BACILLUS ANTHRACIS

• “Anthrax” means “charcoal,” which describes appearance lesions it produces on the skin.
• Is the causative agent of anthrax.
• Resides in the soil, where its spores can persist for years; spores are also found on animals, especially on hides and wool.
• Non-motile
• In clinical specimens, spores are rarely seen; colonies are non-hemolytic.
• Current treatment recommendations for B. anthracis infection include ciprofloxacin or doxycycline with antitoxins
  – Amoxicillin is often used to treat cutaneous anthrax.
• In areas where B. anthracis is endemic, vaccination of humans and animals can help control disease.
• Infected animals should be incinerated, as spores can remain in the soil for many years.

Virulence factors:

• Unique polypeptide capsule comprising D-glutamic acid, which enables host immune evasion (recall most bacterial capsules comprise polysaccharides).
• Exotoxins: Edema toxin and Lethal toxin.
  – Toxins comprise A and B subunits:
  B subunit is Protective antigen (PA); this is the region of the toxin that binds with host cells.
  – A subunits are factors that combine with Protective antigen to form active toxins:
  Edema factor (EF) is an adenylate cyclase that increases intracellular cyclic adenosine monophosphate (cAMP), resulting in edema.
  Lethal factor (LF) is a protease that inactivates mitogen-activated protein kinase (MAPK) pathways, resulting in cell death.

Be aware that these factors are nontoxic on their own; they must combine with protective antigen to enter host cells and cause damage.

Infections:

• B. anthracis primarily infects non-human animals.
• Interaction with contaminated animal products can lead to three types of human infections:
  • Cutaneous, Inhalation, Gastrointestinal.

• Cutaneous anthrax
  – Most common form.
  – Presents as skin lesions with central necrotic eschar surrounded by edema. The pustule is initially painless, but infection can progress to produce systemic signs of edema, and bacteremia can be fatal.
  – Typically, cutaneous anthrax is due to exposure to contaminated soil or animal hides, hair, or wool; however, outbreaks have also been reported among injection drug users.

• Inhalation anthrax
  – Initially presents with nonspecific symptoms, including fever, non-productive cough, and myalgias.
  – However, as the spores travel from the lungs to the nearby lymph nodes, edema and mediastinal lymph node enlargement occurs; in chest X-rays, mediastinal widening is an important diagnostic cue. Respiratory failure can ensue, and, in about half the cases, meningeal symptoms occur.
  – Historically, inhalation anthrax in humans was associated with spore inhalation while working with animal products. However, weaponized anthrax has been used in bioterrorism; person-to-person transmission does not occur, because bacterial replication occurs in the lymph nodes.

• Gastrointestinal anthrax
  – Aka, ingestion anthrax, occurs upon consumption of contaminated meat.
  – Manifests in the upper and lower gastrointestinal tract:
  In the oral cavity, pharynx, and esophagus, anthrax produces lymphadenopathy, edema, sore throat, and can lead to sepsis. In some patients, pseudeomembranes form; these are grayish coverings that comprise fibrin, leukocytes, and other exudates.
  In the lower gastrointestinal tract, particularly the ileum and cecum, infection causes ulcerative lesions and edema, with nausea, vomiting, and bloody diarrhea.

BACILLUS CEREUS

• “Cereus” means “wax-like”; these bacteria produce flat, grayish white colonies on blood agar plates.
• Environmentally ubiquitous, motile, and beta-hemolytic.
• It causes two forms gastroenteritis, aka, food poisoning, upon consumption; symptoms depend on the type of toxin ingested.
  – Symptomatic treatment is given, infection resolves itself.

Emetic food poisoning

• An intoxication caused by pre-formed cereulide, which is heat stable.
• Spores survive initial cooking, and germinate if the food is not refrigerated; importantly, reheating does NOT kill the enterotoxin.
• It is commonly associated with rice and other starchy foods left at room-temperature for long periods of time.
• Toxin produces nausea, abdominal cramps, and vomiting; in rare complications, liver failure occurs when large quantities impair mitochondrial fatty acid metabolism.
• Quick: the incubation period < 6 hours after consumption, and illness duration < 24 hours.

Diarrheal food poisoning

• Infection by vegetative cells that produce heat labile enterotoxin in the intestine.
• Within the intestinal epithelial cells, the toxin increase the concentration of cyclic AMP.
• The bacteria tend to reside on meats and vegetables.
• They multiply in the gastrointestinal tract, where heat labile enterotoxin produces nausea, abdominal cramps, and watery diarrhea.
• Long duration: incubation period is > 6 hours; duration > 24 hours.

Other B. cereus infections:

• Ocular infections associated with trauma, surgery, or bacteremia.
  – At least three toxins are associated with ocular infection: necrotic toxin, cereolysin, and phospholipase C. Interactions of these toxins, and perhaps factors, leads to rapid infection progression and eye loss.
  – Clindamycin or vancomycin is urgent.
• Severe pneumonia* that mimicks inhalation anthrax.
• Intravenous catheter and CNS shunt infections, endocarditis, bacteremia, and meningitis.
  – Immunocompromised patients.

