Crohn's disease (CD) and ulcerative colitis (UC) are chronic relapsing inflammatory bowel disorders (IBD), the pathogenesis of which is uncertain but includes genetic susceptibility factors, immune-mediated tissue injury and environmental influences, most of which appear to act via the gut microbiome. We hypothesized that host-microbe alterations could be used to prognostically stratify patients experiencing relapses up to four years after endoscopy. We therefore examined multiple omics data, including published and new datasets, generated from paired inflamed and non-inflamed mucosal biopsies from 142 patients with IBD (54 CD; 88 UC) and from 34 control (non-diseased) biopsies. The relapse-predictive potential of 16S rRNA gene and transcript amplicons (standing and active microbiota) were investigated along with host transcriptomics, epigenomics and genetics. While standard single-omics analysis could not distinguish between patients who relapsed and those that remained in remission within four years of colonoscopy, we did find an association between the number of flares and a patient's succinotype. Our multi-omics machine learning approach was also able to predict relapse when combining features from the microbiome and human host. Therefore multi-omics, rather than single omics, better predicts relapse within 4 years of colonoscopy, while a patient's succinotype is associated with a higher frequency of relapses.

Metagenomic sequencing deepened our knowledge about the role of the intestinal microbiota in human health, and several studies with various methodologies explored its dynamics during antibiotic treatments. We compared the impact of four widely used antibiotics on the gut bacterial diversity. We used plasma and fecal samples collected during and after treatment from healthy volunteers assigned to a 5-day treatment either by ceftriaxone (1 g every 24 h through IV route), ceftazidime/avibactam (2 g/500 mg every 8 h through IV route), piperacillin/tazobactam (1 g/500 mg every 8 h through IV route) or moxifloxacin (400 mg every 24 h through oral route). Antibiotic concentrations were measured in plasma and feces, and bacterial diversity was assessed by the Shannon index from 16S rRNA gene profiling. The relationship between the evolutions of antibiotic fecal exposure and bacterial diversity was modeled using non-linear mixed effects models. We compared the impact of antibiotics on gut microbiota diversity by simulation, using various reconstructed pharmacodynamic indices. Piperacillin/tazobactam was characterized by the highest impact in terms of intensity of perturbation (maximal [IQR] loss of diversity of 27.3% [1.9; 40.0]), while moxifloxacin had the longest duration of perturbation, with a time to return to 95% of baseline value after the last administration of 13.2 d [8.3; 19.1]. Overall, moxifloxacin exhibited the highest global impact, followed by piperacillin/tazobactam, ceftazidime/avibactam and ceftriaxone. Their AUC between day 0 and day 42 of the change of diversity indices from day 0 were, respectively, -13.2 Shannon unit.day [-20.4; -7.9], -10.9 Shannon unit.day [-20.4; -0.6] and -10.1 Shannon unit.day [-18.3; -4.6]. We conclude that antibiotics alter the intestinal diversity to varying degrees, both within and between antibiotics families. Such studies are needed to help antibiotic stewardship in using the antibiotics with the lowest impact on the intestinal microbiota.

Excess amino acids from a protein-rich diet are mainly catabolized in the liver. However, it is still unclear to what extent the gut microbiota may be involved in the mechanisms governing this catabolism. Therefore, the aim of this study was to investigate whether consumption of different dietary protein concentrations induces changes in the taxonomy of the gut microbiota, which may contribute to the regulation of hepatic amino acid catabolism. Consumption of a high-protein diet caused overexpression of HIF-1α in the colon and increase in mitochondrial activity, creating a more anaerobic environment that was associated with changes in the taxonomy of the gut microbiota promoting an increase in the synthesis of secondary bile acids, increased secretion of pancreatic glucagon. This effect was demonstrated in pancreatic islets, where secondary bile acids stimulated the expression of the PC2 enzyme that promotes glucagon formation. The increase in circulating glucagon was associated with an induction of the expression of hepatic amino acid-degrading enzymes, an effect attenuated by antibiotics. Thus, high protein intake in mice and humans induced the increase of different species in the gut microbiota with the capacity to produce secondary bile acids leading to an increase in secondary bile acids and glucagon levels, promoting amino acid catabolism.

Recent advancements in data technology offer immense opportunities for the discovery and development of new enzymes for the green synthesis of chemicals. Current protein databases predominantly prioritize overall sequence matches. The multi-scale features underpinning catalytic mechanisms and processes, which are scattered across various data sources, have not been sufficiently integrated to be effectively utilized in enzyme mining. In this study, we developed a sequence- and taxonomic-feature evaluation driven workflow to discover enzymes that can be expressed in E. coli and catalyze chemical reactions in vitro, using alcohol oxidase (AOX) for demonstration, which catalyzes the conversion of methanol to formaldehyde. A dataset of 21 reported AOXs was used to construct sequence scoring rules based on features, including sequence length, structural motifs, catalytic-related residues, binding residues, and overall structure. These scoring rules were applied to filter the results from HMM-based searches, yielding 357 candidate sequences of eukaryotic origin, which were categorized into six classes at 85 % sequence similarity. Experimental validation was conducted in two rounds on 31 selected sequences representing all classes. Among these selected sequences, 19 were expressed as soluble proteins in E. coli, and 18 of these soluble proteins exhibited AOX activity, as predicted. Notably, the most active recombinant AOX exhibited an activity of 8.65 ± 0.29 U/mg, approaching the highest activity of native eukaryotic enzymes. Compared to the established UniProt-annotation-based workflow, this feature-evaluation-based approach yielded a higher probability of highly active recombinant AOX (from 8.3 % to 19.4 %), demonstrating the efficiency and potential of this multi-dimensional feature evaluation method in accelerating the discovery of active enzymes.

