WGS-based surveillance of third-generation cephalosporin-resistant Escherichia coli from bloodstream infections in Denmark.

Objectives: To evaluate a genome-based surveillance of all Danish third-generation cephalosporin-resistant Escherichia coli (3GC-R Ec ) from bloodstream infections between 2014 and 2015, focusing on horizontally transferable resistance mechanisms. Methods: A collection of 552 3GC-R Ec isolates were whole-genome sequenced and characterized by using the batch uploader from the Center for Genomic Epidemiology (CGE) and automatically analysed using the CGE tools according to resistance profile, MLST, serotype and fimH subtype. Additionally, the phylogenetic relationship of the isolates was analysed by SNP analysis. Results: The majority of the 552 isolates were ESBL producers (89%), with bla CTX-M-15 being the most prevalent (50%) gene, followed by bla CTX-M-14 (14%), bla CTX-M-27 (11%) and bla CTX-M-101 (5%). ST131 was detected in 50% of the E. coli isolates, with the remaining isolates belonging to 73 other STs, including globally disseminated STs (e.g. ST10, ST38, ST58, ST69 and ST410). Five of the bloodstream isolates were carbapenemase producers, carrying bla OXA-181 (3) and bla OXA-48 (2). Phylogenetic analysis revealed 15 possible national outbreaks during the 2 year period, one caused by a novel ST131/ bla CTX-M-101 clone, here observed for the first time in Denmark. Additionally, the analysis revealed three individual cases with possible persistence of closely related clones collected more than 13 months apart. Conclusions: Continuous WGS-based national surveillance of 3GC-R Ec , in combination with more detailed epidemiological information, can improve the ability to follow the population dynamics of 3GC-R Ec , thus allowing for the detection of potential outbreaks and the effects of changing treatment regimens in the future.

Recurrent Invasive Pneumococcal Disease Serotype 12F in a Vaccinated Splenectomized Patient.

This is the first case report of recurrent invasive pneumococcal disease (IPD), specifically, due to serotype 12F. The patient described here was vaccinated with the 23-valent pneumococcal polysaccharide vaccine (PPV23) due to previous splenectomy, and an anti-pneumococcal IgG test concluded that she had responded sufficiently to vaccination. Still, she had a fulminate recurrent infection with PPV23 serotype 12F. We investigated the anti-pneumococcal IgG test, and it turned out that it is based on the geometric mean value of only 12 of the serotypes included in PPV23; 12F is none of them. The reason is that there are no titer cut-offs available for 11 of the PPV23 serotypes, including 12F, neither nationally nor internationally. Yet, this is not specified in the answer to the clinicians. This case illustrates the need for titer cut-offs for the remaining pneumococcal serotypes in available vaccines, in order to get a more accurate estimation of the vaccination coverage for the individual patient. Therefore, more research on this area is warranted, along with a discussion of whether the laboratory answers to the clinicians should be more detailed.

[Pasteurella multocida septicaemia and pneumonia after chemotherapy in a patient who had no known contact with animals].

Pasteurella multocida inhabits the upper respiratory tract of many animals. It can cause skin and soft tissue infections in humans, usually in association with animal bites. We present a case of a 66-year-old chemotherapy-induced immunocompromised patient with lung cancer, who was treated for pneumonia and septicaemia due to P. multocida. There was no anamnestic contact with animals, which underlines the fact that immunocompromised patients can suffer from serious systemic infections due to P. multocida - even with no known animal contact.

Characterization of carbapenem nonsusceptible Pseudomonas aeruginosa in Denmark: a nationwide, prospective study.

