Prokaryotes, Viruses and Bacterial Structures

• These are study notes on bacteria, viruses, fungi, and parasites.

Bacteria

• Prokaryotes lack a nuclear membrane and membrane-bound organelles.
• Eukaryotes possess a nuclear membrane and membrane-bounded organelles.
• Viruses are acellular, completely parasitic particles composed of nucleic acid and protein, sometimes with lipids.

Prokaryote Characteristics

• Bacteria is a major group.
• Approximate size is 0.5-3.0µm.
• There is no nuclear membrane.
• The chromosome is a single, supercoiled circular DNA haploid genome.
• Mitochondria, Golgi bodies, and Endoplasmic reticulum are absent.
• Ribosomes sediment at 70S (50S + 30S).
• The cytoplasmic membrane does not contain sterols.
• The cell wall is a complex structure containing protein, lipids, and peptidoglycans.
• Reproduction is asexual (binary fission).
• Movement involves a simple flagellum, when present.
• Respiration occurs via the cytoplasmic membrane.

Eukaryote Characteristics

• Major groups are algae, fungi, protozoa, plants, and animals.
• Approximate size is >5µm.
• There is a classic nuclear membrane.
• Chromosomes are strands of DNA with a diploid genome.
• Mitochondria, Golgi bodies, and Endoplasmic reticulum are present.
• Ribosomes sediment at 80S (60S + 40S).
• The cytoplasmic membrane contains sterols.
• The cell wall is present for fungi; otherwise absent.
• Reproduction is sexual and asexual.
• Movement occurs via a complex flagellum, if present.
• Respiration occurs via mitochondria.

Ways to Classify Bacteria

• Bacteria shapes include coccus, bacillus, coccobacillus, fusiform bacillus, spirillum, vibrio, and spirochete.

Bacterial Flagellum Structure and Function

• Flagella, also known as "H Antigen," function in cell motility through the environment.
• It rotates 360 degrees (tumble/clockwise/unorganized and run/counterclockwise/organized).
• Flagella are recognized by TLR5 in innate immune response as a "true antigenic structure".
• There are 3 parts consisting of the filament (of flagellin), hook, and basal body.
• Monotrichous flagella have a single flagellum at one end.
• Lophotrichous flagella have small bunches emerging from the same site.
• Amphitrichous flagella are located at both ends.
• Peritrichous flagella are dispersed over the cell surface and are the slowest.

Chemotaxis in Motile Bacteria

• Chemotaxis involves movement of an organism in response to a chemical stimulus (gradient).

Bacterial vs. Periplasmic Flagellum

• Periplasmic Flagella are internal flagella, that are enclosed in the space between the outer sheath and cell wall peptidoglycan.
• They produce cell motility by contracting and imparting twisting or flexing motion.
• Periplasmic Flagella is important for virulence in bacteria that possess them (spirochetes).

Fimbriae Structure, Function, and Significance

• Fimbriae are fine, proteinaceous, hair-like bristles emerging from the cell surface.
• They are predominantly found in gram- bacteria.
• Function is in adhesion to other cells and surfaces, and can aid in immune evasion.

Pili Structure, Function, and Significance

• Pili are a rigid, tubular structure made of pilin protein, only found in gram- cells.
• Function is to join bacterial cells for partial DNA transfer (conjugation).
• Pili are important in the transfer of antibiotic resistance.

Bacterial Conjugation

• Conjugation is a process where one bacterium transfers genetic material to another through direct contact.
• One bacterium is the donor, and one is the recipient.
• The donor carries a DNA sequence called fertility factor (F-factor).
• The mechanism is step 1 (contact), step 2 (activation of DNA for transfer), step 3 (plasmid transfer), and step 4 (synthesis of functional plasmid).
• A Donor F+ cell transfers the F plasmid to F- cell through pili.

Glycocalyx Types

• Glycocalyx is a coating of molecules external to the cell wall, made of sugars and/or proteins, and composed mostly of polysaccharides.
• Slime Layer is a loosely organized and attached glycocalyx.
• Capsule is a highly organized, tightly attached glycocalyx (K antigen) made of gram+ and gram-.

Glycocalyx Function

• Glycocalyx protects cells from dehydration and nutrient loss.
• It inhibits phagocytosis by WBCs, contributing to pathogenicity, inhibit complement, and attachment (formation of biofilms).
• Glycocalyx is related to quorum sensing.

