BIO 1100 17 + 18

Microbiology

Study of microorganisms (microbes) and their effects on other living organisms

Benefits of Microbes

• Aiding digestion (gut microbiome)

• Immunity

• Bioremediation (e.g., Alcanivorax borkumensis metabolizing oil after spills)

• Food production (yogurt, bread, wine, beer)

Classification of Microbes

• Bacteria and Archaea: All are unicellular prokaryotes lacking a nucleus.

• Eukarya: Includes unicellular eukaryotes like yeast and amoebas.

• Viruses: Acellular entities that do not fall into any domain.

Biological Scale and Size of Microbes

• Viruses: 10 nm to 500 nm (measured in billionths of a meter).

• Bacteria: 1 μm to 10 μm (e.g., Streptococcus is ~1 μm; E. coli is ~2 μm).

• Eukaryotes: Approximately 10 μm to 100 μm (e.g., yeast is ~7 μm).

Structural Components of Viruses

• Acellular parasites consisting of genetic material—either DNA or RNA, but never both—surrounded by a protein coat called a capsid

• Nucleic Acid Core: Can be circular or linear, single-stranded or double-stranded. Most RNA viruses (e.g., SARS-CoV-2, influenza) are single-stranded.

• Capsid: A protein sheath composed of repeated protein subunits.

• Envelope: Found in many animal viruses; derived from the host cell's membrane and embedded with viral proteins.

• Enzymes: Some viruses carry specialized enzymes (e.g., reverse transcriptase in HIV).

Replication and Host Interaction of Viruses

• Inert outside of a host (virions) and must hijack host machinery, specifically ribosomes, for protein synthesis.

• Host Range: The specific types of organisms a virus can infect.

• Tissue Tropism: The tendency of a virus to infect specific tissues within a host based on surface receptors.

• Mutation Rates: Viral replication is often error-prone, leading to high mutation rates. This makes them difficult targets for the immune system and vaccine development.

Viral Lifecycle Cycles (Bacteriophages)

1. Lytic Cycle: The virus infects the cell, replicates immediately, and causes the cell to burst (lyse), releasing new phages.

2. Lysogenic Cycle: The viral DNA integrates into the host genome (prophage). It is passed to daughter cells during division. Under stress (e.g., UV damage), the virus exits the genome and enters the lytic cycle.

Case Studies: HIV (Human Immunodeficiency Virus)

• An RNA retrovirus that uses reverse transcriptase to convert RNA into DNA, which then integrates into host Helper T cells via the enzyme integrase.

• This destroys the immune system, leading to AIDS.

• Treatments include AZT (inhibits reverse transcriptase) and protease inhibitors.

Case Studies: SARS-CoV-2

• The virus causing COVID-19.

  • It uses spike proteins to bind to ACE2 receptors, primarily in lung tissue.

• mRNA Vaccines: Deliver mRNA coding for the spike protein.

  • Host ribosomes produce the protein, triggering an immune response and antibody production without using the actual virus.

Nucleoid Region of Bacteria

Area containing the single, circular bacterial chromosome.

Plasmids of Bacteria

Small, extra-chromosomal circles of DNA often carrying resistance genes.

Plasma Membrane of Bacteria

Regulates transport; may have invaginations to increase surface area.

Cell Wall of Bacteria

Made of peptidoglycan (sugar-peptide mix); provides shape and protection.

Capsule/Glycocalyx of Bacteria

A sugary polysaccharide layer that helps bacteria evade the immune system.

Flagella of Bacteria

Microtubule-based structures providing motility.

Pili of Bacteria

Protein tubes used for DNA transfer (conjugation)

Fimbriae of Bacteria

Hair-like bristles used for attachment to surfaces.

Gram Staining

Bacteria are classified by their reaction

• Gram-Positive: Thick peptidoglycan wall; stains purple; sensitive to penicillin.

• Gram-Negative: Thin peptidoglycan wall plus an outer membrane; stains pink; resistant to penicillin.

Common Shapes of Bacteria

• Coccus: Spherical (e.g., Streptococcus in chains; Staphylococcus in clusters).

• Bacillus: Rod-shaped.

• Spirillum/Spirochete: Spiral-shaped.

Growth Requirements and Conditions of Bacteria

• Oxygen: Obligate aerobes (require O2​), Facultative aerobes (prefer O2​), Obligate anaerobes (killed by O2​), and Facultative anaerobes.

• Temperature:

  • Psychrophiles: Cold-loving.

  • Mesophiles: Prefer moderate temps (~37°C, includes human pathogens like E. coli).

  • Thermophiles: Heat-loving (source of Taq polymerase for PCR).

