[Objective] To study the physicochemical properties and active components of extracellular antibacterial substances from Bacillus subtilis subsp. spizizenii Bspi2104. [Methods] The Oxford cup method was employed to determine the antibacterial activities and physicochemical properties of B. subtilis subsp. spizizenii Bspi2104 with broad-spectrum antibacterial activity and B. subtilis subsp. spizizenii BspiL6 without antibacterial activity. Metabolomics was employed to detect the antibacterial components of the extracellular products of the strain Bspi2104. [Results] The treatment with trypsin, papain, protease K, pepsin, and lipase had no significant effect on the antibacterial activity of extracellular products. Extracellular products lost the antibacterial activity after treatment at 80 ℃ and 100 ℃ and presented decreased antibacterial activity at pH 9.0 and pH 11.0, especially at higher pH values. There was no significant difference in the antibacterial activity between the extracellular products treated with and without UV (P>0.05). The ammonium sulfate precipitates at 60%, 70%, and 80% saturation exerted antibacterial activities, which was the strongest at the saturation of 70%. The extracellular products of the two strains were extracted by hydrochloric acid precipitation, combined with methanol extraction, ethyl acetate extraction, and chloroform extraction, and all the extracts showed antibacterial activities. The ethyl acetate extract had the strongest antibacterial activity. LC-MS/MS was employed to analyze the composition of extracellular products of Bspi2104 and BspiL6 extracted with different methods. There were 35 common differential metabolites in the extracellular products of the two strains extracted with different methods. The differential metabolites belonged to 37 categories of compounds, including carboxylic acids and derivatives, fatty acids, organic oxygen-containing compounds, organic nitrogen-containing compounds, steroids and derivatives, pregnenolone lipids, phenols, alkaloids and derivatives, glycerol phosphates, isoflavonoids, and benzene and substituted derivatives. Some of these compounds, such as kurarinone, and surfactin B, had antibacterial activities. [Conclusion] The extracellular products of B. subtilis subsp. spizizenii Bspi2104 had good physicochemical stability and maintained high antibacterial activity after treatment with various proteases and lipases, and at −20 ℃ to 60 ℃, pH 1.0–11.0, and UV irradiation for 3 h. The ammonium sulfate precipitates and the extracts from hydrochloric acid precipitation combined with methanol extraction, ethyl acetate extraction, and chloroform extraction of the extracellular products of B. subtilis subsp. spizizenii Bspi2104 had antibacterial activities. Among them, the 70% ammonium sulfate precipitate and ethyl acetate extract had the best antibacterial effects. The extracellular products of the strain contained diverse categories of antibacterial compounds. The findings provide theoretical reference for the discovery and screening of antibacterial components of Bacillus, and the related metabolites have research prospects.

[Objective] To investigate the acclimation mechanisms of straw-decomposing microbiomes in response to historically different climate conditions as characterized by extreme temperature distinction, we collected two native forest soil samples from the tropical (annual mean temperature: 25 ℃) and cold-temperate (annual mean temperature: −2 ℃) zones at a distance of 3 860 km. [Methods] Microcosm incubation was conducted at a low temperature (10 ℃), a high temperature (35 ℃), and alternated high and low temperatures (10 ℃/35 ℃). The two native forest soil samples were inoculated for targeted cultivation of straw-decomposing microbiomes. After 12 consecutive weeks of passage, 16S rRNA gene sequencing was carried out to analyze the microbial community composition. [Results] At 10 ℃, higher straw decomposition rate was observed in the forest soil from Changbai Mountain in the cold-temperate zone (15.5%) than that from the tropical zone. At 35 ℃, the decomposition rate in the soil from Sanya in the tropical zone (33.1%) was higher than that from Changbai Mountain The results of linear discriminant analysis effect size (LEfSe) showed that the dominant straw-decomposing genera included Duganella, Pedobacter, Janthinobacterium, and Serratia after 12 weeks of enrichment at 10 ℃ with the forest soil from Changbai Mountain. The dominant genera were Paenibacillus and Rhodanobacter after enrichment at 35 ℃, and Stenotrophomonas, Burkholderia, and Achromobacter after enrichment at 10 ℃/35 ℃. As for the forest soil from the tropical zone, the enriched dominant genera were Pseudomonas, Acinetobacter, and Flavobacterium at 10 ℃, Cupriavidus at 35 ℃, and Enterobacter and Cohnella at 10 ℃/35 ℃. [Conclusion] This study revealed the indicator microbial species for straw decomposition at different temperatures in native forest soils from geographically highly distinct regions with a 3 860 km distance. The results suggest that temperature could have likely played a pivotal role in shaping the microbiomes for straw decomposition. The findings provide a scientific basis for mining the straw-decomposing microbial resources in the cold zone in northeast China and the tropical zone in south China.

