ABSTRACT Invasive fungal diseases are difficult to diagnose because of the limited sensitivity of culture. Panfungal PCR amplicon sequencing assays (targeting ribosomal RNA, such as 18S, 28S, ITS) are recommended for fungal identification in histopathology samples showing fungal elements. However, data describing its overall performance and consistency are lacking. This systematic literature review and meta-analysis assessed the performance of panfungal PCR on formalin-fixed paraffin-embedded (FFPE) and non-fixed (fresh or frozen) tissue samples. A systematic literature search was performed to include studies reporting the use of panfungal PCR for fungal identification in FFPE or non-fixed tissue samples. PCR sensitivity and specificity were assessed using the reference standard of histopathology showing fungal elements. Quality assessment was performed using the Quality Assessment of Diagnostic Accuracy Studies (QUADAS-2) tool. Pooled estimates were obtained using random-effects meta-analysis. Twenty-eight studies were included. In FFPE samples (18 studies, 852 samples), sensitivity and specificity were 75.4% (95% confidence interval [CI], 59.2–86.6) and 93.5% (70.2–98.9), respectively. Sensitivity in non-fixed samples (13 studies, 207 samples) was 86.5% (74.7–93.3), while specificity could not be assessed (insufficient data). Comparative analyses showed a significantly higher sensitivity of panfungal PCR over culture (88.2%; 76–94.7 vs 52.2%; 39–65, P = 0.001). Sub-analyses could not demonstrate the superiority of one PCR target over another due to limited data. Panfungal PCR exhibited adequate sensitivity and good specificity in FFPE samples. Sensitivity was even higher in non-fixed samples and largely superior to culture. Nevertheless, large interstudy variability was observed, warranting interlaboratory studies to define the optimal PCR target and standardized protocols. IMPORTANCE Invasive fungal diseases are difficult to diagnose because of the low sensitivity of culture. Panfungal PCRs are widely used for fungal identification in tissue specimens but suffer from heterogeneous procedures and performance. This meta-analysis shows an acceptable sensitivity (75.4% and 86.5% in fixed and non-fixed samples, respectively) and good specificity (93.5%) of panfungal PCR, supporting its use, not only on histopathology-positive fixed samples but also in non-fixed samples concomitantly with other diagnostic tools (cultures and fungal-specific PCRs if available). These results provide a strong basis for further standardization of panfungal PCR techniques via interlaboratory assays to assess reproducibility and optimize analytical protocols. CLINICAL TRIALS This study is registered with PROSPERO as CRD42023461148 .

Candidozyma auris (C. auris) is an emerging healthcare-associated yeast of major epidemiological concern because of its multidrug resistance and outbreak potential. We report the recovery of a single C. auris isolate from a used face mask collected in May 2025 during a blinded dental medicine quality-control programme assessing microbial contamination in the working environment. To contextualise this finding, we analysed routine diagnostic laboratory data from 2017 to 2025. The isolate underwent whole-genome sequencing for molecular characterisation, including analysis of the ERG11 gene, and antifungal susceptibility testing by EUCAST broth microdilution. In addition, 53,802 patient-related Candida spp. isolates collected between 2017 and 2025 were reviewed retrospectively; species identification had been performed by MALDI-TOF. The environmental isolate belonged to clade III and carried the V125A/F126L substitutions in ERG11, consistent with African clade isolates and associated with intrinsically high fluconazole minimum inhibitory concentrations. No C. auris was detected in routine patient specimens during the study period, whereas Candida albicans remained the predominant species in clinical samples. These findings provide no evidence of ongoing C. auris transmission at the Medical University of Innsbruck, but highlight the need for continued vigilance and robust infection-prevention measures to limit the risk posed by isolated introductions.

