The ClpY-ClpQ protease regulates multicellular development in Bacillus subtilis.

ATP-dependent proteases play essential roles in both protein quality control and the regulation of protein activities in bacteria. ClpYQ (also known as HslVU) is one of several highly conserved ATP-dependent proteases in bacteria. The regulation and biological function of ClpYQ have been well studied in Gram-negative bacteria, but are poorly understood in Gram-positive species. In this study, we showed that in the Gram-positive bacterium Bacillus subtilis, the ΔclpYQ deletion mutant formed early and robust biofilms, while swarming motility was severely impaired. Colonies of the ΔclpYQ mutant were also much less mucoid on agar plates, indicating the loss of the production of secreted γ-poly-dl-glutamic acid (γ-PGA). Global proteomic analysis using isobaric tags for relative and absolute quantification (iTRAQ) confirmed that a number of proteins involved in motility, chemotaxis and the production of γ-PGA were less abundant in the ΔclpYQ mutant. The results from both iTRAQ and Western immunoblotting showed that levels of the biofilm master repressor SinR were modestly reduced in the ΔclpYQ mutant, but probably significantly enough to alter biofilm regulation due to the ultrasensitivity of the expression of biofilm genes to SinR protein levels. Western immunoblotting also showed that the abundance of CodY, whose gene is clustered with clpYQ in the same operon, was not impacted on by ΔclpYQ. Lastly, our results suggested that, unlike in Escherichia coli, ClpYQ does not play an essential role in heat-shock response in both B. subtilis and Bacillus cereus. In conclusion, we propose that the ClpYQ protease is primarily involved in multicellular development in B. subtilis.

Comparative proteomic analysis reveals intracellular targets for bacillomycin L to induce Rhizoctonia solani Kühn hyphal cell death.

Bacillomycin L, a natural iturinic lipopeptide produced by Bacillus amyloliquefaciens, is characterized by strong antifungal activity against a variety of agronomically important filamentous fungi including Rhizoctonia solani Kühn. To further understand its antifungal actions, proteomes were comparatively studied within R. solani hyphal cells treated with or without bacillomycin L. The results show that 39 proteins were alternatively expressed within cells in response to this lipopeptide, which are involved in stress response, carbohydrate, amino acid and nucleotide metabolism, cellular component organization, calcium homeostasis, protein degradation, RNA processing, gene transcription, and others, suggesting that, in addition to inducing cell membrane permeabilization, iturin exhibits antibiotic activities by targeting intracellular molecules. Based on these results, a model of action of bacillomycin L against R. solani hyphal cells was proposed. Our study provides new insight into the antibiotic mechanisms of iturins.

The Rational Engineered Bacteria Based Biohybrid Living System for Tumor Therapy.

Living therapy based on bacterial cells has gained increasing attention for their applications in tumor treatments. Bacterial cells can naturally target to tumor sites and active the innate immunological responses. The intrinsic advantages of bacteria attribute to the development of biohybrid living carriers for targeting delivery toward hypoxic environments. The rationally engineered bacterial cells integrate various functions to enhance the tumor therapy and reduce toxic side effects. In this review, the antitumor effects of bacteria and their application are discussed as living therapeutic agents across multiple antitumor platforms. The various kinds of bacteria used for cancer therapy are first introduced and demonstrated the mechanism of antitumor effects as well as the immunological effects. Additionally, this study focused on the genetically modified bacteria for the production of antitumor agents as living delivery system to treat cancer. The combination of living bacterial cells with functional nanomaterials is then discussed in the cancer treatments. In brief, the rational design of living therapy based on bacterial cells highlighted a rapid development in tumor therapy and pointed out the potentials in clinical applications.

Integrated analysis of tertiary lymphoid structures and immune infiltration in ccRCC microenvironment revealed their clinical significances: a multicenter cohort study.

