Microbial Degradation of Nicotinamide by a Strain Alcaligenes sp. P156.

A novel Alcaligenes sp. strain P156, which can utilize nicotinamide as its sole source of carbon, nitrogen and energy, was enriched and isolated from soil in a solid waste treatment plant. Aerobic growth and degradation with nicotinamide were characterized. Seven nicotinamide degradation-related genes were obtained by sequence alignment from the genome sequence of strain P156. Four genes, designated naaA, naaD, naaE and naaF, were cloned and heterologously expressed. Nicotinamide degradation is initiated by deamination to form nicotinic acid catalyzed by the nicotinamidase NaaA, which shares highest amino acid sequence identity (27.2%) with nicotinamidase from Arabidopsis thaliana. Nicotinic acid is converted to 6-hydroxynicotinic acid, which is further oxidized to 2,5-dihydroxypyridine (2,5-DHP). 2,5-DHP is then transformed to a ring-cleavage product, N-formylmaleamic acid, by an Fe2+ dependent dioxygenase NaaD. N-formylmaleamic acid is transformed to fumaric acid through maleamic acid and maleic acid by NaaE and NaaF, respectively. To our knowledge, this is the first report of the complete microbial degradation of nicotinamide in bacteria. Nicotinamide is considered as a model compound for the study of microbial degradation of pyridinic compounds, and the nicotinamide degrading related genes in strain P156 were distributed differently from the reported similar gene clusters. Therefore, this study contribute to the knowledge on the degradation of pyridinic compounds.

Alcaligenes endophyticus sp. nov., isolated from roots of Ammodendron bifolium.

A Gram-stain-negative, rod-shaped, motile bacterium, designated AER10T, was isolated from the roots of Ammodendron bifolium collected from Takeermohuer desert in Xinjiang Uygur Autonomous Region, northwestern China. Growth was found to occur from 10 to 45 °C, at pH 5.0-9.0, and could tolerate up to 10 % (w/v) NaCl. 16S rRNA gene sequence result indicated that the strain AER10T belongs to the genus Alcaligenes and was closely related to Alcaligenes aquatilis (98.4 %), Alcaligenes faecalissubsp. parafaecalis (98.4 %), Alcaligenes faecalissubsp. faecalis (98.1 %) and Alcaligenes faecalissubsp. phenolicus (97.9 %). However, the DNA-DNA hybridization values between the strain AER10T and the above strains were less than the threshold value (below 70 %) for the delineation of genomic species. The DNA G+C content was 53.3 mol%. Ubiquinone-8 (Q-8) was the only quinone system present. The major fatty acids were summed feature 8 (C18 : 1ω7c, 25 %), C16 : 0 (24.2 %), summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c, 19.3 %) and cyclo-C17 : 0 (10.5 %). The polar lipid profile of the strain AER10T consists of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylserine, two unidentified aminolipids and five unknown polar lipids. On the basis of the evidence presented in this study, strain AER10T is a representative of a novel species in the genus Alcaligenes, for which the name Alcaligenes endophyticus sp. nov. is proposed. The type strain is AER10T (=DSM 100498T=KCTC 42688T).

Characteristics and metabolic pathway of Alcaligenes sp. TB for simultaneous heterotrophic nitrification-aerobic denitrification.

A novel heterotrophic nitrification-aerobic denitrification bacterium, Alcaligenes sp. TB (GenBank accession no. JQ044686), was isolated from a rotating drum biofilter for NO removal. Its characteristics and metabolic pathway for NO removal were comprehensively investigated. Experimental results showed that the nitrification and denitrification efficiency reached 99.42 and 96.44 %, respectively, after 44 h under the conditions of pH 7.2, 30 °C, and 120 rpm. The tests with the addition of Pb2+ and Na2WO4 as the reductase inhibitor revealed that nitrite was the key intermediate to produce the nitrogen gas as the final product in the simultaneous heterotrophic nitrification and denitrification by strain TB. Based on the experimental results, the metabolic pathway of strain TB has been proposed that it carries out shortcut/complete simultaneous nitrification and denitrification with nitrite as an intermediate and nitrogen gas as a final product. The two potential metabolic pathways existing in strain TB can be described as NH4+ â†’ NH2OH â†’ NO2- â†’ N2O â†’ N2 and NH4+ â†’ NH2OH â†’ NO2- â†’ NO3- â†’ NO2- â†’ N2O â†’ N2. This work indicates that the strain TB may be a good candidate for the denitrification of the sewage.