Spore-Producing Rods
Bacillus (aerobic) and Clostridium (anaerobic)

• Bacillus anthracis is the causative agent of anthrax, which takes on cutaneous, respiratory, and gastrointestinal forms.
  – Cutaneous anthrax is the most common and least dangerous form.
  Contracted from handling animal hides or other products that carry anthrax spores; intravenous drug use can also spread cutaneous anthrax.
  Lesions tend to be on exposed areas of the body, including the face, neck, forearms, and hands. Indicate that lesions comprise a red area of edema with a central necrotic eschar.
  – Inhalation anthrax is the most lethal form of anthrax.
  Upon inhalation of the spores, the mediastinal lymph nodes enlarge; pulmonary edema occurs (which produces a bat-wing appearance on x-ray), and, respiratory failure from collapsed lung can result without effective treatment.
  – Gastrointestinal anthrax is the result of consumption of contaminated meats.
  The effects can occur throughout the GI tract, and include lesions and ulcers; patients can experience vomiting and diarrhea.

• Bacillus cereus causes food poisoning, both emetic and diarrheal forms.
  – Infection is especially associated with contaminated rice, meats, and vegetables.

• Clostridium difficile causes colitis, which is characterized by inflammation of the colon and diarrhea.
  – A pseudomembranous covering can develop on the wall of the colon. The covering, which comprises inflamed mucosa with raised yellow plaques, looks like a furry rug in gross images.
  – Colitis is associated with antibiotic use, which can kill off the normal, harmless gastrointestinal bacteria and allow for Clostridium difficile overgrowth.

• Clostridium perfringens can cause soft tissue infections, including myocystis, cellulitis, and myonecrosis.
  – Myonecrosis, which is also called gas gangrene, is characterized by areas of necrosis that produce gaseous bubbles in the tissues.
  – Clostridium perfringens can also cause food poisoning.

• Clostridium tetani, which can be found in the soil, causes tetanus.
  – Tetanus is characterized by spastic paralysis; show that this can produce the characteristic extension of the head, neck, and spine, called opistotonos.

• Clostridium botulinum causes botulism; the bacteria can be found in canned goods, and is the reason that infants under one year of age should not be given honey (by childhood, most individuals are resistant to the pathogen in honey).
  – Botulism is characterized by descending flaccid paralysis and respiratory failure.

Non-Spore-Producing Rods

• Corynebacterium diphtheriae
  – The Diphtheria toxin gene is introduced to the bacteria via a bacteriophage.
  – Respiratory diphtheria is characterized by a sore throat with a firmly adhered pseudomembranous covering.
  Serious complications can arise, including airway obstruction, myocarditis, and cranial neuropathies.

• Listeria monocytogenes causes listeriosis.
  – Individuals become infected when they ingest contaminated foods, most often deli meats or cheeses (this is why pregnant women are advised to avoid these foods).
  – The effects of listeriosis depend on the health and age of the individual:
  In healthy, non-pregnant adults, infection can cause flu-like symptoms with watery diarrhea.
  Immunocompromised adults are at increased risk of meningitis.
  Neonates fare the worst, with the possibility of death, organ damage, or meningitis.

OVERVIEW

• Gram-negative cocci
• Neisseria are paired; they tend to look like coffee beans.
• Catalase and oxidase positive
  – Catalase is an enzyme that protects bacteria from oxidative damage.
• Pathogenic Neisseria with Type IV pili have twitching motility (be aware of intertextual variation regarding the motility of Neisseria).
• Invade host cells; in the image of a fluid sample, we see the tiny Neisseria gonorrhoeae inside the larger neutrophils.

Neisseria meningitidis

• Aka, meningococcus.
• Pathogenic serogroups include A, B, C, Y, and W-135.
  – Different serotypes are responsible for geographically distinct epidemics.
• Neisseria meningitidis can be grown on both chocolate agar and blood agar; under some conditions, it can be grown on nutrient agar.

Neisseria gonorrhoeae

• Aka, gonococcus.
• Require highly specific growth conditions, and can only be grown on chocolate agar.
  – Selective Thayer-Martin chocolate agar has antibiotics to exclude other bacterial types present in the sample.

Neisseria Virulence Factors

• Many virulence factors are part of the cell wall.
• Neisseria endotoxin comprises Lipooligosaccharide (LOS), with toxic lipid A region.
  – During infection, Neisseria can release “blebs” of their cell membranes with the endotoxin.
• Pili have multiple effects:
  – Attach to host cells, which is a key step in colonization.
  – Resist neutrophil bactericidal activities.
  – Type IV pili facilitate twitching motility.
• IgA1 protease degrades Immunoglobulin A, which enables the bacteria to reach the mucous membranes of the respiratory and genital tracts.
• Neisseria have special receptors that bind to host transferrin, lactoferrin, and hemoglobin to acquire iron, which is crucial for bacterial metabolism.
  – Notice that this mechanism is different from the siderophores of other bacteria.
• Opacity proteins (aka, Opa), which bind epithelial and phagocytic cells; they also engage in cell to cell signaling.
  – Strains with these proteins appear opaque in culture, hence their name.
• Reduction-modifiable proteins (Rmp) protect porin and lipooligosaccharide from bactericidal antibodies.
• Porin proteins (aka, Por) insert pores into the bacterial cells to allow for movement of nutrients and wastes; they also prevent phagolysosome fusion within neutrophils, which allows the bacteria to survive intracellularly and aid in the invasion of epithelial cells.
  – There are two porin genes:
  PorA is only active in Neisseria meningitidis
  PorB is active in both N. meningitidis and N. gonorrhoeae.

