J Microbiol, 2004 Dec, 42(4), 346 - 52 Enzymatic and Non-enzymatic Degradation of Poly (3-Hydroxybutyrate-co-3-Hydroxyvalerate) Copolyesters Produced by Alcaligenes sp . MT-16; Choi GG et al.; Poly(3-hydroxybutyrate-co-3-hydroxyvalerate), poly(3HB-co-3HV), copolyesters with a variety of 3HV contents (ranging from 17 to 60 mol%) were produced by Alcaligenes sp . MT-16 grown on a medium containing glucose and levulinic acid in various ratios, and the effects of hydrophilicity and crystallinity on the degradability of the copolyesters were evaluated . Measurements of thermo-mechanical properties and Fourier-transform infrared spectroscopy in the attenuated total reflectance revealed that the hydrophilicity and crystallinity of poly(3HB-co-3HV) copolyesters decreased as 3HV content in the copolyester increased . When the prepared copolyester film samples were non-enzymatically hydrolysed in 0.01 N NaOH solution, the weights of all samples were found to have undergone no changes over a period of 20 weeks . In contrast, the copolyester film samples were degraded by the action of extracellular polyhydroxybutyrate depolymerase from Emericellopsis minima W2 . The overall rate of weight loss was higher in the films containing higher amounts of 3HV, suggesting that the enzymatic degradation of the copolyester is more dependent on the crystallinity of the copolyester than on its hydrophilicity . Our results suggest that the degradability characteristics of poly(3HB-co-3HV) copolyesters, as well as their thermo-mechanical properties, are greatly influenced by the 3HV content in the copolyesters. J Synchrotron Radiat, 2005 Jan 1, 12(Pt 1), 13 - 8 Epub 2004 Dec 23. Heterologous metalloprotein biosynthesis in Escherichia coli: conditions for the overproduction of functional copper-containing nitrite reductase and azurin from Alcaligenes xylosoxidans; Harris RL; This paper reports on the optimization of conditions for the overproduction and isolation of two recombinant copper metalloproteins, originally encoded on the chromosome of the dentrifying soil bacterium Alcaligenes xylosoxidans, in the heterologous host Escherichia coli . The trimeric enzyme nitrite reductase (NiR) contains both type-1 and type-2 Cu centres, whilst its putative redox partner, azurin I, is monomeric and has only a type-1 Cu centre . Both proteins were processed and exported to the periplasm of E . coli, which is consistent with their periplasmic location in their native host A . xylosoxidans . NiR could be readily purified from the periplasmic fraction of E . coli but the enzyme as isolated possessed only type-1 Cu centres . The type-2 Cu centre could be fully reconstituted by incubation of the periplasmic fraction with copper sulfate prior to enzyme purification . Azurin I could only be isolated with a fully occupied type-1 centre when isolated from the crude cell extract but not after isolation from the periplasmic fraction, suggesting loss of the copper due to proteolysis . Based on a number of criteria, including spectroscopic, mass spectrometric, biochemical and structural analyses, both recombinant proteins were found to be indistinguishable from their native counterparts isolated from A . xylosoxidans . The findings of this work have important implications for the overproduction of recombinant metalloproteins in heterologous hosts. Biochem Biophys Res Commun, 2005 Jan 7, 326(1), 74 - 8 Site-directed mutagenesis alters DnaK-dependent folding process; Yoshimune K et al.; The overproduction of d-aminoacylase (A6-d-ANase) of Alcaligenes xylosoxydans subsp . xylosoxydans A-6 (Alcaligenes A-6) is accompanied by aggregation of the overproduced protein, and its soluble expression is facilitated by the coexpression of DnaK-DnaJ-GrpE (DnaKJE) . When the A6-d-ANase gene was expressed in the Escherichia coli dnaK mutant dnaK756, little activity was observed in the soluble fraction, and it was restored by the coexpression of DnaKJE or the substitution of the R354 residue of A6-d-ANase for lysine . These results suggest that the guanidino group of the R354 residue of A6-d-ANase disturbs its proper folding in the absence of DnaK and the disturbance is eliminated by binding of DnaK to the R354 residue in the presence of DnaK . This is the first report that the DnaK-dependent folding process of the enzyme is altered by site-directed mutagenesis. FEMS Microbiol Lett, 2004 Oct 15, 239(2), 285 - 93 Cloning and functional analysis by gene disruption of a novel gene involved in indigo production and fluoranthene metabolism in Pseudomonas alcaligenes PA-10; Alemayehu D et al.; A novel indole dioxygenase (idoA) gene has been cloned from Pseudomonas alcaligenes PA-10, based on its ability to convert indole to indigo . The chromosomally encoded idoA gene exhibits no similarity to previously cloned naphthalene dioxygenases or to aromatic oxygenases from other species at the nucleotide level . Phylogenetic analysis indicates that the idoA gene product is most similar to an acyl-CoA dehydrogenase from Novosphingobium aromaticivorans . The enzyme encoded by the idoA gene is essential for the metabolism of fluoranthene, since a mutant in which the idoA gene has been disrupted looses the ability to degrade this compound . The idoA gene appears to be constitutively expressed in PA-10, but its expression is also subject to regulation following prior exposure to salicylate and to fluoranthene degradative intermediates. Can J Microbiol, 2004 Aug, 50(8), 633 - 44 A survey of the composition and diversity of bacterial populations in bleached kraft pulp-mill wastewater secondary treatment systems; Gilbride KA et al.; Bacterial community compositions from 10 pulp- and paper-mill treatment systems were compared using both traditional and molecular techniques . 16S-RFLP (Random Fragment Length Polymorphisms) analysis was used to examine the genotypic profiles of the whole bacterial community of each treatment system . Although all the communities shared approximately 60% of their DNA band pattern, as determined by computer-assisted cluster analysis, each community displayed a unique profile that was stable over time under normal operating parameters . Reverse Sample Genome Probing (RSGP) and 16S-RFLP were used to compare the culturable bacterial communities of several geographically separated pulp-mill biotreatment system communities . There was little overlap in the composition of the culturable community between mills at the genus level . Furthermore, RSGP variation was almost as high within a mill as between mills . Partial sequences of the 16S rRNA genes from culturable isolates identified Bacillus spp., Pseudomonas spp., and Xanthobacter as some of the dominant species . Finally, several 16S rRNA genes from two whole community 16S RNA gene libraries were partially sequenced and identified as similar to unknown alpha-, beta-, and gamma-Proteobacteria, Ralstonia, Alcaligenes, Nitrospira, Firmicutes, and clones representing the new Holophaga/Acidobacterium phylum . These findings suggest that although these pulp- and paper-mill biotreatment communities perform similar functions, they are populated by unique mixtures of species. J Cyst Fibros, 2004 Aug, 3(3), 151 - 7 Antibiograms of resistant Gram-negative bacteria from Scottish CF patients; Mackenzie FM et al.; Background: Over a 19-month pilot phase, 93 multiply resistant Gram-negative isolates from Scottish cystic fibrosis patients were sent to a referral laboratory for further investigation . Methods: In common with the referring diagnostic laboratories, disc diffusion testing was carried out . Antibiotic susceptibility testing was also established by MIC methodology . NCCLS methods were used throughout . Twenty antibiotics were tested . Results: Comparing disc diffusion results against MIC results, there were 167 (14%) major errors . By MIC, Pseudomonas aeruginosa (n=59), Stenotrophomonas maltophilia (n=16), Burkholderia cepacia (n=10) and Alcaligenes xylosoxidans (n=7) were susceptible to 18%, 11%, 4% and 35% of the antibiotics tested, respectively . Colistin and tobramycin were the most active agents against P . aeruginosa with 60% and 49%, respectively, testing susceptible . Minocycline and gentamicin were most active against S . maltophilia with 58% and 18%, respectively, testing susceptible . B . cepacia were most susceptible to co-trimoxazole (10%) and ciprofloxacin (10%) . Five and six of the seven A . xylosoxidans isolates were susceptible to piperacillin and imipenem, respectively . Conclusions: Improved methods for susceptibility testing of such clinical isolates need to be employed in routine diagnostic laboratories . Levels of resistance in referred isolates were very high and similar to those described in the USA. Biosci Biotechnol Biochem, 2004 Sep, 68(9), 1921 - 8 Purification and characterization of aromatic amine dehydrogenase from Alcaligenes xylosoxidans; Kondo T et al.; Aromatic amine dehydrogenase was purified and characterized from Alcaligenes xylosoxidans IFO13495 grown on beta-phenylethylamine . The molecular mass of the enzyme was 95.5 kDa . The enzyme consisted of heterotetrameric subunits (alpha2beta2) with two different molecular masses of 42.3 kDa and 15.2 kDa . The N-terminal amino acid sequences of the alpha-subunit (42.3-kDa subunit) and the beta-subunit (15.2-kDa subunit) were DLPIEELXGGTRLPP and APAAGNKXPQMDDTA respectively . The enzyme had a quinone cofactor in the beta-subunit and showed a typical absorption spectrum of tryptophan tryptophylquinone-containing quinoprotein showing maxima at 435 nm in the oxidized form and 330 nm in the reduced form . The pH optima of the enzyme activity for histamine, tyramine, and beta-phenylethylamine were the same at 8.0 . The enzyme retained full activity after incubation at 70 degrees C for 40 min . It readily oxidized various aromatic amines as well as some aliphatic amines . The Michaelis constants for phenazine methosulfate, beta-phenylethylamine, tyramine, and histamine were 48.1, 1.8, 6.9, and 171 microM respectively . The enzyme activity was strongly inhibited by carbonyl reagents . The enzyme could be stored without appreciable loss of enzyme activity at 4 degrees C for one month at least in phosphate buffer (pH 7.0). Indian J Exp Biol, 2003 Dec, 41(12), 1469 - 72 Biological control of Fusarium wilt of pigeonpea Cajanus cajan (L.) Millsp with chitinolytic Alcaligenes xylosoxydans; Vaidya RJ et al.; Alcaligenes xylosoxydans protected pigeonpea from Fusarium wilt in a pot experiment and field trials . When seeds of pigeonpea (C . cajan) were treated with A . xylosoxydans and sown in soil infested with Fusarium, the incidence of wilt was reduced by 43.5% and resulted in 58% higher grain yield . The antifungal activity of A . xylosoxydans was based on chitinase production and was comparable in efficacy to commercial antifungal agents such as benlate, monitor WP, thiram and bavistin. Appl Biochem Biotechnol, 2004 Jul-Sep, 118(1-3), 65 - 72 Treatment of dairy wastewater using a selected bacterial isolate, Alcaligenes sp . MMRR7; Rajeshkumar K et al.; Physicochemical and biologic analysis of dairy wastewater showed that the effluent had a high organic load (chemical oxygen demand {COD}: 5095 mg/L), an acidic pH (6.4), and a high probability of coliforms (most probable number {MPN} >1100) . The various bacterial strains isolated and purified were identified as Sporolactobacillus sp., Citrobacter sp., Pseudomonas sp., Alcaligenes sp., Bacillus sp., Staphylococcus sp., and Proteus sp., as per the Bergey's manual of systematic bacteriology . Among the five selected bacterial strains, the strain designated as MMRR7 and identified as Alcaligenes sp . was found to give a maximum reduction in COD (62%) in 5 d of incubation . Chemical coagulation using alum at a concentration of 0.5 g/100 mL was found to be effective in the primary treatment of the effluent . Studies on free-cell treatment of the coagulated effluent with the selected bacterial strain Alcaligenes sp . MMRR7 gave a maximum COD reduction of 91% in 120 h . This study clearly indicates the possibility of using Alcaligenes sp . MMRR7 for the effective treatment of dairy wastewater. Appl Biochem Biotechnol, 1999 Spring, 77-79, 445 - 54 Conversion of industrial food wastes by alcaligenes latus into polyhydroxyalkanoates; Yu PH et al.; Broader usage of biodegradable plastics in packaging and disposable products as a solution to environmental problems would heavily depend on further reduction of costs and the discovery of novel biodegradable plastics with improved properties . As the first step in our pursuit of eventual usage of industrial food wastewater as nutrients for microorganisms to synthesise environmental-friendly bioplastics, we investigated the usage of soya wastes from a soya milk dairy, and malt wastes from a beer brewery plant as the carbon sources for the production of polyhydroxyalkanoates (PHA) by selected strain of microorganism . Bench experiments showed that Alcaligenes latus DSM 1124 used the nutrients from malt and soya wastes to biosynthesise PHAs . The final dried cell mass and specific polymer production of A . latus DSM 1124 were 32g/L and 70% polymer/cells (g/g), 18.42 g/L and 32.57% polymer/cell (g/g), and 28 g/L and 36% polymer/cells (g/g), from malt waste, soya waste, and from sucrose, respectively . These results suggest that many types of food wastes might be used as the carbon source for the production of PHA. Acta Crystallogr D Biol Crystallogr, 1997 Jul, 53(Pt 4), 406 - 18 Structures of a blue-copper nitrite reductase and its substrate-bound complex; Dodd FE; Copper-containing nitrite reductases (NiR's) have been conveniently subdivided into blue and green NiR's which are thought to be redox partners of azurins and pseudo-azurins, respectively . Crystal structures of two green NiR's have recently been determined . Alcaligenes xylosoxidans has been shown to have a blue-copper nitrite reductase (AxNiR) and two azurins with 67% homology both of which donate electrons to it effectively . The first crystal structure of a blue NiR (AxNiR) in its oxidized and nitrite-bound forms, with particular emphasis to the Cu sites, is presented . The Cu-Smet distance is the same as those in the green NiR's . Thus, the length of this interaction is unlikely to be responsible for differences in colour . Crystallographic data presented here taken together with structural data of other single Cu type-1 proteins and their mutants suggest that the displacement of Cu from the strong ligand plane is perhaps the cause for the differences in colour observed for otherwise 'classical' blue Cu centre . Nitrite is observed binding to the catalytic Cu in a bidentate fashion displacing the water molecule, offering a neat rationalization for the XAFS observation that the type-2 Cu-ligand distances increase on nitrite binding as a result of increased coordination . These results are discussed in terms of enzyme mechanism. Acta Crystallogr D Biol Crystallogr, 1996 Jul, 52(Pt 4), 858 - 63 A re-evaluation of the crystal structure of chloromuconate cycloisomerase; Kleywegt GJ; It is shown here that the reported 3 A crystal structure of chloromuconate cycloisomerase from Alcaligenes eutrophus {Hoier, Schlomann, Hammer, Glusker, Carrell, Goldman, Stezowski & Heinemann (1994) . Acta Cryst . D50, 75-84} was refined in the incorrect space group I4 . In addition, a stretch of about 25 residues near the N-terminus is out-of-register with the density in the original structure . From the coordinates and structure factors deposited in the Protein Data Bank (PDB), it was possible to determine the correct space group to be I422 . The structure was then re-refined, using the original data reduced to I422, to a crystallographic free R factor of 0.264 at 3 A resolution (conventional R factor 0.189) . With conservative refinement and rebuilding methods, the errors in the chain tracing could be identified and remedied . Since the two molecules per asymmetric unit in the original structure are actually related by crystallographic symmetry, the observed differences between them are artefacts . In particular, the differences between, and peculiarities of the metal-binding sites are unreal . This case shows the dangers of crystallographic refinement in cases with unfavourable data-to-parameter ratios, and the importance of reducing the number of parameters in such cases to prevent gross errors (for instance, by using NCS constraints) . It also demonstrates how the evaluation and monitoring of model quality during the entire refinement and rebuilding process can be used to detect and remedy serious errors . Finally, it presents a strong case in favour of depositing not only model coordinates, but also experimental data (preferably, both merged and unmerged data). Acta Crystallogr D Biol Crystallogr, 1996 Sep, 52(Pt 5), 937 - 41 Direct-Method Structure Determination of the Native Azurin II Protein Using One-Wavelength Anomalous Scattering Data; Xiao-Feng Z; The one-wavelength anomalous scattering (OAS) X-ray diffraction data of azurin II, a copper-containing protein from Alcaligenes xylosoxidans were collected at the Photon Factory, Japan at a 'routine' wavelength of 0.97 A . The structure had been originally solved by the molecular-replacement method {Dodd, Hasnain, Abraham, Eady & Smith (1995) . Acta Cryst . D51, 1052-1064} . As a technique of ab initio structure determination, the direct method {Fan, Hao, Gu, Qian, Zheng & Ke (1990) . Acta Cryst . A46, 935-939} was attempted to break the phase ambiguity intrinsic to OAS data . The phases were then improved using the solvent-flattening method . The final electron-density map clearly shows most Calpha positions and many side chains and it is traceable without prior knowledge of the structure . It is concluded that the direct method is capable of phasing anomalous scattering data collected at one wavelength from moderate-sized native proteins (M(w) approximately 20 kDa) which contain copper or atoms with a similar scattering power. Acta Crystallogr D Biol Crystallogr, 1994 Jan, 50(Pt 1), 75 - 84 Crystal structure of chloromuconate cycloisomerase from Alcaligenes eutrophus JMP134 (pJP4) at 3 A resolution; Hoier H; Chloromuconate cycloisomerase (E.C . 5.5.1.7) is an enzyme involved in the 2,4-dichlorophenoxyacetate degradation pathway of Alcaligenes eutrophus JMP134 (pJP4) . The crystal structure of this protein was determined at 3 A resolution by molecular-replacement techniques using atomic coordinates from the reported crystal structure of the homologous muconate cycloisomerase (E.C . 5.5.1.1) from Pseudomonas putida as the search model (42% identical positions in the sequences) . Structure refinement by simulated-annealing and restrained least-squares techniques converged at R = 0.195 . In the crystals studied, space group I4, the protein is present as two octamers per unit cell with two subunits per asymmetric unit . Each subunit consists of two globular domains, one of which forms an alpha/beta-barrel . Comparison of this structure with that of muconate cycloisomerase reveals the reasons for the altered substrate specificity of chloromuconate cycloisomerase . Marked differences are observed in polarity, accessibility and hydrogen-bonding potential in the channel leading into the active site as well as in the active center itself. Rev Port Pneumol, 2003 Nov, IX(5 Suppl 1), 35 - 36 {EPIDEMIOLOGICAL SURVEY OF BACTERIA ISOLATED FROM THE RESPIRATORY TRACT OF CYSTIC FIBROSIS PATIENTS}; Quintas S et al.; With the aim of characterizing the evolution of the epidemiological profile of respiratory bacterial infections of patients having Cystic Fibrosis (CF), the authors conducted a retrospective analysis about it's incidence and prevalence in 78 CF patients followed at the CF Specialized Centre, Paediatric Department, Santa Maria Hospital, Lisbon, during a 5 years period (1995-1999) . Pseudomonas aeruginosa was the most frequently isolated bacteria during the first three years of the study (60-73%), being surpassed by Staphylococcus aureus . However, Pseudomonas aeruginosa always remained the principal agent of chronic colonization (44-59%), with a peak of beginning between 0 and 5 years (34%) . A significative increase of the prevalence of intermitent and chronic colonization with Staphylococcus aureus was verified during this five years (48% to 83% and 32% to 54%) . The prevalence of isolations of Meticillin Resistant Staphylococcus aureus and of Burkholderia cepacia almost duplicated during this period . The taxes of isolation and chronic colonization with Alcaligenes xylosoxidans raised sharply beyond 1997 (from 3% and 0% in 1996 to 7% and 5% in 1997 and 10% and 7% in 1999) . Chronic colonization with Haemophilus influenzae kept a median prevalence of 22%, in spite of an increase in isolations (from 42% to 61%) . Fifty five per cent of the patients were chronically colonized by two or more agents . In view of these results, the authors discuss the therapeutic schemes and the measures to limit cross-infection which have been being advocated for CF patients in our centre. Biochemistry, 2004 Jul 13, 43(27), 8670 - 9 Crystal structure of 4-chlorobenzoate:CoA ligase/synthetase in the unliganded and aryl substrate-bound states; Gulick AM et al.; 4-Chlorobenzoate:CoA ligase (CBAL) is a member of a family of adenylate-forming enzymes that catalyze two-step adenylation and thioester-forming reactions . In previous studies, we have provided structural evidence that members of this enzyme family (exemplified by acetyl-CoA synthetase) use a large domain rotation to catalyze the respective partial reactions {A . M . Gulick, V . J . Starai, A . R . Horswill, K . M . Homick, and J . C . Escalante-Semerena, (2003) Biochemistry 42, 2866-2873} . CBAL catalyzes the synthesis of 4-chlorobenzoyl-CoA, the first step in the 4-chlorobenzoate degredation pathway in PCB-degrading bacteria . We have solved the 2.0 A crystal structure of the CBAL enzyme from Alcaligenes sp . AL3007 using multiwavelength anomalous dispersion . The results demonstrate that in the absence of any ligands, or bound to the aryl substrate 4-chlorobenzoate, the enzyme adopts the conformation poised for catalysis of the adenylate-forming half-reaction . We hypothesize that coenzyme A binding is required for stabilization of the alternate conformation, which catalyzes the 4-CBA-CoA thioester-forming reaction . We have also determined the structure of the enzyme bound to the aryl substrate 4-chlorobenzoate . The aryl binding pocket is composed of Phe184, His207, Val208, Val209, Phe249, Ala280, Ile303, Gly305, Met310, and Asn311 . The structure of the 4-chlorobenzoate binding site is discussed in the context of the binding sites of other family members to gain insight into substrate specificity and evolution of new function. Biodegradation, 2004 Jun, 15(3), 205 - 12 Isolation and characterization of polycyclic aromatic hydrocarbons-degrading Sphingomonas sp . strain ZL5; Liu Y et al.; A bacterial strain ZL5, capable of growing on phenanthrene as a sole carbon and energy source but not naphthalene, was isolated by selective enrichment from crude-oil-contaminated soil of Liaohe Oil Field in China . The isolate was identified as a Sphingomonas sp . strain on the basis of 16S ribosomal DNA analysis . Strain ZL5 grown on phenanthrene exhibited catechol 2,3-dioxygenase (C23O) activity but no catechol 1,2-dioxygenase, gentisate 1,2-dioxygenase, protocatechuate 3,4-dioxygenase and protocatechuate 4,5-dioxygenase activities . This suggests that the mode of cleavage of phenanthrene by strain ZL5 could be meta via the intermediate catechol, which is different from the protocatechuate way of other two bacteria, Alcaligenes faecelis AFK2 and Nocardioides sp . strain KP7, also capable of growing on phenanthrene but not naphthalene . A resident plasmid (approximately 60 kb in size), designated as pZL, was detected from strain ZL5 . Curing the plasmid with mitomycin C and transferring the plasmid to E . coli revealed that pZL was responsible for polycyclic aromatic hydrocarbons degradation . The C23O gene located on plasmid pZL was cloned and overexpressed in E . coli JM109(DE3) . The ring-fission activity of the purified C23O from the recombinant E . coli on dihydroxylated aromatics was in order of catechol > 4-methylcatechol > 3-methylcatechol > 4-chlorocatechol >> 3,4-dihydroxyphenanthrene > 3-chlorocatechol. Proteomics, 2004 Jul, 4(7), 2028 - 36 Proteome analysis of gentisate-induced response in Pseudomonas alcaligenes NCIB 9867; Zhao B et al.; Pseudomonas alcaligenes NCIB 9867 (P25X wild-type) is capable of degrading aromatic hydrocarbons via the gentisate pathway . Biochemical characterization of P25X mutants indicated that it has isofunctional enzymes for the mono- and dioxygenase-catalyzed reactions . One set of the enzymes is constitutive whereas the other is strictly inducible . To date, only the gene encoding the constitutively-expressed gentisate dioxygenase had been cloned and characterized . A mutant strain of P25X, designated G56, which had the constitutive copy of the gentisate 1,2-dioxygenase gene interrupted by a streptomycin/spectinomycin resistance gene cassette, was found to express gentisate dioxygenase, but only when the cells were induced by gentisate . The proteome profiles of P . alcaligenes P25X and mutant G56 cells grown in the presence and absence of gentisate were compared after two-dimensional polyacrylamide gel electrophoresis . Eight distinctive protein spots (designated M1-M8) which were observed only in induced cells of strain G56 but absent in noninduced cells were further analyzed by matrix-assisted laser desorption/ionization-time of flight, quadrupole-TOF and N-terminal sequencing . Of the 15 proteins (including seven up-regulated) examined, 13 showed sequence similarities to proteins with assigned functions in other microorganisms . The identification of protein M5 which showed high homology to a gentisate dioxygenase from Ralstonia sp . U2 indicated the putative function of this protein being consistent with the inducible gentisate 1,2-dioxygenase in P . alcaligenes . In addition, the induction of stress proteins and other adaptation phenomena were also observed. J Biol Chem, 2004 Sep 3, 279(36), 37250 - 60 Epub 2004 Jun 25. Biochemical and molecular characterization of a ring fission dioxygenase with the ability to oxidize (substituted) salicylate(s) from Pseudaminobacter salicylatoxidans; Hintner JP et al.; The gene coding for a dioxygenase with the ability to cleave salicylate by a direct ring fission mechanism to 2-oxohepta-3,5-dienedioic acid was cloned from Pseudaminobacter salicylatoxidans strain BN12 . The deduced amino acid sequence encoded a protein with a molecular mass of 41,176 Da, which showed 28 and 31% sequence identity, respectively, to a gentisate 1,2-dioxygenase from Pseudomonas alcaligenes NCIMB 9867 and a 1-hydroxy-2-naphthoate 1,2-dioxygenase from Nocardioides sp . KP7 . The highest degree of sequence identity (58%) was found to a presumed gentisate 1,2-dioxygenase from Corynebacterium glutamicum . The enzyme from P . salicylatoxidans BN12 was heterologously expressed in Escherichia coli and purified as a His-tagged enzyme variant . The purified enzyme oxidized in addition to salicylate, gentisate, 5-aminosalicylate, and 1-hydroxy-2-naphthoate also 3-amino- and 3- and 4-hydroxysalicylate, 5-fluorosalicylate, 3-, 4-, and 5-chlorosalicylate, 3-, 4-, and 5-bromosalicylate, 3-, 4-, and 5-methylsalicylate, and 3,5-dichlorosalicylate . The reactions were analyzed by high pressure liquid chromatography/mass spectrometry, and the reaction products were tentatively identified . For comparison, the putative gentisate 1,2-dioxygenase from C . glutamicum was functionally expressed in E . coli and shown to convert gentisate but not salicylate or 