Type 2 diabetes mellitus (T2DM) stands as a chronic metabolic disorder posing a challenge to global public health, owing to its widespread prevalence. The intricate interplay between gut microbiota and the onset and progression of T2DM, along with the potential therapeutic benefits of modulating gut microbiota, has emerged as a focal point in contemporary research. Recent studies have underscored the capacity of traditional Chinese medicine to ameliorate T2DM by inducing alterations in gut microbiota. Nevertheless, the precise mechanisms underlying the pharmacological actions of traditional Chinese medicine via gut microbiota regulation remain elusive. The diverse bioactive compounds in traditional Chinese medicine play pivotal roles in eliciting its pharmacological effects. This article systematically reviews the advancements in the research concerning the modulation of gut microbiota for T2DM intervention by a spectrum of bioactive components in traditional Chinese medicine, encompassing polysaccharides, alkaloids, flavonoids, saponins, and other compounds. The objective of this review is to furnish a comprehensive theoretical framework supporting the preventive and therapeutic potential of traditional Chinese medicine in T2DM management, thereby significantly contributing to the modernization of traditional Chinese medicine.

[Objective] To investigate the role of WekM, the O-antigen glycosyltransferase of avian pathogenic Escherichia coli (APEC) O₁, in lipopolysaccharide biosynthesis and environmental adaptation. [Methods] The wekM-deleted strain ΔwekM of APEC O₁ was constructed by Red homologous recombination, and then the complementary strain CΔwekM was constructed. The impacts of wekM on bacterial growth and motility were examined. The lipopolysaccharide (LPS) profile and reactivity with rabbit anti-O₁ serum of each strain were identified by silver staining and Western blotting. Real-time fluorescence quantitative PCR was conducted to determine the transcriptional levels of flagellum-related genes, and ethidium bromide was used to measure the bacterial cell membrane permeability. Finally, the drug sensitivity test was carried out to identify the bacterial susceptibility to antibiotics such as ciprofloxacin. [Results] The constructed ΔwekM and CΔwekM were verified by PCR amplification and DNA sequencing. Compared with the wild type, ΔwekM showed incomplete LPS profile and absence of some O-antigen bands. Western blotting results showed that ΔwekM did not react with the anti-O₁ serum, suggesting that the loss of WekM impaired the LPS production. The deletion of wekM reduced the swimming motility and did not impact the bacterial growth rate compared with the wild type. The transcription levels of flagellum-related genes such as flgC were down-regulated in ΔwekM. The results implied that the reduced motility of ΔwekM was caused by the decrease in flagellar production. In addition, ΔwekM demonstrated increased cell membrane permeability compared with the wild type (P < 0.01), and ΔwekM improved bacterial sensitivity to 7 antibiotics including polymyxin. This result suggested that the adaptability of ΔwekM to the environment was inhibited due to the increased cell membrane permeability. [Conclusion] The deletion of wekM in APEC results in diminished swimming motility, increased antibiotic resistance, improved cell membrane permeability, and damaged LPS integrity. The findings lay a foundation for mining the role of wekM and enrich our understanding of the stress resistance mechanism of APEC.

