The effects of geographic origin and antibiotic treatment on the gut symbiotic communities of Bactrocera oleae populations

The olive fruit fly, Bactrocera oleae (Rossi) (Diptera: Tephritidae), is the major insect pest of olive orchards (Olea europaea L.), causing extensive damages on cultivated olive crops worldwide. Due to its economic importance, it has been the target species for a variety of population control approaches including the sterile insect technique (SIT). However, the inefficiency of the current mass‐rearing techniques impedes the successful application of area‐wide integrated pest management programs with an SIT component. It has been shown that insect mass rearing and quality of sterile insects can be improved by the manipulation of the insect gut microbiota and probiotic applications. In order to exploit the gut bacteria, it is important to investigate the structure of the gut microbial community. In the current study, we characterized the gut bacterial profile of two wild olive fruit fly populations introduced in laboratory conditions using next generation sequencing of two regions of the 16S rRNA gene. We compared the microbiota profiles regarding the geographic origin of the samples. Additionally, we investigated potential changes in the gut bacteria community before and after the first exposure of the wild adult flies to artificial adult diet with and without antibiotics. Various genera – such as Erwinia, Providencia, Enterobacter, and Klebsiella – were detected for the first time in B. oleae. The most dominant species was Candidatus Erwinia dacicola Capuzzo et al. and it was not affected by the antibiotics in the artificial adult diet used in the first generation of laboratory rearing. Geographic origin affected the overall structure of the gut community of the olive fruit fly, but antibiotic treatment in the first generation did not significantly alter the gut microbiota community.     

Exploitation of the Medfly Gut Microbiota for the Enhancement of Sterile Insect Technique: Use of Enterobacter sp. in Larval Diet-Based Probiotic Applications

The Mediterranean fruit fly (medfly), Ceratitis capitata, is a pest of worldwide substantial economic importance, as well as a Tephritidae model for sterile insect technique (SIT) applications. The latter is partially due to the development and utilization of genetic sexing strains (GSS) for this species, such as the Vienna 8 strain, which is currently used in mass rearing facilities worldwide. Improving the performance of such a strain both in mass rearing facilities and in the field could significantly enhance the efficacy of SIT and reduce operational costs. Recent studies have suggested that the manipulation of gut symbionts can have a significant positive effect on the overall fitness of insect strains. We used culture-based approaches to isolate and characterize gut-associated bacterial species of the Vienna 8 strain under mass rearing conditions. We also exploited one of the isolated bacterial species, Enterobacter sp., as dietary supplement (probiotic) to the larval diet, and we assessed its effects on fitness parameters under the standard operating procedures used in SIT operational programs. Probiotic application of Enterobacter sp. resulted in improvement of both pupal and adult productivity, as well as reduced rearing duration, particularly for males, without affecting pupal weight, sex ratio, male mating competitiveness, flight ability and longevity under starvation.     

Molecular phylogenetic profiling of gut‐associated bacteria in larvae and adults of flesh flies

Flesh flies of the genus Sarcophaga (Diptera: Sarcophagidae) are carrion‐breeding, necrophagous insects important in medical and veterinary entomology as potential transmitters of pathogens to humans and animals. Our aim was to analyse the diversity of gut‐associated bacteria in wild‐caught larvae and adult flesh flies using culture‐dependent and culture‐independent methods. Analysis of 16S rRNA gene sequences from cultured isolates and clone libraries revealed bacteria affiliated to Proteobacteria, Actinobacteria, Firmicutes and Bacteroidetes in the guts of larval and adult flesh flies. Bacteria cultured from larval and adult flesh fly guts belonged to the genera Acinetobacter, Bacillus, Budvicia, Citrobacter, Dermacoccus, Enterococcus, Ignatzschineria, Lysinibacillus, Myroides, Pasteurella, Proteus, Providencia and Staphylococcus. Phylogenetic analysis showed clone sequences of the genera Aeromonas, Bacillus, Bradyrhizobium, Citrobacter, Clostridium, Corynebacterium, Ignatzschineria, Klebsiella, Pantoea, Propionibacterium, Proteus, Providencia, Serratia, Sporosarcina, Weissella and Wohlfahrtiimonas. Species of clinically significant genera such as Ignatzschineria and Wohlfahrtiimonas spp. were detected in both larvae and adult flesh flies. Sequence analysis of 16S rRNA gene libraries supported culture‐based results and revealed the presence of additional bacterial taxa. This study determined the diversity of gut microbiota in flesh flies, which will bolster the ability to assess microbiological risk associated with the presence of these flies. The present data thereby establish a platform for a much larger study.     

