scholarly journals The Colonization Process of Sunflower by a Green Fluorescent Protein-Tagged Isolate of Verticillium dahliae and its Seed Transmission

Plant Disease ◽  
2018 ◽  
Vol 102 (9) ◽  
pp. 1772-1778 ◽  
Author(s):  
Y. Zhang ◽  
J. Zhang ◽  
J. Gao ◽  
G. Zhang ◽  
Y. Yu ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Verticillium Dahliae ◽  
Fluorescent Protein ◽  
Lateral Roots ◽  
Pollen Grains ◽  
Seed Transmission ◽  
Vascular Bundles ◽  
Long Distance ◽  
Vascular Elements ◽  
Green Fluorescent

Sunflower Verticillium wilt is a widespread and destructive disease caused by the soilborne pathogen Verticillium dahliae. To better understand the process of infection and seed transmission of the fungus, sunflower roots were inoculated with a V. dahliae strain (VdBM9-6) labeled with green fluorescent protein (GFP) and monitored microscopically. After 24 to 96 h postinoculation (hpi), conidia germinated and developed into mycelium on root hairs, elongation zones, and caps of lateral roots. Mycelium colonized vascular bundles of lateral roots and taproots at 7 days postinoculation (dpi). At 10 weeks postinoculation (wpi), the epidermal cells, cortical tissues, and vascular elements of stem, petiole, and leaf veins were colonized by mycelium. By 12 wpi, strong GFP signals were detected not only on different tissues of inflorescence but also on testa of seed and a small fraction of pollen grains. A GFP signal was not observed on cotyledon tissues in the seed. Additionally, the colonization of V. dahliae on testa was also confirmed with MNP-10 selection medium, indicating that the testa of seed is the main carrier for the long distance transmission of sunflower yellow wilt.

scholarly journals Colonization of Resistant and Susceptible Lettuce Cultivars by a Green Fluorescent Protein-Tagged Isolate of Verticillium dahliae

2008 ◽  
Vol 98 (8) ◽  
pp. 871-885 ◽  
Author(s):  
G. E. Vallad ◽  
K. V. Subbarao
Keyword(s):  
Green Fluorescent Protein ◽  
Verticillium Dahliae ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Lateral Roots ◽  
Conidial Suspension ◽  
Root Tips ◽  
Vascular Elements ◽  
Race 1 ◽  
Green Fluorescent

Interactions between lettuce and a green fluorescent protein (GFP)-expressing, race 1 isolate of Verticillium dahliae, were studied to determine infection and colonization of lettuce cultivars resistant and susceptible to Verticillium wilt. The roots of lettuce seedlings were inoculated with a conidial suspension of the GFP-expressing isolate. Colonization was studied with the aid of laser scanning confocal and epi-fluorescence microscopes. Few differences in the initial infection and colonization of lateral roots were observed between resistant and susceptible cultivars. Hyphal colonies formed on root tips and within the root elongation zones by 5 days, leading to the colonization of cortical tissues and penetration of vascular elements regardless of the lettuce cultivar by 2 weeks. By 8 to 10 weeks after inoculation, vascular discoloration developed within the taproot and crown regions of susceptible cultivars well in advance of V. dahliae colonization. Actual foliar wilt coincided with the colonization of the taproot and crown areas and the eruption of mycelia into surrounding cortical tissues. Advance colonization of stems, pedicels, and inflorescence, including developing capitula and mature achenes was observed. Seedborne infection was limited to the maternal tissues of the achene, including the pappus, pericarp, integument, and endosperm; but the embryo was never compromised. Resistant lettuce cultivars remained free of disease symptoms. Furthermore, V. dahliae colonization never progressed beyond infected lateral roots of resistant cultivars. Results indicated that resistance in lettuce may lie with the plant's ability to shed infected lateral roots or to inhibit the systemic progress of the fungus through vascular tissues into the taproot.

scholarly journals Systemic Colonization of Potato Plants by a Soilborne, Green Fluorescent Protein-Tagged Strain of Dickeya sp. Biovar 3

2010 ◽  
Vol 100 (2) ◽  
pp. 134-142 ◽  
Author(s):  
Robert Czajkowski ◽  
Waldo J. de Boer ◽  
Henk Velvis ◽  
Jan M. van der Wolf
Keyword(s):  
Green Fluorescent Protein ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Agar Medium ◽  
Lateral Roots ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Potato Plants ◽  
Soil Inoculation ◽  
Green Fluorescent

