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Microbiology I Step I Review. Devin Gillaspie. First the Basics…. Gram Positive Organisms Stain purple/blue Cell wall + cell membrane Contains techoic acid – induces TNF and IL-1 Gram Negative Organisms Stain red Outer membrane + Periplasmic space + cytoplasmic membrane

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first the basics
First the Basics…
  • Gram Positive Organisms
    • Stain purple/blue
    • Cell wall + cell membrane
    • Contains techoic acid – induces TNF and IL-1
  • Gram Negative Organisms
    • Stain red
    • Outer membrane + Periplasmic space + cytoplasmic membrane
    • LPS in outer membrane
who doesn t gram stain
Who doesn’t Gram stain?
  • Treponema – too thin; use dark field microscopy or fluorescent Ab staining
  • Rickettsia, Legionellaand Chlamydia – intracellular. (Silver stain for Legionella)
  • Mycobacteria – high lipid content, use acid fast stain
  • Mycoplasma – no cell wall
  • Giemsa Stain – Borrelia, Plasmodium, trypanosomes, Chlamydia
  • PAS – Stains glycogen, mucopolysaccharides; to diagnose Whipple’s disease
  • Ziehl-Neelsen – Acid Fast organisms
  • India Ink – Cryptococcus neoformans (thick polysaccharide capsule)
  • Silver stain – Fungi and Legionella
special culture media
Special Culture Media
  • H. influenza – Chocolate agar with factor V and X
  • N. gonorrhoea – Thayer-Martin media
  • B. pertussis – Bordet-Gengou
  • C. diphtheriae – Telluritepalte, Loffler’s media
  • M. pneumoniae – Eaton’s agar
  • Lactose fermenters – MacConkey’s agar (pink colonies); E. coli turns black-blue on EMB agar
  • Legionella– Charcoal yeast extract agar with cystein
  • Fungi – Sabouraud’s agar
obligate aerobes
Obligate Aerobes
  • Use oxygen dependent system to make ATP
  • Examples:
    • Nocardia
    • Pseudamonasaeruginosa
    • Mycobacterium tuberculosis (reactivation TB usually in apices of lungs which have highest PO2)
    • Bacillus
obligate anaerobes
Obligate anaerobes
  • Lack catalase susceptible to superoxide dismutase
  • Foul smelling, difficult to culture, produce gas in tissue
  • Examples:
    • Clostridium
    • Bacteroides (GI tract normal flora)
    • Actinomyces
intracellular organisms
Intracellular Organisms
  • Obligate intracellular
    • Chalmydia, Rickettsia
  • Facultative intracellular
    • Salmonella, Neisseria, Brucella, Mycobacterium, Listeria, Fracncisella, Legionella
encapsulated bacteria
Encapsulated Bacteria
  • Capsules serve as anti-phagocytic virulence factor
  • Positive Quellung reaction – capsule swells when specific anticapsularantisera is added
  • Examples:
    • Klebsiellapneumoniae
    • Salmonella
    • Strep. pneumoniae
    • H. influenza type B
    • Neisseriameningitidis
  • Urease positive bugs
    • Proteus, Klebsiella, H.pylori, Ureaplasma
  • Pigment producing bacteria
    • Actinomycesisralii– yellow “sulfur” granules
    • S. aureus– yellow pigment
    • Pseudamonasaeruginosa– blue-green pigment
    • Serratiamarcescens– red pigment
bacterial virulence factors
Bacterial Virulence Factors
  • Protein A – Binds Fc region of Ig, preventing opsonization and phagocytosis (S. aureus)
  • IgA protease – cleaves IgA, to colonize respiratory mucosa (S. penumoniae, H. flu type B, and Neisseria)
  • M protein – Prevents phagocytosis
  • Certain gram positive rods
  • Form at end of stationary phase – no metabolic activity in spore form
  • Highly resistant to destruction by heat and chemicals – autoclave required to kill spores
exotoxins and you
Exotoxins and you
  • Super antigens
    • S. aureus– TSST-1 causes toxic shock syndrome
    • S. pyogenes – Scarlet fever-erythrogenic toxin
  • A-B toxins
    • C. diphtheriae – inactivates EF2; causespseudomembrane in throat
    • Vibriocholerae – inactivates Gs protein, stimulating adenylylcyclase and increase pumping Cl into gut and decreased Na absorption causing rice water diarrhea
    • E. coli (ETEC) – Heat-labile toxin (stimulates adenylatecyclase); Heat-stable toxin (stimulates Guanylatecyclase)  watery diarrhea
    • B. pertussis – increases cAMP by inhibiting Gαi whooping cough; inhibits chemokine receptor lymphocytosis

