Performance of the eazyplex® BloodScreen GN as a simple and rapid molecular test for identification of Gram-negative bacteria from positive blood cultures.

The LAMP-based eazyplex® BloodScreen GN was evaluated for the detection of frequent Gram-negatives directly from positive blood culture (BC) bottles. A total of 449 BCs were analyzed. Sensitivities and specificities were 100% and 100% for Escherichia coli, 95.7% and 100% for Klebsiella pneumoniae, 100% and 100% for blaCTX-M, 100% and 100% for Klebsiella oxytoca, 100% and 99% for Proteus mirabilis, and 100% and 99.8% for Pseudomonas aeruginosa, respectively. The time to result ranged from 8 to 16 min, plus about 6 min for sample preparation. The eazyplex® BloodScreen GN is a reliable molecular assay for rapid BC testing.

Time to positivity as a prognostic factor in bloodstream infections with Enterococcus spp.

Time to positivity (TTP) is the delay of time from incubation to blood culture positivity. Short TTP can predict mortality and source of infection. The aim of this study was to investigate the value of TTP of patients with bloodstream infections with enterococci (E-BSI).In a single centre retrospective cohort study in Germany, the data of 244 patients with monomicrobial E-BSI were analyzed with hospital mortality as the primary outcome of interest from January 1 2014 to December 31 2016. Mortality rate of patients with bloodstream infections (BSI) with E. faecalis was 16.7%, Vancomycin sensitive E. faecium (VSEfm) 26.7% and Vancomycin resistant E. faecium (VREfm) 38.2%. Cut-offs showed a significantly higher mortality rate when compared to longer TTP (E. faecalis: P=0.047; VSEfm: P=0.02), but were not risk factors in survival analysis (E.faecalis: HR (hazard ratio): 2.73; P=0.17; VSEfm: HR: 1.63; P=0.15; VREfm: HR: 1.24; P=0.63). TTP≤10.5 hours with E. faecalis BSI was a discriminator for cardiovascular source of infection (AUC: 0.75). A short TTP could predict mortality rates and source of infection but was not an independent parameter for risk of death in survival analysis.

Management of a Hospital-Wide COVID-19 Outbreak Affecting Patients and Healthcare Workers.

To the best of our knowledge, here, we describe the first hospital-wide outbreak of SARS-CoV-2 that occurred in Germany in April 2020. We aim to share our experience in order to facilitate the management of nosocomial COVID-19 outbreaks in healthcare facilities. All patients and hospital workers were screened for SARS-CoV-2 repeatedly. An infection control team on the side was installed. Strict spatial separation of patients and intensified hygiene training of healthcare workers (HCW) were initiated. By the time of reporting, 26 patients and 21 hospital workers were infected with a cluster of cases in the geriatric department. Fourteen patients developed COVID-19 consistent symptoms and five patients with severe pre-existing medical conditions died. The outbreak was successfully contained after intensified infection control measures were implemented and no further cases among patients were detected over a period of 14 days. Strict application of standard infection control measures proved to be successful in the management of nosocomial SARS-CoV-2 outbreaks.

Diagnostic accuracy and clinical impact of loop-mediated isothermal amplification for rapid detection of Staphylococcus aureus bacteremia: a retrospective observational study.

The aim of this study was to evaluate the diagnostic accuracy and the clinical impact of isothermal loop-mediated amplification (LAMP; eazyplex® MRSA kits) for rapid diagnosis of Staphylococcus aureus bacteremia (SAB) in comparison with conventional blood culture diagnostics. We performed a retrospective, single-center observational study over the period between November 2016 and December 2018 on patients (and blood cultures) with growth of Gram-positive cocci in clusters in their blood cultures. We quantified diagnostic accuracy with sensitivity and specificity for detection of S. aureus, methicillin-resistant S. aureus (MRSA), and the mecA/C resistance genes in 797 blood cultures. The clinical impact was assessed by time to result reporting, time to appropriate treatment, and length of stay in intensive care unit (ICU) in 190 SAB patients. We observed sensitivity and specificity above 90% for S. aureus detection (sensitivity (95% confidence interval (CI)), 99.57% (97.61%, 99.98%); specificity, 99.12% (97.95%, 99.71%)), for MRSA detection (sensitivity, 100% (89.11%, 100.00%); specificity, 99.72% (99.05, 99.96)), and for mecA/C detection (sensitivity, 94.71% (91.85%, 96.78%); specificity, 95.89% (93.58%, 97.54%)). LAMP testing was associated with shorter median time to result reporting (24.0 h (first and third quartiles (Q1-Q3), 20.0-27.0 h) vs 41.5 h (36.0-46.0 h); p < 0.001) and different distribution of time to appropriate treatment (2.0 days (1.0-3.0) vs 2.0 days (2.0-3.0); p = 0.004). No evidence for differences in length-of-stay in ICU was observed. Our analysis suggests for the application of LAMP (i) a high diagnostic accuracy for detection of S. aureus and the mecA/C genes in blood cultures, (ii) an earlier result reporting, and (iii) a shorter time to appropriate treatment.

