Confocal Laser Microscopy and Digital Image Analysis in Microbial Ecology

  • Douglas E. Caldwell
  • Darren R. Korber
  • John R. Lawrence
Part of the Advances in Microbial Ecology book series (AMIE, volume 12)


Microbial ecologists have extensively explored the potential applications of light microscopy for more than five decades (Henrici and Johnson, 1935; Perfil’ev and Gabe, 1969; Casida, 1969, 1972, 1975, 1976; Staley, 1971; Caldwell and Hirsch, 1973; Caldwell et al., 1973, 1975; Caldwell and Tiedje, 1975a,b; Labeda et al., 1976; Hirsch, 1977, 1980; Geesey et al., 1978; Marshall, 1986). Now traditional microscopy has given way to “microvisualization” (Friedhoff, 1991) greatly accelerating research. Microorganisms are no longer merely photographed; instead, they are digitally “imaged” using fluorescent molecular probes, confocal laser microscopy, and computer image analysis. The chemical and biological relationships between a microorganism and its microenvironment are seen directly, nondestructively, in situ, and in “real time” (Lawrence and Caldwell, 1990). Consequently, it is no longer necessary to disrupt microbial communities when studying the molecular or behavioral aspects of their ecology.


Salt Stress Gray Level Pseudomonas Fluorescens Surface Colonization Fluorescence Recovery After Photobleaching 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Absolom, D. R., Lamberti, F. V., Policova, Z., Zingg, W., van Oss, C. J., and Neumann, A. W., 1983, Surface thermodynamics of bacterial adhesion, Appl. Environ. Microbiol. 46:90–97.PubMedGoogle Scholar
  2. Agard, D. A., Hiraoka, Y., Shaw, P. J., and Sedat, J. W., 1989, Fluorescence microscopy in three dimensions, Methods Cell Biol. 30:353–377.PubMedGoogle Scholar
  3. Anwar, H. M., Dasgupta, Lam. K., and Costerton, J. W., 1989, Tobramycin resistance of mucoid Pseudomonas aeruginosa biofilm grown under iron limitation, J. Antimicrob. Chemother. 24:647–655.PubMedGoogle Scholar
  4. Berg, H. C., 1985, Physics of bacterial Chemotaxis, in: Sensory Perception and Transduction in Aneural Organisms (G. Colombetti, F. Linci, and P.-S. Song, eds.), Plenum Press, New York, pp. 19–30.Google Scholar
  5. Berg, H. C., and Brown, D. A., 1972, Chemotaxis in Escherichia coli analyzed by three-dimensional tracking, Nature 239:500–504.PubMedGoogle Scholar
  6. Bjornsen, P. K., 1986, Automatic determination of bacterioplankton biomass by means of image analysis, Appl. Environ. Microbiol. 51:1199–1204.PubMedGoogle Scholar
  7. Bott, T. L., and Brock, T. D., 1970, Growth and metabolism of periphytic bacteria: Methodology, Limnol. Oceanogr. 15:333–342.Google Scholar
  8. Boyde, A., 1990, Confocal optical microscopy, in: Modern Microscopies: Techniques and Applications (P. J. Duke and A. G. Michette, eds.), Plenum Press, New York, pp. 185–204.Google Scholar
  9. Bradbury, S., 1979, Microscopical image analysis: Problems and approaches, J. Microsc. 115:137–150.PubMedGoogle Scholar
  10. Brakenhoff, G. J., van der Voort, H. T. M., Baarslag, M. W., Mans, B., Oud, J. L., Zwart, R., and van Driel, R., 1988, Visualization and analysis techniques for three dimensional information acquired by confocal microscopy, Scanning Microsc. 2:1831–1838.PubMedGoogle Scholar
  11. Brock, T. D., 1971, Microbial growth rates in nature, Bacteriol. Rev. 35:39–58.PubMedGoogle Scholar
