[Objective] To understand the molecular functions and potential applications of the significantly up-regulated gene cluster chr1_2605-chr1_2604 in response to tetrabromobisphenol A (TBBPA) stress, we investigated the roles of chr1_2605 and chr1_2604 in the specific recognition and efficient degradation of TBBPA. [Methods] Synthetic biology methods were employed to construct Sphingobiumxenophagum C1 (pBBR-2605-HiBiT) and Escherichiacoli BL21(DE3, pET30b-2604) as chassis cells for biosensing and degrading, respectively. The response characteristics of Chr1_2605 in the chassis cells to different pollutants were analyzed by the luciferase activity assay. Additionally, the degradation activity of TBBPA by Chr1_2604 in the chassis cells was determined by high-performance liquid chromatography. [Results] The xenobiotic-responsive element Chr1_2605 exhibited a highly specific response to TBBPA. The Chr1_2605-based chassis cell of S. xenophagum C1 (pBBR-2605-HiBiT) demonstrated high responsivity and sensitivity to TBBPA, with a limit of detection ranging from 0.010 to 0.050 μmol/L. The 2-oxoglutarate/Fe-dependent dioxygenase Chr1_2604 in the chassis cell of E. coli BL21(DE3, pET30b-2604) displayed the degradation rate of 44.415% for 2.0 mg/L TBBPA within 3 d [0.296 mg/(L·d)], which was significantly higher than those of most reported microbial strains under non-co-metabolic conditions. [Conclusion] The chr1_2605-chr1_2604 gene cluster can accurately recognize and degrade TBBPA. Specifically, the xenobiotic-responsive element Chr1_2605 specifically recognizes TBBPA, whereas the 2-oxoglutarate/Fe-dependent dioxygenase Chr1_2604 efficiently degrades TBBPA.

[Objective] Conventional culture methods can merely uncover a fraction of microbial diversity. A vast number of microorganisms remain unculturable under laboratory settings. The advent of in-situ culture technology offers a key solution to this predicament. This study endeavors to innovate the in-situ culture technology, explore microorganisms under hitherto unknown culture conditions, and probe into their potential applications within the realm of uncultured microorganisms. [Methods] PCR tubes were employed as the core of the device, and the idea of sorting prior to culture was introduced. Microorganisms were separated from the environment by means of polymer membranes for independent culture. The efficacy of the device was validated through the pure culture of Escherichia coli. Moreover, this device was applied to in-situ culture in diverse environments such as soil, sewage, and mountain spring water. [Results] The abundance of E. coli increased significantly in the pure-culture device. When the concentration of the polymer membrane reached 15%, the encapsulation effect effectively prevented the escape of E. coli. The co-culture experiments further corroborated this finding. In the in-situ culture experiments, the device designed in this study successfully cultured single-cell microorganisms from sewage, soil, and mountain spring water samples. Sequencing results indicated that this device could culture species recalcitrant to be cultured in the laboratory. Comparison with the NCBI database verified that new species were successfully cultured, which demonstrated the effectiveness of the culture device in various environments. [Conclusion] The culture method designed in this study is suitable for single-cell microbial culture, enrichment of specific microbial communities, and co-culture of multiple microorganisms. The device can isolate and culture richer and more microorganisms than conventional culture methods. This new technology not only isolates and cultures more microorganisms but also manages to culture those previously unculturable under laboratory conditions. It holds great significance for microbiological and ecological research.

