Physiological assessment of bacteria using fluorochromes

McFeters, Gordon A.; Yu, Feipeng Philip; Pyle, Barry H.; Stewart, Philip S.

doi:10.1016/0167-7012(94)00027-5

Cited by 141 publications

(114 citation statements)

References 61 publications

Supporting

Mentioning

110

Contrasting

Unclassified

Order By: Relevance

“…The use of such processing software offers the feasibility of identifying as much as possible metabolic modifications, which are associated to cell shape modifications, allowing the evaluation of a density profile (metabolism) and cell morphology. In addition, the density profile derived from the Acridine Orange (AO) stained cells, also provides information on the metabolic activity (11,12). The dimension index was calculated by delineating the cells.…”

Section: Discussionmentioning

confidence: 99%

“…Stained samples were observed by a phase-contrast Nikon Optiphot microscope (filter set 14; excitation BP 546/12; emission LP 590) coupled with a Hamamatsu 5985 camera, processed by NIH Scion Image 1.61 on a Macintosh 6100/ 66 computer to estimate shape parameters. Bacteria that appear green after AO staining have been considered inactive, while red or orange cells are thought to be active, according to the classical interpretation (12).…”

Section: Acridine Orange Stainingmentioning

confidence: 99%

See 1 more Smart Citation

Use of multiparameter analysis for Vibrio alginolyticus viable but nonculturable state determination

Albertini¹,

Accorsi²,

Teodori³

et al. 2006

Cytometry Pt A

View full text Add to dashboard Cite

Background:Vibrio alginolyticus is known to enter into a viable but nonculturable (VBNC) state in response to environmental conditions unfavorable to the growth. Cells in VBNC condition pose a public health threat because they are potentially pathogenic.Methods:We constructed a pathway for the identification of the most significant variables and the characterization of those variables able to discriminate the groups under investigation. Different parameters measured by the image processing software were chosen as the most representative of V. alginolyticus cell morphology (length index for dimension) and metabolic activity (density profile indexes). To detect relationships between the groups of treatment performed, we carried out a principal components analysis (PCA).Results:The PCA analysis indicated that increasing coccoid shape transformation was related to both metabolic and dimension variations, delineating a well defined graph profile. Indeed, we discovered that specific morphological variations occur when cells in the culturable state pass into VBNC condition, namely comma‐shaped culturable bacteria are converted into coccoid‐shaped VBNC cells. The results were also supported by scanning electron microscopy analysis.Conclusions:This technique allows the analysis of a large number of vibrio samples in a short period of time. The obtained multiparameter information may complement genetic/molecular analyses facilitating, in an automatic fashion, further studies to evaluate the potential risk of this pathogen in the environment. It may also be a useful tool for large‐scale cell biology studies and high content screening. © 2006 International Society for Analytical Cytology

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Acridine Orange Stainingmentioning

confidence: 99%

Use of multiparameter analysis for Vibrio alginolyticus viable but nonculturable state determination

Albertini¹,

Accorsi²,

Teodori³

et al. 2006

Cytometry Pt A

View full text Add to dashboard Cite

show abstract

“…Comas and Vives-Rego, 1997), and characterizing the starvation-survival response of selected bacterial species, usually those with pathogenic relevance (Thorsen et al, 1992;Lebaron and Joux, 1994). The development of fluorescent probes that indicate various aspects of cell metabolism has further stimulated this area of research (McFeters et al, 1995;Porter et al, 1996;Davey et al, 1999).…”

Section: Bacteria and Flow Cytometrymentioning

confidence: 99%

“…Hutter and Eipel (1978) used Erythrosine B to label damaged yeast cells and since then many more viability probes have been put in use for bacteria (Table 3). There are two broad categories of physiological probes currently in use: 1) Those that indicate the state of membrane integrity or energization, and 2) those that are taken up by the viable cells and then modified intracellularly to yield fluorescent products (McFeters et al, 1995;Nybroe, 1995;Nebevon Caron et al, 1998). Among the first group there are exclusion stains, which do not penetrate intact and healthy membranes because of their molecular structure and size, but do penetrate cells with injured membranes and then stain nucleic acids (i.e.…”

Section: Single-cell Activitymentioning

confidence: 99%

Using flow cytometry for counting natural planktonic bacteria and understanding the structure of planktonic bacterial communities

Gasol¹,

Giorgio²

2000

Sci. Mar.

