Microbes and Their Roles

Human Understanding of Microbes

• Old view: Microbes are pathogens that cause diseases.
• New view: Normal microbiota promotes health and may exclude pathogens.

Nomenclature

• Scientific names are either italicized or underlined.
• The genus name is capitalized, and the specific epithet is lowercase.
• Example: Saccharomyces cerevisiae

Microbes and Their Roles

• Most microbes are harmless, some are helpful, and very few are pathogenic.
• Microbes includes bacteria, archaea, fungi, protozoa, microscopic algae, and viruses.
• In ecosystems, cyanobacteria and algae function as primary producers.
• Decomposers are essential for biogeochemical cycles.
• Photosynthetic bacteria and methanogenic archaea impact the carbon cycle.
• Nitrogen-fixing bacteria like Rhizobium affect the nitrogen cycle by converting N₂ to ammonia in legume root nodules and heterocysts in cyanobacteria.
• Microorganisms utilizing sulfate as a final electron acceptor impact the sulfur cycle in anaerobic respiration, reducing sulfate to hydrogen sulfide
• Microbes are used in fermenting foods, industrial production, biomanufacturing, pest control using Bacillus thuringiensis, and bioremediation.
• Humans and microbes have mutualistic relationships, where humans provide microbes with attachment sites, nutrients, stable temperatures, etc.
• Microbes provide hosts with vitamin K and aid in food digestion.
• Microbial antagonism or competitive exclusion is when microbes inhibit pathogen colonization via competition for nutrients and attachment sites, or secretion of inhibitory substances.
• Training of the immune system is guided by microbes.
• The Hygiene Hypothesis states a lack of exposure to microbes and helminths can lead to an improperly trained immune system, potentially causing allergies, asthma, eczema, and autoimmune disorders.

Normal Microbiota

• Formerly called microflora, the resident microbiota remains a part of the normal microbiota throughout life
• Normally found on the skin, gut, upper respiratory tract, and the distal portion of the urethra.
• It mainly feeds on cellular wastes and dead cells without causing harm under normal conditions.

Transient Microbiota

• Transient microbiota remains for hours, days, or months before disappearing.
• It fails to persist due to competition, elimination by defense cells, or physical and chemical changes in the body, like handwashing.

Microscopy Conversions

• 1000 mm = 1 m
• 1000 µm = 1 mm
• 1000 nm = 1 µm

Microscopy

• Total magnification in compound light microscopy is calculated by multiplying the objective lens by the ocular lens.
• Simple lenses (like magnifying glasses) differ from compound lenses (ocular and objective).
• Light microscopy types are Brightfield, Darkfield, Phase-contrast, and Fluorescence.
• Resolution is the ability to distinguish two points.
• Contrast is the difference in illumination between the lightest and darkest parts of a specimen.
• Phase-contrast microscopy allows lighter and darker parts to appear without staining, due to the constructive or destructive interference of light waves; it can be used on living cells.
• Fluorescent substances absorb UV light and emit visible light.

Staining

• Staining uses dyes to increase contrast for observations using a microscope to counter the transparency of microscopic subjects under brightfield microscopy.
• Basic dyes, the chromophore is a cation, includes Crystal Violet, Safranin, and Methylene Blue.
• Acidic dyes, the chromophore is an anion, includes India ink and Congo red, and are often used for Negative Staining and Capsule Staining.

Bacterial Morphology and Arrangements

• Bacterial shapes include coccus (spherical), bacillus (rod-shaped), and spiral (spirillum, vibrio, spirochete).
• Bacterial cell arrangements are strepto-, staphylo-, and diplo-.

Functional Anatomy of Cells

• The three domains of organisms are bacteria, archaea, and eukarya.
• Chitin is found in the cell walls of fungi.
• Cellulose in plant cell walls.
• Peptidoglycan in bacteria.
• Pseudopeptidoglycan in archaea.
• Cell walls provide shape, mechanical protection, and prevent osmotic lysis (cytolysis).
• Cells without cell walls include Mycoplasma, protozoa, animal cells, protoplasts, and spheroplasts.

