Antimicrobial agents have been at the forefront of global efforts to treat bacterial infections. However, their efficacy and reliability have steadily declined due to the widespread emergence of multidrug resistance (MDR). This challenge is particularly critical for Vibrio cholerae, the etiological agent of cholera, a disease that remains a significant global health burden in endemic and outbreak-prone regions. The increasing prevalence of MDR V. cholerae strains, coupled with a stagnating pipeline of new antibiotics, underscores the urgent need for novel antibacterial targets and therapeutic strategies. The sodium-translocating NADH:ubiquinone oxidoreductase (NQR) is a ubiquitous and essential aerobic respiratory enzyme found in several MDR bacterial pathogens, including V. cholerae, Pseudomonas aeruginosa, Chlamydia trachomatis, and Klebsiella pneumoniae. Herein, we report the first systematic investigation of quinolone-based inhibitors targeting NQR, involving the design, synthesis, and biological assessment of a series of 3-substituted quinolones inspired by the antimalarial lead compound endochin and other highly toxic quinolones. Our structure-guided optimization strategy mitigates the high toxicity historically associated with quinolone derivatives, yielding analogs with improved safety profiles. These compounds exhibit enhanced potency and selectivity against NQR, highlighting their potential as a new class of antibacterial agents against MDR pathogens.

The therapeutic efficacy of antibiotics has been significant in extending human life expectancy by combating virulent bacterial infections. Nevertheless, multidrug-resistant (MDR) microorganisms remain a global crisis as these bacteria have developed resistance to conventional antibacterial agents. An unexplored antibiotic target found exclusively in bacteria is the Na+-translocating NADH:ubiquinone oxidoreductase (NQR), which is an indispensable membrane-bound bacterial enzyme complex that enables cellular functionality and is present in many infectious bacterial species, including Vibrio cholerae and H. influenzae. NQR serves as an essential complex in the bacterial electron transport chain (ETC) and operates as a highly conserved primary Na+ pump that drives many bioenergetic functions. This six-subunit protein shuttles electrons from NADH to ubiquinone, which drives the translocation of Na+ ions and creates a gradient that provides the driving force for various cellular processes. We have synthesized and evaluated a series of 1,4-naphthoquinones that exhibit high potency against NQR with minimal cytotoxicity and potential to serve as new, NQR-targeting antibacterial agents for use against V. cholerae.

A series of N-aryl sulfonamides was designed, synthesized, and evaluated for their ability to inhibit the bacterial enzyme N-succinyl-L,L-diaminopimelic acid desuccinylase (DapE, EC 3.5.1.18). Our initial lead compound, N-(o-methoxyphenyl)-toluenesulfonamide (4a), identified through a high-throughput screen, showed modest inhibition, with an IC₅₀ of 72.9 μM. Through lead optimization, we improved the potency by modifying key functional groups. Replacing the o-OCH₃ on the aniline ring and p-CH₃ on the sulfonamide ring with p-CF₃ and p-OCH₃, respectively, while retaining the N-CHF₂ hydrophobic moiety, led to our most potent inhibitor, compound 4g (IC₅₀ = 8.9 ± 5.0 μM, Ki = 1.36 ± 0.61 μM), representing an 8-fold increase in potency over hit 4a, and providing the most potent DapE inhibitor yet reported based on the Ki. Molecular docking studies provided insights into this enhanced activity, revealing key interactions within the DapE active site.

Diaminopimelate desuccinylase (DapE) is a promising antibacterial target involved in lysine biosynthesis and bacterial cell wall formation. Previously, N-aryl sulfonamide hit 1 (IC₅₀ = 72.9 μM) was optimized to 2 with enhanced potency (IC₅₀ = 8.9 ± 5.0 μM, Kᵢ = 1.36 ± 0.61 μM). In this paper we describe the synthesis of N-heterocyclic sulfonamides and N-aryl sultams to explore the effects of heterocycles and conformational constraints on DapE inhibition. Aminopyridine substitutions generally reduced activity, although N-pyridyl sulfonamide 6e retained moderate inhibition (68.5% at 100 μM). Five- and seven-membered sultams showed modest activity, with p-methoxybenzene 7b displaying the highest inhibition (60.4% at 100 μM). Docking studies predicted favorable zinc-binding for several analogues, particularly those with carboxylic acids, but these interactions generally did not improve experimental potency.

