Journal of General and Applied Microbiology最新文献

As the first step toward understanding how NADPH levels are regulated in Bacillus subtilis, we sought to obtain mutant strains with enhanced NADPH levels. Our previous study demonstrated that in a strain of B. subtilis expressing bacterial luciferase derived from Photorhabdus luminescens, artificially enhancing NADPH levels enhanced luciferase luminescence in the colonies. In this study, from a library of ethyl methanesulfonate-treated mutants, those with enhanced luciferase luminescence in colonies were isolated, and five isolates were further selected by luminescence in microplate culture. Finally, we measured intracellular NADPH levels of them and found that all the five strains had significantly enhanced NADPH levels compared to the parental strain. In addition, four strains significantly increased total NADP(H) levels. These results demonstrate the effectiveness of our strategy as a methodology for obtaining mutant strains useful for elucidating the mechanisms for regulation of NADPH levels in B. subtilis.

Disc immuno-immobilization is a simple method for typing flagellar antigens from Salmonella enterica. In this study, we successfully adapted this method for Escherichia coli. All eleven strains tested were determined their antigens within 14 h from inoculation. This method improves the efficiency and speed, highlighting its usefulness in clinical laboratories.

p-Anisaldehyde, a fragrance and flavour with important roles in food, cosmetics, and drug industries, is currently synthesized through chemical methods. Production of p-anisaldehyde by chemical oxidation of trans-anethole in industry gives rise to excessive by-products and adverse environmental impacts, whereas biological process would address such problems. Here, we presented a process of biotransformation of trans-anethole for production of p-anisaldehyde. The tao gene encoding for trans-anethole oxygenase (TAO) from Paraburkholderia sp. MR185 was fused with a solubilization tag GST and ProS2, respectively. GST did not exhibit solubility enhancement effect, whereas fusion with ProS2 significantly improved TAO's soluble expression in E. coli and the fusion protein ProS2-TAO-Sil3K accounted for more than 40% of total soluble proteins. ProS2-TAO-Sil3K was purified by simple silica affinity and its activity did not require addition of NADH, NADPH, and FAD. Metal ions Co²⁺, Zn²⁺, Ni²⁺, and Cu²⁺ displayed significant inhibition effect on TAO activity, and addition of Fe²⁺ improved enzyme activity by 32.6%. After induction, engineered E. coli cells were used as whole-cell biocatalyst for transformation of trans-anethole, and the final concentration of p-anisaldehyde reached 10.18 mM (1.38 g/L), with the volumetric productivity of 0.11 g/L/h and conversion rate of 67.9%. These results reveal that the biosynthesis of p-anisaldehyde has a great potential in practice.

Aureispira marina is a marine bacterium with gliding motility isolated from the southern coastline of Thailand. It contained ceramide as a major cellular lipid composed of saturated or unsaturated branched chain 2-hydroxy-fatty acid and sphingosine. The structure of unsaturated 2-hydroxy-fatty acid was investigated in our previous study, but the geometric configuration of the double bond remained unclear. In the present study, 14-methyl-∆²-pentadecenol (∆²-iso-C_16:1-ol) was prepared from D-2-hydroxy-15-methyl-∆³-hexadecenoic acid (D-2-OH-∆³-iso-C_17:1) of the ceramide component, and analyzed by ¹H and ¹³C NMR in comparison with ∆²-trans-hexadecenol (∆²-trans-n-C_16:1-ol) derived from commercially available D-sphingosine. From the coupling constants of protons in the double bond and the chemical shift value of allylic carbon, the configuration of the double bond was determined as trans. Since the structure of 2-hydroxy-fatty acids was clarified, cellular fatty acids of A. marina and A. maritima, another species of the genus Aureispira, were reexamined, and the description on the cellular fatty acid composition of the genus Aureispira in the previous papers (Hosoya et al., 2006, Int. J. System. Evol. Microbiol., 56, 2931-2935; Hosoya et al., 2007, Int. J. System. Evol. Microbiol., 57, 1948-1951) lacking the description of 2-hydroxy-fatty acids was emended.

Protein trafficking to vacuoles in plants and fungi, and to lysosomes in animals, is essential for the maintenance of cellular homeostasis. In Saccharomyces cerevisiae, the vacuolar protein sorting (VPS) pathway has been well studied by using vacuolar carboxypeptidase Y (CPY) as a model, and many VPS genes have been identified. By contrast, the vacuolar protein trafficking pathway in Schizosaccharomyces pombe remains poorly understood. In this study, we identified a novel VPS gene (SPBC1709.03) in S. pombe that is broadly conserved in fungi, but not in S. cerevisiae. Owing to its DUF3844 domain of unknown function, the gene was named vps3844. Disruption mutants of vps3844 had defects in both CPY sorting and incorporation of FM4-64 dye into the vacuolar membrane. Partial deletion analysis of the Vps3844 protein revealed that, within the DUF3844 domain, the region comprising amino acids 354 to 380 is important for protein trafficking to the vacuole. Our findings represent the first report of a VPS gene involved in vacuolar transport that is conserved in fungi, particularly S. pombe, but lacks representation in S. cerevisiae.

