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Microbiological diagnostics of bloodstream infections in Europe—an ESGBIES survey

Open ArchivePublished:April 10, 2019DOI:https://doi.org/10.1016/j.cmi.2019.03.024

      Abstract

      Objectives

      High-quality diagnosis of bloodstream infections (BSI) is important for successful patient management. As knowledge on current practices of microbiological BSI diagnostics is limited, this project aimed to assess its current state in European microbiological laboratories.

      Methods

      We performed an online questionnaire-based cross-sectional survey comprising 34 questions on practices of microbiological BSI diagnostics. The ESCMID Study Group for Bloodstream Infections, Endocarditis and Sepsis (ESGBIES) was the primary platform to engage national coordinators who recruited laboratories within their countries.

      Results

      Responses were received from 209 laboratories in 25 European countries. Although 32.5% (68/209) of laboratories only used the classical processing of positive blood cultures (BC), two-thirds applied rapid technologies. Of laboratories that provided data, 42.2% (78/185) were able to start incubating BC in automated BC incubators around-the-clock, and only 13% (25/192) had established a 24-h service to start immediate processing of positive BC. Only 4.7% (9/190) of laboratories validated and transmitted the results of identification and antimicrobial susceptibility testing (AST) of BC pathogens to clinicians 24 h/day. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry from briefly incubated sub-cultures on solid media was the most commonly used approach to rapid pathogen identification from positive BC, and direct disc diffusion was the most common rapid AST method from positive BC.

      Conclusions

      Laboratories have started to implement novel technologies for rapid identification and AST for positive BC. However, progress is severely compromised by limited operating hours such that current practice of BC diagnostics in Europe complies only partly with the requirements for optimal BSI management.

      Keywords

      Introduction

      The importance of high-quality diagnosis of bloodstream infections (BSI) is well-recognized [
      • Dubourg G.
      • Lamy B.
      • Ruimy R.
      Rapid phenotypic methods to improve the diagnosis of bacterial bloodstream infections: meeting the challenge to reduce the time to result.
      ,
      • Opota O.
      • Croxatto A.
      • Prod'hom G.
      • Greub G.
      Blood culture-based diagnosis of bacteraemia: state of the art.
      ,
      • Rhodes A.
      • Evans L.E.
      • Alhazzani W.
      • Levy M.M.
      • Antonelli M.
      • Ferrer R.
      • et al.
      Surviving Sepsis campaign: international guidelines for management of sepsis and septic shock: 2016.
      ,
      • Idelevich E.A.
      • Reischl U.
      • Becker K.
      New microbiological techniques in the diagnosis of bloodstream infections.
      ,
      • Habib G.
      • Lancellotti P.
      • Antunes M.J.
      • Bongiorni M.G.
      • Casalta J.P.
      • Del Z.F.
      • et al.
      2015 ESC guidelines for the management of infective endocarditis: the task force for the management of infective endocarditis of the European Society of Cardiology (ESC). Endorsed by: European Association for Cardio-Thoracic Surgery (EACTS), the European Association of Nuclear Medicine (EANM).
      ,
      • Lamy B.
      • Ferroni A.
      • Henning C.
      • Cattoen C.
      • Laudat P.
      How to: accreditation of blood cultures' proceedings. A clinical microbiology approach for adding value to patient care.
      ]; however, European guidelines on microbiological BSI diagnostics do not exist. Unified quality standards are unavailable and current practices of microbiological BSI diagnosis appear heterogeneous across European countries.
      Rapid pathogen identification and antimicrobial susceptibility testing (AST) are crucial for targeted management of BSIs [
      • Idelevich E.A.
      • Silling G.
      • Niederbracht Y.
      • Penner H.
      • Sauerland M.C.
      • Tafelski S.
      • et al.
      Impact of multiplex PCR on antimicrobial treatment in febrile neutropenia: a randomized controlled study.
      ,
      • Kerremans J.J.
      • Verboom P.
      • Stijnen T.
      • Hakkaart-van R.L.
      • Goessens W.
      • Verbrugh H.A.
      • et al.
      Rapid identification and antimicrobial susceptibility testing reduce antibiotic use and accelerate pathogen-directed antibiotic use.
      ,
      • Köck R.
      • Wüllenweber J.
      • Horn D.
      • Lanckohr C.
      • Becker K.
      • Idelevich E.A.
      Implementation of short incubation MALDI-TOF MS identification from positive blood cultures in routine diagnostics and effects on empiric antimicrobial therapy.
      ,
      • Beekmann S.E.
      • Diekema D.J.
      • Chapin K.C.
      • Doern G.V.
      Effects of rapid detection of bloodstream infections on length of hospitalization and hospital charges.
      ,
      • van Belkum A.
      • Bachmann T.T.
      • Ludke G.
      • Lisby J.G.
      • Kahlmeter G.
      • Mohess A.
      • et al.
      Developmental roadmap for antimicrobial susceptibility testing systems.
      ]. However, conventional microbiological methods require 48–72 h to provide a final species identification and AST report [
      • Dubourg G.
      • Lamy B.
      • Ruimy R.
      Rapid phenotypic methods to improve the diagnosis of bacterial bloodstream infections: meeting the challenge to reduce the time to result.
      ,
      • Opota O.
      • Croxatto A.
      • Prod'hom G.
      • Greub G.
      Blood culture-based diagnosis of bacteraemia: state of the art.
      ,
      • Kerremans J.J.
      • Verboom P.
      • Stijnen T.
      • Hakkaart-van R.L.
      • Goessens W.
      • Verbrugh H.A.
      • et al.
      Rapid identification and antimicrobial susceptibility testing reduce antibiotic use and accelerate pathogen-directed antibiotic use.
      ,
      • Tabak Y.P.
      • Vankeepuram L.
      • Ye G.
      • Jeffers K.
      • Gupta V.
      • Murray P.R.
      Blood culture turnaround time in US acute care hospitals and implications for laboratory process optimization.
      ]. Some rapid diagnostic technologies, e.g. molecular methods [
      • Idelevich E.A.
      • Silling G.
      • Niederbracht Y.
      • Penner H.
      • Sauerland M.C.
      • Tafelski S.
      • et al.
      Impact of multiplex PCR on antimicrobial treatment in febrile neutropenia: a randomized controlled study.
      ] and rapid culture methods supported by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) [
      • Idelevich E.A.
      • Schüle I.
      • Grünastel B.
      • Wüllenweber J.
      • Peters G.
      • Becker K.
      Rapid identification of microorganisms from positive blood cultures by MALDI-TOF mass spectrometry subsequent to very short-term incubation on solid medium.
      ] have been suggested to speed up microbiological BSI diagnosis [
      • Dubourg G.
      • Lamy B.
      • Ruimy R.
      Rapid phenotypic methods to improve the diagnosis of bacterial bloodstream infections: meeting the challenge to reduce the time to result.
      ,
      • Opota O.
      • Croxatto A.
      • Prod'hom G.
      • Greub G.
      Blood culture-based diagnosis of bacteraemia: state of the art.
      ,
      • Idelevich E.A.
      • Reischl U.
      • Becker K.
      New microbiological techniques in the diagnosis of bloodstream infections.
      ], but data about their use and acceptance in routine practice are lacking [
      • Arena F.
      • Argentieri M.
      • Bernaschi P.
      • Fortina G.
      • Kroumova V.
      • Manso E.
      • et al.
      Real life turnaround time of blood cultures in the clinical microbiology laboratory: results of the first Italian survey, May 2015.
      ,
      • Schmitz R.P.
      • Keller P.M.
      • Baier M.
      • Hagel S.
      • Pletz M.W.
      • Brunkhorst F.M.
      Quality of blood culture testing—a survey in intensive care units and microbiological laboratories across four European countries.
      ]. The transportation time of microbiological specimens is an underestimated pre-analytical factor, which is critical for the clinical impact of microbiological diagnostics, in particular if rapid tests are applied [
      • Kerremans J.J.
      • Verboom P.
      • Stijnen T.
      • Hakkaart-van R.L.
      • Goessens W.
      • Verbrugh H.A.
      • et al.
      Rapid identification and antimicrobial susceptibility testing reduce antibiotic use and accelerate pathogen-directed antibiotic use.
      ,
      • Blondeau J.M.
      • Idelevich E.A.
      The 24-h clinical microbiology service is essential for patient management.
      ,
      • Morton B.
      • Nagaraja S.
      • Collins A.
      • Pennington S.H.
      • Blakey J.D.
      A retrospective evaluation of critical care blood culture yield—do support services contribute to the "Weekend Effect"?.
      ,
      • Rönnberg C.
      • Mildh M.
      • Ullberg M.
      • Özenci V.
      Transport time for blood culture bottles: underlying factors and its consequences.
      ,
      • Schneiderhan W.
      • Grundt A.
      • Wörner S.
      • Findeisen P.
      • Neumaier M.
      Work flow analysis of around-the-clock processing of blood culture samples and integrated MALDI-TOF mass spectrometry analysis for the diagnosis of bloodstream infections.
      ]. On behalf of the ESCMID Study Group for Bloodstream Infections, Endocarditis and Sepsis (ESGBIES), we performed a survey that aimed to assess the current state of microbiological BSI diagnostics in European laboratories.

