Hfq plays a fundamental role in bacterial cell physiology. It can stimulate or repress the expression of certain target genes, and there is a possibility that Hfq regulates the oxidative stress response. However, how Hfq functions that in Vibrio parahaemolyticus remains speculative. In this paper, we explain the functions Hfq plays in V. parahaemolyticus in the gene expression of superoxide dismutase gene and catalase gene, comparing the hfq deletion mutant strain to the parental strain. The results show that the hfq deletion mutant V. parahaemolyticus has a stronger ability to resist H2O2. Superoxide dismutase (SOD) and catalase (CAT) activities in the hfq deletion mutant were remarkably higher than in the parental strain. Genetic experiments indicated that the gene expression of sod and kat was up-regulated in the mutant strain. These results indicate that Hfq down-regulates CAT and SOD activity, and Hfq is associated with the oxidative stress response.
Transposons play a significant role in the evolution of bacterial genomes. Quantifying frequency of transpositional events caused by a transposon will facilitate understanding its role. Here, we report successful measurement of the frequency of IS1 transposition using “GFP hop-on assay” in which transposition-dependent GFP expression is monitored by FACS. This assay allows easy assessment of IS transposition into the chromosomal DNA on a single-cell scale; this is an advantage over other conventional methods to measure transposition frequency.
Facultatively anaerobic bacterial strains (A48T, RR25 and RR54) were isolated from roots of living rice plants in an irrigated rice-field in Japan. The three strains had identical 16S rRNA gene sequences and showed almost the same phenotypic properties examined. Cells of the strains were Gram-negative, non-spore-forming rods. Reproduction of cells was by binary fission as well as by budding. Cells occurred singly or in pairs arranged angularly. Some cells, including dividing cells, were motile with a single polar flagellum. Cells developed a polar prostheca (stalk) with a holdfast-like structure and the cell with the stalk budded a daughter cell. The strains were chemoorganotrophs and utilized various sugars as growth substrates. The strains fermentatively produced acetate and lactate as well as small amounts of ethanol and H2 from the substrates. Growth temperature and pH ranges for growth were 15-40ºC and pH 5.5-7.3 with optimum growth at 30-35ºC and pH 6.8. NaCl concentration range for growth was 0-1.0% (wt/vol) with an optimum at 0% (wt/vol). Catalase and oxidase activities were not detected. The strains reduced Fe(III) to Fe(II) in the presence of glucose, while they did not reduce nitrate, fumarate, malate or sulfate. The major cellular fatty acids of the strains were C18:1ω7, anteiso-C15:0, iso-C15:0, C16:0 and C18:0. Ubiquinone Q-10 was the major respiratory quinone and the genomic DNA G + C contents were 53.4-55.4 mol%. Phylogenetic analysis based on 16S rRNA gene sequences placed the strains in the class Alphaproteobacteria and the strains formed a novel deep branch in the phylogenetic trees constructed. Based on the differences in 16S rRNA gene sequences and phenotypic properties of the novel strains from those of their relatives, we proposed that the strains be assigned in the novel genus and species as Rhizomicrobium palustre gen. nov., sp. nov. The type strain of the novel species is strain A48T (= JCM 14971T = DSM 19867T).
Traditional brewing of Fukuyama pot vinegar is a process that has been continued in Fukuyama, Kagoshima, Japan, for almost 200years. The entire process proceeds from raw materials, including steamed rice, rice koji (steamed rice grown with a fungus, Aspergillus oryzae) and water, to produce vinegar in roughly capped large pots laid in the open air. No special fermentative manipulation is required, except for scattering dried rice koji (called furi-koji) on the surface of the mash to form a cap-like mat on the surface at the start of brewing. As the biochemical mechanism of the natural transition of the fermentative processes during brewing has not been fully explained, we conducted a microbiological and biochemical study on the transition. First, a distinct biochemical change was observed in the brewing of spring preparation; that is, a sharp decline in pH from 6.5 to 3.5 within the first 5days of brewing was observed due to lactic acid fermentation. Alcoholic fermentation also proceeded with a sharp increase to 4.5% ethanol within the first 5days under the acidic conditions, suggesting that saccharification and both fermentations proceed in parallel. Acidic conditions and ethanol accumulation restricted the growth of most microorganisms in the mash, and in turn provided a favorable growth condition for acetic acid bacteria which are acid resistant and “ethanol-philic.” Acetic acid was detected from day 16 and gradually increased in concentration, reaching a maximum of 7% at day 70 that was maintained thereafter. Empirically furi-koji naturally sinks into the mash after around day 40 by an unknown mechanism, allowing acetic acid bacteria to easily form pellicles on the mash surface and promoting efficient acetic acid fermentation. Dominant microbial species involved in the three fermentations were identified by denaturing gradient gel electrophoresis analysis using PCR-amplified defined-regions of small rDNA from microorganisms in the brewing mash or colony direct PCR applied to isolated microorganisms from the mash.