The floatation of anammox granules can be a serious challenge in practical wastewater treatment, as it can deteriorate reactor performance and cause bacterial loss. To deepen the understanding of floatation mechanism, in this study, both the floating (F-AnGS) and settling anammox granules (S-AnGS) from a high-rate anammox reactor were comparatively investigated. F-AnGS demonstrated 1.6 times higher specific anammox activity compared to S-AnGS, but only 65 % of produced gas could be successfully released, as quantified by anaerobic respirometry. In addition to the overall EPS accumulation, F-AnGS exhibited a heterogeneous microstructure distinct from that of S-AnGS, as revealed by 3D X-ray microscopic imaging at the single granule level. The heterogeneous distribution of EPS, which can form a dense surface layer, was the main cause for granule floatation. The heterogeneous microstructure of F-AnGS can reduce the distance between microorganisms and enhance the metabolic interaction between anammox bacteria and heterotrophs. The abundance of community members did not have a significant variation, but the functional genes related to anammox and partial denitrification pathway were significantly increased, indicating the enhanced nitrite loop in F-AnGS. This study proposed new structural insights into mechanism of anammox granule floatation, suggesting the appropriate activity control of granule-based anammox process.

Tongfu Xiexia Decoction (TFXXD), a traditional Chinese medicine formula, comprises six herbs: semen raphani, Rehmannia glutinosa, rhubarb, Magnolia officinalis, trifoliate orange, and mirabilite. The aim of this study was to investigate the effects of TFXXD on its biotransformation and microbial abundance in the gut of constipated rats.

Ultrahigh-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS/MS) was used to analyze the biotransformation of TFXXD in the colonic contents, and 16S rRNA sequencing was used to assess the structure and diversity of the gut microbiota across various rat groups.

We identified and analyzed 25 biotransformations of TFXXD in the colonic contents, which undergo various reactions such as deglycosylation, ring opening, and hydration in intestinal bacteria, with 14 originating from trifoliate orange, six from rhubarb, three from Rehmannia glutinosa, one from Semen raphani, and one from M. officinalis. 16S rRNA analysis revealed that TFXXD significantly enhanced the relative abundance of beneficial bacteria, such as Lactobacillus and Bacteroides while significantly reducing Oscillospira abundance. Moreover, TFXXD considerably affected the carbohydrate and amino acid metabolism. Correlation analyses revealed a significant negative correlation between Bacteroides and ACH (Acetylcholine), NO (nitric oxide), D-Lac (D-Lactate), IL-6 (Interleukin-6), TNF-α (tumor necrosis factor-α), and IL-1β (Interleukin-1β) and a significant positive correlation between Bacteroides, hesperetin, and rhein.

In conclusion, our findings indicate that TFXXD can modulate the structure and diversity of the gut microbiota and enhance the metabolic balance in constipated rats.

Dissolved organic nitrogen (DON) is often an overlooked form of nitrogen that can leach from the soil into aquifers. The reactive transport and dispersion of DON in aquifers can contribute to regional nitrogen contamination. The current body of research has primarily focused on the vertical leaching process of DON through the vadose zone. However, these studies have largely ignored the broader reactive transport of DON within aquifers under the influence of groundwater flow. In this study, we investigate the reactive transport of DON under groundwater flow conditions. Utilizing molecular biological technologies, we aim to reveal DON's intrinsic role in the nitrogen cycle within aquifers. Our findings reveal that urea exhibits greater mobility compared to amino acids and proteins. The transport of amino acids and proteins reduces the NO3--N concentrations (44.6 % and 89.6 %) compared to the blank control, while urea leads to the accumulation of NO3--N in groundwater (10.1 %). Amino acid and protein columns show higher relative abundances of Pseudomonas (10.1 % and 7.3 %) and Thermomonas (3.9 % and 5.1 %) with denitrification functions, facilitating denitrification in groundwater. Conversely, the presence of urea increases the relative abundances of Nitrosomonadaceae and Nitrophilus (0.33 % and 0.67 %), posing a potential NO3--N contamination risk. Biotransformation has the greatest effect on protein transport (19.6 %), while adsorption mainly influences amino acid transport (12.4 %). The study provides fundamental insights into the reactive transport of different DON components in aquifers, which holds important implications for regional groundwater environment protection.

Bladder cancer is a common malignancy of the genitourinary system. Recent studies have confirmed the existence of microorganisms in urine. This study aimed to characterize changes in the urinary microbiota of Chinese bladder cancer patients and determine differences between patients with muscle-invasive bladder cancer (MIBC) and those with non-muscle-invasive bladder cancer (NMIBC).

Urine samples were collected from 64 patients with bladder cancer and 94 disease-free controls using the clean catch method and sequenced by 16S rRNA gene sequencing. Sequencing reads were filtered by VSEARCH and clustered by UPARSE.

Significant associations were found between urinary microbiota and factors such as sex, age, and disease status. After age adjustment, differences in beta diversity were observed between healthy men and women, cancer patients and healthy controls, and NMIBC and MIBC patients. The cancer patients had an increased abundance of 14 bacterial genera, including Stenotrophomonas, Propionibacterium, and Acinetobacter. Notably, Peptoniphilus spp. were enriched in high-risk MIBC patients, indicating their potential as a risk marker. Functional prediction via PICRUSt analysis suggested enriched metabolic pathways in specific disease groups. Furthermore, molecular ecological network analysis revealed differences based on sex and disease type.

This significant microbial diversity indicates a potential correlation between urinary microbiota dysbiosis and bladder cancer, with implications for risk stratification and disease management. The identified urinary microbiota may serve as noninvasive markers for bladder cancer, warranting further validation in larger cohorts. This study provides a foundation for further research on the mechanisms of bladder cancer progression.

