[Objective] Yersinia enterocolitica is the primary pathogenic agent of yersiniosis, and it is one of the major foodborne pathogens worldwide. The purpose of this study is to isolate and identify a virulent phage against Y. enterocolitica and explore its application potential. [Methods] A novel virulent phage, named Yersinia phage Yen-yong1 (abbreviated as Yen-yong1), was isolated from the sewers of Lulin Market in Beilun District, Ningbo City, Zhejiang Province, China, by the double-layer agar method with Y. enterocolitica CICC 10869 as the indicator host. The morphology, host range, genome sequence, gene functions, phylogenetic position, physical and chemical tolerance, and biofilm removal ability of Yen-yong1 were studied. Additionally, the sterilization effect of Yen-yong1 on Y. enterocolitica in raw pork slices was evaluated. [Results] Yen-yong1 clarified the turbid CICC 10869 culture within 1.5 h. Yen-yong1 was Siphovirus-like, consisting of an elliptical head with a length of (83.32±0.15) nm and a width of (59.57±2.28) nm and a long tail with a length of (168.04±3.21) nm. The double-stranded DNA genome of Yen-yong1 was 51 321 bp, with the G+C content of 47.80% and 93 open reading frames (ORFs). No known lysogeny-associated genes, resistance genes, or virulence genes detrimental to application were found in Yen-yong1. The highest nucleotide sequence similarity between Yen-yong1 and the other phages in public databases was 20.64%, which was <70.00% (the threshold to define a genus). Yen-yong1 shared 20.00% of core genes with the closest relative, which was lower than the ICTV criteria for subfamily classification (27.00%‒79.00%). The results suggested that Yen-yong1 represented a novel genus and a novel subfamily belonging to Caudoviricetes. Yen-Yong1 was species-specific and capable of lysing 3 out of 4 tested Y. enterocolitica strains. Yen-yong1 maintained high bacteriolytic activity at -80‒60 ℃ and pH 2.0‒13.0, as well as in the presence of 0‒3% NaCl. Yen-yong1 can disrupt the biofilms of Y. enterocolitica. At 25 ℃ and MOI=100, the sterilization rates of Yen-yong1 against Y. enterocolitica CICC 10869, CICC 21669, and CMCC 52202 in raw pork slices were 93.48%, 52.77%, and 43.24%, respectively. All the 3 tested Y. enterocolitica strains proliferated at 4 ℃. At 4 ℃ and MOI=100, the sterilization rates of Yen-yong1 against Y. enterocolitica CICC 10869, CICC 21669, and CMCC 52202 in raw pork slices were 98.28%, 78.35%, and 96.33%, respectively. [Conclusion] Yen-Yong1 reveals a previously unknown evolutionary lineage. Yen-Yong1 has excellent biological characteristics and potential applications. This study enriches the knowledge of Y. enterocolitica phages, phage database, and gene database, laying a foundation for the application of phages in the prevention and control of food-borne yersiniosis.

[Objective] To clarify the diversity characteristics of gut microbiota in the important wild pollinator, Xylocopa appendiculata, and provide theoretical basis for the conservation and utilization of wild pollinator resources. [Methods] Utilizing employed 16S rRNA gene high-throughput sequencing, we conducted comparative analyses of gut microbiota diversity in adults X. appendiculata from different habitats and sexes. Functional prediction of gut microbial communities was performed using PICRUSt2. [Results] The gut microbiota in adults X. appendiculata was predominantly composed of Bacillota (relative abundance of 50.17%), Pseudomonadota (relative abundance of 32.04%), Actinomycetota (relative abundance of 12.84%), and Bacteroidota (relative abundance of 3.77%). The genus Lactobacillus was identified as a core microbiota present in all sampled individuals. Significant differences in the relative abundance of gut microbiota were observed across habitats and sexes. The samples from the Sui and Tang Dynasties City Ruins Botanical Garden habitat (HXST) exhibited the highest richness, diversity, and evenness of gut microbiota. In the Sui and Tang Dynasties City Ruins Botanical Garden habitat (ST) and the Caoyao Village habitat (CY), female adults showed higher microbial richness, diversity, and evenness compared to males. In the Shuiquan Village habitat (SQ), male adults showed higher microbial richness, diversity, and evenness compared to females. The genus Gilliamella, as the dominant genus of male samples from Caoyao Village (HXCYM), accounts for a very small proportion in other samples. Functional prediction PICRUSt2 showed that the metabolic gene functions of the gut microbiota of adults X. appendiculata are the most enriched, with an abundance ratio of 74.00%. [Conclusion] The gut microbiota of adults X. appendiculata exhibits ecological adaptive features with social bees (e.g., Apis mellifera and Bombus spp.), characterized by low diversity, highly conservation, and specialized core microbial community structure. Environmental heterogeneity and host sex were identified as key drivers of gut microbiota divergence. These findings suggest that the gut microbiota of X. appendiculata may play a central role in maintaining host energy homeostasis and population adaptability.

