Anti-herbivory defences delivered by Epichloë fungal endophytes: a quantitative review of alkaloid concentration variation among hosts and plant parts.

BACKGROUND AND AIMS: In the subfamily Poöideae (Poaceae), certain grass species possess anti-herbivore alkaloids synthesized by fungal endophytes that belong to the genus Epichloë (Clavicipitaceae). The protective role of these symbiotic endophytes can vary, depending on alkaloid concentrations within specific plant-endophyte associations and plant parts. METHODS: We conducted a literature review to identify articles containing alkaloid concentration data for various plant parts in six important pasture species, Lolium arundinaceum, Lolium perenne, Lolium pratense, Lolium multiflorum|Lolium rigidum and Festuca rubra, associated with their common endophytes. We considered the alkaloids lolines (1-aminopyrrolizidines), peramine (pyrrolopyrazines), ergovaline (ergot alkaloids) and lolitrem B (indole-diterpenes). While all these alkaloids have shown bioactivity against insect herbivores, ergovaline and lolitrem B are harmful for mammals. KEY RESULTS: Loline alkaloid levels were higher in the perennial grasses L. pratense and L. arundinaceum compared to the annual species L. multiflorum and L. rigidum, and higher in reproductive tissues than in vegetative structures. This is probably due to the greater biomass accumulation in perennial species that can result in higher endophyte mycelial biomass. Peramine concentrations were higher in L. perenne than in L. arundinaceum and not affected by plant part. This can be attributed to the high within-plant mobility of peramine. Ergovaline and lolitrem B, both hydrophobic compounds, were associated with plant parts where fungal mycelium is usually present, and their concentrations were higher in plant reproductive tissues. Only loline alkaloid data were sufficient for below-ground tissue analyses and concentrations were lower than in above-ground parts. CONCLUSIONS: Our study provides a comprehensive synthesis of fungal alkaloid variation across host grasses and plant parts, essential for understanding the endophyte-conferred defence extent. The patterns can be understood by considering endophyte growth within the plant and alkaloid mobility. Our study identifies research gaps, including the limited documentation of alkaloid presence in roots and the need to investigate the influence of different environmental conditions.

Small burrowing amphipods cause major damage in a large kelp.

Large herbivores such as sea urchins and fish consume a high proportion of benthic primary production and frequently control the biomass of marine macrophytes. By contrast, small mesograzers, including gastropods and peracarid crustaceans, are abundant on seaweeds but have low per capita feeding rates and their impacts on marine macrophytes are difficult to predict. To quantify how mesograzers can affect macrophytes, we examined feeding damage by the herbivorous amphipods Sunamphitoe lessoniophila and Bircenna sp., which construct burrows in the stipes of subtidal individuals of the kelp Lessonia berteroana in northern-central Chile, southeast Pacific. Infested stipes showed a characteristic sequence of progressive tissue degeneration. The composition of the amphipod assemblages inside the burrows varied between the different stages of infestation of the burrows. Aggregations of grazers within burrows and microhabitat preference of the amphipods result in localized feeding, leading to stipe breakage and loss of substantial algal biomass. The estimated loss of biomass of single stipes varied between 1 and 77%. For the local kelp population, the amphipods caused an estimated loss of biomass of 24-44%. Consequently, small herbivores can cause considerable damage to large kelp species if their feeding activity is concentrated on structurally valuable algal tissue.

Sipha maydis sensitivity to defences of Lolium multiflorum and its endophytic fungus Epichloë occultans.

BACKGROUND: Plants possess a sophisticated immune system to defend from herbivores. These defence responses are regulated by plant hormones including salicylic acid (SA) and jasmonic acid (JA). Sometimes, plant defences can be complemented by the presence of symbiotic microorganisms. A remarkable example of this are grasses establishing symbiotic associations with Epichloë fungal endophytes. We studied the level of resistance provided by the grass' defence hormones, and that provided by Epichloë fungal endophytes, against an introduced herbivore aphid. These fungi protect their hosts against herbivores by producing bioactive alkaloids. We hypothesized that either the presence of fungal endophytes or the induction of the plant salicylic acid (SA) defence pathway would enhance the level of resistance of the grass to the aphid. METHODS: Lolium multiflorum plants, with and without the fungal endophyte Epichloë occultans, were subjected to an exogenous application of SA followed by a challenge with the aphid, Sipha maydis. RESULTS: Our results indicate that neither the presence of E. occultans nor the induction of the plant's SA pathway regulate S. maydis populations. However, endophyte-symbiotic plants may have been more tolerant to the aphid feeding because these plants produced more aboveground biomass. We suggest that this insect insensitivity could be explained by a combination between the ineffectiveness of the specific alkaloids produced by E. occultans in controlling S. maydis aphids and the capacity of this herbivore to deal with hormone-dependent defences of L. multiflorum.