[Objective] To explore the autophagy induced by the African swine fever virus (ASFV) protein E423R. [Methods] The RFP-LC3 fluorescent spots following the transfection of HeLa-Difluo^TM hLC3 cells with pCMV-Myc-E423R was quantified by the High-Content Analysis System, and the autophagic flux was assessed. The expression levels of key autophagy proteins, LC3-Ⅱ and SQSTM1/p62, in HeLa cells were analyzed via Western blotting. Co-localization of autophagosomes and lysosomes was examined by laser confocal microscopy. Additionally, Western blotting was employed to investigate the dose-dependent effect of E423R on LC3-Ⅱ expression and the regulatory effect of E423R on the AKT/mTOR/ULK1 signaling pathway. [Results] The High-Content Analysis System demonstrated a significant increase in RFP-LC3 fluorescence spots in the reporter cells expressing E423R, suggesting that E423R induced the activation of autophagy. Western blotting further confirmed that the expression of E423R significantly elevated the LC3-Ⅱ/β-actin ratio while decreasing the expression level of p62. Confocal microscopy results indicated that E423R enhanced the expression of GFP-LC3, promoted the co-localization of GFP-LC3 with Lyso-Tracker Red, and facilitated the fusion of autophagosomes and lysosomes. Additionally, the expression level of E423R exhibited a positive correlation with the expression level of LC3-Ⅱ in HeLa cells. Furthermore, E423R down-regulated the expression of p-AKT, p-mTOR, and p-ULK1 (Ser757) in the mTOR signaling pathway. [Conclusion] This study demonstrates that the ASFV E423R protein induces complete autophagy via the AKT/mTOR/ULK1 signaling pathway, exhibiting a dose-dependent effect on LC3-Ⅱ expression to a certain degree. These findings provide a foundation for further investigation into the infection and pathogenic mechanisms of ASFV.

Hydroxylamine, as an important intermediate product in the nitrogen cycle, connects ammonia oxidation and nitrite oxidation, influencing the velocities and directions of processes like ammonia oxidation, nitrite oxidation, and denitrification. Because of the close associations with the generation of N₂O through enzymatic reactions and self-decomposition or reactions with other substances, hydroxylamine has become a focus and hotspot of research. This paper summarized the generation and transformation of hydroxylamine in autotrophic and heterotrophic ammonia oxidation, the key role of hydroxylamine in the nitrogen cycle, and the promoting effect of hydroxylamine on N₂O emissions. It analyzed the processes of autotrophic and heterotrophic ammonia oxidation and their enzymatic differences, aiming to provide a theoretical reference for in-depth research on the role of hydroxylamine in the microbial nitrogen cycle and for the research and development of measures to reduce N₂O emissions and protect the atmospheric environment.

This article explores the dynamic regulation of fluid shear stress on the biofilm formation of Streptococcus mutans, aiming to elucidate the pathogenic mechanism of this bacterium and provide references for the prevention of dental caries. S. mutans is a major cariogenic bacterium in the oral cavity, and it destroys the tooth tissue by forming biofilms and producing acids by metabolism. Studies have shown that fluid shear stress can regulate the physical structure, secretion of extracellular polymeric substances, adhesion, and quorum sensing of biofilms, thereby affecting the pathogenicity. This study provides a theoretical basis for deeply understanding the ecological adaptability of pathogens in fluid environments and the development of new intervention measures in clinical practice.

The vpa1443-vpa1445 gene cluster (mfpABC) encoding the membrane fusion protein (MFP) is hypothesized to be involved in the biofilm formation of Vibrio parahaemolyticus (Vp). The gene mfpA (vpa1445) encodes a Ca²⁺-binding extracellular protein containing a repeats-in-toxin (RTX) domain, while its function is still under exploration. [Objective] To study the influences of mfpA mutation on the biofilm formation and motility of Vp. [Methods] The single mutants of three genes in the mfpABC gene cluster were constructed, and the Vp motility was compared between the three mutants and the wild type. Furthermore, the influences of mfpA mutation on bacterial motility and biofilm formation were analyzed in detail, and the expression of related genes was analyzed by RT-qPCR. Moreover, the cytotoxicity of ΔmfpA to HeLa cells was investigated. [Results] The mutation of mfpA significantly reduced the swimming and swarming motility of Vp. Crystal violet staining and scanning electron microscopy results showed that the mutation of mfpA enhanced the biofilm formation and increased the content of extracellular polysaccharides and proteins. RT-qPCR confirmed that the expression levels of flagellar genes were downregulated, while those of extracellular polysaccharide synthesis-related genes were upregulated in ΔmfpA. The cytotoxicity of ΔmfpA significantly decreased compared with that of wild type. [Conclusion] The mutation of mfpA can affect the expression of genes associated with flagella and extracellular polysaccharides to reduce the motility, enhance the biofilm formation, and attenuate the cytotoxicity of Vp.