1-hydroxy-2-naphthoate. Biosci Biotechnol Biochem, 2004 Jun, 68(6), 1353 - 6 Total degradation of pentachloroethane by an engineered Alcaligenes strain expressing a modified camphor monooxygenase and a hybrid dioxygenase; Iwakiri R et al.; We engineered biphenyl-degrading Alcaligenes sp . strain KF711 for total degradation of pentachloroethane (PCA), which expresses a modified camphor monooxygenase and a hybrid dioxygenase consisting of TodC1 (a large subunit of toluene dioxygenase of Pseudomonas putida F1) and BphA2-BphA3-pbhA4 (a small subunit, ferredoxin and ferredoxin reductase of biphenyl dioxygenase, respectively, in strain KF707) . Modified camphor monooxygenase genes (camCAB) were supplied as a plasmid and the todC1 gene was integrated within the chromosomal bph gene cluster by a single crossover recombination . The resultant strain KF711S-3cam dechlorinated PCA to trichloroethene by the action of the modified camphor monooxygenase under anaerobic conditions . The same strain subsequently degraded trichloroethene formed oxidatively by the action of the Tol-Bph hybrid dioxygenase under aerobic conditions . Thus sequential anaerobic and aerobic treatments of the KF711S-3cam resting cells resulted in efficient and total degradation of PCA. Biotechnol Lett, 2004 Apr, 26(8), 617 - 22 Transcription level of granule-associated phaP and phaR genes and granular morphogenesis of poly-beta-hydroxyalkanoate granules in Ralstonia eutropha; Seo MC et al.; The transcription levels of the granule-associated phaP and phaR genes in Ralstonia eutropha were regulated through the transformation of the phbC genes from R . eutropha and Alcaligenes latus into the poly-beta-hydroxybutyrate synthase-negative mutant . The granular morphogenesis of short chain length, poly-beta-hydroxyalkanoate (scl-PHA) was closely associated with the mRNA transcription levels of the phaP and phaR genes, especially with the ratio of phaP/phaR genes . The phasin protein encoded by the phaP gene increased the number of granules, while the PhaR protein of the phaR gene enlarged the size of the scl-PHA granules in R . eutropha. Rev Argent Microbiol, 2004 Jan-Mar, 36(1), 47 - 51 Microbial dehalogenation of polychlorinated biphenyls in aerobic conditions; Araoz B et al.; From soils contaminated with polychlorinated biphenyls (PCBs) a strain of Alcaligenes sp . able to grow in a mineral medium with a commercial mixture of PCBs as carbon source was isolated . This strain consumed up to 200 ppm in seven days in laboratory conditions . After 24 h of incubation, some new congeners of PCBs could be recognized by mass spectrometry . Through the identification of these compounds it was possible to postulate examples of possible transformations by dechlorinations of penta- and tetra-chlorinated congeners into tri-chlorinated ones . The properties of the isolated strain are appropriate for bioremediation of contaminated areas and also for using in bioreactors in order to remove the xenobiotic chemical. Pneumologie, 2004 May, 58(5), 309 - 15 {Isolation measurements for cystic fibrosis patients}; Vonberg R et al.; BACKGROUND: Respiratory tract infections significantly contribute to morbidity and mortality of cystic fibrosis patients . METHODS: We conducted a systematic literature review (Pubmed 01/1966 up to 09/2003) in order to present recommendations for the isolation of CF patients colonized with Burkholderia cepacia spp., Pseudomonas aeruginosa, Stenotrophomonas maltophilia and Alcaligenes spp . Evidence and quality of 64 publications dealing with pathogen transmission or isolation measurements of colonized patients were evaluated . RESULTS: B . cepacia spp . was dealt most often with and 35 of 36 authors recommended the isolation of patients colonized with this pathogen . Isolation of patients colonized with P . aeruginosa was proposed by 21 of 25 authors . Only 5 studies concerned S . maltophilia or Alcaligenes spp . CONCLUSIONS: A) B . cepacia spp . colonized patients need to get a single room for their own . B) P . aeruginosa colonized CF patients should be separated from non-colonized CF patients . C) Patients harbouring even multi drug resistant P . aeruginosa, S . maltophilia or Alcaligenes spp . may not share their room with immunocompromised patients and should also be isolated when treated in intensive care units. J Ind Microbiol Biotechnol, 2004 Mar, 31(3), 137 - 47 Epub 2004 Apr 27. Antibacterial effects of knotwood extractives on paper mill bacteria; Lindberg LE et al.; Hydrophilic knotwood extracts from 18 wood species were assessed in disc diffusion and liquid culture tests for antibacterial effects against three species of paper mill bacteria . The Pinus sylvestris, P . resinosa, P . contorta, and P . banksiana extracts decreased or inhibited bacterial growth . The susceptibility order was P . sylvestris > P . resinosa > P . contorta > P . banksiana, correlating with the concentrations of pinosylvin and pinosylvin monomethyl ether in these wood species . Also, Pseudotsuga menziesii and Thuja occidentalis extracts had a small inhibitory effect . The Gram-positive Bacillus coagulans was more susceptible to the extracts than the Gram-negative Burkholderia multivorans and Alcaligenes xylosoxydans . The main components in the Pinus knotwood extracts were pinosylvin monomethyl ether and pinosylvin, suggesting these to be the active components . Therefore, pure pinosylvin, pinosylvin monomethyl ether, and dihydro-pinosylvin monomethyl ether were also tested . All compounds showed antibacterial effects . However, higher concentrations were needed for these pure compounds than for the knotwood extracts . Pinosylvin had stronger antibacterial effects than pinosylvin monomethyl ether . This work shows that knotwood extracts, especially from Pinus species, have a potential for use as natural biocides in papermaking. Environ Pollut, 1993, 81(1), 47 - 50 Evaluation of Acacia nilotica (L.) del . and Casuarina equisetifolia forst . for tolerance and growth on microbially treated dyestuff wastewater; Kanekar P et al.; Two tree species, Acacia nilotica (L.) Del . and Casuarina equisetifolia Forst., were tested for their tolerance and growth on dyestuff wastewater containing phenol, aniline, and methyl violet . The wastewater was treated microbially by using a culture of Pseudomonas alcaligenes in a fixed-film bioreactor . The plants were watered with untreated and treated wastewater and tap water (control) at the rate of 2 litre per plant per week . A . nilotica exhibited 100% survival with both untreated and treated wastewater, and growth (increase in height) was comparable to that of control plants . However, growth of C . equisetifolia was adversely affected . It exhibited 100% survival with treated wastewater but only 87% survival with untreated waste-water and the percentage increase in height was less with both treated and untreated wastewater . There was no effect on soil except for an increase in chloride content. Extremophiles, 2004 Feb, 8(1), 45 - 51 Epub 2003 Oct 01. Bacterial diversity of the Inner Mongolian Baer Soda Lake as revealed by 16S rRNA gene sequence analyses; Ma Y et al.; Bacterial diversity associated with Baer Soda Lake in Inner Mongolia of China was investigated using a culture-independent method . Bacterial 16S rRNA gene libraries were generated using bacterial oligonucleotide primers, and 16S rRNA gene sequences of 58 clones were analyzed phylogenetically . The library was dominated by 16S rDNAs of Gram-negative bacteria (24% alpha-Proteobacteria, 31% beta-Proteobacteria, 33% gamma-Proteobacteria, and 2% delta-Proteobacteria), with a lower percentage of clones corresponding to Gram-positive bacteria . Forty cloned sequences were similar to that of known bacterial isolates (> 97% sequence similarity), represented by the species of the genera Brevundimonas, Comamonas, Alcaligenes, Stenotrophomonas, and Klebsiella . Eighteen cloned sequences showed less affiliation with known taxa (< 97% sequence similarity) and may represent novel taxa. J Bacteriol, 2004 Apr, 186(8), 2328 - 39 A new amidohydrolase from Bordetella or Alcaligenes strain FB188 with similarities to histone deacetylases; Hildmann C et al.; The full-length gene encoding the histone deacetylase (HDAC)-like amidohydrolase (HDAH) from Bordetella or Alcaligenes (Bordetella/Alcaligenes) strain FB188 (DSM 11172) was cloned using degenerate primer PCR combined with inverse-PCR techniques and ultimately expressed in Escherichia coli . The expressed enzyme was biochemically characterized and found to be similar to the native enzyme for all properties examined . Nucleotide sequence analysis revealed an open reading frame of 1,110 bp which encodes a polypeptide with a theoretical molecular mass of 39 kDa . Interestingly, peptide sequencing disclosed that the N-terminal methionine is lacking in the mature wild-type enzyme, presumably due to the action of methionyl aminopeptidase . Sequence database searches suggest that the new amidohydrolase belongs to the HDAC superfamily, with the closest homologs being found in the subfamily assigned acetylpolyamine amidohydrolases (APAH) . The APAH subfamily comprises enzymes or putative enzymes from such diverse microorganisms as Pseudomonas aeruginosa, Archaeoglobus fulgidus, and the actinomycete Mycoplana ramosa (formerly M . bullata) . The FB188 HDAH, however, is only moderately active in catalyzing the deacetylation of acetylpolyamines . In fact, FB188 HDAH exhibits significant activity in standard HDAC assays and is inhibited by known HDAC inhibitors such as trichostatin A and suberoylanilide hydroxamic acid (SAHA) . Several lines of evidence indicate that the FB188 HDAH is very similar to class 1 and 2 HDACs and contains a Zn(2+) ion in the active site which contributes significantly to catalytic activity . Initial biotechnological applications demonstrated the extensive substrate spectrum and broad optimum pH range to be excellent criteria for using the new HDAH from Bordetella/Alcaligenes strain FB188 as a biocatalyst in technical biotransformations, e.g., within the scope of human immunodeficiency virus reverse transcriptase inhibitor synthesis. FEBS Lett, 2004 Mar 12, 561(1-3), 173 - 6 Met144Ala mutation of the copper-containing nitrite reductase from Alcaligenes xylosoxidans reverses the intramolecular electron transfer; Farver O et al.; Pulse radiolysis has been employed to investigate the intramolecular electron transfer (ET) between the type 1 (T1) and type 2 (T2) copper sites in the Met144Ala Alcaligenes xylosoxidans nitrite reductase (AxCuNiR) mutant . This mutation increases the reduction potential of the T1 copper center . Kinetic results suggest that the change in driving force has a dramatic influence on the reactivity: The T2Cu(II) is initially reduced followed by ET to T1Cu(II) . The activation parameters have been determined and are compared with those of the wild-type (WT) AxCuNiR . The reorganization energy of the T2 site in the latter enzyme was calculated to be 1.6+/-0.2 eV which is two-fold larger than that of the T1 copper center in the WT protein. Biotechnol Appl Biochem, 2004 Dec, 40(Pt 3), 271 - 5 A novel synthetic peptide derivative from lactoferrin exhibiting antimicrobial activity; Ramesh CV et al.; A tetrapeptide derivative Boc-L-Lys(Boc)-L-Arg-L-Asp-L-Ser(Bu(t))-OBu(t) (PEP 1261; Boc is butoxycarbonyl, Bu(t) is t-butyl and OBu(t) is t-butyl ester), synthesized by a solution-phase strategy, exhibited antimicrobial activity against a broad spectrum of micro-organisms at an optimal concentration of 500 mug/ml . Whereas the tetrapeptide salt (L-Lys-L-Arg-L-Asp-L-Ser.HCl) was found to be fairly effective against bacterial cultures, it was not effective against fungal cultures . Comparative growth studies showed that PEP 1261 was equally as potent as the conventional antibiotics kanamycin, streptomycin and actidione for the Gram-negative bacteria Escherichia coli, Pseudomonas alcaligenes and the non-filamentous fungus Saccharomyces cerevisiae (baker's yeast), whereas 62 and 88.9% inhibition were observed for Gram-positive organisms such as Staphylococcus aureus and Bacillus thuringiensis respectively . PEP 1261 might exert its antimicrobial activity by permeabilizing the bacterial membrane, and this was confirmed by an increase in beta-galactosidase activity. Environ Toxicol Chem, 2004 Feb, 23(2), 325 - 30 Biodegradation of 2-chlorophenol in forest soil: effect of inoculation with aerobic sewage sludge; Lallai A et al.; Decontamination of 2-monochlorophenol-containing forest soil was studied in laboratory experiments . We found that in sterile soil, sorption of chlorophenol can occur . Chlorophenol disappearance of approximately 55% was observed in native soil; both soil sorption and degradation by indigenous soil populations caused this disappearance . In native soil, however, the rate of chlorophenol disappearance was enhanced up to slightly more than 90% by inoculation with a sludge taken from the aeration tank of a municipal wastewater treatment plant . In this sludge, the presence of Alcaligenes and Pseudomonas spp . was observed . In other experiments, addition to the soil of a laboratory culture preacclimated to 2-monochlorophenol did not lead to a greater increase in chlorophenol disappearance . In contrast to native soil, inoculation of sterile soil had no effect on disappearance of the chlorophenol . A possible explanation for the lack of cometabolic degradation is that autoclaving of the soil destroys the organic substances within it. J Basic Microbiol, 2004, 44(1), 59 - 65 Isolation and characterization of microorganisms capable of decolorizing various triphenylmethane dyes; Sharma DK et al.; Various soil and sludge samples collected from the vicinity of textile dyeing industries and waste disposal sites were used for enrichment of microbial population in the presence of triphenylmethane (TPM) dye Acid Violet-17 (AV-17) . Twenty-five (25) isolates were screened for their ability to decolorize AV-17 dye added at a rate of 10 mgl(-1) in mineral salts medium (MSM) agar plates . Five bacterial isolates belonging to Bacillus sp., Alcaligenes sp . and Aeromonas sp . were selected on the basis of their higher decolorization ability and were used to develop a bacterial consortium . The consortium was able to efficiently decolorize various TPM dyes viz . Acid Violet-17 (86%), Acid Blue-15 (85%), Crystal Violet (82%), Malachite Green (82%) and Brilliant Green (85%) . The consortium will be further used for designing efficient and cost effective treatment system for effluents of textile processing industries (TPI). Arch Pediatr, 2003 Sep, 10 Suppl 2, 342s - 346s {Pathogenic bacteria in cystic fibrosis}; Mariani-Kurkdjian P et al.; Since the CF gene identification in 1989 and despite the improvement of our knowledge in the physiopathology of the disease, bronchopulmonary infection determines the vital prognosis . Following Staphylococcus aureus infection, patients are colonized or colonized by Pseudomonas aeruginosa, greatly involved in the pulmonary deterioration . Other bacteria may be involved Burkholderia cepacia, Stenotrophomonas maltophilia, Alcaligenes sp . Intensive antibiotic treatment of primocolonisation helps to prevent or delay chronic colonisation . Chronic colonization needs a rational long term antibiotic strategy to prevent the occurrence of multiresistant germs; antibiotic cures are performed every 3 or 4 months before pulmonary exacerbation symptoms. Ukr Biokhim Zh, 2003 Mar-Apr, 75(2), 90 - 3 {Obtaining bioluminescent bacterial strains for detection of thallium ions}; Gruzina TG et al.; The possibility of Alcaligenes eutrophus T1 luxphenotype creation by temperature induced mutagenesis has been shown . These mutants are able to emit light after induction by thallium ions . This biological method of thallium detection possesses high specificity and sensitivity (0.5 microM of metal) . Such microbial cells can be used to quantify bioavailability of thallium part in the environment. Biotechnol Adv, 1986, 4(2), 245 - 59 Microbial polysaccharides with actual potential industrial applications; Paul F et al.; The microbial polysaccharides reviewed include xanthan gum, scleroglucan, PS-10, PS-21 and PS-53 gums, polysaccharides from Alcaligenes sp., PS-7 gum, gellan gum, curdlan, bacterial alginate, dextran, pullulan, Baker's Yeast Glycan, 6-deoxy-hexose-containing polysaccharides and bacterial cellulose . Factors limiting the commercial potential of certain microbial polysaccharides such as availability, rheological properties, and polyvalency are outlined . The polysaccharides are classified according to their uses as viscosity-increasing agents and as gelling agents . A third category includes polysaccharides with specific applications such as tailor-made dextran and pullulan and polysaccharides used as substrates for the preparation of rare sugars . The difficulties encountered in development of a polysaccharide at the industrial level are pointed out. Biotechnol Prog, 2003 Sep-Oct, 19(5), 1487 - 97 Cybernetic model predictive control of a continuous bioreactor with cell recycle; Gadkar KG et al.; The control of poly-beta-hydroxybutyrate (PHB) productivity in a continuous bioreactor with cell recycle is studied by simulation . A cybernetic model of PHB synthesis in Alcaligenes eutrophus is developed . Model parameters are identified using experimental data, and simulation results are presented . The model is interfaced to a multirate model predictive control (MPC) algorithm . PHB productivity and concentration are controlled by manipulating dilution rate and recycle ratio . Unmeasured time varying disturbances are imposed to study regulatory control performance, including unreachable setpoints . With proper controller tuning, the nonlinear MPC algorithm can track productivity and concentration setpoints despite a change in the sign of PHB productivity gain with respect to dilution rate . It is shown that the nonlinear MPC algorithm is able to track the maximum achievable productivity for unreachable setpoints under significant process/model mismatch . The impact of model uncertainty upon controller performance is explored . The multirate MPC algorithm is tested using three controllers employing models that vary in complexity of regulation . It is shown that controller performance deteriorates as a function of decreasing biological complexity. Microbiol Res, 2003, 158(3), 215 - 27 In silico analysis of a flavohemoglobin from Sinorhizobium meliloti strain 1021; Lira-Ruan V et al.; Hemoglobins (Hbs) have been characterized from a wide variety of eubacteria, but not from nitrogen-fixing rhizobia . Our search for Hb-like sequences in the Sinorhizobium meliloti genome revealed that a gene coding for a flavohemoglobin (fHb) exists in S . meliloti (SmfHb) . Computer analysis showed that SmfHb and Alcaligenes eutrophus fHb are highly similar and could fold into the same tertiary structure . A FNR-like box was detected upstream of the smfhb gene and mapping analysis revealed that the smfhb gene is flanked by nos and fix genes . These observations suggest that smjhb is regulated by the concentration of O2 and that SmfHb functions in some aspects of nitrogen metabolism. Biotechnol Lett, 2003 Sep, 25(17), 1421 - 4 Removal and destruction of high concentrations of gaseous toluene in a two-phase partitioning bioreactor by Alcaligenes xylosoxidans; Daugulis AJ et al.; A two-phase bioreactor consisting of hexadecane dispersed in an aqueous, cell-containing medium (organic fraction = 0.33) was used to trap toluene vapours from an air stream . The affinity for toluene by the solvent resulted in high efficiency of removal and transfer to the aqueous phase based on equilibrium transfer . The system was readily able to handle a loading capacity of 748 mg l(-1) h(-1) at a toluene degradation efficiency of greater than 98%. Biotechnol Lett, 2003 Jul, 25(14), 1203 - 7 Delivery of benzene to Alcaligenes xylosoxidans by solid polymers in a two-phase partitioning bioreactor; Daugulis AJ et al.; Toxic levels of benzene were decreased to sub-inhibitory levels in a bioreactor via absorption by polymer beads or cylinders (poly(ethylene-co-vinyl acetate) or poly(styrene-co-butadiene)) . After inoculation with Alcaligenes xylosoxidans, the remaining benzene in the aqueous phase as well as the benzene taken up by the polymers was degraded to completion . The capacity of these polymers for benzene, and benzene diffusivity within the polymers were also determined. Appl Microbiol Biotechnol, 2003 Sep, 62(4), 337 - 41 Epub 2003 Feb 26. Regioselective hydroxylation of quinolinic acid, lutidinic acid and isocinchomeronic acid by resting cells of pyridine dicarboxylic acid-degrading microorganisms; Uchida A et al.; Microorganisms aerobically degrading quinolinic acid, lutidinic acid or isocinchomeronic acid were isolated and the microbial regioselective hydroxylation of these pyridine dicarboxylic acids was studied . Alcaligenes sp . UK21 cells converted quinolinic acid into 6-hydroxypicolinic acid, suggesting the involvement of two enzyme reactions catalyzing hydroxylation at position C6 and decarboxylation at position C3 of quinolinic acid . Resting cells of Alcaligenes sp . UK21 accumulated 94.9 mM 6-hydroxypicolinic acid (13.2 g l(-1)), with a 96% molar conversion yield by 48 h incubation . Rhizobium sp . LA17 and Hydrogenophaga sp . IMA01 catalyzed the regioselective hydroxylation of lutidinic acid and isocinchomeronic acid into 6-hydroxylutidinic acid and 6-hydroxyisocinchomeronic acid, respectively . 6-Hydroxylutidinic acid accumulated up to 95.4 mM (17.5 g l(-1)) by 24 h incubation in the resting cells reaction, using Rhizobium sp . LA17, with a 99% molar conversion yield . Resting cells of Hydrogenophaga sp . IMA01 produced 88.7 mM 6-hydroxyisocinchomeronic acid (16.2 g l(-1)) by 24 h incubation, with a 81% molar conversion yield. Gene, 2003 Jul 17, 312, 239 - 48 Molecular characterization of an inducible gentisate 1,2-dioxygenase gene, xlnE, from Pseudomonas alcaligenes NCIMB 9867; Yeo CC et al.; Pseudomonas alcaligenes NCIMB 9867 (strain P25X) produces isofunctional enzymes of the gentisate pathway that enables the degradation of xylenols and cresols via gentisate . Previous reports had indicated that one set of enzymes is constitutively expressed whereas the other set is strictly inducible by aromatic hydrocarbon substrates . The gene encoding gentisate 1,2-dioxygenase (GDO), the enzyme that catalyzes the cleavage of the gentisate aromatic ring, was cloned from strain P25X . The GDO gene, designated xlnE, is 1,044 bp, and is part of a 5.4 kb operon which consists of six genes, xlnEFGHID . Transcription of this operon was driven by a sigma 70-type promoter, PxlnE, located 123 bp upstream of the xlnE start codon . Primer extension analysis showed that the xlnE transcription start point is located at the -87 adenine residue . In a P25X xlnE knockout mutant, GDO activity could only be detected when cells were grown in the presence of aromatic substrates, suggesting that xlnE encodes for the constitutive copy of GDO . This was verified by constructing a P25X strain with xlnE transcriptionally fused to a promoterless catechol 2,3-dioxygenase gene . In this strain, catechol 2,3-dioxygenase activity was detected in cells that were grown in the absence of aromatic inducers . However, catechol 2,3-dioxygenase activity increased up to four fold when these cells were grown in the presence of aromatic substrates, in particular 3-hydroxybenzoate . Thus, xlnE is in fact, inducible and the constitutive activity observed under non-inducing conditions was due to its relatively high basal levels of expression. Microbiol Res, 2003, 158(2), 195 - 9 Effect of carbon source on pyrimidine biosynthesis in Pseudomonas alcaligenes ATCC 14909; Santiago MF et al.; The effect of carbon source on the regulation of the de novo pyrimidine biosynthetic enzymes in Pseudomonas alcaligenes ATCC 14909 was investigated . The de novo pyrimidine biosynthetic enzymes were measured in extracts of P . alcaligenes ATCC 14909 cells and of cells from an auxotroph deficient for orotate phosphoribosyltransferase activity . Pyrimidine biosynthetic enzyme activities in ATCC 14909 were influenced by pyrimidine supplementation to the culture medium but not by the carbon source present . Pyrimidine limitation of the auxotroph elevated the de novo enzyme activities indicating that this pathway may be controlled at the transcriptional level by a pyrimidine-related compound . Its regulation seemed to be subject to less transcriptional control by a pyrimidine-related compound than what was observed in the closely related species Pseudomonas pseudoalcaligenes. Biotechnol Bioeng, 2003 Sep 30, 83(7), 854 - 63 In silico prediction and validation of the importance of the Entner-Doudoroff pathway in poly(3-hydroxybutyrate) production by metabolically engineered Escherichia coli; Hong SH et al.; The metabolic network of Escherichia coli was constructed and was used to simulate the distribution of metabolic fluxes in wild-type E . coli and recombinant E . coli producing poly(3-hydroxybutyrate) {P(3HB)} . The flux of acetyl-CoA into the tricarboxylic acid (TCA) cycle, which competes with the P(3HB) biosynthesis pathway, decreased significantly during P(3HB) production . It was notable to find from in silico analysis that the Entner-Doudoroff (ED) pathway flux increased significantly under P(3HB)-accumulating conditions . To prove the role of ED pathway on P(3HB) production, a mutant E . coli strain, KEDA, which is defective in the activity of 2-keto-3-deoxy-6-phosphogluconate aldolase (Eda), was examined as a host strain for the production of P(3HB) by transforming it with pJC4, a plasmid containing the Alcaligenes latus P(3HB) biosynthesis operon . The P(3HB) content obtained with KEDA (pJC4) was lower than that obtained with its parent strain KS272 (pJC4) . The reduced P(3HB) biosynthetic capacity of KEDA (pJC4) could be restored by the co-expression of the E . coli eda gene, which proves the important role of ED pathway on P(3HB) synthesis in recombinant E . coli as predicted by metabolic flux analysis . Biotechnol Lett, 2003 Jan, 25(1), 83 - 7 Chemical modification of lipases with various hydrophobic groups improves their enantioselectivity in hydrolytic reactions; Ueji S et al.; Semi-purified lipases from Candida rugosa, Pseudomonas cepacia and Alcaligenes sp . were chemically modified with a wide range of hydrophobic groups such as benzyloxycarbonyl, p-nitrobenzyloxycarbonyl, p-methoxybenzyloxycarbonyl, t-butoxycarbonyl, lauroyl and acetyl moieties . The Candida rugosa lipase MY modified with the benzyloxycarbonyl group (modification ratio = 84%) brought about a 15-fold increase in enantioselectivity (E value) towards the hydrolysis of racemic butyl 2-(4-ethylphenoxy)propionate in an aqueous buffer solution, although the enzymatic activity was decreased . The origin of the enantioselectivity enhancement by chemical modification of the lipase is attributed to a significant deceleration in the initial reaction rate for the incorrectly binding enantiomer. Biotechnol Lett, 2003 May, 25(9), 715 - 7 Purification and characterization of chitinase from Alcaligenes xylosoxydans; Vaidya R et al.; Extracellular chitinase from Alcaligenes xylosoxydans was purified to electrophoretic homogeneity using affinity and gel filtration chromatography . The molecular mass of chitinase was estimated to be 45 kDa and 44 kDa by SDS-PAGE and gel-filtration, respectively . The enzyme was optimally active at 50 degrees C (over 30 min) and pH 5 . Activity staining after PAGE showed a single band . The Km for chitin was 3 g l-1 . Cu2+ and Na+ at 5 mM inhibited chitinase activity to 25% while Ca2+, Mg2+ and Ba2+ had no effect at the same concentration . The purified enzyme