Prokaryotic vs. Eukaryotic Cells

• Key differences include cell size
• Presence of a nucleus and membrane-bound organelles
• Number of chromosomes
• Structure of chromosomes (circular or linear)
• Presence of core histone proteins

Bacterial Glycocalyx

• Composed mainly of polysaccharides and glycoproteins.
• Functions include helping with attachment to surfaces
• Resistance to dehydration
• Acting as a virulence factor by protecting against phagocytosis or camouflaging the cell
• Serving as a source of nutrients
• Forming biofilms on surfaces.
• Streptococcus mutans forms a capsule to create dental plaque.
• Capsules are virulence factors in Streptococcus pneumoniae strains, and are targets for antibacterial vaccines.

Fimbriae and Pili

• Fimbriae are finger-like projections that aid in attachment to surfaces.
• Pili are used for bacterial conjugation, transferring DNA, and twitching motility, shown in Neisseria gonorrhoeae.

Bacterial Flagella

• Consists of a basal body, hook, and filament (flagellin).
• Functions for rotary, propeller-like motility, powered by a proton gradient.
• Its basal body is similar to the ATP Synthase complex.
• Taxis (chemotaxis) involves attractants and repellents, with data-driven decisions in runs and tumbles.
• Flagellar arrangements can be amphitrichous, lophotrichous, monotrichous, and peritrichous.
• Axial Filaments or endoflagella are found only in spirochetes such as Treponema pallidum, Borrelia burgdorferi, and Leptospira interrogans.

Bacterial Cell Wall

• The bacterial cell wall structure contains peptidoglycan.
• Its location differs in Gram-positive and Gram-negative cells and has backbone units of NAG and NAM, with peptide chains connected to NAM forming cross bridges.
• Gram-positive bacteria have thick peptidoglycan layers with teichoic acid and no outer membrane.

Gram-Negative Bacteria

• Characterized by a thin peptidoglycan layer in the periplasmic space, between the plasma membrane and the outer membrane.
• Outer membrane components include porins, lipopolysaccharide, and Lipid A (endotoxin).
• Lipid A release indicates pathogenesis leading to septic and endotoxic shock, vasodilation, inflammation, disseminated intravascular coagulation (DIC), and fever.
• MAMPs and PAMPs are Pathogen Associated Molecular Patterns found on pathogens but not host cells, such as teichoic acid, flagellin, pilin, and Lipid A.

Antibiotics and Cell Wall Targets

• Beta-lactam antibiotics include penicillin and vancomycin.
• Lysozyme target the cell wall with specific mechanisms of action.
• Protoplasts are cells without a cell wall.
• Spheroplasts are gram-negative bacteria that have had their peptidoglycan wall mostly, but not entirely, removed

Gram Staining

• Gram-positive bacteria stain purple.
• Gram-negative bacteria stain pink/red.

Plasma Membrane

• Known as the cytoplasmic/cell membrane, it is a selective barrier that maintains homeostasis through transport systems in a fluid mosaic model.
• Bacterial membranes differ from eukaryotic membranes by lacking cholesterol, except for Mycoplasma.
• Contains integral and peripheral proteins.
• Polymyxin B disrupts lipid membranes.
• 70% alcohol and phenol/Lysol denature membrane proteins and dissolve lipids.
• Antiseptics are for external/topical use only.

Plasma Membrane Functions

• Passive transport includes diffusion, facilitated diffusion, osmosis, and aquaporins.
• Active transport includes active transport, group translocation, antiports, symports, and Na+/K+ pumps.
• Consequences of cells with or without cell walls in hypotonic, hypertonic, or isotonic solutions include turgid, cytolysis, plasmolysis, and crenation.

Bacterial Cytoplasm

• Includes cytosol, DNA, inclusions, endospores, ribosomes, and cytoskeleton.
• Chromosomes and plasmids are typically circular without ends.

Bacterial Ribosomes (70S)

• Translate mRNA information to assemble proteins
• These have large and small subunits composed of ribosomal RNA and proteins.
• They are targets of antibacterial drugs like Aminoglycosides, Tetracyclines, Chloramphenicol, and Macrolides, which do not harm typical eukaryotic ribosomes (80S).