Biofilms

• Biofilms are found on solid substrates, and consist of many species of bacteria and archaea living as a community.
• Glycocalyx holds the cells together.
• Bacteria living in a biofilm can have significantly different properties from free-floating bacteria.
• Bacteria in biofilms are protected from the outside world (increased resistance to detergents, resistance to antibiotics, and host defenses).
• Biofilms are a major factor in quorum sensing.

Quorum Sensing

• Quorum Sensing is regulation of gene expression in response to fluctuations in cell-population density.
• Bacteria produce and release chemical signal molecules (autoinducers) that increase in concentration as a function of cell density

Cell Wall Types

• Gram+ have a cell membrane and a thick layer of peptidoglycan, lipotechoic acid, and techoic acid.
• Gram- have a cell membrane, a thin layer of peptidoglycan, and another outer membrane (3 layers).
• Gram - have more protection and selective of what goes through (porin proteins).
• Gram - are very drug resistant.
• Nontypical Cell Walls: some bacterial groups lack typical cell wall structure (Mycobacterium/Nocardia)

Gram+ Cell Walls

• Gram+ cell walls have lipid mycolic acid (cord factor).
• With Gram+ cell walls, there is pathogenicity and high degree of resistance to certain chemicals and dyes.
• Basis for acid-fast stain used for diagnosis of infections caused by these microorganisms.
• Most familiar species are M. tuberculosis and M. leprae

Mycoplasma

• Mycoplasma has no cell wall.
• The cell wall is stabilized by sterols incorporated from the host.
• Mycoplasma is pleomorphic, the smallest cells, and have a minimal genome.
• They are obligate parasites (mostly).
• Anti-cell wall drugs will not work; most familiar species are M. pneumoniae, M. genitalium, and Ureaplasma urealyticum

New Stuff

• Gut Flora aids in digestion of food, vitamin production, and immune stimulation.
• Gut flora also aides in colonization resistance or microbial antagonism (occupy attachment sites, modify pH, produce bacteriocins)
• Disruption of normal flora almost always leads to problems

Opportunistic Infections

• Opportunistic Infections cause disease when the habitat is changed.
• It may occur due to weakened immune system
• Trauma can introduce flora to a sterile site.
• Immunosuppression and chemotherapy can cause opportunistic Infection.
• Other Manipulations, by medical personnel, that enable normal flora to cause disease (surgery, catheters, antibiotics)

Virulence Factors

• Virulence is a quantitative measure of pathogenecity. Relative ability of a microorganism to cause disease
• 50% lethal dose (LD50) and 50% infectious dose (ID50) are forms of virulence.
• Highly virulent pathogens are paradoxically often less successful.
• Virulence Factors are traits that determine pathogenicity and virulence (capsules, toxins, adhesive fimbriae); often encoded on pathogenicity islands

Toxins

• Endotoxin is lipopolysaccharide (LPS) toxin that is not secreted; integral part of gram- cell wall (lipid A component).
• Exotoxin is a polypeptide toxin molecule that is secreted.
• It is produced by certain species of some gram+ and gram- bacteria.
• Exotoxins have strong specificity of target cell.
• Exotoxins are good antigens (can be used to induce antitoxins).
• Types include A-B toxins, cytotoxins, and superantigens.

Viruses

• Viruses/Virions are obligate/dependent intracellular parasites (unstable outside of host cell); cannot synthesize proteins without a host; genome can be DNA or RNA (single or double stranded); host machinery responsible for viral assembly; non-living
• Structure: has nucleic acid core (DNA/RNA, single/double strand), capsid (protects core and mediates viral absorption, composed of capsomeres), envelope (made of host plasma membrane, acquired during exocytosis, either naked or enveloped)
• dsDNA goes to the nucleus for transcription.
• ssDNA is same as above; recognized by polymerase
• (+)ssRNA: polymerase recognizes protein (but there will be some diff steps)
• (-)ssRNA: NEED TO PACK THEIR OWN POLYERMASE
• dsRNA: polymerase recognizes it (no problem)
• Size is generally smaller than cells (20-150 nm)
• Viral Symmetry: helical (gives rod-like appearance), icosahedral (nucleic acid surrounded by capsomeres arranged in icosahedrons; most are naked)
• Envelope: consists of lipid bilayers from host cell membrane and virally encoded glycoproteins; acquired during late stages of replication as virus undergoes budding; susceptible to denaturation, as desiccation and detergents can destroy
• Naked Virus: lacks bilayer lipid membrane; heat-resistant
• Enveloped Virus: enclosed within lipid membrane; sensitive to heat
• Viral Life Cycle: 1) Adsorption, 2) Penetration, 3) Uncoated, 4) Synthesis (replication and protein prod.), 5) Assembly, 6) Release

