Objectives Although perceived as a rare clinical entity, recent studies have noted the emergence of MDR C. parapsilosis (MDR-Cp) isolates from single patients (resistant to both azole and echinocandins). We previously reported a case series of MDR-Cp isolates carrying a novel FKS1R658G mutation. Herein, we identified an echinocandin-naive patient infected with MDR-Cp a few months after the previously described isolates. WGS and CRISPR-Cas9 editing were used to explore the origin of the new MDR-Cp isolates, and to determine if the novel mutation confers echinocandin resistance. Methods WGS was applied to assess the clonality of these isolates and CRISPR-Cas9 editing and a Galleria mellonella model were used to examine whether FKS1R658G confers echinocandin resistance. Results Fluconazole treatment failed, and the patient was successfully treated with liposomal amphotericin B (LAMB). WGS proved that all historical and novel MDR-Cp strains were clonal and distant from the fluconazole-resistant outbreak cluster in the same hospital. CRISPR-Cas9 editing and G. mellonella virulence assays confirmed that FKS1R658G confers echinocandin resistance in vitro and in vivo. Interestingly, the FKS1R658G mutant showed a very modest fitness cost compared with the parental WT strain, consistent with the persistence of the MDR-Cp cluster in our hospital. Conclusions Our study showcases the emergence of MDR-Cp isolates as a novel threat in clinical settings, which undermines the efficacy of the two most widely used antifungal drugs against candidiasis, leaving only LAMB as a last resort. Additionally, surveillance studies and WGS are warranted to effectively establish infection control and antifungal stewardship strategies.

Vaccines against SARS-CoV-2 protect from critical or severe pathogenesis also against new variants of concern (VOCs) such as BA.4 and BA.5, but immediate interventions to avoid viral transmission and subsequent inflammatory reactions are needed. Here we applied the ColdZyme® medical device mouth spray to fully differentiated, polarized human epithelium cultured at an air-liquid interphase (ALI). We found using VOCs BA.1 and BA.4/5 that this device effectively blocked respiratory tissue infection. While infection with these VOCs resulted in intracellular complement activation, thus enhanced inflammation, and drop of transepithelial resistance, these phenomena were prevented by a single administration of this medical device. Thus, ColdZyme® mouth spray significantly shields epithelial integrity, hinders virus infection and blocks in a secondary effect intrinsic complement activation within airway cultures also in terms of the highly contagious VOCs BA.4/5. Crucially, our in vitro data suggest that ColdZyme® mouth spray may have an impact to protect against SARS-CoV-2 transmission, also in case of the Omicron BA.1, BA.4 and BA.5 variants.

test result in 15-30 min, it still requires up to 2 h of incubation for some isolates and to confirm a negative test result. The rCIM(-A) may already give a positive result at 2 h (30 min of incubation and 1.5 h of growth monitoring of the Escherichia coli ATCC 25922 indicator strain) using standard routine equipment and reagents (carbapenem discs, Trypticase soy broth, a tabletop centrifuge, a vortex and a nephelometer). We agree that the rCIM requires extra hands-on time, thus making it less attractive for high-throughput laboratories, but it may be appropriate for lowthroughput and low-resource settings. 2,3 Finally, both assays have similar turnaround times, as both require overnight bacterial cultures, and the longer detection time of the rCIM is only a small fraction of total reporting time. Moreover, for initiating appropriate antibiotic treatments in critically ill patients, rapid susceptibility testing using techniques that have been developed and endorsed by EUCAST are crucial, even in low-and middle-income countries. 11 Transparency declarationsNone to declare. References1 Nordmann P, Poirel L. Comment on: Optimization of the rapid carbapenem inactivation method for use with AmpC hyperproducers.