degraded mycelia of Aspergillus niger. Biotechnol Lett, 2003 May, 25(9), 665 - 70 Production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) with high molar fractions of 3-hydroxyvalerate by a threonine-overproducing mutant of Alcaligenes sp . SH-69; Choi GG et al.; A threonine overproducing mutant of Alcaligenes sp . SH-69 was isolated and its ability to produce poly(3-hydroxybutyrate-co-3-hydroxyvalerate), poly(3HB-co-3HV), was investigated . The 3HV fraction in poly(3HB-co-3HV) produced from glucose as the sole carbon source exceeded 22 mol%, which is approximately six times higher than that achieved by the wild type under the same culture conditions . Furthermore, the addition of a relatively low concentration (10 mM) of propionic acid, valeric acid or levulinic acid to the glucose medium greatly increased the molar fraction of 3HV in the copolyester, to 38-77 mol% . The results suggest that metabolic engineering of the biosynthetic pathways supplying polyhydroxyalkanoate monomers, such as the threonine biosynthetic pathway, can lead to new poly(3HB-co-3HV)-producing strains. Arch Microbiol, 2003 Aug, 180(2), 81 - 7 Epub 2003 Jul 03. Quinolinate dehydrogenase and 6-hydroxyquinolinate decarboxylase involved in the conversion of quinolinic acid to 6-hydroxypicolinic acid by Alcaligenes sp . strain UK21; Uchida A et al.; In the conversion of quinolinic acid to 6-hydroxypicolinic acid by whole cells of Alcaligenes sp . strain UK21, the enzyme reactions involved in the hydroxylation and decarboxylation of quinolinic acid were examined . Quinolinate dehydrogenase, which catalyzes the first step, the hydroxylation of quinolinic acid, was solubilized from a membrane fraction, partially purified, and characterized . The enzyme catalyzed the incorporation of oxygen atoms of H(2)O into the hydroxyl group . The dehydrogenase hydroxylated quinolinic acid and pyrazine-2,3-dicarboxylic acid to form 6-hydroxyquinolinic acid and 5-hydroxypyrazine-2,3-dicarboxylic acid, respectively . Phenazine methosulfate was the preferred electron acceptor for quinolinate dehydrogenase . 6-Hydroxyquinolinate decarboxylase, catalyzing the nonoxidative decarboxylation of 6-hydroxyquinolinic acid, was purified to homogeneity and characterized . The purified enzyme had a molecular mass of approximately 221 kDa and consisted of six identical subunits . The decarboxylase specifically catalyzed the decarboxylation of 6-hydroxyquinolinic acid to 6-hydroxypicolinic acid, without any co-factors . The N-terminal amino acid sequence was homologous with those of bacterial 4,5-dihydroxyphthalate decarboxylases. FEMS Microbiol Rev, 2003 Jun, 27(2-3), 385 - 410 Ralstonia metallidurans, a bacterium specifically adapted to toxic metals: towards a catalogue of metal-responsive genes; Mergeay M et al.; Ralstonia metallidurans, formerly known as Alcaligenes eutrophus and thereafter as Ralstonia eutropha, is a beta-Proteobacterium colonizing industrial sediments, soils or wastes with a high content of heavy metals . The type strain CH34 carries two large plasmids (pMOL28 and pMOL30) bearing a variety of genes for metal resistance . A chronological overview describes the progress made in the knowledge of the plasmid-borne metal resistance mechanisms, the genetics of R . metallidurans CH34 and its taxonomy, and the applications of this strain in the fields of environmental remediation and microbial ecology . Recently, the sequence draft of the genome of R . metallidurans has become available . This allowed a comparison of these preliminary data with the published genome data of the plant pathogen Ralstonia solanacearum, which harbors a megaplasmid (of 2.1 Mb) carrying some metal resistance genes that are similar to those found in R . metallidurans CH34 . In addition, a first inventory of metal resistance genes and operons across these two organisms could be made . This inventory, which partly relied on the use of proteomic approaches, revealed the presence of numerous loci not only on the large plasmids pMOL28 and pMOL30 but also on the chromosome . It suggests that metal-resistant Ralstonia, through evolution, are particularly well adapted to the harsh environments typically created by extreme anthropogenic situations or biotopes. J Appl Microbiol, 2003, 95(1), 160 - 6 Development of a Gram-negative selective agar (GNSA) for the detection of Gram-negative microflora in sputa in patients with cystic fibrosis; Moore JE et al.; AIMS: To develop a selective agar medium to help detect and quantify Gram-negative flora in the sputum of patients with cystic fibrosis (CF) . METHODS AND RESULTS: A novel Gram-negative Selective Agar (GNSA) medium was developed consisting of tryptone soya broth (30 g), bacteriological agar no.1 (10 g), yeast extract (5 g), crystal violet (2 mg), nisin (48 mg), novobiocin (5 mg), cycloheximide (100 mg), amphotericin (2 mg) and double distilled water (1 l), for the selective culture of all Gram-negative flora from the sputum of patients with CF . GNSA was able to support the proliferation of all 34 Gram-negative organisms examined, including 23 species most commonly associated with CF, but was unable to support the growth of the 12 Gram-positive or seven fungal organisms examined . Sensitivity studies demonstrated that the GNSA medium was able to detect not less than 1.50 x 102 CFU ml-1 sputum Pseudomonas aeruginosa, 2.38 x 102 CFU ml-1 sputum Burkholderia cepacia genomovar IIIb and 6.70 x 103 CFU ml-1 sputum Stenotrophomonas maltophilia . A comparison of the microbial flora detected in the sputa of 12 adult CF patients by employment of routine bacteriological agar media and GNSA, demonstrated that GNSA was able to detect all Gram-negative organisms cultured by routine media, but had the advantage of detecting Alcaligenes xylosoxidans in two CF patients, whom had no previous history of Gram-negative infection . CONCLUSIONS: GNSA was unable to support the proliferation of any Gram-positive organism or yeast/fungi, but was successful in supporting the growth of all Gram-negative organisms challenged . SIGNIFICANCE AND IMPACT OF THE STUDY: Employment of this medium coupled with semi-automated technology may aid in helping to efficiently determine Gram-negative loading of respiratory secretions, particularly in response to antibiotic intervention. Infect Immun, 2003 Jun, 71(6), 3196 - 205 Modulation of virulence by two acidified nitrite-responsive loci of Salmonella enterica serovar Typhimurium; Kim CC et al.; Two acidified nitrite-inducible genes of Salmonella enterica serovar Typhimurium were identified with a green fluorescent protein-based promoter-trap screen . The nitrite-inducible promoters were located upstream of loci that we designated nipAB and nipC, which correspond to hcp-hcr (hybrid cluster protein) of Escherichia coli and norA of Alcaligenes eutrophus, respectively . Maximal induction of the promoters by nitrite was dependent on pH . The nipAB promoter was regulated by oxygen in an Fnr-dependent manner . The nipC promoter was also regulated by oxygen but in an Fnr-independent manner . The promoters were upregulated in activated RAW264.7 macrophage-like cells, which produce NO via the inducible nitric oxide synthase (iNOS), and the induction was inhibited by aminoguanidine, an inhibitor of iNOS . Although the nipAB and nipC mutants displayed no defects under a variety of in vitro conditions or in tissue culture infections, they exhibited lower oral 50% lethal doses (LD(50)s) than did the wild type in C57BL/6J mouse infections . The lower LD(50)s reflected an unexpected increased ability of small inoculating doses of the mutant bacteria to cause lethal infection 2 to 3 weeks after challenge, compared to a similar challenge dose of wild-type bacteria . We conclude that these genes are regulated by physiological nitrogen oxides and that the absence of these bacterial genes in some way diminishes the ability of mice to clear a low dose infection. Biotechnol Bioeng, 2003 Jul 5, 83(1), 104 - 11 Evaluation of spectrofluorometry as a tool for estimation in fed-batch fermentations; Hagedorn A et al.; Native culture fluorescence was investigated as an additional source of information for predicting biomass and glucose concentrations in a fed-batch fermentation of Alcaligenes eutrophus . Partial least squares (PLS) regression and a feed forward neural network (FFNN) coupled with principle component analysis (PCA) were each used to model the kinetics of the fermentation . Data from three fermentations was combined to form a training set for model calibration and data from a fourth fermentation was used as the testing set . The fluorescent soft-sensors were compared with a previously developed feed forward neural network soft-sensor model which used oxygen uptake rate (OUR), carbon dioxide evolution rate (CER), aeration rate, feed rate, and fermentor volume to estimate biomass and glucose concentrations . The best model performance for predicting both biomass and glucose concentrations was achieved using the native fluorescence-based models . Real data predictions of the biomass concentration in the testing set were obtained using both the PLS and FFNN PCA modeling utilizing fluorescence measurements plus the rate of change of the fluorescence measurements . Accurate predictions of the glucose concentration in the testing set were obtained using the FFNN PCA modeling technique utilizing the rate of change of the fluorescence measurements . Substrate exhaustion was indicated qualitatively by a first-order PLS model utilizing the rate of change of fluorescence measurements . These results indicate that native culture fluorescence shows promise for providing additional valuable information to enhance predictive modeling which cannot be extracted from other easily acquired measurements . Int Microbiol, 2003 Mar, 6(1), 57 - 64 Epub 2003 Mar 13. Conjugative plasmid mediated inducible nickel resistance in Hafnia alvei 5-5; Park JE et al.; Hafnia alvei 5-5, isolated from a soil-litter mixture underneath the canopy of the nickel-hyperaccumulating tree Sebertia acuminata (Sapotaceae) in New Caledonia, was found to be resistant to 30 mM Ni(2+) or 2 mM Co(2+) . The 70-kb plasmid, pEJH 501, was transferred by conjugation to Escherichia coli, Serratia marcescens, and Klebsiella oxytoca . Transconjugant strains expressed inducible nickel resistance to between 5 and 17 mM Ni(2+), and cobalt resistance to 2 mM Co(2+) . A 4.8-kb Sal- EcoRI fragment containing the nickel resistance determinant was subcloned, and the hybrid plasmid was found to confer a moderate level of resistance to nickel (7 mM Ni(2+)) even to E . coli . The expression of nickel resistance was inducible by exposure to nickel chloride at a concentration as low as 0.5 mM Ni(2+) . By random Tn phoA'-1 insertion mutagenesis, the fragment was shown to have structural genes as well as regulatory regions for nickel resistance . Southern hybridization studies showed that the nickel-resistance determinant from pEJH501 of H . alvei 5-5 was homologous to that of pTOM9 from Alcaligenes xylosoxydans 31A. J Food Prot, 2003 Apr, 66(4), 592 - 8 Comparison of sodium hypochlorite-based foam and peroxyacetic acid-based fog sanitizing procedures in a salmon smokehouse: survival of the general microflora and Listeria monocytogenes; Bagge-Ravn D et al.; The effects of fog sanitization with peroxyacetic acid (hydrogen peroxide, peracetic acid, and acetic acid in combination) on general hygiene (aerobic plate count) and on Listeria monocytogenes were assessed in a slicing area at a salmon smokehouse and compared with the effects of foam sanitization with sodium hypochlorite (routinely performed at the smokehouse) . Two hundred twenty-three environmental samples were collected with sponges and swabs after each of the sanitization procedures, and 68 samples were collected during production . The total culturable aerobic plate count was determined for each sample, and a total of 288 bacterial strains were randomly isolated and tentatively identified to genus level by physiological and biochemical tests . The microflora was dominated by Neisseriaceae, Enterobacteriaceae, and lactic acid bacteria during production . Foam sanitization caused a change in the composition of the flora, with Pseudomonas spp . and Alcaligenes spp . being the dominant gram-negative bacteria and Kurthia spp . and Bacillus spp . being the surviving gram-positive bacteria . Bacteria were very sensitive to fog sanitization, and yeasts accounted for almost half of the surviving flora . By a selective isolation method, strains of L . monocytogenes were isolated and subsequently characterized by random amplified polymorphic DNA (RAPD) typing . Following foam sanitization, 14 to 42% of the samples contained <10 CFU per site, whereas 29 to 78% of the samples collected after fog sanitization contained this level of bacteria . The prevalence of L . monocytogenes was unchanged, but L . monocytogenes was found only in poorly cleaned areas such as drains . The RAPD types for all positive samples were identical to the type that had persisted in the smokehouse since 1995, indicating the importance of drains as a niche. J Mol Biol, 2003 Apr 25, 328(2), 429 - 38 Atomic resolution structures of native copper nitrite reductase from Alcaligenes xylosoxidans and the active site mutant Asp92Glu; Ellis MJ et al.; We provide the first atomic resolution (<1.20 A) structure of a copper protein, nitrite reductase, and of a mutant of the catalytically important Asp92 residue (D92E) . The atomic resolution where carbon-carbon bonds of the peptide become clearly resolved, remains a key goal of structural analysis . Despite much effort and technological progress, still very few structures are known at such resolution . For example, in the Protein Data Bank (PDB) there are some 200 structures of copper proteins but the highest resolution structure is that of amicyanin, a small (12 