Rapid antibiotic susceptibility testing in blood culture diagnostics performed by direct inoculation using the VITEK®-2 and BD Phoenix™ platforms.

Early availability of microbiological results can improve treatment decisions of patients suffering from bloodstream infections. Direct inoculation of automated susceptibility testing (AST) platforms is an approach to shorten time-to-result in blood culture diagnostics. We performed a comparative evaluation of the two commercial AST systems VITEK®-2 and BD Phoenix™ for the direct inoculation with blood culture samples. Furthermore, two different methods of sample preparation were compared in this study. Positive blood cultures were prepared for direct inoculation by use of serum separator tubes and twofold centrifugation. AST was performed with the VITEK®-2 and the BD Phoenix™ system by the standard method according to the manufacturer's recommendations using subcultures on solid media and by direct inoculation of blood culture samples. A hundred clinical samples from blood cultures were included in this study. Rapid AST by direct inoculation showed inter-test agreement rates ranging from 92.45 to 97.7%. Comparing both AST platforms, the VITEK®-2 system demonstrated a higher test accuracy for direct inoculation. No relevant difference was observed for the two different sample preparation methods. Direct inoculation is an easy and inexpensive approach to obtain early full panel phenotypic AST results in blood culture diagnostics. Sample preparation is sufficiently performed by a simple centrifugation method. Both commercial platforms, the VITEK®-2 and the BD Phoenix™, have proven suitable for the use of direct inoculation.

Dwarfs in disguise: multiple spinal abscesses and spondylodiscitis caused by an Enterococcus faecium small-colony variant.

Small-colony variants are slow-growing subpopulations of bacteria known to be involved in latent or recurrent infections, especially in deep-seated foci. Their atypical growth in small colonies can hamper prompt and correct identification in clinical specimens. Here, we present the first case of multiple spinal abscesses and spondylodiscitis associated with an Enterococcus faecium small-colony-variant in an immunocompetent patient. This case demonstrates the diagnostic challenges when encountering this phenotype in the diagnostic laboratory.

First Time Isolation of Mycobacterium hassiacum From a Respiratory Sample.

We describe the first isolation of Mycobacterium hassiacum, a rapid-growing, partial acid-resistant mycobacterium, in a respiratory specimen from a patient with exacerbated chronic obstructive pulmonary disease. To provide therapeutic recommendation for future cases, antibiotic susceptibility testing of 3 clinical isolates was performed by broth microdilution. All strains tested showed susceptibility to clarithromycin, imipenem, ciprofloxacin, and doxycycline. The role of M hassiacum as a respiratory pathogen remains unclear and needs to be evaluated by future reports.

Lysozyme facilitates adherence of Enterococcus faecium to host cells and induction of necrotic cell death.

The prevalence of infections with enterococci is increasing worldwide. However, little is known about the mechanisms which enable these opportunistic pathogens to cause infections of their host. Here we demonstrate that Enterococcus faecium in the presence of lysozyme induces necrosis in human and mouse cells after 4 h indicated by disrupted cellular membranes of epithelial (HeLa), myeloid (U937, J774A.1) and lymphoid (Jurkat J16, thymocytes), but not intestinal epithelial cells (CaCo-2, CMT-93). Using an appropriate mutant strain it was shown that the enterococcal surface-protein SgrA is involved in cell death induction in mouse cells (J774A.1, thymocytes). Microscopic analyses of epithelial cells 30 min post infection revealed that lysozyme increases adhesion of E. faecium to HeLa, but not CaCo-2 cells. At that time the phalloidin-FITC-stained cytoskeleton of infected cells was still intact, whereas 2 h post infection the F-actin network of HeLa, but not CaCo-2 cells was disrupted. Hence, the early, lysozyme-mediated increase of bacterial adherence plays an important role for cell death induction by E. faecium in HeLa cells. Moreover, bacterial extracellular hydrogen peroxide might contribute to necrosis induction, since the rate of propidium iodide-positive HeLa and J774A.1 cells was lowered after infection with a ROS-deficient E. faecium mutant.