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Fungal Plant Pathology - Science topic

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I was wondering for how much time does a mycelium freshly isolated from plant tissue continue to be "similar to what it used to be" in nature. I can see that morphological changes frequently occur when transferring from plate to plate so in short, my question is: can I keep using a mycelium for a long time, making sure it still is the same? Should I re-isolate it from time to time? And, are there some techniques to prevent changes to happen?
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Dear Alessndro,
It depends on the type of fungus. Generally, you must not preserve the fungus with this method for a long time. You can use another methods for preserving the fungi for a long time such as glycerol 10%, lyophilization, liquid nitrogen.
Regards
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There are many Alternaria pathogens that produce host-specific toxins. Alternaria host-specific toxins are classified in three groups in terms of the primary site action. First group of toxins have in common an epoxy-decatrienoic acid structure and exert their primary effect on the plasma membrane of susceptible cells. The second group is represented by ACR(L)-toxin, which induces changes in mitochondria, including swelling, vesiculation of cristae, decrease in the electron density of the matrix, increase in the rate of NADH oxidation, and inhibition of malate oxidation. The third group consists of AM-toxin, which appears to exert an early effect on both chloroplasts and plasma membranes.  
Ames and bacterial assays has demonstrated that Altertoxins I, II,  III, toxins AOH and AME  were mutagenic. A.N. Samokhvalov ( «A method for producing mutant strains of plant pathogen X. campesrtis». Invention certificate  No 1473360 of 15.12.1988 (USSR) has found that Alternaria brassicola caused similar mutation in different strains of X. campestris affected xanthomonadin synthesis and virulence of  the bacteria.
Is there any information about specific interaction between Alternaria toxins and bacterial/chloroplast or mitochondrial DNA?
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Schrader, T. J., et al. "Further examination of the effects of nitrosylation on Alternaria alternata mycotoxin mutagenicity in vitro." Mutation Research/Genetic Toxicology and Environmental Mutagenesis 606.1-2 (2006): 61-71.
... To examine other targets for mutation, the metabolites Altertoxin I (ATX I), Altenuene (ALT), Alternariol (AOH), Alternariol monomethyl ether (AME), Tentoxin (TENT), Tenuazonic acid (TA) and Radicinin (RAD) were reexamined ± nitrosylation, using Ames Salmonella strain TA97, sensitive to frameshift mutations at a run of C's, as well as strains TA102 and TA104, reverted by base pair mutations at AT sites and more sensitive to oxidative damage. ... These results suggest ATX I, AME and AOH induce mutations at AT sites, possibly through oxidative damage, with nitrosylation enhancing ATX I frameshift mutagenicity at runs of C's. Nitrosylated ATX I was also directly mutagenic in mammalian test systems.
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The type of media and the condition
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I know it would be so late answered.
However, I set a method that supports maximum sporulation while do not decrease the infection percentage.
I move A. solani to new PDA media (pH 5.5) for five days at 12/12 light/dark and 28 C°.
After that I removed surface mysilium gently and keep the plate (without cover) under black light for 8 hours and 48 dark place.
Finally, I collected sporue by dissolving the plate in water.
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I am currently trying to get the Ceratocystis sp. using carrot baiting, there is present of fruiting body on the carrot but after I try to transfer the spore to the MEA, it failed because instead of Cerato there is bacteria growth. I had tried many times and I also tried to add penicillin in my MEA but it seems to have failed again. So, what is the technique for making sure that the spores is successfully transferred to MEA ?
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The medium PCA (Potato Carrot Agar) which consists of 20 gm potato, 20 gm carrot and 15-20 gm agar is more suitable to produce spores successfully.
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Are powdery mildew and downy mildew vascular diseases?
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Both of powdery and downy mildew diseases are caused by obligate parasite fungi, but they are not considered as vascular diseases. The first disease is found on the surface of the leaves (on both parts of the leaf, upper and lower), while the second disease is caused by endogenously parasite fungi (intercellular or intracellular).
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sterile control of a mineral substrat for plants
grow medium: sabouraud agar
sample 2
tia
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In the picture above, the grayish brown mycelium represents the fungus Rhizopus. While the white and greenish blue colonies represent the fungus Penicillium.
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steril control of a mineral substrats for plants
grow medium: sabouraud agar
sample 8
tia
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The above micrograph shows penicillium spp.
