In vitro activity of meropenem-vaborbactam plus aztreonam against metallo-ß-lactamase-producing Klebsiella pneumoniae.

We evaluated the in vitro activity of meropenem-vaborbactam plus aztreonam (MEV-ATM) against 140 metallo-ß-lactamase (MBL)-producing Klebsiella pneumoniae isolates. Among them, 25 isolates (17.9%) displayed minimum inhibitory concentrations (MIC) ≥ 8 µg/mL, while 112 (80.0%) had MIC ≤ 2 µg/mL. Genomic analysis and subsequent gene cloning experiments revealed OmpK36 134-135GD-insertion and increased carbapenemase gene (blaNDM-1 and blaOXA-48-like) copy numbers are the main factors responsible for MEV-ATM non-susceptibility. Notably, MEV-ATM is actively against aztreonam-avibactam-resistant mutants due to CMY-16 mutations.

Within-Host Genotypic and Phenotypic Diversity of Contemporaneous Carbapenem-Resistant Klebsiella pneumoniae from Blood Cultures of Patients with Bacteremia.

It is unknown whether bacterial bloodstream infections (BSIs) are commonly caused by single organisms or mixed microbial populations. We hypothesized that contemporaneous carbapenem-resistant Klebsiella pneumoniae (CRKP) strains from blood cultures of individual patients are genetically and phenotypically distinct. We determined short-read whole-genome sequences of 10 sequence type 258 (ST258) CRKP strains from blood cultures in each of 6 patients (Illumina HiSeq). Strains clustered by patient by core genome and pan-genome phylogeny. In 5 patients, there was within-host strain diversity by gene mutations, presence/absence of antibiotic resistance or virulence genes, and/or plasmid content. Accessory gene phylogeny revealed strain diversity in all 6 patients. Strains from 3 patients underwent long-read sequencing for genome completion (Oxford Nanopore) and phenotypic testing. Genetically distinct strains within individuals exhibited significant differences in carbapenem and other antibiotic responses, capsular polysaccharide (CPS) production, mucoviscosity, and/or serum killing. In 2 patients, strains differed significantly in virulence during mouse BSIs. Genetic or phenotypic diversity was not observed among strains recovered from blood culture bottles seeded with index strains from the 3 patients and incubated in vitro at 37°C. In conclusion, we identified genotypic and phenotypic variant ST258 CRKP strains from blood cultures of individual patients with BSIs, which were not detected by the clinical laboratory or in seeded blood cultures. The data suggest a new paradigm of CRKP population diversity during BSIs, at least in some patients. If validated for BSIs caused by other bacteria, within-host microbial diversity may have implications for medical, microbiology, and infection prevention practices and for understanding antibiotic resistance and pathogenesis. IMPORTANCE The long-standing paradigm for pathogenesis of bacteremia is that, in most cases, a single organism passes through a bottleneck and establishes itself in the bloodstream (single-organism hypothesis). In keeping with this paradigm, standard practice in processing positive microbiologic cultures is to test single bacterial strains from morphologically distinct colonies. This study is the first genome-wide analysis of within-host diversity of Klebsiella pneumoniae strains recovered from individual patients with bloodstream infections (BSIs). Our finding that positive blood cultures comprised genetically and phenotypically heterogeneous carbapenem-resistant K. pneumoniae strains challenges the single-organism hypothesis and suggests that at least some BSIs are caused by mixed bacterial populations that are unrecognized by the clinical laboratory. The data support a model of pathogenesis in which pressures in vivo select for strain variants with particular antibiotic resistance or virulence attributes and raise questions about laboratory protocols and treatment decisions directed against single strains.

Multidrug-Resistant Gram-Negative Bacteria in Burn Patients.