Streptococcus pneumoniae causes serious diseases such as pneumoniae and meningitis in humans. Capsular polysaccharides (CPSs) surrounding bacteria are not only key virulence factors but also major antigens. Therefore, CPSs have been prepared into polysaccharide vaccines and polysaccharide conjugate vaccines, which have greatly reduced the infection of pneumococci. CPSs are formed by polymerization of oligosaccharide repeating units which generally have 2−8 monosaccharides. CPSs present complex structures with diverse antigenic epitopes, being the basis of bacterial serotyping. Currently, 107 serotypes of S. pneumoniae have been identified. Each serotype has a unique CPS structure, a stable genetic basis, and specific serological characteristics. The diversity and constant changes of CPS structures explain the difficulty in the eradication of pneumococci. This review summarizes the known chemical structures of 95 CPSs and discusses the genetic basis, biosynthesis mechanism, and purification methods of CPSs. This review aims to enrich the knowledge about CPS diversity and provide a reference for probing into the functions and evolution of CPSs as well as for preparing polysaccharide vaccines.

[Objective] To explore the effects of cell walls of Saccharomyces cerevisiae on the intestinal microbiota in finishing bulls by 16S rDNA and ITS sequencing. [Methods] A total of 40 simmental crossbred finishing bulls weighing about 550 kg were randomized into 4 groups, with 10 bulls in each group. The control group was fed with a basic diet, and 5, 10, and 15 g cell walls of S. cerevisiae were added to the diet of each bull per day in trial 1, 2, and 3 groups, respectively. The preliminary trial and trial lasted for 10 days and 94 days, respectively. Intestinal feces were collected 7 days before the end of the trial. [Results] 16S rDNA: (1) The Chao and ACE indices in the trial 3 group were higher than those in other groups (P < 0.05); (2) Firmicutes and Bacteroidota were the dominant phyla, and Prevotella_9, Faecalibacterium, Succinivibrio, Bacteroides, and Bifidobacterium were the dominant genera; (3) The linear discriminant analysis effect size (LEfSe) revealed one differential species (LDA≥4.0, P < 0.05) playing an important role in the trial 2 group. ITS: (1) There was no significant difference in the alpha or beta diversity among groups (P > 0.05); (2) Ascomycota with the relative abundance above 50.00% was the dominant phylum. Penicillium, unidentified_Ascomycota_sp., Aspergillus, Orpinomyces, and Eurotium were the dominant genera; (3) LEfSe revealed 8 differential species (LDA≥3.0, P < 0.05), which included 3, 3, and 2 differential species playing an important role in the control, trial 2, and trial 3 groups, respectively. [Conclusion] Under conditions of this study, adding 10–15 g/d cell walls of S. cerevisiae in the basic diet increased the richness of intestinal microbiota and the relative abundance of beneficial bacteria Provetella_9, Tolypocladium, and Torulaspora, which were conducive to improve intestinal microecological environment of finishing bulls.

[Objective] The plasmid interference system of CRISPR/LshCas13a was constructed in Escherichia coli MG1655-ΔrecA and Escherichia coli DH10B to analyze the escape phenomenon in RNA editing experiments by targeting the non-essential gene lacZ and the essential gene polA. [Methods] An inducible CRISPR/LshCas13a RNA editing system- associated plasmid was designed with LshCas13a from Leptotrichia shahii. MG1655-ΔrecA and DH10B were selected as the research objects. The Crisporo algorithm was employed to design the CRISPR RNA (crRNA) sequences targeting lacZ and polA, and the LshCas13a plasmid interference experiment was carried out to study the escape phenomena targeting lacZ and polA. The escape phenomenon of the LshCas13a system was evaluated based on the number and sequences of escaped colonies. PCR and Sanger sequencing were conducted to explore the escape events of the LshCas13a system. The escaped colonies carrying the LshCas13a system disrupted by the insertion sequence (IS) were selected, and OD₆₀₀ was measured to evaluate the growth recovery of the strains. [Results] The LshCas13a system was used to target lacZ and polA in MG1655-ΔrecA and DH10B. MG1655-ΔrecA escaped through point mutation of LshCas13a and IS-mediated transposition when lacZ was targeted. When polA was targeted, MG1655-ΔrecA and DH10B escaped by point mutation of LshCas13a, IS-mediated transposition, and mutation of the direct repeat (DR) sequence of crRNA. The mutation of LshCas13a promoted the recovery of strain growth. [Conclusion] The LshCas13a plasmid interference system successfully revealed the diversified escape phenomena during the RNA editing of E. coli, including IS-mediated the transposition of LshCas13a, point mutation of LshCas13a, and DR sequence mutation or recombination of crRNA. The results laid a foundation for optimization of the CRISPR/LshCas13a gene editing system.

