Introduction: The global spread of antimicrobial resistance (AMR) genes (ARGs) poses a major challenge to bacterial infection treatment. Antibiotic use, while targeting pathogens, also disrupts commensal gut bacteria, with effects varying by antibiotic type. The impact of gut dysbiosis on ARG mobility from foodborne bacteria, especially with or without food matrix microbes, remains unclear.

Methods: Using a murine model, we investigated ARG dissemination via mobile AMR plasmids. Mice were pre-treated with streptomycin, ampicillin, or sulfamethazine to induce varying levels of gut dysbiosis. They were then inoculated with beta-lactam-resistant Salmonella Heidelberg (donor) and beta-lactam-susceptible Salmonella Typhimurium (recipient) with or without additional food matrix microbes. Fecal samples were cultured to detect ARG transfer among Salmonella, E. coli, and other gut bacteria, which were confirmed using whole-genome sequencing. Changes in gut microbiota were assessed using 16S rRNA sequencing.

Results: Without background food matrix microbes, streptomycin caused severe gut dysbiosis, enhancing AMR plasmid transfer from S. Heidelberg to S. Typhimurium and E. coli. Ampicillin induced moderate dysbiosis, allowing S. Heidelberg colonization and plasmid transfer to E. coli. Sulfamethazine caused mild dysbiosis, hindering both Salmonella colonization and plasmid transfer. In contrast, food matrix microbes reduced AMR plasmid transfer, except in streptomycin pre-treated mice, where Enterobacteriaceae enrichment enabled AMR plasmid transfer to Escherichia, Enterobacter, Citrobacter, and Proteus.

Conclusions: Pre-existing gut microbiome disturbances from antibiotics significantly affect ARG dissemination. These findings support more judicious antibiotic use to mitigate resistance spread.

Medicinal plants are one of the groups of economically important species and a valuable natural resource. One of these is thyme (Thymus vulgaris L.), the source of essential oil with high antimicrobial activity against microorganisms from various systematic groups. The study of thyme–microbial interactions was carried out in 2014-2024 for the collection of medicinal plants of the V. I. Edelstein Vegetable Experimental Station by cultural, chromatography, and NGS methods. The data obtained indicated the importance of the phytosphere's physiological and biochemical features for microbial community structure and representation depending on the development stage, since these factors significantly affected the plant–microbial interaction in the thyme–soil system. In other words, the features of the studied plant–microbiological interaction depended on which microorganisms inhabited specific parts of the thyme phytosphere and at what stage of plant development this occurred. Moreover, for bacterial communities, the key roles were played by the features of the phenological stage of the plant (54.3%) and the colonized part (44.7%), while for fungi, the interaction of these two factors was the most significant (75.8%). For microbial biotechnologies, it is necessary to obtain cultured identified forms of microorganisms. So, typical representatives of microbial communities were isolated and sequenced. At least nine phytosphere microorganisms can be recommended as protective and growth-regulating agents for thyme cultivation in the Non-Chernozem zone due to the suppression of root rot pathogens that actively develop during thaws and high humidity conditions.

The rise in antimicrobial resistance underscores the urgent need for novel antimicrobial compounds. Bacteria, including Clostridium botulinum, offer a promising natural reservoir for such discoveries. While C. botulinum is best known for its neurotoxin production, its potential as a source of antimicrobial compounds is largely unexplored. This study integrates next-generation sequencing (NGS), bioinformatics, and antimicrobial assays to identify novel antimicrobial compounds, focusing on bacteriocins—ribosomally synthesized and post-translationally modified peptides (RiPPs) targeting closely related species. The nontoxic C. botulinum HA5 strain exhibited antimicrobial activity against clostridial species, including toxigenic C. botulinum, C. difficile, and C. perfringens. Bioinformatics revealed multiple biosynthetic gene clusters (BGCs). Knockout experiments identified a spontaneous mutant lacking antimicrobial activity due to the loss of a plasmid (pHA5) encoding the botocidin biosynthetic gene cluster (a ranthipeptide or sactipeptide). This confirmed the role of botocidin in antimicrobial activity. The botocidin BGC comprised two operons: one encoding the core peptide, modification enzymes, an ABC transporter, and proteases, and the other, oriented oppositely, encoding an MFS transporter and a potential modification enzyme. The Spo0A transcription factor regulated the production operon, while the MFS transporter conferred immunity without affecting production. Botocidin production was confirmed via TCA precipitation and LC-MS, revealing peptide masses consistent with the predicted compound. Stability tests showed botocidin's heat stability (1 h at 90 °C), pH stability (pH 4–9), moderate UV sensitivity, and protease sensitivity. Time-lapse microscopy revealed bactericidal activity, causing cell lysis by targeting the membrane, as confirmed by propidium iodide staining. While botocidin did not inhibit spore germination, it blocked spore outgrowth sporestatically, as shown by plate counts and microscopy. These findings identify botocidin as the active antimicrobial in C. botulinum HA5, highlighting its potential to address antimicrobial resistance.

