify several genes that could be candidates for explaining Wolbachia-induced CI. Our data form a basis to help elucidate the molecular consequences of CI in embryos.

Bacteria symbionts in herbivores play an important role in host biology and ecology, and are affected by environmental factors such as temperature, diet, habitat, antibiotics and so on. However, the effects of antibiotics on the microbiome of the small brown planthopper Laodelphax striatellus (SBPH) remain unclear. Here, we studied the effects of tetracycline on the diversity and composition of bacterial colonies in different tissues of SBPH using high throughput sequencing of 16S ribosomal RNA amplicons. Our results show that Proteobacteria, Firmicutes, Bacteroidetes, Actinobacteria were most abundant in SBPH, and the genera Asaia and Wolbachia were most abundant in all body parts of SBPH. Antibiotic treatment had persistent effects on the composition of the SBPH microbiome. Tetracycline depleted the population of Firmicutes, Bacteroidetes, Tenericutes and Fusobacteria, and nearly 100% eliminated Wolbachia, Bacteroides and Abiotrophia in SBPH. Together, these results suggest that antibiotic exposure affects the bacteria symbionts of different body parts in SBPH and will facilitate future studies of the bacterial symbionts of arthropod hosts.

Arthropods host a diverse range of microbes. Some symbiotic bacteria are important in influencing the performance of arthropods, especially when facing biotic and abiotic challenges. The symbiont Wolbachia is a genus of intracellular bacteria (Rickettsiaceae) found in about 40% of terrestrial arthropods. It causes a variety of reproductive alterations in arthropod hosts, such as inducing parthenogenesis, killing males, feminizing genetic males, and inducing cytoplasmic incompatibility (CI), and is well known as a reproductive manipulator in arthropods. Recent studies show that Wolbachia can also affect host fitness by mediating interactions between plant and herbivores. But it remains unclear whether saliva proteins are involved in this process. The two-spotted spider mite (Tetranychus urticae), a notorious cosmopolitan agricultural pest, harbors Wolbachia (Xie et al., 2006). In this study, we explored the effects of Wolbachia on biological performance of T. urticae (Bing et al., 2022). We showed that the Wolbachia-infected T. urticae decreased the number of deposited eggs but increased the egg hatching rate compared with Wolbachia-uninfected mites. With transcriptomic analysis data, we further demonstrated that Wolbachia infection upregulated the gene expression levels of many T. urticae salivary proteins, which include a cluster of Tetranychidae-specific, functionally uncharacterized SHOT1s (secreted host-responsive proteins of Tetranychidae). These SHOT1 genes were highly enriched in the proterosomas (Jonckheere et al., 2018) and were expressed more in the feeding stages (nymphs and adults) of mites than in eggs. RNAi experiments showed that knocking down SHOT1s could significantly decrease Wolbachia titer, increase the number of deposited eggs and decrease the egg hatching rate. Altogether, our results suggest that host salivary proteins are related to Wolbachia-mediated manipulations of host reproduction.

