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PslG, a self-produced glycosyl hydrolase, triggers biofilm disassembly by disrupting exopolysaccharide matrix.

ABSTRACT: Biofilms are surface-associated communities of microorganism embedded in extracellular matrix. Exopolysaccharide is a critical component in the extracellular matrix that maintains biofilm architecture and protects resident biofilm bacteria from antimicrobials and host immune attack. However, self-produced factors that target the matrix exopolysaccharides, are still poorly understood. Here, we show that PslG, a protein involved in the synthesis of a key biofilm matrix exopolysaccharide Psl in Pseudomonas aeruginosa, prevents biofilm formation and disassembles existing biofilms within minutes at nanomolar concentrations when supplied exogenously. The crystal structure of PslG indicates the typical features of an endoglycosidase. PslG mainly disrupts the Psl matrix to disperse bacteria from biofilms. PslG treatment markedly enhances biofilm sensitivity to antibiotics and macrophage cells, resulting in improved biofilm clearance in a mouse implant infection model. Furthermore, PslG shows biofilm inhibition and disassembly activity against a wide range of Pseudomonas species, indicating its great potential in combating biofilm-related complications.

SUBMITTER: Yu S

PROVIDER: S-EPMC4670989 | biostudies-literature | 2015 Dec

REPOSITORIES: biostudies-literature

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PslG, a self-produced glycosyl hydrolase, triggers biofilm disassembly by disrupting exopolysaccharide matrix.

Yu Shan S Su Tiantian T Wu Huijun H Liu Shiheng S Wang Di D Zhao Tianhu T Jin Zengjun Z Du Wenbin W Zhu Mei-Jun MJ Chua Song Lin SL Yang Liang L Zhu Deyu D Gu Lichuan L Ma Luyan Z LZ

Cell research 20151127 12

Biofilms are surface-associated communities of microorganism embedded in extracellular matrix. Exopolysaccharide is a critical component in the extracellular matrix that maintains biofilm architecture and protects resident biofilm bacteria from antimicrobials and host immune attack. However, self-produced factors that target the matrix exopolysaccharides, are still poorly understood. Here, we show that PslG, a protein involved in the synthesis of a key biofilm matrix exopolysaccharide Psl in Pse ...[more]

PMID: 26611635

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Project description:Histophilus somni, a gram-negative coccobacillus, is an obligate inhabitant of bovine and ovine mucosal surfaces, and an opportunistic pathogen responsible for respiratory disease and other systemic infections in cattle and sheep. Capsules are important virulence factors for many pathogenic bacteria, but a capsule has not been identified on H. somni. However, H. somni does form a biofilm in vitro and in vivo, and the biofilm matrix of most bacteria consists of a polysaccharide.Following incubation of H. somni under growth-restricting stress conditions, such as during anaerobiosis, stationary phase, or in hypertonic salt, a polysaccharide could be isolated from washed cells or culture supernatant. The polysaccharide was present in large amounts in broth culture sediment after H. somni was grown under low oxygen tension for 4-5 days (conditions favorable to biofilm formation), but not from planktonic cells during log phase growth. Immuno-transmission electron microscopy showed that the polysaccharide was not closely associated with the cell surface, and was of heterogeneous high molecular size by gel electrophoresis, indicating it was an exopolysaccharide (EPS). The EPS was a branched mannose polymer containing some galactose, as determined by structural analysis. The mannose-specific Moringa M lectin and antibodies to the EPS bound to the biofilm matrix, demonstrating that the EPS was a component of the biofilm. The addition of N-acetylneuraminic acid to the growth medium resulted in sialylation of the EPS, and increased biofilm formation. Real-time quantitative reverse transcription-polymerase chain reaction analyses indicated that genes previously identified in a putative polysaccharide locus were upregulated when the bacteria were grown under conditions favorable to a biofilm, compared to planktonic cells.H. somni is capable of producing a branching, mannose-galactose EPS polymer under growth conditions favorable to the biofilm phase of growth, and the EPS is a component of the biofilm matrix. The EPS can be sialylated in strains with sialyltransferase activity, resulting in enhanced density of the biofilm, and suggesting that EPS and biofilm formation may be important to persistence in the bovine host. The EPS may be critical to virulence if the biofilm state is required for H. somni to persist in systemic sites.

Project description:Pseudomonas aeruginosa is an important pathogen that causes chronic infections that involve multicellular aggregates called biofilms. Within biofilms, bacteria are surrounded in a protective extracellular matrix of proteins, exopolysaccharides (EPS), and DNA. A key P. aeruginosa matrix protein is an extracellular adhesin called CdrA, which promotes aggregation by binding to the EPS Psl and via CdrA-CdrA interactions. We hypothesized that because of its ability to bind Psl, CdrA would be important only for strains that use Psl as the primary EPS (e.g., the laboratory strain PAO1). Thus, we predicted that cdrA might be dispensable for biofilm formation by strains that do not utilize Psl (e.g., the laboratory strain PA14). Instead, we observed that cdrA deletion strains exhibited biofilm defects, regardless of their EPS dependencies. We screened a panel of clinical and environmental P. aeruginosa isolates for the presence of the cdrA allele and production of CdrA protein. All isolates that we tested contained the cdrA allele, and these alleles had minimal sequence variation compared to the reference PAO1 cdrA gene. Additionally, all isolates except one produced detectable CdrA protein. We investigated the possible mechanisms of CdrA-promoted biofilm formation in these strains where Psl is not dominant, and we discovered that CdrA binds to Pel. Although Psl and Pel chemical structures are distinct, this appears to be a specific interaction, since previous work has shown that CdrA binds discriminately to other EPS. Our findings provide new understanding of biofilm formation across P. aeruginosa isolates and emphasize the versatility of CdrA.IMPORTANCE Depending upon the strain, Pseudomonas aeruginosa can use different exopolysaccharides (e.g., Psl, Pel, and alginate) to build its biofilm matrix. Previously, we demonstrated that the biofilm matrix protein CdrA binds to Psl, promoting biofilm formation and aggregate stability. As such, it was thought that CdrA might be important for biofilm assembly only in strains that rely upon Psl. However, past studies indicated that CdrA can interact with monosaccharides not present in Psl, including N-acetylglucosamine, a constituent of another EPS called Pel. We discovered that CdrA also binds to Pel and promotes biofilm formation by strains in which Psl is not dominant. Thus, our findings suggest that CdrA plays a common role as a biofilm matrix cross-linker across P. aeruginosa isolates with different EPS.

Dataset Information

PslG, a self-produced glycosyl hydrolase, triggers biofilm disassembly by disrupting exopolysaccharide matrix.

Publications

PslG, a self-produced glycosyl hydrolase, triggers biofilm disassembly by disrupting exopolysaccharide matrix.

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