Antimicrobial resistance surveillance of Bacteroides fragilis isolated from blood cultures, Europe, 2022 (ReSuBacfrag).

BACKGROUND: Bacteroides fragilis is the most frequent cause of anaerobic bacteraemia. Although recent data suggest a rise in antimicrobial resistance (AMR) of this and other anaerobic bacteria, surveillance remains limited due to a lack of both data availability and comparability. However, a newly introduced standardised method for antimicrobial susceptibility testing (AST) of anaerobic bacteria has made larger scale surveillance possible for the first time. AIM: To investigate phenotypic AMR of Bacteroides fragilis isolates from bacteraemia across Europe in 2022. METHODS: In a multicentre approach, clinical microbiology laboratories in Europe were invited to contribute results of AST for Bacteroides fragilis blood culture isolates (including only the first isolate per patient and year). AST of a selection of four antibiotics was performed locally by participating laboratories in a prospective or retrospective manner, using the new EUCAST disc diffusion method on fastidious anaerobe agar (FAA-HB). RESULTS: A total of 16 European countries reported antimicrobial susceptibilities in 449 unique isolates of Bacteroides fragilis from blood cultures in 2022. Clindamycin demonstrated the highest resistance rates (20.9%, range 0 - 63.6%), followed by piperacillin-tazobactam (11.1%, 0 - 54.5%), meropenem (13.4%, 0 - 45.5%), and metronidazole (1.8%, 0 - 20.0%), all with wide variation between countries. CONCLUSION: Considering that the mean resistance rates across Europe were higher than expected for three of the four anti-anaerobic antibiotics under surveillance, both local AST of clinically relevant isolates of Bacteroides fragilis and continued surveillance on an international level is warranted.

Humoral antibody response following mRNA vaccines against SARS-CoV-2 in solid organ transplant recipients; a status after a fifth and bivalent vaccine dose.

Background: In solid organ transplant (SOT) recipients, the humoral response following COVID-19 vaccination is reduced, as a result of their immunosuppressed treatment. In this study, we investigated antibody concentrations after booster vaccinations until the fifth dose, the latter by monovalent or bivalent BA1 or BA4/5 vaccines. In addition, we evaluated the efficacy of vaccination by recording breakthrough infections, hospitalizations, and deaths. Method: This prospective cohort study included 438 SOT recipients (>18 years) vaccinated with mRNA vaccines against COVID-19 from January 2021 until March 2023. Blood samples were drawn before and after each vaccination and tested for SARS-CoV-2 spike RBD IgG antibodies with the lowest and highest cut-off at 7.1 and 5,680 BAU/mL, respectively. Vaccine information, breakthrough infections, and hospitalizations were collected from the medical records. Results: Most participants received BNT162b2 and 61.4% received five vaccine doses. The response proportion in SOT recipients increased from 86.7% after the fourth dose to 93.0% following the fifth dose. Antibody concentration decreased with 142.7 BAU/mL between the third and fourth dose (median 132 days, Quartile 1: 123, Quartile 3: 148) and 234.3 BAU/mL between the fourth and fifth (median 250 days, Quartile 1: 241, Quartile 3: 262) dose among those without breakthrough infection (p=0.34). When comparing the Omicron BA.1 or Omicron BA.4/BA.5 adapted vaccines, no significant differences in antibody concentration were found, but 20.0% of SOT recipients receiving a monovalent fifth vaccine dose had a breakthrough infection compared to 4.0% and 7.9% among those who received BA.1 and BA.4/BA.5 adapted vaccines, respectively (p=0.04). Since January 2021, 240 (54.8%) participants had a breakthrough infection, and 22 were hospitalized, but no deaths were observed. Conclusions: The fifth COVID-19 vaccine dose raised antibody response to 93.0% of the study population. Additional booster doses, as well as bivalent vaccines, led to higher levels of antibody concentration in SOT recipients. We found a lower incidence of breakthrough infections among SOT recipients after receiving a bivalent vaccine as a fifth dose compared to those receiving a monovalent dose. Antibody concentrations did not wane when the time between doses was prolonged from four to eight months.

Performance of automated antimicrobial susceptibility testing for the detection of antimicrobial resistance in gram-negative bacteria: a NordicAST study.

