Int J Syst Evol Microbiol, 2002 Sep, 52(Pt 5), 1669 - 74 Paenibacillus glycanilyticus sp . nov., a novel species that degrades heteropolysaccharide produced by the cyanobacterium Nostoc commune; Dasman et al.; A novel bacterial strain, DS-1T, was isolated that degrades heteropolysaccharide produced by the cyanobacterium Nostoc commune . The isolate was identified by a combination of phenotypic characterization, cellular fatty acid analysis, DNA base composition, DNA-DNA hybridization and 165 rRNA gene sequence analysis . Phylogenetic analysis placed strain DS-1T within the Paenibacillus cluster on a phylogenetic tree and the phenotypic characteristics of this strain appear to be similar to those of Paenibacillus curdlanolyticus IFO 15724T and Paenibacillus kobensis IFO 15729T . The strain was distinguished from P . curdlanolyticus IFO 15724T and P . kobensis IFO 15729T by its ability to degrade the polysaccharide of Nostoc commune, by assimilation of rhamnose, inositol and L-fucose and by its wide range of optimal growth temperature (28-37 degrees C) . Like other Paenibacillus species, this strain contains anteiso-C15:0 as a major cellular fatty acid, and it has a DNA G+C content of 50.5 mol % . Based on these results, it is concluded that this isolate should be placed within a novel species of Paenibacillus, Paenibacillus glycanilyticus sp . nov., with the type strain DS-1T (= IFO 16618T = JCM 11221T = NRRL B-23455T). Int J Syst Evol Microbiol, 2002 Sep, 52(Pt 5), 1597 - 601 Paenibacillus koleovorans sp . nov., able to grow on the sheath of Sphaerotilus natans; Takeda M et al.; Two bacterial strains that are able to grow specifically on the sheath of a sheathed filamentous bacterium, Sphaerotilus natans, were isolated from soil samples . The sheath-degrading organisms, designated strains TB(T) and TK, are facultatively anaerobic and form endospores . The Gram reaction was negative at all stages of cultivation . The optimum growth temperature and pH were 30 degrees C and pH 7 . The DNA G+C content was 54.0-55.8 mol% . MK-7 was the predominant menaquinone and anteiso-C15:0 was the major fatty acid . Phylogenetic analysis based on the 16S rDNA sequences revealed that the isolates were closely related to Paenibacillus chondroitinus, Paenibacillus alginolyticus, Paenibacillus koreensis, Paenibacillus validus, Paenibacillus larvae subsp . larvae and P . larvae subsp . pulvifaciens . The sequences were found to contain consensus sequences characteristic of all Paenibacillus species . The isolates were able to lyse and utilize the purified sheath of S . natans as the sole carbon and energy source . Acid was not produced from common carbon sources, allowing easy distinction from other members of Paenibacillus . It is concluded that the two strains represent a novel Paenibacillus species, for which the name Paenibacillus koleovorans sp . nov . is proposed . The type strain is strain TB(T) (= JCM 11186T = IAM 14926T = KCTC 13912T). Appl Microbiol Biotechnol, 2002 Sep, 59(6), 649 - 57 Epub 2002 Jul 30. Flavin reductase coupling with two monooxygenases involved in dibenzothiophene desulfurization: purification and characterization from a non-desulfurizing bacterium, Paenibacillus polymyxa A-1; Ohshiro T et al.; The dibenzothiophene (DBT) desulfurizing bacterium metabolizes DBT to form 2-hydroxybiphenyl without breaking the carbon skeleton . Of the DBT desulfurization enzymes, DszC and DszA catalyze monooxygenation reactions, both requiring flavin reductase . We searched for non-DBT-desulfurizing microorganisms producing a flavin reductase that couples more efficiently with DszC than that produced by the DBT desulfurizing bacterium Rhodococcus erythropolis D-1, and found Paenibacillus polymyxa A-1 to be a promising strain . The enzyme was purified to complete homogeneity . K(m) values for FMN and NADH were 2.1 microM and 0.57 mM, respectively . Flavin compounds were good substrates, some nitroaromatic compounds were also active, and regarding the electron donor, the activity for NADPH was about 1.5 times that for NADH . In the coupling assay with DszC, only FMN or riboflavin acted as the electron acceptor . The coupling reactions of P . polymyxa A-1 flavin reductase with DszC and DszA proceeded more efficiently (3.5- and 5-fold, respectively) than those of R . erythropolis D-1 flavin reductase when identical enzyme activities of each flavin reductase were added to the reaction mixture . The result of the coupling reaction suggested that, in the microbial DBT desulfurization, flavin reductase from the non-DBT-desulfurizing bacterium was superior to that from the DBT-desulfurizing bacterium. Appl Environ Microbiol, 2002 Sep, 68(9), 4650 - 2 Potential for misidentification of a spore-forming Paenibacillus polymyxa isolate as an endophyte by using culture-based methods; Bent E et al.; While Paenibacillus polymyxa strain Pw-2 has been identified as an endophyte of lodgepole pine (M . Shishido, B . M . Loeb, and C . P . Chanway, Can . J . Microbiol . 41:707-713, 1995), P . polymyxa strain L6 has not, a distinction that could be explained by the differential abilities of these isolates to form spores, rather than the differential abilities to colonize the interior tissues of lodgepole pine . Chemical disinfection was used to destroy bacteria on the root exterior, but bacterial endospores are known for their ability to withstand chemical disinfection, and strain Pw-2 was found to produce 300 to 11,000 times more germinating endospores than strain L6 under the experimental conditions used by Shishido et al . (Can . J . Microbiol . 41:707-713, 1995) . Attempts to identify strain Pw-2 within lodgepole pine root tissues by using confocal microscopy techniques failed . We discuss the possibility that spore-forming bacteria can be mistakenly identified as endophytes when culture-based methods alone are used. Appl Microbiol Biotechnol, 2002 Jul, 59(2-3), 325 - 8 Epub 2002 Apr 12. Biodesulfurization of benzothiophene and dibenzothiophene by a newly isolated Rhodococcus strain; Tanaka Y et al.; Rhodococcus sp . KT462, which can grow on either benzothiophene (BT) or dibenzothiophene (DBT) as the sole source of sulfur, was newly isolated and characterized . GC and GC-MS analyses revealed that strain KT462 has the same BT desulfurization pathway as that reported for Paenibacillus sp . A11-2 and Sinorhizobium sp . KT55 . The desulfurized product of DBT produced by this strain, as well as other DBT-desulfurizing bacteria such as R . erythropolis KA2-5-1 and R . erythropolis IGTS8, was 2-hydroxybiphenyl . A resting cells study indicated that this strain was also able to degrade various alkyl derivatives of BT and DBT. Lett Appl Microbiol, 2002, 35(1), 52 - 6 Evaluation of the diversity of Paenibacillus polymyxa strains by using the DNA of bacteriophage IPy1 as a probe in hybridization experiments; Santos SC et al.; AIMS: To evaluate the genetic diversity within the species Paenibacillus polymyxa . METHODS: Southern hybridization was performed on 102 strains of P . polymyxa using DNA from the phage IPy1 as a probe . Results: All 102 strains hybridized to phage IPy1 DNA . Data from different hybridization patterns obtained were used to construct a dendrogram in which 53 genotypic groups were split into two main clusters . One cluster contained strains from the rhizospheres of sorghum and maize planted in Cerrado soil, Brazil, and the majority of strains received from two culture collections . The other cluster contained strains isolated from different Brazilian soils and rhizospheres and strains deposited in a third culture collection . SIGNIFICANCE AND IMPACT OF THE STUDY: The approach used in this study appears to be a new and a very useful tool to study the diversity within this species. Curr Microbiol, 2002 Jul, 45(1), 5 - 12 Molecular cloning and expression in Escherichia coli of an exo-levanase gene from the endophytic bacterium Gluconacetobacter diazotrophicus SRT4; Menendez C et al.; Gluconacetobacter diazotrophicus produces levan from sucrose by a secreted levansucrase (LsdA) . A levanase-encoding gene ( lsdB), starting 51 bp downstream of the lsdA gene, was cloned from strain SRT4 . The lsdB gene (1605 bp) encodes a protein (calculated molecular mass 58.4 kDa) containing a putative 36-amino-acid signal peptide at the N-terminus . The deduced amino acid sequence shares 34%, 33%, 32%, and 29% identities with levanases from Actinomyces naeslundii, Bacillus subtilis, Paenibacillus polymyxa, and Bacteroides fragilis, respectively . The lsdB expression in Escherichia coli under the control of the T7 RNA polymerase promoter resulted in an active enzyme which hydrolyzed levan, inulin, 1-kestose, raffinose, and sucrose, but not melezitose . Levanase activity was maximal at pH 6.0 and 30 degrees C, and it was not inhibited by the metal ion chelator EDTA or the denaturing agents dithiothreitol and beta-mercaptoethanol . The recombinant LsdB showed a fourfold higher rate of hydrolysis on levan compared to inulin, and the reaction on both substrates resulted in the successive liberation of the terminal fructosyl residues without formation of intermediate oligofructans, indicating a non-specific exo-levanase activity. Biosci Biotechnol Biochem, 2002 Feb, 66(2), 458 - 63 Commensal relationship between a sheath-forming bacterium, Sphaerotilus natans, and a sheath-degrading bacterium, Paenibacillus sp; Takeda M et al.; Paenibacillus sp . strain TB is capable of degrading the sheath prepared from a sheathed bacterium, Sphaerotilus natans . S . natans was able to grow alone on casamino acids but strain TB was not . Cocultivation of strain TB and S . natans was examined in a medium supplemented with casamino acids as a growth substrate . The growth of strain TB was observed when the sheath was supplied to the medium or in cocultivation with S . natans . The phospholipid amount reached a maximum after 24 h of cocultivation and subsequently kept almost the same level for 96 h . The sheath amount also reached a maximum after 24 h and then gradually declined . The cell concentration of strain TB increased throughout the cocultivation . By competitive PCR targeted for amplification of a part of 16S rDNA, the abundance ratio (S . natans/strain TB) of 6.7 was obtained at 72 h . Almost no growth of strain TB was detected in a coculture with a sheath-less mutant of S . natans . The evidence allows the conclusion that strain TB grew by utilizing the intact sheath in coculture with S . natans. Phys Rev E Stat Phys Plasmas Fluids Relat Interdiscip Topics, 1999 Jun, 59(6), 7025 - 35 Lubricating bacteria model for branching growth of bacterial colonies; Kozlovsky Y et al.; Various bacterial strains (e.g., strains belonging to the genera Bacillus, Paenibacillus, Serratia, and Salmonella) exhibit colonial branching patterns during growth on poor semisolid substrates . These patterns reflect the bacterial cooperative self-organization . A central part of the cooperation is the collective formation of a lubricant on top of the agar which enables the bacteria to swim . Hence it provides the colony means to advance towards the food . One method of modeling the colonial development is via coupled reaction-diffusion equations which describe the time evolution of the bacterial density and the concentrations of the relevant chemical fields . This idea has been pursued by a number of groups . Here we present an additional model which specifically includes an evolution equation for the lubricant excreted by the bacteria . We show that when the diffusion of the fluid is governed by a nonlinear diffusion coefficient, branching patterns evolve . We study the effect of the rates of emission and decomposition of the lubricant fluid on the observed patterns . The results are compared with experimental observations . We also include fields of chemotactic agents and food chemotaxis and conclude that these features are needed in order to explain the observations. Can J Microbiol, 2002 Feb, 48(2), 159 - 69 Paenibacillus polymyxa produces fusaricidin-type antifungal antibiotics active against Leptosphaeria maculans, the causative agent of blackleg disease of canola; Beatty PH et al.; A bacterial isolate capable of inhibiting the growth of Leptosphaeria maculans (Desmaz.) Ces . & De Not., the causative agent of blackleg disease of canola (Brassica napus L . and Brassica rapa L.), was identified as a potential biological control agent . This environmental isolate was determined to be Paenibacillus polymyxa based on its (i) biochemical and growth characteristics and (ii) 16S rRNA sequence similarity, and was given the strain designation PKB1 . Antifungal peptides were produced by P . polymyxa PKB1 around the onset of sporulation, with optimal production on potato dextrose broth . The antifungal peptides were extracted from P . polymyxa PKB1 cells and (or) spores using methanol and were purified using size exclusion and reverse-phase chromatography . Characterization of the antifungal peptides was done using amino acid compositional analysis, Edman degradation sequencing from partially hydrolyzed material, and a variety of mass spectrometric methods . The purified antifungal material was found to be a mixture of related peptides of molecular masses 883, 897, 948, and 961 Da, with the most likely structure of the 897 Da component determined to be a cyclic depsipeptide with an unusual 15-guanidino-3-hydroxypentadecanoic acid moiety bound to a free amino group . These compounds are therefore members of the fusaricidin group of cyclic depsipeptides. Int J Syst Evol Microbiol, 2002 Mar, 52(Pt 