Rod-shaped bacteria typically elongate and divide by transverse fission. However, several bacterial species can form rod-shaped cells that divide longitudinally. Here, we study the evolution of cell shape and division mode within the family Neisseriaceae, which includes Gram-negative coccoid and rod-shaped species. In particular, bacteria of the genera Alysiella, Simonsiella and Conchiformibius, which can be found in the oral cavity of mammals, are multicellular and divide longitudinally. We use comparative genomics and ultrastructural microscopy to infer that longitudinal division within Neisseriaceae evolved from a rod-shaped ancestor. In multicellular longitudinally-dividing species, neighbouring cells within multicellular filaments are attached by their lateral peptidoglycan. In these bacteria, peptidoglycan insertion does not appear concentric, i.e. from the cell periphery to its centre, but as a medial sheet guillotining each cell. Finally, we identify genes and alleles associated with multicellularity and longitudinal division, including the acquisition of amidase-encoding gene amiC2, and amino acid changes in proteins including MreB and FtsA. Introduction of amiC2 and allelic substitution of mreB in a rod-shaped species that divides by transverse fission results in shorter cells with longer septa. Our work sheds light on the evolution of multicellularity and longitudinal division in bacteria, and suggests that members of the Neisseriaceae family may be good models to study these processes due to their morphological plasticity and genetic tractability.

Chemoautotrophic endosymbionts are famous for exploiting sulfur oxidization to feed marine organisms with fixed carbon. However, the physiology of thiotrophic bacteria thriving on the surface of animals (ectosymbionts) is less understood.

57Chemosynthetic symbioses occur worldwide in marine habitats. However, physiological 58 studies of chemoautotrophic bacteria thriving on animals are scarce. Stilbonematinae are 59 coated by monocultures of thiotrophic Gammaproteobacteria. As these nematodes migrate 60 through the redox zone, their ectosymbionts experience varying oxygen concentrations. 61Here, by applying omics, Raman microspectroscopy and stable isotope labeling, we 62 investigated the effect of oxygen on the metabolism of Candidatus Thiosymbion oneisti. 63Unexpectedly, sulfur oxidation genes were upregulated in anoxic relative to oxic conditions, 64 but carbon fixation genes and incorporation of 13 C-labeled bicarbonate were not. Instead, 65several genes involved in carbon fixation, the assimilation of organic carbon and 66 polyhydroxyalkanoate (PHA) biosynthesis, as well as nitrogen fixation and urea utilization 67 were upregulated in oxic conditions. Furthermore, in the presence of oxygen, stress-related 68 genes were upregulated together with vitamin and cofactor biosynthesis genes likely 69 necessary to withstand its deleterious effects. 70Based on this first global physiological study of a chemosynthetic ectosymbiont, we 71propose that, in anoxic sediment, it proliferates by utilizing nitrate to oxidize reduced sulfur, 72whereas in superficial sediment it exploits aerobic respiration to facilitate assimilation of 73 carbon and nitrogen to survive oxidative stress. Both anaerobic sulfur oxidation and its 74 decoupling from carbon fixation represent unprecedented adaptations among 75 chemosynthetic symbionts. 76 77 intracorporeal thiotrophic bacteria, it is generally accepted that the endosymbiont's 85 chemosynthetic metabolism serves to provide organic carbon for feeding the animal host 86[reviewed in 1-3]. In addition, some chemosynthetic symbionts have been found capable to 87 fix atmospheric nitrogen, albeit symbiont-to-host transfer of fixed nitrogen remains unproven 88 [4, 5]. As for the rarer chemosynthetic bacterial-animal associations in which symbionts 89 colonize exterior surfaces (ectosymbionts), fixation of inorganic carbon and transfer of 90 organic carbon to the host has only been demonstrated for the microbial community 91 colonizing the gill chamber of the hydrothermal vent shrimp Rimicaris exoculata [6]. 92In this study, we focused on Candidatus Thiosymbion oneisti, a 93Gammaproteobacterium belonging to the basal family of Chromatiaceae (also known as 94 purple sulfur bacteria), which colonizes the surface of the marine nematode Laxus oneistus 95 (Stilbonematinae). Curiously, this group of free-living roundworms represents the only known 96 animals engaging in monospecific ectosymbioses, i.e. each nematode species is 97 ensheathed by a single Ca. Thiosymbion phylotype, and, in the case of Ca. T. oneisti, the 98 bacteria form a single layer on the cuticle of its host [7-11]. Moreover, the rod-shaped 99 representatives of this bacterial genus, including Ca. T. oneisti, divide by FtsZ-based 100 longitudinal fission, a unique ...

Highlights d Symbiont sister ori segregate along the short and the long cell axis (diagonally) d ParB recapitulates ori localization and binds an ori-proximal parS site in vitro d Septal sister ter migrate to midcell concomitantly with septation progression d Bidimensional segregation endows maintenance of chromosome configuration

Summary Less than a handful of cuboid and squared cells have been described in nature, which makes them a rarity. Here, we show how Candidatus Thiosymbion cuboideus, a cube-like gammaproteobacterium, reproduces on the surface of marine free-living nematodes. Immunostaining of symbiont cells with an anti-fimbriae antibody revealed that they are host-polarized, as these appendages exclusively localized at the host-proximal (animal-attached) pole. Moreover, by applying a fluorescently labeled metabolic probe to track new cell wall insertion in vivo , we observed that the host-attached pole started septation before the distal one. Similarly, Ca. T. cuboideus cells immunostained with an anti-FtsZ antibody revealed a proximal-to-distal localization pattern of this tubulin homolog. Although FtsZ has been shown to arrange into squares in synthetically remodeled cuboid cells, here we show that FtsZ may also mediate the division of naturally occurring ones. This implies that, even in natural settings, membrane roundness is not required for FtsZ function.