High salinity levels in wastewater treatment plants (WWTPs) can be attributed to multiple sources, including seawater intrusion in wastewater streams. In coastal WWTPs, this phenomenon is becoming more frequent, causing transient salinity shocks on the microbial populations involved in the treatment process. Aerobic granular sludge (AGS) has emerged as a revolutionary technology that has been adopted worldwide for treating several types of wastewater. Much of its success is related to its great tolerance to extreme environments, including high-saline wastewater. In this study, a laboratory-scale AGS reactor was exposed to different salinity stresses over 286 days. First, over 131 days, the seawater content in wastewater was gradually increased in the feeding regime (1.5 – 15 g/L). For the remainder of the operation, the AGS had to deal with daily salinity oscillations, ranging from high (7.5 g/L) to very high (22.5 g/L) seawater levels in wastewater. Throughout the operation, the removal performance of organic carbon, ammonium, and phosphate was consistently effective, despite the daily fluctuations in the seawater content of the wastewater. This was likely ensured by the nutrient removal-related taxa present in the AGS core microbiome, which was highly diverse and resilient to changes in wastewater composition. Over time, enrichment of the core microbiome with halotolerant taxa and extracellular polymeric substance producers proved crucial for maintaining the integrity and stability of the reactor’s performance. The findings of this work underscore the flexibility and robustness of AGS communities in thriving under diverse environmental challenges and adapting to sustain AGS reactor performance.

Acknowledgments: The authors thank the CBQF scientific collaboration under the FCT project UIDB/50016/2020. C. Miranda thanks the research grant from FCT, Portugal (doi.org/10.54499/2020.06577.BD), and POCH, supported by the European Social Fund and the MCTES national funds.

Human herpesvirus 6 (HHV-6), a widely distributed member of the Herpesviridae family, establishes lifelong latency in monocytes and macrophages. The persistence of HHV-6 under immunosuppressive conditions, such as chronic alcohol consumption, may potentiate liver inflammation and exacerbate tissue damage by synergising with the hepatotoxic effects of ethanol. This interaction could raise significant concerns in the context of infectious diseases, as it poses a heightened risk to individuals with a history of heavy alcohol use, particularly those with underlying comorbidities or compromised immune function. The transcription factor NF-κB, essential for transactivating target genes involved in immune and inflammatory responses and CD163, expressed on anti-inflammatory macrophages, provides valuable insights into tissue damage during the presence of HHV- 6 infection.

Fifty-four liver tissue specimens were collected from the following groups: control (n=11), young alcohol users (n=15), and chronic alcohol users (n=28). Specimens were immunohistochemically stained with anti-CD163, anti-NF-κB, and anti-HHV-6 antibodies and analysed via light microscopy. HHV-6- and CD163-positive cells were counted quantitatively, while both the intensity and distribution of NF-κB expression were analysed semi-quantitatively. Statistical analysis was performed using SPSS 28.0.

HHV-6-positive liver lobules were identified in 48.75% of the controls, 63.89% of young alcohol users, and 72.04% of chronic alcohol users. The mean CD163-positive cell count in the lobular area increased significantly in young alcohol users (mean=135 ± 43 SEM) and chronic alcohol users (mean=226 ± 38 SEM) compared to the controls (mean=59 ± 14 SEM). NF-κB expression intensity in the lobular area was significantly higher in young alcohol users (p<0.005), and both intensity and distribution were notably increased in chronic alcoholics (p<0.001, p=0.02) compared to the controls.

Chronic alcohol consumption increases liver inflammation and damage, potentially exacerbated by HHV‑6 persistence. Further studies are needed to confirm these interactions and explore the mechanisms driving the synergistic effects of HHV-6 and ethanol on liver tissue damage.

