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SUMO modification is a vital post-translational regulation process in eukaryotes, in which the SUMO protease is responsible for the maturation of the SUMO precursor and the deconjugation of the SUMO protein from modified proteins by accurately cleaving behind the C-terminal Gly–Gly motif. To promote the understanding of the high specificity of the SUMO protease against the SUMO protein as well as to clarify whether the conserved Gly–Gly motif is strictly required for the processing of the SUMO precursor, we systematically profiled the specificity of the S. cerevisiae SUMO protease (Ulp1) on Smt3 at the P2–P1↓P1’ (Gly–Gly↓Ala) position using the YESS–PSSC system. Our results demonstrated that Ulp1 was able to cleave Gly–Gly↓ motif-mutated substrates, indicating that the diglycine motif is not strictly required for Ulp1 cleavage. A structural-modeling analysis indicated that it is the special tapered active pocket of Ulp1 conferred the selectivity of small residues at the P1–P2 position of Smt3, such as Gly, Ala, Ser and Cys, and only which can smoothly deliver the scissile bond into the active site for cleavage. Meanwhile, the P1’ position Ala of Smt3 was found to play a vital role in maintaining Ulp1’s precise cleavage after the Gly–Gly motif and replacing Ala with Gly in this position could expand Ulp1 inclusivity against the P1 and P2 position residues of Smt3. All in all, our studies advanced the traditional knowledge of the SUMO protein, which may provide potential directions for the drug discovery of abnormal SUMOylation-related diseases.

Wolbachia infection is known to affect host reproduction and development. To date, however, the underlying mechanism related to the effects of Wolbachia on host development has not yet been reported. Here, we compared the developmental duration and body weight in different instars of Wolbachia-positive (W+ ) and Wolbachia-negative (W- ) spiders (Hylyphantes graminicola) and detected the relative expression levels of 6 insulin-related genes and 3 ecdysone-related genes using reverse transcription qPCR. Results showed that the developmental duration was significantly shortened in W+ spiders compared with W- spiders. Furthermore, W+ spiders were significantly heavier than W- spiders at the 3rd and 4th instars, although no significant differences in body weight were observed after maturity. We also found that the expression levels of insulin-like growth factor-2 mRNA-binding protein-1, insulin-degrading enzyme, and ecdysone-inducible protein-1 genes were significantly down-regulated in W+ spiders compared with W- spiders, whereas the expression levels of insulin-like growth factor-binding protein-1, insulin-like peptide receptor, insulin receptor substrate 2-B, insulin-like, ecdysone-induced protein-2, and ecdysone receptor genes were significantly up-regulated in W+ spiders. Our results suggest that Wolbachia may influence host development by affecting insulin and ecdysone signaling pathways.

DNA methylation plays an important role in biological regulation in vertebrates, while its presence is variable among insects. However, only a few studies have reported such methylation in spiders. Here, we detected DNA methylation levels and patterns in the small-sized spider Hylyphantes graminicola (Araneae: Linyphiidae) through methylation-sensitive amplified fragment length polymorphism (MSAP). Given the prevailing rate of Wolbachia infection in H. graminicola and the potential impact on host DNA methylation, four H. graminicola groups, i.e., Wolbachia-uninfected (W−) males, Wolbachia-infected (W+) males, W− females, and W+ females, were used for detection of DNA methylation. In addition, quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of the DNA methyltransferase 1 (Dnmt1) gene in the four groups. Results demonstrated the existence of genomic DNA methylation in H. graminicola. The hemi- or fully methylated ratios at the internal cytosine (HMeCG or MeCG) was significantly higher than the hemi-methylated ratio at the external cytosine (HMeCCG) in both W− and W+ spiders. Compared to W− male and female spiders, hemi- or fully methylated ratios at the internal cytosine in W+ males and females were significantly lower, and Dnmt1 expression was significantly lower in W+ males and females than in W− males and females. These results suggest that a potential association may exist between Wolbachia infection and DNA methylation in H. graminicola.

Many arthropods exhibit flower-visiting behavior, including a variety of spider species. However, as spiders are assumed to be strictly predatory, flower-visiting spiders are an often neglected group. We conducted a systematic biodiversity study of flower-visiting spiders based on published papers and field surveys. Most previous studies have focused on the herbivorous behavior of flower-visiting spiders (nectivory or pollinivory) and their effects on host flowers (tritrophic interactions with flower-visiting insects). In our field survey, we utilized standard transect walks (active sampling) and colored pan traps (passive sampling) to investigate species occurrence, diurnal and seasonal variation, and flower color preference of flower-visiting spiders. From the transect walks, crab spider species were found to be the dominant flower-visiting spiders and, based on all spider species, juvenile visitors were significantly more common than adults. Furthermore, in terms of spider number and species richness, tulips were the preferred flower to visit. For the pan traps, wolf spiders were found to be the dominant spider species. No significant differences were observed in the number of spiders caught in different colored pans, suggesting that color may not be an important flower trait in regard to spider preference. To the best of our knowledge, this study is the first to propose the term 'flower-visiting spiders' and conduct a systematic investigation of their diversity. However, this is preliminary research and further studies are required, especially as biodiversity is often closely linked to survey sites and ecotopes.1. Generally speaking, flower-visiting spiders focuses on the herbivorous behavior (nectivory or pollinivory) and their effects on host flowers (tritrophic interactions with flower-visiting insect). 2. The authenticity of flower-visiting spiders collected by active sampling was higher than passive trapping; juvenile spider visitors were significantly more common than adults. 3. Our study is the first to propose the term ‘flower-visiting spiders’, and further research is needed.