Contaminants of emerging concern (CECs) have gathered significant public attention due to their widespread occurrence, high persistence, and increasing exposure potential. In this study, we used polyethylene biocarriers for acclimating biofilms from singular or combined activated sludges collected from three wastewater treatment plants (R, P, and L) over 5 month-long cycles. The acclimated biofilms achieved an average removal rate at 0.333, 0.313, and 0.185 week-1 for N, N-diethyl-meta-toluamide (DEET), sulfamethoxazole (SMX), and carbamazepine (CBZ), respectively, when external carbon was supplemented, which were significantly higher (p < 0.05) than biofilms that did not receive external carbons. Metabolite screening revealed SMX transformation through ipso-hydroxylation and acetyl conjugation, while CBZ degradation could be initiated by epoxidation. Significant but slower degradation rates (0.024∼0.031 week-1) were observed for aminotriazole (AMT), lidocaine (LDC), and trimethoprim (TMP), whereas atrazine (ATZ) exhibited minimal removal, highlighting its high recalcitrance. Biofilms acclimated from individual R and P sludges, with external carbon supplementation, attained the greatest removal efficiencies for 7 CECs. Multivariate statistical correlations (p < 0.05) identified potential degraders, including Sphingomonas and Zoogolea for AMT, Labrys and Koazkia for CBZ, and Asprobacter, unclassified Cyclobacteriaceae (ELB16-189) and Bryobacteraceae (Fen-178) for LDC. Abundance distribution of potential degraders among biofilms revealed that Sludge R favored the enrichment of key degraders for AMT, CBZ and LDC, while Sludge P was more conducive to acclimating CBZ degraders. This study advances our understanding of strategies in biofilm acclimation to improve CEC removal and provides insights into degradation pathways and associated microbial communities for future research.

The accumulation of pipeline sediments within drinking water distribution systems (DWDS) has garnered widespread attention because of their roles as microbial reservoirs. However, previous investigations predominantly concentrated on microbial occurrence in unchlorinated DWDS sediments but largely overlooked chlorinated systems and failed to characterize the spatial distribution patterns of potential pathogens along the DWDS. This study systematically examined bacterial communities in both the sediment and bulk water phases across a chlorinated DWDS through the seasonal collection of 96 samples. Physicochemical water quality parameters, such as turbidity and residual chlorine, exhibited relative stability throughout the network. As anticipated, sediment samples showed substantial particulate accumulation (summer: 1.13 ± 0.61 Log10 NTU; winter: 1.07 ± 0.45 Log10 NTU). Microbial biomass proved significantly elevated in sediments (summer: 4.78 ± 0.65 Log10 gene copies/mL; winter: 4.99 ± 0.42 Log10 gene copies/mL) than water samples (summer: 3.98 ± 0.50 Log10 gene copies/mL; winter: 4.06 ± 0.57 Log10 gene copies/mL; p < 0.05), with similar patterns emerging for the potentially pathogenic fungi, Mycobacterium spp., and Legionella spp. Notably, no longitudinal accumulation gradient of microbial biomass was detected along the pipeline network in either the sediment or water samples. Interestingly, the winter sediment samples displayed peak microbial biomass levels. Seasonal variation exerted a substantial effect on microbial community composition, with turbidity and residual chlorine demonstrating stronger correlations with biomass in summer than in winter. These findings underscore the necessity for regular sediment removal from chlorinated DWDS as a critical preventive measure against waterborne pathogen proliferation and disease transmission.

Shipping scrubber effluents, containing a cocktail of Polycyclic Aromatic Hydrocarbons (PAHs), show undisputed effects at single-species experiments while PAHs fate in the marine environment after effluent discharge is still investigated. Bacterioplankton, composed of abundant diverse taxa with xenobiotic-degrading capabilities, are the first responders to scrubber emissions and can affect PAHs impacts on marine life. This work aims to examine the fate of scrubber effluent PAHs and alkyl-PAHs in mesocosms of coastal bacterioplankton communities from a pristine (phytoplankton carbon biomass was 8.16 μg C L-1) and a eutrophic (105.35 μg C L-1) coastal site. High-throughput 16S rRNA metabarcoding revealed differential responses of the bacterioplankton linked to their initial community structure and population abundances. Taxa known for their PAHs-degrading capacity were retrieved, including the genera Roseobacter, Porticoccus, Marinomonas, Arcobacter, Lentibacter, Lacinutrix, Pseudospirillum, Glaciecola, Vibrio, Marivita, and Mycobacterium, and were found to have increased roles in shifted communities by increasing their relative abundances at least 5-fold in treatments with high scrubber effluent additions. Additionally, metagenomic analysis of shotgun sequencing, indicated an increase on the number of Clusters of Orthologous Genes (COGs) associated with pathways involved in PAHs degradation. Up to 198 more COGs involved in signal transduction were retrieved in scrubber effluent enriched mesocosms compared to controls, while 15, 86, and 136 more COGs associated with naphthalene, aromatic compound, and benzoate degradation, respectively, were detected in the pristine mesocosms after effluent additions. In both experiments, bacterioplankton responses towards xenobiotic degradation under increased PAHs and alkyl-PAHs were coupled with a drop in their concentrations, below the limit of detection by Day 3 of the experiment in the eutrophic community, and by half in Day 6 in the pristine environment's community. Our findings indicate that PAHs and alkyl-PAHs impacts can be rapidly reduced in natural systems of high bacterial activity.

Wastewater is a hotspot for viral diversity, harboring various microbial, plant, and animal viruses, including those that infect humans. However, the dynamics, resilience, and ecological roles of viral communities during treatment are largely unknown. In this study, we explored RNA virus ecogenomics using metagenomics from influent and effluent samples across three wastewater catchment areas in Chile, with a population of 7.05 million equivalent inhabitants. We identified 14,212 RNA-dependent RNA polymerase (RdRP)-coding sequences from the Orthornavirae kingdom, clustering into 4989 viral species. Using extensive databases of 14,150 family-level representative sequences, we classified 90 % of our sequences at the family level. Our analysis revealed that treatment reduced viral richness and evenness (Shannon index), but phylogenetic diversity remained unchanged. Effluents showed lower richness and evenness than influents with similar phylogenetic diversity. Species turnover, influenced by catchment area and treatment, accounted for 54 % of sample dissimilarities (Weighted Unifrac). Biomarker analysis indicated that families like Astroviridae and Fiersviridae were more abundant in influents, while Reoviridae and Virgaviridae dominated effluents. This suggests that viral resistance to treatment varies and cannot be solely attributed to genome type, size, or morphology. We traced viral genomes through time and space, identifying sequences like the Pepper Mild Mottle Virus (PMMoV) from the Virgaviridae family over large distances and periods, highlighting its wastewater marker potential. High concentrations of human pathogens, such as Rotavirus (Reoviridae) and Human Astrovirus (Astroviridae), were found in both influents and effluents, stressing the need for continuous monitoring, especially for treated wastewater reuse.

