Features Diseases
Hunting for tiny, targeted weapons

Mycoviruses – viruses that infect and replicate in fungal cells – offer an intriguing possibility as biocontrol agents for fungal crop diseases. Andriy Bilichak is spearheading an exploratory project aimed at identifying mycoviruses that could effectively control wheat rust fungi.

The first mycoviruses were discovered more than 50 years ago, in the 1960s. However, recent advances in DNA sequencing technologies, which have significantly reduced sequencing costs and increased analytical capacity, are now opening up the world of mycoviruses to researchers like Bilichak.

“Recent discoveries indicate that mycoviruses are very common in fungi,” notes Bilichak, a research scientist with Agriculture and Agri-Food Canada (AAFC) in Morden, Man. Researchers estimate that 30 to 80 per cent of fungal species may be infected by mycoviruses. Moreover, mycovirus hosts include important crop pathogens such as Fusarium and rust species.

Bilichak’s interest in the potential use of mycoviruses to control fungal diseases began during his Ph.D. research. “I was working on using plant viruses to modify the expression of plant genes through the virus-induced gene silencing (VIGS) process. Depending on the target gene in plants, different phenotypes could be achieved by ‘silencing,’ or turning off, the gene.”

He explains that almost every organism can get infected with viruses, and viruses often weaken or harm their hosts. However, mycoviruses must have hosts to replicate, so they have evolved ways to live within their host without causing excessive damage to the fungus, which would threaten their own survival.

The impact of a mycovirus on its host fungus varies depending on the specific virus and host species involved. Sometimes a mycovirus infection does not produce any symptoms in its host; sometimes infection causes hypovirulence, meaning the fungus becomes less effective at causing disease; and sometimes it causes hypervirulence, meaning the fungus becomes more potent in causing disease.

“We are now starting to understand how mycoviruses interact with their host and if they can be used as biocontrol agents,” says Bilichak. “Since many fungal species carry mycoviruses in their cells, sequencing fungal isolates from different geographic regions allows us to discover new viruses and correlate them with fungal virulence.”

He adds, “Searching through the literature, I came across some examples of viruses that can infect pathogenic fungi and reduce fungal infection, causing hypovirulence.” He mentions a recent study on Sclerotinia sclerotiorum, the fungus causing stem rot in canola, where the application of a specific mycovirus reduced disease severity and improved yields.

Seeking rust mycoviruses
Bilichak notes that researchers have found and described a few mycoviruses in rust. However, little research has been conducted to identify specific mycoviruses that could help control rust diseases.

His mycovirus project focuses on leaf rust (Puccinia triticina) and stripe rust (Puccinia striiformis f. sp. tritici), two major wheat diseases affecting both Western and Eastern Canada.

This project, initiated in 2022, utilizes rust samples and sequencing data provided by his plant pathologist colleagues Brent McCallum and Xiben Wang at AAFC-Morden and Guus Bakkeren at AAFC-Summerland.

“Brent McCallum is leading a biovigilance study to monitor the distribution and appearance of novel rust races across the Prairies,” says Bilichak. “His group receives about 200 to 300 isolates annually, mostly from Manitoba and Saskatchewan. Over the past 10 years, his rust collection has grown to about 800 to 1,000 different isolates, allowing us to conduct a systematic study to evaluate the distribution of mycoviruses in leaf and stripe rust isolates across the Prairies.”

Bakkeren has sequenced the genomes and transcriptomes of many of these rust isolates. A transcriptome represents the entire set of RNA molecules expressed by an organism, revealing which genes in the rust fungus’s DNA have been activated.

“Since most rust mycoviruses are RNA viruses, we search the transcriptomes of rust isolates for the presence of mycoviruses using bioinformatics tools,” Bilichak explains.

His research group has started by examining the leaf rust isolates. They are currently analyzing a vast amount of transcriptome data to create a database of mycoviruses infecting the leaf rust isolates.

“In the first year of the project, we initiated the screening of the wheat leaf rust transcriptome for the presence of mycoviral transcripts. Zhen Yao [bioinformatician at AAFC-Morden] has developed the bioinformatics pipeline based on published protocols for targeted mining of the fungal transcriptome for mycoviral RNA,” he explains.

“We successfully identified viral transcripts in the initial screening of pure isolates’ transcriptomes. We are currently working on expanding the search to other isolates from infected wheat tissues.”

Once Bilichak and his team complete cataloging the mycoviruses in the leaf rust samples, they plan to correlate the presence of specific mycoviruses in the leaf rust transcriptome with the virulence of rust on wheat.

They also intend to begin searching for mycoviruses in the stripe rust isolates and develop a strategy for isolating mycoviruses from rust spores.

Looking ahead
Bilichak is optimistic that identifying mycoviruses responsible for inducing hypovirulence in rust fungi may lead to the development of “virocontrol” biopesticides for rusts in the future.

“The rust diseases of wheat are significant, long-term endemic pests that have caused substantial damage to wheat production in Canada,” he says. “The use of fungicides at the flag leaf and anthesis [flowering] stages has increased over the years to prevent fungal diseases.”

However, concerns regarding pathogens developing resistance to these chemical products are on the rise. Additionally, studies have shown that fungicides can have unintended impacts on non-target organisms and ecosystems.

These factors are driving interest in bio-fungicides. “The demand for bio-fungicides is growing due to their environmental friendliness, greater safety for workers, and exemption from residue tolerances on edible crops.” Furthermore, bio-fungicides employ different mechanisms of action compared to chemical products, offering crop growers more options for rotating disease control products to reduce the risk of resistance.

Mycoviruses also pose a low risk to non-target organisms. Bilichak notes, “Mycoviruses are considered ideal biological control agents due to their narrow host ranges.”

While this research is in its early stages, mycovirus products for controlling leaf and stripe rust may ultimately contribute to more sustainable wheat production systems.

This project receives funding from the Manitoba Crop Alliance, Saskatchewan Wheat Development Commission, Alberta Wheat Commission (now Alberta Grains) and Western Grains Research Foundation.