Ruiling Zhang

The sesquiterpenoid juvenile hormone (JH) plays a central role in insect life and is involved in the regulation of multiple biological functions. Precise control of JH biosynthesis is crucial for normal insect development. JH acid O‐methyltransferase (JHAMT) is a key regulatory enzyme that functions in the final step of JH biosynthesis, which is highly specific to insects. Significant progress has been made in identifying and characterizing the features of the JHAMT. Functional studies have confirmed that JHAMT is involved in metamorphosis, development, reproduction, and a wide range of other biological processes, such as diapause, behavior, and vitellogenin gene expression in different insect species through regulating titers of JH. Herein, we summarized the current knowledge regarding the diverse functions of JHAMT and the interactions of JHAMT with other genes, translation factors, and signaling networks governing the growth and development of insects, and the structures of JHAMT. This wealth of knowledge will contribute to novel insights into the role of JHAMT in insects and its practical implications for pest management.

Background: House fly gut microbiota plays a major role in regulating larval development and immune responses. As the house fly gut microbiota is complex with different functions, the role of many gut microbiota in the host remains unknown. The differential oxygen content in the midgut and hindgut of house fly intestines creates distinct gut microenvironments, resulting in variations in the composition of gut microbiota. Research on the distribution of aerobic and facultative anaerobic bacteria in different parts of the intestine—and how they interact with each other—is helpful for exploring the interaction between intestinal bacteria and the host in aerobic and anaerobic states. However, there have been no mechanistic studies on the interactions of gut bacteria in different oxygen microenvironments within the house fly. Methods: In this study, aerobic Enterobacter hormaechei EhX and facultative anaerobic E. hormaechei EhY were isolated from the guts of house fly larvae. The specific phages, EhXP and EhYP, were isolated against these two bacteria, with phage EhXP and EhYP specifically targeting EhX and EhY, respectively. Furthermore, the role of the aerobic–anaerobic interaction in E. hormaechei during larval development was investigated using phage and bacterial feeding experiments. Results: The results showed that the addition of aerobic E. hormaechei EhX or facultative anaerobic E. hormaechei EhY alone to the diet promoted larval growth and phenoloxidase activity. However, in the phage-treated groups, the opposite effect was observed. In contrast, EhY was the dominant bacteria in larval hindgut. Conversely, EhX was the main bacteria in the midgut of larvae. Aerobic/facultative anaerobic gut bacteria competed and cooperated to maintain a stable composition of larval gut microbiota, forming a negative correlational “pedal effect” promoting larval development. Through high-throughput sequencing it was demonstrated that the abundance of beneficial bacteria increased in the bacteria treatment group. The number of harmful bacteria increased in the phage treatment group. Conclusions: The results revealed the crucial role of aerobic and facultative anaerobic E. hormaechei in the house fly. These results highlight the potential of future applications of multiple phages for the study of insect intestinal flora. Graphical

The prevalence, evolution, and disease risk of pathogenic tick-borne viruses (TBVs) remains poorly understood in northwest China and adjacent countries, which are endemic for several TBVs of public health importance. Herein, we perform meta-transcriptomic sequencing of >9600 ticks collected across this vast geographic area to identify 92 RNA viruses and assemble 1567 viral genomes from 28 different tick species, including ten human- and mammal-infecting TBVs. Tacheng tick virus 1 (TcTV-1), Tacheng tick virus 2 (TcTV-2), Tamdy virus (TAMV), and Crimean-Congo hemorrhagic fever virus (CCHFV) are the most common human-infecting TBVs detected. We also report several pathogenic TBVs not previously identified in China (Burana virus and Bhanja virus), Kazakhstan (TcTV-1), and northwest China (Wad Medani virus and Alongshan virus). We predict a significant increase in the number of high-risk regions for the four major tick vectors of pathogenic TBVs, and the distribution of TAMV, TcTV-1, TcTV-2, and CCHFV, with 65.4% of the counties in this region identified as high risk for CCHFV. Therefore, the real disease burden caused by TBVs in northwest China and adjacent countries may be more underestimated than appreciated, and we call for strengthened field surveys and epidemiological surveillance in this vast region.