Inclusions

• Storage granules or areas in the bacterial cytoplasm
• Metachromatic granules (volutin) store phosphate reserves.
• Polysaccharide granules and lipid inclusions store energy reserves.
• Sulfur granules store energy reserves.
• Carboxysomes contain ribulose 1,5-diphosphate carboxylase for CO₂ fixation.
• Gas vacuoles are protein-covered cylinders.
• Magnetosomes contain iron oxide.

Endospores

• These are tough, dormant structures in Gram-positive bacteria like Clostridium and Bacillus.
• They are highly resistant to dehydration, radiation, and toxic chemicals.
• They form due to unfavorable conditions such as desiccation, nutrient deprivation, or waste accumulation.
• The endospore coat is thick, keratinized, and contains DNA, RNA, and ribosomes.

Eukaryotic Cells

• Larger than bacterial cells and contain membrane-bound organelles such as the nucleus, rough endoplasmic reticulum, smooth ER, Golgi, mitochondria, chloroplasts, peroxisomes, and lysosomes.
• DNA is linear with telomere ends, wrapped around core histone proteins (chromatin), and located in the nucleus.
• Flagella are larger than bacterial flagella, undulating, based on microtubules and dynein, and use ATP.
• Cilia are numerous and shorter than flagella; the ciliary escalator is found in the human respiratory tract.

Endosymbiotic Hypothesis

• Suggests that eukaryotic mitochondria originated from bacteria.
• Chloroplasts may be derived from cyanobacteria.

Eukaryotic Organisms: Fungi

• Eukaryotes with cell walls typically made of chitin, glucan, or mannan.
• They can be unicellular (yeast), aseptate (coenocytic), or septate (cells divided by cell walls).
• Hyphae are thread-like filaments, while mycelium is a mass of hyphae.
• Dimorphism: Individual species may have yeast-like or mold-like morphology under different conditions.

Fungi Reproduction and Genetics

• Haploid cells have one copy of each chromosome.
• Fungi are mostly haploid, with a brief diploid stage before meiosis/sexual reproduction.
• Fungal spores are haploid and used for reproduction and dispersion.
• Asexual spores are clones, while sexual spores increase genetic variation.

Sexual Reproduction in Fungi

• Involves the fusion of haploid nuclei from two opposite mating strains in three phases:
• Plasmogamy: a haploid donor cell nucleus (+) penetrates the cytoplasm of a recipient cell (-).
• Karyogamy: (+) and (-) nuclei fuse to form a diploid zygote.
• Meiosis: a diploid nucleus produces haploid nuclei (sexual spores).

Saccharomyces cerevisiae

• Baker's/brewer's yeast used to make bread and wine.
• Well-studied model system in molecular biology.
• Mycoses are diseases caused by fungal pathogens.
• Tinea, Candida, and Pneumocystis are fungal genera to know.
• Candida can cause superinfections when the normal microbiota is removed by antibacterial drugs.

Lichens and Algae

• Lichens are a mutualistic combination of a photosynthetic microbe (alga or cyanobacterium) and fungus
• Alga produce and secrete carbohydrates with fungus providing a holdfast.
• Algae are photosynthetic eukaryotes with chloroplasts that aren't plants
• Alage lack roots, stems, and true leaves, are mostly aquatic, and produce most of the planet's O₂.
• Examples: brown algae (kelp, Sargasso), diatoms, dinoflagellates, red algae, and green algae.
• Red algae produces agar and carrageenan.
• Green algae are direct ancestors of land plants with cellulose cell walls, store carbohydrates as starch, and use chlorophylls a and b.
• Dinoflagellates are unicellular algae that may produce neurotoxins, causing paralytic shellfish poisoning

Protozoa

• Unicellular eukaryotes without cell walls that ingest food.
• Examples include amoeba, paramecium, and euglena.
• Trophozoites are a feeding and growing form.
• Some produce cysts for survival under adverse conditions.
• Protozoan parasites may be ingested via contaminated water, transmitted by arthropod vectors, or sexually transmitted.
• Giardia intestinalis features multiple flagella, no mitochondria, and causes oily diarrhea/gas upon ingestion from contaminated water.
• Trichomonas vaginalis is sexually transmitted without a cyst stage, causing itchy, smelly discharge.
• Leishmania is transmitted by sandflies and causes skin lesions.
• Trypanosoma cruzi causes Chagas' disease and is transmitted by kissing bugs.
• Trypanosoma brucei causes sleeping sickness and is transmitted by the tse-tse fly.