Opa, Por, and Rmp proteins are also referred to as outer membrane protein classes I-V, particularly in Neisseria meningitidis.

• Virulence factors associated with specific species.
  – Neisseria meningitidis:
  Outer polysaccharide capsule that allows it to resist phagocytosis. Several vaccines use the capsule as immunogen, however, this is not effective for Group B meningococcus because its capsule is not immunogenic.
  Factor H binding protein (FHBP) binds Factor H to inhibit complement factor C3b; thus, this protein inhibits opsonization and membrane attack complex formation. And, because it is present on Group B meningococci, it is used as a vaccine immunogen.
  – Neisseria gonorrhoeae:
  Some strains produce beta-lactamase (aka, penicillinase), which promotes penicillin resistance.

NEISSERIA MENINGITIDIS

• Transmitted in airborne respiratory droplets.
• Within the host, pili and other adhesins promote adherence to the pharynx; from here, N. meningitidis can enter the bloodstream.
• Individuals with complement deficiencies are especially at risk for meningococcal disease.
• Some people are chronic carriers that can spread infection to others.
• Meningococcal diseases are largely preventable with vaccines.
• Infections can be treated with Cephalosporin or Penicillin G.

Infections
Meningitis:
– Inflammation of the meninges that cover the brain and/or spinal cord.
– Neisseria meningitidis is the most common cause of meningitis in individuals ages 2-18; Streptococci pneumoniaeis also major cause of bacterial meningitis in this age group.
– Patients have fever, stick neck, headache, and increased neutrophils in their cerebrospinal fluid.

Meningococcemia:
– Can occur with or without meningitis.
– Characterized by fever, headache, nausea, vomiting, and chills.
– Skin lesions typically begin as small petechiae-like spots that can coalesce into larger hemorrhagic areas; vascular damage can lead to gangrene.
– Musculoskeletal pain, particularly of the joints, is common.
– In severe cases, adrenal gland hemorrhage (aka, Waterhouse-Friderichsen syndrome) and thrombosis with vessel and organ damage can be fatal.

Pneumonia, particularly in older adults.

NEISSERIA GONORRHOEAE

• No vaccine.
• Sexually transmitted
• Can cause local infections of the genitals, rectum, and pharynx; within the blood, it causes disseminated infection.
• Asymptomatic carriers are key reservoirs

Infections

Localized

• In males, the most common site of infection is the urethra; purulent discharge is common.
  – Complications include epididymitis, prostatitis, and development of periurethral abscesses.

• In females, the cervix is the primary site of infection.
  – Many women are asymptomatic; others experience cervical discharge, abdominopelvic pain, and bleeding.
  – Complications occur when infection ascends to the uterus and uterine tubes, resulting in pelvic inflammatory disease.
  – As a result of scarring, ectopic pregnancies and infertility can occur.

• Opthalmia neonatorum
  – Mothers can pass gonorrheal infection to offspring during passage through the birth canal. Affected neonates acquire opthalmia neonatorum, a form of conjunctivitis. Prophylactic use of erythromycin ointment shortly after birth can prevent this.

Systemic

• Gonococcemia
  – Produces fever, chills, and skin pustules on the extremities; septic arthritis can also occur.

OVERVIEW

• Streptococcal agalactiae is beta-hemolytic; it is also referred to as Group B streptococcus.
• Enterococci faecalis and Enterococci faecium, which were formerly classified as members of Group D streptococcus; they are alpha and gamma hemolytic.
• Viridans group streptococci, include several alpha-hemolytic subgroups.
• Streptococcus pneumoniae, aka, pneumococcus, which some authors categorize as members of the Viridans group.

DETAILS

Group B streptococcus

• Virulence factors that contribute to its pathogenicity:
  – Polysaccharide capsule inhibits phagocytosis.
  – Sialic acids in the capsule inhibit activation of the alternative complement pathway and neutrophilic activities, thus facilitating host immune evasion.

• Colonization and Infection:
  – Group B streptococcus commonly colonizes the colon and genitourinary tracts of women.
  – Group B streptococcus can be passed to the neonate, particularly if labor is prolonged; administration of antibiotics to infected mothers can help prevent bacterial transmission.
  – Early-onset diseases occur within the first 7 days after birth, and include bacteremia, pneumonia, and meningitis. This is typically the result of transmission during labor and birth.
  – Late-onset diseases occur between 1 week and 3 months of age, and include bacteremia with meningitis.
  – In women, Group B streptococcus is associated with post-partum endometritis, which is characterized by infiltration of plasma cells and inflammatory cells to the endometrium; it also causes wound and urinary tract infections.
  – In both men and women, Group B streptococcus is associated with bacteremia leading to pneumonia and infections of the bones, joints, skin, and soft tissues; immunocompromised, diabetic, and alcoholic patients are at higher risk of Group B streptococcal infections.
  – Penicillin G is the preferred antibiotic to treat Group B streptococcal infections.

Enterococci faecalis and Enterococci faecium

• Virulence factors:
  – Easily adhere to host surfaces via surface proteins and pili, which promote biofilm formation.
  – Are resistant to most antibiotics; thus, although not as virulent as some other Gram-positive cocci, established infections are difficult to eradicate.

