Abstract:Plant rhizosphere refers to the interface between plant roots and soil, where the life activity and metabolism of root system have the most direct and strong influence on soil. Physical, chemical and biological properties of rhizosphere are different from bulk soil. Many microorganisms that interact with plants in this region are known as rhizosphere microorganisms. Rhizosphere microorganisms play an important role in plant growth and biological control of plant diseases and insect pests. In this article, we first summarize the research status of rhizosphere microorganisms. Second, we introduce the classic and the latest research methods used in the rhizosphere microbiome, such as root box method, isotope techniques, high-throughput sequencing, quantitative analysis of microbial abundance, high-throughput isolation and culture. Third, we discuss the influencing factors on rhizosphere microorganisms, such as plant root exudates (carbohydrate, amino acids, flavonoids, phenols, hormone and its signal substance), soil physical and chemical properties. Fourth, we discuss the interaction between rhizosphere microorganisms and plant, and introduce the effects of rhizosphere microorganisms on plant growth stress resistance. Last, we address the problems in rhizosphere microbiome study and future development in this field.

Abstract:African swine fever (ASF) is a highly contagious, fatal infectious disease of pigs caused by African swine fever virus (ASFV). There is no commercialized vaccine or economical control strategy other than slaughter, which poses a serious threat to the healthy development of the swine industry in China. A major difficulty in the development of ASF vaccines is insufficient knowledge about the virulence-associated genes, the pathogenesis and immunoevasion mechanisms of ASFV. This review summarizes the recent progress in ASFV immunoevasion, discusses the immunoevasion-related functions of ASFV genes and the encoded proteins, which will help understand immunoevasion strategies and mechanisms of ASFV, and provide insights into pathogenesis and vaccine development.

Abstract:Staphylococcus argenteus was identified and proposed to be a novel species closely related to S. aureus in 2015, and considered as a genetically divergent lineages within S. aureus. Most of S. argenteus strains are isolated from humans, could cause the same infections and foodborne illnesses as S. aureus, and harbor homologs of most virulence genes found in S. aureus. Because of its associations with human health, S. argenteus is a focus in more and more studies and reported in more than 20 countries/regions. It is still unclear about its global distribution, original habitat and dissemination, because this species is just recognized several years ago and is difficult to be distinguished from S. aureus. This paper reviews the recent progress on the emergence, distribution, pathogenicity and identification methods of S. argenteus.

Abstract:Lactulose, a reducing disaccharide, is composed of D-galactosyl and D-fructosyl moieties via the ligation of a β-1,4 glycosidic bond. The concentrated lactulose solution can be used to treat chronic constipation and hepatic encephalopathy. It is an over-the-counter drug (OTC) worldwide, resulting in a massive requirement for its production. Lactulose is also a prebiotic that benefits human intestinal flora. Current industrial production of lactulose relies on chemical catalysis, with harmful catalyst and difficult in the down-stream processing. Recently, cellobiose 2-epimerase (CE) is recognized as an efficient biocatalyst for lactulose production using lactose as the substrate. This technique is environmentally friendly and composed of simpler procedures, showing promising future for an industrial application. This paper reviews recent developments in CE enzyme research, and the biotechnological route of lactulose synthesis.

Abstract:Celiac disease is an autoimmune-mediated enteropathy caused by ingestion of gluten in genetically susceptible population. Accumulating evidence has shown that the second human genome, intestinal microbiota, plays an important role in the development of celiac disease. Compared with the healthy population, patients with celiac disease show higher diversity in intestinal microbiota, but more proinflammatory bacteria and less beneficial bacteria in the gut, paralleled by the altered function and metabolism of gut microbiota. However, it is unclear how this dysbiosis of microorganism occurs and whether this change promotes celiac disease. This review is aimed at summarizing current evidence on the relationship between the gut dysbiosis and celiac disease, thus contributing the prevention and treatment of this disorder by regulating microbiota.

