The toxin-antitoxin (TA) system is ubiquitous in bacteria and archaea. A typical TA system generally consists of a toxin that inhibits bacterial growth and an antitoxin that neutralizes toxin toxicity. At present, TA systems are classified into types I-VIII, of which type II system is the most extensively studied. In addition, researchers have found that there are atypical TA systems such as monocistronic TA systems and three-component TA systems. After the discovery of the first TA system (CcdB/CcdA) in the 1980s, TA systems have been shown to play a key role in the physiological processes of microorganisms. In this paper, we review the research results about the roles of TA systems in resisting bacteriophage infections in recent years and summarize the neutralization mechanisms. In particular, we brief how TA systems specifically sense the invading bacteriophages and the underlying molecular mechanisms, aiming to provide reference for the research on the roles and regulation mechanisms of unknown TA systems in the future.

In the post-antibiotic era, phage therapy becomes a candidate for treating drug-resistant bacteria. Phages are diverse, and jumbo phages are a class of phages with genomes longer than 200 kb. Their large genomes carry abundant functional genes with scattered distribution. Jumbo phages have a variety of unique biological characteristics, such as oversized phage particles, unique replication cycles, and unique structures such as the nucleus-like compartment, the “inner body”, and the long-wavy curly tail fiber. This paper reviews the research progress in jumbo phages, focusing on their biological characteristics, genomes, evolution, and special replication mechanisms and structures. Furthermore, this review explores the application potential of jumbo phages in the treatment of drug-resistant bacterial infection, environmental management, aquaculture, and biocontrol, aiming to provide reference and implications for the research and application of jumbo phages.

Heavy metal pollution has attracted increasing attention because of its toxicity, environmental persistence, bioaccumulation, and potential contribution to antibiotic resistance. The accurate, rapid, efficient, and sensitive detection of heavy metals in the environment is of great significance for environmental protection and human health. A whole-cell microbial biosensor (WCMB), integrating a biorecognition module and a signal processing module, provides a new strategy for heavy metal monitoring. Great progress has been achieved in the transcription factor-based WCMBs for the monitoring of heavy metals in recent years. This paper introduced the basic composition and design principles of WCMBs, summarized the construction and application of WCMBs for heavy metal detection that were developed in recent years, analyzed the optimization strategies of WCMBs by synthetic biology, and finally prospected the future research directions of WCMBs. This paper is expected to be an important reference for the effective prevention and control of heavy metal pollution in the environment.

[Objective] To elucidate the phylogenetic position and mine the gene resources for synthesis of secondary metabolites from Streptomyces sp. YH02, a Gram-positive actinomycete strain isolated from the soil sediment of Yuncheng Salt Lake in Shanxi. [Methods] Illumina and PacBio platforms were used for whole genome sequencing of YH02, which was followed by gene prediction, functional annotation, prediction of secondary metabolite synthetic gene clusters (BGCs), comparative genomic analysis, and morphological, physiological, and biochemical characterization. [Results] The YH02 genome was a linear chromosome spanning 8 285 116 bp, with the G+C content of 71.77% and 7 237 open reading frames. Gene annotations in the GO, COG, KEGG, and CAZy identified 2 829, 5 478, 4 805, and 279 genes, respectively. The subcellular localization analysis predicted various secretion system-related proteins and 1 030 transporters. Additionally, 32 secondary metabolite BGCs were predicted in strain YH02, involving the synthesis of various natural products such as terpenoids, non-ribosomal peptides, polyketides, and ribosomally synthesized and post-translationally modified peptides. The comparative genomic analysis revealed 15 739 pan-genome orthologous gene clusters and 4 267 core genome orthologous gene clusters. The phylogenetic analysis based on the 16S rRNA gene sequence revealed a proximate phylogenetic affiliation between strain YH02 and Streptomycesvenezuelae ATCC 10712 as well as Streptomyceszaomyceticus NBC 00278. However, the average nucleotide identity (ANI) value was below the threshold of 95.00%, and the digital DNA-DNA hybridization (dDDH) value was less than 70.00%. YH02 exhibited light pink aerial mycelia on the ISP 2 medium. It showed significant differences in tolerance to pH, sodium chloride, and growth temperature compared with its closely related strains. Additionally, this strain demonstrated weak starch hydrolysis activity, positive gelatin liquefaction, positive nitrate reduction, and slow milk coagulation. [Conclusion] Based on the findings from genomic, physiological, and biochemical analyses, strain YH02 is confirmed as a potential new species of Streptomyces. This study not only enriches the microbial resource pool but also provides a theoretical basis and potential genetic resources for mining the natural products with unique mechanisms of action.