  12. Buskey, E. J., and Stoecker, D. K., 1989, Behavioral responses of the marine tintinnid Favella sp. to phytoplankton: Influence of chemical, mechanical and photic stimuli, J. Exp. Mar. Biol. Ecol. 132:1–16.Google Scholar
  13. Busscher, H. J., Bellon-Fontaine, M.-N., Mozes, N., van der Mei, H. C., Sjollema, J., and Rouxhet, P. G., 1990a, Deposition of Leuconostoc mesenteroides and Streptococcus thermophilus to solid substrata in a parallel plate flow cell, Biofouling 2:55–63.Google Scholar
  14. Busscher, H. J., Sjollema, J., and van der Mei, H. C., 1990b, Relative importance of surface free energy as a measure of hydrophobicity in bacterial adhesion to solid surfaces, in: Microbial Cell Surface Hydrophobicty (R. J. Doyle and M. Rosenberg, eds.), ASM, Washington, D.C., pp. 335–359.Google Scholar
  15. Caldwell, D. E., 1985, New developments in computer-enhanced microscopy, J. Microbiol. Methods 4:117–125.Google Scholar
  16. Caldwell, D. E., 1987, Microbial colonization of solid-liquid interfaces, Ann. N.Y. Acad. Sci. 506:274–280.PubMedGoogle Scholar
  17. Caldwell, D. E., and Caldwell, S. J., 1978, A Zoogloea sp. associated with blooms of Anabaena flosaquae, Can J. Microbiol. 24:922–931.PubMedGoogle Scholar
  18. Caldwell, D. E., and Germida, J. J., 1985, Evaluation of difference imagery for visualizing and quantitating microbial growth, Can J. Microbiol. 31:35–44.Google Scholar
  19. Caldwell, D. E., and Hirsh, P., 1973, Growth of microorganisms in two-dimensional steady-state diffusion gradients, Can. J. Microbiol. 19:53–58.PubMedGoogle Scholar
  20. Caldwell, D. E., and Lawrence, J. R., 1986, Growth kinetics of Pseudomonas fluorescens microcolonies within the hydrodynamic boundary layers of surface microenvironments, Microb. Ecol. 12:299–312.Google Scholar
  21. Caldwell, D. E., and Lawrence, J. R., 1988, Study of attached cells in continuous-flow slide culture, in: CRC Handbook of Laboratory Model Systems for Microbial Ecosystems (J. W. T. Wimpenny, ed.), CRC Press, Boca Raton, pp. 117–138.Google Scholar
  22. Caldwell, D. E., and Lawrence, J. R., 1989, Microbial growth and behavior within surface microenvironments, in: Proceedings of ISME-5 (T. Hattori, Y. Ishida, Y. Maruyama, R. Y. Morita, and A. Uchida, eds.), JSS Press, Tokyo, pp. 140–145.Google Scholar
  23. Caldwell, D. E., and Tiedje, J. M., 1975a, The structure of anaerobic bacterial communities in the hypolimnion of several Michigan lakes, Can. J. Microbiol. 21:377–385.PubMedGoogle Scholar
  24. Caldwell, D. E., and Tiedje, J. M., 1975b, A morphological study of anaerobic bacteria from the hypolimnion of two Michigan lakes, Can. J. Microbiol. 21:362–376.PubMedGoogle Scholar
  25. Caldwell, D. E., Lai, S. H., and Tiedje, J. M., 1973, A two-dimensional steady-state diffusion gradient for ecological studies, Bull. Ecol. Res. Commun. 17:151–158.Google Scholar
  26. Caldwell, D. E., Caldwell, S. J., and Tiedje, J. M., 1975, An ecological study of the sulfur-oxidizing bacteria from the littoral zone of a Michigan lake and a sulfur spring in Florida, Plant Soil 43:101–114.Google Scholar
  27. Caldwell, D. E., Korber, D. R., and Lawrence, J. R., 1992, Imaging of bacterial cells by fluorescence exclusion using scanning confocal laser microscopy, J. Microbiol. Methods 15:249–261.Google Scholar
  28. Carlsson, K., and Lileborg, A., 1989, A confocal laser microscope scanner for digital recording of optical serial sections, J. Microsc. 153:171–180.PubMedGoogle Scholar