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sterile control of a mineral substrate for plants
grow medium: sabouraud agar
tia
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The colony which limited with yellow marker with white mycelium may be belong to the fungus Beauveria bassiana, but not exactly. It needs more images to clearly identified.
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The crop hosts and other plant hosts of Rhizoctonia solani AG3.
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and Tobacco
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Thanks in advance.
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I realise this reply is very late down the line, but did you have much success?
I am looking to isolate fungal dna and also barcode for any interacting metazoan dna.
Thank you in advance,
Kirsty.
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I am looking for few common tobacco pathogens as well as other general plant fungal pathogens. If anyone will be willing to provide it then please contact me. 
Thanks
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Lot of work has been done by Tobacco Research Centre at Nipani, Dist-Belgavi, Karnatak state(India). You may contact Director of the Research centre .
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Dear All,
I'm still relatively new to genetic diversity analysis of plant pathogens. I'd like to ask for suggestions on how to produce a dendrogram consisting of a combination of several genetic markers using the NTSYS software.
I'm using SSR and ISSR markers to analyse the diversity of various isolates of Fusarium wilt pathogen. I'm confident of generating a data matrix using a single genetic marker (using binary codes) and then producing a dendrogram. However, I'm not entirely sure how the data matrix would look like when more than one genetic markers are used.
May I ask for your suggestions on how a data matrix consisting of more than one genetic markers would look like?
Also, is it necessary to test the combinability of data sets from different genetic markers using partition homogeneity test using PAUP software? Does anyone have any experience in handling such test using this software?
Thank you in advance.
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Binary data matrix of two different markers has to be combined into one input file. Genelyx software is much easier to analyse. After getting result of Nei's genetic diversity of populations, u can transfer the data to mega for dendrogram construction.
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When I was studying the feeding regime of the collembolan Pseudosinella alba, I reared it on a variety of fungal strains I had previously isolated from a black rendzina soil. This species exhibited clear preferences for some strains but was able to feed and reproduce on the majority of them, including the well-known pathogenic fungi Verticillium dahliae. The paper (in French) is here:
Pseudosinella alba lives in litter and feeds exclusively (and abundantly) on micro-fungi, both mycelia and spores, the germination of which becomes negligible once gut transit is completed and faeces are deposited. This property, scarce in Collembola (known for their opportunist feeding regime), is shared by other members of the genera Pseudosinella and Willemia. In this article I suggested that such species could be used for preventing pathogenic fungi to invade greenhouses and hydroponics, where pesticides are still currently used for the protection of cultures. Several attempts have been made by various authors to use springtails for the biological control of pathogenic fungi, but to my knowledge they are still not employed at this usage. However, these animals can be easily grown in boxes filled with plaster of Paris and fed with baker’s yeast, where their populations can reach very high densities providing they are fed ad libitum, as this is currently achieved in academic laboratories.
Catch as catch can…
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@Xin Zhang
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Hi everyone,
I'm firstly interested in identifying Alternaria cultures to species level. After this, I will have a look at a specific pathotype. In this regard, would it be sufficient to use the ITS region for the initial identification to species level? I'll be happy if it can just tell me if a specific culture is Alternaria alternate or not. I appreciate your time.
Thanks,
Pieter
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ITS region along with Inter simple sequence repeat markers (ISSR) can be used to identify and differentiate between Alternaria sp. Just sequencing the ITS region and comparing from NCBI database (blast) can also be done for identification.
You can refer the following publications.
1. Molecular identification and genetic variation of Alternaria species isolated from tomatoes using ITS1 sequencing and inter simple sequence repeat methods (Mohammadi A and Seifollah B, 2019).
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I have isolated about 20 different kinds of Fungals from the fruit peel and now I would like to measure the spore concentration,
1- How can prepare the spore suspension solution for spore counting, as not sure which kind of fungi?
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Marta Filipa Simões Thanks a lot ,
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Many efforts have been done by the researchers to culture the fungus extracted from Taxus plants in different mediums but the success rate was less. Hence, the Taxus plants remains the only source for paclitaxel extracts which is important for preparation of Taxol used in diagnosing cancer patients.
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Please see the following RG link.
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Macrophomina phaseolina is a fungal pathogen with a host range of more than 500 plant families; inciting a stem cancer disease in many crops that is often referred to as charcoal rot, due to the charcoal type coloration imparted to the colonized plant tissues.
How effective is biological control among other management strategies for Macrophomina??
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.
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Please have a look at this useful RG link.
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our isolate has stopped sporulation .What is the easiest way to revive it?
