Mycoviruses are widely distributed across major fungal taxa, including yeasts, edible mushrooms, and pathogenic fungi (Zhang et al., 2022). Since their initial discovery in Agaricus bisporus in the mid-20th century, viruses or virus-like particles have been reported in several economically important edible fungi, such as Pleurotus ostreatus, Flammulina velutipes, Auricularia heimuer, and Lentinula edodes (Kim et al., 2013, Kim et al., 2008, Li et al., 2020; Magae and Sunagawa, 2010). In recent years, advances in high-throughput sequencing and bioinformatics have markedly accelerated the discovery and characterization of novel mycoviruses (Lu et al., 2024, Regnault et al., 2021). To date, more than 60 mycoviruses have been identified in over 15 edible mushroom species, some of which adversely affect the cultivation performance (Villan Larios et al., 2023, Zhang et al., 2022). However, unlike animal and plant viruses, mycoviral infections in mushrooms are typically latent, exhibiting inconspicuous symptoms that may only become prominently visible at specific cultivation stages. Once established, such infections can potentially facilitate cross-infection with other pathogens and degrade the production environment, thereby reducing yield and quality and leading to notable economic losses (Eastwood et al., 2015, Song et al., 2020; Wu and Li, 2025; Zhang et al., 2022). Nevertheless, due to coinfections with multiple mycoviruses, latent infections, and the limited availability of effective elimination methods, elucidating the pathogenic mechanisms and ecological roles of mycoviruses remains challenging (Hough et al., 2023, Mehetre et al., 2021; Xie and Jiang, 2024).

In contrast to well-studied cultivated mushrooms such as Lentinula edodes, Pleurotus spp., and Agaricus bisporus, research on mycoviruses in morels (Morchella spp.) remains limited. The first mycovirus in Morchella importuna was reported in 2019 by Gilbert et al., who identified several viruses (Morchella importuna endornavirus 1 [MiEV1], Morchella importuna endornavirus 2 [MiEV2], Morchella importuna endornavirus 3 [MiEV3], Morchella importuna fusarivirus 1 [MiFV1], and Morchella importuna rhabdovirus 1 [MiRV1]) from transcriptomic data using bioinformatics (Gilbert et al., 2019). Among these, MiEV1, MiEV2, and MiEV3 are proposed members of the family Endornaviridae, with double-stranded RNA genomes of 16.5 kb, 15.3 kb, and 14.3 kb, respectively (GenBank accessions: MK279477–MK279479). Subsequently, two additional viruses, Morchella esculenta fusarivirus 1 (MeFV1) and Morchella esculenta fusarivirus 2 (MeFV2), were identified in wild Morchella esculenta populations, indicating potential diversity among morel-associated viruses and their possible impact on cultivation (Sahin et al., 2021). To date, none of these viruses, including MiEV1, has been formally ratified as an approved viral species by the International Committee on Taxonomy of Viruses (ICTV). Despite these recent discoveries, several key gaps remain: (1) most identified viruses, including MiEV1, MiEV2 and MiEV3, have been detected primarily through bioinformatics analyses, without experimental validation in cultivated strains; (2) the incidence, transmission patterns, and interspecies transmissibility of these viruses in cultivated Morchella germplasm resources remain largely unexplored, owing to the scarcity of epidemiological data; and (3) effective methods for eliminating mycoviruses from morel cultures are currently unavailable, and the effects of virus removal on Morchella growth performance have yet to be systematically assessed. Collectively, these shortcomings constrain disease management strategies in edible fungi and obstruct deeper mechanistic investigations into virus-host interactions.

China is the world’s leading producer of morels, yet the rapidly expanding cultivation has intensified disease concerns in morels (Xu et al., 2022). These pathogens may heighten strain sensitivity to environmental stress and contribute to yield instability or even total crop failure (Liu et al., 2017, Peng et al., 2025). While reports on morel diseases have primarily focused on fungal pathogens, research on viral infections remains limited (Shi et al., 2022, Zhu et al., 2023). Building on these gaps, this study screened five mycoviruses in cultivated Morchella spp. strains from China, assessed seven antiviral agents for efficacy against MiEV1, and established a protocol for generating virus-free strains through successive treatments. We also evaluated MiEV1 elimination's effects on mycelial growth and biomass to elucidate its host impacts. These efforts provide initial epidemiological insights into mycovirus prevalence in cultivated Morchella, along with evidence for ribavirin's potential as an antiviral agent. Overall, the findings contribute to strategies for virus management in morel cultivation and support further exploration of mycovirus-host interactions in edible fungi.