From January 1st 2011 through June 30th 2011, 116 nonreplicate, noncystic fibrosis-related Pseudomonas aeruginosa isolates with reduced carbapenem susceptibility were collected from 12 out of 13 Danish departments of clinical microbiology. The presence of acquired ß-lactamases was assessed with combination tablet-diffusion methodology and polymerase chain reaction. In addition, antimicrobial susceptibility testing, an efflux pump inhibitor assay, and pulsed-field gel electrophoresis (PFGE) were performed. Isolates producing acquired ß-lactamases were further investigated by serotyping and multi locus sequence typing. Eight isolates produced the metallo-ß-lactamase (MBL) VIM-2, and one isolate produced OXA-10 and VEB-1-like extended-spectrum beta-lactamase (ESBL). Phenotypic indications of derepressed AmpC and efflux pump were seen in 56 and 43 isolates, respectively. Overall, the results indicate that mutational factors related to permeability--often combined with derepressed, chromosomal AmpC--is the main factor behind carbapenem nonsusceptibility in Danish P. aeruginosa isolates. The ESBL producer and all the VIM producers belonged to international clones. PFGE revealed that most of the isolates were unrelated, but clonal spread was seen; the 116 isolates distributed in 97 PFGE types, with the largest cluster consisting of 4 isolates (including three isolates from the same hospital with 100% similarity). Thirty-two isolates were pair-wise related, while the remaining isolates were clonally unrelated, as were all nine ESBL/MBL producers.

Extended-spectrum ß-lactamase (ESBL) in Danish clinical isolates of Escherichia coli and Klebsiella pneumoniae: prevalence, ß-lactamase distribution, phylogroups, and co-resistance.

BACKGROUND: Most Gram-negative community-acquired and nosocomial infections are caused by Escherichia coli and Klebsiella pneumoniae, among which increasing resistance due to extended-spectrum ß-lactamase (ESBL) is a major problem. We present data from the first Danish nationwide prevalence study on ESBL-producing E. coli, K. pneumoniae, and Proteus mirabilis in blood and urine cultures from hospitals and the community. METHODS: During September and October 2007, 13 of 15 Danish departments of clinical microbiology collected data and strains. Confirmatory ESBL test-positive isolates were sent to a central laboratory for species and ESBL-phenotype confirmation, extended susceptibility testing, phylogenetic grouping of E. coli strains, and ESBL gene characterization. RESULTS: During the study, blood samples from 18,259 patients and urine samples from 47,504 patients were subjected to culture. Among 14,674 cultured isolates, 352 were confirmed to be ESBL-producers. Thus, the crude ESBL prevalence was 2.4% (range 1.5% of E. coli in community urine to 6.6% of K. pneumoniae in hospital urine). An average of 7.2 ESBL-producers per 100,000 consumed bed-days was calculated. Of the 352 reported ESBL-producers, 205 E. coli, 73 K. pneumoniae, and 1 P. mirabilis, were available for testing. CTX-M enzymes dominated, both in hospitals and in the community, occurring in 92% of E. coli and 88% of K. pneumoniae, and with CTX-M-15 constituting 60% and 77%, respectively. CONCLUSIONS: Compared to 2003 data the ESBL prevalence in Denmark has increased significantly. In the ESBL-producers, reduced susceptibility towards both gentamicin and ciprofloxacin was seen among 43% E. coli and 55% K. pneumoniae, leaving clinicians in these cases with only a carbapenem for the treatment of serious infections. Part of this study was presented at the 20(th) European Congress of Clinical Microbiology and Infectious Diseases, abstract P-1617.

[Possible daptomycin-induced organizing pneumonia]. / Mulig daptomycininduceret pneumoni.

We report a possible and rare case of drug-induced organizing pneumonia in a patient receiving daptomycin. A 75-year-old man with acute infective aorta valve endocarditis due to Enterococcus faecalis developed organizing pneumonia following treatment with intravenous daptomycin and ceftriaxone. The patient could have acquired an infectious agent which was resistant to ceftriaxone, such as Legionella, and he was thus treated with ciprofloxacin and rifampicin. However, no antigen was found in the urine, and the patient's symptoms subsided after daptomycin was discontinued.

Vancomycin-resistant Enterococcus spp.: validation of susceptibility testing and in vitro activity of vancomycin, linezolid, tigecycline and daptomycin.

