Measurement of marine picoplankton cell size by using a cooled, charge-coupled device camera with image-analyzed fluorescence microscopy

Viles, Charles L.; Sieracki, Michael E.

doi:10.1128/aem.58.2.584-592.1992

Cited by 64 publications

(28 citation statements)

References 21 publications

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“…Total bacteria counts were done by flow cytometry using PicoGreen (Molecular Probes, Inc.) for the October cruise (CH-1301) and by image analysis from DAPI images (Viles and Sieracki 1992) for March (CH-0402), because of PicoGreen staining problems. Cell counts by microscopy and flow cytometry have been shown to be comparable (Sieracki et al 1999;Ducklow et al 2001).…”

Section: Methodsmentioning

confidence: 99%

“…Cell counts by microscopy and flow cytometry have been shown to be comparable (Sieracki et al 1999;Ducklow et al 2001). Cell biovolumes were measured by image analysis using the DAPI images and previously described algorithms (Sieracki et al 1989;Viles and Sieracki 1992). AAP cell sizes were measured in their corresponding DAPI images to compare with total bacteria.…”

Section: Methodsmentioning

confidence: 99%

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Distribution of planktonic aerobic anoxygenic photoheterotrophic bacteria in the northwest Atlantic

et al. 2006

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Aerobic anoxygenic photoheterotrophic (AAP) bacteria can use both dissolved organic matter and light for energy production, but their photosynthesis does not produce oxygen. We measured AAP bacterial cell and bacteriochlorophyll distributions in the northwest Atlantic, from the coast of the Gulf of Maine to the Sargasso Sea, in October 2001 and March 2002. The abundance of AAPs ranged from 7 ϫ 10 3 to 9.8 ϫ 10 4 cells mL Ϫ1 (mean, 2.9 ϫ 10 4 mL Ϫ1 ) in surface waters, or between 1% and 9% (mean, 2.3%) of total bacteria. Mean abundances in October in the Gulf of Maine (6.6 ϫ 10 4 mL Ϫ1 ) were about five times higher than those measured in March (1.3 ϫ 10 4 mL Ϫ1 ), whereas the mean Sargasso Sea values were not different between October and March. AAP cells were larger than other bacteria, so AAP biomass ranged from 2% to 13% of total bacterial biomass. AAP cells were higher in abundance, biomass, and proportion of total bacteria in productive coastal and shelf waters than in the Sargasso Sea. Cell quotas of bacteriochlorophyll were low and quite variable, ranging from 0.02 to 0.17 fg cell Ϫ1 (mean, 0.08 fg cell Ϫ1 ). Our results indicate possible control by temperature and organic and inorganic nutrients on the distribution of planktonic AAPs, but they do not support the idea that they are specifically adapted to oligotrophic conditions. A unique type of bacterial phototrophic metabolism has recently been shown to be widespread in the ocean (Kolber et al. 2000(Kolber et al. , 2001Goericke 2002). These bacteria, termed aerobic anoxygenic photoheterotrophs (AAPs), use bacteriochlorophyll (BChl) a but are differentiated from purple and green anoxygenic phototrophs by their requirements for oxygen, synthesis of BChl a pigments under dark aerobic conditions, and by their dependence on respiration for energy, with photosynthesis having a stimulatory effect on growth. These cells can use light to fix carbon (Koblízek et al. 2003) and/or to synthesize adenosine triphosphate and nicotinamide adenine dinucleotide phosphate (NADPH), but they do not produce oxygen. They were first isolated from a variety of marine environments by Shiba et al. (1979). Direct cultivation from water samples was rarely successful (0.9% of isolates) relative to isolation from seaweeds and beach AcknowledgmentsWe thank Capt. Dick Hogus and the crew of the R/V Cape Hatteras. Brian Thompson, Rich Entel, Renée Lagasse, and Helga Gomes prepared samples and measured Chl a on the cruises. Zbignew Kolber provided microscope advice. Robert Guillard provided a critical review of the manuscript.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Distribution of planktonic aerobic anoxygenic photoheterotrophic bacteria in the northwest Atlantic

et al. 2006

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“…A Gaussian 5 ¥ 5 filter was then applied to remove any noise created by the Laplacian filter. Images were then thresholded as described by Viles and Sieracki (1992) using the global visual threshold. Cells were then counted and cell parameters measured.…”