Automated testing of antimicrobial susceptibility is common in clinical microbiology laboratories but their ability to detect low-level resistance has been questioned. This Nordic multicentre study aimed to evaluate the performance of commercially available automated AST systems. A phenotypically well-characterised collection of gram-negative bacilli (Escherichia coli (n = 7), Klebsiella pneumoniae (n = 6) and Pseudomonas aeruginosa (n = 7)) with and without resistance mechanisms was examined by Danish (n = 1), Finnish (n = 6), Norwegian (n = 16) and Swedish (n = 5) laboratories. Minimum inhibitory concentrations (MICs) were determined for 12 antimicrobials with automated systems and compared with MICs obtained with gold standard broth microdilution. The automated systems used were VITEK 2 (n = 23), Phoenix (n = 4), MicroScan (n = 1), and ARIS (n = 1). Very major errors were identified for six antimicrobials; cefotaxime (6.9%), meropenem (0.4%), ciprofloxacin (0.7%), ertapenem (4.3%), amikacin (3.4%) and colistin (6.4%). Categorical agreement of MIC for the automated systems compared to broth microdilution ranged from 83% for imipenem to 100% for ampicillin and trimethoprim-sulfamethoxazole. The analysis revealed several important antimicrobials where resistance was underestimated, potentially with significant consequences in patient treatment. The results cast doubt on the use of automated AST in the management of patients with serious infections and suggests that more work is needed to define their limitations.

Antimicrobial susceptibility testing of Bacteroides species by disk diffusion: The NordicAST Bacteroides study.

OBJECTIVES: Antimicrobial susceptibility testing (AST) of anaerobic bacteria has until recently been done by MIC methods. We have carried out a multi-centre evaluation of the newly validated EUCAST disk diffusion method for AST of Bacteroides spp. METHODS: A panel of 30 Bacteroides strains was assembled based on reference agar dilution MICs, resistance gene detection and quantification of cfiA carbapenemase gene expression. Nordic clinical microbiology laboratories (n = 45) performed disk diffusion on Fastidious Anaerobe Agar with 5% mechanically defibrinated horse blood (FAA-HB) for piperacillin-tazobactam, meropenem and metronidazole. RESULTS: A total of 43/45 (95.6%) laboratories carried out disk diffusion per protocol. Intraclass correlation coefficients were 0.87 (0.80-0.93) for piperacillin-tazobactam, 0.95 (0.91-0.97) for meropenem and 0.89 (0.83-0.94) for metronidazole. For metronidazole, one media lot yielded smaller zones and higher variability than another. Piperacillin-tazobactam and meropenem zone diameters correlated negatively with cfiA expression. A meropenem zone diameter of <28 mm in B. fragilis indicated presence of cfiA. Piperacillin-tazobactam had the most false susceptible results. Categorical errors for this antimicrobial were particularly prevalent in cfiA-positive strains, and piperacillin-tazobactam had the highest number of comments describing zone reading difficulties. CONCLUSIONS: Inter-laboratory agreement by disk diffusion was good or very good. The main challenges were media-related variability for metronidazole and categorical disagreement with the reference method for piperacillin-tazobactam in some cfiA-positive strains. An area of technical uncertainty specific for such strains may be warranted.

Characterisation of carbapenemase-producing Acinetobacter baumannii isolates from danish patients 2014-2021: detection of a new international clone - IC11.

OBJECTIVES: This study aimed to characterise carbapenemase-producing Acinetobacter baumannii (A. baumannii) isolates from Danish patients using whole genome sequencing (WGS). It also compared typing and epidemiological data for further investigation of the spread and origin of the carbapenemase-producing A. baumannii isolates. METHODS: From 1 January 2014 to 30 September 2021, 141 carbapenemase-producing A. baumannii isolates, received at the national reference laboratory at Statens Serum Institut, were investigated using WGS. Multilocus sequence typing (MLST) and cgMLST data, obtained by SeqSphere+ software, were linked to data related to source of isolation, patient age and sex, hospital admission and travel history. RESULTS: Most of the carbapenemase-producing A. baumannii isolates were from males (n = 100, 71%). Most patients (n = 88, 63%) had travelled outside Scandinavia before admission to a Danish hospital. The most prevalent carbapenemase gene was blaOXA-23 (n = 124). Isolates belonging to the dominating international clone IC2 accounted for 78% of all isolates. A new international ST164/OXA-91 clone, proposed to be named IC11, was recognised and described. The cgMLST analysis revealed 17 clusters, reflecting both sporadic travel to similar geographical areas and confirmed outbreaks in Danish hospitals. CONCLUSIONS: The occurrence of carbapenemase-producing A. baumannii in Denmark was still low; however, isolates belonging to major international clones with a high potential to spread within hospitals, mainly IC2, dominated. OXA-23 was by far the most prevalent carbapenemase detected. Sporadic and travel-related introductions to Danish hospitals, also intra-hospital transmission, could be confirmed, emphasising the need for continuing vigilance.

