Microbial Cell Structure and Function Lecture 2024 PDF

Microbial cell structure and function (08203) Lectures 2024-2025 Assist. Prof. Asmaa Mohamed Youssif  Contents Microscopes Cells of Bacteria and archeae  Microscopes  The microscope is the microbiologist’s oldest and most ba...

Microbial cell structure and function (08203) Lectures 2024-2025 Assist. Prof. Asmaa Mohamed Youssif  Contents Microscopes Cells of Bacteria and archeae  Microscopes  The microscope is the microbiologist’s oldest and most basic tool for studying microbial structure.  Many types of microscopy are used and some are extremely powerful. So as a prelude to our study of cell structure, let’s first take a look at some common tools for visualizing cells with a goal of understanding how they work and what they can tell us.  To see microorganisms, one needs a microscope of some sort, either a light microscope or an electron microscope.  In general, light microscopes are used to examine cells at relatively low magnifications, and electron microscopes are used to examine cells and cell structures at very high magnification.  Microscopes  All microscopes employ lenses that magnify the image. Magnification, however, is not the limiting factor in our ability to see small objects. It is instead resolution—the ability to distinguish two adjacent objects as distinct and separate—that governs our ability to see the very small.  Although magnification can be increased virtually without limit, resolution cannot, because resolution s a function of the physical properties of light.  We begin with the light microscope, for which the limits of resolution are about 0.2 mm (mm is the abbreviation for micrometer, 10-6 m).  We then proceed to the electron microscope, for which resolution is considerably greater.  Microscopes I.The Compound Light Microscope  The light microscope uses visible light to illuminate cell structures. Several types of light microscopes are used in microbiology: bright-field, phase-contrast, differential interference contrast, dark-field, and fluorescence. (A) Bright-field microscope  specimens are visualized because of the slight differences in contrast that exist between them and their surroundings, differences that arise because cells absorb or scatter light to varying degrees.  Microscopes I. The Compound Light Microscope (A) Bright-field microscope  The modern compound light microscope contains two lenses, objective and ocular, that function in combination to form the image.  The light source is focused on the specimen by the condenser  Microscopes I. The Compound Light Microscope (A) Bright-field microscope  Bacterial cells are typically difficult to see well with the bright-field microscope because the cells themselves lack significant contrast with their surrounding medium.  Cells visualized by a form of light microscopy called phase-contrast overcome these limitations.  Pigmented microorganisms are also an exception because the color of the organism itself adds contrast, which makes them easier to visualize by bright-field optics  Microscopes I. The Compound Light Microscope (B) Phase contrast microscope  The phase-contrast microscope in particular is widely used in teaching and research for the observation of living preparations.  Phase-contrast microscopy is based on the principle that cells differ in refractive index (a factor by which light is slowed as it passes through a material) from their surroundings.  Light passing through a cell thus differs in phase from light passing through the surrounding liquid.  This subtle difference is amplified by a device in the objective lens of the phase-contrast microscope called the phase ring, resulting in a dark image on a light background  Microscopes I.The Compound Light Microscope C) Dark field microscope  the dark-field microscope, light reaches the specimen from the sides only. The only light that reaches the lens is that scattered by the specimen, and thus the specimen appears light on a dark background.  Resolution by dark-field microscopy is often better than by light microscopy, and some objects can be resolved by dark-field that cannot be resolved by bright-field or even by phase-contrast microscopes.  Dark-field microscopy is a particularly good way to observe microbial motility, as bundles of flagella (the structures responsible for swimming motility) are often resolvable with this technique  Microscopes I.The Compound Light Microscope  Microscopes I. The Compound Light Microscope D) Fluorescence Microscopy  The fluorescence microscope is used to visualize specimens that fluoresce, emitting light of one color after absorbing light of another color  Cells fluoresce because they either contain naturally fluorescent substances such as chlorophyll or other fluorescing components, or because they have been stained with a fluorescent dye.  DAPI (4′,6-diamidino-2-phenylindole) is a widely used fluorescent dye. DAPI stains cells bright blue because it complexes with the cell’s DNA  DAPI can be used to visualize cells in their natural habitats, such as soil, water, food, or a clinical specimen.  Fluorescence microscopy using DAPI is therefore widely used in clinical diagnostic microbiology and also in microbial ecology for enumerating bacteria in a natural environment or in a cell suspension.  Microscopes I. The Compound Light Microscope D) Fluorescence Microscopy  Microscopes I.The Compound Light Microscope (E) Differential Interference Contrast Microscopy  Differential interference contrast (DIC) microscopy is a form of light microscopy that employs a polarizer in the condenser to produce polarized light (light in a single plane).  