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Van Leeuwen, G., Hobb, I, and M. Jeger. 2002. Factors affecting mummification. Mol. Plant Path. Open Access December 20, 2002. Prepare you nectarine dextrose agar using diced nectarines that are cooked and strained. Use half the dextrose of PDA and agar to your solification need. Hope the information is useful. In the Dutch study the low temperature favoring sporulation was apparent this is outside the normal media growth temperatures. UV light can be very important as Alex noted.
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Dear researchers,
What are the black structures that appear on Botrytis cinerea mycellium after two weeks of in vitro culture??
I tried to observe them under binocular microscope and I think that they are fruiting bodies but I am not sure.
Can anyone cofirm this ??
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Hi, These are sclerotia , The fungus produces highly resistant sclerotia as survival structures in older cultures.
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I have tried aniline blue and cotton blue staining for root sections. They're not effective, in this case.
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wheat germ agglutinin (WGA) staining
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Dear researchers,
During the field harvest, I've managed to collect infected oak leaves with fungal hyphae, without any visual cues to identify the fungal species. I'm worried that I might confuse other species with actual pathogen if isolation failed.
Is there any clever way to boost the probability to isolate this unknown fungus solely? (weather condition were rainy / humid / 20'C to 30'C recently)
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To Jawad's point, I think you;'ll need to develop a reproduction of the disease in the lab so you can identify the etiologic agent(s).   Think of Koch's postulates..
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Hello, I want to preserve wheat blast fungus in glycerol for long term storage. However, I do not know the procedure for this particular fungus. What should be the concentration of glycerol? Do I need to make conidial suspension and then store it? or What is the whole procedure? please give me some suggestions.
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I am interested to study the genetic diversity of a plant fungal pathogen. I have gathered potential genetic markers (around 30 sets) from journals to investigate their % polymorphism before selecting the suitable ones for further analysis. I have a total of 50 isolates of fungal pathogen. Do I have to screen 50 isolates at one time using one set of genetic marker? 
In other words, can I just use 10 representative isolates, instead of 50 isolates, to screen for suitable genetic markers with high % of polymorphism? How many bands is needed to produce reliable genetic diversity results?
Thank you in advance.
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I remembered when I did my SSR screening, my supervisor asked me to choose 5 individuals from each population, in order to test the polymorphism of a specific marker. So i guess using 10 isolates out of 50 should be enough. For SSR, I guess markers showing more than 3 genotypes in your population would be good. Hope this helps.
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I am interested in the life cycle of the dinoderus minutus as it consumes cut bamboo.  It's digestion is dependent on symbiotic fungus.
I notice the bugs seem to have specific preferences for which bamboo it will infest first.
Are there known resources about what kinds of fungus are naturally included in bamboo?  Are the on the external surface of the bamboo, or are they already incorporated into the bamboo wood? 
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Thank you for these resources.  Fungi associated with roots of bamboo, as well as other plants is an interesting topic with obvious economic benefits.
My interest is in fungi that may be in the arboreal portions of the bamboo culm rather than the roots, as the dioderus is not known to eat the roots.
According to the 2005 PhD dissertation of Carlos M. Grarcia at Oregon State U, the gut of the Dinoderus contained several species of fungus, notably mucur and sporothrix.
Currently, I am the world's only dust farmer, making Bamboo Snow (bamboosnow.com). It is the frass of the dinoderus, and has many unique and useful properties.
Attached is a photo of an artificial rock formulated with Bamboo Snow that absorbs so much water from the air that it spontaneously weeps.  It could permanently and continuously hold and add moisture to dry and barren soil. Instant oasis?
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Olive (Olea europaea L.) is one of the most important crops in the Mediterranean countries.  Olive leaves, available throughout the year, are one of the byproducts of olive farming; they accumulate during the pruning of the olive trees and can be found in large amounts in olive oil industries after being separated from fruits before processing (about 10% of the weight of olives). Several reports have shown that olive leaves have antioxidant activity and anti-fungal properties, So, can be used dry leaves powder  as protectants against  phytopathogenic.
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green leaves are used as organic amendment which have shown nematicidal activity. Dry leaf powder is also rich in polyphenols, flavonols and glucosides are active secondary metabolite, these are present in dry powder also, so you can try..
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My lab is drafting a field trial in mildew resistance. Can someone suggest a good metered dose sprayer that would allow uniform area and fungal spore count onto a leaf surface?
