As farmland represents the largest nitrous oxide (N2O) emitter, widely used chiral pesticides frequently coexist with N2O. To explore the underlying molecular mechanisms of chiral pesticides regulating N2O emissions, this study experimented with Paracoccus denitrificans PD1222 on anaerobic/aerobic denitrification under cis-epoxiconazole (EPO) stress. The (+)-EPO increased 13.51-fold N2O emissions under anaerobic denitrification by inhibiting the denitrification genes, while (-)-EPO merely enhanced 4.22-fold N2O emissions through improving electron transfer and nitrous oxide reductase assembly. Anaerobic conditions suppressed glucose metabolism and NADH production, disrupting the energy supply for denitrification under EPO stress. Aerobic conditions hindered electron transfer and intracellular iron transport, reducing denitrifying enzyme activity and causing a further 1.29-10.47-fold increase in N2O emissions and extra nitrite accumulation. These findings revealed that oxygen intensified the ecological risk of cis-epoxiconazole monomers stimulating N2O emissions and underscore the potential risks of agrochemical applications to climate change and ecosystem stability.

The dissemination of antibiotic resistance genes (ARGs) through plasmid-mediated conjugative transfer poses a serious public health threat. Although non-antibiotic pharmaceuticals are known to influence horizontal gene transfer (HGT), their stereoselective effects remain poorly understood. In this study, we investigated the enantioselective effects of naproxen (NAP) on RP4 plasmid-mediated conjugation between Escherichia coli and Pseudomonas aeruginosa. The R- and S-enantiomers of NAP (R-NAP and S-NAP) exhibited markedly different influences on antibiotic resistance gene (ARG) transfer, with R-NAP inducing a substantially higher conjugation frequency than S-NAP. This observation was consistent with transcriptomic and RT-qPCR analyses, which showed stronger upregulation of key conjugation-related genes (e.g., traF, traJ, trfAp, and trbBp) and membrane-associated genes (ompA, ompF, and oprF) under R-NAP exposure. Proteomic analysis further confirmed the upregulation of pilus- and membrane-associated proteins in response to R-NAP exposure. Molecular docking suggested that this stereoselective effect arises from distinct binding interactions between NAP enantiomers and conjugation-associated proteins, with R-NAP forming more stable conformations with core proteins such as TraF, TraJ, OmpA, and OprF. Collectively, these results demonstrate that R-NAP enhances RP4-mediated conjugative transfer by stereoselectively modulating gene and protein expression and strengthening protein-ligand interactions. This study highlights the overlooked role of chiral pharmaceuticals in accelerating ARG dissemination and underscores the need to consider stereoselectivity in environmental risk assessments of non-antibiotic pharmaceuticals.

The enrichment of antibiotic resistance genes (ARGs) and pathogenic microorganisms on microplastics may pose potential risks to public health. In the context of ongoing global warming, however, the mechanisms governing the occurrence and dissemination of ARGs within the riverine plastisphere remain poorly understood. In this study, we investigated the effects of temperature on the plastisphere resistome by cultivating biofilms on PVC (φ = 3 mm) surfaces in river water at 5, 15, 25, and 35 °C. Our results demonstrate that temperature drives shifts in microbial community structure and thereby indirectly regulates ARG composition. Both increases and decreases in temperature enhanced the abundance and potential mobility of ARGs. At low temperatures, human-associated pathogens such as Pseudomonas fluorescens were enriched and predominantly carried ARGs associated with antibiotic efflux, contributing to elevated pathogenic risk. Under these conditions, ARG dissemination was largely governed by stochastic processes mediated by transposases. In contrast, higher temperatures favoured the enrichment of high-risk ARGs and promoted more stable dissemination through the increased prevalence of site-specific recombinases. Although pathogen diversity and abundance declined at elevated temperatures, the remaining hosts harboured ARGs spanning multiple resistance mechanisms. Collectively, these findings demonstrate that temperature is a key driver of both the composition and dissemination potential of the plastisphere resistome, and reveal a shift in ecological risk patterns under ongoing climate warming.