Section: Image Analysismentioning

confidence: 99%

Bacterial community shifts in organically perturbed sediments

Bissett

Burke

Cook

et al. 2006

Environmental Microbiology

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Bacterial abundance, diversity and sediment function were investigated in organically perturbed sediments under Tasmanian salmon (Salmo salar) farms and adjacent reference sites. Bacterial numbers increased as farming and organic loading progressed through the farm stocking cycle and declined during the fallow period, although not to prestocking levels. Bacterial numbers ranged between approximately 2 x 10(8) and 3 x 10(9) cells per gram of sediment and were higher at cage sites than reference sites. Microelectrode and respiration data also demonstrated a clear effect of organic loading on sediments. Denaturing gradient gel electrophoresis (DGGE) showed that bacterial communities shifted both in response to farm loading and its cessation. A seasonal effect on microbial communities was also evident. Although bacterial communities did shift again during the fallowing period, this shift was not necessarily a return to preloading communities. The complexity of community shifts may be affected by the vast functional redundancy of bacterial groups. All bacterial communities, including those at reference sites, were highly dynamic. Respiration studies of amended sediments indicated that fish farm sediments were at least as resilient and diverse as reference site communities. The results of this study indicate that the functional redundancy of highly complex bacterial communities contributes to their robustness. The relationship between diversity and stability in bacterial communities remains unclear and requires further investigation before an understanding of bacterial response to perturbation is possible.

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“…In complex images, locating and sizing objects of interest are often best achieved by edge detection [9,10]. This is particularly true for images where the brightness of objects of interest varies over a wide range.…”

Section: Green Channel Processingmentioning

confidence: 99%

“…Setting a suitable threshold value for bright bacteria resulted in fainter bacteria not being detected. On the other hand, setting a threshold value suitable for detecting faint bacteria resulted in the size of bright bacteria being overestimated [9] and non-bacterial features being selected and retained, at least in part, after con¢rmation with the blue channel. It then proved di¤cult in subsequent analytical steps to distinguish bacteria from non-bacterial features on morphological grounds because these features were often in the bacterial shape and size range.…”

Section: Validation Of Image Analysis Proceduresmentioning

confidence: 99%

Quantification of the in situ distribution of soil bacteria by large-scale imaging of thin sections of undisturbed soil

Nunan¹

2001

FEMS Microbiology Ecology

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A method for determining the number and in situ spatial distribution of bacterial cells over spatial scales ranging from micrometres to centimetres in mineral soils is described. Biological thin sections of undisturbed cores of soil were prepared in order to preserve the spatial distribution of bacterial cells. Composite (tessellated) images in which individual bacteria can be resolved within an area of 0.282 mm 2 were acquired by means of a motorised scanning microscope stage. An image processing and analysis procedure was developed to determine the numbers and locations of bacterial cells in the composite images. The image processing procedure first homogenised the background of the images and then discriminated between bacteria and non-bacterial features using the colour and morphological properties of the images of the bacterial cells. Feature edges were detected in the green channel of colour (red, green, blue) images and bacterial cell edges were confirmed in the blue channel after elimination of autofluorescent features in the red channel. No significant difference was found between the number of bacteria or associated distributions determined automatically and control values derived interactively on individual fields of view. Data relating to total bacterial counts in thin sections and in paired dispersed samples suggested that all soil bacteria were being visualised in thin sections. Significant differences between samples taken from a depth profile of a fallow arable soil were found for both cell numbers and for cell distribution as measured by an index of dispersion. ß

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Measurement of marine picoplankton cell size by using a cooled, charge-coupled device camera with image-analyzed fluorescence microscopy

Cited by 64 publications

References 21 publications

Distribution of planktonic aerobic anoxygenic photoheterotrophic bacteria in the northwest Atlantic

Distribution of planktonic aerobic anoxygenic photoheterotrophic bacteria in the northwest Atlantic

Bacterial community shifts in organically perturbed sediments

Quantification of the in situ distribution of soil bacteria by large-scale imaging of thin sections of undisturbed soil

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