We have considered a paint sprayer, but the lack of a 1-pump spray lead to variation in the amount of spores sprayed between people.
Additionally, nebulizers do not allow particle sizes between 15-30 microns, and are thus too small.
We have considered a nasal sprayer, but that shoots more of a spray over a uniform mist.
I cannot find any purchasable and empty metered dose inhalers that could be used for this online.
Thanks so much and I appreciate any help!
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We usually measure spore concentration and volume of inoculum and then spray all from the sprayer. Suitable concentration and sufficient (high) volume will make the inoculation quite uniform. In that way, we can use the cheapest sprayer bottles we want. Would this be suitable solution for you?
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Hi all,
I am trying to extract DNA from Alternaria spores. They prove to be very tough to open. Does anyone have a lab protocol they are using to obtain a high yield of Alternaria spore DNA? Currently, I am using the E.Z.N.A. Universal Pathogen Kit and bead milling with a range of bead sizes from 0.1-0.5 mm. I am thinking of trying a pre-DNA extraction step of incubating with lyticase overnight at 30C, or trying a freeze thaw step with LN2.
Any help would be much appreciated!
Cheers,
Melissa
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If you want to quantify fungal spores, why do you not count them with an optical microscope as real spores from a known volume and with a grid instead of with the more expensive and laborious technique of qPCR? DNA methods are not the answer to everything. Simple is best.
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I am currently establishing protocols for identification of plant pathogenic species by PCR. I tried primer pair BC108/BC563 published in 2006 by Rigottii and collegues, but it does not work with me (while I get excellent amplification with ITS1/ITS4 primer pair).  Any suggestions or any experiences with BC108/BC563 ?
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Hei,
To obtain a sensitive and specific assay, I would recommend the use of a primer/probe -based real-time PCR assay. It is probably very challenging to use the ITS rDNA gene cluster to design a specific assay for Botrytis cinerea. One option is to use betatubulin -see e.g. New Phytol (2005) 168: 465-474.
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The objective is to study the genetic variations of atoxigenic A.flavus strains using VCG method,
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Thanks my brother Masoud Latifan
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I’ve tried to isolating Phytophthora of avocado soil or any Solanaceae soil with different media culture like V8 or Carrot media, or even potato media and ornamental leaves tramp but not success yet, ¿does anybody know the best way to isolate this oomycete from soil?
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Dear Hyacint  Mtz
,I think that the best method to isolate Phytophtora from soil is to use apple fruit trap. In order to get that you have to chose apple fruit, wash it carfully, then sterilize it in suitable solution, wash it again in sterilized distilled water then immersed it until its half.  Irrigate the soil and leave the fruit till appearance white mycelium in area of the fruit between soil and air, then you can isolate   on suitable medium.
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I am working with pathogenic oomycetes in grasses which are asymptomatic and rarely produce sporangia growing out from the host tissue. The grass samples were fixed in 5% Glutaraldehyde in PBS (pH 7.5) and thus difficult to proceed with oomycete-specific probes for FISH (GA creates a very strong fluorescent background that even my negative controls reacts with the different filters). Apparently, I tried using multiple fluorescent stains and combinations (Aniline Blue, Trypan Blue, and Uvitex2b) although these stains also binds to chitin. The problem is the host plant may have both fungal and oomycete pathogens together. Likewise, it is already sure that my target parasite is present since specific primers were designed (sequences obtained were 99-100% identical and forms a clade with the same species with >95% support by RaxML) and the samples for microscopy where derived from the same batch. I had also considered morphological aspects like the presence and absence of septation, but reports of septation in the hyphae of oomycetes had been widely observed in different genera (Plasmopara, Peronospora, etc). Trying chitin degradation had been an option too, but the procedure is too tedious, likewise the chances of degrading cellulose and glucans of oomycete cell walls are highly possible. Considering my little amount of samples, this risk is also quite the least option for me. Sorry for the long detail, I hope someone can offer some suggestions. Thanks!
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ohhh. Thanks a lot. sorry for my delayed response. But these are really helpful. I will check them! Thanks again
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I need about 2000-5000 R. solani sclerotia for my experiments. I usually grew R. solani in Petri plates but I just get 10-12 sclerotia. It has wasted my time. I need a substrate on which I cultivate R. solani so that It may produce a lot of sclerotia for my experiments. Looking forward for your suggestions. 
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Dear colleague,
I would like to ask first about the purpose for getting high count of R. solani sclerotia?
