Effects of feeding on different parts of Ailanthus altissima on the intestinal microbiota of Eucryptorrhynchus scrobiculatus and Eucryptorrhynchus brandti (Coleoptera: Curculionidae).

Eucryptorrhynchus brandti and Eucryptorrhynchus scrobiculatus (Coleoptera: Curculionidae) are two monophagous weevil pests that feed on Ailanthus altissima (Mill.) Swingle but differ in their diet niche. In the field, adults of E. brandti prefer to feed on the trunk of A. altissima, whereas adults of E. scrobiculatus prefer to feed on the tender parts. We conducted Illumina sequencing of 16S rRNA to examine changes in bacterial diversity in the adults of these two weevil species after they fed on different parts of A. altissima (trunk, 2-3-year-old branches, annual branches, and petioles). Proteobacteria, Tenericutes, and Firmicutes were the dominant phyla in E. brandti (relative abundance was 50.64, 41.56, and 5.63%, respectively) and E. scrobiculatus (relative abundance was 78.63, 11.91, and 7.41%, respectively). At the genus level, Spiroplasma, endosymbionts2, Unclassified Enterobacteriaceae, and Lactococcus were dominant in E. brandti, and Unclassified Enterobacteriaceae, Wolbachia and Spiroplasma, and endosymbionts2 were dominant in E. scrobiculatus. Linear discriminant analysis effect size analysis revealed microbial biomarkers in the different treatment group of adults of both weevil species. Adults of E. brandti may require the trunk, and adults of E. scrobiculatus may require the petioles and annual branches to maintain the high diversity of their gut microbes. The results of this study indicate that feeding on different parts of A. altissima affects the composition and function of the microbes of E. brandti and the microbial composition of E. scrobiculatus. Variation in the abundance of Wolbachia and Spiroplasma in E. brandti and E. scrobiculatus is associated with dietary niche changes, and this might explain the evolution of reproductive isolation between these two sibling weevil species.

Population differentiation and intraspecific genetic admixture in two Eucryptorrhynchus weevils (Coleoptera: Curculionidae) across northern China.

Increasing damage of pests in agriculture and forestry can arise both as a consequence of changes in local species and through the introduction of alien species. In this study, we used population genetics approaches to examine population processes of two pests of the tree-of-heaven trunk weevil (TTW), Eucryptorrhynchus brandti (Harold) and the tree-of-heaven root weevil (TRW), E. scrobiculatus (Motschulsky) on the tree-of-heaven across their native range of China. We analyzed the population genetics of the two weevils based on ten highly polymorphic microsatellite markers. Population genetic diversity analysis showed strong population differentiation among populations of each species, with F ST ranges from 0.0197 to 0.6650 and from -0.0724 to 0.6845, respectively. Populations from the same geographic areas can be divided into different genetic clusters, and the same genetic cluster contained populations from different geographic populations, pointing to dispersal of the weevils possibly being human-mediated. Redundancy analysis showed that the independent effects of environment and geography could account for 93.94% and 29.70% of the explained genetic variance in TTW, and 41.90% and 55.73% of the explained genetic variance in TRW, respectively, indicating possible impacts of local climates on population genetic differentiation. Our study helps to uncover population genetic processes of these local pest species with relevance to control methods.

Reproductive Dormancy in Overwintering Adult Eucryptorrhynchus brandti (Coleoptera: Curculionidae).

Dormancy is important for overwintering insects to resist and adapt to adverse conditions. Dormancy generally contains quiescence and diapause. Eucryptorrhynchus brandti Harold (Coleoptera: Curculionidae), tree-of-heaven trunk weevil (TTW), is a destructive pest and highly host-specific to Ailanthus altissima in China. TTW has one generation per year and overwinters as both larvae and adults. In this study, to examined dormancy type of adults and find a method to store overwintering adults, we collected adults from 20 October 2018 to 13 March 2019. We studied the behavior and reproductive development of adults under field cold conditions for 0 and 10 d and laboratory warm conditions for 5 and 10 d. We recorded developing eggs in females, and the clarity of the testis edge, the yellow point in the testis lobe, the ratio of the inner content in the accessory gland, and the accessory gland color in males. Adults transferred from the field to the laboratory had resumed reproductive development directly. Results indicated that the dormancy type of TTW adults was quiescence. Adults stored in the field were still in a dormant state and the field-storage method was effective. Current study provided basic data for controlling overwintering TTW adults and solve the storage of insect sources during the winter.

Red imported fire ants (Hymenoptera: Formicidae) cover inaccessible surfaces with particles to facilitate food search and transportation.