• Infection:
  – Enterococci are particularly problematic in hospital settings, where they are associated with:
  Urinary tract infections, Bacteremia, Endocarditis, and Peritonitis (inflammation of the peritoneum).

Viridans group streptococci

Subgroups: S. mutans, S. salivarius, S. angionosus, S. mitis, and S. sanguinis.
– Fortunately, this group is mostly vulnerable to antibiotic treatments, although S. mitis is an important exception.
– Viridans group streptococci are associated with: Deep tissue abscesses (S. sanguinis); Bacteremia (S. mitis infection in cancer patients with neutropenia); Subacute endocarditis (S. mitis and S. salivarius); and, Dental caries (S. mutans).

Streptococcus pneumoniae

• Virulence factors:
  – Its polysaccharide capsule inhibits phagocytosis, and recombination of capsular genes facilitates antibiotic resistance.
  – Surface proteins bind respiratory tract epithelia, which facilitates respiratory infections.
  – IgA protease keeps the bacteria trapped within the mucin and inhibits clearance.
  – Pneumolysin lyses phagocytic cells and ciliated respiratory epithelium, which also prevents bacterial clearance.
  – Complement activation produces inflammation, and is triggered by pneumolysin, teichoic acid, and peptidoglycan fragments.
• Hydrogen peroxide damages host tissues.
• Phosphorylcholine facilitates entry into host cells, which protects bacteria from removal and promotes migration to deeper tissues.

• Infections:
  – Pneumonia, particularly in the lower lobes: In the sample, we can see that bacteria, red blood cells, and white blood cells infiltrate the alveolar tissue. Symptoms include chest pain (aka, pleurisy) with fever and chills with coughing that produces rust-colored sputum. Be aware that patients with dysfunctional or absent spleens can have severe forms of pneumococcal pneumonia.
  – Otitis media and sinusitis can occur after viral infections that obstruct sinus drainage and allow bacteria to proliferate.
  – Pneumococcal meningitis can occur when Streptococcus pneumoniae spreads to the central nervous system. This can happen as a result of bacteremia or when trauma produces a connection between the nasopharynx and subarachnoid space.

• Vaccine:
  Given the severity of Streptococcus pneumoniae infections, the CDC recommends a pneumococcal vaccine for all children younger than 2 years old, all adults 65 years or older, and for anyone at increased risk of infection.

• Streptococci grow in pairs and chains.
• They are facultative anaerobes, and are catalase-negative.

Streptococci classification:

• Hemolysis of blood agar:
  – Alpha-hemolytic species incompletely lyse red blood cells; this casts a greenish hue around the bacteria, which include Viridans group and S. pneumoniae.
  – Beta-hemolytic streptococci completely lyse red blood cells, which produces a clear outline around the bacteria; this group includes S. pyogenes and S. agalactiae.
  – Gamma-hemolytic streptococci do not lyse red blood cells; thus, there is no outline around the bacteria. This group includes the Enterococci.

• Lancefield Serologic groupings categorize the beta-hemolytic streptococci based upon the C-carbohydrates in their cell walls.

STREPTOCOCCUS PYOGENES

Streptococcus pyogenes is beta-hemolytic; it is also known as the Group A streptococci (GAS).

Virulence factors

• M-protein is one of the most important virulent factors of S. pyogenes.
  – It extends from the inner cytoplasmic membrane to block complement activation and phagocytosis. Strains with different M proteins are associated with different kinds of infections.
• Outer capsule comprises hyaluronic acid, which also prevents phagocytic clearance (in some strains).
• Lipoteichoic acid and F proteins bind fibronectin, thus adhering the bacteria to the host’s extracellular matrix.
• M and F proteins also facilitate epithelial cell invasion, which promotes persistent infection and migration to deeper tissues.
• M-like surface proteins bind fibronectin in the ECM; they also bind antibodies to block phagocytosis.
• Surface C5a peptidase inactivates C5a, which would otherwise attract neutrophils and other phagocytes.

• Toxins & Enzymes:
  – Streptococcal pyrogenic exotoxins (formerly called erythrogenic toxins) are superantigens that stimulate T cellsand macrophages to release IL-1, IL-2, and TNF.
  – Streptolysins S and O lyse blood cells.
  – Streptokinases A and B are enzymes that break up blood clots to promote bacterial spread.
  – DNases decrease pus viscosity, which also promotes bacterial spread, and aids in the evasion of Neutrophil Extracellular Traps (NET’s).

