Pea Aphid (Acyrthosiphon pisum) Host Races Reduce Heat-Induced Forisome Dispersion in Vicia faba and Trifolium pratense.

Although phloem-feeding insects such as aphids can cause significant damage to plants, relatively little is known about early plant defenses against these insects. As a first line of defense, legumes can stop the phloem mass flow through a conformational change in phloem proteins known as forisomes in response to Ca2+ influx. However, specialized phloem-feeding insects might be able to suppress the conformational change of forisomes and thereby prevent sieve element occlusion. To investigate this possibility, we triggered forisome dispersion through application of a local heat stimulus to the leaf tips of pea (Pisum sativum), clover (Trifolium pratense) and broad bean (Vicia faba) plants infested with different pea aphid (Acyrthosiphon pisum) host races and monitored forisome responses. Pea aphids were able to suppress forisome dispersion, but this depended on the infesting aphid host race, the plant species, and the age of the plant. Differences in the ability of aphids to suppress forisome dispersion may be explained by differences in the composition and quantity of the aphid saliva injected into the plant. Various mechanisms of how pea aphids might suppress forisome dispersion are discussed.

Species-Specific and Distance-Dependent Dispersive Behaviour of Forisomes in Different Legume Species.

Forisomes are giant fusiform protein complexes composed of sieve element occlusion (SEO) protein monomers, exclusively found in sieve elements (SEs) of legumes. Forisomes block the phloem mass flow by a Ca2+-induced conformational change (swelling and rounding). We studied the forisome reactivity in four different legume species-Medicago sativa, Pisum sativum, Trifolium pratense and Vicia faba. Depending on the species, we found direct relationships between SE diameter, forisome surface area and distance from the leaf tip, all indicative of a developmentally tuned regulation of SE diameter and forisome size. Heat-induced forisome dispersion occurred later with increasing distance from the stimulus site. T. pratense and V. faba dispersion occurred faster for forisomes with a smaller surface area. Near the stimulus site, electro potential waves (EPWs)-overlapping action (APs), and variation potentials (VPs)-were linked with high full-dispersion rates of forisomes. Distance-associated reduction of forisome reactivity was assigned to the disintegration of EPWs into APs, VPs and system potentials (SPs). Overall, APs and SPs alone were unable to induce forisome dispersion and only VPs above a critical threshold were capable of inducing forisome reactions.

The Physiological and Biochemical Effects on Napier Grass Plants Following Napier Grass Stunt Phytoplasma Infection.

Napier grass stunt (NGS) phytoplasma, a phloem-limited bacterium, infects Napier grass leading to severe yield losses in East Africa. The infected plants are strongly inhibited in growth and biomass production. In this study, phytoplasma-induced morphological changes of the vascular system and physiological changes were analyzed and compared with uninfected plants. The study showed that the phytoplasmas are more abundant in source leaves and range from 103 bacteria/µg total DNA in infected roots to 106 in mature Napier grass leaves. Using microscopical, biochemical, and physiological tools, we demonstrated that the ultrastructure of the phloem and sieve elements is severely altered in the infected plants, which results in the reduction of both the mass flow and the translocation of photoassimilates in the infected leaves. The reduced transport rate inhibits the photochemistry of photosystem II in the infected plants, which is accompanied by loss of chloroplastic pigments in response to the phytoplasma infection stress eventually resulting in yellowing of diseased plants. The phytoplasma infection stress also causes imbalances in the levels of defense-related antioxidants, glutathione, ascorbic acid, reactive oxygen species (ROS), and-in particular-hydrogen peroxide. This study shows that the infection of NGS phytoplasma in the phloem of Napier grass has an impact on the primary metabolism and activates a ROS-dependent defense response.

Measurement of Electropotential Waves in Intact Phloem Sieve Elements Using Microelectrodes.

Electropotential waves (EPW) are involved in plant responses to both abiotic and biotic stresses. Three different types of EPWs have been identified: action potential, variation potential, and system potential, all of which have been indicated to participate in phloem-based communication between plant organs. In this chapter we describe in detail how to measure EPWs in plants, including how to access the phloem, and how to insert microelectrodes. Such experiments can be used, for example, to study the local and systemic signaling in response to diverse stimuli like microbial threat or herbivore attack.

Comparison of intracellular location and stimulus reaction times of forisomes in sieve tubes of four legume species.

Forisomes in legumes are responsible for fast sieve-element occlusion in response to injury to the vascular system. This prevents uncontrolled leakage of phloem sap and protects against invasion of pathogens. Here we compared forisomes of four different legumes (Pisum sativum, Vicia faba, Trifolium pratense and Medicago sativa) by their location (basal, central, apical) in the sieve element and reactivity to a distant heat stimulus. In each species, the majority of forisomes was located basally. Yet, we found differences in intracellular location: forisomes are distributed more evenly in the sieve elements of Pisum. After burning, basally located forisomes of the four species reacted with dispersion, followed by a spontaneous recondensation with similar reaction times. The results suggest universal forisome behaviour in fabacean species.

Barley yellow dwarf virus Infection Leads to Higher Chemical Defense Signals and Lower Electrophysiological Reactions in Susceptible Compared to Tolerant Barley Genotypes.

Barley yellow dwarf virus (BYDV) is a phloem limited virus that is persistently transmitted by aphids. Due to huge yield losses in agriculture, the virus is of high economic relevance. Since the control of the virus itself is not possible, tolerant barley genotypes are considered as the most effective approach to avoid yield losses. Although several genes and quantitative trait loci are known and used in barley breeding for virus tolerance, little is known about molecular and physiological backgrounds of this trait. Therefore, we compared the anatomy and early defense responses of a virus susceptible to those of a virus-tolerant cultivar. One of the very early defense responses is the transmission of electrophysiological reactions. Electrophysiological reactions to BYDV infection might differ between susceptible and tolerant cultivars, since BYDV causes disintegration of sieve elements in susceptible cultivars. The structure of vascular bundles, xylem vessels and sieve elements was examined using microscopy. All three were significantly decreased in size in infected susceptible plants where the virus causes disintegration of sieve elements. This could be associated with an uncontrolled ion exchange between the sieve-element lumen and apoplast. Further, a reduced electrophysiological isolation would negatively affect the propagation of electrophysiological reactions. To test the influence of BYDV infection on electrophysiological reactions, electropotential waves (EPWs) induced by leaf-tip burning were recorded using aphids as bioelectrodes. EPWs in infected susceptible plants disappeared already after 10 cm in contrast to those in healthy susceptible or infected tolerant or healthy tolerant plants. Another early plant defense reaction is an increase in reactive oxygen species (ROS). Using a fluorescent dye, we found a significant increase in ROS content in infected susceptible plants but not in infected tolerant plants. Similar results were found for the phytohormones abscisic acid and three jasmonates. Salicylic acid levels were generally higher after BYDV infection compared to uninfected plants. Heat stimulation caused an increase in jasmonates. By shedding light on the plant defense mechanisms against BYDV, this study, provides further knowledge for breeding virus tolerant plants.