Development and Clinical Application of a Multilocus Sequence Typing Scheme for Bacteroides fragilis Based on Whole-Genome Sequencing Data.

Bacteroides fragilis is among the most abundant and pathogenic bacterial species in the gut microbiota and is associated with diarrheal disease in children, inflammatory bowel disease, and the development of colorectal cancer. It is increasingly resistant to potent antimicrobial agents such as carbapenems and metronidazole, making it among the most resistant anaerobic bacteria. These factors combined call for increased monitoring of B. fragilis and its population structure on a worldwide scale. Here, we present a possible solution through the development of a multilocus sequence typing scheme (MLST). The scheme is based on seven core gene fragments: groL (hsp60), rpoB, recA, dnaJ, rprX, prfA, and fusA. These gene fragments possess high discriminatory power while retaining concordance with whole core genome-based phylogenetic analysis. The scheme proved capable of differentiating B. fragilis isolates at the strain level. It offers a standardized method for molecular typing and can be applied to isolates from various sampling backgrounds, such as patient isolates, environmental samples, and strains obtained from food and animal sources. In total, 567 B. fragilis genomes were sequence typed and their isolate data collected. The MLST scheme clearly divided the B. fragilis population into two divisions based on the presence of the cfiA and cepA resistance genes. However, no other specific subpopulations within the analyzed genomes were found to be associated with any specific diseases or geographical location. With this MLST scheme, we hope to provide a powerful tool for the study and monitoring of B. fragilis on an international scale. IMPORTANCE Here, we present the first MLST scheme for Bacteroides fragilis, one of the most abundant pathogenic bacteria in the human gut microbiota. The scheme enables standard classification and monitoring of B. fragilis on a worldwide scale and groups the majority of current isolate data in one place. A more unified approach to the collection and analysis of B. fragilis data could provide crucial insights into how the pathogen operates and develops as a species. Close monitoring of B. fragilis is especially relevant, as it is increasingly resistant to potent antimicrobial agents and engages in horizontal gene transfer with other bacteria. Hopefully, this approach will guide new discoveries into how B. fragilis evolves and interacts with its human host. Additionally, the scheme could potentially be applied to other species of the genus Bacteroides.

Invasive procedures and risk of brain abscess: a nationwide, population-based case-control study.

OBJECTIVES: It is unknown whether invasive procedures are associated with brain abscess. METHODS: Nationwide, population-based, matched case-control study of patients with culture verified brain abscess in Denmark from 1989 to 2016. Exposure was invasive procedures 0-6 months before study inclusion. RESULTS: We identified 435 patients and 3909 controls. The level of comorbidity was higher among patients with brain abscess than among controls. A total of 48 cases (11%) had one or more invasive procedures 0-6 months before study inclusion (adjusted odds ratios (aOR) of 3.6 (95% confidence interval (CI): 2.5-5.1), a population attributable fractions of 8% (95% CI: 7-9)). In primary care, ear, nose and throat (ENT) procedures were associated with brain abscess (aOR of 4.0 (95% CI: 2.0-8.0)), but gastrointestinal endoscopies were not (aOR of 1.0 (95% CI: 0.3-3.2)). No bronchoscopies were performed in primary care. In the hospital-based setting, ENT procedures, bronchoscopies and gastrointestinal endoscopies were associated with an increased risk of brain abscess (aOR of 14.5 (95% CI: 4.8-43.8), 20.3 (95% CI: 3.8-110.1) and 3.4 (95% CI: 2.0-5.6), respectively). CONCLUSIONS: The association between invasive procedures and brain abscess was more pronounced in the hospital-based setting than in primary care.

Humoral immune response following a third SARS-CoV-2 mRNA vaccine dose in solid organ transplant recipients compared with matched controls.