Chemical signals are important to all living organisms, especially to plants which don’t have vision or mobility. Plant volatiles participate in the competition between plants, herbivores and predators; meanwhile, the tritrophic interactions promote the evolution of plant volatiles. To our knowledge, spiders are common predators in the interactions, but only few studies have begun to investigate what floral volatiles do to spiders. Here, we used Ebrechtella tricuspidata and their common host flowers (tulips) found in our previous field survey to determine the volatile scent preference of spiders for host flowers. Eight main floral volatile scent compounds of tulips were picked out and tested in dual choice behavioural assays. E. tricuspidata showed a significant avoidance of α-pinene and no preference for limonene, β-ocimene, linalool, benzaldehyde, benzyl-alcohol, methyl-salicylate, or 2-phenylethanol. The results illustrate that facultative flower visitor E. tricuspidata is not popular among some kinds of tulip. This probably represents a typical response: facultative flower visitors cannot tolerate all defensive floral volatile scents. Compared with previous studies, we detect that the same floral volatile scent compound has different effects on different spiders, indicating that floral volatile scent compounds attract and reject spiders with specificity and they evolved for different functions and targets. However, this is a preliminary study on the effects of chemical signals on plant-spider interactions, both chemical and physical signals should be considered in future research.

Many studies have explored the effects of plastic particles on aquatic organisms. To date, however, few studies have reported on the effects of plastic particles on terrestrial invertebrates. Here, Culex quinquefasciatus (southern house mosquito, prey) and Pardosa pseudoannulata (wolf spider, predator) were used to explore the transmission of nanoplastics (NPs) from aquatic to terrestrial invertebrates and to verify the effects of NPs in prey on predators. Mosquito larvae were exposed to 0, 200, and 1000 NPs mL

Hylyphantes graminicola is a resident spider species found in maize and cotton fields and is an important biological control agent of various pests. Previous studies have demonstrated that stress from elevated CO2 and Wolbachia infection can strongly affect spider species. Thus, based on CO2 levels (400 ppm, current atmospheric CO2 concentration and 800 ppm, high CO2 concentration) and Wolbachia status (Wolbachia-infected, W+ and Wolbachia-uninfected, W- ), we divided H. graminicola individuals into four treatment groups: W- 400 ppm, W- 800 ppm, W+ 400 ppm, and W+ 800 ppm. To investigate the effects of elevated CO2 levels (W- 400 vs W- 800), Wolbachia infection (W- 400 vs W+ 400), and the interactions between these two factors (W- 400 vs W+ 800), high-throughput transcriptome sequencing was employed to characterize the de novo transcriptome of the spiders and identify stress-related differentially expressed genes (DEGs). De novo assembly of complementary DNA sequences generated 86 688 unigenes, 23 938 of which were annotated in public databases. A total of 84, 21, and 157 DEGs were found among W- 400 vs W- 800, W- 400 vs W+ 400, and W- 400 vs W+ 800, respectively. Functional enrichment analysis revealed that metabolic processes, signaling, and catalytic activity were significantly affected by elevated CO2 levels and Wolbachia infection. Our findings suggest that the impact of elevated CO2 levels and Wolbachia infection on the H. graminicola transcriptome was, to a large extent, on genes involved in metabolic processes. This study is the first description of transcriptome changes in response to elevated CO2 levels and Wolbachia infection in spiders.

Microorganisms in insect guts have been recognized as having a great impact on their hosts' nutrition, health, and behavior. Spiders are important natural enemies of pests, and the composition of the gut microbiota of spiders remains unclear. Will the bacterial taxa in spiders be same as the bacterial taxa in insects, and what are the potential functions of the gut bacteria in spiders? To gain insight into the composition of the gut bacteria in spiders and their potential function, we collected three spider species, Pardosa laura, Pardosa astrigera, and Nurscia albofasciata, in the field, and high‐throughput sequencing of the 16S rRNA V3 and V4 regions was used to investigate the diversity of gut microbiota across the three spider species. A total of 23 phyla and 150 families were identified in these three spider species. The dominant bacterial phylum across all samples was Proteobacteria. Burkholderia, Ralstonia, Ochrobactrum, Providencia, Acinetobacter, Proteus, and Rhodoplanes were the dominant genera in the guts of the three spider species. The relative abundances of Wolbachia and Rickettsiella detected in N. albofasciata were significantly higher than those in the other two spider species. The relative abundance of Thermus, Amycolatopsis, Lactococcus, Acinetobacter Microbacterium, and Koribacter detected in spider gut was different among the three spider species. Biomolecular interaction networks indicated that the microbiota in the guts had complex interactions. The results of this study also suggested that at the genus level, some of the gut bacteria taxa in the three spider species were the same as the bacteria in insect guts.