Several European countries reported a marked rise in paediatric invasive group A streptococcal (iGAS) infections from mid-2022 onwards. We describe the clinical spectrum and intensive care course of children with iGAS admitted to Austrian paediatric intensive care units (PICUs) during this period. We conducted a nationwide, multicentre, retrospective observational cohort study across all 13 Austrian PICUs. Children aged 0–18 years admitted between 1 September 2022 and 31 March 2024 with microbiologically confirmed iGAS were identified using a standardised, pseudonymised case report form. Data on demographics, clinical presentation, organ support, co-infections, microbiology, and outcomes were collected. All 13 Austrian PICUs participated, contributing 70 cases. The most frequent initial symptoms were pleural empyema (51.4

Dermatophytes are fungi that infect the human skin and its appendages. With new pathogenic species emerging and resistance to first-line drugs rising, microbiologic diagnosis and species identification are becoming even more important. In this study, the DermaGenius® 2.0/3.0 Complete multiplex real-time PCR and the EUROArray Dermatomycosis kits were compared to fungal culture and with each other; 78 reference strains and 124 clinical samples were analyzed. Both the DermaGenius® kit (97%; 95%CI 89–100%) and the EUROArray assay (91%; 95% CI: 82–96%) were sensitive when analyzing on-panel reference strains. In clinical samples, the DermaGenius® assay provided a positive result in 63 out of 124 (51%) samples and the EUROArray assay in 74 out of 124 (60%) samples. Both kits supported the diagnosis and species identification of culture-negative samples, and samples with growth of unconventional species. However, there was suspicion of false-positive results with F. solani in the EUROArray kit both in clinical and reference strains. The most common conventional dermatophytes in this study combining all methods were T. rubrum/soudanense (n = 40) and T. interdigitale/mentagrophytes (n = 11). In summary, both PCR kits were sensitive for the diagnosis and species identification of dermatophytoses. Combining culture and a PCR-based method can increase the diagnostic yield and compensate for the weakness of the other methods. The optimal PCR-based kit, and especially the optimal panel size, depends on the local epidemiology of dermatophytes.

Nakaseomyces glabratus (formerly Candida glabrata) is a leading cause of invasive candidiasis and rapidly develops antifungal drug resistance during treatment. An increasing number of clinical isolates show reduced susceptibility to echinocandins and azoles, leaving amphotericin B (AMB) as a last therapeutic option. Resistance of N. glabratus to this drug is rare, and its underlying mechanisms are still not fully understood. Here, we describe two independent multidrug-resistant bloodstream isolates displaying resistance to AMB and anidulafungin (ANF), as well as a reduced susceptibility to azoles. We performed whole-genome sequencing and sterol profiling on nine clinical N. glabratus isolates, which were resistant to ANF and displayed resistance or low susceptibility to fluconazole (FLU) and AMB. We identified loss-of-function mutations in the genes ERG3 and ERG4, which could be linked to ergosterol depletion and AMB resistance. The transcriptional response of the reference strain CBS138 and an AMBR + ANFR isolate was analyzed by RNA-seq, revealing that ergosterol depletion also contributed to upregulation of ERG and ABC transporter genes, which might explain the low FLU susceptibility. Surprisingly, the AMBR isolates displayed severe fitness defects, and one of them was fully virulent in a Galleria mellonella infection model. Our results indicate that ergosterol depletion in N. glabratus leads to AMB resistance without affecting fitness or virulence. IMPORTANCE:The major human fungal pathogen Nakaseomyces glabratus is well known for its fast development of antifungal drug resistance, especially against commonly used azoles. However, it can also acquire resistance to echinocandins, leading to multidrug resistance (MDR) and leaving amphotericin B (AMB) as the last therapeutic option. AMB resistance is rare, mainly caused by ergosterol depletion, and is normally associated with severe fitness costs for the pathogen. However, we found N. glabratus bloodstream isolates with stable AMB resistance without apparent fitness and virulence defects. The underlying ergosterol depletion contributed to low azole susceptibility and was associated with anidulafungin resistance. These findings demonstrate how fast MDR can evolve in N. glabratus and underline the need for close resistance monitoring.

