Biology subjects

Fall, M. L.

Publications and source records attributed to Fall, M. L..

5 recordsLinked to original sources

bioRxiv2026

Botrytis virus X ORF2 suppresses RNA silencing in a trigger-restricted manner and modulates is associated with altered Dicer-like gene expression in Botrytis cinerea

RNA interference (RNAi) is a central antiviral defense mechanism in fungi, yet relatively few mycoviral suppressors of RNA silencing (VSRs) have been functionally characterized, particularly in phytopathogenic hosts. Botrytis virus X (BVX), a positive-sense RNA virus in the family Alphaflexiviridae, infects Botrytis cinerea and encodes five predicted open reading frames (ORFs), most of which have unknown functions. Here, we screened BVX ORFs 2-5 for RNA silencing suppressor activity using complementary GFP-based assays in Nicotiana benthamiana and examined the leading candidate in the fungal host B. cinerea. BVX ORF2 (X2) enhanced GFP transcript and protein accumulation in assays where silencing is triggered by sense RNA but failed to suppress silencing triggered by hairpin-derived siRNAs or miRNA-guided targeting, indicating a trigger-restricted suppressor phenotype. In B. cinerea, transgenic expression of X2 was associated with reduced induction of the RNAi associated genes BcDCL1 and BcDCL2 compared to empty vector controls, with the strongest effect observed on BcDCL1. In a virus-infected fungal background, X2 expression was also associated with increased viral RNA accumulation. Together, these results identify BVX X2 as a BVX-encoded, trigger-restricted suppressor of RNA silencing and link its expression to altered RNAi-related gene induction and increased viral RNA accumulation in B. cinerea.

microbiology↗

bioRxiv2025

Exploring the mycovirome: novel and diverse mycoviruses in Botrytis cinerea

Botrytis cinerea is a necrotrophic fungal pathogen that causes significant economic losses to many crops, including vegetables, fruits, and ornamental plants. The management of B. cinerea is difficult due to a rise in fungicide resistance. Harnessing mycoviruses that cause reduced virulence (hypovirulence) in B. cinerea is a promising alternative. Over 100 mycoviruses have been identified in Botrytis spp. to date, including several hypovirulence-inducing mycoviruses. This research aimed to further explore, for the first time in Canada, the mycovirome of B. cinerea and identify potential hypovirulence-inducing mycoviruses. Isolates of B. cinerea were collected from fruits and vegetables in the province of Quebec. Fitness and pathogenicity criteria, including sclerotia production, colony morphotype, and lesion size were evaluated. A double-stranded RNA (dsRNA) extraction protocol tailored to the detection of mycoviruses was used to sequence dsRNA from 45 isolates with low fitness/pathogenicity, and an in-house bioinformatics workflow was used to profile the mycovirome. Mycoviruses were identified in 44/45 isolates. Most of these had positive single-stranded RNA or dsRNA genomes, and a small number had negative single-stranded RNA, single-stranded DNA, or reverse transcriptase RNA genomes. Following deep analysis of RNA-dependent RNA polymerase and replication initiation proteins, a total of 62 unique contigs were identified belonging to new strains of mycovirus species. Furthermore, four putative novel mycovirus species belonging to Endornaviridae, Botybirnaviridae, Peribunyaviridae, and Bunyavirales taxa were identified. Several mycovirus species positively and/or negatively co-occurred with B. cinerea isolates collected from strawberry or raspberry. This study revealed a high degree of diversity in the mycovirome of B. cinerea. Species accumulation curve analysis indicated that, with the number of isolates characterized, we were unable to capture the full extent of expected diversity. Nevertheless, we identified potential hypovirulence-inducing mycoviruses, including Botrytis cinerea mitovirus 1, Botrytis cinerea hypovirus 1, and Botrytis porri botybirnavirus 1. Some of these novel mycoviruses belonged to taxa known to produce viral particles, which can be an interesting feature for their use as biocontrol agents (BCA). ImportanceThis study provides the first comprehensive profiling of mycoviruses infecting Botrytis cinerea in Canada, a significant step in understanding how these viruses can naturally limit crop disease. Due to growing resistance against conventional fungicides, new biological methods to control B. cinerea are crucial. By profiling mycoviruses in fungal samples collected in Quebec, we identified several novel viruses that appear to reduce the pathogenicity of B. cinerea. These viruses, known as hypovirulence-inducing mycoviruses, could be used to develop biocontrol agents (BCA), offering a more sustainable disease management alternative. Notably, we found virus families with extracellular potential, which may enable easier application as BCAs in agriculture. This research not only broadens the understanding of fungal virology but also holds promise for innovative, eco-friendly approaches to managing Botrytis cinerea in Canada.

genomics↗

bioRxiv2023

dsRNA-based viromics: A novel tool unveiled hidden soil viral diversity and richness

