Introduction: The accelerating threat of multidrug-resistant bacteria (MRB) forces health care providers to use antibiotics more rationally. Antibiotic stewardship programs (ASP) are a proven and safe way to achieve that goal. They have been comprehensively studied in adults but data from secondary care pediatric hospitals are lacking. Material and Methods: In our study an ASP with standard operating procedures (SOPs), audits, a weekly ward round with experts in pediatric infectious diseases and an antibiotic pocket-card for selected infectious diseases was established in July 2017 in a Munich municipal secondary care children's hospital. All antibiotic prescriptions on general pediatric wards were reviewed each in the first quarter of 2017 and 2018. The primary outcome was adherence to treatment guidelines. Secondary outcomes were substance consumption, duration of therapy and death. Results: After the ASP was implemented guideline adherence increased significantly from 33 to 63%. The consumption of cephalosporins decreased significantly (-60%), whereas aminopenicillin use increased accordingly (+120%). Neither in the pre- nor in the post-intervention group deaths occurred. Discussion: Data on ASP in pediatric secondary care hospitals are scarce. Most previous studies have been performed at tertiary care/university children's hospitals. We demonstrate a significant improvement in guideline adherence regarding antibiotic treatments after the implementation of an ASP. Cephalosporin consumption decreased which might be relevant for the selection of MRB (e.g., vancomycin-resistant enterococci). Results are limited by the single-center design and the short observation period. The study encourages the implementation of ASPs in secondary care children's hospitals.

Meningitis caused by Salmonella enteritidis is an uncommon infection, linked with great mortality and neurological problems, which makes prompt diagnosis and treatment very crucial. Patients of bacterial meningitis present with common symptoms such as headache, fever, altered level of consciousness, and neck rigidity. A positive gram stain or culture of cerebrospinal fluid (CSF) leads to its diagnosis. In this report, we present a case of a 16-year-old male with extensively drug-resistant (XDR) Salmonella typhi meningitis, which was not responsive to initial medical intervention. He was treated with meropenem, imipenem, azithromycin, and metronidazole. Immediate tracheostomy and intubation were performed in the surgical intensive care unit (ICU), as the patient had developed stridor, shortness of breath, tachypnea, tachycardia, and had severely decreased O2 saturation of 60%. As far as treatment is concerned, third-generation cephalosporins are considered the treatment of choice. In addition, the use of fluoroquinolones and carbapenems, mainly meropenem, has also been described as a therapeutic alternative.

Multidrug-resistant Klebsiella pneumoniae is an increasing cause of infant mortality in developing countries. We aimed to develop a quantitative understanding of the drivers of this epidemic by estimating the effects of antibiotics on nosocomial transmission risk, comparing competing hypotheses about mechanisms of spread, and quantifying the impact of potential interventions. Using a sequence of dynamic models, we analysed data from a one-year prospective carriage study in a Cambodian neonatal unit with hyperendemic third-generation cephalosporin-resistant K. pneumoniae. All widely-used antibiotics except imipenem were associated with an increased daily acquisition risk, with an odds ratio for the most common combination (ampicillin + gentamicin) of 1.96 (95% CrI 1.18, 3.36). Models incorporating genomic data found that colonisation pressure was associated with a higher transmission risk, indicated sequence type heterogeneity in transmissibility, and showed that within-ward transmission was insufficient to maintain endemicity. Simulations indicated that increasing the nurse-patient ratio could be an effective intervention.

Clavulanic acid is a bacterial specialized metabolite, which inhibits certain serine β-lactamases, enzymes that inactivate β-lactam antibiotics to confer resistance. Due to this activity, clavulanic acid is widely used in combination with penicillin and cephalosporin (β-lactam) antibiotics to treat infections caused by β-lactamase-producing bacteria. Clavulanic acid is industrially produced by fermenting Streptomyces clavuligerus, as large-scale chemical synthesis is not commercially feasible. Other than S. clavuligerus, Streptomyces jumonjinensis and Streptomyces katsurahamanus also produce clavulanic acid along with cephamycin C, but information regarding their genome sequences is not available. In addition, the Streptomyces contain many biosynthetic gene clusters thought to be "cryptic," as the specialized metabolites produced by them are not known. Therefore, we sequenced the genomes of S. jumonjinensis and S. katsurahamanus, and examined their metabolomes using untargeted mass spectrometry along with S. clavuligerus for comparison. We analyzed the biosynthetic gene cluster content of the three species to correlate their biosynthetic capacities, by matching them with the specialized metabolites detected in the current study. It was recently reported that S. clavuligerus can produce the plant-associated metabolite naringenin, and we describe more examples of such specialized metabolites in extracts from the three Streptomyces species. Detailed comparisons of the biosynthetic gene clusters involved in clavulanic acid (and cephamycin C) production were also performed, and based on our analyses, we propose the core set of genes responsible for producing this medicinally important metabolite.

