The global surge in antimicrobial resistance presents a critical threat to public health, emphasizing the urgent need for the development of new and more effective bacterial vaccines. Since the success of mRNA vaccines during the COVID-19 pandemic, this vaccine strategy has rapidly advanced, with most efforts focused on cancer immunotherapy and targeting viral pathogens. Recently, mRNA vaccines have entered the early phases of clinical development for bacterial diseases. However, bacteria present greater biological complexity compared with viruses, posing additional challenges for vaccine design, such as antigen selection, immune response and mRNA construct design. Here, we discuss critical aspects in the development of bacterial mRNA vaccines, from antigen selection to construct design. We also highlight the current preclinical landscape and discuss remaining translational challenges and future potential for mRNA vaccines against bacterial infections.

Recent CDC guidelines for treating systemic anthrax recommend a combination of a Tetracycline (Minocycline or Doxycycline) with Meropenem or a Fluoroquinolone. These recommendations differ from the 2001 or 2014 updates, and in the absence of significant clinical data, rely mainly on animal studies. Herein we present high efficacy of treating late-stage systemic anthrax with Doxycycline as a monotherapy or in combination with Levofloxacin in NHP.

The relation between the variability of the mid and upper tropospheric levels and the boundary layer profile is investigated by the aid of self-organising maps (SOM) clustering. For this purpose, two separate analyses of synoptic parameters and boundary layer profile are devised. This is the first comprehensive study to reveal this relation over the eastern Mediterranean region. The variability of the synoptic regimes is first analysed by the aid of potential vorticity (PV) at 320 K isentropic level and geopotential height at 1000 hPa (GPH1000). Projection against previous boundary layer profile SOM clustering shows the ability to infer the boundary layer profile from synoptic data according to the vertical coupling along the troposphere. Multi-variable SOM clustering of PV and GPH1000 presents 20 clusters which follow the variability of surface winter lows, highs, and Red Sea troughs. The active Red Sea trough relates to the north easterly and relatively narrow PV stream. Clusters presenting strong winter highs or lows with strong PV gradients have the highest persistence and directly indicate specific boundary layer profile patterns due to strong coupling along the troposphere. Under shallow GPH1000 gradients, when the dominating processes in the BL are local and the synoptic contribution is minimal and/or rapidly changing, indication of the boundary layer profile pattern is less predictable. This clustering framework offers further studies of the synoptic regimes dynamics, particularly the vertical interactions between the boundary layer and the free troposphere under pronounced surface pressure and PV gradients. Moreover, understanding these interactions has implications for the dispersion of anthropogenic pollutants across the eastern Mediterranean region.

Metabolite identification remains a significant challenge in mass spectrometry (MS)-based metabolomics research. To address this issue, we combined a triple-labeled precursor-based isotope tracing approach (TLEMMA) with high-resolution liquid chromatography-MS for metabolite identification and metabolic network construction. As a demonstration, we fed duckweed (Spirodela polyrhiza) with four forms of phenylalanine (Phe) including unlabeled Phe, Phe-⁵H₂, Phe-⁸H₂, and Phe-¹³C₉¹⁵N₁. The distinctive isotopic pattern obtained from MS spectra greatly facilitated data processing, enabling comprehensive extraction of all Phe-derived metabolites. Importantly, the labeling pattern allowed efficient metabolite identification by significantly reducing the number of structural and positional isomers. Using this approach, 47 phenylalanine-derived metabolites were putatively identified. To further evaluate the efficiency of metabolite identification in relation to the number of differently labeled precursors used, we compared the number of filtered candidates based solely on the labeling patterns obtained from unlabeled, single, dual, and triple isotope-labeled precursor tracing experiments. On average, TLEMMA eliminates the number of false candidates by 99.1% compared with unlabeled samples, 95% compared with single isotope-labeled samples, and 66.7% compared with dual isotope-labeled samples. This significant reduction in the number of false positives, along with the ability to identify previously unreported metabolites, demonstrates the power of TLEMMA in advancing the field of metabolomics and metabolic network reconstruction.