      Materials and methods

       Design and sampling strategy

      The survey was designed and performed according to the recent methodological recommendations for surveys [
      • Pulcini C.
      • Leibovici L.
      CMI guidance for authors of surveys.
      ]. ESGBIES was the primary platform to recruit national coordinators. The cross-sectional survey used a snowballing technique for sampling, with each national coordinator contacting approximately ten laboratories within the country. At least half of the included laboratories should be university hospital laboratories with the remainder being municipal/regional/peripheral hospital laboratories, or those serving several hospitals and/or outpatient facilities. The participating laboratories within each country had to have a reasonable geographical distribution.

       Questionnaire and analysis

      A questionnaire comprising 34 items (see Supplementary material, Text S1) was developed and tested for content validity by the project coordinators. It included single choice, multiple choice and open questions on laboratory set-up and on current practices of microbiological diagnostics of BSI. The internet-based questionnaire was created using Google Forms and was completed online. The responses were anonymized. Descriptive statistics were performed using Excel (Microsoft, Redmond, WA, USA) for all items of the questionnaire. Analysis for individual countries was performed only when at least seven laboratories per country provided a response to a particular question.

      Results

       Recruited laboratories and response rates

      National coordinators were initially recruited from 28 countries; responses were ultimately received from 25 countries (Fig. 1a), resulting in a response rate of 89.3% among the initially invited countries. Of 238 laboratories from those 25 countries, 209 submitted their responses (Fig. 1a) between 14 October 2016 and 23 November 2017. This accounts for a response rate of 87.8% of the initially invited laboratories. The distribution of laboratories according to their types is shown in Fig. 1b.
      Fig. 1
      Fig. 1Participating laboratories (n = 209) per country (n = 25) (a) and distribution of laboratories (n = 209) according to their types (b).

