Abstract:Medical microbiology focuses on the biological characteristics and pathogenesis of pathogenic microorganisms, mechanisms of human immune responses to infections, and pathogen diagnosis and prevention of infectious diseases. This discipline has evolved from morphology to molecular microbiology, microbiomics, microbial systems biology, and reverse microbial etiology at the knowledge level, while at the application level it encompasses genetic engineering, microbial gene manipulation, and microbial synthetic biology. This article reviews the recent advances in medical microbiology in terms of biological characteristics, pathogenicity, immunity, clinical pathogen diagnosis, prevention, and treatment. We should re-examine the relationship between microorganisms and human health, as well as microorganisms themselves, pay attention to microbiome data and knowledge system reshaping, and enhance public awareness of microorganisms. This review provides directions for the future development of medical microbiology.

Abstract:Plants are subjected to biotic and abiotic stresses during growth, of which pathogen infection is a major biotic stress. Since the plant microbiome co-evolves with plants, the plants exposed to pathogen stress will fight against pathogens by special secretions or recruitment of beneficial microorganisms. This paper reviews the research progress in the “cry for help” strategy of plants under pathogen stress from the changes in plant microbiome and metabolites, plant-microbe interactions, the disease-suppressive soil formed after the plant “cry for help”, with a view to providing a basis for the prevention and control of soil-borne diseases in plants.

Abstract:Aromatic amino acid derivatives are widely distributed in the nature and possess antioxidant, anti-inflammatory, neuroprotective, and anticancer activities. They play important roles in various fields such as food, cosmetics, nutritional supplements, and chemistry, particularly in the pharmaceutical industry. In recent years, the unconventional model organism Yarrowia lipolytica has emerged as a promising candidate in biosynthesis due to its unique physiological and biochemical characteristics. Specifically, it has shown significant potential in the synthesis of aromatic amino acid derivatives, including resveratrol and naringenin. With a distinctive cytosolic environment and high flux of acetyl-CoA, Y. lipolytica stands out as an ideal host for the synthesis of aromatic amino acid derivatives. This review summarizes the biosynthesis strategies of Y. lipolytica in the production of aromatic amino acid derivatives and reviews the current progress in this field. Furthermore, it discusses the future prospects and challenges of applying Y. lipolytica as a chassis cell in synthetic biology.

Abstract:Viral gastroenteritis is an acute infectious disease caused by multiple different viruses. Postbiotics can be used for the prevention and treatment of viral gastroenteritis, and the specific clinical feasibility and treatment strategy remain to be studied. This article reviews the latest research progress in the application and mechanism of postbiotic components in treating gastroenteritis caused by rotavirus and norovirus and discusses the current status and future research directions of postbiotics in inhibiting enteroviruses. This review is conducive to the research on the application of postbiotics in the prevention and treatment of enteroviruses.

Abstract:As one of the common and deadly malignant tumors worldwide, colorectal cancer (CRC) poses a serious threat to public health and causes a heavy burden to the society. The influences of microorganisms on the occurrence and development of CRC has received much attention from researchers, whereas most of the available studies have focused on gut microorganisms. In recent years, with the rapid development of next-generation sequencing, intratumoral microorganisms of CRC have been discovered and gradually opened up new research areas. We review the research progress in intratumoral microorganisms of CRC and their effects on the course of CRC and compare the relative abundance and community composition of intratumoral microorganisms of CRC with different molecular subtypes, at different stages of adenoma-carcinoma progression, and at different locations. Intratumoral microorganisms mainly affect CRC development by influencing colonic epithelial cells, tumor cells, and immune cells, with the pathogenic mechanisms including DNA damage, metabolic reprogramming, and oncogenic non-coding RNAs. Different intratumoral microorganisms exert dual effects on CRC. In the future, comprehensive experimental protocols and suitable in vitro models should be established to overcome the limitations of existing studies and elucidate the functioning mechanisms of intratumoral microorganisms, which will lay a foundation for the clinical application of microorganisms-targeted therapies for CRC and even other tumors.

Abstract:Atmospheric and room temperature plasma (ARTP), a new approach of mutagenesis, is characterized by fast mutagenesis speed, mild mutagenesis conditions, and environmental friendliness. The ARTP-induced mutants have high mutagenesis frequency and good genetic stability. Therefore, ARTP is widely used in microbial breeding. This paper reviews the progress in ARTP in microbial breeding in recent years, the screening strategy of mutants after mutagenesis, and the mechanism of ARTP-induced mutagenesis in microorganisms. Finally, we discuss the existing problems of ARTP in microbial breeding.

Abstract:Influenza virus continues to pose a threat to global public health and safety, leading to infectious respiratory diseases and causing hundreds of thousands of deaths worldwide every year. New strains are evolved through antigenic shift and drift, thereby developing resistance to existing drugs and vaccines. To address this issue and develop novel antiviral drugs, researchers need to explore new antiviral targets. The genetic products of influenza virus are widely modified through phosphorylation by host kinases. The reversible phosphorylation of serine, threonine, or tyrosine residues dynamically regulates the structures, functions, and subcellular localization of viral proteins at different stages of the viral life cycle. In addition, kinases affect a large number of signaling pathways, which influence virus transmission by regulating the host cell environment, thereby establishing critical virus-host relationships. The dependence on host kinases provides a theoretical basis for developing kinase inhibitors as the next generation of anti-influenza virus drugs. To fully utilize this potential, we need to clarify the influenza virus-host kinase interaction network. The focus of this review is to outline the molecular mechanisms by which host kinases regulate different stages, from adsorption to assembly and budding, in the life cycle of influenza virus. Evaluating the contributions of different host kinases and their specific phosphorylation throughout the influenza virus life cycle can provide a comprehensive overview of the virus-host kinase interaction network, which may help reveal potential targets for developing novel antiviral drugs.

