Conventional invasive pneumococcal disease (IPD) definition using laboratory confirmation lacks sensitivity. Using a vaccine-probe design, the FinIP trial showed that IPD disease burden and vaccine-preventable disease incidence were fourfold higher when a more sensitive outcome, clinically suspected IPD, was used.

Vaccine-preventable disease incidence (ie, absolute reduction due to PCV10 vaccination) during routine vaccination program was threefold with the more sensitive outcome of clinically suspected IPD compared with the conventional IPD definition. This has major implications for cost-effectiveness of PCVs. (Read the full article)

Small-for-gestational-age neonates are at risk for thrombocytopenia during the first days and weeks after birth. However, the incidence, duration, severity, responsible mechanism, value of platelet transfusions, and risk of death from this variety of neonatal thrombocytopenia are unknown.

Ten percent of thrombocytopenic small-for-gestational-age neonates have a recognized cause for low platelets (aneuploidy, extracorporeal membrane oxygenation, disseminated intravascular coagulation); they have a high mortality rate (65%). Ninety percent have a moderate, transient (2 weeks), hyporegenerative thrombocytopenia with a low mortality rate (2%). (Read the full article)

Increasing percentages of youth are living with chronic medical conditions. Although adolescents face peak risks for onset and intensification of alcohol and marijuana use, we know little about these behaviors and their associations with treatment adherence among chronically ill youth.

This study quantifies alcohol and marijuana use behaviors among a heterogeneous sample of chronically ill youth in aggregate and by condition, and measures associations between alcohol use/binge drinking and knowledge about alcohol interactions with medications/laboratory tests and also treatment nonadherence. (Read the full article)

Small for gestational age neonates (weight <10th percentile) are at risk for neutropenia during the first days after birth. However, the duration, responsible mechanism, and outcomes of this variety of neonatal neutropenia are not precisely known.

Six percent of small for gestational age neonates had neutrophils <1000/μL, with an average neutropenia duration of 7 days. Neutropenia was more closely linked with small for gestational age status than maternal hypertension. This neutropenia is associated with elevated nucleated red blood cell count and increased odds of necrotizing enterocolitis. (Read the full article)

With Pennsylvania Gov. Tom Wolf’s March 25 “Stay at Home” order, Penn State is updating the status of its Abington, Brandywine and Great Valley campuses.

Ashkan Negahban, assistant professor of engineering management, and Satish Srinivasan, assistant professor of information science, will lead projects that help address the COVID-19 crisis, thanks to seed grants from the Huck Institutes of the Life Sciences.

Source: www.brooklynpaper.com - Saturday, May 09, 2020
The owner of the iconic Park Slope events venue Grand Prospect Hall, Michael Halkias, died from COVID-19 on Wednesday. He was 82. Halkias’ death sent shockwaves throughout Brooklyn, where community leaders and friends remember him as a passionate, generous figure. “ He was a Brooklyn character for sure in the best sort of way,” said Randy Peers, president and CEO of the Brooklyn Chamber of Commerce. “He was larger than life.” Halkias and his wife Alice bought Grand Prospect Hall in 1984 and turned the extravagant Prospect Avenue building into an opulent catering hall. The space became a New York icon because of the its long-running, popular commercials, where Alice Halkias declares in a Greek accent, “We make your dreams come true!” Saturday Night Live spoofed the famous commercial in February of 2019, and the pair appeared on Jimmy Kimmel Live in October to remake the ad with Mets player Pete Alonso. Grand Prospect Hall, a Victorian banquet hall built in 1892, attracted big names such as dancer Fred Astaire and mafioso Al Capone during its heyday in the early 20th century — and boasted some of the borough’s oldest treasures, such as Brooklyn’s first reported elevator, which functions to this day. But by the 1980s, the landmarked building had fallen into disrepair: its walls had been painted black, molding had been stripped off the walls, and the chandeliers were gone, Halkias told Brooklyn Paper in 2004. The couple spent 20 yea

Makeda Brome, Pia Hansen, Linda Gojak, Marian Small, Kenneth Baum and David Krulwich share their thoughts on the biggest challenges facing math teachers.