       Survey results

      The numbers of blood culture (BC) bottles and sets, respectively, received by laboratories per 100 hospitalizations and per 100 beds/year are depicted in the Supplementary material (Fig. S1). Only 39.7% (83/209) of respondents indicated that at least two BC sets are taken for the initial diagnostic evaluation of a fever episode in the vast majority of cases (defined as ≥75% of cases) (Fig. 2a). The majority (91.9%; 192/209) of laboratories use a single pair of aerobic and anaerobic vials as a standard BC set (Fig. 2b).
      Fig. 2
      Fig. 2Estimated number of blood cultures taken initially per fever episode (a) and the composition of standard blood culture sets in the majority (>80%) of cases (b).
      Only 33.5% (70/209) of all laboratories regularly monitor the volume of blood in BC bottles, with the mean estimated blood volume being 8.1 ± 2.2 (standard deviation, SD) mL/bottle. The most common incubation time for standard BC bottles was 5 days, implemented by 69.9% of laboratories (146/209; including one laboratory at 5.5 days), followed by 7 days (20.1%; 42/209) and 6 days (6.7%; 14/209; including one laboratory at 6.5 days). Other laboratories reported 4 days (1.0%; 2/209) or 10 and 14 days, respectively (each 0.5%; 1/209); no response was received from 1.4% of laboratories (3/209). The BC positivity rates are shown in Table 1.
      Table 1Blood culture positivity rates (any organism, potential contaminants included)
      DataBlood culture positivity rates
      As proportion of positive BC setsAs proportion of positive BC bottles
      Mean BC positivity rate14.1%13.8%
      Number (%) of laboratories which provided data for that particular analysis162/209 (77.5%)159/209 (76.1%)
      Abbreviation: BC, blood culture.
      Of laboratories that provided data, 42.2% (78/185) are able to start blood cultivation in automated BC incubators around-the-clock (Table 2). Laboratories with time-restricted coverage can insert the BC bottles into automated BC incubators on average only 10.5 ± 2.5 (SD) h/day on weekdays, with coverage on Saturdays and Sundays/holidays being even less (Table 2). Overall, 9.3% (17/183) of laboratories do not accept BC specimens on Sundays and holidays at all (Table 2); 59.3% (124/209), 26.3% (55/209) and 10.5% (22/209) of respondents assumed that the majority of BC (>80%) arrive in their laboratories within 4, 12 or 24 h of BC draw, respectively (see Supplementary material, Fig. S2a).
      Table 2Percentage of laboratories with 24-h, time-restricted or no (0-h) service for blood culture diagnostics
      Calculated as % of all laboratories which provided data for that particular question.
      Day of the weekStarting incubation of BC bottlesProcessing of positive BCsValidation of ID and AST results
      Validating and transmitting the results of identification and antimicrobial susceptibility testing of blood culture isolates to the clinicians.
      n
      n, number of laboratories that provided data for that particular question.
      Percentage of laboratories withCoverage of time-restricted service
      Mean time coverage per day for laboratories with time-restricted service.
      N
      n, number of laboratories that provided data for that particular question.
      Percentage of laboratories withCoverage of time-restricted service
      Mean time coverage per day for laboratories with time-restricted service.
      n
      n, number of laboratories that provided data for that particular question.
      Percentage of laboratories withCoverage of time-restricted service
      Mean time coverage per day for laboratories with time-restricted service.
      24-h serviceTime-restricted serviceNo service24-h serviceTime-restricted serviceNo service24-h serviceTime-restricted serviceNo service
      Monday–Friday18542.2%57.8%0.0%10.5 h19213.0%87.0%0.0%10.3 h1904.7%95.3%0.0%9.3 h
      Saturday18441.3%58.7%0.0%8.1 h19112.6%85.9%1.6%7.8 h1904.2%91.6%4.2%7.2 h
      Sunday/holiday18341.5%49.2%9.3%7.4 h19112.6%72.3%15.2%7.1 h1904.2%74.2%21.6%6.3 h
      Abbreviation: AST, antimicrobial susceptibility testing; BC, blood culture; ID, identification.
      a Calculated as % of all laboratories which provided data for that particular question.
      b n, number of laboratories that provided data for that particular question.
      c Validating and transmitting the results of identification and antimicrobial susceptibility testing of blood culture isolates to the clinicians.
      d Mean time coverage per day for laboratories with time-restricted service.
      Only 13% (25/192) of laboratories provide 24-h service to start immediate processing of BC bottles that have flagged positive. Laboratories that have only limited service times can take action on positive BCs 10.3 ± 2.5 (SD) h/day on weekdays with even shorter coverage on Saturdays and Sundays/holidays (Table 2); 15.2% (29/191) of laboratories do not handle positive BCs on Sundays and holidays at all (Table 2). Fig. S3 (see Supplementary material) provides an overview of the service coverage of BC diagnostics in laboratories of different countries. Of 23 laboratories with 24-h service for both starting incubation in the automated systems and for processing of positive BCs, six (26.1%) estimated that >40% of BCs are received or processed during the out-of-hours period (e.g. between 7 pm and 7 am) and five (21.7%) estimated that the percentage of BCs received or processed out-of-hours is 30%–40% (see Supplementary material, Fig. S2b).
      Fewer than 5% of laboratories validate and transmit the results of identification and AST of BC pathogens to the clinicians 24 h/day; most laboratories report findings only during a limited time period; 21.6% (41/190) of laboratories generally do not forward results to the clinicians on Sundays (Table 2).
      Of all respondents, 94.7% (198/209) and 3.3% (7/209) use a semi-automated continuously monitoring BC system as the only or main type of BC diagnostics, respectively. Only 15.3% (32/209) of laboratories have the facility to load BC bottles into a semi-automated BC incubator outside the laboratory, whereas 9.1% (19/209) provide this opportunity only in some facilities served and 75.6% (158/209) do not offer this option at all. Semi-automated systems are not used by 1.9% (4/209) of laboratories. The other blood culturing methods reported included the lysis-centrifugation method with subsequent sediment culturing on agar, which was employed by 5/209 laboratories (3.8%), broth-based culturing with a positive result indicated by a signal device through pressure increase was used by 2/209 laboratories (1.0%) and one laboratory performed the Castaneda method.
      Gram staining remains standard in the vast majority of laboratories (always, 97.1% (203/209); in particular circumstances, 1.9% (4/209)) and the Gram stain result is immediately reported to clinicians by most laboratories (always, 90.8% (188/207); in particular circumstances, 4.3% (9/207)). In most laboratories (81.8%; 171/209), the Gram stain result is also the first finding that is usually reported to the clinicians. Of note, 14.8% (31/209) of laboratories first communicate results to clinicians only after rapid species identification has been obtained (Fig. 3).
      Fig. 3
      Fig. 3Providing the first report on a positive blood culture to the ward (a) and the ways to communicate (b, multiple answers were allowed for the latter part).
      Although 32.5% (68/209) of laboratories still use the classical diagnostic approach characterized by sub-cultivation of positively flagged BC bottles on solid media followed by overnight incubation and use of mature colonies for identification and AST the next day, 67.5% (141/209) apply various rapid diagnostic strategies. Both rapid identification and rapid AST are applied by 43.5% (91/209) of laboratories, 18.7% (39/209) apply only rapid identification and 5.3% (11/209) only rapid AST. MALDI-TOF MS from briefly incubated sub-cultures on solid media is currently the most used strategy (37.3%; 78/209) to achieve rapid pathogen identification from positive BCs, followed by direct latex agglutination for particular pathogens, direct MALDI-TOF identification (e.g. lysis-centrifugation procedure) and inoculation of semi-automated biochemical identification systems after a short sub-cultivation on agar plates (Fig. 4a). Direct disc diffusion is by far the most commonly used AST method from positive BCs, followed by direct inoculation of automated AST systems (Fig. 4b). For species identification from sub-cultivated positive BCs, 36.4% (76/209) of laboratories use MALDI-TOF MS preferentially to biochemical methods, 34.9% (73/209) use only biochemical methods and 24.9% (52/209) use only MALDI-TOF MS (Fig. 5). Direct identification of microbial DNA from whole blood is undertaken for all patients in 1.0% (2/209) of laboratories and for selected patients in 10.0% (21/209) of laboratories (Fig. 6).
      Fig. 4
      Fig. 4Rapid blood culture diagnostics. (a) Rapid identification technologies used after the blood culture bottle becomes positive (multiple answers were allowed). (b) Rapid antimicrobial susceptibility testing technologies used after the blood culture bottle becomes positive (multiple answers were allowed).
      Fig. 5
      Fig. 5Methods applied for species identification from sub-cultivated positive blood cultures.
      Fig. 6
      Fig. 6Use of direct identification of microbial DNA from whole blood (a) and reasons for selecting a patient for this method (b, among laboratories that apply this method for selected patients; multiple responses were allowed).
      The proportion of multidrug-resistant organisms among BC isolates is shown in Fig. S4 (see Supplementary material).
      Advice on antibiotic treatment is provided directly by laboratory staff in 79.4% (166/209) of microbiology laboratories; however, the practice of result communication to the infectious diseases department seems to vary greatly between the laboratories (see Supplementary material, Fig. S5).
      Most of the respondents (83.7%; 175/209) think that European guidelines for BC diagnostics would be useful, 12.4% (26/209) do not know whether it would be useful, and 3.8% (8/209) think it would not be useful. Most respondents (99.5%; 208/209) responded positively to the question of whether they would be interested in participating again.