2,2',4,4'-tetrabromodiphenyl ether (BDE-47) is a persistent organic pollutant that poses significant environmental challenges. Microbial transformation plays a crucial role in removing BDE-47. While most studies to date have focused on its degradation using single microbial strains under specific conditions, research on combined degradation approaches remains limited. This study developed a stable, reciprocal synthetic microbial community (SynCom) comprising aerobic and facultative anaerobic bacteria. By integrating experimental validation with molecular sequencing, the study analyzed degradation efficiency, pathways, interspecies interactions, and key genes involved in the process. The findings reveal mechanisms underlying enhanced biotransformation efficiency. In the SynCom, two bacterial strains complemented each other's degradation pathways and gene functions, working synergistically through cross-feeding. This collaboration resulted in a more complete, efficient, and environmentally friendly degradation process compared to individual strain degradation. Specifically, Enterobacter sp. strain XM utilized hydroxylation products generated by Stenotrophomonas sp. strain WZN-1 to cleave ether bonds, producing smaller monocyclic compounds that entered the TCA cycle. This research introduces a novel and highly efficient SynCom for BDE-47 bioremediation and identifies key genes involved in its degradation. These findings offer valuable insights and potential strategies for the bioremediation of BDE-47-contaminated environments.

Antibiotic resistance poses a global threat to public health, with recent studies highlighting the role of nonantibiotic pharmaceuticals in the transmission of antibiotic resistance genes (ARGs). This study provides insights into the comprehensive profile, horizontal gene transfer potential, hosts, and public health risks associated with antibiotic resistomes in river ecosystems exposed to chiral naproxen (NAP). Our findings demonstrate that NAP stress selectively enriches ARGs and mobile genetic elements (MGEs), thereby bolstering bacterial resistance to specific antibiotics. Importantly, the spatial variation of NAP chiral enantiomers influences the enantioselective response of bacterial communities to antibiotics. While (S)-NAP and (R)-NAP exhibit differing degrees of horizontal transfer potential, (S/R)-NAP notably facilitates microbial aggregation and DNA transport via type IV secretion system (T4SS)-related functional genes, promoting the conjugation of sul1. Moreover, (S/R)-NAP promotes the horizontal transfer of ARGs by stimulating ROS production and altering cell membrane permeability. Chiral NAP exposure induces pathogens to acquire ARGs and accelerates the proliferation of Burkholderia. ARG-Rank analysis indicates that the health risk posed by (R)-NAP exposure surpasses that of (S)-NAP, with the highest risk observed when both enantiomers are present. This study elucidates the horizontal transfer and transmission mechanisms of ARGs under chiral NAP stress, underscoring the potential health hazards associated with NAP chiral enantiomers.

Agricultural soil is a major source of N2O emissions due to soil denitrification. However, the influence of pesticides, particularly chiral pesticides, on soil denitrification has often been overlooked. In this study, we investigated the effects and regulatory mechanisms of the widely used chiral pesticide cis-epoxiconazole on denitrification and N2O emissions. The 2R, 3S-(+)-cis-epoxiconazole ((+)-EPO) enantiomer reduced nitrate removal efficiency by 15.04%-54.49% after 3 days and increased N2O emissions by 109.27%-163.68% after 35 days of exposure in a microcosm system, whereas the 2S, 3R-(-)-cis-epoxiconazole ((-)-EPO) enantiomer had minimal impact. At a concentration of 0.5 mg/kg, (+)-EPO inhibited β-glucosidase activity by 34.80% and decreased electron transport system activity by 43.70%, thereby impairing electron generation and transfer during denitrification. Expression of the nosZ gene was suppressed by 53.34%, while nitric oxide reductase and nitrous oxide reductase activities were upregulated by 118.89% and downregulated by 46.70%, respectively. These key regulatory changes directly contribute to the increased N2O emissions observed with (+)-EPO treatment. Additionally, although (+)-EPO caused only minor changes in denitrifying microbial taxa, it increased the relative abundance of Nitrososphaerota and Euryarchaeota by 0.08%-0.10% and 0.40%-0.45%, respectively, after 35 days, enhancing nitrification and indirectly stimulating N2O emissions. Overall, this study deepens our understanding of how cis-epoxiconazole disrupts denitrification in agricultural soils through microbial, genetic, electronic, and enzymatic pathways.