Eusocial insects have evolved diverse particle-use behaviors. A previous study reported that red imported fire ants, Solenopsis invicta Buren, deposited soil particles on substances treated with essential balm, a fire ant repellent. We hypothesized that S. invicta modifies inaccessible surfaces by covering them with soil particles to facilitate food search and transportation. Here, laboratory experiments were conducted to study the particle-covering behavior of S. invicta in response to viscose surfaces or surfaces treated with essential balm or liquid paraffin in the presence of real food (sausage) or non-food objects (acrylic plates). S. invicta workers deposited significantly more soil particles on these three types of treated surfaces than on untreated surfaces. In addition, significantly more particles were relocated on viscose and paraffin-smeared surfaces in the presence of food than in the presence of non-food objects. The particle-covering behavior on viscose surfaces was also observed in the field. Interestingly, when no soil particles were available, ants searched and transported food on viscose surfaces only if the surfaces were artificially covered with sufficient quantities of soil particles but could not do so on viscose surfaces without soil particles or with insufficient quantities of soil particles. In addition, ants actively relocated particles to cover viscose surfaces if the transportation distance was within 200 mm, whereas significantly fewer particles were relocated at longer transportation distances (400 mm). Our study provides a novel example of particle use by fire ants during foraging.

Oviposition Behavior and Distribution of Eucryptorrhynchus scrobiculatus and E. brandti (Coleoptera: Curculionidae) on Ailanthus altissima (Mill.).

(1) Eucryptorrhynchus scrobiculatus Motschulsky (Coleoptera: Curculionidae: Cryptorrhychinae) is a major quarantine forest pest in China. It often co-occurs with E. brandti (Coleoptera: Curculionidae: Cryptorrhychinae) on a single host Ailanthus altissima (Mill.) Swingle (tree of heaven). (2) In this study, to explain the coexistence of the two weevils on a single host, we investigated the oviposition behavior of E. scrobiculatus and oviposition sites of E. scrobiculatus and E. brandti under afield and laboratory conditions. (3) The characteristic behaviors of E. scrobiculatus females prior to oviposition included searching, locating, excavation, turning, locating the oviposition cavity, egg deposition, and hiding. (4) The oviposition sites used by E. scrobiculatus and E. brandti differed. Eucryptorrhynchus scrobiculatus females laid eggs in the soil near A. altissima and compound leaf petioles, while E. brandti females laid eggs in A. altissima trunks. The eggs in compound leaf petioles did not hatch in the field. (5) Eucryptorrhynchus scrobiculatus and E. brandti utilized different oviposition sites and these differences in habitat use may reduce the competition for resources between species during the larval period, thus facilitating their coexistence on A. altissima.

Effects of Trap Color and Shape on the Capture of Eucryptorrhynchus scrobiculatus (Coleoptera: Curculionidae).

Eucryptorrhynchus scrobiculatus (Coleoptera:Curculionidae) is an important pest in China that specifically damages Ailanthus altissima (Mill.) Swingle (Sapindales: Simaroubaceae). Trembler grid lamps and food attractant traps frequently were used to monitor and control E. scrobiculatus; however, the effect of these methods is poor in actual application. The purpose of this study was to develop a new monitoring method. Traps of seven colors and two shapes were compared with respect to the attraction of E. scrobiculatus adults, and a field trapping test was performed. We found that E. scrobiculatus adults were most sensitive to red (16.11 ± 7.72) and black traps (14.44 ± 8.07) and to tall vertical black shapes in the laboratory. In the field, red (70.50 ± 5.74) and black traps (60.75 ± 8.22) were most effective at catching E. scrobiculatus, and traps with color and attractant still were more attractive to E. scrobiculatus than traps with colors only. These results provide a reference for monitoring E. scrobiculatus adults.

Transcriptomic Analysis of Eucryptorrhynchus chinensis (Coleoptera: Curculionidae) Using 454 Pyrosequencing Technology.

Eucryptorrhynchus chinensis Olivier (Coleoptera: Curculionidae) is one of the most important pests of Ailanthus altissima; however, so far, no studies on the genome or transcriptome of E. chinensis have been reported. Using the Roche 454 FLX Titanium platform, an RNA pool obtained from E. chinensis eggs, larva, pupae, and adults was sequenced and assembled de novo to achieve maximum diversity of sampled transcripts. We obtained 1,441,137 (∼518 Mb) raw reads with an average length of 360 bp. After trimming, 89% qualified reads were produced and assembled into 35,509 isotigs with an average length of 440 bp, N50 of 1,048 bp, and 111,643 singletons. We generated 87,894 unigenes following a cluster analysis of the isotigs and singletons, and then functionally annotated the unigenes with gene descriptions. We obtained 23,363 GO assignments, and 12,724 unigenes were assigned to KOG. Based on these annotations, 294 biochemical pathways involved in growth, reproduction, and stress or immune responses were predicted. A total of 659,026 single nucleotide variants and 6,112 simple sequence repeats were detected. Our data provide comprehensive information on the sequence and possible functions of E. chinensis transcripts.