Do you need it for pot experiment? If so, I do not prefer to increase inoculum using biotic   carrier. Simply if you want to get high amount of sclerotia you can grow the fungus in large Petri dishes ( 15 cm in diameter for at least 15 dayes then you can find high number of sclertia.
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Dear colleagues,
pathogenicity test  needs seed clear from pathogen ; but  they are mostly not available, so we some times use dusting seeds.
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I think, to avoid fungicides you have to wash the seeds with sterilized water intermittently. It would not harm the seeds as well. 
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what is the simplest method to identify fusarium wilt in banana at entry level,
is there any instant method available?  
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Dear
You may try the same as similar method we use
for Field evaluations, the symptoms on cutting longitudinal or horizontal of stem bases and incidental, variegated or dark brown color with or without scent enough to distinguish symptoms of fungal, bacterial and viruses . symptoms on the leaves and vegetative growth,  distance between internodes in plants such as chickpeas and cotton.. other crops, and we usually used a degree for resistant or tolerance to differentiate between cultivars, also the patches distribution in the fields. but in banana I did not test them in the field .
Pls. find the attached files, look at page 117,  in downloadPDF.pdf  
Pls. find the attached files
Hoping this will be helpful
Regards
Prof. Houda Kawas
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Plant apoplast contains various antimicrobial peptides e.g., defensins. As part of their antifungal mode of action, these defensins (not all defensins) are known to interact specifically with membrane-resident bioactive phospholipids, such as PA, PI(4,5)P2 with high affinity.  We know that these pathogens have different life style. Very importantly, when biotrophic pathogens thrive in the apoplst, they encounter various antifungal proteins and fungus needs to protect itself from these proteins which subsequently determine the outcome.  However, necrotrophic pathogens produce different cell wall-degrading enzymes (CWDEs) and toxins to kill the host plant cell.  When necrotrophic pathogens damage the cell wall and plasma membrane, the antimicrobial peptides (e.g., defensins) which are present in the apoplast, may encounter plant membrane-resident bioactive phospholipids. Do you think these defensins highly interact with plant phospholipids (instead of fungal membrane-resident bioactive phospholipids) and indirectly regulate its own plant lipid metabolism? If so, why does it prefer binding to its own phospholipids? Does this non-specific interaction leads to toxicity to the plant cells? What about binding to fungal membrane-resident bioactive phospholipids? Does defensin loses it ability to bind to fungal phospholipids, subsequently paving the way to infect the plant? How do transgenic (and also non-transgenic) plants over expressing phospholipid-binding proteins (e.g., defensins) control itself of these non-specific interactions?  Do you think the change in the environment (e.g., pH) can limit these non-specific interactions in both non-transgenic and overexpressing plants ?
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Dear Siva, You have a lot of very interesting questions there and I don't think they can be answered just yet without resorting to conjecture. what is known at present is that there are general reasons as to why fungal membranes are more sensitive to defensins and this will include the presence of differing sterols and the presentation of charged surfaces as well as the distribution of various lipids frim one leaflet to another. For those defensins that do have a specific interactions with e.g. PI lipids then the accessibility of these lipids will be important. You may also consider that the mechanism of action of plant defensins has no been investigated in much detail and that they may have other roles beyond a simple fungicidal activity (i.e. signalling). I think you have devised an interesting research programme with your questions and could spend many fruitful years providing a much more detailed understanding than what is currently available.
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could you please tell me what the problem is?
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thanks for your follow 
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 Hi i am looking for bioagent(fungi )active against Hyacinth water,a very destructive water weed.
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Dear Prof. Mohammed Fayyadh.
You can see the following articles.
Best Greetings
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I place some of  R. solani cultured Petriplates under fluorescent tubes and some of the Petri dishes in complete dark. The results were surprising, the color of R. solani under flouresent tubes as whitish while the color of R. solani which were placed in dark as completely dark. What is the reason? 
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Hello Aqleem, 
You observed differences morphology of the same fungi under different environmental conditions, its simply because of phenotypic plasticity of the fungi. It is the ability of a genotype to produce more than one phenotype under different environmental conditions, it may be specific or general. 
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These fungi are recovered from a Neogene wood of North-east India
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its not clear .
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I want to do a research project about host induced gene silencing, so I need to know if Arabidopsis can be infected by F.fujikuroi?
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96
Hi Sharon Hou
Can not guess like this, you can do pathogenicity test and then give your consideration
Ok, Good luck
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Hello everyone, I am interested in infecting eggplants with microsclerotia of V. dahliae and I need a clear method on how to do it. Thank you!