Valve's tweet describes the upcoming title Half-Life: Alyx as the company's 'flagship VR game.' So you'll probably need a virtual headset to play it. The company will reveal more details on Thursday.

An underground stall in Seoul's Yongsan district holds one of the biggest classic video game malls you'll ever see. Take a tour through our photos.

Lyon will play their eighth final in a decade while Barcelona are there for the first time when they meet in Budapest on Saturday.

All posts from the past seven years on race & gender challenges - in one place!

The Education Department's office for civil rights will investigate whether a Georgia elementary school's policy for transgender students contributed to the alleged assault of a girl.

All Classroom Q&A posts on race and gender challenges from the past eight years are described and linked to in this compilation post.

The Dallas String Quartet will deliver its passionate fusion of classical and contemporary music to the Community Arts Center at 7:30 p.m. Thursday, March 19.

Most districts have the technology to support the basic digital textbooks of today, but not the interactive, multimedia-rich ones of the future.

A drill focused on improving movement, teamwork and making the most of scoring chances.

We visit Walderslade FC, where a Manchester United FC regular first made his mark.

Many outside PD opportunities still separate the "haves" from the "have-nots" and uphold systemic oppression.

A panel of experts—including a national teacher's union president and an official from the Department of Education—discussed how to make teaching a more attractive profession.

One OMer’s obedience to God’s plan results in a multiplying and tireless ministry.

United Kingdom and European Union negotiators were working through Sunday as the outcome of Brexit talks hung in the balance.

For a moment, Wall Street seemed to be inching away from Saudi Arabia. Now, it’s already inching back.

The HUB-Robeson Galleries are proud to present "moments," an online thesis exhibition of photography by Zsuzsanna Nagy, a second-year master of fine arts student in photography.

The recent business shutdowns and stay-at-home orders resulting from the coronavirus pandemic haven’t stopped a Penn State student artist from helping her peers show their work remotely after galleries closed across the United States.

The McGregor BFA table showed surprising ‘all-rounder’ listings in the top 30 rankings.

Read an extract from business author and coach Douglas Kruger's latest book, Virus-proof your small business.

State building in the Democratic Republic of Congo is at risk of failing without a new impetus to support democratic consolidation in 2010.

François Hollande semble hésiter à se rendre à Kinshasa en octobre pour le sommet de la francophonie. Il serait bien inspiré de ne pas offrir au président, Joseph Kabila, une telle occasion de s’afficher réconcilié avec les démocraties après les élections présidentielle et législative calamiteuses en République démocratique du Congo (RDC) qui privent le régime d’une véritable légitimité démocratique. En se rendant à Kinshasa, François Hollande enverrait un message plus que trouble aux pays d’Afrique où la démocratisation est encore une lutte quotidienne que l’on paie au prix du sang.

Sensible, inclusive regulation of pastoralism that has mitigated tension in parts of the Sahel should be extended to the Democratic Republic of Congo (DRC) and the Central African Republic (CAR), where conflicts have worsened with the southward expansion of pastoralism.

Background: This study summarizes drug resistance analyses in 4 recent phase 2b trials of the respiratory syncytial virus (RSV) fusion inhibitor presatovir in naturally infected adults.

Methods: Adult hematopoietic cell transplant (HCT) recipients, lung transplant recipients, or hospitalized patients with naturally acquired, laboratory-confirmed RSV infection were enrolled in 4 randomized, double-blind, placebo-controlled studies with study-specific presatovir dosing. Full-length RSV F sequences amplified from nasal swabs obtained at baseline and postbaseline were analyzed by population sequencing. Substitutions at RSV fusion inhibitor resistance-associated positions are reported.