Purpose

The dynamic course of sulfur mustard (SM)-induced ocular insult involves an acute phase, which may progress to a chronic phase or a quiescent period, followed by late pathology. Visualizing pathological corneal changes in vivo could enhance understanding of this process and aid treatment development.

Methods

SM burn was induced in the right eyes of three transgenic mouse strains—expressing RFP under the VE-Cadherin promoter (blood vessels and hematopoietic cells), GFP under the keratin 15 promoter (limbal stem cells), and YFP under the Thy-1 promoter (mid-stromal nerve fibers, MSNFs)—by vapor exposure. Cell infiltration, neovascularization (NV), innervation loss, and stem cell (SC) depletion were monitored in vivo by stereomicroscopy for up to 8 weeks. Corneal whole-mounts were used to assess 360° structures, infiltrating cells, and subbasal nerve plexus (SNP) loss. Histology included H&E, Masson-Trichrome, and periodic acid-Schiff staining.

Results

A 35-s exposure caused minor ocular insult with moderate SNP changes, corneal cell infiltration, and reversible SC loss, mostly resolving by 4 weeks. A 120-s exposure caused severe insult with NV, extensive MSNF and SNP loss, marked CD45⁺ and Iba1⁺ infiltration, and irreversible SC depletion. NV, stromal inflammation, edema, epithelial changes, and goblet cells were seen in histology and correlated with fluorescence imaging.

Conclusions

This study demonstrates the utility of transgenic mice as powerful models for studying SM-induced ocular injury and for developing novel therapeutic strategies.

RNA-based therapeutics have revolutionized precision medicine due to their unprecedented potency, specificity, and adaptability. However, the inherent limited stability of RNA, including mRNA used in vaccines, is a major obstacle to the full realization of their potential. This instability, coupled with the centralized nature of vaccine production, currently limits the generation of RNA therapeutics at the point of care, which will otherwise fully harness the potential of these agents. Here, a microfluidic platform is presented for on-demand, personalized synthesis of modified mRNA stabilized by lipid nanoparticles. The design includes trapped biotinylated DNA, tagged T7 RNA polymerase, and a Tesla mixer, allowing the on-chip synthesis, purification, and encapsulation of mRNA in uniform lipid nanoparticles (LNPs), all conducted seamlessly on the same microfluidic device. This on-chip microfluidic synthesis approach is found to match standardized mRNA production yields, yet surpasses typical purification methods. Furthermore, as a proof-of-concept, the versatility and efficacy of the platform are demonstrated by generating diverse RNA sequences and structures, exhibiting functionality in human cell lines and mouse models. Moreover, an active SARS-CoV-2 vaccine is successfully engineered, highlighting the platform's potential for personalized vaccination strategies and offering a promising avenue for high throughput, decentralized vaccine delivery, reduced cold chain dependence, and even advancing current personalized medicine approaches through custom RNA therapeutics.

Regulatory T cells (T_regs) maintain immune homeostasis and their adoptive transfer is being widely explored to mitigate inflammatory and autoimmune conditions. Here a biomaterial is developed to accumulate T_regs at a specific anatomic location to bypass the need for ex vivo T_reg isolation and adoptive transfer. It is first shown that eliglustat, an FDA-approved inhibitor of UDP-glucose ceramide glucosyltransferase, promotes T_regs from both naïve and activated CD4⁺ T cells in vitro. Click-crosslinked cryogels fabricated from alginate and collagen allow for a sustained release of CXCL10 or CXCL11, and when injected in subcutaneous tissues led to the enrichment of effector and memory T cells to the scaffolds. Loading eliglustat into these cryogels significantly enhances the local accumulation of T_regs in vivo. These findings demonstrate that eliglustat-loaded cryogels offer a simple yet effective biomaterial strategy to boost T_reg directly in vivo, potentially providing a targeted method to treat various inflammatory and autoimmune diseases.