PAX (antiProtonic Atom X-ray spectroscopy) is a new experiment with the aim to test strong-field quantum electrodynamics (QED) effects by performing high-precision x-ray spectroscopy of antiprotonic atoms. By utilizing advanced microcalorimeter detection techniques and a low-energy antiproton beam provided by the ELENA ring at CERN, gaseous targets will be used for the creation of antiprotonic atoms, and the measurement of transitions between circular Rydberg states will be conducted with up to two orders of magnitude improved accuracy over previous studies using high-purity germanium detectors. Our approach eliminates the longstanding issue of nuclear uncertainties that have hindered prior studies using highly charged ions, thus enabling direct and purely QED-focused measurements. By precisely probing atomic systems with electric fields up to two orders of magnitude above the Schwinger limit, PAX will test vacuum polarization and second-order QED corrections, opening new frontiers in fundamental physics and uncovering potential pathways to physics beyond the Standard Model.

Based on the inhibitory potencies from earlier reported tetrazole thioether analogs, we now describe the synthesis and inhibition of pyrazole-based inhibitors of N-succinyl-l,l-2,6-diaminopimelic acid desuccinylase (DapE) from Haemophilus influenzae (HiDapE). The most potent pyrazole analog 7d bears an aminopyridine amide with an IC50 of 17.9 +/- 8.0 mu M, and the single enantiomer of alpha-methyl analog 7q has an IC50 of 18.8 mu M, with potency residing in the (R)-enantiomer. Thermal shift revealed strong stabilization upon binding inhibitor (R)-7q with Tm = 50.2 degrees C and a Ki of 17.3 +/- 2.8 mu M. Enzyme kinetic experiments confirm competitive inhibition, and docking reveals key active site interactions.

A Super-Conducting ENergetic x-ray Telescope (ASCENT) is a concept for a future balloon-borne high-energy X-ray telescope in the energy range 60–85 keV to study gamma-ray emissions of 67.87 keV and 78.32 keV from the radioactive isotope 44Ti. For the focal plane instrumentation, ASCENT will use Mo-Cu/Mo-Au bilayer transition edge sensor (TES) microcalorimeter gamma-ray detectors with tin (Sn) absorbers. ‘Spectrometer to Leverage Extensive Development of Gamma-ray TESs for Huge Arrays using Microwave Multiplexed Enabled Readout’ (SLEDGEHAMMER), a detector development project at the National Institute of Standards and Technology, acts as the basis for the detector arrays for ASCENT. SLEDGEHAMMER has tin (Sn) absorbers attached to the SU-8 epoxy posts, lithographically placed on the detectors, but we are also considering other geometries for the chips where the absorbers are attached to the chips separated from the TESs, which could help to avoid parallel path for a current flow around the detectors with these BiSn sphere attachments. In this work, we are reporting on developing a method to attach Tin (Sn) absorbers to the transition edge sensors (TESs) with 0.2 mm diameter BiSn solder spheres replacing epoxy. The goal is to improve the thermal conductivity between the absorbers and the TESs compared to what was achieved using epoxy, potentially reducing the presence of an athermal component in the tails of signal pulses. We describe our efforts toward finding optimal temperature and pressure conditions for making this contact and the progress toward contact resistance measurements of these joints.

