1887

Abstract

The aim of this study was to evaluate the survival responses of two strains of , both in natural and in defined media. For this purpose, freshwater and defined media containing different salinities (3·3–0·9 %) and nutrient concentrations (17–0·005 mg l) were assayed. The incubation temperatures were established at 4, 10 and 22 °C. The acridine orange staining technique was used for total cell enumeration and the number of viable cells was determined using two direct assays, nalidixic acid and tetrazolium salt reduction and plate spreading. Resuscitation assays of viable but non-culturable (VBNC) cells were conducted. According to the counting procedures employed, at least four different subpopulations were found: (i) active (positive response in both nalidixic acid and tetrazolium assays) culturable cells; (ii) active non-culturable cells; (iii) tetrazolium-salt-responsive non-culturable cells and (iv) non-active (responsive to none of the direct viable assays) non-culturable cells. Long-term survival was found at salinities and nutrient concentrations of seawater environments (3·3 % and 5 mg l or 1 g l), whereas the strains entered a VBNC state in freshwater and in brackish (0·9 or 1·6 % salinities) or high nutrient content (17 g l) defined medium. The recovery of VBNC cells was not achieved.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.25574-0
2003-02-01
2020-04-04
Loading full text...

Full text loading...

/deliver/fulltext/micro/149/2/mic149369.html?itemId=/content/journal/micro/10.1099/mic.0.25574-0&mimeType=html&fmt=ahah

