Respiratory Infections by Gram-Negative Bacteria

• Gram-negative rods that relate to the respiratory tract include Bordetella pertussis and Acinetobacter baumannii.
• Symptoms of respiratory tract infection: sore throat, loss of voice, voice change, and the need to clear your throat.
• Treatment for respiratory infections: use of a dehumidifier, vocal rest, gargling salt water, drinking fluids, and avoiding smoking or alcohol.

Medically Important Gram-Negative Rods

• Four medically important Gram-negative rods are typically associated with the respiratory tract: Haemophilus influenzae, Bordetella pertussis, Acinetobacter baumannii, and Legionella pneumophila.
• H. influenzae and B. pertussis are found only in humans.
• L. pneumophila is found primarily in environmental water sources.
• A. baumannii is found in environmental water sources but also colonizes skin and the upper respiratory tract.

Diseases and Diagnosis

• Haemophilus influenzae causes meningitis, otitis media, sinusitis, pneumonia, and epiglottitis; diagnosed by culture, capsular polysaccharide in serum or spinal fluid and requires Factors X and V for growth with a vaccine available, prophylaxis for contacts is Rifampin.
• Bordetella pertussis causes whooping cough and is diagnosed by fluorescent antibody on secretions, and there is a vaccine available, requires Azithromycin as precaution for close contacts.
• Legionella pneumophila causes pneumonia and is diagnosed by serology or urinary antigen culture.
• Acinetobacter baumannii causes ventilator-associated pneumonia and can be diagnosed by culture and no vaccine available.

Bordetella Pertussis Etiology

• Bordetella pertussis causes Whooping Cough (Pertussis).
• B. pertussis is a small, coccobacillary, encapsulated Gram-negative rod.
• B. pertussis (pathogen only for humans) is transmitted by airborne droplets produced during severe coughing episodes.
• Organisms attach to ciliated epithelium of the upper respiratory tract, but do not invade underlying tissue.
• Decreased cilia activity and subsequent death of ciliated epithelial cells are important aspects of pathogenesis.
• Pertussis is a highly contagious disease that occurs primarily in infants and young children and has worldwide distribution.
• The number of cases has declined in the US because vaccine use is widespread.
• Outbreaks of pertussis during 2005, 2010, and 2012 have led to concern about waning immunity to the vaccine and to the recommendation that an additional booster immunization be given.

Pertussis Toxin And Pathogenesis

• Attachment of organism to cilia of epithelial cells is mediated by protein on pili called filamentous hemagglutinin.
  • Antibody against filamentous hemagglutinin inhibits attachment and protects against disease.
• Pertussis toxin stimulates adenylate cyclase by catalyzing the addition of adenosine diphosphate ribose (a process called ADP-ribosylation) to inhibitory subunit of G protein complex (Gi protein).
  • This results in prolonged stimulation of adenylate cyclase and a consequent rise in cAMP and cyclic AMP-dependent protein kinase activity.
  • This results in edema of the respiratory mucosa that contributes to the severe cough of pertussis.
  • Toxin also has a domain that mediates its binding to receptors on the surface of respiratory tract epithelial cells and is an A-B subunit toxin.
  • Pertussis toxin also causes striking lymphocytosis in the blood of patients with pertussis.
  • Toxin inhibits signal transduction by chemokine receptors, resulting in failure of lymphocytes to enter lymphoid tissue (Spleen & Lymph Nodes).
  • Because lymphocytes do not enter lymphoid tissue, there is an increase in their number in blood.
  • Inhibition of signal transduction by chemokine receptors is also caused by ADP-ribosylation of Gi protein
• Organisms also synthesize & export Adenylate Cyclase
  • This enzyme (when taken up by phagocytic cells [e.g., neutrophils]) can inhibit their bactericidal activity.
  • Bacterial mutants that lack cyclase activity are Avirulent.
• Tracheal cytotoxin is a fragment of the bacterial peptidoglycan that damages ciliated cells of respiratory tract
  • Tracheal cytotoxin act in concert with endotoxin to induce nitric oxide, which kills the ciliated epithelial cells.

Clinical Finds

• Whooping cough is an acute tracheobronchitis that begins with mild URT tract symptoms followed by severe paroxysmal cough (lasts from 1 to 4 weeks).
• Paroxysmal Pattern is characterized by: a series of hacking coughs, accompanied by the production of copious amounts of mucus, that ends with an inspiratory "whoop" as air rushes past the narrowed glottis.
• Despite the severity of symptoms, the organism is restricted to the respiratory tract (blood cultures are negative).
• Pronounced Leukocytosis with up to 70% lymphocytes is seen.
• Although Central Nervous System Anoxia and Exhaustion can occur as a result of the severe coughing, Death is due mainly to Pneumonia.
• Classic findings of Whooping Cough (described earlier) occur primarily in young children.
• In adults, B. pertussis infection often manifests as a paroxysmal cough of varying severity lasting weeks.
• The characteristic Whoop is often absent, leading to difficulty in recognizing the cough as caused by this organism.
• In the correct clinical setting, adults with cough lasting several weeks (often called the 100-day cough) should be evaluated for infection with B. pertussis.