Abstract:Intercellular heterogeneity in bacterial populations is a crucial factor determining bacterial antibiotic resistance and virulence and plays a key role in host-pathogen interactions, posing a major challenge to the control of bacterial infections. Single-cell sequencing is a powerful tool for detecting cellular heterogeneity. However, bacteria have unique biological characteristics such as small genomes, low cellular mRNA contents, and high proportion of rRNAs in the transcriptome, short half-life, polyA tail-lacking mRNA transcripts, and thick cell walls, making the eukaryote-based single-cell sequencing methods impossible to be applied in bacteria. To overcome these obstacles and promote the development of single-cell technology in bacterial research, researchers have successfully developed the single-cell sequencing methods applicable to bacteria. This review systematically summarizes the bacterial single-cell transcriptome sequencing and genome sequencing technologies proposed in recent years, describes their characteristics, and discusses the application prospects of bacterial single-cell sequencing in revealing bacterial drug resistance, aiming to shed light on the subsequent the related research. Bacterial single-cell can deeply reveal bacterial heterogeneity, improve the accuracy of disease diagnosis and treatment, and support microbiological and infectious disease research.

Abstract:Inflammatory bowel disease (IBD) is a complex and chronic inflammatory disease that is challenging to be cured and prone to carcinogenesis. Existing medications for IBD have limited efficacy and often come with adverse effects, which necessitates the development of new therapies. Clostridium butyricum and its metabolite, butyrate, have recently been identified as a probiotic and a metabolite, respectively, that offer protective benefits to the intestinal mucosal barrier. However, the specific mechanisms underlying their actions remain to be elucidated. Therefore, this article comprehensively reviews the research progress in the roles of C. butyricum and butyrate in the pathogenesis of IBD via influencing the intestinal barrier function, as well as their potential clinical values in the treatment of IBD. The findings will give novel insights into the research on the pathogenesis and innovation of the therapies of IBD.

Abstract:The research on the disease-inducing mechanisms of pathogens is an important way to reveal the regularity of host infections. Streptococcus suis (SS) is one of the major pathogens causing economic losses in the global pig industry. Among the different SS serotypes, SS serotype 2 (SS2) is the most pathogenic and widespread. This paper briefly describes the crucial virulence factors of SS2 and elaborates on the mechanisms of SS2 in inducing streptococcal toxic shock-like syndrome and meningitis, with a view to providing new ideas for the prevention and control of porcine streptococcal diseases.

Abstract:[Background] Hospital sewage contains abundant pathogenic bacteria and antibiotic resistance genes (ARGs), which can escape into the air along with the sewage treatment process and lead to infections and emergence of infectious diseases. [Objective] To investigate the presence and seasonal changes of antibiotic resistant bacteria and ARGs in hospital sewage and the bioaerosols. [Methods] Bioaerosol samples were collected by an Anderson eight-stage sampler, and the antimicrobial susceptibility test and PCR were employed to assess the resistance profile of Escherichia coli. The abundance and structure of ARGs in the sewage and air were examined by high-throughput quantitative PCR. [Results] The bioaerosol concentration was the highest in summer. The air of the biochemical tank had higher bacterial concentration ((250±170) CFU/m³) and higher percentage of small-size (<2.1 μm) bacterial aerosols than that at other sampling sites. A total of seven strains of E. coli were isolated and identified in the samples from the biochemical tank. These strains showed the highest resistance (85.71%) to cefoxitin and imipenem, and the detection rates of parC, bla_TEM, and dhps were 100%. The abundance of ARGs in the air of the biochemical tank was the highest (26.18 copies/m³) in winter, and the mean ARGs abundance in different seasons was 14.82 copies/m³. intI1 had higher abundance in the air in spring and autumn, and tnpA-05 and strB in the air had the highest abundance in summer and winter, respectively. The source tracking results showed that biochemical tank effluent was the main source of ARGs. According to the multiple antibiotic resistance index and exposure dose, the sewage of the biochemical tank was seriously polluted by antibiotics, and the exposure dose of ARGs in the air of the biochemical tank was high in spring and winter. [Conclusion] The hospital sewage treatment stations, as the potential sources with a high risk of antibiotic exposure, should be well managed for the prevention and control of infections.

Abstract:[Background] Perchlorate is a new water pollutant, and using microorganisms for harmless treatment of industrial wastewater containing perchlorate has high efficiency, low costs, and environmental friendliness. [Objective] However, the resources of local perchlorate- reducing bacteria with fast growth, strong activity, and high reduction efficiency are extremely scarce and urgently need to be explored. [Methods] In this study, we isolated and screened perchlorate-reducing bacteria from activated sludge, identified their species using molecular biology techniques, and explored the physiological and metabolic characteristics of the isolated strains through microbial culture experiments. [Results] Finally, a perchlorate-reducing bacterial strain QD19-16 was isolated and identified as Brucella sp. by 16S rRNA gene analysis. Strain QD19-16 was an anaerobic facultative bacterium, with optimal growth at 30 ℃ and pH 7.0. Strain QD19-16 demonstrated a variety of metabolic activities and showed the ability to use acetate, yeast extract, and lactic acid as electron donors and perchlorate, chlorate, sulfate, and oxygen as electron acceptors. Under anaerobic conditions, the maximum specific growth rate (μ_max) of strain QD19-16 using perchlorate as an electron acceptor was 0.279 d⁻¹, with a half-saturation constant (K_s) of 0.294 mmol/L for perchlorate. [Conclusion] Brucella sp. QD19-16 is the first reported perchlorate-reducing bacterium of Brucella, expanding the distribution of perchlorate-reducing bacteria of Proteobacteria. Strain QD19-16 can be applied in assembling the microbial communities for perchlorate reduction in anaerobic bioreactors.

