Skip to main content
SpringerLink
Log in

Practical Methods for Determining Phage Growth Parameters

  • Protocol
Book cover Bacteriophages

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 501))

Abstract

Bacteriophage growth may be differentiated into sequential steps: (i) phage collision with an adsorption-susceptible bacterium, (ii) virion attachment, (iii) virion nucleic acid uptake, (iv) an eclipse period during which infections synthesize phage proteins and nucleic acid, (v) a “post-eclipse” period during which virions mature, (vi) a virion release step, and (vii) a diffusion-delimited period of virion extracellular search for bacteria to adsorb (1). The latent period begins at the point of virion attachment (ii) and/or nucleic acid uptake (iii) and ends with infection termination, spanning both the eclipse (iv) and the post-eclipse maturation (v) periods. For lytic phages, latent-period termination occurs at lysis, i.e., at the point of phage-progeny release (vi). A second compound step is phage adsorption, which, depending upon one’s perspective, can begin with virion release (vi), may include the virion extracellular search (vii), certainly involves virion collision with (i) and then attachment to (ii) a bacterium, and ends either with irreversible virion attachment to bacteria (ii) or with phage nucleic acid uptake into cytoplasm (iii). Thus, the phage life cycle, particularly for virulent phages, consists of an adsorption period, virion attachment/nucleic acid uptake, a latent period, and virion release ((2), p. 13, citing d’Herelle). The duration of these steps together define the phage generation time and help to define rates of phage population growth. Also controlling rates of phage population growth is the number of phage progeny produced per infection: the phage burst size. In this chapter we present protocols for determining phage growth parameters, particularly phage rate of adsorption, latent period, eclipse period, and burst size.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 189.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 239.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 249.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Abedon, S.T. (2006) Phage ecology, in The Bacteriophages (Calendar, R. and Abedon, S T eds.), Oxford University Press, Oxford, pp. 37–46.

    Google Scholar 

  2. Adams, M.H. (1959). Bacteriophages. Interscience, New York.

    Google Scholar 

  3. d’H’érelle, F. and Smith, G.H. (1926). The Bacteriophage and Its Behavior [English translation]. The Williams & Wilkins Co., Baltimore.

    Google Scholar 

  4. Abedon, S.T. (1992) Lysis of lysis inhibited bacteriophage T4 infected cells. J. Bacteriol. 174, 8073–8080.

    CAS  PubMed  Google Scholar 

  5. d’Herelle, F. (1922). The Bacteriophage: Its Role in Immunity. Williams and Wilkins Co./Waverly Press, Baltimore.

    Google Scholar 

  6. Eisenstark, A. (1967) Bacteriophage techniques. Meth. Virol. 1, 449–524.

    Google Scholar 

  7. Cairns, J., Stent, G. and Watson, J.D. (1966). Phage and the Origins of Molecular Biology. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.

    Google Scholar 

  8. Ellis, E.L. and Delbrück, M. (1939) The growth of bacteriophage. J. Gen. Physiol. 22, 365–384.

    Article  CAS  PubMed  Google Scholar 

  9. Carlson, K. (2005) Working with bacteriophages: common techniques and methodological approaches, in Bacteriophages: Biology and Application (Kutter, E. and Sulakvelidze, A eds.), CRC Press, Boca Raton, Florida, pp. 437–494.

    Google Scholar 

  10. Carlson, K. and Miller, E.S. (1994) Working with T4, in Molecular Biology of Bacteriophage T4 (Karam, J.D. ed.), ASM Press, Washington, DC, pp. 421–426.

    Google Scholar 

  11. Stent, G.S. (1963). Molecular Biology of Bacterial Viruses. WH Freeman and Co., San Francisco, CA.

    Google Scholar 

  12. Heller, K.J. and Bryniok, D. (1984) O antigen-dependent mutant of bacteriophage T5. J. Virol. 49, 20–25.

    CAS  PubMed  Google Scholar 

  13. Prehm, P., Jann, B., Schmidt, G. and Stirm, S. (1976) On a bacteriophage T3 and T4 receptor region within the cell wall lipopolysaccharide of Escherichia coli B. J. Mol. Biol. 101, 277–281.

    Article  CAS  PubMed  Google Scholar 

  14. Brumfitt, W. (1960) Studies on the mechanism of adsorption and penetration by bacteriophage. J. Pathol. Bacteriol. 79, 1–9.

