​Monday Plenary Speaker

Marin Brewer

• Question: Marin, thanks for an excellent talk. The genetic diversity figure showed higher genetic diversity for multi-drug resistant isolates​ that clustered together. What could explain this?
• Answer: Thanks! I think we would have to do further analyses to determine if this clade is more diverse. Others have found less genetic diversity in pan-azole resistant isolates that is likely due to selective sweeps.
• Question: Are there any evolutionary costs associated with Azole (or multi-fungicide) resistance? Could these populations decline over time or are they with us forever?
• Answer: We hope so and are currently investigating this! Probably not to benzimidazoles and QoIs since we do not see costs to these in most plant-pathogenic fungi where it has been investigated, but there are often costs to azole resistance.
• Question: Marin, thank you for sharing this very interesting, yet quite disturbing story! I think you mentioned there are three azoles and maybe one that is on label for human infections. Do you know whether there are efforts underway to identify more fungicides for these infections that target different modes of action?
• Answer: There are only three classes (triazoles, polyenes, echinocandins) of antifungals used in medicine. There are several azole drugs with varying uses in medicine and a handful that are used to treat aspergillosis. Yes, there are efforts to develop alternative moa antifungals.
• Question: What is the residence time of azoles in the substrates that are hotspots? What is the mechanism of degration of azole in the environment (if there is one)?
• Answer: They are persistent in these environments. There are some studies being conducted on how long they persist and if they breakdown products are also selcting for resistant isolates.
• Question: Are fungicides used more frequently in flower beds than in agricultural fields?  Is there any research investing Aspergillus flavus fungicide resistance in human patients?
• Answer: Those are great questions. Bulbs are dipped in azoles and they are used in very high amounts in Europe, especially in flower production. I don't know offhand about A. flavus azole resistance, but it would be interesting to look into.
• Question: Are there genetic backgrounds of Aspergillus that are more favorable for acquisition of fungicide/asole resistance?
• Answer: It appears that there are based on the pan-azole cluster, but we are not sure what it is.
• Question: In compost and hotspots where you find resistant isolates, is A. fumigatus the dominant fungus present? Also do you see azole resistance in other fungi in these places?
• Answer: Great question! A. fumigatus is dominant. We do see some resistant mucorales fungi when we isolate at 45C, but it would be very interesting to look at others that are not necessarily thermotolerant.
• Question: Genetic mutations are contributing to fungicide resistance. Do you think there is potential for genetics to contribute novel control strategies so that we rely less on fungicides?
• Answer: Absolutely! Host resistance is one of the best control strategies. 
• Question: Is there any indication that bacterial biocontrol species can help control Aspergillus fumigatus?
• Answer: I have not seen any research on this topic.
• Question: Does proper hot composting destroy A. fumigatus? So do we have a problem iwth improper composting?
• Answer: Some studies have shown that A. fum is destroyed by the composting process, so it could be an issue with improper composting
• Question: Could you explain more about the air? how did those samples were collected and analyzed?
• Answer: We use an air sampler.
• Question: Could you please explain what you mean by fitness cost
• Answer: The fungus is less fit meaning it's growth and reproduction are impacted (in the absence of azoles) compared to susceptible isolates.
• Question: Has comparison in ARAf between compost piles from organic vs conventional farms?
• Answer: That is something we plan to do more.
• Question: From Jay pscheidt orginally - The ornamental industry can use a wider variety of this class of fungicide (group 3), could one of them be driving more of the resistance development?
• Answer: Answered live.
• Question: Don't you think that the “Ag Environment" label is too broad and misleading, given that these things seem to be coming predominantly from flower farms?
• Answer: We do not know where the multifungicide resistant isolates emerged infections are coming from, although you are correct that most of the environmental detection is in these flower and compost hotspots. However, we do find these ARAf in multiple ag environments aside from flowers and compost. We are interested in sampling more of the ag environment to better identify hot spots, but we may eventually narrow it specific ag environments. ​
  

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Tuesday Plenary Speaker

Liying Sun 

• Question: What degree of disease severity can be anticipated when viruses move from one kingdom to another? Any instance of increase in disease severity if virus moves from plant to human?
• Answer: We only could answer this question based on limited examples of virus cross-kingdom transfers between plants and fungi observed in our studies. Infection of tobacco mosaic virus (TMV) and cucumber mosaic virus (CMV) in fungi does not induce symptoms. Likewise, infection of a mycovirus, Cryphonetria hypovirus 1 (CHV1), in plants is also asymptomatic. More observations on cross-kingdom infection involving a wide range of viruses would reveal whether viruses can actually induce symptoms or disease in their novel unconventional hosts. It is interesting to note that CMV infection in Rhizoctonia solani is asymptomatic, but CMV infection enhances the virulence of R. solani in plants. Thus, the outcome of cross-kingdom virus infection can also be anticipated on the change of pathogenicity or performance of the novel host. In the case of plant viruses that also infect and replicate in their insect vectors such as plant reoviruses, rhabdoviruses and bunyavirus, infection could cause diseases in both plant and insect hosts.
•  So far, there is no evidence that plant viruses can infect human. Some plant viruses were detected in human lungs, saliva or stools, although there is no direct evidence of potential risks to human health.
  
  
• Question: Do you have any example where mycoviruses cause plant disease and produce symptoms without presence of helper plant virus?
• Answer: To our knowledge, there is no example of a common mycovirus that causes disease in plants. However, many plant viruses are closely related to mycoviruses. It is necessary to investigate whether these plants viruses are originated from the fungi and have been transferred to the plants.
   
• Question: Those apperently latent dsRNA viruses that infects plants and fungi (like amalgavirus and partitivirus) could be transmited by fungi to plants? Is that why the capsid of some of them were not idenfied yet?
• Answer: This is an interesting view, but it should be demonstrated experimentally. Considering that those latent plant dsRNA viruses are only known to be transmitted via seeds, transmission through the fungal vectors may also be a potential route for the spread of this kind of viruses. Our findings support the view of the presence of active transport mechanisms of molecules, including viral components between fungi and plants, such as recently known the transfer of small RNAs from fungi to plants that involves vesicles.
   
• Question: Have you done any study of viruses infecting oomycetes ?
• Answer: No, we do not do any works on viruses infecting Oomycetes.
  
  
• Question: Why do you think extracellular viruses are so rare for mycoviruses?  Do fungi have the best cellular barriers?
• Answer: Because mycoviruses generally have been adapted to intracellular transmission through spores or hypal anastomosis. However, our findings on cross-kingdom virus infection provide insight for an alternative route for mycovirus transmission through the plants.
  
  
• Question: From your experiences, what is your thoughts on the longevity of such fungal acquired plant virus or vice versa?
• Answer: In our study, we observed that under laboratory conditions, some plant viruses could be quite stable in particular compatible fungal hosts. Perhaps plant virus infection in fungi is also relatively stable under natural conditions because we have found a plant virus (CMV) infecting R. solani isolated from the field. However, more findings of natural cross-kingdom virus infection are necessary to support this view.
  
  
• Question: Is there any changes of expression of genes for mycoviruses  between plant and fungi ?
• Answer: Currently, we do not have any data comparing the gene expression of mycoviruses in plants and fungi.​