Viral Replication

• 1. Recognition of/attachment to a target host cell, 2) Internalization (penetration) and uncoating of virus, 3) Macromolecular synthesis (early phase, replication of viral genome, late phase), 4) Virus assembly, 5) Release of virions, 6) Viral maturation (in some viruses)
• DNA Virus Replication: 1) Early period (nonstructural proteins; transcription is initiated; only part of genome transcribed/translated; regulatory proteins, polymerases, matrix proteins); 2) Late period (progeny nucleic acid; late proteins, capsid protein, envelope, glycoproteins)
• dsDNA Replication: DNA polymerase enzymes copy both the + and – DNA strands producing dsDNA viral genome; then use mRNA polymerase enzymes (mRNA); then translation
• ssDNA Replication: Parvovirus; DNA polymerase copy + strand of genome producing dsDNA intermediate; DNA polymerase then copies – DNA strand into ss+ DNA genomes; RNA polymerase enzymes copy – DNA strand into + viral mRNA; + viral mRNA then translated
• +RNA Virus Replication: CORONAVIRUS; directly translated by cellular polyribosomes into large proteins; RNA polymerase, protease; endocytosis
• -RNA Virus Replication: Filovirus; need to be transcribed into +RNA strands before synthesized (doesn't have RNA-dependent RNA polymerase, so virus packs its own polymerases)
• Retrovirus Replication: reverse transcription, transport, integration, transcription, assembly and release
• Host Specificity: viruses can infect only certain species of hosts and only certain cells within that host; viral receptor must be found on host cell surface for virus to attach; some have broad range (rabies), some narrow (bacteriophages)
• Antigenic Shift: mix of 2 diff strains of virus in which new surface proteins (or combination of proteins) are expressed; "leads to new sub-type; a little more dramatic; can lead to pandemic"
• Virus A + Virus B → Virus C
• Antigenic Drift: mutations occur in proteins that bind to Abs (antigenic drift is one of main reasons why vaccine development is hard); "only have small mutations; easier to contain"
• Virus A → Virus B

Fungi

• Fungi have 2 forms (unicellular/yeast or multicellular/mold/hyphae); cell walls have chitin and glucan; cell membranes have ergosterol; Cryptococcus is only encapsulated fungus (highly diagnostic); mostly aerobic; dimorphic fungi
• Fungal Disease: skin infections (superficial, cutaneous, subcutaneous), pneumonias and general, disseminated mycoses (typically from pneumonias), opportunistic infections
• Superficial Skin Infections: cosmetic effects; easily treatment; Tinea versicolor (Malassezia furfur, "spaghetti and meatballs"); dimorphic, normal flora; humidity, poor hygiene contribute to pathology)
• Cutaneous Skin Infections: dermatophytes (Tineas, Ringworm); molds only (no yeast form); named for body site affected; generally restricted to keratinized layers of skin (keratinolytic); Microsporum, Trichophyton, Epidermophyton; topical treatment

KOH Wet Mount for Fungal Infections

• The mount includes a 10-20% KOH Wet Mount of skin scraping that dissolves human cells and leaves fungal cells.
• These can be stained in same prep with various fungal stains

Subcutaneous Skin Infections

• These infections require trauma to allow entry to deeper layers of skin.
• Sporothrix Schenckii ("rose-gardener's disease) that is dimorphic with cigar-shaped yeast form.
• Lesion at site of entry with travel along lymphatics
• Disseminate to visceral organs in immunocompromised patients

Pneumonias/General Infections

• These begin in the lungs and disseminate to other body sites (often involve granulomas).
• Respiratory exposure to spores or other fungal repro structures that become airborne (no person-to-person spread).
• Dimorphic fungi; geography and soil characteristics become much more important with these clinical presentations
• Histoplasmosis: MS and OH River valleys (moist soils with guano); "histo hides"; often found inside macrophage
• Blastomycosis: Eastern/Central US; "broad-based budding yeast"; disseminates to skin, mimics squamous cell carcinoma
• Coccidioidomycosis: SW US (arid soils are best); spherules filled with spores; dissemination to skin and bones; erythema nodosum
• Paracoccidioidomycosis: Latin America; "Captain's Wheel" or "Mariner's Wheel"; presents similarly to blastomycosis

