New Derivatives of Nonactic and Homononactic Acids from Bacillus pumilus Derived from Breynia fruticosa

Six new nonactic and homononactic acid derivatives, ethyl homononactate ( 1 ), ethyl nonactate ( 2 ), homononactyl homononactate ( 6 ), ethyl homononactyl nonactate ( 7 ), ethyl homononactyl homononactate ( 8 ), and ethyl nonactyl nonactate ( 9 ), as well as four known compounds, homononactic acid ( 3 ), nonactic acid ( 4 ), homononactyl nonactate ( 5 ), and bishomononactic acid ( 10 ), were isolated from culture broth of Bacillus pumilus derived from Breynia fruticosa. The structures of new compounds were elucidated by spectroscopic analysis and chemical methods. The optical purities of 1 – 6 were determined by HPLC/MS after treatment with L ‐phenylalanine methyl ester. The dimeric compounds 5 – 9 showed weak cytotoxic activities against five human cancer cell lines (IC₅₀ 19–100 μg/ml).     

Molecular analysis of autochthonous microbiota along the digestive tract of juvenile grouper Epinephelus coioides following probiotic Bacillus pumilus administration

Aims: To evaluate the diversity of dominant autochthonous microbiota along the digestive tract of juvenile Epinephelus coioides following the dietary administration of probiotic Bacillus pumilus for 60 days. Methods and Results: Polymerase chain reaction‐denaturing gradient gel electrophoresis (PCR‐DGGE) with subsequently sequencing analysis was used to assess the gut microbiota. Generally similar DGGE patterns were observed in the foregut, midgut and hindgut of E. coioides, while the similarity dendrogram clearly revealed three different clusters depending on the three compartments of the GI tract. Dietary administration of B. pumilus stimulated its colonization in each compartment of the digestive tract. Samples collected from the probiotic group and the control group showed similar DGGE patterns, and no significant difference in the total number of bands and the Shannon index were detected between the probiotic group and the control group, suggested that B. pumilus exert no significant effect on the gut microbiota. However, various potentially beneficial bacteria, such as uncultured Bacillus sp. clone QJNY94‐like, Nitratireductor sp. YCSC5‐like, Methylobacterium hispanicum‐like and Microbacterium sp. YACS1‐like bacteria were stimulated by probiotic B. pumilus, while the potential harmful Staphylococcus saprophyticus‐like bacterium was depressed. Conclusions: Autochthonous gut microbiota of E. coioides was modulated to some degree, not significant, by probiotic B. pumilus, various potentially beneficial bacteria were selectively stimulated, while one potential harmful species was depressed. Significance and Impact of the Study: This work represents the first report that dietary administration of probiotic B. pumilus modulated the gut microbiota of E. coioides. These findings broaden our understanding of probiotic effects at the gut level, which is helpful in understanding the mechanisms that underpin host benefits.     

环介导恒温扩增法快速检测短小芽孢杆菌

短小芽孢杆菌(Bacillus pumilus)是一种能引起食源性疾病的腐败菌,对其进行快速检测具有重要意义。针对短小芽孢杆菌木聚糖(xynA)基因,设计了4条特异性引物(两条内引物和两条外引物),通过条件优化,首次将一种新颖的核酸扩增技术——环介导恒温扩增技术应用于短小芽孢杆菌的快速检测。采用该技术,63℃温育1h的条件下扩增短小芽孢杆菌DNA,琼脂糖凝胶电泳得到特异性梯度条带。PCR和LAMP的检测灵敏度分别约为162和16.2拷贝每反应。结果表明,该方法检测短小芽孢杆菌特异性强、灵敏度高、操作简便、检测成本低,1h即可完成,有望发展成为快速检测短小芽孢杆菌的有效手段。     

Screening and mutagenesis of a novel Bacillus pumilus strain producing alkaline protease for dehairing

AIMS: To characterize and optimize a novel Bacillus pumilus strain isolated from biological waste which produces protease with excellent dehairing effect. This newly isolated strain could be utilized in the industrial leather dehairing process. METHODS AND RESULTS: Bacterial strains secreting proteases were screened from biological wastes. Positive clones were further characterized by analysing their efficacy in dehairing and effects on collagen integrity. Among 171 colonies tested, a strain BA06, identified as B. pumilus, was picked owing to its efficient dehairing capabilities with minimal impact on collagen. By combined mutagenesis using UV, N-methyl-N'-nitro-N-nitrosdguanidine and Co(60)-gamma-rays, this strain was further improved with regard to its alkaline protease production. The alkaline protease activity of the mutant strain SCU11was greatly improved up to 6000 U ml(-1), in comparison with its parent strain BA06 of 1200 U ml(-1). CONCLUSIONS: By using screening and mutagenesis methods, we have successfully created a B. pumilus strain that can produce high levels of alkaline proteases that are able to efficiently remove hair from skin with minimal damage on the collagen. SIGNIFICANCE AND IMPACT OF THE STUDY: This strain could be used in commercial alkaline protease production for leather dehairing.     