Antimicrobial resistance is a significant cause of mortality globally due to infections, a trend that is expected to continue to rise. As existing treatments fail and new drug discovery slows, the urgency to develop novel antimicrobial therapeutics grows stronger. One promising strategy involves targeting bacterial systems exclusive to pathogens, such as the transcription regulator protein GabR. Expressed in diverse bacteria including Escherichia coli, Bordetella pertussis, and Klebsiella pneumoniae, GabR has no homolog in eukaryotes, making it an ideal therapeutic target. Bacillus subtilis GabR (bsGabR), the most studied variant, regulates its own transcription and activates genes for GABA aminotransferase (GabT) and succinic semialdehyde dehydrogenase (GabD). This intricate regulatory system presents a compelling antimicrobial target with the potential for agonistic intervention to disrupt bacterial gene expression and induce cellular dysfunction, especially in bacterial stress responses. To explore manipulation of this system and the potential of this protein as an antimicrobial target, an in-depth understanding of the unique PLP-dependent transcription regulation is critical. Herein, we report the successful structural modification of the cofactor PLP and demonstrate the biochemical reactivity of the PLP analog pyridoxal-5 '-tetrazole (PLT). Through both spectrophotometric and X-ray crystallographic analyses, we explore the interaction between bsGabR and PLT, together with a synthesized GABA derivative (S)-4-amino-5-phenoxypentanoate (4-phenoxymethyl-GABA or 4PMG). Most notably, we present a crystal structure of the condensed, external aldimine complex within bsGabR. While PLT alone is not a drug candidate, it can act as a probe to study the detailed mechanism of GabR-mediated function. PLT employs a tetrazole moiety as a bioisosteric replacement for phosphate in PLP. In addition, the PLP-4PMG adduct observed in the structure may serve as a novel chemical scaffold for subsequent structure-based antimicrobial design.

The proposed Pandemic Agreement must ensure that COVID-19 vaccine nationalism is never repeated; 290 scientists call for action. The proposed Pandemic Agreement must ensure that COVID-19 vaccine nationalism is never repeated; 290 scientists call for action.

We explore the role of Australian black flying foxes ( Pteropus alecto ) as reservoir hosts of potentially zoonotic Borrelia bacteria. Across six sites, 2% of 840 bats were infected with one of two novel Borrelia haplotypes. Phylogenetic reconstruction indicated these infections are distinct from Lyme or relapsing fever clades. ### Competing Interest Statement The authors have declared no competing interest.

There is an urgent need for new antibiotics given the rise of antibiotic resistance, and succinyl-diaminopimelate desuccinylase (DapE, E.C. 3.5.1.18) has emerged as a promising bacterial enzyme target. DapE from Haemophilus influenzae (HiDapE) has been studied and inhibitors identified, but it is essential to explore DapE from different species to assess selective versus broad-spectrum therapeutics. We have determined the structure of DapE from the ESKAPE pathogen Acinetobacter baumannii (AbDapE) and studied inhibition by known inhibitors of HiDapE. AbDapE is inhibited by captopril and sulfate comparable to HiDapE, but AbDapE was not significantly inhibited by a known indoline sulfonamide HiDapE inhibitor. Captopril and sulfate both stabilize HiDapE by increasing the thermal melting temperature (Tm) in thermal shift assays. By contrast, sulfate decreases the stability of the AbDapE enzyme, whereas captopril increases the stability. Further, we report two crystal structures of selenomethionine-substituted AbDapE in the closed conformation, one with AbDapE in complex with succinate derived from enzymatic hydrolysis of N6-methyl-l,l-SDAP substrate and acetate (PDB code 7T1Q, 2.25 Å resolution), and a crystal structure of AbDapE with bound succinate along with l-(S)-lactate, a product of degradation of citric acid from the crystallization buffer during X-ray irradiation (PDB code 8F8O, 2.10 Å resolution).

