Impact of plant nutrients on the relationship between a herbivorous insect and its symbiotic bacteria

The interactions between herbivorous insects and their symbiotic micro-organisms can be influenced by the plant species on which the insects are reared, but the underlying mechanisms are not understood. Here, we identify plant nutrients, specifically amino acids, as a candidate factor affecting the impact of symbiotic bacteria on the performance of the phloem-feeding aphid Aphis fabae. Aphis fabae grew more slowly on the labiate plant Lamium purpureum than on an alternative host plant Vicia faba, and the negative effect of L. purpureum on aphid growth was consistently exacerbated by the bacterial secondary symbionts Regiella insecticola and Hamiltonella defensa, which attained high densities in L. purpureum-reared aphids. The amino acid content of the phloem sap of L. purpureum was very low; and A. fabae on chemically defined diets of low amino acid content also grew slowly and had elevated secondary symbiont densities. It is suggested that the phloem nutrient profile of L. purpureum promotes deleterious traits in the secondary symbionts and disturbs insect controls over bacterial abundance.     

Sources of variation in dietary requirements in an obligate nutritional symbiosis

The nutritional symbiosis between aphids and their obligate symbiont, Buchnera aphidicola, is often characterized as a highly functional partnership in which the symbiont provides the host with essential nutrients. Despite this, some aphid lineages exhibit dietary requirements for nutrients typically synthesized by Buchnera, suggesting that some aspect of the symbiosis is disrupted. To examine this phenomenon in the pea aphid, Acyrthosiphon pisum, populations were assayed using defined artificial diet to determine dietary requirements for essential amino acids (EAAs). Six clones exhibiting dependence on EAAs in their diet were investigated further. In one aphid clone, a mutation in a Buchnera amino acid biosynthesis gene could account for the clone''s requirement for dietary arginine. Analysis of aphid F₁ hybrids allowed separation of effects of the host and symbiont genomes, and revealed that both affect the requirement for dietary EAAs in the clones tested. Amino acid requirements were minimally affected by secondary symbiont infection. Our results indicate that variation among pea aphids in dependence on dietary amino acids can result from Buchnera mutation as well as variation in the host genotype.     

How the insect immune system interacts with an obligate symbiotic bacterium

The animal immune system provides defence against microbial infection, and the evolution of certain animal–microbial symbioses is predicted to involve adaptive changes in the host immune system to accommodate the microbial partner. For example, the reduced humoral immune system in the pea aphid Acyrthosiphon pisum, including an apparently non-functional immune deficiency (IMD) signalling pathway and absence of peptidoglycan recognition proteins (PGRPs), has been suggested to be an adaptation for the symbiosis with the bacterium Buchnera aphidicola. To investigate this hypothesis, the interaction between Buchnera and non-host cells, specifically cultured Drosophila S2 cells, was investigated. Microarray analysis of the gene expression pattern in S2 cells indicated that Buchnera triggered an immune response, including upregulated expression of genes for antimicrobial peptides via the IMD pathway with the PGRP-LC as receptor. Buchnera cells were readily taken up by S2 cells, but were subsequently eliminated over 1–2 days. These data suggest that Buchnera induces in non-host cells a defensive immune response that is deficient in its host. They support the proposed contribution of the Buchnera symbiosis to the evolution of the apparently reduced immune function in the aphid host.     

Genetic and metabolic determinants of nutritional phenotype in an insect-bacterial symbiosis

