Osmotic stimulation of microcolony development by Nitrosomonas europaea

In liquid cultures low levels of salt (0.1 M NaCl) had no effect on the growth rate of proliferating cells of Nitrosomonas europaea (NCIMB 11850), while at higher salt concentrations the growth rate was progressively reduced. A technique which enabled enumeration of microcolony-forming cells after just a few days incubation on the surface of membrane filters was subsequently used to investigate the effects of medium osmolarity and osmoprotectants on the ability to initiate cell division in single cells from liquid cultures. Typically, only ∼50% of the cells from early stationary phase cultures in basal medium was able to initiate cell division and form microcolonies during subsequent incubation on filters on the same medium. However, the fraction of cells forming microcolonies was stimulated by low levels of salt, while higher concentrations were inhibitory. A range of bacterial compatible solutes (betaine, trehalose, proline, MOPS, taurine and GABA) were unable to enhance microcolony formation at inhibitory salt concentrations. Other solutes (KCl, LiCl, mannitol, sucrose) also stimulated microcolony development, demonstrating that the effect was osmotic. A low salt (0.1 M NaCl) shock prior to incubation at inhibitory salt levels dramatically increased the number of cells forming microcolonies. A comparative study of several Nitrosospira spp. indicated that osmotic stimulation of microcolony development may be specific to N. europaea, although the proportion of cells able to form microcolonies was much smaller in the Nitrosospira spp. examined. Taken together these results suggest that while inhibition of growth in N. europaea by high osmolarity may be similar to that observed in other bacteria, the osmotic stimulation observed for initiation of cell division may have important implications for the successful establishment and recovery of this nitrifier in biofilms and in soil.