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Group A streptococcal (GAS; aka Streptococcus pyogenes) meningitis is a fulminant disease associated with high morbidity and mortality. To elucidate the mechanisms underlying the invasiveness of GAS in meningitis, we compared GAS isolates derived from five cases of meningitis to otitis and colonizing isolates. We did not observe differences in adherence to and invasion of human brain microvascular endothelial cells, virulence factors activity, or barrier disruption. Whole genome sequencing did not reveal particular invasiveness traits. Most patients previously suffered from otitis media suggesting that meningitis likely resulted from a continuous spread of the infection rather than being attributable to changes in the pathogen's virulence.

Staphylococcus aureus carriage is a risk factor for invasive infections. Unique genetic elements favoring the transition from colonizing to invasive phenotype have not yet been identified, and phenotypic adaptation traits are understudied. We therefore assessed phenotypic and genotypic profiles of 11 S. aureus isolate pairs sampled from colonized patients simultaneously suffering from invasive S. aureus infections. Ten out of 11 isolate pairs displayed the same spa and multilocus sequence type, suggesting colonization as an origin for the invasive infection. Systematic analysis of colonizing and invasive isolate pairs showed similar adherence, hemolysis, reproductive fitness properties, antibiotic tolerance, and virulence in a Galleria mellonella infection model, as well as minimal genetic differences. Our results provide insights into the similar phenotypes associated with limited adaptation between colonizing and invasive isolates. Disruption of the physical barriers of mucosa or skin was identified in the majority of patients, further emphasizing colonization as a major risk factor for invasive disease. IMPORTANCE S. aureus is a major pathogen of humans, causing a wide range of diseases. The difficulty to develop a vaccine and antibiotic treatment failure warrant the exploration of novel treatment strategies. Asymptomatic colonization of the human nasal passages is a major risk factor for invasive disease, and decolonization procedures have been effective in preventing invasive infections. However, the transition of S. aureus from a benign colonizer of the nasal passages to a major pathogen is not well understood, and both host and bacterial properties have been discussed as being relevant for this behavioral change. We conducted a thorough investigation of patient-derived strain pairs reflecting colonizing and invasive isolates in a given patient. Although we identified limited genetic adaptation in certain strains, as well as slight differences in adherence cap

Background: Pseudomonas aeruginosa and other Gram-negative bacteria have the ability to persist in moist environments in healthcare settings, but their spread from these areas can result in outbreaks of healthcare-associated infections. Methods: This study reports the investigation and containment of a multi-drug-resistant P. aeruginosa outbreak in three intensive care units of a Swiss university hospital. In total, 255 patients and 276 environmental samples were screened for the multi-drug-resistant P. aeruginosa outbreak strain. The environmental sampling and molecular characterization of patient and environmental strains, and control strategies implemented, including waterless patient care, are described. Results: Between March and November 2019, the outbreak affected 29 patients. Environmental sampling detected the outbreak strain in nine samples of sink siphons of three different intensive care units with a common water sewage system, and on one gastroscope. Three weeks after replacement of the sink siphons, the outbreak strain re-grew in siphon-derived samples and newly affected patients were identified. The outbreak ceased after removal of all sinks in the proximity of patients and in medication preparation areas, and minimization of tap water use. Multi-locus sequence typing indicated clonality (sequence type 316) in 28/29 patient isolates and all 10 environmental samples. Conclusions: Sink removal combined with the introduction of waterless patient care terminated the multi-drug-resistant P. aeruginosa outbreak. Sinks in intensive care units may pose a risk for point source outbreaks with P. aeruginosa and other bacteria persisting in moist environments. Keywords: Intensive care; Multi-drug resistance; Outbreak; Pseudomonas aeruginosa; Sink; Siphon; Waterless patient care.

Healthcare-associated outbreaks of vancomycin-resistant Enterococcus faecium (VREfm) are a worldwide problem with increasing prevalence. The genomic plasticity of this hospital-adapted pathogen contributes to its efficient spread despite infection control measures. Here, we aimed to identify the genomic and phenotypic determinants of health care-associated transmission of VREfm. We assessed the VREfm transmission networks at the tertiary-care University Hospital of Zurich (USZ) between October 2014 and February 2018 and investigated microevolutionary dynamics of this pathogen. We performed whole-genome sequencing for the 69 VREfm isolates collected during this time frame and assessed the population structure and variability of the vancomycin resistance transposon. Phylogenomic analysis allowed us to reconstruct transmission networks and to unveil external or wider transmission networks undetectable by routine surveillance. Notably, it unveiled a persistent clone, sampled 31 times over a 29-month period. Exploring the evolutionary dynamics of this clone and characterizing the phenotypic consequences revealed the spread of a variant with decreased daptomycin susceptibility and the acquired ability to utilize N-acetyl-galactosamine (GalNAc), one of the primary constituents of the human gut mucins. This nutrient utilization advantage was conferred by a novel plasmid, termed pELF_USZ, which exhibited a linear topology. This plasmid, which was harbored by two distinct clones, was transferable by conjugation. Overall, this work highlights the potential of combining epidemiological, functional genomic, and evolutionary perspectives to unveil adaptation strategies of VREfm. IMPORTANCE Sequencing microbial pathogens causing outbreaks has become a common practice to characterize transmission networks. In addition to the signal provided by vertical evolution, bacterial genomes harbor mobile genetic elements shared horizontally between clones. While macroevolutionary studies hav

Treatment failure in biofilm-associated bacterial infections is an important healthcare issue. In vitro studies and mouse models suggest that bacteria enter a slow-growing/non-growing state that results in transient tolerance to antibiotics in the absence of a specific resistance mechanism. However, little clinical confirmation of antibiotic tolerant bacteria in patients exists. In this study we investigate a Staphylococcus epidermidis pacemaker-associated endocarditis, in a patient who developed a break-through bacteremia despite taking antibiotics to which the S. epidermidis isolate is fully susceptible in vitro. Characterization of the clinical S. epidermidis isolates reveals in-host evolution over the 16-week infection period, resulting in increased antibiotic tolerance of the entire population due to a prolonged lag time until growth resumption and a reduced growth rate. Furthermore, we observe adaptation towards an increased biofilm formation capacity and genetic diversification of the S. epidermidis isolates within the patient.