Abstract:[Background] Extensive studies have been carried out on strain mutagenesis, especially chemical mutagenesis, in strain breeding. However, most of these studies employ conventional mutagenesis, which usually does not involve any special treatment of target cells and has low mutagenic efficiency due to random and uncontrollable mutagenic process. [Objective] To screen thermotolerant strains for simultaneous sulfide and nitrate removal, we improved the mutagenic scheme on the target strain and studied the entry of mutagen into the cells. [Methods] Both conventional mutagenesis and improved mutagenesis were carried out for Thiobacillus denitrificans. In the improved mutagenesis, piperazine-1,4-diethylsulfonic acid (PESA) was used to treat the exponential-phase cells into a susceptible state before mutagenesis to change the permeability of cytoplasmic membrane, and then different volumes (0.5‰–3.0‰) of the mutagen diethyl sulfate (DES) were separately added into the cell suspension for mutagenesis at a set temperature of 45 ℃ for 20 min. The thermotolerance and sulfide and nitrate removal performance were tested at 47 ℃ to obtain the thermotolerant mutants. During the process of strain mutagenesis, we used S₂O₃²⁻, which could terminate DES mutagenesis, to observe the distribution of mutagen inside and outside the cells and compared the experimental results under different mutagenic schemes. [Results] We obtained the thermotolerant mutants capable of simultaneously removing sulfide and nitrate. The comparison of the two mutagenic schemes proved that changing the permeability of cytoplasmic membrane promoted the entry of mutagen into the cells. To achieve the same mutagenic effect, the improved mutagenesis only needed 1/10–1/9 of mutagen required by conventional mutagenesis. The percentage of mutagen entering the cells was 41.8%–40.4% of the total DES added in the improved mutagenesis and only 5.6%–4.4% in the conventional mutagenesis. The forward mutation efficiency might be closely related to the dosage of mutagen entering the cells, and the dosage in the improved mutagenesis was more suitable for achieving higher forward mutation efficiency. However, the dosage of mutagen did not determine under which circumstance or when individual mutation, such as thermotolerance or higher sulfide removal efficiency of mutants, probably occurred. The maximum dosage of mutagen entering cells depended on cell’s loading capacity and was independent of mutagenic schemes. The product analysis showed that S₂O₃²⁻ could be used to terminate the mutagenesis triggered by DES. [Conclusion] After treating the cells into a susceptible state, mutagenesis was carried out. And then through temperature-rise screening, 7 thermotolerant mutants with high-performance for sulfide and nitrate removal were obtained, providing new ideas for research on strain mutagenesis.

Abstract:[Background] Efficient strains are the key to the biodegradation of oil pollutants. [Objective] To screen an efficient oil-degrading strain, optimize its immobilization conditions, and evaluate the oil degradation efficiency of the immobilized strain. [Methods] A highly efficient oil-degrading strain was isolated from traditional bacon in Pingxiang by the plate culture method and identified by 16S rRNA gene sequencing. The safety of the strain was evaluated based on three pathogenic indicators. The immobilization conditions of the strain were optimized by single factor tests and an orthogonal design. The degradation effects of the immobilized strain and free strain on soybean oil were compared. Furthermore, the reuse effect and the degradation effect of the immobilized strain on the oil in sewage were evaluated. [Results] The strain was identified as Staphylococcus sp., and it was not pathogenic. The optimal immobilization conditions for the strain were 4.0% sodium alginate, 2.0% CaCl₂, sodium alginate solution:strain suspension volume ratio of 1:1, and crosslinking for 8 h. In the case of degrading soybean oil at pH 7.0, 30.0 ℃, and 180.0 r/min for 3 days, the immobilized strain achieved the highest degradation rate of 65.7%, which was 14.1% higher than that of the free strain. Moreover, the immobilized strain had tolerance to a wider pH range and stronger thermal stability than the free strain, and it demonstrated high oil-degrading ability after being repeatedly used for 4 times. [Conclusion] The degradation rate of the immobilized strain reached 41.6% for the oil in sewage. The immobilized strain can degrade both soybean oil and sewage oil, demonstrating a broad application prospect.

Abstract:[Background] Phosphorus in soil generally exists in the form of insoluble phosphate, which limits the direct absorption and utilization by plants. Phosphate-solubilizing bacteria can convert the insoluble phosphate in the environment into phosphorus that can be absorbed and utilized by plants. [Objective] To isolate and identify a highly efficient phosphate-solubilizing bacterial strain, and explore its phosphate-solubilizing capacity, plant growth-promoting effect, and related genes, so as to provide elite strain resources and theoretical support for the research and development of phosphate-solubilizing bacterial fertilizer. [Methods] The strain with significant phosphate-solubilizing effects was isolated from soil and identified based on physiological and biochemical characteristics as well as molecular evidence. The phosphate-solubilizing capability of the strain was determined by the transparent circle method and the Mo-Sb colorimetric method. Single-factor experiments were carried out to study the effects of different carbon sources, nitrogen sources, and insoluble phosphates on the phosphate-solubilizing capability of the strain. Further, the whole genome of the strain was sequenced by the next-generation sequencing platform, and the genes related to phosphate solubilizing and siderophore synthesis and transport in the genome were mined. Finally, the plant growth-promoting effect of the strain was assessed by the hydroponic experiment. [Results] Strain PSB-K was identified as Acinetobacter baumannii, with a phosphate-solubilizing index of 2.58. Strain PSB-K exhibited the highest phosphate-solubilizing capability in the case of glucose as the carbon source, potassium nitrate as the nitrogen source, and tricalcium phosphate as the insoluble phosphate, under which the soluble phosphorus concentration in the fermentation liquid reached 560.35 μg/mL. The whole genome of strain PSB-K was 3 983 883 bp in length, containing 3 849 coding genes, including various genes involved in organic acid synthesis, phosphate transport, and siderophore synthesis and transport. The results of the hydroponic experiment showed that strain PSB-K applied at 2.50×10⁷, 2.50×10⁷, and 1.25×10⁷ CFU/mL demonstrated the strongest promoting effects on the leaf area, fresh weight, and root length of oilseed rape seedlings, which were increased by 72.15%, 72.08%, and 197.81%, respectively, compared with the control. [Conclusion] Strain PSB-K possessed the capability to solubilize a variety of insoluble phosphates, and carbon and nitrogen sources directly influenced the phosphate-solubilizing performance of the strain. With a growth-promoting effect on oilseed rape plants, this strain serves as a candidate for the development of microbial fertilizer. In addition, the whole-genome data provides a reference for understanding the phosphate-solubilizing mechanisms and mining functional genes of phosphate-solubilizing bacteria.