  29. Carlsson, K., Wallen, P., and Brodin, L., 1989, Three-dimensional imaging of neurons by confocal fluorescence microscopy, J. Microsc. 155:15–26.PubMedGoogle Scholar
  30. Casida, L. E., 1969, Observation of microorganisms in soil and other natural habitats, Appl. Microbiol. 18:1065–1071.PubMedGoogle Scholar
  31. Casida, L. E., 1972, Interval scanning photomicrography of microbial cell populations, Appl. Microbiol. 23:190–192.PubMedGoogle Scholar
  32. Casida, L. E., 1975, Infrared color photomicrography of soil microorganisms, Can. J. Microbiol. 21:1892–1893.PubMedGoogle Scholar
  33. Casida, L. E., 1976, Continuously variable amplitude contrast microscopy for the detection and study of microorganisms in soil, Appl. Environ. Microbiol. 31:605–608.PubMedGoogle Scholar
  34. Cheng, K.-J., Ingram, J. M., and Costerton, J. W., 1970, Alkaline phosphatase localization and spheroplast formation of Pseudomonas aeruginosa, Can J. Microbiol. 16:1319–1324.PubMedGoogle Scholar
  35. Cooksey, B., and Cooksey, K. E., 1988, Chemical signal-response in diatoms of the genus Amphora, J. Cell Sci. 91:523–529.Google Scholar
  36. Costello, P. J., and Monk, P. R., 1985, Image analysis method for the rapid counting of Saccharomyces cerevisiae cells, Appl. Environ. Microbiol. 49:863–866.PubMedGoogle Scholar
  37. Costerton, J. W., 1988, Structure and plasticity at various organization levels in the bacterial cell, Can. J. Microbiol. 34:513–521.PubMedGoogle Scholar
  38. Costerton, J. W., Ingram, J. M., and Cheng, K.-J., 1974, Structure and function of the cell envelope of gram-negative bacteria, Bacteriol. Rev. 38:87–110.PubMedGoogle Scholar
  39. Costerton, J. W., Cheng, K.J., Geesey, G. G., Ladd, T., Nickel, J. C., Dasgupta, M., and Marrie, T. J., 1987, Bacterial biofilms in nature and disease, Annu. Rev. Microbiol. 41:435–464.PubMedGoogle Scholar
  40. Davenport, D., 1973, Studies of the behavior of microorganisms by computerized television, in: Behavior of Micro-organisms (A. Perez-Miravete, ed.), Proceedings of the 10th International Congress, Plenum Press, New York.Google Scholar
  41. DeYoung, H. G., 1988, Microscopy and image analysis, Bio/Technology 6:78–79.Google Scholar
  42. Donovan, R. M., Goldstein, E., Kim, Y., Lippert, W., Kailath, E., Aoki, K. T., Cheung, A. T. W., Miller, M. E., and Chang, D. P., 1987, A computer-assisted image-analysis system for analyzing polymorphonuclear leukocyte chemotaxis in patients with Diabetes mellitus, J. Infect. Dis. 155:737–741.PubMedGoogle Scholar
  43. Drake, B., Prater, C. B., Weisenhorn, A. L., Gould, S. A. C., Albrecht, T. R., Quate, C. F., Cannell, D. S., Hansma, H. G., and Hansma, P. K., 1989, Imaging crystals, polymers, and processes in water with the atomic force microscope, Science 241:1586–1589.Google Scholar
  44. Edgar, L. A., 1979, Diatom locomotion: Computer-assisted analysis of cine film, Br. Phycol. J. 14:83–101.Google Scholar
  45. Eighmy, T. T., Maratea, D., and Bishop, P. L., 1983, Electron microscopic examination of wastewater biofilm formation and structural components, Appl. Environ. Microbiol. 45:1921–1931.PubMedGoogle Scholar
  46. Ellwood, D. C., Keevil, C. W., Marsh, P. D., Brown, C. M., and Wardell, J. N., 1982, Surface-associated growth, Philos. Trans. R. Soc. London Ser. B 297:517–532.Google Scholar
  47. Emmett, A., 1991, In search of the miracle hologram: Spatial image researchers strive to achieve stereoscopic reality, Comput. Graphics World 14:44–52.Google Scholar