[Objective] To identify mazEF family type II toxin-antitoxin systems of Leuconostocpseudomesenteroides L64 and to elucidate the molecular roles of the mazEF systems in the host exposed to environmental acid stress. [Methods] Putative MazF toxins were induced alone or co-expressed with their cognate antitoxins in Escherichiacoli. The toxic effect of MazF on bacterial growth and the antitoxic effects of cognate antitoxins were examined. The lacZ reporter system and electrophoretic mobility shift assay (EMSA) were used to decipher the auto-regulation mechanism of the mazEF system invivo and invitro. The putative target genes regulated by MazE were predicted and validated through invivo and invitro experiments. [Results] Among the three putative mazEF systems in L. pseudomesenteroides L64, mazEF1-Leup (OYT_01690-OYT_01685) encoded a functional type II toxin-antitoxin system. MazE1-Leup (OYT_01685) inhibited mazEF1-Leup transcription by binding to the palindromic sequence (TAACAaaatgTGTTA) in the promoter. In addition, MazE1-Leup inhibited transcription of the dlt-acpS-alr operon by binding to the similar palindromic sequence (TAACAtattgaaatatatgTGTTA) in the promoter of dlt-acpS-alr. [Conclusion] mazEF1-Leup (OYT_01690-OYT_01685) encodes a functional mazEF family type II toxin-antitoxin system. Beyond regulating its own operon, MazE1-Leup regulates the transcription of dlt-acpS-alr and finally assists L. pseudomesenteroides L64 in response to low acid stress.

[Objective] The biological regulatory mechanisms of soil microorganisms in response to single heavy metals or organic pollutants have been extensively studied, while the biological mechanisms of microbial responses to combined pollution remain unclear. This study aims to reveal the biological mechanisms of Enterococcus faecalis HHT-1 in response to single and combined stress of cadmium (Cd) and aniline (AN) by transcriptomics. [Methods] Transcriptomics was employed to explore the transcriptional regulation of HHT-1 under single (Cd: 150 mg/L; AN: 2 g/L) and combined stress (150 mg/L Cd and 2 g/L AN) at half inhibitory concentrations (IC₅₀). [Results] Under single Cd stress, HHT-1 upregulated the expression of genes encoding metal-binding proteins and transporters to promote the sequestration or efflux of Cd²⁺. High concentrations of Cd induced oxidative stress in cells, and HHT-1 upregulated the expression of ribosome-related genes and nucleotide-related genes to repair the protein damage and DNA damage caused by oxidative stress, and cleared intracellular reactive oxygen species (ROS) by producing catalase. Moreover, Cd stress upregulated the expression of genes related to virulence and antibiotic resistance in HHT-1. Under the single stress of AN, HHT-1 activated the expression of AN-degrading enzyme genes to reduce its toxicity and upregulated the expression of genes encoding efflux pumps to excrete AN from the cell. AN also caused intracellular oxidative stress, and HHT-1 cleared the ROS by upregulating the expression of glutathione synthase genes. Under the combined stress of Cd and AN, HHT-1 exhibited a complex array of adaptive mechanisms. Initially, HHT-1 upregulated the expression of genes encoding metal transporters and efflux pumps to transport Cd and AN out of the cell, thereby mitigating their toxic effects on the cell. At the same time, HHT-1 cleared intracellular ROS via the dual pathway of catalase and glutathione. In addition, HHT-1 upregulates the expression of virulence and antibiotic resistance genes under combined stress, which may enhance its pathogenicity and resistance. It is worth noting that HHT-1 did not significantly upregulate the expression of AN-degrading enzyme-related genes under the combined stress. [Conclusion] The transcriptional regulation mechanism of E. faecalis HHT-1 in response to the combined stress of Cd and AN is a combination of the transcriptional regulation mechanisms observed under single Cd stress and single AN stress, and it is more similar to the mechanism observed under single Cd stress. HHT-1 mainly responds to the combined stress of Cd and AN by enhancing cell wall synthesis, Cd²⁺ efflux, DNA repair, and ROS scavenging. Genes associated with efflux pumps and glutathione under AN stress continue to be expressed under the combined stress. Both single Cd stress and combined stress of Cd and AN may increase the potential pathogenicity and drug resistance of HHT-1.