Bacterial endosymbionts, such as Wolbachia, are widespread in arthropods, with vertical transmission (from mother to offspring) deemed to be the dominant route. Although horizontal transmission of Wolbachia is considered possible as a conveyance route for infections to be introduced to new individuals, evidence for such transmission remains limited. Thus, to elucidate the horizontal transmission of Wolbachia among hosts, we conducted parasitism, predation, cannibalism, and mating experiments in small spider species Hylyphantes graminicola. While no direct evidence was found for horizontal transmission via parasitism or predation, cannibalism and mating behavior were shown to be potential Wolbachia transmission pathways, with copulatory transmission identified from Wolbachia-positive females to Wolbachia-negative males (and vice versa). Based on these findings, our study suggests that horizontal transmission of Wolbachia is more likely to occur within species than between species.

Characterizing protein–protein interactions (PPIs) is an effective method to help explore protein function. Here, through integrating a newly identified split human Rhinovirus 3 C (HRV 3 C) protease, super-folder GFP (sfGFP), and ClpXP-SsrA protein degradation machinery, we developed a fluorescence-assisted single-cell methodology (split protease-E. coli ClpXP (SPEC)) to explore protein–protein interactions for both eukaryotic and prokaryotic species in E. coli cells. We firstly identified a highly efficient split HRV 3 C protease with high re-assembly ability and then incorporated it into the SPEC method. The SPEC method could convert the cellular protein-protein interaction to quantitative fluorescence signals through a split HRV 3 C protease-mediated proteolytic reaction with high efficiency and broad temperature adaptability. Using SPEC method, we explored the interactions among effectors of representative type I-E and I-F CRISPR/Cas complexes, which combining with subsequent studies of Cas3 mutations conferred further understanding of the functions and structures of CRISPR/Cas complexes., Wang et al. developed a fluorescence-assisted single-cell methodology (split protease-E. coli ClpXP (SPEC)) to characterise protein-protein interactions for both eukaryotic and prokaryotic species in E. coli cells. Their method can quantify these interactions with high sensitivity, easy manipulation, and broad temperature adaptability

Elevated CO

Using specific primers, we measured the infection status of Wolbachia, WO, Cardinium, and Spiroplasma in Chilo suppressalis (Walker) (Lepidoptera: Crambidae) collected from 14 locations in China, and conducted phylogenetic analysis of the three endosymbionts. The results showed that eight geographical populations of C. suppressalis were infected by Wolbachia. The infection rates of Wolbachia in four geographical populations had no significant difference between 2011 and 2012 (P>0.05). There were five geographical populations infected with Cardinium and seven geographical populations infected with Spiroplasma. Seven populations of C. suppressalis carrying Wolbachia were also infected with WO. The maximum parsimony tree shows that Wolbachia from C. suppressalis, Pieris rapae, Coliaserate poliographus and Tetranychus urticae fall into the same clade. Cardinium from C. suppressalis joins together with Cardinium from Oppiella nova. Spiroplasma in C. suppressalis has a close relationship with that of Fannia manicata. In conclusion, infections of three endosymbionts showed regional differences, and there were eight geographical populations infected with more than one endosymbiont. Our study found, for the first time, double infection with Wolbachia and Spiroplasma in one sample from Xinzhou. Phylogenetic analysis of the three endosymbionts proved that the endosymbionts can transfer horizontally between different individuals of the same or different hosts.

Spiders, as predators of insects and other invertebrates, are an important part of the natural enemies, and they are recognized as an important biological control agent. Plutellaxylostella (Linnaeus, 1758), the diamondback moth (DBM), is a well-known and destructive insect pest of brassicaceous crops worldwide. Here, we analyzed the functional responses of four spiders (Araneae) – Ebrechtellatricuspidata (Fabricius, 1775) (Thomisidae), Pardosalaura (Karsch, 1879) (Lycosidae), Pardosaastrigera (Koch, 1878) (Lycosidae), and Pardosapseudoannulata (Bösenberg & Strand, 1906) (Lycosidae) – on P.xylostella larvae. We also analyzed intraspecific disturbances in the predation reaction and the intensity of scrambling competition of the spiders to P.xylostella larvae. Our results demonstrated that the functional responses of four spiders of different genera were in line with the Holling II model. Two Lycosidae spiders (P.astrigera and P.pseudoannulata) had the potential to control P.xylostella, and female and male spiders that belonged to the same species had different functional responses to P.xylostella. The functional responses of female predation of P.astrigena, P.laura, and P.pseudoannulata was stronger than the males, but male E.tricuspidatus had stronger functional responses to predation than females. We used the Hassell model to describe the intraspecific disturbance experiments of four spiders. There were intraspecific disturbances in the predation reactions of spiders, indicating that the predation ratio of spiders decreased in relation to the increase of its density, and with the increase of spider density, the intensity of scrambling competition of the spider increased.

