Surfactin production is not essential for pellicle and root-associated biofilm development ofBacillus subtilis

Abstract

20Secondary metabolites have an important impact on the biocontrol potential of soil-21 derived microbes. In addition, various microbe-produced chemicals have been 22 suggested to impact the development and phenotypic differentiation of bacteria, 23including biofilms. The non-ribosomal synthesized lipopeptide of Bacillus subtilis, 24 surfactin, has been described to impact the plant promoting capacity of the bacterium. 25Here, we investigated the impact of surfactin production on biofilm formation of B. 26 subtilis using the laboratory model systems; pellicle formation at the air-medium 27 interface and architecturally complex colony development, in addition to plant root-28 associated biofilms. We found that the production of surfactin by B. subtilis is not 29 essential for pellicle biofilm formation neither in the well-studied strain, NCIB 3610, nor 30 in the newly isolated environmental strains, but lack of surfactin reduces colony 31 expansion. Further, plant root colonization was comparable both in the presence or 32 absence of surfactin synthesis. Our results suggest that surfactin-related biocontrol and 33 plant promotion in B. subtilis strains are independent of biofilm formation. 34 35 Keywords: Bacillus subtilis, biofilm, surfactin, plant root colonization, pellicle 36 39 Several species from the "Bacillus subtilis complex" are well-characterized plant growth-40 promoting rhizobacteria (PGPRs), providing various beneficial activities for plants and 41 inhibiting fungal and bacterial pathogens [1]. Many strains of Bacillus subtilis, Bacillus 42 amyloliquefaciens and Bacillus velezensis are currently used in organic and traditional 43 agriculture to prevent infection and/or increase yields of various crops [2-4]. These 44 species are of particular interest because they can form stress-resistant endospores, a 45 cell-type ideal for product formulation. Most PGPR Bacillus spp. also produce a wide 46 range of bioactive molecules, such as lipopeptides, which directly influences plant growth 47 and defence [5]. 48 49 Many of these molecules are synthesized by multienzyme-complexes called non-50 ribosomal peptide synthetases (NRPS) [6]. B. subtilis NCIB3610 possesses 3 NRPS 51 clusters and one NRPS/polyketide synthetase (PKS) cluster, which is few compared to 52 the bioactive molecule synthesis capacity of B. velezensis strains [1]. Bacillaene, a broad 53 spectrum antibiotic, is synthesized by proteins encoded in 80 kB pksA-S cluster [7]. The 54ppsA-E encodes for the peptide synthetase responsible for the synthesis of plipastatin 55 (fengycin family), a strong antifungal molecule [5,8], while the siderophore bacillibactin is 56 synthesized by the product of the dhbA-F operon [9]. Finally, SrfAA-AD produces versatile 57 molecules from the surfactin family [10]. 58 59Surfactin molecules are composed of a heptapeptide, i.e. two acidic and five nonpolar 60 amino acids, interlinked with a β-hydroxy fatty acid, and condensed in a cyclic lactone 61 right structure [10,11]. The amino acid sequence, the length, and the ...

The ComX Quorum Sensing Peptide of Bacillus subtilis Affects Biofilm Formation Negatively and Sporulation Positively

“…Our results are in accordance with recent findings, indicating that surfactin is not generally essential for the formation of pellicle biofilms [ 73 ] with surfactin mutants exhibiting similar shifts in the expression of epsA and tapA operons, as we show here for the QS mutant. However, the results presented here suggest that the main regulatory path of biofilm development may include ComX and only subsequently surfactin.…”

Surfactin stimulated by pectin molecular patterns and root exudates acts as a key driver ofBacillus-plant mutualistic interaction

“…This could be of clear ecological benefit for the bacterium since surfactin is known to favor motility of multicellular communities and biofilm formation [ 16,28,29 ]. However, a recent study questioned the real role of surfactin in these key functions, since it production appears as non-essential for pellicle biofilm formation in B. subtilis NCIB 3610, suggesting a strain dependant role [ 30 ]. We previously reported that motility and biofilm formation are boosted upon growth on root exudates [ 27 ].…”

Deletion of Rap-Phr systems inBacillus subtilisinfluencesin vitrobiofilm formation and plant root colonization

“…These experiments thus support that biofilm formation in vitro as well as on plant roots is influenced by additional factors than just matrix gene expression. For example surfactin production, which is regulated by ComA -a target of several Rap phosphatases -was shown to influence the colony structure of B. subtilis 3610 on MSgg, though this secondary metabolite was not essential for pellicle formation and root colonization (Thérien et al, 2020).…”