Herbivores and plants have been engaged in a tight co-evolutionary arms race, and saliva played important roles in this process. Some herbivore-derived small molecules or proteins called herbivore-associated molecular patterns (HAMPs) can be recognized by plants and thus trigger plant immune responses. Another type of salivary proteins, called effectors, utilize various strategies to suppress plant defense responses to establish successful feeding. The tomato red spider mite Tetranychus evansi is a worldwide pest of Solanaceous crops and causes enormous economic damage in many regions of the world. During the feeding process, T. evansi secretes saliva into plant cells through cheliceral stylets. Secreted saliva plays crucial roles in modulating plant-mite interaction, such as effectors Te28 and Te84 (Villarroel et al., 2016). We previously identified 136 salivary proteins from T. evansi by transcriptome and LC-MS/MS analyses (Huang et al., 2018). However, the role of these salivary proteins in mite-plant interaction was unknown. Here, we identified a salivary protein involved in the mite-plant interaction (Cui et al., 2022). This protein encodes a protein disulfide isomerase (TePDI) and acts as a HAMP that triggers plant defenses by inducing ROS burst, callose deposition and plant defense-related genes in Nicotiana benthamiana. TePDI can be recognized by multiple Solanaceae plants such as tomato, pepper, and eggplant. TePDI-mediated cell death in N. benthamiana is dependent on the plant signaling molecules SGT1 (suppressor of the G2 allele of skp1) and HSP90 (heat shock protein 90). To better feed on plants, T. evansi inhibited TePDI-triggered cell death and plant defense responses by secreting effectors Te28 and Te84. Further analysis revealed that PDI from phylogenetically distinct herbivorous and non-herbivorous arthropods all triggered cell death and immune response in N. benthamiana. Moreover, silencing PDI gene in spider mites and whiteflies resulted in the reduced survival rate of both pests. Altogether, our study revealed that PDI is a double-edged sword. On the one hand, it is functionally conserved in herbivores and required for their survival; on the other hand, it is recognized by Solanaceae plants and enhances plant resistance against herbivores. Our findings indicate that plants utilize evolutionarily conserved HAMPs to activate plant defense and resist pest damage, providing a potential strategy for pest management.

Gut microbes play important roles in host physiology; however, the mechanisms underlying their impact remain poorly characterized. Here, we demonstrate that microbes not only influence gut physiology but also alter its epithelial composition. The microbiota and pathogens both influence intestinal stem cell (ISC) differentiation. Intriguingly, while the microbiota promotes ISC differentiation into enterocytes (EC), pathogens stimulate entero-endocrine (EE) cell fate and long-term accumulation of EEs in the midgut epithelium. Importantly, the evolutionarily conserved Drosophila NFKB (Relish) pushes stem cell lineage specification toward ECs by directly regulating differentiation factors. Conversely, the JAK-STAT pathway promotes EE fate in response to infectious damage. We propose a model in which the balance of microbial pattern recognition pathways, such as Imd-Relish, and damage response pathways, such as JAK-STAT, influence ISC differentiation, epithelial composition, and gut physiology.

The structures of arthropod bacterial communities are complex. These microbiotas usually provide many beneficial services to their hosts, whereas occasionally they may be parasitical. To date, little is known about the bacterial communities of Tetranychus truncatus and the factors contributing to the structure of its bacterial communities are unexplored yet. Here, we used four symbiont-infected T. truncatus strains—including one Wolbachia and Spiroplasma co-infected strain, two symbiont singly-infected strains and one symbiont uninfected strain—to investigate the influence of endosymbionts on the structure of the host mites’ microbiota. Based on 16S rRNA genes sequencing analysis, we found Wolbachia and Spiroplasma were the two most abundant bacteria in T. truncatus and the presence of both symbionts could not change the diversity of bacterial communities (based on alpha-diversity indexes such as ACE, Chao1, Shannon and Simpson diversity index). Symbiont infection did alter the abundance of many other bacterial genera, such as Megamonas and Bacteroides. The structures of bacterial communities differed significantly among symbiont-infected strains. These results suggested a prominent effect of Wolbachia and Spiroplasma on bacterial communities of the host T. truncatus. These findings advance our understanding of T. truncatus microbiota and will be helpful for further study on bacterial communities of spider mites.

Wolbachia are intracellular bacteria that manipulate host reproduction by several mechanisms including cytoplasmic incompatibility (CI). However, the underlying mechanisms of Wolbachia-induced CI are not entirely clear. Here, we monitored the Wolbachia distribution in the male gonads of the small brown planthopper ( Laodelphax striatellus , SBPH) at different development stages, and investigated the influence of Wolbachia on male gonads by a quantitative proteomic analysis. A total of 276 differentially expressed proteins were identified, with the majority of them participating in metabolism, modification, and reproduction. Knocking down the expression of outer dense fiber protein (ODFP) and venom allergen 5-like (VA5L) showed decreased egg reproduction, and these two genes might be responsible for Wolbachia improved fecundity in infected L. striatellus; whereas knocking down the expression of cytosol amino-peptidase-like (CAL) significantly decreased the egg hatch rate in Wolbachia-uninfected L. striatellus, but not in the Wolbachia-infected one. Considering that the mRNA/protein level of CAL was downregulated by Wolbachia infection and dsCAL treatment closely mimicked Wolbachia-induced CI, we presumed that CAL might be one of the factors determining the CI phenotype.