[Objective] To isolate and screen the salt-tolerant strains with good plant growth-promoting effect from the saline-alkali soil of the Yellow River Delta, thus providing strain resources for the efficient cultivation of crops in saline-alkali soil. [Methods] Bacillus strains were isolated by the dilution coating method, and the strains with good plant growth-promoting effects were further selected by the seed soaking test. The growth-promoting characteristics of the selected strains were measured, and the growth-promoting effects of the strains on Sesbania cannabina under salt stress was evaluated by pot experiments. The strain with the strongest plant growth-promoting effect was identified based on morphological characteristics, physiological and biochemical characteristics, and molecular biological evidence. Through the whole genome sequence analysis, the genes related to the plant growth-promoting function were discovered. [Results] A total of 60 Bacillus strains were isolated, from which strains M4, M5, B5, L3, and Q17 were screened out by the seed soaking test. These strains showed robust plant growth-promoting properties, being capable of solubilizing inorganic phosphorus, solubilizing potassium, and producing inole-3-acetic acid. The pot experiment results showed that under normal culture conditions and under low salt stress (NaCl concentration of 100 mmol/L), inoculation with Bacillus increased the plant height, maximum leaf area, stem dry weight, and root dry weight of S. cannabina seedlings (P<0.05). Under high salt stress (NaCl concentration of 200 mmol/L), the fresh and dry weights of stem and leaves of S. cannabina seedlings were increased by inoculation with five strains of Bacillus (P<0.05). In addition, inoculation with Bacillus enhanced the activities of catalase, superoxide dismutase, and peroxidase while reducing the content of malondialdehyde in the leaves of S. cannabina seedlings (P<0.05). Strain M4 with the strongest plant growth-promoting effect was identified as Bacillus thuringiensis. [Conclusion] All the five isolates have various plant growth-promoting properties, being capable of promoting the growth of S. cannabina seedlings under salt stress and alleviating the inhibitory effect of salt on the seedlings. Strain M4 with the robust plant growth-promoting effect is identified as B. thuringiensis, and it has the potential to be developed as plant growth-promoting bacterial fertilizer for saline-alkali soil.

[Objective] To isolate and identify a strain of foot-and-mouth disease virus (FMDV) from bovine oropharyngeal fluid (OPF) and subsequently analyze the amino acid mutations in antigenic proteins and the proliferative properties of the isolated virus. [Methods] RT-PCR and sequencing were performed to identify the FMDV-positive OPF samples. The FMDV strain was isolated from bovine OPF samples by cell inoculation and identified by indirect immunofluorescence assay. The whole genome sequence of the strain was obtained by fragment amplification and sequencing. The amino acid sequences were compared to reveal the amino acid mutations in the antigenic proteins of the strain. The proliferative properties of the isolated FMDV strain were analyzed by the plaque forming assay and one-step growth curve. [Results] A FMDV strain O/FMDV/OP/2022/B was successfully isolated from bovine OPF, belonging to serotype O and Ind-2001 lineage. Amino acid mutations were identified in the antigenic proteins VP1, VP2, and VP3 of O/FMDV/OP/2022/B. In addition, the plaque of O/FMDV/OP/2022/B in BHK-21 cells was smaller than that of the vaccine strain O/FMDV/HN/93. The one-step growth curves showed that O/FMDV/OP/2022/B had significantly lower replication titer than O/FMDV/HN/93 in BHK-21 cells. [Conclusion] A strain of FMDV was isolated from bovine OPF, and it displayed amino acid mutations in antigenic proteins and had lower proliferative capacity than the vaccine strain O/FMDV/HN/93. This study contributes to a better understanding of the mechanism of persistent FMDV infection in the bovine oropharynx and provides a reference for the prevention and control of FMD.