Introduction: Type 2 diabetes mellitus (T2DM) is an age-related metabolic disease that is often considered inflammatory, as various cytokine profiles are associated with its progression. T2DM typically follows a prediabetic stage, during which insulin resistance develops. This stage can often be corrected early in life, potentially preventing or delaying the onset of T2D. Intestinal dysbiosis has been linked to metabolic disorders, including insulin resistance and T2D. A healthy, diverse gut microbiome, composed primarily of four phyla—Bacteroidetes, Firmicutes, Actinobacteria, and Proteobacteria—is essential for maintaining the gut epithelial barrier. Phylogenetically related bacterial groups, such as Proteobacteria and Enterobacteriaceae, have been linked to poor glycaemic control and negative metabolic outcomes, including obesity, insulin resistance, and an impaired lipid profile. Methods: This study involved 73 older adults (ages 76–83) from the randomized controlled trial entitled the Generation 100 Study. We performed high-throughput sequencing of the bacterial 16S rRNA gene to obtain metagenomic microbial profiles for all participants. These profiles were then correlated with clinical measures. Results: We observed distinct patterns of microbial beta diversity between the high- and normal-glucose groups. Overall, the microbial diversity was significantly reduced in the high-glucose group. At the highest taxonomic level (Phylum), we found that Synergistes, Elusimicobia, Euryarchaeota, Verrucomicrobia, and Proteobacteria were all significantly decreased in participants with high blood glucose. Additionally, P. copri was significantly elevated in the high-glucose (10-fold increase) and high-CRP (777-fold increase) groups, suggesting that it may serve as an early inflammatory and diabetic marker. These findings are consistent with previous research identifying P. copri as a pro-inflammatory pathogen. We also found that the Fusobacterium genus was significantly increased in the normal glucose group, with a 151-fold increase compared to the high-glucose group (p < 0.005). Conclusions: Our results indicate significant changes in the microbiome that may provide valuable insights for early intervention in pre-diabetic states.

Minced meat contains spoilage microorganisms, reducing shelf-life and causing economic and environmental impacts. It can also harbour pathogens that pose risks to public health, leading to foodborne illnesses and product recalls. The aim of this study was to evaluate the deteriorative and pathogenic microbiota of minced meat obtained in hypermarkets. Thirty samples of minced meat from bovine (n=15), swine (n=6), and poultry (n=9), produced from hypermarkets in Vila Real, were analysed for the presence of three important foodborne pathogenic bacteria, i.e., L. monocytogenes, S. aureus, and E. coli. In parallel, total aerobic microorganisms at 30ºC (mesophiles), Enterobacteriaceae, Lactic Acid Bacteria (LAB), and Pseudomonas spp., were also enumerated as hygiene / safety indicator organisms. Microbial concentrations (log10 cfu/g) of 6,01 ± 0,19 (total mesophiles), 3,79 ± 1,27 (Enterobacteriaceae), 4,14 ± 0,96 (LAB), 1,35 ± 1,31 (Pseudomonas spp.), 0,07 ± 0,29 (L. monocytogenes); 0,21 ± 0,66 (S. aureus), and 0,52 ± 0,81 (E. coli) were found. Significant differences in microbial counts were found only for total mesophiles and E. coli, with poultry showing higher levels than beef and pork (p < 0.05). Although poultry meat had the overall highest microbial counts, which is assocaited with its lower acceptability, followed by beef and pork minced meat,this difference was not statistically significant (p > 0.05). Enterobacteriaceae was the microorganism with the lowest acceptability, contributing to a reduction in the general product acceptability (83.3%) compared to the higher rates for other microorganisms. These findings emphasize the role of minced meat in the transmission of pathogenic and deteriorative microorganisms, highlighting the importance of proper handling and thorough cooking to prevent foodborne diseases.

Cultivated soybean is a vital source of protein and oil. Soybean obtains nitrogen (N) primarily from two sources: symbiotic nitrogen fixation (SNF) facilitated by microbes like Bradyrhizobium japonicum, and mineral nitrogen from soil or nitrate fertilizers. While nitrate fertilizers enhance plant growth, high nitrate levels inhibit nodule formation and SNF activity, reducing the proportion of nitrogen derived from SNF.

This study investigated the potential of nitrate-reducing bacteria (Bacillus subtilis and Priestia megaterium) to mitigate the nitrate-induced inhibition of SNF when co-inoculated with B. japonicum USDA 110 or USDA 6^T strains. Nitrogenase activity, nodulation, root elongation, and plant biomass were evaluated under hydroponic conditions at 1 mM and 10 mM nitrate levels, with nitrate assimilation analyzed using the ¹⁵N stable isotope. The growth responses of B. japonicum to varying nitrate concentrations were also examined. The growth of B. japonicum peaked at 7 mM nitrate but declined at 10 mM, suggesting inhibitory effects at higher concentrations.

