Virus analysis requires many different tools for different steps. While many effective software options exist for automating these, they usually follow an end…

Ambiviruses are fungal-infecting circular RNA viruses that uniquely combine viroid-like and viral features, yet the function of their conserved ORF-B protein and their effects on hosts remain unknown,...

Metagenomic profiling algorithms commonly rely on genomic differences between lineages, strains, or species to infer the relative abundances of sequences prese…

Tomato spotted wilt virus (TSWV) is a generalist plant virus that infects over 1000 plant species, causing significant damage to cultivated crops, inc…

Viroids are small, circular non-coding RNAs that autonomously replicate in plants, exploiting host cellular machinery for replication and spread. Recent studies reveal that viroid-like agents can infect filamentous fungi, suggesting cross-kingdom interactions. In this study, we report the discovery and the characterization of TsvlRNA1 in Trichoderma spirale , a transmissible viroid-like RNA containing a hammerhead ribozyme in one polarity strand. Bioinformatic data, molecular validation, and reverse genetics experiments demonstrate that TsvlRNA1 is circular with an active ribozyme essential for replication. TsvlRNA1 replicates autonomously and transmits horizontally between Trichoderma species, eliciting 21–23 nt viroid-derived small RNAs consistent with RNA silencing targeting. The biocontrol capacity of Trichoderma against Rhizoctonia solani is variably modulated by TsvlRNA1, with effects ranging from positive to negative depending on host strain. In T. spirale , data suggests genotype-by-agent interactions influence antagonistic potential negatively. TsvlRNA1 transmission via horizontal routes is prevalent, and the viroid-like RNA fails to infect plant hosts experimentally. These results highlight so-far the underappreciated ecological and functional diversity of viroid-like agents in fungi, with implications for fungal biology, biocontrol, and genotype-phenotype relationships in eukaryotes.

In phylogenomics, gene tree reconstruction depends on multiple sequence alignment (MSA) and tree inference, and ongoing work continues to improve inference quality. Denser taxon sampling has been associated with improved gene tree inference, suggesting that adding homologs could be a practical route to higher accuracy as sequence databases continue to expand. However, adding sequences can influence multiple steps of typical inference pipelines, and little is known on its specific effect on the multiple sequence alignment, tree reconstruction, and rooting steps. We performed a large-scale empirical benchmark to quantify how homolog enrichment affects alignment and phylogenetic inference. Using an enrichment-impoverishment design and a measure of tree accuracy based on taxonomic congruence, we found that enrichment consistently improves tree inference quality, while effects on alignment quality are marginal. We show that this improvement is associated with accurate root placement on enriched trees when sensitive homolog search is accompanied. Notably, much of the benefit can be retained with relatively compact alignments produced by sequence addition. Building on these observations, we provide a tool, AmpliPhy, which efficiently improves phylogenetic reconstruction of protein families through homolog enrichment. The AmpliPhy open-source pipeline software is available at https://github.com/DessimozLab/ampliphy. ### Competing Interest Statement The authors have declared no competing interest. Swiss National Science Foundation, https://ror.org/00yjd3n13, 216623, 10005715

Since the development of the Minimum Information about an Uncultured Viral Genome (MIUViG) standard (Roux et al. 2019), there has been an explosion in the number of genomic data generated on unculture...