Diseases

• Streptococcal pharyngitis (“strep throat”) is characterized by throat and tonsil inflammation with swollen cervical lymph nodes, in addition to fever and headache. In the image, we can see whitish tonsillar exudate.
  – Antibiotic treatment, comprising penicillin and amoxicillin, is important to prevent later complications.
• Scarlet fever is a complication of streptococcal pharyngitis; within days of the initial infection, affected patients experience a rash that first appears on the chest, then the limbs.
  – Initially the tongue may be covered in a yellowish coating; this sheds, leaving behind a bright red “Strawberry tongue” surface.
• Pyoderma, aka, impetigo, is characterized by pustules that crust upon rupture; it is most common among children.
  – Pyoderma is also associated with other bacterial strains, including Staphylococcus aureus.
• Erysipelas (erythros = red; pella = skin) is characterized by raised red lesions, typically on the face but sometimes on the legs, and is accompanied by pain, fever, and chills.
  – It is most common in young children or older adults.
• Cellulitis involves the skin and underlying tissues; affected areas are red, swollen, and warm to the touch. It is usually painful and can spread within the deep tissues.
  – Be aware that Staphylococcus aureus is also associated with cellulitis; determination of the causative agent is an important part of effective treatment.
• Necrotizing fasciitis, aka, streptococcal gangrene. In our image, the skin is significantly damaged; in more severe cases, infection spreads to deeper tissues, where it can cause multi-organ failure and death. Diabetes is thought to be a risk factor for necrotizing fasciitis.
  – This disease lends S. pyogenes its nickname: “the flesh-eating bacteria.”
• Toxic Shock Syndrome via production of specific pyogenic exotoxins. Patients experience fever, chills, vomiting and diarrhea, and, as with Staphylococcus aureus-induced toxic shock syndrome, the disease can affect multiple organ systems.
  – Additionally, streptococcal toxic shock syndrome is associated with bacteremia and necrotizing fasciitis.
  – Treatment includes administration of intravenous penicillin and clindamycin; intravenous fluids and/or vasopressors can correct hypotension, and surgical removal of necrotic tissue may be required.
  • Review Shock
• Immune-mediated diseases that can occur after Streptococcal pyogenes infection:
  – Acute glomerulonephritis, which is characterized by deposition of antigen-antibody complexes in the glomerular basement membrane and infiltration of neutrophils. Inflammation leads to edema, hypertension, hematuria, and proteinuria.
  – Rheumatic fever and heart disease can occur after streptococcal pharyngitis; inflammation cardiac tissues can damage the cardiac valves and impede blood flow. Other manifestations include inflammation of the joints and blood vessels. Patients with pharyngitis should be given antibiotics within the first 10 days of infection to prevent later development of rheumatic fever.

OVERVIEW

• Staphylococcus grow in grape-like clusters.
• They are catalase-positive.
  – Catalase is an enzyme that converts hydrogen peroxide to water and oxygen; this allows the bacteria to resist oxidative stress.
• Staphylococci are non-motile and do not form spores.
• Staphylococcus aureus is the most virulent strain of staphylococcus and is a leading cause of infectious disease.
  – As we can see in the petri dish, carotenoid pigments give S. aureus a distinctive golden color.

S. AUREUS

Virulence factors

• Capsule of S. aureus inhibits phagocytosis; additionally, it disrupts chemotaxis and mononuclear cell proliferation.
• Teichoic acids, which are anchored to the peptidoglycan of the cell wall, bind S. aureus to fibronectin of the host extracellular matrix.
• Lipoteichoic acid and the peptidoglycan layer have endotoxin-like effects: they trigger macrophage release of IL-1 and Tumor Necrosis Factor, which induce hypotension and cause septic shock.
• Cell wall Protein A binds antibodies to block complementactivation and inhibit phagocytosis.
• Biofilm firmly adheres bacterial colonies and debris to host tissues. The biofilm reinforces adhesion to the host and shields the bacteria from immune cells and antibiotics.
• Cytotoxins (alpha, beta, delta, gamma, and Panton-Valentine leukocidin) that lyse red and white blood cells.
• Exfoliative toxins (A & B) are proteases that destroy the stratum granulosum of the epidermis.
• Enterotoxins stimulate T-cell and macrophage release of cytokines and trigger Mast cell degranulation, which results in peristalsis and vomiting.
• Toxic shock syndrome toxin -1 stimulates T cell proliferation and T cell and macrophage release of IL1, IL-2, and TNF, which causes blood vessel leakage.

Exfoliative toxin A, Enterotoxin, and TSS-1 are superantigens that induce massive immune responses that cause significant damage to the host.

• Coagulase converts fibrinogen to fibrin; by promoting clot formation and clumping, it is thought that S. aureus protects itself from host defenses.
• Fibrinolysin, aka, staphylokinase, has the opposite effects: it dissolves fibrin clots, potentially allowing S. aureus to spread to new niches within the host.
• Hyaluronidase degrades hyaluronic acids, which are present in host extracellular matrix.
• Lipases free fatty acids; it is thought that lipases inhibit host granulocytes, inactivate bactericidal lipids, and promote biofilm formation.
• Nucleases hydrolyze DNA and aid in bacterial evasion of Neutrophil Extracellular Traps (NETs).

Diseases

• Purulent skin infections include the following:
  – Impetigo tends to occur on the limbs and face of children; it is characterized by flat, reddened areas with pustules that crust upon rupture. Be aware that Group A streptococci also cause some forms of impetigo.
  – Folliculitis, as its name suggests, is infection of the hair follicle; “styes” are infections of eyelash follicles.
  – Furuncles, aka, boils, are larger, raised pus-filled nodules that can be quite painful; surgical drainage is sometimes necessary.
  – Carbuncles are furuncles that coalesce and affect the deeper subcutaneous tissues; bacteremia leads to chills and fever.
  – Wound infections can also be caused by S. aureus,particularly in patients with compromised immune systems.