The pervasive influence of resident microorganisms on the phenotype of their hosts is exemplified by the intracellular bacterium Buchnera aphidicola, which provides its aphid partner with essential amino acids (EAAs). We investigated variation in the dietary requirement for EAAs among four pea aphid (Acyrthosiphon pisum) clones. Buchnera-derived nitrogen contributed to the synthesis of all EAAs for which aphid clones required a dietary supply, and to none of the EAAs for which all four clones had no dietary requirement, suggesting that low total dietary nitrogen may select for reduced synthesis of certain EAAs in some aphid clones. The sequenced Buchnera genomes showed that the EAA nutritional phenotype (i.e. the profile of dietary EAAs required by the aphid) cannot be attributed to sequence variation of Buchnera genes coding EAA biosynthetic enzymes. Metabolic modelling by flux balance analysis demonstrated that EAA output from Buchnera can be determined precisely by the flux of host metabolic precursors to Buchnera. Specifically, the four EAA nutritional phenotypes could be reproduced by metabolic models with unique profiles of host inputs, dominated by variation in supply of aspartate, homocysteine and glutamate. This suggests that the nutritional phenotype of the symbiosis is determined principally by host metabolism and transporter genes that regulate nutrient supply to Buchnera. Intraspecific variation in the nutritional phenotype of symbioses is expected to mediate partitioning of plant resources among aphid genotypes, potentially promoting the genetic subdivision of aphid populations. In this way, microbial symbioses may play an important role in the evolutionary diversification of phytophagous insects.     

Effects of bacterial secondary symbionts on host plant use in pea aphids

Aphids possess several facultative bacterial symbionts that have important effects on their hosts'' biology. These have been most closely studied in the pea aphid (Acyrthosiphon pisum), a species that feeds on multiple host plants. Whether secondary symbionts influence host plant utilization is unclear. We report the fitness consequences of introducing different strains of the symbiont Hamiltonella defensa into three aphid clones collected on Lathyrus pratensis that naturally lack symbionts, and of removing symbionts from 20 natural aphid–bacterial associations. Infection decreased fitness on Lathyrus but not on Vicia faba, a plant on which most pea aphids readily feed. This may explain the unusually low prevalence of symbionts in aphids collected on Lathyrus. There was no effect of presence of symbiont on performance of the aphids on the host plants of the clones from which the H. defensa strains were isolated. Removing the symbiont from natural aphid–bacterial associations led to an average approximate 20 per cent reduction in fecundity, both on the natural host plant and on V. faba, suggesting general rather than plant-species-specific effects of the symbiont. Throughout, we find significant genetic variation among aphid clones. The results provide no evidence that secondary symbionts have a major direct role in facilitating aphid utilization of particular host plant species.     

The central role of the host cell in symbiotic nitrogen metabolism

Symbiotic nitrogen recycling enables animals to thrive on nitrogen-poor diets and environments. It traditionally refers to the utilization of animal waste nitrogen by symbiotic micro-organisms to synthesize essential amino acids (EAAs), which are translocated back to the animal host. We applied metabolic modelling and complementary metabolite profiling to investigate nitrogen recycling in the symbiosis between the pea aphid and the intracellular bacterium Buchnera, which synthesizes EAAs. The results differ from traditional notions of nitrogen recycling in two important respects. First, aphid waste ammonia is recycled predominantly by the host cell (bacteriocyte) and not Buchnera. Host cell recycling is mediated by shared biosynthetic pathways for four EAAs, in which aphid transaminases incorporate ammonia-derived nitrogen into carbon skeletons synthesized by Buchnera to generate EAAs. Second, the ammonia substrate for nitrogen recycling is derived from bacteriocyte metabolism, such that the symbiosis is not a sink for nitrogenous waste from other aphid organs. Host cell-mediated nitrogen recycling may be general among insect symbioses with shared EAA biosynthetic pathways generated by the loss of symbiont genes mediating terminal reactions in EAA synthesis.     

Interaction between host genotype and environmental conditions affects bacterial density in Wolbachia symbiosis

Regulation of microbial population density is a necessity in stable symbiotic interactions. In Wolbachia symbiosis, both bacterial and host genotypes are involved in density regulation, but environmental factors may also affect bacterial population density. Here, we studied the interaction between three strains of Wolbachia in two divergent homozygous lines of the wasp Leptopilina heterotoma at two different temperatures. Wolbachia density varied between the two host genotypes at only one temperature. Moreover, at this temperature, reciprocal-cross F1 insects displayed identical Wolbachia densities, which were intermediate between the densities in the two parental lines. While these findings confirm that the host genotype plays an important role in Wolbachia density, they also highlight its interaction with environmental conditions, making possible the evolution of local adaptations for the regulation of Wolbachia density.     

