Table 2 Discrepant clinical specimens resolved in favour of molecular assay (true positive or negative by PCR/sequencing)
Specimen No. | Specimen typea | Stain results | Standard methodsb | Initial molecular results | Sequence results | Results of resolution (repeat PCR and clinical review) |
|---|---|---|---|---|---|---|
1 | BAL LUL | CW = fungal elements | C. albicans, A. fumigatus, A. flavus | ITS/LSU targets POS, β-globin POS | ITS = C. albicans LSU = C. albicans | True positive PCR Fungal culture contaminated |
2 | BAL LLL | Gram stain = Hvy WBCs + mixed bacteria; CW = NEG | Aspergillus spp. | ITS/LSU targets NEG, β-globin POS | N/A | True negative PCR No pulmonary disease Fungal culture contaminated |
3 | BAL RLL | Gram stain – Hvy WBCs; CW = NEG | Aspergillus spp., Penicillium spp. | ITS/LSU targets NEG, β-globin POS | N/A | True negative PCR No pulmonary disease Fungal culture contaminated |
4 | BAL RUL | Gram stain − Hvy WBCs; CW = NEG | A. fumigatus | ITS/LSU targets NEG, β-globin POS | N/A | True negative PCR No pulmonary disease Fungal culture contaminated |
5 | BAL LLL | Gram stain = Hvy WBCs; CW = Hvy yeast seen | A. fumigatus, dermatiaceous fungus | ITS/LSU targets POS, β-globin POS | ITS = Exophiala dermatitidis LSU = Exophiala dermatitidis | True positive PCR Other BAL samples grew A. fumigatus, E. jeanselmi Interstitial pneumonia |
6 | BAL RLL | Gram stain =  Hvy WBCs + mixed bacteria; CW = NEG | A. fumigatus | ITS/LSU targets POS (weak), β-globin POS | ITS = A. fumigatus LSU = A. fumigatus | True positive PCR Clinical diagnosis of pulmonary Aspergillosis |
7 | BAL LUL | Gram stain = Sct. WBCs + mixed bacteria; CW = NEG | C. albicans + oropharyngeal flora | ITS/LSU targets NEG, β-globin POS | N/A | True negative PCR Pneumocystis jirovecii PCR POS |
8 | BAL RLL | Gram stain = Hvy WBCs; CW = Hvy yeast seen | C. albicans, yeast not C. albicans | ITS/LSU targets POS, β-globin POS | Mixed sequence: ITS = C. albicans and C. glabrata LSU = C. albicans and C. glabrata | True positive PCR Consistent with oropharyngeal colonization and overgrowth of Candida spp. |
9 | BAL RLL | Gram stain = Hvy WBCs + mixed bacteria; CW = NEG | A. niger, A. fumigatus | ITS/LSU targets POS, β-globin POS | ITS = A. niger LSU = A. niger | True positive PCR Other BAL samples grew both Aspergillus spp. Clinical diagnosis of airway colonization |
10 | BAL RLL | Gram stain =  Hvy WBCs + mixed bacteria; CW = NEG | C. albicans, A. fumigatus | ITS/LSU targets POS, β-globin POS | ITS = C. albicans LSU = C. albicans | True positive PCR Clinical diagnosis of metapneumovirus/enterovirus infection C. albicans consistent with airway colonization; A. fumigatus contaminant |
11c | Brain tissue | Gram stain − Hvy WBCs; CW = NEG | No growth after 4 weeks | ITS POS LSU NEG β-globin POS | ITS: Rhizopus oryzae LSU: No data | True positive PCR Pathology sections positive for broad aseptate hyphae Clinical diagnosis of rhinocerebral mucormycosis |
12c | Cheek tissue | CW = fungal elements | No growth after 6 weeks | ITS POS LSU POS β-globin POS | ITS: Rhizopus oryzae LSU: Rhizopus oryzae | True positive PCR Pathology PAS and GMS section stains showed broad aseptate hyphae Clinical diagnosis of rhinocerebral mucormycosis |
13c | Parotid gland tissue | CW = no fungal elements | C. albicans | ITS POS LSU POS β-globin POS | ITS: Rhizopus oryzae LSU: Rhizopus oryzae | True positive PCR Clinical diagnosis of rhinocerebral mucormycosis |
14 | Sinus tract fluid | Gram stain = Hvy WBCs with mixed bacteria including yeast | C. albicans | ITS POS LSU POS β-globin POS | ITS: C. glabrata LSU: C. glabrata | True positive PCR MALDI-TOF MS confirmed C. glabrata and isolate had elevated fluconazole MIC. Fungal culture initially mis-identified |
15d | Abdominal abscess tissue/fluid | Gram stain = Hvy WBCs + mixed bacteria; CW = NEG | No growth after 6 weeks | ITS POS LSU POS β-globin POS | ITS: C. albicans LSU: C. albicans | True positive PCR Clinical diagnosis of intra-abdominal abscess |
16 | Shoulder tissue (intermedullary) | Gram stain − = Hvy WBCs; CW = NEG | Alternaria spp. | ITS NEG LSU NEG β-globin POS | N/A | True negative PCR Clinical diagnosis of Cutibacterium acnes joint infection Fungal culture contaminated |
17 | R hip tissue | Gram stain = No WBCs; CW = NEG | Environmental fungus isolated (not further identified at reference laboratory) | ITS weak band LSU weak band β-globin POS | ITS: poor sequence LSU: poor sequence | True negative PCR No evidence of IFD Fungal culture contaminated |
18e | Liver aspirate | Gram stain = no WBCS; CW = NEG | No growth after 4 weeks | ITS POS LSU POS β-globin POS | ITS: Rhizomucor pusillus LSU: Rhizomucor pusillus | True positive PCR Pathology sections positive for broad aseptate hyphae Clinical diagnosis of hepatosplenic mucormycosis |
19f | Lung tissue/fluid | Gram stain = Few WBCs; CW = NEG | No growth after 6 weeks | ITS POS LSU POS β-globin POS | ITS: Histoplasma capsulatum LSU: No data | True positive PCR Pathology of lung tissue showed necrotizing granulomas with yeast morphologically consistent Clinical diagnosis of Histoplasmosis |