Comparison of whole blood, serum, and plasma for early detection of candidemia by multiplex-tandem PCR

Abstract

We applied multiplex-tandem PCR (MT-PCR) to 255 EDTA whole-blood specimens, 29 serum specimens, and 24 plasma specimens from 109 patients with Candida bloodstream infection (candidemia) to determine whether a diagnosis could be expedited in comparison with the time to diagnosis by the use of standard blood culture. Overall, the MT-PCR performed better than blood culture with DNA extracted from whole blood from 52/74 (70%) patients, accelerating the time to detection (blood culture flagging) and determination of the pathogenic species (by use of the API 32C system [bioMérieux, Marcy l'Etoile, France]) by up to 4 days (mean, 2.2 days; range, 0.5 to 8 days). Candida DNA was detected more often in serum (71%) and plasma (75%) than in whole blood (54%), although relatively small numbers of serum and plasma specimens were tested. The sensitivity, specificity, positive predictive value, and negative predictive value of the assay with whole blood were 75%, 97%, 95%, and 85%, respectively. Fungal DNA was not detected by MT-PCR in 6/24 (25%) whole-blood samples drawn simultaneously with the positive blood culture sample. MT-PCR performed better with whole-blood specimens stored at -20 degrees C (75%) and when DNA was extracted within 1 week of sampling (66%). The molecular and culture identification results correlated for 61 of 66 patients (92%); one discrepant result was due to misidentification by culture. All but one sample from 53 patients who were at high risk of candidemia but did not have proven disease were negative by MT-PCR. The results demonstrate the good potential of MT-PCR to detect candidemia, to provide Candida species identification prior to blood culture positivity, and to provide improved sensitivity when applied to with serum and plasma specimens.

FIG. 1.

Results of MT-PCR detection for 109 candidemic patients which have been stratified for time of collection relative to the time of collection of the first blood sample that flagged positive by culture (T = 0); i.e., those obtained before the time of collection of the blood sample that flagged positive by blood culture (BC) (T < 0), simultaneously with the time of collection of the blood sample that flagged positive by blood culture (T = 0), or after collection of the blood sample that flagged positive by blood culture (T > 0). T > 0 flag+, blood samples drawn after the blood culture had flagged positive. +ve, positive; WB, whole blood. Note that samples were collected from some patients at multiple time points.

FIG. 2.

Percentage of MT-PCR-positive whole-blood samples stratified by collection times relative to the time that the first positive blood culture (BC) sample was drawn (T = 0) for 255 specimens collected from 109 candidemic patients.

FIG. 3.

Effect of storage on MT-PCR detection of fungi from 85 whole-blood specimens from 62 candidemic patients.