Other exotoxins

    • Clostridium perfringens – α toxin  gas gangrene
    • C. tetani – blocks release of inhibitory neurotransmitters GABA and glycine lockjaw
    • C. botulinum – Blocks release of Ach, causes anticholninergic symptoms, CNS paralysis
    • Bacillus anthracis – Edema factor, and adenylatecyclase
    • Shigella – Shiga toxin cleaves host cell rRNA (inactivates 60S ribosome), and enhances cytokine release  HUS
    • S. pyogenes – Streptolysin O, a hemolysin.
bacterial genetics
Bacterial Genetics
  • Transformation
    • Take up DNA from environment
    • Especially encapsulated bacteria
  • Conjugation
    • F+ plasmid contains genes required for conjugation (bacteria w/o plasmid are F-)
    • Plasmid is replicated and transferred through pilus to F- bacteria (no chromosomal DNA)
    • If F+ plasmid is incorporated into chromosomal DNA = Hfr
      • Replication of plasmid DNA may include some flanking genes
bacterial genetics1
Bacterial Genetics
  • Transduction
    • Generalized – LyticBacteriophage infects bacteria  cleavage of host DNA  production of viral proteins  some bacterial DNA may be packaged in viral capsid
    • Specialized – Lysogenic phage infects bacteria  viral DNA incorporated into bacterial chromosome  phage DNA excised with flanking genes
  • Transposition
    • Segment of DNA that “jumps” from one location to another
    • Plasmid to chromosome and vice versa
gram positives