The European Confederation of Medical Mycology Candida III was a pan-European, multicenter observational study of adult patients with blood culture-proven candidemia. Among a total of 632 patients with candidemia across 64 institutions in 20 European countries, a subanalysis of 396 (63%) cases occurring outside the intensive care unit (ICU) was conducted. Compared with ICU patients, non-ICU patients had a higher comorbidity burden (median Charlson comorbidity index [CCI] 6 vs 5 in ICU patients, P = .006). Hematologic and oncologic malignancies were more frequent among non-ICU cases (45.5% vs 28.4%, P < .001), whereas both chronic kidney and cardiovascular disease were more prevalent in ICU patients (P < .001). Non-ICU patients had significantly lower mortality in Kaplan-Meier survival analysis (P > .001). Postsurgical non-ICU patients (n = 45) had the highest survival rate (73.3%, P = .003) and the longest hospital stay, even after excluding all cases with a fatal outcome before day 30. In non-ICU patients, older age, hemato-oncologic malignancies, chronic liver disease, and COVID-19 were all independently associated with mortality risk, while treatment consultation by an infectious disease or clinical microbiology consultant, and initial treatment with an echinocandin, respectively, higher EQUAL Candida scores were associated with lower mortality risk in the multivariable Cox regression models. In conclusion, despite higher comorbidity rates, non-ICU patients with candidemia had higher survival rates.

Dermatophytes cause a wide range of superficial infections of the skin, hair and nails, with high global prevalence and considerable public health relevance. Treatment regimens for dermatophytosis are limited to few antifungal drug classes, and rising resistance—particularly to terbinafine—further compromises therapeutic efficacy. Topical antiseptics such as octenidine (OCT) may represent a promising option for infection control and local therapy. This study investigated the in vitro antifungal activity of the pure antiseptic OCT (at final assay concentrations of 0.1

Fungal infections of the skin are a frequent infectious skin disease in childhood. In particular, the close skin contact in communal facilities, contact to animals, the still immature skin barrier and immune defence as well as certain hygienic and climatic conditions facilitate the occurrence. Global migration movements and international travelling activities also contribute to the distribution of new or previously rare pathogens with elevated resistance profiles. The spectrum of cutaneous mycoses ranges from harmless superficial infections up to extensive inflammatory or chronic recurrent trajectories. A differentiated consideration of the spectrum of pathogens, age-related clinical signs, appropriate diagnostic procedures and individually adapted treatment options is essential.

Topical antiseptics are crucial for preventing infections and reducing transmission of pathogens. However, commonly used antiseptic agents have been reported to cause cross-resistance to other antimicrobials in bacteria, which has not yet been described in yeasts. This study aims to assess the in vitro efficacy of antiseptics against clinical and reference isolates of Candida albicans and Nakaseomyces glabratus, and whether prolonged exposure to antiseptics promotes the development of antifungal (cross)resistance. A high-throughput approach for in vitro resistance development was established to simultaneously expose 96 C. albicans and N. glabratus isolates to increasing concentrations of a given antiseptic – chlorhexidine, triclosan or octenidine. Susceptibility testing and whole genome sequencing of yeast isolates pre- and post-exposure were performed. Long-term exposure to antiseptics does not result in the development of stable resistance to the antiseptics themselves. However, 50 N. glabratus isolates acquired resistance to azole antifungals after long-term exposure to triclosan or chlorhexidine, revealing newly acquired mutations in the PDR1 and PMA1 genes. Chlorhexidine as well as triclosan, but not octenidine, were able to introduce selective pressure promoting resistance to azole antifungals. Although we assessed this phenomenon only in vitro, these findings warrant critical monitoring in clinical settings.