2), 607 - 16 Paenibacillus graminis sp . nov . and Paenibacillus odorifer sp . nov., isolated from plant roots, soil and food; Berge O et al.; Sixteen gram-positive endospore-forming bacteria previously isolated from soil, plant rhizospheres, plant roots and pasteurized pureed vegetables were studied to determine their taxonomic positions . The isolates were formerly identified as Bacillus circulans based on their biochemical characters using API galleries . Two of these strains, RSA19T and TOD45T, were recently assigned to the genus Paenibacillus based on phylogenetic analysis of their 16S rRNA (rrs) gene sequence . In the present work, the sixteen isolates were assigned to two genomospecies using DNA-DNA hybridization, in agreement with rrs gene sequence analysis . These genomospecies can also be differentiated on the basis of their cultural and biochemical characters into two novel species, for which the names Paenibacillus graminis sp . nov . (type strain RSA19T = ATCC BAA-95T = LMG 19080T) and Paenibacillus odorifer sp . nov . (type strain TOD45T = ATCC BAA-93T = LMG 19079T) are proposed. Int J Syst Evol Microbiol, 2002 Mar, 52(Pt 2), 415 - 21 Paenibacillus chinjuensis sp . nov., a novel exopolysaccharide-producing bacterium; Yoon JH et al.; A novel exopolysaccharide-producing bacterium (WN9T) was isolated from Chinju, Korea, and was identified as a member of the genus Paenibacillus on the basis of phenotypic characteristics and phylogenetic inference based on 16S rDNA sequence . This organism is a facultatively anaerobic, endospore-forming rod . The diamino acid found in the peptidoglycan is meso-diaminopimelic acid . The predominant menaquinone is MK-7 . The major cellular fatty acid is anteiso-C15:0 . The G+C content is 53 mol% . The phylogenetic tree shows that strain WN9T falls within the radiation of a cluster comprising the Paenibacillus species . The levels of 16S rDNA similarity between strain WN9T and the type strains of validly described Paenibacillus species are 92.1-95.8% . Strain WN9T is clearly distinguishable from some phylogenetically related Paenibacillus species on the basis of DNA-DNA relatedness data and phenotypic characters . Therefore, on the basis of these data, a novel species of the genus Paenibacillus, Paenibacillus chinjuensis sp . nov., is proposed . The type strain is strain WN9T (= KCTC 8951PT = JCM 10939T). J Appl Microbiol, 2002, 92(3), 475 - 81 Cytometric monitoring of growth, sporogenesis and spore cell sorting in Paenibacillus polymyxa (formerly Bacillus polymyxa); Comas-Riu J et al.; AIMS: Formation of bacterial endospores is a basic process in Gram-positive bacteria and has implications for health, industry and the environment . Flow cytometry offers a practical alternative for the rapid detection, enumeration and characterization of bacterial endospores . METHODS AND RESULTS: Paenibacillus polymyxa was chosen for this study because its spores cause sporangium deformation and have thick walls with a star-shaped section . Sporulating populations were analysed with a particle analyser and a flow cytometer after labelling with propidium iodide and Syto-13 . Flow cytometric detection of single spores was confirmed by optical and scanning electron microscopy after cell sorting . Four cell sub-populations were cytometrically detected in P . polymyxa cultures grown in liquid sporulation medium . Two sub-populations consisted of vegetative cells differing in both morphology and viability; the other two sub-populations consisted of spores differing in their viability . CONCLUSIONS: This work has shown that flow cytometry is a simple and fast method (less than 15 minutes for sample preparation and analysis) for the study of the sporulation in P . polymyxa . The use of this technique allowed both detection and quantification of sporulation inside a culture, and distinguished cells that differed in viability despite being morphologically identical under microscopic observation . SIGNIFICANCE AND IMPACT OF THE STUDY: Flow cytometry has been proved to be a valuable tool for the analysis of sporulation in P . polymyxa cultures, with the unique capacity of distinguishing between endospores and vegetative cells, and between live and dead cells, in the same analysis . An important percentage of non-viable endospores has been found in aged cultures using this method. J Biol Chem, 2002 Apr 26, 277(17), 14695 - 702 Epub 2002 Feb 19. Biochemical and genetic properties of Paenibacillus glycosyl hydrolase having chitosanase activity and discoidin domain; Kimoto H et al.; Cells of "Paenibacillus fukuinensis" D2 produced chitosanase into surrounding medium, in the presence of colloidal chitosan or glucosamine . The gene of this enzyme was cloned, sequenced, and subjected to site-directed mutation and deletion analyses . The nucleotide sequence indicated that the chitosanase was composed of 797 amino acids and its molecular weight was 85,610 . Unlike conventional family 46 chitosanases, the enzyme has family 8 glycosyl hydrolase catalytic domain, at the amino-terminal side, and discoidin domain at the carboxyl-terminal region . Expression of the cloned gene in Escherichia coli revealed beta-1,4-glucanase function, besides chitosanase activity . Analyses by zymography and immunoblotting suggested that the active enzyme was, after removal of signal peptide, produced from inactive 81-kDa form by proteolysis at the carboxyl-terminal region . Replacements of Glu(115) and Asp(176), highly conserved residues in the family 8 glycosylase region, with Gln and Asn caused simultaneous loss of chitosanase and glucanase activities, suggesting that these residues formed part of the catalytic site . Truncation experiments demonstrated indispensability of an amino-terminal region spanning 425 residues adjacent to the signal peptide. Int J Syst Evol Microbiol, 2002 Jan, 52(Pt 1), 131 - 9 PAH-degradation by Paenibacillus spp . and description of Paenibacillus naphthalenovorans sp . nov., a naphthalene-degrading bacterium from the rhizosphere of salt marsh plants; Daane LL et al.; Bacteria belonging to the genus Paenibacillus were isolated by enrichment from petroleum-hydrocarbon-contaminated sediment and salt marsh rhizosphere using either naphthalene or phenanthrene as the sole carbon source, and were characterized using phenotypic, morphological and molecular techniques . The isolates were grouped by their colony morphologies and polyaromatic hydrocarbon-degradation patterns . Phenanthrene-degrading isolates produced mottled colonies on solid media and were identified as P . validus by fatty acid methyl ester and 16S rRNA gene sequence analyses . In contrast, the naphthalene-degrading isolates with mucoid colony morphology were distantly related to Paenibacillus validus, according to fatty acid methyl ester and 16S rRNA gene sequence analyses . The predominant fatty acids of the mucoid isolates were 15:0 anteiso, 16:1omega11c, 16:0 and 17:0 anteiso, constituting, on average, 50.5, 12.0, 11.2 and 6.5% of the total, respectively . The G+C contents of their DNA ranged from 47 to 52 mol% . The 16S rDNA sequence analysis revealed the highest (< or = 94%) similarity to P . validus . In addition, phylogenetic analyses based on 16S rDNA sequences showed that the mucoid isolates formed a distinct cluster within Paenibacillus . DNA-DNA hybridization experiments showed only a 6% DNA similarity between the type strain of P . validus and mucoid strain PR-N1 . On the basis of the morphological, phenotypic and molecular data, the naphthalene-degrading isolates merit classification as a new Paenibacillus species, for which the name Paenibacillus naphthalenovorans sp . nov . is proposed, with PR-N1T (= ATCC BAA-206T = DSM 14203T) as the type strain. Appl Environ Microbiol, 2002 Feb, 68(2), 772 - 83 Osmotically regulated synthesis of the compatible solute ectoine in Bacillus pasteurii and related Bacillus spp; Kuhlmann AU et al.; By using natural-abundance (13)C-nuclear magnetic resonance spectroscopy and high-performance liquid chromatography (HPLC) analysis we have investigated the types of compatible solutes that are synthesized de novo in a variety of Bacillus species under high-osmolality growth conditions . Five different patterns of compatible solute production were found among the 13 Bacillus species we studied . Bacillus subtilis, B . licheniformis, and B . megaterium produced proline; B . cereus, B . circulans, B . thuringiensis, Paenibacillus polymyxa, and Aneurinibacillus aneurinilyticus synthesized glutamate; B . alcalophilus, B . psychrophilus, and B . pasteurii synthesized ectoine; and Salibacillus (formerly Bacillus) salexigens produced both ectoine and hydroxyectoine, whereas Virgibacillus (formerly Bacillus) pantothenticus synthesized both ectoine and proline . Hence, the ability to produce the tetrahydropyrimidine ectoine under hyperosmotic growth conditions is widespread within the genus Bacillus and closely related taxa . To study ectoine biosynthesis within the group of Bacillus species in greater detail, we focused on B . pasteurii . We cloned and sequenced its ectoine biosynthetic genes (ectABC) . The ectABC genes encode the diaminobutyric acid acetyltransferase (EctA), the diaminobutyric acid aminotransferase (EctB), and the ectoine synthase (EctC) . Together these proteins constitute the ectoine biosynthetic pathway, and their heterologous expression in B . subtilis led to the production of ectoine . Northern blot analysis demonstrated that the ectABC genes are genetically organized as an operon whose expression is strongly enhanced when the osmolality of the growth medium is raised . Primer extension analysis allowed us to pinpoint the osmoregulated promoter of the B . pasteurii ectABC gene cluster . HPLC analysis of osmotically challenged B . pasteurii cells revealed that ectoine production within this bacterium is finely tuned and closely correlated with the osmolality of the growth medium . These observations together with the osmotic control of ectABC transcription suggest that the de novo synthesis of ectoine is an important facet in the cellular adaptation of B . pasteurii to high-osmolarity surroundings. Syst Appl Microbiol, 2001 Nov, 24(3), 417 - 22 16S rDNA sequence analysis of bacterial isolates from biodeteriorated mural paintings in the Servilia tomb (Necropolis of carmona, Seville, Spain); Heyrman J et al.; Bacteria were isolated from damaged mural paintings of the Servilia tomb (necropolis of Carmona, Seville, Spain) . Selected strains, representative for different clusters of isolates with similar fatty acid profiles, were analysed by 16S rDNA sequence analysis . Bacillus is the dominant genus among the isolates: members of the rRNA species complexes of B . megaterium, B . pumilus and B . firmus were found as well as several other Bacillus species . One group of halotolerant isolates falls in the Bacillus sensu lato group, with closest relatedness to the genera Salibacillus and Virgibacillus . Other genera found are Artbrobacter, Micrococcus, Streptomyces, Sphingomonas, Paenibacillus, and a genus closely related to Paracraurococcus . Many isolates showed low 16S rDNA sequence similarities with the closest related database entries, a strong indication for the presence of several new species among the isolates. Biotechnol Prog, 2001 Nov-Dec, 17(6), 1020 - 5 Biphasic inactivation kinetics of Escherichia coli in liquid whole egg by high hydrostatic pressure treatments; Lee DU et al.; Kinetic studies on the isothermal high hydrostatic pressure (HHP) inactivation of Escherichia coli in liquid whole egg (LWE) were performed at 5 and 25 degrees C in the pressure range of 250-400 MPa . The characteristic tailing inactivation curves were described by a first-order biphasic model . As compared to a previous rheological study, it is suggested that the phase change of LWE during pressure treatment affects the inactivation rate of E . coli . Within the processing criteria where the rheological properties of LWE were still comparable to those of fresh LWE, HHP treatments at 5 degrees C induced more E . coli inactivations than those at 25 degrees C . From the results of approximately 3 log reductions of E . coli and over 5 log reductions of Pseudomonas and Paenibacillus, HHP treatment of LWE at 5 degrees C is regarded to be as effective as conventional thermal pasteurization . However, no post-process contamination and the consistency of temperature during preparation, HHP treatment, and storage provide clear processing advantages. Can J Microbiol, 2001 Sep, 47(9), 793 - 800 Alterations in plant growth and in root hormone levels of lodgepole pines inoculated with rhizobacteria; Bent E et al.; The presence of other soil microorganisms might influence the ability of rhizobacterial inoculants to promote plant growth either by reducing contact between the inoculant and the plant root or by interfering with the mechanism(s) involved in rhizobacterially mediated growth promotion . We conducted the following experiments to determine whether reductions in the extent of growth promotion of lodgepole pine mediated by Paenibacillus polymyxa occur in the presence of a forest soil isolate (Pseudomonas fluorescens M20) and whether changes in plant growth promotion mediated by P . polymyxa (i) are related to changes in P . polymyxa density in the rhizosphere or (ii) result from alterations in root hormone levels . The extent of plant growth, P . polymyxa rhizosphere density, and root hormone concentrations were determined for lodgepole pine treated with (i) a single growth-promoting rhizobacterial strain (P . polymyxa L6 or Pw-2) or (ii) a combination of bacteria: strain L6 + strain M20 or strain Pw-2 + strain M20 . There was no difference in the growth of pines inoculated with strain L6 and those inoculated with strain L6 + strain M20 . However, seedlings inoculated with strain Pw-2 had more lateral roots and greater root mass at 12 weeks after inoculation than plants inoculated with strain Pw-2 + strain M20 . The extent of growth promotion mediated by P . polymyxa L6 and Pw-2 in each treatment was not correlated to the average population density of each strain in the rhizosphere . Bacterial species-specific effects were observed in root hormone levels: indole-3-acetic acid concentration was elevated in roots inoculated with P . polymyxa L6 or Pw-2, while dihydrozeatin riboside concentration was elevated in roots inoculated with P . fluorescens M20. Int J Syst Evol Microbiol, 2001 Sep, 51(Pt 5), 1681 - 5 Paenibacillus azoreducens sp . nov., a synthetic azo dye decolorizing bacterium from industrial wastewater; Meehan C et al.; An azo-dye-reducing, endospore-forming bacterium isolated from textile industry wastewater has been taxonomically studied . Particularly interesting was the ability of this organism to decolorize the azo dye Remazol Black B by 98% within 24 h . Levels of 16S rRNA similarity between the isolate and Paenibacillus species ranged from 92.1 to 95.0% . The DNA G+C content was 46.8 mol % and anteiso-branched C15:0 was the major fatty acid . Based upon the phenotypic properties and the phylogenetic inference, it is proposed that the bacterium should be designated Paenibacillus azoreducens sp . nov . The type strain of Paenibacillus azoreducens is CM1T (= DSM 13822T = NCIMB 13761T). J Appl Microbiol, 2001 Aug, 91(2), 212 - 6 Disinfection of wooden structures contaminated with Paenibacillus larvae subsp . larvae spores; Dobbelaere W et al.; AIMS: The aim of the study is to examine the disinfection of wood contaminated with Paenibacillus larvae subsp . larvae spores, in order to find a practical decontamination method for hive materials . METHODS AND RESULTS: The number of viable spores recovered after the treatment, on the surface by swabbing, and in the deeper parts of the wood by scraping, was used to test the efficiency of the disinfection . Our results indicate that chemical disinfection is only complete when high concentrations (> 50%) of the disinfectant are used . Heat treatment in general was found to be very effective . The scorching of wood was not satisfactory as it only killed spores at the surface . CONCLUSION: Complete disinfection is only possible with some heat treatments or by using high concentrations of chemical disinfectants . SIGNIFICANCE AND IMPACT OF THE STUDY: This study puts forward some methods that can provide complete decontamination, which is necessary for an effective control of American foulbrood disease. Appl Environ Microbiol, 2001 Aug, 67(8), 3557 - 63 Effect of primers hybridizing to different evolutionarily conserved regions of the small-subunit rRNA gene in PCR-based microbial community analyses and genetic profiling; Schmalenberger A et al.; Genetic profiling techniques of microbial communities based on PCR-amplified signature genes, such as denaturing gradient gel electrophoresis or single-strand-conformation polymorphism (SSCP) analysis, are normally done with PCR products of less than 500-bp . The most common target for diversity analysis, the small-subunit rRNA genes, however, are larger, and thus, only partial sequences can be analyzed . Here, we compared the results obtained by PCR targeting different variable (V) regions (V2 and V3, V4 and V5, and V6 to V8) of the bacterial 16S rRNA gene with primers hybridizing to evolutionarily conserved flanking regions . SSCP analysis of single-stranded PCR products generated from 13 different bacterial species showed fewer bands with products containing V4-V5 (average, 1.7 bands per organism) than with V2-V3 (2.2 bands) and V6-V8 (2.3 bands) . We found that the additional bands (>1 per organism) were caused by intraspecies operon heterogeneities or by more than one conformation of the same sequence . Community profiles, generated by PCR-SSCP from bacterial-cell consortia extracted from rhizospheres of field-grown maize (Zea mays), were analyzed by cloning and sequencing of the dominant bands . A total of 48 sequences could be attributed to 34 different strains from 10 taxonomical groups . Independent of the primer pairs, we found proteobacteria (alpha, beta, and gamma subgroups) and members of the genus Paenibacillus (low G+C gram-positive) to be the dominant organisms . Other groups, however, were only detected with single primer pairs . This study gives an example of how much the selection of different variable regions combined with different specificities of the flanking "universal" primers can affect a PCR-based microbial community analysis. Lett Appl Microbiol, 2001 Aug, 33(2), 117 - 21 Characterization of pectin lyase produced by an endophytic strain isolated from coffee cherries; Sakiyama CC et al.; AIMS: The effect of endophytic bacterial activity on the quality of coffee beverage was studied . METHODS AND RESULTS: A survey of the micro-organisms in coffee cherries was performed before harvesting, and their growth on the main nutrients available in coffee cherries was determined in vitro . CONCLUSION: Many endophytic bacteria were isolated from surface-sterilized coffee cherries . One of the pectinolytic strains was physiologically and phenotypically characterized, and was tentatively identified by partial 16S rDNA sequencing as Paenibacillus amylolyticus . This endophytic strain produced an extracellular pectinase with maximal activity at 40 degrees C and pH 7.9, and was thermostable up to 45 degrees C . EDTA and metal ions had little effect on pectin lyase activity . Km and Vmax values were 4.6 mg ml(-1) and 94.0 10(-8) mol min(-1) ml(-1), respectively . SIGNIFICANCE AND IMPACT OF THE STUDY: Pectin lyases have been found in fungi but rarely in bacteria, and this isolate is a promising tool for regulation studies of these enzymes. J Invertebr Pathol, 2001 May, 77(4), 231 - 6 Adult honeybee's resistance against Paenibacillus larvae larvae, the causative agent of the American foulbrood; Riessberger-Galle U et al.; American foulbrood is a widespread disease of honeybee larvae caused by the spore-forming bacterium Paenibacillus larvae subsp . larvae . Spores represent the infectious stage; when ingested by a larva they germinate in the midgut . The rod-shaped vegetative forms penetrate the larva's intestinal tissue and start multiplying rapidly, which finally kills the larva . Spores fed to adult honeybees, however, do not harm the bees . We investigated this phenomenon . Specifically, we studied the influence of the adult honeybee midgut on the vegetative growth and on the germination of spores of P . larvae larvae . We focused on two groups of adult workers that are likely to have large numbers of spores in their gastrointestinal tracts in infected colonies: middle-aged bees, which are known to remove or cannibalize dead larvae and clean brood cells, and winterbees, which do not have frequent chances to defecate . We found that midgut extract from winterbees and worker-aged bees of different colonies almost completely inhibited the growth of the vegetative stage of P . larvae larvae and suppressed the germination of spores . The inhibiting substance or substances from the adult midgut are very temperature stable: they still show about 60% of their growth-inhibiting capacity against this bacterium after 15 min at 125 degrees C . We established a method to test growth-inhibiting factors against P . larvae larvae in vitro . Clin Diagn Lab Immunol, 2001 Jul, 8(4), 706 - 10 Biological response modifier activity of an exopolysaccharide from Paenibacillus jamilae CP-7; Ruiz-Bravo A et al.; An extracellular polysaccharide was purified from culture supernatants of Paenibacillus jamilae CP-7, a gram-positive bacillus that was isolated from compost prepared with olive mill wastewaters . The extracellular polysaccharide was produced under aerobic conditions in a medium containing olive mill wastewaters (80% {vol/vol}) . This exopolymer had a low level of acute toxicity when it is administered to BALB/c mice by the intraperitoneal route . Interesting immunomodulatory effects were detected when mice were given 10 mg of exopolysaccharide per kg of body weight; the proliferative responses of splenocytes to B-cell and T-cell mitogens were suppressed, the in vitro levels of production of gamma interferon and granulocyte-macrophage colony-stimulating factor by unstimulated and lipopolysaccharide-stimulated splenocytes were enhanced, and the levels of resistance to the intracellular pathogen Listeria monocytogenes was increased in mice . Also, the exopolysaccharide was able to induce lymphocyte proliferation in vitro . We conclude that P . jamilae produces an exopolysaccharide with interesting immunomodulatory properties. Glycoconj J, 2000 Oct, 17(10), 681 - 90 A pyrophosphate bridge links the pyruvate-containing secondary cell wall polymer of Paenibacillus alvei CCM 2051 to muramic acid; Schaffer C et al.; The peptidoglycan, the secondary cell wall polymer (SCWP), and the surface layer (S-layer) glycoprotein are the major glycosylated cell wall components of Paenibacillus alvei CCM 2051 . In this report, the complete structure of the SCWP, its linkage to the peptidoglycan layer, and its physicochemical properties have been investigated . From the combined evidence of chemical and structural analyses together with one- and two-dimensional nuclear magnetic resonance spectroscopy, the following structure of the SCWP-peptidoglycan complex is proposed: {(Pyr4,6)-beta-D-ManpNAc-(1-->4)-beta-D-GlcpNAc-(1-->3)}n-11-(Pyr4,6)-beta-D-ManpNAc-(1-->4)-alpha-D-GlcpNAc-(1-->O)-PO2-O-PO2-(O-->6)-MurNAc- Each disaccharide unit is substituted by 4,6-linked pyruvic acid residues . Under mild acidic conditions, up to 50% of them are lost, leaving non-substituted ManNAc residues . The anionic glycan chains constituting the SCWP are randomly linked via pyrophosphate groups to C-6 of muramic acid residues of the peptidoglycan layer . 31P NMR reveals two signals that, as a consequence of micelle formation, experience different line broadening . Therefore, their integral ratio deviates significantly from 1:1 . By treatment with ethylenediaminetetraacetic acid, sodium dodecyl sulfate, and sonication immediately prior to NMR measurement, this ratio approaches unity . The reversibility of this behavior corroborates the presence of a pyrophosphate linker in this SCWP-peptidoglycan complex . In addition to the determination of the structure and linkage of the SCWP, a possible scenario for its biological function is discussed. Appl Microbiol Biotechnol, 2001 May, 55(5), 632 - 5 Screening for microorganisms that produce only endo-inulinase; Gern RM et al.; Sixteen fungal strains reported in the literature as endo-inulinase producers and three bacterial strains, isolated from the dahlia rhizosphere, were analysed for endo-inulinase production . From four isolated strains (one fungus and three bacteria) the results were evaluated in terms of substrate consumption, cell growth and production of endo-inulinases . All three bacterial strains were sole endo-inulinase producers and, among these, strain Paenibacillus sp . CDB 003 was the most suitable for endo-inulinase production, as this enzyme produced inulobiose as the principal substrate as well as inulo-oligosaccharides with polymerisation degrees of 3-5. Appl Environ Microbiol, 2001 Jun, 67(6), 2683 - 91 Isolation and characterization of polycyclic aromatic hydrocarbon-degrading bacteria associated with the rhizosphere of salt marsh plants; Daane LL et al.; Polycyclic aromatic hydrocarbon (PAH)-degrading bacteria were isolated from contaminated estuarine sediment and salt marsh rhizosphere by enrichment using either naphthalene, phenanthrene, or biphenyl as the sole source of carbon and energy . Pasteurization of samples prior to enrichment resulted in isolation of gram-positive, spore-forming bacteria . The isolates were characterized using a variety of phenotypic, morphologic, and molecular properties . Identification of the isolates based on their fatty acid profiles and partial 16S rRNA gene sequences assigned them to three main bacterial groups: gram-negative pseudomonads; gram-positive, non-spore-forming nocardioforms; and the gram-positive, spore-forming group, Paenibacillus . Genomic digest patterns of all isolates were used to determine unique isolates, and representatives from each bacterial group were chosen for further investigation . Southern hybridization was performed using genes for PAH degradation from Pseudomonas putida NCIB 9816-4, Comamonas testosteroni GZ42, Sphingomonas yanoikuyae B1, and Mycobacterium sp . strain PY01 . None of the isolates from the three groups showed homology to the B1 genes, only two nocardioform isolates showed homology to the PY01 genes, and only members of the pseudomonad group showed homology to the NCIB 9816-4 or GZ42 probes . The Paenibacillus isolates showed no homology to any of the tested gene probes, indicating the possibility of novel genes for PAH degradation . Pure culture substrate utilization experiments using several selected isolates from each of the three groups showed that the phenanthrene-enriched isolates are able to utilize a greater number of PAHs than are the naphthalene-enriched isolates . Inoculating two of the gram-positive isolates