Transplantation of engineered B cells has shown efficacy in HIV disease models. B cell engineering may also be utilized for treatment of cancer. Recent studies highlighted that the activity of B cells is associated with favorable clinical outcomes in oncology. In mice, polyclonal B cells were shown to elicit anti-cancer responses. As a potential novel cell therapy, we demonstrate that engineering B cells to target a tumor-associated antigen potentiates polyclonal anti-tumor responses. We observe that engineered B cells expressing an anti-HPV B cell receptor internalize the antigen to allow subsequent activation of oncoantigen-specific T cells. Secreted antibodies from engineered B cells form immune complexes which are taken up by antigen-presenting cells to further promote T cell activation. Engineered B cells hold potential as novel, multi-modal cell therapies and open new avenues in solid tumor targeting.

Organophosphorus (OP) pesticides are a class of chemicals that are extensively used worldwide. The exposure to and use of organophosphates can be assessed by analyzing their metabolites and degradation products, such as dialkyl phosphate (DAP), dialkyl thiophosphate (DATP), and dialkyl dithiophosphate (DADTP). However, since these metabolites/hydrolysis products can result from the metabolism or breakdown of several organophosphorus pesticide families, they serve as nonspecific biomarkers and do not indicate the specific pesticide involved in exposure. In an earlier study, chemical derivatization using N-(2-(bromomethyl)benzyl)-N,N-diethylethanaminium bromide (CAX-B) was described to improve the signal intensity of numerous organophosphorus (OP) acids in liquid chromatography tandem mass spectrometry (LC-ESI-MS/MS) analysis. In the present study, CAX-B was employed to derivatize a set of seven phenolic compounds corresponding to the complementary portion of OP pesticides. The derivatization process using CAX-B was performed in acetonitrile with potassium carbonate at 50 °C for 30 min. LC-Orbitrap-ESI-MS/MS was used to analyze the resulting phenol derivatives and their fragmentation patterns were studied. Notably, the derivatized phenols were markedly more sensitive than the underivatized phenols when LC-ESI-MS/MS was used in MRM technique, without being affected by the sample matrix (soil or plant extracts). This derivatization technique aids in identifying OP pesticides, offers insights into their subfamily, and pinpoints a specific compound through the analysis of corresponding phenol derivative.

Ricin is a highly potent toxin that causes severe lung injury upon inhalation by initiating a complex cascade of cellular responses that ultimately leads to cell death. The low-density lipoprotein receptor-related protein 1 (LRP1) is a multifunctional receptor involved in various physiological processes, including ricin-mediated toxicity. This study explores the role of LRP1 shedding in the development of ricin-induced lung injury. Analysis of bronchoalveolar lavage fluid (BALF) from ricin-intoxicated mice and swine showed a significant increase in soluble LRP1 (sLRP1) levels, whereas serum LRP1 levels remained largely unchanged, suggesting the lungs are the primary source of sLRP1 release. In vitro assays demonstrated the formation of ricin-sLRP1 complexes, indicating that sLRP1 in BALF retained ricin-binding capability. Flow cytometric analysis of lung cells revealed a reduction in both the percentage and total number of LRP1-expressing cells following ricin exposure. Further investigation of specific lung cell populations showed that alveolar epithelial type II (AT-II) cells, despite experiencing significant injury, exhibited minimal LRP1 shedding. No shedding of LRP1 occurred in neutrophils. In contrast, fibroblasts, which were resistant to ricin-induced cell death, exhibited increased shedding of LRP1 and a corresponding decrease in membrane-bound LRP1 expression. This shedding of the LRP1 ectodomain was mediated by metalloproteinases. Immunohistochemical staining further confirmed decreased LRP1 expression in fibroblasts from ricin-exposed mice. Macrophages also showed substantial LRP1 shedding, despite undergoing significant depletion. These findings highlight the complex cell-specific nature of LRP1 shedding in response to ricin intoxication and suggests the potential role of LRP1 in modulation of cellular susceptibility and resistance to ricin-induced lung injury.