      Discussion

      Rapid pathogen identification and AST are crucial for the appropriate management of BSIs [
      • Idelevich E.A.
      • Reischl U.
      • Becker K.
      New microbiological techniques in the diagnosis of bloodstream infections.
      ,
      • Idelevich E.A.
      • Silling G.
      • Niederbracht Y.
      • Penner H.
      • Sauerland M.C.
      • Tafelski S.
      • et al.
      Impact of multiplex PCR on antimicrobial treatment in febrile neutropenia: a randomized controlled study.
      ,
      • Kerremans J.J.
      • Verboom P.
      • Stijnen T.
      • Hakkaart-van R.L.
      • Goessens W.
      • Verbrugh H.A.
      • et al.
      Rapid identification and antimicrobial susceptibility testing reduce antibiotic use and accelerate pathogen-directed antibiotic use.
      ,
      • Köck R.
      • Wüllenweber J.
      • Horn D.
      • Lanckohr C.
      • Becker K.
      • Idelevich E.A.
      Implementation of short incubation MALDI-TOF MS identification from positive blood cultures in routine diagnostics and effects on empiric antimicrobial therapy.
      ,
      • Beekmann S.E.
      • Diekema D.J.
      • Chapin K.C.
      • Doern G.V.
      Effects of rapid detection of bloodstream infections on length of hospitalization and hospital charges.
      ,
      • van Belkum A.
      • Bachmann T.T.
      • Ludke G.
      • Lisby J.G.
      • Kahlmeter G.
      • Mohess A.
      • et al.
      Developmental roadmap for antimicrobial susceptibility testing systems.
      ]. However, numerous recent technological innovations are making it increasingly difficult to assess their implementation in routine diagnostics, necessitating respective surveys as the basis for future benchmarking and guideline development.
      The use of one aerobic and one anaerobic vial as standard BC set by the majority of laboratories is in accordance with the current recommendations [
      • Rhodes A.
      • Evans L.E.
      • Alhazzani W.
      • Levy M.M.
      • Antonelli M.
      • Ferrer R.
      • et al.
      Surviving Sepsis campaign: international guidelines for management of sepsis and septic shock: 2016.
      ]. However, the recommendation to take at least two initial BC sets [
      • Rhodes A.
      • Evans L.E.
      • Alhazzani W.
      • Levy M.M.
      • Antonelli M.
      • Ferrer R.
      • et al.
      Surviving Sepsis campaign: international guidelines for management of sepsis and septic shock: 2016.
      ] seems to be much less observed. As blood volume monitoring represents an important measure for quality control of BC practices [
      • Lamy B.
      • Ferroni A.
      • Henning C.
      • Cattoen C.
      • Laudat P.
      How to: accreditation of blood cultures' proceedings. A clinical microbiology approach for adding value to patient care.
      ], efforts should be made by laboratories to implement this procedure. The BC positivity rate was slightly higher than that reported elsewhere [
      • Tabak Y.P.
      • Vankeepuram L.
      • Ye G.
      • Jeffers K.
      • Gupta V.
      • Murray P.R.
      Blood culture turnaround time in US acute care hospitals and implications for laboratory process optimization.
      ,
      • Gubbels S.
      • Nielsen J.
      • Voldstedlund M.
      • Kristensen B.
      • Schønheyder H.C.
      • Vandenbroucke-Grauls C.M.
      • et al.
      Utilization of blood cultures in Danish hospitals: a population-based descriptive analysis.
      ]. The annual numbers of BCs received by laboratories were separately calculated using data for BC sets and BC bottles per 100 hospitalizations and per 100 beds. These data provide comparisons between the countries that can be used for international and national benchmarking of BC sampling rates [
      • Gubbels S.
      • Nielsen J.
      • Voldstedlund M.
      • Kristensen B.
      • Schønheyder H.C.
      • Vandenbroucke-Grauls C.M.
      • et al.
      Utilization of blood cultures in Danish hospitals: a population-based descriptive analysis.
      ,
      • Karch A.
      • Castell S.
      • Schwab F.
      • Geffers C.
      • Bongartz H.
      • Brunkhorst F.M.
      • et al.
      Proposing an empirically justified reference threshold for blood culture sampling rates in intensive care units.
      ,
      • Laupland K.B.
      • Niven D.J.
      • Pasquill K.
      • Parfitt E.C.
      • Steele L.
      Culturing rate and the surveillance of bloodstream infections: a population-based assessment.
      ,
      European Centre for Disease Prevention and Control
      Surveillance of antimicrobial resistance in Europe 2016.
      ].
      Our data demonstrate the most serious problem in current BC diagnostics, namely the insufficient service coverage by laboratories. BC diagnostics have been considered urgent [
      • Idelevich E.A.
      • Reischl U.
      • Becker K.
      New microbiological techniques in the diagnosis of bloodstream infections.
      ,
      • Habib G.
      • Lancellotti P.
      • Antunes M.J.
      • Bongiorni M.G.
      • Casalta J.P.
      • Del Z.F.
      • et al.
      2015 ESC guidelines for the management of infective endocarditis: the task force for the management of infective endocarditis of the European Society of Cardiology (ESC). Endorsed by: European Association for Cardio-Thoracic Surgery (EACTS), the European Association of Nuclear Medicine (EANM).
      ], which implies 24/7 availability of laboratory services [
      • Dubourg G.
      • Lamy B.
      • Ruimy R.
      Rapid phenotypic methods to improve the diagnosis of bacterial bloodstream infections: meeting the challenge to reduce the time to result.
      ,
      • Blondeau J.M.
      • Idelevich E.A.
      The 24-h clinical microbiology service is essential for patient management.
      ,
      • Schneiderhan W.
      • Grundt A.
      • Wörner S.
      • Findeisen P.
      • Neumaier M.
