Accumulation Of Fluorescent Compounds Research Articles

Verticillium dahliae's VdNEP acts both as a plant defence elicitor and a pathogenicity factor in the interaction with Helianthus annuus

The soil-borne fungus Verticillium dahliae Kleb. causes a serious wilt disease in many economic crops worldwide, including sunflower. We investigated the role of V. dahliae's necrosis and ethylene-inducing proteins (VdNEP) in the interaction with sunflower. We inoculated two highly and two weakly aggressive V. dahliae isolates onto moderately resistant and susceptible sunflower hybrids. We also synthesized VdNEP protein in vitro and infiltrated it into sunflower control plants to compare its effects with those of the fungus. VdNEP induced wilting symptoms, i.e. chlorosis, necrosis and vascular discoloration but also triggered host defence responses, i.e. hypersensitive response in Nicotiana benthamiana leaves and sunflower cotyledons. It activated the production of reactive oxygen species and the accumulation of fluorescent compounds in sunflower leaves, as well as pathogenesis-related genes (Ha-PR-3 and Ha-PR-5), two defensin genes (Ha-PDF and Ha-CUA1) and those encoding Ha-ACO, Ha-CHOX, Ha-GST and Ha-SCO. The latter suggested that more than one signalling pathway may be involved in the V. dahliae-sunflower interaction. Two other genes (Ha-PAL and Ha-NML1), related to the salicylic acid pathway, were slightly downregulated by VdNEP, suggesting a possible involvement of VdNEP in affecting sunflower defences. The phenotypic changes induced by VdNEP indicate that this protein acts both as a defence elicitor and a pathogenicity factor.

Host and Nonhost Resistance in Medicago–Colletotrichum Interactions

Medicago truncatula lines resistant (A17) or susceptible (F83005.5) to the alfalfa pathogen Colletotrichum trifolii were used to compare defense reactions induced upon inoculation with C. trifolii or with the nonadapted pathogens C. lindemuthianum and C. higginsianum. Nonadapted Colletotrichum spp. induced a hypersensitive response (HR)-like reaction similar to the one induced during the host-incompatible interaction. Molecular analyses indicated an induction of PR10 and chalcone synthase genes in host and nonhost interactions but delayed responses were observed in the F83005.5 line. The clste12 penetration-deficient C. lindemuthianum mutant induced an HR and defense gene expression, showing that perception of nonadapted strains occurs before penetration of epidermal cells. Cytological and transcriptomic analyses performed upon inoculation of near-isogenic M. truncatula lines, differing only at the C. trifolii resistance locus, Ct1, with the nonadapted Colletotrichum strain, showed that nonhost responses are similar in the two lines. These included a localized oxidative burst, accumulation of fluorescent compounds, and transient expression of a small number of genes. Host interactions were characterized by a group of defense and signaling-related genes induced at 3 days postinoculation, associated with an accumulation of salicylic acid. Together, these results show that M. truncatula displays a rapid and transient response to nonadapted Colletotrichum strains and that this response is not linked to the C. trifolii resistance locus.

PHENOTYPIC CHARACTERIZATION OF THE Rvi15 (Vr2) APPLE SCAB RESISTANCE

Recently, a new apple scab resistance was found in the apple accession GMAL 2473 and named Rvi15 (Vr2). The resistance reaction mediated by Rvi15 (Vr2) appeared on the upper surface of young infected leaves 7 days post inoculation (dpi) as browning of epidermal cells. Starting from 11 dpi, browning of mesophyll cells was also observed suggesting a gradual involvement of the different leaf tissues. The resistance reaction resulted in pinpoint pits similar to those observed in Malus accessions carrying the Rvi5 (Vm) or Rvi4 (Vh4) resistance, which are considered to induce the classical hypersensitive response (symptoms appearing at both epidermal and mesophyll level 2-3 dpi). When the timing of the reaction is considered, Rvi15 (Vr2) seems more similar to the Rvi7 (Vfh) resistance found in Malus floribunda 821, which results in pinpoint pits 6 dpi. However, size of the pinpoint pits, number of cells involved and accumulation of fluorescent compounds at the site of pathogen penetration are very similar between all four resistance reactions.

Age estimation of Mexican fruit fly (Diptera: Tephritidae) based on accumulation of pterins.

A common method of aging adult flies, fluorescence spectrometry, was used to monitor the increase of overall pterine titer in head extracts of Anastrepha ludens (Loew). Accumulation of fluorescent compounds was measured as a function of chronological age of flies maintained at 17 and 27 degrees C. Although relative fluorescence increased with age, field studies revealed that this phenomenon could not be used for accurate age estimation, as relative fluorescence did not increase predictably with age over the entire life span. Accumulation of individual pterins, deoxysepiapterin and sepiapterin, were studied in a similar manner. These two specific compounds were separated by high-pressure liquid chromatography and their accumulation was followed at 15 and 30 degrees C in the laboratory and under caged field conditions. While titer of deoxysepiapterin increased steadily in a curvilinear fashion, sepiapterin quickly reached a maximum and then maintained a constant level for the rest of the life of the flies. Based on the physiological response of deoxysepiapterin to chronological time and ambient thermal conditions, this compound was determined to be an age specific biological parameter for the Mexican fruit fly and should allow age estimation in field-collected flies.