Wild bacterial probiotics fed to larvae of mass‐reared Queensland fruit fly [Bactrocera tryoni (Froggatt)] do not impact long‐term survival,mate selection,or locomotor activity

Queensland fruit fly [Bactrocera tryoni (Froggatt), Diptera, Tephritidae] is the most devastating insect pest impacting Australian horticulture. The Sterile Insect Technique (SIT) is an important component of tephritid pest management programs. However, mass‐rearing and irradiation (to render insects sterile) may reduce the fitness and performance of the insect, including the ability of sterile males to successfully compete for wild females. Manipulation of the gut microbiome, including the supplementation with bacterial probiotics shows promise for enhancing the quality of mass‐reared sterile flies, however there are fewer published studies targeting the larval stage. In this study, we supplemented the larval stage of mass‐reared B. tryoni with bacterial probiotics. We tested several individual bacteria that had been previously isolated and characterized from the gut of wild B. tryoni larvae including Asaia sp., Enterobacter sp., Lactobacillus sp., Leuconostoc sp. We also tested a consortium of all four of these bacterial isolates. The fitness parameters tested included adult survival in field cages, laboratory mate selection of bacteria supplemented males by bacteria nonsupplemented females, and laboratory locomotor activity of adult flies. None of the bacterial probiotic treatments in the current study was significantly different to the control for field survival, mate selection or locomotor activity of adult B. tryoni, which agree with some of the other studies regarding bacterial probiotics fed to the larval stage of tephritids. Future work is needed to determine if feeding the same, and/or other probiotics to adults, as opposed to larvae can positively impact survival, mating performance, mating competitiveness and locomotor activity of B. tryoni. The bacterial group(s) and function of bacterial species that increase fitness and competitiveness is also of interest to tephritid mass‐rearing programs.     

Ingestion and digestion of epilithic algae by larval insects in a heavily grazed montane stream

1. Epilithic algae grown on elevated or non-elevated ceramic tiles were exposed (to produce assemblages with different grazing histories) in a heavily grazed, montane stream in New Mexico, U.S.A. to Ameletus nymphs (Ephemeroptera) and Ecclisomyia larvae (Trichoptera) and the algal composition in insect faeces was compared to that on the tiles. Differences in grazing and digestion efficiency between grazers were then assessed and also differences in susceptibility to ingestion and digestibility among common algae. 2. Ordination of tile and faecal samples, using the relative abundance of common algae, revealed that: (i) algal assemblages on elevated vs. non-elevated tiles differed only slightly; (ii) the taxonomic composition of algae in faeces of both caddis and mayflies differed substantially from that on the tiles, indicating low grazing efficiency for some algal taxa; and (iii) the algal composition of faeces produced by caddis larvae and mayflies was similar, indicating little difference in grazing efficiency between them. However, some algal taxa were more susceptible to ingestion by caddisfly larvae when occurring on elevated tiles than on non-elevated tiles, suggesting that previous exposure to caddis grazing influenced assemblage attributes. 3. Although Ameletus and Ecclisomyia differed little in grazing efficiency, the percentage of diatoms that were dead after passage through the gut was greatest in the mayfly treatment, suggesting that mayflies digested diatoms more efficiently than the caddis. Analyses of differences in the condition of chloroplasts within diatoms in tile and faecal samples showed that losses of ’live‘ diatom cells (i.e. those containing full chloroplasts) during gut passage through mayflies equalled the increase, in faeces, of ’dead‘ (empty frustules) cells of all common diatoms. In contrast, some diatoms were digested inefficiently by caddis larvae. 4. Algae on elevated tiles contained a higher proportion of dead diatoms than those on non-elevated tiles, possibly because mayflies visited raised tiles more often and, consequently, ingested and defaecated cells at a higher rate in the absence of caddis larvae. Moreover, diatom taxa differed in the percentage of cells that were dead within tile assemblages, with populations of typically grazer-resistant taxa (e.g. Achnanthidium minutissimum, Planothidium lanceolatum and Cocconeis placentula var. euglypta) containing significantly more dead cells than grazer-susceptible taxa [e.g. small, chain-forming Fragilaria (= Staurosirella)]. This result suggests that a trade-off exists between ingestion vs. digestion resistance of microalgae. Both the ingestion and digestion efficiency of algivorous macroinvertebrates could influence the structure and function of algal assemblages. In heavily grazed systems, where algal cells are probably processed through grazer guts repeatedly, differential resistance to digestion among algae may be particularly important.     

