Genetics and Molecular Biology Quiz

Course Information

• Course title: CSF-6250 Integrative Pathophysiology & Biomechanics in Health Care 2
• Module: Microbes in Human Health and Disease
• Unit: 03 - Bacteria

Unit 3 Objectives

• Differentiate prokaryotic and eukaryotic cells
• Recognize structural and functional features of bacterial species useful for diagnostic purposes
• Recognize methods for measuring microbial growth
• List factors affecting microbial growth
• Name the three processes of horizontal gene transfer in bacteria
• Recognize the purpose and applications of PCR and CRISPR

Lesson 1: Prokaryotic Cells

• Salmonella typhimurium invades cultured cells

The Prokaryotic Cell

• Prokaryotes are small and simple (0.1-5 mm)

• They are unicellular

• They lack a nucleus

• They have circular DNA

• They lack membrane-bound organelles

• Components of a prokaryotic cell include: plasma membrane, cell wall, capsule, pilus, flagellum, cytoplasm, ribosome, nucleoid, inclusion, and plasmid.

Prokaryotic and Eukaryotic Cells

• Prokaryotes: small, simple cells (0.1-5 mm)
• Unicellular
• Lack a nucleus, circular DNA, and membrane-bound organelles.
• Eukaryotes: large, complex cells (10-100 mm)
• Uni- or multicellular
• Possess a nucleus, linear DNA, and membrane-bound organelles.

Bacterial Cell Shapes and Arrangements

• Bacterial shapes are species-specific and useful in diagnosing infections.
• Common shapes include coccus (round), bacillus (rod), vibrio (curved rod), coccobacillus (short rod), spirillum (spiral), and spirochete (long, loose, helical spiral). Common arrangements include single coccus, diplococcus, tetrad, streptococcus, staphylococcus, and chain of rods.

CM and Cytoplasm of Prokaryotic Cells

• Cell membrane (CM): lipid bilayer, without cholesterol, high protein content
• Ribosomes: 70S (vs 80S in eukaryotic cells); target for certain antibiotics
• Inclusions: storage of resources in polymerized form (e.g., volutin in Corynebacterium diphtheriae)

External Structures of Prokaryotic Cells: Cell Wall

• Major component: peptidoglycan
• Protects from osmotic pressure
• Several types of antibiotics inhibit peptidoglycan synthesis (e.g., penicillin)

External Structures of Prokaryotic Cells: Gram-positive and Gram-negative Cell Wall

• Gram-positive: thicker peptidoglycan layer, lacks outer membrane, and contains teichoic acid.
• Gram-negative: thinner peptidoglycan layer, has an outer membrane, and contains lipopolysaccharide (LPS).
• LPS is an endotoxin.

External Structures of Prokaryotic Cells: Gram-positive and Gram-negative Stain

• Gram staining differentiates between Gram-positive and Gram-negative bacteria.
• Gram-positive bacteria stain purple; Gram-negative bacteria stain pink.

External Structures of Prokaryotic Cells: Clinical Significance of Cell Wall Type

• Gram-positive: high susceptibility to penicillin and lysozyme, low sensitivity to heat and disinfectants, and lack of endotoxin.
• Gram-negative: low susceptibility to penicillin and lysozyme, high sensitivity to heat and disinfectants, and presence of endotoxin.
• Various species produce exotoxins.

External Structures of Prokaryotic Cells: Acid Fast Cell Wall

• Gram-positive bacteria with a waxy mycolic acid layer
• Stained pink using acid-fast stain
• Clinically important acid-fast bacteria include Mycobacterium tuberculosis and Mycobacterium leprae

External Structures of Prokaryotic Cells: Capsule

• Found in some species; outside the cell wall
• Cannot be stained; visualized by negative staining
• Glycocalyx (sugar coat) consists of polysaccharides or proteins
• Functions: water retention and biofilm formation

External Structures of Prokaryotic Cells: Flagella, Fimbriae

• Flagella: motility structures; presence and arrangement are species-specific
• Fimbriae: sticky appendages, often numerous, contribute to biofilm formation

External Structures of Prokaryotic Cells: Pilus/Pili

• Hollow tubes (made of protein)
• Longer than fimbriae, shorter than flagella
• Function: DNA transfer between bacterial cells (conjugation)

Endospores

• Found in some gram-positive genera of bacteria (Bacillus and Clostridium)
• Dormant stage under unfavorable conditions
• Resist sterilization
• Species-specific morphology (e.g. terminal, central, or lateral)

Commonly Used Stains

• Different stains used to visualize different bacterial components (e.g., Gram stain, acid-fast stain, endospore stain, flagella stain, capsule stain)

Lesson 2: Bacterial Metabolism

• Comprehensive overview of bacterial metabolic processes, including central pathways, respiration, and fermentation.

