Molecular Basis of Wrinkled Variants Isolated From Pseudoalteromonas lipolytica Biofilms

Abstract

Many Pseudoalteromonas species are dominant biofilm-forming Gammaproteobacteria in the ocean. The formation of Pseudoalteromonas biofilms is often accompanied by the occurrence of variants with different colony morphologies that may exhibit increased marine antifouling or anticorrosion activities. However, the genetic basis of the occurrence of these variants remains largely unexplored. In this study, we identified that wrinkled variants of P. lipolytica mainly arose due to mutations in the AT00_08765, a wspF-like gene, that are associated with decreased swimming motility and increased cellulose production. Moreover, we found that the spontaneous mutation in flhA, encoding a flagellar biosynthesis protein, also caused a wrinkled colony morphology that is associated with cellulose overproduction, indicating that flhA plays a dual role in controlling flagellar assembly and polysaccharide production in P. lipolytica. Investigation of wrinkled variants harboring spontaneous mutation in dgcB, encoding a GGDEF domain protein, also demonstrated dgcB plays an important role in regulating cellulose production and swimming motility. In addition, by screening the suppressor of the AT00_08765 variant strain, we also identified that the spontaneous mutation in cheR and bcsC directly abolished the wrinkled phenotype of the AT00_08765 variant strain, suggesting that the chemosensory signaling transduction and cellulose production are crucial for the determination of the wrinkled phenotype in P. lipolytica. Taken together, this study provides insights into the genetic variation within biofilms of P. lipolytica.

Evolution of cyclic di-GMP signalling on a short and long term time scale

“…Intriguingly, the Wsp chemotactic signalling pathway and the YfiBNR system are also major targets of mutational events leading to the development of small colony variants of P. aeruginosa upon colonization of the cystic fibrosis lung [ 150, 151 ] and in rhizosphere-colonizing Pseudomonas chlororaphis [ 152 ]. Similarly, mutations in the Wsp chemotactic pathway arise readily upon chronic infection in a full-thickness burn wound model [ 153 ], the challenge of P. aeruginosa with sub-lethal concentrations of hydrogen peroxide, the subjection of Burkholderia cenocepacia to a heterogeneous microcosm [ 154 ], other infections where hyper biofilm producers with elevated cyclic di-GMP levels are observed [ 155 ] and also upon biofilm formation of oceanic Pseudoalteromonas lipolytica [ 156 ]. An unexpected twist is a point mutation in the 5′ untranslated region of the fatty acid biosynthesis accBC gene cluster which might activate the Wsp signalling pathway by alteration of the fatty acid composition of the membrane [ 157 ].…”

Molecular basis of the phenotypic variants arising from a Pseudoalteromonas lipolytica mutator

“…We previously also found that wrinkled variants isolated from P. lipolytica biofilms harbour spontaneous mutations in the flagellar regulatory gene wspF or flagellar biosynthesis gene flhA , leading to a reduction in swimming activity [21, 25]. However, there is still limited knowledge linking reduced swimming motility to the specific hot mutation targets introduced by the mutS deletion.…”

“…For example, during long-term colonization in adult mice, spontaneous nonmotile variants of V. cholerae were highly produced at a rate of approximately 10 %, in which the mutations occurred primarily in conserved regions of the flagellar regulatory genes flrA and flrC [40]. We previously also found that wrinkled variants isolated from P. lipolytica biofilms harbour spontaneous mutations in the flagellar regulatory gene wspF or flagellar biosynthesis gene flhA, leading to a reduction in swimming activity [21,25]. However, there is still limited knowledge linking reduced swimming motility to the specific hot mutation targets introduced by the mutS deletion.…”

“…It has gained considerable attention due to its ecological significance by producing a large number of active compounds [19,20]. In a previous study, we demonstrated that the motile marine bacterium Pseudoalteromonas lipolytica produced different genetic variants during biofilm formation, such as wrinkled, pigment production and translucent colony morphology variants, which had significant marine antifouling and anticorrosion applications [21][22][23][24][25][26]. The alteration of colony morphology is often tightly linked to the bacterial mutator phenotype [27].…”

Stimulation of the Biofilms of a marine bacterium Pseudoalteromonas agarivorans by Fucoidan extracted from macroalgae

“…Bio lm is a special matrix of bacteria that live as a community in close contact enabling several advantages like cell to cell communication, digestion of nutrients and protection for vital activities including growth, reproduction, and functional activity in the marine environment (Drescher et al 2014; Zeng et al 2022). The microbial cells inside the bio lm structure are supported by the secretion of extracellular enzymes and polymeric substances to digest and take nutrients, respectively, from the marine environment (Flemming and Wingender 2010).…”