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Schubert, S.

Publications and source records attributed to Schubert, S..

7 recordsLinked to original sources

Cycles of contamination and recovery: Combined sewer overflows drive acute but transient antimicrobial resistance exposure in an urban stream

Combined sewer overflows (CSOs) are a major pathway for untreated wastewater into urban streams, yet their role in shaping antimicrobial resistance (AMR) dynamics remains poorly understood. Here, we used high-frequency, time-resolved sampling during two storm-triggered CSO events across two monitoring locations and one stormwater-only control site in an urban stream to quantify how these disturbances affect microbial communities, antibiotic resistance genes (ARGs), and mobile genetic elements (MGEs) in an urban stream. CSO events caused rapid, up to two orders of magnitude, increases in bacterial, pathogen, and ARG abundance, with multiple transient peaks occurring within single overflow episodes. However, these increases were largely proportional to the total bacterial load, and most ARGs and MGEs did not change in relative abundance, indicating that CSOs primarily act as mass-transfer events rather than drivers of in situ selection. Downstream attenuation was governed by hydrological dilution despite additional CSO inputs: Both microbial and resistance signals largely returned to baseline within short time frames. This demonstrates that CSOs function as hydrologically driven pulse disturbances that generate acute but transient AMR exposure. Because CSO events lack the sustained pressure associated with continuous wastewater discharges, rapid washout prevents the long-term establishment of sewage-derived resistance. These findings highlight that AMR risk in CSO-impacted systems is driven primarily by short-term exposure rather than by persistent ecological transformation, with important implications for urban water management under increasingly extreme rainfall conditions.

microbiology↗

Lineage-associated tumor cell states predict outcome and guide immunotherapeutic target selection in pediatric osteosarcoma

Osteosarcoma is a highly heterogeneous primary bone malignancy that predominantly affects adolescents and young adults, and robust prognostic biomarkers and effective targeted therapies remain lacking. While single-cell transcriptomics has begun to resolve tumor composition, a tumor cell-intrinsic framework linking transcriptional states to clinical outcome and therapeutic targeting is not established. Here, we present a single-cell transcriptomic analysis of pediatric osteosarcoma, integrating 22 samples across 18 patients spanning six diagnostic, six post-treatment, and ten metastatic lesions profiled using complementary sequencing platforms. Using non-negative matrix factorization, we identify recurrent lineage-associated transcriptional programs that define osteoblastic-like, chondroblastic-like, and fibroblastic-like tumor cell states. These programs show overlapping activity across tumor cells, indicating that tumor cell states are not strictly discrete. Strikingly, combined osteoblastic-like and chondroblastic-like program activity is associated with poorer overall survival, a finding validated across two independent bulk RNA-sequencing cohorts. These tumor cell states are further linked to distinct immune microenvironmental compositions. Mapping candidate immunotherapeutic targets at single-cell resolution reveals variable expression across tumor cell states, supporting state-informed strategies for CAR T cell targeting. Together, our findings establish a tumor cell state framework that links osteosarcoma heterogeneity to clinical outcome and provides a basis for patient stratification and the development of state-informed therapeutic strategies in pediatric osteosarcoma.

Cancer Biology↗

Exploring lipid nanoparticle design spaces using self-regulating microfluidic machines and multiplexed in vivo biodistribution

Delivering therapeutic mRNA relies on lipid nanoparticles (LNPs). Finding optimal process parameters for new lipid combinations in LNP formulations remains a challenge. In our work, we used an automated, self-regulating microfluidic platform that actively changes process parameters to tune LNP formulations for preset, desired quality standards. We tested new LNPs by swapping poly(ethylene glycol) (PEG) lipopolymers for alternatives based on poly(2-methyl-2-oxazoline) (PMeOx) and poly(2-ethyl-2-oxazoline) (PEtOx). For each lipopolymer variant, the platform independently identified optimal production conditions in four or fewer iterative cycles, yielding particles of the preset size and high mRNA encapsulation. Small-angle X-ray scattering revealed that smaller LNPs modified with PEtOx had more structural surface variety and higher mRNA loading efficiency. When multiplexing these formulations in mice, the PEtOx-containing LNPs accumulated more in bone marrow compared to those with PEG, indicating trends that the chemistry of the lipopolymer affects the biodistribution of the resulting LNPs. By combining automated formulation and in vivo multiplexed testing, our approach provides a practical way to rapidly plan, formulate, and evaluate large pharmaceutical design spaces, to select excipients and process parameters yielding optimal biological performance of LNPs. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=76 SRC="FIGDIR/small/729611v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1a08b68org.highwire.dtl.DTLVardef@120724corg.highwire.dtl.DTLVardef@12fed98org.highwire.dtl.DTLVardef@197aa41_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

Relationships of crystallinity and reaction rates for enzymatic degradation of poly (ethylene terephthalate), PET

Biocatalytic degradation of plastic waste is anticipated to play an important role in future recycling systems. However, enzymatic degradation of crystalline poly (ethylene terephthalate) (PET) remains consistently poor. Herein, we employed functional assays to elucidate the molecular underpinnings of this limitation. This included utilizing complementary activity assays to monitor the degradation of PET disks with varying crystallinity (XC), as well as kinetic parameters for soluble PET fragments. The results indicate that a proficient PET-hydrolase, LCCICCG, operates through an endolytic mode of action, and that its activity is limited by conformational constraints in the PET polymer. Such constraints become more pronounced at high XC values, and this limits the density of productive sites on the PET surface. Endolytic chain-scissions are the dominant reaction type in the initial stage, and this means that little or no soluble organic product occurs here. However, endolytic cuts gradually and locally promote chain mobility and hence the density of attack sites on the surface. This leads to an upward concave progress curve; a behavior sometimes termed lag-phase kinetics.

biochemistry↗

Hepatoblastoma exhibits a predominantly myeloid immune landscape and reveals opportunities for macrophage targeted immunotherapy

Background & AimsHepatoblastoma (HB) is a rare form of pediatric liver cancer which is currently treated with chemotherapy and surgery. The side effects of chemotherapy pose a major problem in HB and underline the need for an alternative treatment option. We aimed to characterize the immune landscape of HB to improve our understanding of the immunologic contribution to this disease and explore immunotherapeutic options. MethodsAn imaging mass cytometry panel of 36 antibodies was used on tissue of treatment-naive HB (n=5), and chemotherapy-treated HB (n=3), with paired distal normal liver tissue. Immunofluorescence was used to stain HB and normal liver tissue for Kupffer cell marker MARCO. A public single-cell RNA-sequencing (scRNA-seq) dataset was analyzed consisting of 9 chemotherapy-treated HB and paired normal liver tissue. ResultsHB showed a heterogeneous immune landscape predominantly comprising macrophages and monocytes with high expression of immune checkpoints CD47, SIRP, and VISTA, whereas T cells were limited. Chemotherapy increased influx of macrophages and CD8+ T cells in HB. Transcriptome profiling demonstrated an early activated phenotype of CD8+ T cells in chemotherapy-treated HB and absence of an exhaustion signature and immune checkpoint expression. Furthermore, tumor-associated macrophages had low MARCO expression, upregulated inflammatory markers and a high liver tissue residency score while expressing other Kupffer cell markers, such as CD5L, to a variable degree. ConclusionsThe absence of immune checkpoints and exhaustion markers in CD8+ T cells prohibits T cell-targeting by immune checkpoint blockade in HB patients. Instead, HB tumors contain a large myeloid compartment which provide opportunities for macrophage targeting, thereby paving the way for the development of improved treatment strategies for HB patients. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/546852v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@3748e2org.highwire.dtl.DTLVardef@191f9d9org.highwire.dtl.DTLVardef@aa4172org.highwire.dtl.DTLVardef@1aed5f9_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

Reaction pathways for the enzymatic degradation of poly(ethylene terephthalate): What characterizes an efficient PET-hydrolase?

Bioprocessing of polyester waste has emerged as a promising tool in the quest for a cyclic plastic economy. One key step is the enzymatic breakdown of the polymer, and this entails a complicated pathway with substrates, intermediates, and products of variable size and solubility. We have elucidated this pathway for poly(ethylene terephthalate) (PET) and four enzymes. Specifically, we combined different kinetic measurements and a novel stochastic model, and found that the ability to hydrolyze internal bonds in the polymer (endo-lytic activity) was a key parameter for overall enzyme performance. Endo-lytic activity promoted the release of soluble PET fragments with two or three aromatic rings, which, in turn, were broken down with remarkable efficiency (kcat/KM-values of about 105 M-1s-1) in the aqueous bulk. This meant that about 70% of the final, monoaromatic products was formed via soluble di-or tri-aromatic intermediates.

biochemistry↗

Major role of the high-pathogenicity island (HPI) in the intrinsic extra-intestinal virulence of Escherichia coli revealed by a genome-wide association study

The genus Escherichia is composed of several species and cryptic clades, including E. coli, which behave as a vertebrate gut commensal, but also as an opportunistic pathogen involved in both diarrheic and extra-intestinal diseases. To characterize the genetic determinants of extra-intestinal virulence within the genus, we carried out an unbiased genome-wide association study (GWAS) on 370 commensal, pathogenic and environmental strains representative of the Escherichia genus phylogenetic diversity and including E. albertii (n=7), E. fergusonii (n=5), Escherichia clades (n=32) and E. coli (n=326), tested in a mouse model of sepsis. We found that the high-pathogenicity island (HPI), a [~]35 kbp gene island encoding the yersiniabactin siderophore, is highly associated with death in mice, surpassing other associated genetic factors also related to iron uptake, such as the aerobactin and the sitABCD operons. We validated the association in vivo by deleting key components of the HPI in E. coli strains in two phylogenetic backgrounds, and found that virulence is correlated in E. coli with growth in the presence of various stressors including several antimicrobials, which hints at collateral sensitivities associated with intrinsic virulence. This study points to the major role of iron capture systems in the extra-intestinal virulence of the genus Escherichia and the collateral effects on cell growth of such systems.

microbiology↗