Manganese (Mn) oxides are crucial for degrading organic pollutants and driving biogeochemical cycles. Microorganisms drive Mn(II) oxidation, but traditional cultivation-dependent identification methods are inefficient and error-prone. To overcome these limitations, we developed MnOxGeneTool, a bioinformatics tool for identifying and quantifying Mn(II)-oxidizing genes from genomic and metagenomic data. MnOxGeneTool consists of three main components: (1) a curated database of known Mn(II)-oxidizing proteins and their homologues, (2) a hidden Markov models (HMMs) database derived from this protein data set, and (3) a computational pipeline that integrates bioinformatics tools (e.g., HMMER and BLASTX) to identify and quantify Mn(II)-oxidizing genes. We assessed the accuracy and sensitivity of these HMMs through cross-validation, demonstrating their effectiveness in identifying Mn(II)-oxidizing genes in bacterial genomes. Using MnOxGeneTool, we explored the phylogenetic diversity of Mn(II)-oxidizers and identified 824 bacterial genera containing Mn(II)-oxidizing genes, significantly expanding previous knowledge in this field. Additionally, we analyzed metagenomic data from various environments to explore environmental drivers of Mn(II)-oxidizing genes, identifying two potential drivers: oligotrophic conditions and alkaline environments. These findings enable targeted discovery of novel Mn(II)-oxidizers and genetic determinants through identification of their ecological niches and expression optima, thereby expanding MnOxGeneTool's predictive coverage of uncatalogued Mn(II)-oxidizing proteins. By providing an innovative bioinformatics tool that enables efficient identification and quantification of Mn(II)-oxidizing genes from both genomic and metagenomic data, this study offers significant advancements in the research of biogenic Mn(II) oxidation. The tool is available at https://github.com/wyh19990121/MnOxGeneTool.

Human microbiome plays a crucial role in host health and disease by mediating the impact of environmental factors on clinical outcomes. Mediation analysis is a valuable tool for dissecting these complex relationships. However, existing approaches are primarily designed for cross-sectional studies. Modern clinical research increasingly utilizes long follow-up periods, leading to complex data structures, particularly in metagenomic studies. To address this limitation, we introduce a novel mediation framework based on structural equation modeling that leverages linear mixed-effects models using penalized quasi-likelihood estimation with a debiased lasso. We applied this framework to a 16S rRNA sputum microbiome data set collected from patients with cystic fibrosis over 10 years to investigate the mediating role of the microbiome in the relationship between clinical states, disease aggressiveness phenotypes, and lung function. We identified richness as a key mediator of lung function. Specifically, Streptococcus was found to be significantly associated with mediating the decline in lung function on treatment compared to exacerbation, while Gemella was associated with the decline in lung function on recovery. This approach offers a powerful new tool for understanding the complex interplay between microbiome and clinical outcomes in longitudinal studies, facilitating targeted microbiome-based interventions.

Understanding the mechanisms by which the microbiome influences clinical outcomes is paramount for realizing the full potential of microbiome-based medicine, including diagnostics and therapeutics. Identifying specific microbial mediators not only reveals potential targets for novel therapies and drug repurposing but also offers a more precise approach to patient stratification and personalized interventions. While traditional mediation analyses are ill-equipped to address the complexities of longitudinal metagenomic data, our framework directly addresses this gap, enabling robust investigation of these increasingly common study designs. By applying this framework to a decade-long cystic fibrosis study, we have begun to unravel the intricate relationships between the sputum microbiome and lung function decline across different clinical states, yielding insights that were previously unknown.

Ecological restoration is a promising approach to alleviate eutrophication. However, its impacts on greenhouse gas (GHG) emissions and the underlying microbial mechanisms in different habitats of lakes remain unclear. To address this knowledge gap, we measured carbon dioxide (CO2), methane (CH4) and nitrous oxide (N2O) fluxes at both water-air and sediment-water interfaces of eutrophic (Caohai) and restored area (Dapokou) of Dianchi Lake, a typical eutrophic lake in China. Meanwhile, we investigated the responses of planktonic and sedimentary bacterial and fungal communities by high-throughput sequencing. Our results indicated that 6 years of ecological restoration significantly reduced CO2 and N2O fluxes by 1.0-3.6 and 2.2-2.8 folds respectively, with more pronounced variations at the water-air interface than the sediment-water interface. Ecological restoration also shifted the structures of planktonic bacterial and fungal communities remarkably, leading to a significant reduction in the relative abundances of Actinobacteriota (by 70.94%), Bacteroidota (by 61.65%), Planctomycetota (by 74.18%) and Chytridiomycota (by 95.44%). Correlation analyses further suggested that GHG fluxes at the water-air interface were significantly correlated with planktonic microbial community composition (P < 0.05), and the significant reduction of CO2 and N2O fluxes under ecological restoration could be attributed to the decreased abundances of organic matter decomposers (such as hgcI_clade, Sporichthyaceae and Acidibacter) and increased abundances of autotrophs (such as Hydrogenophaga and Cyanobium_PCC-6307) in water. Collectively, our findings verify the importance of ecological restoration in reducing GHG emissions in inland lake ecosystems, providing new insights for addressing global climate change and advancing carbon neutrality.

