Extracellular DNA in Single- and Multiple-Species Unsaturated Biofilms

ABSTRACT The extracellular polymeric substances (EPS) of bacterial biofilms form a hydrated barrier between cells and their external environment. Better characterization of EPS could be useful in understanding biofilm physiology. The EPS are chemically complex, changing with both bacterial strain and culture conditions. Previously, we reported that Pseudomonas aeruginosa unsaturated biofilm EPS contains large amounts of extracellular DNA (eDNA) (R. E. Steinberger, A. R. Allen, H. G. Hansma, and P. A. Holden, Microb. Ecol. 43:416-423, 2002). Here, we investigated the compositional similarity of eDNA to cellular DNA, the relative quantity of eDNA, and the terminal restriction fragment length polymorphism (TRFLP) community profile of eDNA in multiple-species biofilms. By randomly amplified polymorphic DNA analysis, cellular DNA and eDNA appear identical for P. aeruginosa biofilms. Significantly more eDNA was produced in P. aeruginosa and Pseudomonas putida biofilms than in Rhodococcus erythropolis or Variovorax paradoxus biofilms. While the amount of eDNA in dual-species biofilms was of the same order of magnitude as that of of single-species biofilms, the amounts were not predictable from single-strain measurements. By the Shannon diversity index and principle components analysis of TRFLP profiles generated from 16S rRNA genes, eDNA of four-species biofilms differed significantly from either cellular or total DNA of the same biofilm. However, total DNA- and cellular DNA-based TRFLP analyses of this biofilm community yielded identical results. We conclude that extracellular DNA production in unsaturated biofilms is species dependent and that the phylogenetic information contained in this DNA pool is quantifiable and distinct from either total or cellular DNA.

[1]  L. Forney,et al.  Molecular microbial ecology: land of the one-eyed king. , 2004, Current opinion in microbiology.

[2]  A. Agnelli,et al.  Distribution of microbial communities in a forest soil profile investigated by microbial biomass, soil respiration and DGGE of total and extracellular DNA , 2004 .

[3]  Ursula Obst,et al.  Investigation of natural biofilms formed during the production of drinking water from surface water embankment filtration. , 2004, Water research.

[4]  J. Lawrence,et al.  Microbial exopolymers provide a mechanism for bioaccumulation of contaminants , 1994, Microbial Ecology.

[5]  P. Holden,et al.  Macromolecular composition of unsaturated Pseudomonas aeruginosa biofilms with time and carbon source , 2004 .

[6]  Zhongtang Yu,et al.  Stability of the bacterial community in a pulp mill effluent treatment system during normal operation and a system shutdown. , 2003, Water research.

[7]  Sebastian Maassen,et al.  Microbial diversity and functional characterization of sediments from reservoirs of different trophic state. , 2003, FEMS microbiology ecology.

[8]  W. Hop,et al.  DNase in stable cystic fibrosis infants: a pilot study. , 2003, Journal of cystic fibrosis : official journal of the European Cystic Fibrosis Society.

[9]  T. Vogel,et al.  Extraction of DNA from soil , 2003 .

[10]  Gordon Webster,et al.  Assessment of bacterial community structure in the deep sub-seafloor biosphere by 16S rDNA-based techniques: a cautionary tale. , 2003, Journal of microbiological methods.

[11]  S. Kjelleberg,et al.  Cell Death in Pseudomonas aeruginosa Biofilm Development , 2003, Journal of bacteriology.

[12]  J. Schimel,et al.  Comparison of subsurface and surface soil bacterial communities in california grassland as assessed by terminal restriction fragment length polymorphisms of PCR-amplified 16S rRNA genes , 2003, Microbial Ecology.

[13]  Stefan Bertilsson,et al.  Heterotrophic Bacterial Growth Efficiency and Community Structure at Different Natural Organic Carbon Concentrations , 2003, Applied and Environmental Microbiology.

[14]  S. Edwards,et al.  Soil fungal community structure in a temperate upland grassland soil. , 2003, FEMS microbiology ecology.