BACKGROUND: Tertiary lymphoid structures (TLSs) serve as organized lymphoid aggregates that influence immune responses within the tumor microenvironment. This study aims to investigate the characteristics and clinical significance of TLSs and tumor-infiltrating lymphocytes (TILs) in clear cell renal cell carcinoma (ccRCC). METHODS: TLSs and TILs were analyzed comprehensively in 754 ccRCC patients from 6 academic centers and 532 patients from The Cancer Genome Atlas. Integrated analysis was performed based on single-cell RNA-sequencing datasets from 21 ccRCC patients to investigate TLS heterogeneity in ccRCC. Immunohistochemistry and multiplex immunofluorescence were applied. Cox regression and Kaplan-Meier analyses were used to reveal the prognostic significance. RESULTS: The study demonstrated the existence of TLSs and TILs heterogeneities in the ccRCC microenvironment. TLSs were identified in 16% of the tumor tissues in 113 patients. High density (>0.6/mm2) and maturation of TLSs predicted good overall survival (OS) (p<0.01) in ccRCC patients. However, high infiltration (>151) of scattered TILs was an independent risk factor of poor ccRCC prognosis (HR=14.818, p<0.001). The presence of TLSs was correlated with improved progression-free survival (p=0.002) and responsiveness to therapy (p<0.001). Interestingly, the combination of age and TLSs abundance had an impact on OS (p<0.001). Higher senescence scores were detected in individuals with immature TLSs (p=0.003). CONCLUSIONS: The study revealed the contradictory features of intratumoral TLSs and TILs in the ccRCC microenvironment and their impact on clinical prognosis, suggesting that abundant and mature intratumoral TLSs were associated with decreased risks of postoperative ccRCC relapse and death as well as favorable therapeutic response. Distinct spatial distributions of immune infiltration could reflect effective antitumor or protumor immunity in ccRCC.

Decreased expression of DAB2IP in pancreatic cancer with wild-type KRAS.

BACKGROUND: KRAS mutation plays an important role in the pathogenesis of pancreatic cancer. However, the role of wild-type KRAS in the progression of pancreatic cancer remains unknown. The present study was to investigate the expression of the Ras GTPase activating protein (DAB2IP) in pancreatic cancer and its clinical significance. METHODS: The expression of DAB2IP in pancreatic cancer cell lines and normal human pancreatic ductal epithelial cells was analyzed by Western blotting and real-time quantitative reverse transcription-PCR (qRT-PCR). The KRAS mutational types of pancreatic cancer tissues obtained from pancreatic cancer patients (n=20) were also analyzed. Subsequently, DAB2IP expression was detected in pancreatic cancer tissues, adjacent and normal pancreatic tissues (n=2) by immunohistochemistry, and the relationship between DAB2IP expression and the clinical characteristics of patients was evaluated. RESULTS: Western blotting and qRT-PCR results showed that DAB2IP expression in pancreatic cancer cells with wild-type KRAS was lower than that in those with mutation-type KRAS and normal human pancreatic ductal epithelial cells (P<0.05). Immunohistochemistry showed that DAB2IP expression was lower in pancreatic cancer tissues than that in adjacent and normal pancreatic tissues (Z=-4.000, P=0.000). DAB2IP expression was lower in pancreatic cancer patients with the wild-type KRAS gene than that in those with KRAS mutations (WilcoxonW=35.000, P=0.042). Furthermore, DAB2IP expression in patients with perineurial invasion was lower than that in those without invasion (WilcoxonW=71.500, P=0.028). DAB2IP expression was lower in patients with more advanced stage than that in those with early clinical stage (WilcoxonW=54.000, P=0.002). CONCLUSIONS: DAB2IP expression was reduced in patients with pancreatic cancer compared with those with no cancer. DAB2IP expression was correlated with the KRAS gene, perineurial invasion and clinical stage of the disease. Our data indicated that DAP2IP expression can be used as a potential prognostic indicator and a promising molecular target for therapeutic intervention in patients with pancreatic cancer.

[Identification of lactic acid bacteria in commercial yogurt and their antibiotic resistance].

