Micro- unit 3 SLG

Ziehl-Neelsen / Kinyoun stain target organism

Mycobacterium tuberculosis & Nontuberculous Mycobacteria (NTM) 

Auramine-Rhodamine stain target organism

Mycobacteria species (Fluorescent screening stain) 

modified Acid-Fast stain target organisms

Nocardia species & Rhodococcus 

Calcofluor White / KOH target organism

Fungal elements (Aspergillus, Blastomyces, Histoplasma, Coccidioides) 

Direct Fluorescent Ab (DFA) target organism

Legionella pneumophila, Bordetella pertussis, Respiratory Viruses 

Buffered Charcoal Yeast Extract (BCYE) target organism

Legionella pneumophila 

Buffered Charcoal Yeast Extract (BCYE) key constituents and growth characteristics

Contains L-cysteine and ferric pyrophosphate. Colonies appear white-blue with ground-glass appearance. 

Bordet-Gengou / Regan-Lowe target organism

Bordetella pertussis 

Bordet-Gengou / Regan-Lowe  key constituents and growth characteristics

Regan-Lowe (charcoal horse blood agar) recovers B. pertussis. Colonies look like shiny mercury droplets. 

Cystine-Tellurite / Tinsdale Medium target organism

Corynebacterium diphtheriae 

key constituents and growth characteristics

Potassium tellurite inhibits normal flora. C. diphtheriae forms black/brown colonies with dark halos (Tinsdale). 

Lowenstein-Jensen (LJ) / Middlebrook target organism

Mycobacterium tuberculosis 

Lowenstein-Jensen (LJ) / Middlebrook key constituents and growth characteristics

Egg-based (LJ) or synthetic agar (Middlebrook 7H10/7H11). M. tuberculosis shows rough, buff, non-pigmented colonies. 

sabouraud dextrose/ mycosel agar target organism

Fungal Respiratory Pathogens 

Sabouraud Dextrose / Mycosel Agar key constituents and growth characteristics

SAB supports yeast and mold growth; Mycosel adds cycloheximide and chloramphenicol to suppress bacteria/saprophytes. 

routine respiratory cultures incubation

35°C–37°C in a capneic atmosphere (5%–10% CO2) for 24–48 hours to support fastidious capsular pathogens

special cultures incubation

require extended incubation up to 4–6 weeks.

Normal Oropharyngeal Flora

Viridans group streptococci, non-pathogenic Neisseria spp., Coagulase-negative Staphylococci, Corynebacterium spp. (diphtheroids), Haemophilus parainfluenzae, and Rothia mucilaginosa.

Potential / Primary Pathogens

Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, Staphylococcus aureus (including MRSA), Streptococcus pyogenes (Group A), Pseudomonas aeruginosa, Klebsiella pneumoniae, Legionella pneumophila, Mycoplasma pneumoniae, Mycobacterium tuberculosis.

Streptococcus pneumoniae clinical significance

Leading cause of community-acquired pneumonia (CAP), otitis media, sinusitis, and bacteremic pneumonia. 

haemophilus influenzae clinical significance and disease states

Causes CAP, acute exacerbations of COPD, epiglottitis, otitis media. Serotype b (Hib) major pediatric pathogen. 

moraxella catarrhalis key biochemical and phenotypic reactions

Gram-negative diplococci (kidney-bean). Oxidase (+), Catalase (+), Butyrate Esterase (+), DNase (+). 'Hockey puck' colony mobility.

moraxella catarrhalis clinical significance and disease states

Causes otitis media, sinusitis, bronchitis, pneumonia in elderly and COPD patients. 

Staphylococcus aureus / MRSA clinical significance and disease states

Post-viral secondary bacterial pneumonia, nosocomial/ventilator-associated pneumonia (VAP), necrotizing pneumonia. 

Pseudomonas aeruginosa  key biochemical and phenotypic reactions

Gram-negative non-fermenting rod. Oxidase (+), Pyocyanin/Pyoverdin green pigment, grape-like odor, Growth at 42°C. 

pseudomonas aeruginosa clinical significance and disease states

Major cause of VAP, chronic pulmonary infection in Cystic Fibrosis (CF) patients, healthcare-associated pneumonia. 

klebsiella pneumoniae key biochemical and phenotypic reactions

Gram-negative rod. Lactose fermenter (pink on MAC), Mucoid colonies (hypercapsule), Indole (-), VP (+), Citrate (+), Motility (-). 

klebsiella pneumoniae clinical significance and disease states

Severe lobar pneumonia with 'currant jelly' sputum; frequent cause of hospital-acquired pneumonia

legionella pneumophila key biochemical and phenotypic reactions

Fastidious Gram-negative rod. Requires BCYE medium (L-cysteine dependent). Direct fluorescent antibody (+) or Urinary Antigen (+). 

Legionella pneumophila clinical significance and disease states

Legionnaires' disease (severe atypical pneumonia) and Pontiac fever. Water-borne environmental pathogen. 

kirby bauer disk diffusion

Standardized inoculum (0.5 McFarland) on Mueller-Hinton Agar (MHA) or MHA with 5% Sheep Blood (for S. pneumoniae). Measured zone sizes correlated with CLSI breakpoints.

broth microdilution/ MIC

Determines Minimum Inhibitory Concentration (μg/mL) using automated or manual broth systems (e.g., HTM broth for Haemophilus).

E test (gradient strip)

Quantitative MIC determination for fastidious pathogens like S. pneumoniae (penicillin/cefotaxime MICs).

Altered Penicillin-Binding Proteins (PBP2b/2x)  target organism

Streptococcus pneumoniae (PRSP) 

Altered Penicillin-Binding Proteins (PBP2b/2x) detection protocol and clinical implications

Screened using 1µg Oxacillin disk (zone <20mm indicates resistance; requires quantitative E-test MIC for Penicillin/Ceftriaxone). 

Beta-Lactamase Production target organisms

Haemophilus influenzae & Moraxella catarrhalis 

Beta-Lactamase Production detection protocol and clinical implications

Rapid Chromogenic Cephalosporin (Cefinase/Nitrocefin) test. Positive test confers resistance to ampicillin/amoxicillin. 

mecA Gene / PBP2a (MRSA) detection protocol and clinical implications

Detected via Cefoxitin disk diffusion (surrogate marker) or meCA PCR. Confers resistance to all beta-lactams. 

D-test (inducible Clindamycin resistance) target pathogens

Erythromycin disk placed adjacent to Clindamycin disk. Flattening of clindamycin zone ('D' shape) indicates inducible erm gene resistance. 

ESBL, ampC, CRE target organisms

Enterobacteriaceae & Pseudomonas aeruginosa 

ESBL, ampC, CRE detection protocol and clinical implications

Extended-spectrum beta-lactamases and carbapenemases in hospital-acquired pneumonia. Requires CLSI phenotypic screening and confirmation