to a marine sediment slurry spiked with a mixture of PAHs (naphthalene, fluorene, phenanthrene, and pyrene) and biphenyl resulted in rapid transformation of pyrene, in addition to the two- and three-ringed PAHs and biphenyl . This study indicates that the rhizosphere of salt marsh plants contains a diverse population of PAH-degrading bacteria, and the use of plant-associated microorganisms has the potential for bioremediation of contaminated sediments. Int J Syst Evol Microbiol, 2001 Mar, 51(Pt 2), 535 - 45 Paenibacillus borealis sp . nov., a nitrogen-fixing species isolated from spruce forest humus in Finland; Elo S et al.; Seven spore-forming, nitrogen-fixing bacterial isolates from spruce forest humus in Finland were studied using the polyphasic approach . PCR amplification of 16S rRNA gene fragment with specific primers showed that the isolates were members of Paenibacillus . Levels of 16S rDNA similarity between the isolates were 97.3-100.0% and those between the isolates and other Paenibacillus species were 90.3-96.5% . The highest similarities were observed with Paenibacillus azotofixans and Paenibacillus durus . Ribotyping with EcoRI and PvuII restriction showed a high diversity in the Paenibacillus species and distinguished the isolates from these closely related species . The main whole-cell fatty acids were anteiso-C15:0 (33-48%), straight-chain C14:0 (7-21%) and C16:0 (9-20%), and iso-C15:0 (6-15%) . Electron microscopy revealed a unique striped morphology of the spore surfaces . Based on phylogenetic inference and phenotypic and chemotaxonomic characteristics, these isolates are proposed as a new species, Paenibacillus borealis sp . nov., the type strain of which is KK19T (= DSM 13188T = CCUG 43137T). Am J Infect Control, 2001 Apr, 29(2), 126 - 9 Paenibacillus macerans pseudobacteremia resulting from contaminated blood culture bottles in a neonatal intensive care unit; Noskin GA et al.; Paenibacillus species are gram-positive, rod-shaped, spore-forming aerobes that are abundant in nature and closely related to Bacillus . Between June 24 and June 30, 1999, 8 neonates in our neonatal intensive care unit had positive blood cultures for Paenibacillus macerans . This cluster of positive blood cultures with an unusual pathogen suggested a pseudoepidemic . Investigation revealed that the most likely etiology of the pseudobacteremia was environmental contamination of the rubber stoppers in blood culture bottles . This was confirmed by environmental sampling and simulated inoculation studies . This pseudobacteremia outbreak highlights the importance of adhering to well-established methods for blood culture collection and ongoing infection control surveillance. Appl Microbiol Biotechnol, 2001 Jan, 55(1), 61 - 8 Molecular cloning and characterization of a multidomain endoglucanase from Paenibacillus sp BP-23: evaluation of its performance in pulp refining; Pastor FI et al.; The gene celB encoding an endoglucanase from Paenibacillus sp . BP-23 was cloned and expressed in Escherichia coli . The nucleotide sequence of a 4161 bp DNA fragment containing the celB gene was determined, revealing an open reading frame of 2991 nucleotides that encodes a protein of 106,927 Da . Comparison of the deduced amino acid sequence of endoglucanase B with known beta-glycanase sequences showed that the encoded enzyme is a modular protein and exhibits high homology to enzymes belonging to family 9 cellulases . The celB gene product synthesized in E . coli showed high activity on carboxymethyl cellulose and lichenan while low activity was found on Avicel . Activity was enhanced in the presence of 10 mM Ca2+ and showed its maximum at 53 degrees C and pH 5.5 . The effect of the cloned enzyme in modifying the physical properties of pulp and paper from Eucalyptus was tested (CelB treatment) . An increase in mechanical strength of paper and a decrease in pulp dewatering properties were found, indicating that CelB treatment can be considered as a biorefining . Treatment with CelB gave rise to an improvement in paper strength similar to that obtained with 1,000 revolutions increase in mechanical refining . Comparison with the performances of recently developed endoglucanase A from the same strain and with a commercial cellulase showed that CelB produced the highest refining effect. Environ Microbiol, 2000 Jun, 2(3), 333 - 42 Disruption of the Paenibacillus polymyxa levansucrase gene impairs its ability to aggregate soil in the wheat rhizosphere; Bezzate S et al.; Inoculation of wheat roots with Paenibacillus (formerly Bacillus) polymyxa CF43 increases the mass of root-adhering soil . We tested the role of levan, a fructosyl polymer produced by strain CF43, in the aggregation of soil adhering to wheat roots . The P . polymyxa gene homologous to the Bacillus subtilis sacB gene encoding levansucrase was cloned and sequenced . The corresponding gene product synthesises high molecular weight levan . A P . polymyxa mutant strain, SB03, whose sacB gene is disrupted, was constructed using heterogramic conjugation . Effects of wheat inoculation with the wild type and the mutant strain were compared using two different cultivated silt loam soils in four independent pot experiments . Roots of wheat plantlets inoculated with CF43 or SB03 were colonized after 7-14 days at the same level, and root and shoot masses were not significantly different from those of the non-inoculated control plants . The ratio of root-adhering soil dry mass to root tissue dry mass was significantly higher for plants inoculated with strain CF43 than for those inoculated with mutant strain SB03: +30% in Orgeval soil and +100% in Dieulouard soil . Thus the levan produced by P . polymyxa is implicated in the aggregation of root-adhering soil on wheat. J Appl Microbiol, 2000 Nov, 89(5), 801 - 6 Heat resistance of Paenibacillus polymyxa in relation to pH and acidulants; Casadei MA et al.; The efficacy of different organic acids in decreasing the heat resistance of Paenibacillus polymyxa spores was assessed . The relationship between concentration of the undissociated form of different organic acids and decrease in heat resistance was also investigated . The heat resistance of P . polymyxa spores was tested in distilled water at 85, 90 and 95 degrees C, at pH4 and in the presence of 50, 100 and 200 mmol l(-1) of the undissociated form of lactic, citric or acetic acid and sodium citrate or acetate . The undissociated form of organic acids was responsible for increasing the heat sensitivity of spores . The most effective acid was lactic acid . The D values of the spores decreased rapidly (between 74 and 43%) in the presence of 50 mmol l(-1) of the undissociated form of organic acid, and increasing concentrations of these forms affected the heat resistance of spores less than proportionally . The heat resistance of the spores in milk was approximately threefold lower than in distilled water . This work has shown that the undissociated fraction of organic acids increases, albeit non-linearly, the sensitivity of spores to heat, even in complex substrates such as milk . By knowing the amount of organic acids added to a given substrate, their dissociation constants and the final pH, it could be possible to estimate the concentration of undissociated forms and the corresponding increase in lethality of heat treatments . This would help the food industry to maximize the lethality achieved by heat processes and/or safely reduce the heat treatments already in use. Syst Appl Microbiol, 2000 Oct, 23(3), 344 - 8 Paenibacillus granivorans sp . nov., a new Paenibacillus species which degrades native potato starch granules; van der Maarel MJ et al.; From a native potato starch-degrading enrichment culture, strain A30 had been isolated and had tentatively been identified as a member of the Bacillus firmus/lentus group (Wijbenga et al . Appl . Microbiol . Biotechnol . 35, 180-184, 1991) . In this paper the isolate A30 is further characterized using phylogentic analysis of the 16S rDNA and determination of a number of additional phenotypic characteristics . These data are compared to those of Paenibacillus amylolyticus, P . chibensis, and P . thiaminolyticus . It is concluded that strain A30 is a new Paenibacillus species, for which the name Paenibacillus granivorans is suggested. Bioseparation, 2000, 9(3), 185 - 8 Purification, characterization and crystallization of enzymes for dibenzothiophene desulfurization; Ohshiro T et al.; DszC and DszA, DBT monooxygenase and DBT sulfone monooxygenase, respectively, involved in dibenzothiophene (DBT) desulfurization, were purified to homogeneity from Rhodococcus erythropolis D-1 . The two enzymes were crystallized and enzymologically characterized . We found a high activity of flavin reductase in the non-DBT-desulfurizing bacterium, Paenibacillus polymyxa A-1, which is essential for DszC and A activities, and purified to homogeneity and characterized the enzyme. Carbohydr Res, 2000 Oct 20, 329(1), 57 - 63 Two-step enzymatic synthesis of maltooligosaccharide esters; Degn P et al.; Glucose and maltose esters were synthesised in organic media by employing a lipase (E.C . 3.1.1.3) from Candida antarctica . In a second reaction step, a transglycosylation catalysed by a cyclodextrin glycosyltransferase (E.C . 2.4.1.19) from either Paenibacillus sp . F8 or Bacillus sp . strain no . 169 (DSM 2518) extended the degree of polymerisation (DP) of the carbohydrate moieties of the carbohydrate esters . The donor substrates used were either a cyclodextrin, a maltooligosaccharide or starch . The highest rate of low DP maltooligosaccharide ester formation was obtained when starch was used as glycosyl donor and caproyl maltose as glycosyl acceptor . The structures of two of the products were identified by 1H and 13C NMR and MALDI-TOF MS as capronate monoesters of maltotriose and maltotetraose, with the ester bond at C-6 of the second glucose unit from the reducing end. Appl Environ Microbiol, 2000 Nov, 66(11), 4998 - 5004 Purification and properties of an enzyme capable of degrading the sheath of Sphaerotilus natans; Takeda M et al.; Microorganisms which can degrade and grow on the purified sheath of a sheathed bacterium Sphaerotilus natans were collected from soil and river water . Two bacterial strains were isolated from the soil and designated strains TB and TK . Both strains are rod shaped, negatively stained by gram staining, facultatively anaerobic, and formed ellipsoidal endospores . These characteristics suggested that the isolates belong to the genus Paenibacillus, according to Ash et al . (C . Ash, F . G . Priest, and M . D . Collins, Antonie Leeuwenhoek 64:253-260, 1993) . Phylogenetic analysis based on the 16S rDNA supported this possibility . Purification of the sheath-degrading enzyme was carried out from the culture broth of strain TB . The molecular weight of the enzyme was calculated to be 78,000 and 50, 000 by sodium dodecyl sulfate-polyacrylamide electrophoresis and gel filtration chromatography, respectively . Enzyme activity was optimized at pH 6.5 to 7.0 and 30 to 40 degrees C . The reaction was accelerated by the addition of Mg(2+), Ca(2+), Fe(3+), and iodoacetamide, whereas it was inhibited by the addition of Cu(2+), Mn(2+), and dithiothreitol . The enzyme acted on the polysaccharide moiety of the sheath, producing an oligosaccharide the size of which was between the sizes of maltopentaose and maltohexaose . As the reaction proceeded, the absorbance at 235 nm of the reaction mixture increased, suggesting the generation of unsaturated sugars . Incorporation of unsaturated sugars was also suggested by the thiobarbituric acid reaction . It is possible that the enzyme is not a hydrolytic enzyme but a kind of polysaccharide eliminase which acts on the basic polysaccharide. J Appl Microbiol, 2000 Oct, 89(4), 595 - 8 Direct microscopy of bacillus endospore germination in soil microcosms; Thrane C et al.; Antagonistic endospore-forming Bacillus spp . offer a large potential as seed inoculants for control of soil-borne pathogens . In the soil, however, inoculated Bacillus endospores may remain dormant without germination, and plant protection can therefore be inefficient and unpredictable . A method based on direct fluorescence microscopy in soil microcosms was used to determine whether low-cost organic additives incorporated into seed coating material could stimulate endospore germination . Complex organic additives supported a high level of endospore germination of the fungal antagonist Paenibacillus polymyxa CM5-5 . Skim milk is a low-cost additive that may be incorporated into seed coating material for efficient induction of Bacillus endospore germination in soil. J Invertebr Pathol, 2000 Oct, 76(3), 169 - 75 Paenibacillus associated with milky disease in Central and South American scarabs; Harrison H et al.; Thirty-one isolates of bacteria causing milky disease in scarab larvae collected in Central and South America were identified as Paenibacillus popilliae or Paenibacillus lentimorbus by use of DNA similarity analysis . The isolates were more similar to each other than to the North American isolates that are the type strains of the species . All of the bacteria of both species produced parasporal bodies, a characteristic previously believed to be unique to P . popilliae . Screening of the bacteria using PCR with parasporal protein primers revealed differences among the parasporal protein genes of P . popilliae isolates and between the parasporal genes of P . popilliae and P . lentimorbus . In contrast to P . popilliae from North America, none