OBJECTIVE: To identify lactic acid bacteria (LAB) in commercial yogurts and investigate their antibiotic resistance. METHODS: LABs were cultured from 5 yogurt brands and the isolates were identified at the species level by 16S rRNA sequence. Genotyping was performed by repetitive extragenic palindromic PCR (rep-PCR). The sensitivity to 7 antibiotics was tested for all LAB isolates by Kirby-Bauer paper diffusion (K-B method). Meanwhile, 9 antibiotic resistance genes (ARGs), including erythromycin resistance genes (ermA and ermB) and tetracycline resistance genes (tetM, tetK, tetS, tetQ, tetO, tetL and tetW), were detected by PCR amplification in the identified LAB isolates. The PCR products were confirmed by sequencing. RESULTS: Total 100 LABs were isolated, including 23 Lactobacillus delbrueckii ssp. bulgaricus, 26 Lactobacillus casei, 30 Streptococcus thermophilus, 5 Lactobacillus acidophilus, 6 Lactobacillus plantarum, and 10 Lactobacillus paracasei. The drug susceptibility test shows that all 100 isolates were resistant to gentamicin and streptomycin, 42 isolates were resistant to vancomycin, and on the contrary all were sensitive to cefalexin, erythromycin, tetracycline and oxytetracycline. Moreover, 5 ARGs were found in the 28 sequencing confirmed isolates, ermB gene was detected in 8 isolates, tet K in 4 isolates, tetL in 2 isolates, tetM in 4 isolates, tetO in 2 isolates. erm A, tet S, tet Q and tet W genes were not detected in the isolates. Antibiotic resistance genes were found in 53.57% (15/28) sequenced isolates, 2 -3 antibiotic resistance genes were detected in 4 isolates of L. delbrueckii ssp. bulgaricus. CONCLUSIONS: Some LABs were not labeled in commercial yogurt products. Antibiotic resistance genes tend to be found in the starter culture of L. delbrueckii ssp. Bulgaricus and S. thermophilus. All the LAB isolates were sensitive to erythromycin and tetracycline, even though some carried erythromycin and/or tetracycline resistance genes. We proved again that LAB could carry antibiotic resistance gene(s) though it is sensitive to antibiotics.

Shoutai pills for threatened abortion: A protocol for systematic review and meta-analysis.

BACKGROUND: Threatened abortions are a serious health risk for women. Deferiprone tablets are commonly used in the treatment of clinical delivery. Traditional Chinese medicine, a characteristic medical system inherited for thousands of years, often applies Shoutai pills in the treatment of Threatened abortion and has achieved good results. This systematic review and meta-analysis aimed to evaluate the efficacy and safety of Shoutai pills combined with dedrogesterone tablets for the treatment of early preterm abortion. METHODS: Electronic searches of clinical randomized controlled trials in PubMed, Web of Science, MEDLINE, EMBASE, China National Knowledge Infrastructure, Wanfang database, and China Scientific Journal Database (VIP) were conducted. References to the included literature, gray literature in Open Grey, and other relevant literature such as clinical studies registered in ClinicalTrials.gov, were also manually searched. Relevant data were extracted, and a meta-analysis was performed using Reviewer Manager 5.4. RESULTS: The results of this study will be submitted to peer-reviewed journals. CONCLUSION: This study provides high-quality evidence on the efficacy and safety of Shoutai pills in combination with dedrogesterone tablets for the treatment of preterm abortion.

Integrated network pharmacology and molecular docking analyses of the mechanisms underlying the antihypertensive effects of lotusine.

Hypertension is a modifiable cardiovascular risk factor and cause of death worldwide. Lotusine, an alkaloid extracted from a plant used in traditional Chinese Medicine, has shown anti-hypertensive effects. However, its therapeutic efficacy requires further investigation. We adopted integrated network pharmacology and molecular docking approaches with the aim of investigating lotusine's antihypertensive effects and mechanisms of action in rat models. After identifying the optimal intravenous dosage, we observed the effects of lotusine administration on two-kidney, one-clip (2K1C) rats and spontaneously hypertensive rats (SHRs). Based on network pharmacology and molecular docking analyses, we measured renal sympathetic nerve activity (RSNA) to evaluate lotusine's effect. Finally, an abdominal aortic coarctation (AAC) model was established to evaluate lotusine's long-term effects. The network pharmacology analysis identified 21 intersection targets; of these, 17 were also implicated by the neuroactive live receiver interaction. Further integrated analysis showed high lotusine affinity for the cholinergic receptor nicotinic alpha 2 subunit, adrenoceptor beta 2, and adrenoceptor alpha 1B. Blood pressure of the 2K1C rats and SHRs decreased after treatment with 2.0 and 4.0 mg/kg of lotusine (P < 0.001 versus saline control). We also observed RSNA decreases consistent with the network pharmacology and molecular docking analysis results. Results from the AAC rat model indicated that myocardial hypertrophy was decreased with lotusine administration, demonstrated by echocardiography and hematoxylin and eosin and Masson staining. This study provides insights into the antihypertensive effects and underlying mechanisms of lotusine; lotusine may exert long-term protective effects against myocardial hypertrophy caused by elevated blood pressure.