Most spiders are natural enemies of pests, and it is beneficial for the biological control of pests to learn the relationships between symbionts and their spider hosts. Research on the bacterial communities of insects has been conducted recently, but only a few studies have addressed the bacterial communities of spiders. To obtain a complete overview of the microbial communities of spiders, we examined eight species of spider (Pirata subpiraticus, Agelena difficilis, Artema atlanta, Nurscia albofasciata, Agelena labyrinthica, Ummeliata insecticeps, Dictis striatipes, and Hylyphantes graminicola) with high‐throughput sequencing based on the V3 and V4 regions of the 16S rRNA gene. The bacterial communities of the spider samples were dominated by five types of endosymbionts, Wolbachia, Cardinium, Rickettsia, Spiroplasma, and Rickettsiella. The dominant OTUs (operational taxonomic units) from each of the five endosymbionts were analyzed, and the results showed that different spider species were usually dominated by special OTUs. In addition to endosymbionts, Pseudomonas, Sphingomonas, Acinetobacter, Novosphingobium, Aquabacterium, Methylobacterium, Brevundimonas, Rhizobium, Bradyrhizobium, Citrobacter, Arthrobacter, Pseudonocardia, Microbacterium, Lactobacillus, and Lactococcus were detected in spider samples in our study. Moreover, the abundance of Sphingomonas, Methylobacterium, Brevundimonas, and Rhizobium in the spider D. striatipes was significantly higher (p

Elevated CO2 concentrations can change the multi-level nutritional relationship of the ecosystem through the cascading effect of the food chain. To date, few studies have investigated the effects of elevated CO2 concentration on the Araneae species through the tritrophic system. Hylyphantes graminicola (Araneae: Linyphiidae) is distributed widely in Asia and is a dominant predator in cotton fields. This study investigated chemical components in the food chain of cotton (Gossypium hirsutum)—cotton aphid (Aphis gossypii)—predator (H. graminicola) and compared the development, reproduction, and predation of H. graminicola under ambient (400 ppm) and elevated concentration of CO2 (800 ppm). The results showed that the elevated CO2 concentration increased the chemicals of cotton and cotton aphid, but it did not affect the nutrients, development, reproduction, and predation of the spider. However, the survival rate of the spider was significantly decreased in elevated CO2. The results will further our understanding of the role of natural enemies in an environment with elevated CO2 concentration.

The ladybeetle Propylea japonica is a widely distributed natural enemy in many agricultural systems. P. japonica is often used as a test organism for safety assessments of transgenic Bacillus thuringiensis crops. Plant varieties expressing the Vip3Aa insecticidal protein are not currently commercially available in China. In this study, protease inhibitor E-64 was used as a positive control to examine the responses of P. japonica larvae to a high concentration of Vip3Aa proteins. Larvae that were fed E-64 had increased mortality and prolonged developmental period, but these parameters were unaffected when larvae were fed Vip3Aa. The epithelial cells of midguts were intact and closely connected with the basal membrane when larvae were fed Vip3Aa, but the epithelial cells degenerated in the E-64 treatment. The activities of antioxidative enzymes and expression levels of detoxification-related genes in P. japonica larvae were not altered after exposure to Vip3Aa; however, these biochemical and molecular parameters were significantly changed in the E-64 treatment. The results demonstrate that Vip3Aa protein is not harmful to the predator P. japonica.

Temperature is a critical abiotic factor that causes physiological changes in arthropods. However, little is known about the effect of heat stress on the antioxidant responses of Araneae species. Hylyphantes graminicola is a dominant predator in many cropping systems in China. In the present study, the effect of short-term heat stress (36, 38, 40 or 42 °C) on the reactive oxygen species (ROS) levels, the activities of antioxidant enzymes (superoxide dismutase [SOD], catalase [CAT], peroxidases [POD] and glutathione-S-transferases GST]), total antioxidant capacity (TAC), malondialdehyde (MDA) concentrations and survival of H. graminicola spiderlings and adults were investigated. The results showed that H. graminicola adults had a significantly higher survival rate compared to spiderlings at 40 °C. The heat stress increased ROS contents in H. graminicola. The SOD, CAT, POD and GST activities increased in spiderlings and adults under heat stress. These data suggest a defensive function for these enzymes in alleviating oxidative damage. Specifically, SOD plays a key role in reducing the high level of superoxide radicals in spiderlings and adults. Moreover, the POD and CAT capabilities for scavenging H2O2 in spiderlings were similar, and CAT may play a more important role than POD in scavenging H2O2 in adults at 42 °C. The spiderling TAC increased significantly at 40 and 42 °C, and the adult TAC was stable at 36–40 °C but decreased at 42 °C. These data suggest that TAC was insufficient in H. graminicola adults under more severe stress conditions. These results further our understanding of the physiological response of Araneae species exposed to heat stress.