Colonization of potato plants by soilborne, green fluorescent protein (GFP)-tagged Dickeya sp. IPO2254 was investigated by selective plating, epifluorescence stereo microscopy (ESM), and confocal laser scanning microscopy (CLSM). Replicated experiments were carried out in a greenhouse using plants with an intact root system and plants from which ca. 30% of the lateral roots was removed. One day after soil inoculation, adherence of the pathogen on the roots and the internal colonization of the plants were detected using ESM and CLSM of plant parts embedded in an agar medium. Fifteen days post-soil inoculation, Dickeya sp. was found on average inside 42% of the roots, 13% of the stems, and 13% of the stolons in plants with undamaged roots. At the same time-point, in plants with damaged roots, Dickeya sp. was found inside 50% of the roots, 25% of the stems, and 25% of the stolons. Thirty days postinoculation, some plants showed true blackleg symptoms. In roots, Dickeya sp. was detected in parenchyma cells of the cortex, both inter- and intracellularly. In stems, bacteria were found in xylem vessels and in protoxylem cells. Microscopical observations were confirmed by dilution spread-plating the plant extracts onto agar medium directly after harvest. The implications of infection from soilborne inoculum are discussed.

scholarly journals Human Cytomegalovirus Labeled with Green Fluorescent Protein for Live Analysis of Intracellular Particle Movements

2005 ◽  
Vol 79 (5) ◽  
pp. 2754-2767 ◽  
Author(s):  
Kerstin Laib Sampaio ◽  
Yolaine Cavignac ◽  
York-Dieter Stierhof ◽  
Christian Sinzger
Keyword(s):  
Green Fluorescent Protein ◽  
Human Cytomegalovirus ◽  
Fluorescent Protein ◽  
Rare Event ◽  
Single Step ◽  
Wild Type Virus ◽  
Cell Cortex ◽  
Long Distance ◽  
Infected Cells ◽  
Green Fluorescent

ABSTRACT Human cytomegalovirus (HCMV) replicates in the nuclei of infected cells. Successful replication therefore depends on particle movements between the cell cortex and nucleus during entry and egress. To visualize HCMV particles in living cells, we have generated a recombinant HCMV expressing enhanced green fluorescent protein (EGFP) fused to the C terminus of the capsid-associated tegument protein pUL32 (pp150). The resulting UL32-EGFP-HCMV was analyzed by immunofluorescence, electron microscopy, immunoblotting, confocal microscopy, and time-lapse microscopy to evaluate the growth properties of this virus and the dynamics of particle movements. UL32-EGFP-HCMV replicated similarly to wild-type virus in fibroblast cultures. Green fluorescent virus particles were released from infected cells. The fluorescence stayed associated with particles during viral entry, and fluorescent progeny particles appeared in the nucleus at 44 h after infection. Surprisingly, strict colocalization of pUL32 and the major capsid protein pUL86 within nuclear inclusions indicated that incorporation of pUL32 into nascent HCMV particles occurred simultaneously with or immediately after assembly of the capsid. A slow transport of nuclear particles towards the nuclear margin was demonstrated. Within the cytoplasm, most particles performed irregular short-distance movements, while a smaller fraction of particles performed centripetal and centrifugal long-distance movements. Although numerous particles accumulated in the cytoplasm, release of particles from infected cells was a rare event, consistent with a release rate of about 1 infectious unit per h per cell in HCMV-infected fibroblasts as calculated from single-step growth curves. UL32-EGFP-HCMV will be useful for further investigations into the entry, maturation, and release of this virus.

scholarly journals Dynamics of Bean Dwarf Mosaic Geminivirus Cell-to-Cell and Long-Distance Movement in Phaseolus vulgaris Revealed, Using the Green Fluorescent Protein

1998 ◽  
Vol 11 (4) ◽  
pp. 277-291 ◽  
Author(s):  
M. R. Sudarshana ◽  
H. L. Wang ◽  
W. J. Lucas ◽  
R. L. Gilbertson
Keyword(s):  
Green Fluorescent Protein ◽  
Phaseolus Vulgaris ◽  
Particle Bombardment ◽  
Fluorescent Protein ◽  
Fluorescence Analysis ◽  
Reporter System ◽  
Long Distance ◽  
Viral Dna ◽  
Green Fluorescent ◽  
Long Distance Movement

The cell-to-cell and long-distance movement of the bipartite geminivirus, bean dwarf mosaic (BDMV), in Phaseolus vulgaris plants was examined with the noninvasive reporter, the green fluorescent protein (GFP). A modified GFP gene (mGFP4) was inserted into the BDMV DNA-A component in place of the coat protein gene (BDMVA-mGFP4), and particle bombardment was used to introduce viral DNA into bean seedlings (radicle and hypocotyl tissues). Fluorescence analysis of GFP expressed from BDMVA-mGFP4 established that particle bombardment introduced viral DNA only into epidermal cells, and the requirement for the DNA-B-encoded proteins (BV1 and BC1) in the cell-to-cell movement of BDMVA-mGFP4. This GFP reporter system was used to follow the viral infection process from the seedling stage throughout the entire plant life cycle. In inoculated hypocotyls, BDMV moved from cell to cell through the cortex and showed a striking phloem tropism. Upon entry into phloem tissues, BDMV moved rapidly toward the root via the long-distance transport system, and toward the shoot apex by a combination of cell-to-cell and long-distance movement. Analysis of GFP distribution in systemically infected tissues revealed that BDMV was restricted to phloem cells in both roots and stems. In systemically infected primary and trifoliolate leaves, BDMV infected phloem cells associated with all vein orders (first through fifth), and the capacity of BDMV to exit from phloem tissue into nonphloem cells was correlated with the stage of plant development. Finally, fluorescence analysis of GFP in reproductive tissues established that BDMV infected flower, pod, and seed-coat tissues, but was excluded from the embryo.