Genome size estimation with quantitative real‐time PCR in two Tephritidae species: Ceratitis capitata and Bactrocera oleae

The medfly Ceratitis capitata and the olive fruit fly Bactrocera oleae belong to the Tephritidae family of Diptera, a family whose members cause severe damages in agriculture worldwide. For such insect pests, the utmost concern is their population control. The sterile insect technique (SIT) has been used in the Tephritidae family with varying degrees of success. Its efficient use usually depends on the development of genetic sexing strains and the release of only male flies. However, such advances are based on modern genetic, molecular and genomic tools. The medfly is clearly the prototype of the family, since such tools have advanced considerably, which has resulted in effective SIT efforts around the world. A whole‐genome sequencing project of this insect is already underway. In contrast, similar tools in the olive fly lag behind, even though the insect is considered a promising candidate for a next SIT target. An accurate estimate of genome size provides a preliminary view of genome complexity and indicates possible difficulties in genome assembly in whole‐genome projects. Taking advantage of a quantitative real‐time PCR approach, we determined the genome size of these two species C. capitata and B. oleae as 591 Mb (CI range: 577–605 Mb) and 322 Mb (CI range: 310–334 Mb) respectively.     

Elicitation of alkaloids in in vitro PLB (protocorm-like body) cultures of Pinellia ternata

The objective of this study was to determine the effect of two endophytic bacterial elicitors (Pseudomonas sp. and Enterobacter sp.) on the production of alkaloids in protocorm-like bodies (PLBs) of Pinellia ternata Breit. Both bacterial strains increased the growth rate of P. ternata PLBs. Pseudomonas sp. promoted the differentiation of the PLBs, whereas Enterobacter sp. inhibited PLB differentiation. The bacterial strains increased guanosine production in PLBs by 9–166%, inosine production by 2–33%, and trigonelline production by 114–1140% compared to the control. For Pseudomonas sp., guanosine and trigonelline production was greater when bacterial extracts were added to the PLB suspension cultures rather than living cells (co-culture treatment). Inosine production was similar in both the bacterial extract and co-culture treatments. For the Enterobacter sp., guanosine, inosine, and trigonelline production tended to be greatest when living cells were added to the PLB suspension cultures rather than bacterial extracts. These results suggest that Pseudomonas sp. and Enterobacter sp. could increase alkaloid yield from P. ternata under field or tissue culture conditions. We also observed that Pseudomonas sp. and Enterobacter sp. produced some of the same alkaloids as their host plants. Additional study needs to be done to determine if these endophytic bacteria could be used to produce alkaloids in the fermentation industry.     

