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The oleaginous bacterium  Rhodococcus opacus  PD630 is metabolically diverse and can be cultivated on various renewable resources to serve as a sustainable triacylglycerol (TAG) feedstock for biodiesel production. Current methods for TAG extraction are costly, but infection of cultures by lytic bacteriophages (phages) may be a viable approach for achieving release of intracellular lipid from oleaginous bacteria such as  R. opacus . This study reports the novel tectiviral phage Toil capable of releasing intracellular contents including a fluorescent protein marker and TAGs into the supernatant after phage infection of  R. opacus  PD631, a domesticated derivative of strain PD630. Phage Toil is placed in the  Tectiviridae  by its morphology, the presence of a lipid membrane, its genome architecture and the presence of terminal covalently-linked proteins. Toil is the first tectivirus capable of infecting a member of the  Actinobacteria . Microscopy shows that infected cells do not undergo sudden lysis but instead maintain their original shape for several hours, with the cellular morphology gradually deteriorating. Approximately 30% of intracellular TAGs could be recovered from the culture supernatants of Toil-infected PD631 cells. Phage Toil has potential to be used as an agent in extraction of TAGs from oleaginous bacterium  R. opacus .  Importance : This study reported the first tectivirus (Phage Toil) capable of infecting a member of the  Actinobacteria . In this study, we showed that Phage Toil can infect oleaginous bacterium  Rhodococcus opacus  to release intracellular contents such as a fluorescent protein marker and TAG lipid granules, which can serve as a starting material for biodiesel production. This study demonstrates a new method to extract TAGs by using this phage. Additionally, Phage Toil can be a new model phage to advance knowledge regarding phage infection mechanisms in  Rhodococcus  and other mycolic acid-containing bacteria such as  Mycobacterium .

Characterization of a Novel Tectivirus Phage Toil and Its Potential as an Agent for Biolipid Extraction