  48. Escher, A. R., and Characklis, W. G., 1988, Microbial colonization of a smooth substratum: A kinetic analysis using image analysis, Water Sci. Technol. 20:45–51.Google Scholar
  49. Fernandes, M. A., Jackman, P. J., Clark, S. A., and Gunard, S. R., 1988, Detection and quantification of microorganisms in a heterogenous foodstuff by image analysis, Comput. Appl. Biosci. 4:291–295.PubMedGoogle Scholar
  50. Friedhoff, R. M., 1991, Microvisualization, Comput. Graphics World 14:38–44.Google Scholar
  51. Fry, J. C., 1988, Determination of biomass, in: Methods in Aquatic Bacteriology (B. Austin, ed.), Wiley, New York, pp. 27–72.Google Scholar
  52. Fry, J. C., and Davies, A. R., 1985, An assessment of methods for measuring volumes of planktonic bacteria, with particular reference to television image analysis, J. Appl. Bacteriol. 58:105–112.Google Scholar
  53. Gaju, N., Guerrero, R., and Pedros-Alio, C., 1989, Measurement of cell volume of phototrophic bacteria in pure cultures and natural samples: Phase contrast, epifluorescence and particle sizing, FEMS Microbiol. Ecol. 62:295–302.Google Scholar
  54. Geesey, G. C., Mutch, R., and Costerton, J. W., 1978, Sessile bacteria: An important component of the microbial population in small mountain streams, Limnol. Oceanogr. 23:1214–1223.Google Scholar
  55. Getliff, J. M., and Fry, J. C., 1989, Using the solitaire plus image analyser for direct estimates of bacterial volume, Binary 1:93–100.Google Scholar
  56. Gonzalez, R. C., and Wintz, P., 1977, Digital Image Processing. Addison-Wesley, Reading, Mass.Google Scholar
  57. Gratton, E., and van deVen, M. J., 1990, Laser sources for confocal microscopy, in Handbook of Biological Confocal Microscopy (J. B. Pawley, ed.), Plenum Press, New York, pp. 53–67.Google Scholar
  58. Gualtieri, P., Colombetti, G., and Lend, F. 1985, Automatic analysis of the motion of microorganisms, J. Microsc. 139:57–62.Google Scholar
  59. Gualtieri, P., Francesco, G., Passarelli, V., and Barsanti, L., 1988, Microorganism track reconstruction: An image processing approach, Comput. Biol. Med. 18:57–63.PubMedGoogle Scholar
  60. Hansma, P. K., Drake, B., Mari, O., Gould, S. A. C., and Prater, C. B., 1989a, The scanning ionconductance microscope, Science 243:641–643.PubMedGoogle Scholar
  61. Hansma, P. K., Elings, V. B., Mari, O., and Bracker, C. E., 1989b, Scanning tunneling microscopy and atomic force microscopy: Application to biology and technology, Science 241:209–216.Google Scholar
  62. Harris, R., 1991, Confocal microscopy, Microsc. Soc. Can. Bull. 19:33–35.Google Scholar
  63. Haugland, R. P., 1989, Handbook of Fluorescent Probes and Research Chemicals, Molecular Probes Inc., Eugene, Oreg.Google Scholar
  64. Henrici, A. T., and Johnson, D. E., 1935, Studies of freshwater bacteria. II. Stalked bacteria, a new order of Schizomycetes, J. Bacteriol. 30:61–93.PubMedGoogle Scholar
  65. Hirsch, P., 1974, Budding bacteria, Annu. Rev. Microbiol. 28:391–433.PubMedGoogle Scholar
  66. Hirsch, P., 1977, Distribution and pure culture studies of morphologically distinct solar lake microorganisms, in: Hypersaline Brines and Evaporinc Environments (A. Nissenbaum, ed.), Elsevier, Amsterdam, pp. 41–60.Google Scholar
  67. Hirsch, P., 1980, Some thoughts on and examples of microbial interactions in the natural environment, in: Aquatic Microbial Ecology (R. R. Colwell and A. J. Foster, eds.), University of Maryland, pp. 36–54.Google Scholar
