LightCycler SeptiFast technology in patients with solid malignancies clinical utility for rapid etiologic diagnosis of sepsis

LightCycler SeptiFast technology in patients with solid malignancies clinical utility for rapid etiologic diagnosis of sepsis Dubská et al. Critical Care 2012, 16:404 LightCycler SeptiFast technology in patients with solid malignancies: clinical utility for rapid etiologic diagnosis of sepsis Lenka Dubská *, Martina Vysko?ilová , Dagmar Mina?íková , Petr Jelínek , Renata Tejkalová and Dalibor Valík 1 1 2 2 3 1 Recent development of molecular tools for pathogen analysis every working day during a day shift. In such detection introduced the possibility of early targeted conditions, the turnaround time of SeptiFast results for antimicrobial treatment that, in turn, may improve the samples delivered to the laboratory on a workday outcome of patients with sepsis [1]. between 7 am and 2 pm was as follows: 100% of samples, We retrospectively evaluated 54 results of the SeptiFast 10 hours; 97% of samples, 8 hours; 59% of samples, test from patients with solid malignancy admitted to the 6 hours e average turn around time for tests performed ICU between June 2009 and August 2011. Specimens with manual DNA isolation was 6 hours 11 minutes; by from suspected bloodstream infection were analyzed implementation of automated isolation of DNA using the using LightCycler SeptiFast (Roche Molecular Diagnostics, MagNA Pure Compact System [4], the mean turnaround Prague, Czech Republic) according to the manufacturer’s time was shortened to 5 hours 17 minutes. In conclusion, detection of pathogen by the LightCycler instructions and evaluated in comparison with blood culture results obtained from blood sampled no longer SeptiFast system is generally not inferior to the results than 24 hours before or after sampling for SeptiFast. from blood cultur e added value of multiplex DNA Blood culturing and identi, cation were performed ampli, cation-based pathogen detection is the shorter according to the routine diagnostic procedures e total test turnaround time and probably the increased true number of complementary blood cultures analyzed was sensitivity of detection of certain pathogens. Moreover, 85. Consistently, negative results from SeptiFast and in clinical settings pathogen detection may be required blood culture were obtained in 21 (39%) cases. after an antibiotic administration; in such conditions, To assess the true positivity of both microbiological cultivation independent methods are of clear bene, t [5]. methods, discrepant cases were evaluated in the context Competing interests of clinical and laboratory , ndings by two independent Reimbursement from Roche Diagnostics unrelated to the present work: physicians experienced in critical care (Figure e in 2008, DV and LD for a lecture on a pharmacogenomic and predictive oncology topic, respectively, and in 2010, MV and LD for an expertise work clinically relevant presence of a pathogen detected by blood culture but not by SeptiFast was recorded for on a protocol for automated DNA isolation from formalin-, xed para, n- embedded tissues. Klebsiella pneumoniae/oxytoca, and in both cases the presence of another member of Enterobacteriaceae was Authors’ contributions LD, MV, RT and DV participated in the design of the study. LD and MV reported by SeptiFast, Escherichia coli and Enterobacter carried out the molecular genetics part of the study. DM and PJ evaluated cloacae/aerogenes – thus misidenti, cation of strains with the microbiological , ndings in the context of clinical status. All authors participated in writing the report. atypical phenotype cannot be excluded [3]. Our results also show that SeptiFast is more e, cient in detection of Acknowledgements The authors received a contribution for the reagents, equipment from Roche clinically relevant infection by E. coli and Pseudomonas Diagnostics for this project, and were supported by the European Regional aeruginosa. Development Fund and the State budget of the Czech Republic for RECAMO e turnaround time for blood culture analysis is 24 to (CZ.1.05/2.1.00/03.0101). 48 hours e work, ow in our laboratory allows SeptiFast Author details 1 Department of Laboratory Medicine, Masaryk Memorial Cancer Institute, Zluty kopec 7, 656 53 Brno, Czech Republic. Department of Anesthesiology *Correspondence: dubska@mou.cz 1 2 Department of Laboratory Medicine, Masaryk Memorial Cancer Institute, Zluty and Intensive Care, Masaryk Memorial Cancer Institute, Zluty kopec 7, 656 53 kopec 7, 656 53 Brno, Czech Republic 3 Full list of author information is available at the end of the article Brno, Czech Republic. Department of Microbiology, St. Anne’s Faculty Hospital and Faculty of Medicine, Masaryk University, Pekarska 53, 664/53, 656 91 Brno, ? 2010 BioMed Central Ltd ? 2012 BioMed Central Ltd Published: 17 January 2012Czech Republic. Page 2 of 3 Dubská et al. Critical Care 2012, 16:404 G- unclear unclear E. coli P. aeruginosa K. pneumoniae/oxytoca unclear unclear E. cloacae/aerogenes S. maltophilia G+ unclear S. aureus unclear unclear unclear CoNS unclear E. faecalis F unclear unclear C. albicans BC only BC+SF SF only Figure 1. Scheme summarizing detection of pathogens together with clinical evaluation of discrepant ,ndings. Totally 47 pathogens were detected in 29 SeptiFast (SF) and/or 43 blood culture (BC) samples. From 20 pathogens listed on the SF menu [2], nine were detected in the present study and all pathogens detected by blood culture were on the SF menu, 18 microorganisms were detected by both BC and SF, 12 by BC only, and 18 by SF only. The clinical evaluation is as follows: each pathogen detected by either method was classi,ed as possible infection (based on clinical data; for example, bene,t from antimicrobial therapy), as probable infection (based on clinical data and cultivation of the pathogen from other site), as highly probable infection (based on clinical data and presence of the pathogen in blood within the relevant time window), as unlike infection (for example, contamination) or as undetermined. To obtain orientation in false positive and false negative results in pathogen detection by SF and cultivation, we assigned a score value to each positive classi,cation as follows: possible infection (score 2), probable infection (score 3), and highly probable infection (score 4). Black boxes represent summary of cases when pathogen was detected by both methods, SF and BC. Red boxes left of concordant events show detection by BC only, and blue objects right of black squares represent events of pathogen detection by SF only. Empty boxes, cases evaluated by both ICU experts as unlike infection; unclear, cases evaluated by ICU experts discrepantly (for example, unlike vs. possible or probable infection). The size of ,lled objects together with abbreviated classi,cation describe the evaluation of ICU experts for each case: h.pr., highly probable infection; pr., probable infection; po., possible infection. Coagulase-negative staphylococci (CoNS) were detected far more often by BC than SF, probably as a result of contamination of blood culture sample by staphylococci from normal skin ,ora. Regarding CoNS, SF is able to re,ect the amount of CoNS DNA template; if its level is below the discrimination threshold (crossing point (CP) value of ampli,cation growth curve >20), the result is reported as negative. In this context, we retrospectively evaluated all SF raw data ,les for the CoNS ampli,cation curve and its CP. The CoNS curve with CP >20 was present in 15 out of 44 (34%) SF staphylococci-negative with a lowest CP of 21.4, supporting the necessity for the discrimination threshold in the SF evaluation algorithm for commensal bacteria. Page 3 of 3 Dubská et al. Critical Care 2012, 16:404 5. Yanagihara K, Kitagawa Y, Tomonaga M, Tsukasaki K, Kohno S, Seki M, References 1. 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