Abstract:Tuberculosis caused by Mycobacterium tuberculosis (Mtb) remains a serious global infectious disease. The world health organization estimates that 1.5 million people died from the disease in 2019. The biology of Mtb can inform new measures against tuberculosis. To summarize the progress of CRISPR/Cas-associated genes (CRISPR/Cas) system in Mtb for better tuberculosis control tools development and biology study of Mtb. Current publications and progress in our lab were retrieved and compared. Clustered regularly interspaced short palindromic repeats, CRISPR/Cas system, well known bacterial adaptive immunity system widespread in Mycobacteria including M. tuberculosis, was developed as gene editing tool. We summarized the biology of the endogenous type III-A CRISPR-Cas systems in Mycobacteria, as well as CRISPR/Cas gene editing tool application in Mtb basic and applied studies, with focus on its potential for novel measures against tuberculosis. CRISPR/Cas is burgeoning focus in M. tuberculosis study and promising tool for better tuberculosis control.

Abstract:The discovery of complete ammonia oxidation (comammox) bacteria was one of the major hotspots in microbial nitrogen cycle fields, and thus caused widespread concern about their global distribution, phylogenetic characteristics, physiological and biochemical properties. This review first summarizes the distribution pattern of comammox bacteria in soil, surface water, wastewater treatment systems, etc, then expounds the mechanism of interaction and niche separation between comammox bacteria and traditional nitrifying microorganisms from the aspects of substrate affinity and metabolic diversity. Based on the above, coupling of comammox bacteria with other denitrifying microorganisms, e.g, denitrifying bacteria, anaerobic ammonia oxidizing bacteria, and anaerobic methane oxidizing bacteria, is proposed to achieve advanced wastewater nitrogen removal under low ammonia and dissolved oxygen conditions, thereby saving energy consumption and reducing greenhouse gas emission. Future research should focus on identifying the physiological and biochemical characteristics of comammox bacteria, evaluating the ecological function and contribution to the biogeochemical nitrogen cycle, and then sequentially tapping their application potential in biological wastewater treatment process and other fields.

Abstract:The species of microalgae are very rich and diverse. Microalgae generally contains chloroplasts for photosynthesis. Studies have shown that microalgae's ability to fix CO₂ is 10 times that of land plants. Microalgae is widely used in renewable energy, biomedicine, food industry and environmental monitoring due to its rich metabolites and unique physiological characteristics. However, how to scale-up microalgae under the premise of controlling cost has become a difficult problem in the application. Therefore, we review the research progress of microalgae scale culture in the recent ten years from the biochemical characteristics of microalgae and its application in various fields, the three growth modes of microalgae, the application of photobioreactors and the harvesting of microalgae, with a view to providing a basis for the further development and utilization of microalgae.

Abstract:Listeria monocytogenes is an important foodborne pathogenic microorganism. Its oxidative stress tolerance is the key ability to survive and mediate pathogenesis. Increasing of intracellular Reactive Oxygen Species (ROS) leads to imbalance of redox homeostasis, oxidative stress, and further damage of proteins as well as other large biological molecules. Cysteine and sulfur-containing amino acid of proteins are especially sensitive to ROS, and formation of disulfide bonds is an oxidation reaction, with two electrons removed from cysteine residues, which contributes to stabilizing protein structures and preventing protein damages. Antioxidant repair usually refers to the oxidation reaction of cysteine residues, consisting of disulfide bonds formation and breaking. Thioredoxin (Trx) family, including thioredoxin, glutaredoxin, and Dsb-like system, is a widely-distributed system responsible for oxidation repair in microorganisms. Here, we review the most recent research development of the Trx family from bacterial species and present our opinions of this family in L. monocytogenes, to better understand the regulatory network of Listeria during environmental adaption and host infection.

Abstract:As a key post-translational modification involved in regulation of gene expression, protein phosphorylation plays critical roles in bacterial physiological processes. Here we summarize the types of phosphorylation modification in bacteria, two-component signal transduction, tyrosine phosphorylation as well as serine/threonine phosphorylation, and discuss the regulatory mechanisms and functions of phosphorylation modifications in bacterial cellular processes in recent years. This review is helpful to understand the phosphorylation regulation of bacterial post-translational modifications and its relationship with the control of bacterial infections in the future.

