1. Basic Structures and Staining Flashcards
Bacterial Structures: Peptidoglycan. Function and Chemical compositions
Gives rigid support, protects against osmotic pressure; Sugar backbone w/ cross-linked peptide side chains.
Bacterial Structures: Cell wall / cell membrane (G+’s). Function and Chemical compositions
Major surface Ag; Peptidoglycan for support. Lipoteichoic acid induces TNF and IL-1.
Bacterial Structures: Outer membrane (G-). Function and Chemical compositions
Site of endotoxin (Lipopolysaccharide, LPS), major surface Ag; Lipid A induces TNF and IL-1 Polysaccharide is the Ag
Bacterial Structures: Plasma membrane. Function and Chemical compositions
Site of oxidative and transport enzymes; Lipoprotein bilayer
Bacterial Structures: Ribosome. Function and Chemical compositions.
Protein synthesis; 50S and 30S subunits
Bacterial Structures: Periplasm. Function and Chemical compositions
Space btw the cytoplasmic membrane and the peptidoglycan wall in G- bacteria; Contains many hydrolytic enzymes, including beta-lactamases
Bacterial Structures: Capsule Function and Chemical compositions (except which bacteria?)
Protects against phagocytosis; Polysaccharide (except in Bacillus anthracis , which contains D-glutamate)
Bacterial Structures: Pilus/fimbria. Function and Chemical compositions.
Mediate adherence of bacteria to cell surface, sex pilus forms attachment btw 2 bacteria during conjugation; Glycoprotein
Bacterial Structures: Flagellum. Function and Chemical compositions
Motility; Protein
Bacterial Structures: Spore. Function and Chemical compositions
Provides resistance to dehydration, heat, and chemicals; Keratin-like coat, dipiclonic acid
Bacterial Structures: Plasmid. Function and Chemical compositions
Contains a variety of genes for antibiotic resistance, enzymes, toxins; DNA
Bacterial Structures: Glycocalyx. Function and Chemical compositions
Mediates adherence to surfaces, especially foreign surfaces (e.g. indwelling catheters); Polysaccharide
Structures unique to gram (+) organisms
Teichoic acid Cell wall
Structures common to Gram +/- organisms
Flagellum, pilus, capsule, PDG, cytoplasmic membrane
Features unique to G(-) organisms
Endotoxin/LPS (outer membrane) Periplasmic space (location of many beta-lactamases)
What are the bacteria w/ unusual cell membranes/walls (2)?
Mycoplasma: contain sterols and have no cell wall , G+ rod
Mycobacteria: Contain mycolic acid. High lipid content.
Name the G(+) cocci (2)
Staphylococcus Streptococcus
Name the G(-) cocci (1)
Neisseria
Name the G(+) Rods (5)
My cobacterium (acid-fast) List eria B acillus C lostridium C orynebacterium
What happened when you were sending that email to Gram + Rod ? My List B ecame CC ‘d
Bacterial taxonomy of: Mycobacterium (acid fast), Listeria, Bacillus, Cornybacterium, Clostridium
G(+) Rods
Name the G(-) Rods (9)
H ave Y ou E ver L istened to GNR (Guns n’ Roses = Gram (-) Rods)? I P refer F un G reat B ig B oy B ands.
Haemophilus Yersinia Enterics (separate card) Legionella (silver stain) Bordatella Brucella Bartonella Francisella Pasteurella Gardnerella (gram variable)
Bacteria taxonomy of: Haemophilus, Yersinia, Enterics, Legionella, Pasteurella, Francisella, Gardnerella (gram variable), Bordetella, Brucella, Bardetella
G(-) Rods
Name the G+ Branching Filamentous bacteria (2)
Actinomyces Nocardia (weakly acid-fast)
Bacteria taxonomy: Actinomyces, Nocardia (weakly acid-fast)
Branching Filamentous bacteria (G+)
Name the Pleomorphic (G-) bacteria (2)
Rickettsiae
Chlamydiae (Giemsa stain)
Bacteria taxonomy: Rickettsiae, Chlamydiae (Giemsa stain)
Pleomorphic (G-) bacteria
Name the Spirochetes (G-) (3)
Leptospira Borrelia (Giemsa stain) Treponema
Bacterial taxonomy: Leptospira, Borrelia (Giemsa stain), Treponema
Spirochetes (G-)
Neither G+ nor G- (b/c no cell wall)
Mycoplasma
Bugs that won’t Gram stain (6)
These Rascals May Microscopically Lack Color Treponema (too thin to be visualized) Rickettsia (intracellular parasite) Mycobacteria (high-lipid-content cell wall requires acid-fast stain) Mycoplasma (no cell wall) Legionella pneumophilia (primarily intracellular) Chlamydia (intracellular parasite; lacks muramic acid cell wall)
Visualizing Treponemes
Darkfield microscopy and fluorescent Ab staining
Visualizing Mycobacteria
Acid-fast stain
Visualizing Legionella
Silver stain
Giemsa’s stain is used to visualize…?