Hexabromocyclododecane (HBCD), a flame retardant classified as a Persistent Organic Pollutant (POP), undergoes stereoisomer-specific microbial transformation with significant environmental and health implications. However, the underlying mechanisms of this stereoisomer-specific microbial transformation remain poorly understood. In this study, high-purity HBCD chiral isomers were isolated using an optimized high-performance liquid chromatography (HPLC) method and their transformation by Acinetobacter hemolyticum sp. strain HW-2 was investigated through transcriptomic analysis. Within three days, strain HW-2 removed (+) α-, (-) α-, (+) β-, (-) β-, (+) γ-, and (-) γ-HBCD with respective removal efficiencies of 52.38 %, 71.08 %, 71.07 %, 63.34 %, 47.47 %, and 77.05 %. Transcriptomic data revealed stereoisomer-specific processes in HBCD transport, response, and transformation. Strain HW-2 upregulated major facilitator superfamily (MFS) transport genes for HBCD uptake, with distinct genes activated for different diastereoisomers. Compared to γ-HBCD, α- and β-HBCD exerted greater stress on strain HW-2, leading to increased expression of efflux genes and antioxidant-related genes. The transformation of HBCD stereoisomers involved distinct functional enzymes, with only (-) γ-HBCD metabolized via the aromatic compound metabolic pathway. This study elucidates the stereoisomeric-specific transformation mechanisms underlying HBCD transformation by strain HW-2, offering valuable insights for theoretical and practical applications in HBCD remediation.

The dissemination of antibiotic resistance genes (ARGs) is a growing global health concern. This study investigates how the chiral enantiomers of the non-antibiotic drug naproxen (NAP) influence ARG dissemination in biofilms. Metagenomic sequencing and binning analyses revealed that NAP enantiomers selectively enriched ARGs and their bacterial hosts, enhancing resistance to specific antibiotics. Notably, the stereoselective effects of NAP enantiomers not only shaped microbial community composition but also affected the potential for ARG spread. Mechanistically, exposure to R-NAP, in comparison to S-NAP, resulted in a 1.53-fold increase in reactive oxygen species (ROS) production, an 18.20 % enhancement in cell membrane permeability, and a 1.93-fold rise in the abundance of genes associated with the type IV secretion system (T4SS). These physiological and genetic changes promoted microbial aggregation and DNA conjugation, particularly enhancing the transfer of the sul1 gene within the Aquabacter genus through the coordinated action of T4SS, two-component systems (TCS), and quorum sensing (QS). Molecular docking and qRT-PCR analyses further revealed that the stereoselectivity of NAP enantiomers stemmed from their distinct binding interactions with proteins involved in horizontal gene transfer, shedding light on the molecular mechanisms underlying ARG dissemination under chiral NAP exposure.

The stereochemistry of antibiotics significantly affects resistance development. With rising antibiotic resistance, nanotechnology is emerging as a promising solution, potentially increasing the likelihood of nanoparticle coexistence with antibiotics in wastewater treatment plants (WWTPs). However, the influence of nanoparticles on chiral antibiotics induced resistance remains unclear. This study employed chemical characterization, metagenomics, and molecular docking to comprehensively elucidate the resistance mechanisms induced by the combination of silver nanoparticles (AgNPs) and chiral antibiotics (ofloxacin: OFL and levofloxacin: LEV). AgNPs significantly altered the abundance and composition of antibiotic and metal resistance genes, as well as the conjugative transfer of the RP4 plasmid among activated sludge communities, with the OFL_AgNPs combination exhibiting a higher resistance potential than LEV_AgNPs. Mechanistic studies revealed that chiral antibiotics selectively interact with AgNPs, promoting Ag⁺ release, enhancing antibiotic uptake, and forming stable complexes. These interactions reshaped microbial functions, including cell membrane permeability, oxidative stress response, quorum sensing, and extracellular secretion, thereby shaping resistance profiles. This study reveals previously overlooked enantiomer-specific risks associated with nanoparticle-antibiotic interactions and provides a foundation for strategies to mitigate ARG dissemination in wastewater treatment and broader environmental systems.