Abstract:Noble metal-based nanomaterials (NMNs) have a broad spectrum of antibacterial properties. NMNs can inhibit bacteria by means of nano-perforation, disrupting membrane stability, stimulating the generation of reactive oxygen species, binding to biological molecules such as DNA, and releasing toxic ions. Under the selective pressure of NMNs, bacteria evolve the resistance to NMNs inhibition. The resistance of bacteria to NMNs can be exhibited through electrostatic repulsion, secreting adsorption protein, activation of antioxidant enzymes, efflux of toxic particles, and quorum sensing. Here we summarize the antibacterial properties of NMNs, discuss bacterial resistance to NMNs, and indicate the shortage of current researches on bacterial resistance to NMNs, to assist rational development and application of antimicrobial NMNs.

Abstract:[Objective] The purpose of this study is to express Mb0950c protein by prokaryotic expression system, and to evaluate its immunogenicity by mouse model, further to establish a serological indirect enzyme-linked immunosorbent assay (ELISA) for clinical detection of bovine tuberculosis. [Methods] The prokaryotic expression plasmid of pET32a-Mb0950c was constructed and transformed into BL21(DE3) to induce protein expression. The immunogenicity of the protein in mice was analyzed by flow cytometry (FCM) and ELISA. And then an indirect ELISA method based on Mb0950c was established and used in clinical trial. [Results] SDS-PAGE and Western blotting showed that Mb0950c protein was successfully obtained and had good immunogenicity. The analysis of FCM showed that Mb0950c protein upregulated the expression of CD69 on the surface of T cells. The results of ELISA showed that the protein could induce the secretion of IFN-γ and IL-4, at the same time, it could induce the body to secrete specific antibodies, and it mainly belongs to IgG1. Total 192 serum samples were detected by indirect ELISA based on Mb0950c and the results showed that the positive coincidence rate, negative coincidence rate and total coincidence rate were 65.7%, 97.9% and 72.4% respectively. [Conclusion] Mb0950c protein is expressed in the prokaryotic expression system and it induces Th1-and Th2-type immune response in mouse models. Based on this protein, an indirect ELISA method for serological detection of bovine tuberculosis has been established.

Abstract:[Objective] Aconitate isomerase (AI) can mediate the biosynthesis of trans-aconitic acid (TAA), a multi-bioactive small molecule with many application potentials. Although the first AI gene (tbrA) was just identified in the bacterium Bacillus thuringiensis, the property of AI remains unclear. Here we characterized TbrA to understand the AI catalysis. [Methods] We expressed His₆-tagged TbrA protein in Escherichia coli Rosetta strain and purified it with Ni²⁺ affinity chromatography. We characterized TbrA in vitro and applied HPLC to determine product formation and enzyme activity. [Results] The optimal pH, temperature and NaCl ionic strength for TbrA activity were determined to be 8.0, 37℃ and 25 mmol/L, respectively. TbrA exhibited a cold tolerance, retaining nearly 60% of its enzymatic activity at 10℃. Notably, TbrA activity was significantly enhanced by Mg²⁺, Ca²⁺ and DTT, but strongly inhibited by Fe²⁺, Cu²⁺, Zn²⁺ and Mn²⁺. K_m, V_max, k_catand k_cat/K_m for TbrA forward reaction[from cis-aconitic acid (CAA) to TAA] were 6.25 mmol/L, 1.39 μmol/(L·s), 4.08 1/s and 0.65 L/(mmol·s), and for TbrA reverse reaction (from TAA to CAA) were 71.50 mmol/L, 4.17 μmol/(L·s), 12.25 1/s and 0.17 L/(mmol·s), respectively. [Conclusion] The AI enzyme TbrA can achieve maximum activity under mild conditions, and favors for TAA formation. This study quantitatively described the catalytic properties of TbrA protein, which provided the basis for its potential application in TAA industrial bioproduction.