BACKGROUND The whitefly Bemisia tabaci (Gennadius) is considered one of the main pests for agriculture. One important problem with the whitefly is its notorious status as a vector for plant viruses, primarily begomoviruses. We have previously identified a defensin-like antimicrobial peptide, Btdef, from the whitefly B. tabaci MEAM1. However, the function of Btdef in the immune system of the insect vector and begomovirus transmission has yet to be explored. RESULTS To explore the role of Btdef during begomovirus transmission, we firstly investigated the transcriptional response of Btdef following acquisition of Tomato yellow leaf curl China virus (TYLCCNV). The expression of Btdef was up-regulated in the viruliferous whiteflies. After RNA silencing of the Btdef gene in adult whiteflies fed with dsRNA, they were allowed to feed on TYLCCNV-infected plants and then quantified for TYLCCNV DNA titre. Unexpectedly, silencing the Btdef gene reduced both the abundance and expressions of TYLCCNV genes in the whiteflies. In the meantime, the density of the endosymbiont Rickettsia was significantly reduced in dsBtdef-fed whiteflies. CONCLUSION Our data provide evidence that Btdef is involved in begomovirus infection, possibly through symbiont-mediated alteration of begomovirus–whitefly interactions. These findings indicate that Btdef may be targeted for the development of new technology for the control of whitefly-transmitted begomoviruses. © 2016 Society of Chemical Industry

Rice planthoppers (Hemiptera: Delphacidae) are notorious pests for rice (Oryza sativa) in Asia, posing a serious threat to rice production and grain security. Rice planthoppers harbor diverse bacterial symbionts, including Wolbachia, Cardinium, Spiroplasma, and Arsenophonus, which are known to manipulate reproduction in arthropod hosts. This microreview is to introduce current knowledge of bacterial reproductive manipulators in rice planthoppers, including their diversity, population dynamics, localization, transmission, and biological functions.

Surviving infection requires immune and repair mechanisms. Developing organisms face the additional challenge of integrating these mechanisms with tightly controlled developmental processes. The larval Drosophila midgut lacks dedicated intestinal stem cells. We show that, upon infection, larvae perform limited repair using adult midgut precursors (AMPs). AMPs differentiate in response to damage to generate new enterocytes, transiently depleting their pool. Developmental delay allows for AMP reconstitution, ensuring the completion of metamorphosis. Notch signaling is required for the differentiation of AMPs into the encasing, niche-like peripheral cells (PCs), but not to differentiate PCs into enterocytes. Dpp (TGF-β) signaling is sufficient, but not necessary, to induce PC differentiation into enterocytes. Infection-induced JAK-STAT pathway is both required and sufficient for differentiation of AMPs and PCs into new enterocytes. Altogether, this work highlights the constraints imposed by development on an organism's response to infection and demonstrates the transient use of adult precursors for tissue repair.