Plant-ants feed their host plant, but above all a fungal symbiont to recycle nitrogen

In ant-plant symbioses, plants provide symbiotic ants with food and specialized nesting cavities (called domatia). In many ant-plant symbioses, a fungal patch grows within each domatium. The symbiotic nature of the fungal association has been shown in the ant-plant Leonardoxa africana and its protective mutualist ant Petalomyrmex phylax. To decipher trophic fluxes among the three partners, food enriched in (13)C and (15)N was given to the ants and tracked in the different parts of the symbiosis up to 660 days later. The plant received a small, but significant, amount of nitrogen from the ants. However, the ants fed more intensively the fungus. The pattern of isotope enrichment in the system indicated an ant behaviour that functions specifically to feed the fungus. After 660 days, the introduced nitrogen was still present in the system and homogeneously distributed among ant, plant and fungal compartments, indicating efficient recycling within the symbiosis. Another experiment showed that the plant surface absorbed nutrients (in the form of simple molecules) whether or not it is coated by fungus. Our study provides arguments for a mutualistic status of the fungal associate and a framework for investigating the previously unsuspected complexity of food webs in ant-plant mutualisms.     

Gibberellin regulates infection and colonization of host roots by arbuscular mycorrhizal fungi

Arbuscular mycorrhiza (AM) is established by the entry of AM fungi into the host plant roots and the formation of symbiotic structures called arbuscules. The host plant supplies photosynthetic products to the AM fungi, which in return provide phosphate and other minerals to the host through the arbuscules. Both partners gain great advantages from this symbiotic interaction, and both regulate AM development. Our recent work revealed that gibberellic acids (GAs) are required for AM development in the legume Lotus japonicus. GA signaling interact with symbiosis signaling pathways, directing AM fungal colonization in host roots. Expression analysis showed that genes for GA biosynthesis and metabolism were induced in host roots around AM fungal hyphae, suggesting that the GA signaling changes with both location and time during AM development. The fluctuating GA concentrations sometimes positively and sometimes negatively affect the expression of AM-induced genes that regulate AM fungal infection and colonization.     

Cellular and molecular remodelling of a host cell for vertical transmission of bacterial symbionts

Various insects require intracellular bacteria that are restricted to specialized cells (bacteriocytes) and are transmitted vertically via the female ovary, but the transmission mechanisms are obscure. We hypothesized that, in the whitefly Bemisia tabaci, where intact bacteriocytes (and not isolated bacteria) are transferred to oocytes, the transmission mechanism would be evident as cellular and molecular differences between the nymph (pre-adult) and adult bacteriocytes. We demonstrate dramatic remodelling of bacteriocytes at the developmental transition from nymph to adulthood. This transition involves the loss of cell–cell adhesion, high division rates to constant cell size and onset of cell mobility, enabling the bacteriocytes to crawl to the ovaries. These changes are accompanied by cytoskeleton reorganization and changes in gene expression: genes functioning in cell–cell adhesion display reduced expression and genes involved in cell division, cell motility and endocytosis/exocytosis have elevated expression in adult bacteriocytes, relative to nymph bacteriocytes. This study demonstrates, for the first time, how developmentally orchestrated remodelling of gene expression and correlated changes in cell behaviour underpin the capacity of bacteriocytes to mediate the vertical transmission and persistence of the symbiotic bacteria on which the insect host depends.     

Complete Genome sequence of Burkholderia phymatum STM815T,a broad host range and efficient nitrogen-fixing symbiont of Mimosa species

Burkholderia phymatum is a soil bacterium able to develop a nitrogen-fixing symbiosis with species of the legume genus Mimosa, and is frequently found associated specifically with Mimosa pudica. The type strain of the species, STM 815^T, was isolated from a root nodule in French Guiana in 2000. The strain is an aerobic, motile, non-spore forming, Gram-negative rod, and is a highly competitive strain for nodulation compared to other Mimosa symbionts, as it also nodulates a broad range of other legume genera and species. The 8,676,562 bp genome is composed of two chromosomes (3,479,187 and 2,697,374 bp), a megaplasmid (1,904,893 bp) and a plasmid hosting the symbiotic functions (595,108 bp).     