kDa) protein, which has been resolved to 1.30 A . Here, we present the structures of wild-type copper nitrite reductase (wtNiR) from Alcaligenes xylosoxidans (36.5 kDa monomer), the "half-apo" recombinant native protein and the D92E mutant at 1.04, 1.15 and 1.12A resolutions, respectively . These structures provide the basis from which to build a detailed mechanism of this important enzyme . J Basic Microbiol, 2003, 43(1), 75 - 9 Comparison of aspartate transcarbamoylase regulation in Pseudomonas alcaligenes and Pseudomonas mendocina; Santiago MF et al.; The regulation of aspartate transcarbamoylase activity in cell extracts of Pseudomonas alcaligenes ATCC 14909 and Pseudomonas mendocina ATCC 25411 was compared . Under saturating substrate concentrations, pyrophosphate, CTP, UDP and ADP were highly inhibitory of the P . alcaligenes transcarbamoylase activity while pyrophosphate, UDP, ADP, ATP and GTP were the most effective inhibitors of the P . mendocina transcarbamoylase . By examining transcarbamoylase inhibition by ribonucleotide triphosphates, it was possible to differentiate these species assigned to different DNA homology groups and such an analysis might prove useful in the reclassification of Pseudomonas species. Lett Appl Microbiol, 2003, 36(3), 129 - 34 The novel method for isolating chitinolytic bacteria and its application in screening for hyperchitinase producing mutant of Alcaligenes xylosoxydans; Vaidya RJ et al.; AIMS: To develop a novel, rapid and effective screening method for chitinase producing bacteria . METHODS AND RESULTS: A simple and rapid technique for screening of potential chitinolytic bacteria has been developed using the chitin binding dye calcofluor white M2R in chitin agar . Microorganisms possessing high chitinolytic potential gave a clear zone under ultraviolet light after 24-48 h of incubation . This method was successfully applied for isolating the hyperchitinase mutant of Alcaligenes xylosoxydans . The mutant Alc . xylosoxydans EMS 33 was found to produce 3.4 times more chitinase than the wild type . CONCLUSIONS: In this study, the screening method for chitinase producing bacteria has been developed and it was applied to screen chitinase-overproducing mutant of Alc . xylosoxydans . SIGNIFICANCE AND IMPACT OF THE STUDY: The novel screening method for chitinase producer is more sensitive, rapid, user-friendly and reliable, which can also be used for screening of recombinants having chitinase gene. Wei Sheng Wu Xue Bao, 2002 Apr, 42(2), 153 - 62 {Cloming, sequence analysis of imidase gene from Alcaligenes eatrophus and its expression in E . coli}; Wang Y et al.; A hydantoin-cleaving microorganism 112R4 is screened and identified to be Alcaligenes eutrophus . The resting cell of Alcaligenes eutrophus 112R4 can catalyze the hydrolysis of hydantoin, dihydropyrimidine and succinimide effectively, but not function to 5-monosubstituted hydantoins or 5,5'-disubstituted hydantoins . The microorganism can utilize succinimide as a sole carbon source and nitrogen source, which indicates the presence of a complete transformation pathway of succinimide, and a hydantoin-cleaving enzyme, imidase, is suggested to be contained in this metabolic pathway . A 6 kb EcoRI-EcoRI fragment isolated from the genome DNA of Alcaligenes eutrophus 112R4 is shown to be correlative with the transformation of succinimide . A 2 kb DNA fragment containing the gene of imidase is subcloned and sequenced . Deletion analysis verifies that one open reading frame of 876 nucleotides, which encodes a peptide of 291 amino acids, with a calculated molecular weight of 33688, is responsible for the encoding of imidase . This is the first report of the nucleotide and amino acid sequences of imidase (GenBank accession number: AF373287) . A homology search performed in protein database reveals an identity of 14% with polysaccharide deacetylase conserved domain, an identity of 60% with N-terminal 20 amino acids of Blastobacter sp . A17p-4, but no apparent similarity with all known cyclic-amide-cleaving enzymes . This result suggested that the imidase should be classified as a new member of cyclic amidases . Under the control of lac promoter and IPTG induction, the imidase activity of transformed E . coli reached 3200 U/L, which is about 7-fold higher than that of gene donor strain. Wei Sheng Wu Xue Bao, 1999 Jun, 39(3), 247 - 54 {Optimization of fermentation conditions for the production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) with Alcaligenes eutrophus}; Du G et al.; A close relationship, between the initial addition time and concentration of propionate and HV fraction, was observed in shaking culture of Alcaligenes eutrophus for the production of Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) . The optimal initial addition time of propionate was determined at the onset of PHBV formation period . Although relatively high HV unit could be obtained under high propionate concentration, the growth and product synthetic activities were inhibited obviously . Different ratios of glucose to propionate were fed to stimulate the formation of HV unit and the results were compared . The optimizing feeding strategy of propionate was proposed based on the consideration of several fermentation index . The final cell dry weight, PHB concentration, PHB content and HV fraction in PHBV reached 52.1 g/L, 40.8 g/L, 78.3% and 16.2 mol%, respectively, Yield coefficient of HV unit to propionate and PHBV productivity were obtained to be 0.5 g/g and 0.74 g/(L/h), respectively. Wei Sheng Wu Xue Bao, 1999 Apr, 39(2), 160 - 3 {Studies on the parasites of Paris polyphylla var . yunnanensis}; Wang S et al.; Two bacteria and three fungi were isolated from the hypogeal stems of Paris polyphylla var . yunnanensis . The bacteria were identified as Bacillus cereus and Pseudomonas alcaligenes, and the fungi were identified as Periconia sp., Pachnocybe albida and Hormomyces paridiphilus . The results on liquid culture of B . cereus, P . alcaligenes and H . paridiphilus indicated that the colloidization and polysaccharide content increasing in hypogeal stems of P . polyphylla var . yunnanensis were due to the extracellular polysaccharide secretion of the parasitical fungus H . paridiphilus. Wei Sheng Wu Xue Bao, 1998 Feb, 38(1), 52 - 6 {Studies on microbial factor on color change of Dunhuang mural . I . Classification of microbes on color-changed mural and property of some typical species}; Feng Q et al.; Microbial strains were isolated from 51 typical color changed samples of mural in 6 grottos of Dunhuang Mogao Gretto . After identification, it is proved that belong to 6 genera of bacteria and 5 genera of mould . Bacillus, Alcaligenes and Penicillium were dominant species in all of them . In the imitative experiment, it is found that Cladosporium sp., A . niger and two strains of bacteria have a great effect on the color change of red pigment of mural and aging of sizing agent. Wei Sheng Wu Xue Bao, 2000 Dec, 40(6), 579 - 85 {Cloning of a new catechol 1,2-dioxygenase gene (tfd C) from Plesiomonas and its expression in the E . coli}; Ma Z et al.; A new catechol-1,2-dioxygenase gene (tfd C) was cloned from the Plesiomonas using the PCR method . Primers were designed according to the reported sequence of Catechol-1,2-dioxygenase (C120) gene from Alcaligenes eutroplus . The amplified fragment contained a 765 bp open reading frame (ORF), encoding a protein of 255 amino acids . The new tfd C gene shared a high homology with the one cloned from Alcaligenes eutroplus, showing only one base difference at 693 site (C-->A) and consequently one amino acid difference at 228 site (P-->T) . The ORF was cloned to the plasmid pBluescriptII KS, which was transferred to E . coli JM109 and a positive clone, pBt2G, was then selected . A significant activity of C120 was detected in the positive clone . When the ORF was cloned to the plasmid pET-30a, which was transferred to E . coli BL21(DE3) plysS, the expected 33 kD protein was detected from a positive clone, pET30A, by SDS-PAGE . The C120 is a key enzyme in degrading aromatic pollutants in the environment . In order to use plants to degrade aromatic pollutants, the gene will be introduced into the turfgrass . To express the gene properly in plants, its translation initiation codon was modified from GTG to ATG . A similar activity of C120 was obtained following the modification. Wei Sheng Wu Xue Bao, 2000 Jun, 40(3), 290 - 5 {Effects of nitrogen feeding on the accumulation of poly-beta-hydroxybutyrate with Alcaligenes eutrophus}; Du G et al.; On the basis of analysis of PHB fermentation processes, the effects of ammonium sulfate feeding rate at PHB formation period on the PHB accumulation by Alcaligenes eutrophus were investigated . It was shown that the complete absence of nitrogen source at PHB formation phase would lead to the decline of PHB synthetic activity, and the obvious influences of different nitrogen feeding rate on PHB synthesis were observed . Higher PHB content, but relative lower cell dry weight, PHB concentration and PHB productivity could be obtained at slower nitrogen feeding rate . The excessive nitrogen feeding rate resulted in the drop of PHB content, which led to the decrease of PHB concentration and PHB productivity . The better results could be achieved when the ammonium sulfate feeding rate was set at around 0.5 g/h. Wei Sheng Wu Xue Bao, 2000 Feb, 40(1), 26 - 31 {The construction of a recombined E . coli strain with stable and high production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate)}; Tian J et al.; Plasmid pJMC2 was constructed by cloning the parDE fragment of RK2 into pTZ18U-PHB which harbored phaCAB from Alcaligenes eutrophus and was transferred into E . coli HMS174 and E . coli JM107 separately . It is very stable in its hosts cultured in medium without ampicillin . E . coli HMS 174(pTZ18U-PHB) and E . coli JM107(pTZ18U-PHB) produced P(3HB-co-3HB) in a low phosphate concentration medium(18 mmol/L) . The proportion of 3-hydroxyvalerate(3HV) in the polymer was 5%-8% . A fed-batch culture of E . coli HMS174(pJMC2) was conduct in a 5 L automatically controlled fermentor, the final dry cell weight, P(3HB-co-3HV) content, and th 3HV proportion were 42.5 g/L, 70% and 4.9% respectively. J Clin Microbiol, 2003 Jan, 41(1), 492 - 4 Evaluation of MicroScan Autoscan for identification of Pseudomonas aeruginosa isolates from cystic fibrosis patients; Saiman L et al.; Accurate identification of gram-negative bacilli from cystic fibrosis (CF) patients is essential . Only 57% (108 of 189) of nonmucoid strains and 40% (24 of 60) of mucoid strains were definitively identified as Pseudomonas aeruginosa with MicroScan Autoscan . Most common misidentifications were Pseudomonas fluorescens-Pseudomonas putida (i.e., the strain was either P . fluorescens or P . putida, but the system did not make the distinction and yielded the result P . fluorescens/putida) and Alcaligenes spp . Extending the incubation to 48 h improved identification, but 15% of isolates remained misidentified . The MicroScan Autoscan system cannot be recommended for the identification of P . aeruginosa isolates from CF patients. J Gen Appl Microbiol, 1998 Aug, 44(4), 269 - 274 Purification and characterization of beta-1,3-xylanase from a marine bacterium, Alcaligenes sp . XY-234; Araki T et al.; A beta-1,3-xylanase-producing bacterium, Alcaligenes sp . XY-234, was isolated from the marine environment . The organism produced endo-1,3-beta-xylanase at a high level in the culture fluid . The enzyme was purified 292-fold by ammonium sulfate precipitation and several column chromatographies . The final enzyme preparation appeared to be homogeneous on disc gel electrophoresis and SDS-PAGE with a molecular mass of 59 kDa, and the pI was 4.0 . The enzyme hydrolyzed beta-1,3-xylan and larger xylooligosaccharides than xylobiose to give several xylooligosaccharides, but it could not hydrolyze xylobiose, p-nitrophenyl-beta-D-xyloside, and beta-1,4-xylan . The Km of the enzyme was 4.0 mg/ml . Optimal pH and temperature were 7.5 and 40 degrees C, respectively . It was stable from pH 6.0 to 10 and at a temperature of less than 40 degrees C . The enzyme was strongly inhibited by 1 mM HgCl(2)., AlCl(3), CuCl(2), FeCl(3), HgCl(2), Pb(CH(3)COO) (2), and N-bromosuccinimide. J Environ Sci (China), 2002 Oct, 14(4), 445 - 50 A reactor system combining reductive dechlorination with co-metabolic oxidation for complete degradation of tetrachloroentylene; Lee TH et al.; A laboratory sequential anaerobic-aerobic bioreactor system, which consisted of an anaerobic fixed film reactor and two aerobic chemostats, was set up to degrade tetrachloroethylene (PCE) without accumulating highly toxic degradation intermediates . A soil enrichment culture, which could reductively dechlorinate 900 microM (ca . 