Pharmacokinetics of isavuconazole at different target sites in healthy volunteers after single and multiple intravenous infusions.

BACKGROUND: Invasive aspergillosis is a severe fungal infection that affects multiple organ systems including the CNS and the lungs. Isavuconazole, a novel triazole antifungal agent, has demonstrated promising activity against Aspergillus spp. However, data on the penetration of isavuconazole into the CNS and ELF and intracellular accumulation remain limited. MATERIALS AND METHODS: We conducted a prospective single-centre pharmacokinetic (PK) study in 12 healthy volunteers. Subjects received seven doses of 200 mg isavuconazole to achieve an assumed steady-state. After the first and final infusion, plasma sampling was conducted over 8 and 12 h, respectively. All subjects underwent one lumbar puncture and bronchoalveolar lavage, at either 2, 6 or 12 h post-infusion of the final dose. PBMCs were collected in six subjects from blood to determine intracellular isavuconazole concentrations at 6, 8 or 12 h. The AUC/MIC was calculated for an MIC value of 1 mg/L, which marks the EUCAST susceptibility breakpoint for Aspergillus fumigatus and Aspergillus flavus. RESULTS: C max and AUC0-24h of isavuconazole in plasma under assumed steady-state conditions were 6.57 ±â€Š1.68 mg/L (mean ±â€ŠSD) and 106 ±â€Š32.1 h·mg/L, respectively. The average concentrations measured in CSF, ELF and in PBMCs were 0.07 ±â€Š0.03, 0.94 ±â€Š0.46 and 27.1 ±â€Š17.8 mg/L, respectively. The AUC/MIC in plasma, CSF, ELF and in PBMCs under steady-state conditions were 106 ±â€Š32.1, 1.68 ±â€Š0.72, 22.6 ±â€Š11.0 and 650 ±â€Š426 mg·h/L, respectively. CONCLUSION: Isavuconazole demonstrated moderate penetration into ELF, low penetrability into CSF and high accumulation in PBMCs. Current dosing regimens resulted in sufficient plasma exposure in all subjects to treat isolates with MICs ≤ 1 mg/L.

Unraveling novel mutation patterns and morphological variations in two dalbavancin-resistant MRSA strains in Austria using whole genome sequencing and transmission electron microscopy.

BACKGROUND: The increasing prevalence of methicillin-resistant Staphylococcus aureus (MRSA) strains resistant to non-beta-lactam antimicrobials poses a significant challenge in treating severe MRSA bloodstream infections. This study explores resistance development and mechanisms in MRSA isolates, especially after the first dalbavancin-resistant MRSA strain in our hospital in 2016. METHODS: This study investigated 55 MRSA bloodstream isolates (02/2015-02/2021) from the University Hospital of the Medical University of Vienna, Austria. The MICs of dalbavancin, linezolid, and daptomycin were assessed. Two isolates (16-33 and 19-362) resistant to dalbavancin were analyzed via whole-genome sequencing, with morphology evaluated using transmission electron microscopy (TEM). RESULTS: S.aureus BSI strain 19-362 had two novel missense mutations (p.I515M and p.A606D) in the pbp2 gene. Isolate 16-33 had a 534 bp deletion in the DHH domain of GdpP and a SNV in pbp2 (p.G146R). Both strains had mutations in the rpoB gene, but at different positions. TEM revealed significantly thicker cell walls in 16-33 (p < 0.05) compared to 19-362 and dalbavancin-susceptible strains. None of the MRSA isolates showed resistance to linezolid or daptomycin. CONCLUSION: In light of increasing vancomycin resistance reports, continuous surveillance is essential to comprehend the molecular mechanisms of resistance in alternative MRSA treatment options. In this work, two novel missense mutations (p.I515M and p.A606D) in the pbp2 gene were newly identified as possible causes of dalbavancin resistance.

Differences in oxazolidinone resistance mechanisms and small colony variants emergence of Staphylococcus aureus induced in an in vitro resistance development model.

Invasive Staphylococcus aureus infections are associated with a high burden of disease, case fatality rate and healthcare costs. Oxazolidinones such as linezolid and tedizolid are considered potential treatment choices for conditions involving methicillin resistance or penicillin allergies. Additionally, they are being investigated as potential inhibitors of toxins in toxin-mediated diseases. In this study, linezolid and tedizolid were evaluated in an in vitro resistance development model for induction of resistance in S. aureus. Whole genome sequencing was conducted to elucidate resistance mechanisms through the identification of causal mutations. After inducing resistance to both linezolid and tedizolid, several partially novel single nucleotide variants (SNVs) were detected in the rplC gene, which encodes the 50S ribosome protein L3 in S. aureus. These SNVs were found to decrease the binding affinity, potentially serving as the underlying cause for oxazolidinone resistance. Furthermore, in opposite to linezolid we were able to induce phenotypically small colony variants of S. aureus after induction of resistance with tedizolid for the first time in literature. In summary, even if different antibiotic concentrations were required and SNVs were detected, the principal capacity of S. aureus to develop resistance to oxazolidinones seems to differ between linezolid and tedizolid in-vivo but not in vitro. Stepwise induction of resistance seems to be a time and cost-effective tool for assessing resistance evolution. Inducted-resistant strains should be examined and documented for epidemiological reasons, if MICs start to rise or oxazolidinone-resistant S. aureus outbreaks become more frequent.

Reproducibility of LPS-Induced ex vivo Cytokine Response of Healthy Volunteers Using a Whole Blood Assay.

Introduction: Lipopolysaccharide (LPS) stimulation of human whole blood ex vivo has been widely used to investigate human innate immune responses. However, there are uncertainties regarding the reproducibility and reliability of this assay. Methods: In this prospective, single-center study, cytokine responses (interleukin 8, interferon-α, interferon-γ, interleukin 10, interleukin 1-ß, interleukin 6, and tumor necrosis factor-α) to ex vivo whole blood LPS stimulation were assessed in 12 healthy volunteers. Cytokine levels were measured at 0, 2, and 4 h using a multiplex immunoassay (Luminex ®). Stimulation was repeated after six weeks. We examined reproducibility across technical and biological replicates at baseline and between repeated experiments after 6 weeks based on the area under the curve (AUC) of the individual cytokines using Pearson's correlation coefficient and the mean coefficient of variation. Results: The lowest mean coefficients of variation were observed for the technical replicates (5.4 to 9.2%), followed by the biological replicates (8.1 to 24.8%), and the repeated experiments after 6 weeks (17 to 31.2%). Between the baseline and 6-week AUCs, the following Pearson correlation coefficients R were observed: interleukin 10, 0.97; interferon-α, 0.84; interleukin 1-ß, 0.83; interleukin 8, 0.79; interleukin 6, 0.73; interferon-γ, 0.73; and tumor necrosis factor-α, 0.63. Discussion: The level of agreement between the baseline and week-6 cytokine response to ex vivo LPS stimulation was high across the seven cytokines analyzed. While interleukin 10 exhibited the lowest level of variability over time, tumor necrosis factor-α showed the highest variability in repeated experiments, which should be considered in the design and interpretation of future studies.