S. viridans

S. pneumo

S. pyogenes(GAS)

S. agalactiae(GBS)

S. aureus

S. epidermidis

S. saprophyticus

staphylococcus sp staph aureus
Staphylococcus sp. – Staph aureus
  • Causes:
    • Inflammatory disease – skin infections, organ abscesses, pneumonia, etc.
    • Toxin-mediated disease – toxic shock syndrome (TSST-1), scalded skin syndrome (exfoliative toxin), rapid-onset food poisoning (enterotoxins)
    • MRSA – nosocomial and community acquired infections. Resistant to β-lactams due to altered penicillin binding protein
  • Virulence factors –
    • Protein A binds Fc-IgG, inhibiting complement fixation and phagocytosis
staphylococcus sp staph epidermidis
Staphylococcus sp. – Staph epidermidis
  • Infects prosthetic devices and IV catheters
  • Biofilm
  • Normal skin flora – contaminates blood cultures
  • Endocarditis in IVDA
streptococcus sp
Streptococcus sp.
  • Streptococcus pneumoniae– encapsulated, IgA protease
    • Most common causes of:
      • Meningitis
      • Otitis media (in kids)
      • Pneumonia
      • Sinusitis
    • Viridans group strep - α-hemolytic
      • Normal flora of oropharynx – dental carries
      • Subacute bacterial endocarditis
streptococcus sp1
Streptococcus sp.
  • Streptococcus pyogenes(GAS)
    • Causes
      • Pyogenic – pharyngitis, cellulitis, impetigo
      • Toxigenic – scarlet fever, toxic shock syndrome
      • Immunologic – rheumatic fever, acute glomerulonephritis
    • ASO titer detects recent S. pyogenesinfection
  • Streptococcus agalactiae (GBS)
    • β hemolytic, bacitracin resistance
    • Causes – pneumonia, meningitis, and sepsis mainly in babies
group d strep
Group D Strep
  • Enteroccocus
    • E. faecalisand E. faecium
      • Normal colonic flora
      • Penicillin G resistant
      • Cause UTIs
    • VRE - important cause of nosocomial infections
  • Streptococcus bovis
    • Colonizes gut
    • Causes – bacteremia and subacuteendocarditis
    • Colonizes 15% of colon cancer patients
  • Corynebacteriumdiphtheriae
  • Gram positive Rod
  • Exotoxin inhibits EF-2
  • Pseudomembranouspharyngitis with lymphadenopathy
  • Lab Dx – Gram - rods with metachromatic granules
  • Toxoid vaccine
clostridium sp
Clostridium sp.
  • Gram positive rods, spore forming, obligate anaerobes
  • C. tetani
    • Tetanospamin (tetanus exotoxin)  tetanus, spastic paralysis
  • C. botulinum
    • Heat-labile toxin inhibits ACh release at neuromuscular junction  botulism
    • In adults  ingestion of preformed toxin
    • In babies  ingestion of spores (floppy baby syndrome)
clostridium sp1
Clostridium sp.
  • C. perfringens
    • α-toxin myonecrosis (gas gangrene) and hemolysis
  • C. difficile
    • Cytotoxin kills enterocytespseudomembranous colitis
    • Often 2° to antibiotic use especially clindamycin or ampicillin
    • Dx: check for C. diff toxin in stool
    • Tx: metronidazole
  • Bacillus anthracis
  • Spore forming, gram positive rod, produces anthrax toxin
  • Capsule contains D-glutamate
  • Cutaneous anthrax – contact, black eschar formation; can progress to bacteremia, death
  • Pulmonary anthrax – inhalation of spores; flu-like symptoms rapidly progress to fever, pulmonary hemorrhage, mediastinitis, and shock.
listeria monocytogenes
  • Gram positive rod, facultative intracellular
  • “actin rockets” to move cell to cell
  • Tumbling motility
  • The only Gram + with endotoxin
  • Ingestion of unpasteurized milk/cheese and deli meats (avoid in pregnancy) or by vaginal transmission
  • Dz: amnionitis, septicemia, spontaneous abortion, granulomatosisinfantiseptica; neonatal meningitis; meningitis in immune compromised; mildgastroenteritis in healthy individuals
actinomyces vs nocardia
Actinomyces vs. Nocardia
  • Gram positive rods in long branching filaments resembling fungi
  • Actinomycesisraelii
    • Anaerobe, normal oral flora
    • Oral/facial abscesses
    • Drain through sinus tracts in skin – yellow sulfur granules
  • Nocardiaasteroides
    • Weakly acid fast aerobes
    • In soil
    • Pulmonary infection in immunocompromised patients
  • Mycobacterium tuberculosis
  • Primary TB – (non-immune host)
    • Ghon complex = Ghon focus (calcified scar in lower lobe) + lobar and perihilar node involvement
    • Healing with fibrosis (+ PPD), OR
    • Progressive lung disease (in HIV, etc) rarely death, OR
    • Severe bacteremia (miliary TB)  Death, OR
    • Pre-allergic lymphatic or hematogenous dissemination  dormant TB in organs  reactivation TB
  • Secondary TB – Reactivation TB
    • Fibrocaseouscavitary lesion in upper lobes
  • Extrapulmonary TB
    • CNS, Vertebral body (Potts disease), lymphadenitis, Renal, GI
  • Mycobacterium leprae
  • Acid-fast bacillus
  • Likes cool temps – infects skin and superficial nerves
  • US reservoir in armadillos
  • 2 forms of disease
    • Lepromatous – leonine facies = loss of eyebrows, nasal collapse, lumpy earlobes
    • Tuberculous – diffusely over skin, is communicable; lose sensation in fingers and toes (spontaneous amputations)
  • Treatment – long term oral dapsone; or rifampin + clofazimine + dapsone
  • 6 months to 2 years of antibiotics
other mycobacteria
Other Mycobacteria
  • M. kansasii
    • Pulmonary TB-like symptoms
  • M. avium-intracellulare
    • Often multidrug resistant
    • Disseminated disease in AIDS
gram negatives
Gram Negatives

