0 Intra-species Competition in Serratia marcescens is Mediated by Type VI Secretion Rhs 1 Effectors and a Conserved Effector-Associated Accessory Protein 2 3

ABSTRACT 13 The Type VI secretion system (T6SS) is widespread in Gram-negative bacteria and can deliver 14 toxic effector proteins into eukaryotic cells or competitor bacteria. Anti-bacterial T6SSs are 15 increasingly recognised as key mediators of interbacterial competition and may contribute to the 16 outcome of many polymicrobial infections. Multiple anti-bacterial effectors can be delivered by 17 these systems, with diverse activities against target cells and distinct modes of secretion. 18 Polymorphic toxins containing Rhs repeat domains represent a recently-identified and as-yet 19 poorly characterised class of T6SS-dependent effector. Previous work had revealed that the 20 potent anti-bacterial T6SS of the opportunistic pathogen Serratia marcescens promotes intra-21 species as well as inter-species competition. In this study, two new Rhs family anti-bacterial 22 effectors delivered by this T6SS have been identified. One of these was shown to act as a DNase 23 toxin whilst the other contains a novel, cytoplasmic-acting toxin domain. Importantly, using S. 24 marcescens, it has been demonstrated for the first time that Rhs proteins, rather than other T6SS-25 secreted effectors, can be the primary determinant of intra-species competition. Furthermore, a 26 new family of accessory proteins associated with T6SS effectors has been identified, exemplified 2