    Article  CAS  PubMed  Google Scholar 

  15. Fildes, P. and Kay, D. (1957) Tryptophan as a bacteriophage adsorption factor. Brit. J. Exp. Path. 38, 563–572.

    CAS  PubMed  Google Scholar 

  16. Dandekar, A.M. and Modi, V.V. (1978) Interaction between Rhizobium japonicum phage M-1 and its receptor. Can. J. Microbiol. 24, 685–688.

    Article  CAS  PubMed  Google Scholar 

  17. Landry, E.F. and Zsigray, R.M. (1980) Effects of calcium on the lytic cycle of Bacillus subtilis phage 41c. J. Gen. Virol. 51, 125–135.

    Article  CAS  PubMed  Google Scholar 

  18. Garen, A. and Puck, T.T. (1951) The first two steps of the invasion of host cells by bacterial viruses. II. J. Exp. Med. 94, 177–189.

    Article  CAS  PubMed  Google Scholar 

  19. Rountree, P.M. (1951) The role of certain electrolytes in the adsorption of staphylococcal bacteriophages. J. Gen. Microbiol. 5, 673–680.

    CAS  PubMed  Google Scholar 

  20. Daniels, L.L. and Wais, A.C. (1990) Ecophysiology of bacteriophage S5100 infecting Halobacterium cutirubrum. Appl. Environ. Microbiol. 56, 3605–3608.

    CAS  PubMed  Google Scholar 

  21. Conley, M.P. and Wood, W.B. (1975) Bacteriophage T4 whiskers: A rudimentary environment-sensing device. Proc. Natl. Acad. Sci. USA 72, 3701–3705.

    Article  CAS  PubMed  Google Scholar 

  22. Abedon, S.T. The Ecology of Bacteriophage T4. 1990. University of Arizona. Ph.D. dissertation

    Google Scholar 

  23. Takumi, K., Takeoka, A., Kinouchi, T. and Kawata, T. (1985) Solubilization and partial properties of receptor substance for bacteriophage alpha 2 induced from Clostridium botulinum type A 190L. Microbiol. Immunol. 29, 1185–1195.

    CAS  PubMed  Google Scholar 

  24. Watanabe, K., Takesue, S. and Ishibashi, K. (1977) Reversibility of the adsorption of bacteriophage PL-1 to the cell walls isolated from Lactobacillus casei. J. Gen. Virol. 34, 189–194.

    Article  CAS  PubMed  Google Scholar 

  25. Rossi, P. and Aragno, M. (1999) A fast method for assessing rapid inactivation and adsorption kinetics of bacteriophages using batch agitation experiments and colloidal clay particles. Can. J. Microbiol. 45, 9–17.

    Article  CAS  Google Scholar 

  26. Lipson, S.M. and Stotzky, G. (1985) Specificity of virus adsorption to clay minerals. Can. J. Microbiol. 31, 50–53.

    Article  CAS  PubMed  Google Scholar 

  27. Fildes, P. and Kay, D. (1963) The conditions which govern the adsorption of a tryptophan-dependent bacteriophage to kaolin and bacteria. J. Gen. Microbiol. 30, 183–191.

    CAS  PubMed  Google Scholar 

  28. Carlson, K. and Miller, E.S. (1994) Enumerating phage, in Molecular Biology of Bacteriophage T4 (Karam, J.D. ed.), ASM Press, Washington, DC, pp. 427–429.

    Google Scholar 

  29. Hadas, H., Einav, M., Fishov, I. and Zaritsky, A. (1997) Bacteriophage T4 development depends on the physiology of its host Escherichia coli. Microbiology 143, 179–185.

    Article  CAS  PubMed  Google Scholar 

  30. Delbrück, M. (1942) Bacterial viruses (bacteriophages). Adv. Enzymol. 2, 1–32.

    Google Scholar 

  31. Delbrück, M. (1940) Adsorption of bacteriophage under various physiological conditions of the host. J. Gen. Physiol. 23, 631–642.

    Article  PubMed  Google Scholar 

  32. Rosner, A. and Gutstein, R. (1981) Adsorption of actinophage Pal 6 to developing mycelium of Streptomyces albus. Can. J. Microbiol. 27, 254–257.

    Article  CAS  PubMed  Google Scholar 

  33. Preissner, W.C., Maier, S., Volker, H. and Hirsch, P. (1988) Isolation and partial characterization of a bacteriophage active on Hyphomicrobium sp. WI-926. Can. J. Microbiol. 34, 101–106.