Defining the Potential Targets for Biological Activity of Isoegomaketone Based on Network Pharmacology and Molecular Docking Methods.

Isoegomaketone is a water-soluble natural ketone compound that is commonly present in Rabdosia angustifolia and Perilla frutescens. At present, it is known that isoegomaketone has a wide range of pharmacological activity, but there has been no thorough investigation of its potential targets. As a result, we examined the potential targets of isoegomaketone using the network pharmacology approach. In our study, the TCM Database@Taiwan was utilized to search for the chemical formula. The pharmacological characteristics of isoegomaketone were then evaluated in silico using the Swiss Absorption, Distribution, Metabolism, and Excretion (Swiss ADME) and Deep Learning-Acute Oral Toxicity (DL-AOT) methods, and the potential isoegomaketone target genes were identified using a literature study. Additionally, using the clusterProfiler R package 3.8.1, the Gene Ontology (GO) enrichment analysis and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of target genes were performed. In order to obtain the protein interaction network, we simultaneously submitted the targets to the STRING database. After this, we performed molecular docking with respect to targets and isoegomaketone. Finally, we created visual networks of protein-protein interactions (PPI) and examined these networks. Our results showed that isoegomaketone had good drug-likeness, bioavailability, medicinal chemistry friendliness, and acceptable toxicity. Subsequently, through the literature analysis, 48 target genes were selected. The bioinformatics analysis and network analysis found that these target genes were closely related to the biological processes of isoegomaketone, such as atherosclerotic formation, inflammation, tumor formation, cytotoxicity, bacterial infection, virus infection, and parasite infection. These findings show that isoegomaketone may interact with a wide range of proteins and biochemical processes to form a systematic pharmacological network, which has good value for the creation and use of drugs.

The effect of promoting hydrogen bond aggregation based on PEMTC on the mechanical properties and shape memory function of polyurethane elastomers.

In this work, small molecule diols named PEMTC were synthesized from isophorone diisocyanate, N-(2-hydroxyethyl)acrylamide and trimethylolpropane by a semi-directional method. PEMTC (2-(prop-2-enamido)ethyl N-{3-[({[2-ethyl-3-hydroxy-2(hydroxymethyl)propoxy]carbonyl}amino)methyl]-3,5,5-trimethylcyclohexyl}carbamate) contains hydrogen bond active site and light-initiated C=C. We introduced it as a branch chain block into poly(ε-caprolactone) (PCL). By feeding and monitoring the reaction process, we synthesized a large number of polyurethane elastomers, hydrogen bonds PCL-based elastomer (HPE), which contain a large number of dynamic hydrogen bonds. Under UV irradiation, PEMTC can make HPE molecules aggregate and cross-link, improve the degree of internal hydrogen bonding interaction of HPE materials and endow HPE materials with good elasticity, toughness, heat resistance and shape memory ability. After 270 nm UV irradiation, the elongation at break of HPE materials decreased from 607.14-1463.95% to 426.60-610.36%, but the strength at break of HPE materials increased from 3.36-13.52 to 10.28-41.52 MPa, and the toughness increased from 16.36-129.71 to 40.48-172.22 MJ m-3. In addition, the highest shape fixation rate of HPE after UV irradiation was 98.0%, and the recovery rate was 93.7%.

Bacillus subtilis Cell Differentiation, Biofilm Formation and Environmental Prevalence.