Characteristics of an endophytic pyrene-degrading bacterium of Enterobacter sp. 12J1 from Allium macrostemon Bunge

One pyrene-degrading endophytic bacterium was isolated from plants grown in polycyclic aromatic hydrocarbon-contaminated soils and identified as Enterobacter sp. 12J1 based on the 16S rDNA gene sequence analysis. Heavy metal and antibiotic resistance, degradation of pyrene, solubilization of inorganic phosphate and cell surface hydrophobicity characteristics of the isolate were further characterized. The isolate was also evaluated for promoting plant growth of wheat and maize and pyrene removal from pyrene-amended soil in pot experiments. High-performance liquid chromatograph (HPLC) analysis showed that the degradation rate of pyrene (5 mg l⁻¹) by the endophytic bacterial strain 12J1 was 83.8% under 28 °C for 7 days. The Enterobacter sp. 12J1 could produce indole acetic acid (IAA), siderophore and solubilize inorganic phosphate. The Enterobacter sp. 12J1 also has a cell surface hydrophobicity. In the live bacterial inoculation experiment, an increase in pyrene removal varying from 60% to 107% was observed in the planted soils treated with 100 mg kg⁻¹ of pyrene compared with the unplanted soils. The rate of pyrene removal increased by 43–65% in the live bacterium-inoculated planted soils compared with the dead bacterium-inoculated planted soils. Although there were no significant differences in the total culturable bacterial numbers between live and dead bacterial inoculation, the numbers of pyrene-degrading bacteria were significantly greater in the live bacterium-inoculated planted or unplanted soils. The isolate could colonize the tissue (root and stem) interiors and rhizosphere soils of wheat and maize after root inoculation.     

Removal and transformation of high molecular weight polycyclic aromatic hydrocarbons in water by live and dead microalgae

The removal and transformation of seven high molecular weight polycyclic aromatic hydrocarbons (PAHs), namely benz[a]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, dibenzo[a,h]anthracene, indeno[1,2,3-c,d]pyrene and benzo[g,h,i]perylene, by a freshwater microalga Selenastrum capricornutum under gold and white light irradiation was studied. The two light sources did not result in significant differences in the biodegradation of the selected PAHs in live algal cells, but white light was more effective in promoting photodegradation than was gold light in dead cells. The removal efficiency of seven PAHs, as well as the difference between live and dead microalgal cells, was PAH compound-dependent. Benz[a]anthracene and benzo[a]pyrene were highly transformed in live and dead algal cells, and dead cells displayed greater transformation levels than live cells. Further investigation comparing the transformation of single PAH compound, benzo[a]pyrene, by S. capricornutum and another green microalgal species, Chlorella sp., demonstrated that the transformation in dead cells was similar, indicating the process was algal-species independent. Dead algal cells most likely acted as a photosensitizer and accelerated the photodegradation of PAHs.     

Flow Cytometry Demonstrates Bacteriocin-Induced Injury to Listeria monocytogenes

Flow cytometry was used to study the effect of the bacteriocin leucocin B-TA11a on Listeria (L.) monocytogenes. Mixed proportions of dead and live control populations were analyzed by flow cytometry to determine detection limits of the Dead/Live Baclight Bacterial Viability Kit^TM. High correlations for flow cytometric detection of defined proportions of live or dead cells in mixtures between 10 and 100% of dead (r² = 0.97) or live (r² = 0.99) cells were obtained. However, mixtures containing less than 10% of either live or dead control cells gave correlations below 0.72. The growth of L. monocytogenes in the absence and presence of leucocin B-TA11a was analyzed by flow cytometry with Baclight, plate counts, and optical density measurements. Although leucocin B-TA11a initially inhibited listerial growth, the uptake of both Baclight dyes suggested that cells remained viable but became leaky, possibly indicating bacteriocin-induced pore formation in the target membranes. Received: 30 June 1997 / Accepted: 20 October 1997     

Biochemical and nutritional characterization of the medfly gut symbiont Enterobacter sp. AA26 for its use as probiotics in sterile insect technique applications

