Description of Bacillus naganoensis sp. nov.

Abstract

A new species, BaciZZus naganoensis, is proposed for an obligately aerobic, moderately acidophilic, endospore-forming bacterium that produces a thermostable, aciduric pullulanase (EC 3.2.1.41). The organism was isolated from soil by selection on solid, pullulan-containing medium at pH 4.0 and 30°C. The isolate required a medium pH of less than 6.5 for growth initiation. Fatty acid composition studies revealed that the major fatty acid of cells grown in nutrient broth supplemented with 1 % starch was 14-methylpentadecanoic acid (iso-C,,) at 45 mol%. The guanine-plus-cytosine content of the DNA of this organism was 45 f 2 mol%. A type culture has been deposited with the American Type Culture Collection, Rockville, Md., as strain ATCC 53909.Pullulanases (EC 3.2.1.41) are enzymes which specifically cleave the alpha-1,6-glycosidic linkages of pullulan, starch, glycogen, and amylopectin. Recently, these enzymes have found commercial utility in glucose-and maltose-manufacturing processes (10). Yeasts, higher plants, and microorganisms are known to be producers of pullulanase (17). Within the genus Bacillus, several species and strains have been reported to make pullulanase, including Bacillus sp. strain 202-1 (17), "Bacillus acidopullulyticus" (lo), Bacillus cereus subsp. mycoides (23), Bacillus macerans (l), Bacillus p l ymyxa (9), and Bacillus stearothermopkilus (13,22). Of these, only the "B. acidopullulyticus' ' pullulanase has properties that allow its use in industrial saccharification processes.In our search for a pullulanase that could operate under conventional glucose-manufacturing conditions (i .e., pH 4.3 and 60"C), we assumed that bacteria capable of growth at low pH values or high temperatures or both would produce aciduric or thermostable enzymes. In this paper, we describe the isolation and characterization of a mesophilic, moderately acidophilic bacterium, designated Bacillus naganoensis, that makes a pullulanase with desirable acid and temperature stability characteristics. (20), and 7.5 g of red-colored pullulan. The pH was adjusted with 0.2 N sulfuric acid to pH 4.0. The color of the plates was dark violet. Incorporation of the two colored substrates into the plate medium allowed for preliminary selection of organisms which (i) produced aamylases (enzymes that hydrolyzed the blue-colored starch but not the red-colored pullulan, leaving a red background around a colony); (ii) produced pullulanases (enzymes that hydrolyzed the red-colored pullulan but not the blue-colored starch, leaving a blue background around a colony); or (iii) produced both a-amylases and pullulanases (enzymes that hydrolyzed both the blue-colored starch and the red-colored pullulan, leaving a clear zone of hydrolysis around a colony) * Corresponding author. MATERIALS AND METHODS(12). Red-colored pullulan was prepared by dissolving 100 g of pullulan PFlO (Hayashibara Co., Ltd., Okayama, Japan) in 2 liters of distilled water. The solution temperature was increased to 50"C, and 10 g of Mikacion Brilliant Red 5BS (Mitsubishi Kasei ...

Tuberibacillus calidus gen. nov., sp. nov., isolated from a compost pile and reclassification of Bacillus naganoensis Tomimura et al. 1990 as Pullulanibacillus naganoensis gen. nov., comb. nov. and Bacillus laevolacticus Andersch et al. 1994 as Sporolactobacillus laevolacticus comb. nov.

“…S1) and its physiological and chemotaxonomic characteristics mostly agreed with those of other species of this genus (Table 1). Therefore, we also propose that Bacillus naganoensis (Tomimura et al, 1990) is reclassified as Pullulanibacillus naganoensis gen. nov., comb. nov. and Bacillus laevolacticus is reclassified as Sporolactobacillus laevolacticus comb.…”

“…B. naganoensis was isolated as a producer of pullulanase and proposed by Tomimura et al (1990). However, this species was established mainly on the basis of the results of physiological characterization and not analysed according to its phylogenetic position based on 16S rRNA gene sequence.…”

Texcoconibacillus texcoconensis gen. nov., sp. nov., alkalophilic and halotolerant bacteria isolated from soil of the former lake Texcoco (Mexico)

“…On the basis of phenotypic characteristics (Table 1), polar lipid profiles, DNA G+C contents and 16S rRNA gene T (data from this study); 2, Bacillus agaradhaerens PN-105 (Chen et al, 2009b;Lim et al, 2006;Nielsen et al, 1995); 3, Bacillus neizhouensis JSM 071004 T (Chen et al, 2009b); 4, Bacillus locisalis CG1 T (Márquez et al, 2011); 5, Pullulanibacillus naganoensis ATCC 53909 T (Hatayama et al, 2006;Tomimura et al, 1990); 6, Tuberibacillus calidus 607 T (Hatayama et al, 2006); 7, Sporolactobacillus laevolacticus NRIC 0373 (Andersch et al, 1994;Fujita et al, 2010;Hatayama et al, 2006); 8, Geomicrobium halophilum BH1 T (Echigo et al, 2010). +, Positive; 2, negative;…”

Scopulibacillus darangshiensis gen. nov., sp. nov., isolated from rock

“…Colony morphology was investigated by using a dissecting microscope (Olympus, Japan) and its color was observed after incubation for 3 days on YMG agar. Differential characteristics of strains DLS-06 T from the type strains of phylogenetic neighbors Taxa: 1, Strain DLS-06 T ; 2, Marinococcus (data from Hao et al, 1984;Li et al, 2005); 3, P. naganoensis (data from Tomimura et al, 1990;Hatayama et al, 2006, except for major cellular fatty acids); 4, Sinobaca (data from Li et al, , 2008; 5, Sporolactobacillus (data from Andersch et al, 1994;Yanagida et al, 1997;Hatayama et al, 2006;Chang et al, 2008); 6, T. calidus (data from Hatayama et al, 2006, except for major cellular fatty acids) +, Positive; -, negative; w, weakly positive; v, variable; ND, not determined or no data available; i, iso; ai, anteiso. manufacturer.…”