Bacillus subtilis is a soil-dwelling, spore-forming Gram-positive bacterium capable of cell differentiation. For decades, B. subtilis has been used as a model organism to study development of specialized cell types. In this minireview, we discuss cell differentiation in B. subtilis, covering both past research and recent progresses, and the role of cell differentiation in biofilm formation and prevalence of this bacterium in the environment. We review B. subtilis as a classic model for studies of endospore formation, and highlight more recent investigations on cell fate determination and generation of multiple cell types during biofilm formation. We present mechanistic details of how cell fate determination and mutually exclusive cell differentiation are regulated during biofilm formation.

Advances in the Pharmacological Activities and Effects of Perilla Ketone and Isoegomaketone.

As components of a traditional Chinese herbal medicine with many physiological activities, perilla ketone and isoegomaketone isolated from perilla essential oil are important active components of Perilla frutescens. Recent studies have shown that these two compounds have promising antitumor, antifungal, antirheumatoid arthritis, antiobesity, anti-inflammatory, healing-promoting, and other activities and can be used to combat toxicity from immunotherapy. Therefore, the multitude of pharmacological activities and effects demonstrate the broad research potential of perilla ketone and isoegomaketone. However, no reviews have been published related to the pharmacological activities or effects of perilla ketone and isoegomaketone. The purpose of this review is as follows: (1) outline the recent advances made in understanding the pharmacological activities of perilla ketone and isoegomaketone; (2) summarize their effects; and (3) discuss future research perspectives.

Genotype-Specific Recruitment of Rhizosphere Bacteria From Sandy Loam Soil for Growth Promotion of Cucumis sativus var. hardwickii.

The composition and structure of the rhizosphere microbiome is affected by many factors, including soil type, genotype, and cultivation time of the plant. However, the interaction mechanisms among these factors are largely unclear. We use culture-independent 16S rRNA amplicon sequencing to investigate the rhizosphere bacterial composition and the structure of cultivated cucumber Xintaimici (XT) and wild-type cucumber Cucumis sativus var. hardwickii (HD) in four kinds of soils. We found that soil type, cultivation time, and genotype affected the composition and structure of cucumber rhizosphere bacterial communities. Notably, HD showed better physiological features in sandy soil and sandy loam soil than it did in black soil and farm soil at 50 days post-sowing, which was due to its stronger recruitment ability to Nitrospira, Nocardioides, Bacillus, and Gaiella in sandy soil, and more Tumebacillus, Nitrospira, and Paenibacillus in sandy loam soil. Meanwhile, we also found that HD showed a better recruiting capacity for these bacterial genera than XT in both sandy soil and sandy loam soil. Functional predictions indicated that these bacteria might have had stronger root colonization ability and then promoted the growth of cucumbers by enhancing nitrogen metabolism and active metabolite secretion. In this study, our findings provided a better insight into the relationship between cucumber phenotype, genotype, and the rhizosphere bacterial community, which will offer valuable theoretical references for rhizosphere microbiota studies and its future application in agriculture.

Root-secreted bitter triterpene modulates the rhizosphere microbiota to improve plant fitness.

Underground microbial ecosystems have profound impacts on plant health1-5. Recently, essential roles have been shown for plant specialized metabolites in shaping the rhizosphere microbiome6-9. However, the potential mechanisms underlying the root-to-soil delivery of these metabolites remain to be elucidated10. Cucurbitacins, the characteristic bitter triterpenoids in cucurbit plants (such as melon and watermelon), are synthesized by operon-like gene clusters11. Here we report two Multidrug and Toxic Compound Extrusion (MATE) proteins involved in the transport of their respective cucurbitacins, a process co-regulated with cucurbitacin biosynthesis. We further show that the transport of cucurbitacin B from the roots of melon into the soil modulates the rhizosphere microbiome by selectively enriching for two bacterial genera, Enterobacter and Bacillus, and we demonstrate that this, in turn, leads to robust resistance against the soil-borne wilt fungal pathogen, Fusarium oxysporum. Our study offers insights into how transporters for specialized metabolites manipulate the rhizosphere microbiota and thereby affect crop fitness.

