Determination of Active Marine Bacterioplankton: a Comparison of Universal 16S rRNA Probes, Autoradiography, and Nucleoid Staining - PubMed (original) (raw)
Determination of Active Marine Bacterioplankton: a Comparison of Universal 16S rRNA Probes, Autoradiography, and Nucleoid Staining
M Karner et al. Appl Environ Microbiol. 1997 Apr.
Abstract
We compared several currently discussed methods for the assessment of bacterial numbers and activity in marine waters, using samples from a variety of marine environments, from aged offshore seawater to rich harbor water. Samples were simultaneously tested for binding to a fluorescently labeled universal 16S rRNA probe; (sup3)H-labeled amino acid uptake via autoradiography; nucleoid-containing bacterial numbers by modified DAPI (4(prm1),6-diamidino-2-phenylindole) staining; staining with 5-cyano-2,3-ditolyl tetrazolium chloride (CTC), a compound supposed to indicate oxidative cell metabolism; and total bacterial counts (classical DAPI staining), taken as a reference. For the universal-probe counts, we used an image intensifying and processing system coupled to the epifluorescence microscope. All of the above-mentioned methods yielded lower cell counts than DAPI total counts. Universal-probe counts averaged about half of the corresponding DAPI count and were highly correlated to autoradiography counts (r(sup2) = 0.943; n = 7). Nucleoid-containing cell counts could be lower than DAPI counts by as much as 1 order of magnitude but sometimes matched autoradiography or probe counts. CTC counts were 2 orders of magnitude below DAPI counts. Universal 16S rRNA probe counts correlated well with autoradiography results, indicating a population with at least minimal metabolic activity. The greater variability of the nucleoid-containing cell counts calls for further investigation of the processes involved, and CTC counts were well below the range of the other methods tested.
Similar articles
- Increase in Fluorescence Intensity of 16S rRNA In Situ Hybridization in Natural Samples Treated with Chloramphenicol.
Ouverney CC, Fuhrman JA. Ouverney CC, et al. Appl Environ Microbiol. 1997 Jul;63(7):2735-40. doi: 10.1128/aem.63.7.2735-2740.1997. Appl Environ Microbiol. 1997. PMID: 16535648 Free PMC article. - Optimal staining and sample storage time for direct microscopic enumeration of total and active bacteria in soil with two fluorescent dyes.
Yu W, Dodds WK, Banks MK, Skalsky J, Strauss EA. Yu W, et al. Appl Environ Microbiol. 1995 Sep;61(9):3367-72. doi: 10.1128/aem.61.9.3367-3372.1995. Appl Environ Microbiol. 1995. PMID: 16535124 Free PMC article. - Use of a fluorescent redox probe for direct visualization of actively respiring bacteria.
Rodriguez GG, Phipps D, Ishiguro K, Ridgway HF. Rodriguez GG, et al. Appl Environ Microbiol. 1992 Jun;58(6):1801-8. doi: 10.1128/aem.58.6.1801-1808.1992. Appl Environ Microbiol. 1992. PMID: 1622256 Free PMC article. - Dual staining of natural bacterioplankton with 4',6-diamidino-2-phenylindole and fluorescent oligonucleotide probes targeting kingdom-level 16S rRNA sequences.
Hicks RE, Amann RI, Stahl DA. Hicks RE, et al. Appl Environ Microbiol. 1992 Jul;58(7):2158-63. doi: 10.1128/aem.58.7.2158-2163.1992. Appl Environ Microbiol. 1992. PMID: 1379029 Free PMC article. - Direct estimate of active bacteria: CTC use and limitations.
Créach V, Baudoux AC, Bertru G, Rouzic BL. Créach V, et al. J Microbiol Methods. 2003 Jan;52(1):19-28. doi: 10.1016/s0167-7012(02)00128-8. J Microbiol Methods. 2003. PMID: 12401223 Review.
Cited by
- Environmental Management of Legionella in Domestic Water Systems: Consolidated and Innovative Approaches for Disinfection Methods and Risk Assessment.
Sciuto EL, Laganà P, Filice S, Scalese S, Libertino S, Corso D, Faro G, Coniglio MA. Sciuto EL, et al. Microorganisms. 2021 Mar 11;9(3):577. doi: 10.3390/microorganisms9030577. Microorganisms. 2021. PMID: 33799845 Free PMC article. Review. - Visualization is crucial for understanding microbial processes in the ocean.
Sebastián M, Gasol JM. Sebastián M, et al. Philos Trans R Soc Lond B Biol Sci. 2019 Nov 25;374(1786):20190083. doi: 10.1098/rstb.2019.0083. Epub 2019 Oct 7. Philos Trans R Soc Lond B Biol Sci. 2019. PMID: 31587650 Free PMC article. - Determination of viable legionellae in engineered water systems: Do we find what we are looking for?
Kirschner AKT. Kirschner AKT. Water Res. 2016 Apr 15;93:276-288. doi: 10.1016/j.watres.2016.02.016. Epub 2016 Feb 12. Water Res. 2016. PMID: 26928563 Free PMC article. Review. - Dysbiotic bacteria translocate in progressive SIV infection.
Klase Z, Ortiz A, Deleage C, Mudd JC, Quiñones M, Schwartzman E, Klatt NR, Canary L, Estes JD, Brenchley JM. Klase Z, et al. Mucosal Immunol. 2015 Sep;8(5):1009-20. doi: 10.1038/mi.2014.128. Epub 2015 Jan 14. Mucosal Immunol. 2015. PMID: 25586559 Free PMC article. - Substrates specialization in lipid compounds and hydrocarbons of Marinobacter genus.
Bonin P, Vieira C, Grimaud R, Militon C, Cuny P, Lima O, Guasco S, Brussaard CP, Michotey V. Bonin P, et al. Environ Sci Pollut Res Int. 2015 Oct;22(20):15347-59. doi: 10.1007/s11356-014-4009-y. Epub 2015 Jan 6. Environ Sci Pollut Res Int. 2015. PMID: 25561256
References
- Appl Environ Microbiol. 1977 May;33(5):1225-8 - PubMed
- Biotechniques. 1992 Dec;13(6):928-34 - PubMed
- Appl Environ Microbiol. 1992 Jul;58(7):2158-63 - PubMed
- Appl Environ Microbiol. 1992 Jun;58(6):1801-8 - PubMed
- J Bacteriol. 1990 Feb;172(2):762-70 - PubMed
LinkOut - more resources
Full Text Sources