Deletion of Rap‐Phr systems in Bacillus subtilis influences in vitro biofilm formation and plant root colonization

Abstract

Natural isolates of the soil‐dwelling bacterium Bacillus subtilis form robust biofilms under laboratory conditions and colonize plant roots. B. subtilis biofilm gene expression displays phenotypic heterogeneity that is influenced by a family of Rap‐Phr regulatory systems. Most Rap‐Phr systems in B. subtilis have been studied independently, in different genetic backgrounds and under distinct conditions, hampering true comparison of the Rap‐Phr systems’ impact on bacterial cell differentiation. Here, we investigated each of the 12 Rap‐Phr systems of B.subtilis NCIB 3610 for their effect on biofilm formation. By studying single ∆ rap ‐ phr mutants, we show that despite redundancy between the cell–cell communication systems, deletion of each of the 12 Rap‐Phr systems influences matrix gene expression. These Rap‐Phr systems therefore enable fine‐tuning of the timing and level of matrix production in response to specific conditions. Furthermore, some of the ∆ rap ‐ phr mutants demonstrated altered biofilm formation in vitro and colonization of Arabidopsis thaliana roots, but not necessarily similarly in both processes, indicating that the pathways regulating matrix gene expression and other factors important for biofilm formation may be differently regulated under these distinct conditions.

Adaptation of Bacillus thuringiensis to Plant Colonization Affects Differentiation and Toxicity

“…EE of B. thuringiensis 407 Cry − (Bt407) was initiated using six parallel lineages colonizing the roots of 7-day-old A. thaliana seedlings under hydroponic conditions generally applied for Bacilli ( 46 – 50 ). Initially, inoculated planktonic bacterial cells developed biofilms on plant roots.…”

Experimental evolution of Bacillus subtilis on Arabidopsis thaliana roots reveals fast adaptation and improved root colonization

“…This setup was inspired by a long-term EE on polystyrene beads ( Poltak and Cooper, 2011 ), but instead of beads, A. thaliana roots were used for successive root colonization by B. subtilis . The EE on plant seedlings was carried out under axenic, hydroponic conditions optimized for B. subtilis ( Beauregard et al., 2013 ; Dragoš et al., 2018a ; Gallegos-Monterrosa et al., 2016 ; Nordgaard et al., 2021 ; Thérien et al., 2020 ). Seven parallel populations were initiated by inoculating A. thaliana seedlings, placed in 300 μL MSNg medium in 48-well plates, with B. subtilis DK1042 at a starting OD 600 of 0.02.…”

Experimental evolution ofBacillus subtilisonArabidopsis thalianaroots reveals fast adaptation and improved root colonization in the presence of soil microbes

“…This setup was inspired by a longterm EE on polystyrene beads (Poltak and Cooper, 2011), but instead of beads, A. thaliana roots were used for successive root colonization by B. subtilis. The EE on plant seedlings was carried out under axenic, hydroponic conditions optimized for B. subtilis (Beauregard et al, 2013;Dragoš et al, 2018a;Gallegos-Monterrosa et al, 2016;Nordgaard et al, 2021;Thérien et al, 2020). Seven parallel populations were initiated by inoculating A. thaliana seedlings, placed in 300 µL MSNg medium in 48-well plates, with B.…”