Abstract:[Objective] To investigate antibiotic resistance and epidemiological characteristics of bla_CTX-M gene of Escherichia coli from retail livestock and chicken meat in Guangzhou. [Methods] E. coli was isolated from retail livestock and chicken meat samples in different regional retail markets and fresh supermarket in Guangzhou. Those isolates were identified by the gene phoA and sequencing. Drug susceptibility was determined by agar diffusion and broth microdilution method. The bla_CTX-M gene was detected using PCR amplification. Whole genome sequencing was performed in all bla_CTX-M-positive E. coli.[Results] A total of 241 strains were isolated from 323 meat samples. The drug susceptibility test shows that those strains were highly resistant to ampicillin (63.07%), doxycycline (47.72%) and sulfamethoxazole/trimethoprim (43.15%). The prevalence of bla_CTX-M gene was 3.32% (n=8), which contained 4 bla_CTX-M-14-positive strains, 3 bla_CTX-M-65-positive strains and 1 bla_CTX-M-55-positive strain. Eight CTX-M-producing E. coli were divided into four distinct ST types, which carried multiple resistance genes and virulence genes.[Conclusion] The contamination of E. coli was serious in retail meat. All CTX-M-producing E. coli were multi-resistant strains, and the dissemination of bla_CTX gene was mainly due to horizontal transfer, which needs further surveillance.

Abstract:[Objective] In this study, we investigated the antibacterial properties of 4,6-diamino-2mercaptopyrimidine-functionalized gold nanoparticles (AuNPs) against Gram-negative multidrug-resistant (MDR) bacteria. [Methods] Minimum inhibitory concentration (MIC) of AuNPs against 4 clinically isolated Gram-negative MDR bacteria was determined by the broth dilution method. Time-kill curves within 24 h were drawn by plate counting after treatment with different concentrations of AuNPs. Employing Pseudomonas aeruginosa as a representative strain, the damage of AuNPs to bacterial cell components was studied using laser scanning confocal microscopy, transmission electron microscopy, and gel electrophoresis. The evolution of potential drug resistance of bacteria to AuNPs was evaluated by repeatedly exposed to AuNPs at a sublethal concentration. The biocompatibility of AuNPs to mammalian cells was evaluated by MTT analysis. [Results] The average size of synthesized AuNPs was 6.8 nm, and the zeta potential was +38.4 mV. The AuNPs showed timeand concentration-dependent antibacterial activity against the 4 Gram-negative MDR bacteria, with MIC values ranging from 4 to 8 μg/mL. The AuNPs killed these MDR bacteria through a combined mechanism including cell membrane destruction and DNA damage. No resistance development toward the AuNPs was demonstrated even after continuous passage for 30 d. Preliminary cytotoxicity assay results showed that AuNPs have excellent biocompatibility to mammalian cells, and the survival rate of normal L02 liver cells and normal AT II lung cells is still more than 85% when the concentration reaches 256 μg/mL. [Conclusion] Small molecule-mediated AuNPs have excellent antibacterial activity against Gram-negative MDR bacteria, and potential application value in dealing with severe MDR bacterial infection.

Abstract:[Objective] To optimize the transformation system of a phosphate-solubilizing fungus Penicillium brocae and modify the strain with molecular tag for root colonization analysis; to evaluate the growth promotion effects of P. brocae. [Methods] We optimized ATMT (Agrobacterium tumefaciens-mediated transformation) parameters to get P. brocae transformants, we confirmed the transformants and its root colonization with molecular tags; we used TAIL-PCR (thermal asymmetric interlaced-PCR) to characterize T-DNA insertion site; we evaluated the growth promotion effects of P. brocae by Petunia hybrida pot experiments with treatments of T1 (10 mL water), T2 (10 g/L phosphate rock, 10 mL), T3 (10⁶ spores/mL, 10 mL) or T4 (10 g/L phosphate rock and 10⁶ spores/mL, 10 mL).[Results] We got 44 transformants per 10⁶ spores by parameters optimization and confirmed the transformants with gfp and hyg gene cloning, GFP fluorescence detecting and GUS staining. We observed the colonization of P. brocae on P. hybrida root surface by GUS staining. The insertion site flanking sequence of a transformant characterized with phosphate solubilization capacity significantly decreasing was obtained by TAIL-PCR. P. hybrida fresh weight of T4 were 35.4%, 35.6% and 21.4% higher than that of T1, T2 and T3, the dry weight of T4 were 25.1%, 37.8% and 26.0% higher than that of T1, T2 and T3, respectively. There was no significant difference among the fresh or dry weight of T1, T2 and T3 (P ≤ 0.05). [Conclusion] We optimized the ATMT system and observed the root colonization of P. brocae. The application of P. brocae spores and phosphate rock together but not alone, could promote the growth of P. hybrida and this gave a hint that P. brocae may enhance the fertilizer efficiency of phosphate rock in soil.