Opportunistic Infections

• Pneumocystis Jiroveci: PCP; immunosuppression leads to severe disease
• Candida Albicans: superficial (oral, GU) or systemic (bloodstream) infections
• Aspergillus Fumigatus: NOT dimorphic (hyphae only); aspergillomas in existing cavities
• Cryptococcus Neoformans: NO dimorphic (yeast only); encapsulated; hematogenous spread from lungs to meninges
• Mucor/Rhizopus: NOT dimorphic (hyphae only); invasion of sinuses and brain in patients with predisposing conditions

Parasites

• Parasites are Protozoa and Helminths
• Protozoa are single-celled eukaryotes; trophozoite vs cyst forms; motility can be characteristic; can have multiple hosts (intermediate/asexual vs. definitive/sexual vs. dead-end/accidental hosts); can have unique methods of cell repro (schizogony, conjugation, etc.)
• Protozoan Disease can effect the GI, CNS, blood, other (cutaneous, systemic, STI)
• GI Infections are from fecal-oral contaminationi
  • Giardia: giardiasis; fatty, foul-smelling diarrhea
  • Entamoeba: amebiasis; dysentery; liver abscess
  • Cryptosporidium: severe diarrhea in immune patients; mild otherwise
  • CNS Infections
  • Toxoplasma: contaminated meat, cat feces; crosses placenta; reactivation in immune patients; brain abscess
  • Naegleria: warm freshwater habitate; enters via cribriform plate; rapidly fatal meningoencephalitis
  • African Sleeping Sickness: "Trypanosoma brucei"; tsetse fly transmission; fever, somnolence, coma
• Blood Infections: both of these highly diagnostic in blood smears; both highly geographic
• Plasmodium: malaria, transmitted by Anopheles; fevers and chills (cyclical), headache, anemia, splenomegaly
• Babesia: tick-borne (Ixodes tick); fever and hemolytic anemia; "Maltese Cross"
• Other Infections
• Chagas Disease: "Trypanosoma cruzi"; unilateral periorbital swelling; cardiomyopathy; predominantly South America; "kissing bug" vector
• Leishmaniasis: cutaneous and visceral (kala-azar forms); Sandfly vector
• Trichomonas: sexually transmitted (no cyst form); vaginitis with foul smelling discharge
• Helminths: parasitic worms and flatworms; major target is Gl tract, followed by skin and bloodstream; various modes of transmission including insects
  • Ingested: "You'll get sick if you EATT these"
  • Enterobius, Ascaris, Toxocara, Tichinella, Trichuris
  • Enterobius: pinworms; perianal itching in children; "scotch tape test"
  • Ascaris: intestinal roundworms; travel through lungs can cause resp symptoms
  • Toxocara: dog roundworm; visceral larva migrans (targets liver, eyes, CNS, heart)
  • Trichinella: pork tapeworm; larvae encyst in striated muscle
  • Trichuris: whipworm; heavy infestations cause anemia and rectal prolapse in children

Parasites cont…

• Cutaneous: "These get into your feet from the SANd"
  • Strongyloides, Ancylostoma, Necator
• Strongyloids: threadworm; larvae in soil penetrate skin; can cause pulmonary, Gl, cutaneous symptoms
  • Ancylostoma/Necator: hookworks (old world and new); larvae in soil penetrate skin; causue microcytic anemia in children; cutaneous larva migrans cused by dog and cat hookworms
• Bites: "Law LOW to avoid getting bitten"
  • Loa loa, Onchocerca volvulus, Wuchereria bancrofti
  • Loa Loa: "African Eye Worm"; deer flies and horse flies; skin swelling and invasion of sclera of eye
  • Onchocerca: black flies; river blindness and "lizard skin"
  • Wuchereria: mosquitoes; lymphatic filariasis (elephantiasis)

Explore prokaryotes, viruses, bacterial flagella, and innate immune receptors. Learn about lipotechoic acid, F-factors, biofilms and cell walls and structures of bacteria.