[Objective] To explore the role of S1PP in the morphological differentiation, pathogenic process, and maintenance of sphingolipid balance of Magnaporthe oryzae. [Methods] We employed homologous recombination to delete the S1PP gene MoLCB3 from M. oryzae and characterized the obtained mutant ΔMolcb3 was by phenotypic analysis, gene complementation, and lipid metabolomics. Furthermore, we deleted the sphingosine kinase (SK) gene MoLcb4 from ΔMolcb3 to explore the relationship between MoLcb3 and MoLcb4. [Results] The deletion of MoLCB3 resulted in significant decreases in the mycelial growth rate and spore production and affected conidial malformation and initial appressorium formation. ΔMolcb3 completely lost the pathogenicity to barley. Moreover, the ΔMolcb3 mutant were significantly different from the wild type in responding to hyperosmic stress, cell wall integrity stress, high temperature stress, and fungal lipid synthesis inhibitors triadimefon and myriocin, suggesting that MoLcb3 was involved in these stress responses and lipid anabolism. Interestingly, the double mutant ΔMolcb3ΔMolcb4 basically compensated for all phenotypic defects of ΔMolcb3. In addition, lipid metabolomics showed that compared with the wild type, ΔMolcb3 presented significantly different levels of lipids, such as free fatty acids, ceramides, and phosphatidyl inositol. [Conclusion] MoLcb3 plays an important role in the mycelial growth, sporulation, spore germination, pathogenicity, stress responses, and lipid homeostasis. In addition, knockout of MoLCB4 can cushion the effects of MoLcb3 deletion. The results of this study provide new ideas for elucidating the sphingolipid metabolic pathway of M. oryzae and the development of inhibitors of fungal lipid biosynthesis.

2-phenylethanol (2-PE) is a rose-scented aromatic alcohol commonly used in the food, cosmetic, and pharmaceutical industries. The physical and chemical production methods of 2-PE are not suitable for industrial application due to the low yields. As a single-celled eukaryotic microorganism, yeast has the potential to efficiently synthesize natural 2-PE. Therefore, the strategy of using yeast as a chassis microorganism to synthesize 2-PE is favored by researchers. However, during the fermentation for 2-PE production, the yeast is inevitably affected by the toxic effects of 2-PE. Therefore, there is an urgent need to investigate the mechanisms of yeast tolerance to 2-PE, which will provide a theoretical basis for production practice and help to select yeast strains with high tolerance to 2-PE. In this paper, we review the research advances in 2-PE tolerance of yeast from the synthetic pathways of 2-PE and yeast tolerance mechanisms and introduce the methods for improving the 2-PE tolerance of yeast. Deciphering the mechanism of yeast tolerance to 2-PE for improving the yield and conversion efficiency of 2-PE in yeast is a top priority for the future research.

Pyrroloquinoline quinone (PQQ), the third oxidoreductase coenzyme discovered in the nature after nicotinamide and riboflavin, is ubiquitous in bacteria, fungi, plants, and animals. PQQ participates in a variety of life activities and has anti-inflammation, anti-oxidation, cell metabolism-enhancing, and cardioprotective activities, demonstrating broad application prospects in pharmaceuticals, agriculture, food and other fields. Therefore, the large-scale production of PQQ is the primary problem that needs to be solved at present. Microbial fermentation is a primary production method of PQQ. Deciphering the biosynthesis pathway and regulatory mechanism of PQQ is essential for the screening and breeding of strains with short production periods and high yields by metabolic engineering, which has been a hot topic in this field. This paper summarizes the synthesis pathways, strain screening and breeding, microbial production, and purification processes of PQQ, aiming to provide a reference for further research and application of PQQ.