Rationally designed structure of mesoporous carbon hollow microspheres to acquire excellent microwave absorption performance.

In this study, we used a novel and facile hard-template etching method to manufacture mesoporous carbon hollow microspheres (MCHMs). We prove that the dielectric ability and microwave absorption of MCHMs can be adjusted by structural characteristics. When the average particle size of MCHMs is 452 nm, the paraffin composite material mixed with 10 wt% MCHMs can achieve a maximum reflection loss value of -51 dB with a thickness of 4.0 mm at 7.59 GHz. When the average particle size of MCHMs is 425 nm, the effective absorption bandwidth of the paraffin composite material mixed with 10 wt% MCHMs can achieve a broad bandwidth of 7.14 GHz with a thickness of 2.5 mm. Compared with other microwave absorbers, MCHMs possess high microwave absorption capacity and broad microwave absorption bandwidth with as low as a 10 wt% filler ratio. This excellent microwave absorption performance is due to the internal cavity and the mesoporous shell of MCHMs. By rationally designing the structure of MCHMs, excellent microwave absorption performance can be acquired. Meanwhile, this design concept based on a rational design of spherical structure can be extended to other spherical absorbers.

Lightweight MWCNT/hollow mesoporous carbon/WPU composite material with excellent electromagnetic shielding performance.

With the rapid increase of intelligent communication equipment, electromagnetic pollution is becoming more and more serious, and the research and application of high-performance electromagnetic shielding materials have attracted great attention from the academic and engineering circles. Traditional metal-based electromagnetic shielding materials have high reflection loss, high density, and are difficult to process. Polymer-based materials with carbon materials as fillers have the advantages of flexibility, light weight, corrosion resistance and low processing costs. They have become the most important materials in the field of electromagnetic shielding in recent years. However, the conductivity of conductive polymer materials is not high. Therefore, improving the electromagnetic shielding performance and the proportion of absorption loss under low density conditions have become key issues for polymer-based electromagnetic shielding materials. MWCNT/MCHMs/WPU composites were prepared by a solution mixing method, with 20 wt%, 40 wt%, 60 wt% MWCNTs and 40 wt% MWCNT/10 wt% MCHMs as fillers. By comparing the effects of different MWCNT content and MCHMs on the dielectric properties, electromagnetic shielding properties and mechanical properties of the MWCNT/MCHMs/WPU composites, the relationship between the structure and properties of the composites has been explored. The 0.6 mm WPU/60 wt% MWCNT composite has an electrical conductivity of 95.4 S m-1 and an electromagnetic shielding effectiveness of 40 dB in the X band. Adding 10 wt% MCHMs to the WPU/40 wt% MWCNT composite material can significantly improve the composite. The δ of the material increased from 51.2 S m-1 to 55.4 S m-1, and the SE increased from 30 dB to 33 dB. The research results show that the increase in MWCNT content and MCHMs is beneficial to improving the electrical conductivity and electromagnetic shielding performance of the composite materials.

Genomic insights into biocontrol potential of Bacillus stercoris LJBS06.

Bacillus spp. have been widely reported with the ability to control plant diseases. In this work, we analyzed the whole genome of LJBS06, which was isolated from grapevine rhizosphere soil. In view of physiological and biochemical characteristics, genome data, and phylogenetic analysis of 16S rRNA, LJBS06 was affiliated with Bacillus stercoris. LJBS06 showed antagonistic activities against a variety of plant pathogens. The inhibition rate of Magnaporthe oryzae was up to 75.05% and the inhibition rates of Colletotrichum gloeosporioides, Coniothyrium diplodiella, and Botrytis cinerea were all above 50% in the plate assays. The genome of LJBS06 had a 4,154,362-bp circular chromosome, with an average GC content of 43.96%, containing an 82,935-bp plasmid with a GC content of 35.18%. The circular chromosome of LJBS06 contained 4231 protein-coding genes, 30 rRNA genes, and 87 tRNA genes, including genes related to the synthesis of plant defense-related enzymes and the promotion of plant growth. Meanwhile, 11 gene clusters involved in biosynthesis of secondary metabolites were present in the genome of LJBS06. In conclusion, our findings indicated that LJBS06 strain had the necessary genetic machinery to control plant pathogens and provided insights for future studies of the biocontrol mechanisms of B. stercoris LJBS06. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-03000-6.