[Objective] To improve the fermentation performance by optimizing the fermentation medium and conditions of Bacillus subtilis subsp. spizizenii YC25, thereby providing a theoretical basis for the subsequent industrial development and application of the strain. [Methods] With the spore yield as the indicator, the optimal fermentation conditions were screened by single factor tests. On this basis, the response surface methodology was employed to optimize the fermentation medium. Pot experiments were conducted to evaluate the control effects of the optimized fermentation broth on tobacco black shank and tobacco Fusarium root rot. [Results] The optimum fermentation conditions were as follows: 30 ℃, 180 r/min, an inoculation amount of 2%, a liquid-loading volume of 20%, and fermentation time 36 h. The optimum fermentation medium was as follows: corn starch 15.53 g/L, soybean meal 8.70 g/L, KH₂PO₄ 0.50 g/L, MgSO₄ 0.92 g/L, and CaCO₃ 0.50 g/L. Under these conditions, the spore yield reached 1.423 3×10¹⁰ CFU/mL. The results of pot experiments showed that the control effects of the fermentation broth diluted by 20 times on tobacco black shank and tobacco Fusarium root rot were 77.88% and 62.09%, respectively. [Conclusion] The optimized medium could significantly increase the spore yield of strain YC25, and the disease indexes of tobacco black shank and tobacco Fusarium root rot were significantly reduced after the application of YC25 fermentation broth. This study provides a theoretical basis for the development and application of the biocontrol strain YC25 and the prevention and control of field diseases in the future.

[Objective] Stenotrophomonas can promote the growth of plants and aquatic algae. However, limited studies have focused on the bacteriophages infecting Stenotrophomonas. In this study, we isolated bacteriophages infecting Stenotrophomonas from water and studied their biological and genomic characteristics, aiming to improve our understanding of the ecological functions of Stenotrophomonas and its bacteriophages. [Methods] The double-layer plate method was employed to isolate bacteriophages infecting S. indicatrix EB12 from lake water. The biological characteristics, whole genome sequence characteristics, gene functions, phylogenetic relationship, and protein structures of the isolates were analyzed. [Results] Transmission electron microscopy showed that two isolates Ste-X and Ste-D were short-tailed bacteriophages. The incubation period of both strains was 20 min. Ste-X and Ste-D showed the optimal multiplicities of infection being 1 000 and 10, burst sizes of 28.8 PFU/cell and 131.1 PFU/cell, temperature tolerance ranges of 20-60 °C and 20-70 °C, and pH tolerance ranges of pH 4.0-13.0 and pH 3.0-12.0, respectively. Ste-X and Ste-D were deactivated by UV irradiation for 60 min and 120 min, respectively. Their host ranges were narrow. The genomes of both strains showed the lengths of 39 429 bp, with the G+C content of 57.80% and 57.85%, respectively, and both of them contained 65 open reading frames (ORFs). The genomes of the two strains contained only three different bases. However, their homology with other bacteriophages was low, which suggested that the two strains were novel short-tailed bacteriophages. The structures of the proteins encoded by ORF34 and ORF52 were predicted. Result showed thant the proteins encoded by ORF34 were acidic, stable, and hydrophilic, containing two conserved domains, ChtBD3 and PHA00661, with the three-dimensional structures dominated by random coils. The proteins encoded by ORF52 were alkaline, unstable, hydrophilic, with no conserved domain and the three-dimensional structures dominated by α-helixes. [Conclusion] Two novel strains of short-tailed bacteriophages with high genome similarity but distinct biological characteristics were isolated from water.