Background

Symbiotic bacteria contribute to a multitude of important biological functions such as nutrition and reproduction and affect multiple physiological factors like fitness and longevity in their insect hosts. The melon fly, Zeugodacus cucurbitae (Coquillett), is an important agricultural pest that affects a variety of cultivated plants belonging mostly to the Cucurbitaceae family. It is considered invasive and widespread in many parts of the world. Several approaches are currently being considered for the management of its populations including the environmentally friendly and effective sterile insect technique (SIT), as a component of an integrated pest management (IPM) strategy. In the present study, we examined the effect of diet and radiation on the bacterial symbiome of Z. cucurbitae flies with the use of Next Generation Sequencing technologies.

Results

Melon flies were reared on two diets at the larval stage, an artificial bran-based diet and on sweet gourd, which affected significantly the development of the bacterial profiles. Significant differentiation was also observed based on gender. The effect of radiation was mostly diet dependent, with irradiated melon flies reared on the bran diet exhibiting a significant reduction in species diversity and richness compared to their non-irradiated controls. Changes in the bacterial symbiome of the irradiated melon flies included a drastic reduction in the number of sequences affiliated with members of Citrobacter, Raoultella, and Enterobacteriaceae. At the same time, an increase was observed for members of Enterobacter, Providencia and Morganella. Interestingly, the irradiated male melon flies reared on sweet gourd showed a clear differentiation compared to their non-irradiated controls, namely a significant reduction in species richness and minor differences in the relative abundance for members of Enterobacter and Providencia.

Conclusions

The two diets in conjunction with the irradiation affected significantly the formation of the bacterial symbiome. Melon flies reared on the bran-based artificial diet displayed significant changes in the bacterial symbiome upon irradiation, in all aspects, including species richness, diversity and composition. When reared on sweet gourd, significant changes occurred to male samples due to radiation, only in terms of species richness.

     

The internal microbial flora of the elm bark beetle, Scolytus scolytus, at all stages of its development

Adult Scolytus scolytus emerged from the pupal stage with sterile guts although microorganisms were present within the pupal body. Gut sterility was maintained in a large proportion of newly formed adults while they were boring out of an elm log. The adult gut microbial flora is mainly acquired during subsequent feeding. Differences were found between the microbial gut flora of both larvae and adult S. scolytus, and of both male and female beetles.     

Diversity of bacteria in different life stages and their impact on the development and reproduction of Zeugodacus tau (Diptera: Tephritidae)

Fruit flies usually harbor diverse communities of bacteria in their digestive systems,which are known to play a significant role in their fitness.However,little information is available on Zeugodacus tau,a polyphagous pest worldwide.This study reports the first extensive analysis of bacterial communities in different life stages and their effect on the development and reproduction of laboratory-reared Z tan.Cultured bacteria were identified using the conventional method and all bacteria were identified by highthroughput technologies(16S ribosomal RNA gene sequencing of V3-V4 region).A total of six bacterial phyla were identified in larvae,pupae,and male and female adult flies,which were distributed into 14 classes,32 orders,58 families and 96 genera.Proteobacteria was the most represented phylum in all the stages except larvae.Enterobacter,Klebsiella,Providencia,and Pseudomonas were identified by conventional and next-generation sequencing analysis in both male and female adult flies,and Enterobacter was found to be the main genus.After being fed with antibiotics from the first instar larvae,bacterial diversity changed markedly in the adult stage.Untreated flies laid eggs and needed 20 days before oviposition while the treated flies showed ovary development inhibited and were not able to lay eggs,probably due to the alteration of the microbiota.These findings provide the cornerstone for unexplored research on bacterial function in Z tau,which will help to develop an environmentally friendly management technique for this kind of harmful insect.     