Metabolism

• Metabolism = catabolism + anabolism
• Ultimate function: growth and reproduction
• Catabolism: breaking down large molecules to release energy
• Anabolism: using energy to assemble small molecules into larger ones

Carbohydrate Catabolism

• Aerobic respiration (with oxygen)
• Anaerobic respiration (without oxygen)
• Fermentation (no electron transport chain)

Fermentation

• Incomplete oxidation of glucose (or other nutrients) in the absence of oxygen
• Produces organic acids, alcohol, and/or gases (e.g., lactic acid, ethanol).

Carbohydrate Catabolism: Aerobic vs Anaerobic

• Aerobic respiration requires oxygen, produces more ATP, and completely oxidizes glucose
• Fermentation does not require oxygen, produces less ATP, and incompletely oxidizes glucose

Fermentation Products

• Various products resultant from various types of fermentations.
• Different microbes create various commercial products.

Use of Microbial Metabolism in Medicine

• Identifying bacteria using metabolic tests that are quick, inexpensive, and simple.
• Different metabolic products help identify bacterial species/strains.

Lesson 3: Bacterial Growth and Nutrition

• Define and explain microbial growth (increase in cell number, not cell size)
• Growth rates vary with type of microbe
• Optimal conditions for growth in a species vary.

Microbial Growth

• Rapid, exponential growth under optimal conditions
• Generations (time to double) vary
• Factors affecting growth include:
• Chemicals: macro/micronutrients, oxygen, water
• Physical: temperature, pH, osmolarity

Microbial Growth and Nutrition: Chemical Requirements

• Key nutrients for bacterial growth include nitrogen (needed in higher concentrations than in humans; peptidoglycan in cell wall).
• Oxygen utilization varies
• Obligate/facultative aerobes.

Microbial Growth and Nutrition: Chemical Requirements (Cont'd)

• Macronutrients: needed in large quantities, provide carbon, energy, protein, and elements.
• Micronutrients: needed in smaller quantities, some are coenzymes in metabolic reactions.
• Growth factors: vitamins, vitamin-like molecules.

Microbial Growth and Nutrition: Physical Factors – Osmotic Pressure

• High osmotic pressure (high salt or sugar environment) pulls water out of cells, inhibiting bacterial growth.
• Water is a critical solvent for cellular functions.

Microbial Growth and Nutrition: Microbial Growth Curve

• Stages of bacterial growth include lag phase, log phase, stationary phase, and death phase.

Microbial Growth and Nutrition: Measuring Microbial Growth - Direct

• Direct microscopic cell counts and viable plate counts using serial dilutions, and membrane filtration.

Microbial Growth and Nutrition: Measuring Microbial Growth - Indirect

• Indirect methods like turbidity with a spectrophotometer measure microbial growth based on cloudiness.

Lesson 4: Bacterial Genetics

• Describe prokaryotic vs eukaryotic genomes
• Explain plasmids, which are small, extrachromosomal circular DNA molecules.

Prokaryotic versus Eukaryotic Genomes

• Prokaryotes have one circular chromosome in cytoplasm. Plasmids are present in some cells.
• Eukaryotes have multiple linear chromosomes in the nucleus and in cytoplasm.

What are Plasmids?

• Small circular DNA molecules that replicate independently of chromosomal DNA
• Contain genes that confer advantages, such a resistance to antibiotics or heavy metals. Other examples include fertility or virulence factors

The Central Dogma of Molecular Biology

• DNA replication, transcription (conversion to RNA), translation (conversion to proteins)
• Processes are similar in prokaryotes and eukaryotes

Genetic Recombination and Transfer

• Genetic recombination exchanges segments of DNA between molecules
• Recombinants are cells possessing new nucleotide sequences, which results from vertical or horizontal transfer.
• Vertical gene transfer is from one generation to another.
• Horizontal gene transfer is within generations, which contributes to genetic diversity.

Horizontal Gene Transfer in Prokaryotes

• Transformation: uptake of free DNA
• Transduction: transfer via bacteriophage
• Conjugation: direct transfer via a pilus

Gene Technology: PCR

• Polymerase chain reaction (PCR) amplifies DNA (or RNA)
• Areas of application include diagnostics, forensics, and biomedicine

Gene Technology: CRISPR

• Clustered, Regularly Interspaced Short Palindromic Repeats (CRISPR)
• CRISPR-associated proteins (Cas) are used to edit genomes
• Applications include medicine, biotechnology, agriculture, and epidemiology

Summary

• Course content summarizes differences between prokaryotic and eukaryotic cells; bacterial metabolism, growth, and genetics; and principles for two important gene technologies.

Test your knowledge on genetic recombination, polymerase chain reaction (PCR), and plasmids. This quiz covers key concepts in genetics, techniques used in molecular biology, and the significance of various factors in microorganisms. Challenge yourself and enhance your understanding of these fundamental topics.