Differential Biofilm Formation and Motility Associated with Lipopolysaccharide/Exopolysaccharide-Coupled Biosynthetic Genes inStenotrophomonas maltophilia

Abstract

Stenotrophomonas maltophilia WR-C is capable of forming biofilm on polystyrene and glass. The lipopolysaccharide/exopolysaccharide-coupled biosynthetic genes rmlA, rmlC, and xanB are necessary for biofilm formation and twitching motility. Mutants with mutations in rmlAC and xanB display contrasting biofilm phenotypes on polystyrene and glass and differ in swimming motility.Microorganisms can develop biofilms or clogging mats, causing the failure of septic tanks, systems for on-site wastewater disposal. If water in these clogged systems were contaminated by pathogens, it would pose a threat to human health. We isolated Stenotrophomonas maltophilia strain WR-C from a clogged septic tank system that consistently formed biofilms on sand grains, produced exopolysaccharides (EPS), and caused clogging in sand columns (33). S. maltophilia is a gram-negative, rod-shaped, and obligate aerobic bacterium with polar flagella in the ␥-␤ subdivision of Proteobacteria (3,14). It is found in various environments and recently has emerged as an important human pathogen. Very little is known about the mechanisms of biofilm formation by S. maltophilia. It has been shown to adhere to HEp-2 cells as well as abiotic surfaces, such as plastic, glass, and Teflon (7-10, 16).To identify genes that are involved in biofilm formation by S. maltophilia WR-C, about 4,500 transposon mutants generated with the EZ::TN Ͻ R6K␥ori/KAN-2 Ͼ Tnp transposome (Epicenter, Madison, Wis.) were screened for defects in biofilm formation in 96-well polystyrene plates (Becton-Dickinson Labware, Franklin Lakes, N.J.) by using a modified microtiter plate assay (11,29). Briefly, wells containing 200 l Trypticase soy (TS) broth were inoculated with overnight cultures and incubated at 30°C, 50 rpm for 2 days. Biofilm cells were stained with 0.1% crystal violet and washed, the stain remaining in the cells was solubilized with 70% ethanol, and the optical density at 590 nm was determined. Three mutants, TPH7, TPH11, and TPH13, whose growth was similar to that of the wild type but which were deficient in biofilm formation, were selected for further characterization.The transposon flanking regions were rescued by "rescue cloning" as described by the manufacturer and sequenced by using the primers KAN-2 FP-1 and R6KAN-2 RP-1 (Table 1). The transposon-inserted genes are homologous to three genes involved in the biosynthesis of nucleotide sugar precursors of lipopolysaccharide (LPS) and EPS in Xanthomonas campestris pv. campestris ATCC 33913 (GenBank accession no. AE008922) (6). The genes are rmlC (for TPH7, 74% identity), rmlA (for TPH11, 78% identity), and xanB (for TPH13, 80% identity), which are located in the rml and xan operons of X. campestris pv. campestris (19,20). As no genome sequence is available and the gene cluster involved in LPS/EPS biosynthesis has not been reported for S. maltophilia, the DNA segments flanking the transposon in TPH7, TPH11, and TPH13 were sequenced. The complete sequences of the rmlBACD and xanAB operons and their flanking genes were deter...

Biofilm Production by Multi Drug Resistant Bacterial Pathogens Isolated From Patients in Intensive Care Units in Egyptian Hospitals

“…Mutants for the glucose-1-phosphate thymidyltransferase rmlA gene and for the cis-11-methyl-2-dodecenoic acid rpfF gene were reported to decreases biofilm formation [12,13]. Further, the spgM gene, encoding a bifunctional enzyme with both phosphoglucomutase (PGM) and phosphomannomutase activities, is involved in biofilm-forming ability in S. maltophilia [14].…”

Molecular detection of transcriptionally active bacteria from failed prosthetic hip joints removed during revision arthroplasty

“…In fact, it has recently been demonstrated that various Lysobacter-type species have the ability to readily form biofilms on various substrates. These species include Stenotrophomonas maltophilia, Xylella fastidiosa and Xanthomonas axonopodis [40][41][42]. Therefore, it is unsurprising that these species were identified on the prostheses of the patients in our study.…”

Surface Polysaccharide Mutants Reveal that Absence of O Antigen Reduces Biofilm Formation of Actinobacillus pleuropneumoniae

“…While other groups have shown that the LPS core oligosaccharide is essential for biofilm formation by E. coli, Klebsiella pneumoniae, and S. maltophilia (42)(43)(44), our LPS core mutant, strain CG3, was still able to produce biofilms similarly to the parental A. pleuropneumoniae strain (Fig. 1).…”

“…citri O-antigen LPS) impaired biofilm formation on plastic surfaces and host plant leaves (41). The formation of Stenotrophomonas maltophilia biofilms requires an intact LPS, and a mutation in O-antigen biosynthesis affects EPS production and biofilm formation (42). Clifford and coworkers showed that the O antigen of Xylella fastidiosa is needed for biofilm maturation and surface attachment (50).…”

“…Surface polysaccharides, including LPS, CPS, and PGA, have been linked to biofilm formation in several bacterial species (17,25,(40)(41)(42)(43)(44)(45)(46)(47)(48)(49)(50)(51); thus, investigation of their unassessed role in the biofilm formation of A. pleuropneumoniae serotype 1 was the focus of our present study. LPS and CPS have been implicated in both the adhesion and virulence of A. pleuropneumoniae, but their role in biofilm formation is still unclear.…”