Wolbachia is the most prevalent symbiont described in arthropods to date. Wolbachia can manipulate host reproduction, provide nutrition to insect hosts and protect insect hosts from pathogenic viruses. So far, 13 supergroups of Wolbachia have been identified. The whitefly Bemisia tabaci is a complex containing more than 28 morphologically indistinguishable cryptic species. Some cryptic species of this complex are invasive. In this study, we report a comprehensive survey of Wolbachia in B. tabaci and its relative B. afer from 1658 insects representing 54 populations across 13 provinces of China and one state of Australia. Based on the results of PCR or sequencing of the 16S rRNA gene, the overall rates of Wolbachia infection were 79.6% and 0.96% in the indigenous and invasive Bemisia whiteflies, respectively. We detected a new Wolbachia supergroup by sequencing five molecular marker genes including 16S rRNA, groEL, gltA, hcpA, and fbpA genes. Data showed that many protein-coding genes have limitations in detecting and classifying newly identified Wolbachia supergroups and thus raise a challenge to the known Wolbachia MLST standard analysis system. Besides, the other Wolbachia strains detected from whiteflies were clustered into supergroup B. Phylogenetic trees of whitefly mitochondrial cytochrome oxidase subunit I and Wolbachia multiple sequencing typing genes were not congruent. In addition, Wolbachia was also detected outside the special bacteriocytes in two cryptic species by fluorescence in situ hybridization, indicating the horizontal transmission of Wolbachia. Our results indicate that members of Wolbachia are far from well explored.

The whitefly Bemisia tabaci complex contains many cryptic species, of which the Middle East-Asia Minor 1 (MEAM1) and Mediterranean (MED) are notorious invasive pests. In our field-collected whitefly samples, MEAM1 harbors an obligate primary symbiont "Candidatus Portiera aleyrodidarum" and two secondary symbionts, "Candidatus Hamiltonella defensa" and Rickettsia sp., whereas MED has only "Ca. Portiera aleyrodidarum" and "Ca. Hamiltonella defensa." Both "Ca. Portiera aleyrodidarum" and "Ca. Hamiltonella defensa" are intracellular endosymbionts residing in the bacteriomes, whereas Rickettsia sp. has a scattered distribution throughout the host body cavity. We examined responses of these symbionts to adverse temperatures as well as survival of the host insects. After cold treatment at 5 or 10 °C or heat treatment at 35 or 40 °C for 24 h, respectively, the infection rates of all symbionts were not significantly decreased based on diagnosis PCR. However, quantitative PCR assays indicated significant reduction of "Ca. Hamiltonella defensa" at 40 °C, and the reduction became greater as the duration increased. Compared with "Ca. Hamiltonella defensa," "Ca. Portiera aleyrodidarum" was initially less affected in the first day but then showed more rapid reduction at days 3-5. The density of Rickettsia sp. fluctuated but was not reduced significantly at 40 °C. Meanwhile, the mortality rates of the host whiteflies elevated rapidly as the duration of exposure to heat treatment increased. The differential responses of various symbionts to adverse temperatures imply complex interactions among the symbionts inside the same host insect and highlight the importance of taking the whole bacterial community into account in studies of symbioses.

Trehalases (Tres) have been demonstrated to be the key enzymes that are involved in various trehalose-associated physiological processes in insects. However, little attention has been devoted to the Tres in the whitefly, Bemisia tabaci. In this study, a soluble Tre (BtTre-1) and a membrane-bound Tre (BtTre-2) were cloned in the invasive cryptic species Middle East-Asia Minor 1 (MEAM1) of the whitefly B. tabaci complex. Alignment of deduced amino acids sequences of both BtTres revealed that they share common consensus regions and residues with Tres of other insect species. Levels of BtTres expression in various stages and tissues of the whitefly suggested that BtTre-2 may play a key role in trehalose catabolism during development of the whitefly, especially for oocyte development, while BtTre-1 may prevent trehalose in salivary gland from leaking and entering into plants along with saliva. Potential roles of trehalose catabolism in response to direct and/or plant-mediated indirect effects of Tomato Yellow Leaf Curl China Virus (TYLCCNV) were also detected. Whiteflies feeding on virus-infected tobacco plants showed higher BtTres expressions and accordingly higher BtTres activity but lower trehalose content than those feeding on uninfected plants. The enhanced trehalose catabolism may be beneficial to oocyte development in ovary and attenuate plant defensive responses induced by trehalose in saliva. Viruliferous and nonviruliferous whiteflies feeding on cotton, a nonhost plant for TYLCCNV, differed significantly only in trehalose content. The higher trehalose content in viruliferous whiteflies may be conducive to resisting the stress inflicted by TYLCCNV.