  68. Inoué, S., 1986, Video Microscopy, Plenum Press, New York, pp. 263–307.Google Scholar
  69. Inoué, S., 1990, Foundations of confocal scanned imaging in light microscopy, in: Handbook of Confocal Laser Microscopy (J. B. Pawley, ed.), Plenum Press, New York, pp. 1–14.Google Scholar
  70. Jenkinson, D. S., Powlson, D. S., and Wedderburn, R. M. W., 1976, The effects of biocidal treatments on metabolism in soil. III. The relationship between soil biovolume, measured by optical microscopy and the flush of decomposition cause by fumigation, Soil Biol. Biochem. 8:189–202.Google Scholar
  71. Kesterson, J., and Richardson, M., 1991, Confocal microscope capability with desktop affordability, Advanced Imaging 6:23–25.Google Scholar
  72. Kinner, N. E., Balkwill, D. L., and Bishop, P. L., 1983, Light and electron microscopic studies of microorganisms growing in rotating biological contactor biofilms, Appl. Environ. Microbiol. 45:1659–1669.PubMedGoogle Scholar
  73. Kjelleberg, S., Humphrey, B., and Marshall, K. C., 1982, The effect of interfaces on small starved marine bacteria, Appl. Environ. Microbiol. 43:1166–1172.PubMedGoogle Scholar
  74. Kohen, E., and Hirschberg, J. G., 1989, Cell Structure and Function by Microspectrofluorometry, Academic Press, New York.Google Scholar
  75. Korber, D. R., Lawrence, J. R., Sutton, B., and Caldwell, D. E., 1989a, Effect of laminar flow velocity on the kinetics of surface recolonization by mot+ and mot- Pseudomonas fluorescens, Microb. Ecol. 18:1–19.Google Scholar
  76. Korber, D. R., Lawrence, J. R., Cooksey, K. E., Cooksey, B., and Caldwell, D. E., 1989b, Computer image analysis of diatom Chemotaxis, Binary 2:335–350.Google Scholar
  77. Korber, D. R., Lawrence, J. R., Zhang, L., and Caldwell, D. E., 1990, Effect of gravity on bacterial deposition and orientation in laminar flow environments, Biofouling 2:335–350.Google Scholar
  78. Krambeck, C., Krambeck, H. J., Schroder, D., and Newell, S. Y., 1990, Sizing bacterioplankton: A juxtaposition of bias due to shrinkage, halos, subjectivity in image interpretation and asymmetric distributions, Binary 2:5–14.Google Scholar
  79. Kuhn, D. A., and Starr, M. P., 1970, Effects of microscope illumination on bacterial development, Arch. Mikrobiol. 74:292–300.Google Scholar
  80. Labeda, D. P., Liu, K., and Casida, L. E., 1976, Colonization of soil by Arthrobacter and Pseudomonas under varying conditions of water and nutrient availability as studied by plate counts and transmission electron microscopy, Appl. Environ. Microbiol. 31:551–561.PubMedGoogle Scholar
  81. Lappin-Scott, H. M., and Costerton, J. W., 1990, Bacterial biofilms and surface fouling, Biofouling 1:323–342.Google Scholar
  82. Lawrence, J. R., and Caldwell, D. E., 1987, Behavior of bacterial stream populations within the hydrodynamic boundary layers of surface microenvironments, Microb. Ecol. 14:15–27.Google Scholar
  83. Lawrence, J. R., and Caldwell, D. E., 1990, Scanning confocal laser microscopy of biofilms, Can. Lab. 2:12.Google Scholar
  84. Lawrence, J. R., Delaquis, P. J., Korber, D. R., and Caldwell, D. E., 1987, Behavior of Pseudomonas fluorescens within the hydrodynamic boundary layers of surface microenvironments, Microb. Ecol. 14:1–14.Google Scholar
  85. Lawrence, J. R., Korber, D. R., and Caldwell, D. E., 1989a, Computer-enhanced darkfield microscopy for the quantitative analysis of bacterial growth and behavior on surfaces, J. Microbiol. Methods 10:123–138.Google Scholar