The fluorescence assay for Maillard product accumulation does not correlate with seed viability

AbstractA recent hypothesis states that the accumulation of amino-carbonyl reaction products of reducing sugars with proteins (Maillard products) is related to the loss of seed vigour and viability during ageing. Since Maillard products are biochemical end products, their accumulation should provide an index of seed deterioration. A simple fluorescence assay has been used to estimate the quantity of Maillard products in aqueous seed extracts. We tested whether an accumulation of fluorescent compounds correlates with the loss of seed viability during ageing. Fluorescence of extracts from lettuce (Lactuca sativa) embryonic axes increased after controlled deterioration, but was not correlated with ageing period. Fluorescence of extracts from whole broccoli (Brassica oleracea var. italica) seeds was either unchanged or declined during controlled deterioration. Extracts from naturally aged whole seeds of carrot (Daucus carota), tomato (Lycopersicon esculentum), and cauliflower (B. oleracea var. botrytis) had higher fluorescence than controls, but there was no consistent increase in fluorescence in proportion to the loss of viability. Extracts of naturally aged onion (Allium cepa) seeds exhibited less fluorescence than extracts from seeds with higher viability. We conclude that the fluorescence assay does not have general utility as an index of seed deterioration.

Changes in fluorescent lipid peroxidation products of room-ripened and vine-ripened tomato fruits in relation to other ripening parameters

The changes in ripening parameters and in lipid peroxidation products such as aldehydes, malondialdehyde (MDA), and fluorescent compounds (FCs), were examined at different maturity stages of tomato fruits during ripening on the vine. These changes were compared with the corresponding changes occurring in room-ripened tomato fruits, harvested at the mature green (MG) stage and stored at 12 or 22°C. Some of the harvested fruits were treated with 600 μl l −1 ethylene for 2 days prior to their storage at 22°C. Except for aldehydes and MDA, all changes in the other parameters observed with vine-ripened fruits, were enhanced in room-ripened fruits stored at 22°C or treated with ethylene. However, in contrast with fruits stored at 22°C, the ripening pattern of fruits stored at 12°C resembled that of vine-ripened fruits. These results suggest that the slow ripening rate during storage at 12°C reflects the rates of natural ripening occurring on the vine. Among the lipid peroxidative processes assayed, accumulation of FCs proved to be a more sensitive parameter than accumulation of aldehydes and MDA for assessing the extent of lipid oxidation during tomato fruit ripening.

Ethylene-increased Accumulation of Fluorescent Lipid-peroxidation Products Detected during Senescence of Parsley by a Newly Developed Method

A newly developed rapid and convenient method was used for fractionation and analysis of fluorescent compounds (FCs) formed during lipid peroxidation in parsley (Petroselinum crispum Mill.) leaves. These lipofuscin-like FCs [which arise in vivo from reaction of malondialdehyde (MDA) with amino acids] were found to increase during the senescence of detached parsley leaves, following the commencement of chlorophyll degradation and proteolysis. However, accumulation of FCs in response to exogenous ethylene coincided with the onset of chlorophyll loss and proteolysis on day 2 and was accelerated markedly later. Unlike FC accumulation, levels of aldehydes and MDA in control leaves increased more drastically during senescence, but were not affected significantly by exogenous ethylene. The results suggest that the accumulation of FCs in detached parsley leaves exposed to exogenous ethylene is an early senescence-associated process.

Increased Formation of Fluorescent Lipid-peroxidation Products in Avocado Peels Precedes Other Signs of Ripening

Fluorescent products (lipofuscin-like compounds) of lipid peroxidation, which accumulate with age, were extracted from `Fuerte' avocado (Persea americana Mill.) peels during ripening. Fractionation and analysis of these fluorescent compounds (FCs) was carried out by an improved method, based on separation of FCs from-chlorophyll by Sep-Pak silica cartridges. A sharp rise in FCs content was found 2 days after harvest in avocado fruits stored at 22C, and ethylene enhanced this rise 3-fold on the 4th day. The accumulation of FCs preceded by at leasts days the onset of climacteric ethylene and respiration and by 2 days the decrease in fruit firmness. Moreover, a 6-foId increase in the FCs concentration occurred during 1 to 2 weeks of storage at SC, but the avocado fruits did not show any other detectable signs of ripening. These results suggest that lipid peroxidation may be regarded as one of the earliest detectable processes occurring during fruit ripening. Thus, an increase of FCs in peel may be employed as a horticultural characteristic for estimating initiation of ripening in avocado fruit.

Microscopic Studies of the Interaction Between Barley and the Saprophytic Fungus, Cladosporium macrocarpum

Abstract Scanning electron microscopy of the saprophytic fungus Cladosporium macrocarpum on barley leaves revealed characteristic bar‐shaped appressoria. They were almost always located either above a stomatal opening or above the anticlinal cell walls. It was shown with light microscopy that the fungus was able to penetrate via stomatal openings. No successful direct penetrations were observed. The barley tissue responded to direct penetration attempts by formation of papillae and accumulation of fluorescent compounds. Fluorescence microscopy revealed fluorescent compounds in papillae, haloes, hypersensitively reacting epidermal cells, mesophyll cells, and in stomatal areas. The induced resistance reactions are similar to those described earlier for, pathogenic fungi, indicating that they are part of general resistance phenomena.