Historic methicillin-resistant Staphylococcus aureus: expanding current knowledge using molecular epidemiological characterization of a Swiss legacy collection.

BACKGROUND: Few methicillin-resistant Staphylococcus aureus (MRSA) from the early years of its global emergence have been sequenced. Knowledge about evolutionary factors promoting the success of specific MRSA multi-locus sequence types (MLSTs) remains scarce. We aimed to characterize a legacy MRSA collection isolated from 1965 to 1987 and compare it against publicly available international and local genomes. METHODS: We accessed 451 historic (1965-1987) MRSA isolates stored in the Culture Collection of Switzerland, mostly collected from the Zurich region. We determined phenotypic antimicrobial resistance (AMR) and performed whole genome sequencing (WGS) using Illumina short-read sequencing on all isolates and long-read sequencing on a selection with Oxford Nanopore Technology. For context, we included 103 publicly available international assemblies from 1960 to 1992 and sequenced 1207 modern Swiss MRSA isolates from 2007 to 2022. We analyzed the core genome (cg)MLST and predicted SCCmec cassette types, AMR, and virulence genes. RESULTS: Among the 451 historic Swiss MRSA isolates, we found 17 sequence types (STs) of which 11 have been previously described. Two STs were novel combinations of known loci and six isolates carried previously unsubmitted MLST alleles, representing five new STs (ST7843, ST7844, ST7837, ST7839, and ST7842). Most isolates (83% 376/451) represented ST247-MRSA-I isolated in the 1960s, followed by ST7844 (6% 25/451), a novel single locus variant (SLV) of ST239. Analysis by cgMLST indicated that isolates belonging to ST7844-MRSA-III cluster within the diversity of ST239-MRSA-III. Early MRSA were predominantly from clonal complex (CC)8. From 1980 to the end of the twentieth century, we observed that CC22 and CC5 as well as CC8 were present, both locally and internationally. CONCLUSIONS: The combined analysis of 1761 historic and contemporary MRSA isolates across more than 50 years uncovered novel STs and allowed us a glimpse into the lineage flux between Swiss-German and international MRSA across time.

Quality of MALDI-TOF mass spectra in routine diagnostics: results from an international external quality assessment including 36 laboratories from 12 countries using 47 challenging bacterial strains.

OBJECTIVES: Matrix assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) is a widely used method for bacterial species identification. Incomplete databases and mass spectral quality (MSQ) still represent major challenges. Important proxies for MSQ are the number of detected marker masses, reproducibility, and measurement precision. We aimed to assess MSQs across diagnostic laboratories and the potential of simple workflow adaptations to improve it. METHODS: For baseline MSQ assessment, 47 diverse bacterial strains, which are challenging to identify by MALDI-TOF MS, were routinely measured in 36 laboratories from 12 countries, and well-defined MSQ features were used. After an intervention consisting of detailed reported feedback and instructions on how to acquire MALDI-TOF mass spectra, measurements were repeated and MSQs were compared. RESULTS: At baseline, we observed heterogeneous MSQ between the devices, considering the median number of marker masses detected (range = [2-25]), reproducibility between technical replicates (range = [55%-86%]), and measurement error (range = [147 parts per million (ppm)-588 ppm]). As a general trend, the spectral quality was improved after the intervention for devices, which yielded low MSQs in the baseline assessment as follows: for four out of five devices with a high measurement error, the measurement precision was improved (p-values <0.001, paired Wilcoxon test); for six out of ten devices, which detected a low number of marker masses, the number of detected marker masses increased (p-values <0.001, paired Wilcoxon test). DISCUSSION: We have identified simple workflow adaptations, which, to some extent, improve MSQ of poorly performing devices and should be considered by laboratories yielding a low MSQ. Improving MALDI-TOF MSQ in routine diagnostics is essential for increasing the resolution of bacterial identification by MALDI-TOF MS, which is dependent on the reproducible detection of marker masses. The heterogeneity identified in this external quality assessment (EQA) requires further study.

[A very slow growing ankle swelling in a healthy male]. / Ein seltsamer subkutaner Knöcheltumor.

We describe the case report of a 66-year-old man with a very slow growing ankle tumour caused by a subcutaneous fungal abscess. Phaeoacremonium inflatipes, a member of the Dematiaceae family, was identified by needle puncture and culture of the non-odorous creamy yellow brown fluid. The fungal pseudocyst was surgically removed in toto and no further fungicidal drug therapy was required. Human infections by dematiaceous fungi causes subcutaneous phaeohyphomycosis, a rare, deep fungal infection of the skin and subcutaneous tissues usually acquired through traumatic skin lesions. In addition, systemic infections are reported, predominantly in immunosuppressed individuals.

Identification of dermatophytes by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

In this study we evaluated the suitability of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the identification of dermatophytes in diagnostic laboratories. First, a spectral database was built with 108 reference strains belonging to 18 species of the anamorphic genera Epidermophyton, Microsporum and Trichophyton. All strains were well characterized by morphological criteria and ITS sequencing (gold standard). The dendrogram resulting from MALDI-TOF mass spectra was almost identical with the phylogenetic tree based on ITS sequencing. Subsequently, MALDI-TOF MS SuperSpectra were created for the identification of Epidermophyton floccosum, Microsporium audouinii, M. canis, M. gypseum (teleomorph: Arthroderma gypseum), M. gypseum (teleomorph: A. incurvatum), M. persicolor, A. benhamiae (Tax. Entity 3 and Am-Eur. race), T. erinacei, T. interdigitale (anthropophilic and zoophilic populations), T. rubrum/T. violaceum, T. tonsurans and T. terrestre. Because T. rubrum and T. violaceum did not present enough mismatches, a SuperSpectrum covering both species was created, and differentiation between them was done by comparison of eight specific peptide masses. In the second part of this study, MALDI-TOF MS with the newly created SuperSpectra was tested using 141 clinical isolates representing nine species. Analyses were done with 3-day-old cultures. Results were compared to morphological identification and ITS sequencing; 135/141 (95.8%) strains were correctly identified by MALDI-TOF MS compared to 128/141 (90.8%) by morphology. Therefore, MALDI-TOF MS has proven to be a useful and rapid identification method for dermatophytes.