[15]  Tim Tolker-Nielsen,et al.  Gene transfer occurs with enhanced efficiency in biofilms and induces enhanced stabilisation of the biofilm structure. , 2003, Current opinion in biotechnology.

[16]  Dick B Janssen,et al.  Efficient recovery of environmental DNA for expression cloning by indirect extraction methods. , 2003, FEMS microbiology ecology.

[17]  Z. Kawabata,et al.  Release of Extracellular Transformable Plasmid DNA from Escherichia coli Cocultivated with Algae , 2003, Applied and Environmental Microbiology.

[18]  D. Sponza.  Investigation of extracellular polymer substances (EPS) and physicochemical properties of different activated sludge flocs under steady-state conditions , 2003 .

[19]  Noah Fierer,et al.  Variations in microbial community composition through two soil depth profiles , 2003 .

[20]  S. Haruta,et al.  Microbial community changes during organic solid waste treatment analyzed by double gradient-denaturing gradient gel electrophoresis and fluorescence in situ hybridization , 2002, Applied Microbiology and Biotechnology.

[21]  R. Danovaro,et al.  Quantification, base composition, and fate of extracellular DNA in marine sediments , 2002 .

[22]  J. Niemeyer,et al.  Determination of free DNA in soils , 2002 .

[23]  M. Ballmann,et al.  Hypertonic saline and recombinant human DNase: a randomised cross-over pilot study in patients with cystic fibrosis. , 2002, Journal of cystic fibrosis : official journal of the European Cystic Fibrosis Society.

[24]  J. Mattick,et al.  Extracellular DNA required for bacterial biofilm formation. , 2002, Science.

[25]  P. Holden,et al.  Comparison of free-living and particle-associated bacterial communities in a coastal Lagoon , 2003, Microbial Ecology.

[26]  H. Hansma,et al.  Elongation Correlates with Nutrient Deprivation in Pseudomonas aeruginosa Unsaturated Biofilms , 2002, Microbial Ecology.

[27]  R. Kolter,et al.  DNA as a Nutrient: Novel Role for Bacterial Competence Gene Homologs , 2001, Journal of bacteriology.

[28]  M. Firestone,et al.  Water Content Mediated Microaerophilic Toluene Biodegradation in Arid Vadose Zone Materials , 2001, Microbial Ecology.

[29]  M. Franchi,et al.  Effect of molecular characteristics of DNA on its adsorption and binding on homoionic montmorillonite and kaolinite , 2001, Biology and Fertility of Soils.

[30]  K. Lewis,et al.  Programmed Death in Bacteria , 2000, Microbiology and Molecular Biology Reviews.

[31]  S. Lory,et al.  Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic pathogen , 2000, Nature.

[32]  C. Kuske,et al.  Assessment of Microbial Diversity in Four Southwestern United States Soils by 16S rRNA Gene Terminal Restriction Fragment Analysis , 2000, Applied and Environmental Microbiology.

[33]  H. Hansma,et al.  Physical Morphology and Surface Properties of Unsaturated Pseudomonas putida Biofilms , 2000, Journal of bacteriology.

[34]  X. Guan,et al.  Relationship between dsDNA, chloroform labile C and ergosterol in soils of different organic matter contents and pH. , 2000 .

[35]  W. Liesack,et al.  Use of the T-RFLP technique to assess spatial and temporal changes in the bacterial community structure within an agricultural soil planted with transgenic and non-transgenic potato plants. , 2000, FEMS microbiology ecology.

[36]  E. Madsen.  Nucleic-acid characterization of the identity and activity of subsurface microorganisms , 2000 .

[37]  K. Timmis,et al.  An evaluation of terminal-restriction fragment length polymorphism (T-RFLP) analysis for the study of microbial community structure and dynamics. , 2000, Environmental microbiology.

[38]  S. Lory,et al.  Complete genome sequence of Pseudomonas aeruginosa PAO 1 , an opportunistic pathogen , 2000 .

[39]  V. Ramisse,et al.  Quantification of Bias Related to the Extraction of DNA Directly from Soils , 1999, Applied and Environmental Microbiology.

[40]  Jost Wingender,et al.  What are Bacterial Extracellular Polymeric Substances , 1999 .

[41]  Jost Wingender,et al.  Microbial extracellular polymeric substances : characterization, structure, and function , 1999 .

[42]  P. Stewart,et al.  A review of experimental measurements of effective diffusive permeabilities and effective diffusion coefficients in biofilms , 1998, Biotechnology and bioengineering.

[43]  G. Stotzky,et al.  Binding of DNA on humic acids: Effect on transformation of Bacillus subtilis and resistance to DNase , 1998 .

[44]  M. Khanna,et al.  Amplification of DNA bound on clay minerals , 1998 .

[45]  P. Simonet,et al.  The fate of recombinant plant DNA in soil , 1998 .

[46]  Hans H. Cheng,et al.  Characterization of microbial diversity by determining terminal restriction fragment length polymorphisms of genes encoding 16S rRNA , 1997, Applied and environmental microbiology.

[47]  Donald A. Jackson.  COMPOSITIONAL DATA IN COMMUNITY ECOLOGY: THE PARADIGM OR PERIL OF PROPORTIONS? , 1997 .

[48]  P. Stewart,et al.  Theoretical aspects of antibiotic diffusion into microbial biofilms , 1996, Antimicrobial agents and chemotherapy.

[49]  M E Campbell,et al.  Random amplified polymorphic DNA typing of Pseudomonas aeruginosa isolates recovered from patients with cystic fibrosis , 1996, Journal of clinical microbiology.

[50]  J. Tiedje,et al.  DNA recovery from soils of diverse composition , 1996, Applied and environmental microbiology.

[51]  J. Lawrence,et al.  Bioaccumulation of the Herbicide Diclofop in Extracellular Polymers and Its Utilization by a Biofilm Community during Starvation , 1995, Applied and environmental microbiology.

[52]  A. Chakrabarty,et al.  Isolation and assay of Pseudomonas aeruginosa alginate. , 1994, Methods in enzymology.

[53]  M. Palmer.  PUTTING THINGS IN EVEN BETTER ORDER: THE ADVANTAGES OF CANONICAL CORRESPONDENCE ANALYSIS' , 1993 .

[54]  J. D. van Elsas,et al.  Rapid DNA extraction protocol from soil for polymerase chain reaction‐mediated amplification , 1993 .

[55]  R. Gibson,et al.  Efficacy and safety of short-term administration of aerosolized recombinant human deoxyribonuclease in patients with cystic fibrosis. , 1993, The American review of respiratory disease.

[56]  W. Liesack,et al.  Occurrence of novel groups of the domain Bacteria as revealed by analysis of genetic material isolated from an Australian terrestrial environment , 1992, Journal of bacteriology.

[57]  M. Firestone,et al.  Relationship between Desiccation and Exopolysaccharide Production in a Soil Pseudomonas sp , 1992, Applied and environmental microbiology.

[58]  V. Torsvik,et al.  High diversity in DNA of soil bacteria , 1990, Applied and environmental microbiology.

[59]  J. Tiedje,et al.  DNA Probe Method for the Detection of Specific Microorganisms in the Soil Bacterial Community , 1988, Applied and environmental microbiology.

[60]  G. Sayler,et al.  The extraction and purification of microbial DNA from sediments , 1987 .

[61]  J. Paul,et al.  Dynamics of extracellular DNA in the marine environment , 1987, Applied and environmental microbiology.

[62]  M. G. Lorenz,et al.  Protection of Sediment-Adsorbed Transforming DNA Against Enzymatic Inactivation , 1983, Applied and environmental microbiology.

[63]  T. Hara,et al.  A study on the mechanism of DNA excretion from P. aeruginosa KYU-1.Effect of mitomycin C on extracellular DNA production.:Effect of Mitomycin C on Extracellular DNA Production , 1981 .

[64]  M. G. Lorenz,et al.  Interaction of marine sediment with DNA and DNA availability to nucleases , 1981 .

[65]  M. P. Greaves,et al.  The degradation of nucleic acids and montmorillonite-nucleic-acid complexes by soil microorganisms , 1970 .

[66]  S. Colowick,et al.  Methods in Enzymology , Vol , 1966 .