Staphylococcal enterotoxin B-derived haptens promote sensitization.

T helper 2 (Th2) polarization is a major pathological feature in allergic diseases; its etiology is not fully understood. This study aims to elucidate the adjuvant effect of the microbial product-derived small peptides in the initiation of antigen-specific Th2 polarization. In this study, a clinical survey of patients with chronic rhinosinusitis (CRS) and food allergy (FA) was carried out. The Staphylococcal enterotoxin B (SEB)-derived small peptides (Ssps) were examined in the human stool extracts. The formation of Ssp/antigen adducts was tested in a protein-protein combination assay. The bone marrow-derived dendritic cells (BMDCs) were employed to test the role of Ssp/ovalbumin (OVA) adducts in the dendritic cell (DC) maturation. A mouse model was developed to test the role of Ssp/OVA adducts in the initiation of Th2 polarization in the intestine. The results showed that 54 (18.2%) patients with FA were diagnosed among 296 patients with SEB(+) CRS; only eight (2.9%) FA patients were identified among 272 patients with SEB(-) CRS. Ssps were detected in the stool protein extracts from FA patients with SEB(+) CRS, but not in those with SEB(-) CRS. Ssp/OVA adducts induced DC maturation, speeded up DC migration, activated CD4(+) T cells in the regional lymph nodes and induced skewed Th2 polarization in the local tissue. We conclude that patients with SEB(+) CRS are prone to suffering from FA. SEB can be degraded to Ssps in the gastrointestinal tract. The Ssps can bind macromolecular antigens to form adducts to promote the antigenicity of the antigens and induction of the antigen-specific Th2 polarization and inflammation in the local tissue.

Suppression of CB1 cannabinoid receptor by lentivirus mediated small interfering RNA ameliorates hepatic fibrosis in rats.

It is recognized that endogenous cannabinoids, which signal through CB1 receptors in hepatic stellate cells (HSCs), exert a profibrotic effect on chronic liver diseases. In this study, we suppressed CB1 expression by lentivirus mediated small interfering RNA (CB1-RNAi-LV) and investigated its effect on hepatic fibrosis in vitro and in vivo. Our results demonstrated that CB1-RNAi-LV significantly inhibited CB1 expression, and suppressed proliferation and extracellular matrix production in HSCs. Furthermore, CB1-RNAi-LV ameliorated dimethylnitrosamine induced hepatic fibrosis markedly, which was associated with the decreased expression of mesenchymal cell markers smooth muscle α-actin, vimentin and snail, and the increased expression of epithelial cell marker E-cadherin. The mechanism lies on the blockage of Smad signaling transduction induced by transforming growth factor ß1 and its receptor TGF-ß RII. Our study firstly provides the evidence that CB1-RNAi-LV might ameliorate hepatic fibrosis through the reversal of epithelial-to-mesenchymal transition (EMT), while the CB1 antagonists AM251 had no effect on epithelial-mesenchymal transitions of HSCs. This suggests that CB1 is implicated in hepatic fibrosis and selective suppression of CB1 by small interfering RNA may present a powerful tool for hepatic fibrosis treatment.

Phylogenetic relationship of seven Dendrolimus (Lepidoptera: Lasiocampidae) species based on the ultrastructure of male moths' antennae and antennal sensilla.