[Objective] To study the effect of Alistipesfinegoldii (AF) on inflammatory bowel disease (IBD) and the underlying mechanism. [Methods] Six-week-old male C57BL/6J mice were administrated with streptomycin for three days and then randomly assigned into the control, phosphate buffered saline (PBS), and AF groups. Mice were administrated with AF suspension (1×10⁹ CFU, 200 μL per mouse) or PBS by gavage for two weeks, followed by drinking of the water containing 2.5% dextran sulfate sodium (DSS) for one week for the modeling of colitis. The weight loss fraction percentage, fecal characteristics, blood fecesstools, and colon length were determined. The colon tissue was stained with hematoxylin-eosin for the scoring of histopathological changes, and feces samples were collected at the beginning and end of the experiment for sequencing of 16S rRNA gene amplicons at the beginning and end of the experiment. The mRNA levels of colon tissue-associated intestinal barrier proteins and inflammatory mediators were determined by qPCR. [Results] The mice in the AF group had severer disease conditions than those in the PBS group regarding the weight loss percentage, disease activity index, colon shortening, and histopathological score. Compared with the PBS group, the AF group showed down-regulated mRNA levels of occludin and claudin 5 and up-regulated mRNA level of interleukin (IL)-17A. The AF group had lower alpha diversity of intestinal flora than the PBS group, and the beta diversity showed significant differences between AF and PBS groups. The linear discriminant analysis effect size (LEfSe) results revealed that the significantly differential bacteria between AF and PBS groups were Bacilli, Erysipelotrichales, Erysipelotichaceae, Odoribacter, Marinifilaceae, Dubosiella, and Dubosiellanewyorkensis. [Conclusion] AF promotes the secretion of inflammatory mediators, impairs the permeability of the intestinal mucosa, and alters the structure and diversity of the intestinal flora, thereby promoting the development of IBD.

[Objective] To identify the active components in Glycyrrhiza uralensis Fisch. that inhibit methicillin-resistant Staphylococcus epidermidis (MRSE) infections and explore their potential antibacterial mechanisms. [Methods] The half-dilution method was employed to assess the inhibitory activities of pharmacological components from G. uralensis against MRSE. The anti-MRSE phenotype of this medicinal herb was evaluated by microbial adhesion to hydrocarbons, crystal violet staining, scanning electron microscopy, and integrated cell imaging. Additionally, metabolomic analysis was conducted via gas chromatography-mass spectrometry (GC-MS), and the activity of intracellular oxidative dehydrogenase was measured by a commercially available reagent kit. The propidium iodide and laurdan dyes were utilized to assess the membrane damage and fluidity of cells. The challenge test was conducted with the larvae of Galleria mellonella to determine the antibacterial activities of tested pharmacological components in vivo. [Results] Licochalcone A, licochalcone C, and glabridin from G. uralensis demonstrated significantly inhibitory activities against MRSE. Among these compounds, licochalcone A exhibited the strongest inhibitory effect on MRSE, with a minimum inhibitory concentration (MIC) of 6.0 μg/mL and a minimum bactericidal concentration (MBC) of 12.0 μg/mL. The metabolomic analysis indicated that licochalcone A primarily influenced the metabolic pathways, secondary metabolite biosynthesis, and ATP-binding cassette (ABC) transport systems of MRSE. This compound impeded the biosynthesis of ornithine, lysine, and niacin, leading to the accumulation of 1,3-dipalmitin in the cells. Phenotypic experiments corroborated that licochalcone A downregulated the tricarboxylic acid (TCA) cycle flux and reduced the intracellular ATP level in MRSE. Furthermore, it inhibited the biofilm formation and intracellular protein expression, thereby preventing MRSE from adhering to HaCaT cells. Additionally, licochalcone A disrupted the structural integrity of the MRSE cell membrane, resulting in cell collapse, deformation, and even rupture and increased the survival rate of G. mellonella larvae following MRSE infection. [Conclusion] Exposure to licochalcone A alters the metabolism of sugars, lipids, and amino acids in MRSE cells, thereby influencing the biofilm formation, biosynthesis of secondary metabolites such as proteins, and the remodeling of cell membranes. Consequently, this alteration results in an antimicrobial phenotype characterized by decreased ATP production, impaired transporter function, and reduced adhesion and infection of MRSE.