      Work flow analysis of around-the-clock processing of blood culture samples and integrated MALDI-TOF mass spectrometry analysis for the diagnosis of bloodstream infections.
      ]. Insufficient staffing certainly affects patient care, as the onset of BSI obviously does not necessarily match laboratories' working hours. Morton et al. reported significantly lower yields for BCs taken on weekends caused by the delays in BC incubation and processing [
      • Morton B.
      • Nagaraja S.
      • Collins A.
      • Pennington S.H.
      • Blakey J.D.
      A retrospective evaluation of critical care blood culture yield—do support services contribute to the "Weekend Effect"?.
      ]. Hence, laboratories probably contribute to the ‘weekend effect’, which is defined as an increase in adverse outcomes for patients admitted during weekends [
      • Morton B.
      • Nagaraja S.
      • Collins A.
      • Pennington S.H.
      • Blakey J.D.
      A retrospective evaluation of critical care blood culture yield—do support services contribute to the "Weekend Effect"?.
      ]. Furthermore, long periods of storage may lead to reduced detection rates [
      • Lamy B.
      • Ferroni A.
      • Henning C.
      • Cattoen C.
      • Laudat P.
      How to: accreditation of blood cultures' proceedings. A clinical microbiology approach for adding value to patient care.
      ,
      • Morton B.
      • Nagaraja S.
      • Collins A.
      • Pennington S.H.
      • Blakey J.D.
      A retrospective evaluation of critical care blood culture yield—do support services contribute to the "Weekend Effect"?.
      ]. Studies that exactly measure time from venepuncture to the arrival of a sample in the laboratory are scarce, but available publications report remarkably long transit times [
      • Idelevich E.A.
      • Silling G.
      • Niederbracht Y.
      • Penner H.
      • Sauerland M.C.
      • Tafelski S.
      • et al.
      Impact of multiplex PCR on antimicrobial treatment in febrile neutropenia: a randomized controlled study.
      ,
      • Kerremans J.J.
      • Verboom P.
      • Stijnen T.
      • Hakkaart-van R.L.
      • Goessens W.
      • Verbrugh H.A.
      • et al.
      Rapid identification and antimicrobial susceptibility testing reduce antibiotic use and accelerate pathogen-directed antibiotic use.
      ,
      • Rönnberg C.
      • Mildh M.
      • Ullberg M.
      • Özenci V.
      Transport time for blood culture bottles: underlying factors and its consequences.
      ]. Considerable service interruptions revealed in this survey in most of European laboratories allow the assumption that the proportion of BCs arriving substantially delayed in microbiology laboratories must be high. Köck et al. reported that 70% of BCs signalled positive by the automated incubator outside the laboratories' service hours [
      • Köck R.
      • Wüllenweber J.
      • Horn D.
      • Lanckohr C.
      • Becker K.
      • Idelevich E.A.
      Implementation of short incubation MALDI-TOF MS identification from positive blood cultures in routine diagnostics and effects on empiric antimicrobial therapy.
      ]. Another study revealed that, in 92% of cases, either the arrival of BC specimens in the laboratory or their positivity occurred outside regular duty hours [
      • Schneiderhan W.
      • Grundt A.
      • Wörner S.
      • Findeisen P.
      • Neumaier M.
      Work flow analysis of around-the-clock processing of blood culture samples and integrated MALDI-TOF mass spectrometry analysis for the diagnosis of bloodstream infections.
      ].
      Although slightly more than 40% of laboratories allowed 24/7 insertion of BC bottles into automated incubators, e.g. by placing these systems outside the laboratory, the percentage of laboratories that are able to start processing positive BC bottles around-the-clock is much lower. This fact is a matter of serious concern. Extended periods exist during which no attention is paid to the bottles that have already had growth detected and could provide critical information. Not even Gram-staining is performed during these long diagnostic pauses. No time benefit for detection of positive BCs is gained with continuously monitoring systems if loading and processing of bottles is organized discontinuously [
      • Blondeau J.M.
      • Idelevich E.A.
      The 24-h clinical microbiology service is essential for patient management.
      ,
      • Schneiderhan W.
      • Grundt A.
      • Wörner S.
      • Findeisen P.
      • Neumaier M.
      Work flow analysis of around-the-clock processing of blood culture samples and integrated MALDI-TOF mass spectrometry analysis for the diagnosis of bloodstream infections.
      ,
      • Riest G.
      • Linde H.J.
      • Shah P.M.
      Comparison of BacT/Alert and BACTEC NR 860 blood culture systems in a laboratory not continuously staffed.
      ]. It can thus be assumed that the reality of practice of BC diagnostics in Europe currently does not match the requirements of modern BSI management [
      • Rhodes A.
      • Evans L.E.
      • Alhazzani W.
      • Levy M.M.
      • Antonelli M.
      • Ferrer R.
      • et al.
      Surviving Sepsis campaign: international guidelines for management of sepsis and septic shock: 2016.
      ,
      • Blondeau J.M.
      • Idelevich E.A.
      The 24-h clinical microbiology service is essential for patient management.
      ]. Efforts should be made to increase 24-h coverage for both starting incubation of BCs and for BC processing after flagging positive.
      Another crucial diagnostic step is to validate and report the results of identification and AST to the attending clinician. The restricted duration of the working shift has tremendous impact on the availability of the AST result. Although some manufacturers have implemented shortened testing protocols [
      • Eigner U.
      • Schmid A.
      • Wild U.
      • Bertsch D.
      • Fahr A.M.