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Dear Jawad,
Thank you greatly! 
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What can be precise phenotypic parameters of partial resistance to stripe rust rust pathogen in wheat at seedling stage. There are numerous such latency period 1, latency peiord 50, infection type, uredinium size , infection frequency etc. Suggest two or three?
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Hi Dr.Hadi
You are welcome and good luck
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Trichoderma are promising biological bio-protectants agent against plant pathogenic fungi. Producing large number of spores and unaffected to environmental variabilities  and and have a long shelf life  are definitely required.
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Hi Jawad: There are a master thesis about formalization and spore production of Trichoderma  in the department of plant protection - University of Baghdad. I dont remember the title of the thesis but the name of the MS student is Mohammad Al-Hittar, his supervisor is Dr Farkad Al-Rawi. I think there is an electronic copy of the thesis, if you have any contact with any one in plant protection department - university of Baghdad, ask him to provide it to you. Unfortunately i am now in sabbatical leave, other whys I will send it to you.
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(a) Fungal spp. - Hypholoma spp., Phlebiopsis giganteaArmillaria ostoyae, Heterobasidion irregulare. (b) observing interaction pairings
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I need the detail list of techniques used in the screening of fusarium wilt in Musk melon and even the scale required for evaluation.
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Hi Shridhar
You can follow this paper "Effect of root feeding by striped cucumber beetle larvae on the incidence and severity of Fusarium wilt of muskmelon"
Good Luck
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how to measure the rate of growth of endophytic fungi in mangrove?
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you can measure the colony growth on the petri dish and also can check the condia growth under microscopic
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In my project which deals with how certain fungicides affect certain fungi, one of the aspects of data collection is daily spore count after the fungi has been met with fungicides and are being studied.
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First of you need to perform normality test. Based on the outcome then you will decide to transform or not. I will recommend the log transformation in case the normality test indicates that transformation is necessary.
ANOVA has assumptions and some data do not fit. In that case transformation is necessary
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Hi,
I am working with wild relatives of rice.
I have been facing an issue with the germination as the seeds are highly contaminated.
I tried to use ethanol 70% 2 minutes, bleach 4% for 30 minutes, mercury chloride 0.1% for 3 minutes.
Still, the blueish fungi appear on the seeds (in the petri dish).  
Any recommendations? maybe try to use vaccum infiltration in order to help penetrate the seed's wax? Have anybody done it before?
Many thanks in advance
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"1. When you included PronTech Floriculture (benzyl ammonium chloride) into the medium, how did you sterilize it? By filter sterilization?"  Autoclaving.  One of the first tests we ran was the effect of autoclaving vs filter sterilization on controlling contamination - no difference.  Now we just add into the medium prior to autoclaving.  One thing I have not tested is if the material is inherently sterile and could be added directly to sterilized media.
One thing about PronTech the benzyl ammonium chloride (BAC) is chelated to urea.  I think this form of BAC allows for much higher levels of BAC to be used (PronTech is 40% BAC - this is much higher than the BAC levels in cleaning products I have looked at).
"2. If the control of this product is not so good on bacteria, did you include other antibiotics in the medium with BAC to eliminate bacteria?"  Streptomycin.  We tested a number of antibiotics and found that streptomycin worked well (and was the cheapest of the antibiotics that worked).
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antifungal activity
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Hello, Maliha
I think it is better to repeat your experiment; indeed, some fungi do not growth uniformly (such as Penicillium, Aspergillus) and it is difficult to obtain a radial growth; as your control and T4 looks good so try to check possible contamination, or start your cultures with mycelial plugs (I don't know if you have started with spore suspensions - if you do, you must pay attention when you spread). In my opinion, you must try get the cultures from mycelial plugs, cutted from nice monosporal cultures). What fungus is in your test?
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  1. I harvest some infected leaves with S. turcicum patches.
  2. I dried them during 4 days at 30-35°C. 
In which form do you think it is better to conserve it ? Leafs, powder ?
In which conditions ? In a cold chamber ? (6-8°C and H2O%=50%)?
How long can I preserve theses spores in well-adapted storage conditions?
Many thanks in advance for all the answers!