of the isolates from Central and South America was resistant to vancomycin, an indication of an interesting geographic distribution of the resistance genes . Phytochemistry, 2000 Sep, 55(1), 29 - 34 Two polyisoprenylated benzophenones from the trunk latex of Clusia grandiflora (Clusiaceae); Lokvam J et al.; The polyisoprenylated benzophenones, chamones I and II, were isolated from the trunk latex of Clusia grandiflora (Clusiaceae) growing in southeastern Venezuela . A third benzophenone, nemorosone II, was isolated from the pollinator reward resin of the female flowers of the same plant . Chamone I and nemorosone II are structurally similar, differing only in the degree of prenylation . Bioassays of chamone I and nemorosone II using the honeybee pathogens, Paenibacillus larvae and Paenibacillus alvei, demonstrate that both have potent antibacterial activity, and that their structural differences affect both their bactericidal efficacies and their aqueous mobilities. Appl Environ Microbiol, 2000 Sep, 66(9), 3945 - 50 A novel gene encoding xanthan lyase of Paenibacillus alginolyticus strain XL-1; Ruijssenaars HJ et al.; Xanthan-modifying enzymes are powerful tools in studying structure-function relationships of this polysaccharide . One of these modifying enzymes is xanthan lyase, which removes the terminal side chain residue of xanthan . In this paper, the cloning and sequencing of the first xanthan lyase-encoding gene is described, i . e., the xalA gene, encoding pyruvated mannose-specific xanthan lyase of Paenibacillus alginolyticus XL-1 . The xalA gene encoded a 100, 823-Da protein, including a 36-amino-acid signal sequence . The 96, 887-Da mature enzyme could be expressed functionally in Escherichia coli . Like the native enzyme, the recombinant enzyme showed no activity on depyruvated xanthan . Compared to production by P . alginolyticus, a 30-fold increase in volumetric productivity of soluble xanthan lyase was achieved by heterologous production in E . coli . The recombinant xanthan lyase was used to produce modified xanthan, which showed a dramatic loss of the capacity to form gels with locust bean gum. Int J Syst Evol Microbiol, 2000 Jul, 50 Pt 4, 1495 - 500 Paenibacillus koreensis sp . nov., a new species that produces an iturin-like antifungal compound; Chung YR et al.; A bacterial strain, YC300T, that produces an iturin-like antifungal antibiotic was isolated from compost and identified as member of the genus Paenibacillus . Gram reaction of the strain was variable depending upon growth stages and culture media . Three different types of colonies were developed on tryptic soy agar . The organism was facultatively anaerobic and grew at 50 degrees C . The DNA G+C content was 54 mol % and anteiso-C15:0 was the major fatty acid . A 0.9 kb fragment was produced by PCR amplification of strain YC300T DNA using primers PAEN515F and 1377R . Levels of 16S rDNA similarity between strain YC300T and other Paenibacillus species were between 89.8 and 94.8% . Phylogenetically, strain YC300T formed a significant monophyletic clade with Paenibacillus validus . It is clear from polyphasic evidence that the isolate should be classified as Paenibacillus koreensis sp . nov., the type strain of which is YC300T (= KCTC 2393T, KCCM 40903T). Res Microbiol, 2000 Jun, 151(5), 369 - 81 Diversity of Paenibacillus polymyxa strains isolated from the rhizosphere of maize planted in Cerrado soil; von der Weid I et al.; Paenibacillus polymyxa populations present in the rhizosphere of maize (cultivar BR-201) planted in Cerrado soil were investigated in order to assess their diversity at four stages of plant growth . A total of 67 strains were isolated and all strains were identified as P . polymyxa by classical biochemical tests, API 50CH tests and a set of species-specific primers based on the 23S rDNA sequence . To compare the isolated strains, phenotypic characteristics (utilization of different carbohydrates, resistance to antibiotics and production of antimicrobial substances) and genetic approaches (hybridization with a Klebsiella pneumoniae nifKDH probe and BOX-PCR) were used . Fermentation of glycerol, arabinose, xylose and rhamnose varied among the isolates and these data divided the strains into five groups . Fifty strains (75%) showed homology to plasmid pSA30 (containing the nifKDH genes) resulting in five different hybridization patterns . Using BOX-PCR, 18 groups were observed . Phenetic analyses were applied based on the unweighted pair group method with arithmetic means using the phenotypic and genetic data, separately . All P . polymyxa isolates could be divided into two main clusters at approximately 52% and into 18 groups at approximately 89% of similarity, when phenotypic data were used . Also, two main clusters were formed at 65% of similarity when genetic data were used . In this dendrogram, clusters were further split into 10 and 22 groups, at about 88 and 97% of similarity, respectively . Finally, all phenotypic and genetic data, or just the genetic data, were used in a multivariate analysis of variance (MANOVA) in order to address the heterogeneity among P . polymyxa populations during the different stages of maize growth . The resulting data showed that strains isolated 10, 30, 60 and 90 days after maize sowing were statistically different. Res Microbiol, 2000 May, 151(4), 303 - 12 estA, a gene coding for a cell-bound esterase from Paenibacillus sp . BP-23, is a new member of the bacterial subclass of type B carboxylesterases; Prim N et al.; Screening of a gene library from Paenibacillus sp . BP-23 generated in Escherichia coli led to identification of a clone that directed the production of lipolytic activity . From the sequencing data, we found an open reading frame encoding a protein of 485 amino acids with an estimated molecular mass of 53 kDa and a pI of 5.1 . Absence of a signal peptide indicated that it was a cell-bound protein . Sequence analysis showed that the protein contained the signature G-XI-S-X2-G included in most serine-esterases and lipases . The cloned protein showed high homology with enzymes belonging to the bacterial subclass of type B carboxylesterases . The enzyme had a significant preference for esters of short-chain fatty acids and showed the kinetics behaviour of a true esterase . Maximum activity was found at pH 7.5 and 37 degrees C, although the enzyme was active in the pH range 6.0- 9.0 and at temperatures up to 45 degrees C . As expected for a serine-esterase, activity was inhibited by phenylmethylsulphonyl fluoride. J Biol Chem, 2000 Sep 15, 275(37), 28843 - 8 Increased thermal resistance and modification of the catalytic properties of a beta-glucosidase by random mutagenesis and in vitro recombination; Arrizubieta MJ et al.; The bglB gene from Paenibacillus polymyxa was subjected to random mutagenesis mediated by error prone polymerase chain reaction amplification and DNA shuffling . After this treatment, mutant variants of the encoded beta-glucosidase with enhanced thermal resistance were selected . We identified five amino acid substitutions at four different positions of the sequence that increased the resistance of the enzyme to heat denaturation . Four of the mutations, H62R, M319V, M319I, and M361I, did not change the kinetic parameters of the enzyme . However, mutant N223Y, which caused only a marginal increase in thermoresistance, showed an 8-fold decrease in K(m) . Copies of the bglB gene carrying each one of the individual mutations were recombined in vitro by DNA shuffling . As a result, we obtained an enzyme that simultaneously exhibited a 20-fold increase in heat resistance and an 8-fold increase in the catalytic efficiency . The structural basis of the properties conferred by the mutations was analyzed using homology-based structural models . The four mutations causing a more pronounced effect on thermoresistance were located in loops, on the periphery of the (alpha/beta)(8) barrel that conforms the structure of the protein . Mutation N223Y, which modifies the catalytic properties of the enzyme, was on one of the barrel beta-strands that shape the active center. Curr Microbiol, 2000 Aug, 41(2), 84 - 8 Phylogeny of marine Bacillus isolates from the Gulf of Mexico; Siefert JL et al.; The phylogeny of 11 pigmented, aerobic, spore-forming isolates from marine sources was studied . Forty-two biochemical characteristics were examined, and a 16S rDNA sequence was obtained for each isolate . In a phylogenetic tree based on 16S sequencing, four isolates (NRRL B-14850, NRRL B-14904, NRRL B-14907, and NRRL B-14908) clustered with B . subtilis and related organisms; NRRL B-14907 was closely related to B . amyloliquefaciens . NRRL B-14907 and NRRL B-14908 were phenotypically similar to B . amyloliquefaciens and B . pumilus, respectively . Three strains (NRRL B-14906, NRRL B-14910, and NRRL B-14911) clustered in a clade that included B . firmus, B . lentus, and B . megaterium . NRRL B-14910 was closely related phenotypically and phylogenetically to B . megaterium . NRRL B-14905 clustered with the mesophilic round spore-producing species, B . fusiformis and B . sphaericus; the isolate was more closely related to B . fusiformis . NRRL B-14905 displayed characteristics typical of the B . sphaericus-like organisms . NRRL B-14909 and NRRL B-14912 clustered with the Paenibacillus species and displayed characteristics typical of the genus . Only NRRL B-14851, an unusually thin rod that forms very small spores, may represent a new Bacillus species. J Econ Entomol, 2000 Apr, 93(2), 199 - 209 Comparative laboratory toxicity of neem pesticides to honey bees (Hymenoptera: Apidae), their mite parasites Varroa jacobsoni (Acari: Varroidae) and Acarapis woodi (Acari: Tarsonemidae), and brood pathogens Paenibacillus larvae and Ascophaera apis; Melathopoulos AP et al.; Laboratory bioassays were conducted to evaluate neem oil and neem extract for the management of key honey bee (Apis mellifera L.) pests . Neem pesticides inhibited the growth of Paenibacillus larvae (Ash, Priest & Collins) in vitro but had no effect on the growth of Ascophaera apis (Olive & Spiltoir) . Azadirachtin-rich extract (neem-aza) was 10 times more potent than crude neem oil (neem oil) against P . larvae suggesting that azadirachtin is a main antibiotic component in neem . Neem-aza, however, was ineffective at controlling the honey bee mite parasites Varroa jacobsoni (Ouduemans) and Acarapis woodi (Rennie) . Honey bees also were deterred from feeding on sucrose syrup containing > 0.01 mg/ml of neem-aza . However, neem oil applied topically to infested bees in the laboratory proved highly effective against both mite species . Approximately 50-90% V . jacobsoni mortality was observed 48 h after treatment with associated bee mortality lower than 10% . Although topically applied neem oil did not result in direct A . woodi mortality, it offered significant protection of bees from infestation by A . woodi . Other vegetable and petroleum-based oils also offered selective control of honey bee mites, suggesting neem oil has both a physical and a toxicological mode of action . Although oils are not as selective as the V . jacobsoni acaricide tau-fluvalinate, they nonetheless hold promise for the simultaneous management of several honey bee pests. Cell Biol Int, 2000, 24(5), 319 - 24 Histochemical characterization of cell death in honeybee larvae midgut after treatment with Paenibacillus larvae, Amitraz and Oxytetracycline; Gregorc A et al.; A number of techniques were employed to assess cell death induced in honeybee larvae midgut after per os inoculation of bacterium Paenibacillus larvae var . larvae, the causative agent of American foulbrood disease, and separately with acaricide Amitraz and antibiotic Oxytetracycline . In honeybee larvae exposed to Amitraz, which demonstrates both necrosis and apoptosis, cell death was found in 82% of midgut columnar and in 50% of regenerative epithelial cells, 24 h after treatment . Cell death reduced to 36% in the epithelial cells, 48 h after treatment . In Oxytetracycline-treated larvae, cell death was identified in 40% of midgut epithelial cells, 24 h after inoculation and increased to 55% over the next 24 h . In Paenibacillus -infected larvae, all midgut epithelial cells died . Using ApopTag (Oncor) to label the multiple DNA ends generated by DNA fragmentation showed programmed cell death in 49% of columnar midgut cells 24 h after Amitraz application . Cell death was reduced to 9% over the next 24 h . Our data indicate that cell death could be identified and quantified in situ, using TUNEL techniques . This study also shows that the acaricide Amitraz is a trigger for programmed cell death in the midgut epithelial cells of honeybee larvae, unlike Paenibacillus which induces necrosis only . The data show that immunohistochemical methods are useful for studying in situ tissue pathology, and indicate possibilities for monitoring the effects of infective and chemical environmental stressors on cell death in honeybee larvae tissue . Int J Syst Evol Microbiol, 2000 Mar, 50 Pt 2, 939 - 40 What is the type species of the genus Paenibacillus? Request for an opinion; Tindall BJ; The taxonomic status of the type species of the genus Paenibacillus cannot be easily determined according to the rules of the Bacteriological Code since the rules may be interpreted in an ambiguous way . Depending upon how the rules are applied the type species may be either Paenibacillus polymyxa or Paenibacillus durus . In addition, depending upon the way in which the Bacteriological Code is interpreted, the question of whether the name P . durus (Collins et al . 