Viruses with single-stranded DNA (ssDNA) genomes infect hosts in all three domains of life and include economically, medically, and environmentally important viral pathogens. Until recently, ssDNA viruses were classified into 10 different taxa—nine families and one genus not assigned to a family [14, 15]. In 2016, the International Committee on Taxonomy of Viruses (ICTV) created two new families for classification of ssDNA viruses, Pleolipoviridae and Genomoviridae [1]. The family Pleolipoviridae includes viruses infecting hyperhalophilic archaea, and it has recently been described elsewhere [25]. Here, we introduce the family Genomoviridae and provide a short overview of the properties of Sclerotinia sclerotiorum hypovirulence-associated DNA virus 1 (SsHADV-1), the only cultivated member of the new family, and putative SsHADV-1-like viruses.

SsHADV-1 is the first—and thus far the only—ssDNA virus known to infect fungi (all other fungal viruses have RNA genomes) [10, 33]. SsHADV-1 was isolated from a plant-pathogenic fungus, Sclerotinia sclerotiorum [34]. Another unique property of SsHADV-1 that has not been described for other fungal viruses is its ability to establish infection when applied extracellularly in the form of purified viral particles [35]. Virions can infect the hyphae of virus-free S. sclerotiorum directly when applied to hyphae or sprayed on leaves of Arabidopsis thaliana and Brassica napus infected with S. sclerotiorum. When applied to S. sclerotiorum-infected leaves, the virus can suppress the development of S. sclerotiorum-induced lesions. SsHADV-1 has a narrow host range and, in addition to S. sclerotiorum, can infect the related species S. minor and S. nivalis, but not other relatively closely related fungi, such as Botrytis cinerea [35]. Furthermore, SsHADV-1 has been identified in New Zealand and the USA in environmental samples and insects; however, this is not surprising given the near global distribution of S. sclerotiorum [6, 11].

SsHADV-1 virions are non-enveloped, isometric, 20-22 nm in diameter, and constructed from one capsid protein (CP) [34]. The genome is a circular ssDNA molecule of 2,166 nucleotides and contains two genes – for CP and replication initiation protein (Rep) (Fig. 1). The prediction of the cp gene has been validated by N-terminal sequencing of the CP purified from the virions. The large intergenic region contains a potential stem-loop structure with a nonanucleotide (TAATATTAT) motif at its apex, which is likely to be important for rolling-circle replication. The CP of SsHADV-1 is not recognizably similar to the corresponding proteins from viruses in other taxa.

Fig. 1
figure 1

Genome map of SsHADV-1. Genes encoding the replication-initiation protein (Rep) and capsid protein (CP) are shown with blue and red arrows, respectively. The position of the nonanucleotide (TAATATTAT) at the apex of a potential stem-loop structure is also indicated

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Although SsHADV-1 remains the only isolated member of the group, genomes of more than 100 SsHADV-1-like putative viruses have been reported (see Table 1). These genomes have been sequenced from different environmental samples, and many were identified associated with plant material as well as various animal-associated samples, including cerebrospinal fluid and blood of humans (Table 1 and references therein). Even though the hosts of these putative viruses remain unknown, their diversity suggests that SsHADV-1-like viruses are abundant and widespread in the environment.

Table 1 Genome sequences of putative SsHADV-1-like viruses
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All putative SsHADV-1-like viruses encode homologous Rep and CP, and in phylogenetic analyses form monophyletic clades with SsHADV-1 (Figures 2 and 3). Their genomes are of similar size, in the range of 2,089-2,290 nucleotides. Structural and genomic features of SsHADV-1 differ considerably from those of all other currently classified viruses. The Rep of SsHADV-1 is most closely related to the corresponding proteins of members of the family Geminiviridae. It shares with geminiviral proteins two conserved domains, namely geminivirus Rep catalytic domain (Gemini_AL1; PF00799) and geminivirus Rep protein central domain (Gemini_AL1_M; PF08283), with conserved motifs for rolling-circle replication. Notably, similar to some geminiviruses [31], certain SsHADV-1-like viruses also contain introns within their Rep-encoding genes [6, 7, 12, 13, 18, 19, 26–28, 30]. However, Rep-based phylogenetic analysis shows that SsHADV-1 and other related putative viruses form a well-supported, monophyletic clade, which branches as a sister group to geminiviruses (Fig. 2). Unlike the Rep, the CP of SsHADV-1 (and related viruses) does not display any recognizable sequence similarity to proteins of geminiviruses or any other group of known viruses. Furthermore, all geminiviruses possess distinctive geminate virions constructed from two incomplete T = 1 icosahedra [2, 36], whereas the virion of SsHADV-1 is isometric [34]. Finally, the number of genes and size of the genome differ considerably between SsHADV-1-like viruses and geminiviruses. In particular, all putative SsHADV-1-like viruses lack the movement protein, which is essential for the plant geminiviruses.

Fig. 2
figure 2

Phylogenetic analysis of the replication-initiation proteins (Reps) of ssDNA viruses. The type species of the proposed genus Gemycircularvirus within the family Genomoviridae is highlighted in boldface. For phylogenetic analysis, protein sequences were aligned using PROMALS3D [23], and columns containing gaps were removed from the alignment using trimAl (strict mode) [3]. Maximum-likelihood phylogenetic analysis was carried out using PhyML 3.1 [8] with the RtREV +G6 +I +F model, which was selected by PhyML as the best-fitting model. Numbers at the branch points represent aBayes local support values. Branches with lower than 60 % support were collapsed. The scale bar represents the number of substitutions per site. For clarity, the tree was mid-point rooted. All taxa are indicated with the corresponding GenBank accession numbers, followed by abbreviated virus names (full virus names are provided in Table 1)

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Fig. 3
figure 3

Phylogenetic analysis of the SsHADV-1-like capsid proteins. The type species of the proposed genus Gemycircularvirus within the family Genomoviridae is highlighted in boldface. For phylogenetic analysis, protein sequences were aligned using PROMALS3D [23], and columns containing gaps were removed from the alignment using trimAl (strictplus mode) [3]. Maximum-likelihood phylogenetic analysis was carried out using PhyML 3.1 [8] with the LG +G6 +F model, which was selected by PhyML as the best-fitting model. Numbers at the branch points represent aBayes local support values. Branches with lower than 60 % support were collapsed. The scale bar represents the number of substitutions per site. The tree was rooted with the capsid proteins of geminiviruses. All taxa are indicated with the corresponding GenBank accession numbers, followed by abbreviated virus names (full virus names are provided in Table 1)

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In recognition of the unique features described above, SsHADV-1 has been classified into the species Sclerotinia gemycircularvirus 1 within the new genus Gemycircularvirus (Gemini-like myco-infecting circular virus) [26] within the new family Genomoviridae (sigil: Ge- for geminivirus-like, nomo- for no movement protein).

Although Genomoviridae currently includes only a single representative, new members, possibly including uncultivated viruses, are expected to be added to the family in the near future. Based on the available genetic data and phylogenetic analyses (Figures 2 and 3), it is already clear that many new genera will have to be introduced to adequately reflect the diversity of SsHADV-1-like viruses.