Increase in invasive group A streptococcal infections and emergence of novel, rapidly expanding sub-lineage of the virulent Streptococcus pyogenes M1 clone, Denmark, 2023.

A highly virulent sub-lineage of the Streptococcus pyogenes M1 clone has been rapidly expanding throughout Denmark since late 2022 and now accounts for 30% of the new invasive group A streptococcal infections. We aimed to investigate whether a shift in variant composition can account for the high incidence rates observed over winter 2022/23, or if these are better explained by the impact of COVID-19-related restrictions on population immunity and carriage of group A Streptococcus.

Taxonomic reassessment of the genus Pseudocitrobacter using whole genome sequencing: Pseudocitrobacter anthropi is a later heterotypic synonym of Pseudocitrobacter faecalis and description of Pseudocitrobacter vendiensis sp. nov.

The taxonomic status of all Pseudocitrobacter species was re-evaluated by comparative genomics based on whole genome sequencing. As a result, it is obvious that Pseudocitrobacter anthropi is a later heterotypic synonym of Pseudocitrobacter faecalis. In addition, genome-based analysis of strain CPO20170097T, isolated from a patient in northern Denmark was allocated to the genus Pseudocitrobacter. This strain showed significant genotypic and phenotypic differences from P. faecalis and it is proposed that this strain represents a novel species of the genus, for which the name Pseudocitrobacter vendiensis sp. nov. is proposed with the type strain CPO20170097T (=CCUG 73096T=LMG 31042T).

Carrier prevalence and risk factors for colonisation of multiresistant bacteria in Danish emergency departments: a cross-sectional survey.

OBJECTIVES: The aim of this study was to describe the carrier prevalence and demographic variation of four different multiresistant bacteria (MRB) among acute patients in Danish emergency departments (EDs): methicillin-resistant Staphylococcus aureus (MRSA), carbapenemase-producing enterobacteria (CPE), extended-spectrum beta-lactamase-producing enterobacteria (ESBL) and vancomycin-resistant enterococci (VRE), and to analyse the association of MRB carriage to a range of potential risk factors. DESIGN: Multicentre descriptive and analytic cross-sectional survey. SETTING: Eight EDs and four clinical microbiology departments in Denmark. PARTICIPANTS: Adults visiting the ED. MAIN OUTCOME MEASURES: Swabs from nose, throat and rectum were collected and analysed for MRSA, ESBL, VRE and CPE. The primary outcome was the prevalence of MRB carriage, and secondary outcomes relation to risk factors among ED patients. RESULTS: We included 5117 patients in the study. Median age was 68 years (54-77) and gender was equally distributed. In total, 266 (5.2%, 95% CI 4.6 to 5.8) were colonised with at least one MRB. No significant difference was observed between male and female patients, between age groups and between university and regional hospitals. Only 5 of the 266 patients with MRB were colonised with two of the included bacteria and none with more than two. CPE prevalence was 0.1% (95% CI 0.0 to 0.2), MRSA prevalence was 0.3% (95% CI 0.2 to 0.5), VRE prevalence was 0.4% (95% CI 0.3 to 0.6) and ESBL prevalence was 4.5% (95% CI 3.9 to 5.1). Risk factors for MRB carriage were previous antibiotic treatment, previous hospital stay, having chronic respiratory infections, use of urinary catheter and travel to Asia, Oceania or Africa. CONCLUSION: Every 20th patient arriving to a Danish ED brings MRB to the hospital. ESBL is the most common MRB in the ED. The main risk factors for MRB carriage are recent antibiotic use and travel abroad. TRIAL REGISTRATION NUMBER: NCT03352167;Post-results.

Rare Elizabethkingia anophelis meningitis case in a Danish male.

INTRODUCTION: Elizabethkingia anophelis is a Gram-negative, aerobic, non-motile rod belonging to the family Flavobacteriaceae. Over the last 5 years, it has emerged as an opportunistic human pathogen involved in neonatal meningitis and sepsis, as well as nosocomial outbreaks. It has been isolated from the midgut of the Anopheles gambiae mosquito, but there is no evidence for a role of the mosquito in human infections, and very little is known regarding the routes of transmission to humans. Recent studies, primarily from South-East Asia, suggest that E. anophelis, and not Elizabethkingia meningoseptica, is the predominant human pathogen of this genus. However, identification to the species level has been difficult due to the limitations of the current MALDI-TOF MS (matrix-associated laser desorption ionization-time of flight MS) systems for correct species identification. CASE PRESENTATION: Here, we present a rare case of E. anophelis meningitis in a Danish male, who had a travel exposure to Malaysia 7 weeks before hospitalization. A multidrug-resistant Elizabethkingia species was isolated from blood and cerebrospinal fluid, and genomic sequencing was used to characterize the phylogenetic position of the isolate, which was determined as associated with previously described sublineage 11. The patient was successfully treated with intravenous moxifloxacin and rifampicin for 2 weeks with no major sequelae, but we did not find the source of transmission. CONCLUSION: All clinical microbiologists should be aware of the present limitations of the MALDI-TOF MS systems for correct species identification, and therefore we recommend the use of genome sequencing for the correct identification at the species and sublineage level.

