Bacteriology Use of multiplex PCR in diagnosis of bloodstream infections in kidney patients Natalia Fernández-Romero a, 1 , Inmaculada Quiles a, 1 , Carlos Jiménez b , María Ovidea Lopez Oliva b , Begoña Rivas b , Jesús Mingorance a , María Pilar Romero-Gómez a, ⁎ a Servicio de Microbiología, Hospital Universitario La Paz, IdiPAZ, Paseo de la Castellana, 261, 28046, Madrid, Spain b Servicio de Nefrología, Hospital Universitario La Paz, IdiPAZ, Paseo de la Castellana, 261, 28046, Madrid, Spain abstract article info Article history: Received 26 January 2014 Received in revised form 5 June 2014 Accepted 2 July 2014 Available online 16 July 2014 Keywords: Multiplex PCR for diagnostic of sepsis in kidney patients The LightCycler® SeptiFast Test (Roche Diagnostics GmbH, Mannheim, Germany) was prospectively compared with the standard blood culture technique in a series of 86 kidney patients. The sensitivity of the PCR compared with the culture was 71%, and the specificity was 88%. All the species identified by culture in these patients were in the SeptiFast panel. The median time to results was 1 day for the PCR, 3 days for positive cultures, and 5 days for negative cultures. © 2014 Elsevier Inc. All rights reserved. Bloodstream infections (BSIs) are an important cause of morbidity and mortality in kidney patients (Abbott et al., 2001), septic shock being the most severe complication and increasing overall mortality (Harbarth et al., 2002). The case fatality rate from BSI causing organ dysfunction ranges from 25% to 50% (Palmer et al., 2000). BSI is frequently a consequence of urinary tract infection, with a high prevalence of gram-negative bacilli (Chuang et al., 2005; Merçon et al., 2010). Microbiological and epidemiological data indicate that a limited number of bacteria and fungi are responsible for the majority of all BSIs in kidney transplant recipients and hemodialysis patients (Al-Hasan et al., 2009, 2011; Li and Chow, 2012; Silva et al., 2010). Early detection and adequate treatment of causative pathogens within the first 6–12 h is critical for a favorable outcome in patients with BSI (Garnacho-Montero et al., 2003; Morrell et al., 2005), but the microbiological “gold standard” for the detection and identification of bacterial and fungal BSIs is the blood culture (BC), which is limited by high volume requirements and prolonged incubation times. In an effort to address some of these limitations, many advances have been developed to improve the sensitivity and reduce the time to identification of pathogens in blood cultures (Romero-Gómez, 2011; Romero-Gómez et al., 2012). Molecular amplification techniques have been developed to detect the pathogens directly on blood (Haag et al., 2013; Loonen et al., 2014; Samuel et al., 2013; Schreiber et al., 2013; Wojewoda et al., 2013), replacing the incubation step in blood culture, but their use is limited by some important technical drawbacks: 1) detection and identification can be achieved in one step by amplification of universal targets (usually 16S, 23S rRNA genes or their associated ITS regions) and multiplexing, but it is difficult to span the broad range of pathogens that can be found in blood cultures and maintain accuracy in species identification without adding lengthy sequenc- ing or hybridization steps; 2) bacterial loads in blood may be as low as a few colony-forming units per milliliter, so the methods must be very sensitive, but then environmental and reagent contamination with bacterial DNA becomes a serious concern. The LightCycler® SeptiFast Test (Roche Diagnostics GmbH, Mannheim, Germany) is a commercial multiplex real-time PCR assay that uses DNA-free reagents (M Grade) to detect directly in whole blood any of a panel of 25 of the most frequent pathogens (bacteria and fungi) found in blood cultures (Table S1) (Lehmann et al., 2008). The assay has been tested in blood samples from diverse patient groups, including intensive care patients (Lodes et al., 2012; Regueiro et al., 2010; Yanagihara et al., 2010), transplant recipients (Rath et al., 2012), pediatric patients (Torres-Martos et al., 2012), and even in samples different from blood (Fernández et al., 2010; Mencacci et al., 2011). The microorganisms detected by SeptiFast include the most common pathogens found in positive blood cultures from kidney patients suggesting that the technique might be especially well suited for this patient group. On this ground, the objective of this study was to evaluate the utility of the LightCycler® SeptiFast multiplex real- time PCR assay compared to those of conventional blood culture for the diagnosis of BSIs in renal patients. Diagnostic Microbiology and Infectious Disease 80 (2014) 93–96 ⁎ Corresponding author. Tel.: +34-912071842; fax: +34-917277372. E-mail address: mromerogomez@salud.madrid.org (M.P. Romero-Gómez). 1 These 2 authors contributed equally. http://dx.doi.org/10.1016/j.diagmicrobio.2014.07.001 0732-8893/© 2014 Elsevier Inc. All rights reserved. Contents lists available at ScienceDirect Diagnostic Microbiology and Infectious Disease journal homepage: www.elsevier.com/locate/diagmicrobio