Utility of a Commercially Available Multiplex Real-Time PCR Assay To Detect Bacterial and Fungal Pathogens in Febrile Neutropenia

Abstract

Infection is the main treatment-related cause of mortality in cancer patients. Rapid and accurate diagnosis to facilitate specific therapy of febrile neutropenia is therefore urgently warranted. Here, we evaluated a commercial PCR-based kit to detect the DNA of 20 different pathogens (SeptiFast) in the setting of febrile neutropenia after chemotherapy. Seven hundred eighty-four serum samples of 119 febrile neutropenic episodes (FNEs) in 70 patients with hematological malignancies were analyzed and compared with clinical, microbiological, and biochemical findings. In the antibiotic-naïve setting, bacteremia was diagnosed in 34 FNEs and 11 of them yielded the same result in the PCR. Seventy-three FNEs were negative in both systems, leading to an overall agreement in 84 of 119 FNEs (71%). During antibiotic therapy, positivity in blood culture occurred only in 3% of cases, but the PCR yielded a positive result in 15% of cases. In six cases the PCR during antibiotic treatment detected a new pathogen repetitively; this was accompanied by a significant rise in procalcitonin levels, suggestive of a true detection of infection. All patients with probable invasive fungal infection (IFI; n ‫؍‬ 3) according to the standards of the European Organization for Research and Treatment of Cancer had a positive PCR result for Aspergillus fumigatus; in contrast there was only one positive result for Aspergillus fumigatus in an episode without signs and symptoms of IFI. Our results demonstrate that the SeptiFast kit cannot replace blood cultures in the diagnostic workup of FNEs. However, it might be helpful in situations where blood cultures remain negative (e.g., during antimicrobial therapy or in IFI).While systemic infection is the most common cause of a febrile neutropenia episode (FNE) with significant effects on morbidity and mortality, only 30% of blood cultures taken at the onset of fever are positive (11,15). Nonetheless, patients with FNEs are treated with broad-spectrum antimicrobial agents regardless of the result of their blood culture (7) because potentially life-threatening infections need early treatment to ensure better clinical outcome. Noninfective causes of a systemic reaction culminating in a rise in temperature such as tumor fever, drug fever, or transfusion reactions complicate the diagnostic challenge in cancer patients. In addition, the etiology of a deterioration of an FNE during antimicrobial therapy is often difficult to elucidate, since blood cultures are infrequently positive once effective antimicrobial therapy has started (4). Pathogens such as molds which are rarely found in blood cultures are not uncommon in patients with FNEs, particularly if they suffer from hematological malignancies. For these reasons, FNE is one of the conditions where new diagnostic tools to distinguish an infection from a nonmicrobial cause for fever or to identify rare pathogens are most urgently needed. In the past, raised levels of indirect markers such as procalcitonin (PCT) and interleukin 6 (3, 16) have been shown to be ...

Microbiological Diagnosis of Sepsis: Polymerase Chain Reaction System Versus Blood Cultures

“…In some of these studies, the SeptiFast assay was found to be clearly superior, 9,11,21,22 whereas in other studies, both methods displayed either similar sensitivities or the SeptiFast assay compared unfavorably with respect to blood culture. 12,[23][24][25] Differences between studies in terms of patients' characteristics, the spectrum of microorganisms detected, the number of blood culture sets collected, the volume of blood collected for culture, and the presence or absence of ongoing antibiotic therapy at the time of sampling may account for these discrepancies. Inappropriate antimicrobial treatment or delays in starting appropriate treatment are both associated with increased morbidity and mortality in sepsis, particularly in severe cases.…”

Detection, Identification, and Distribution of Fungi in Bronchoalveolar Lavage Specimens by Use of Multilocus PCR Coupled with Electrospray Ionization/Mass Spectrometry

“…Among them, fungal antigen testing is convenient and fast, but it lacks sensitivity and specificity, restricting its use in clinical laboratories. Molecular methods using PCR and real-time PCR techniques are useful for the diagnosis of fungal infection but can detect only one or a few specific pathogens (7,8).…”

Molecular diagnosis of polymicrobial newborn sepsis by multiplex real-time PCR using a small volume of blood sample

“…Recently available molecular assays aid rapid detection of micro-organisms and improve the diagnostic flow-chart (Peters et al, 2004;Gaibani et al, 2009). Among these, a commercial multiplex real-time PCR (LightCycler SeptiFast Test; Roche Molecular Systems) uses a novel technology that enables the direct detection from blood samples of a wide panel of bacterial and fungal microorganisms commonly involved in systemic infections (Lehmann et al, 2008;von Lilienfeld-Toal et al, 2009). This technique was found to give promising results in the neonatal population with sepsis, particularly when antibiotic treatment is initiated (Mussap et al, 2007;Paolucci et al, 2009;Lucignano et al, 2011).…”