Microplastics are widespread environmental pollutants that pose risks to ecosystems, yet their effects on bacterial and archaeal communities in aquatic ecosystems remain understudied. In this study, we performed a 14-day microcosm experiment combined with metagenomic sequencing to compare bacterial and archaeal responses to a biodegradable microplastic (polylactic acid, PLA) and a non-biodegradable microplastic (polyvinyl chloride, PVC). Microplastics selectively enriched distinct microbial assemblages, with Pseudomonadota and Euryarchaeota identified as the dominant bacterial and archaeal phyla, accounting for 67.83 % and 15.95 %, respectively. Archaeal community in surrounding water were more sensitive to colonization time than bacterial community. Compared to the surrounding water, the plastisphere displayed simpler and more loosely connected microbial networks. Notably, co-occurrence networks of both bacteria and archaea in the PVC plastisphere were predominantly shaped by symbiotic interactions. Both bacteria and archaea carried diverse antibiotic resistance genes (ARGs), but PLS-PM indicated that bacteria were the primary drivers of ARG dissemination (path coefficient = 0.952). While the PVC plastisphere showed higher ARG abundance than the PLA plastisphere, elevated intI1 expression in the PLA plastisphere suggests a potentially greater risk of ARG dissemination associated with PLA microplastics. These findings reveal the distinct effects of PLA and PVC microplastics on microbial communities and highlight the role of microplastics in ARG dissemination, emphasizing their ecological risks in aquatic ecosystems.

Misuse of antibiotics drives the spread of antibiotic resistance genes (ARGs). Although reducing residual antibiotic concentrations can help curb ARG proliferation, the biodegradation and transformation of antibiotic stereoisomers may exacerbate resistance development. However, the impact of antibiotic enantiomers on ARG proliferation remains poorly understood. This study employed metagenomic analysis to investigate the enantiomer-specific selection and resistance risks of chiral antibiotic ofloxacin (OFL) and its (S)-enantiomer, levofloxacin (LEV), in activated sludge. Results showed that LEV primarily promoted the enrichment of ARGs related to aminoglycoside and mupirocin resistance by selecting for pathogenic bacteria carrying virulence factors under high toxicity stress. OFL-driven ARG proliferation involved more diverse mechanisms, including increased gene mobility, co-selection with heavy metals, broader host range, and elevated pathogenicity. The antibiotic resistome risk index (ARRI) further demonstrated a higher environmental risk under OFL treatment than LEV. These findings offer critical insights into the enantioselective resistance risks posed by chiral antibiotics.

Temperate and virulent phages coexist in natural environments and can collaboratively contribute to the lysis of bacterial biofilms. However, their therapeutic potential and the dynamics of phage-biofilm interactions, particularly in clinical contexts, remain poorly understood. In this study, we demonstrated the strong biofilm-lysing capabilities of the temperate phage PaoP1 and virulent phage PaoP5 against Pseudomonas aeruginosa biofilms, highlighting their potential for phage therapy. RNA-seq analysis revealed a shared host resistance mechanism involving the downregulation of flagellar biosynthesis and enhanced biofilm formation. Despite this common host response, the two phages exhibited distinct infection strategies: PaoP1 integrated quiescently into the host genome, while PaoP5 rapidly and abundantly expressed its genes, potentially hijacking the host transcriptional machinery through an as-yet-unknown mechanism. These findings deepen our understanding of phage-biofilm interactions and support the development of phage-based approaches to treat biofilm-associated infections.