Ciliates, Apicomplexa, and Plasmodium

• Ciliates move by cilia in precise rows.
• Balantidium coli is the only human ciliate parasite, causing dysentery.
• Apicomplexa are nonmotile, obligate intracellular parasites with complex life cycles.
• Toxoplasma gondii is transmitted by cats, crosses the placenta, and causes fetal infections.
• Cryptosporidium is transmitted via feces/water, causing waterborne illness.
• Plasmodium causes malaria, reproduces sexually in Anopheles mosquito.
• Slime molds are eukaryotic decomposers with an amoeba-like stage, being cellular or plasmodial.

Parasites and Hosts

• Definitive hosts support the mature form of parasites.
• Intermediate hosts support the immature/non-reproductive forms.
• Helminths are parasitic worms lacking digestive systems but possess complex reproductive systems for producing thousands of eggs.
• Platyhelminthes (flatworms) include Trematodes (flukes) and Cestodes (tapeworms).
• Clonorchis sinensis is a Chinese liver fluke ingested from contaminated fish.
• Schistosoma penetrates skin from contaminated water.
• Cestodes (tapeworms) feature a scolex head and proglottids, that are body segments containing reproductive organs.
• Taenia solium is a pork tapeworm contracted by ingestion.
• Nematoda (roundworms) exhibit a cylindrical shape and a complete digestive system.
• Ascaris lumbricoides infect intestines; eggs are consumed by humans.
• Enterobius vermicularis or pinworm eggs are consumed by humans.
• Ancylostoma duodenale or hookworm larvae enter skin.
• Dirofilaria immitis larvae spread by mosquitos, causing heartworm.
• Trichinella spiralis causes trichinellosis, contracted from undercooked pork.

Vectors and Transmission

• Vectors spread pathogens without causing disease themselves.
• Mechanical transmission: pathogens don't reproduce in the vector
• Biological transmission: pathogens multiply in the vector.

Microbial Metabolism

• Metabolism is the sum of chemical reactions in an organism
• Catabolism breaks down complex molecules, providing energy and building blocks for anabolism (exergonic).
• Anabolism uses energy and building blocks to build complex molecules (endergonic).
• Metabolic Pathway – a sequence of enzymatically catalyzed chemical reactions; the products of one reaction serve as the substrates for the next
• Activation Energy – is the energy required to break old chemical bonds (in the reactants) to allow new chemical bonds to form (in the products)

Enzymes

• Protein biological catalysts that are specific for a chemical reaction and are not used up in that reaction.
• They lower amount of activation energy needed to break the old chemical bonds in the reactants and allow new chemical bonds to form in the products
• Apoenzymes are protein components.
• Cofactors are nonprotein components, coenzymes are organic cofactors and holoenzymes are apoenzyme plus cofactor.

Factors Influencing Enzyme Activity

• Influencing factors includes Temperature; denaturation happen as temperatures increase past optimal temperature
• pH – optimum varies, and can denature when pH levels reaches extreme environments
• Substrate Concentration increase leads to substrate level increasing
• Inhibitors slow or stop enzyme from functioning

Enzyme Inhibition

• Competitive inhibitors occupy the same active site as the substrate.
• Sulfonamides and Trimethoprim inhibit folic acid production by inhibiting the action of PABA.
• Noncompetitive inhibitors (allosteric) bind to an allosteric site, causing a conformational change that prevents substrate binding, which leads to reversible on/off switch for reactions.
• Feedback inhibition involves the final product inhibiting the first enzyme.
• Ribozymes catalyze chemical reactions like protein enzymes.

Oxidation-Reduction Reactions

• Redox reactions pair oxidation and reduction reactions.
• Oxidation is Losing electrons and protons.
• Reduction is Gaining electrons and protons.