Making a Better Home: Modulation of Plant Defensive Response by Brevipalpus Mites.

False-spider mites of the genus Brevipalpus are highly polyphagous pests that attack hundreds of plant species of distinct families worldwide. Besides causing direct damage, these mites may also act as vectors of many plant viruses that threaten high-value ornamental plants like orchids and economically important crops such as citrus and coffee. To better understand the molecular mechanisms behind plant-mite interaction we used an RNA-Seq approach to assess the global response of Arabidopsis thaliana (Arabidopsis) plants along the course of the infestation with Brevipalpus yothersi, the main vector species within the genus. Mite infestation triggered a drastic transcriptome reprogramming soon at the beginning of the interaction and throughout the time course, deregulating 1755, 3069 and 2680 genes at 6 hours after infestation (hai), 2 days after infestation (dai), and 6 dai, respectively. Gene set enrichment analysis revealed a clear modulation of processes related to the plant immune system. Co-expressed genes correlated with specific classes of transcription factors regulating defense pathways and developmental processes. Up-regulation of defensive responses correlated with the down-regulation of growth-related processes, suggesting the triggering of the growth-defense crosstalk to optimize plant fitness. Biological processes (BPs) enriched at all time points were markedly related to defense against herbivores and other biotic stresses involving the defense hormones salicylic acid (SA) and jasmonic acid (JA). Levels of both hormones were higher in plants challenged with mites than in the non-infested ones, supporting the simultaneous induction of genes from both pathways. To further clarify the functional relevance of the plant hormonal pathways on the interaction, we evaluated the mite performance on Arabidopsis mutants impaired in SA- or JA-mediated response. Mite oviposition was lower on mutants defective in SA biosynthesis (sid2) and signaling (npr1), showing a function for SA pathway in improving the mite reproduction, an unusual mechanism compared to closely-related spider mites. Here we provide the first report on the global and dynamic plant transcriptome triggered by Brevipalpus feeding, extending our knowledge on plant-mite interaction. Furthermore, our results suggest that Brevipalpus mites manipulate the plant defensive response to render the plant more susceptible to their colonization by inducing the SA-mediated pathway.

Indirect effects of mutualism: ant-treehopper associations deter pollinators and reduce reproduction in a tropical shrub.

Animal-pollinated plants can be susceptible to changes in pollinator availability. Honeydew-producing treehoppers frequently occur on inflorescences, potentially enhancing ant-mediated negative effects on pollination services. However, the effect of ant-attended, honeydew-producing insects on plant reproduction remains uncertain. We recorded the abundance of treehoppers and ants on Byrsonima intermedia (Malpighiaceae), and monitored floral visitors in a Brazilian cerrado savanna. We manipulated the presence of ants and ant-treehopper associations on inflorescences to assess their effect on pollination and fruit formation. We used dried ants pinned to inflorescences to evaluate the effect of ant presence and ant identity on potential pollinators. Results show that the presence of treehoppers increases ant abundance on flowers and disrupts pollination by oil-collecting bees, decreasing the frequency and duration of floral visits and reducing fruit and seed set. Treehopper herbivory has no direct effect on fruit or seed production, which are independent of treehopper density. Pinned ants promote avoidance by floral visitors, reducing the number of visits. Ant identity mediates visitation decisions, with Ectatomma brunneum causing greater avoidance by floral visitors than Camponotus rufipes. Field videos show that pollinating bees are harassed by ants near flowers, prompting avoidance behavior by the bees. This is the first demonstration of indirect effects by honeydew-gathering ants, via disrupted pollination, on plant reproduction in tropical cerrado savanna. Our results highlight the importance of studying other interactions near flowers, in addition to just observing pollinators, for a proper understanding of plant reproduction.

