Microscope Types and Culture Media

• Culture media are classified based on physical state, chemical composition, and functional type.

Types of Microscopes

• Microscopes use lenses to enlarge small objects for viewing.

Light Microscopes

• Light microscopes use visible light and a system of lenses for magnification.
• They use ocular and objective lenses.
• To calculate final magnification, one must multiply the enlarging power of both the ocular and objective lenses.
• For example, a 10x ocular lens and a 4x objective lens yields a 40x final magnification.

Types of Light Microscopes

• There are several types of light microscopes, each with unique properties and uses:
• Bright-field microscopes have a light background and typically require fixing and staining of the specimen.
• Dark-field microscopes are used to view unfixed and unstained specimens, such as living organisms, rendering a dark background and a bright specimen.
• Phase-contrast microscopes are useful for viewing unfixed, transparent specimens and effective for observing cell division, cell motility, and dynamic states of cell organelles.
• Fluorescence microscopes are used to visualize specimens stained with fluorescent substances.
• Confocal microscopes can produce sharper images than traditional microscopes, allow for visualization of different planes of a specimen, and can display images three-dimensionally using electronic staining.

Electron Microscopes

• Electron microscopes use electrons as a source of energy.
• Transmission electron microscopes (TEM) transmit electron beams through the specimen, requiring special preparation, sectioning, and staining.
• TEM produces two-dimensional images with good internal detail.
• Scanning electron microscopes (SEM) scan the surface of an object without sectioning, producing three-dimensional images.

The Chemistry of Life

• Matter is anything that occupies space and has mass, existing as solid, liquid, or gas, and is composed of elements.
• Elements combine to form chemical compounds.
• Atoms are the smallest particles of an element.
• The atomic nucleus contains protons and neutrons.
• Protons are positively charged particles.
• Neutrons are particles without charge.
• Electrons are negatively charged particles.
• Ions are electrically charged atoms, molecules, or particles.
• Cations are positively charged due to electron loss.
• Anions are negatively charged due to electron gain.
• Electrolytes are free ions present in a solvent.

Chemical Reactions

• Synthesis (dehydration synthesis) reactions form larger molecules from smaller ones, removing water and requiring energy (endergonic).
• Hydrolysis reactions break down large molecules, requiring water and releasing energy (exergonic).
• Reduction reactions involve a gain of electrons, while oxidation reactions involve a loss of electrons; both occur simultaneously in redox (reduction-oxidation) reactions.

Compounds

• Hydrophilic compounds are water-soluble (water-loving).
• Hydrophobic compounds are insoluble in water (water-repelling).
• Inorganic compounds do not contain carbon (except CO and CO2), such as acids, bases, salts (which dissolve in water and do not release hydrogen or hydroxyl ions), and water.
• Organic compounds contain carbon and hydrogen.

Organic Compounds

• Important organic compounds include carbohydrates, proteins, and lipids (fats and oils), and nucleic acids (pentose sugar, phosphate, nitrogen base).
• Purines: Adenine (A) and guanine (G).
• Pyrimidines: Cytosine (C), thymine (T), and uracil (U).
• DNA (deoxyribonucleic acid) stores genetic information.
• RNA (ribonucleic acid) is involved in various cellular processes
• ATP (adenosine triphosphate) is the cell's primary energy source.

Carbohydrates

• Monosaccharides include glucose, fructose (fruit sugar), and galactose.
• Disaccharides include sucrose (table sugar, sweeteners), maltose, and lactose (milk sugar).
• Polysaccharides include glycogen (storage form of sugar in animals) and starch (storage form of sugar in plants).

Solutions

• Solutions are mixtures of one or more substances (solutes) dispersed in a dissolving medium (solvent).
• Isotonic solutions have equal solute concentrations inside and outside the cell.
• Hypertonic solutions have a higher solute concentration outside the cell.
• Hypotonic solutions have a lower solute concentration outside the cell.
• Body fluids make up 60% of total body weight, with 2/3 within intracellular fluid (40%) and 1/3 in extracellular fluid (20%).
• In the ECF, 3/4 is in the interstitial fluid (15%) and 1/4 in plasma (5%) which is intravascular.

Electrolytes

• Electrolytes: In ECF, Sodium (Na+) is the main cation and Chloride is the main anion.
• In ICF, Potassium (K+) is the main cation and Phosphate is the main anion.

pH Scale

• The pH scale measures the acidity or alkalinity of a solution.
• It is based on the hydrogen ion concentration and ranges from 0 to 14.
• Pure water is neutral with a pH of 7.
• Substances that dissociate in water can form acids, bases, or salts.
• Acids have a pH less than 7 and release hydrogen ions.
• Bases have a pH greater than 7.

Culture Media

• A pure culture has one type of organism.
• A mixed culture has two or more species.

Physical States of Culture Media

• Liquid media (broth) are water-based solutions.
• Semisolid media clot at room temperature.
• Solid media are solid at room temperature and become liquid at boiling temperature.

Chemical Composition of Culture Media

• Chemically defined (synthetic) media contain pure compounds with a chemical content specified by an exact formula.
• Complex media (nonsynthetic) contain one or more components that are not chemically defined and cannot be represented by an exact chemical formula.

Functional Types of Media

• General purpose media are for microbes that do not have special growth requirements, with nutrient agar and trypticase soy agar (TSA) as examples.
• Enriched media contain complex organic substances such as blood, including blood agar (detects the ability to produce hemolysins), chocolate agar (for respiratory bacteria), and Thayer-Martin agar (for Neisseria).
• Selective media contain one or more agents that inhibit the growth of a certain microbe, favoring or selecting a certain microbe and allowing it to grow, such as phenylethyl alcohol agar (selective for gram-positive bacteria).
• Differential media contain neutral red (yellow when neutral, becomes red when acidic by E. coli, a lactose fermenter, while Salmonella is non-lactose fermenter) and MacConkey agar (selective & differential).
• Mannitol Salt Agar is selective & differential media - its 7.5% NaCl inhibits most organisms, except staphylococcus, which can stand high salt). Live Media
• Certain bacteria require host animals to grow.
• Mycobacterium Leprae on mice footpad

Basic Procedures to Examine & Characterize Microbes (The Five I's)

• Inoculation involves introducing a sample into a medium.
• Incubation occurs at a temperature generally between 20° C and 40° C.
• Isolation can be achieved using a spread plate, which is used to quantify the number of bacteria in a solution.
• Colonies are evenly distributed over the agar surface and are easily counted.
• Inspection involves observing colonies macroscopically, including growth patterns in broths.
• Bacteria grown in broth exhibit growth patterns, e.g., pellicle (a thick growth at the top of the tube), sediment (at the bottom of the tube), or turbid growth (throughout the tube).
• Identification involves determining the type of microbe using morphological, physiological, and biochemical characteristics.