Abstract:Bovine lactoferrin peptides are short peptides with broad spectrum antimicrobial activity produced by hydrolysis of bovine lactoferrin by pepsin. As a food-grade microorganism, Lactococcus lactis has a natural probiotic effect, and it is an ideal bacterial vector for expressing bovine lactoferrin peptide. [Objective] To investigate the antibacterial activity of bovine lactoferrin peptide expressed by the recombinant L. lactis pAMJ399-LFcinBA/MG1363. [Methods] We used the standard of bovine lactoferrin peptide to determine a quantitative standard curve to determine the concentration of the bovine lactoferrin peptide expressed in the supernatants of the recombinant strain, and used the Oxford Cup method to determine the inhibitory effects of the recombinant bovine lactoferrin peptide on 35 strains of bacteria including Escherichia coli and Staphylococcus aureus. And we used the broth microdilution method to determine the minimal inhibitory concentrations. We explored the effects of the recombinant lactoferrin peptide on a number of pathogenic bacteria using scanning electron microscopy, transmission electron microscopy, and fluorescence microscopy. We used gel retardation assay and adhesion test to investigate the effects of the recombinant lactoferrin peptide on bacterial DNA and adhesion. We used CCK-8 to detect its toxic effects on RAW 264.7 cells and to determine the hemolytic rate of red blood cells in mice. [Results] The results showed that the concentration of the bovine lactoferrin peptide expressed in the supernatants of the recombinant strain was 24.39 μg/mL and the recombinant lactoferrin peptide showed inhibition on 25 strains of pathogenic bacteria used in the experiment. Its antibacterial concentration range was 16-128 μg/mL, but had no obvious inhibitory effect on 9 kinds of lactic acid bacteria and 1 strain of Enterococcus faecalis. It destroyed the integrity of E. coli, Staph. aureus, Pasteurella multocida and Salmonella pullorum, targeting the bacterial membranes. It could bind to bacterial DNA and inhibit bacterial adhesion to Caco-2 and IPEC cells. The recombinant lactoferrin peptide had no obvious cytotoxic effect on mouse red blood cells and RAW 264.7 cells.[Conclusion] The recombinant bovine lactoferrin peptide had the same antibacterial activity as the standard of bovine lactoferrin peptide. It could inhibit or kill bacteria by targeting the membranes of bacteria, intracellular nucleic acids and inhibiting the adhesion of bacteria to normal cells, exerting its broad-spectrum antibacterial activity, and had no obvious cytotoxicity to eukaryotic cells.

Abstract:[Objective] To investigate antimicrobial activity and metabolites of Streptomyces sp. from the termite nest. [Methods] Antimicrobial activities of fermentation broth against four pathogenic microbes were tested by Oxford cup method. The bioactive strain T12 was identified by molecular biology 16S rRNA sequence analysis. The active metabolites of T12 were isolated and purified from ethyl acetate crude extracts by various chromatographic methods, and their chemical structures were identified by mass spectrometry and nuclear magnetic resonance spectroscopy, and their antimicrobial activities were determined by filter paper method.[Results] T12 was identified as Streptomyces sp.. Under the concentration of 30 μg/filter paper, the ethyl acetate extract of T12 had potent antibacterial activities against Staphylococcus aureus and Candida albicans with inhibition zone diameters of 20.1 mm and 17.4 mm, respectively. Two metabolites including geldanamycin (1) and 17-O-demethylgeldanamycin (2), were isolated and characterized from fermentation broth. Both metabolites 1 and 2 exhibited good antimicrobial activity at the concentration of 30 μg/filter paper. The inhibition diameters of compounds 1 and 2 against S. aureus were 14.6 mm and 14.5 mm respectively, which were equivalent to those of gentamicin sulfate as positive control. The inhibition diameters of compounds 1 and 2 against C. albicans were 10.9 mm and 13.9 mm respectively, which were comparable to those of the referenced amphotericin. [Conclusion] Strain T12 could be potentially developed as a new microbial microbicide.