  86. Lawrence, J. R., Malone, J. A., Korber, D. R., and Caldwell, D. E., 1989b, Computer image enhancement to increase depth of field in phase contrast microscopy, Binary 1:181–185.Google Scholar
  87. Lawrence, J. R., Delaquis, P. J., Zanyk, B. N., Korber, D. R., and Caldwell, D. E., 1990, Computer-enhanced microscopy study of Pseudomonas fragi biofilm development, Abstracts of the ASM Conference on Multicellular Behavior of Bacteria in Nature, Industry, and the Laboratory, Woods Hole Marine Biological Laboratory.Google Scholar
  88. Lawrence, J. R., Korber, D. R., Hoyle, B. D., Costerton, J. W., and Caldwell, D. E., 1991, Optical sectioning of microbial biofilms, J. Bacteriol. 173:6558–6567.PubMedGoogle Scholar
  89. Linfoot, E. H., and Wolfe, E., 1953, Diffraction images in systems with an annular aperture, Proc. Phys. Soc. B 66:145–149.Google Scholar
  90. Luby-Phelps, K., Lanni, F., and Taylor, D. L., 1988, The submicroscopic properties of cytoplasm as a determinant of cellular function, Annu. Rev. Biophys. Chem. 17:369–396.Google Scholar
  91. McLean, R. J. C., Lawrence, J. R., Korber, D. R., and Caldwell, D. E., 1991, Proteus mirabilis biofilm protection against struvite crystal dissolution and its implications in struvite urolithiasis, J. Urol. 146:1130–1142.Google Scholar
  92. Malone, J. A., 1988, Colonization of surface microenvironments by Rhizobium spp., M.Sc. thesis, University of Sask.Google Scholar
  93. Marshall, K. C., 1986, Microscopic methods for the study of bacterial behavior at inert surfaces, J. Microbiol. Methods 4:217–227.Google Scholar
  94. Marshall, K. C., 1988, Adhesion and growth of bacteria at surfaces in oligotrophic habitats, Can J. Microbiol. 34:503–506.Google Scholar
  95. Marshall, K. C., and Cruickshank, R. H., 1973, Cell surface hydrophobicity and the orientation of certain bacteria at interfaces, Arch. Mikrobiol. 91:29–40.PubMedGoogle Scholar
  96. Marshall, K. C., Stout, R., and Mitchell, R., 1971, Mechanisms of the initial events in the sorption of marine bacteria to solid surfaces, J. Gen. Microbiol. 68:337–348.Google Scholar
  97. Marshall, K. C., Cruikshank, R. H., and Bushby, H. V. A., 1975, The orientation of certain root-nodule bacteria at interfaces, including legume root-hair surfaces, J. Gen. Microbiol. 91:198–200.PubMedGoogle Scholar
  98. Martin, Y., Williams, C. C., and Wickramasinghe, H. K., 1988, Tip techniques for microcharacterization of materials, Scanning Microsc. 2:3–8.Google Scholar
  99. Meadows, P. S., 1971, The attachment of bacteria to solid surfaces, Arch. Mikrobiol. 75:374–381.PubMedGoogle Scholar
  100. Meijer, B.C., Kootstra, G. J., and Wilkinson, M. H. F., 1990, A theoretical and practical investigation into the characterization of bacterial species by image analysis, Binary 2:21–31.Google Scholar
  101. Pawley, J. B. (ed.), 1990, Handbook of Biological Confocal Microscopy, Plenum Press, New York.Google Scholar
  102. Perfil’ev, B. V., and Gabe, D. R., 1969, Capillary Methods of Investigating Microorganisms (J. M. Shewan, trans.), University of Toronto Press, Toronto.Google Scholar
  103. Peters, A. C., 1990, Using image analysis to map bacterial growth on solid media. Binary 2:73–75.Google Scholar
  104. Peters, A.C., Wimpenny, J. W. T., Thomas, L. V., and Griffiths, J., 1991, Mapping bacterial growth on gradient plates using image analysis, Binary 3:147–154.Google Scholar
