Single-cell genomics reveals new rozellid lineages and supports their sister relationship to Microsporidia.

The phylum Rozellomycota has been proposed for a group of early-branching holomycotan lineages representing obligate parasites and hyperparasites of zoosporic fungi, oomycotes or phytoplankton. Given their predominantly intracellular lifestyle, rozellids are typically known from environmental ribosomal DNA data, except for the well-studied Rozella species. To date, the phylogenetic relationship between rozellids and microsporidians (Microsporidia) is not fully understood and most reliable hypotheses are based on phylogenomic analyses that incorporate the only publicly available rozellid genome of Rozella allomycis . Here, we provide genomic data of three new rozellid lineages obtained by single-cell sequencing from environmental samples and show with a phylogenomic approach that rozellids form a monophyletic group that is sister to microsporidians, corroborating the previously proposed phylum Rozellomycota. Whereas no mitochondrial genes coding for the respiratory Complex I could be found, we discovered a gene coding for a nucleotide phosphate transporter in one of the three draft genomes. The scattered absence of Complex I genes and scattered presence of nucleotide transporter genes across diverse microsporidian and rozellid lineages suggest that these adaptations to a parasitic lifestyle, which reduce the parasite's capability to synthesize ATP but enables it to steal ATP from its host, evolved independently in microsporidians and rozellids.