150 mg/L) of PCE stoichiometrically into cis-1,2-dichloroethylene (cis-DCE), was attached to ceramic media in the anaerobic fixed film reactor . A phenol degrading strain, Alcaligenes sp . R5, which can efficiently degrade cis-DCE by co-metabolic oxidation, was used as inoculum for the aerobic chemostats consisted of a transformation reactor and a growth reactor . The anaerobic fixed film bioreactor showed more than 99% of PCE transformation into cis-DCE in the range of influent PCE concentration from 5 microM to 35 microM at hydraulic retention time of 48 h . On the other hand, efficient degradation of the resultant cis-DCE by strain R5 in the following aerobic system could not be achieved due to oxygen limitation . However, 54% of the maximum cis-DCE degradation was obtained when 10 mumol of hydrogen peroxide (H2O2) was supplemented to the transformation reactor as an additional oxygen source . Further studies are needed to achieve more efficient co-metabolic degradation of cis-DCE in the aerobic reactor. Res Microbiol, 2002 Nov, 153(9), 579 - 84 Molecular characterization of a salt-tolerant bacterial community in the rice rhizosphere; Tripathi AK et al.; The diversity of salt-tolerant bacteria present in the rhizosphere of Oryza sativa was investigated . Fourteen bacterial strains, isolated after enrichment in nitrogen-free, semi-solid medium and showing tolerance to 3% NaCl, were analyzed by restriction patterns produced by amplified DNA coding for 16S rDNA (ARDRA) with enzymes Sau3AI, AluI and RsaI which showed that they were represented by 4 ARDRA types . Biodiversity among the 14 strains was also analyzed by the random amplified polymorphic DNA (RAPD) technique with a 10-mer primer . Partial nucleotide sequence of 16S rDNA assigned these clusters to Serratia marcescens, Pseudomonas aeruginosa, Alcaligenes xylosoxidans and Ochrobactrum anthropi . Notably, all four bacterial species are potential human pathogens that infect immunocompromised patients. Appl Environ Microbiol, 2002 Dec, 68(12), 5925 - 32 A novel high-cell-density protein expression system based on Ralstonia eutropha; Srinivasan S et al.; We describe the development of a novel protein expression system based on the industrial fermentation organism Ralstonia eutropha (formerly known as Alcaligenes eutrophus) NCIMB 40124 . This new system overcomes some of the shortcomings of traditional Escherichia coli-based protein expression systems, particularly the propensity of such systems to form inclusion bodies during high-level expression . Using a proteomics approach, we identified promoters that can be induced by simple process parameters or medium compositions in high-density cell culture or shake flasks, respectively . By combining newly developed molecular biological tools with a high-cell-density fermentation process, we were able to produce high levels (>1 g/liter) of soluble, active organophosphohydrolase, a model enzyme prone to inclusion body formation in E . coli. Zh Mikrobiol Epidemiol Immunobiol, 2002 Jul-Aug, (4), 60 - 2 {Persistent properties of protozoa-associated bacteria}; Plotnikov AO et al.; The species structure and persistent properties (antilysozyme and antihistone activity) of bacteria forming associations with protozoa is revealed . Among them, 68.9% of the isolates were enterobacteria, the remaining organisms belonged to the families Aeromonas, Alcaligenes, Pseudomonas, Vibrio, etc . Within the family Enterobacteriaceae bacteria of the Escherichia group prevailed . 50.4% of the isolates were found to have antilysozyme activity and 97%--antihistone activity . The level of persistent properties in the representatives of allochthonous microflora was higher than that in the representatives of autochthonous microflora . In addition to antilysozyme activity antihistone activity was noted in protozoa-associated bacteria, which could be of importance for the formation of symbiotic links in natural associations . These data may be used in sanitary and hygienic practice for microecological monitoring of the environment. Biodegradation, 2002, 13(2), 141 - 7 Degradation of aliphatic polyester films by commercially available lipases with special reference to rapid and complete degradation of poly(L-lactide) film by lipase PL derived from Alcaligenes sp; Hoshino A et al.; Commercial lipases were examined for their degradation efficiency of aliphatic polyester films . In 100 days immersion of polyester films in lipase solutions at 37 degrees C at pH 7.0, Lipase Asahi derived from Chromobacterium viscosum degraded polybutylene succinate-co-adipate (PBSA), poly (e-caprolactone) (PCL) and polybutylene succinate (PBS), and Lipase F derived from Rhizopus niveus degraded PBSA and PCL during 4-17 days . Lipase F-AP15 derived from Rhizopus orizae could degrade PBSA in 22 days . In these cases, PBS and PBSA were mainly degraded to dimers, whereas PCL was mainly degraded to monomers . Only poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHB/V) and poly (L-lactide) (PLA) were not degraded in the experiments . However, PLA degraded completely at 55 degrees C, pH 8.5 with Lipase PL during 20 days . This result could be explained with the sequential reactions of the chemical hydrolysis of the polymer to oligomers at higher pH and temperature, and the succeeding enzymatic hydrolysis of oligomers to the monomers. Environ Pollut, 2002, 120(3), 691 - 700 Runoff rates, chemical speciation and bioavailability of copper released from naturally patinated copper; Karlen C et al.; The release of copper, induced by atmospheric corrosion, from naturally patinated copper of varying age (0 and 30 years) has been investigated together with its potential ecotoxic effect . Results were generated in an interdisciplinary research effort in which corrosion science and ecotoxicology aspects were combined . The aim of the investigation was to elucidate the situation when copper-containing rainwater leaves a roof in terms of runoff rate, chemical speciation, bioavailability and ecotoxicity effects . Data have been collected during a three-year field exposure conducted in the urban environment of Stockholm, Sweden . The potential environmental effects have been evaluated using a combination of a copper specific biosensor test with the bacterium Alcaligenes eutrophus and the conventional 72-h growth inhibition test with the green alga Raphidocelis subcapitata . The results show annual runoff rates between 1.0 and 1.5 g/m2 year for naturally patinated copper of varying age . The runoff rate increased slightly with patina age, which mainly is attributed to the enhanced first flush effect observed on thicker patina layers . The total copper concentration in investigated runoff samplings ranged from 0.9 to 9.7 mg/l . Both computer modeling and experimental studies revealed that the majority (60-100%) of released copper was present as the free hydrated cupric ion, Cu(H2O)6(2+), the most bioavailable copper species . However, other copper species in the runoff water, such as, e.g . Cu(OH)+ and Cu2(OH)2(2+), were also bioavailable . The copper-containing runoff water, sampled directly after release from the roof, caused significant reduction in growth rate of the green alga . It should be emphasized that the results describe the runoff situation immediately after release from the copper roof and not the real environmental ecotoxicity . Therefore the data should only be used as an initial assessment of the potential environmental effect of copper runoff from building applications . Future risk assessments should also consider dilution effects of copper, changes in its chemical speciation and bioavailability during environmental entry, and type and sensitivity of the receiving ecosystem. Eur J Biochem, 2002 Oct, 269(19), 4868 - 78 Identification and characterization of a new gene from Variovorax paradoxus Iso1 encoding N-acyl-D-amino acid amidohydrolase responsible for D-amino acid production; Lin PH et al.; An N-acyl-d-amino acid amidohydrolase (N-D-AAase) was identified in cell extracts of a strain, Iso1, isolated from an environment containing N-acetyl-d-methionine . The bacterium was classified as Variovorax paradoxus by phylogenetic analysis . The gene was cloned and sequenced . The gene consisted of a 1467-bp ORF encoding a polypeptide of 488 amino acids . The V . paradoxusN-D-AAase showed significant amino acid similarity to the N-acyl-d-amino acid amidohydrolases of the two eubacteria Alcaligenes xylosoxydans A-6 (44-56% identity), Alcaligenes facelis DA1 (54% identity) and the hyperthermophilic archaeon Pyrococcus abyssi (42% identity) . After over-expression of the N-D-AAase protein in Escherichia coli, the enzyme was purified by multistep chromatography . The native molecular mass was 52.8 kDa, which agreed with the predicted molecular mass of 52 798 Da and the enzyme appeared to be a monomer protein by gel-filtration chromatography . A homogenous protein with a specific activity of 516 U.mg-1 was finally obtained . After peptide sequencing by LC/MS/MS, the results were in agreement with the deduced amino acid sequence of the N-D-AAase . The pI of the enzyme was 5.12 and it had an optimal pH and temperature of 7.5 and 50 degrees C, respectively . After 30 min heat treatment at 45 degrees C, between pH 6 and pH 8, 80% activity remained . The N-D-AAase had higher hydrolysing activity against N-acetyl-d-amino acid derivates containing d-methionine, d-leucine and d-alanine and against N-chloroacetyl-d-phenylalanine . Importantly, the enzyme does not act on the N-acetyl-l-amino acid derivatives . The enzyme was inhibited by chelating agents and certain metal ions, but was activated by 1 mm of Co2+ and Mg2+ . Thus, the N-D-AAase from V . paradoxus can be considered a chiral specific and metal-dependent enzyme. Syst Appl Microbiol, 2002 Aug, 25(2), 183 - 8 Accumulation of poly(3-hydroxybutyrate) from octanoate in different pseudomonas belonging to the rRNA homology group I; Diard S et al.; It is admitted that one of the characteristics of pseudomonads is their inability to accumulate poly(3-hydroxybutyrate) . In this paper, we show that poly(3-hydroxyoctanoate) synthesis is restricted to Pseudomonas rRNA homology group I, which includes both fluorescent and nonfluorescent species . However, within the genus Pseudomonas, the P . aeruginosa complex can be subdivided into two groups: the "P . aeruginosa group", which includes P . aeruginosa, P . alcaligenes, P . citronellolis, P . mendocina, produce poly(3-hydroxyoctanoate) from octanoate and the "P . oleovorans group" which includes the type strain of P . oleovorans, P . pseudoalcaligenes and two Pseudomonas sp., produce poly(3-hydroxybutyrate) during cultivation on octanoate . Strain GPo1 (ATCC 29347) formely identified as P . oleovorans and known to produce various medium-side-chain PHAs such as poly(3-hydroxyoctanoate) has been reclassified in the P . putida complex. Prikl Biokhim Mikrobiol, 2002 Jul-Aug, 38(4), 401 - 4 {Use of maleic acid by mixed cultures of microorganisms}; Safronova IIu et al.; In a mixed batch culture, Alcaligenes xylosoxidans subsp . xylosoxidans 260 transformed maleic acid into malic acid . Bacillus subtilis 271 used malic acid as a substrate, thus stimulating further transformation of maleic acid . Both bacterial cultures dissociated with the formation of R, S, and M forms . At a concentration of 5.0 g/l, maleic acid was utilized maximally by RS and SS forms of the association A . xylosoxidans and Bacillus subtilis . At concentrations 15.0 and 25.0 g/l, maleic acid was utilized maximally by SS and MS forms of the mixed culture, respectively . Association of bacteria A . xylosoxidans and B . subtilis was not stable under flow conditions water. J AOAC Int, 2002 Jul-Aug, 85(4), 917 - 24 Characterization of copolymer hydroxybutyrate/hydroxyvalerate from saponified vernonia, soybean, and "spent" frying oils; Saeed KA et al.; Poly(beta-hydroxyalkanoate)s (PHAs) were biosynthesized by Ralstonia eutropha (formerly known as Alcaligenes eutrophus) by using saponified soybean, vernonia, and "spent" frying oils . These PHAs were isolated and characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS), gas chromatography/mass spectrometry (GC/MS), proton nuclear magnetic resonance spectrometry (1H NMR), and 2-dimensional homonuclear (1H-1H) correlation spectroscopy (COSY) . The analytical results revealed that the PHAs produced from saponified vernonia and soybean oils were copolymers of hydroxybutyrate (HB) and hydroxyvalerate (HV), that is, P(HB/HV)s, whereas the saponified "spent" frying oil produced only poly(beta- hydroxybutyrate) (PHB) homopolymer . MALDI-MS, GC/MS, and NMR independently confirmed the composition of the PHAs . Saponified soybean oil and vernonia oil PHAs contained approximately 4 and 1% HV units, respectively . For comparison, commercial PHB and P(HB/HV), produced by R . eutropha by using glucose and a cosubstrate of glucose and propionic acid, respectively, as carbon sources, were similarly characterized. Zh Mikrobiol Epidemiol Immunobiol, 2002 May-Jun, (3), 60 - 3 {Intestinal microflora in rats under the conditions of a chronic toxic lesion of the liver}; Sozinov AS et al.; Intestinal microflora in healthy rats and its changes under the conditions of experimental chronic toxic hepatitis were studied . The study revealed that in intact animals the microflora of the small intestine was represented by bacteria of the genera Escherichia, Enterobacter, Moraxella, Alcaligenes, Staphylococcus, Streptococcus . Bacteria of the genera Escherichia, Enterobacter, Moraxella, Alcaligenes, Staphylococcus, Corynebacterium and Clostridium were isolated from the large intestine . No bacteria were found in the systemic blood, the contents of the portal vein, as well as in the liver parenchyma and the mesenterial lymph nodes . As the result of dysbiosis induced by the introduction of kanamycin and in chronic hepatitis caused by carbon tetrachloride the sharp decrease in the species composition of microbial communities (up to 2-3 species) in the small intestine and was observed along with penetration of bacteria into the blood stream, the mesenterial lymph nodes and the liver parenchyma . The tendency towards the restoration of the quantitative and qualitative microflora composition was noted following administration into experimental animals of bactisubtil and amixin--an inductor of interferonogenesis. Chemosphere, 2002 Jun, 47(10), 1073 - 80 Bioavailability of zinc in runoff water from roofing materials; Heijerick DG et al.; Corrosion and runoff from zinc-coated materials and outdoor structures is an important source for the dispersion of zinc in the environment . Being part of a large inter-disciplinary research project, this study presents the bioavailability of zinc in runoff water immediately after release from the surface of 15 different commercially available zinc-based materials exposed to the urban environment of Stockholm, Sweden . Runoff water was analysed chemically and evaluated for its