Verticilliun dahliae Verticillium and other species - Wilt ..., (Verticillium) - Molecular diagnosis quantitative and quantitative (PCR and QRTPCR) and molecular identification (sequencing).
Verticillium species of fungi are Ascomycetes class, which can be saprophytes and parasites of higher plants, insects, nematodes, mollusks, etc. They have differentiated three ecological groups: mycopathogens, entomopathogenic and phytopathogenic. Ente phytopathogenic included to Verticillium dahliae, V. longisporum, V. albo-atrum, V. and V. nubilum tricorpus. Of these, the best known are the first three cited, causing the blighted by Verticillium. This disease affects more than 350 plant species, including strawberries, tomatoes, potatoes, peppers, eggplants, artichokes, sunflowers, strawberries (Fragaria x ananassa), watermelons, lettuce, cauliflower, radishes, cotton, and many trees, such as olive in the Mediterranean area, stone fruits (avocado, plum, cherry, ...), with the exception of citrus and apple trees.
Verticillium species are very diverse, though their life cycle is similar. These species develop survival structures varies in some species. As an example:
- Alboatrum Verticillium is characterized by developing a melaninizado mycelium, which can remain viable for up to 4 years in the soil without susceptible plants where the fungus could be introduced multiply.
- Verticillium dahliae, is characterized by forming microsclerocios, which can remain viable 12 or 15 years in the soil, in the absence of a susceptible host in which they can infect and multiply again. Microsclerotia (microsclerotia) are formed by a mass of fungal mycelium surrounded by tissue rich in melanin that protects against ultraviolet radiation.
- Verticillium Verticillium nubilum negrescens and develop clamidosporas.
- Verticilum tricorpus, can develop any of the above three formations: melaninizado mycelium and chlamydospores Microsclerotia.
When there next roots (about 2 mm) to one of survival structures, the root exudate promotes germination survival structure and the fungus grows into the plant, reaching the internal vascular system of the plant, for distributed throughout the plant. Penetration is facilitated by the existence of cuts or abrasions on the roots, made frequent them as a result of the action of arthropods, nematodes, kitchen utensils , farming, plowing, etc.
Once inside the plant, the fungus is directed to the vascular system, and specifically the xylem, the vascular conduits lignified. By the vascular conduits of the plant it is distributed as hyphae, mycelia and forming colonies which the conidia, which when released, are carried by the sap, new mycelia formed in other locations are generated. Verticillium conidia on conidiophores are generated arranged vertices (whorls). When the plant dies, shaped structures of each species to be found where the plant has fallen survival, inoculating the dump. This fungus can infect the seeds of plants, which can survive at least 13 months, so it has been recommended seed treatment.
Affectation occurs because the leaves curl and discolor and can lead to death of the plant. Wilting is due to blocking xylem, vascular lignified plant tissues leading gross wise from roots to leaves. In small plants and seedlings, Verticillium spp. highly lethal while in older plants, the severity varies. Sometimes only it affects one part of the plant, because once introduced in vascular tissues, migrates upward and not radially. Other symptoms include stunting, chlorosis and leaf yellowing, necrosis and defoliation tissue. In the trees it can be located in some branches.
The spread of this fungus occurs through air currents, irrigation water, agricultural implements, or insects.
It is essential to detect and identify early, quickly and specifically to establish adequate control.
Among the proposals and possible actions include:
- Deep plowing.
- Crop rotation to introduce resistant crops, and not to plant susceptible species in soils where previously the disease is detected.
- Using modified to make them resistant plants.
- Wilted branches and remove diseased trees.
- Seed treatment.
- Proper irrigation control.
- Balanced fertilization, avoiding excessive nitrogen or lack of potassium.
- Weed control that can multiply the fungus, and soil nematodes.
- Disinfection axes, saws with bleach.
- Treatment of land, perhaps the least economic measure.
Possible detection tests
The detection of the fungus in soil or plants affected may be performed by conventional methods or molecular methods.
The conventional method used has been growing in semi -selective media. However , these conventional methods require time and taxonomic expertise, and count colonies of Verticillium dahliae after a few weeks of incubation. Culturing encounters the drawback that it is difficult or impossible, Verticillium dahliae Verticillium differentiate longisporum and sometimes tricorpus Verticillium.
Molecular methods may be qualitative (species specific PCR Verticillium), or quantitative (real - time quantitative PCR) -qrtPCR-). Molecular tests avoid the problems inherent in conventional cultivation and identification, and also are more specific and faster.
Tests conducted at the IVAMI
- Culture isolation: it can be done but we advise against by requiring several days, both for development and for microscopic identification, as well as the difficulty in differentiating between some species.
- Detection and molecular identification: recommended for PCR methods for species specific genomic sequences, confirmed by sequencing. Useful to corroborate the disease in a plant, and to know its presence on land.
- Molecular quantitative detection: recommended when desired to know the density of the fungus in a soil.
- Sick branch fragment about 1 cm ..
- Ground (earth), 5 grams Octoberfest.
- Introduced into sterile plastic container as possible be cooled (white- polystyrene container -corcho with -frigolines- pack frozen).
- Quantitative cultures in selective medium: 14 days.
- 24 to 48 hours for qualitative PCR: molecular methods.
- 48 to 72 hours for quantitative PCR (QRTPCR): molecular methods.
Cost of the test: