The involvement of cell-to-cell signals in the development of a bacterial biofilm

Science

Volumn 280, Issue 5361, 1998, Pages 295-298

  Davies, David G ^a   Parsek, Matthew R ^b   Pearson, James P ^c   Iglewski, Barbara H ^c   Costerton, J W ^a   Greenberg, E P ^b  

^a MONTANA STATE UNIVERSITY   (United States)

^b UNIVERSITY OF IOWA   (United States)

^c UNIVERSITY OF ROCHESTER MEDICAL CENTER   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

ANTIBIOTIC RESISTANCE; ARTICLE; BACTERIAL GROWTH; BACTERIAL VIRULENCE; BACTERIUM MUTANT; BIOFILM; CELL COMMUNICATION; NONHUMAN; PRIORITY JOURNAL; PSEUDOMONAS AERUGINOSA;

4-BUTYROLACTONE; BACTERIAL ADHESION; BACTERIAL PROTEINS; BIOFILMS; HOMOSERINE; LIGASES; MUTATION; POLYSACCHARIDES, BACTERIAL; PSEUDOMONAS AERUGINOSA; SODIUM DODECYL SULFATE; TRANSCRIPTION FACTORS;

BACTERIA (MICROORGANISMS); PSEUDOMONAS; PSEUDOMONAS AERUGINOSA;

EID: 0032502811     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.280.5361.295     Document Type: Article

References (37)

1. M. Dworkin, in Microbial Cell-Cell interactions, M. Dworkin, Ed. (American Society for Microbiology, Washington, DC, 1991), pp. 179-216.
2. R. Losick and D. Kaiser, Sci. Am. 276, 68 (February 1997).
3. J. R. Lawrence, D. R. Korber, B. D. Hoyle, J. W. Costerton, D. E. Caldwell, J. Bacteriol. 173, 6558 (1991).
4. D. DeBeer, P. Stoodley, Z. Lewandowski, Biotech. Bioeng. 44, 636 (1994).
5. J. W. Costerton, Z. Lewandowski, D. E. Caldwell, D. R. Korber, H. M. Lappin-Scott, Annu. Rev. Microbiol. 49, 711 (1995).
6. A. Latifi et al., Mol. Microbiol. 17, 333 (1995).
7. L. Passador, J. M. Cook, M. J. Gambello, L. Rust, B. H. Iglewski, Science 260, 1127 (1993).
8. U. A. Ochsner, A. K. Koch, A. Fiechter, J. Reiser, J. Bacteriol. 176, 2044 (1994).
9. U. A. Ochsner and J. Reiser, Proc. Natl. Acad. Sci. U.S.A. 92, 6424 (1995).
10. M. J. Gambello and B. H. Iglewski, J. Bacteriol. 173, 3000 (1991).
11. J. P. Pearson et al., Proc. Natl. Acad. Sci. U.S.A. 91, 197 (1994).
12. E. C. Pesci, J. P. Pearson, P. C. Seed, B. H. Iglewski, J. Bacteriol. 179, 3127 (1997).
13. E. C. Pesci and B. H. Iglewski, Trends Microbiol. 5, 132 (1997).
14. A. Latifi, M. Foglino, K. Tanaka, P. Williams, A. Lazdunski. Mol. Microbiol. 21, 1137 (1996).
15. J. P. Pearson, L. Passador, B. H. Iglewski, E. P. Greenberg, Proc. Natl. Acad. Sci. U.S.A. 92, 1490 (1995).
16. M. K. Winson et al., ibid., p. 9427.
17. W. C. Fuqua, S. C. Winans, E. P. Greenberg, J. Bacteriol. 176, 269 (1994).
18. R. J. C. McLean, M. Whiteley, D. J. Stickler, W. C. Fuqua, FEMS Microbiol. Lett. 154, 259 (1997).
19. J. P. Pearson, E. C. Pesci, B. H. Iglewski, J. Bacteriol. 179, 5756 (1997).
20. J. M. Brint and D. E. Ohman, ibid. 177, 7155 (1995).
21. We constructed the plasmid pMRP9-1 by cloning an ∼0.8-kb Kpn I-Hind III fragment containing the Aequorea victoria GFP gene from pGFPmut2 [B. P. Cormack, R. H. Valdivia, S. Falkow, Gene 173, 33 (1996)] into Kpn I-Hind III-digested pUCP18. This places the GFP gene under control of the lac promoter and the T7gene10 ribosomal binding site. pMRP9-1 confers resistance to carbenicillin. The excitation maximum for this mutant GFP protein is 481 nm and the emission maximum is 507 nm.
22. T. A. Fassel, M. J. Schaller, C. C. Remsen, Microbiol. Res. Tech. 20, 87 (1992).
23. 12-HSL.
24. I. W. Sutherland, in Surface Carbohydrates of the Prokaryotic Cell, I. W. Sutherland, Ed. (Academic Press, London, 1977), pp. 27-96.
25. P. K. I. Kintner and J. P. V. Buren, J. Food Sci. 47, 756 (1982).
26. L. R. Evans and A. Linker, J. Bacteriol. 116, 915 (1973).
27. M. K. Dubois, A. Gilles, J. K. Hamilton, P. A. Rebers, F. Smith, Anal. Chem. 28, 350 (1956).
28. M. L. Brown, H. C. Aldrich, J. L. Gautier, Appl. Environ. Microbiol. 61, 187 (1995).
29. A. E. Khoury, K. Lam, B. D. Ellis, J. W. Costerton, Am. Soc. Artif. Intern. Organs J. 38, 174 (1992).
30. R. G. Dogget, G. M. Harrison, R. N. Stillwell, E. S. Wallis, J. Pediatr. 68, 215 (1966).
31. 4 (pH 7.0), 0.001% Hutner salts [G. Cohen-Bazire, W. R. Sistrom, R. Y. Stanier, J. Cell. Comp. Physiol. 49, 35 (1957)], 0.1% glucose, and 0.001% L-histidine. Carbenicillin (150 μg/ml) was used to maintain pMRP9-1. A continuous-culture bioreactor was configured as a once-flow-through system [ D. G. Davies, A. M. Chakrabarty, G. G. Geesey, Appl. Environ. Microbiol. 59, 1181 (1993)]. Medium was pumped from the reservoir through the bioreactor at a flow rate of 0.13 ml/min. The flow was laminar with a Reynolds number of 0.17 and a fluid resistance time of 0.43 min. The bioreactor was composed of polycarbonate with a glass coverslip affixed to its top. This glass coverslip served as a substratum. Pseudomonas aeruginosa was introduced into the stream through a septum located 1 cm upstream of the bioreactor.
32. 4 (pH 7.0), 0.001% Hutner salts [G. Cohen-Bazire, W. R. Sistrom, R. Y. Stanier, J. Cell. Comp. Physiol. 49, 35 (1957)], 0.1% glucose, and 0.001% L-histidine. Carbenicillin (150 μg/ml) was used to maintain pMRP9-1. A continuous-culture bioreactor was configured as a once-flow-through system [ D. G. Davies, A. M. Chakrabarty, G. G. Geesey, Appl. Environ. Microbiol. 59, 1181 (1993)]. Medium was pumped from the reservoir through the bioreactor at a flow rate of 0.13 ml/min. The flow was laminar with a Reynolds number of 0.17 and a fluid resistance time of 0.43 min. The bioreactor was composed of polycarbonate with a glass coverslip affixed to its top. This glass coverslip served as a substratum. Pseudomonas aeruginosa was introduced into the stream through a septum located 1 cm upstream of the bioreactor.
33. An Olympus BH2 microscope with a 60 X S PlanApo objective lens was used for epifluorescence and phase-contrast microscopy. Transmitted light images were collected with an Optronics charge-coupled device (Optronics Engineering, Goleta, CA) and the imaging program Image-Pro Plus 3.0 for Windows 95 (Media Cybernetics, Silver Spring, MD). Scanning confocal microscopy was performed with a Bio-Rad MRC600 confocal microscope (Hercules, CA). The excitation band was 488 nm with a 514-nm cutoff. The 50 X ULWD Olympus objective lens was used for scanning confocal microscopy. The imaging software was Comos 7.0 (Bio-Rad). The images were constructed with the SLICER imaging program (Fortner Research, Sterling, VA). The Kalman for each cross section was 5.
34. The nearest-neighbor analyses were on transmitted-light images of bacteria attached to the substratum after 10 days of continous culture. A minimum of 3000 cells in 10 fields were analyzed for each population. The distance of the nearest cell centroid to each study cell centroid was measured, and the average nearest-neighbor distance was calculated. This analysis was developed by G. Harkin (Montana State University, Bozeman, MT).
35. Biofllms for chemical analysis were grown in silicone tubing in a once-through continuous-flow system. Size 15 silicone tubing with a flow rate of 0.13 ml/min or size 15 tubing with a flow rate of 0.8 ml/min was used. After biofilms matured, the tubing was sliced lengthwise and biofilm cells were collected. The collected material was centrifuged at 13.000 rpm (Eppendorf Microfuge) for 10 mm and the sedimented material was analyzed for total carbohydrates, uronic acids, and protein as described (25, 27).
36. SDS (0.2% w/v) in 10 ml of EPRI medium was filtered through a 0.2-μm polycarbonate filter. After the flow of medium into the bioreactor was stopped, the SDS was injected through the septum into the flow chamber. The SDS remained in the bioreactor for 30 min, and then the flow of fresh medium was reinitiated.
37. 12-HSL, L. Loetterle for technical assistance, and T. de Kievit for helpful discussions. Supported by Office of Naval Research grant N00014-5-0190 and a grant from the Cystic Fibrosis Foundation (E.P.G.), NIH grant AI33713 (B.H.I.), and cooperative agreement ECO-8907039 with the Engineering Research Centers and Education Division of NSF (J.W.C.). M.R.P. is an NIH Postdoctoral Fellow (GM 18740), and J.P.P. is an NIH Predoctoral Trainee (5T32AI07362).