use for: Borrelia Plasmodium Trypanosomes Chlamydia
PAS (periodic acid-Schiff) stain
stains glycogen, mucopolysaccharides*; Used to diagnose Whipple’s dz, Tropheryma whippelii (*PASs the sugar)
Ziehl-Neelsen stain (carbol fuchsin)
Use to stain Acid-fast bacteria
India ink
used to visualize Cryptococcus neoformans
Silver stain
used to visualize: fungi (e.g. pneumocystics), Legionella
Media/Special culture requirements for: H. influenzae
Chocolate agar w/ factors V (NAD) and X (hematin) (Media used for isolation)
Chocolate agar w/ factors V (NAD) and X (hematin)
H. influenzae
Media/Special culture requirements for: N. gonorrheae
Thayer-Martin media (or VPN)
Vancomycin (inh. G+), Polymyxin (inh G-), Nystatin (inh. fungi); “to connect to Neiserria, please use your vpN client”
Thayer-Martin media (VPN Vancomycin-Polymyxin-Nystatin)
N. gonorrheae
Media/Special culture requirements for: B. pertussis
Bordet-Gengou (potato) agar (Bordet for Bordetella)
Bordet-Gengou (potato) agar
B. pertussis
Media/Special culture requirements for: C. diphtheriae
Tellurite plate, Loffler’s media (don’t Laugh and double Dip and Tell)
Tellurite plate, Loffler’s media
C. diphtheriae
Media/Special culture requirements for: M. tuberculosis
Lowenstein-Jensen agar (Media used for isolation)
Lowenstein-Jensen agar
M. tuberculosis
Media/Special culture requirements for: M. pneumoniae
Eaton’s agar (Media used for isolation)
Eaton’s agar
M. pneumoniae
Media/Special culture requirements for: E. coli
Eosin-methylene Blue (EMB) agar (green colonies w/ metallic sheen) (Media used for isolation)
Eosin-methylene Blue (EMB) agar (green colonies w/ metallic sheen)
E. coli
Media/Special culture requirements for: Lactose-fermenting enterics
Pink colonies on MacConkey’s agar; fermentation produces acid, turning plate pink (Media used for isolation)
Pink colonies on MacConkey’s agar
Lactose-fermenting enterics
Media/Special culture requirements for: Legionella
Charcoal yeast extract agar buffered w/ increased iron and cysteine (Media used for isolation)
Charcoal yeast extract agar buffered w/ increased iron and cysteine
Legionella
Media/Special culture requirements for: Fungi
Sabouraud’s agar (Media used for isolation)
Sabouraud’s agar
Fungi
G(-) Enterics (rod-shaped) (13)
E. coli Shigella Salmonella Yersinia Klebsiella Proteus Enterobacter Serratia Vibrio Campylobacter Helicobacter Pseudomonas Bacteroides
Obligate aerobes
Use an O2-dependent system to generate ATP Examples: (Nagging Pests Must Breathe) Nocardia Pseudomonas Mycobacterium tuberculosis (w/ prediliction for apices of lungs, where PO2 is highest) Bacillus Also: to help remember Pseudomonas aeruginosa - AERuginosa is an AERobe
Pseudomonas aeruginosa O2 tolerance? Where does it show up?
P. AERuginosa is an AERobe seen in burn wounds, nosocomial pneumonia, and pneumonias in cystic fibrosis pts.
Obligate Anaerobes
Lack catalase and/or superoxide dismutase, and thus are susceptible to oxidative damage. Generally foul-smelling (short-chain FA’s), are difficult to culture, and produce gas in tissue (CO2 and H2). Normal flora in GI tract, pathogenic elsewhere. Examples: AminO2 glycosides are ineffective against anaerobes because the antibiotics require O2 to enter bacterial cell. (anaerobes Can’t Breathe Air) Clostridium Bacteroides Actinomyces
Aminoglycosides and anaerobes
AminO2 glycosides are ineffective against anaerobes b/c these ABX require O2 to enter into the bacterial cell.