The invasive species Aedes albopictus is a major vector of several arboviruses. The global spread of this species seriously threatens human health. Insecticide resistance is an increasing problem worldwide that limits the efficacy of mosquito control. As the major structural component of cuticles, chitin is indispensable to insects. Chitin synthase (CHS) is the enzyme that catalyzes the biosynthesis of chitin at the final step. In this study, two CHS genes of Aedes albopictus (AaCHS1 and AaCHS2) were identified and their basic characteristics were evaluated via bioinformatics analysis. The highest abundance of AaCHS1 transcripts was detected in pupae, whereas that of AaCHS2 transcripts was detected in females; the highest expression levels of AaCHS1 and AaCHS2 were found in the epidermis and the midgut of pupae, respectively. The survival and emergence rates of pupae were significantly reduced after the injection of double‐stranded RNA of AaCHS1 or AaCHS2, indicating that both AaCHS1 and AaCHS2 play crucial roles in the pupal development. In addition, the chitin content of pupae was obviously decreased after the suppression of AaCHS1 expression by RNA interference (RNAi) treatment. This influence of the RNAi treatment was further supported by the reduced chitin thickness and weakened chitin fluorescence signal in the new cuticle. The midgut of pupae presented a reduced intensity of the chitin fluorescence signal along with RNAi treatment specific to AaCHS2 expression. The results of this study indicate that CHS genes may be suitable as molecular targets used for controlling mosquitoes.

Background: Houseflies, Musca domestica L., are an ubiquitous pest that can transmit numerous diseases and threaten human health. Increasing insecticide resistance shown by houseflies necessitates the develop new control alternatives. The housefly gut is densely colonized with microorganisms that interact with each other dynamically and benefit the host’s health. However, the impact of multiple symbiotic bacteria on the composition of housefly gut microbiota and the host’s activities remains unclear. Methods: We isolated and cultured 12 bacterial species from the intestines of housefly larvae. We also isolated seven bacteriophages to precisely target the regulation of certain bacterial species. Using 16S rRNA high-throughput gene sequencing, we analyzed the bacterial diversity after orally administering bacteria/phage cocktails to houseflies. Results: Our results showed that larval growth was promoted, the abundance of beneficial bacteria, such as Klebsiella and Enterobacter, was increased and the abundance of harmful bacteria, such as Providencia, Morganella and Pseudomonas, was decreased in housefly larvae fed with the beneficial bacteria cocktail. However, oral administration of both beneficial and harmful bacterial phage cocktails inhibited larval growth, probably due to the drastic alteration of gut flora. Untargeted metabolomics using liquid chromatography–mass spectrometry showed that disturbances in gut microbiota changed the larval metabolite profiles. Feeding experiments revealed that disrupting the intestinal flora suppressed the beneficial bacteria and increased the harmful bacteria, causing changes in the metabolites and inhibiting larval growth. Conclusions: Based on our results, bacteria/phage cocktails are effective tools for regulating the intestinal flora of insects and have a high potential as a biological control agent for incorporation into an integrated pest management program. Graphical

Background: Aedes albopictus is a vector of numerous devastating arboviruses and places heavy burdens on global public health. Chitin is one of the important components of cuticles and targeting chitin metabolism is a promising strategy for preventing mosquito dispersal and mosquito-borne diseases. Increasing evidence suggests that microRNAs (miRNAs) play crucial roles in various physiological processes of insects. Methods: A previous analysis suggested that the microRNA miR-989 is potentially involved in chitin metabolism in Ae. albopictus pupae. In the present study, we found that the expression level of miR-989 was significantly overexpressed after injection of agomir. A dual-luciferase assay was used to determine the direct target of miR-989. Survival rate, eclosion rate and malformation rate were statistically analyzed to evaluate the potential effect of miR-989. Hematoxylin–eosin staining and chitin staining were used to evaluate the microstructural changes in the cuticles of Ae. albopictus pupae. Results: Overexpression of miR-989 resulted in a significantly reduced survival rate and eclosion rate of pupae and an elevated malformation rate of adults. The results suggested that miR-989 acted as a regulator of chitin metabolism in Ae. albopictus pupae by affecting the transcript levels of the Ae. albopictus genes encoding chitin synthase 1 (AaCHS1) and chitinase 10 (AaCht10). The altered expression levels of the two chitin metabolism-related enzymes (CHS1 and Cht10, respectively) caused the structural changes in cuticles and further affected the pupal-adult transition process of Ae. albopictus. XM_029863591.1 was proven to be the target gene of miR-989 and displayed similar effects on pupae as miR-989. Conclusions: The microRNA miR-989 was found to be essential for chitin metabolism in old and new cuticles of Ae. albopictus pupae. The results of the current study suggested that miR-989 could be used as a potential target to control Ae. albopictus. Graphical