Septic arthritis (SA) in adults can lead to serious complications if not diagnosed promptly. Conventional synovial fluid culture (CC) remains the gold standard for identifying the causing microorganism but is time-consuming and often insensitive, particularly in patients receiving antimicrobial therapy. This study aimed to evaluate the diagnostic performance of a novel automated multiplex PCR (mPCR) system and compared it to conventional culture (CC) in adults with suspected acute native joint infections. In this retrospective single-centre study, adult patients with suspected SA (February 2023-May 2024) were included. Diagnosis was based on institutional criteria incorporating clinical signs, synovial fluid cytological, microbiology, and histology. Agreement between mpCR and CC was assessed using overall percentage agreement and Cohen's Kappa coefficient. Diagnostic performance metrics were calculated for mPCR, CC, and their combined use. Of 143 included patients, 96 (67%) were diagnosed with SA. When considering mPCR-specific microorganisms, mPCR identified 13 additional microorganisms compared to CC. Nine of these (9/13) were diagnosed with SA and six of these (6/9, 67%) were on antibiotics prior to aspiration. Overall agreement between mPCR and CC was 91%, with a positive agreement of 100%, negative agreement of 88% and a Cohen's Kappa coefficient of 0.780. Considering all microorganisms (including off-panel organisms), the overall agreement was 89%, the positive agreement 92%, the negative agreement 88%, and the Cohen's Kappa 0.735. The mPCR demonstrated a sensitivity of 45% and specificity of 89%, while conventional culture showed a sensitivity of 40% and specificity of 100%. No significant difference in performance was observed between the two methods (p = 0.183). Moreover, the combined use (mPCR + CC) yielded a sensitivity of 48% and specificity of 89% (AUC = 0.686). The novel automated mPCR system demonstrated a diagnostic performance similar to that of conventional synovial fluid culture, offering the added benefit of a quicker turnaround time, which can be crucial for patient care. Its use is especially evident in patients who have received prior antibiotic treatment, where conventional cultures may be less reliable.

Aim: The aim of this study was to investigate the diagnostic performance of a novel rapid multiplex polymerase chain reaction (mPCR) in adults with suspected acute native joint infection. Methods: This retrospective single-centre study included 143 patients with suspected acute native joint infection from February 2023 to May 2024. A septic arthritis was classified based on institutional criteria. The agreement between mPCR and conventional culture of synovial fluid (SF) was assessed by calculating the Cohen's κ coefficient. The diagnostic performance of mPCR was calculated, and the area under the curve (AUC) was compared with conventional culture of synovial fluid by using the z test. Results: When considering only microorganisms targeted by mPCR, this method detected 13 novel microorganisms in 13 cases compared to conventional culture, resulting in an overall agreement of 91 %, a positive agreement of 100 %, a negative agreement of 88 %, and a Cohen's κ coefficient of 0.780. Of these 13 cases, 9 were classified as septic, with 6 (n=6/9, 67 %) on antibiotics prior to aspiration. When considering all microorganisms (including off-panel microorganisms), the overall percentage agreement between mPCR and conventional culture was 89 %, with a Cohen's κ coefficient of 0.735, indicating substantial agreement. Sensitivity, specificity, PPV, NPV, LR+, LR−, accuracy, and AUC of mPCR were 45 %, 89 %, 90 %, 44 %, 4.21, 0.62, 59 %, and 0.671, and those of conventional culture were 40 %, 100 %, 100 %, 45 %, 0.60, 59 %, and 0.698. No difference in performance was observed between both methods (p=0.183). The combination of both techniques showed a sensitivity, specificity, PPV, NPV, LR+, LR−, accuracy, and AUC of 48 %, 89 %, 90 %, 46 %, 4.5, 0.58, 62 %, and 0.686. Conclusion: Given its comparable diagnostic performance and faster turnaround time relative to conventional synovial fluid culture, this novel mPCR can be recommended as a valuable adjunct in the diagnosis of septic arthritis in adults, particularly in patients with prior antimicrobial treatment.