1994) has been validly published must also be addressed. Appl Environ Microbiol, 2000 Mar, 66(3), 1098 - 106 Genetic and biochemical diversity among isolates of Paenibacillus alvei cultured from Australian honeybee (Apis mellifera) colonies; Djordjevic SP et al.; Twenty-five unique CfoI-generated whole-cell DNA profiles were identified in a study of 30 Paenibacillus alvei isolates cultured from honey and diseased larvae collected from honeybee (Apis mellifera) colonies in geographically diverse areas in Australia . The fingerprint patterns were highly variable and readily discernible from one another, which highlighted the potential of this method for tracing the movement of isolates in epidemiological studies . 16S rRNA gene fragments (length, 1,416 bp) for all 30 isolates were enzymatically amplified by PCR and subjected to restriction analysis with DraI, HinfI, CfoI, AluI, FokI, and RsaI . With each enzyme the restriction profiles of the 16S rRNA genes from all 30 isolates were identical (one restriction fragment length polymorphism {RFLP} was observed in the HinfI profile of the 16S rRNA gene from isolate 17), which confirmed that the isolates belonged to the same species . The restriction profiles generated by using DraI, FokI, and HinfI differentiated P . alvei from the phylogenetically closely related species Paenibacillus macerans and Paenibacillus macquariensis . Alveolysin gene fragments (length, 1, 555 bp) were enzymatically amplified from some of the P . alvei isolates (19 of 30 isolates), and RFLP were detected by using the enzymes CfoI, Sau3AI, and RsaI . Extrachromosomal DNA ranging in size from 1 to 10 kb was detected in 17 of 30 (57%) P . alvei whole-cell DNA profiles . Extensive biochemical heterogeneity was observed among the 28 P . alvei isolates examined with the API 50CHB system . All of these isolates were catalase, oxidase, and Voges-Proskauer positive and nitrate negative, and all produced acid when glycerol, esculin, and maltose were added . The isolates produced variable results for 16 of the 49 biochemical tests; negative reactions were recorded in the remaining 30 assays . The genetic and biochemical heterogeneity in P . alvei isolates may be a reflection of adaptation to the special habitats in which they originated. Antimicrob Agents Chemother, 2000 Mar, 44(3), 705 - 9 The biopesticide Paenibacillus popilliae has a vancomycin resistance gene cluster homologous to the enterococcal VanA vancomycin resistance gene cluster; Patel R et al.; We have previously identified, in Paenibacillus popilliae, a 708-bp sequence which has homology to the sequence of the enterococcal vanA gene . We have performed further studies revealing five genes encoding homologues of VanY, VanZ, VanH, VanA, and VanX in P . popilliae . The predicted amino acid sequences are similar to those in VanA vancomycin-resistant enterococci: 61% identity for VanY, 21% for VanZ, 74% for VanH, 77% for VanA, and 79% for VanX . The genes in P . popilliae may have been a precursor to or have had ancestral genes in common with vancomycin resistance genes in enterococci . The use of P . popilliae biopesticidal preparations in agricultural practice may have an impact on bacterial resistance in human pathogens. Cell Biol Int, 1999, 23(3), 211 - 8 In situ localization of heat-shock and histone proteins in honey-bee (Apis mellifera l.) larvae infected with Paenibacillus larvae; Gregorc A et al.; The immunohistochemical localization of the heat shock proteins (Hsp70 and Hsp90) and histone protein in healthy and Paenibacillus larvae infected honeybee (Apis mellifera L.) larvae has been studied . Hsp70 was found in the nuclei and the cytoplasm of infected midgut, salivary gland cells and haemocytes, but not in uninfected larvae . Hsp90 was localized in both infected and uninfected cells . Exposed histone proteins were localized in the nuclei of dying uninfected cells undergoing programmed cell death . The distribution of histone protein in uninfected cells of midgut, salivary gland, and other tissues was nuclear and indicative of normal programmed cell death at levels between 1 and 5%.After applying histone protein antibodies to P . larvae infected honeybee larvae, the DAB based reaction product was located in the nuclei or immediate surroundings of all larval cells . The Hsp70, Hsp90 and histone protein distribution patterns are discussed in relation to the morphological, cytochemical and immunocytochemical characteristics of programmed cell death and pathological necrosis . Results produced by methyl green-pyronin staining confirm an elevation of RNA levels in normal programmed cell death and a reduced staining for RNA in necrotic infected cells . Appl Environ Microbiol, 1999 Nov, 65(11), 5148 - 50 Isolation from the Sorghum bicolor mycorrhizosphere of a bacterium compatible with arbuscular mycorrhiza development and antagonistic towards soilborne fungal pathogens; Budi SW et al.; A gram-positive bacterium with antagonistic activity towards soilborne fungal pathogens has been isolated from the mycorrhizosphere of Sorghum bicolor inoculated with Glomus mosseae . It has been identified as Paenibacillus sp . strain B2 based on its analytical profile index and on 16S ribosomal DNA analysis . Besides having antagonistic activity, this bacterium stimulates mycorrhization. Microbiology, 1999 Jul, 145 ( Pt 7), 1731 - 41 Differential detection of key enzymes of polyaromatic-hydrocarbon-degrading bacteria using PCR and gene probes; Meyer S et al.; Bacteria with ability to degrade polyaromatic hydrocarbons (PAHs), isolated from wastewater and soil samples, were investigated for their taxonomic, physiological and genetic diversity . Eighteen isolates able to metabolize naphthalene or phenanthrene as sole carbon source were taxonomically affiliated to different subclasses of the Proteobacteria (Sphingomonas spp., Acidovorax spp., Comamonas spp . and Pseudomonas spp.) and to phyla of Gram-positive bacteria with low and high DNA G + C content (Paenibacillus sp . and Rhodococcus spp., respectively) . Representatives of the genera Pseudomonas and Sphingomonas formed a remarkably high fraction of these isolates; 9 out of 18 strains belonged to these groups . Tests for enzyme activities showed that the majority of the isolates growing with PAHs as sole sources of carbon and energy had an active catechol 2,3-dioxygenase (C230) . C230 specific activities were very diverse, ranging from 0.1 to 650 mU (mg protein)-1 . Pseudomonas and Sphingomonas strains showed considerably higher activities than the other isolates . All PAH degraders were examined for the presence of an initial PAH dioxygenase and C230, which catalyse key steps of PAH degradation, by PCR amplification of gene fragments and subsequent hybridization . PCR primers and internal oligonucleotide probes were developed for the specific detection of the genes of Pseudomonas and Sphingomonas strains. Int J Syst Bacteriol, 1999 Jul, 49 Pt 3, 1083 - 90 Proposal of Virgibacillus proomii sp . nov . and emended description of Virgibacillus pantothenticus (Proom and Knight 1950) Heyndrickx et al . 1998; Heyndrickx M et al.; A polyphasic study of strains originally received as Bacillus (now Virgibacillus) pantothenticus, along with strains representing species belonging to Bacillus, Halobacillus and Paenibacillus, was undertaken using amplified rDNA restriction analysis (ARDRA), fatty acid methyl ester (FAME) analysis, SDS-PAGE of whole-cell proteins and routine diagnostic characters comprising 61 biochemical tests in the API system and 15 observations of vegetative cell and sporangial morphology . It revealed the presence within Virgibacillus of an as yet undescribed new species, for which the name Virgibacillus proomii is proposed; V . proomii can be distinguished from V . pantothenticus and members of Bacillus sensu stricto, and from members of Paenibacillus and other aerobic endospore-forming bacteria, by routine phenotypic tests . The type strain of Virgibacillus proomii is LMG 12370T. Appl Environ Microbiol, 1999 Jun, 65(6), 2446 - 52 A pyruvated mannose-specific xanthan lyase involved in xanthan degradation by Paenibacillus alginolyticus XL-1; Ruijssenaars HJ et al.; The xanthan-degrading bacterium Paenibacillus alginolyticus XL-1, isolated from soil, degrades approximately 28% of the xanthan molecule and appears to leave the backbone intact . Several xanthan-degrading enzymes were excreted during growth on xanthan, including xanthan lyase . Xanthan lyase production was induced by xanthan and inhibited by glucose and low-molecular-weight enzymatic degradation products from xanthan . A xanthan lyase with a molecular mass of 85 kDa and a pI of 7.9 was purified and characterized . The enzyme is specific for pyruvated mannosyl side chain residues and optimally active at pH 6.0 and 55 degrees C. Int J Syst Bacteriol, 1999 Apr, 49 Pt 2, 531 - 40 Transfer of Bacillus lentimorbus and Bacillus popilliae to the genus Paenibacillus with emended descriptions of Paenibacillus lentimorbus comb . nov . and Paenibacillus popilliae comb . nov; Pettersson B et al.; Almost complete 16S rRNA gene sequences were generated for the type strains of the obligate insect pathogens Bacillus lentimorbus and Bacillus popilliae and a second strain of Bacillus popilliae (NRRL B-4081) received as 'Bacillus popilliae var . melolonthae' . A phylogenetic tree was constructed which grouped these strains into a well defined subcluster within the genus Paenibacillus . Bacillus popilliae NRRL B-4081 occupied an intermediate position between the type strains of Bacillus lentimorbus and Bacillus popilliae but with a marked clustering to the latter . The phylogenetic assignment of these strains to Paenibacillus is in contrast to earlier studies which placed these bacteria in the genus Bacillus, close to Bacillus subtilis . Indeed, the rRNA sequences generated in this study share less than 88% similarity to the deposited sequences for Bacillus popilliae ATCC 14706T and Bacillus lentimorbus ATCC 14707T . The results obtained by using different tree algorithms, bootstrap analysis, branch lengths and verification by signature nucleotide analysis supported the reclassification of these species in the genus Paenibacillus as Paenibacillus lentimorbus comb . nov . and Paenibacillus popilliae comb . nov. Appl Environ Microbiol, 1999 May, 65(5), 2243 - 5 A PCR detection method for rapid identification of Paenibacillus larvae; Govan VA et al.; American foulbrood is a disease of larval honeybees (Apis mellifera) caused by the bacterium Paenibacillus larvae . Over the years attempts have been made to develop a selective medium for the detection of P . larvae spores from honey samples . The most successful of these is a semiselective medium containing nalidixic acid and pipermedic acid . Although this medium allows the growth of P . larvae and prevents the growth of most other bacterial species, the false-positive colonies that grow on it prevent the rapid confirmation of the presence of P . larvae . Here we describe a PCR detection method which can be used on the colonies that grow on this semiselective medium and thereby allows the rapid confirmation of the presence of P . larvae . The PCR primers were designed on the basis of the 16S rRNA gene of P . larvae and selectively amplify a 973-bp amplicon . The PCR amplicon was confirmed as originating from P . larvae by sequencing in both directions . Detection was specific for P . larvae, and the primers did not hybridize with DNA from closely related bacterial species. J Appl Microbiol, 1999 Jan, 86(1), 13 - 21 Antibiosis plays a role in the context of direct interaction during antagonism of Paenibacillus polymyxa towards Fusarium oxysporum; Dijksterhuis J et al.; Interaction of Fusarium oxysporum and Paenibacillus polymyxa starts with polar attachment of bacteria to the fungal hyphae followed by the formation of a large cluster of non-motile cells embedded in an extracellular matrix in which the bacteria develop endospores . Enumeration of fungal viable counts showed that less than one of 36,000 colony-forming units survived in paired cultures for 71 h . Effective antagonism was not observed below pH5 and was specific for the bacterial species . Development of F . oxysporum was inhibited in cell-free filtrates derived from cultures of P . polymyxa, but was much more strongly repressed in the presence of living bacteria . Furthermore, recovery of fungal growth started immediately after addition of antibiotics to paired cultures . Restoration of fungal growth was enhanced in filtrates that were supplemented with MgCl2, which suggests that anti-fungal compounds produced by the bacteria were counteracted by magnesium ions . In paired cultures, fungal counts remained very low, even in the presence of the magnesium salt . This study clearly showed that P . polymyxa antagonizes the plant pathogenic fungus F . oxysporum in liquid medium by means of an interaction process in which the presence of living bacteria is a prerequisite for continuous suppression of fungal growth. Int J Syst Bacteriol, 1999 Jan, 49 Pt 1, 239 - 46 Paenibacillus dendritiformis sp . nov., proposal for a new pattern-forming species and its localization within a phylogenetic cluster; Tcherpakov M et al.; A new strain capable of forming distinctive patterns during colony development was identified by using a combination of phenotypic characterization, fatty acid analysis and analysis of the 16S rRNA gene sequence . The strain formed either a branched, tip-splitting colony morphology (referred to as the T morphotype) or a chiral pattern exhibiting thinner branches with distinctive curling patterns (referred to as the C morphotype) . Isolates of the T morphotype exhibited sequence identities greater than 97% to Paenibacillus thiaminolyticus JCM 7540 . Phylogenetic analysis placed the T morphotype within the Paenibacillus cluster on a phylogenetic tree . On the basis of unique colony morphology and distinctive phenotypic characteristics, it is proposed that the pattern-forming isolates should be placed within a new species of Paenibacillus, Paenibacillus dendritiformis sp . nov., the type strain of which is T168T (= 30A1T). Appl Environ Microbiol, 1998 Oct, 64(10), 3860 - 8 Comparison of paenibacillus azotofixans strains isolated from rhizoplane, rhizosphere, and non-root-associated soil from maize planted in two different brazilian soils Seldin L, Rosado AS, da Cruz DW, Nobrega A, van Elsas JD, Paiva E. Paenibacillus azotofixans is a nitrogen-fixing bacterium often found in soil and in the rhizospheres of different grasses . In this study, two Brazilian clay soils were planted with cross-hybrid maize (BR-201) and four stages of plant growth were analyzed to characterize the P . azotofixans populations present in the rhizoplanes, rhizospheres, and non-root-associated soils (herein called nonrhizospheres) . A total of 106 strains were isolated and identified as P . azotofixans with an API 50CH kit, by classical biochemical tests, and via the use of specific primers based on the 16S rRNA gene in PCRs . To compare the isolated strains, phenotypic characteristics were determined and three different probes were used in hybridization experiments: two nif probes and one probe comprising a 0.58-kb fragment cloned from the P . azotofixans C3L4 genome . These results were used to construct a dendrogram, in which two main clusters could be observed . One cluster contained exclusively strains from Varzea soil, and the other contained the majority of strains from Cerrado soil . The 60 strains from Varzea soil and the 46 strains from Cerrado soil were further analyzed with REP and BOX primers, respectively . Based on the patterns obtained, it was possible to identify 21 different groups among strains from Varzea soil and 4 different groups among strains from Cerrado soil . These different patterns were tested by multivariate analysis of variance, and differences in the populations of P . azotofixans during the four stages of plant growth were demonstrated . Moreover, strains isolated from the rhizoplanes, rhizospheres, and nonrhizospheres of maize planted in Cerrado and Varzea soils were shown to be statistically different; the diversity of P . azotofixans strains was affected by the soil type. J Appl Microbiol, 1998 Sep, 85(3), 463 - 71 Quantitative detection of Sphingomonas chlorophenolica in soil via competitive polymerase chain reaction; van Elsas JD et al.; The 16S ribosomal RNA gene sequence of the pentachlorophenol degrader Sphingomonas chlorophenolica strain RA2 was used to generate specific polymerase chain reaction (PCR) primers for the detection of this strain in soil, whereas a region internal to the two primers was used to provide an S . chlorophenolica strain RA2-specific oligonucleotide probe . The PCR detection system resulted in a 727 bp product detectable via gel electrophoresis and hybridization . It was specific for strain RA2 and its close relative, S . chlorophenolica ATCC 39723, as evidenced by PCR amplifications of a range of bacterial genomic DNAs . Tests of total microbial community DNA obtained from five uninoculated and two RA2-inoculated soils confirmed this specificity for introduced S . chlorophenolica RA2 . Strain RA2 could be detected in soil down to a level of 10(3) cfu g-1 soil . Two strategies were followed to generate internal standard DNA for competitive PCR . First, a 479 bp MIMICS fragment was obtained based on a previously constructed gene cassette; however, this standard did not reliably quantify RA2 targets . Low stringency PCR performed with a range of bacterial genomic DNAs resulted in the generation of an amplicon with a Paenibacillus azotofixans strain that was slightly smaller than the RA2-derived product . Both products were easily separable via conventional gel electrophoresis . The use of this competitor in a threefold dilution scheme applied to the target DNA allowed for the quantitative detection of RA2-specific target DNA molecules from pure culture and from soil . The fate of strain RA2 in pentachlorophenol-contaminated soil was described using this competitive PCR approach, and the organism was shown to persist at two inoculum levels over prolonged periods of time. Int J Syst Bacteriol, 1998 Jul, 48 Pt 3, 833 - 7 Paenibacillus campinasensis sp . nov., a cyclodextrin-producing bacterium isolated in Brazil; Yoon JH et al.; An alkaliphilic, endospore-forming bacterium isolated from Brazilian soil was taxonomically studied and is proposed as a new Paenibacillus species . This organism (strain 324T) was particularly distinguishable from other Paenibacillus species by its ability to grow optimally at pH 10 and 40 degrees C . The DNA G+C content was 5.0 mol% . The diamine acid of the cell-wall peptidoglycan was meso-diaminopimelic acid . MK-7 was the predominant menaquinone and anteiso-C15:0 was the major fatty acid . Levels of 16S rDNA similarity between strain 324T and other Paenibacillus species were 90.6-95.9% . Phylogenetically, strain 324T formed an evolutionary lineage distinct from other species within the evolutionary radiation encompassing the genus Paenibacillus . Based on phenotyic and chemotaxonomic properties, and phylogenetic inference, it is proposed that strain 324T should be placed in the genus Paenibacillus as a new species is strain 324T should be placed in the genus Paenibacilus as a new species, Paenibacillus campinasensis . This type strain of the new species is strain 325T (= KCTC 0364BP). Arch Microbiol, 1998 Oct, 170(4), 304 - 8 Expression of the succinate dehydrogenase genes (sdhCAB) from the facultatively anaerobic paenibacillus macerans during aerobic growth Schirawski J, Hankeln T, Unden G. Paenibacillus (formerly Bacillus) macerans is capable of succinate oxidation under oxic conditions and fumarate reduction under anoxic conditions . The reactions are catalyzed by different enzymes, succinate dehydrogenase (Sdh) and fumarate reductase (Frd) . The genes encoding Sdh (sdhCAB) were analyzed . The gene products of sdhA and sdhB were similar to the subunits of known Sdh and Frd enzymes . The hydrophobic subunit SdhC showed close sequence similarity to the class of Sdh/Frd enzymes containing diheme cytochrome b . From the sdhCAB gene cluster two transcripts were produced, one comprising sdhCAB, the other sdhAB . The transcripts were found only during aerobic growth, and the amount was directly proportional to Sdh activity, but inversely proportional to Frd activity. J Vet Pharmacol Ther, 1998 Aug, 21(4), 269 - 73 Disposition of mirosamicin in honeybee hives; Nakajima C et al.; Disposition of mirosamicin, a macrolide antibiotic, to honeybee adults, larvae, honey and royal jelly in the beehive after in-feed administration to adult bees was studied . Treatment was initiated at the end of July when the availability of natural pollen and nectar was poor . The drug was mixed with pollen-substitute paste and administered to honeybee colonies continuously for a week at a dosage of 200 mg/hive/week . High distributions in adult bees, jelly, larvae and a relatively low distribution in honey, of mirosamicin were observed . One day dosing of microsamin in sucrose syrup, a nectar substitute, resulted in a very high and long lasting residue in honey . Both royal and worker jelly, secreted from the jelly glands of adult bees, are acidic, so that a high distribution of a basic drug, such as mirosamicin, in jelly can be expected . This mechanism was considered to be responsible for a high concentration of mirosamicin in honeybee larvae, the host of Paenibacillus-larvae infection (American foulbrood), as primary larval food is jelly. Appl Environ Microbiol, 1998 Aug, 64(8), 2770 - 9 Genetic diversity of nifH gene sequences in paenibacillus azotofixans strains and soil samples analyzed by denaturing gradient gel electrophoresis of PCR-amplified gene fragments; Rosado AS et al.; The diversity of dinitrogenase reductase gene (nifH) fragments in Paenibacillus azotofixans strains was investigated by using molecular methods . The partial nifH gene sequences of eight P . azotofixans strains, as well as one strain each of the close relatives Paenibacillus durum, Paenibacillus polymyxa, and Paenibacillus macerans, were amplified by PCR by using degenerate primers and were characterized by DNA sequencing . We found that there are two nifH sequence clusters, designated clusters I and II, in P . azotofixans . The data further indicated that there was sequence divergence among the nifH genes of P . azotofixans strains at the DNA level . However, the gene products were more conserved at the protein level . Phylogenetic analysis showed that all nifH cluster II sequences were similar to the alternative (anf) nitrogenase sequence . A nested PCR assay for the detection of nifH (cluster I) of P . azotofixans was developed by using the degenerate primers as outer primers and two specific primers, designed on the basis of the sequence information obtained, as inner primers . The specificity of the inner primers was tested with several diazotrophic bacteria, and PCR revealed that these primers are specific for the P . azotofixans nifH gene . A GC clamp was attached to one inner primer, and a denaturing gradient gel electrophoresis (DGGE) protocol was developed to study the genetic diversity of this region of nifH in P . azotofixans strains, as well as in soil and rhizosphere samples . The results revealed sequence heterogeneity among different nifH genes . Moreover, nifH is probably a multicopy gene in P . azotofixans . Both similarities and differences were detected in the P . azotofixans nifH DGGE profiles generated with soil and rhizosphere DNAs . The DGGE assay developed here is reproducible and provides a rapid way to assess the intraspecific genetic diversity of an important functional gene in pure cultures, as well as in environmental samples. J Invertebr Pathol, 1998 Jul, 72(1), 21 - 7 Characterization of isolates of Paenibacillus larvae subsp . larvae from diverse geographical origin by the polymerase chain reaction and BOX primers; Alippi AM et al.; Ninety-nine strains of Paenibacillus larvae subsp . larvae, the causal agent of American Foulbrood disease (AFB) of honeybees, were isolated from different regions of Argentina and other countries . The isolates were characterized on the basis of DNA fingerprints by a polymerase chain reaction technique (PCR) with BOX sequence-specific primers . Isolates from Argentina generated three groups of patterns (designated A, B, and C), while P . larvae subsp . larvae strains obtained from other countries yielded two distinguishable patterns (coincident with A and B) . Strains from U.S . A . and Germany were identical and related to Group A, while all Czech and English isolates belonged to Group B . Strains from France, Poland, Italy, Sweden, and New Zealand showed two different patterns (A and B) . Comparisons of the biochemical type and genotype of isolates rendered no obvious linkage between both features . These results suggest that AFB in Argentina resulted from multiple sources of contaminated material . J Invertebr Pathol, 1998 Jan, 71(1), 73 - 81 Effect of Soil Texture and Soil Sterilization on Susceptibility of Ovipositing Grasshoppers to Beauveria bassiana Inglis GD, Johnson DL, Kawchuk LM, Goettel MS. The effect of conidial concentration, soil texture, and soil sterilization on the efficacy of Beauveria bassiana against ovipositing grasshoppers (Melanoplus sanguinipes) was investigated in a controlled environment . In the first experiment, mortality of female grasshoppers ovipositing into a sterile loamy-sand soil containing conidia of B . bassiana was measured . The prevalence of mortality increased as the concentration of conidia in soil increased, and a median lethal concentration of 10(4) colony-forming units (CFU) per gram of soil (dry weight) was observed . Conidia (10(2.9) to 10(3) CFU per abdomen) were recovered from the abdomens of grasshoppers ovipositing into sand containing 10(5.5) and 10(6) conidia per gram . Similar numbers of eggs were laid among treatments during the first oviposition period (1 to 7 days), but an effect of conidial concentration on eggs laid was observed during the second oviposition period (8 to 14 days) . This was attributed to reduction in female numbers and not to reduction in fecundity independent of mortality . In a second experiment, grasshoppers oviposited into soils of three different textures (loamy-sand, sandy-loam, or clay-loam) that were amended with 10(5) B . bassiana conidia per gram and possessed either a viable or heat-killed microflora . There was no effect of soil texture on mortality of ovipositing grasshoppers, on the number of eggs laid, on positioning of egg pods in the soil profile, or on numbers of B . bassiana CFU recovered from female abdomens . However, a higher prevalence of mortality was observed for females ovipositing into the sterilized than nonsterilized sandy-loam and clay-loam soils . Substantial populations of fungi and bacteria were recovered from nonsterilized soils . The predominant fungi isolated from these soils were members of