M-699. Multiplex-tandem PCR (MT-PCR) for Rapid Diagnosis of Fungemia
Session: Poster Session: Diagnostic Mycology
Saturday, October 25, 2008: 12:00 AM
Room: Hall C
Background: Invasive candidiasis (IC) is the major fungal disease in the critically ill. Blood cultures (BC) require ≥ 48 h for fungal detection and identification and are negative in up to 60% of cases. We designed, developed and tested a novel, real-time, MT-PCR assay that simultaneously identifies 12 fungi (including 7 Candida spp.) to determine whether the time to diagnosis could be expedited. Methods: EDTA blood samples from patients obtained before or at (T≤0) and/or after (T>0) blood that was culture [+] for Candida spp. were retrieved from the diagnostic haematology laboratory. DNA was extracted and MT-PCR performed. Results were correlated with standard yeast identification. Results: (Table) The MT-PCR assay detection limit was 10 cfu/ml of blood with a turn around time ≤ 2 h. Samples were available from 43 patients (n=65 time points). MT-PCR was positive in 24/42 (57%) patients with blood drawn at T≤0 (up to -4 days) and 28/43 (65%) before BC flagged positive. 7/14 (50%) patients on antifungal (AF) therapy and 31/51 (61%) not, were MT-PCR positive. MT-PCR sensitivity may have been compromised by storage of blood specimens, suboptimal sample volumes and variable fungal load in IC. Sensitivity was improved by increasing the blood sample volume. Unexplained discrepancies in species identification occurred in 3 patients (7%). Since there was no cross-reactivity of MT-PCR with 100 control specimens and DNA from 48 clinically relevant organisms, discrepancies are likely due to culture misidentification. Conclusions: MT-PCR is a cost effective and novel diagnostic tool for early diagnosis of fungemia.
No. patients (%)
Timing of blood draws relative to BC [+] sampleTotalMT-PCR posProven IC at time of blood drawnOn AFOn AF
& MT-PCR pos
T<02916 (55)0 (0)4 (14)2 (50)
T=0138 (62)0 (0)1 (8)0 (0)
T>02314 (61)13 (57)9 (39)5 (56)
Total6538 (58)13 (20)14 (22)7 (50)
Anna Lau1, Catriona Halliday2, E Playford3, Keith Stanley4, Sharon Chen, MD5, Tania Sorrell1 and  A. Lau, None., (1)PSIFIC Study Group, (2)CIDM Westmead Hosp., (3)Princess Alexandra Hosp., (4)AusDiagnostics, (5)Westmead Hospital, Westmead, NSW, Australia

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