790

717

View full text Add to dashboard Cite

SUMMARY: Flow cytometry is rapidly becoming a routine methodology in aquatic microbial ecology. The combination of simple to use bench-top flow cytometers and highly fluorescent nucleic acid stains allows fast and easy determination of microbe abundance in the plankton of lakes and oceans. The different dyes and protocols used to stain and count planktonic bacteria as well as the equipment in use are reviewed, with special attention to some of the problems encountered in daily routine practice such as fixation, staining and absolute counting. One of the main advantages of flow cytometry over epifluorescence microscopy is the ability to obtain cell-specific measurements in large numbers of cells with limited effort. We discuss how this characteristic has been used for differentiating photosynthetic from non-photosynthetic prokaryotes, for measuring bacterial cell size and nucleic acid content, and for estimating the relative activity and physiological state of each cell. We also describe how some of the flow cytometrically obtained data can be used to characterize the role of microbes on carbon cycling in the aquatic environment and we prospect the likely avenues of progress in the study of planktonic prokaryotes through the use of flow cytometry.

show abstract

“…In many ecological and environmental studies, uorochromes have been coupled with various heterocyclic tetrazolium dyes and nalidixic acid to determine not only total microorganism numbers, but also the fraction of metabolically active microorganisms (Bianchi and Giuliano 1996;Bitton and Koopman 1982;Hernandez et al 1994;Huang et al 1995;Jarnagin and Luchsinger 1980;Maki and Remsen 1981;McFeters et al 1995;Rodriguez et al 1992;Soderstrom 1977;Tabor and Neihof 1982;Trevors 1984;Zimmerman et al 1978). However, in bioaerosol studies, the detection and quantitation of metabolically active microorganisms has been primarily based on plate count assays in which sample collection methods as well as microorganism nutritional requirements and culturability bias the results (Burge 1990;Buttner et al 1993;Flannigan 1993;Heinsohn 1994;Marchand et al 1995;Miller 1992;Stewart et al 1995;Terzieva et al 1996;Thorne et al 1992).…”

Section: Introductionmentioning

confidence: 99%

A Combined Fluorochrome Method for Quantitation of Metabolically Active and Inactive Airborne Bacteria

Hernandez

1999

Aerosol Science and Technology

View full text Add to dashboard Cite

ABSTRACT. To better understand the ecology of microorganisms in the environment and to quantify the concentrations of airborne microorganisms, methods are needed to count all microbial cells that are present and differentiate between those that are metabolically competent and those that are nonviable as they exist in situ. We developed and tested a direct epi uorescent method to estimate the quantity and activity of airborne bacteria aerosolized into full-scale rooms. Midget impingers, lled with the uorescing redox dye 5-cyano-2, 3-ditolyl tetrazolium chloride (CTC), were used to capture and stain metabolically active bacteria from room air. Both active and inactive bacteria were counterstained with a uorescein derivative and counted with an epi uorescent microscope. The choice of uorochrome stains allowed concurrent identi cation of active and inactive bacteria in the same microscopic eld. Airborne bacterial numbers determined by epi uorescence microscopy were compared to standard, nonselective colony counts cultured from midget impingers operated under identical conditions. Direct epi uorescent estimates of total airborne bacteria were higher than concurrent plate counts. However, estimates of active airborne bacteria were lower than concurrent plate counts. Variability experiments showed that the direct-count method was repeatable to within 35%.

show abstract

Physiological assessment of bacteria using fluorochromes

Cited by 141 publications

References 61 publications

Use of multiparameter analysis for Vibrio alginolyticus viable but nonculturable state determination

Use of multiparameter analysis for Vibrio alginolyticus viable but nonculturable state determination

Using flow cytometry for counting natural planktonic bacteria and understanding the structure of planktonic bacterial communities

A Combined Fluorochrome Method for Quantitation of Metabolically Active and Inactive Airborne Bacteria

Contact Info

Product

Resources

About