Three Gram-negative, pale-pink-pigmented, spherical, chemoheterotrophic bacteria were isolated from seawater and a dystrophic leaf in the Republic of Palau. Phylogenetic analyses based on 16S rRNA gene sequences indicated that the novel isolates YM31-114T, YM31-066T and YM31-067 shared approximately 97-100% sequence similarity with members of the genus Cerasicoccus of the family Puniceicoccaceae within the phylum ‘Verrucomicrobia.’ The hybridization values for DNA-DNA relatedness between the novel isolates and Cerasicoccus arenae YM26-026T were less than 70%, which is accepted as a phylogenetic definition of a species. β-Lactam antibiotic susceptibility test and amino acid analysis of cell-wall hydrolysates revealed that the novel isolates did not contain muramic acid or diaminopimelic acid in their cell walls, suggesting that these strains lack peptidoglycan. The DNA G+C contents of the three strains were 55-56 mol%; MK-7 was the major menaquinone. The presence of C14:0 and C18:1ω9c as the major cellular fatty acids supported the identification of the novel isolates as members of the genus Cerasicoccus. On the basis of polyphasic taxonomic evidence, it was concluded that these strains should be classified as representing two novel, separate species in the genus Cerasicoccus within the phylum ‘Verrucomicrobia,’ for which the names Cerasicoccus maritimus sp. nov. (type strain YM31-114T=MBIC24844T) and Cerasicoccus frondis sp. nov. (type strain YM31-066T=MBIC24796T) are proposed. Proposal for designation of the Verrucomicrobia phyl. nov., nom. rev. is also presented.
The gene encoding a glycoside hydrolase (GH) family 45 endoglucanase (Cel45A) was cloned from P. decumbens 114-2 and expressed in Pichia pastoris. To our knowledge, this is the first report of characterization of a GH family 45 protein from Penicillium species. The purified recombinant enzyme showed a higher activity on konjac glucomannan (KGM) than on sodium carboxymethyl cellulose (CMC-Na) or phosphoric acid swollen cellulose (PASC). The highest hydrolytic activity was detected at pH5.0 on KGM and pH 3.5 on CMC-Na, indicating the mode of action on the two substrates may be different for Cel45A. The optimum temperatures on the two substrates were both 60ºC and about 90% relative activities were retained at 70ºC. Products released from PASC and CMC-Na were mainly cellobiose, cellotriose and cellotetraose. The protein with higher glucomannanase activity might help the efficient degradation of lignocellulose by P. decumbens in the natural state.
A 15-kDa RNase was purified from Pleurotus djamor using ion exchange chromatography and gel filtration. Its N-terminal amino acid sequence was different from previously reported RNase sequences of mushrooms belonging to the genus of Pleurotus and other genera. The RNase exhibited maximal RNase activity at pH 4.6 and 60ºC. Its activity toward polyhomoribonucleotides was poly(U) > poly(C) > poly(A) > poly(G). It inhibited proliferation of hepatoma cells and breast cancer cells. The ranking of inhibitory potencies of metal ions on RNase activity was Fe3+> Al3+> Ca2+> Hg2+. The isolated RNase had a distinctive N-terminal sequence and optimum pH. It exhibited antiproliferative activity on tumor cells.
A novel Gram-negative, aerobic, rod-shaped and pesticide (chlorpyrifos and atrazine) tolerant strain, designated CW-D 3T, was isolated from a heavily polluted soil sample in Jiangsu Province, China. The strain could tolerate about 800μg ml-1 atrazine or chlorpyrifos when added in TYB medium. A polyphasic taxonomy including phylogenetic and phenotypic analysis was performed on the new isolate. Phylogenetic analysis and chemotaxonomic characteristics revealed that strain CW-D 3T belongs to the genus Klebsiella. Its closest phylogenetic neighbors were K. singaporensis (strain LX3T), three subspecies of K. pneumoniae (strains ATCC 13883T, ATCC 13884T and ATCC 11296T) and K. granulomatis, according to 16S rRNA and rpoB gene analysis. The 16S rRNA gene sequence similarity was about 94.8-97.6% and the rpoB gene sequence similarity was about 96.8-98.6% to its phylogenetic neighbors. Strain CW-D 3T also showed low DNA-DNA reassociation values (less than 21%) with respect to the three subspecies of K. pneumoniae and Klebsiella singaporensis. Its major fatty acids were C14:0, C16:0, c-C17:0 and c-C19:0 ω8c. The G+C content of the DNA was 58.3 mol%. Phylogenetic and phenotypic results supported the assignment of CW-D 3T as a novel species of the genus Klebsiella, for which the name Klebsiella alba sp. nov. is proposed. The type strain is CW-D 3T (=LMG 24441T=KCTC 12878T =CCTCC AB 206144T).
A parathione-degrading bacterium isolated from rice field in the Philippines, Flavobacterium sp. ATCC 27551 (Sethunathan and Yoshida, 1973, Can. J. Microbiol., 19, 873-875), was re-examined chemotaxonomically and phylogenetically. The strain contained 2-hydroxymyristic acid (2-OH 14 : 0), cis-vaccenic acid (18 : 1 ω7c), and palmitic acid (16 : 0) as major cellular fatty acids, two kinds of glycosphingolipids, and ubiquinone-10 as a sole quinone component. The G+C content of genomic DNA of the strain was 65.9 mol%. The phylogenetic analyses of the 16S rRNA gene indicated that the strain was included in the family Sphingomonadaceae, and most closely related to Sphingobium fuliginis (98.0% similarity) and Sphingobium herbicidovorans (97.3%). The strain showed similar physiological characteristics and a moderate value of DNA-DNA relatedness to S. fuliginis. These data suggested it reasonable to conclude that strain ATCC 27551 was identified as S. fuliginis.