The effects of elevated CO2 concentration on the growth and development of the funnel-web weaving spider Agelena labyrinthica (Clerck) (Araneae: Agelenidae) were studied in climate chambers with low (370 μl l−1) or high (750 μl l−1) CO2 concentration. Seventh-instar A. labyrinthica cultured under each of these CO2 concentrations were randomly selected to determine nutrient composition (total protein, total amino acid, and free fatty acid) and digestive or detoxification enzymes activity (peroxidase, amylase, and superoxide dismutase) using test kits. When reared under high CO2 concentration, total development of A. labyrinthica was significantly faster. Carapace length and width and body weight did not differ between CO2 concentrations, nor the levels of protein and total amino acids in seventh-instar A. labyrinthica. However, free fatty acid levels were significantly lower under high CO2 concentration. Specific activities of peroxidase and superoxide dismutase in seventh-instar A. labyrinthica did not differ between CO2 concentrations. The specific activity of amylase under high CO2 concentration was higher than that of the low CO2 group. The effects of elevated CO2 on A. labyrinthica varied from those on the wandering spider Pardosa astrigera L Koch, as found in an earlier study. Apparently, elevated CO2 has a species-specific impact on spiders.

In nature, male wolf spiders may encounter silk of conspecific females, males, and/or both. It is generally accepted that males are able to discriminate maturity, sex, and mating status through silk-mediated chemical information of females. However, whether males can discriminate silk by chemical cues among virgin females of different post-maturation ages and between female silk with and without previously being occupied by other males remains largely unexplored. Given that female sexual receptivity increases in post-maturation, males are predicted to prefer old over young virgin females, and are able to discriminate silk of females which have and have not been previously being occupied by other males. In this study, we used the monandrous wolf spider, Pardosa astrigera, to test this prediction experimentally. When males were exposed to the silk of virgin females of different post-maturation ages (1-, 3-, and 6-day-old females), we found no significant differences in male courtship latency and courtship intensity to female silk across the different ages. However, males displayed significantly higher courtship intensity to the silk of females without being previously occupied by other males than to the silk of females with being previously occupied by other males. When males were directly exposed to virgin females, significantly higher courtship intensity was detected in males in response to 6-day-old females compared with 1-day-old ones. Thus, our results suggest that P. astrigera males can discriminate females using chemical cues from silk that has been previously occupied by other males and to older females with higher sexual receptivity, whereas they invest less to silk cues from virgin females regardless of their post-maturation ages.

Phage WO is a bacteriophage found in Wolbachia. Herein, we represent the first phylogenetic study of WOs that infect spiders (Araneae). Seven species of spiders (Araneus alternidens, Nephila clavata, Hylyphantes graminicola, Prosoponoides sinensis, Pholcus crypticolens, Coleosoma octomaculatum, and Nurscia albofasciata) from six families were infected by Wolbachia and WO, followed by comprehensive sequence analysis. Interestingly, WO could be only detected Wolbachia-infected spiders. The relative infection rates of those seven species of spiders were 75, 100, 88.9, 100, 62.5, 72.7, and 100 %, respectively. Our results indicated that both Wolbachia and WO were found in three different body parts of N. clavata, and WO could be passed to the next generation of H. graminicola by vertical transmission. There were three different sequences for WO infected in A. alternidens and two different WO sequences from C. octomaculatum. Only one sequence of WO was found for the other five species of spiders. The discovered sequence of WO ranged from 239 to 311 bp. Phylogenetic tree was generated using maximum likelihood (ML) based on the orf7 gene sequences. According to the phylogenetic tree, WOs in N. clavata and H. graminicola were clustered in the same group. WOs from A. alternidens (WAlt1) and C. octomaculatum (WOct2) were closely related to another clade, whereas WO in P. sinensis was classified as a sole cluster.

The effects of elevated CO2 concentration on spiders were studied using third-instar spiderlings of the wolf spider Pardosa astrigera L. Koch 1878 in CO2 climate chambers with two different concentrations of CO2 (low, 370 ppm and high, 750 ppm). The food intake and total developmental period of spiderlings reared at high CO2 concentration increased significantly, and the body length and weight of adult spiders decreased compared to those in the low-CO2 group. The oviposition rate of female P. astrigera and the hatching rate of eggs did not differ between the high- and low-CO2 groups, but the number of egg sacs and the total number of eggs produced by females from the high-CO2 group decreased. These results suggest that elevated CO2 concentrations are harmful to the growth, development, and reproduction of P. astrigera.

Symbiosis, the close association of unrelated organisms, has been pivotal in biological diversification. In the obligate symbioses found in many insect hosts, organisms that were once independent are permanently and intimately associated, resulting in expanded ecological capabilities. The primary model for this kind of symbiosis is the association between the bacterium Buchnera and the pea aphid (Acyrthosiphon pisum). A longstanding obstacle to efforts to illuminate genetic changes underlying obligate symbioses has been the inability to experimentally disrupt and reconstitute symbiont–host partnerships. Our experiments show that Buchnera can be experimentally transferred between aphid matrilines and, furthermore, that Buchnera replacement has a massive effect on host fitness. Using a recipient pea aphid matriline containing Buchnera that are heat sensitive because of an allele eliminating the heat shock response of a small chaperone, we reduced native Buchnera through heat exposure and introduced a genetically distinct Buchnera from another matriline, achieving complete replacement and stable inheritance. This transfer disrupted 100 million years (∼1 billion generations) of continuous maternal transmission of Buchnera in its host aphids. Furthermore, aphids with the Buchnera replacement enjoyed a dramatic increase in heat tolerance, directly demonstrating a strong effect of symbiont genotype on host ecology.