Results: Genotypic analyses were performed on 233 presatovir-treated and 149 placebo-treated subjects. RSV F variant V127A was present in 8 subjects at baseline. Population sequencing detected treatment-emergent substitutions in 10/89 (11.2%) HCT recipients with upper and 6/29 (20.7%) with lower respiratory tract infection, 1/35 (2.9%) lung transplant recipients, and 1/80 (1.3%) hospitalized patients treated with presatovir; placebo-treated subjects had no emergent resistance-associated substitutions. Subjects with substitutions at resistance-associated positions had smaller decreases in viral load during treatment relative to those without, but similar clinical outcomes.

Conclusions: Subject population type and dosing regimen may have influenced RSV resistance development during presatovir treatment. Subjects with vs without genotypic resistance development had decreased virologic responses but comparable clinical outcomes.

QPX7728 is a new ultra-broad-spectrum inhibitor of serine and metallo beta-lactamases from a class of cyclic boronates that gave rise to vaborbactam. The spectrum and mechanism of beta-lactamase inhibition by QPX7728 were assessed using purified enzymes from all molecular classes. QPX7728 inhibits class A ESBLs (IC₅₀ range 1-3 nM) and carbapenemases such as KPC (IC₅₀ 2.9±0.4 nM) as well as class C P99 (IC₅₀ of 22±8 nM) with a potency that is comparable or higher than recently FDA approved BLIs avibactam, relebactam and vaborbactam. Unlike those other BLIs, QPX7728 is also a potent inhibitor of class D carbapenemases such as OXA-48 from Enterobacteriaceae and OXA enzymes from A. baumannii (OXA-23/24/58, IC₅₀ range 1-2 nM) as well as MBLs such as NDM-1 (IC₅₀ 55±25 nM), VIM-1 (IC₅₀ 14±4 nM) and IMP-1 (IC₅₀ 610±70 nM). Inhibition of serine enzymes by QPX7728 is associated with progressive inactivation with a high efficiency k₂/K ranging from of 6.3 x 10⁴ (for P99) to 9.9 x 10⁵ M^-1 s^-1 (for OXA-23). This inhibition is reversible with variable stability of the QPX7728-beta-lactamase complexes with target residence time ranging from minutes to several hours: 5-20 minutes for OXA carbapenemases from A. baumanii, ~50 minutes for OXA-48 and 2-3 hours for KPC and CTX-M-15. QPX7728 inhibited all tested serine enzymes at 1:1 molar ratio. Metallo-beta-lactamases NDM, VIM, and IMP were inhibited by a competitive mechanism with fast-on-fast-off kinetics, with K_is of 7.5±2.1 nM, 32±14 nM and 240±30 nM for VIM-1, NDM-1 and IMP-1, respectively. QPX7728 ultra-broad-spectrum of BLI inhibition combined with its high potency enables combinations with multiple different beta-lactam antibiotics.

Malaria parasites invade and replicate within red blood cells (RBCs), extensively modifying their structure and gaining access to the extracellular environment by placing the plasmodial surface anion channel (PSAC) into the RBC membrane. Expression of members of the cytoadherence linked antigen gene 3 (clag3) family is required for PSAC activity, a process that is regulated epigenetically. PSAC is a well-established route of uptake for large, hydrophilic antimalarial compounds and parasites can acquire resistance by silencing clag3 gene expression, thereby reducing drug uptake. We found that exposure to sub-IC50 concentrations of the histone methyltransferase inhibitor chaetocin caused substantial changes in both clag3 gene expression and RBC permeability, reversing acquired resistance to the antimalarial compound blasticidin S that is transported through PSAC. Chaetocin treatment also altered progression of parasites through their replicative cycle, presumably by changing their ability to modify chromatin appropriately to enable DNA replication. These results indicate that targeting histone modifiers could represent a novel tool for reversing epigenetically acquired drug resistance in P. falciparum.