Abstract:[Background] Gene expression is often studied by real-time quantitative reverse transcription PCR (RT-qPCR), the stability of which largely depends on the choice of reference genes. Cordyceps blackwelliae is a fungus of development potential, and it is thus vital to understand the expression patterns of functional genes in this fungus. [Objective] To identify the optimal reference genes that express stably across different developmental stages (mycelia from liquid culture, mycelia from wheat grain media, and fruiting bodies) of C. blackwelliae. [Methods] We utilized RT-qPCR to investigate the expression of eight reference genes (18S rRNA, gapdh, ꞵ-tubulin, cyclophilin A, γ-actin, rpl10, tef1α, and ubiquitin). The expression stability of these genes across different developmental stages was evaluated using geNorm, NormFinder, BestKeeper, and ∆C_t algorithms. [Results] The results indicated that γ-actin and 18S rRNA exhibited more stable expression than the other genes, making them suitable reference genes for the transcriptional analysis of functional genes in C. blackwelliae. [Conclusion] This study successfully identified appropriate reference genes for RT-qPCR in C. blackwelliae, aiding future research on gene expression dynamics.

Abstract:[Background] Pruned branches of citrus are rich and renewable biomass resources, in which the stable and complex lignocellulose structure limits the utilization of the resources. [Objective] To screening and identify efficient lignin-degrading bacteria from citrus waste compost and optimize the lignin-degrading conditions of strains, thus providing strain resources for the utilization of citrus waste. [Methods] We isolated lignin-degrading strains from citrus waste compost by the aniline blue method and compared the laccase, manganese peroxidase, and lignin peroxidase activities between different strains. Then, we optimized the lignin degradation conditions of the target strain (LDB1) by single factor tests, orthogonal test, and response surface methodology. Finally, we verified the degradation effect of LDB1 on the lignin derived from three plant species by laboratory experiments. [Results] We isolated eight lignin-degrading bacterial strains from the citrus waste compost. Among them, strain LDB1 had the strongest enzyme activity and was identified as Bacillus amyloliquefaciens (GenBank No. OR244332). The lignin degradation by LDB1 was affected by pH, inoculum amount, and C/N ratio. The fermentation with LDB1 at pH 6.9, inoculum amount of 2.9%, and C/N ratio of 29.0 achieved the lignin degradation rate of (14.9±0.9)% after 4 days, which increased by 2.0% compared with the orthogonal test result. After 30 days, the lignin degradation rates of LDB1 for citrus branches, maize straw, and rice straw were 12.0%, 36.8%, and 30.6%, respectively. [Conclusion] B. amyloliquefaciens LDB1 can simultaneously secrete three lignin-degrading enzymes to destroy the lignocellulose structure, demonstrating strong lignocellulose-degrading ability.

Abstract:[Background] Composting is an effective way of treating agricultural waste, and improving the phosphorus conversion efficiency during the composting process is receiving increasing attention. [Objective] To study the effect of corn steep liquor addition on the transformation of phosphorus fractions and microbial community in the composting process of spent mushroom substrate. [Methods] Two treatments of CK (100% spent mushroom substrate) and CP (spent mushroom substrate added with 0.5% corn steep liquor) were designed for aerobic composting for 36 d. The Hedley method was used for phosphorus fractionation. High-throughput sequencing was performed to reveal the microbial community composition, and the data were analyzed by bioinformatics tools. [Results] The addition of corn steep liquor significantly increased the temperature of the heap, prolonged the thermophilic phase, promoted the decomposition of organic matter, and improved the nutrient status of the heap by increasing active phosphorus content and promoting phosphorus activation and accumulation. The addition of corn steep liquor increased the diversity and richness of bacteria and fungi. It significantly increased the relative abundance of Acinetobacter, Pseudomonas, Mycothermus, and Myceliophthora. The addition of corn steep liquor reduced the complexity of the microbial network and promoted the cooperation and symbiosis among microorganisms. In addition, it changed the correlations between phosphorus fractions and some microbial genera. Acinetobacter and Sphingobacterium in the CP treatment greatly promoted phosphorus activation and accumulation. In the CK treatment, Agaricus promoted phosphorus activation, and the rest fungal genera promoted phosphorus accumulation. More fungi were positively correlated with active phosphorus after the addition of corn steep liquor. [Conclusion] The addition of corn steep liquor affected the microbial community structure and phosphorus transformation by changing the trophic modes and physicochemical indexes during the composting process. This study provides a reference for improving the availability of compost phosphorus.

Abstract:[Background] Cyclocybe chaxingu has high edible and medicinal values. Strengthening the breeding of elite strains is the premise for promoting the development of the C. chaxingu industry. At present, crossing is a common method in the breeding of edible fungi, and the acquisition of monokaryons and identification of the mating type is the basic work in the breeding of edible fungi. [Objective] To identify the mating type of C. chaxingu and evaluate the influence of mating type on the mycelial growth rate of monokaryons, thus laying a theoretical foundation for the crossing of C. chaxingu. [Methods] The protoplast monokaryons of C. chaxingu Gancha AS-5 were obtained by protoplast monokaryotation, and the standard tester strains were ascertained by mutual mating of monokaryotized protoplasts. Furthermore, basidiospores were collected. The basidiospore-derived monokaryons were matched with standard tester strains and with each other, respectively, and then the mating type of monokaryons was classified. The mating type of monokaryons was identified by oak wood extract-squeezed orange juice agar (OWE-SOJ), nuclear migration test, and molecular identification. [Results] A total of 420 monokaryons were collected and classified into four types, including 69, 126, 148, and 77 of T1, T2, T3, and T4, which accounted for 16.43%, 30.00%, 35.24%, and 18.33%, respectively. The results of OWE-SOJ, nuclear migration test, and molecular identification showed that the mating types of T1, T2, T3, and T4 were A1B1, A2B2, A2B1, and A1B2, respectively. The mating combinations of monokaryons randomly selected from the identification results could form fruiting bodies. Both A and B factors had significant effects on the mycelial growth rate of monokaryons, and the effect of B factor was greater than that of A factor. [Conclusion] OWE-SOJ, nuclear migration, and molecular techniques can accurately identify the mating type of monokaryons. Partial segregation appeared in the mating of monokaryons. There is an association between the mycelial growth rate and mating type of C. chaxingu monokaryons.

