Microbiology: Koch's Postulates, Staining, and Bacterial Structures

What are Koch's postulates?

four criteria developed by Robert Koch to establish that a specific microbe causes a specific infectious disease, requiring the organism to be present in all cases, isolated and grown in pure culture, cause the disease when introduced to a healthy host, and be re-isolated from the new host

Koch postulate 1

The microorganism must be found in abundance in all organisms suffering from the disease, but not in healthy organisms

Koch postulate 2

The microorganism must be isolated from the diseased organism and grown in pure culture.

Koch postulate 3

The pure culture of the microorganism must cause the disease when introduced into a healthy, susceptible host.

Koch postulate 4

The microorganism must be re-isolated from the experimentally infected host and identified as the original organism. 

What is resolution in microscopy?

the ability to distinguish two very close points as separate entities, defined by the shortest distance at which they remain distinct

What is darkfield illumination used for?

It is used to examine specimens that cannot be distinguished from the background, such as unstained, living specimens.

What does heat fixing preserve?

preserves overall morphology but not internal structures.

What are the two groups of microorganisms based on Gram stain?

Gram-positive (Gm+) and Gram-negative (Gm−).

What is acid-fast staining used for?

It is used for Mycobacterium, such as M. tuberculosis and M. leprae, which have a high lipid content in their cell walls.

What is negative staining used to visualize?

It is used to visualize capsules, which appear colorless against a stained background.

What does flagellar staining do?

It uses a mordant to increase the thickness of the flagella for visualization.

What are the shapes of prokaryotic cells?

Cocci, bacili, vibrio, spirilla, and spirochetes.

Cocci

Sphere shaped bacteria

Bacili

Rob shaped bacteria

Vibrio

Comma shaped bacteria

Spirllia

Rigid helicial shaped bacteria

Spirochetes

Flexible helical shaped bacteria

What are halophiles?

Halophiles grow optimally in high salt concentrations, defined as >0.2 M. Extreme halophiles require >2 M salt.

What are macronutrients required for prokaryotic cells?

C, O, H, N, S, P, K+, Ca2+, Mg2+, and Fe2+/3+.

What is the role of the plasma membrane in prokaryotic cells?

It is crucial for metabolic processes such as respiration and lipid synthesis.

How does passive diffusion work?

It moves substances like H2O, O2, and CO2 from higher to lower concentration.

What is facilitated diffusion?

It involves carrier molecules (permeases) facilitating transport, requiring a smaller concentration gradient than passive diffusion.

What is active transport?

It is an energy-dependent process that moves substances against the concentration gradient.

What is group translocation?

It is a process where molecules are modified during transport, such as the phosphoenolpyruvate:sugar phosphotransferase system (PTS) in E. coli and Salmonella.

What aids in iron uptake in prokaryotes?

Siderophores, such as enterobactin in E. coli, aid in iron uptake by complexing with Fe3+.

What is the bacterial cell wall composed of?

Peptidoglycan (murein), the synthesis of which is inhibited by penicillin.

What is the Gram stain reaction based on?

It is probably due to cell wall structure.

What color do Gram-positive cells appear after Gram staining?

Purple.

What color do Gram-negative cells appear after Gram staining?

Pink.

What is the peptidoglycan backbone made of?

Alternating sugars: N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM).

What is the outer membrane of Gram-negative bacteria composed of?

Lipids, lipoproteins, and lipopolysaccharide (LPS), but no teichoic acids.

What is the function of the O side chain of LPS?

It provides protection from host defenses and is immunogenic.

What stabilizes the outer membrane structure in Gram-negative bacteria?

Lipid A.

What is the permeability of the outer membrane compared to the plasma membrane?

The outer membrane is more permeable due to the presence of porins.

What are capsules in prokaryotes?

Usually polysaccharides, well organized, not easily removed, and resist phagocytosis.

What are slime layers?

Diffuse, unorganized polysaccharides that are easily removed.

What are S-layers?

Structured layers of protein or glycoprotein common among Archaea.

What is unique about some archaeal plasma membranes?

They have a monolayer structure instead of a bilayer structure.

Why is the Gram stain not useful for Archaea?

Because they lack peptidoglycan.

What is the composition of prokaryotic ribosomes?

70S, composed of a large 50S subunit and a small 30S subunit.

What is the nucleoid?

The location of the chromosome, which is usually circular and not membrane-bound.

What are plasmids?

Usually small, closed circular DNA molecules that are extrachromosomal and carry genes for selective advantage.

What are fimbriae?

Short, thin, proteinaceous appendages used for attachment to surfaces.

What are sex pili?

Longer, thicker, and less numerous appendages required for mating (e.g., conjugation).

How is bacterial flagella movement driven?

By a H+ gradient.

What happens during flagellum rotation?

Counterclockwise results in forward motion (run); clockwise disrupts the run, causing a tumble.

How do spirochetes achieve motility?

Via periplasmic axial fibrils, resulting in a flexing/spinning movement.