The whitefly Bemisia tabaci (Gennadius) is considered one of the main pests for agriculture. One important problem with the whitefly is its notorious status as a vector for plant viruses, primarily begomoviruses. We have previously identified a defensin-like antimicrobial peptide, Btdef, from the whitefly B. tabaci MEAM1. However, the function of Btdef in the immune system of the insect vector and begomovirus transmission has yet to be explored.To explore the role of Btdef during begomovirus transmission, we firstly investigated the transcriptional response of Btdef following acquisition of Tomato yellow leaf curl China virus (TYLCCNV). The expression of Btdef was up-regulated in the viruliferous whiteflies. After RNA silencing of the Btdef gene in adult whiteflies fed with dsRNA, they were allowed to feed on TYLCCNV-infected plants and then quantified for TYLCCNV DNA titre. Unexpectedly, silencing the Btdef gene reduced both the abundance and expressions of TYLCCNV genes in the whiteflies. In the meantime, the density of the endosymbiont Rickettsia was significantly reduced in dsBtdef-fed whiteflies.Our data provide evidence that Btdef is involved in begomovirus infection, possibly through symbiont-mediated alteration of begomovirus-whitefly interactions. These findings indicate that Btdef may be targeted for the development of new technology for the control of whitefly-transmitted begomoviruses. © 2016 Society of Chemical Industry.

Tsetse flies are biological vectors of African trypanosomes, the protozoan parasites responsible for causing human and animal trypanosomiases across sub-Saharan Africa. Currently, no vaccines are available for disease prevention due to antigenic variation of the Variant Surface Glycoproteins (VSG) that coat parasites while they reside within mammalian hosts. As a result, interference with parasite development in the tsetse vector is being explored to reduce disease transmission. A major bottleneck to infection occurs as parasites attempt to colonize tsetse’s midgut. One critical factor influencing this bottleneck is the fly’s peritrophic matrix (PM), a semipermeable, chitinous barrier that lines the midgut. The mechanisms that enable trypanosomes to cross this barrier are currently unknown. Here, we determined that as parasites enter the tsetse’s gut, VSG molecules released from trypanosomes are internalized by cells of the cardia—the tissue responsible for producing the PM. VSG internalization results in decreased expression of a tsetse microRNA (mir-275) and interferes with the Wnt-signaling pathway and the Iroquois/IRX transcription factor family. This interference reduces the function of the PM barrier and promotes parasite colonization of the gut early in the infection process. Manipulation of the insect midgut homeostasis by the mammalian parasite coat proteins is a novel function and indicates that VSG serves a dual role in trypanosome biology—that of facilitating transmission through its mammalian host and insect vector. We detail critical steps in the course of trypanosome infection establishment that can serve as novel targets to reduce the tsetse’s vector competence and disease transmission.

The invasive Middle East-Asia Minor 1 (MEAM1) cryptic species (formerly referred to as the B ‘biotype’) of the whitefly Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) complex has become a major crop pest worldwide due to its global invasion as well as epidemics of begomoviruses it transmits. Previous studies demonstrated that MEAM1 performed better on tobacco [Nicotiana tabacum L. (Solanaceae)] plants infected by Tomato yellow leaf curl China virus (TYLCCNV) than on uninfected plants. However, mechanisms underlying the interactions are not completely understood. In this study, we compared the composition of sugars and amino acids in the phloem sap of uninfected and TYLCCNV-infected tobacco plants with or without whiteflies feeding on the plants. No significant differences were detected in terms of amino acid profiles, percentage of essential amino acids, and sugars:amino acids ratio. However, whiteflies feeding on TYLCCNV-infected plants excreted comparatively a lower percentage of amino acids, especially essential amino acids, or proportionally a higher level of sugars than those feeding on uninfected plants, suggesting that whiteflies feeding on virus-infected plants are able to achieve a more balanced nutritional assimilation. We also compared the honeydew excreted by viruliferous and non-viruliferous whiteflies feeding on cotton [Gossypium hirsutum L. (Malvaceae)], a non-host plant for TYLCCNV, to test the direct effects of the virus on whiteflies. Although the amino acids profiles in honeydew were similar between viruliferous and non-viruliferous whiteflies, the sugars:amino acids ratio in honeydew excreted by viruliferous whiteflies was lower than that of non-viruliferous whiteflies, suggesting that the viruliferous whiteflies assimilated a more imbalanced nutrition with proportionally more sugar. Taken together, our data indicate that although a plant virus may exert negative direct effects on its vector, feeding by the vector on virus-infected plants may improve nutritional assimilation and consequently performance of the vector.