For centuries, quinoline alkaloids from the tree bark of Cinchona ledgeriana (C. ledgeriana) have been used in the treatment of malaria. However, unsustainable harvesting and poor growth conditions greatly limit its use as raw materials. Since plant endophytes are known to contribute to the physiology of the host and its metabolism for survival, this study showed the potential of endophytes isolated from C. ledgeriana roots in promoting the germination of Catharathus roseus (C. roseus) seedlings and the biosynthesis of quinoline alkaloid. In this present study, we found that the Enterobacteriaceae family comprised the majority of the bacterial community, with Klebsiella pneumoniae being the most abundant species at the C. ledgeriana roots. Characterization of culturable bacterial endophytes from the C. ledgeriana roots showed that all the isolates displayed plant growth-promoting factors and antifungal activities. Interestingly, chromatographic analyses led to the identification of the quinoline alkaloids producing Achromobacter xylosoxidans (A. xylosoxidans) A1. Moreover, the co-cultures of A. xylosoxidans A1, Cytobacillus solani (C. solani) A3, and Klebsiella aerogenes A6 increased the fresh and dry weight of the C. roseus seedlings. These results suggest that these bacterial endophytes may enhance quinine and quinidine production as well as the growth of the plant host.

Rapid sand filters (RSFs) are employed in a drinking water treatment to remove undesirable elements such as suspended solids and dissolved metal ions. At a closed uranium (U) mine site, two sets of tandemly linked paired RSF systems (RSF1-RSF2 and RSF1-RSF3) were utilized to remove iron and manganese from mine water. In this study, a 16S rRNA-based amplicon sequencing survey was conducted to investigate the core microbes within the RSF system treating the mine water. In RSF1, two operational taxonomic units (OTUs) related to methanotrophic bacteria, Methylobacter tundripaludum (relative abundance: 18.1%) and Methylovulum psychrotolerans (11.5%), were the most and second most dominant species, respectively, alongside iron-oxidizing bacteria. The presence of these OTUs in RSF1 can be attributed to the microbial community in the inlet mine water, as the three most abundant OTUs in the mine water also dominated RSF1. Conversely, in both RSF2 and RSF3, Nevskia sp., previously isolated from the Ytterby mine manganese oxide producing ecosystem, became dominant, although known manganese-oxidizing bacterial OTUs were not detected. In contrast, a unique OTU related to Rhodanobacter sp. was the third most abundant (8.0%) in RSF1, possibly due to selective pressure from the radionuclide-contaminated environment during RSF operation, as this genus is known to be abundant at nuclear legacy waste sites. Understanding the key bacterial taxa in RSF system for mine water treatment could enhance the effectiveness of RSF processes in treating mine water from closed U mines.

Naphthalene is a persistent environmental pollutant for its potential teratogenic, carcinogenic and mutagenic effects. In this study, 10 strains of bacteria capable of degrading naphthalene were isolated from crude-oil contaminated soil. Among them, Pseudomonas plecoglossicida 2P exhibited prominent growth with 1000 mg/L naphthalene as the sole carbon source and degraded 94.15% of naphthalene in 36 h. Whole genome sequencing analysis showed that P. plecoglossicida 2P had a total of 22 genes related to naphthalene degradation, of which 8 genes were related to the salicylic acid pathway only, 5 genes were related to the phthalic acid pathway only, 8 genes were common in both the salicylic acid and phthalic acid pathways, and 1 gene was related to the gentisic acid pathway. P. plecoglossicida 2P was applied in a two-phase partition bioreactor (TPPB) to degrade naphthalene in wastewater. The optimal operating conditions of the reactor were obtained through response surface optimization: initial naphthalene concentration (C₀) =1600 mg/L, bacterial liquid concentration (OD₆₀₀) = 1.3, and polymer-to-wastewater mass ratio (PWR) = 2%. Under these conditions, the naphthalene degradation rate was 98.36% at 24 h. The degradation kinetics were fitted using the Haldane equation with a high coefficient of determination (R²=0.94). The present study laid foundations for naphthalene degradation mechanism of genus Pseudomonas and its potential application in TPPB.

We have successfully isolated two novel compounds, 24R005A (1, C₁₃H₁₄O₄) and 24R005B (2, C₁₃H₁₃ClO₄), from Streptomyces sp. 24R005, using fish (anchovy) powder as a medium. In this study, we evaluated the use of fish (anchovy) powder as a fermentation material for producing bioactive compounds. Spectroscopic analyses revealed that the two compounds share a common skeletal structure. However, each compound contains unique branched side chains. Furthermore, compounds 1 and 2 exhibit moderate radical-scavenging activity for 1,1-diphenyl-2-picrylhydrazyl (DPPH), with ED₅₀ values of 200 and 130 μM, respectively.

Polyamide 4 (PA4) is expected to solve the issue of marine plastic pollution due to its excellent mechanical properties and biodegradability. In this study, to reveal the mechanism of PA4 biodegradation in the marine environment, we isolated 5 strains of PA4-degrading bacteria belonging to Aliiglaciecola, Dasania, and Pseudophaeobacter from a marine environment. The isolated 5 strains are novel PA4-degrading bacteria that are phylogenetically distinct from those isolated in previous studies. In addition, we compared the PA4-degrading activities and structures of the PA4-degrading enzymes secreted by the 5 strains and PA4-degrading strains isolated in our previous study. The PA4-degrading activity in the supernatant of the cultivation solutions differed among the strains. Native-PAGE and zymography using a polyacrylamide gel containing a PA4 emulsion demonstrated that PA4-degrading enzymes are classified into no less than three types of structures. These results suggested that marine PA4-degrading bacteria have multiple PA4-degrading enzymes. Our findings will contribute to a better understanding of the microbial degradation of PA4 in the marine environment.