Cross sectional study of multiresistant bacteria in Danish emergency departments: prevalence, patterns and risk factors for colonization (AB-RED project).

BACKGROUND: Multiresistant bacteria (MRB) is an increasing problem. Early identification of patients with MRB is mandatory to avoid transmission and to target the antibiotic treatment. The emergency department (ED) is a key player in the early identification of patients who are colonized with MRB. There is currently sparse knowledge of both prevalence and risk factors for colonization with MRSA, ESBL, VRE, CPE and CD in acutely admitted patients in Western European countries including Denmark. To develop evidence-based screening tools for identifying carriers of resistant bacteria among acutely admitted patients, systematic collection of information on risk factors and exposures is required. Since a geographical variation is suspected, it is desirable to include emergency departments across the country. The aim of this project is to provide a comprehensive overview of prevalence and risk factors for MRSA, ESBL, VRE, CPE and CD colonization in patients admitted to Danish ED's. The objectives are to describe the prevalence and demography of resistance, co-infections, to identify risk factors for carrier state and to develop and validate a screening tool for identification of carriers. METHODS: Multicenter descriptive and analytic cross-sectional survey from January-May 2018 of around 10.000 acutely admitted patients > 18 years in 8 EDs for carrier state and risk factors for antibiotic resistant bacteria. Information about the background and possible risk factors for carrier status together with swabs from the nose, throat and rectum is collected and analyzed for MRSA, ESBL, VRE, CPE and CD. The prevalence of the resistant bacteria are calculated at hospital level, regional level and national level and described with relation to residency, sex, age and risk factors. A screening model for identification of carrier stage of resistant bacteria is developed and validated. DISCUSSION: The study will provide the prevalence of colonized patients with resistant bacteria on arrival to the ED and variation in demographic patterns, and will develop a clinical tool to identify certain risk groups. This will enable the clinician to target antibiotic treatments and to reduce the in-hospital spreading of resistant bacteria. This knowledge is important for implementing and evaluating antimicrobial stewardships, screening and infection control strategies. TRIAL REGISTRATION: Clinicaltrials.gov : NCT03352167 (registration date: 20. November 2017).

Clostridium hastiforme bacteraemia secondary to pyometra in a 64-year-old woman.

The Gram-negative, rod-shaped, anaerobe bacteria Tissierella praeacuta was first described in 1908 by Tissier. However, during the past decade, Clostridium hastiforme has been recognised as a later synonym of T. praeacuta. C. hastiforme/T. praeacuta has only rarely been described in previous literature as a cause of human infection. We present here a case report of C. hastiforme/T. praeacuta bacteraemia secondary to pyometra in a 64-year-old woman with a history of multiple sclerosis and an intrauterine device inserted three decades earlier. C. hastiforme/T. praeacuta was isolated from blood as well as pus from the site of infection. The patient was cured with a combination of drainage and antibiotic therapy.

First report of metronidazole resistant, nimD-positive, Bacteroides stercoris isolated from an abdominal abscess in a 70-year-old woman.

We here present the first case of a metronidazole resistant nimD positive Bacteroides stercoris. The isolate originated from a polymicrobial intra-abdominal abscess in a 70-year-old woman. The nimD gene was detected by use of whole-genome shotgun sequencing and the subsequent use of the ResFinder 2.1 web service.

[Epidemic increase in the incidence of carbapenemase-producing Enterobacteriaceae in Denmark].

Carbapenemase-producing Enterobacteriaceae (CPE) are extremely multiresistant bacteria with few or no treatment options. Infections with CPE are associated with a mortality of 40-50%. In Denmark, CPE were first detected in 2008. Prior to 2013 the CPE incidence was low, but since then the incidence has increased significantly. Seven outbreaks have occurred, and at least 25 patients have been infected or colonized. The rise in CPE incidence emphasizes the need for a national intervention to reduce the spread. This can be obtained through systematic surveillance, infection control and reduction of antibiotic consumption.

Use of WGS data for investigation of a long-term NDM-1-producing Citrobacter freundii outbreak and secondary in vivo spread of blaNDM-1 to Escherichia coli, Klebsiella pneumoniae and Klebsiella oxytoca.