• S. aureus bacteremia:
  – Acute endocarditis occurs when bacteria and cellular debris accumulate in vegetations and damage the cardiac valves. Blood flow can be significantly impaired, and vegetations that break free can embolize.
  – In the lungs, S. aureus can cause pneumonia with infiltrates and consolidation or abscesses caused by cell-damaging toxins and enzymes. Some patients go on to develop empyema, which is the accumulation of pus in the pleural cavity.
  –  Osteomyelitis occurs when S. aureus infects the bones; early onset is characterized by pain and fever. In children, infection involves the metaphyseal area of long bones. In adults, infection tends to occur in the vertebral bodies. In subacute osteomyelitis, localized infection within the bone can produce Brodie’s abscesses.
  – Septic arthritis typically affects the large joints.

• Toxin-mediated diseases:
  – Scalded skin syndrome, aka, Ritter’s disease, is caused by exfoliative toxins, and primarily affects newborns and young children. Disease onset is abrupt, and begins with perioral inflammation followed by superficial cutaneous blistering, then epithelial desquamation. Antibodies appear within 7-10 days, scarring is unusual, and mortality rate is low.
  – Food poisoning is caused by ingestion of enterotoxins, which produces nausea, vomiting, and diarrhea.
  – Toxic shock syndrome toxin -1 penetrates mucosal barriers and induces fever, hypotension and shock, and rash. Because it travels in the bloodstream, the toxin causes damage to multiple organ systems.

COAGULASE-NEGATIVE STAPHYLOCOCCAL STRAINS (CONS)

The following are often associated with infections of prosthetic joints and valves as well as catheters and shunts:

• Staphylococcus epidermidis
• Staphylococcus saprophytic is associated with urinary tract infections in sexually active young women.
• Staphylococcus lugdunensis is particularly associated with native valve endocarditis.
• Staphylococcus haemolyticus

• Gram-positive cocci can first be categorized as catalase positive or catalase negative.
  — Catalase-positive cocci include species of Staphylococcus.
  — Catalase-negative cocci include species of Streptococcusand Enterococcus.

Catalase-positive cocci: Staphylococcus

• Species of Staphylococcus can be categorized based on the presence of coagulase, which is a bacterial enzyme that induces blood or plasma coagulation:
  — The coagulase-positive group comprises Staphylococcus aureus.
  — Coagulase-negative species include Staphylococcus epidermidis and Staphylococcus saphrophyticus.

Staphylococcus aureus

• Named for its golden color.
• Some strains are resistant to the antibiotic Methicillin (these strains are called MRSA); infections caused by these strains are difficult to treat.
• Inflammatory Conditions caused by S. aureus
  — Skin infections include various purulent conditions such as impetigo, furuncles, and others.
  — Serious organ infections include endocarditis, pneumonia, and infections of the bones and joints that lead to osteomyelitis and septic arthritis.

• Toxin-mediated conditions caused by S. aureus
• Toxic shock syndrome
  — Formerly associated with tampon use, septic shock now occurs at least as often, if not more often, in non-menstruating individuals.
  — Toxic shock syndrome is characterized by acute onset of fever, gastrointestinal upset, sore throat, and diffuse erythroderma; desquamation occurs when the skin begins to peel and flake away.
• Scalded skin syndrome* is also a desquamating condition, is most common in infants and young children.
  — In our illustration, we’ve shown the characteristic red and flaky areas of skin.
• Food Poisoning

Staphylococcus epidermis

• An important source of medical device infections, particularly in individuals with prosthetic joints and valves or catheters and shunts; it is a significant cause of bacterial sepsis.

Staphylococcus saphrophyticus

• Common cause of urinary tract infections.

Catalase-negative cocci: Streptococci & Enterococci

• Hemolysis can be used to distinguish between species that are alpha-hemolytic, beta-hemolytic, and gamma-hemolytic.

Alpha-hemolytic strains
Can be further distinguished by their sensitivity to optochin:

• Streptococcus pneumoniae is sensitive to optochin, whereas Viridans group Streptococci are not.
  — Streptococcus pneumoniae is associated with pneumonia, otitis media and sinusitis, as well as meningitis.
  — Viridans group Streptococci are associated with subacute endocarditis and dental caries, as well as some other infections not listed here.

Beta-hemolytic strains
Can be distinguished by their sensitivity to Bacitracin:

• Group A Streptococcus is sensitive, whereas Group B streptococcus is not.
  — Group A Streptococcus, also known as Streptococcus pyogenes, causes pharyngitis with white exudate in the tonsils (strep throat); some people will also develop a rash over their bodies, called Scarlet fever.
  — Group A Streptococcus is also responsible for a variety of soft tissue infections, which can range from impetigo and erysipelas to the more serious cellulitis or even necrotizing fasciitis.
  — Group A streptococcus can cause toxic shock syndrome.
  — Delayed, anti-body mediated reaction to Group A Streptococcus infection can occur in some patients, and may produce post-streptococcal glomerulonephritis or rheumatic fever.
  — Group B Streptococcus, also called Streptococcus agalactiae, is associated with neonatal infections including meningitis, pneumonia, and bacteremia; because the neonates acquire the bacteria from their mothers, prenatal care should include screening for Group B Streptococcus. Post-pregnancy infections can also have serious consequences for the mother.
  — Adult infections can manifest similarly, including bacteremia, pneumonia, and bone, joint, and soft tissue infections.

Gamma-hemolytic strains
Strains that can grow in relatively high concentrations of salt and bile are categorized as Enterococcus
— These bacteria were formerly categorized as Group D Streptococcus, and are common commensals of the GI tract.