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Dear Cyprien Gerard
From attached pdf. = downloadPDF  = Single germinated conidia of S. turcica were transferred to lactose casein hydrolysate agar (LCA) (44). Cultures grew under 12 h lighting for 10 to 14 days at 20°C until abundant conidia were produced. Conidia of each isolate were harvested  in sterile 15% (vol/vol) glycerol solution, placed in duplicate labeled vials, and stored at –80°C for future use
Pls. find the attached files
Hoping this will be helpful
Regards
Prof. Houda Kawas
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Dear Friends
Please let me know what is this on apple?
See the picture.
Thanks
Best
Adalbert
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Hi Zviadi,
Thanks for the explanation.
I remember when I was a little boy, I saw a lot of cicadas on the trees singing. This kind of big cicadas (see picture #1). I did not know Buffalo treehopper cicada (see attached picture #2). Now I know. It is an interesting discussion, and thank you for your inputs.
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i apply my treatment and no fungus grow after 1 day
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Maliha,
Before inoculation, all  fruits should be surface disinfected. Even those in control.
Franci
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In my case, I'm working with entire root systems of Medicago truncatula infected by an oomycete.
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Hi Pierre-Marc,
The qPCR has been done :) I just need to see the localization of the expression now. I will have a look at your fixation proposition.
Thanks
Justine
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I want to know the methods and procedures for inoculating sawdust at a certain weight and the parameters to measure or take to know if degradation is enhanced.
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first make 5 to 6 sets of saw dust soaked with double distilled water ( including a control set without any inoculation). then add the microbes aseptically to each set after serial dilution with .01% MgSO4 solution ( upto 10-5/-6 level) and take your desired data after certain period of inoculation. 
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experimental methode and medium for isolation of Bipolaris sorokiniana from infested wheat roots
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I'm now working with Bipolaris sorokiniana. It was isolated from Eleusine indica grass disease....for now...I'm culturing it in PDA media...and it grows well...
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Which protocol i may follow to preserve plant pathogens at -20 and at -80 degree Celsius specially Rhizoctonia, Alternaria, Bipolaris, Curvularia, Sclerotium, Colletotrichum and Fusarium? we are not using liquid nitrogen based preservation more. 
Thanks in advance
Shuvrah Rehman
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Preservation of the pathogenic spore suspension at -80°C in glycerol is the best way as suggested by Dr. Latif. 
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207
I would like to inquire whether someone has been treated fungal cultures by vacuum in a Petri dish? We have found that the agar dries suddenly. This is normal? How can we detect the effect of vacuum and not the desiccation?
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73
Thank you. But I am not Levente :)
In this experiment we used 4mbar pressure (~0.004 atm). This vacuum cause the drying and cooling of the petridish and substrate (perhaps the evaporation). But it seems to the fungi (Fusarium sp.) can survive this treatment.
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I'm working towards a research on Pterocarpus trees on the Palmas del Mar forest at Humacao, we've noticed some white leaves on the seedlings of these trees, I'm having trouble pin-pointing a protocol to start this project. Any suggestions?
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Also
Houda
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Wheat blast caused by Magnaporthe oryzae Triticum pathotype is a fearsome disease first emerged in Bangladesh. It poses serious threat to food and nutritional security of Bangladesh. Scientist fears that this disease may spread to neighbouring Asian countries. We need to isolate the pathogen in infected plants and alternate hosts and thus needs a cheaper and faster protocol.
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Hi: You can use water agar (WA) to isolate the pathogen from the tissue after surfers sterilize, WA allows the pathogen to grow very slowly that give you time to purify it in new culture
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I want to test the inoculation factor of these 3 fungus againts eleusine indica leaves.
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If I am not confused with your question, I could suggest you to go for Ethyl acetate extraction method followed by agar well diffusion technique for the bioassay. 
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I want to check chitinase activity of bacteria. Is any indicator dye can be added to observe the result of chitinase production by bacteria easily? If yes, then what should be the pH to be maintained? 
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Hi, if you try to dissolve colloidal chitin into the media reducing sugars will be released into the media and enzyme activity through the enzyme assays will become over estimated. Alternatively, you can use specific substrates such as p-NAG to determine activity.
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Plants are young (tree nursery). Apical leaves are affected.