Characteristics of nutrient dynamics in Lake Sagate,Japan, a shallow sand dune lake

We examined differences in digestibility and viability following gut passage through water penny larvae (Psephenus herricki) of Synedra ulna and Achnanthidium lanceoloatum, two common diatom taxa that differ in growth habit and autecological characteristics. Prior to the experiment, diatoms were cultured in Chu-10 media in petri plates to establish a monospecific biofilm to offer grazers. After collection, insects were left to clear their guts over night, allowed to graze for 3 hours on diatom biofilms, and then placed in vials over 1-mm mesh to defecate. Samples from source material and from insect feces were mounted in syrup media and the ratio of chloroplast-containing to empty diatom frustules was microscopically assessed. In addition, subsamples from source material and feces were sprayed onto agar plates prepared with Chu-10 and individual cells were mapped and tracked for 5 days to quantify reproduction. Cells of both S. ulna and A. lanceolatum taken from source material formed colonies on agar. Achnanthidium lanceolatum cells from insect feces also formed colonies, but with lower densities than those from source material. In contrast, none of the S. ulna cells tracked from fecal cultures formed colonies, and the percentage of S. ulna cells that were dead was significantly greater in feces relative to source material. Dead cell percentages of A. lanceolatum were also higher in feces relative to source material, but to a lesser degree than observed for S. ulna. These findings have potential implications for linking patterns of energy transfer in stream ecosystems and the structure and dynamics of benthic microalgal communities.     

Hormonal and nutritional regulation of insect fat body development and function

The insect fat body is an organ analogue to vertebrate adipose tissue and liver and functions as a major organ for nutrient storage and energy metabolism. Similar to other larval organs, fat body undergoes a developmental “remodeling” process during the period of insect metamorphosis, with the massive destruction of obsolete larval tissues by programmed cell death and the simultaneous growth and differentiation of adult tissues from small clusters of progenitor cells. Genetic ablation of Drosophila fat body cells during larval‐pupal transition results in lethality at the late pupal stage and changes sizes of other larval organs indicating that fat body is the center for pupal development and adult formation. Fat body development and function are largely regulated by several hormonal (i.e. insulin and ecdysteroids) and nutritional signals, including oncogenes and tumor suppressors in these pathways. Combining silkworm physiology with fruitfly genetics might provide a valuable system to understand the mystery of hormonal regulation of insect fat body development and function. © 2009 Wiley Periodicals, Inc.     

Bacteria abundance and diversity of different life stages of Plutella xylostella (Lepidoptera: Plutellidae), revealed by bacteria culture‐dependent and PCR‐DGGE methods

Microbial abundance and diversity of different life stages (fourth instar larvae, pupae and adults) of the diamondback moth, Plutella xylostella L., collected from field and reared in laboratory, were investigated using bacteria culture‐dependent method and PCR‐DGGE analysis based on the sequence of bacteria 16S rRNA V3 region gene. A large quantity of bacteria was found in all life stages of P. xylostella. Field population had higher quantity of bacteria than laboratory population, and larval gut had higher quantity than pupae and adults. Culturable bacteria differed in different life stages of P. xylostella. Twenty‐five different bacterial strains were identified in total, among them 20 strains were presented in larval gut, only 8 strains in pupae and 14 strains in adults were detected. Firmicutes bacteria, Bacillus sp., were the most dominant species in every life stage. 15 distinct bands were obtained from DGGE electrophoresis gel. The sequences blasted in GenBank database showed these bacteria belonged to six different genera. Phylogenetic analysis showed the sequences of the bacteria belonged to the Actinobacteri, Proteobacteria and Firmicutes. Serratia sp. in Proteobacteria was the most abundant species in larval gut. In pupae, unculturable bacteria were the most dominant species, and unculturable bacteria and Serratia sp. were the most dominant species in adults. Our study suggested that a combination of molecular and traditional culturing methods can be effectively used to analyze and to determine the diversity of gut microflora. These known bacteria may play important roles in development of P. xylostella.     

