Immunology, Human Microbiome, and Microbial Ecology

Fundamental Definitions in Immunology

• Hematopoiesis: The biological process responsible for the formation of all new blood cells, including both white blood cells (leukocytes) and red blood cells.
• Leukocytes: The scientific term for white blood cells. Note: Do not confuse this general category with lymphocytes, which are a specific subtype of leukocyte.
• Innate Immune System: The body's first-line defense system. It is characterized by responding first, having no immunological memory, and possessing a limited ability to "see" (recognize) microbes.
• Adaptive Immune System: The second line of defense. It possesses immunological memory and a vast, highly specific ability to "see" microbes through specialized receptors.
• Antigen: Any substance or molecule that triggers an immune response.
• Epitope: The specific chemical or physical part of an antigen that an immune cell receptor recognizes as foreign.

The Innate Immune System and Localized Response

• Physical Barriers: The skin serves as a primary barrier. Keratinocytes comprise the outer layer. Underneath this layer, macrophages and mast cells reside.
• Complement and Antimicrobial Proteins: These proteins can opsonize bacteria, marking them for destruction.
• Early Response Mechanism:   - Bacteria enter through a breach in the skin (keratinocyte layer).   - Macrophages recognize the bacteria via receptors and initiate phagocytosis.   - Mast Cells detect the pathogen and undergo degranulation, releasing chemical mediators.
• Chemical Mediators:   - Histamine: Released by mast cells; triggers vasodilation of blood vessels and increases vascular permeability.   - Cytokines: Released by macrophages and mast cells; signal other immune cells and induce the expression of adhesion molecules on blood vessel walls.
• Leukocyte Adhesion Cascade: The process by which leukocytes (such as neutrophils and monocytes) in the blood vessel adhere to the endothelium via adhesion molecules and migrate into the tissue to fight infection.
• Recruited Cells:   - Neutrophils: Early responders that arrive at the site of infection.   - Monocytes: Differentiate into macrophages upon entering the tissue.   - NK cells (Natural Killer cells): Release Interferon-gamma ($IFN-\gamma$) to activate macrophages and help kill infected or abnormal cells.

Mechanisms of Phagocytosis and Microbial Killing

• Phagocyte Receptors: Microbes bind to specific receptors including the Mannose-receptor, Mac-1 integrin, and Scavenger receptor.
• Ingestion: The phagocyte membrane "zips up" around the microbe, forming a phagosome.
• Phagolysosome Formation: The phagosome fuses with a lysosome, which contains degradative enzymes.
• Respiratory Burst (Oxidative Killing): Phagocytes produce reactive oxygen species (ROS) to destroy ingested bacteria.   - NADPH Oxidase: Converts $O_2$ and $NADPH$ into $O_2^-$ (superoxide) and $NADP^+$.   - Superoxide Dismutase (SOD): Converts $O_2^-$ into $H_2O_2$ (hydrogen peroxide).   - Myeloperoxidase: Converts $H_2O_2$ and chloride ions into Hypochlorous acid ($HCIO$), which is highly toxic to bacteria.
• Nitric Oxide (NO) Killing: The enzyme iNOS converts Arginine into Nitric Oxide ($NO$) in the presence of $O_2$.
• Extracellular Killing (Large Parasites): Helminths (worms) are too large for phagocytosis.   - Eosinophils bind to the parasite via IgE antibodies attached to Fc receptors.   - This triggers the release of granule proteins and reactive oxygen species onto the surface of the helminth.

Adaptive Immunity: Receptors and Somatic Recombination

• Innate vs. Adaptive Recognition:   - Innate cells use pattern recognition receptors (PRRs). There are only a few hundred varieties.   - Adaptive cells (lymphocytes) have a repertoire of over $10 \text{ trillion}$ ($10^{13}$) unique receptors.
• Somatic Recombination: A process unique to B and T cells to generate receptor diversity from a limited genome of approximately $25,000$ genes.   - Unlike normal gene expression where the gene remains constant and introns are spliced out of mRNA, somatic recombination physically rearranges the DNA gene segments.   - Gene Segments: Multiple segments (e.g., Segment 1, 2, 3, 4) are recombined into a new, unique gene configuration.   - This process is irreversible; the original germline version of the gene is lost in that specific cell.
• Clonal Expansion: Once a specific lymphocyte recognizes its target epitope, it undergoes rapid division to create a large population of effector cells.