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Dear Cecilia Lutz
a candidate for some environmental stress rather than a pathological or disease related issue. The affected areas on the leaves may be related to a nutritional imbalance (phosphorus deficiency, yellow due to nutrient deficiencies, Several Deficiency Symptoms Iron, Manganese and Zinc together) or a root malfunction in the plant or even a contact type injury from something applied/sprayed to the soil or directly to the leaves.  You may consider having the soil tested for nutrient availability and reviewing any chemicals applied to the foliage (such insecticides against whitefly, picture show traces).  Watch the new leaves as they form for the same symptoms.  If they look OK, then this fading and curling may be temporary.  Also check closely on both sides of the leaves for any insects.  You may also snip off a few affected leaves and show them to your local plant nursery or garden center for their comments since these folks may have seen these symptoms before. Also freezing weather , possibly caused by poor drainage, overwatering. For  all disease Symptoms http://www.bioversityinternational.org/uploads/tx_news/FAO_lBPGR_technical_guidelines_for_the_safe_movement_of_citrus_germplasm_501.pdf
Hoping this will be helpful            
Regards
Prof. Houda Kawas
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I got fungal contamination every time (photo attached) during isolation of Phytophthora infestans. I tried culturing on potato tuber slices, for first few days culture is pure but after 1 week some other fungus grows on it. What is this fungi? What are the causes for this type of contamination and how to avoid this? Thanks in advance.
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I normally isolated Phytophthora infestans by plating sporangia or small pieces of infected tissue onto antibiotic rye agar including the anti-fungal antibiotic pimaricin (natamycin) to suppress fungal contamination as well as an antibacterial antibiotic (rifamycin).  The media are described in the CIP Manual referenced above.
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Hi All, 
Please here are some other pictures that I got. May somebody help me with the name of this fungus? I am working on water hyacinth and these pictures come from damage leaves by pathogens
Thanks
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Dear Sonia is may be possible Rhizoctonia sp. The hyphae of Rhizoctonia have  the following characteristics: 1) some shade of brown; 2) a special type of cross wall within the hyphae, called a dolipore septum; 3) each cell is multinucleate (has many nuclei) rather than binucleate; 4) branches that are produced at right angles; 5) no asexual spores are formed by the mycelium. You can isolate DNA of your culture and test some primer for identification. 
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We want to check the effect on DNA due to infection of Plant Pathogen. What could be proper procedure to isolate DNA from an infected Plant? Is the procedure of isolation of DNA same for both healthy as well as disease Plants?
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Thank You 
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During my experiment (cut stem inoculation technique) to identify the isolate aggressiveness of Macrophomina phaseolina and out of 15, 3 isolates found very aggressive.
Can we go for race identification. If yes, then what procedure i should follow.
Thank you.
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Dear Folks, Note all pathogenic fungi forming pathological races have as Oadi Matny points differentiated response in susceptibility and resistance divulged by screening a range of cultivars and looking at the response pattern. 
Pathogenicity testing under standard uniform condition will be necessary for  your study.
I would select a range of isolates and look to distinguishing cultural morphology many times these show differences in their pathogenicity focusing on stem inoculation and response with a toothpick inoculation technique. 
As for a inoculation technique I would use toothpick inoculation and have the plants under favorable temperature and humidity to look for differences in symptom expressions.
Many times different pathological races show distinctive symptomatology. Beside looking at cultivars within soybean looking at distinctive species of host plants will reveal potential grouping and differences. 
Ultimately these can be confirmation from a fungal genomic DNA analysis. 
Macrophomina phaselina as a facultative pathogen may show less distinctive pathological races as commonly expressed in obligate parasites such as rusts, powdery mildews for instance. 
Good luck keep us posted on your results. 
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The photos were taken in northern Italy at the beginning of September of the current year. The vines are placed in an experimental area and are grown in pots. The lower surface of the leaves is normal. Some entomologists and acarologists exclude that the damage is caused by mites. The damage has a negative impact on the quality of grapes. Did someone note similar damage elsewere and does he know  or can hypothesize the cause?
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Perhaps, the phytotoxicity of a plant protection product or combination of products being used. For instance, some varieties (in particularly American hybrids) may be susceptible on wettable sulphur, especially when used at higher temperatures exceeding 30 oC and/or in higher doses.  I would exclude biotic causers like fungi or mites.
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Rapid potato stem damage caused by Phytophthora is spreading this year on some potato cultivars. Is it common symptoms comparing to late blight? Some plants are infected by Pectobacterium spp. together with oomycete, increasing soft rot symptoms. Is it common for green part of potato plant?
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Jan, 
I think you are right. It plays a special role at warm weather with short nights (middle of  summer). 
Alex
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Hi, can anyone give me the detail for SOD assay calculation (SOD U/mL) and SOD inhibition %?
Thanks in advance.
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