[Objective] To study the response relationship of the biological properties of bacterial communities to long-term climate change. [Methods] The loess (YL, PL, and RL) and paleosol (YS, PS, and RS) sequences, which have persisted for approximately 500 000 years at Shimao (Y), Potou (P), and Renjiapo (R), were sampled. The structures and functions of the soil bacterial communities were predicted by bioinformatics approaches, including high-throughput sequencing and FAPROTAX. [Results] The physical and chemical characteristics associated with the loess-paleosol alternation in the three regions reflected the shifts in climatic conditions, including dry, cold, warm, and wet phases during soil development. Over this period, the climate at Renjiapo was characterized by the highest temperatures and precipitation levels, with strong influences from summer winds. Shimao experienced the driest conditions and the lowest temperatures. Potou exhibited the intermediate climatic conditions between Renjiapo and Shimao. The abundance and diversity indices of the bacterial communities in Shimao were higher than those in Potou and Renjiapo. Across all the three regions, the dominant bacterial phyla were Proteobacteria, Actinobacteria, and Acidobacteria although their relative abundance varied significantly. The bacterial communities in the loess and paleosol layers of Shimao showed greater compositional similarity, forming a network structure driven by synergistic interactions. The ecological functions of the bacterial communities in the loess-paleosol sequences were primarily associated with carbon, nitrogen, and sulfur cycling. Carbon and nitrogen cycling was weakly expressed in Shimao, while nitrogen cycling was more prominent in Potou. Renjiapo exhibited the most pronounced carbon cycling. [Conclusion] Soil bacterial communities in warm and humid climates tend to exhibit more complex network structures and greater functional diversity. In contrast, bacterial communities in dry and cold climates are characterized by similar composition and synergistic patterns, which enable these communities to adapt to harsh conditions by increasing their abundance and diversity, compensating for nutrient limitations associated with reduced carbon and nitrogen cycling.

[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.

[Objective] To extract and purify extracellular polysaccharides (EPS) from Sphingomonas echinoides 1K04342 and investigate the ameliorating effects and mechanisms of the purified EPS on glucose and lipid metabolism disorders induced by oleic acid (OA) in human hepatoma cells (HepG2). [Methods] A response surface methodology was employed to optimize the ultrasound power, ultrasound time, and incubation time for maximizing the EPS yield of S. echinoides 1K04342 from ultrasound-assisted extraction. The crude EPS underwent deproteinization and was further purified through a DEAE-Sepharose Fast Flow anion exchange column. The effects of the purified EPS on the proliferation, glucose uptake, and α-glucosidase inhibition and pancreatic lipase inhibition rates in HepG2 cells were evaluated. The most effective EPS component for mitigating OA-induced glucose and lipid metabolism disorders in IR-HepG2 cells was selected for in-depth physicochemical and bioactivity analyses. [Results] The optimal EPS extraction parameters were as follows: ultrasound power at 59.39 W, extraction for 42.08 s, and incubation for 9.24 h, under which the estimated yield of EPS reached 2.13 g/L. Under the adjusted parameters (60 W, 42 s, and 9.2 h), the EPS yield was 2.10 g/L, which was statistically consistent with the theoretical value (2.13 g/L). The optimal component for ameliorating glucose and lipid metabolism disorders in IR-HepG2 cells was EPS-2, which showed the total sugar content of 74.3%, with a monosaccharide profile comprising fucose, rhamnose, arabinose, galactose, glucose, and mannose. EPS-2 alleviated OA-induced glucose and lipid metabolic disorders in IR-HepG2 cells. EPS-2 reversed OA-induced decreases in cell viability. EPS-2 may activate the IRS-1/PI3K/AKT pathway, up-regulating the protein levels of GSK3β and FoxO1 and down-regulating the mRNA and protein levels of PEPCK and G6Pase, thereby mitigating glucose metabolism disorders. EPS-2 may regulate OA-induced lipid metabolism disorders by activating the AMPK/ACC1/SREBP-1C pathway, lowering the triacylglycerol (TG), total cholesterol (TC), and low-density lipoprotein (LDL) levels, and increasing the high-density lipoprotein (HDL) level. [Conclusion] EPS-2 derived from S. echinoides 1K04342 could effectively ameliorate glucose and lipid metabolism disorders in IR-HepG2 cells.

[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] Using Sphingomonas paucimobilis as the starting strain, a high-yield gellan gum-producing engineered strain was constructed through metabolic engineering, and fermentation process optimization was performed, providing both theoretical support and technical foundations for the efficient biosynthesis of gellan gum with this bacterium. [Methods] A CRISPR-Cas9-based gene editing system was developed for S. paucimobilis, and subsequently employed to genomically integrate two key gellan gum biosynthesis genes: the regulatory protein gene (gelA) and the β-1,4-glucuronosyltransferase gene (gelK), both under the control of constitutive promoters. Building upon this foundation, fermentation parameters including carbon source, nitrogen source, pH, and dissolved oxygen were systematically optimized through single-factor experiments. [Results] The engineered strain FMME-GG08 achieved a gellan gum yield of 10.8 g/L in shake-flask cultivation, representing a 130.2% enhancement over the parental strain. Following fermentation process optimization, the production level reached 20.1 g/L in 15 L scale bioreactors, with a sucrose conversion efficiency of 0.50 g/g. [Conclusion] This study not only successfully constructed a high-yield gellan gum-producing strain and established an efficient fermentation process, providing a reliable technical solution for industrial production, but also developed a genetic editing strategy that serves as an important reference for metabolic engineering of non-model microorganisms to produce other high-value exopolysaccharides.