SARS-CoV-2 continues to evolve, with new variants emerging that evade pre-existing immunity and limit the efficacy of existing vaccines. One approach toward developing superior, variant-proof vaccines is to engineer immunogens that preferentially elicit antibodies with broad cross-reactivity against SARS-CoV-2 and its variants by targeting conserved epitopes on spike. The inner and outer faces of the receptor binding domain (RBD) are two such conserved regions targeted by antibodies that recognize diverse human and animal coronaviruses. To promote the elicitation of such antibodies by vaccination, we engineered "resurfaced" RBD immunogens that contained mutations at exposed RBD residues outside the target epitopes. In the context of pre-existing immunity, these vaccine candidates aim to disfavor the elicitation of strain-specific antibodies against the immunodominant receptor binding motif (RBM) while boosting the induction of inner and outer face antibodies. The engineered resurfaced RBD immunogens were stable, lacked binding to monoclonal antibodies with limited breadth, and maintained strong interactions with target broadly neutralizing antibodies. When used as vaccines, they limited humoral responses against the RBM as intended. Multimerization on nanoparticles further increased the immunogenicity of the resurfaced RBD immunogens, thus supporting resurfacing as a promising immunogen design approach to rationally shift natural immune responses to develop more protective vaccines.IMPORTANCESARS-CoV-2 is the third coronavirus to cause significant human disease over the last two decades. Despite their success in preventing serious disease, current SARS-CoV-2 vaccines must be updated regularly to match the circulating strains for continued protection. Therefore, it would be advantageous to develop vaccines that protect more broadly against SARS-CoV-2, its variants, and other pre-emergent coronaviruses. This may be achieved by preferentially eliciting antibodies against conserved regions of the spike protein that decorates the virus. Toward this goal, we engineered vaccine candidates to target the conserved inner and outer domains of the Receptor Binding Domain of SARS-CoV-2, by altering the surface of the wild-type protein such that strain-specific antibodies that bind outside these regions are no longer recognized. When used in animals with pre-existing SARS-CoV-2 immunity, these molecules reduce the elicitation of variant-specific antibodies, thus providing a blueprint to alter the natural immunodominance hierarchies of SARS-CoV-2 proteins.

Norway has low prevalence of resistance in the clinics, the contribution of hospital effluent to the spread of antimicrobial resistance (AMR) in the environment is largely unknown. The aim of our study was to determine the role of hospital sewage in dissemination of AMR by defining resistome of hospital effluent, and influent and treated-effluent from the receiving sewage treatment plant (STP) using culture-based methods and metagenomics. Around 94 % E. coli strains (n = 66) were multidrug-resistant (MDR), while 92.3 % of the Klebsiella spp. strains (n = 55) showed MDR phenotype, with some strains carrying carbapenemases, such as NDM-5 (n = 3) and KPC-3 (n = 3). Identical clones of Klebsiella michiganensis were detected in hospital effluent, influent and STP treated effluent. From approximately 238 Gigabases of sequence data, we assembled 1205 antibiotic resistance genes (ARGs) using fARGene method, of which 349 genes represented novel ARGs (< 90 % amino acid identity against known ARGs). Both known and novel ARGs (n = 54) were shared between hospital effluent and the treated effluent of the receiving STP. We assembled 523 metagenome assembled genomes (MAGs) with several representing novel taxa, of which 138 (26 %) MAGs carried 429 ARGs with > 83 % representing putative novel ARGs. Potential pathogens accounted for 60 % of the detected ARGs. Around 15.4 % MAGs were shared between hospital effluent and STP treated effluent. We demonstrate that hospital effluent in Norway has a high diversity of both known and novel ARGs. We show that hospital effluent contributes to the dissemination of not only clinically relevant pathogens but also known and novel ARGs into the receiving marine environment in Norway.

Rice tillering is an important agronomic trait regulated by plant genetic and environmental factors. However, the role and mechanism of the root microbiota in modulating rice tillering have not been explored. Here, we examined the root microbiota composition and tiller numbers of 182 genome-sequenced rice varieties grown under field conditions and uncovered a significant correlation between root microbiota composition and rice tiller number. Using cultivated bacterial isolates, we demonstrated that various members of the root microbiota can regulate rice tillering in both laboratory and field conditions. Genetic, biochemical, and structural analyses revealed that cyclo(Leu-Pro), produced by the tiller-inhibiting bacterium Exiguobacterium R2567, activates the rice strigolactone (SL) signaling pathway by binding to the SL receptor OsD14, thus regulating tillering. The present work provides insight into how the root microbiota regulates key agronomic traits and offers a promising strategy for optimizing crop growth by harnessing the root microbiota in sustainable agriculture.

As one of the top predators in the Yellow Sea, the Tanaka's snailfish (Liparis tanakae) plays an important ecological role in maintaining the structure and function of the ecosystem. This species also has fast and strong adaptability to external pressures such as climate change and fishing activities. To facilitate further molecular evolution researches of L. tanakae, we generated a chromosome-scale genome assembly in this study. The final assembly yielded 574.44 Mb in total length, with a scaffold N50 of 24.64 Mb, and anchored 97.87% of the sequences into 24 pseudo-chromosomes. Our assembly was 20.18 Mb longer than the reference genome (Tanakav1) in total length, with higher scaffold N50 and fewer scaffolds. The BUSCO score of 97.3% and Merqury quality value of 36.98 revealed high completeness and accuracy of our assembly. The genome contained 20,933 predicted protein-coding genes and 28.28% of the assembly was annotated as repetitive sequences. This study significantly advances the genomic resources for L. tanakae and facilitates future adaptation and evolution researches of this species.

Israel's Mediterranean biogeographical region is characterized by high habitat diversity and stark seasonal changes in forage composition, availability and quality. Managers of protected areas in this region advocate livestock ranching to mitigate fire risk and enhance conservation merits. However, competition between livestock and endangered, native ungulates in these areas might impair their functioning as refugia. We used fecal DNA metabarcoding to study the diets of native mountain gazelles (Gazella gazella) and domestic cattle (Bos taurus), in two nature reserves with distinct vegetation types (shrubland vs grassland), and during different seasons. Dietary overlap was ubiquitously low, and seasonal changes in the diets of both ungulates translated into differences in their dietary overlap, with the highest overlap found in grassland during winter. This generally low overlap may be attributed to the extreme differences in their body size or may also result from long-lasting sympatry of gazelles and cattle - first wild and later domesticated-shaping a robust dietary separation. Yet, since cattle biomass is typically much higher than gazelles', a low dietary overlap in key food items of gazelles may result in their depletion which might negatively affect gazelles, especially during the fawning season and drought years. Our results highlight the need to cover diverse conditions when studying herbivore dietary composition and overlap.