H. flu






N. gonorrhoeae

N. meningitidis








E. Coli


neisseria sp
Neisseria sp.
  • Gram negative cocci – diplococci
  • Gonococci
    • No capsule, maltose non-fermenter, no vaccine, STD
    • Causes gonorrhea, septic arthritis, neonatal conjunctivitis (give erythromycin eye drops at birth), PID, and Fitz-Hugh-Curtis Syndrome
  • Meningococci
    • Capsule, maltose fermenter, vaccine, spread by respiratory and oral secretions
    • Causes meningococcemia and meningitis, Waterhouse Friderichsen syndrome
legionella pneumophila
  • Gram negative rod
  • Silver stain
  • Grows on charcoal yeast extract w/ cystein
  • Dx: antigens in urine
  • Transmission: aerosol from water source
  • Legionnaire’s disease = severe pneumonia and fever
  • Pontiac fever = mild flu-like syndrome
  • Treatment - erythromycin
pseudomonas aeruginosa
Pseudomonas aeruginosa
  • Gram – rod, aerobic, blue-green pigment, grapelike odor
  • Associated with wound and burn infections
  • Also causes:
    • Pneumonia (CF)
    • Sepsis
    • External otitis (swimmer’s ear)
    • UTI
    • Drug use (endocarditis)
    • Diabetic osteomyelitis
    • Hot tub folliculitis
    • Malignant otitisexterna (in DM)
  • Tx: Aminoglycoside + extended spectrum penicillin
  • Family that includes E. coli, Salmonella, Shigella, Klebsiella, Enterobacter, Serratia, and Proteus
  • All have O antigen (polysaccharide of endotoxin)
  • Capsular K antigen  virulence of bacteria
  • Flagella H antigen in motile species
  • Glucose fermenters
  • Oxidase negative
e coli
E. coli
  • EIEC
    • Invade intestinal mucosa
    • Shiga-like toxin
    • Dysentery
  • EHEC (O157:H7)
    • Shiga-like toxin
    • Dysentery; HUS
  • ETEC
    • Labile toxin
    • Traveler’s Diarrhea
  • EPEC
    • No toxin, adheres to apical surface, flattens villi, prevents absorption
    • Diarrhea
  • Intestinal flora - causes aspiration pneumonia in alcoholics and diabetics
    • Abscess in lung
  • Red current jelly sputum
  • NosocomialUTIs
salmonella vs shigella
Salmonella vs.Shigella
  • Both:
    • Non-lactose fermenters
    • Invade intestinal mucosa
    • Can cause bloody diarrhea
  • Salmonella
    • Flagella
    • Disseminate hematogenously
    • Produce hydrogen sulfide (H2S)
    • S. typhi only in humans, other species have animal reservoir
    • Antibiotics can prolong carrier state
  • Shigella
    • No flagella, move w/in cell using actin
    • More virulent
    • Bacterial dysentery
yersinia enterocolitica
  • Transmission
    • Pet feces
    • Contaminated milk
    • Pork
  • Outbreaks in day-care centers
  • Mimics Crohn’s or appendicitis (severe RLQ or LLQ pain)
helicobacter pylori
Helicobacter pylori
  • Gram – rod, urease positive (urease breath test)
  • Causes
    • Gastritis
    • 90% of duodenal ulcers
  • Risk factor for:
    • Peptic ulcers
    • Gastric adenocarcinoma
    • Lymphoma
  • Tx: Triple therapy (metronidazole, bismuth, and tetracycline or amoxicillin; or metronidazole, omeprazole, and clarithromycin)
leptospira interrogans
  • Question-mark shaped spirochete
  • In water contaminated with animal urine
  • Leptospirosis
    • Flu-like sx, fever, HA, abdominal pain, jaundice, photophobia with conjunctivitis
    • Mostly in tropics
  • Weil’s disease
    • Icterohemorrhagicleptospirosis
    • Severe form with jaundice and azotemia from liver and kidney dysfunction; fever, hemorrhage and anemia
lyme disease
Lyme disease
  • Borreliaburgdorferi
  • Transmitted by tick Ixodes
  • Prevalent in the northeastern US