Patients with burn injuries are at high risk for infectious complications, and infections are the most common cause of death after the first 72 h of hospitalization. Hospital-acquired infections caused by multidrug resistant (MDR) Gram-negative bacteria (GNB) in this population are concerning. Here, we evaluated carriage with MDR GNB in patients in a large tertiary-care burn intensive care unit. Twenty-nine patients in the burn unit were screened for intestinal carriage. Samples were cultured on selective media. Median time from admission to the burn unit to first sample collection was 9 days (IQR 5 - 17 days). In 21 (72%) patients, MDR GNB were recovered; the most common bacterial species isolated was Pseudomonas aeruginosa, which was found in 11/29 (38%) of patients. Two of these patients later developed bloodstream infections with P. aeruginosa. Transmission of KPC-31-producing ST22 Citrobacter freundii was detected. Samples from two patients grew genetically similar C. freundii isolates that were resistant to ceftazidime-avibactam. On analysis of whole-genome sequencing, blaKPC-31 was part of a Tn4401b transposon that was present on two different plasmids in each C. freundii isolate. Plasmid curing experiments showed that removal of both copies of blaKPC-31 was required to restore susceptibility to ceftazidime-avibactam. In summary, MDR GNB colonization is common in burn patients and patient-to-patient transmission of highly resistant GNB occurs. These results emphasize the ongoing need for infection prevention and antimicrobial stewardship efforts in this highly vulnerable population.

Ceftazidime-avibactam based combinations against carbapenemase producing Klebsiella pneumoniae harboring hypervirulence plasmids.

The combination of carbapenem resistance and hypervirulence in Klebsiella pneumoniae is an emerging and urgent threat due to its potential to resist common antibiotics and cause life-threatening infections in healthy hosts. This study aimed to evaluate the activity of clinically relevant antibiotic regimens against carbapenem-resistant K. pneumoniae with hypervirulence plasmids and to identify pathways associated with antibiotic tolerance using transcriptomics. We studied two carbapenem-resistant K. pneumoniae isolates, CDI694 and CDI231, both harboring hypervirulence plasmids. Time-kill and dynamic one-compartment pharmacokinetic/pharmacodynamic assays were used to assess ceftazidime/avibactam-based therapies. RNAseq was performed following 48 h of antibiotic exposure. Closed genomes of CDI694 and CDI231 were obtained; each isolate harbored carbapenem-resistance and hypervirulence (containing rmpA/rmpA2 and iut genes) plasmids. Ceftazidime/avibactam-based regimens were bactericidal, though both isolates continued to grow in the presence of antibiotics despite no shifts in MIC. Transcriptomic analyses suggested that perturbations to cell respiration, carbohydrate transport, and stress-response pathways contributed to the antibiotic tolerance in CDI231. Genes associated with hypervirulence and antibiotic resistance were not strongly impacted by drug exposure except for ompW, which was significantly downregulated. Treatment of carbapenem-resistant K. pneumoniae harboring hypervirulence plasmids with ceftazidime/avibactam-based regimens may yield a tolerant population due to altered transcription of multiple key pathways.

In vitro antibacterial activity of cefiderocol against recent multidrug-resistant carbapenem-nonsusceptible Enterobacterales isolates.

Cefiderocol, a siderophore-containing cephalosporin with broad-spectrum antimicrobial activity against many ß-lactam-resistant Gram-negative bacteria, was tested by broth microdilution against 104 carbapenem-non-susceptible Enterobacterales clinical isolates from 2011 to 2018. Carbapenemase identification was determined using PCR followed by targeted gene sequencing or whole genome sequencing (WGS). All isolates were multidrug-resistant, 89.4% (93/104) and produced a serine (KPC or SME) carbapenemase, with as many as four ß-lactamases present. A VIM-1 or NDM-1 metallo-ß-lactamase was confirmed in 6.7% of the isolates (N = 5 and 2, respectively). All isolates were susceptible to cefiderocol, unlike the comparator agents. Susceptibility for comparators ranged from 24.0% for meropenem to 91.3%, 92.3% and 96.1% for imipenem-relebactam, ceftazidime-avibactam and meropenem-vaborbactam, respectively; 48.1%, 75.2% and 79.8% of the isolates were susceptible to omadacycline, colistin and eravacycline, respectively. Two isolates with cefiderocol MICs of 2 mg/L had mutations or deletions of the iron transport genes fhuA/E or fepA, as determined by WGS.