Lymphatic System and Adaptive Cell Activation

• Antigen Presentation: Dendritic cells in the skin capture bacteria and travel through the Lymphatic System to a Lymph Node (LN).   - Antigens in the blood are processed by the spleen.
• T Cell Subsets:   - Cytotoxic T cells (CD8+): Target and kill infected cells, cancer cells, or cells containing intracellular pathogens by inducing apoptosis (programmed cell death).   - Helper T cells (CD4+): Coordinate the immune response by releasing cytokines to help other cells (B cells, macrophages, neutrophils) function more effectively.
• B Cells: Produce Antibodies.
• Immunological Memory:   - Primary Response: Occurs after first infection/exposure. Time lag is $5-10$ days. Peak response is smaller. Primarily uses IgM.   - Secondary Response: Occurs upon repeat infection. Time lag is $1-3$ days. Peak response is much larger and more rapid. Primarily uses IgG.   - Memory Cells: Long-lived B and T cells that persist in the bone marrow and lymphoid tissues.

Specialized T Helper Cell Functions and Antibody Isotypes

• Th1: Releases $IFN-\gamma$; activates macrophages to kill intracellular pathogens; linked to autoimmunity and chronic inflammation.
• Th2: Releases IL-4, IL-5, and IL-13; activates eosinophils and mast cells for defense against helminths; linked to allergies.
• Th17: Releases IL-17 and IL-22; recruits neutrophils for extracellular bacteria and fungi; linked to autoimmunity.
• Tfh (T follicular helper): Releases IL-21; helps B cells produce high-affinity antibodies.
• Antibody Isotypes:   - IgG: Opsonization, complement activation, neonatal immunity (crosses placenta), and ADCC by NK cells.   - IgM: Early response; potent activator of the classical complement pathway.   - IgA: Mucosal immunity; secreted into the gastrointestinal and respiratory tracts.   - IgE: Defense against helminths; triggers mast cell degranulation; involved in allergies.

The Human Microbiome and Development

• Colonization: Bodies provide warm, nutrient-rich environments with stable pH. Colonization begins at mucosal barriers (epithelial cells).
• Sterile Womb Hypothesis: Historically, the fetus was thought to be sterile. Recent DNA sequencing detected bacteria like Micrococcus luteus, Staphylococcus, and Lactobacillus in the fetal environment (placenta, amniotic fluid, meconium), suggesting early immune priming.
• Birth and Feeding Influence:   - Vaginal Birth: Infants acquire maternal vaginal and fecal microbiota (e.g., Bacteroides, Bifidobacterium).   - C-section: Infants often lack these, showing higher levels of pathobionts. Some studies use maternal FMT (Fecal Microbiota Transplant) to correct this.   - Breastfeeding: Promotes Bifidobacterium, which competes with pathogens like Clostridium difficile. Bottle-fed infants are $4\times$ as likely to harbor $C. difficile$.