  • 3 Stages
    • 1 = erythemachronicummigrans, flulike sx
    • 2 = weeks to months after tick bite, Bell’s Palsy and AV nodal block
    • 3 = chronic monoarthritis, and migratory polyarthritis
  • Mice are reservoir
  • Deer are part of tick life cycle
  • Tx: Doxycycline (stages 1+2), ceftriaxone (stage 3)
tremonemal diseases
Tremonemal diseases
  • Treponemapallidum causes Syphilis
  • Treponemapertenuecauses yaws
    • Infection of skin, bones and joints
    • Heals with keloid scars  severe limb deformities
    • After years, see facial deformities, especially affecting nasal cartilage
    • Disease of the tropics, not STD
    • VDRL positive
  • Treponemapallidum
  • 1° - painless chancre, localized dz
  • 2° - Disseminated disease with constitutional symptoms, maculopapular rash (palms and soles), condylomalata
  • 3° - Gummas (chronic granulomas), aortitis, neurosyphilis, Argyll Robertson pupil
  • Congenital Syphilis – Saber shins, CN VIII deafness, Hutchinson’s teeth, mulberry molars
vdrl vs fta abs
  • FTA-ABS is specific for Treponemes, turns positive earliest in the disease, and remains positive longest
  • VDRL detects non-specific Ab that reacts with beef cardiolipin. Can have false positives
zoonotic bacteria
Zoonotic bacteria
  • Bartonella spp.
    • Cat scratch disease; Bacillary angiomatosis in immune compromised (looks like Kaposi’s)
  • Borreliaburgdorferi
    • Lyme disease
  • Brucella spp.
    • Brucellosis/undulant fever; dairy products (unpasteurized) and contact with animals
  • Francicellatularensis
    • Tularemia; tick bites, rabbits, and deer
  • Yersiniapestis
    • Plague; flea bite; rodents, especially prairie dogs
  • Pasteurellamultocida
    • Cellulits; animal bites – cats and dogs
gardnerella vaginalis
  • Gram variable rod – pleomorphic
  • Causes
    • Bacterial Vaginosis (overgrowth of certain bact)
    • Gray vaginal discharge
    • Fishy smell
    • Non-painful
  • Transmission - Sexually
  • Treatment – metronidazole
  • Dx – Clue cells
  • Obligate intracellular
  • Transmitted by arthopods and cause HA, fever, rash
    • (except Coxiellaburnettii– transmitted by aerosols and causes pneumonia Q fever)
  • Weil-Felix reaction – patient’s serum is mixed with Proteus antigens, if present, anti-Rickettsial Antibodies will cross-react and agglutinate
  • Tx: tetracycline
rocky mountain spotted fever
Rocky Mountain Spotted Fever
  • Rickettsiarickettsii
  • Sx:
    • Rash on palms and soles, migrating to wrists and ankles, then trunk
    • Headache
    • Fever
  • Endemic to east coast
  • Obligate intracellular organisms (can’t make own ATP)
  • 2 forms
    • Elementary body – infection, enters cell by endocytosis
    • Reticulocyte body – replicates in cell
  • Chlamydia trachomatis
    • Reactive arthritis, conjuctivitis, nongonococcalurethritis, and PID
  • C. pneumonia and C. psittaci
    • Atypical pneumonia; aerosol transmission
    • Psittacosis – parrots; severe lung disease
  • Tx: erythromycin or tetracycline
chlamydia trachomatis serotypes
Chlamydia trachomatisserotypes
  • A, B and C = chronic infections; blindness in Africa
  • D – K = urethritis/PID, ectopic pregnancy, neonatal pneumonia, or neonatal conjunctivitis
  • L1, L2, and L3 – lymphogranulomavenerum (primary ulcers then inguinal lymphadenitis)
  • Atypical “walking” pneumonia
  • Symptoms
    • Insidious onset
    • HA
    • Non-productive cough
    • Diffuse interstitial infiltrate (CXR looks worse than pt)
  • Treatment – tetracycline or erythromycin