      Analysis of the comparative workflow and performance characteristics of the VITEK 2 and Phoenix systems.
      ], the testing will usually not be finalized during working hours of the laboratory. Knowledge of the actual working time patterns is important to underscore the need for the development of rapid AST methods that can be accomplished within a working shift. Laboratories equipped with MALDI-TOF MS are at least able to report the identification results on the same day that they set up the test. Laboratories that have to rely on biochemical identification methods will usually receive the identification result only the next day and therefore encounter the same problem as described above for AST.
      Although this survey focused on European laboratories, published data indicate that clinical microbiology in the USA is facing similar problems [
      • Tabak Y.P.
      • Vankeepuram L.
      • Ye G.
      • Jeffers K.
      • Gupta V.
      • Murray P.R.
      Blood culture turnaround time in US acute care hospitals and implications for laboratory process optimization.
      ,
      • Savinelli T.
      • Parenteau S.
      • Mermel L.A.
      What happens when automated blood culture instrument detect growth but there are no technologists in the microbiology laboratory?.
      ,
      • Schifman R.B.
      • Meier F.A.
      • Souers R.J.
      Timeliness and accuracy of reporting preliminary blood culture results: a College of American Pathologists Q-probes study of 65 institutions.
      ]. Worldwide, optimization of workflow and increasing service time have the potential to considerably reduce turnaround times.
      Rapid species identification reflects the recognition of the benefits to the clinicians of knowing the causative agents' species identification. This development is generally encouraging, but one-third of all laboratories still use the classical diagnostic approach, i.e. sub-cultivation on solid media and working with mature colonies the next day. As many rapid diagnostic techniques are now available and affordable [
      • Dubourg G.
      • Lamy B.
      • Ruimy R.
      Rapid phenotypic methods to improve the diagnosis of bacterial bloodstream infections: meeting the challenge to reduce the time to result.
      ,
      • Opota O.
      • Croxatto A.
      • Prod'hom G.
      • Greub G.
      Blood culture-based diagnosis of bacteraemia: state of the art.
      ,
      • Idelevich E.A.
      • Reischl U.
      • Becker K.
      New microbiological techniques in the diagnosis of bloodstream infections.
      ,
      • Idelevich E.A.
      • Becker K.
      Identification and susceptibility testing from shortly incubated cultures accelerate blood culture diagnostics at no cost.
      ,
      • EUCAST
      EUCAST rapid AST directly from positive blood culture bottles. Version 1.0.
      ], efforts should be made to implement them into clinical routine. The broad implementation of MALDI-TOF MS identification from briefly incubated sub-cultures on solid media is notable and understandable as this method provides accurate identification without additional expense within a few hours and only requires re-organization of laboratory workflow [
      • Idelevich E.A.
      • Schüle I.
      • Grünastel B.
      • Wüllenweber J.
      • Peters G.
      • Becker K.
      Rapid identification of microorganisms from positive blood cultures by MALDI-TOF mass spectrometry subsequent to very short-term incubation on solid medium.
      ]. The inoculation of semi-automated biochemical systems after a short sub-cultivation on agar plates is most probably applied as an equivalent by laboratories without access to MALDI-TOF MS.
      Far fewer laboratories reported AST from briefly incubated sub-cultures on agar. The use of this method may be expected to increase, as it has been shown to provide accurate final results one day sooner [
      • Ballestero-Téllez M.
      • Recacha E.
      • de Cueto M.
      • Pascual A.
      Identification and antimicrobial susceptibility testing of positive blood culture isolates from briefly incubated solid medium cultures.
      ,
      • Idelevich E.A.
      • Schüle I.
      • Grünastel B.
      • Wüllenweber J.
      • Peters G.
      • Becker K.
      Acceleration of antimicrobial susceptibility testing of positive blood cultures by inoculation of Vitek 2 cards with briefly incubated solid medium cultures.
      ,
      • Van den Poel B.
      • Klak A.
      • Desmet S.
      • Verhaegen J.
      How small modifications in laboratory workflow of blood cultures can have a significant impact on time to results.
      ] and can be easily combined with the identification from the same briefly incubated sub-cultures, both being easy to use without any additional investment [
      • Idelevich E.A.
      • Becker K.
      Identification and susceptibility testing from shortly incubated cultures accelerate blood culture diagnostics at no cost.
      ].
      One limitation of our study is that the inclusion might have been biased to some degree by the potential selection of laboratories that are particularly interested in BC diagnostics and so more likely to provide enhanced services. This bias might have been introduced in spite of efforts to include laboratories of different kinds, as described in the Methods section. A further limitation is that the participants were permitted to estimate some parameters when exact data were not available, e.g. the number of BC sets taken initially per fever episode. Future surveys could be expanded to include questions regarding contamination rate, storage of BC isolates and clinician attitudes to 24-h service versus restrictive services.
      This study assessed microbiological practices for the diagnosis of BSI in Europe and provided benchmarking for national and international comparisons. Furthermore, this project can set the basis for the development of European guidelines on BSI diagnostics.