IlvA1, a pyridoxal phosphate-dependent (PLP) enzyme, catalyzes the deamination of l-threonine and l-serine to yield 2-ketobutyric acid or pyruvate. To gain insights into the function of IlvA1, we determined its crystal structure from Pseudomonas aeruginosa to 2.3 Å. Density for a 2-ketobutyric acid product was identified in the active site and a putative allosteric site. Activity and substrate binding assays confirmed that IlvA1 utilizes l-threonine, l-serine, and L-allo-threonine as substrates. The enzymatic activity is regulated by the end products l-isoleucine and l-valine. Additionally, the efficiency of d-cycloserine and l-cycloserine inhibitors on IlvA1 enzymatic activity was examined. Notably, site-directed mutagenesis confirmed the active site residues and revealed that Gln165 enhances the enzyme activity, emphasizing its role in substrate access. This work provides crucial insights into the structure and mechanism of IlvA1 and serves as a starting point for further functional and mechanistic studies of the threonine deaminase in P. aeruginosa.

Regulated intramembrane proteolysis (RIP) is a fundamentally conserved mechanism involving sequential cleavage by a membrane-bound Site-1 protease (S1P) and a transmembrane Site-2 protease (S2P). In the opportunistic pathogen Pseudomonas aeruginosa, the alternate sigma factor σ22 activates alginate production and in turn is regulated by the MucABCD system. The anti-sigma factor MucA, which inhibits σ22, is sequentially cleaved via RIP by AlgW (S1P) and MucP (S2P) respectively. In this study, we report high-resolution crystal structures of the MucP PDZ1 and PDZ2 domains. Structural and binding analysis confirms that MucP PDZ2 recognizes the carboxy-terminal Ala136 residue of MucA following Site-1 cleavage by AlgW, while the peptide binding groove of PDZ1 is obstructed by a short α-helix. A structure of MucP PDZ2 with bound MucA peptide shows how PDZ2 binds the newly exposed carboxyl terminus of MucA following AlgW cleavage. The ability of a ΔmucP strain of P. aeruginosa to form biofilms was reduced to a similar extent as a ΔalgW strain. This work paves the way for further studies of MucP and other PDZ-containing S2Ps in regulated intramembrane proteolysis.

Human activities have caused an imbalance in the input nitrogen and phosphorus (N/P) in the biosphere. The imbalance of N/P is one of the characteristics of water eutrophication, which is the fundamental factor responsible for the blooms. The effects of the N/P imbalance on diatom and phycospheric bacteria in blooms are poorly understood. In this study, the N/P molar ratio in real water (14:1) and the predicted N/P molar ratio in future water (65:1) were simulated to analyze the response of Cyclotella sp. and phycospheric bacteria to the N/P imbalance. The results showed that the N/P imbalance inhibited the growth of Cyclotella sp., but prolonged diatom bloom duration. The resistance of Cyclotella sp. to the N/P imbalance is related to phycospheric bacteria, and there are dynamic regulatory mechanisms within the phycospheric bacteria community to resist the N/P imbalance: (1) the increase of HNA bacterial density, the decrease of LNA bacterial density, (2) the increase of phycospheric bacterial diversity and eutrophic bacteria abundance, and the change of denitrifying bacteria abundance, (3) the activity of nitrogen and phosphorus metabolism of HNA bacteria enhanced, while that of LNA bacteria decreased. And the gene hosts of nitrogen and phosphorus metabolism were most enriched in Proteobacteria, indicating that Proteobacteria played an important role in maintaining the stability of phycospheric bacteria and was the dominant phylum resistant to the N/P imbalance. This study clarified that the algal-bacteria system was resistant to the N/P imbalance and implied that the N/P imbalance had little effect on the occurrence of diatom bloom events due to the presence of phycospheric bacteria.

