Patterns of reporter gene expression in the phase diagram of Bacillus subtilis colony forms

Abstract

Factors governing the morphogenesis of Bacillus subtilis colonies as well as the spatial-temporal pattern of expression of a reporter gene during colony development were examined by systematically varying the initial nutrient levels and agar concentrations (wetness), the relative humidity throughout incubation, and the genotype of the inoculum. A relationship between colony form and reporter gene expression pattern was found, indicating that cells respond to local signals during colony development as well as global conditions. The most complex colony forms were produced by motile strains grown under specific conditions such that cells could swim within the colony but not swarm outward uniformly from the colony periphery. The wetness of the growth environment was found to be a critical factor. Complex colonies consisted of structures produced by growth of finger-like projections that expanded outward a finite distance before giving rise to a successive round of fingers that behaved in a similar fashion. Finger tip expansion occurred when groups of cells penetrated the peripheral boundary. Although surfactin production was found to influence similar colony forms in other B. subtilis strains, the strains used here to study reporter gene expression do not produce it. The temporal expression of a reporter gene during morphogenesis of complex colonies by motile strains such as M18 was investigated. Expression arose first in cells located at the tips of fingers that were no longer expanding. The final expression pattern obtained reflects the developmental history of the colony.

The interaction of thin-film flow, bacterial swarming and cell differentiation in colonies of Serratia liquefaciens

“…The biosurfactant allows a continuous expansion of the wetting-agent and thus the bacterial colony. Mendelson & Salhi (1996) demonstrate that Bacillus subtilis mutants that are unable to produce surfactin are defective in swarming motility. Furthermore, Eberl et al (1999) state that &a non-#agellated mutant of S. liquefaciens MG1 is capable of colonizing the surface of low percentage agar by means of spreading motility (which is solely driven by the biosurfactant) as has been observed previously with S.…”

Branched swarming patterns on a synthetic medium formed by wild-type Bacillus subtilis strain 3610: detection of different cellular morphologies and constellations of cells as the complex architecture develops

“…An antibiotic surfactant, whose synthesis is subject to control by quorum sensing (Solomon et al, 1995;Solomon & Grossman, 1996), has also been implicated in swarming in this organism (Kearns & Losick, 2003;Bees et al, 2000; see also Wakita et al, 1998). However, opposing results have been obtained concerning the importance of surfactin in swarming (Mendelson & Salhi, 1996;Dixit et al, 2002). In fact, the mechanism of all aspects of swarming, including the production and nature of presumed 'swarmer' cells, as found in various Gram-negative bacteria, remains poorly understood in B. subtilis.…”

“…Patterns with pronounced chirality, or repeating concentric zones, were also observed. Whilst the concentration of nutrients influenced patterns, the agar concentration appeared most important in determining bacterial swarming per se (Mendelson & Salhi, 1996), leading to the conclusion that swarming requires the extraction of water from the agar or reduction of surface friction, processes in which surfactants presumably play a role (Matsuyama et al, 1989;Matsuyama & Matsushita, 2001; Bees et al, 2000).…”

“…Bacillus subtilis, inoculated from a central point, is able rapidly to colonize large areas of nutrient agar (0?6-1?5 %) by a coordinated, symmetrical migration or swarming over the surface (Ben-Jacob et al, 1994;Matsushita, 1997;Mendelson & Salhi, 1996; see also Shapiro, 1995). Swarming may take several forms, including the production of highly branched, fractal patterns (Matsuyama et al, 1989;Vicsek, 1992).…”

“…In order to analyse the effect of swarming on the interplay between nutrient concentration and percentage of agar, most previous studies have used complex medium with laboratory strains of B. subtilis, in particular strain 168, grown on peptone (Ben-Jacob et al, 1994;Matsushita, 1997), beef extract/tryptose (Mendelson & Salhi, 1996) and LB (Dixit et al, 2002). These studies revealed different swarming patterns, from amorphous, largely confluent growth zones, through relatively simple radial branching, to highly complex, densely branched structures.…”