Molecular analysis of autochthonous microbiota along the digestive tract of juvenile grouper Epinephelus coioides following probiotic Bacillus pumilus administration

Aims: To evaluate the diversity of dominant autochthonous microbiota along the digestive tract of juvenile Epinephelus coioides following the dietary administration of probiotic Bacillus pumilus for 60 days. Methods and Results: Polymerase chain reaction‐denaturing gradient gel electrophoresis (PCR‐DGGE) with subsequently sequencing analysis was used to assess the gut microbiota. Generally similar DGGE patterns were observed in the foregut, midgut and hindgut of E. coioides, while the similarity dendrogram clearly revealed three different clusters depending on the three compartments of the GI tract. Dietary administration of B. pumilus stimulated its colonization in each compartment of the digestive tract. Samples collected from the probiotic group and the control group showed similar DGGE patterns, and no significant difference in the total number of bands and the Shannon index were detected between the probiotic group and the control group, suggested that B. pumilus exert no significant effect on the gut microbiota. However, various potentially beneficial bacteria, such as uncultured Bacillus sp. clone QJNY94‐like, Nitratireductor sp. YCSC5‐like, Methylobacterium hispanicum‐like and Microbacterium sp. YACS1‐like bacteria were stimulated by probiotic B. pumilus, while the potential harmful Staphylococcus saprophyticus‐like bacterium was depressed. Conclusions: Autochthonous gut microbiota of E. coioides was modulated to some degree, not significant, by probiotic B. pumilus, various potentially beneficial bacteria were selectively stimulated, while one potential harmful species was depressed. Significance and Impact of the Study: This work represents the first report that dietary administration of probiotic B. pumilus modulated the gut microbiota of E. coioides. These findings broaden our understanding of probiotic effects at the gut level, which is helpful in understanding the mechanisms that underpin host benefits.     

Activities of plant cell wall-degrading enzymes by bacterial soft rot of orchid

Soft rot by bacterial pathogens is one of the most widespread and destructive diseases on various plants including orchids throughout the world. The pathogenicity of the pathogens is reported to be mainly determined by massive production of plant cell wall-degrading enzymes (PCDE). In the previous work, we have isolated 20 isolates of bacterial soft rot from orchids collected in Yogyakarta Special Region and West Java province, Indonesia. In this study, we further confirmed them as pathogens by hypersensitive reaction assay on tobacco leaves followed by pathogenicity test on Phalaenopsis sp. The production of four major PCDE by qualitative plate assays including pectate lyase, polygalacturonase, cellulase and protease was also evaluated. Even though all the isolates were able to initiate soft rot symptom, our results showed two distinct groups which clustered as producing and non-producing PCDE. The 16S rDNA analysis revealed that the isolates belonged to the genera Pectobacterium, Klebsiella, Serratia, Enterobacter, Citrobacter, Providencia and Pseudomonas.     

Taking insight into the gut microbiota of three spider species: No characteristic symbiont was found corresponding to the special feeding style of 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.     

Effects of two insect growth regulators and a neonicotinoid on various life stages of <Emphasis Type="Italic">Aphytis melinus</Emphasis> (Hymenoptera: Aphelinidae)

The effect of two insect growth regulators and a neonicotinoid insecticide were tested on immature stages and adults of the parasitoid Aphtyis melinus DeBach (Hymenoptera: Aphelinidae), a key natural enemy of California red scale, Aonidiella aurantii (Maskell) (Hemiptera: Diaspididae), in California. No significant effects of the insect growth regulators on survival or development to the adult stage were found when the parasitoid was treated at any of the egg, larval, or pupal stages. The broad-spectrum neonicotinoid acetamiprid also showed no significant effect on the development of A. melinus to the pupal stage, probably because immature stages of this ectoparasitoid are protected under the cover of its armored scale host. However, 48 h exposure of adults to acetamiprid residues following emergence resulted in high levels of wasp mortality. Aphytis melinus adults treated with either of the two insect growth regulators as larvae survived 48 h exposure to pesticide residues as adults and showed levels of fecundity comparable with control insects. We conclude that the two insect growth regulators are compatible with augmentative releases of A. melinus but that treatments of acetamiprid should be avoided in situations where biological control by this parasitoid is important.