• Of particular concern are strains resistant to Vancomycin.
• Enterococci are a significant cause of nosocomial infections.
• Enterococci are commonly associated with urinary tract infections, as well as endocarditis, peritonitis, and bacteremia.
• Streptococcus bovis, which is also gamma-hemolytic but cannot thrive in high salt concentrations, causes similar illnesses as Enterococcus, and is also associated with colorectal cancer.

OVERVIEW

• Bacteria produce toxins to break down host tissues and promote their own growth.
• Toxins facilitate invasion, release nutrients from host cells, and resist destruction by the immune system.
• Furthermore, as bacteria colonize the host, they trigger immune and inflammatory responses; in fact, the symptoms of many infections are the result of host immune response.

ENDOTOXINS

• Endotoxins are part of the cell wall of Gram-negative bacteria
  – The lipopolysaccharide endotoxin extends from the outer membrane
  – Three important regions of the endotoxin, starting at the cell membrane: Lipid A, Core polysaccharides, and the O-antigen
• They have relatively low toxicity
• Upon infection and endotoxin release, the Lipid A portion of the lipopolysaccharide interacts with Toll-Like Receptor 4 on macrophage surfaces and triggers cytokine release, potentially inciting cytokine storms.
• Indicate four important consequences:
  – Complement activation, which results in neutrophil chemotaxis and inflammation.
  – Cytokines IL-1 and IL-6 induce fever.
  – Tissue factor activation leads to coagulation.
  – Tumor necrosis factor, nitric oxide, and bradykinin induce hypotension (low blood pressure) via vasodilation.

• Recognize that, in acute, local conditions, these reactions can protect the host from infection; however, in large quantities, endotoxin can be fatal.
• Endotoxin is a major cause of septic shock.

EXOTOXINS

• Polypeptides secreted by both gram-positive and gram-negative bacteria.
• Many are dimeric in structure, with A and B subunits.
  – The A subunit is the toxic active Portion; the B subunit is the Binding portion that attaches to the host cell.
• Highly toxic, even in small quantities.
• Directly kill or alter host cell functions.

Mechanisms:

• ADP-ribosylation adds ADP-ribose to proteins in the host cell.
  – Diptheria toxin inhibits protein synthesis, leading to cell death; other toxins that act via ADP-ribosylation can hyperactivate protein synthesis.

• Increase Cyclic AMP
  – In the case of heat-labile enterotoxin, this results in fluid and electrolyte loss into the lumen of the gastrointestinal tract, which causes watery diarrhea.

• Proteases
  – Botulinum toxin is a neurotoxin that blocks acetylcholine release, producing paralysis.

• Super antigens
  – For example, Toxic Shock Syndrome toxin overstimulates T cells, triggering cytokine storms.

INFLAMMATION

Two main types of inflammation associated with bacterial pathogens:

• Purulent inflammation is characterized by neutrophilinfiltration and pus formation from liquefied tissues.
• Granulomatous inflammation, which occurs in chronic inflammation, is characterized by aggregates of macrophages and epithelioid cells, called granulomas, aka, tubercles (as in tuberculosis).

Recall that inflammation is characteristic of early immune responses:

• When controlled and acute, it has protective effects for the host: eradication of microbes and tissue healing.
• However, when uncontrolled or chronic, inflammatory and immune responses cause significant damage to the host.
  – For example, let’s consider two disease states that can occur after Group A streptococcus (Streptococcus pyogenes) infection.

1. Post-streptococcal glomerulonephritis:
   Neutrophil infiltration and deposition of antigen-antibody complexes in the basement membrane, which damage the renal filtration system. Be aware that the initial infection occurred elsewhere in the body, such as the skin or throat, leading to circulating immune complexes that became fixed to the glomeruli.
2. Rheumatic fever and heart disease, which develop in the weeks following untreated pharyngeal infection by Group A streptococcus. The host’s innate and adaptive responses lead to valve thickening, which can cause severe valvular stenosis. In the histological sample, we can see areas of calcification and fibrosis have damaged the valve tissue.

OVERVIEW

• The host provides shelter, warmth, moisture, and food for bacteria; as we learn elsewhere, there are several microorganisms that take advantage of these benefits without harming the host – these commensals comprise the microbiome.
• Virulence factors increase a bacterial strain’s ability to colonize and cause disease.
  – The genes for virulence factors are often clustered together in pathogenicity islands; thus, they are easily transferred via plasmids, bacteriophages, and other gene-sharing mechanisms.
  – Furthermore, the genes for many virulence factors are regulated via quorum sensing; as we learn elsewhere, quorum sensing allows for bacterial behaviors to change with group density.

ADHESION TO HOST CELLS & ECM

This early step in colonization unleashes specific pathogen behaviors and host responses.

• Adhesins are molecules that facilitate adhesion to other pathogens or host structures; indicate that they can be located on the tips of pili or on the bacterial cell surface.
• A bacterium can have one or several types of pili and surface adhesions.
  – Different strains of the same bacteria can have different pili types, which can influence their virulence in different host environments.

Gram-Negative bacteria:

• P pili, Type I pili, Curli pili, and Type IV pili.
  – Uropathogenic strains of Escherichia coli use both P pili and Type I pili to adhere to the urothelium of the urinary tract; without these pili, the bacteria would be physically removed by the flow of urine.
  – Some strains of E. coli have curli pili, which, in addition to adhesion, provoke the host inflammatory response.
  – Type IV pili confer twitching motility to some species, independent of flagella; Neisseria gonorrhoeae and Pseudomonas aeruginosa are examples of bacteria that “walk” via retraction of Type IV pili.

