Theme: Microbiological diagnosis of whooping cough and Haemophilus influenzae infections Morphology Bordetella pertusis, Gram stain Bordetella parapertusis, Gram stain • Pathogenesis of whooping cough Virulence factors Cough plate method Per nasal swab Laboratory diagnosis of whooping cough • Specimen: nasopharyngeal aspirate (cough plate method), nasopharyngeal swab. • 1. IF (detection of antigen). • 2. Culture on Bordet-Gengou medium. • Identification: – slide AT. – Biochemical tests: • B.pertusis: urease-negative, citrate-negative, • B.parapertusis: urease-positive, citrate-negative. • 3. Serology (detection of antibodies): AT, CFT. Scheme of laboratory diagnosis of whooping cough Sputum, nasopharyngeal swab Microscopy 1 step Culture Immufluorescense Gram stain Agglutination test, CFT Presumptive diagnosis Study of colonies 3 step Serology Cough plate method: Borde-Gengou agar Presumptive diagnosis 2 step Serum Gram stain Seeding on slant agar Pigment production Slide agglutination test Urease test Definitive diagnosis Morphology of Haemophilus influenzae, Gram stain Growth on chocolate agar Identification of Haemophilus influenzae Grows on Mueller-Hinton with XV factor disk X disk only V disk only Satellite around various colonies on blood agar Laboratory diagnosis of H. influenzae infections. • Specimen: liquor. • 1. IF (detection of capsular antigen). • Specimen: nasopharyngeal swab, pus, sputum, blood, liquor. • 2. Culture: Chocolate agar. • 3. Serology (detection of capsular antigen): AT, ELISA, immunoelectrophoresis. Nasopharyngeal swab, pus, sputum, blood, cerebrospinal fluid Microscopy 1 step Culture Immufluorescense Gram stain PCR Chocolate agar Cerebrospinal fluid Serology (detection of capsular antigen). IF, immunoelectrophoresis Presumptive diagnosis Study of colonies 2 step Gram stain Seeding on slant agar Biochemical tests 3 step catalasepositive ureasepositiv e indolpositive nitrate reductionpositive H2Snegative Serology(detection of capsular antigen) oxydasenegative Definitive diagnosis nonhemolytic AT ELISA immunoelectrophoresis • • • • • • • • Biochemical tests: catalase-positive, urease-positive, indol-positive, nitrate reduction-positive, nonhemolytic, H2S-negative, oxydase-negative. Haemophilus influenzae treatment • • • • • • • • • • • • • • Penicillins ampicillin amoxicillin ampicillin/sulbactam amoxicillin/clavulanate Cephalosporins cefuroxime ceftriaxone cefotaxime Alternative: Trimethoprim-Sulfomethoxazole(TMP-SMX) azithromycin clarithromycin doxycicline