A heavy-metal tolerant novel bacterium, Alcaligenes pakistanensis sp. nov., isolated from industrial effluent in Pakistan.

Two strains, NCCP-650(T) and NCCP-667, were isolated from industrial effluent and their taxonomic positions were investigated using a polyphasic taxonomic approach. The strains were found to be Gram-stain negative, strictly aerobic, motile short rods, which are tolerant to heavy-metals (Cr(+2), As(+2), Pb(+2) and Cu(+2)). Cells were observed to grow at a temperature range of 10-37 °C (optimal 25-33 °C), pH range of 5.5-10.0 (optimal 6.5-7.5) and can tolerate 0-7 % NaCl (w/v) (optimum 0-1 %) in tryptic soya agar medium. Sequencing of the 16S rRNA gene and two housekeeping genes, gyrB and nirK, of the isolated strains revealed that both strains belong to the Betaproteobacteria showing highest sequence similarities with members of the genus Alcaligenes. The chemotaxonomic data [major quinones as Q-8; predominant cellular fatty acids as summed features 3 (C16 :1 ω7c/iso-C15 :0 2OH) and C16:0 followed by Summed features 2 (iso-C16 :1 I/C14 :0 3OH), C17:0 Cyclo and C18:1 ω7c; major polar lipids as diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and one unidentified aminolipid] also supported the affiliation of the isolated strains with the genus Alcaligenes. DNA-DNA hybridizations between the two strains and with closely related type strains of species of the genus Alcaligenes confirmed that both isolates belong to a single novel species within the genus Alcaligenes. On the basis of phylogenetic analyses, physiological, biochemical characteristics and DNA-DNA hybridization, the isolated strains can be differentiated from established Alcaligenes species and thus represent a novel species, for which the name Alcaligenes pakistanensis sp. nov. is proposed with the type strain NCCP-650(T) (=LMG 28368(T) = KCTC42083(T) = JCM 30216(T)).

Characterization of bacterial isolates from rubber dump site and their use in biodegradation of isoprene in batch and continuous bioreactors.

Bacterial isolates from contaminated soil of a waste rubber dumping site were isolated and characterized using biochemical and molecular approaches. Isoprene degradation kinetics in batch mode (isoprene concentration: 100-1000 ppm) revealed the degradation efficiency of isolates as: Pseudomonas sp. (83%)>Alcaligenes sp. (70%)>Klebsiella sp. (68.5%). The most efficient isolate Pseudomonas sp. was finally inoculated in a specifically designed bioreactor system comprising a bioscrubber and a biofilter packed with polyurethane foam connected in series. The bioscrubber and biofilter units when operated in a series showed more than 90% removal efficiency up to the inlet loading rate (IL) of 371.1g/m(3)/h. Maximum elimination capacity (EC) of biofilter was found to be an order of magnitude greater than that for bioscrubber. Oxidative cleavage of the double bond of isoprene has been revealed through IR spectra of the leachate.

Microbial abundance and community composition influence production performance in a low-temperature petroleum reservoir.

Enhanced oil recovery using indigenous microorganisms has been successfully applied in the petroleum industry, but the role of microorganisms remains poorly understood. Here, we investigated the relationship between microbial population dynamics and oil production performance during a water flooding process coupled with nutrient injection in a low-temperature petroleum reservoir. Samples were collected monthly over a two-year period. The microbial composition of samples was determined using 16S rRNA gene pyrosequencing and real-time quantitative polymerase chain reaction analyses. Our results indicated that the microbial community structure in each production well microhabitat was dramatically altered during flooding with eutrophic water. As well as an increase in the density of microorganisms, biosurfactant producers, such as Pseudomonas, Alcaligenes, Rhodococcus, and Rhizobium, were detected in abundance. Furthermore, the density of these microorganisms was closely related to the incremental oil production. Oil emulsification and changes in the fluid-production profile were also observed. In addition, we found that microbial community structure was strongly correlated with environmental factors, such as water content and total nitrogen. These results suggest that injected nutrients increase the abundance of microorganisms, particularly biosurfactant producers. These bacteria and their metabolic products subsequently emulsify oil and alter fluid-production profiles to enhance oil recovery.