    Article  CAS  PubMed  Google Scholar 

  34. Ohshima, Y., Schumacher-Perdreau, F., Peters, G. and Pulverer, G. (1988) The role of capsule as a barrier to bacteriophage adsorption in an encapsulated Staphylococcus simulans strain. Med. Microbiol. Immunol. 177, 229–233.

    CAS  PubMed  Google Scholar 

  35. Wilkinson, B.J. and Holmes, K.M. (1979) Staphylococcus aureus cell surface: capsule as a barrier to bacteriophage adsorption. Infect. Immun. 23, 549–552.

    CAS  PubMed  Google Scholar 

  36. Schwartz, M. (1976) The adsorption of coliphage lambda to its host: Effect of variation in the surface density of the receptor and in phage-receptor affinity. J. Mol. Biol. 103, 521–536.

    Article  CAS  PubMed  Google Scholar 

  37. Chai, T.-J. (1983) Characteristics of Escherichia coli grown in bay waters as compared with rich medium. Appl. Environ. Microbiol. 45, 1316–1323.

    CAS  PubMed  Google Scholar 

  38. Koch, A.L. (1960) Encounter efficiency of coliphage-bacterium interaction. Biochim. Biophys. Acta 39, 311–318.

    Article  Google Scholar 

  39. Anderson, T.F. (1949) On the mechanism of adsorption of bacteriophages on host cells, in The Nature of the Bacterial Surface (Miles, A.A. and Pirie, N W eds.), Blackwell Scientific Publications, Oxford, pp. 76–95.

    Google Scholar 

  40. Delbrück, M. (1940) The growth of bacteriophage and lysis of the host. J. Gen. Physiol. 23, 643–660.

    Article  PubMed  Google Scholar 

  41. Abedon, S.T. (1994) Lysis and the interaction between free phages and infected cells, in The Molecular Biology of Bacteriophage T4 (Karam, J.D. ed.), ASM Press, Washington, DC, pp. 397–405.

    Google Scholar 

  42. Carlson, K. and Miller, E.S. (1994) General procedures, in Molecular Biology of Bacteriophage T4 (Karam, J.D. ed.), ASM Press, Washington, DC, pp. 427–437.

    Google Scholar 

  43. S’échaud, J. and Kellenberger, E. (1956) Lyse precoce, provoqu’ée par le chloroforme, chez les bact’éries infect’és par du bact’ériophage. Ann. Inst. Pasteur 90, 102–106.

    Google Scholar 

  44. Doermann, A.H. (1992) The eclipse in the bacteriophage life cycle, in Phage and the Origins of Molecular Biology (expanded edition) (Cairns, J., Stent, G S and Watson, J D eds.), Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp. 79–87.

    Google Scholar 

  45. Cumming, G., Fidler, F. and Vaux, D.L. (2007) Error bars in experimental biology. J. Cell Biol. 177, 7–11.

    Article  CAS  PubMed  Google Scholar 

  46. Jarvis, A.W. (1993) Analysis of phage resistance mechanisms encoded by lactococcal plasmid pAJ2074. Can. J. Microbiol. 39, 252–258.

    Article  CAS  Google Scholar 

  47. Gupta, B.M., Chandra, K. and Clifford, I. (1963) Human intestinal bacteriophage. V. Effect of yeast adenylic acid on virus adsorption and plaque formation by coli-dysentery phage CVX-5 on purine-deficient Escherichia coli. Ann. Biochem. Exp. Med. 23, 395–400.

    CAS  PubMed  Google Scholar 

  48. Sanders, M.E. and Klaenhammer, T.R. (1983) Characterization of phage-sensitive mutants from a phage-insensitive strain of Streptococcus lactis: Evidence for a plasmid determinant that prevents phage adsorption. Appl. Environ. Microbiol. 46, 1125–1133.

    CAS  PubMed  Google Scholar 

  49. Chakrabarti, B.K., Si, K. and Chattopadhyay, D. (1996) Characterization of Vibrio cholerae EIT or typing phage D10. J. Gen. Virol. 77, 2881–2884.

    Article  CAS  PubMed  Google Scholar 

  50. Gründling, A., Manson, M.D. and Young, R. (2001) Holins kill without warning. Proc. Natl. Acad. Sci. USA 98, 9348–9352.

    Article  PubMed  Google Scholar 

  51. Carlson, K. (1994) Single-step growth, in Molecular Biology of Bacteriophage T4 (Karam, J.D. ed.), ASM Press, Washington, pp. 434–437.