Endosymbionts are important components of arthropod biology. The whitefly Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a cryptic species complex composed of ≥ 28 putative species. In addition to the primary endosymbiont Portiera aleyrodidarum, six secondary endosymbionts (S-endosymbionts), Hamiltonella, Rickettsia, Wolbachia, Cardinium, Arsenophonus and Fritschea, have been identified in B. tabaci thus far. Here, we tested five of the six S-endosymbiont lineages (excluding Fritschea) from 340 whitely individuals representing six putative species from China. Hamiltonella was detected only in the two exotic invaders, Middle East-Asia Minor 1 (MEAM1) and Mediterranean (MED). Rickettsia was absent in Asia II 1 and MED, scarce in Asia II 3 (13%), but abundant in Asia II 7 (63.2%), China 1 (84.7%) and MEAM1 (100%). Wolbachia, Cardinium and Arsenophonus were absent in the invasive MEAM1 and MED but mostly abundant in the native putative species. Furthermore, phylogenetic analyses revealed that some S-endosymbionts have several clades and different B. tabaci putative species can harbor different clades of a given S-endosymbiont, demonstrating further the complexity of S-endosymbionts in B. tabaci. All together, our results demonstrate the variation and diversity of S-endosymbionts in different putative species of B. tabaci, especially between invasive and native whiteflies.

Arthropod vectors have multiple physical and immunological barriers that impede the development and transmission of parasites to new vertebrate hosts. These barriers include the peritrophic matrix (PM), a chitinous barrier that separates the blood bolus from the midgut epithelia and modulates vector-pathogens interactions. In tsetse flies, a sleeve-like PM is continuously produced by the cardia organ located at the fore- and midgut junction. African trypanosomes, Trypanosoma brucei, must bypass the PM twice; first to colonize the midgut and secondly to reach the salivary glands (SG), to complete their transmission cycle in tsetse. However, not all flies with midgut infections develop mammalian transmissible SG infections—the reasons for which are unclear. Here, we used transcriptomics, microscopy and functional genomics analyses to understand the factors that regulate parasite migration from midgut to SG. In flies with midgut infections only, parasites fail to cross the PM as they are eliminated from the cardia by reactive oxygen intermediates (ROIs)—albeit at the expense of collateral cytotoxic damage to the cardia. In flies with midgut and SG infections, expression of genes encoding components of the PM is reduced in the cardia, and structural integrity of the PM barrier is compromised. Under these circumstances trypanosomes traverse through the newly secreted and compromised PM. The process of PM attrition that enables the parasites to re-enter into the midgut lumen is apparently mediated by components of the parasites residing in the cardia. Thus, a fine-tuned dialogue between tsetse and trypanosomes at the cardia determines the outcome of PM integrity and trypanosome transmission success., Author summary Insects are responsible for transmission of parasites that cause deadly diseases in humans and animals. Understanding the key factors that enhance or interfere with parasite transmission processes can result in new control strategies. Here, we report that a proportion of tsetse flies with African trypanosome infections in their midgut can prevent parasites from migrating to the salivary glands, albeit at the expense of collateral damage. In a subset of flies with gut infections, the parasites manipulate the integrity of a midgut barrier, called the peritrophic matrix, and reach the salivary glands for transmission to the next mammal. Either targeting parasite manipulative processes or enhancing peritrophic matrix integrity could reduce parasite transmission.