Cefepime is a fourth-generation cephalosporin antibiotic with an extended spectrum of activity against many Gram-positive and Gram-negative bacteria. There is a growing need to develop sensitive, small volume assays, along with less invasive sample collection to facilitate pediatric pharmacokinetic clinical trials and therapeutic drug monitoring. The volumetric absorptive microsampling (VAMS™) approach provides an accurate and precise collection of a fixed volume of blood (10 μL), reducing or eliminating the volumetric blood hematocrit assay-bias associated with the dried blood spotting technique. We developed a high-performance liquid chromatographic method with tandem mass spectrometry detection for quantification of cefepime. Sample extraction from VAMS™ devices, followed by reversed-phase chromatographic separation and selective detection using tandem mass spectrometry with a 4 min runtime per sample was employed. Standard curves were linear between 0.1-100 μg/mL for cefepime. Intra- and inter-day accuracies were within 95.4-113% and precision (CV) was < 15 % based on a 3-day validation study. Recoveries ranged from 40.8 to 62.1% and the matrix effect was within 89.5-96.7% for cefepime. Cefepime was stable in human whole blood under assay conditions (3 h at room temperature, 24 h in autosampler post-extraction). Cefepime was also stable for at least 1 week (7 days) at 4 °C, 1 month (39 days) at -20 °C and 3 months (91 days) at -78 °C as dried microsamples. This assay provides an efficient quantitation of cefepime and was successfully implemented for the analysis of whole blood microsamples in a pediatric clinical trial.

Carbapenemase-producing Enterobacterales expressing OXA-48, KPC, NDM, VIM or IMP enzymes are increasingly reported worldwide. We have characterized LMB-1, a novel metallo-β-latamase (MBL) of Ambler class B3 from Citrobacter freundii 164 (Cf164) clinical isolate from Buenos Aires, Argentina. Cf164 displayed reduced susceptibility to carbapenems but gave inconsistent results with carbapenemase confirmatory tests, suggesting the presence of a weak carbapenemase. Analysis of WGS of Cf164 using Resfinder revealed four β-lactamase genes coding for CTX-M-8, PER-2, TEM-1 and CMY-150, a novel chromosomally-encoded CMY variant. Kinetic parameters of purified CMY-150 did not reveal any carbapenemase activity. However, CMY-150 conferred to E. coli higher MIC values for ceftazidime and aztreonam as compared to CMY-2. The in-house developed β-lactamase search software (ResMINER) in WGS data, revealed a novel subclass B3 MBL named LMB-1. LMB-1 conferred to E. coli, resistance to penicillins, to expanded-spectrum cephalosporins and reduced susceptibility to carbapenems. The blaLMB-1 gene was located on a 176-kb IncA/C2 plasmid. LMB-1 shared 99% of amino acid sequence identity with the MBL encoded in the chromosome of Rheinheimera pacifica, it's likely progenitor. Despite repeated attempts, LMB-1 could not be purified, thus only specific activities could evidence hydrolysis of carbapenems. Here we report CMY-150, a novel CMY-2 variant that confers increased ceftazidime and aztreonam MICs to E. coli and the first description of LMB-1 in Argentina. This work underlines the need for several CPE confirmatory tests, as this novel enzyme might have been missed using only one.

Cephalosporins are among the most frequently used broad-spectrum antimicrobial agents. Ceftazidime is a semisynthetic β-lactam antibiotic for parenteral administration widely used in the clinical practice, which has been categorized as a third-generation cephalosporin antibiotic. This drug crystallizes as a pentahydrate and, as all cephalosporin, it is unstable and subject to hydrolytic degradation. Taking this into account, this study investigates the stability under two different conditions (high temperature and exposition to vacuum) by using various techniques, as thermal analysis, Raman spectroscopy and powder X-ray diffraction supported by Multivariate Curve Resolution (MCR). It was proved that ceftazidime pentahydrate is unstable under the studied variables, exhibiting several transformation processes, which were discussed in terms of the crystalline structure.