  105. Pettipher, G. L., and Rodrigues, U. M., 1982, Semi-automated counting of bacteria and somatic cells in milk using epifluorescence microscopy and television image analysis, J. Appl. Bacteriol. 53:323–329.PubMedGoogle Scholar
  106. Power, K., and Marshall, K. C., 1988, Cellular growth and reproduction of marine bacteria on surface-bound substrate, Biofouling 1:163–174.Google Scholar
  107. Read, N. D., Knight, H., and Trewas, A. J., 1992, Fluorescence ratio imaging and confocal microscopy in filamentous fungi, Binary 4:50–52.Google Scholar
  108. Robinson, R. W., Akin, D. E., Nordstedt, R. A., Thomas, M. V., and Aldrich, H. C., 1984, Light and electron microscopic examinations of methane-producing biofilms from anaerobic fixed-bed reactors, Appl. Environ. Microbiol. 48:127–136.PubMedGoogle Scholar
  109. Russ, J. C., 1990, Computer-Assisted Microscopy: The Measurement and Analysis of Images, Plenum Press, New York.Google Scholar
  110. Shaw, P. J., and Rawlins, P. J., 1991, The point-spread function of a confocal microscope: Its measurement and use in deconvolution of 3-D data, J. Micros. 163:151–165.Google Scholar
  111. Shotton, D. M., 1989, Confocal scanning optical microscopy and its applications for biological specimens, J. Cell Sci. 94:175–206.Google Scholar
  112. Shotton, D., and White, N., 1989, Confocal scanning microscopy: Three dimensional biological imaging, Trends Biochem. Sci. 14:435–438.PubMedGoogle Scholar
  113. Sieracki, M. E., and Webb, K. L., 1986, A color video image analysis system for studying pico-and nanoplanktonic bacteria, EOS Trans. Am. Geophys. Union 66:1298.Google Scholar
  114. Sieracki, M. E., Johnson, P. W., and Sieburth, J. M., 1985, Detection, enumeration, and sizing of planktonic bacteria by image-analzyed epifluorescence microscopy, Appl. Environ. Microbiol. 49:799–810.PubMedGoogle Scholar
  115. Sieracki, M. E., Reichenback, S. E., and Webb, K. L., 1989, Evaluation of automated threshold selection methods for accurately sizing microscopic fluorescent cells by image analysis, Appl. Environ. Microbiol. 55:2762–2772.PubMedGoogle Scholar
  116. Silverman, M., and Simon, M., 1974, Flagellar rotation and the mechanism of bacterial motility, Nature 249:73–74.PubMedGoogle Scholar
  117. Singh, A., Yu, F.-P., and McFeters, G. A., 1990, Rapid detection of chlorine-induced bacterial injury by the direct viable count method using image analysis, Appl. Environ. Microbiol. 56:389–394.PubMedGoogle Scholar
  118. Sjollema, J., Busscher, H. J., and Weerkamp, A. H., 1988, Deposition of oral streptococci and polystyrene lattices onto glass in a parallel plate flow cell, Biofouling 1:101–112.Google Scholar
  119. Sjollema, J., Busscher, H. J., and Weerkamp, A. H., 1989, Real-time enumeration of adhering microorganisms in a parallel plate flow cell using automated image analysis, J. Microbiol. Methods 9:73–78.Google Scholar
  120. Sjollema, J., van der Mei, H. M., and Busscher, H. J., 1990a, Direct observations of cooperative effects in oral streptococcal adhesion to glass by analysis of the spatial arrangement of adhering bacteria, FEMS Microbiol. Ecol. Lett. 69:263–270.Google Scholar
  121. Sjollema, J., van der Mei, H. M., and Busscher, H. J., 1990b, The influence of collector and bacterial cell surface properties on the deposition of oral streptococci in a parallel plate flow cell. J. Adhesion Sci. Technol. 4:765–777.Google Scholar