Background Tea is one of the most economically important crops in China. However, the tea geometrid (Ectropis obliqua), a serious leaf-feeding pest, causes significant damage to tea crops and reduces tea yield and quality. Spiders are the most dominant predatory enemies in the tea plantation ecosystem, which makes them potentially useful biological control agents of E. obliqua. These highlight the need for alternative pest control measures. Our previous studies have shown that tea saponin (TS) exerts insecticidal activity against lepidopteran pests. Here, we investigate whether TS represents a potentially new alternative insecticide with no harm to spiders. Methods We investigated laboratory bioactivities and the field control properties of TS solution against E. obliqua. (i) A leaf-dip bioassay was used to evaluate the toxicity of TS to 3rd-instar E. obliqua larvae and effects of TS on the activities of enzymes glutathione-S-transferase (GST), acetylcholinesterase (AChE), carboxylesterase (CES) and peroxidase (POD) of 3rd-instar E. obliqua larvae in the laboratory. (ii) Topical application was used to measure the toxicity of 30% TS (w/v) and two chemical insecticides (10% bifenthrin EC and 50% diafenthiuron SC) to two species of spider, Ebrechtella tricuspidata and Evarcha albaria. (iii) Field trials were used to investigate the controlling efficacy of 30% TS against E. obliqua larvae and to classify the effect of TS to spiders in the tea plantation. Results The toxicity of TS to 3rd-instar E. obliqua larvae occurred in a dose-dependent manner and the LC50 was 164.32 mg/mL. Activities of the detoxifying-related enzymes, GST and POD, increased in 3rd-instar E. obliqua larvae, whereas AChE and CES were inhibited with time by treatment with TS. Mortalities of E. tricuspidata and E. albaria after 48 h with 30% TS treatment (16.67% and 20%, respectively) were significantly lower than those with 10% bifenthrin EC (80% and 73.33%, respectively) and 50% diafenthiuron EC (43.33% and 36.67%, respectively). The highest controlling efficacy of 30% TS was 77.02% at 5 d after treatment, which showed no difference to 10% bifenthrin EC or 50% diafenthiuron SC. 30% TS was placed in the class N (harmless or slightly harmful) of IOBC (International Organization of Biological Control) categories for natural enemies, namely spiders. Conclusions Our results indicate that TS is a botanical insecticide that has a good controlling efficacy in E. obliqua larvae, which suggests it has promise as application in the integrated pest management (IPM) envisaged for tea crops.

Arthropods are associated with various microorganisms which confer benefits to their hosts. Recently, research has been conducted on bacterial communities of insects to provide an insight into the potential interactions of the symbiotic bacteria and their hosts. Spiders are interesting to study as they are perceived to be natural enemies of pests. The effect of endosymbionts on spiders has been reported, but little is known about the overall bacterial communities present in spiders. Here, we report on the characterization of bacterial communities present in the whole body of the spider Marpiss magister using Illumina sequencing of 16S rRNA amplicons. Our study shows that the most abundant phyla of bacteria included Proteobacteria, Tenericutes, Bacteroidetes and Actinobacteria. At the genus level, the most abundant genera included Rickettsia, Wolbachia, Spiroplasma, and Cardinium. Besides these dominant endosymbionts, our study also showed the existence of bacteria in the genera Arthrobacter, Novosphingobium, Acinetobacter, Pseudomonas, Aquabacterium and Sphingomonas at an abundance ranging from 0.65 to 0.84%, and the existence of bacterial in genera Lactobacillus, Sphingobium, Methylobacterium, Bradyrhizobium, Propionibacterium, Brevundimonas, Achromobacter, Microbacterium, Corynebacterium, and Flavobacterium at a slightly lower abundance ranging from 0.1 to 0.5%. Therefore, our finding indicates that endosymbionts are not the only microbiota present in the spider M. magister, and other bacterial taxa also exist in its bacterial community.

