Objectives: While Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA), defined as CC398, is a well-known pathogen among those working with livestock, there are indications... Show moreObjectives: While Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA), defined as CC398, is a well-known pathogen among those working with livestock, there are indications that LA-MRSA prevalence among the general population is increasing. However, the clinical impact in urban areas remains unknown. The aim of this study was to assess the genetic epidemiology and clinical characteristics of LA-MRSA in an urban area with a limited livestock population. Methods: In this retrospective study, we evaluated LA-MRSA strains that were collected between 2014 and 2018 from patients who received clinical care in a single urban area in Netherlands. Patient files were assessed for livestock exposure data, clinical findings, and contact tracing information. Next-generation sequencing (NGS) analysis in combination with wgMLST was conducted to assess genetic diversity and relatedness and to detect virulence and resistance genes. Results: LA-MRSA strains were cultured from 81 patients, comprising 12% of all the MRSA strains found in seven study laboratories between 2014 and 2018. No livestock link was found in 76% of patients (n = 61), and 28% of patients (n = 23) had an infection, mostly of the skin or soft tissue. Contact tracing had been initiated in 14 cases, leading to the identification of two hospital transmissions: a cluster of 9 cases and one of 2 cases. NGS data were available for 91% (n = 75) of the patients. wgMLST confirmed the clusters detected via contact tracing (n = 2) and identified 5 additional clusters without a known epidemiological link. Relevant resistance and virulence findings included the PVL virulence gene (3 isolates) and tetracycline resistance (79 isolates). Conclusion: LA-MRSA may cause a relevant burden of disease in urban areas. Surprisingly, most infections in the present study occurred in the absence of a livestock link, suggesting inter-human transmission. These findings and the presence of PVL and other immune evasive complex virulence genes warrant future surveillance and preventative measures. Show less
Objectives: While Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA), defined as CC398, is a well-known pathogen among those working with livestock, there are indications... Show moreObjectives: While Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA), defined as CC398, is a well-known pathogen among those working with livestock, there are indications that LA-MRSA prevalence among the general population is increasing. However, the clinical impact in urban areas remains unknown. The aim of this study was to assess the genetic epidemiology and clinical characteristics of LA-MRSA in an urban area with a limited livestock population.Methods: In this retrospective study, we evaluated LA-MRSA strains that were collected between 2014 and 2018 from patients who received clinical care in a single urban area in Netherlands. Patient files were assessed for livestock exposure data, clinical findings, and contact tracing information. Next-generation sequencing (NGS) analysis in combination with wgMLST was conducted to assess genetic diversity and relatedness and to detect virulence and resistance genes.Results: LA-MRSA strains were cultured from 81 patients, comprising 12% of all the MRSA strains found in seven study laboratories between 2014 and 2018. No livestock link was found in 76% of patients (n = 61), and 28% of patients (n = 23) had an infection, mostly of the skin or soft tissue. Contact tracing had been initiated in 14 cases, leading to the identification of two hospital transmissions: a cluster of 9 cases and one of 2 cases. NGS data were available for 91% (n = 75) of the patients. wgMLST confirmed the clusters detected via contact tracing (n = 2) and identified 5 additional clusters without a known epidemiological link. Relevant resistance and virulence findings included the PVL virulence gene (3 isolates) and tetracycline resistance (79 isolates).Conclusion: LA-MRSA may cause a relevant burden of disease in urban areas. Surprisingly, most infections in the present study occurred in the absence of a livestock link, suggesting inter-human transmission. These findings and the presence of PVL and other immune evasive complex virulence genes warrant future surveillance and preventative measures. Show less
Clostridioides difficile is the most common cause of antibiotic-associated gastrointestinal infections. Capillary electrophoresis (CE)-PCR ribotyping is currently the gold standard for C. difficile... Show moreClostridioides difficile is the most common cause of antibiotic-associated gastrointestinal infections. Capillary electrophoresis (CE)-PCR ribotyping is currently the gold standard for C. difficile typing but lacks the discriminatory power to study transmission and outbreaks in detail. New molecular methods have the capacity to differentiate better and provide standardized and interlaboratory exchangeable data. Using a well-characterized collection of diverse strains (N = 630; 100 unique ribotypes [RTs]), we compared the discriminatory power of core genome multilocus sequence typing (cgMLST) (SeqSphere and EnteroBase), whole-genome MLST (wgMLST) (EnteroBase), and single-nucleotide polymorphism (SNP) analysis. A unique cgMLST profile (more than six allele differences) was observed in 82 of 100 RTs, indicating that cgMLST could distinguish most, but not all, RTs. Application of cgMLST in two outbreak settings with RT078 and RT181 (known to have low intra-RT allele differences) showed no distinction between outbreak and nonoutbreak strains in contrast to wgMLST and SNP analysis. We conclude that cgMLST has the potential to be an alternative to CE-PCR ribotyping. The method is reproducible, easy to standardize, and offers higher discrimination. However, adjusted cutoff thresholds and epidemiological data are necessary to recognize outbreaks of some specific RTs. We propose to use an allelic threshold of three alleles to identify outbreaks. Show less
Wegner, F.; Roloff, T.; Huber, M.; Cordey, S.; Ramette, A.; Gerth, Y.; ... ; Egli, A. 2022
