Exploring the nitrogen ingestion of aphids--a new method using electrical penetration graph and (15)N labelling.

Studying plant-aphid interactions is challenging as aphid feeding is a complex process hidden in the plant tissue. Here we propose a combination of two well established methods to study nutrient acquisition by aphids focusing on the uptake of isotopically labelled nitrogen ((15)N). We combined the Electrical Penetration Graph (EPG) technique that allows detailed recording of aphid feeding behaviour and stable isotope ratio mass spectrometry (IRMS) to precisely measure the uptake of nitrogen. Bird cherry-oat aphids Rhopalosiphum padi L. (Hemiptera, Aphididae) fed for 24 h on barley plants (Hordeum vulgare L., cultivar Lina, Poaceae) that were cultivated with a (15)N enriched nutrient solution. The time aphids fed in the phloem was strongly positive correlated with their (15)N uptake. All other single behavioural phases were not correlated with (15)N enrichment in the aphids, which corroborates their classification as non-feeding EPG phases. In addition, phloem-feeding and (15)N enrichment of aphids was divided into two groups. One group spent only short time in the phloem phase and was unsuccessful in nitrogen acquisition, while the other group displayed longer phloem-feeding phases and was successful in nitrogen acquisition. This suggests that several factors such as the right feeding site, time span of feeding and individual conditions play a role for the aphids to acquire nutrients successfully. The power of this combination of methods for studying plant-aphid interactions is discussed.

Independent responses to ultraviolet radiation and herbivore attack in broccoli.

The plant responses to ultraviolet-B radiation (UV-B) and to insect herbivory are believed to be partially similar. In this study, responses to these factors were investigated in the crop species broccoli (Brassica oleracea L. convar. botrytis, Brassicaceae). Plants were first grown under three UV-B regimes (80%, 23%, and 4% transmittance of ambient UV-B) in greenhouses covered with either innovative materials (high and medium transmittance) or conventional glass (low transmittance). Half of the plants then remained under these conditions, but the other half were transferred to the field with ambient light and herbivore access for up to 3 d. The plant responses to distinct environmental conditions were examined by analysing the morphological and chemical parameters of plants kept inside and plants exposed in the field. Furthermore, suitability of field-exposed plants to naturally occurring insects was investigated in relation to UV-B pretreatment. High levels of UV-B radiation led to increased flavonoid concentrations, but to a lower biomass accumulation in broccoli. These patterns remained after outdoor exposure. However, UV-induced changes of plant traits did not alter attractiveness to herbivorous insects: thrips, whiteflies, and aphids attacked plants independently of UV-B pretreatment. A 3-fold increase of indolyl glucosinolate concentrations occurred in above-ground tissue of all the plants, most likely due to massive herbivore attack after 3 d of field exposure. The results show that plants respond with high specificity to different abiotic and biotic impacts, demonstrating the separate perception and processing of stress factors.

Revised determination of free and complexed myrosinase activities in plant extracts.

The enzyme myrosinase (thioglucoside glucohydrolase, EC 3.2.1.147, formerly EC 3.2.3.1) catalyzes the hydrolysis of glucosinolates after tissue damage in plants of the order Brassicales. The various myrosinase isoforms occur either as free soluble dimers or as insoluble complexes. We propose a reliable method for determination of both soluble and insoluble myrosinase activity concentrations in partially purified plant extracts. The procedure requires the removal of endogenous glucosinolates through ion-exchange columns previous to enzyme measurements. Myrosinase activity was assayed in continuous mode by photometric quantification of the released glucose using glucose-oxidase with peroxidase and colorimetric indicators. The measurement of the colored product at 492nm has a favorable signal to noise ratio both in clear extract solutions (free dimers) and in turbid pellet suspensions (insoluble complexes). No interferences by ascorbic acid were found in continuous analyses. With the recommended sample preparation methods and assay conditions potential activities in damaged plant tissues can be characterized which are involved in plant defense mechanisms.