Development of a PCR-based method for monitoring the status of Alcaligenes species in the agricultural environment.

To analyze the status of the genus Alcaligenes in the agricultural environment, we developed a PCR method for detection of these species from vegetables and farming soil. The selected PCR primers amplified a 107-bp fragment of the 16S rRNA gene in a specific PCR assay with a detection limit of 1.06 pg of pure culture DNA, corresponding to DNA extracted from approximately 23 cells of Alcaligenes faecalis. Meanwhile, PCR primers generated a detectable amount of the amplicon from 2.2×10(2) CFU/ml cell suspensions from the soil. Analysis of vegetable phylloepiphytic and farming soil microbes showed that bacterial species belonging to the genus Alcaligenes were present in the range from 0.9×10(0) CFU per gram (or cm(2)) (Japanese radish: Raphanus sativus var. longipinnatus) to more than 1.1×10(4) CFU/g (broccoli flowers: Brassica oleracea var. italic), while 2.4×10(2) to 4.4×10(3) CFU/g were detected from all soil samples. These results indicated that Alcaligenes species are present in the phytosphere at levels 10-1000 times lower than those in soil. Our approach may be useful for tracking or quantifying species of the genus Alcaligenes in the agricultural environment.

[Isolation, identification and characterization of a high-efficient heterotrophic nitrifier].

OBJECTIVE: In order to improve the rate of the heterotrophic nitrification, we screened and identified a high-efficient heterotrophic nitrifier, as well as studied its nitrification characteristics and nitrification conditions. METHODS: We obtained activated sludge samples from sewage and chemical fertilizer factories and farmland. We then utilized sodium citrate and ammonium chloride as carbon and nitrogen source. We used methods including domestication, gradient dilution of domestication liquid, isolation from streaking plate and color indicator as rapid nitrification detection. Finally a high-efficient heterotrophic nitrifier was obtained. We identified this strain according to its physiological, biochemical properties and the sequence analysis of 16S rDNA. After inoculating the strain into artificial ammonia-nitrogen wastewater, changes of nitrogen compounds were measured in order to understand the nitrification characteristics. Nitrification condition was also optimized by changing the carbon source, dissolved oxygen, C/N ratio, temperature and pH of the medium. RESULTS: The heterotrophic nitrifier was a gram-negative bacilli. It neither fermented glucose, nor produced indole. Oxidase and catalase tests were positive. It could produce alkali if organic salt was provided. The strain shared 99.7% sequence identity of its 16S rDNA with ES-SDK-3 of Alcaligenes sp. In the artificial wastewater with 182.30 mg/L ammonia nitrogen as initial concentration, the removal efficiency by the strain was 99. 8% after 30h cultivation. The average nitrogen removal rate was 9. 61 mg-N/L/h in its exponential phase. It produced almost no NO(2-)-N and NO(3-)-N in the entire nitrification process. The optimal carbon source is sodium citrate. Higher dissolved oxygen and C/N ratio favor its nitrification. When temperature is ranged from 30 degrees C to 35 degrees C and pH is ranged from 5.0 to 9.0, it can completely remove ammonia nitrogen. CONCLUSIONS: The strain was identified as Alcaligenes genus, and named as Alcaligenes sp. HN-S. Our research confirmed that the Alcaligenes sp. HN-S had significant advantages over heterotrophic nitrifiers that were screened previously with aspect of ammonia nitrogen removal rate. The research of its nitrification condition definitely provided necessary theory support for a new biology process to remove nitrogen with high efficiency.

Diversity of microbes and potential exoelectrogenic bacteria on anode surface in microbial fuel cells.