References

  1. Angulo L, López J. E., Vicente J. A. 1994a; Epizootic outbreaks by a protozoan and a bacterium in reared turbot ( Scophthalmus maximus ). Bull Eur Ass Fish Pathol14:39–40
    [Google Scholar]
  2. Angulo L, López J. E, Vicente J. A., Saborido A. M. 1994b; Haemorrhagic areas in the mouth of farmed turbot, Scophthalmus maximus (L.). J Fish Dis17:163–169
    [Google Scholar]
  3. Balebona M. C, Zorrilla I, Moriñigo M. A., Borrego J. J. 1998; Survey of bacterial pathologies affecting farmed gilt-head sea bream ( Sparus aurata L.) in southwestern Spain. Aquaculture166:19–35
    [Google Scholar]
  4. Barcina I, Arana I, Santorum P, Iriberri J., Egea L. 1995; Direct viable count of Gram-positive and Gram-negative bacteria using ciprofloxacin as inhibitor of cellular division. J Microbiol Methods22:139–150
    [Google Scholar]
  5. Bianchi A., Giuliano L. 1996; Enumeration of viable bacteria in the marine pelagic environment. Appl Environ Microbiol62:174–177
    [Google Scholar]
  6. Colwell R. R, Brayton P, Herrington B, Tall B, Huq A., Levine M. M. 1996; Viable but nonculturable Vibrio cholerae 01 revert to a cultivable state in the human intestine. World J Microbiol Biotechnol12:28–31
    [Google Scholar]
  7. Comas J., Vives-Rego J. 1998; Enumeration, viability and heterogeneity in Staphylococcus aureus cultures by flow cytometry. J Microbiol Methods32:45–53
    [Google Scholar]
  8. Farto R, Montes M, Pérez M. J, Nieto T. P, Larsen J. L., Pedersen K. 1999; Characterization by numerical taxonomy and ribotyping of Vibrio splendidus biotype I and Vibrio scophthalmi strains associated with turbot cultures. J Appl Microbiol86:796–804
    [Google Scholar]
  9. Fegatella F., Cavicchioli R. 2000; Physiological responses to starvation in the marine oligotrophic ultramicrobacterium Sphingomonas sp. strain RB2256. Appl Environ Microbiol66:2037–2044
    [Google Scholar]
  10. Ghezzi J. I., Steck T. R. 1999; Induction of the viable but nonculturable condition in Xanthomonas campestris pv. campestris in liquid microcosms and sterile soil. FEMS Microbiol Ecol30:203–208
    [Google Scholar]
  11. Grimes D. J, Mills A. L., Nealson K. H. 2000; The importance of viable but nonculturable bacteria in biogeochemistry. In Nonculturable Microorganisms in the Environment pp209–227 Edited by Colwell R. R., Grimes D. J.. Washington, DC: ASM Press;
    [Google Scholar]
  12. Hoff K. A. 1989; Survival of Vibrio anguillarum and Vibrio salmonicida at different salinities. Appl Environ Microbiol55:1775–1786
    [Google Scholar]
  13. Huq A., Colwell R. R. 1995; A microbiological paradox: viable but nonculturable bacteria with special reference to Vibrio cholerae. J Food Prot59:96–101
    [Google Scholar]
  14. Huq A, Rivera I. N. G., Colwell R. R. 2000; Epidemiological significance of viable but nonculturable microorganisms. In Nonculturable Microorganisms in the Environment pp301–323 Edited by Colwell R. R., Grimes D. J.. Washington, DC: ASM Press;
    [Google Scholar]
  15. Jiang X., Chai T. 1996; Survival of Vibrio parahaemolyticus at low temperatures under starvation conditions and subsequent resuscitation of viable, nonculturable cells. Appl Environ Microbiol62:1300–1305
    [Google Scholar]
  16. Joux F, Lebaron P., Troussellier M. 1997; Succession of cellular states in a Salmonella typhimurium population during starvation in artificial seawater microcosms. FEMS Microbiol Ecol22:65–76
    [Google Scholar]
  17. Kaprelyants A. S, Mukamolova G. V, Davey H. M., Kell D. B. 1996; Quantitative analysis of the physiological heterogeneity within starved cultures of Micrococcus luteus by flow cytometry and cell sorting. Appl Environ Microbiol62:1311–1316
    [Google Scholar]
  18. Kim Y.-M., Kwon J.-Y. 1997; Effects of salinity and temperature on the survival of Vibrio vulnificus. J Korean Fish Soc30:367–376
    [Google Scholar]
  19. Kogure K, Simidu U., Taga N. 1979; A tentative direct microscopic method for counting living marine bacteria. Can J Microbiol25:415–420
    [Google Scholar]
  20. Lebaron P, Bernard L, Baudart J., Courties C. 1999; The ecological role of VBNC cells in the marine environment. In Proceedings of the 8th International Symposium on Microbial Ecology Edited by Bell C. R., Brylinsky M., Johnson-Green P.. Halifax, Canada: Atlantic Canada Society for Microbial Ecology;
    [Google Scholar]
  21. López J. E., Angulo L. 1995; Survival of Vibrio splendidus biotype I in seawater. Bull Eur Ass Fish Pathol15:70–72
    [Google Scholar]
  22. Marco-Noales E, Biosca E. G., Amaro C. 1999; Effects of salinity and temperature on long-term survival of the eel pathogen Vibrio vulnificus biotype 2 (serovar E). Appl Environ Microbiol65:1117–1126
    [Google Scholar]
  23. McDougald D, Rice S. A, Weichart D., Kjelleberg S. 1998; Nonculturability: adaptation or debilitation?. FEMS Microbiol Ecol25:1–9
    [Google Scholar]
  24. Miranda C. D., Rojas R. 1996; Vibriosis in the flounder Paralichthys adspersus (Steindachner, 1867) in captivity. Rev Biol Mar31:1–9
    [Google Scholar]
  25. Montes M, Pérez M. J., Nieto T. P. 1999; Numerical taxonomy of Gram-negative, facultative aerobic bacteria isolated from the skin of turbot ( Scophthalmus maximus ) and surrounding water. Syst Appl Microbiol22:604–618
    [Google Scholar]
  26. Morr N, Resnick M, Silbaq F, Bercovier H., Levy L. 1988; Reduction of tellurite and deesterification of fluorescein diacetate are not well correlated with the viability of mycobacteria. Ann Inst Pasteur139:279–288
    [Google Scholar]
  27. Nwoguh C. E, Harwood C. R., Barer M. R. 1995; Detection of induced galactosidase activity in individual non-culturable cells of pathogenic bacteria by quantitative cytological assay. Mol Microbiol17:545–554
    [Google Scholar]
  28. Nybroe O. 1995; Assessment of metabolic activity of single bacterial cells – new developments in microcolony and dehydrogenase assays. FEMS Microbiol Ecol17:77–84
    [Google Scholar]
  29. Oliver J. D. 1995; The viable but nonculturable state in the human pathogen Vibrio vulnificus. FEMS Microbiol Lett133:203–208
    [Google Scholar]
  30. Oliver J. D., Bockian R. 1995; In vivo resuscitation, and virulence towards mice, of viable but nonculturable cells of Vibrio vulnificus. Appl Environ Microbiol61:2620–2623
    [Google Scholar]
  31. Ortigosa M, Garay E., Pujalte M. J. 1994; Numerical taxonomy of Vibrionaceae isolated from oysters and seawater along an annual cycle. Syst Appl Microbiol17:216–225
    [Google Scholar]
  32. Pommepuy M, Butin M, Derrien A, Gourmelon M, Colwell R. R., Cormier M. 1996; Retention of enteropathogenicity by viable but nonculturable Escherichia coli exposed to seawater and sunlight. Appl Environ Microbiol62:4621–4626
    [Google Scholar]
  33. Rahman I, Shahamat M, Kirchman P. A, Russek-Cohen E., Colwell R. R. 1994; Methionine uptake and cytopathogenicity of viable but nonculturable Shigella dysenteriae. Appl Environ Microbiol60:3573–3578
    [Google Scholar]
  34. Reichalt J. L, Baumann P., Baumann L. 1976; Study of genetic relationships among marine species of the genera Beneckea and Photobacterium by means of in vitro DNA/DNA hybridization. Arch Microbiol110:101–120
    [Google Scholar]
  35. Roszak D. B., Colwell R. 1987; Survival strategies of bacteria in the natural environment. Microbiol Rev51:365–379
    [Google Scholar]
  36. Smith J. C. 1990; Potential-sensitive molecular probes in membranes of bioenergetic relevance. Biochim Biophys Acta1016:1–28
    [Google Scholar]
  37. Villarino A, Bouvet O. M. M, Regnault B, Martin-Delautre S., Grimont P. A. D. 2000; Exploring the frontier between life and death in Escherichia coli : evaluation of different viability markers in live and heat- or UV-killed cells. Res Microbiol151:755–768
    [Google Scholar]
  38. Weichart D., Kjelleberg S. 1996; Stress resistance and recovery potential of culturable and viable but nonculturable cells of Vibrio vulnificus. Microbiology142:845–853
    [Google Scholar]
  39. West P. A, Brayton P. R, Bryant T. N., Colwell R. R. 1986; Numerical taxonomy of vibrios isolated from aquatic environments. Int J Syst Bacteriol36:531–543
    [Google Scholar]
  40. Wolf P. W., Oliver J. D. 1992; Temperature effects on the viable but nonculturable state of Vibrio vulnificus. FEMS Microbiol Ecol101:33–39
    [Google Scholar]
  41. Zimmermann R, Iturriaga R., Becker-Birck J. 1978; Simultaneous determination of the total number of aquatic bacteria and the number thereof involved in respiration. Appl Environ Microbiol36:926–935
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.25574-0
Loading
/content/journal/micro/10.1099/mic.0.25574-0
Loading

Data & Media loading...

Most cited this month

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error