The morphology and ultrastructure of the antennae and antennal sensilla of seven male Dendrolimus species and a male Trabala vishnou gigantina (Yang) (Lepidoptera: Lasiocampidae) were examined by light microscope and scanning electron microscope. Six morphological types of antennal sensilla were identified: Sensilla trichodea, Sensilla chaetica, Sensilla styloconica, Sensilla coeloconica, Böhm bristles, and foot-like sensilla. Six of the Dendrolimus moths and Trabala vishnou gigantina Yang share the same antennal sensilla type, as do various geographic populations of the same species. The exception, Dendrolimus spectabilis Butler, has foot-like sensilla. However, the antennal sensilla subtypes were significantly different among species and/or populations. There were no remarkable differences in the width of the scape, pedicel, subflagellum, and the side-branches between the eight male species studied. However, we observed significant differences in the number of flagellomere and the length of scape, pedicel, subflagellums, and side-branches. The length and basal diameter of various types of antennal sensilla did not vary significantly among Dendrolimus moths. Beyond that, there were no differences among populations of the same kind of species. Hierarchical cluster analysis found two clusters: the first contained D. punctata punctata (Walker), D. punctata wenshanensis (Tsai et Liu), D. tabulaeformis (Tsai et Liu), and D. spectabilis and D. superans (Butler), and the second contained D. grisea (Moore) and D. kikuchii kikuchii (Matsumura). Trabala vishnou gigantina was placed separately from the two clusters. We conclude that D. punctata wenshanensis,D. tabulaeformis, and D. spectabilis are geographic subspecies of D. punctata punctata.

Integrin alphavbeta6 promotes tumor tolerance in colorectal cancer.

Tumor immune tolerance plays a critical role in tumor cell survival; the establishment of tumor immune tolerance is incompletely understood yet. Integrin alphavbeta6 (avb6) is involved in tumor growth and metastasis. This study aimed to observe the effect of avb6 on the development of tumor tolerance in colorectal cancer (CRC). In this study, 28 CRC patients were recruited. The frequencies of tolerogenic dendritic cells (TolDC), regulatory T cells (Treg), and CD8+ T cells in surgically removed CRC tissue were assessed by flow cytometry. The levels of avb6 in CRC tissue were measured by enzyme-linked immunoassay (ELISA). The effect of avb6 on inducing TolDCs and Tregs was evaluated with the cell culture model. The results showed that in surgically removed CRC tissue, we detected higher frequencies of TolDC and Tregs, lower frequency CD8+ T cells and high levels of avb6 as compared with non-CRC tissue. CRC protein extracts could induce TolDC development that could be blocked by anti-avb6 antibody. CRC-derived DCs could convert naïve CD4+ T cells to Tregs. Peripheral CD8+ T cells from CRC patients still retained the ability to produce granzyme B and to proliferate in response to CRC tumor antigen in culture that was abolished by the presence of CRC-derived Tregs. We conclude that CRC-derived avb6 is involved in the establishment of tumor immune tolerance in local tissues.

AAV-HGFK1 and Ad-p53 cocktail therapy prolongs survival of mice with colon cancer.

This study tried to evaluate the application of a novel cancer gene therapy using recombinant adeno-associated virus (AAV) carrying the kringle 1 domain of human hepatocyte growth factor (AAV-HGFK1) in combination with recombinant adenovirus carrying p53 gene (Ad-p53). BALB/c and nude mice models of colon cancer were established and the mice were treated with AAV-HGFK1 alone or in combination with Ad-p53. Combination of AAV-HGFK1 and Ad-p53 significantly prolonged the survival of the mice and also significantly inhibited primary and secondary tumor growth. Histochemical examination of the tumors revealed that AAV-HGFK1+Ad-p53 combinatorial treatment not only induced necrosis and apoptosis in the tumors but also suppressed tumor angiogenesis. The antiangiogenesis effect could likely be attributed to the ability of AAV-HGFK1+Ad-p53 viral cocktail to inhibit endothelial cell migration and proliferation. AAV-HGFK1+Ad-p53 also inhibited tumor cell growth in vitro by inhibiting epidermal growth factor receptor phosphorylation. Therefore, AAV-HGFK1+Ad-p53 cocktail therapy has a significantly higher therapeutic effect than AAV-HGFK1 or Ad-p53 alone and is a novel promising gene therapy for colon cancer.

[Establishment and comparison of liver metastasis models of two colorectal carcinoma cell lines in mice].

OBJECTIVE: To establish hepatic metastasis models of two colorectal carcinoma cell lines in mice for studying mechanism involved in colorectal carcinoma metastasis and its potential countermeasures. METHODS: Murine and human colorectal carcinoma CT26 and LoVo cells were inoculated into the spleen of Balb/c mice and Balb/c nude mice, respectively. The conditions of all the mice were observed, and the survival time and liver metastases were recorded. RESULTS: All mice inoculated with CT26 cells and a few with LoVo cells developed liver metastases without metastases in any other organs. Pathological examination identified the liver metastatic foci as poorly differentiated colonic adenocarcinoma. Compared with the mice inoculated with LoVo cells, those with CT26 cells had a higher rate of liver metastasis and a shorter survival time. CONCLUSION: The mouse model has been established successfully, which well mimics the pathological process of liver metastasis of colorectal cancer.