Scanning electron microscope study of Saccharomyces cerevisiae spheroplast formation

Pringle, Alastair T.; Forsdyke, J; Rose, Anthony H.

doi:10.1128/jb.140.1.289-293.1979

Cited by 19 publications

(5 citation statements)

References 20 publications

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“…Note that the border of the cell is surrounded by an artifactual structure resulting from the contact between the AFM probe and the pore edges. The cell surface is smooth and shows a ∼1 µm circular protrusion which can be attributed to a bud scar, in agreement with previous electron microscopy observations (Pringle et al , 1979; Koch and Rademacher, 1980). Note that birth scars were sometimes observed.…”

Section: Resultssupporting

confidence: 91%

“…Previous cytological and chemical investigations have shown that the cell walls of S. cerevisiae consist primarily of a microfibrillar array of β1–3 glucans overlaid by an outer layer of β1–6 glucans and mannoproteins (Pringle et al , 1979; Fleet, 1991; Kreutzfeldt and Witt, 1991; Osumi, 1998). Accordingly, enzyme digestion studies have revealed that the cell wall is sensitive to protease (Zlotnik et al , 1984) and β‐glucanase (Fleet, 1991).…”

Section: Resultsmentioning

confidence: 99%

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Real‐time imaging of the surface topography of living yeast cells by atomic force microscopy

Ahimou

Touhami

Dufrêne

2002

Yeast

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Atomic force microscopy (AFM) was used to image the surface topography of living Saccharomyces cerevisiae cells at high resolution and to monitor enzyme digestion of the cell wall in real time. Apart from the presence of bud scars, the surface of native cells imaged in aqueous solution was homogeneous and smooth. Topographic images of the surface were recorded to a lateral resolution of 2 nm without significant modification of the surface morphology. Successive images of single cells were collected at fixed time intervals following addition of protease and amyloglucosidase solutions. Protease caused a progressive increase of surface roughness. Large depressions surrounded by protruding edges, ∼50 nm in height, were formed and attributed to the erosion of the mannoprotein outer layer. By contrast, no modification of the cell surface was noted upon addition of amyloglucosidase, which was consistent with the cell wall biochemical composition. These results indicate that AFM is a complementary tool to electron microscopy in that it allows the surface of living cells to be explored directly in real time.

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Section: Resultssupporting

confidence: 91%

Section: Resultsmentioning

confidence: 99%

Real‐time imaging of the surface topography of living yeast cells by atomic force microscopy

Ahimou

Touhami

Dufrêne

2002

Yeast

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“…Similar features have been reported for fungal spores and bacterial cells . They can be ascribed to the stretching of flexible cell surface macromolecules, i.e., glucans and mannoproteins in the present work. , Interestingly, we also note that the mean rupture length was correlated with the adhesion force value, i.e., largest rupture lengths being observed at pH 4, suggesting that the adhesiveness of cell surface polymer chains may depend on their conformation.…”

Section: Resultssupporting

confidence: 89%

Probing Microbial Cell Surface Charges by Atomic Force Microscopy

et al. 2002

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Most microorganisms possess a negative surface charge under physiological conditions due to the presence of anionic carboxyl and phosphate groups. Cell surface charge plays an important role in controlling cell adhesion and aggregation phenomena, as well as antigen−antibody, cell−virus, cell−drug, and cell−ions interactions. We have used atomic force microscopy (AFM) with chemically functionalized probes to investigate the surface charges of yeast cells. Force−distance curves and adhesion maps recorded with probes terminated with ionizable carboxyl groups were strongly influenced by pH: while no adhesion was measured at neutral/alkaline pH, multiple adhesion forces were recorded at pH ≤ 5. Three pieces of evidence indicated that these changes were related to differences in the ionization state of the cell surface functional groups. First, the adhesion force vs pH curve was correlated with microelectrophoresis data, the pH of the largest adhesion force corresponding to the cell isoelectric point, i.e., pH 4. Second, treating the cells with Cu(II) ions caused a reversal of the cell surface charge at neutral pH and promoted the adhesion toward the negatively charged probe. Third, control experiments using nonionizable hydroxyl-terminated probes indicated that the changes in adhesion forces were not simply due to the titration of the probe surface charges. This study shows that AFM with chemically modified probes is a valuable approach in microbiology and biophysics for probing the local electrostatic properties of microbial cell surfaces.

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“…To minimize the risk of contamination by reusable equipment and maximize the fungal DNA yield from blood specimens containing small quantities of infecting fungus, we chose an enzymatic method of fungal cell wall disruption for the extraction procedure. ␤-1,3-Glucanase enzymes hydrolyze glucose polymers at ␤-1,3glucan linkages in fungi, releasing laminaryipentaose and resulting in fungal spheroplasts, which are modified organisms that have partially lost the cell wall and that have increased osmotic sensitivity (37). Until the 1970s, snail gut enzyme was the prototype enzyme used for fungal cell wall lysis, but the preparation had variable activity from batch to batch (26,27).…”

Section: Discussionmentioning

confidence: 99%

Panfungal PCR Assay for Detection of Fungal Infection in Human Blood Specimens

Burik

Myerson

Schreckhise³

et al. 1998

J Clin Microbiol

233

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A novel panfungal PCR assay which detects the small-subunit rRNA gene sequence of the two major fungal organism groups was used to test whole-blood specimens obtained from a series of blood or bone marrow transplant recipients. The 580-bp PCR product was identified after amplification by panfungal primers and hybridization to a 245-bp digoxigenin-labeled probe. The lower limit of detection of the assay was approximately four organisms per milliliter of blood. Multiple whole-blood specimens from five patients without fungal infection or colonization had negative PCR results. Specimens from 11 infected patients had positive PCR results. Blood from three patients with pulmonary aspergillosis had positive PCR results: one patient’s blood specimen obtained in the week prior to the diagnosis of infection by a positive bronchoalveolar lavage fluid culture result was positive by PCR, and blood specimens obtained from two patients 1 to 2 days after lung biopsy and which were sterile by culture were positive by PCR. The blood of four patients with candidemia, three patients with mixed fungal infections, and one patient with fusariosis also had positive PCR signals. The panfungal PCR assay can detect multiple fungal genera and may be used as an adjunct to conventional methods for the detection of fungal infection or for describing the natural history of fungal infection. Further studies are needed to define the sensitivity and specificity of this assay for the diagnosis of fungal infection prior to the existence of other clinical or laboratory indications of invasive fungal infection.

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Scanning electron microscope study of Saccharomyces cerevisiae spheroplast formation

Cited by 19 publications

References 20 publications

Real‐time imaging of the surface topography of living yeast cells by atomic force microscopy

Real‐time imaging of the surface topography of living yeast cells by atomic force microscopy

Probing Microbial Cell Surface Charges by Atomic Force Microscopy

Panfungal PCR Assay for Detection of Fungal Infection in Human Blood Specimens

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