In recent years there has been an increase in the diagnoses of sexually transmitted infections (STI) among men who have sex with men (MSM) using human immunodeficiency virus (HIV) pre-exposure prophylaxis (PrEP); however, data on PrEP users in Austria are limited. In June 2020, we initiated a prospective observational cohort study at Vienna General Hospital including PrEP users from Vienna. Participants underwent STI testing quarterly and provided behavioral information using a questionnaire. Between June 2020 and December 2023 a total of 360 individuals (99

Benzylpenicillin is regaining attention as a treatment option for susceptible S. aureus, including in severe invasive diseases such as blood stream infections. Timely and reliable susceptibility determination is essential to support its use in clinical practice. In this study, we assessed the EUCAST-recommended methodology of interpreting zone edges in a national multicenter trial. In total, nine microbiology laboratories in Austria participated. Each center received 10 isolates in blinded duplicates, all with inhibition zones of ≥ 26 mm. Three were blaZ-positive with sharp edges and seven were blaZ-negative with fuzzy edges. Benzylpenicillin susceptibility testing according to EUCAST guidelines using 1 unit discs was performed by two independent technicians in duplicate on two separate days. All plates were interpreted by two different assessors generating a total of 1440 data points. Overall, 85.5

Abstract Background Despite advances in antifungals, Candida infections still have a high mortality rate of up to 40%. The ECMM Candida III study in Europe investigated the changing epidemiology and outcomes, highlighting the need to understand and manage these infections. Methods In this observational cohort study, participating hospitals enrolled the first ten consecutive adults with confirmed candidemia. Data collected included patient demographics, risk factors, hospital stay length (with a 90-day follow-up), diagnostic procedures, Candida species, treatment details, and outcome. Controls were matched in a 1:1 ratio from the same hospitals, ensuring similarity in age, underlying illness, ICU versus normal ward stay, and recent major surgery. The study described overall and attributable mortality and assessed survival probability for both cases and controls. Results The study included 171 pairs consisting of patients with candidemia and matched controls from 28 institutions. In those with candidemia, overall mortality was 40.4%. The attributable mortality was 18.1% overall but differed among the causative Candida species (7.7% for Candida albicans, 23.7% for Candida glabrata, 7.7% for Candida parapsilosis, and 63.6% for Candida tropicalis). Regarding risk factors, the presence of central venous catheter, total parenteral nutrition, and acute or chronic kidney disease were significantly more common in cases versus controls. Length of hospitalization and ICU stay were significantly longer in candidemia cases (20 days (IQR 10-33) vs. 15 days (IQR 7-28); p=0.004). Conclusion Although overall mortality remains high in this