Mutations in Gordonia amarae mycolic acid biosynthetic pathway confer resistance to Patescibacteria parasite Mycosynbacter amalyticus.

The obligate necrotrophic parasite, Candidatus Mycosynbacter amalyticus, a member of the Patescibacteria has been isolated from wastewater. Subsequent efforts have been directed toward unravelling its biological lifecycle and attachment mechanism facilitating infection and subsequent lysis of its Actinobacterial host, Gordonia amarae. Here, using electron cryo-tomography (CryoET), we reveal the molecular anatomy of parasitic Mycosynbacter amalyticus cells, uncovering an unusual infection process. Through laboratory-based evolution experiments, we generated eleven slow-growing independent spontaneous Gordonia amarae resistant mutants. Mycolic acids (MA) are key components of the outer cellular envelope of G. amarae and other Actinobacteria, with MA being the physical attribute implicated in G. amarae associated wastewater foaming. CryoET and genome sequencing exposed absence of intact MA and an associated suite of mutations predominantly occurring within the pks13 and pptT genes of the MA biosynthetic pathway. Our findings suggest that MA structural integrity is critical for attachment of Ca. Mycosynbacter amalyticus to its host.