Abstract:[Objective] To compare the structural characteristics of intestinal flora between patients with polycystic ovary syndrome and healthy controls, and to explore the relationship between differential flora and biochemical immune indicators in patients with polycystic ovary syndrome. [Methods] From March 2018 to May 2019, 23 patients with polycystic ovary syndrome and 23 healthy women who were treated at the gynecological clinic of The Second Affiliated Hospital of Guangzhou University of Chinese Medicine were selected. Each patient retained a stool specimen and a blood specimen at the initial diagnosis. Fecal specimens were extracted from DNA for metagenomic sequencing, and serum was used to detect sex hormones and inflammatory factors. Biological information analysis was used to compare the structural characteristics of intestinal flora between patients with polycystic ovary syndrome and healthy controls. [Results] The intestinal flora of patients with polycystic ovary syndrome and healthy controls were significantly different at the Phylum, Genus and Species levels. There are 4 species with a marked difference between the two groups. The relative abundance of Bacteroides vulgatus and Escherichia fergusonii in the polycystic ovary syndrome group is higher than that in the control group, and these two bacteria are closely related to the pathogenesis of polycystic ovary syndrome. While the relative abundance of Eubacterium ventriosum and Subdoligranulum unclassified is significantly lower than that of the healthy control group. Among them, Bacteroides vulgatus, Eubacterium ventriosum and Subdoligranulum unclassified are correlated with each other, and there are correlations between different species and biochemical immune indexes. It was found that the combined of five species of Ruminococcus bromii, Gordonibacter pamelaeae, Desulfovibrio piger, Eubacterium ventriosum, Prevotella stercorea had good efficacy in the early diagnosis of polycystic ovary syndrome patients. Genetic analysis found that PWY 7228 super-pathway of guanosine nucleotides de novo biosynthesis I gene pathway is enriched in the polycystic ovary syndrome population. [Conclusion] The intestinal flora and biochemical immune parameters of patients with polycystic ovary syndrome are significantly different from those of the control group, and have a certain correlation, which provides a direction for the comprehensive diagnosis and treatment of patients with polycystic ovary syndrome.

Abstract:[Objective] To explore the structure, function and relationship of bacterial community in sclerotia, cortices and habitat soil of Cordyceps cicadae under the natural ecology. [Methods] High-throughput sequencing of bacterial 16S rRNA amplification fragments was performed to analyze the composition, diversity and potential functions of bacterial community in sclerotia, cortices and habitat soil of Cordyceps cicadae in Mount Dajiang and Guiyang Forest Park. [Results] A total of 562 bacterial genera were detected in the sclerotia samples, 521 genera were detected in the cortices samples, and 578 genera were detected in the hyphosphere soil samples. The bacterial community structure of each group of samples from the two places was similar. In the sclerotia samples, Pseudomonas and Sphingobacterium were the dominant populations. Pseudomonas and Aminobacter were the dominant populations in the cortices samples. The norank_c_Acidobacteria and norank_f_Xanthobacteraceae were the dominant genera in the soil samples. Venn analysis showed that the hyphosphere soil samples included most genera of the cortices samples, and the sclerotia samples had more endemic genera, such as Wolbachia and Rickettsia. Furthermore, the PICRUSt function prediction results showed that there are a total of 24 gene function families, which are mainly related to the metabolism and transportation of material and energy, as well as the occurrence and regulation of cell behavior. [Conclusion] Bacteria in the Cordyceps cicadae and its microhabitat had rich diversity. Their potential function may be related to the metabolism of nutrients and played important roles in the individual growth and development of the Cordyceps cicadae. The results have reference value for supplementing the ecological information data of Cordyceps cicadae and imitateing wild cultivation.