Methylmercury (MeHg) is recognized as a deleterious neurotoxin with the traits of biomagnification through the food chain and accumulation in edible aquatic products. However, the in situ production of MeHg in aquaculture environments has not been well understood. Herein, the sediments were collected from aquaculture ponds with different rearing operations. Isotope-based tracer analysis showed that Hg methylation and MeHg demethylation rate constants in the aquaculture sediments were 0.001-0.022 d-1 and 0.11-0.40 d-1, respectively. Most of bacterial Hg methylators (> 97.0 %) in aquaculture sediments were assigned to Firmicutes and Actinobacteria phyla. Four functional genes responsible for Hg transformation (hgcAB and merAB) could be detected in the aquaculture sediments. In particular, Hg methylation rate constants were positively and significantly correlated with the levels of hgcAB genes (p < 0.05). Inhibitive reagent addition assays and correlation analysis consistently demonstrated that sulfate-reducing bacteria (SRB) were the main methylators in aquaculture sediments, and antibiotic use could fortify the resistance of Hg methylators to antibiotics. These findings suggest that the in situ production of MeHg in aquaculture sediments may be effectively reduced via inhibiting SRB activities, and both hgcAB genes are useful markers of MeHg production in aquaculture environments.

The global prevalence of food allergies, particularly IgE-mediated responses, is increasing at an alarming rate. This trend is likely driven by environmental factors such as nanoplastics (NPs) ingestion and the westernization of dietary and lifestyle habits. This study examines the impact of polystyrene nanoplastics (PS-NPs) on ovalbumin (OVA)-induced food allergies in mice subjected to either a normal diet (ND) or a high-fat diet (HFD). BALB/c mice were stratified into eight groups based on dietary regimen, NP exposure, and OVA sensitization. Food allergy was induced via OVA administration, and multiple physiological and immunological parameters were evaluated, including body weight, intestinal permeability, cytokine profiles, gut microbiota composition, and small intestinal gene expression. Mice in the HFD + OVA + NP group exhibited significant increases in intestinal permeability, diarrhea severity, and serum OVA-specific IgE levels compared to other groups. Flow cytometric analysis revealed an expansion of innate lymphoid cells (ILC2 and ILC1) within the lamina propria of the small intestine. Shotgun metagenomic sequencing demonstrated gut microbiota dysbiosis, characterized by a reduction in beneficial bacterial populations in the HFD + OVA + NP cohort. Weighted Gene Co-Expression Network Analysis (WGCNA) identified a negative correlation between NPs exposure or OVA sensitization and the expression of Slc1a1, Slc5a8, and Mep1a, while a positive correlation was observed with Aa467197 expression. These findings indicate that oral exposure to PS-NPs exacerbates OVA-induced food allergies, particularly in the context of an HFD, through mechanisms involving increased intestinal permeability, gut microbial dysbiosis, and gene expression modulation. This study highlights the potential health hazards posed by environmental microplastic contamination and its possible contribution to the escalating incidence of food allergies.

Viruses alter the ecological and evolutionary trajectories of bacterial host communities. Plant grafting is a technique that integrates two species or varietiies and have consequences on the rhizosphere functioning. The grafting effects on the taxonomic and functional assembly of viruses and their bacterial host in the plant rhizosphere remain largely elusive. Using shotgun metagenome sequencing, we recover a total of 1441 viral operational taxonomic units from the rhizosphere of grafted and ungrafted plants after 8-year continuous monoculture. In the grafted and ungrafted rhizosphere, the Myoviridae, Zobellviridae and Kyanoviridae emerged as the predominant viral families, collectively representing around 40% of the viral community in each respective environment. Grafting enriched the members in viral family Kyanoviridae, Tectiviridae, Peduoviridae and Suoliviridae, and auxiliary metabolic genes related to pyruvate metabolism and energy acquisition (e.g., gloB, DNMT1 and dcyD). The virus-bacterial interactions increased the rapid growth potential of bacteria, which explains the strong increase in abundance of specific bacterial hosts (i.e., Chitinophagaceae, Cyclobacteriaceae and Spirosomaceae) in the grafted-plant rhizosphere. Overall, these results deepen our understanding of microbial community assembly and ecological services from the perspective of virus-host interactions.

Although the Antarctic continent represents one of the most hostile environments on earth, microbial life has adapted to cope with these extreme conditions. Lichens are one of the most successful groups of organisms in Antarctica, where they serve as unique niches for microbial diversification. We have selected eight epilithic lichen species growing in Victoria Land (three cosmopolitan and five endemic to Antarctica) to describe with amplicon sequencing the diversity of the associated fungal and bacterial communities. The lichen mycobiota is predominantly composed of Ascomycota belonging to the classes Chaetothyriomycetes and Dothideomycetes, while a few key representative taxa were recognised as basidiomycetous yeasts. Bacteria associated with lichens were represented by Pseudomonadota, Cyanobacteria, and Bacteroidota in which psychrophilic genera were identified. The microbiota was diverse among the lichen species, and their variation was driven by the lichen species itself and their endemic or cosmopolitan distribution. There was a strong association of the microbial communities linked to the lichen itself, rather than to the specific characteristics of the collecting site. The lichen thallus, thus, plays an important role in microbial diversification and may potentially act as a selective biodiversity filter in which different fungal and bacterial communities thrive in it.

Probiotic microbes such as Lactobacillus may reduce serum total cholesterol (TC) and low-density lipoprotein (LDL) cholesterol. The objective of this study was to assess the effect of Lactobacillus plantarum strains CECT7527, CECT7528, and CECT7529 (LP) on the serum lipids, cardiovascular parameters, and fecal gut microbiota composition in patients with mild hypercholesterolemia. A randomized, double-blinded, placebo-controlled clinical trial with 86 healthy adult participants with untreated elevated LDL cholesterol ≥ 160 mg/dl was conducted. Participants were randomly allocated to either placebo or LP (1.2 × 109 CFU/d) for 12 weeks. LDL, HDL, TC, and triglycerides (TG), cardiovascular parameters (blood pressure, arterial stiffness), and fecal gut microbiota composition (16S rRNA gene sequencing) were assessed at baseline and after 12 weeks. Both groups were comparable regarding age, sex, and LDL-C at baseline. LDL-C decreased (mean decrease - 6.6 mg/dl ±  - 14.0 mg/dl, Ptime*group = 0.006) in the LP group but not in the placebo group. No effects were observed on HDL, TG, or cardiovascular parameters or overall gut microbiota composition. Responders to LP intervention (> 5% LDL-C reduction) were characterized by higher BMI, pronounced TC reduction, higher abundance of fecal Roseburia, and lower abundance of Oscillibacter. In conclusion, 12 weeks of L. plantarum intake moderately reduced LDL-C and TC as compared to placebo. LDL-C-lowering efficacy of L. plantarum strains may potentially be dependent on individual difference in the gut microbiota. Trial registration: DRKS00020384, dated 07/01/2020.