Carbapenem-resistant Gram-negative pathogens are a critical public health threat and there is an urgent need for new treatments. Carbapenemases (β-lactamases able to inactivate carbapenems) have been identified in both serine β-lactamase (SBL) and metallo β-lactamase (MBL) families. The recent introduction of SBL carbapenemase-inhibitors has provided alternative therapeutic options. Unfortunately, there are no approved inhibitors of MBL-mediated carbapenem-resistance and treatment options for infections caused by MBL-producing Gram-negatives are limited. Here, we present ZN148, a zinc-chelating MBL-inhibitor capable of restoring the bactericidal effect of meropenem and in vitro clinical susceptibility to carbapenems in >98% of a large international collection of MBL-producing clinical Enterobacterales strains (n=234). Moreover, ZN148 was able to potentiate the effect of meropenem against NDM-1-producing Klebsiella pneumoniae in a murine neutropenic peritonitis model. ZN148 showed no inhibition of the human zinc-containing enzyme glyoxylase II at 500 μM and no acute toxicity was observed in an in vivo mouse model with cumulative dosages up to 128 mg/kg. Biochemical analysis showed a time-dependent inhibition of MBLs by ZN148 and removal of zinc ions from the active site. Addition of exogenous zinc after ZN148 exposure only restored MBL activity by ~30%, suggesting an irreversible mechanism of inhibition. Mass-spectrometry and molecular modelling indicated potential oxidation of the active site Cys221 residue. Overall, these results demonstrate the therapeutic potential of a ZN148-carbapenem combination against MBL-producing Gram-negative pathogens and that ZN148 is a highly promising MBL inhibitor, capable of operating in a functional space not presently filled by any clinically approved compound.

Antimicrobial peptides and proteins play critical roles in the host defense against invading pathogens. We recently discovered that recombinantly expressed human and mouse serum amyloid A1 (rhSAA1 and rmSAA1) proteins have potent antifungal activities against the major human fungal pathogen Candida albicans. At high concentrations, rhSAA1 disrupts C. albicans membrane integrity and induces rapid fungal cell death. In the current study, we find that rhSAA1 promotes cell aggregation and targets the C. albicans cell wall adhesin Als3. Inactivation of ALS3 in C. albicans leads to a striking decrease in cell aggregation and cell death upon rhSAA1 treatment, suggesting that Als3 plays a critical role in SAA1 sensing. We further demonstrate that deletion of the transcriptional regulators controlling the expression of ALS3, such as AHR1, BCR1, and EFG1 in C. albicans results in similar effects to that of the als3/als3 mutant upon rhSAA1 treatment. Global gene expression profiling indicates that rhSAA1 has a discernible impact on the expression of cell wall- and metabolism-related genes, suggesting that rhSAA1 treatment could lead to a nutrient starvation effect on C. albicans cells.

Multi-drug resistance among Gram-negative bacteria is a major global public health threat. Metallo-β-lactamases (MBLs) target the most widely-used antibiotic class, the β-lactams, including the most recent-generation carbapenems. Interspecies spread renders these enzymes a serious clinical threat and there are no clinically-available inhibitors. We present crystal structures of IMP-13, a structurally-uncharacterized MBL from Gram-negative Pseudomonas aerugionasa found in clinical outbreaks globally, and characterize the binding using solution NMR-spectroscopy and molecular-dynamics simulations. Crystal structures of apo IMP-13 and bound to four clinically-relevant carbapenem antibiotics (doripenem, ertapenem, imipenem and meropenem) are presented. Active site plasticity and the active-site loop, where a tryptophan residue stabilizes the antibiotic core scaffold, are essential to the substrate-binding mechanism. The conserved carbapenem scaffold plays the most significant role in IMP-13 binding, explaining the broad substrate specificity. The observed plasticity and substrate-locking mechanism provide opportunities for rational drug design of novel metallo-β-lactamase inhibitors, essential in the fight against antibiotic resistance.

Resistance to amoxicillin-clavulanate, a widely used beta-lactam/beta-lactamase inhibitor combination antibiotic, is rising globally, yet susceptibility testing remains challenging. To test whether whole-genome sequencing (WGS) could provide a more reliable assessment of susceptibility than traditional methods, we predicted resistance from WGS for 976 E. coli bloodstream infection isolates from Oxfordshire, UK, comparing against phenotypes from the BD Phoenix (calibrated against EUCAST guidelines). 339/976 (35%) isolates were amoxicillin-clavulanate resistant. Predictions based solely on beta-lactamase presence/absence performed poorly (sensitivity 23% (78/339)) but improved when genetic features associated with penicillinase hyper-production (e.g. promoter mutations, copy number estimates) were considered (sensitivity 82% (277/339); p<0.0001). Most discrepancies occurred in isolates with peri-breakpoint MICs. We investigated two potential causes; the phenotypic reference and the binary resistant/susceptible classification. We performed reference standard, replicated phenotyping in a random stratified subsample of 261/976 (27%) isolates using agar dilution, following both EUCAST and CLSI guidelines, which use different clavulanate concentrations. As well as disagreeing with each other, neither agar dilution phenotype aligned perfectly with genetic features. A random-effects model investigating associations between genetic features and MICs showed that some genetic features had small, variable and additive effects, resulting in variable resistance classification. Using model fixed-effects to predict MICs for the non-agar dilution isolates, predicted MICs were in essential agreement (±1 doubling dilution) with observed (BD Phoenix) MICs for 691/715 (97%) isolates. This suggests amoxicillin-clavulanate resistance in E. coli is quantitative, rather than qualitative, explaining the poorly reproducible binary (resistant/susceptible) phenotypes and suboptimal concordance between different phenotypic methods and with WGS-based predictions.

Objectives: Carbapenemase-producing Enterobacterales and specifically KPC-producing Klebsiella pneumoniae (KPC-Kp) are rapidly spreading worldwide. The prognosis of ventilator-associated pneumonia (VAP) caused by KPC-producing Klebsiella pneumoniae (KPC-Kp) is not well known. Our study tries to assess whether ventilator-associated pneumonia caused by a KPC-Kp strain is associated with higher all-cause mortality than if caused by carbapenem-susceptible isolates.

Study design and methods: This is a retrospective cohort study of patients with VAP due to K. pneumoniae from a 35-bed polyvalent Intensive Care Unit in a university hospital (> 40,000 annual admissions) between January 2012 and December 2016. Adjusted multivariate analysis was used to study the association of KPC-Kp with 30-day all-cause mortality (Cox regression).

Results. We analyze 69 cases of K. pneumoniae VAP of which 39 were produced by a KPC-Kp strain with high-level resistance to meropenem (MIC > 16 mg/mL). All-cause mortality at 30 days was 41% in the KPC-Kp group (16/39) and 33.3% in the carbapenem-susceptible cases (10/30). KPC-Kp etiology was not associated with higher mortality when controlled for confounders (adjusted hazard ratio [lsqb]HR[rsqb] 1.25; 95% CI: 0.46–3.41). Adequate targeted therapy (HR 0.03; 95% CI: <0.01–0.23) was associated with all-cause mortality.

Conclussion. Assuming the limitations due to the available sample size, the prognosis of VAP caused by KPC-Kp is similar to VAPs caused by carbapenem-susceptible K. pneumoniae when appropriate treatment is used.