Abstract:[Background] The continuous cropping and poor fertilization habits in the tobacco-planting areas of Panzhihua have brought serious stress to the nutrient balance and microbial diversity in the soil, leading to declines in the yield and quality of tobacco and seriously affecting the development of the tobacco industry. [Objective] To study the effects of applying rhizobium-inoculated leguminous green manure on the microbial diversity and community composition in the soil cultivated with tobacco for years in Panzhihua and provide a theoretical basis for scientific fertilization and establishment of a reasonable planting pattern of flue-cured tobacco in Panzhihua. [Methods] We collected soil samples from the newly reclaimed tobacco field and the field planted with tobacco for years (both with the application of green manure) in Pingshan Township, Miyi County of Panzhihua. The physical and chemical properties of the soil samples were determined, and the bacterial community composition and diversity were analyzed by high-throughput sequencing. [Results] Inoculation with rhizobia increased the number of nodules and roots of pea seedlings, especially in the newly reclaimed tobacco field. The application of green manure increased the content of organic carbon and inorganic nutrients in soil and reduced soil pH and volume weight. In addition, the application of pea green manure changed the bacterial diversity and community composition in the soil. Specifically, it elevated the Simpson and Shannon indexes of bacteria in the newly reclaimed tobacco field and reduced the relevant diversity indexes in the field cultivated with tobacco for years. The predominant bacterial phyla of the soil samples planted with tobacco for different years included Proteobacteria, Chloroflexi, and Actinobacteria, while the dominant bacterial genera were Acinetobacter, Ochrobactrum, Sphingomonas, and Streptomyces. The redundant analysis showed that soil pH, available potassium (AK), and alkali-hydrolyzed nitrogen (AN) had strong correlations with the microbial community structure (P<0.05). [Conclusion] The application of green manure improves the nutrients and changes the bacterial diversity and community composition of the tobacco-growing soil.

Abstract:[Background] Ophiopogon japonicus-maize intercropping is the main planting mode in Ophiopogonis Radix producing areas. Revealing the microbiome structure and functional changes in the rhizosphere soil of this mode is crucial for ecological intercropping. [Objective] To investigate the diversity and functions of rhizosphere microbiome in the O. japonicus-maize intercropping system. [Methods] We employed Illumina-NovaSeq high-throughput sequencing of the 16S rRNA gene of bacteria and the ITS rDNA of fungi to study the microbiome structure and functions in the rhizosphere soil samples of the O. japonicus-maize intercropping system and monoculture systems. [Results] Intercropping significantly increased the plant height and yield of O. japonicus but had little effect on maize. The bacterial diversity in rhizosphere soil of both plants had no significant difference between intercropping and monoculture patterns. However, compared with monoculture, intercropping increased the fungal diversity in the rhizosphere soil of O. japonicus and decreased the fungal diversity in the rhizosphere soil of maize. Compared with monoculture, intercropping reduced the relative abundance of Actinobacteriota, Mortierellomycota, Xenomyrothecium, and Gibberella but enriched Ascomycota, Mucoromycota, Pyrenochaetopsis, Rhizopus, Fusarium, Alternaria, Lysobacter, and Rhodanobacter in the rhizosphere of O. japonicus. In the rhizosphere of maize, intercropping increased the relative abundance of Ascomycota, Desulfobacterota, Myxococcota, Coprinellus, and Fusarium, while decreasing the relative abundance of ten families including Cladosporiaceae and two genera of Xenomyrothecium and Gibberella. No significant differences were observed in the relative abundance of KEGG pathways of rhizosphere microbiome between monoculture and intercropping modes. Intercropping significantly increased the relative abundance of plant pathogens, soil saprotrophs, and wood saprotrophs in the rhizosphere of both O. japonicus and maize. However, it decreased the relative abundance of fungal parasites in the rhizosphere of maize while increasing that in the rhizosphere of O. japonicus. Furthermore, intercropping reduced the relative abundance of plant pathogens in the rhizosphere of maize but significantly increased that in the rhizosphere of O. japonicus. [Conclusion] The intercropping of O. japonicus and maize promotes the growth and increases the yield of O. japonicus. It has mild effects on the structure and functions of rhizosphere bacterial community of O. japonicus and maize, and most bacterial genera displayed no significant difference between the two cropping modes. However, the structure and functions of fungal community were greatly influenced by intercropping, and most fungal genera showed significant convergence or divergence.

Abstract:[Background] Thermophilic strains are ubiquitous in the brewing environment of Baijiu and are a microbial group indispensable for the brewing of Baijiu. [Objective] To isolate and identify thermophilic strains in the samples collected from the brewing environment of Baijiu and characterize the growth of the strains, thereby laying a theoretical foundation for the application of the strains in the production of Baijiu. [Methods] Gradient dilution was carried out for the samples of Jiuqu (starter), Jiupei (fermented grains), and Jiuzao (distilled grains), respectively. The samples with suitable dilution factors were then selected and coated on the Luria-Bertani, yeast extract peptone dextrose, and potato dextrose agar media, respectively. The plate streaking method was employed to isolate the strains. [Results] A total of 22 strains of thermophilic bacteria and 18 strains of thermophilic fungi were screened out. Among them, 4 species of bacteria and 4 species of fungi with distinct colony morphology were selected and identified by morphological observation, physiological and biochemical tests, and phylogenetic analysis. The four species of bacteria were identified as Streptomyces albulus, Bacillus subtilis, B. amyloliquefaciens, and B. rugosus, and the four species of fungi were identified as Rhizomucor pusillus, Thermomyces lanuginosus, Thermoascus crustaceus, and Malbranchea cinnamomea. It is worth noting that B. rugosus and M. cinnamomea had not been reported in the research on Baijiu. The eight species showcased differences in the growth temperature range, pH range, and NaCl tolerance. [Conclusion] We screened out, identified, and characterized the growth of four species of thermophilic bacteria and four species of thermophilic fungi, which provided data support for studying the roles and applications of these microorganisms in the brewing of Baijiu.

Abstract:[Background] The Chinese monal (Lophophorus lhuysii) is a protected wild bird endemic to China, with a declining population due to human activities and diseases. Enterococcus, a genus of common conditional pathogenic bacteria in the nature, is known for strong drug resistance and difficult treatment, posing challenges to wildlife conservation. [Objective] Therefore, it is necessary to pay attention to the enterococcal infection or carrier status and study the characteristics of enterococci in order to better protect the Chinese monal population, which is of great significance to the survival and protection of Chinese monal population. [Methods] Fecal samples were collected from captive Chinese monal at the Fengtongzhai National Nature Reserve in four different months of a year. The bacteria were isolated and then identified by morphological observation and biochemical and molecular analyses. Additionally, drug susceptibility tests were conducted to assess the resistance patterns of the isolated strains. [Results] A total of 17 strains were successfully isolated. These strains exhibited grayish opaque colonies with smooth surfaces and diameters not exceeding 1 mm on the culture media. Positive results were obtained for the sugar fermentation test, and bile esculin hydrolysis test. Negative results were observed for the catalase test and gas production test. All these characteristics aligned with those of Enterococcus, and 16S rRNA gene sequencing further validated the result. The strains of Enterococcus from Chinese monal showed strong resistance to tetracycline, ciprofloxacin, norfloxacin, and levofloxacin. [Conclusion] We successfully isolated Enterococcus strains from Chinese monal and comprehensively analyzed their drug resistance. The findings reveal the current status and drug resistance profile of Enterococcus carried by Chinese monal, giving insights into the disease prevention and protection of Chinese monal.

Abstract:[Background] 2,2'-bipyridines, a class of antimicrobial compounds produced by actinomycetes, have significant antifungal activities and are featured by the 2,2'-bipyridine moiety. No deactivated compound after post-translational modification of the known 2,2'-bipyridines has been discovered, and the self-resistance mechanisms of microorganisms to these compounds remain to be reported. [Objective] To isolate, identify, and determine the antifungal activities of 2,2'-bipyridines from Streptomyces sp. HKIB0006, evaluate the effect of post-translational modification on the antifungal activity, and decipher the self-resistance mechanism of the strain to 2,2'-bipyridines. [Methods] We screened out a Streptomyces strain with anti-fungal activity by the disk diffusion method and then identified the strain by genome sequencing. The target compounds were separated by silica gel column chromatography and HPLC, and their structures were elucidated by MS and nuclear magnetic resonance spectroscopy. The antifungal activities of the isolated compounds were evaluated, and a glycosyltransferase responsible for the self-resistance was revealed by sequence alignment. [Results] Two 2,2'-bipyridines were isolated from the fermentation broth of Streptomyces sp. HKIB0006. One was a new compound named as 11-O-gluconicacid-collismycin A (1), and the other was the known antibiotic collismycin A (2). The antifungal assay of 1 and 2 showed that 1 was inactive and 2 was active. [Conclusion] The antibacterial activity of compounds 1 and 2 was tested. The compound 1 is a gluconic acid modification product of collismycin A, and it is inactive due to the gluconic acid modification. According to genetic analysis, we propose that the gluconic acid modification may represents one of the self-resistance mechanisms of strain HKIB0006 against collismycin A.

Abstract:[Background] Streptococcus mutans stands as the primary pathogen responsible for dental caries. Despite the definite efficacy in treating dental caries, fluoride-containing products pose potential risks to human health. [Objective] We investigated the effects of kuwanon G on the growth and biofilm formation of S. mutans, aiming to facilitate the development of plant-derived anti-caries medications. [Methods] The microdilution and plate counting methods were employed to study the effects of kuwanon G on the viability and acid tolerance of planktonic S. mutans. Furthermore, the influence of kuwanon G on the acid production by glycolysis in S. mutans was gauged by a pH meter. Quantification of biofilm formation was executed via crystal violet staining, and the content of water-insoluble polysaccharides was determined by the phenol-sulfuric acid method. Additionally, laser scanning confocal microscopy was employed to observe the quantity of extracellular DNA, elucidating the effects of kuwanon G on biofilm development. Real-time fluorescence quantitative PCR was employed to assess the mRNA levels of virulence genes in the biofilm. [Results] Kuwanon G showed the minimum inhibitory concentration of 8 μg/mL and the minimum bactericidal concentration of 64 μg/mL against S. mutans. Notably, at higher concentrations, kuwanon G exhibited a potent bactericidal effect on S. mutans. Kuwanon G suppressed the expression of gtfB, gtfC, gtfD, ldh, comD, vicK, atpF, and gbpB in the biofilm milieu, thereby diminishing the synthesis of water-insoluble polysaccharides and impeding biofilm formation. Furthermore, it reduced the content of extracellular DNA in the biofilm matrix and curtailed the overall extent of biofilm formation. Moreover, kuwanon G adversely affected the acid production and acid tolerance of S. mutans. [Conclusion] Kuwanon G can down-regulate the mRNA levels of virulence genes to inhibit the adhesion and loosen the structure of the biofilm, thus exerting inhibitory effects on biofilm formation. This study offers a theoretical basis for further applications of plant-derived natural products.

Abstract:[Background] Autophagy plays a pivotal role in regulating the cell lifespan and aging. Our previous study showed that deletion of the autophagy-related gene ATG8 significantly decreased the replicative lifespan of yeast cells, while the specific mechanism remains to be explored. [Objective] To study the effect and potential mechanism of ATG8 in regulating the cellular lifespan of Saccharomyces cerevisiae. [Methods] The yeast strain overexpressing ATG8 (ATG8 OE) was constructed by homologous recombination. The chronological lifespans (CLS) of ATG8-deleted (atg8Δ) and ATG8 OE strains were measured. The proliferative rates of yeast cells were analyzed by a Bioscreen C MB instrument based on growth curves. The production of reactive oxygen species (ROS) was measured by flow cytometry. The expression levels of oxidative stress-related genes were determined by quantitative RT-PCR (RT-qPCR). The activity of the key apoptotic enzyme, Caspase-3, was measured by the colorimetric method. [Results] The deletion of ATG8 shortened the chronological lifespan and weakened the proliferation of yeast cells. Meanwhile, it increased the production of intracellular ROS, down-regulated the expression levels of oxidative stress-related genes, and enhanced the activity of caspase-3. The overexpression of ATG8 decreased the production of intracellular ROS and up-regulated the expression levels of oxidative stress-related genes in yeast cells. The ATG8 OE strain showed no significant change in the chronological lifespan, proliferation, or caspase-3 activity. [Conclusion] The autophagy-related gene ATG8 plays a role in regulating the chronological lifespan and oxidative stress response of yeast cells.