    Google Scholar 

  52. Dulbecco, R. (1949) On the reliability of the Poisson distribution as a distribution of the number of phage particles infecting individual bacteria in a population. Genetics 34, 122–125.

    Google Scholar 

  53. Gots, J.S. (1959) Chemical interference with phage growth, in Bacteriophages (Hershey, A.D., Hotchkiss, R D, Pappenheimer, A M, Jr. and Racker, E eds.), Interscience Publishers, Inc., New York, pp. 265–287.

    Google Scholar 

  54. Smart, K.M. and Steinberg, B.M. (1977) Simultaneous presence of antiviral activity and its degrader in Bacillus extracts. Can. J. Microbiol. 23, 726–732.

    Article  CAS  PubMed  Google Scholar 

  55. Doermann, A.H. and Dissosway, C. (1948) Intracellular growth and genetics of bacteriophage. Year Book Carnegie Inst. Wash. 48, 170–176.

    Google Scholar 

  56. Kerr, B., West, J. and Bohannan, B.J.M. (2008) Bacteriophage: models for exploring basic principles of ecology, in Bacteriophage Ecology (Abedon, S.T. ed.), Cambridge University Press, Cambridge, UK, pp. 31–63.

    Google Scholar 

  57. Heineman, R.H. and Bull, J.J. (2007) Testing optimality with experimental evolution: lysis time in a bacteriophage. Evolution 61, 1695–1709.

    Article  PubMed  Google Scholar 

  58. Doermann, A.H. (1952) The intracellular growth of bacteriophages. I. Liberation of intracellular bacteriophage T4 by premature lysis with another phage or with cyanide. J. Gen. Physiol. 35, 645–656.

    Article  CAS  PubMed  Google Scholar 

  59. Underwood, N. and Doermann, A.H. (1947) A photoelectric nephelometer. Rev. Scient. Instr. 18, 665–672.

    Article  CAS  Google Scholar 

  60. Doermann, A.H. (1948) Lysis and lysis inhibition with Escherichia coli bacteriophage. J. Bacteriol. 55, 257–275.

    Google Scholar 

  61. Abedon, S.T. (2000) The murky origin of Snow White and her T-even dwarfs. Genetics 155, 481–486.

    CAS  PubMed  Google Scholar 

  62. Young, R. (1992) Bacteriophage Lysis: Mechanisms and regulation. Microbiol. Rev. 56, 430–481.

    CAS  PubMed  Google Scholar 

  63. Young, R. and Wang, I.-N. (2006) Phage Lysis, in The Bacteriophages (Calendar, R. and Abedon, S T eds.), Oxford University Press, Oxford, pp. 104–125.

    Google Scholar 

  64. Abedon, S.T. (1990) Selection for lysis inhibition in bacteriophage. J. Theor. Biol. 146, 501–511.

    Article  CAS  PubMed  Google Scholar 

  65. Abedon, S.T. (1999) Bacteriophage T4 resistance to lysis-inhibition collapse. Genet. Res. 74, 1–11.

    Article  CAS  PubMed  Google Scholar 

  66. Abedon, S.T., Hyman, P. and Thomas, C. (2003) Experimental examination of bacteriophage latent-period evolution as a response to bacterial availability. Appl. Environ. Microbiol. 69, 7499–7506.

    Article  CAS  PubMed  Google Scholar 

  67. Paddison, P., Abedon, S.T., Dressman, H.K., Gailbreath, K., Tracy, J., Mosser, E., Neitzel, J., Guttman, B. and Kutter, E. (1998) Lysis inhibition and fine-structure genetics in bacteriophage T4. Genetics 148, 1539–1550.

    CAS  PubMed  Google Scholar 

  68. Sarimo, S.S., Hartiala, M. and Aaltonen, L. (1976) Preparation and partial characterization of a Lactobacillus lactis bacteriophage. Arch. Microbiol. 107, 193–197.

    Article  Google Scholar 

  69. Brown, A. (1956) A study of lysis in bacteriophage-infected Escherichia coli. J. Bacteriol. 71, 482–490.

    CAS  PubMed  Google Scholar 

  70. Ackermann, H.-W., Roy, R., Martin, M., Murthy, M.R.V. and Smirnoff, W.A. (1978) Partial characterization of a cubic Bacillus phage. Can. J. Microbiol. 24, 986–993.

    Article  CAS  PubMed  Google Scholar 

  71. Wollman, E. and Wollman, E. (1937) Les “phases” des bact’ériophages (facteurs lysog‘énes). Compt. Rend. Soc. Biol. 124, 931–934.