We report here the sequence of a complete mitochondrial genome (mitogenome) of a population of Bemisia afer species complex, which was collected from Abutilonaviecnnae gaerner at LinYi, Shan Dong province of China (hereafter B. afer_China_Ag) (GenBank accession number: KR819174). The mitogenome of B. afer_China_Ag is 15 300 bp and contains 13 protein-coding genes, 22 transfer RNAs, 2 ribosomal RNAs, and 2 control regions. The content and arrangement of mitochondrial genes of B. afer_China_Ag exhibit remarkable differences from the mitogenome of a B. afer population collected from African cassava (KF734668). Phylogenetic analyses based on 13 protein-coding genes support the close relationship of the two sequenced B. afer mitogenomes and confirm the species status of our specimen.

Trehalases (Tres) have been demonstrated to be the key enzymes that are involved in various trehalose-associated physiological processes in insects. However, little attention has been devoted to the Tres in the whitefly, Bemisia tabaci. In this study, a soluble Tre (BtTre-1) and a membrane-bound Tre (BtTre-2) were cloned in the invasive cryptic species Middle East-Asia Minor 1 (MEAM1) of the whitefly B. tabaci complex. Alignment of deduced amino acids sequences of both BtTres revealed that they share common consensus regions and residues with Tres of other insect species. Levels of BtTres expression in various stages and tissues of the whitefly suggested that BtTre-2 may play a key role in trehalose catabolism during development of the whitefly, especially for oocyte development, while BtTre-1 may prevent trehalose in salivary gland from leaking and entering into plants along with saliva. Potential roles of trehalose catabolism in response to direct and/or plant-mediated indirect effects of Tomato Yellow Leaf Curl China Virus (TYLCCNV) were also detected. Whiteflies feeding on virus-infected tobacco plants showed higher BtTres expressions and accordingly higher BtTres activity but lower trehalose content than those feeding on uninfected plants. The enhanced trehalose catabolism may be beneficial to oocyte development in ovary and attenuate plant defensive responses induced by trehalose in saliva. Viruliferous and nonviruliferous whiteflies feeding on cotton, a nonhost plant for TYLCCNV, differed significantly only in trehalose content. The higher trehalose content in viruliferous whiteflies may be conducive to resisting the stress inflicted by TYLCCNV.

Endosymbionts are important components of arthropod biology. The whitefly Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a cryptic species complex composed of ≥ 28 putative species. In addition to the primary endosymbiont Portiera aleyrodidarum, six secondary endosymbionts (S-endosymbionts), Hamiltonella, Rickettsia, Wolbachia, Cardinium, Arsenophonus and Fritschea, have been identified in B. tabaci thus far. Here, we tested five of the six S-endosymbiont lineages (excluding Fritschea) from 340 whitely individuals representing six putative species from China. Hamiltonella was detected only in the two exotic invaders, Middle East-Asia Minor 1 (MEAM1) and Mediterranean (MED). Rickettsia was absent in Asia II 1 and MED, scarce in Asia II 3 (13%), but abundant in Asia II 7 (63.2%), China 1 (84.7%) and MEAM1 (100%). Wolbachia, Cardinium and Arsenophonus were absent in the invasive MEAM1 and MED but mostly abundant in the native putative species. Furthermore, phylogenetic analyses revealed that some S-endosymbionts have several clades and different B. tabaci putative species can harbor different clades of a given S-endosymbiont, demonstrating further the complexity of S-endosymbionts in B. tabaci. All together, our results demonstrate the variation and diversity of S-endosymbionts in different putative species of B. tabaci, especially between invasive and native whiteflies.

“ Candidatus Hamiltonella defensa” is a facultative endosymbiont of the whitefly Bemisia tabaci . Herein, we report the first draft genome sequence of “ Candidatus Hamiltonella defensa” from the invasive Mediterranean cryptic species of the B. tabaci complex. The 1.84-Mbp genome sequence comprises 404 contigs and contains 1,806 predicted protein-coding genes.