Abstract:[Background] Soil harbors abundant filamentous fungi which are of great significance for the research on biodiversity and the development of new bioactive compounds. [Objective] To investigate the soil fungal diversity in some areas of China and reserve biological resources. [Methods] Fungal strains were isolated by the dilution-plate method from soil samples. The colony characteristics and microscopic morphology of these strains were observed, measured, described and graphed. The genomic DNA was extracted from mycelia, and the ITS and LSU regions were sequenced. Bayesian inference and maximum likelihood methods were employed to construct phylogenetic trees. [Results] Based on the morphological characteristics and molecular data, three Hypocreales species, Neoacremonium minutisporum, Parasarocladium wereldwijsianum and Sarocladium sasijaorum were identified. [Conclusion] Referring to the reported Hypocreales species in China, we confirmed these three species as new records in China. The results enrich the species diversity of fungi in China.

Abstract:[Background] The bovine diarrheal syndrome, caused by single or multiple pathogen infections, significantly restricts the development of the cattle industry. Rapid and accurate pathogen diagnosis is crucial for disease control. [Objective] To establish a one-step multiplex PCR/RT-PCR method for detecting 11 bacterial and viral pathogens causing bovine diarrhea, facilitating rapid pathogen diagnosis. [Methods] We identified the main pathogens causing bovine diarrhea and appropriate high-coverage primers by literature searching. According to the searching results, we selected primers for the following target genes: Clostridium perfringens α-toxin, Salmonella enterica invA, Escherichia coli K99, bovine enterovirus (BEV) 5'-UTR, bovine astrovirus (BAstV) ORF1a, bovine rotavirus (BRoV) VP6, bovine kobuvirus (BKoV) D4, bovine norovirus (BNoV) ORF1, bovine coronavirus (BCoV) N, bovine torovirus (BToV) N, and bovine viral diarrhea virus (BVDV) 5'-UTR. After optimization of the annealing temperature with the temperature gradient PCR method as well as the primer concentration and cycle number with the single factor test method, we established a one-step multiplex PCR/RT-PCR method for detecting the 11 pathogens. The specificity, sensitivity, and repeatability of the method were evaluated, and then the method was employed to detect clinical samples. [Results] The optimal annealing temperature was 54.4 ℃. The optimal concentrations of primers for the target genes in C. perfringens, S. enterica, E. coli, BEV, BAstV, BRoV, BKoV, BNoV, BCoV, BToV, and BVDV were 0.20, 0.25, 0.25, 0.20, 0.25, 0.25, 0.35, 0.50, 0.25, 0.25, and 0.30 μmol/L, respectively. The optimal number of cycles was 35. The established method demonstrated high specificity, yielding positive results only for the target pathogens and negative results for Mannheimia haemolytica, Streptococcus pyogenes, and Mycoplasma bovine. This method exhibited high sensitivity, with the lowest limits of detection of the recombinant plasmid standards being 7.5×10³, 7.5×10⁴, 7.5×10³, 7.5×10¹, 7.5×10⁴, 7.5×10², 7.5×10³, 7.5×10², 7.5×10², 7.5×10⁴, and 7.5×10³ copies/μL, respectively. The method showed good reproducibility, with consistent inter-batch and intra-batch test results. We then used the established method to detect 490 clinical samples from Jiangsu. The positive rates for BEV, BAstV, BRoV, BKoV, BNoV, BCoV, BToV and BVDV were 0.61%, 0.41%, 0.61%, 0.21%, 27.14%, 3.27%, 0.21%, and 1.02%, respectively. The results of the one-step multiplex PCR/RT-PCR method in clinical samples were consistent with those of single PCR. Sequencing verification of 50 randomly selected positive PCR products confirmed the presence of genes corresponding to the identified pathogens. [Conclusion] We successfully established a one-step multiplex PCR/RT-PCR method for the simultaneous detection of 11 major pathogens causing bovine diarrhea.

Abstract:[Background] High-temperature Daqu is the basis of brewing and fermentation of Maotai-flavor liquor, playing an important role in the whole brewing process. The ester-producing yeast in high-temperature Daqu is associated with the formation of flavor components in liquor. [Objective] To isolate and optimize the fermentation conditions of the ester-producing yeast in high-temperature Daqu. [Methods] We employed the YPD medium, the dilution-coating method, and morphological observation to screen the ester-producing yeast strains in high-temperature Daqu and determine the ester production. After the strain was identified based on molecular evidence, single factor tests and response surface methodology were employed to optimize the fermentation conditions (soluble starch, peptone, and potassium dihydrogen phosphate) of the strain for ester production. [Results] Five ester-producing yeast strains were obtained, in which strain EY3 showcased the highest ester production of 2.16 g/L. Strain EY3 was identified as Wickerhamomyces anomalus, and it demonstrated the highest ester production at 30 ℃, pH 5.0, ethanol concentration of 7%, and inoculation amount of 3%. The optimum carbon source, nitrogen source, and inorganic salt in the medium for ester production by this strain were soluble starch, peptone, and potassium dihydrogen phosphate, respectively. The fermentation conditions for maximum ester production were optimized as 11.4 g/L soluble starch, 3.0 g/L peptone, and 0.4 g/L potassium dihydrogen phosphate, under which the ester production reached (3.43±0.22) g/L. [Conclusion] We screened out a high-yielding ester yeast strain W. anomalus from high-temperature Daqu and optimized the fermentation conditions of this strain, which enriched the diversity of ester-producing yeast.