Background: Aedes albopictus is an increasingly serious threat in public health due to it is vector of multiple arboviruses that cause devastating human diseases, as well as its widening distribution in recent years. Insecticide resistance is a serious problem worldwide that limits the efficacy of chemical control strategies against Ae. albopictus. Chitinase genes have been widely recognized as attractive targets for the development of effective and environmentally safe insect management measures. Methods: Chitinase genes of Ae. albopictus were identified and characterized on the basis of bioinformatics search of the referenced genome. Gene characterizations and phylogenetic relationships of chitinase genes were investigated, and spatio-temporal expression pattern of each chitinase gene was evaluated using qRT-PCR. RNA interference (RNAi) was used to suppress the expression of AaCht10, and the roles of AaCht10 were verified based on phynotype observations, chitin content analysis and hematoxylin and eosin (H&E) stain of epidermis and midgut. Results: Altogether, 14 chitinase-related genes (12 chitinase genes and 2 IDGFs) encoding 17 proteins were identified. Phylogenetic analysis showed that all these AaChts were classified into seven groups, and most of them were gathered into group IX. Only AaCht5-1, AaCht10 and AaCht18 contained both catalytic and chitin-binding domains. Different AaChts displayed development- and tissue-specific expression profiling. Suppression of the expression of AaCht10 resulted in abnormal molting, increased mortality, decreased chitin content and thinning epicuticle, procuticle and midgut wall of pupa. Conclusions: Findings of the present study will aid in determining the biological functions of AaChts and also contribute to using AaChts as potential target for mosquito management. Graphical

Background: The Asian tiger mosquito,Aedes albopictus, is one of the most invasive species and a vector of numerous arboviruses. The deleterious effects of long-term and inappropriate use of chemical pesticides have stimulated the exploration of new, environmentally friendly control strategies. Non-coding RNAs (ncRNAs) have been proven to participate in almost all biological processes of insects.Methods: In this study, circular RNAs (circRNAs) and microRNAs (miRNAs) covering five developmental stages [egg, early larvae, late larvae, pupae, adult (female and male)] ofA. albopictuswere obtained using whole-transcriptome sequencing technology. Combined with long non-coding RNAs (lncRNAs) from previous research, circRNA/lncRNA‒miRNA‒mitochondrial RNA (mRNA) networks were constructed.Results: A total of 1434 circRNAs and 208 miRNAs were identified. More differentially expressed circRNAs (DE circRNAs) and miRNAs (DE miRNAs) were found in the egg versus early larvae comparison group. Functional enrichment analysis demonstrated that most of the circRNA/lncRNA‒miRNA‒mRNA networks were involved in chitin metabolism. Hub genes of each circRNA/lncRNA‒miRNA‒mRNA network were screened out, which can be used as novel targets to disturb the molting process ofA. albopictus.Conclusions: Regulatory relationships obtained from competing endogenous RNA (ceRNA) networks provide more information to manipulate the metamorphosis process and are helpful for developing effective and sustainable methods to control mosquitoes.Graphical

Culex pipiens molestus and Culex pipiens pallens are two distinct bioforms in the Culex pipiens complex that are important vectors of several pathogens and are widely distributed around the world. In the current study, we present a high-quality chromosome-level genome of Cx. pipiens f. molestus and describe the genetic characteristics of this genome. The assembly genome was 559.749 Mb with contig and scaffold N50 values of 200.952 Mb and 0.370 Mb, and more than 94.78% of the assembled bases were located on 3 chromosomes. A total of 19,399 protein-coding genes were predicted. Many gene families were expanded in the genome of Cx. pipiens f. molestus, particularly those of the chemosensory protein (CSP) and gustatory receptor (GR) gene families. In addition, utilizing Hi-C data, we improved the previously assembled draft genome of Cx. pipiens f. pallens, with scaffold N50 of 186.195 Mb and contig N50 of 0.749 Mb, and more than 97.02% of the assembled bases were located on three chromosomes. This reference genome provides a foundation for genome-based investigations of the unique ecological and evolutionary characteristics of Cx. pipiens f. molestus, and the findings in this study will help to elucidate the mechanisms involved in species divergence in the Culex pipiens complex.