QPX7728 is an ultra-broad-spectrum boronic acid beta-lactamase inhibitor with potent inhibition of key serine and metallo beta-lactamases observed in biochemical assays. Microbiological studies using characterized strains were used to provide a comprehensive characterization of the spectrum of beta-lactamase inhibition by QPX7728. The MIC of multiple IV only (ceftazidime, piperacillin, cefepime, ceftolozane and meropenem) and orally bioavailable (ceftibuten, cefpodoxime, tebipenem) antibiotics alone and in combination with QPX7728 (4 μg/ml), as well as comparator agents, were determined against the panels of laboratory strains of P. aeruginosa and K. pneumoniae expressing over 55 diverse serine and metallo beta-lactamases. QPX7728 significantly enhanced the potency of antibiotics against the strains expressing Class A extended spectrum beta-lactamases (CTX-M, SHV, TEM, VEB, PER) and carbapenemases (KPC, SME, NMC-A, BKC-1), consistent with beta-lactamase inhibition demonstrated in biochemical assays. It also inhibits both plasmidic (CMY, FOX, MIR, DHA) and chromosomally encoded (P99, PDC, ADC) Class C beta-lactamases and Class D enzymes including carbapenemases such as OXA-48 from Enterobacteriaceae and OXA enzymes from Acinetobacter baumannii (OXA-23/24/72/58). QPX7728 is also a potent inhibitor of many class B metallo beta-lactamases (NDM, VIM, CcrA1, IMP, GIM but not SPM or L1). Addition of QPX7728 (4 μg/ml) reduced the MICs in a majority of strains to the level observed for the vector alone control, indicative of complete beta-lactamase inhibition. The ultra-broad-spectrum beta-lactamase inhibition profile makes QPX7728 a viable candidate for further development.

QPX7728 is an ultra-broad-spectrum boronic acid beta-lactamase inhibitor that demonstrates inhibition of key serine and metallo beta-lactamases at a nano molar range in biochemical assays with purified enzymes. The broad-spectrum inhibitory activity of QPX7728 observed in biochemical experiments translates into enhancement of the potency of many beta-lactams against strains of target pathogens producing beta-lactamases. The impact of bacterial efflux and permeability on inhibitory potency were determined using isogenic panels of KPC-3 producing isogenic strains of K. pneumoniae and P. aeruginosa and OXA-23-producing strains of A. baumannii with various combinations of efflux and porin mutations. QPX7728 was minimally affected by multi-drug resistance efflux pumps in either Enterobacteriaceae, or in non-fermenters such as P. aeruginosa or A. baumannii. In P. aeruginosa, the potency of QPX7728 was further enhanced when the outer membrane is permeabilized. The potency of QPX7728 in P. aeruginosa is not affected by inactivation of the carbapenem porin OprD. While changes in OmpK36 (but not OmpK35) reduced the potency of QPX7728 (8-16-fold), QPX7728 (4 μg/ml) nevertheless completely reversed KPC-mediated meropenem resistance in strains with porin mutations, consistent with a lesser effect of these mutations on the potency of QPX7728 compared to other agents. The ultra-broad-spectrum beta-lactamase inhibition profile combined with enhancement of the activity of multiple beta-lactam antibiotics with varying sensitivity to the intrinsic resistance mechanisms of efflux and permeability indicate QPX7728 is a useful inhibitor for use with multiple beta-lactam antibiotics.

Carbapenem pharmacokinetic profiles are significantly changed in critically ill patients because of the drastic variability of the patients' physiological parameters. Published population PK studies have mainly focused on specific diseases and the majority of these studies had small sample sizes. The aim of this study was to develop a population PK model of imipenem in critically ill patients that estimated the influence of various clinical and biological covariates and the use of Extracorporeal Membrane Oxygenation (ECMO) and Continuous Renal Replacement Therapy (CRRT). A two-compartment population PK model with Creatinine clearance (CrCL), body weight (WT), and ECMO as fixed effects was developed using the non-linear mixed effect model (NONMEM). A Monte Carlo simulation was performed to evaluate various dosing schemes and different levels of covariates based on the pharmacokinetic/pharmacodynamic index (f%T>MIC) for the range of clinically relevant minimum inhibitory concentrations(MICs). The results showed that there may be insufficient drug use in the clinical routine drug dose regimen, and 750mg Q6h could achieve a higher treatment success rate. The blood concentrations of imipenem in ECMO patients were lower than that of non-ECMO patients, therefore dosage may need to be increased. The dosage may need adjustment for patients with CrCL ≤ 70ml/min, but dose should be lowered carefully to avoid the insufficient drug exposure. Dose adjustment is not necessary for patients within the WT ranging from 50-80 kg. Due to the large variation in PK profile of imipenem in critically ill patients, TDM should be carried out to optimize drug regimens.