Cheaper is not always worse: strongly protective isolates of a defensive symbiont are less costly to the aphid host

Defences against parasites are typically associated with costs to the host that contribute to the maintenance of variation in resistance. This also applies to the defence provided by the facultative bacterial endosymbiont Hamiltonella defensa, which protects its aphid hosts against parasitoid wasps while imposing life-history costs. To investigate the cost–benefit relationship within protected hosts, we introduced multiple isolates of H. defensa to the same genetic backgrounds of black bean aphids, Aphis fabae, and we quantified the protection against their parasitoid Lysiphlebus fabarum as well as the costs to the host (reduced lifespan and reproduction) in the absence of parasitoids. Surprisingly, we observed the opposite of a trade-off. Strongly protective isolates of H. defensa reduced lifespan and lifetime reproduction of unparasitized aphids to a lesser extent than weakly protective isolates. This finding has important implications for the evolution of defensive symbiosis and highlights the need for a better understanding of how strain variation in protective symbionts is maintained.     

Metagenomic and metaproteomic insights into bacterial communities in leaf-cutter ant fungus gardens

Herbivores gain access to nutrients stored in plant biomass largely by harnessing the metabolic activities of microbes. Leaf-cutter ants of the genus Atta are a hallmark example; these dominant neotropical herbivores cultivate symbiotic fungus gardens on large quantities of fresh plant forage. As the external digestive system of the ants, fungus gardens facilitate the production and sustenance of millions of workers. Using metagenomic and metaproteomic techniques, we characterize the bacterial diversity and physiological potential of fungus gardens from two species of Atta. Our analysis of over 1.2 Gbp of community metagenomic sequence and three 16S pyrotag libraries reveals that in addition to harboring the dominant fungal crop, these ecosystems contain abundant populations of Enterobacteriaceae, including the genera Enterobacter, Pantoea, Klebsiella, Citrobacter and Escherichia. We show that these bacterial communities possess genes associated with lignocellulose degradation and diverse biosynthetic pathways, suggesting that they play a role in nutrient cycling by converting the nitrogen-poor forage of the ants into B-vitamins, amino acids and other cellular components. Our metaproteomic analysis confirms that bacterial glycosyl hydrolases and proteins with putative biosynthetic functions are produced in both field-collected and laboratory-reared colonies. These results are consistent with the hypothesis that fungus gardens are specialized fungus–bacteria communities that convert plant material into energy for their ant hosts. Together with recent investigations into the microbial symbionts of vertebrates, our work underscores the importance of microbial communities in the ecology and evolution of herbivorous metazoans.     

4种野生蔬菜的氨基酸含量及其营养价值评价

野生蔬菜具有独特的口味,丰富的营养,有些还具有防病治病的功能,对人体有良好的保健作用.白子菜[Gynura divaricata (L.) DC.]茎叶有消热、舒筋及止血祛瘀的作用,用于镇咳、风湿性关节痛、骨折、创伤止血,痈肿疮疥等症,在民间还作抗癌草使用[1].红凤菜(Gynura bicolor DC.)茎叶有活血止血,解毒消肿的功效,治痛经,血崩,咳血,创伤出血,溃疡久不收口[2].诸葛菜[Orychophragmus violaceus (L.) Schulz]作为野生蔬菜在江苏、安徽民间有很长的食用历史,测定结果表明,该种含有黄酮类化合物[3].马齿苋(Portulaca oleracea L.)具有清热解毒、消炎、利尿、消肿的功效,民间还用于白发症、糖尿病、心血管疾病的治疗[4],中国卫生部将它列为药食同用的植物.     