What is twitching motility?

Involves Type IV pili and is observed in groups of cells that are contacting each other.

What is chemotaxis?

Movement towards or away from a chemical, detected by cell surface chemoreceptors.

What is the bacterial endospore?

A dormant and resistant structure to numerous environmental conditions.

What contributes to endospore resistance?

Calcium complexed with dipicolinic acid, small acid-soluble DNA-binding proteins (SASPs), a dehydrated core, the spore coat, and DNA repair enzymes.

When does sporulation commence?

When growth ceases due to lack of nutrients.

What is sterilization?

The elimination of all microorganisms and spores.

What is disinfection?

The killing, inhibition, or removal of pathogens, but not necessarily sterilizing.

What are disinfectants used on?

Inanimate objects.

What is antisepsis?

The prevention of infection of living tissue.

What are antiseptics?

Applied to tissue and are less toxic.

What are -cidal agents?

Agents that kill, such as germicides.

What are -static agents?

Agents that inhibit growth.

What is filtration in microbiology?

A mechanical method that physically removes microorganisms from liquids and air.

What are HEPA filters used for?

They are used in laminar flow biological safety cabinets to provide a clean working environment.

What is moist heat sterilization effective against?

All types of microorganisms.

What are the effects of moist heat?

Degrading nucleic acids, denaturing proteins, and disrupting membranes.

What is dry heat sterilization?

Less effective than moist heat and requires higher temperatures and longer exposures.

What does pasteurization do?

Involves controlled heating at temperatures below boiling to reduce the total microbial population and increase shelf life.

What is the decimal reduction time (D value)?

The time to kill 90% of microorganisms or spores at a specific temperature.

What is pasteurization?

Controlled heating at temperatures below boiling to reduce microbial population and increase shelf life.

What does the D value represent?

The time to kill 90% of microorganisms or spores at a specific temperature.

What is the z value?

The increase in temperature required to reduce the D value by 1/10.

What is the purpose of UV radiation at 260nm?

Used for surface sterilization, kills microorganisms, and inactivates viruses.

What type of radiation can penetrate deep and destroy cells?

Ionizing radiation, usually gamma rays.

What do phenolics do?

Denature proteins and disrupt cell membranes; they are tuberculocidal.

What is the effect of alcohols as disinfectants?

Bactericidal and fungicidal, but not sporicidal; they denature proteins.

What is iodine used for?

A skin antiseptic that oxidizes cell constituents and iodinates proteins.

What does chlorine do?

Oxidizes cell constituents.

What is the role of heavy metals in microbial control?

They are usually toxic and inactivate proteins.

What are cationic detergents effective against?

Most bacteria by disrupting membranes and denaturing proteins.

What are aldehydes used for?

They are chemical sterilants and sporicidal, inactivating nucleic acids and proteins.

What is the Minimum Inhibitory Concentration (MIC)?

The smallest amount of an agent needed to inhibit the growth of a microorganism.

What is the zone of inhibition?

The area of no growth around the disc in the Disc diffusion technique.

Who developed the concept of selective toxicity?

Paul Ehrlich in 1904.

Who discovered penicillin?

Alexander Fleming in 1928.

What is the therapeutic dose?

The drug level required for clinical treatment.

What is the toxic dose?

The drug level at which the drug becomes too toxic for the patient.

What are narrow-spectrum drugs?

Drugs that attack only a few different pathogens.

What are broad-spectrum drugs?

Drugs that attack many different pathogens.

What is the main mechanism of action for penicillins?

They block the enzyme that catalyzes transpeptidation.

What is a key feature of penicillins?

The β-lactam ring.

What do sulfonamides inhibit?

Folic acid synthesis enzymes.

What is the function of the TCA cycle?

To oxidize acetyl-CoA and produce energy carriers.

What is produced in the TCA cycle for each acetyl-CoA oxidized?

2 CO2, 3 NADH, 1 FADH2, and 1 GTP.

What is the role of the electron transport chain (ETC)?

To produce most ATP by oxidizing NADH and FADH2.

What are amphibolic pathways?

Pathways that are both catabolic and anabolic, such as glycolysis and the TCA cycle.

What are the three common pathways for glucose catabolism?

Glycolysis, pentose phosphate pathway, and Entner-Doudoroff pathway.

What is the net summary of glycolysis?

Glucose + 2 ADP + 2 Pi + 2 NAD+ → 2 pyruvate + 2 ATP + 2 NADH + 2 H+.

What does fermentation use as an electron acceptor?

An endogenous electron acceptor.

What is the difference between aerobic and anaerobic respiration?

Aerobic uses O2 as an electron acceptor, while anaerobic uses other exogenous electron acceptors.

What is the significance of acquired resistance?

It arises from spontaneous resistant mutants and can be transmitted via horizontal gene transfer.

What is drug inactivation?

A mechanism of resistance where enzymes like penicillinases inactivate the drug.