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New Single-Stranded DNA Virus With a Unique Genomic Structure That Infects Marine Diatom Chaetoceros Setoensis

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New Single-Stranded DNA Virus With a Unique Genomic Structure That Infects Marine Diatom Chaetoceros Setoensis

Yuji Tomaru et al. Sci Rep.

Abstract

Diatoms are among the most abundant organisms in nature; however, their relationships with single-stranded DNA (ssDNA) viruses have not yet been defined in detail. We report the isolation and characterisation of a virus (CsetDNAV) that lytically infects the bloom-forming diatom Chaetoceros setoensis. The virion is 33 nm in diameter and accumulates in the nucleus of its host. CsetDNAV harbours a covalently closed-circular ssDNA genome comprising 5836 nucleotides and eight different short-complementary fragments (67-145 nucleotides), which have not been reported in other diatom viruses. Phylogenetic analysis based on the putative replicase-related protein showed that CsetDNAV was not included in the monophyly of the recently established genus Bacilladnavirus. This discovery of CsetDNAV, which harbours a genome with a structure that is unique among known viruses that infect diatoms, suggests that other such undiscovered viruses possess diverse genomic architectures.

Figures

Figure 1
Figure 1. Chaetoceros setoensis.
(A) Micrograph of intact cells. (B) Micrograph of CsetDNAV-infected cells at 72 hpi. Bars indicate 100 μm.
Figure 2
Figure 2. TEM micrographs of ultra-thin sections of C. setoensis and negatively stained CsetDNAV particles.
(A) Uninfected cell. (B) Nucleus of the uninfected cell in panel A. (C) CsetDNAV infected cell at 48 hpi. (D) Nucleus in panel C. (E) Higher magnification of the infected host cell nucleus in panel D. (F) Negatively stained CsetDNAV particles in the culture lysate. Ch: chloroplast; M: mitochondrion; N: nucleus; F: fibrous structure; V: virus-like particles. Arrows indicate electron dense particles, which are smaller than the mature virus-like particles.
Figure 3
Figure 3. Nucleic acids of CsetDNAV untreated (lane 1) or treated with S-1 nuclease (lane 2), DNase I (lane 3), RNase A (lane 4), and heated at 100°C for 2 min (lane 5).
Figure 4
Figure 4. PCR products amplified with primer sets CsetD_1 and CsetD_2 (a), CsetD_3 and CsetD_4 (b), CsetD_1 and CsetD_4 (c), and CsetD_2 and CsetD_3 (d).
The expected sizes of the products in lanes a–d are 5857 bp, 5856 bp, 2724 bp, and 3153 bp, respectively. The most intense bands correspond to target amplicons. The faint bands were likely generated due to overlapping nucleotide sequences.
Figure 5
Figure 5. Structure of the CsetDNAV genome.
Open arrows indicate PCR primers (see Fig. 4). Grey arrows indicate the positions of the VP1, VP2, and VP3 ORFs. Grey boxes indicate the positions of the small, complementary ssDNA fragments.
Figure 6
Figure 6. ML tree based on deduced amino acid sequences of the putative replication protein.
NJ bootstrap values (%) from 1000 samples are shown at the nodes, followed by bootstrap values (%) based on the ML analysis of 1000 samples. The ML distance scale bar is shown. Amino acid sequences and their accession numbers are as follows: C. debilis DNA virus (CdebDNAV), AB504376; C. lorenzianus DNA virus (ClorDNAV), AB553581; C. salsugineum DNA virus (CsalDNAV), AB193315; C. setoensis DNA virus (CsetDNAV), AB781089; C. tenuissimus DNA virus (CtenDNAV), AB597949; Chaetoceros sp. strain TG07-C28 DNA virus (Csp05DNAV), AB647334; Thalassionema nitzschioides DNA virus (TnitDNAV), AB781284.
Figure 7
Figure 7. Structural proteins of CsetDNAV.
Figure 8
Figure 8. Changes in cell numbers of virus-infected (closed squares) or virus-uninfected cultures (open squares) of C. setoensis and viral titers (open circles).
The error bars indicate 95% confidence intervals.

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