Background: Solid organ transplant (SOT) recipients have shown suboptimal antibody response following COVID-19 vaccination. Several risk factors for the diminished response have been identified including immunosuppression and older age, but the influence of different comorbidities is not fully elucidated. Method: This case-control study consisted of 420 Danish adult SOT recipients and 840 sex- and age-matched controls, all vaccinated with a third homologous dose of either BNT162b2 (Pfizer-BioNTech) or mRNA-1273 (Moderna) vaccine. The primary outcome was differences in humoral immune response. The secondary outcome was breakthrough infections. Additionally, we looked for factors that could predict possible differences between the two groups. Results: Response rate increased from 186/382 (49%) to 275/358 (77%) in SOT recipients and remained on 781/790 (99%) to 601/609 (99%) in controls following a third vaccine dose. SOT recipients had significantly lower median antibody concentrations after third dose compared to controls (332.6 BAU/ml vs 46,470.0 BAU/ml, p <0.001). Lowest median antibody concentrations were seen in SOT recipients with liver disease (10.3 BAU/ml, IQR 7.1-319) and diabetes (275.3 BAU/ml, IQR 7.3-957.4). Breakthrough infections occurred similarly frequent, 150 (40%) among cases and 301 (39%) among controls (p = 0.80). Conclusion: A third COVID-19 vaccine dose resulted in a significant increase in humoral immunogenicity in SOT recipients and maintained high response rate in controls. Furthermore, SOT recipients were less likely to produce antibodies with overall lower antibody concentrations and humoral immunity was highly influenced by the presence of liver disease and diabetes. The prevalence of breakthrough infections was similar in the two groups.

Omicron BA.5 Neutralization among Vaccine-Boosted Persons with Prior Omicron BA.1/BA.2 Infections.

Worldwide, millions of persons have received multiple COVID-19 vaccinations and subsequently recovered from SARS-CoV-2 Omicron breakthrough infections. In 2 small, matched cohorts (n = 12, n = 24) in Denmark, we found Omicron BA.1/BA.2 breakthrough infection after 3-dose BNT162b2 vaccination provided improved Omicron BA.5 neutralization over 3-dose vaccination alone.

Machine learning-based typing of Salmonella enterica O-serogroups by the Fourier-Transform Infrared (FTIR) Spectroscopy-based IR Biotyper system.

BACKGROUND: Salmonella enterica is among the major burdens for public health at global level. Typing of salmonellae below the species level is fundamental for different purposes, but traditional methods are expensive, technically demanding, and time-consuming, and therefore limited to reference centers. Fourier transform infrared (FTIR) spectroscopy is an alternative method for bacterial typing, successfully applied for classification at different infra-species levels. AIM: This study aimed to address the challenge of subtyping Salmonella enterica at O-serogroup level by using FTIR spectroscopy. We applied machine learning to develop a novel approach for S. enterica typing, using the FTIR-based IR Biotyper® system (IRBT; Bruker Daltonics GmbH & Co. KG, Germany). We investigated a multicentric collection of isolates, and we compared the novel approach with classical serotyping-based and molecular methods. METHODS: A total of 958 well characterized Salmonella isolates (25 serogroups, 138 serovars), collected in 11 different centers (in Europe and Japan), from clinical, environmental and food samples were included in this study and analyzed by IRBT. Infrared absorption spectra were acquired from water-ethanol bacterial suspensions, from culture isolates grown on seven different agar media. In the first part of the study, the discriminatory potential of the IRBT system was evaluated by comparison with reference typing method/s. In the second part of the study, the artificial intelligence capabilities of the IRBT software were applied to develop a classifier for Salmonella isolates at serogroup level. Different machine learning algorithms were investigated (artificial neural networks and support vector machine). A subset of 88 pre-characterized isolates (corresponding to 25 serogroups and 53 serovars) were included in the training set. The remaining 870 samples were used as validation set. The classifiers were evaluated in terms of accuracy, error rate and failed classification rate. RESULTS: The classifier that provided the highest accuracy in the cross-validation was selected to be tested with four external testing sets. Considering all the testing sites, accuracy ranged from 97.0% to 99.2% for non-selective media, and from 94.7% to 96.4% for selective media. CONCLUSIONS: The IRBT system proved to be a very promising, user-friendly, and cost-effective tool for Salmonella typing at serogroup level. The application of machine learning algorithms proved to enable a novel approach for typing, which relies on automated analysis and result interpretation, and it is therefore free of potential human biases. The system demonstrated a high robustness and adaptability to routine workflows, without the need of highly trained personnel, and proving to be suitable to be applied with isolates grown on different agar media, both selective and unselective. Further tests with currently circulating clinical, food and environmental isolates would be necessary before implementing it as a potentially stand-alone standard method for routine use.