Treatment of dogs naturally infected with Leishmania infantum using meglumine antimoniate (MA) encapsulated in conventional liposomes (LC) in association with allopurinol has been previously reported to promote marked reduction in the parasite burden in the main infection sites. Here, a new assay in naturally infected dogs was performed using a novel liposome formulation of MA consisting of a mixture of conventional and long-circulating (PEGylated) liposomes (LCP), with expected broader distribution among affected tissues of the mononuclear phagocyte system. Experimental groups of naturally infected dogs were as follows: LCP+Allop, receiving LCP intravenously as 2 cycles of 6 doses (6.5 mg Sb/kg/dose) at 4-day intervals, plus allopurinol at 30 mg/kg/12 h p.o. during 130 days; LC+Allop, receiving LC intravenously as 2 cycles of 6 doses (6.5 mg Sb/kg/dose), plus allopurinol during 130 days; Allop, treated with allopurinol only; non-treated control. Parasite loads were evaluated by quantitative PCR in liver, spleen and bone marrow and by immunohistochemistry in the ear skin, before, just after treatment and 4 months later. LCP+Allop and LC+Allop groups, but not the Allop group, showed significant suppression of the parasites in the liver, spleen and bone marrow 4 months after treatment, compared to the pre-treatment period or the control group. Only LCP+Allop group showed significantly lower parasite burden in the skin, in comparison to the control group. On the basis of clinical staging and parasitological evaluations, LCP formulation exhibited a more favorable therapeutic profile, when compared to LC one, being therefore promising for treatment of canine visceral leishmaniasis.

Pharmacokinetic changes are often seen in patients with severe infections. Administration by continuous infusion has been suggested to optimize antibiotic exposure and pharmacokinetic/pharmacodynamic (PK/PD) target attainment for β-lactams. In an observational study, unbound piperacillin concentrations (n=196) were assessed in 78 critically ill patients following continuous infusion of piperacillin/tazobactam (ratio 8:1). The initial dose of 8, 12 or 16 g (piperacillin component) was determined by individual creatinine clearance (CRCL). Piperacillin concentrations were compared to the EUCAST clinical breakpoint MIC for Pseudomonas aeruginosa (16 mg/L), and the following PK/PD targets were evaluated: 100% fT>1xMIC and 100% fT>4xMIC. A population pharmacokinetic model was developed using NONMEM 7.4.3 consisting of a one-compartment disposition model with linear elimination separated into non-renal and renal (linearly increasing with patient CRCL) clearances. Target attainment was predicted and visualized for all individuals based on the utilized CRCL dosing algorithm. The target of 100% fT>1xMIC was achieved for all patients based on the administered dose, but few patients achieved the target of 100% fT>4xMIC. Probability of target attainment for a simulated cohort of patients showed, that increasing the daily dose by 4 g increments (piperacillin component) did not result in substantially improved target attainment for the 100% fT>4xMIC target. To conclude, in patients with high CRCL combined with high-MIC bacterial infections, even a CI regimen with a daily dose of 24 g may be insufficient to achieve therapeutic concentrations.

Despite the worldwide re-emergence of the chikungunya virus (CHIKV) and the high morbidity associated with CHIKV infections, there is no approved vaccine or antiviral treatment available. We here aim to identify the target of a novel class of CHIKV inhibitors i.e. CHVB series. CHVB compounds inhibit the in vitro replication of CHIKV isolates with 50% effective concentrations in the low micromolar range. A CHVB-resistant variant (CHVB^res) was selected that carried two mutations in the gene encoding nsP1 (responsible for viral RNA capping), one mutation in nsP2 and one mutation in nsP3. Reverse genetics studies demonstrated that both nsP1 mutations were necessary and sufficient to achieve ~18-fold resistance, suggesting that CHVB targets viral mRNA capping. Interestingly, CHVB^res was cross-resistant to the previously described CHIKV capping inhibitors from the MADTP series, suggesting they share a similar mechanism of action. In enzymatic assays, CHVB inhibited the methyltransferase and guanylyltransferase activities of alphavirus nsP1 proteins. To conclude, we identified a class of CHIKV inhibitors that targets the viral capping machinery. The potent anti-CHIKV activity makes this chemical scaffold a potential candidate for CHIKV drug development.