The complete genome sequence of Bacillus velezensis LPL061, an exopolysaccharide-producing bacterium.

Bacillus velezensis LPL061, which shows strong exopolysaccharide (EPS) producing capacity, was isolated from carnations in Beijing, China. The complete genome of LPL061 comprised a single circular chromosome (3,907,268 bp; G+C content of 46.7%) with 3,737 coding DNA sequences, 26 rRNA, and 89 tRNA. According to genome analysis, 12 protein-coding genes which related to polysaccharide biosynthesis in LPL061 were identified. Comparative genome analysis revealed that the EPS biosynthetic gene cluster was relatively conserved among Bacillus species. EPS showed approximately 60% inhibitory activity on the α-glucosidase at 100 µg/mL. The results of quantitative reverse transcription PCR further demonstrated that compared to insulin-resistant model with insulin (500 µg/mL) (without EPS treatment), the insulin-resistant HepG2 cells treated with EPS decreased the expression of phosphoenolpyruvate carboxykinase (PEPCK) from 4.425 to 0.1587, glucose-6-phosphatase (G6Pase) decreased from 4.272 to 0.1929, and glycogen synthase kinase3ß (GSK(3)ß) decreased from 2.451 to 0.993, respectively. Meanwhile, EPS treatment increased GS expression and resulted in intracellular glycogen concentration increased from 28.30% to 86.48%, which further supported that EPS form LPL061 could reduce the concentration of blood glucose effectively. These results could be beneficial for better understanding of the hypoglycemic mechanism of B. velezensis LPL061 EPS and developing an EPS-based anti-diabetic agent in the future.

Complete genome sequence of Bacillus velezensis LPL-K103, an antifungal cyclic lipopeptide bacillomycin L producer from the surface of lemon.

Bacillus velezensis LPL-K103, which shows strong antifungal function, was isolated from the surface of lemon in Beijing, China. The complete genome of B. velezensis LPL-K103 contains a circular chromosome of 3,933,292 bp (46.61% G+C content). According to genomic analysis, 4080 protein-coding genes, 113 RNAs (27 rRNAs + 86 tRNAs), and a non-ribosomal peptide synthase gene cluster involved in antifungal cyclic lipopeptide bacillomycin L biosynthesis were identified. Here, we propose that the biosynthesis pathway of bacillomycin L in LPL-K103 depends on its genome information.

Negative Interplay between Biofilm Formation and Competence in the Environmental Strains of Bacillus subtilis.

Environmental strains of the soil bacterium Bacillus subtilis have valuable applications in agriculture, industry, and biotechnology; however, environmental strains are genetically less accessible. This reduced accessibility is in sharp contrast to laboratory strains, which are well known for their natural competence, and a limitation in their applications. In this study, we observed that robust biofilm formation by environmental strains of B. subtilis greatly reduced the frequency of competent cells in the biofilm. By using model strain 3610, we revealed a cross-pathway regulation that allows biofilm matrix producers and competence-developing cells to undergo mutually exclusive cell differentiation. We further demonstrated that the competence activator ComK represses the key biofilm regulatory gene sinI by directly binding to the sinI promoter, thus blocking competent cells from simultaneously becoming matrix producers. In parallel, the biofilm activator SlrR represses competence through three distinct mechanisms involving both genetic regulation and cell morphological changes. Finally, we discuss the potential implications of limiting competence in a bacterial biofilm.IMPORTANCE The soil bacterium Bacillus subtilis can form robust biofilms, which are important for its survival in the environment. B. subtilis also exhibits natural competence. By investigating competence development in B. subtilis in situ during biofilm formation, we reveal that robust biofilm formation often greatly reduces the frequency of competent cells within the biofilm. We then characterize a cross-pathway regulation that allows cells in these two developmental events to undergo mutually exclusive cell differentiation during biofilm formation. Finally, we discuss potential biological implications of limiting competence in a bacterial biofilm.