Abstract:[Background] Cordyceps militaris, as the model species of the genus Cordyceps in the family Cordycipitaceae, has been receiving global attention from researchers. [Objective] This study employs Bibliometric methods to comprehensively examine the current state and future trends of C. militaris research from multiple dimensions. [Methods] we comprehensively retrieved, organized, analyzed, and visualized the papers related to Cordyceps militaris from Web of Science core collection that were published from 2005 to 2024. [Results] In the past two decades, the research on C. militaris expanded beyond cultivation characteristics to interdisciplinary fields, with a particular focus on its active components and pharmacological effects. The artificial cultivation of C. militaris was the main research direction from 2005 to 2009. From 2010 to 2014, the research topics encompassed the pharmacology of the fruiting bodies. After 2015, research topics became diversified, covering optimization, expression, oxidative stress, fungi, antioxidant, chemical-composition, Nf-κB and cell cycle arrest. This shift indicated a transition from cultivation techniques to the in-depth research on biological and medical mechanisms. [Conclusion] The research on C. militaris evolved from conventional cultivation to multidisciplinary fields. The future research is expected to focus mainly on the functional mechanisms, pharmacological effects, and potential medical applications of bioactive substances, providing a scientific basis for the in-depth study and development of C. militaris.

Abstract:[Background] Long-term continuous cropping deteriorates the soil environment, disrupts the soil microbial community balance, and impacts soil ecosystem health. [Objective] This paper employed a bibliometric approach to explore the current research status, hotspots, and development trends related to the effects of continuous cropping on soil microbial communities. [Methods] CiteSpace was used for the visual analysis of the studies on the effects of continuous cropping on soil microbial communities that were published from 2003 to 2023 in the Web of Science (WOS) core collection. The analysis involved publishing countries/regions, institutions, authors, and keywords. [Results] A total of 1 652 publications were screened out. The annual number of publications showed a general increasing trend. China’s annual publications in this research field kept growing, with the number of total publications during 2003–2023 ranking first globally. The Chinese Academy of Sciences was the institution with the largest number of publications. The research team led by Liang Aizhen was highly influential. The Applied and Environmental Microbiology was deemed as the core journal in this field. The research hotspots in this field included the interaction mechanisms between microbial communities and the soil carbon and nitrogen cycling during continuous cropping. The responses of soil microbial communities to tillage measures were also a key research area. [Conclusion] The bibliometric analysis suggests that future research should focus on the integrated use of multiple microbiological methods to investigate how soil microbial communities mediate changes in soil environments and food webs during continuous cropping. Additionally, the regulation and improvement of soil microbial communities by agricultural management practices should be a primary focus in the future studies.

Abstract:Under the background of digital transformation in education, higher education presents a trend of diversification, personalization, and intelligence. Conventional teaching models cannot fully meet the diverse learning needs of students. Modern Engineering Microbiology, as a course leveraging the strengths of the discipline of electronics and information at Xidian University, aims to cultivate outstanding interdisciplinary and innovative talents in biomedical engineering with a distinctive electronic information character, as well as future leaders in the field. The course is application-oriented, interdisciplinary, and challenging in depth. In the current context of artificial intelligence (AI)+education, it is of great significance to integrate AI into the teaching reform. We investigate the blended teaching practice of this course for the students majoring in biomedical engineering at Xidian University, with focus on the application of human-machine collaborative learning based on the large language model and the team’s self-developed deep network intelligent evaluation model throughout the entire process in course design and teaching process. We encourage students to utilize AI to assist learning and fully leverage the role of AI in assisted teaching. The human-machine collaborative learning model enables the interactive learning of students, who can obtain the necessary knowledge and feedback and thus improve the learning efficiency. At the same time, the intelligent evaluation platform throughout the entire process plays a guiding and supervisory role to ensure the quality of learning. The research and practice results indicate that this teaching model improves the quality and effectiveness of teaching and learning, providing a paradigm and experience for the reform of higher education curriculum in the context of digital transformation.

Abstract:Water Treatment Microbiology, a basic course for the undergraduates majoring in water supply and drainage science and engineering, plays an important role in the cultivation of innovative talents in emerging engineering. In response to the problems in the theoretical teaching, practical teaching, and ideological and political teaching of this course, our group carried out teaching research and built a “2+2+N” teaching system for reform and innovation. The system includes visual teaching to consolidate theoretical foundations, innovation of the “pre class-in class-after class” teaching process to guide active learning, and integration of in-class and after-class competitions into the course to strengthen students’ practice and innovative capacity. In addition, the N-dimensional affinity is added to the course to guide the fostering of value. After teaching practice and continuous improvement, this course has been approved as a school-level ideological and political demonstration course, with one provincial educational reform project finished and five educational reform papers published in the past three years. It has received good feedback in terms of enriching the professional knowledge, improving the practice skills, and enhancing the research innovation capacity of students. This innovative system not only enables students to integrate into the classroom but also encourages them to step out of it, the students taught with this system have won third prize or above in seven national level discipline competitions. Moreover, this system helps to form a harmonious teacher-student relationship during the process of mutual communication, achieving entertaining and mutual teaching and learning between teachers and students. This teaching reform and innovation system can provide reference for the current teaching reform of engineering courses in local colleges and universities.

Abstract:Universities, as the cradle for cultivating students’ innovation capability, regard nurturing high-quality talents with professional knowledge, critical thinking, and innovation capability as the fundamental purpose and mission of higher education. For a long time, the higher education in China has lacked the concept of individualized and innovative education, and a comprehensive system for cultivating college students’ innovative thinking and practical skills remains to be established. Following the concept of outcome-based education (OBE), this study takes students as the center to train their innovation capability by virtue of the undergraduate microbiological research interest group and stimulate their innovative thinking by a series of activities. According to the philosophy of combination of teaching and research and the application needs, we guide group students to participate in scientific research and production practice. By participating in the “Challenge Cup”, “Internet+”, national life science competition and other projects, students enhance their research innovation capability and practical skills. Our team combines the training process with the feedback mechanism to perfect the training and evaluation system of students’ innovation capability, providing the train of thought and method for the fostering the innovation capability of science talents in normal universities and ordinary universities.