scholarly journals Early Events in the Fusarium verticillioides-Maize Interaction Characterized by Using a Green Fluorescent Protein-Expressing Transgenic Isolate

2003 ◽  
Vol 69 (3) ◽  
pp. 1695-1701 ◽  
Author(s):  
Liat Oren ◽  
Smadar Ezrati ◽  
David Cohen ◽  
Amir Sharon
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Lateral Roots ◽  
Fusarium Verticillioides ◽  
Systemic Infection ◽  
Fungal Mycelium ◽  
Infested Soil ◽  
Disease Symptoms ◽  
Fungal Hyphae ◽  
Green Fluorescent

ABSTRACT The infection of maize by Fusarium verticillioides can result in highly variable disease symptoms ranging from asymptomatic plants to severe rotting and wilting. We produced F. verticillioides green fluorescent protein-expressing transgenic isolates and used them to characterize early events in the F. verticillioides-maize interaction that may affect later symptom appearance. Plants grown in F. verticillioides-infested soil were smaller and chlorotic. The fungus colonized all of the underground parts of a plant but was found primarily in lateral roots and mesocotyl tissue. In some mesocotyl cells, conidia were produced within 14 to 21 days after infection. Intercellular mycelium was detected, but additional cells were not infected until 21 days after planting. At 25 to 30 days after planting, the mesocotyl and main roots were heavily infected, and rotting developed in these tissues. Other tissues, including the adventitious roots and the stem, appeared to be healthy and contained only a small number of hyphae. These results imply that asymptomatic systemic infection is characterized by a mode of fungal development that includes infection of certain tissues, intercellular growth of a limited number of fungal hyphae, and reproduction of the fungus in a few cells without invasion of other cells. Development of visibly rotted tissue is associated with massive production of fungal mycelium and much less organized growth.

scholarly journals Analysis of Cell-to-Cell and Long-Distance Movement of a Bean Dwarf Mosaic Geminivirus-Green Fluorescent Protein Reporter in Host and Nonhost Species: Identification of Sites of Resistance

1999 ◽  
Vol 12 (4) ◽  
pp. 345-355 ◽  
Author(s):  
H. L. Wang ◽  
M. R. Sudarshana ◽  
R. L. Gilbertson ◽  
W. J. Lucas
Keyword(s):  
Green Fluorescent Protein ◽  
Viral Infection ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Systemic Infection ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Long Distance ◽  
Green Fluorescent ◽  
Long Distance Movement

A bean dwarf mosaic geminivirus-green fluorescent protein (BDMV-GFP) reporter system was employed to analyze the viral infection process in host and nonhost species. Five classes of BDMV/host interaction were identified: (i) adapted hosts (susceptible Phaseolus vulgaris cultivars) permissive for systemic infection; (ii) adapted hosts (resistant P. vulgaris cv. Othello) displaying the development of a hypersensitive response (HR) associated with resistance to systemic infection; (iii) adapted (resistant P. vulgaris cv. Black Turtle Soup T-39) and nonadapted (Vigna unguiculata) hosts in which cell-to-cell, but not long-distance, movement was permitted; (iv) nonadapted hosts (Glycine max) in which systemic infection was coat protein-dependent; and (v) nonhosts (Cucurbita maxima, Gossypium barbadense, and Zea mays) in which the virus was confined to inoculated cells. Confocal laser scanning microscopy, fluorescence microscopy, and histochemical analyses were used to identify the cellular distribution of BDMV-GFP and the host response to viral infection. With this approach, the HR in P. vulgaris cv. Othello was visualized within cells of the epidermis, cortex, and phloem of inoculated hypocotyls. Infection studies performed with four begomoviruses and infectious BDMV/tomato mottle geminivirus pseudorecombinants revealed that the HR determinant(s) mapped to the BDMV DNA-B component.

Colonization in cotton plants by a green fluorescent protein labelled strain of Verticillium dahliae

2012 ◽  
Vol 135 (4) ◽  
pp. 867-876 ◽  
Author(s):  
Wen-Wei Zhang ◽  
Teng-Fei Jiang ◽  
Xiao Cui ◽  
Fang-Jun Qi ◽  
Gui-Liang Jian
Keyword(s):  
Green Fluorescent Protein ◽  
Verticillium Dahliae ◽  
Fluorescent Protein ◽  
Cotton Plants ◽  
Green Fluorescent
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