The green peach aphid,Myzus persicae,acquires a LIPOXYGENASE5-derived oxylipin from Arabidopsis thaliana,which promotes colonization of the host plant

Oxylipins derived from lipoxygenase (LOX) activity play important roles in plant growth, development and stress response. In a recent study, we provided evidence that infestation of Arabidopsis thaliana foliage by the green peach aphid (GPA; Myzus persicae), a phloem sap-consuming insect, was promoted by plant LOX5-derived oxylipins. In comparison to the wild-type (WT) plant, GPA population was smaller on the Arabidopsis lox5 mutant. The insect spent less time feeding from the sieve element and xylem of the lox5 mutant compared with the WT plant. In addition, compared with insects feeding on the WT plant, when on the lox5 mutant, the GPA was unable to suppress an antibiotic activity that is present in Arabidopsis vascular sap. Roots are the critical source of a LOX5-derived oxylipin(s) that promotes colonization of the foliage by GPA. Here we show that the 9-hydoxy-10E, 12Z-octadecadienoic acid (9-HOD), a LOX5-derived oxylipin, accumulated in GPA that were reared on the WT, but not the lox5 mutant plant. However, 9-HOD accumulated in insects reared on lox5 mutant plants that were irrigated with 9-HOD, thus indicating that the insect ingests oxylipins from the host plant. We further demonstrate that the host plant requires LOX5 function to promote expression of the defense regulatory gene PHYTOALEXIN-DEFICIENT4 in the foliage. Taken together, our previous observations and results presented here indicate that while the host plant utilizes LOX5-dependent factors for promoting defense mechanisms, GPA has evolved to utilize plant 9-LOX-derived oxylipins as cues to facilitate infestation, thus suggesting a complex involvement of oxylipins in Arabidopsis interaction with GPA.     

Responses of the coastal bacterial community to viral infection of the algae Phaeocystis globosa

The release of organic material upon algal cell lyses has a key role in structuring bacterial communities and affects the cycling of biolimiting elements in the marine environment. Here we show that already before cell lysis the leakage or excretion of organic matter by infected yet intact algal cells shaped North Sea bacterial community composition and enhanced bacterial substrate assimilation. Infected algal cultures of Phaeocystis globosa grown in coastal North Sea water contained gamma- and alphaproteobacterial phylotypes that were distinct from those in the non-infected control cultures 5 h after infection. The gammaproteobacterial population at this time mainly consisted of Alteromonas sp. cells that were attached to the infected but still intact host cells. Nano-scale secondary-ion mass spectrometry (nanoSIMS) showed ∼20% transfer of organic matter derived from the infected ¹³C- and ¹⁵N-labelled P. globosa cells to Alteromonas sp. cells. Subsequent, viral lysis of P. globosa resulted in the formation of aggregates that were densely colonised by bacteria. Aggregate dissolution was observed after 2 days, which we attribute to bacteriophage-induced lysis of the attached bacteria. Isotope mass spectrometry analysis showed that 40% of the particulate ¹³C-organic carbon from the infected P. globosa culture was remineralized to dissolved inorganic carbon after 7 days. These findings reveal a novel role of viruses in the leakage or excretion of algal biomass upon infection, which provides an additional ecological niche for specific bacterial populations and potentially redirects carbon availability.     

Quorum-sensing blockade as a strategy for enhancing host defences against bacterial pathogens

Conventional antibiotics target the growth and the basal life processes of bacteria leading to growth arrest and cell death. The selective force that is inherently linked to this mode of action eventually selects out antibiotic-resistant variants. The most obvious alternative to antibiotic-mediated killing or growth inhibition would be to attenuate the bacteria with respect to pathogenicity. The realization that Pseudomonas aeruginosa, and a number of other pathogens, controls much of their virulence arsenal by means of extracellular signal molecules in a process denoted quorum sensing (QS) gave rise to a new 'drug target rush'. Recently, QS has been shown to be involved in the development of tolerance to various antimicrobial treatments and immune modulation. The regulation of virulence via QS confers a strategic advantage over host defences. Consequently, a drug capable of blocking QS is likely to increase the susceptibility of the infecting organism to host defences and its clearance from the host. The use of QS signal blockers to attenuate bacterial pathogenicity, rather than bacterial growth, is therefore highly attractive, particularly with respect to the emergence of multi-antibiotic resistant bacteria.