Vancomycin-resistant enterococci (VRE) have emerged to become a significant nosocomial pathogen. However, detection may be challenging and treatment possibilities are limited. Reports of resistance to linezolide, daptomycin and tigecycline underline the need for reliable susceptibility testing with respect to these compounds. We evaluated the in vitro activity of vancomycin, linezolid, daptomycin and tigecycline against a panel of VRE and vancomycin-susceptible enterococci by broth microdilution (BMD). Etest for determination of minimum inhibitory concentration of these four antibiotics and two disc diffusion assays for detecting VRE and for susceptibility testing against tigecycline and linezolid were evaluated. Before susceptibility testing, all isolates were classified by polymerase chain reaction as vanA or vanB gene positive or vanA/B gene negative. Linezolid, daptomycin and tigecycline had excellent in vitro activity towards all isolates. For daptomycin and tigecycline, the overall agreement between BMD and Etest was suboptimal. For both disc diffusion assays, use of current break points was inadequate to detect vancomycin resistance for isolates carrying the vanB gene. Inspection of the inhibition zone for a diffuse edge, as recommended, accurately predicted presence of the vanB gene.

Community outbreak of perianal group A streptococcal infection in Denmark.

BACKGROUND: Perianal group A streptococcal infection (PASI) occurs primarily in children. There is limited information on the incidence, transmission and treatment of PASI. We report a cluster of cases connected to a Danish kindergarten and observations of the incidence of PASI in the local population. SETTING: A Danish rural community with 1765 children 15 years and younger registered with two general practice clinics. METHODS: After being alerted of a possible cluster of PASI cases, all isolates of group A beta-hemolytic streptococci were collected and subjected to T typing and pulsed field gel electrophoresis (PFGE) if grown from either a rectal swab or an accompanying throat swab obtained in the offices of local general practitioners during the ensuing 4-month period. Clinical data were obtained from the files of the local general practitioners. RESULTS: Twelve cases of PASI were caused by group A beta-hemolytic streptococci T type 28 with an identical PFGE profile: 6 of the cases were in children attending the same kindergarten, 4 were connected otherwise to the cluster and 2 cases seemed to be unrelated. Five cases of PASI with different T types and PFGE profiles were diagnosed during the same period giving an estimated annual incidence of 2 to 7 per 1000 children. Penicillin V was ineffective in 3 cases, and no recurrence was seen after change of the treatment to oral clarithromycin. CONCLUSIONS: A clone of T type 28 seemed to be the cause of the largest cluster of PASI cases described thus far. Clarithromycin was effective as second line treatment. An estimated annual baseline incidence of 2 to 7 per 1000 in the local population indicates that PASI may not be as rare as previously estimated.

Significance of low-level resistance to ciprofloxacin in Klebsiella pneumoniae and the effect of increased dosage of ciprofloxacin in vivo using the rat granuloma pouch model.

This study was designed to compare the killing effect of ciprofloxacin on strains of Klebsiella pneumoniae with different MICs of ciprofloxacin in vivo using the rat granuloma pouch infection model. Five different strains were used: one ciprofloxacin-susceptible strain (MIC 0.06 mg/L); one strain highly resistant to ciprofloxacin (MIC 8 mg/L); and three nalidixic acid-resistant strains with low-level resistance to ciprofloxacin (MIC 0.25-0.5 mg/L). The efficacy of ciprofloxacin was evaluated 3 h after bacterial challenge (treating an acute infection) or after 3 days (treating a late infection) with a single intraperitoneal injection of ciprofloxacin (40 and 200 mg/kg). Ciprofloxacin was bactericidal against both growing K. pneumoniae (acute infection model) and non-growing K. pneumoniae (late infection model), but the extent of killing was significantly higher on growing bacteria and against ciprofloxacin-susceptible K. pneumoniae. A peak concentration of ciprofloxacin, at the infection site, <3 x MIC was not sufficient for optimal bacterial elimination. However, it was possible to compensate for the lower killing in low-level ciprofloxacin-resistant K. pneumoniae by increasing the dosage of ciprofloxacin from 40 to 200 mg/kg, consistent with the higher MIC.