Bacteremia With Anaerobic Bacteria and Association With Colorectal Cancer: A Population-based Cohort Study.

BACKGROUND: There is a well-described association between bacteremia with bovis group streptococci or Clostridium septicum and an increased probability of a colorectal cancer (CRC) diagnosis. We wanted to investigate the existence of a similar association between CRC and bacteremia with other bacteria belonging to the gut microbiota.. METHODS: A population based cohort study in a population about 2 million people including 45 774 bacteremia episodes and 231 387 blood culture negative cases was performed in the Region of Southern Denmark and Region Zealand from 2007-2016. Episodes of bacteremia were combined with the Danish central register for CRC. We performed Cox's regression analysis with hazard ratios (HRs) and 95% confidence intervals (CIs). RESULTS: The study results confirmed previous findings of an increased risk of a CRC diagnosis after bacteremia with the bovis group streptococci (risk within a year: 4.3%; HR [95% CI]: 8.46 [3.51-20.4]) or C. septicum (20.8%; 76.2 [42.0-138]). Furthermore, Bacteroides ovatus (6.7%; 20.3 [5.04-81.8]), Bacteroides uniformis (5.4%; 16.2 [4.02-65.7]), Clostridium tertium (3.6 %; 13.9 [1.96-99.4]), Fusobacterium spp. (excluding F. necrophorum) (3.0 %; 8.51 [2.73-26.5]), and Gram-positive anaerobic cocci (3.6 %; 10.9 [4.50-26.3]) were also associated with an increased risk of a CRC diagnosis compared to patients with negative blood cultures (0.4%). CONCLUSIONS: Bacteremia with specific gut microbiota anaerobic bacteria is associated with a high risk of a diagnosis of CRC, indicating the need for colorectal workup. Importantly, this strategy also holds the possible additional benefit of detecting adenomas or other premalignant conditions, which were not included in the present study.

Serum Neutralization of SARS-CoV-2 Omicron BA.1 and BA.2 after BNT162b2 Booster Vaccination.

The SARS-CoV-2 Omicron variant BA.2 sublineage is rapidly replacing earlier Omicron lineages, suggesting BA.2 has increased vaccine evasion properties. We measured neutralization titers of authentic BA.1 and BA.2 isolates in serum samples from persons who received the BNT162b2 booster vaccine. All samples neutralized BA.1 and BA.2 at equal median values.

Dentist's Visits and Risk of Brain Abscess: A Nationwide, Population-Based Case-Control Study.

BACKGROUND: Brain abscesses are frequently caused by oral cavity bacteria, but whether dental status and invasive dental procedures are important risk factors is unknown. METHODS: A nationwide, population-based, case-control study examined the association between dentist's visits and invasive dental procedures and risk of brain abscess caused by oral cavity bacteria from 1989 through 2016. Date of brain abscess diagnosis was considered the index date. Using risk-set sampling, 10 population controls per case were individually matched by age, sex, and residential area. Conditional logistic regression was used to compute odds ratios with 95% confidence intervals (CIs), adjusted for comorbidity. RESULTS: We identified 362 patients with culture-proven brain abscess caused by oral cavity bacteria. The median age was 53 years (interquartile range, 39-65 years) and 220 (61%) were male. Invasive dental procedures within 6 months before the index date was observed in 21 of 362 (6%) patients with brain abscess and 179 of 3257 (5%) population controls (adjusted odds ratio [aOR], 1.07 [95% CI, .67-1.70]). Two hundred thirteen of 362 (59%) patients with brain abscess had visited their dentist within 1 year before the index date compared with 1944 of 3257 (60%) of population controls (aOR, 0.99 [95% CI, .77-1.26]). Using no dentist's visits as reference, we observed aORs of 0.95 (95% CI, .64-1.40) for 1-2 visits within 3 years of the index date and 1.01 (95% CI, .76-1.35) for 3 or more visits. CONCLUSIONS: Recent invasive dental procedures and number of dentist's visits were not associated with culture-verified brain abscess caused by oral cavity bacteria.