Short incubation of disc diffusion for Streptococcus pneumoniae and Haemophilus influenzae to reduce time to susceptibility report.

BACKGROUND: Rapidly instituted antimicrobial therapy is important in severe infections, and reduced time to the antimicrobial susceptibility testing (AST) report is thus of importance. Disc diffusion (DD) is a cheap, rapidly adaptable, flexible and comprehensive method for phenotypic AST. Previous studies have shown that early reading of inhibition zones for non-fastidious species is possible. OBJECTIVES: To evaluate zone reading after short incubation of DD in Haemophilus influenzae (n = 73) and Streptococcus pneumoniae (n = 112). METHODS: The readability was evaluated and susceptibility interpretation (SIR) was performed, using the EUCAST 18 ±â€Š2 h incubation breakpoint table (version 12.0), after 6 and 8 h of incubation. Categorical agreement (CA) and error rates were calculated using standard DD and broth microdilution as reference. RESULTS: The proportion of readable zones in H. influenzae was 19% (6 h) and 89% (8 h). The CA was 98% after 8 h. The corresponding readability in S. pneumoniae was 63%/98% and CA was 95%/97% after 6 and 8 h, respectively. Early reading of the screening discs (benzylpenicillin 1 unit in H. influenzae and oxacillin 1 µg in S. pneumoniae) correctly identified 18/22 of the H. influenzae isolates and all the readable S. pneumoniae isolates with reduced ß-lactam susceptibility. For non-ß-lactam agents, very major errors were most common for quinolones in S. pneumoniae. Introduction of areas of technical uncertainty (ATUs) reduced the error rate to ≤1.1%. CONCLUSIONS: We conclude that shortened incubation is feasible for H. influenzae and S. pneumoniae. To reduce the risk of false categorization a buffer zone (i.e. ATU) near the breakpoints must be used.

Analytical Performance and Potential Clinical Utility of EUCAST Rapid Antimicrobial Susceptibility Testing in Blood Cultures after Four Hours of Incubation.

EUCAST rapid antimicrobial susceptibility testing (RAST) provides antibiotic susceptibility results after 4 to 8 h of incubation. This study assessed the diagnostic performance and clinical usefulness of EUCAST RAST after 4 h. This was a retrospective clinical study performed on blood cultures with Escherichia coli and Klebsiella pneumoniae complex (K. pneumoniae and Klebsiella variicola) at Karolinska University Laboratory (Stockholm, Sweden). The rate of categorized RAST results and the categorical agreement (CA) of RAST with the standard EUCAST 16-to-20-h disk diffusion (DD) method for piperacillin-tazobactam, cefotaxime, ceftazidime, meropenem, and ciprofloxacin were analyzed, as well as the utility of RAST for adjusting the empirical antibiotic therapy (EAT) and the combination of RAST with a lateral flow assay (LFA) for extended-spectrum ß-lactamase (ESBL) detection. A total of 530 E. coli and 112 K. pneumoniae complex strains were analyzed, generating 2,641 and 558 readable RAST zones, respectively. RAST results categorized according to antimicrobial sensitivity/resistance (S/R) were obtained for 83.1% (2,194/2,641) and 87.5% (488/558) of E. coli and K. pneumoniae complex strains, respectively. The RAST result categorization to S/R for piperacillin-tazobactam was poor (37.2% for E. coli and 66.1% for K. pneumoniae complex). CA with the standard DD method was over 97% for all tested antibiotics. Using RAST, we detected 15/26 and 1/10 of the E. coli and K. pneumoniae complex strains that were resistant to the EAT. For patients treated with cefotaxime, RAST was used to detect 13/14 cefotaxime-resistant E. coli strains and 1/1 cefotaxime-resistant K. pneumoniae complex strain. ESBL positivity was reported the same day as blood culture positivity with RAST and LFA. EUCAST RAST provides accurate and clinically relevant susceptibility results after 4 h of incubation and can accelerate the assessment of resistance patterns. IMPORTANCE Early effective antimicrobial treatment has been shown to be crucial for improving the outcome of bloodstream infections (BSI) and sepsis. In combination with the rise of antibiotic resistance, this calls for accelerated methods for antibiotic susceptibility testing (AST) for effective treatment of BSI. This study assesses EUCAST RAST, an AST method that yields results in 4, 6, or 8 h after blood culture positivity. We analyzed a high number of clinical samples of Escherichia coli and Klebsiella pneumoniae complex strains and confirm that the method delivers reliable results after 4 h of incubation for the relevant antibiotics for treating E. coli and K. pneumoniae complex bacteremia. Furthermore, we conclude that it is an important tool for antibiotic treatment decision-making and early detection of ESBL-producing isolates.