Gram-Positive bacteria

• Also have pili-like structures; though assembled differently the pili of Gram-negative bacteria, they perform similar functions.
• Spa, GAS M1, PI-1, PI-2
  – Spa pili, which are long and flexible, facilitate adherence of Corynebacterium diphtheriae, the causative agent of diphtheria, to epithelial cells of the pharynx.
  – Similarly, GAS M1 facilitates adherence of Group A Streptococcus (aka, Streptococcus pyogenes) to pharyngeal epithelial cells.
  – PI-1 and PI-2 facilitate adherence of Group B Streptococcus (Streptococcus agalactiae) to the cells of the lungs.
  PI = Pilus Island, which refers to the gene loci. Group B streptococcus causes neonatal sepsis, pneumonia, and meningitis.

MSCRAMMs

Non-pilus adhesins on the bacterial cell surface that attach pathogens to host structures.

• MSCRAMMs – Microbial Surface Components Recognizing Adhesive Matrix Molecules – are proteins that facilitate colonization by Gram positive bacteria.
• Staphylococcus aureus adheres to fibrinogen via Clumping factor A, and to fibronectin via Fibronectin Binding Protein (FnBP).
• In turn, these ECM components make their own connections to platelets and host cells, thereby establishing secure associations between S. aureus and the host.
• Furthermore, S. aureus can take advantage these associations and enter host cells to either lie latent or act as a super-antigen (for more, see our tutorial on bacterial endocarditis).

Biofilm

• To further secure adherence to the host, and to protect themselves from the immune system and antibiotics, pili and surface adhesins contribute to the formation of biofilms.
• Comprises bacterial cells, in some cases of multiple strains or species, surrounded by matrix. Biofilm formation is an example of a virulence factor regulated by quorum sensing.
  – Dental plaque is an example of a biofilm, which we can see in the image as purple-stained areas.
  – Biofilm production by S. aureus in endocarditis facilitates the growth of large bacterial vegetations, which can damage the heart or, if they break free, cause stroke.

ENTRY INTO HOST CELLS

2 examples of how some bacteria enter into host cells.

Complement Opsonization

• Mycobacterium tuberculosis, the causative of tuberculosis, makes use of complement opsonization.
• Recall that opsonization by C3b typically results in phagocytosis and microbe destruction; however, M. tuberculosis, once taken up by macrophages, avoids destruction and instead replicates inside the host cell.
• Ultimately, pathogen-host interactions result in the formation of granulomas, aka, tubercles, which harbor M. tuberculosis.

Type III secretion system

• The Type III secretion system uses a needle-like structure to inject effectors into host cells.
  – The effectors and their actions vary by bacterial strain.
• In the case of salmonella, the effectors trigger cytoskeleton reorganization of host cells such that the pathogen can enter it; once inside, the bacteria can make use of host cell machinery and replicate.

EVASION OF HOST IMMUNE SYSTEM

Mechanisms to evade phagocytosis

• The polysaccharide capsule on Gram positive bacteria inhibits phagocyte adhesion.
  – Thus, anticapsular antibodies are important preventative measures against infection by Streptococcus pneumoniae and Neisseria meningitides.
• The M protein of Group A Streptococcus (aka, Streptococcus pyogenes) resists opsonization and phagocytosis.
• Protein A, found in the cell wall of Staphylococcus aureus, binds immunoglobulins M and G, preventing complement activation and, therefore, phagocytosis.
• Leukocidins, which are pore-forming cytotoxins released by staphylococcus bacteria, kill leukocytes, including phagocytic neutrophils and macrophages.

Immunoglobin A protease degrades IgA

• This allows the causative agents of bacterial meningitis, Streptococcus pneumoniae, Neisseria meningitidis, and Haemophilus influenzae, to adhere to mucous membranes.

Operon

Cluster of genes that are transcribed into one long mRNA allowing the genes of a single pathway to be controlled with a single on/off switch

Operator

Segment of DNA that acts as a switch to control access of RNA polymerase to the gene

Activator

Protein that binds to DNA and stimulates gene transcription

Negative regulation

Bound repressor protein blocks transcription

Positive regulation

Bound activator protein promotes transcription

LAC OPERON

• Set of genes that code for proteins necessary for the bacterium to use the sugar lactose as an energy source

Structure

• 3 genes: lacZ (beta-galactosidase), lacY (lactose permease) and lacA (galactoside acetyltransferase)
• Promoter region:

1. CAP-binding site
2. Operator

• lacI gene – prior to CAP-binding site; codes for repressor protein; under control of a different promoter

High glucose, no lactose

• CAP-binding site empty (inactive catabolite activator protein due to low cAMP levels)
• Repressor is bound to operator (no allolactose present to inactivate repressor)
• No transcription

No glucose, no lactose

• CAP is bound to CAP-binding site (low glucose means high levels of cAMP)
• Repressor is bound to operator
• No transcription

High glucose, lactose available

• Cap-binding site empty
• Operator is empty (allolactose present inactivates repressor protein)
• Low-level transcription

No glucose, lactose available

• CAP is bound to CAP-binding site
• Operator is empty
• High levels of transcription