Bacteria are becoming increasingly resistant to antibiotics, therefore there is an urgent need for new classes of antibiotics to fight antibiotic resistance. Mammals do not express Nɑ -acetyl-L-ornithine deacetylase (ArgE), an enzyme that is critical for bacterial survival and growth, thus ArgE represents a promising new antibiotic drug target, as inhibitors would not suffer from mechanism-based toxicity. A new ninhydrin-based assay was designed and validated that included the synthesis of the substrate analog N5, N5-di-methyl Nα-acetyl-L-ornithine (kcat/Km = 7.32 ± 0.94 × 104 M−1s−1). This new assay enabled the screening of potential inhibitors that absorb in the UV region, and thus is superior to the established 214 nm assay. Using this new ninhydrin-based assay, captopril was confirmed as an ArgE inhibitor (IC50 = 58.7 μM; Ki = 37.1 ± 0.85 μM), and a number of phenylboronic acid derivatives were identified as inhibitors, including 4-(diethylamino)phenylboronic acid (IC50 = 50.1 μM). Selected inhibitors were also tested in a thermal shift assay with ArgE using SYPRO Orange dye against Escherichia coli ArgE to observe the stability of the enzyme in the presence of inhibitors (captopril Ki = 35.9 ± 5.1 μM). The active site structure of di-Zn EcArgE was confirmed using X-ray absorption spectroscopy, and we reported two X-ray crystal structures of E. coli ArgE. In summary, we describe the development of a new ninhydrin-based assay for ArgE, the identification of captopril and phenylboronic acids as ArgE inhibitors, thermal shift studies with ArgE + captopril, and the first two published crystal structures of ArgE (mono-Zn and di-Zn).

Based on our previous success in using cyclobutanone derivatives as enzyme inhibitors, we have designed and prepared a 37-member library of α-aminocyclobutanone amides and sulfonamides, screened for inhibition of the bacterial enzyme diaminopimelate desuccinylase (DapE), which is a promising antibiotic target, and identified several inhibitors with micromolar inhibitory potency. Molecular docking suggests binding of the deprotonated hydrate of the strained cyclobutanone, and thermal shift analysis with the most potent inhibitor (3y, IC50 = 23.1 µM) enabled determination of a Ki value of 10.2 +/− 0.26 µM and observed two separate Tm values for H. influenzae DapE (HiDapE).

Thousands of scientists and practitioners conduct research on infectious diseases of wildlife. Rapid and comprehensive data sharing is vital to the transparency and actionability of their work, but unfortunately, most efforts designed to publically share these data are focused on pathogen determination and genetic sequence data. Other facets of existing surveillance data – particularly negative results – are often withheld or, at best, summarized in a descriptive table with limited metadata. As a result, very few datasets on wildlife disease dynamics over space and time are publicly available for synthesis research or applied uses in conservation or public health. Here, we propose a minimum data and metadata reporting standard for wildlife disease studies. Our checklist identifies a minimum set of 30 fields required to standardize and document a dataset consisting of records disaggregated to the finest possible spatial, temporal, and taxonomic scale. We illustrate how this standard is applied to an example study, which documented a novel alphacoronavirus found in bats in Belize. Finally, we outline best practices for how data should be formatted for optimal re-use, and how researchers can navigate potential safety concerns around data sharing.

The Eulipotyphla (true insectivores) is the third largest mammalian order, comprising over 500 species, and could be an important source of human infectious diseases. However, relatively little is known about the diversity of microbes in insectivores and the contribution of insectivores to virus transmission more specifically among wild hosts. In this study, we compiled a comprehensive dataset containing over 400,000 records of insectivores and their associated microbes from 1903 to 2023. Our analyses showed that insectivores host a wide spectrum of 941 microbes, 60% of which are viruses and are predominantly found in the shrew and hedgehog families. Notably, human-associated viruses harbored by shrews and hedgehogs were phylogenetically closely related to those found in humans, suggesting potential bidirectional transmission between insectivores and humans. Moreover, virus-sharing networks revealed that insectivores held the second-most central position for virus sharing, just second to bats, among all mammalian orders. Insectivores had a high proportion of cross-order transmitted viruses, including many human-associated viruses. Dietary diversity, habitat diversity, and distributional traits (e.g. geographical range size, mean latitude, and urban adaptation status) emerged as the key ecological factors contributing to this cross-species virus transmission. Our findings highlight the microbial diversity present in insectivores, indicating this order may act as potential incubators for novel viruses capable of infecting mammals and spreading viruses of public health concern. ### Competing Interest Statement The authors have declared no competing interest.