Arthropods are associated with various microorganisms which confer benefits to their hosts. Recently, research has been conducted on bacterial communities of insects to provide an insight into the potential interactions of the symbiotic bacteria and their hosts. Spiders are interesting to study as they are perceived to be natural enemies of pests. The effect of endosymbionts on spiders has been reported, but little is known about the overall bacterial communities present in spiders. Here, we report on the characterization of bacterial communities present in the whole body of the spider Marpiss magister using Illumina sequencing of 16S rRNA amplicons. Our study shows that the most abundant phyla of bacteria included Proteobacteria, Tenericutes, Bacteroidetes and Actinobacteria. At the genus level, the most abundant genera included Rickettsia, Wolbachia, Spiroplasma, and Cardinium. Besides these dominant endosymbionts, our study also showed the existence of bacteria in the genera Arthrobacter, Novosphingobium, Acinetobacter, Pseudomonas, Aquabacterium and Sphingomonas at an abundance ranging from 0.65 to 0.84%, and the existence of bacterial in genera Lactobacillus, Sphingobium, Methylobacterium, Bradyrhizobium, Propionibacterium, Brevundimonas, Achromobacter, Microbacterium, Corynebacterium, and Flavobacterium at a slightly lower abundance ranging from 0.1 to 0.5%. Therefore, our finding indicates that endosymbionts are not the only microbiota present in the spider M. magister, and other bacterial taxa also exist in its bacterial community.

According to sexual selection theory, age affects the preference of mate choice, and this preference ultimately influences the fecundity of the female. Pardosa pseudoannulata (Araneae: Lycosidae) is a valued predator in many cropping systems. By determining oviposition rate, egg hatching rate, and also the number and carapace width of the 2nd instar spiderlings of the F1 generation, we explored the effects of age on fecundity of the female spider. There were no significant effects of age on courtship duration, sexual cannibalism rate, mating rate, oviposition rate, egg hatching rate, or the number and carapace width of 2nd instar spiderings of P. pseudoannulata. However, age had a significant effect on courtship latency, courtship intensity, and mating duration of the spider. Courtship latency decreased significantly with an increase in the age of the male, and courtship intensity of the low-age male increased with increasing female age. Increasing age of male and female spiders was associated with significantly prolonged mating duration. The results indicated that low-age male spiders were more inclined to mate with high-age females, and age had no significant effect on sexual cannibalism rate or the fecundity of the female.

Associations between endosymbiotic bacteria and their hosts are widespread in nature and have been demonstrated extensively; however, only a few studies have examined how facultative symbionts affect host nutrition and metabolism. To gain insight into the associations between facultative symbionts and host nutrition and metabolic activity, we detected endosymbiotic infection in a small spider species, Hylyphantes graminicola, and established two infectious strains, i.e., W−C+ (Wolbachia negative, Cardinium positive) and W+C+ (Wolbachia positive, Cardinium positive). We then determined the content of fat and free amino acids in W−C+ and W+C+ spiders, respectively. We also detected the transcriptome of H. graminicola and the expression of genes involved in fat and amino acid metabolism at different host ages. Results showed that fat content in W+C+ spiders was higher than that in W−C+ spiders, and free amino acid content was higher in W+C+ males than W−C+ males, with no difference observed in females. Transcriptome analysis identified 144 (W−C+ vs W+C+) differentially expressed genes (DEGs). Moreover, the expression of five genes involved in fat and amino acid metabolism were significantly up-regulated in the third, fourth, and fifth instar stages in W+C+ spiders. This study indicated that Wolbachia and Cardinium co-infection had a pivotal effect on fat and amino acid synthesis in hosts. Moreover, our results provide strong evidence explaining the long-term coexistence of hosts and endosymbionts.

Pardosa pseudoannulata is one of the most common wandering spiders in agricultural fields and a potentially good bioindicator for heavy metal contamination. However, little is known about the mechanism by which spiders respond to heavy metals at the molecular level. In this study, high-throughput transcriptome sequencing was employed to characterize the de novo transcriptome of the spiders and to identify differentially expressed genes (DEGs) after cadmium exposure. We obtained 60,489 assembled unigenes, 18,773 of which were annotated in the public databases. Ultimately, 3450 cDNA simple sequence repeats were identified and validated as potential molecular markers in the unigenes. A total of 2939, 2491 and 3759 DEGs were detected among the three libraries of two Cd-treated groups and the control. Functional enrichment analysis revealed that metabolism processes and digestive system function were predominately enriched in response to Cd stress. At the cellular and molecular levels, significantly enriched pathways in lysosomes and phagosomes as well as replication, recombination and repair demonstrated that oxidative damage resulted from Cd exposure. Based on the selected DEGs, certain critical genes involved in defence and detoxification were analysed. These results may elucidate the molecular mechanism underlying spiders' responses to heavy metal stress.

Pardosa pseudoannulata is one of the most common wandering spiders in agricultural fields and a potentially good bioindicator for heavy metal contamination. However, little is known about the mechanism by which spiders respond to heavy metals at the molecular level. In this study, high-throughput transcriptome sequencing was employed to characterize the de novo transcriptome of the spiders and to identify differentially expressed genes (DEGs) after cadmium exposure. We obtained 60,489 assembled unigenes, 18,773 of which were annotated in the public databases. Ultimately, 3450 cDNA simple sequence repeats were identified and validated as potential molecular markers in the unigenes. A total of 2939, 2491 and 3759 DEGs were detected among the three libraries of two Cd-treated groups and the control. Functional enrichment analysis revealed that metabolism processes and digestive system function were predominately enriched in response to Cd stress. At the cellular and molecular levels, significantly enriched pathways in lysosomes and phagosomes as well as replication, recombination and repair demonstrated that oxidative damage resulted from Cd exposure. Based on the selected DEGs, certain critical genes involved in defence and detoxification were analysed. These results may elucidate the molecular mechanism underlying spiders' responses to heavy metal stress.