[Objective] To study the cytopathic effects (CPE) and replication characteristics of HCoV-OC43 VR1558 strain in cell lines derived from different species. [Methods] Thirteen cell lines from humans (MRC-5, HRT-18, Huh7, Huh7.5, RD, and HeLa), non-human primates (LLC-MK2 and Vero), and rodents (17Cl-1, Mv.1Lu, BHK-21, BHK-21-APN, and Neuro 2a) were selected and infected with HCoV-OC43 VR1558 strain. CPE were observed for several consecutive days. Virus-infected cells and supernatants were collected daily. RT-qPCR was conducted to monitor the changes in viral RNA copy number. The load of viruses with infectious ability was determined based on the tissue culture infectious dose 50% (TCID₅₀), and the kinetic curves of viral replication in different cell lines were established. [Results] Following infection with HCoV-OC43 VR1558 strain, CPE were observed in all the 13 cell lines. CPE primarily manifested as cell aggregation, shrinkage, rounding, and detachment. CPE appeared early in MRC-5, Mv.1Lu, HeLa, and 17Cl-1 cells, being noticeable within 72 h post-infection (hpi). The virus induced CPE in other cell lines after 120 hpi, and CPE were the mildest in HRT-18 and Huh7.5 cells. RT-qPCR results indicated that the viral RNA copy number increased most significantly within 24 hpi, although the time to reach the peak and the peak copy number varied among cell lines. Specifically, the RNA copy number in Huh7.5 cells reached the peak (10⁷ copies/mL) at 24 hpi, and that in 17Cl-1, BHK-21-APN, Mv.1Lu, and BHK-21 cells reached the peaks (10⁷ to 10⁹ copies/mL) at 48 hpi. In MRC-5, LLC-MK2, Neuro 2a, and Vero cells, the replication peaks (10⁶ to 10⁹ copies/mL) occurred at 72 hpi. In HRT-18, HeLa, Huh7, and RD cells, the viral RNA copy number peaked after 96 hpi, reaching 10⁸ to 10⁹ copies/mL. TCID₅₀ assay results demonstrated rapid viral proliferation within 24 hpi, while the time to reach the peak titer and the peak titers varied. The peak titer (2.68× 10⁷ TCID₅₀/mL) in BHK-21-APN cells was observed at 48 hpi. In BHK-21 and Neuro 2a cells, the peak titers (10⁶ to 10⁷ TCID₅₀/mL) were observed at 72 hpi. In MRC-5, 17Cl-1, HeLa, Huh7, Huh7.5, Mv.1Lu, LLC-MK2, and RD cells, the peak titers (10⁶ to 10⁸ TCID₅₀/mL) were observed at 96 hpi. In Vero cells, the virus strain reached the peak titer (10⁵ TCID₅₀/mL) at 120 hpi, while the strain reached the peak titer of 10⁸ TCID₅₀/mL in HRT-18 cells at 144 hpi. [Conclusion] HCoV-OC43 VR1558 strain exhibits a wide spectrum of cell tropism, demonstrating rapid replication and proliferation within 24 hpi across 13 cell lines derived from various species. However, the time to reach the replication peak varied among different cell lines. The highest viral titer achieved was 10⁸ TCID₅₀/mL, observed in MRC-5 and HRT-18 cells. This study provides experimental reference for further investigation of the replication characteristics, infection mechanism, and pathogenicity of HCoV-OC43, as well as for the screening and evaluation of antiviral drugs.

Currently, the drugs available for treating fungal infections show limited number and off-target effects. Thus, there is an urgent need to develop new antifungal drugs. Fungal apoptosis-like cell death (ALCD) is a cell death phenomenon that occurs during the normal development stage of organisms. This article summarizes the characteristics, signaling pathways, and key factors involved in fungal ALCD and introduces the natural and synthetic drugs that can induce fungal apoptosis. The natural drugs include lipid peptides, farnesol, statins, and alkaloids from microorganisms, organic acids and essential oils from plants, and melittins from insects. Furthermore, this article establishes the basic molecular landscape of drug-induced fungal ALCD. This article provides a theoretical basis for formulating a new strategy for resisting pathogenic fungi and developing targeted antifungal drugs.