Obligate intracellular bugs
Rickettsia, Chlamydia (Stay inside [cells] when it’s R eally C old) *Can’t make their own ATP
Facultatively intracellular bugs (8)
Some Nasty Bugs May Live FacultativeLY Salmonella Neisseria Brucella Mycobacterium Listeria Francisella Legionella Yersinia
Quellung Reaction
Postive quellung: If encapsulated bug is present, the capsule swells when specific anti-capsular antisera are added (Quellung = capsular swellung)
Capsule and vaccines
Capsule is Ag for vaccines (e.g. Pneumovax, H. influenzae B, meningococcal vaccines) Conjugation w/ protein increases immunogenicity and T-cell dependent response
Quellung (+) Bacteria
SHiN SKiS (Strep pneumo, H Influenzae type B, Neisseria, Salmonella, Klebsiella, group B Strep) Polysaccharide capsule is an antiphagocytic virulence factor for above bacteria. Asplenic individuals have decreased ability to opsonize encapsulated pathogens so are at risk of severe infections (give s. pneumo, h. influenza and N. mening vaccines)
Urease (+) bugs
Proteus, Ureaplasma, Nocardia, Cryptococcus, H.pylori, Klebsiella
“PUNCH the Kid who pee on you”
Pigment-producing bacteria (hint: what is yellow (2)? red? blue-green?)
Actinomyces israelii - yellow “sulfur” granules, composed of mass of filaments and formed in pus (Israel has yellow sand)
S. aureus (gold) - yellow pigment
Pseudomonas aeruginosa - blue-green (arugula is green)
Serratia marcescens - red (Mars is red)
Virulence factors
Promote evasion of host immune response
Important virulence factors in S. aureus
protein A: Binds Fc region of Ig, disrupts opsonization and phagocytosis.
Who makes Protein A? What is it for?
S. aureus. Virulence factor that binds Fc region of Ig to prevent opsonization and phagocytosis.
Who secretes IgA protease and what is it for?
S. pneumo, H. influenzae type B, Neisseria (same capsulated SHiN); virulence factor to colonize respiratory mucosa (mucosal immunity)
Important virulence factor for SHiN group (S. pneumoniae, H.influenzae type B, Neisseria)
IgA protease. Enzyme that cleaves IgA to colonize respiratory mucosa.
Group A streptococcus virulence factors
M protein: Helps prevent phagocytosis
(A.M. A for Staph aureus and M. for Group A Strep)
Who has M protein and what is it for?
Group A streptococcus virulence factors; helps prevent phagocytosis
Exotoxin vs. Endotoxin: Source
Certain species of some G(+) and G(-) organisms; Outer cell membrane of most G(-) bacteria and Listeria
Exotoxin vs. Endotoxin: Is it ecreted from cell?
Exotoxin is secreted from cell. Endotoxin is not.
Exotoxin vs. Endotoxin: Chemistry
Polypeptide; Lipopolysaccharide (structural part of the bacteria. Released when lysed)
Exotoxin vs. Endotoxin: Location of genes
Plasmid or bacteriophage; bacterial chromosome (Exo ; Endo)
Exotoxin vs. Endotoxin: Toxicity
High (fatal dose on the order of 1 microgram); Low (fatal dose on the order of hundreds of micrograms) (Exo ; Endo)
Exotoxin vs. Endotoxin: Clinical effects
Various; Fever, shock (Exo ; Endo)
Exotoxin vs. Endotoxin: Mode of action
Various; Includes TNF and IL-1 (Exo ; Endo)
Exotoxin vs. Endotoxin: Antigenicity
Induces high-titer Abs called antitoxins ; Poorly antigenic (Exo ; Endo)
Exotoxin vs. Endotoxin: Vaccines
Toxoids useful as vaccines; No toxoids formed and no vaccine available (Exo ; Endo)
Exotoxin vs. Endotoxin: Heat stability
Destroyed rapidly at 60C (except staphylococcal enterotoxin); Stable at 100C for 1 hour (Exo ; Endo)
Exotoxin vs. Endotoxin: Typical diseases
Tetanus, botulism, diphtheria; Meningococcemia, sepsis by G(-) rods (Exo ; Endo)
Superantigens
Bind directly to MHCII and T-cell receptor simultaneously, activating large numbers of T-cells to stimulate release of IFN-gamma and IL-2
Toxins: TSST-1
Organism, toxin function: S. aureus SuperAg that causes toxic shock syndrome (fever, rash, shock).