A 60 year-old woman comes to the physician because of fevers, chills, and a cough for the past 2 weeks. She lives at home with her husband, has no chronic medical conditions, and has not travelled recently. Her temperature is 38.2° C (100.8° F). Physical examination reveals altered breath sounds. Chest X-Ray is abnormal. Which of the following is the most likely causal organism?

A. Chlamydia pneumoniae

B.Haemophilus influenza


D. Staphylococcus aureus

E. Streptoccuspneumoniae


Correct answer is E. The most common cause of community acquired pneumonia is Sterptococcuspneumoniae. Choices A, B, and C are important causes of community-acquired pneumonia but are NOT the most frequent cause. Choice D (Staph aureus) is an important cause of CAP particularly in elderly patients with comorbidities, but is not the most common cause

Take home point: COMMON IS COMMON!!!


A three-week-old boy develops seizures, lethargy, and vomiting. Examination shows, a bulging fontanelle, and nuchal rigidity. Which of the following organisms should be suspected?

A. E. coli and Streptococcus agalactiae

B. H. influenza and Neisseriameningitidis

C. H. influenza and Streptococcus pneumoniae

D. Listeriamonocytogenes and Neisseriameningitidis

E. Staphylococcus aureus and Staphylococcus epidermidis


Correct answer is A. Microbial causes of meningitis varies by patient’s age and clinical setting. The most common causes of neonatal meningitis are Streptococcus agalactiae(Group B Strep), and E. coli.

H. Influenza was the most common cause of menigitis in ages 1-5 before 1990, but is much less common today after the introduction of an effective vaccine.

Neisseriamenigitidis is the most common in ages 5-40 (35% of children 5-15 yo, and 20% adults)

Streptococcus pneumoniae occurs in all ages (40% of adult cases, 15% of childhood cases) but is uncommon in neonates

Listeriamonocytogenes accounts for only 1% of bacterial menigits; seen in infants, elderly, and immunosuppressed.

Staphylococcus aureus is seen in post-surgical and post-traumatic meningitis

Staphylococcus epidermidis accounts for 75% of shunt-related meningitis


A 47 year-old man with a history of sickle cell disease has had numerous hospitalizations requiring the placement of IV lines. The patient has poor peripheral venous access, and a catheter is placed in the right subclavian vein. The patient subsequently develops right arm discomfort and swelling, and a temperature of 40.1°C with chills. Multiple blood cultures were taken and gram-positive cocci are isolated. The organism is catalase positive, and produces whitish colonies on mannitol salt agar. The colonies are gamma-hemolytic on blood sheep agar. Which of the following organisms is the most likely cause of this patient’s symptoms?

A. Enterococcusfaecalis

B. Staphylococcus aureus

C. Staphylococcus epidermidis

D. Streptococcus agalactiae

E. Streptococcus pyogenes


Correct answer is C. The patient has developed bacteremia. The description of the causative agent is consistent with a staphylococcal organism (catalase +, gram + cocci that grow on mannitol salt agar). The organism is therefore most likely Staphylococcus epidermidis because it was able to ferment mannitol and was not hemolytic (these characteristics rule out S. aureus).

Enteroccoccus and Streptococcus organisms are catalase negative. Both Strep species are Beta hemolytic and would not grow on mannitol salt agar, enterococcus is gamma hemolytic.

gram positives1







S. viridans

S. pneumo

S. pyogenes(GAS)

S. agalactiae(GBS)

S. aureus

S. epidermidis

S. saprophyticus


A 58 year old alcoholic male with multiple dental carries develops a pulmonary abscess and is treated with antibiotics. Several days later, he develops nausea, vomiting, abdominal pain, and voluminous green diarrhea. Which of the following antibiotics is most likely responsible for this patient’s symptoms?

A. Chloramphenicol

B. Clindamycin

C. Gentamycin

D. Metronidazole

E. Vancomycin


Correct answer is B. Any time you see the development of diarrhea in the same question stem with “treated with antibiotics,” you should immediately think of pseudomembranous colitis. This is caused by Clostridium difficile and typically occurs as a result of treatment with clindamycin or ampicillin. You would confirm you suspicion by sending stool sample to be tested for C. diff toxin.

Chloramphenicol is not frequently used in the US and the most common side effect is aplastic anemia.

The key side effects of gentamycin are ototoxicity and nephrotoxiciy

Metronidazole and vancomycin are used to treat pseudomembranous colitis


A 16 year-old boy with sickle cell disease is hospitalized for a severe infection. His spleen has autosplenectomized, and he has had other minor infections in the past. His symptoms include fever, chills, cough, and chest pain. Bacteria from the patient’s sputum are optochin-sensitive organisms with a positive Quellung reaction. Which of the following is the most likely pathogen?

A. E. coli

B. H. influenza

C. Klebsiella pneumonia

D. Neisseriagonorrheae

E. Streptococcus pneumoniae


Correct answer is E. The combination of optochin sensitivity and positive Quellung reaction is characteristic of a single organism, Sterptococcuspneumoniae.

H. influenza, Klebsiella, and N. meningitidisare all encapsulated organisms and have positive Quellung tests, but none are optochin sensitive.