Next-generation sequencing for hypothesis-free genomic detection of invasive tropical infections in poly-microbially contaminated, formalin-fixed, paraffin-embedded tissue samples – a proof-of-principle assessment

Table 6 Comparison of the NGS Results in Terms of Percentage of Species-specific Reads of Investigated Fungal Species Per Total Number of Fungal Reads Only in the Sample with the Respective Largest Standard Deviation above the Mean of All Samples in Multiples of the Standard Deviation (SD), Depicted for the Samples from Patients with Invasive Mycoses. “Positive” = matching of the molecular results with histology. “Negative” = inconsistency of molecular results with histology. “Partially positive” = detection of both matching and nonmatching molecular results compared with histology. “Contaminated” = Detection of environmental fungi only. “Match” = NGS result matching the histological findings. “Mismatch” = NGS results not matching the histological findings, “No match” = No evidence for relevant fungal species above the respective standard deviation (SD), “Not performed” in “Specific PCR” = No specific PCR performed. Highest standard deviations (SD) are shown

Sample	Histological diagnosis	Specific PCR [13]	Panfungal PCRs [13]	NGS > 1st SD	NGS > 2nd SD	NGS > 3rd SD	NGS > 4th SD
1	Chromoblastomycosis	Not performed	Contaminated	No match	Match^d	No match	No match
2	Mucormycosis	Negative	Contaminated	Mismatch^e	No match	No match	No match
3	Histoplasmosis	Negative	Contaminated	Mismatch^f	Mismatch^g	Mismatch^h	No match
4	Mucormycosis	Positive^a	Contaminated	No match	No match	No match	Matchⁱ
5	Histoplasmosis or cryptococcosis	Negative	Contaminated	No match	No match	No match	Match^j
6	Chromoblastomycosis	Not performed	Contaminated	No match	Match^k	No match	No match
7	Rhinosporidiosis	Not performed	Contaminated	No match	No match	No match	No match
8	Mycetoma	Not performed	Contaminated	No match	No match	No match	Match^l
9	Rhinosporidiosis	Not performed	Contaminated	Mismatch^m	No match	No match	No match
10	Mycetoma	Not performed	Contaminated	Mismatchⁿ	No match	No match	No match
11	Histoplasmosis	Negative	Contaminated	No match	No match	No match	No match
12	Histoplasmosis	Negative	Contaminated	No match	No match	No match	No match
13	Chromoblastomycosis	Not performed	Contaminated	No match	No match	No match	No match
14	Histoplasmosis	Positive	Partially positive^b	Mismatch^o	No match	No match	Match^p
15	Coccidioidomycosis	Not performed	Contaminated	No match	No match	No match	No match
16	Coccidioidomycosis	Not performed	Contaminated	No match	No match	No match	No match
17	Mycetoma	Not performed	Partially positive^c	Mismatch^q	Mismatch^r	No match	Match^s

^aLichtheimia / Absidia corymbifera, ^bHistoplasma capsulatum in 2 out of 5 panfungal PCRs, ^cMadurella mycetomatis in 1 out of 5 panfungal PCRs, ^d4% Cladophialophora yegresii, ^e3% Cladophialophora yegresii, ^f0.2% Cryptococcus neoformans, 0.6% Chaetomium globosum, ^g1% Capronia epimyces, ^h0.6% Cladophialophora carrionii, ⁱ0.05% Rhizopus oryzae, ^j0.01% Cryptococcus carnescens, ^k0.8% Chaetomium globosum, 0.4% Cladophialophora psammophila, ^l16% Fusarium pseudograminearum, ^m3% Cladophialophora yegresii, ⁿ3% Cladophialophora yegresii, ^o0.09% Cryptococcus gattii, 0.4% Paracoccidioides brasiliensis, ^p34% Histoplasma capsulatum, ^q0.7% Capronia epimyces, ^r0.2 Exophiala dermatitidis, 0.5% Chaetomium thermophilum var. Thermophilum, 1% Chaetomium globosum, ^s4% Madurella mycetomatis, Matches with the histological findings are depicted in underlined print