Nitrogenase activity significantly decreased under high-nitrate conditions. However, co-inoculation with B. subtilis partially alleviated this inhibition, enhancing nitrogenase activity by 15.23% (USDA 110) and 72.62% (USDA 6^T) at 10 mM nitrate. Conversely, P. megaterium co-inoculation further reduced nitrogenase activity in some treatments. Nodulation was similarly inhibited by high nitrate levels but improved with the co-inoculation of B. subtilis under both nitrate conditions, while the effects of P. megaterium were strain-dependent. Root length and plant biomass responded positively to co-inoculation with both bacteria, although nitrate-induced reductions were observed in nodule and root N concentrations.

This study highlights the potential of B. subtilis to counteract the nitrate-induced inhibition of SNF and improve soybean growth under nitrogen-rich conditions. The findings provide insights into microbial interactions that could inform sustainable agricultural practices aimed at enhancing legume productivity in nitrogen-rich soils such as those found in intensively managed agricultural systems.

Introduction

There is growing evidence that cancer patients have altered metabolism due to disruption of the microbiome. Serum concentrations of aromatic microbial metabolites may reflect microbiota dysfunction.

Aim

The aim of this study was to evaluate the serum concentrations of aromatic microbial metabolites in patients undergoing treatment for malignant oncological diseases.

Methods

Two cohorts of patients were studied: patients with pancreatic cancer before surgery (n=64) and children with various malignant oncological diseases (leukemia, lymphoma, nephroblastoma, ependymoma, etc.) (n=40). The study used two comparison groups: healthy donors (n=18) and practically healthy children referred for preventive examination (n=18). Aromatic microbial metabolites such as phenyllactic acid, hydroxyphenyllactic acid and hydroxyphenylacetic acid were identified by GC-MS.

Results

The concentration of metabolites in the serum of patients with cancer in both studies is statistically significantly lower than that of healthy subjects. The sum of concentrations of the aromatic microbial metabolites was 1.9 (1.5; 2.2) µmol/L in healthy donors and 1.4 (0.9; 2.0) µmol/L in pancreatic cancer patients before surgery (p-value<0.001). The sum of concentrations of the aromatic microbial metabolites in children was 2.2 (1.5; 2.6) µmol/L in the control group vs 1.5 (1.1; 1.8) µmol/L in the cancer group (p-value=0.001).

Conclusions

Our study reveals the profound metabolic dysfunction of microbiota in cancer, as serum aromatic microbial metabolites were significantly different in patients compared to healthy subjects. It is promising to study the relationship between metabolic profiles and gut microbiota composition for a deep understanding of cancer pathogenesis.

Creating robust and long-lasting storage solutions presents a major challenge in developing microbial inoculants for eco-friendly farming. Well-developed techniques for anchoring bacteria onto solid supports are available, and growing interest is focusing on the implementation of sustainable carrier materials. This work aims to examine the utilization of cellulose functional fibers, chemically altered by the integration of natural polymers derived from the pulp industry. The fibers were assessed as a carrier for a consortium of plant growth-promoting bacteria. The consortium was immobilized on fibers culturing the strains in a combined fermentation with the addition of fibers (1% w/v) to allow for microbial self-adhesion to the surface. The fibers were added at three specific stages of the bioreaction (0, 24 hours, and 48 hours), utilizing two separate culture mediums. Desiccation was executed using freeze-drying and heat-drying techniques. Cell viability was assessed in never-dried functional fibers and dried fibers until one month from inoculation, after drying. Immobilized bacteria were also assessed for plant growth-promoting (PGP) traits, encompassing indoles, ammonia production, and phosphate solubilization. Never-dried fibers demonstrated a favorable microbial load (between 6.78 and 8.22 Log CFU g-1), although the cell viability after drying diminished more than 1 Log CFU g-1 relative to the never-dried matter. Notable findings were achieved when comparing the two distinct growth conditions and the timing of functional fiber inoculation. Post-drying disparities between unmodified fibers and functional fibers were observed, underscoring the possible role of natural polymers in enhancing cellular protection. Plant growth-promoting testing exhibited advantageous results relative to the non-immobilized consortia. These results establish a foundation for additional research, concentrating on shelf-life assessments and suitable applications in plant and greenhouse investigations.