Introduction to Microbiology

Pt. 2
Dr. Linroy Christian

Microscopes and Microscopy

Microscopic examination of microorganisms makes use of either the light microscope or the electron microscope The compound light microscope is used for most routine examinations The electron microscope is used for special research purposes All microscopes use lenses to magnify the image of the cell

Microscopes and Microscopy

In general, the highest magnification obtained with the light microscope is x1000 and for the electron microscope x 100,000 Several applications of light microscopy are:
Bright-field microscopy Dark-ground microscopy Phase-contrast microscopy Fluorescence microscopy

Electron microscopes can be used in two basic ways:

Transmission electron microscope Scanning electron microscope

Bright-Field

Dark-Ground

Phase Contrast

Fluorescence

The Compound Light Microscope

SEM

TEM

Scanning Transmission Electron Microscope

The Electron Microscope

General Staining in Light Microscopy

Various dyes used to increase contrast:
Methylene blue Crystal violet Safranin

These dyes are positively charged and bind to negatively charged structures such as DNA and various structures on the cell surface

Differential Staining in Light Microscopy

The Gram stain is a very important differential staining technique in microbiology Bacterial cells can be split into two major groups based on gram staining:
Gram positive (stain purple) Gram negative (pink-red)

Bacterial cells are viewed with the x100 (oil immersion) lens in light microscopy

The Gram Stain

Add a drop of water to the slide Add bacteria and smear Air dry Heat fix Add crystal violet for 1 min Wash Add iodine/potassium iodide for 1 min Wash Decolourise for 10-15 Sec Wash Add safranin for 1 min Wash Blot dry

Size of Microbial Cells

Bacterial size is generally measured in micrometers (m) 1 m = 1 thousand times smaller than a millimeter (mm) Typical bacterial cell like Escherichia coli (E. coli) measures 1 x 3 m Typical eukaryotic cell ranges from 2 200 m Complex eukaryotes are made up of many cells

Overview of Cell Structure

Prokaryotes
Cytoplasmic membrane Cell wall Ribosomes Genetic material (DNA) Inclusions (carbon, nitrogen, sulfur, phosphorous)

Eukaryotes
Cytoplasmic membrane Cell wall Ribosomes Nucleus (chromosomes) Various organelles vacuoles

Nomenclature of Microbes
Full name is comprised of a genus and a species name First letter of genus name capitalised Species name in common letters Full name should be italicised