matched case/control analysis, attributable mortality has decreased compared to historical cohorts. This may be due to a better prognosis for candidemia caused by Candida albicans, which has an attributable mortality of 7.7%, while candidemia cases caused by non-albicans Candida exhibit higher attributable mortality. Disclosures Oliver A. Cornely, Prof. Dr., Abbott: Honoraria|Abbvie: Advisor/Consultant|Abbvie: Honoraria|AiCuris: Advisor/Consultant|Akademie fur Infektionmedizin: Honoraria|Al-Jazeera Pharmaceuticals/Hikma: Honoraria|amedes: Honoraria|AstraZeneca: Honoraria|Basilea: Advisor/Consultant|Biocon: Advisor/Consultant|BMBF: Grant/Research Support|Boston Strategic Partners: Advisor/Consultant|CIdara: Advisor/Consultant|CIdara: Expert Testimony|CIdara: Grant/Research Support|CIdara: Participation on a DRC or DSMB|CoRe Consulting: Stocks/Bonds (Private Company)|Deutscher Arzteverlag: Honoraria|DZIF: Grant/Research Support|EasyRadiology: Stocks/Bonds (Private Company)|EU-DG RTD: Grant/Research Support|F2G: Grant/Research Support|Gilead: Advisor/Consultant|Gilead: Grant/Research Support|Gilead: Honoraria|Grupo Biotoscana/United Medical/Knight: Honoraria|GSK: Advisor/Consultant|GSK: Honoraria|IQVIA: Advisor/Consultant|IQVIA: Participation on a DRC or DSMB|Janssen: Advisor/Consultant|Janssen: Participation on a DRC or DSMB|Matinas: Advisor/Consultant|MedPace: Advisor/Consultant|MedPace: Grant/Research Support|MedPace: Participation on a DRC or DSMB|Medscape/WebMD: Honoraria|MedUpdate: Honoraria|Menarini: Advisor/Consultant|Moderna: Honoraria|Molecular Partners: Advisor/Consultant|MSD: Grant/Research Support|MSD: Honoraria|MSG-ERC: Advisor/Consultant|Mundipharma: Advisor/Consultant|Mundipharma: Grant/Research Support|Mundipharma: Honoraria|Noscendo: Honoraria|Noxxon: Advisor/Consultant|Octapharma: Advisor/Consultant|Octapharma: Grant/Research Support|Pardes: Advisor/Consultant|Partner Therapeutics: Advisor/Consultant|Patent: US18/562644|Paul-Martini-Stiftung: Honoraria|Pfizer: Advisor/Consultant|Pfizer: Grant/Research Support|Pfizer: Honoraria|PSI: Advisor/Consultant|PSI: Participation on a DRC or DSMB|Pulmocide: Participation on a DRC or DSMB|Sandoz: Honoraria|Scynexis: Advisor/Consultant|Scynexis: Grant/Research Support|Seqirus: Advisor/Consultant|Seqirus: Honoraria|Seres: Advisor/Consultant|Shionogi: Advisor/Consultant|Shionogi: Honoraria|streamedup!: Honoraria|The Prime Meridian Group: Advisor/Consultant|Touch Independent: Honoraria|Vitis: Honoraria Jean-Pierre Gangneux, Prof., Gilead: Honoraria|MundiPharma: Advisor/Consultant|MundiPharma: Honoraria|Pfizer: Honoraria|Shionogi: Honoraria Matteo Bassetti, PhD, Angelini: Advisor/Consultant|Angelini: Honoraria|Astellas: Advisor/Consultant|Astellas: Honoraria|bioMerieux: Advisor/Consultant|bioMerieux: Honoraria|Cidara: Advisor/Consultant|Cidara: Honoraria|Gilead: Advisor/Consultant|Gilead: Honoraria|Menarini: Advisor/Consultant|Menarini: Honoraria|MSD: Advisor/Consultant|MSD: Honoraria|Nabriva: Advisor/Consultant|Nabriva: Honoraria|Pfizer: Advisor/Consultant|Pfizer: Honoraria|Tetraphase: Advisor/Consultant|Tetraphase: Honoraria