Evaluation of fluorescence in situ hybridization to detect encapsulated Bacillus pumilus SAFR-032 spores released from poly(methylmethacrylate)

Bacillus pumilus SAFR‐032 spores originally isolated from the Jet Propulsion Laboratory spacecraft assembly facility clean room are extremely resistant to UV radiation, H₂O₂, desiccation, chemical disinfection and starvation compared to spores of other Bacillus species. The resistance of B. pumilus SAFR‐032 spores to standard industrial clean room sterilization practices is not only a major concern for medical, pharmaceutical and food industries, but also a threat to the extraterrestrial environment during search for life via spacecraft. The objective of the present study was to investigate the potential of Alexa‐FISH (fluorescence in situ hybridization with Alexa Fluor® 488 labeled oligonucleotide) method as a molecular diagnostic tool for enumeration of multiple sterilant‐resistant B. pumilus SAFR‐032 spores artificially encapsulated in, and released via organic solvent from, a model polymeric material: poly(methylmethacrylate) (Lucite, Plexiglas). Plexiglas is used extensively in various aerospace applications and in medical, pharmaceutical and food industries. Alexa‐FISH signals were not detected from spores via standard methods for vegetative bacterial cells. Optimization of a spore permeabilization protocol capitalizing on the synergistic action of proteinase‐K, lysozyme, mutanolysin and Triton X‐100 facilitated efficient spore detection by Alexa‐FISH microscopy. Neither of the Alexa‐probes tested gave rise to considerable levels of Lucite‐ or solvent‐associated background autofluorescence, demonstrating the immense potential of Alexa‐FISH for rapid quantification of encapsulated B. pumilus SAFR‐032 spores released from poly(methylmethacrylate).     

产细菌素的细菌筛选鉴定及其产物抑菌性能研究

从水体改善制剂样品中筛选出一株产细菌素的芽孢杆菌XDF-2,经生理生化分析和分子生物学鉴定,确定为短小芽孢杆菌。发现该菌产细菌素受发酵时间和培养的影响。对该粗提的细菌素进行特性研究,发现该细菌素只对金黄色葡萄球菌和藤黄微球菌具有抑菌活性。但是该细菌素在不同pH、100℃以下的温度、表面活性剂和有机溶剂的环境下稳定。经酶解试验所得,推测该细菌素具有多肽结构和羧基酯的结构。     

Production of prednisolone by Bacillus pumilus E601 cells incorporated in radiation induced poly(vinyl alcohol g-2 hydroxyethylmethacrylate) cryogels

Immobilization of Bacillus pumilus E601 in poly(vinyl alcohol) (PVA) cryogels at a concentration of 10% grafted with hydroxy-ethylmethacrylate (HEMA) at a concentration of 0.4% using a radiation polymerization technique lead to an increase in the prednisolone yield (46%) compared with the prednisolone yield (38%) produced by immobilized B. pumilus E601 carrier on the surface of the polymer at the same concentrations. The Δ¹-dehydrogenase of B. pumilus E601 was affected by the molecular weight, the irradiation dose and the diameter of the polymer. The storage of immobilized B. pumilus E601 in poly(PVA)/HEMA at −4 °C for 30 days shows a higher yield of prednisolone (80%) as compared with prednisolone yield (75%) at 25 °C at the time of storage.     

玉米浆对转酮酶缺陷型短小芽孢杆菌菌株成链的影响

在研究D 核糖发酵过程中发现 ,培养基中玉米浆的含量直接影响着菌体的形态及D 核糖产量。在不同培养基中加入不同浓度的玉米浆 ,镜检菌株的生长情况 ,并测定发酵培养基中D 核糖的产量。研究表明 ,转酮酶缺陷是突变株在菌体生长过程中出现链状的内因 ,而玉米浆中所含的芳香族氨基酸是转酮酶缺陷型突变株在生长过程中出现链状的外因。     

Biotransformation of isoeugenol to vanillin by a newly isolated Bacillus pumilus strain: identification of major metabolites

A bacterial strain S-1 capable of transforming isoeugenol to vanillin was isolated. The strain was identified as Bacillus pumilus based on biochemical tests, cellular fatty acid composition, riboprint pattern and 16S rRNA gene sequence analyses. In the biotransformation of isoeugenol, vanillin was the main product. With the growing culture of B. pumilus S-1, 10 g l⁻¹ isoeugenol was converted to 3.75 g l⁻¹ vanillin in 150 h, with a molar yield of 40.5% that is the highest up to now. Dehydrodiisoeugenol, a dimer of isoeugenol, was separated by preparative thin layer chromatography and identified by gas chromatography–mass spectrometry. Based on the accurate masses obtained from gas chromatography–high resolution mass spectrometry, two key intermediates, isoeugenol-epoxide (IE) and isoeugenol-diol (ID), were identified by mass spectra interpretations. The biotransformation with resting cells showed that vanillin was oxidized to vanillic acid and then to protocatechuic acid before the aromatic ring was broken. These findings suggest that isoeugenol is degraded through an epoxide-diol pathway.