• Endospores: Thick-walled, dormant structures (e.g., Anthrax) that survive boiling and harsh conditions.

Horizontal Gene Transfer (HGT)

Bacteria transfer genes within the same generation via:

1. Transformation: Taking up free DNA from the environment.

2. Transduction: DNA transfer mediated by a virus (bacteriophage).

3. Conjugation: Direct transfer of a plasmid through a pilus.

DNA Cloning and Recombinant DNA

Recombinant DNA (rDNA) contains DNA from two or more different sources.

• Cloning Vectors: Usually plasmids used to carry foreign genes into host cells.

• Restriction Enzymes: "Molecular scissors" that cut DNA at specific restriction sites.

• Sticky Ends: Overhanging single-stranded ends created by restriction enzymes that allow for base-pairing between different DNA fragments.

• DNA Ligase: The "molecular glue" that seals the sugar-phosphate backbone.

• Bacterial Transformation: Introducing rDNA into competent E. coli via heat shock or electric shock.

Critical Laboratory Techniques

• PCR (Polymerase Chain Reaction): Amplifies specific DNA sequences.

• Gel Electrophoresis: Separates DNA fragments by size. DNA is negatively charged and moves toward the positive electrode; smaller fragments travel faster through the porous agarose gel.

• DNA Sequencing: Determining the exact order of nucleotides (A, T, C, G).

• DNA Microarrays: "DNA chips" used to monitor the expression of thousands of genes simultaneously by hybridizing fluorescently labeled cDNA to known DNA sequences on the chip.

• cDNA (Complementary DNA): Made from mRNA using reverse transcriptase. It lacks introns, allowing eukaryotic genes to be expressed in bacteria.

Genome Editing: CRISPR/Cas9

• A system using a guide RNA to target specific DNA sequences and the Cas9 enzyme to cut the DNA.

• This allows for precise gene deletion or replacement (e.g., potentially fixing the sickle cell mutation).

Genetically Modified Organisms (GMOs)

Contain genes from other species.

Agricultural and Industrial GMOs

• Transgenic Bacteria: Used for bioremediation or producing human proteins like insulin (Humulin) and growth hormone.

• Bt Corn: Contains a bacterial gene that produces a toxin to kill caterpillars, reducing pesticide use.

• Golden Rice: Engineered to produce vitamin A to prevent blindness in malnourished populations.

Medical and Forensics Applications of GMOs

• Gene Pharming: Using transgenic farm animals (e.g., goats) to produce pharmaceuticals in their milk.

• Mouse Models/PDX: Injecting human cancer cells into mice (Patient-Derived Xenografts) to test drug efficacy.

• Knockout Mice: Deleting specific genes to study their function or model diseases like cystic fibrosis.

• DNA Profiling (Fingerprinting): Uses PCR to amplify Short Tandem Repeats (STRs)—non-coding regions where sequences repeat. The FBI uses 13 specific STR sites to distinguish individuals with near-certainty.

In Vitro Fertilization (IVF) and Screening

• Involves fertilizing eggs in a dish.

  • Blastocyst: A five-day-old embryo consisting of an outer trophoblast (becomes placenta) and an Inner Cell Mass (ICM).

  • Preimplantation Genetic Diagnosis (PGD): Testing a single cell from an IVF embryo using DNA probes to screen for genetic diseases before implantation.

Stem Cell Potency

Can self-renew or differentiate into specialized cells.

1. Totipotent: Can become any cell type plus the placenta (early zygote).

2. Pluripotent: Can become any cell in the body but not the placenta (ICM of blastocyst).

3. Multipotent: Limited to a specific category (e.g., adult bone marrow stem cells).

Advanced Cloning and Therapeutic Potential

• Somatic Cell Nuclear Transfer (SCNT): Moving a somatic cell nucleus into an enucleated egg to restart development. This was used to clone Dolly the sheep.

• Induced Pluripotent Stem Cells (iPS): Reprogramming adult differentiated cells (like skin) back into pluripotent stem cells using transcription factors. This avoids the ethical issues of using embryos and prevents transplant rejection.

Gene Therapy

Delivering healthy genes into a patient to treat a disorder.

• Ex vivo involves treating cells outside the body and returning them; in vivo involves delivering the gene directly (e.g., inhaled virus for cystic fibrosis)

Proteomics

• The study of the proteome (all proteins expressed by a species). T

• he proteome is larger than the genome due to alternative splicing and post-translational modifications

Bioinformatics

Using computer software and statistics to analyze biological data (A, T, C, G sequences) to find patterns and relationships to disease.