GRAZING ON GIANT KELP MICROSCOPIC PHASES AND THE RECRUITMENT SUCCESS OF ANNUAL POPULATIONS OF MACROCYSTIS PYRIFERA (LAMINARIALES, PHAEOPHYTA) IN SOUTHERN CHILE(1).

The giant kelp Macrocystis pyrifera (L.) C. Agardh is widely distributed in the Northern Hemisphere and Southern Hemisphere, yet it exhibits distinct population dynamics at local to regional spatial scales. Giant kelp populations are typically perennial with the potential for year-round reproduction and recruitment. In southern Chile, however, annual giant kelp populations exist and often persist entirely on secondary substrata (e.g., shells of the slipper limpet Crepipatella fecunda [Gastropoda, Calyptraeidae]) that can cover up to 90% of the rocky bottom. In these populations, the macroscopic sporophyte phase disappears annually during winter and early spring, leaving a 3-4 month period in which a persistent microscopic phase remains to support the subsequent year's recruitment. We tested the effects of a suite of grazers on the recruitment success of this critical microscopic phase at two sites in southern Chile. Field experiments indicated that the snail Tegula atra negatively impacted M. pyrifera sporophyte recruitment, but that recruitment was highest in the presence of sessile female limpets, C. fecunda. Conversely, small male C. fecunda (biofilm grazers) did not regulate kelp recruitment. Laboratory observations showed that C. fecunda males only grazed on microscopic kelp gametophytes and small (<250 µm) sporophytes, rejecting larger sporophytes, whereas T. atra grazed on all the kelp stages. Recruitment to the C. fecunda treatments far exceeded that to bare rock in the absence of grazers but was not due to the physical presence of C. fecunda shells. We concluded that the key to M. pyrifera recruitment success in southern Chile is its capacity to colonize secondary substrates provided by the slipper limpet C. fecunda.

High performance liquid chromatography determination of cucurbitacins in the roots of Wilbrandia ebracteata Cogn. / Determinação de cucurbitacinas em raízes de Wilbrandia ebracteata Cogn. por cromatografia líquida de alta eficiência

Roots of Wilbrandia ebracteata Cogn., Cucurbitaceae, used in folk medicine for treatment of rheumatic disease, are rich in cucurbitacins. Dihydrocucurbitacin B is the most abundant cucurbitacin while cucurbitacin B is a minor component. A reverse-phase HPLC system was developed for simultaneous quantitative assay of these cucurbitacins in the roots. The optimised experimental conditions were acetonitrile/H(2)0 40:60, flow-rate 1.2 mL/min., detection at 230 nm and isocratic elution. A variety of sample preparation modes were tested and the extraction with dichloromethane under reflux gave better results. The validation process included linearity, accuracy, repeatability and intermediate precision. The calibration curve of dihydrocucurbitacin B was linear from 40.00 to 400 μg/mL, the recovery was 95.5±3.01 percent, the intermediate precision was found to be 1.64 percent and the repeatability varied between 1.30 to 2.05 percent. The calibration curve of cucurbitacin B was linear from 4.00 to 240 μg/mL, intermediate precision was found to be 2.29 percent and repeatability varied between 1.03 to 2.95 percent. Analysis of the same specimen of W. ebracteata every year from 2002 to 2005 revealed a great rise on the cucurbitacin B concentration after the root was attacked by an herbivore.

Raízes de Wilbrandia ebracteata Cogn. (Cucurbitaceae), tradicionalmente empregada no tratamento de doenças reumáticas, contém cucurbitacinas, sendo di-hidrocucurbitacina B a mais abundante, enquanto cucurbitacina B está presente em menor quantidade. Foi desenvolvido um método para determinação quantitativa destas cucurbitacinas. Os parâmetros selecionados foram: eluente isocrático acetonitrila/H(2)0 40:60, fluxo 1,2 mL/min. e detecção em 230 nm. Diversas formas de preparo da amostra foram testadas, sendo que extração com diclorometano sob refluxo forneceu o melhor resultado. O processo de validação incluiu: linearidade, exatidão, repetibilidade e precisão intermediária. A curva de calibração para a di-hidrocucurbitacina B foi linear de 40.00 to 400 μg/mL, a recuperação foi 95,5±3.01 por cento, a precisão intermediária, 1,64 por cento e a repetibilidade variou entre 1,30 a 2,05 por cento. A curva de calibração da cucurbitacina B foi linear de 4,00 to 240 μg/mL, a recuperação encontrada foi igual a 96,6±2.45 por cento, a precisão intermediária, 2,29 por cento e a repetibilidade variou entre 1,03 a 2,95 por cento. Análise do mesmo espécime de W. ebracteata uma vez por ano de 2002 a 2005 revelou grande aumento no teor de cucurbitacina B após a raiz ter sido atacada por herbívoro.