Salvia miltiorrhiza Bunge, a renowned medicinal herb in traditional Chinese medicine, displays distinctive root texture and high phenolic acid content, traits influenced by genetic and environmental factors. However, the underlying regulatory networks remain unclear. Here, we performed multi-omics analyses on ecotypes from four major Chinese regions, focusing on environmental impacts on root structure, phenolic acid accumulation and lignin composition. Lower temperatures and increased UV-B radiation were associated with elevated rosmarinic acid (RA) and salvianolic acid B (SAB) levels, particularly in the Sichuan ecotype. Structural models indicated that the radial arrangement of xylem conduits contributes to greater root hardness. Genomic assembly and comparative analysis of the Sichuan ecotype revealed a unique phenolic acid metabolism gene cluster, including SmWRKY40, a WRKY transcription factor essential for RA and SAB biosynthesis. Overexpression of SmWRKY40 enhanced phenolic acid levels and lignin content, whereas its knockout reduced root hardness. Integrating high-throughput (DNA affinity purification sequencing) and point-to-point (Yeast One-Hybrid, Dual-Luciferase and Electrophoretic Mobility Shift Assay) protein-DNA interaction detection platform further identified SmWRKY40 binding sites across ecotypes, revealing specific regulatory networks. Our findings provide insights into the molecular basis of root texture and bioactive compound accumulation, advancing breeding strategies for quality improvement in S. miltiorrhiza.

Human activities, such as agriculture, environmental manipulation, and city development, have impacted the distribution of flora, fauna, and microbes (including potential human pathogens) at the global level. This study focused on the bacterial genus Leptospira, an organism causing leptospirosis that is prevalent in tropical and subtropical regions. We hypothesized that although only a few cases of leptospirosis have been reported in the urban region of main island of Okinawa Prefecture (Okinawa Island, OKI), Japan, Leptospira is present in these regions.

Thirty-four samples were collected from rivers in urban OKI and rural Ishigaki Island (ISG) and analyzed to determine the distribution of Leptospira and mammals using environmental DNA (eDNA) metabarcoding. High-throughput sequencing analysis was performed to sequence the polymerase chain reaction products of partial leptospiral 16S rRNA and vertebrate mitochondrial 12S rRNA genes from 16 and 18 river samples of OKI and ISG, respectively, including the waters collected from Leptospira-endemic areas in ISG.

Leptospira noguchii and L. interrogans-related, two Leptospira species of the P1+ clade that are pathogenic to humans and mammals, were repeatedly detected in OKI and ISG, supporting our hypothesis. The sequence numbers of the five Leptospira species of P1- and P2 clades showed significant correlations with those of cattle (Bos taurus) in OKI; however, the potential host animals for P1+ species remain unclear. The total number of leptospiral sequences obtained from the ISG samples was correlated with the distance from the mountainous woodlands.

The pathogenic P1+ Leptospira was distributed in urban OKI, in addition to rural ISG. The factors correlated with leptospiral detection, that is, cattle eDNAs and the distance from mountainous forests in OKI and ISG, respectively, suggest the silent prevalence of Leptospira in urban and developing regions related to human activities. The findings of the present study provide insights into public health in cities with respect to climate change and possible flood damage.

Cephalopods play crucial roles in marine ecosystems, acting as both predators and prey for apex predators, thereby contributing to the distribution of energy and nutrients across the food web. Traditional net capture methods are often ineffective for studying cephalopods owing to their wide distribution in marine environments, necessitating the development of simple and efficient surveying techniques to assess cephalopod diversity. Therefore, in this study, we aimed to establish universal polymerase chain reaction primers specifically targeting mitochondrial 16S rRNA genes for environmental DNA metabarcoding in cephalopods. Two primer sets, Cep16S_D and Cep16S_O, were designed for squids and octopuses, respectively. Taxonomic specificity, resolution, and coverage of these primers were evaluated via in silico and in vitro analyses. Additionally, efficiency of these primer sets was assessed using tissue samples and mock communities. Finally, their applicability and performance were tested at various depths. The developed primers exhibited a relatively large amplification size with mixed bases that enhanced their amplification efficiency and sensitivity for cephalopod detection. We successfully identified cephalopod species with different body sizes, from small species, such as Heteroteuthis dagamensis, to large species, such as Architeuthis dux, at varying water depths. Overall, the primer sets established in this study serve as powerful tools to study cephalopod diversity and exhibit great potential for barcoding and genetic diversity investigations.

Viromics produces millions of viral genomes and fragments annually, overwhelming traditional sequence comparison methods. Here we introduce Vclust, an approach that determines average nucleotide identity by Lempel-Ziv parsing and clusters viral genomes with thresholds endorsed by authoritative viral genomics and taxonomy consortia. Vclust demonstrates superior accuracy and efficiency compared to existing tools, clustering millions of genomes in a few hours on a mid-range workstation.