OBJECTIVES: An outbreak of NDM-1-producing Citrobacter freundii and possible secondary in vivo spread of blaNDM-1 to other Enterobacteriaceae were investigated. METHODS: From October 2012 to March 2015, meropenem-resistant Enterobacteriaceae were detected in 45 samples from seven patients at Aalborg University Hospital, Aalborg, Denmark. In silico resistance genes, Inc plasmid types and STs (MLST) were obtained from WGS data from 24 meropenem-resistant isolates (13 C. freundii, 6 Klebsiella pneumoniae, 4 Escherichia coli and 1 Klebsiella oxytoca) and 1 meropenem-susceptible K. oxytoca. The sequences of the meropenem-resistant C. freundii isolates were compared by phylogenetic analyses. In vitro susceptibility to 21 antimicrobial agents was tested. Furthermore, in vitro conjugation and plasmid characterization was performed. RESULTS: From the seven patients, 13 highly clonal ST18 NDM-1-producing C. freundii were isolated. The ST18 NDM-1-producing C. freundii isolates were only susceptible to tetracycline, tigecycline, colistin and fosfomycin (except for the C. freundii isolates from Patient 2 and Patient 7, which were additionally resistant to tetracycline). The E. coli and K. pneumoniae from different patients belonged to different STs, indicating in vivo transfer of blaNDM-1 in the individual patients. This was further supported by in vitro conjugation and detection of a 154 kb IncA/C2 plasmid with blaNDM-1. Patient screenings failed to reveal any additional cases. None of the patients had a history of recent travel abroad and the source of the blaNDM-1 plasmid was unknown. CONCLUSIONS: To our knowledge, this is the first report of an NDM-1-producing C. freundii outbreak and secondary in vivo spread of an IncA/C2 plasmid with blaNDM-1 to other Enterobacteriaceae.

Characterization of third-generation cephalosporin-resistant Escherichia coli from bloodstream infections in Denmark.

The aim of the study was to investigate the molecular epidemiology of 87 third-generation cephalosporin-resistant Escherichia coli (3GC-R Ec) from bloodstream infections in Denmark from 2009. Sixty-eight of the 87 isolates were extended-spectrum beta-lactamase (ESBL) producers, whereas 17 isolates featured AmpC mutations only (without a coexpressed ESBL enzyme) and 2 isolates were producing CMY-22. The majority (82%) of the ESBL-producing isolates in our study were CTX-M-15 producers and primarily belonged to phylogroup B2 (54.4%) or D (23.5%). Further, one of the two CMY-22-producing isolates belonged to B2, whereas only few of the other AmpCs isolates belonged to B2 and D. Pulsed-field gel electrophoresis revealed that both clonal and nonclonal spread of 3GC-R Ec occurred. ST131 was detected in 50% of ESBL-producing isolates. The remaining ESBL-producing isolates belonged to 17 other sequence types (STs), including several other internationally spreading STs (e.g., ST10, ST69, and ST405). The majority (93%) of the ESBL-producing isolates and one of the CMY-22-producing isolates were multiresistant. In conclusion, 3GC-R in bacteriaemic E. coli in Denmark was mostly due to ESBL production, overexpression of AmpC, and to a lesser extent to plasmid-mediated AmpC. The worldwide disseminated CTX-M-15-ST131 was strongly represented in this collection of Danish, bacteriaemic E. coli isolates.

The use of Gram stain and matrix-assisted laser desorption ionization time-of-flight mass spectrometry on positive blood culture: synergy between new and old technology.

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) is promising as an alternative to more costly and cumbersome methods for direct identifications in blood cultures. We wanted to evaluate a simplified pre-treatment method for using MALDI-TOF-MS directly on positive blood cultures using BacT/Alert blood culture system, and to test an algorithm combining the result of the initial microscopy with the result suggested by MALDI-TOF-MS. Using the recommended cut-off score of 1.7 the best results were obtained among Gram-negative rods with correct identifications in 91% of Enterobacteriaceae, 83% in aerobic/non-fermentative Gram-negative rods, whereas results were more modest among Gram-positive cocci with correct identifications in 52% of Staphylococci, 54% in Enterococci and only 20% in Streptococci. Combining the results of Gram stain with the top reports by MALDI-TOF-MS, increased the sensitivity from 91% to 93% in the score range from 1.5 to 1.7 and from 48% to 85% in the score range from 1.3 to 1.5. Thus, using this strategy and accepting a cut-off at 1.3 instead of the suggested 1.7, overall sensitivity could be increased from 88.1% to 96.3%. MALDI-TOF-MS is an efficient method for direct routine identification of bacterial isolates in blood culture, especially when combined with the result of the Gram stain.

Positive blood cultures and diagnosis of bacterial meningitis in cases with negative culture of cerebrospinal fluid.

The diagnosis of bacterial meningitis may be established through positive blood cultures in cases with negative culture of cerebrospinal fluid (CSF). However, information is sparse regarding how often the aetiological diagnosis is determined by blood culture alone. Therefore, we undertook a retrospective study of patients with meningitis and a blood culture-based bacterial diagnosis in North Jutland County, Denmark in 1997-2005. Patients were included if they had 1) a microscopy- and culture-negative CSF specimen with a leukocyte count > or =100x10(6)/l and an erythrocyte count < or =10,000x10(6)/l and 2) a positive blood culture within an interval of 3 d. A total of 20 patients fulfilled these criteria: 7 with Streptococcus pneumoniae, 4 with Neisseria meningitidis, 4 with Staphylococcus aureus, and 5 with miscellaneous bacteria. Suitable antibiotic therapy was delayed up to 48 h after lumbar puncture, and in-hospital mortality was 25%. A discharge diagnosis of bacterial meningitis was missing in 6/20 cases, and 12/20 cases had not been reported to Danish health authorities. Thus, patients with CSF-culture negative bacterial meningitis are at risk of delayed therapy, a poor outcome, and incomplete notification to health authorities.