Salinity is a critical environmental factor in marine ecosystems and has complex and wide-ranging biological effects. However, the effects of changing salinity on diversity and ecological functions of high nucleic acid (HNA) and low nucleic acid (LNA) bacteria are not well understood. In this study, we used 16S rRNA sequencing and metagenomic sequencing analysis to reveal the response of HNA and LNA bacterial communities and their ecological functions to salinity, which was decreased from 26 %o to 16 %o. The results showed that salinity changes had significant effects on the community composition of HNA and LNA bacteria. Among LNA bacteria, 14 classes showed a significant correlation between relative abundance and salinity. Salinity changes can lead to the transfer of some bacteria from HNA bacteria to LNA bacteria. In the network topology relationship, the complexity of the network between HNA and LNA bacterial communities gradually decreased with decreased salinity. The abundance of some carbon and nitrogen cycling genes in HNA and LNA bacteria varied with salinity. Overall, this study demonstrates the effects of salinity on diversity , ecological functions and suggests the importance of salinity in regulating HNA and LNA bacterial communities and functions.

In recent years, micro/nanoplastics have garnered widespread attention due to their ecological risks. In this study, we investigated the effects of polystyrene nanoparticles (PS-NPs) of different sizes on the growth and biofilm formation of Pseudomonas aeruginosa PAO1. The results demonstrated that exposure to certain concentrations of PS-NPs significantly promoted bacterial biofilm formation. Meanwhile, we comprehensively revealed its mechanism whereby PS-NPs induced oxidative stress and altered bacterial membrane permeability by contacting or penetrating bacterial membranes. To counteract the stimulation by PS-NPs and reduce their toxicity, bacteria enhanced biofilm formation by upregulating the expression of biofilm-related genes, increasing EPS and virulence factors secretion, and enhancing bacterial motility through the participation of the quorum sensing (QS) system. Additionally, we also found that exposure to PS-NPs enhanced bacterial antibiotic resistance, posing a challenge to antimicrobial therapy. Our study reveals the toxic effects of nanoplastics and the defense mechanisms of bacteria, which has important implications for the risk assessment and management of environmental nanoplastics.

In eukaryotic cells, the synthesis, processing, and degradation of mRNA are important processes required for the accurate execution of gene expression programmes. Fully processed cytoplasmic mRNA is characterised by the presence of a 5′cap structure and 3′poly(A) tail. These elements promote translation and prevent non-specific degradation. Degradation via the deadenylation-dependent 5′-3′ degradation pathway can be induced by trans-acting factors binding the mRNA, such as RNA-binding proteins recognising sequence elements and the miRNA-induced repression complex. These factors recruit the core mRNA degradation machinery that carries out the following steps: i) shortening of the poly(A) tail by the Ccr4-Not and Pan2-Pan3 poly (A)-specific nucleases (deadenylases); ii) removal of the 5′cap structure by the Dcp1-Dcp2 decapping complex that is recruited by the Lsm1-7-Pat1 complex; and iii) degradation of the mRNA body by the 5′-3′ exoribonuclease Xrn1. In this review, the biochemical function of the nucleases and accessory proteins involved in deadenylation-dependent mRNA degradation will be reviewed with a particular focus on structural aspects of the proteins and enzymes involved.

Pseudomonas aeruginosa is capable of causing acute and chronic infections in various host tissues, which depends on its abilities to effectively utilize host-derived nutrients and produce protein virulence factors and toxic compounds. However, the regulatory mechanisms that direct metabolic intermediates towards production of toxic compounds are poorly understood. We previously identified a regulatory protein PvrA that controls genes involved in fatty acid catabolism by binding to palmitoyl-coenzyme A (CoA). In this study, transcriptomic analyses revealed that PvrA activates the Pseudomonas quinolone signal (PQS) synthesis genes, while suppressing genes for production of polyhydroxyalkanoates (PHAs). When palmitic acid was the sole carbon source, mutation of pvrA reduced production of pyocyanin and rhamnolipids due to defective PQS synthesis, but increased PHA production. We further solved the co-crystal structure of PvrA with palmitoyl-CoA and identified palmitoyl-CoA-binding residues. By using pvrA mutants, we verified the roles of the key palmitoyl-CoA-binding residues in gene regulation in response to palmitic acid. Since the PQS signal molecules, rhamnolipids and PHA synthesis pathways are interconnected by common metabolic intermediates, our results revealed a regulatory mechanism that directs carbon flux from carbon/energy storage to virulence factor production, which might be crucial for the pathogenesis.