This first pilot trial on external quality assessment (EQA) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) whole-genome sequencing, initiated by the European Society of Clinical... Show moreThis first pilot trial on external quality assessment (EQA) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) whole-genome sequencing, initiated by the European Society of Clinical Microbiology and Infectious Diseases (ESCMID) Study Group for Genomic and Molecular Diagnostics (ESGMD) and the Swiss Society for Microbiology (SSM), aims to build a framework between laboratories in order to improve pathogen surveillance sequencing. Ten samples with various viral loads were sent out to 15 clinical laboratories that had free choice of sequencing methods and bioinformatic analyses. The key aspects on which the individual centers were compared were the identification of (i) single nucleotide polymorphisms (SNPs) and indels, (ii) Pango lineages, and (iii) clusters between samples. The participating laboratories used a wide array of methods and analysis pipelines. Most were able to generate whole genomes for all samples. Genomes were sequenced to various depths (up to a 100-fold difference across centers). There was a very good consensus regarding the majority of reporting criteria, but there were a few discrepancies in lineage and cluster assignments. Additionally, there were inconsistencies in variant calling. The main reasons for discrepancies were missing data, bioinformatic choices, and interpretation of data. The pilot EQA was overall a success. It was able to show the high quality of participating laboratories and provide valuable feedback in cases where problems occurred, thereby improving the sequencing setup of laboratories. A larger follow-up EQA should, however, improve on defining the variables and format of the report. Additionally, contamination and/or minority variants should be a further aspect of assessment. Show less
Simple SummaryAlthough granulosa cell tumors can occur in rare syndromes and one familial case of a granulosa cell tumor has been described, a genetic predisposition for granulosa cell tumors has... Show moreSimple SummaryAlthough granulosa cell tumors can occur in rare syndromes and one familial case of a granulosa cell tumor has been described, a genetic predisposition for granulosa cell tumors has not been identified. Through our collaborations with patients, we identified four families in which two women of each family were diagnosed with an adult granulosa cell tumor. Although predicted deleterious variants in PIK3C2G, BMP5, and LRP2 were found, we could not identify an overlapping genetic variant or affected locus that may explain a genetic predisposition for granulosa cell tumors. The age of onset in the familial patients was significantly lower (median 38 years, range from 17 to 60) than in sporadic patients (median between 50 and 55 years). Furthermore, breast cancer, polycystic ovary syndrome, and subfertility were seen in these families.Adult granulosa cell tumor (AGCT) is a rare ovarian cancer subtype, with a peak incidence around 50-55 years. Although AGCT can occur in specific syndromes, a genetic predisposition for AGCT has not been identified. The aim of this study is to identify a genetic variant in families with AGCT patients, potentially contributing to tumor evolution. We identified four families, each including two women diagnosed with AGCT. Whole-genome sequencing was performed to identify overlapping germline variants or affected genes. Familial relationship was evaluated using genealogy and genomic analyses. Patient characteristics, medical (family) history, and pedigrees were collected. Findings were compared to a reference group of 33 unrelated AGCT patients. Mean age at diagnosis was 38 years (range from 17 to 60) versus 51 years in the reference group, and seven of eight patients were premenopausal. In two families, three first degree relatives were diagnosed with breast cancer. Furthermore, polycystic ovary syndrome (PCOS) and subfertility was reported in three families. Predicted deleterious variants in PIK3C2G, BMP5, and LRP2 were identified. In conclusion, AGCTs occur in families and could potentially be hereditary. In these families, the age of AGCT diagnosis is lower and cases of breast cancer, PCOS, and subfertility are present. We could not identify an overlapping genetic variant or affected locus that may explain a genetic predisposition for AGCT. Show less
Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare and highly aggressive hematological malignancy with a poorly understood pathobiology and no effective therapeutic options. Despite a... Show moreBlastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare and highly aggressive hematological malignancy with a poorly understood pathobiology and no effective therapeutic options. Despite a few recurrent genetic defects (eg, single nucleotide changes, indels, large chromosomal aberrations) have been identified in BPDCN, none are disease-specific, and more importantly, none explain its genesis or clinical behavior. In this study, we performed the first high resolution whole-genome analysis of BPDCN with a special focus on structural genomic alterations by using whole-genome sequencing and RNA sequencing. Our study, the first to characterize the landscape of genomic rearrangements and copy number alterations of BPDCN at nucleotide-level resolution, revealed that IKZF1, a gene encoding a transcription factor required for the differentiation of plasmacytoid dendritic cell precursors, is focally inactivated through recurrent structural alterations in this neoplasm. In concordance with the genomic data, transcriptome analysis revealed that conserved IKZF1 target genes display a loss-of-IKZF1 expression pattern. Furthermore, up-regulation of cellular processes responsible for cell-cell and cell-ECM interactions, which is a hallmark of IKZF1 deficiency, was prominent in BPDCN. Our findings suggest that IKZF1 inactivation plays a central role in the pathobiology of the disease, and consequently, therapeutic approaches directed at reestablishing the function of this gene might be beneficial for patients. Show less