As resistance to artemisinins (current frontline drugs in malaria treatment) emerges in south East Asia, there is an urgent need to identify the genetic determinants and understand the molecular mechanisms underpinning such resistance. Such insights could lead to prospective interventions to contain resistance and prevent the eventual spread to other malaria endemic regions. Artemisinin reduced susceptibility in South East Asia (SEA) has been primarily linked to mutations in P. falciparum Kelch-13, which is currently widely recognised as a molecular marker of artemisinin resistance. However, 2 mutations in a ubiquitin hydrolase, UBP-1, have been previously associated with artemisinin reduced susceptibility in a rodent model of malaria and some cases of UBP-1 mutation variants associating with artemisinin treatment failure have been reported in Africa and SEA. In this study, we have employed CRISPR-Cas9 genome editing and pre-emptive drug pressures to test these artemisinin susceptibility associated mutations in UBP-1 in P. berghei sensitive lines in vivo. Using these approaches, we have shown that the V2721F UBP-1 mutation results in reduced artemisinin susceptibility, while the V2752F mutation results in resistance to chloroquine and moderately impacts tolerance to artemisinins. Genetic reversal of the V2752F mutation restored chloroquine sensitivity in these mutant lines while simultaneous introduction of both mutations could not be achieved and appears to be lethal. Interestingly, these mutations carry a detrimental growth defect, which would possibly explain their lack of expansion in natural infection settings. Our work has provided independent experimental evidence on the role of UBP-1 in modulating parasite responses to artemisinin and chloroquine under in vivo conditions.

Treatment of exacerbations of chronic Pseudomonas aeruginosa infections in patients with cystic fibrosis (CF) is highly challenging due to hypermutability, biofilm formation and an increased risk of resistance emergence. We evaluated the impact of ciprofloxacin and meropenem as monotherapy and in combination in the dynamic in vitro CDC biofilm reactor (CBR). Two hypermutable P. aeruginosa strains, PAOmutS (MIC_{ciprofloxacin} 0.25 mg/L, MIC_meropenem 2 mg/L) and CW44 (MIC_{ciprofloxacin} 0.5 mg/L, MIC_meropenem 4 mg/L), were investigated for 120h. Concentration-time profiles achievable in epithelial lining fluid (ELF) following FDA-approved doses were simulated in the CBR. Treatments were ciprofloxacin 0.4g every 8h as 1h-infusions (80% ELF penetration), meropenem 6 g/day as continuous infusion (CI; 30% and 60% ELF penetration) and their combinations. Counts of total and less-susceptible planktonic and biofilm bacteria and MICs were determined. Antibiotic concentrations were quantified by UHPLC-PDA. For both strains, all monotherapies failed with substantial regrowth and resistance of planktonic (≥8log₁₀ CFU/mL) and biofilm (>8log₁₀ CFU/cm²) bacteria at 120h (MIC_{ciprofloxacin} up to 8 mg/L, MIC_meropenem up to 64 mg/L). Both combination treatments demonstrated synergistic bacterial killing of planktonic and biofilm bacteria of both strains from ~48h onwards and suppressed regrowth to ≤4log₁₀ CFU/mL and ≤6log₁₀ CFU/cm² at 120h. Overall, both combination treatments suppressed amplification of resistance of planktonic bacteria for both strains, and biofilm bacteria for CW44. The combination with meropenem at 60% ELF penetration also suppressed amplification of resistance of biofilm bacteria for PAOmutS. Thus, combination treatment demonstrated synergistic bacterial killing and resistance suppression against difficult-to-treat hypermutable P. aeruginosa strains.