    Google Scholar 

  72. Rees, P.J. and Fry, B.A. (1981) The morphology of staphylococcal bacteriophage K and DNA metabolism in infected Staphylococcus aureus. J. Gen. Virol. 53, 293–307.

    Article  CAS  PubMed  Google Scholar 

  73. Hooper, I., Woods, W.H. and Egan, B. (1981) Coliphage 186 Replication is delayed when the host cell is UV irradiated before infection. J. Virol. 40, 341–349.

    CAS  PubMed  Google Scholar 

  74. Zachary, A. (1976) Physiology and ecology of bacteriophages of the marine bacterium Beneckea natriegens: salinity. Appl. Environ. Microbiol. 31, 415–422.

    CAS  PubMed  Google Scholar 

  75. de Siqueira, R.S., Dodd, C.E.R. and Rees, C.E.D. (2006) Evaluation of the natural virucidal activity of teas for use in the phage amplification assay. Int. J. Food Microbiol. 111, 259–262.

    Article  PubMed  Google Scholar 

  76. Josslin, R. (1971) Physiological studies on the t gene defect in T4-infected Escherichia coli. Virology 44, 101–107.

    Article  CAS  PubMed  Google Scholar 

  77. Josslin, R. (1970) The lysis mechanism of phage T4: Mutants affecting lysis. Virology 40, 719–726.

    Article  CAS  PubMed  Google Scholar 

  78. Reddy, A.B. and Gopinathan, K.P. (1987) Characterization of mycobacteriophage I8 and its unrelatedness to mycobacteriophages I1, I3 and I5. J. Gen. Virol. 68, 949–956.

    Article  CAS  PubMed  Google Scholar 

  79. Stevens, R.H., Hammond, B.F. and Lai, C.H. (1982) Characterization of an inducible bacteriophage from a leukotoxic strain of Actinobacillus actinomycetemcomitans. Infect. Immun. 35, 343–349.

    CAS  PubMed  Google Scholar 

  80. Benzer, S. and Jacob, F. (1953) ’étude du d’éveloppement du bact’ériophage au moyen d’irradiations par la lumi‘ére ultra-violette. Ann. Inst. Pasteur 84, 186–204.

    CAS  Google Scholar 

  81. Schlesinger, M. (1932) Absorption [sic?] of bacteriophages to homologous bacteria [translation], in Bacterial Viruses, Little, Brown and Co., Boston, pp. 26–36.

    Google Scholar 

  82. Gerba, C.P. (1984) Applied and theoretical aspects of virus adsorption to surfaces. Adv. Appl. Microbiol. 30, 133–168.

    Article  CAS  PubMed  Google Scholar 

  83. Kasman, L.M., Kasman, A., Westwater, C., Dolan, J., Schmidt, M.G. and Norris, J.S. (2002) Overcoming the phage replication threshold: a mathematical model with implications for phage therapy. J. Virol. 76, 5557–5564.

    Article  CAS  PubMed  Google Scholar 

  84. Abedon, S.T. (2008) Phage population growth: constraints, games, adaptations, in Bacteriophage Ecology (Abedon, S.T. ed.), Cambridge University Press, Cambridge, UK, pp. 64–93.

    Chapter  Google Scholar 

  85. Goodridge, L.D. (2008) Phages, bacteria, and food, in Bacteriophage Ecology (Abedon, S.T. ed.), Cambridge University Press, Cambridge, UK, pp. 302–331.

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. MedCentral College of Nursing, Mansfield, OH, USA

    Paul Hyman

  2. Department of Microbiology, The Ohio State University, Mansfield, OH, USA

    Stephen T. Abedon

Authors
  1. Paul Hyman
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Stephen T. Abedon
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. University of Leicester, Leicester, UK

    Martha R.J. Clokie

  2. Public Health Agency of Canada, Guelph, Ontario, Canada

    Andrew M. Kropinski

Rights and permissions

Reprints and permissions

Copyright information

© 2009 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Hyman, P., Abedon, S.T. (2009). Practical Methods for Determining Phage Growth Parameters. In: Clokie, M.R., Kropinski, A.M. (eds) Bacteriophages. Methods in Molecular Biology™, vol 501. Humana Press. https://doi.org/10.1007/978-1-60327-164-6_18

Download citation

  • DOI: https://doi.org/10.1007/978-1-60327-164-6_18

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-682-5

  • Online ISBN: 978-1-60327-164-6

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation