Johnson, Stephen

Background: Staphylococcus aureus causes bloodstream and skin infections in humans. The prevalence of multidrug-resistant S. aureus strains means new antibiotics are needed. A novel antimicrobial drug named SK-03-92, a synthetic aromatic organic stilbenoid compound, kills S. aureus cells within 30 min, but an increase in both biofilm formation and persister cells occurs. SK-03-92 treatment downregulates transcription of the biofilm regulating protein regulator (brpR) gene and biofilm regulating protein sensor (brpS) gene in S. aureus. BrpR/BrpS system may be a LytTR regulatory system tied to biofilm formation, creation of persister cells, and late-stage competence in S. aureus. The aim of this study was to determine what biofilm, late-stage competence, and persister-associated genes were regulated in a brpR mutant compared to wild-type strains. Methods: In this study, involvement of BrpR in regulating other genes was assessed by comparing transcriptional changes in a brpR mutant strain to the S. aureus parent strain via RNA sequencing (RNA-Seq). Bioinformatic analysis was then performed on the RNA-Seq data to assess what biochemical pathways might be involved. Results: From these analyses, 440 genes were identified that had significant differences in transcript abundance when comparing the brpR mutant to wild-type strains. Quantitative reverse transcription polymerase chain reaction analysis confirmed bacA, icd, metE, and pdhA transcript levels were lower, whereas alr and mraY were higher in the brpR mutant versus wild-type strain. Furthermore, an enzymatic assay targeting NADH production from the pyruvate dehydrogenase complex showed lower levels in the mutant compared to wild-type strain. Conclusions: Overall, the study demonstrated several biosynthetic pathways tied to biofilm formation and late-stage competency may be regulated by BrpR and some potential leads for the mechanism of action of the SK-03-92 drug were uncovered.

(1) Background: Mammographic breast density (MBD) is a well-established predictor of breast cancer risk, yet the biological mechanisms underlying this association remain incompletely understood. MBD is characterized by alterations in breast stromal architecture, including increased collagen deposition and changes in immune cell composition. Given emerging evidence that the breast harbors a resident microbiome, we investigated whether the breast tissue microbiome correlates with MBD. (2) Methods: Adjacent normal breast tissue was collected under sterile conditions from 33 women undergoing surgery for benign or malignant breast disease. DNA was extracted and subjected to 16S rRNA gene sequencing (Illumina MiSeq). (3) Results: We observed a non-significant trend toward lower α-diversity in high-MBD samples compared to low-MBD samples, p = 0.13. β-Diversity analyses identified a modest association between MBD and microbial community composition (MiRKAT p = 0.049). A random forest-based model incorporating genus-level relative abundances improved prediction of MBD over clinical characteristics alone, identifying Corynebacterium (Actinobacteria) and other genera as key predictors. (4) Conclusions: Breast tissue microbial features vary with mammographic breast density, suggesting a potential association with density-associated breast cancer risk. These exploratory findings warrant validation in larger cohorts to better elucidate biological mechanisms and clinical relevance.

ABSTRACT Aims To investigate whether the association between the nitrite‐generating capacity of the subgingival microbiome and early cardiometabolic risk biomarkers varies by dietary nitrate intake. Materials and Methods Cross‐sectional data from 668 participants (mean age 31 ± 9 years, 73% women) were analysed. Dietary nitrate intake was calculated from food frequency questionnaires. Subgingival 16S rRNA sequencing (Illumina, MiSeq) and PICRUSt2 estimated microbial genes. The Microbiome‐Induced Nitric Oxide Enrichment Score (MINES) was calculated as a ratio of microbial gene abundances representing enhanced net capacity for NO generation. Adjusted multivariable linear models regressed cardiometabolic risk biomarkers (HbA1c, glucose, insulin, insulin resistance (HOMA‐IR), blood pressure) on nitrate intake and MINES together with a MINES × nitrate intake interaction term. Results Mean nitrate intake was 190 ± 171 mg/day. Significant interactions of MINES and nitrate intake were observed for insulin and HOMA‐IR ( p < 0.05). Among participants with a low MINES, higher nitrate intake was associated with lower HOMA‐IR (1.2 [1.1–1.4] vs. 1.5 [1.3–1.6]; p = 0.002), but levels were similar in those with high MINES ( p = 0.84). Conclusions A biomarker of higher microbial NO‐generating capacity in subgingival plaque is associated with lower insulin and insulin resistance among individuals with lower dietary nitrate intake. Future trials evaluating the cardiometabolic benefits of nitrate‐rich diets should incorporate measures of the entire oral microbiome.

Three previously uncharacterized bacterial species, designated strains BD586 T , BD613 T and BD626 T , isolated from human clinical specimens, were identified at Mayo Clinic, Rochester, Minnesota, USA. Initial identification efforts using matrix-assisted laser desorption ionization-time-of-flight MS and partial 16S rRNA gene sequencing proved inconclusive. Comprehensive analysis involving phenotypic characterization, biochemical assays and whole-genome sequencing was undertaken. The isolates were Gram-negative, motile, facultatively anaerobic rods, occurring singly, in pairs and in short chains; BD613 T additionally formed small aggregates. The isolates tested positive for catalase and negative for oxidase. Their colonies appeared smooth, white, opaque and non-haemolytic. Growth was observed at 35 °C under aerobic, anaerobic and CO₂-enriched conditions, as well as in media with NaCl concentrations up to 10% and at pH 7-9. Phylogenetic relationships were inferred from core gene alignments, average nucleotide identity and digital DNA–DNA hybridization comparisons. Results confirmed the placement of BD586 T , BD613 T and BD626 T within the recently established Dryocola genus, while also indicating their novelty as distinct species. The major cellular fatty acids were C 16:0 and C 17:0 cyclo. Based on these findings, a formal description of three new species, Dryocola mayonis sp. nov. (type strain BD586 T =TSD 474 T , =NCTC 15089 T , =DSM 119465 T ), Dryocola sharpae sp. nov. (type strain BD613 T =TSD 475 T , =NCTC 15090 T , =DSM 119466 T ) and Dryocola baronae sp. nov. (type strain BD626 T =TSD 476 T , =NCTC 15091 T , =DSM 119479 T ), is proposed, and the genus description of Dryocola is emended to refine its genomic, morphological and chemotaxonomic boundaries.

Background Neoadjuvant treatment has become standard for patients with high-risk operable stage III melanoma, but the optimal regimen is unknown. Targeted therapy approaches yield high pathological response rates, while immunotherapy regimens show favorable recurrence-free survival (RFS). NeoACTIVATE was designed to address whether a neoadjuvant combination of both targeted therapy and immunotherapy might leverage the benefits of each. Methods We tested neoadjuvant treatment with 12 weeks of vemurafenib, cobimetinib, and atezolizumab for patients with BRAF-mutated (BRAFm) melanoma (cohort A) and cobimetinib and atezolizumab for patients with BRAF-wild-type (BRAFwt) melanoma (cohort B), regimens which we have shown generate a substantial major pathological response. After therapeutic lymph node dissection, patients received 24 weeks of adjuvant atezolizumab. Here, we report survival outcomes and their association with biomarkers assayed among the gut microbiome and peripheral blood immune subsets. Results With 49 months median follow-up, the median RFS was not reached for cohort A and was 40.8 months for cohort B. At 24 months after operation, 2 of 14 cohort A patients and 4 of 13 cohort B patients had experienced distant relapse. Key findings from correlative analyses included diversity, taxonomic and functional metagenomic gut microbiome signals associated with distant metastasis-free survival at 2 years. Notably, we observed a strong correlation between low microbial arginine biosynthesis (required for T-cell activation and effector function) and early distant recurrence (p=0.0005), which correlated with taxonomic differential abundance findings. Peripheral blood immune monitoring revealed increased double-positive (CD4+CD8+) T cells in patients with early recurrence. Conclusions Neoadjuvant treatment with cobimetinib and atezolizumab±vemurafenib was associated with a low rate of distant metastasis in patients with high-risk stage III melanoma. Freedom from early distant metastasis was highly associated with taxonomic differences in gut microbiome structure and with functional pathway alterations known to modulate T cell immunity. Identification of predictive biomarkers will permit optimization of neoadjuvant therapy regimens for individual patients. Trial registration number NCT03554083.