Slide 1Slide 2Slide 3Slide 4Slide 5Bordetella pertussisEpidemiology of PertussisSlide 8Epidemiology of PertussisSlide 10Slide 11Virulence factors of Bordetella pertussisVirulence factors of Bordetella pertussisSlide 14Slide 15Pertussis: Clinical ManifestationsDiagnosisQuestions?Slide 19Slide 20Haemophilus influenzaeVirulence Factors of H. influenzaeSlide 23Nontypeable H. influenzae (NTHI)NTHI –associated Otitis MediaSlide 26Slide 27Slide 28Slide 29Slide 30Slide 31Slide 32Slide 33Two serious Bacterial pathogensof theUpper Respiratory TractDr. Terry D. Connell: [email protected] influenzaeBordetella spp.Bordetella spp.Five pathogenic species of Bordetella B. bronchiseptica Kennel cough (dogs)Necrotic rhinitis (pigs)URT infection (human)B. avium Coryza (birds)B. parapertussis Mild whooping cough (human)B. homesii Pneumonia, bacteremiaBordetella pertussis Whooping cough (human)DPT Vaccine !!Bordetella pertussisExtremely small, gram-negative coccobacilliStrictly aerobicObligate human pathogen. No animal reservoir (unlike Salmonella enteriditis)Fastidious, slow growing Three to six days for colonies on Bordet-Gengou agarInfects the upper respiratory tract (URT)T) - ciliated respiratory epitheliumElaborates powerful toxins which elicit most of the symptoms of the disease“pertussis” - “violent cough”Epidemiology of PertussisWorldwide problem - 60 million cases, 600,000 deaths/yrDeveloped countries - Dramatic recent increase in cases● Decreased vaccine use● Rise of strains resistant to the vaccineHighly contagious - attack rates of 50 -100%Transmission - aerosol droplets from coughingFemales - Higher attack rate, morbidity and mortality (???)Epidemiology of PertussisWorldwide problem - 60 million cases, 600,000 deaths/yrDeveloped countries - Dramatic recent increase in cases● Decreased vaccine use● Rise of strains resistant to the vaccineHighly contagious - attack rates of 50 -100%Transmission - aerosol droplets from coughingFemales (???) - Higher attack rate, morbidity, mortality Age distribution - Recent shifts in age groups19881998Shift in age distributionPre-vaccine - predominately infected young children► Young children (<1 - 3 years of age)Post-vaccine - increased rate of infection in two groups ► Older children (5 to 14) ► Young adults (18 to 25 yrs) A SHIFT in age-distribution of infectionVaccines exerts STRONG SELECTIVE PRESSURES on pathogensSelects for variants for which the vaccine doesn’t evoke protectionVaccination has “pushed” bacterium into other age groups. WHY?- fewer maternal antibodies remaining in older children - protection by vaccination in young children is not lifelongVirulence factors of Bordetella pertussisPili Attachment to host cellsCapsule Antiphagocytic, adherenceFilamentous hemagglutinin Adherence to glycolipidsPertactin Binds to host cellsBinds to: Ciliated epithelium of RTMultiple ADHESINS:Virulence factors of Bordetella pertussisPertussis toxin (Ptx) A1-B5 class of toxin (CT) B pentamer – binding to receptors on cells A polypeptide – enzyme which increases cAMP cAMP = cyclic adenosine monophosphate Adenlyate cyclase toxin Increases cAMP in infected URT cellsLymphocytosis-promoting Increases lymphocyte numbers in URTTracheal cytotoxin Fragment of the peptidoglycan cell wall- destroys ciliated epithelial cellsDermonecrotic toxin Causes skin lesions and fatality in miceLPS (endotoxin) Activates alternative complement pathwayMultiple TOXINS:Pertussis: Clinical Manifestations2. Catarrhal (or Prodromal) stage: Lasts from 7 to 14 days Nonspecific symptoms Malaise, rhinorrhea, lacrimation,low grade fever, anorexia (flu-like!) Dry cough develops, worse at night1. Incubation stage:No overt symptomsLasts 7 to 10 days (individual is already infectious!)Pertussis: Clinical ManifestationsPertussis: Clinical Manifestations3. Paroxysmal stage:Paroxysmal coughing -Series of repetitive coughs followed by a characteristic inspiratory “whoop” (cyanosis; convulsions; seizures)-Patient looks normal between paroxysms, with minimal fever Tenacious mucus Ciliostasis - Death of URT ciliated epithelial cells; trachael cytotoxin)- Failure of the respiratory escalator to move mucus from lungs to throatLymphocytosis - Neutrophil count in the tissues - to 200,000 cells/mlSymptoms last 1-2 weeks ! Ciliated cells re-differentiate from basal cells Treatment – Erythromycin Antibiotic treatment doesn’t ameliorate the symptoms. Why not?Convalescence - 3 to 4 weeks; lymphocytes ↓ gradually; cough subsidesDiagnosis1. How one handles and cultures the sputum specimen is critical 2. Culturing bacteria from respiratory secretions (Not always successful!!)-Viable, but non-culturable bacteria - “VBNC” -Cannot use cotton-swabs for sampling the throat (fatty acids in cotton kill B. pertussis bacteria)4. Immunofluorescence assay on secretions Antibodies to specific proteins5. Agglutination reaction on secretions Antibodies to specific proteins6. Diagnosis based on clinical diagnosis: Whoop-type coughing and lymphocytosisAgglutination reactionAbcamQuestions?Infection of the:Upper Respiratory TractLower Respiratory Tract Middle EarHaemophilus influenzaeInfection of the:Upper Respiratory TractLower Respiratory Tract Middle EarHaemophilus influenzaeAerobic, gram-negative bacteriumCoccobacillus or pleomorphic rodsMay be encapsulated or non-encapsulated.Required for growth:- Hemin- Nicotinamide adenine dinucleotide (NAD)Fastidious bacterium to culture Chocolate agar (heat-treated erythrocytes)Virulence Factors of H. influenzaeCapsule Secreted polysaccharide “coat”; anti-phagocytic activity (resists killing bymacrophages and polymorphonuclearneutrophils)Pilus Rod-like appendage that promotes attachment to target cells of theURT or middle earHAP proteins Surface proteins that promotes more intimate adherence of bacterium to cellsEndotoxin (LPS) Inflammatory, pyrogenic, impairsciliary function of URT cellsIgA1 protease Enzyme that facilitates colonization of themucosal surface; destroys IgAIgAHow do we differentiate between strains of Haemophilus influenzae ?Six (6) serotypes ; expression of capsular POLYSACCHARIDE = A, B, C, D, E, … Serotype B (Hib) - the most prominent disease-causing strainuntil development of the very successful vaccine