Catabolic Metabolism and ATP

• Its purpose is to produce ATP from ADP and Phosphate.
• Cellular ATP uses (the most widely used of the nucleotide triphosphates equals energy currency of the cell).
• In coupled reactions, enzymes couple the removal of the last phosphate with unfavorable reactions.
• Polymerases (DNA synthesis, transcription) require Nucleotide TriPhosphates.
• Anabolic reactions (like Protein synthesis (on Ribosomes)) also require them.
• Some forms of active transport involves Transmembrane “pumps” (Na+/K+ ATPase).
• Phosphorylation (add inorganic phosphate) to other molecules.

Methods of Converting to ATP

• Substrate-level ATP Synthesis occurs in Glycolysis and the Krebs Cycle.
• ATP Synthase Complex is powered by a proton gradient and requires a membrane.

Carbohydrate Catabolism

• Glycolysis oxidizes glucose to pyruvic acid.
• The net output of glycolysis per glucose input: 2 ATP, 2 NADH, and 2 pyruvic acids.

Fermentation

• Glycolysis plus another step Releases energy from oxidation of organic molecules
• It does not use the Krebs cycle or ETC
• It uses an organic molecule as the final electron acceptor.
• Its purpose is to regenerate NAD+ to allow glycolysis to proceed without cellular respiration.

Fermentation Products

• Saccharomyces produces ethanol and carbon dioxide.
• Lactobacillus, Streptococcus, and animals produce lactic acid.
• Lactobacillales: Firmicutes that includes Lactobacillus, Streptococcus, and Enterococcus.
• Some are aerotolerant anaerobes, lack an electron transport chain, and are obligate fermenters.

Transition Rxn and Krebs Cycle

• In transition reactions, pyruvic acid from glycolysis is oxidized and decarboxylated, where NADH, carbon dioxide, and Acetyl-CoA are produced.
• Acetyl-CoA then enters the Krebs Cycle.
• Oxidation of Acetyl CoA produces NADH and FADH₂.
• NADH, FADH₂, and carbon dioxide are produced.
• GTP is produced by substrate-level synthesis.

Coenzymes and the ETC

• NADH and FADH₂ are derived from B-vitamins, carrying electrons and protons from carbohydrates to the ETC, then returning to their oxidized state.

Electron Transport Chain

• A series of carrier molecules exchanges electrons
• Energy released makes H+ gradient
• H+ produce ATP
• ETC transports protons from low gradient to high gradient
• High concentration of protons turns ATP
• Complex which puts phosphate onto ADP to make ATP
• Active Transport requires energy

Aerobic versus Anaerobic Respiration

• If the final electron acceptor = oxygen, then aerobic happens
• If final electron acceptor not oxygen, that produces less ATP than Aerobic
• If the final electron acceptor is carbonate, methane gas produces
• Methanogenic Archaea only produce methane

ATP Synthase Complex

• Located in membranes and powered by a proton gradient to join ADP and Phosphate to make ATP.
• Located in bacterial plasma membranes and eukaryotic cristae membranes within mitochondria.

Basal Body of Flagella

• This is also in membranes
• Both that and atp synth are powered by proton gradients
• Rotate in membranes

Cellular Respiration

• Active transport requires energy.
• Aerobic ETC pump more protons and receive more energy.
• Energy used to transport them actively comes from H+ electron carriers
• Also comes from from final receptor which creats water

Good Reasons for Membranes ETC rxn to happen

• Separates concentrations of protons and concentration
• Allows carriers electrons pass easily

Bacteria, Mitochondria and ATP

• Bacteria produce up to 38ATP
• Mitochondria products up to 36ATP
• Turning ATP, bacterial cell and also H+ symports and antiports make proton gradient good.
• H20 produced along gradient as well which powers ETC.

Catabolism

• Lipids broke down into Acetyl-CoA through digestive enzymes
• Energy produced from fats in calories

Deamination

• Removal of an amino acid makes ammonia and releases environment from microbial amino acids.
• Transamination is moving of groups molecule to molecule

Photosynthesis

• Cyanobacteria generate oxygen from photosynthesis
• Purple and green sulfur bacteria create sulfur with anoxygenic methods.

Anabolic Reactions

• Energy makes large blocks from smaller blocks
• Such as proteins to acids.