Viruses play a crucial role in agroecosystem functioning. However, few studies have examined the diversity of the soil virome, especially when it comes to RNA viruses. Despite the great progress in viral metagenomics and metatranscriptomics (metaviromics) toward RNA viruses characterization, soil RNA viruses ecology is embryonic compared to DNA viruses. We currently lack a wet lab. method to accurately unhide the true soil viral diversity. To overcome this limitation, we developed dsRNA-based methods capitalizing on our expertise in soil RNA extraction and dsRNA extraction ported from studies of phyllosphere viral diversity. This proposed method detected both RNA and DNA viruses and is proven to capture a greater soil virus diversity than existing methods, virion-associated nucleic enrichment, and metaviromics. Indeed, using this method we detected 284 novel RNA-dependent RNA polymerases and expanded the diversity of Birnaviridae and Retroviridae viral families to agricultural soil, which, to our knowledge, have never been reported in such ecosystem. The dsRNA-based method is cost-effective in terms of affordability and requirements for data processing, facilitating large-scale and high-throughput soil sample processing to unlock the potential of the soil virome and its impact on biogeochemical processes (e.g. carbon and nutrient cycling). This method can also benefit future studies of viruses in complex environments, for example, to characterize RNA viruses in the human gut or aquatic environment where RNA viruses are less studied mainly because of technical limitations.

ecology↗

bioRxiv2023

Nanovirseq: dsRNA sequencing for plant virus and viroid detection by Nanopore sequencing

Worldwide, there is a need for certified clean plant materials to limit viral diseases spread. In order to design a robust and proactive viral-like disease certification, diagnostics, and management program, it is essential to have a fast, inexpensive, and user-friendly tool. The purpose of this study was to determine whether dsRNA-based nanopore sequencing can be a reliable method for the detection of viruses and viroids in grapevines or not. Compared to direct RNA sequencing from rRNA-depleted total RNA (rdTotalRNA), direct-cDNA sequencing from dsRNA (dsRNAcD) yielded more viral reads and detected all grapevine viruses and viroids detected using Illumina MiSeq sequencing (dsRNA-MiSeq). With dsRNAcD sequencing it was possible to detect low abundance viruses (e.g., Grapevine red globe virus) where rdTotalRNA sequencing failed to detect them. Indeed, even after removing rRNA, rdTotalRNA sequencing yielded low viral read numbers. rdTotalRNA sequencing was not sensitive enough to detect all the viruses detected by dsRNA-MiSeq. In addition, there was a false positive identification of a viroid in the rdTotalRNA sequencing that was due to misannotation of a host-driven read. For quick and accurate reads classification, two different taxonomical classification workflows based on protein and nucleotide homology were evaluated in this study, namely DIAMOND&MEGAND (DIA&MEG) and Centrifuge&Recentrifuge (Cent&Rec), respectively. Virome profiles from both workflows were similar except for grapevine endophyte endornavirus (GEEV), which was only detected using DIA&MEG. However, because DIA&MEGs classification is based on protein homology, it cannot detect viroid infection despite giving more robust results. Even though Cent&Recs virus and viroid detection workflow was faster (30 minutes) than DIA&MEGs (two hours), it could not provide the details and information DIA&MEG was able to provide. As demonstrated in our study, nanopore dsRNAcD sequencing and the proposed data analysis workflows are suitable and reliable for viruses and viroids detection, especially in grapevine where viral mixed infection is common.

pathology↗

bioRxiv2023

A decade of hidden phytoplasmas unveiled through citizen science

Climate change is impacting agriculture in many ways, and a contribution from all is required to reduce the imminent loses related to it. Recently, it has been showed that citizen science could be a way to trace the impact of climate change. However, how can citizen science be applied in plant pathology? Here, using as an example a decade of phytoplasma-related diseases reported by growers, agronomists, citizens in general, and confirmed by a government laboratory, we explore a new way of valuing plant pathogens monitoring data deriving from land-users or stakeholders. Through this collaboration we found that in the last decade thirty-four hosts have been affected by phytoplasmas, nine, thirteen and five of these plants were, for the first time, reported phytoplasma hosts in Eastern Canada, in Canada and worldwide, respectively. Another finding of great impact is the first report of a Ca. P. phoenicium-related strain in Canada, while Ca. P. pruni and Ca. P. pyri was reported for the first time in Eastern Canada. These findings will have a great impact in the management of phytoplasmas and their insect vectors. Using these insect-vectored bacterial pathogens, we show the needs of new strategies that allow a fast and accurate communication between concerned citizens and those institutions confirming their observations. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/524422v1_ufig1.gif" ALT="Figure 1"> View larger version (107K): org.highwire.dtl.DTLVardef@f676eborg.highwire.dtl.DTLVardef@f964bforg.highwire.dtl.DTLVardef@14576acorg.highwire.dtl.DTLVardef@1721bfb_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