      Transparency declaration

      BL reports a research grant on blood culture practice in France. DZ has received payments for lectures from Becton Dickinson and bioMérieux outside the submitted work. EAI has received grants or research support from the German Bundesministerium für Bildung und Forschung and is co-inventor of pending patents, which are owned by the University of Münster and licensed to Bruker, and received speaker honoraria from Bruker. HS has received grants or research support from the Bundesministerium für Bildung und Forschung, Germany , the German Centre for Infection Research (DZIF) , and Accelerate Diagnostics, has been a consultant for Becton Dickinson, ThermoFisher, and 3M, and has received payments for lectures from Becton Dickinson, bioMérieux and ThermoFisher. KB has received grants or research support from the German Bundesministerium für Bildung und Forschung and is co-inventor of pending patents, which are owned by the University of Münster and licensed to Bruker, and received speaker and consultation honoraria from Becton Dickinson, bioMérieux, Bruker Daltonik, Hain Lifescience, Roche Molecular Systems and ThermoFisher. LS has no financial conflict of interest to disclose; she is current ESCMID medical guidelines director. MS has received reimbursement for labour and material to collect samples in BSI studies outside the submitted work from Abbott Diagnostics and Q-linea AB. WJW receives funding from the Netherlands Organization for Scientific Research and the EU through a Marie Skłodowska-Curie Innovative Training Network grant. YC has received grants or research support from MSD, Pfizer, Allecra Therapeutics, Rempex Pharmaceuticals and Shionogi, and has received payments for lectures or consulting from MSD , Pfizer , Allecra Therapeutics , Rempex Pharmaceuticals , Nabriva , Achaogen and Roche . Other authors have nothing to disclose.

      Funding

      No external funding was received for this work.

      Acknowledgements

      We are grateful to the following persons who filled in the questionnaire or otherwise contributed to the project: Albania: Denada Lacej, Doloresa Mullaliu, Selaudin Jaupllari; Belgium: Michel Delmée, Alexia Verroken, Geert Claeys, Daniel Huang, Pierrette Melin, Eric Nulens, Michael Boudewijns, Annelies De Bel, Catherine Potvliege, Bruno Van Herendael, Koen Magerman; Bulgaria: Emma Keuleyan, Ivanka Gergova, Magdalena Lesseva, Rossitza Vatcheva-Dobrevska, Dobrinka Ivanova, Mariya Sredkova, Mariana Murdjeva, Grozdanka Lazarova, Temenuga Stoeva, Tatyana Malakova; Croatia: Arjana Tambić Andrašević, Irina Pristaš, Jasenka Škrlin, Marija Tonkić, Maja Abram, Iva Košćak, Ivanka Matas, Marina Payerl Pal, Edita Sušić, Ines Jajić-Benčić; Czech Republic: Lenka Ryskova, Pavel Cermak, Tamara Bergerova, Milan Kolar, Eva Simeckova, Miloslava Rumlerova, Helena Skacani, Natasa Bartonikova, Josef Scharfen; Denmark: Lars Lemming; Estonia: Krista Lõivukene, Anna Tisler-Sala, Valentina Kolesnikova, Irina Zolotuhhina, Leila Farah, Marika Jürna-Ellam, Ülle Laaring; Finland: Anu Patari-Sampo, Kaisu Rantakokko-Jalava, Jari Hirvonen, Jari Kauranen, Kerttu Saha, Maritta Kauppinen, Tamara Tuuminen, Pauliina Karpanoja, Jaakko Uksila; France: Raymond Ruimy, Jean-Winoc Decousser, Jocelyne Caillon, Olivier Barraud, Christophe Burucoa, Jean-Philippe Lavigne, Damien Fournier, Béatrice Pangon, Christian Cattoen; Germany: Can Imirzalioglu, Jan Rupp, Hanna Gölz, Ariane Dinkelacker, Inka Schüttert, Andreas Pennekamp, Frank Hünger, Stefanie Deinhardt-Emmer, Eberhard Kniehl, Winfried V. Kern; Greece: Aikaterini Tarpatzi, Efi Protonotariou, Evaggelia Kouskouni, Maria Panopoulou, Konstantina Gartzonika, Aggeliki Poulou, Vassilios Thomogloy, Olympia Zarkotou, Martha Nepka, Konstantina Kontopoulou, Dimitra Krikou; Israel: Tsilia Lazarovitch, Yuval Geffen, Avi Peretz, Gill Smollan, Sarit Freimann, Jacob Moran-Gilad, Orli Sagi, Meirav Strauss; Italy: Maurizio Sanguinetti, Claudio Scarparo, Giuseppe Cornaglia, Giuliana Lo Cascio, Paola Salvatore, Antonella Mencacci, Pierangelo Clerici, Carla Fontana, Piero Marone, Giuseppe Miragliotta, Francesco Luzzaro, Claudio Francesco Farina, Cristina Giraldi; Latvia: Irina Grave, Solvita Selderina, Dace Rudzite, Antonina Muizzemniece, Tatjana Djundika, Larisa Goldsteina, Inta Umbraško, Aļa Jegorova, Rita Levcenko; Lithuania: Silvija Kiveryte, Anzelika Jasiulioniene, Gintarė Sinkute, Gintaras Makstutis, Ieva Rutkauskienė, Jelena Kopeykiniene, Astra Vitkauskiene, Rita Velaviciene, Jurate Kirsliene, Greta Vizuje; Malta: Nina Nestorova; the Netherlands: Wieke Freudenburg, Miquel Ekkelenkamp, Karin van Dijk, Greetje Kampinga, Myrte Tielemans, Jacobien Veenemans; Norway: Truls Michael Leegaard, Gunnar Skov Simonsen, Jan Egil Afset, Elling Ulvestad, Ståle Tofteland, Nils Grude, Fredrik Müller, Iren Löhr; Poland: Lukasz Naumiuk, Anna Mol, Maciej Mikolajczyk, Anna Karolina Jurczak, Marta Wroblewska, Beata Muraszko, Jolanta Szumielewicz, Krzysztof Golec, Agnieszka Bys, Danuta Kedzia; Slovenia: Slavica Lorenčič Robnik, Barbara Zdolšek, Irena Grmek Košnik, Martina Kavčič, Viktorija Tomič, Jerneja Fišer; Spain: Marina de Cueto, Jorge Calvo, Enrique Ruiz de Gopegui, Maria Rodriguez Mayo, Manuel Lamela, Elena Loza, Jose María Navarro, Marta Arias Temprano, Concepcion Gimeno, Fatima Galan; Sweden: Kerstin Frölander, Volkan Özenci, Johan Rydberg, Torbjörn Kjaerstadius, Claes Henning, Anna Heydecke, Anna Åkerlund, Tor Monsen; Switzerland: Stephen Leib, Adrian Egli, Reinhard Zbinden, Hans Siegrist, Dieter Burki, Stefan Pfister; Turkey: Kerim Parlak, Tugba Cuhadar, Cemal Bulut, Gulay Okay, Yasemin Cag, Nurcan Baykam, Semsi Nur Karabela, Mustafa Guney; United Kingdom: Cressida Auckland, Gillian Orange, Mike Weinbren, Andree Evans, Bob Baker, Steve Glass, Tom Lewis, Aaron Nagar, Nidhika Berry, Rebecca Bamber, Andrew Kirby, Julian Bendle, Fahed Bekri.