Lab Testing for B. Pertussis

• Diagnosis of whooping cough can usually be made clinically.
• Only occasionally is the laboratory required to investigate B. pertussis infection.
• Specimens: Preferably Nasopharyngeal Secretions collected by Aspiration or a correctly taken Pernasal Swab taken during the paroxysmal stage.
• Morphology: B. pertussis is a small, non-motile, capsulated Gram-negative coccobacillus.
• It may occur singly or in chains and may show bipolar staining.
• Bordetella species are strict aerobes
• Specimens for the isolation of B. pertussis must be cultured as soon as possible.
• The medium of choice for Bordetellae is Bordet-Gengou medium (contains a high percentage of blood [20%-30%] to inactivate inhibitors in agar).
• Culture: BGM or Charcoal Cephalexin Blood Agar when incubated for 2–6 days at 35-37 ºC in a Moist Aerobic Atmosphere, and they produce small pearly-gray, shiny (mercury-like), usually mucoid colonies.
• Identification can be made by : agglutination with specific antiserum or by fluorescent antibody staining.
• The organism grows very slowly in culture, so direct fluorescent-antibody staining of nasopharyngeal specimens can be used for diagnosis.
• PCR-based tests are highly specific and sensitive and should be used if available.
• Isolation in patients with prolonged cough is often difficult: Serologic tests that detect antibody in a patient's serum can be used for diagnosis in those patients.
• Bordetella parapertussis grows more rapidly and forms larger colonies than B. pertussis.
• It produces pigment in the Medium and is able to grow aerobically on blood agar and nutrient agar.
• B. pertussis is Oxidase Positive and Urease Negative.
• B. parapertussis is Oxidase Negative and Slowly Urease Positive (after 24 h).
• Bordetella species can be serotyped in a specialist microbiology laboratory.
• Antimicrobials with activity against B. pertussis include: Erythromycin, Chloramphenicol, Tetracycline, and Cotrimoxazole.
• Protection against whooping cough is by prophylactic vaccination.

Treatment and Preventatives

• Azithromycin is the drug of choice, will reduce organisms in the throat and decrease risk of secondary complications.
• But has little effect on the course of the disease at the "prolonged cough" stage (because the toxins already damaged the respiratory mucosa).
• Supportive care (e.g., oxygen therapy & suction of mucus) during the paroxysmal stage is important (especially in infants).
• Acellular Vaccine contains purified proteins from Bordetellae.
• Killed Vaccine contains inactivated Bordetella pertussis
• The Acellular Vaccine contains 5 antigens purified from Bordetellae (used currently in the US). -Main Immunogen in this vaccine is Inactivated Pertussis Toxin (Pertussis Toxoid). -Toxoid in the vaccine is Pertussis Toxin that is inactivated genetically by introducing two amino acid changes which (eliminates its ADP-ribosylating activity but retains its antigenicity).
• It is the first vaccine to contain a genetically inactivated toxoid.
• Other Pertussis Antigens in Acellular Vaccine are: Filamentous Hemagglutinin, Pertactin, and Fimbriae Types 2 & 3
• Acellular Vaccine has fewer side effects than the killed Vaccine.
• Acellular Vaccine has a shorter duration of immunity than the killed Vaccine.
• The pertussis vaccine is usually given combined with diphtheria and tetanus toxoids (DTaP) in three doses beginning at 2 months of age.
• A booster at 12 to 15 months of age and another at the time of entering school are recommended.
• Because outbreaks of pertussis usually occur among teenagers, a Booster for those between 10 and 18 years old is recommended.
• This vaccine (called Boostrix) contains Diphtheria and Tetanus Toxoids also
• Another vaccine (called Adacel) also contains Diphtheria and Tetanus Toxoids
• A Pertussis Booster Dose is recommended for adults as well
• To Protect newborns, pregnant women should receive Pertussis Vaccine.
• Anti-pertussis IgG will pass the placenta & protect the newborn.
• The killed Vaccine is no longer used in the US (because it is suspected of causing various side effects, including Post-Vaccine Encephalopathy at a rate of about one case per million doses administered).
• The killed Vaccine is in use in many other countries
• Azithromycin is useful in the prevention of disease in exposed, unimmunized individuals.
• It should also be given to immunized children younger than 4 years who have been exposed (because vaccine-induced immunity is not completely protective).

Acinetobacter Baumannii Specifics

• Acinetobacter species are Gram-negative coccobacillary rods.
• Found commonly in soil and water, but they can be part of the normal flora.
• They are Opportunists that readily colonize patients with compromised host defenses
• Acinetobacter baumannii is the species usually involved in human infection.
• It causes disease mostly in a hospital setting (associated with respiratory therapy equipment [ventilator-associated pneumonia] & indwelling catheters).
• Sepsis, Pneumonia, and UTIs are the most frequent manifestations.
• A. baumannii is remarkably Antibiotic Resistant.
• Some isolates are resistant to all known antibiotics
• Imipenem is the drug of choice for infections caused by susceptible strains.
• Colistin is useful in Carbapenem-Resistant Strains.
• Previous Genus names for this organism include Herellea & Mima.

Questions focus on identifying Gram-negative bacteria, such as Bordetella pertussis and Haemophilus influenzae, commonly associated with respiratory infections. Emphasis is on differentiating them based on characteristics, reservoirs, and mechanisms of infection. The questions also cover diagnosis and potential outbreaks.