Browsing by Author "Assimakopoulou, Irini Jr"

The genetic and morphological diversity of viruses and more specifically membrane-containing bacteriophages (phages) with single-stranded DNA (ssDNA) genomes is largely unexplored. It can be difficult to detect evolutionary relationships of viruses using solely sequence-based methods due to their rapid sequence evolution. However, more distant evolutionary connections of viruses have been observed based on structure data. Here we introduce an icosahedral tailless ssDNA phage, Cellulophaga phage phi48:2, isolated from the Baltic Sea that has not been assigned to any virus family or taxa. Phage phi48:2 has been previously linked to the family Finnlakeviridae whose members are icosahedral, internal membrane-containing phages with circular ssDNA genomes. However, the presence of lipids in phi48:2 virion has not been studied. In this study, different buffer conditions were tested for infectivity and stability of phi48:2 allowing us to optimize the purification of the phage particles by rate zonal and equilibrium ultracentrifugation in sucrose. Solvent tests in chloroform and ether, as well as low buoyant density of the virion suggested the presence of lipids in the phi48:2 virion. Analysis of the phi48:2 lipids extracted from highly purified virions by thin-layer chromatography revealed that phi48:2 is a membrane-containing phage and acquires its lipids unselectively from its host bacterium Cellulophaga baltica. Lastly, cryogenic electron microscopy of the purified virions also proposed that lipids form a membrane vesicle under the capsid. Altogether our results show that phi48:2 is an icosahedral membrane-containing phage, thus connecting it further with FLiP, which is the sole member of family Finnlakeviridae. Moreover, FLiP and phi48:2 virions are both ~60 nm in diameter and showed some similarity in their major capsid protein sequences (~21% amino acid identity). To conclude, even though phi48:2 and FLiP share various similarities they cannot be placed within the same family due to the low similarity in their genome sequences. However, for now we can assume they are possible distant relatives. The diversity and abundancy of membrane-containing ssDNA phages is gradually starting to uncover and through their characterization and classification we might consequently understand better their significance in microbial ecology.