Blood culture diagnostics: a Nordic multicentre survey comparison of practices in clinical microbiology laboratories.

OBJECTIVES: Accurate and rapid microbiological diagnostics are crucial to tailor treatment and improve outcomes in patients with severe infections. This study aimed to assess blood culture diagnostics in the Nordic countries and to compare them with those of a previous survey conducted in Sweden in 2013. METHODS: An online questionnaire was designed and distributed to the Nordic clinical microbiology laboratories (CMLs) (n = 76) in January 2018. RESULTS: The response rate was 64% (49/76). Around-the-clock incubation of blood cultures (BCs) was supported in 82% of the CMLs (40/49), although in six of these access to the incubators around the clock was not given to all of the cabinets in the catchment area, and 41% of the sites (20/49) did not assist with satellite incubators. Almost half (49%, 24/49) of the CMLs offered opening hours for ≥10 h during weekdays, more commonly in CMLs with an annual output ≥30 000 BCs. Still, positive BCs were left unprocessed for 60-70% of the day due to restrictive opening hours. Treatment advice was given by 23% of CMLs (11/48) in ≥75% of the phone contacts. Rapid analyses (species identification and susceptibility testing with short incubation), performed on aliquots from positive cultures, were implemented in 18% of CMLs (9/49). Compared to 2013, species identification from subcultured colonies (<6 h) had become more common. CONCLUSIONS: CMLs have taken action to improve aspects of BC diagnostics, implementing satellite incubators, rapid species identification and susceptibility testing. However, the limited opening hours and availability of clinical microbiologists are confining the advantages of these changes.

EUCAST rapid antimicrobial susceptibility testing (RAST) in blood cultures: validation in 55 European laboratories.

OBJECTIVES: When bloodstream infections are caused by resistant bacteria, rapid antimicrobial susceptibility testing (RAST) is important for adjustment of therapy. The EUCAST RAST method, directly from positive blood cultures, was validated in a multi-laboratory study in Europe. METHODS: RAST was performed in 40 laboratories in northern Europe (NE) and 15 in southern Europe (SE) from clinical blood cultures positive for Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Staphylococcus aureus or Streptococcus pneumoniae. Categorical results at 4, 6 and 8 h of incubation were compared with results for EUCAST standard 16-20 h disc diffusion. The method, preliminary breakpoints and the performance of the laboratories were evaluated. RESULTS: The total number of isolates was 833/318 in NE/SE. The number of zone diameters that could be read (88%, 96% and 99%) and interpreted (70%, 81% and 85%) increased with incubation time (4, 6 and 8 h). The categorical agreement was acceptable, with total error rates in NE/SE of 2.4%/4.9% at 4 h, 1.1%/3.5% at 6 h and 1.1%/3.3% at 8 h. False susceptibility at 4, 6 and 8 h of incubation was below 0.3% and 1.1% in NE and SE, respectively, and the corresponding percentages for false resistance were below 1.9% and 2.8%. After fine-tuning breakpoints, more zones could be interpreted (73%, 89% and 93%), with only marginally affected error rates. CONCLUSIONS: The EUCAST RAST method can be implemented in routine laboratories without major investments. It provides reliable antimicrobial susceptibility testing results for relevant bloodstream infection pathogens after 4-6 h of incubation.

[Response times can be shortened in microbiological diagnosis of sepsis. Better opening hours in laboratories and active advice enables faster therapy]. / Svarstiderna kan kortas vid mikrobiologisk diagnostik av sepsis--Bättre öppettider på laboratorier och aktiv rådgivning ger snabbare terapi.

Early and adequate antimicrobial therapy is lifesaving in patients with bloodstream infections (BSI). Continuous incubation and 24h availability of microbial pathogen identification in clinical microbiology laboratories shortens the time to report. To describe the current status in Swedish clinical microbiology laboratories, a web-based survey was performed. The survey showed major differences in availability both regarding laboratory opening hours and ability to incubate blood culture bottles outside working hours. It also showed differences in what was conveyed verbally to the clinician and in time to report of the bacterial species. Based on this survey, we debate how Swedish healthcare can improve the care of patients with BSI by offering a more rapid diagnostic process. This could be achieved through longer opening hours of the laboratory, better transport systems and blood culture cabinets positioned closer to the patient.