Hyaluronan enhances bone marrow cell therapy for myocardial repair after infarction.

Hyaluronan (HA) has been shown to play an important role during early heart development and promote angiogenesis under various physiological and pathological conditions. In recent years, stem cell therapy, which may reduce cardiomyocyte apoptosis, increase neovascularization, and prevent cardiac fibrosis, has emerged as a promising approach to treat myocardial infarction (MI). However, effective delivery of stem cells for cardiac therapy remains a major challenge. In this study, we tested whether transplanting a combination of HA and allogeneic bone marrow mononuclear cells (MNCs) promotes cell therapy efficacy and thus improves cardiac performance after MI in rats. We showed that HA provided a favorable microenvironment for cell adhesion, proliferation, and vascular differentiation in MNC culture. Following MI in rats, compared with the injection of HA alone or MNC alone, injection of both HA and MNCs significantly reduced inflammatory cell infiltration, cardiomyocyte apoptosis, and infarct size and also improved cell retention, angiogenesis, and arteriogenesis, and thus the overall cardiac performance. Ultimately, HA/MNC treatment improved vasculature engraftment of transplanted cells in the infarcted region. Together, our results indicate that combining the biocompatible material HA with bone marrow stem cells exerts a therapeutic effect on heart repair and may further provide potential treatment for ischemic diseases.

Intramyocardial peptide nanofiber injection improves postinfarction ventricular remodeling and efficacy of bone marrow cell therapy in pigs.

BACKGROUND: Growing evidence suggests that intramyocardial biomaterial injection improves cardiac functions after myocardial infarction (MI) in rodents. Cell therapy is another promising approach to treat MI, although poor retention of transplanted cells is a major challenge. In this study, we hypothesized that intramyocardial injection of self-assembling peptide nanofibers (NFs) thickens the infarcted myocardium and increases transplanted autologous bone marrow mononuclear cell (MNC) retention to attenuate cardiac remodeling and dysfunction in a pig MI model. METHODS AND RESULTS: A total of 40 mature minipigs were divided into 5 groups: sham, MI+normal saline, MI+NFs, MI+MNCs, and MI+MNCs/NFs. MI was induced by coronary occlusion followed by intramyocardial injection of 2 mL normal saline or 1% NFs with or without 1×10(8) isolated autologous MNCs. NF injection significantly improved diastolic function and reduced ventricular remodeling 28 days after treatment. Injection of MNCs alone ameliorated systolic function only, whereas injection of MNCs with NFs significantly improved both systolic and diastolic functions as indicated by +dP/dt and -dP/dt (1214.5±91.9 and -1109.7±91.2 mm Hg/s in MI+NS, 1693.7±84.7 and -1809.6±264.3 mm Hg/s in MI+MNCs/NFs, respectively), increased transplanted cell retention (29.3±4.5 cells/mm(2) in MI+MNCs and 229.4±41.4 cells/mm(2) in MI+MNCs/NFs) and promoted capillary density in the peri-infarct area. CONCLUSIONS: We demonstrated that NF injection alone prevents ventricular remodeling, whereas cell implantation with NFs improves cell retention and cardiac functions after MI in pigs. This unprecedented combined treatment in a large animal model has therapeutic effects, which can be translated to clinical applications in the foreseeable future.

A bistable polarizer-free electro-optical switch using a droplet manipulation on a liquid crystal and polymer composite film.

A bistable, polarizer-free, and reflective electro-optical switch based on a droplet manipulation on a liquid crystal and polymer composite film (LCPCF) is demonstrated. A color droplet on LCPCF can be manipulated by a wettability gradient owning to the distribution of LC directors anchored among the polymer grains on LCPCF. The contrast ratio is around 8:1 in a reflective mode. The potential applications of droplet manipulation are electronic papers and reflective displays.