The polarized light then passes through a prism that generates two distinct beams. These beams pass through the specimen and enter the objective lens where they are recombined into one. Because the two beams pass through substances that differ in refractive index, the combined beams are not totally in phase but instead interfere with each other, and this effect enhances subtle differences in cell structure.  Thus, by DIC microscopy, cellular structures such as the nucleus of eukaryotic cells (endospores, vacuoles, and inclusions of bacterial cells) appear more three- dimensional.  DIC microscopy is typically used on unstained cells as it can reveal internal cell structures that are nearly invisible by brightfield without the need for staining  Microscopes I.The Compound Light Microscope (E) Differential Interference Contrast Microscopy  Microscopes II. Electron Microscope  Electron microscopes use electrons instead of visible light (photons) to image cells and cell structures. In the electron microscope, electromagnets function as lenses, and the whole system operates in a vacuum.  Electron microscopes are fitted with cameras to allow a photograph, called an electron micrograph, to be taken. Two types of electron microscopy are in routine use in microbiology: transmission and scanning.  Microscopes II. Electron Microscope (A) Transmission Electron Microscopy  The transmission electron microscope (TEM) is used to examine cells and cell structure at very high magnification and resolution. The resolving power of a TEM is much greater than that of the light microscope, even allowing one to view structures at the molecular level.  This is because the wavelength of electrons is much shorter than the wavelength of visible light, and, as we have learned, wavelength affects resolution  For example, whereas the resolving power of a light microscope is about 0.2 micrometer, the resolving power of a TEM is about 0.2 nanometer, a thousand fold improvement.  With such powerful resolution, objects as small as individual protein and nucleic acid molecules can be visualized by transmission electron microscopy  Microscopes II. Electron Microscope (A) Transmission Electron Microscopy  Unlike photons, electrons are very poor at penetrating; even a single cell is too thick to penetrate with an electron beam. Consequently, to view the internal structure of a cell, thin sections of the cell are needed, and the sections must be stabilized and stained with various chemicals to make them visible.  A single bacterial cell, for instance, is cut into extremely thin (20–60 nm) slices, which are then examined individually by TEM  To obtain sufficient contrast, the sections are treated with stains such as osmic acid, or permanganate, uranium, lanthanum, or lead salts. Because these substances are composed of atoms of high atomic weight, they scatter electrons well and thus improve contrast.  If only the external features of an organism are to be observed, thin sections are unnecessary. Intact cells or cell components can be observed directly in the TEM by a technique called negative staining  Microscopes II. Electron Microscope (A) Transmission Electron Microscopy  Microscopes II. Electron Microscope (B) Scanning Electron Microscopy  For optimal three-dimensional imaging of cells, a scanning electron microscope (SEM) is used  In scanning electron microscopy, the specimen is coated with a thin film of a heavy metal, typically gold. An electron beam then scans back and forth across the specimen.  Electrons scattered from the metal coating are collected and projected on a monitor to produce an image  In the SEM, even fairly large specimens can be observed, and the depth of field (the portion of the image that remains insharp focus) is extremely good.  A wide range of magnifications can be obtained with the SEM, from as low as 15* up to about 100,000*, but only the surface of an object is typically visualized.  Electron micrographs taken by either TEM or SEM are originally taken as black- and-white images.  2. Cells of bacteria & Archaea  Two features of prokaryotic cells that are immediately obvious upon microscopic examination are their shape and small size.  A variety of shapes are possible, and in general, prokaryotes are extremely small relative to eukaryotic cells.  Cell shape can be useful for distinguishing different cells and undoubtedly has some ecological significance, but cell shape rarely has phylogenetic relevance.  By contrast, the typically small size of prokaryotes affect many aspects of their biology.  2. Cells of bacteria & Archaea I. Cell morphology  In microbiology, the term morphology means cell shape. (A) Major cell morphologies  A cell that is spherical or ovoid in morphology is called a coccus (plural, cocci).  A cylindrically shaped cell is called a rod or a bacillus.  Some rods form spiral shapes and are called spirilla.  The cells of some prokaryotes remain together in groups or clusters after cell division, and the arrangements are often characteristic. For instance, some cocci form long chains (for example, the bacterium Streptococcus), others occur in three-dimensional cubes (Sarcina), and still others in grapelike clusters (Staphylococcus).  2. Cells of bacteria & Archaea I. Cell morphology (A) Major cell morphologies  A few bacterial groups are immediately recognizable by the unusual shapes of their individual cells.  Examples include the spirochetes, which are tightly coiled bacteria; appendaged bacteria, which possess extensions of their cells as long tubes or stalks; and filamentous bacteria, which form long, thin cells or chains of cells  The cell morphologies described here should only be considered representative; many variations of these morphologies are known.  For example, there are fat rods, thin rods, short rods, and long rods, a rod simply being a cell that is longer in one dimension than in the other.  2. Cells of bacteria & Archaea I. Cell morphology (A) Major cell morphologies Thank you