[Objective] To develop a safe, effective, and identifiable new vaccine. [Methods] The gene deletion strain S2Δbp26 was constructed by homologous recombination with S2 as the parental strain. The stability, safety, and efficacy of S2Δbp26 were then evaluated. [Results] The phenotype and genotype of S2Δbp26 did not change after 20 successive passages invitro. The experiments in mice and guinea pigs showed that there was no difference in biosafety between S2Δbp26 and S2. The bacterial load was less than 2×10⁵ CFU per gram of spleen in guinea pigs inoculated with S2Δbp26 at different passages and S2, which suggested that the virulence of the mutant strain was attenuated. The mice inoculated with S2ΔBP26 had a shorter 50% recovery time than those inoculated with S2. Meanwhile, the mice immunized with S2Δbp26 could successfully resist the challenge with M28 wild-type strain at a dose of 2×10⁵ CFU/mouse. [Conclusion] S2Δbp26 was successfully constructed, with excellent safety and immunoprotective capacity, which provided technical reserves for the development of identifiable vaccines for brucellosis.

[Objective] To explore the potential mechanism by which Prevotella copri promotes atherosclerosis (AS) from the perspective of host-gut microbiota-metabolism. [Methods] ApoE^-/- mice were randomized into four groups (n=8): control group (Chow group, fed with a normal diet), model group (AS group, fed with a high-fat diet), low-concentration P. copri group (P. copri-low group, administrated with P. copri at 10⁹ CFU/mL by oral gavage daily from the first day of feeding with the high-fat diet), and high-concentration P. copri group (P. copri-high group, administrated with P. copri at 10¹¹ CFU/mL by oral gavage daily from the first day of feeding with the high-fat diet). The body weight was measured and recorded weekly to evaluate the weight gain trend. After 5 weeks, oil red O staining was employed to evaluate the aortic plaque area, and enzyme-linked immunosorbent assay (ELISA) was employed to measure lipid levels, on the basis of which the impact of P. copri on AS progression was assessed. Additionally, qPCR was used to detect the abundance of P. copri in the gut, and untargeted metabolomics was employed to analyze the metabolite changes in the feces of mice. [Results] Compared with the Chow group, the AS group showed increases in the body weight, aortic plaque area, and plasma levels of low-density lipoprotein cholesterol (LDL-C), total cholesterol (TC), and triglycerides (TG) and a decline in the high-density lipoprotein cholesterol (HDL-C) level. The abundance of P. copri in the gut showed no significant difference between the P. copri-low group and the P. copri-high group, indicating that P. copri successfully colonized the gut in both groups. Based on this, the P. copri-low group was selected as the standard concentration group (P. copri group) for further analysis. Compared with the AS group, P. copri colonization in the gut significantly increased the body weight and aortic plaque area and exacerbated dyslipidemia. Metabolomic analysis revealed that P. copri transplantation led to significant increases in the content of several metabolites, including Cer(d18:1/18:1(9Z)), N-palmitoylsphingosine, genistein, adenine, and linoleic acid. KEGG pathway enrichment analysis further indicated that P. copri might contribute to the development and progression of AS through key pathways such as the regulation of ABC transporters, bile acid metabolism, and neuroactive ligand-receptor interactions. [Conclusion] P. copri may exacerbate inflammation and lipid metabolism imbalance by regulating sphingolipid signaling, purine metabolism, and linoleic acid metabolism, thereby promoting the progression of AS.