      Appendix A. Supplementary data

      The following are the Supplementary data to this article:
      • Multimedia component 1

        Fig. S1. The annual numbers of blood cultures received by laboratories. A: blood culture sets received per 100 patient hospitalisations; B: blood culture sets received per 100 beds; C: blood culture bottles received per 100 patient hospitalisations; D: blood culture bottles received per 100 beds. In the analysis for all countries (left column on each figure) all laboratories are considered which provided response to that particular question (A, n=162; B, n=169; C, n=154; D, n=154). In the analysis for individual countries only countries are included for which at least 7 laboratories per country provided response to that particular question. Median values are presented.

      • Multimedia component 2

        Fig. S2. Estimated time to arrival of the majority of blood cultures (>80%) in laboratory after blood culture draw, % of respondents (A) and estimated percentage of blood cultures received and processed during off-hours for laboratories with 24/7 service for both starting incubation in the automated systems and for processing of positive BCs (n=23), % of respondents (B).

      • Multimedia component 3

        Fig. S3. Coverage of blood culture diagnostics in laboratories of different countries. A. Hours during which the laboratory can start incubation of blood culture bottles in a semi-automated blood culture system. B. Hours during which the laboratory can start processing blood culture bottles that have flagged positive in a semi-automated blood culture system. C. Hours during which the laboratory can validate and transmit the results of identification and susceptibility testing of blood culture isolates to the clinicians. In the analysis for all countries (left column on each graph) all laboratories are considered which provided response to that particular question (A, during the week: n=185; A, on Saturdays: n=184; A, on Sundays and holidays: n=183; B, during the week: n=192; B, on Saturdays: n=191; B, on Sundays and holidays: n=191; C, during the week: n=190; C, on Saturdays: n=190; C, on Sundays and holidays: n=190). For the analysis of individual countries only countries are included for which at least 7 laboratories per country provided response to that particular question. The analysis includes all laboratories independent of whether they provide 24 h-, part-time or no (0 h) respective service on respective days. Mean values of time coverage per 24 hours are presented.

      • Multimedia component 4

        Fig. S4. Proportion of multidrug-resistant organisms among blood culture isolates. A: methicillin-resistant S. aureus (MRSA) among all S. aureus blood culture isolates; B: ESBL-producing or third-generation cephalosporin-resistant E. coli among all E. coli blood cultures isolates; C: carbapenem-resistant Klebsiella spp. among all Klebsiella spp. blood cultures isolates; D: carbapenem-resistant P. aeruginosa among all P. aeruginosa blood cultures isolates; E: vancomycin-resistant E. faecium (VRE) among all E. faecium blood cultures isolates. In the analysis for all countries (left column on each figure) all laboratories are considered which provided response to that particular question (A, n=188; B, n=191; C, n=188; D, n=186; E, n=192). In the analysis for individual countries only countries are included for which at least 7 laboratories per country provided response to that particular question. Mean values are presented.

      • Multimedia component 5

        Fig. S5. Consultations by microbiology laboratory staff and infectious diseases department. A. Is advice for antimicrobial treatment provided by the laboratory staff for patients with detected bacteremia? B. Are blood culture results communicated to the infectious diseases department staff or consultation service? C. Reasons for selecting cases for presentation to infectious diseases department staff or consultation service (among laboratories which communicate only selected positive results to the infectious diseases department; multiple responses were allowed). * The following multidrug-resistant organisms were most commonly specified: methicillin-resistant S. aureus (MRSA) (8 laboratories), carbapenem-resistant bacteria (8 laboratories), vancomycin-resistant enterococci (VRE) (7 laboratories), ESBL-producing bacteria (4 laboratories). ** The following wards were most commonly specified: intensive care unit (5 laboratories), oncology/haematology (3 laboratories), paediatrics (1 laboratory).

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