Microplastics (MPs) are critical vectors for the dissemination of antibiotic resistance genes (ARGs); however, the prevalence and ecological risks of high-risk ARGs in mangrove ecosystems-globally vital yet understudied coastal habitats-remain poorly understood. To address this gap, this study investigated polyethylene, polystyrene, and polyvinyl chloride incubated in mangrove sediments for one month, focusing on high-risk ARGs, virulence gene (VGs), and pathogenic antibiotic-resistant bacteria within the mangrove plastisphere. High-throughput PCR and metagenomic analyses revealed that high-risk ARGs, VGs, and mobile genetic elements (MGEs) were significantly enriched on MPs compared to surrounding sediments. Pathogenic bacteria and MGEs were also more abundant in the plastisphere, highlighting its role as a hotspot for ARG dispersal. Metagenome-assembled genome analysis identified Pseudomonas and Bacillus as key hosts for ARGs, MGEs, and VGs, particularly multidrug resistance genes, integrase genes, and adherence factors. Notably, polystyrene harbored the highest abundance of pathogenic bacteria carrying ARGs, MGEs, and VGs, and mangrove root exudates were found to amplify horizontal gene transfer on MPs, uncovering a previously overlooked mechanism driving antibiotic resistance in coastal ecosystems. These findings not only elucidate how MPs accelerate the spread of ARGs, but also underscore the urgent need for targeted mitigation strategies to address the adverse impacts microplastic pollution on human, animal, and environmental health.

Artisanal and small-scale gold mining (ASGM) expansion in the Madre de Dios region of the Peruvian Amazon has transformed primary forests into a novel wetland complex of thousands of abandoned mining ponds. Despite their ecological relevance, post-mining recovery of these systems remains understudied, particularly regarding fish biodiversity and recolonisation. In this study, we evaluate fish community richness and composition in mining ponds of different dimensions, years post abandonment, physicochemical properties and degree of pulse flood connectivity using traditional collection-based methods and environmental DNA (eDNA) with the 12S and COI markers. We compared these two methods of biodiversity inventory and contrasted results from ASGM waterbodies with those obtained from nearby pristine oxbow lakes. Overall, we registered more fish richness at all sites using eDNA versus traditional methods, especially with the 12S marker. We identified 14 and 13 unique genera using traditional methods and eDNA, respectively, with 40 genera detected by both approaches, evidencing their complementarity. Notably, we found that the degree of pulse flooding connectivity was the main predictor of species richness among the abandoned mining ponds (p-value < 0.05). We registered 11-22, 23-71 and 56 morphospecies in non-flooded mining ponds, pulse flooded mining ponds and nearby oxbow lakes, respectively. Furthermore, the fish community composition of mining ponds most influenced by pulse flooding was similar to that of pristine lakes. Our findings highlight the role of hydrological connectivity in ecological recovery within mining-impacted wetlands. Future restoration efforts should enhance aquatic connectivity to accelerate recovery in post-mining environments.

Sperm mosaicism, the presence of a pathogenic variant in a subset of sperm, is an important cause of heritable genetic disease. However, clinical testing for sperm mosaicism outside research has been limited by the lack of Clinical Laboratory Improvement Amendments (CLIA)-validated results deliverable to patients. We developed the Sensitive Assay for Mosaicism (SAM), a two-phase method for sperm mosaicism detection. In phase 1, sperm DNA undergoes deep sequencing using next-generation sequencing or nanopore-based sequencing with unique molecular identifiers (UMIs) to improve accuracy. In phase 2, PCR primers specific to UMI sequences generate amplicons for CLIA-validated Sanger sequencing, providing patient-ready results. SAM's performance was characterized and tested on semen samples from 14 participants, each with a prior offspring with a de novo pathogenic variant. SAM demonstrated a detection limit of approximately 0.005%. The UMI strategy improved sequencing accuracy on next-generation sequencing and nanopore platforms from 99.9% to >99.999%, and from 93% to >99.99%, respectively. Sperm mosaicism was identified in two tested cases: FAM111A (5.51%) and FGFR3 (0.0129%), with FGFR3 exhibiting selfish mutation validated in unrelated individuals showing varying mosaicism levels. SAM provides sensitive detection of low-level sperm mosaicism with CLIA-validated results for patients, enabling recurrence risk assessment and guiding risk mitigation strategies such as in vitro fertilization with preimplantation genetic testing for monogenic disease, sperm donation, and prenatal diagnosis.

Protein developability is an important, yet often overlooked, aspect of protein discovery campaigns that is a key driver of utility. Recent advances have improved developability screening capacity, making it an increasingly viable option in early-stage discovery. Here, we engineered one component of developability, stability, of two affibody proteins-one that targets death receptor 5 and another that targets tumor necrosis factor receptor 1-previously evolved to bind receptor and non-competitively inhibit signaling via conformational modulation. Starting from an error-prone PCR library of each affibody, variants were screened via yeast surface display binder selections, including depletion of non-specific binders, followed by developability assessment using the on-yeast protease and yeast display level assays. Multiplex deep sequencing identified variants for further evaluation. Purified variants exhibited elevated stability-8°C to 14°C increase in Tm,app-with maintained 1-2 nM affinity for the TNFR1 affibody and 30-fold improvement in the DR5 affibody affinity to 0.8 nM.

Anti-CRISPR (Acr) proteins play a key role in phage-host interactions and hold great promise for advancing genome-editing technologies. However, finding new Acrs has been challenging due to their low sequence similarity. Recent advances in protein structure prediction have opened new pathways for Acr discovery by using 3D structure similarity. This study presents an updated AcrDB, with the following new features not available in other databases: (1) predicted Acrs from human gut virome databases, (2) Acr structures predicted by AlphaFold2, (3) a structural similarity search function to allow users to submit new sequences and structures to search against 3D structures of experimentally known Acrs. The updated AcrDB contains predicted 3D structures of 795 candidate Acrs with structural similarity (TM-score ≥0.7) to known Acrs supported by at least two of the three non-sequence similarity-based tools (TM-Vec, Foldseek, AcrPred). Among these candidate Acrs, 121 are supported by all three tools. AcrDB also includes 3D structures of 122 experimentally characterized Acr proteins. The 121 most confident candidate Acrs were combined with the 122 known Acrs and clustered into 163 sequence similarity-based Acr families. The 163 families were further subject to a structure similarity-based hierarchical clustering, revealing structural similarity between 44 candidate Acr (cAcr) families and 119 known Acr families. The bacterial hosts of these 163 Acr families are mainly from Bacillota, Pseudomonadota, and Bacteroidota, which are all dominant gut bacterial phyla. Many of these 163 Acr families are also co-localized in Acr operons. All the data and visualization are provided on our website: https://pro.unl.edu/AcrDB.