Five cDNA fragments (I–V) representing five P450 isoforms were cloned and a novel P450 cDNA was isolated from Ummeliata insecticeps (Bosenberg & Strand), using the reverse transcription-polymerase chain reaction and rapid amplification of cDNA ends. The full cDNA is 2708 bp in length and contains an open reading frame (ORF) encoding 521 amino acid residues. The putative protein contains the conserved structural and functional regions characteristic of all P450 enzymes. The P450 shows the highest percentage of amino acid identities (47%) with CYP4V18 from Litopenaeus vannamei (Boone, 1931). Phylogenetic analysis showed that the P450 was grouped in the lineage containing some of the CYP4 family members. The GenBank accession number is FN773088. This is the first report of P450 genes cloned in spiders (Araneae).

Members of the spider families Thomisidae, Salticidae, Miturgidae, Anyphaenidae, and Corinnidae have been observed on floral and extrafloral nectaries of plants, where they are presumably feeding on nectar. Nectar feeding is a phenomenon that has long intrigued biologists. However, few studies have focused on the effect of nectar on biological characteristics of these spiders. The aim of this study was to determine whether crab spiders, Ebrechtella tricuspidata (Fabricius) (Araneae: Thomisidae), show an active preference for honey solution (a simulated nectar source) and to test the effect of honey solution on the survival and development time of spiderlings and the fecundity of adult E. tricuspidata. The results showed that the number of females feeding on the 10, 20, and 30% honey solution was significantly higher than those feeding on water alone, and there was no difference in the number of females feeding on the three concentrations of honey solution. Significantly more E. tricuspidata preferred feeding on the 20% honey solution, and they spent significantly more time feeding on 20% honey solution than on water, regardless of whether they were males or females, adults or spiderlings. Ingestion of honey solution significantly increased the survival and shortened the development time of E. tricuspidata. Female spiders that fed on honey solution had a shorter pre-oviposition period and laid more eggs than those given only water. Our results suggest that nectar could be a high-quality supplementary food to maintain normal growth and metabolism in spiderlings and adult female spiders in nature.

The Wolbachia endosymbiont of spiders has not been extensively examined. In order to investigate the distribution, evolutionary history, and reproductive phenotype of Wolbachia in spiders in China, we tested 11 geographic populations of Hylyphantes graminicola. Wolbachia infection has been detected in each population. 10 Wolbachia strains have been characterized by multilocus sequence typing (MLST). Phylogenetic analyses indicated that eight Wolbachia strains in H. graminicola belonged to supergroup B, and two belonged to supergroup A. No correlation existed between Wolbachia diversity and host's geographic distance. The significant correlation was observed between pairwise distance of H. graminicola COI and genetic divergence of associated Wolbachia strains. We also found that Wolbachia infection frequencies in hosts varied over geographic space.

The objective of the present study was to assess the ecotoxicological responses of Pardosa pseudoannulata to a common environmental pollutant, cadmium. Third-instar spiderlings and adult spiders were exposed to sublethal concentrations of CdCl2 solution in their drinking water. The Cd content in P. pseudoannulata adults increased significantly with the number of days of exposure to a 0.2 mM CdCl2 solution, when exposed to 2 mM CdCl2 solution, the Cd content in the spiders increased sharply in the first two (male) or three (female) weeks, and then no significant changes were recorded following with the next three (male) or two (female) weeks exposure. Exposure of spiders to Cd contaminated drinking water resulted in reduced body mass, delayed development, fewer eggs and increased mortality. Significantly higher activities of superoxide dismutase, catalase and glutathione-s-transferase were recorded in the spiders after 7 day exposure to 0.2 mM CdCl2 solution. However, longer-term exposures or increased Cd concentrations did not result in significantly higher antioxidant enzyme activity relative to control treatment.

This study investigated the bioaccumulation and effects of mercury (Hg) in funnel-web spiders, Agelena labyrinthica, following exposure to sublethal concentrations of Hg(NO3)2 in their drinking water. The results showed that the Hg content in adult A. labyrinthica increased rapidly with the number of days exposed to the Hg(NO3)2 solution, and the mortality of adult spiders within 30 days increased with increased concentrations of Hg(NO3)2 in the drinking water. The total developmental duration of A. labyrinthica exposed to Hg(NO3)2 was significantly longer than in the control spiders, but there were no significant differences in the total developmental duration of spiders among the three treatment groups (exposed to 10, 20 and 50 mg/L Hg(NO3)2 solution). We also compared the web-weaving of the control and treated spiders, and found no significant differences in shape, structure, color, or size of the webs between the control and treated spiders; however, there was a significant difference in web placement between the treatment and control groups. The spiders in the control group appeared to have an episodic-like memory, choosing to weave their five webs in the same corner in the five time periods allowed.