Invasive candidiasis, including candidaemia, is associated with high morbidity and mortality. Diagnosis is traditionally based on blood culture, which lacks sensitivity. Therefore, additional tools such as PCR-based diagnostic methods are increasingly used. The T2MR technology is based on polymerase chain reaction and detection of the PCR product involving magnetic resonance technology. In this study, we compare the T2Candida in a clinical routine setting to conventional blood culture in order to explore its usefulness, strengths and weaknesses in its daily application. This retrospective analysis was performed at the Vienna University Hospital with clinical routine samples submitted between April 2021 and May 2024. Sensitivity, specificity, positive predictive value, negative predictive value and accordance were calculated with blood culture as reference method. Patients with a positive T2Candida result but a negative result in blood culture were assessed according to a clinical case definition. Based on direct detection in blood by alternative methods (e.g. blood culture, alternative PCR), 1-3-beta-D-Glucan, patient risk factors and detection of the same species in other sample materials, each result was categorised as “proven”, “probable”, “possible”, “improbable” or “not assessable”. 2105 samples from 1447 unique patients were submitted for analysis during the study period. 94 samples were positive (4.5

Fungi associated with humans include several Candida species that rely on phenotypic plasticity for persistence and pathogenicity. Key adaptive traits, such as adherence, stress resistance, and biofilm formation, enable survival in diverse host niches. However, the degree of intra- and interspecific phenotypic variation across human-associated Candida species has not been systematically characterized. We analyzed 1,366 clinical isolates representing 13 Candida species using high-throughput quantitative fitness profiling under controlled environmental stressors, antifungal exposure, and biofilm-inducing conditions. The resulting data set revealed both conserved and species-specific adaptive signatures. Isolates consistently segregated into three phenotypic archetypes: heat-resistant fast growers, osmo-sensitive strains, and slow growers. A robust inverse correlation was detected between basal growth rate and stress resistance, reflecting a fundamental physiological trade-off. In addition, distinct resistance profiles against antifungal agents and environmental stressors highlighted species-specific adaptive trajectories and ecological specialization. Despite genetic homogeneity, C. parapsilosis isolates displayed striking phenotypic heterogeneity. By contrast, the closely related C. albicans and C. dubliniensis exhibited divergent stress-response profiles. High-resolution fitness mapping of C. glabrata isolates revealed that temperature stress progressively disrupts multiple cellular functions, whereas osmotic stress exerts more discrete, pathway-specific effects. Our systematic phenotypic landscape analysis delineates conserved versus species-specific adaptive properties among human-associated Candida species, providing a comparative framework to interrogate evolutionary trends, ecological specialization, and pathogenic potential. IMPORTANCE:Human-associated fungi include multiple Candida species whose persistence relies on phenotypic plasticity enabling adherence, stress resistance, and biofilm formation. Yet, the extent of phenotypic variation within and across species remains poorly defined. We profiled 1,366 clinical isolates from 13 Candida species using high-throughput quantitative fitness assays under environmental stress, antifungal exposure, and biofilm-inducing conditions. The analysis uncovered both conserved and species-specific adaptive traits. Isolates segregated into three major phenotypic archetypes: heat-resistant fast growers, osmo-sensitive strains, and slow growers. A consistent inverse correlation emerged between basal growth rate and stress resistance, revealing a fundamental physiological trade-off. Species-specific resistance signatures further reflected ecological specialization and divergent adaptive trajectories. Our quantitative framework establishes, for the first time, a comparative phenotypic landscape across a multispecies collection of human-associated Candida, providing new insights into their ecological specialization and adaptive strategies.

BACKGROUND:Identification of filamentous fungi still poses a major challenge to laboratories. Matrix assisted laser desorption/ionisation time of flight mass spectrometry (MALDI-TOF MS) is a promising tool, since it offers low-cost and fast results. Different MALDI-TOF MS systems are available for routine laboratories. This is a comprehensive head-to-head comparison of three devices and the respective reference spectrum databases. MATERIALS AND METHODS:A set of 77 pre-characterised isolates of filamentous fungi was measured (in duplicates) parallelly with three MALDI-TOF MS systems after 24 h, 48 h and 72 h of incubation: (1) Biotyper smart ('BT', Bruker Daltonics),(2) EXS2600 ('EXS', Zybio) and (3) VITEK MS PRIME ('VITEK', bioMérieux). RESULTS:After three measurements, no valid results at the species level ('green category') were obtained for 18%, 21% and 14% of isolates by BT, EXS and VITEK. Depending on the MALDI-TOF MS system, validity rates ranged from 58%-82% for the different time points. Correct and valid results were obtained for 82%, 73% and 81% of isolates by BT, EXS and VITEK. BT was the system that required the most duplicate measurements. EXS displayed the highest rate of misidentification events. VITEK had the highest rate of unidentified isolates, which were featured within its database. CONCLUSION:Based on our experience, all three devices proved to be suitable for routine diagnostics. The timepoint of measurement had a major impact on the quality of analysis and should be considered by the user. Lowering the validity cut-off levels might increase the performance of the EXS system.