S. aureus SuperAg that causes toxic shock syndrome (fever, rash, shock).
TSST-1
Toxins: Exfoliatin
Organism, toxin function: S. aureus Causes staphylococcal scalded skin syndrome
S. aureus Causes staphylococcal scalded skin syndrome
Exfoliatin
Toxins: Enterotoxins
Organism, toxin function: S. aureus (and others) Cause food poisoning
S. aureus (and others) Cause food poisoning
Enterotoxins
Toxins: Scarlet fever-erythrogenic toxin
Organism, toxin function: S. pyogenes SuperAg that causes toxic shock-like syndrome
S. pyogenes SuperAg that causes toxic shock-like syndrome
Scarlet fever-erythrogenic toxin
ADP ribosylating exotoxins
Interfere w/ host cell function. B (binding) component binds to a receptor on the surface of the host cell, enabling endocytosis. A (active) component then attaches an ADP-ribosyl to a host cell protein (ADP ribosylation), altering protein function.
Toxins: Diphtheria exotoxin
Organism, toxin function: Corynebacterium diphtheriae ADP-ribosylating A-B exotoxin that inactivates EF-2 (similar to Psudomonas exotoxin A) Causes pharyngitis and pseudomembrane in throat.
Corynebacterium diphtheriae ADP-ribosylating A-B exotoxin that inactivates EF-2 (similar to Psudomonas exotoxin A) Causes pharyngitis and pseudomembrane in throat.
Diphtheria exotoxin
Toxins: Cholera toxin
Organism, toxin function: Vibrio Cholerae ADP ribosylation of G-protein stimulates adenylyl cyclase Increased pumping of Cl- into gut and decreased Na+ absorption Water moves into gut lumen Causes voluminous rice-water diarrhea
Vibrio Cholerae ADP ribosylation of G-protein stimulates adenylyl cyclase Increased pumping of Cl- into gut and decreased Na+ absorption Water moves into gut lumen Causes voluminous rice-water diarrhea
Cholera toxin
Toxins: Heat-labile toxin, Heat-stabile toxin
Organism, toxin function: E. coli ADP-ribosylating A-B toxins Heat-labile: stimulates Adenylyl cyclase Heat-stabile: stimulates Guanylate cyclase Both: cause watery diarrhea Labile like the A ir, Stable like the G round
Toxins: Pertussis toxin (PT)
Organism, toxin function: Bordetella pertussis ADP-ribosylating A-B toxin that increases cAMP by inhibiting G-alpha1 Causes whooping cough Inhibits chemokine receptor –> causes lymphocytosis
Toxins: alpha toxin
Organism, toxin function: Clostridium perfringens Causes gas gangrene Get double zone of hemolysis on blood agar
Clostridium perfringens Causes gas gangrene Get double zone of hemolysis on blood agar
alpha toxin
Toxins: Tetanus toxin (tetanospasmin)
Organism, toxin function: C. tetani Blocks the release of inhibitory neurotransmitters GABA and glycine Causes lockjaw
C. tetani Blocks the release of inhibitory neurotransmitters GABA and glycine Causes lockjaw
Tetanus toxin (tetanospasmin)
Toxins: Botulinum toxin (aka Botox)
Organism, toxin function: C. botulinum Blocks release of ACh Causes anticholinergic symptoms, CNS paralysis (especially cranial nerves) Spores found in canned food, honey (causes floppy baby)
C. botulinum Blocks release of ACh Causes anticholinergic symptoms, CNS paralysis (especially cranial nerves) Spores found in canned food, honey (causes floppy baby)
Botulinum toxin (aka Botox)
Toxins: Anthrax toxin
Organism, toxin function: Bacillus anthracis Edema factor, part of the toxin complex, is an adenylate cyclase
Toxins: Shiga toxin
Organism, toxin function: Shigella, and E. coli O157:H7 Cleaves host cell rRNA (inactivates 60S ribosome) Enhances cytokine release, causing HUS
Shigella, and E. coli O157:H7 Cleaves host cell rRNA (inactivates 60S ribosome) Enhances cytokine release, causing HUS
Shiga toxin
Toxins: Streptolysin O
Organism, toxin function: S. pyogenes a hemolysin. Ag for ASO Ab, which is used in Dx of rheumatic fever
S. pyogenes a hemolysin. Ag for ASO Ab, which is used in Dx of rheumatic fever
Streptolysin O
cAMP inducers (list)
Vibrio cholerae B. pertussis E. coli Bacillus anthracis* *Cholera, pertussis, and E. coli toxins act via ADP ribosylation to permanently activate endogenous adenylate cyclase (increasing cAMP), While anthrax edema factor is itself an adenylate cyclase
Vibrio Cholerae toxin and cAMP
Toxin permanently activates Gs, Causing rice-water diarrhea (Cholera turns the on on)
B. pertussis and cAMP
Pertussis toxin permanently disables Gi, causing whooping cough (Pertussis toxin turns the off off) *Pertussis toxin also promotes lymphocytosis by inhibiting chemokine receptors.