Cytochrome P450 2D6 (CYP2D6) is one of the most important enzymes involved in drug metabolism. Genetic polymorphism can influence drug metabolism by CYP2D6 such that a therapy is seriously affected by under- or overdosing of drugs. However, a general explanation at the atomistic level for poor activity is missing so far. Here we show for the 20 most common single nucleotide polymorphisms (SNPs) of CYP2D6 that poor metabolism is driven by four mechanisms. We found in extensive all-atom molecular dynamics simulations that the rigidity of the I-helix (central helix), distance between central phenylalanines (stabilizing bound substrate), availability of basic residues on the surface of CYP2D6 (binding of Cytochrome P450 reductase), and position of arginine 132 (electron transfer to heme) are essential for an extensive function of the enzyme. These results were applied to SNPs with unknown effects and potential SNPs that may lead to poor drug metabolism were identified. The revealed molecular mechanisms might be important for other drug-metabolizing Cytochrome P450 enzymes.

Growing antibiotic resistance by pathogenic bacteria has led to a global crisis. The bacterial enzyme N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) provides a very attractive target for the discovery of a new class of antibiotics, as it resides exclusively in many pathogenic bacterial strains and is a key enzyme in the lysine biosynthetic pathway. This pathway is responsible for the production of lysine as well as meso-diaminopimelate (m-DAP), both of which are required for peptidoglycan cell-wall synthesis, and lysine for peptide synthesis. The enzyme DapE catalyzes the hydrolysis of N-succinyl-l,l-diaminopimelic acid (l,l-SDAP) to succinate and l,l-diaminopimelic acid (l,l-DAP), and due to its absence in humans, inhibition of DapE avoids mechanism-based side effects. We have executed the asymmetric synthesis of N,N-dimethyl-SDAP, an l,l-SDAP substrate analog and an analog of the synthetic substrate of our previously described DapE assay. Previous modeling studies advocated that N,N-dimethyl-SDAP might function as an inhibitor, however the compound behaves as a substrate, and we have demonstrated the use of N,N-dimethyl-SDAP as the substrate in a modified ninhydrin-based DapE assay. Thermal shift experiments of DapE in the presence of N,N-dimethyl-SDAP are consistent with a melt temperature (Tm) shifted by succinate, the product of enzymatic hydrolysis.

Core collapse supernovae are thought to be one of the main sources in the galaxy of elements heavier than iron. Understanding the origin of the elements is thus tightly linked to our understanding of the explosion mechanism of supernovae and supernova nucleosynthesis. X-ray and gamma-ray observations of young supernova remnants, combined with improved theoretical modeling, have resulted in enormous improvements in our knowledge of these events. The isotope Ti-44 is one of the most sensitive probes of the innermost regions of the core collapse engine, and its spatial and velocity distribution are key observables. Hard x-ray imaging spectroscopy with the Nuclear Spectroscopic Telescope Array (NuSTAR) has provided new insights into the structure of the supernova remnant Cassiopeia A (Cas A), establishing the convective nature of the supernova engine. However, many questions about the details of this engine remain. We present here the concept for a balloon-borne follow-up mission called A SuperConducting ENergetic x-ray Telescope (ASCENT). ASCENT uses transition edge sensor gamma-ray microcalorimeter detectors with a demonstrated 55-eV full-width half maximum energy resolution at 97 keV. This 8- to 16-fold improvement in energy resolution over NuSTAR will allow for high-resolution imaging and spectroscopy of the Ti-44 emission. This will allow for a detailed reconstruction of gamma-ray line redshifts, widths, and shapes, allowing us to address questions such as, What is the source of the neutron star kicks? What is the dominant production pathway for Ti-44? Is the engine of Cas A unique?