  122. Soderstrom, B., 1977, Vital staining of fungi in pure cultures and in soil with fluorescein diacetate, Soil Biol. Biochem. 9:59–63.Google Scholar
  123. Soderstrom, B. E., 1979, Some problems in assessing the fluorescein diacetate-active fungal biomass in the soil, Soil Biol. Biochem. 11:147–148.Google Scholar
  124. Staley, J. T., 1971, Growth rates of algae determined in situ using an immersed microscope, J. Phytol. 7:13–17.Google Scholar
  125. Tsien, R. Y., 1989, Fluorescent indicators of ion concentrations, Methods Cell Biol. 30:127–156.PubMedGoogle Scholar
  126. Tsien, R. Y., and Waggoner, A., 1990, Fluorophores for confocal microscopy: Photophysics and photochemistry, in: Handbook of Confocal Microscopy (J. B. Pawley, ed.), Plenum Press, New York, pp. 169–178.Google Scholar
  127. Vaitzus, Z., and Doetsch, R. N., 1969, Motility tracks: Technique for quantitative study of bacterial movement, Appl. Environ. Microbiol. 17:584–588.Google Scholar
  128. van Loosdrecht, M. C. W., Lyklema, J., Norde, W., and Zehnder, A. J. B., 1990, Influence of interfaces on microbial activity, Microbiol. Rev. 54:75–87.PubMedGoogle Scholar
  129. van Veen, J., and Paul, E. A., 1979, Conversion of biovolume measurements of soil organisms, grown under various moisture tensions, to biomass and their nutrient content, Appl. Environ. Microbiol. 37:686–692.PubMedGoogle Scholar
  130. Verran, J., and Rocliffe, M. D., 1986, Feasibility of using automatic image analysis for measuring dental plaque in situ, J. Dent., 14:11–13.PubMedGoogle Scholar
  131. Wells, K. S., Sandison, D. R., Strickler, J., and Webb, W. W., 1990, Imaging with laser scanning confocal microscopy, in: Handbook of Biological Confocal Microscopy (J. B. Pawley, ed.), Plenum Press, New York, pp. 27–39.Google Scholar
  132. White, J. G., Amos, W. B., and Fordham, M., 1987, An evaluation of confocal versus conventional imaging of biological structure by fluorescence light microscopy, J. Cell Biol. 105:41–48.PubMedGoogle Scholar
  133. Wimpenny, J. W. T., Waters, P., and Peters, A. C., 1988, Gel-plate methods in microbiology, in: Handbook of Laboratory Model Systems for Microbial Ecosystems, Vol. 1 (J. W. T. Wimpenny, ed.), CRC Press, Boca Raton, pp. 229–251.Google Scholar
  134. Wolfaardt, G. M., Lawrence, J. R., Hendry, M. J., Roberts, R. D., and Caldwell, D. E., 1992, The use of model diffusion gradients for the isolation of degradative microbial consortia, Abstracts of the Annual Meeting of the Canadian Society of Microbiologists.Google Scholar
  135. Wynn-Williams, D. D., 1988a, Television image analysis of microbial communities in Antarctic fellfields, Polarforschung 58:239–249.Google Scholar
  136. Wynn-Williams, D. D., 1988b, Microbial colonization processes in Antarctic fellfield soils—An experimental overview, in: Proceedings of the NIPR Symposium on Polar Biology, National Institute of Polar Research, Tokyo, Vol. 3, pp. 164–178.Google Scholar
  137. Zanyk, B. N., Korber, D. R., Lawrence, J. R., and Caldwell, D. E., 1991, 4-D visualization of biofilm development by Pseudomonas fragi, Binary 3:24–29.Google Scholar

Copyright information

© Plenum Press, New York 1992

Authors and Affiliations

  • Douglas E. Caldwell
    • 1
  • Darren R. Korber
    • 1
  • John R. Lawrence
    • 2
  1. 1.Department of Applied Microbiology and Food ScienceUniversity of SaskatchewanSaskatoonCanada
  2. 2.National Hydrology Research InstituteEnvironment CanadaSaskatoonCanada

Personalised recommendations