Attraction of Bucephalogonia xanthophis (Hemiptera: Cicadellidae) to volatiles of its natural host Vernonia condensata (Asteraceae)

Plant volatiles are important cues for the orientation of herbivorous insects. It is possible that these compounds indicate whether the plant is suitable for feeding and larval development, or for mating aggregation. Vernonia condensata (Asteraceae) is known to attract species of leafhoppers, most of them important vectors of the citrus variegated chlorosis (CVC). In this study, we evaluated the role of volatiles of V. condensata on the orientation of Bucephalogonia xanthophis (Hemiptera: Cicadellidae). Four-arm olfactometer bioassays showed that only males were attracted to the volatiles of the host-plants Citrus sp. and V. condensata. Furthermore, fresh leaves of V. condensata induced a stronger response than volatiles from hexane-extracted leaves. This study opens the possibility to utilize V. condensata volatiles for pest management programs of B. xanthopis.

Os voláteis de plantas são sinais importantes para a orientação de insetos herbívoros. É possível que esses compostos indiquem que a planta é apropriada para alimentação e desenvolvimento larval, ou para agrupamento reprodutivo. Vernonia condensata (Asteraceae) atrai espécies de cigarrinhas, muitas delas vetores da clorose variegada dos citros (CVC). Neste trabalho, foi avaliado o papel dos voláteis de V. condensata na orientação de Bucephalogonia xanthophis (Hemiptera: Cicadellidae). Bioensaios em olfatômetro de quatro braços mostraram que somente os machos foram atraídos pelos voláteis das plantas hospedeiras Citrus sp. e V. condensata, sendo que os voláteis das folhas frescas de V. condensata induziram uma maior resposta quando comparada aos extratos hexânicos da planta. A possibilidade de usar os voláteis de V. condensata abre novas perspectivas para programas e alternativas para o manejo de pragas.

Attraction of Bucephalogonia xanthophis (Hemiptera: Cicadellidae) to volatiles of its natural host Vernonia condensata (Asteraceae)

Plant volatiles are important cues for the orientation of herbivorous insects. It is possible that these compounds indicate whether the plant is suitable for feeding and larval development, or for mating aggregation. Vernonia condensata (Asteraceae) is known to attract species of leafhoppers, most of them important vectors of the citrus variegated chlorosis (CVC). In this study, we evaluated the role of volatiles of V. condensata on the orientation of Bucephalogonia xanthophis (Hemiptera: Cicadellidae). Four-arm olfactometer bioassays showed that only males were attracted to the volatiles of the host-plants Citrus sp. and V. condensata. Furthermore, fresh leaves of V. condensata induced a stronger response than volatiles from hexane-extracted leaves. This study opens the possibility to utilize V. condensata volatiles for pest management programs of B. xanthopis.

Os voláteis de plantas são sinais importantes para a orientação de insetos herbívoros. É possível que esses compostos indiquem que a planta é apropriada para alimentação e desenvolvimento larval, ou para agrupamento reprodutivo. Vernonia condensata (Asteraceae) atrai espécies de cigarrinhas, muitas delas vetores da clorose variegada dos citros (CVC). Neste trabalho, foi avaliado o papel dos voláteis de V. condensata na orientação de Bucephalogonia xanthophis (Hemiptera: Cicadellidae). Bioensaios em olfatômetro de quatro braços mostraram que somente os machos foram atraídos pelos voláteis das plantas hospedeiras Citrus sp. e V. condensata, sendo que os voláteis das folhas frescas de V. condensata induziram uma maior resposta quando comparada aos extratos hexânicos da planta. A possibilidade de usar os voláteis de V. condensata abre novas perspectivas para programas e alternativas para o manejo de pragas.