Regional Microbiota

• Skin:   - Dominated by four phyla: Actinobacteria ($36-51\%$), Firmicutes ($24-34\%$), Proteobacteria ($11-16\%$), and Bacteroidetes ($6-9\%$).   - Moist sites: Staphylococcus and Corynebacterium.   - Oily (Sebaceous) sites: Cutibacterium (formerly Propionibacterium).   - Cutibacterium acnes: Common skin commensal; linked to acne; consumes sebum; often resistant to macrolides (erythromycin) but sensitive to tetracyclines and benzoyl peroxide.   - Skin Fungi: Malassezia accounts for $80\%$ of skin fungi.
• Oral Cavity:   - Nutrient-rich. Biofilms like dental plaque form on teeth in five stages: 1. Reversible attachment, 2. Irreversible attachment, 3. First maturation, 4. Second maturation (channel formation), 5. Dispersal.
• Gastrointestinal (GI) Tract:   - Stomach: pH is approximately $2$. Resident microbes include Lactobacillus and Helicobacter.   - Large Intestine: High density (up to $10^{13} \text{ cells/gram}$). Acts as a fermentation vessel. Dominant phyla: Bacteroidetes and Firmicutes.   - Obesity: Associated with a reduction in Bacteroidetes and an increase in Firmicutes and methanogenic Archaea.

Pathogens and Disease Correlations

• Helicobacter pylori: Spiral-shaped, Gram-negative bacterium. Causes gastritis and ulcers; classified as a Class I Carcinogen (gastric cancer). Uses cagA as a genetic risk factor for cancer.
• Fusobacterium nucleatum: Gram-negative obligate anaerobe. A pathobiont that promotes colorectal cancer, IBD, and and even linked to Alzheimer’s via the oral-brain axis. It uses the FadA adhesin to activate pro-inflammatory pathways.
• Dysbiosis: Imbalance in the microbiome linked to Type 2 diabetes, asthma, cancer, and IBD.
• Lactobacillus rhamnosus GG: Probiotic used to stabilize gut flora and reduce sensitivity to cow's milk (casein) allergies.

Microbial Ecology and Environment

• Sergei Winogradsky: Discovered chemolithotrophy through studies on Beggiatoa, which oxidizes $H_2S$ to sulfur granules for respiration.
• Winogradsky Column Zones:   - Oxic Zone: Top layer; contains Cyanobacteria (oxygenic photolithoautotrophs) and heterotrophic bacteria.   - Suboxic Zone: Iron-oxidizing bacteria and purple non-sulfur bacteria.   - Anoxic Zone: Bottom layer; contains purple and green sulfur bacteria and sulfate-reducing bacteria.
• Ecological Methods:   - Culture-Dependent: Enrichment approach (e.g., using N-free media to find $N_2$ fixers).   - Culture-Independent (-omics):     - Genomics: Cell potential.     - Transcriptomics: Cell effort/intent.     - Proteomics: Cell action.     - Metabolomics: Current cell state.

Symbiosis Examples

• Squid-Vibrio Symbiosis: Euprymna scolopes (squid) and Aliivibrio fischeri (bioluminescent bacteria).   - Diel Cycle: Squid expel $95\%$ of symbionts at dawn and repopulate them daily.   - Specificity: Only A. fischeri can colonize the light organ, overcoming host oxidative stressors (NO and hypophalous acid).
• Coral-Microbe Symbiosis: Coral and Symbiodinium (dinoflagellate).   - Coral Bleaching: Loss of symbionts due to temperature increases ($0.5-1.5\,^{\circ}C$ above optimal).   - Probiotic Hypothesis: Corals adapt by changing their microbiome to resist pathogens like Vibrio sholloi.
• Elysia chlorotica: A sea slug that performs kleptoplasty by stealing chloroplasts from the alga Vaucheria litorea. Evidence suggests horizontal gene transfer (HGT) of algal genes into slug chromosomes.

Bacterial Predation and Wildlife Microbiomes

• Predation Modes:   - Epibiotic: Vampirovibrio and Vampirococcus attach to the outside of prey.   - Endobiotic: Bdellovibrio (periplasm) and Daptobacter (cytoplasm) enter prey cells.   - Wolf Pack: Myxococcus xanthus swarms prey using social behavior and high enzyme density (fruiting bodies).
• Shark/Bat Studies:   - Sharks: Microbiomes on gills, teeth, skin, and cloacae are distinct from the surrounding seawater.   - Bats: Gut microbiomes are dominated by Proteobacteria and Firmicutes. Composition is heavily influenced by diet (insectivore vs. frugivore).