the genera Chrysosporium, Fusarium, Gliocladium, Penicillium, Rhizopus, and Trichoderma, whereas Bacillus, Paenibacillus, and Pseudomonas species were the most commonly isolated bacteria . This study demonstrates that ovipositing grasshoppers are susceptible to relatively low densities of conidia in soils of varying textures, but the soil microflora may have an adverse affect on the efficacy of B . bassiana in field soils . J Vet Med Sci, 1997 Oct, 59(10), 953 - 4 Sporicidal activities of disinfectants on Paenibacillus larvae; Okayama A et al.; Sporicidal activities of glutaraldehyde, sodium hypochlorite, povidone iodine, ethylene oxide gas, chlorhexidine gluconate, and didecyl dimethylammonium chloride on wet and dry spores of Paenibacillus larvae (basonym: Bacillus larvae) were evaluated for control of honeybee American foulbrood . Glutaraldehyde was found to have a strong and rapid effect on both the wet and the dry spores among the disinfectants tested. J Vet Med Sci, 1997 Sep, 59(9), 765 - 7 Disposition of ampicillin in honeybees and hives; Nakajima C et al.; Disposition profile of ampicillin (ABPC) among honeybees, larvae, honey and royal jelly in a hive after oral dosing to adult bees was studied . Four honeybee colonies were administered the single dose of ABPC at the rate of 30 mg/hive by addition to sugar syrup or pollen substitute (paste) for 1 day intake . The colonies received ABPC in syrup showed high drug residue levels in honey and it lasted over 14 days beyond the detection limit of residual analysis . In the hives given ABPC in paste, relatively low honey residues were found, however, the distributions of the drug in young larvae and jelly which was the food of the larvae were very low . ABPC was considered to be a promising drug for the control of American foulbrood, an important bacterial disease of honeybee larvae, because of its high antibacterial activity to the pathogen, Paenibacillus larvae, and instability of residue in honey as human food . The low distribution in young larvae, the target of the disease, threw a doubt on the efficacy of ABPC for American foulbrood control. J Invertebr Pathol, 1997 Sep, 70(2), 79 - 87 The Proteases of American Foulbrood Scales Dancer BN, Chantawannakul P. The gross protease activity of pathological samples of American foulbrood-infected cadavers from several UK sources was studied . In all cases the bulk of the activity is caused by neutral protease(s) (optimum pH ca . 6.8) that are inhibited by chelating agents such as EDTA and 1,10 phenanthroline (indicating metalloproteases) but not by inhibitors of other classes of proteolytic enzymes . The proteases, which derive from the infectious agent of AFB, Paenibacillus larvae, were unusual in being insensitive to phosphoramidon and in not degrading FAGLA, the artificial substrate specific for most Bacillus metalloproteases . The enzymes in AFB ropes and scales had temperature optima of 60-65&deg;C and were inactivated quickly on incubation at 80&deg;C . Activity at moderate temperatures (37&deg;C) was great on general substrates such as casein, gelatin, and hide powder azure, slight on elastin-Congo red, and nonexistent on collagen . In SDS-polyacrylamide gels the enzymes from the various sources all had molecular weights about 24 kDa . The proteases could be detected only zymographically after brief washing to remove SDS . On silver-stained gels no bands corresponding to the enzymes' activities could be detected . On native polyacrylamide gels enzyme activity was resolved zymographically as at least three metalloprotease bands with samples from different sources showing a variety of patterns. Gene, 1997 Jul 18, 194(1), 25 - 33 Sequencing and phylogenetic analysis of the spoIIA operon from diverse Bacillus and Paenibacillus species; Park SG et al.; In order to clone the spoIIA operon from three different Bacillus and Paenibacillus species, we designed two sets of PCR primers based on three previously published Bacillus spoIIA sequences . One set of primers corresponded to the C-terminal region of SpoIIAB and a region near the middle of SpoIIAC . These primers were used to amplify the corresponding region of spoIIA from Bacillus stearothermophilus and Paenibacillus polymyxa (previously called Bacillus polymyxa {see Ash, C., Priest, F.G., Collins, M.D., 1993 . Molecular identification of ribosomal-RNA group 3 bacilli using a PCR probe test - proposal for the creation of a new genus Paenibacillus . Antonie van Leeuwenhoek Int . J . Gen . Mol . Microbiol . 64, 253-260} . The other set of primers, corresponding to an N-terminal and a C-terminal region of SpoIIAC, was used for B . sphaericus . The PCR products were used as probes for Southern blotting of homologous chromosomal DNA . DNA corresponding to spoIIA from the three organisms was identified by screening chromosomal DNA libraries, and cloned . Sequence analysis showed that all spoIIA sequences were conserved, but conservation was strongest in SpoIIAC and least strong in SpoIIAA . In the promoter the -35 region was conserved well but the -10 region rather poorly . Within the proteins, certain regions were particularly strongly conserved, suggesting that they are essential to the function of the protein . Phylogenetic analysis of spoIIA suggested that B . stearothermophilus is close to B . subtilis and B . licheniformis, but that P . polymyxa and B . sphaericus are remote from B . subtilis. Int J Syst Bacteriol, 1997 Apr, 47(2), 299 - 306 Emended description of Paenibacillus amylolyticus and description of Paenibacillus illinoisensis sp . nov . and Paenibacillus chibensis sp . nov; Shida O et al.; The taxonomic position of unidentified group 6 of Bacillus circulans as described by Nakamura and Swezey (L.K . Nakamura and J . Swezey, Int . J . Syst . Bacteriol . 33:46-52, 1983) was determined, and the taxonomy of Paenibacillus amylolyticus was reexamined . The results of PCR amplification of a 16S rRNA gene fragment with a specific primer and comparative analysis of 16S rRNA gene sequences warranted placing the two taxa in the genus Paenibacillus . The levels of DNA reassociation among the strains revealed four groups (designated groups I, II, III, and 6), each with a high level of intragroup relatedness (> 72%) . Clustering based on phenotypic characteristics correlated well with DNA relatedness grouping . P . amylolyticus strains were scattered in groups I, II, and III . Strains labeled the type strain of P . amylolyticus from different culture collections appeared in groups I and III . Strains found in group I were identified as P . amylolyticus sensu stricto, and the one strain found in group III was identified as Paenibacillus lautus . Group 6 encompassed strains formerly assigned to B . circulans group 6, and group II contained other strains identified as P . amylolyticus . Groups 6 and II were phenotypically and genetically distinct taxa that were distinguishable from the previously described species . These findings showed that groups 6 and II were new species, for which we propose the names Paenibacillus illinoisensis and Paenibacillus chibensis, respectively. Int J Syst Bacteriol, 1997 Apr, 47(2), 289 - 98 Transfer of Bacillus alginolyticus, Bacillus chondroitinus, Bacillus curdlanolyticus, Bacillus glucanolyticus, Bacillus kobensis, and Bacillus thiaminolyticus to the genus Paenibacillus and emended description of the genus Paenibacillus; Shida O et al.; We determined the taxonomic status of six Bacillus species (Bacillus alginolyticus, Bacillus chondroitinus, Bacillus curdlanolyticus, Bacillus glucanolyticus, Bacillus kobensis, and Bacillus thiaminolyticus) by using the results of 16S rRNA gene sequence and cellular fatty acid composition analyses . Phylogenetic analysis clustered these species closely with the Paenibacillus species . Like the Paenibacillus species, the six Bacillus species contained anteiso-C15:0 fatty acid as a major cellular fatty acid . The use of a specific PCR primer designed for differentiating the genus Paenibacillus from other members of the Bacillaceae showed that the six Bacillus species had the same amplified 16S rRNA gene fragment as members of the genus Paenibacillus . Based on these observations and other taxonomic characteristics, the six Bacillus species were transferred to the genus Paenibacillus . In addition, we propose emendation of the genus Paenibacillus. Antonie Van Leeuwenhoek, 1997 Mar, 71(3), 195 - 200 Evaluation of Biolog system for identification of strains of Paenibacillus azotofixans; Pires MN et al.; Biolog system was evaluated for the identification of strains of Paenibacillus azotofixans as no data concerning this species were in the list of Bacillus currently identified using Biolog data base . The P . azotofixans type strain P3L5 was first tested with the results recorded manually or using the automatic plate reader . In both cases, P3L5 utilized 22 carbon sources and when the results obtained were compared to data of Biolog software (Release 3.50), P3L5 was identified as B . azotoformans with a similarity coefficient of 0.913 (data recorded manually) and of 0.791 (data recorded automatically) . Metabolic profiles of P3L5 were also compared after readings of 4 and 24 h using the computer-driven automatic plate reader . No significant difference was observed in both cases and P3L5 was identified again as B . azotoformans with indices of similarity considered only for 'excellent identification' . Besides P3L5, other 15 P . azotofixans strains were tested with the Biolog system and all were identified as B . azotoformans with similarity coefficients varying from 0.511 to 0.927 . Phenotypic and genetic characteristics of B . azotoformans were compared to those described for P . azotofixans to explain the misidentification of the latter species . We could conclude that these two species are quite different and that data of Biolog software are from P . azotofixans and not from B . azotoformans. Rev Argent Microbiol, 1996 Oct-Dec, 28(4), 197 - 203 {Characterization of isolates of Paenibacillus larvae with biochemical type and oxytetracycline resistance}; Alippi AM; A collection of 91 isolates from different geographical origins of Paenibacillus larvae, the etiologic agent of American Foulbrood disease of honey bees, was characterized according to its biochemical type and susceptibility to oxytetracycline hydrochloride (OTC), the most commonly used antibiotic for the control of the disease . The majority of the Argentinian strains corresponded to the biochemical type II while only one culture from Rio Negro (Argentina), one from Buenos Aires (Argentina) and one from Cordoba (Argentina) presented characteristics of type V . In relation to their response to OTC it was found a 48% resistance within the collection of Argentinian strains; for this group, the values of minimal inhibitory concentration (MIC) were 10-15 micrograms/ml, while the susceptible ones presented MIC values under 5 micrograms/ml . All the isolates from France, Italy, New Zealand, Sweden, USA, Poland, Czech Republic and Germany were susceptible with MIC values under 5 micrograms/ml. J Appl Bacteriol, 1996 Oct, 81(4), 445 - 58 Bacterial contaminants in liquid packaging boards: assessment of potential for food spoilage; Pirttijarvi TS et al.; Liquid packaging boards and blanks were examined for microbial contaminants . A total of 218 strains were identified and representatives of the most frequent species were characterized for their potential for food spoilage . Contaminants found were aerobic spore-forming bacteria, mostly Bacillus megaterium, B . licheniformis, B . cereus group, B . pumilus, Paenibacillus macerans, P . polymyxa, P . pabuli and B . flexus . Production of amylolytic, proteolytic, lipolytic and phospholipolytic enzymes was common . Approximately 50% of the B . cereus group strains were positive in the diarrhoeal enterotoxin immunoassay test or in the enterotoxin reversed passive latex agglutination test . Strains capable of growth at 6 degrees C were found among B . cereus group, P . pabuli, P . validus, B . megaterium and P . polymyxa . All b . licheniformis strains grew at 55 degrees C . The spores of B . licheniformis were most resistant to hydrogen peroxide . The B . cereus group strains were recognizable by fatty acid components not present in any of the other paperboard strains, 11-methyldodecanoic acid (13:0 iso) and trans-9-hexadecenoic acid (16:1 omega 7 trans), each contributing 7% or more to the total cellular fatty acids. Int J Syst Bacteriol, 1996 Oct, 46(4), 988 - 1003 A polyphasic reassessment of the genus Paenibacillus, reclassification of Bacillus lautus (Nakamura 1984) as Paenibacillus lautus comb . nov . and of Bacillus peoriae (Montefusco et al . 1993) as Paenibacillus peoriae comb . nov., and emended descriptions of P . lautus and of P . peoriae; Heyndrickx M et al.; Seventy-seven strains representing 10 species in the Paenibacillus polymyxa 16S rRNA group and 3 other species that exhibit phenetic relatedness to members of this group, Bacillus lautus, "Bacillus longisporus," and Bacillus peoriae, were characterized genotypically and phenotypically by performing an amplified ribosomal DNA restriction analysis, a randomly amplified polymorphic DNA analysis, a fatty acid methyl ester analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of whole-cell proteins, pyrolysis mass spectrometry, and API and other routine phenotypic tests . These analyses revealed distinct clusters representing Paenibacillus alvei, Paenibacillus amylolyticus, Paenibacillus azotofixans, Paenibacillus durum, Paeniba