E. coli and cAMP
Heat-labile toxin stimulates adenylyl cyclase
Bacillus anthracis and cAMP
Anthrax toxin includes edema factor, a bacterial adenylate cyclase (increases cAMP)
Endotoxin
A Lipopolysaccharide found in the cell wall of G(-) bacteria N-dotoxin is an integral part of the gram-Negative cell wall *Endotoxin is heat stable
Endotoxin and Macrophages
Activates Macs: IL-1 causes fever TNF causes fever, hemorrhagic tissue necrosis NO causes hypotension (shock)
Endotoxin and complement
Activates the complement (alternative pathway): C3a causes hypotension, edema C5a causes PMN chemotaxis
Endotoxin and Hageman factor (factor XII)
Activates Hageman factor This activates coagulation cascade, causing DIC
Bacterial growth curve
Lag phase: metabolic activity w/o division Log phase: rapid cell division Stationary phase: nutrient depletion slows growth. Spore formation in some bacteria. Death phase: prolonged nutrient depletion and buildup of waste products leads to death.
Transformation
DNA taken up directly from environment by competent prokaryotic and eukaryotic cells. Any DNA can be used
F+ x F- Conjugation
F+ plasmid contains genes for conjugation process. Bacteria w/o this are termed F-. Plasmid is replicated and transferred through pilus from the F+ cell. Plasmid DNA only, no transfer of chromosomal genes.
Hfr x F- Conjugation
F+ plasmid can become incorporated into bacterial chromosomal DNA, now termed Hfr cell. Replication of incoporated plasmid DNA may include some flanking chromosomal DNA. Transfer of plasmid and chromosomal genes.
Generalized transduction
Lytic phage infects bacterium, leading to cleavage of bacterial DNA and synthesis of viral proteins. Parts of bacterial chromosomal DNA may become packaged in viral capsid. Phage infects another bacterium, transferring these genes.
Specialized transduction
Lysogenic phage infects bacterium; viral DNA incorporated into bacterial chromosome. When phage DNA is excised, flanking bacterial gnees may be excised w/ it. DNA is packaged into phage viral capsid and can infect another bacterium.
Transposition (transposons)
Segment of DNA can jump (excision and reincorporation) from one location to another, can transfer genes from plasmid to choromosome and vice versa. When excision occurs, may include some flanking chromosomal DNA, which can be incorporated into a plasmid and transferred to another bacterium.
5 bacterial toxins encoded in a lysogenic phage
ABCDE ShigA -like toxin B otulinum toxin (certain strains) C holera toxin D iphtheria toxin E rythrogenic toxin of Streptococcus pyogenes
Which bugs are catalase-positive?
S.aureus, Serratia, Pseudomonas, Actinomyces, Candida, E.coli (SSPACE for you CATs)
Degrades H2O2 before it can be converted to microbicidal products by enzyme myeloperoxidase. People with chronic granulomatous disease (NADPH oxidase deficiency) have recurrent infections with these microbes because they degrade the limited H2O2.
How do vaccines containing polysaccharide work?
A polysaccharide antigen alone would not be recognized and presented by T cells (only IgM antibodies produced). A protein is conjugated to polysaccharide antigen to promote T-cell activation and subsequent class switching A.
What are examples of polysaccharide vaccines?
Pneumovax (polysaccharide vaccine with no conjugated protein), H. influenzae type B (conjugated vaccine), Meningococcal vaccines