possible environmental impact, using both a biosensor test with the bacteria Alcaligenes eutrophus (Biomet) and the conventional 72 h growth inhibition test with the green alga Raphidocelis subcapitata . Chemical speciation modelling revealed that most zinc (94.3-99.9%) was present as the free Zn ion, the most bioavailable speciation form . These findings were confirmed by the results of the biosensor test (Biomet) which indicated that all zinc was indeed bioavailable . Analysis of the ecotoxicity data also suggested that the observed toxic effects were due to the presence of Zn2+ ions . Finally, regression analysis showed that, for this type of runoff samples, the rapid screening biosensor was capable of predicting (a) the total amount of zinc present in the runoff samples (R2 of 0.93-0.98; p < 0.05) and (b) the observed 72 h-EbC50s (R2 of 0.69-0.97; p < 0.05). Pediatr Pulmonol, 2002 Aug, 34(2), 101 - 4 Alcaligenes infection in cystic fibrosis; Tan K et al.; The aim of this study was to investigate the effect of chronic Alcaligenes species infection of the respiratory tract on the clinical status of patients with cystic fibrosis . We conducted a retrospective case-controlled study . The microbiological records of all patients attending the Leeds Regional Pediatric and Adult Cystic Fibrosis Units from 1992-1999 were examined . Chronic Alcaligenes infection was defined as a positive sputum culture on at least three occasions over a 6-month period . These patients were compared with controls matched for age, gender, respiratory function, and Pseudomonas aeruginosa infection status . Respiratory function tests, anthropometric data, Shwachman-Kulczycki score, Northern chest x-ray score, intravenous and nebulized antibiotic treatment, and corticosteroid treatment were compared from 2 years before to 2 years after Alcaligenes infection.From a clinic population of 557, 13 (2.3%) fulfilled the criteria for chronic infection . The median age at acquisition of infection was 17.2 years (range, 6.5-33.6) . There was no significant difference in the changes of percentage predicted values for FEV(1), FVC, FEF(25-75), or Shwachman-Kulczycki and Northern chest x-ray scores, or in weight, height, and body mass index z-scores between Alcaligenes-infected cases and controls . There was also no significant difference in the use of antibiotics (intravenous and nebulized) or corticosteroids (inhaled and oral).We conclude that in our clinic, chronic infection with Alcaligenes species was uncommon . Chronically infected patients showed no excess deterioration in clinical or pulmonary function status from 2 years before to 2 years after primary acquisition . Aviakosm Ekolog Med, 2002, 36(2), 28 - 32 {Species composition and microbial contamination of atmospheric humidity condensate and potable water from the Mir water supply systems during Mir main missions 4 through 27}; Skuratov VM et al.; Analyzed were quantitative and qualitative characteristics, and processes of formation and transformation of microflora in the atmospheric humidity condensate (AHC) and potable water (PW) regenerated from the condensate by the long-operating system (SRV-K) aboard space station Mir . The paper presents data on species composition of microflora of AHC, and PW regenerated by SRV-K in the period from 1989 through to 1999 . The following microbial species were identified in AHC: Alcaligenes (14.8%), Bacillus (3.7%), Citrobacter (7.4%), Clostridium (11.1%), Enterobacter (7.4%), Proteus (7.4%), Pseudomonas (18.5%), and Staphylococcus (100%) . There were two PW cocks in SRV-K--"hot" and "cold" . "Hot" PW microflora included Alcaligenes (15%), Bacillus (5%), Citrobacter (20%), Clostridium (5%), Enterobacter (15%), Proteus (10%), Pseudomonas (15%), and Staphylococcus (75%) . "Cold" PW microflora included Aeromonas (14.3%), Alcaligenes (21.4%), Bacillus (7.1%), Citrobacter (7.1%), Clostridium (7.1%), Enterobacter (7.1%), Moraxella (7.1%), Pseudomonas (7.1%), and Staphylococcus (78.5%) . Microbial content averaged 8.9 x 10(2) +/- 1.3 (0.01-4) x 10(3) CFU/ml in condensate and 1.5 +/- 0.1 (0-3.0) CFU/ml in regenerated potable water. Folia Microbiol (Praha), 2002, 47(3), 247 - 54 Characterization of polychlorinated biphenyl-degrading bacteria isolated from contaminated sites in Czechia; Totevova S et al.; Biphenyl-utilizing polychlorinated biphenyls (PCB)-degrading bacteria were isolated from sites highly contaminated by PCBs, and their degradation abilities were determined using GC for typical commercial PCB mixtures (Delor 103 and Delor 106) . Out of twelve strains which utilized biphenyl as a sole source of carbon and energy, strains Pseudomonas alcaligenes KP2 and P . fluorescens KP12, characterized by the BIOLOG identification system and the NEFERM test, were shown to significantly co-metabolize the PCB mixture Delor 103 . DNA-DNA hybridization was used to compare both strains with well-known PCB-degraders Burkholderia cepacia strain LB400 and Ralstonia eutropha strain H850 . The strain KP12 employs the same meta-fission route for degradation of chlorobenzoates as a chlorobiphenyl degrader Pseudomonas cepacia P166 . Both isolates KP2 and KP12 belong to different phylogenetic groups, which indicates that the same geographical location does not ensure the same ancestor of degradative enzymes . We confirmed that also highly chlorinated and the most toxic congeners, which are contained in commercial PCB mixtures, can be biotransformed by members of indigenous bacterial-soil community under aerobic conditions. Biodegradation, 2001, 12(6), 393 - 400 Fluoranthene degradation in Pseudomonas alcaligenes PA-10; Gordon L et al.; Pseudomonas alcaligenes strain PA-10 degrades the four-ring polycyclic aromatic hydrocarbon fluoranthene, co-metabolically . HPLC analysis of the growth medium identified four intermediates, 9-fluorenone-1-carboxylic acid; 9-hydroxy-1-fluorene carboxylic acid; 9-fluorenone and 9-fluorenol, formed during fluoranthene degradation . Pre-exposure of PA-10 to 9-fluorenone-1-carboxylic acid and 9-hydroxy-1-fluorene-carboxylic acid resulted in increases in fluoranthene removal, while pre-exposure to 9-fluorenone and 9-fluorenol resulted in a decrease in fluoranthene degradation . The rate of indole transformation was similarly affected by pre-exposure to these metabolic intermediates, indicating a link between fluoranthene degradation and indigo formation in this strain. Microb Ecol, 2002 Mar, 43(2), 280 - 9 Epub 2002 Feb 20. Diversity of Pseudomonas spp . isolated from rice rhizosphere populations grown along a salinity gradient; Rangarajan S et al.; Along the coastline of Tamil Nadu, five sites were chosen to assess the diversity of Pseudomonas populations isolated from rice (Oryza sativa) cultivated along a salinity gradient . One of these sites was under organic farming while the other four were under inorganic farming . A total of 256 Pseudomonas strains isolated from these five sites were analyzed using both phenotypic (substrate utilization patterns and antibiotic resistance assay) and genotypic (PCR-RFLP of 16S rDNA) characteristics . The results derived from this study indicate that soil salinity affects rhizosphere Pseudomonas populations . It was observed that increasing salinity led to decreasing diversity . Fluorescent pseudomonads were the dominant species found in the non-saline site, while in the saline sites they were replaced by salt-tolerant species, in particular Pseudomonas alcaligenes and P . pseudoalcaligenes . An interesting observation was the increase in diversity found in the saline site under organic farming . Organic farming was found to be capable of mitigating the harmful effects of saline stress to a large extent, and restoring the Pseudomonas diversity, thereby making it comparable with the diversity encountered in the non-saline site. Microb Ecol, 2002 Mar, 43(2), 199 - 216 Epub 2002 Jan 22. Autecological properties of 3-chlorobenzoate-degrading bacteria and their population dynamics when introduced into sediments; Bott TL et al.; Ecologically significant properties of wild-type and genetically engineered bacteria capable of degrading 3-chlorobenzoate (3-CB) were compared in the laboratory, and isolates were introduced into streambed sediments in microcosms to observe their population dynamics . 3-CB metabolism, growth on algal extract, temperature optima, and ingestion by protozoa were ecological properties considered relevant to the persistence of these bacteria if introduced into nature . Cell-specific Vmax for 3-CB metabolism and cell-specific mineralization rates each spanned approximately 2 orders of magnitude, but isolates did not rank consistently . The Ks for 3-CB metabolism for Alcaligenes sp . BR60 was approximately 40-fold lower than the mean value for the other isolates, which differed only approximately 4-fold among themselves . All isolates grew on an algal extract nearly as well as on tryptone-yeast extract, implying potential for survival on natural metabolic substrates in situ . Most isolates had temperature optima that were 3-15 degrees C higher than maximum stream water temperature (22 degrees C) . Ciliates preferentially ingested P . acidovorans M3GY, and either P . putida RC-4(pSI30) or its parent strain were least preferred, but microflagellates did not exhibit consistent preferences . Fluorescent antibodies were prepared against isolates to permit detection of target cells in natural communities . In three different microcosm experiments the cell densities of introduced isolates declined over a period of days . In one experiment, 3-CB additions (100 mg/L) led to increases of P . alcaligenes C-0 and P . acidovorans M3GY cell densities within 1 day, although P . putida RC-4(pSI30) took 4 days . In a second experiment, the persistence of P . putida RC-4(pSI30) and its parent strain P . putida RC-4 were compared and rates of initial population decline were not statistically different . 3-CB addition stimulated the growth of other organisms while densities of the P . putida strains further declined . In a third experiment exposure to 100 mg 3-CB/L slowed the rate of decline of P . acidovorans M3GY densities compared to a 10 mg/L concentration or unamended control . Competition with the native flora was a significant factor affecting the persistence of introduced 3-CB degraders. Antimicrob Agents Chemother, 2002 Jun, 46(6), 2014 - 6 Detection of a variant metallo-beta-lactamase, IMP-10, from two unrelated strains of Pseudomonas aeruginosa and an alcaligenes xylosoxidans strain; Iyobe S et al.; The gene bla(IMP-10) of a variant metallo-beta-lactamase, IMP-10, had a single base replacement of G by T at nucleotide 145, which led to an amino acid alteration of Val49 to Phe compared to the IMP-1 enzyme, indicating that IMP-10 was a point mutation derivative of IMP-1 . Highly purified enzymes revealed that IMP-10 was different from IMP-1 in its extremely low hydrolyzing activities for penicillins, such as benzylpenicillin, ampicillin, and piperacillin. Can J Microbiol, 2002 Mar, 48(3), 189 - 99 Response of spring rape (Brassica napus var . oleifera L.) to inoculation with plant growth promoting rhizobacteria containing 1-aminocyclopropane-1-carboxylate deaminase depends on nutrient status of the plant; Belimov AA et al.; Responses of rape (Brassica napus var . oleifera L.) to inoculation with plant growth promoting rhizobacteria, Pseudomonas putida Am2, Pseudomonas putida Bm3, Alcaligenes xylosoxidans Cm4, and Pseudomonas sp . Dp2, containing 1-aminocyclopropane-l-carboxylate (ACC) deaminase were studied using growth pouch and soil cultures . In growth pouch culture, the bacteria significantly increased root elongation of phosphorus-sufficient seedlings, whereas root elongation of phosphorus-deficient seedlings was not affected or was even inhibited by the bacteria . Bacterial stimulation of root elongation of phosphorus-sufficient seedlings was eliminated in the presence of a high ammonia concentration (1 mM) in the nutrient solution . Bacterial effects on root elongation of potassium-deficient and potassium-sufficient seedlings were similar . The bacteria also decreased inorganic phosphate content in shoots of potassium- and phosphorus-sufficient seedlings, reduced ethylene production by phosphorus-sufficient seedlings, and inhibited development of root hairs . The effects of treatment with Ag+, a chemical inhibitor of plant ethylene production, on root elongation, ethylene evolution, and root hair formation were similar to bacterial treatments . The number of bacteria on the roots of phosphorus-deficient seedlings was not limited by phosphorus deficiency . In pot experiments with soil culture, inoculation of seeds with bacteria and treatment with aminoethoxyvinylglycine, an inhibitor of ethylene biosynthesis in plants, increased root and (or) shoot biomass of rape plants . Stimulation of plant growth caused by the bacteria was often associated with a decrease in the content of nutrients, such as P, K, S, Mo, and Ba, in shoots, depending on the strain used . The results obtained show that the growth-promoting effects of ACC-utilizing rhizobacteria depend significantly on the nutrient status of the plant. J Biol Chem, 2002 Jun 28, 277(26), 23725 - 32 Epub 2002 Apr 18. The X-ray structure of ferric Escherichia coli flavohemoglobin reveals an unexpected geometry of the distal heme pocket; Ilari A et al.; The x-ray structure of ferric unliganded lipid-free Escherichia coli flavohemoglobin has been solved to a resolution of 2.2 A and refined to an R-factor of 19% . The overall fold is similar to that of ferrous lipid-bound Alcaligenes eutrophus flavohemoglobin with the notable exception of the E helix positioning within the globin domain and a rotation of the NAD binding module with respect to the FAD-binding domain accompanied by a substantial rearrangement of the C-terminal region . An inspection of the heme environment in