Single-chamber microbial fuel cells (MFCs), inoculated with anaerobic sludge and continuously run with two kinds of organic wastewater influents, were systemically investigated. The diversity of microbes, determined by 16S rDNA analysis, was analyzed on three anodes under different conditions. One anode was in a closed circuit in synthetic wastewater containing glucose. The other two anodes, in open or closed circuits, were fed effluent from an anaerobic reactor treating starch wastewater. The chemical oxygen demand (COD) removal efficiency was about 70%, and the exported voltages were about 450 mV. The 16S rDNA molecular clones of microbes on anode surfaces showed significant changes in Eubacterial structure under different conditions. gamma-Proteobacteria and the high G+C gram-positive groups were predominant in the synthetic wastewater, while epsilon-Proteobacteria predominated in the anaerobic reactor effluent. Known exoelectrogenic bacterial species composition also changed greatly depending on substrate. On the artificial substrate, 28% of the bacterial sequences were affiliated with Aeromonas, Pseudomonas, Geobacter, and Desulfobulbus. On the anaerobic effluent, only 6% were affiliated with Geobacter or Clostridium. Because only a few exoelectrogenic bacteria from MFCs have been directly isolated and studied, we compared the community structures of two bacterial anodes, in open and closed circuits, under the same substrate of anaerobic effluent in order to identify additional exoelectrogenic bacterial strains. Alcaligenes monasteriensis, Comamonas denitrificans, and Dechloromonas sp. were found to be potential exoelectrogenic bacteria worthy of further research.

[Alcaligenes xylosoxidans bacteremia in a patient with acute lymphoblastic leukemia]. / Akut lenfoblastik lösemili bir olguda Alcaligenes xylosoxidans bakteriyemisi.

Alcaligenes xylosoxidans which is an aerobic, non-fermentative gram-negative bacillus found in aqueous environments and human flora, can lead to opportunistic infections. It causes infections in elderly, immunocompromised patients, patients with chronic disorders or premature infants. In this report, a case of A. xylosoxidans bacteremia that developed in a child with acute lymphoblastic leukemia (ALL) was presented. Four-years-old male patient under ALL induction therapy was admitted with symptoms of lethargy, headache, somnolence, and fever (39 degrees C). Cerebrospinal fluid, blood, throat and urine cultures were taken from the patient and empirical treatment with sulbactam cefoperazon and amikacin was initiated. Blood cultures in BacT Alert 3D (Bio Merieux, France) revealed the growth of a gram-negative coccobacillus. The agent which was non-fermentative, indol and H2S negative, was identified as A. xylosoxidans by API 20 NE (Bio Merieux, France). Since fever continued under the current antibiotic treatment, the therapy was switched to imipenem (90 mg/kg 3x/day) and the patient's condition improved markedly after 24 hours. Disc diffusion susceptibility testing of the isolate revealed that it was resistant to ampicillin, cephalothin, cefuroxime, cefoxitin, cefotaxime, amikacin, netilmicin and gentamicin; susceptible to amoxicillin clavulanate, piperacillin tazobactam, seftazidime, cefepime, imipenem and ciprofloxacin. Following 14 days of imipenem therapy, the patient recovered and discharged from the hospital on routine follow-up. It is important to consider A. xylosoxidans as a possible causative agent particularly in the infections that develop in high risk patients at oncology, dialysis and neonatal intensive care units.

[Production of biodemulsifier by Alcaligenes sp. XJ-T-1 with waste frying oil].

As a new member of demulsifier family, biodemulsifier is applied in oil-water emulsion breaking. A strain, XJ-T-1, was isolated from petroleum-contaminated soil and identified as Alcaligenes sp.. Its physicochemical properties, demulsification capability and utilization of waste oil were further investigated. With waste frying oil (WFO) as carbon source, the demulsifier produced by Alcaligenes sp. showed high demulsifying capability. Moreover, the production of demulsifier was 4.6 times of that generated with paraffine as the carbon source. The CMC(-1) of biodemulsifier produced by paraffine and waste frying oil was achieved at 10 and 20, respectively. The biodemulsifier cultured with paraffine as the carbon source achieved 96% and 50% of emulsion breaking ratio in W/O (water in oil) and O/W (oil in water) model emulsion, while the demulsifier cultured on waste frying oil II as carbon source exhibited 97.8% and 65% demulsification ratio in the two model emulsions correspondingly. From the dynamic change of kerosene breaking ratio, emulsion breaking ratio and water breaking ratio during demulsification process, it was found that this biodemulsifier reacted with the continuous phase of the emulsion prior to its reaction with the dispersed phase. It was identified the valid part of biodemulsifier produced by WFO II was lipopeptide by TLC and IR.

Evaluation of screening methods for demulsifying bacteria and characterization of lipopeptide bio-demulsifier produced by Alcaligenes sp.

In this paper, surface tension measurement, oil-spreading test and blood-plate hemolysis test were attempted in the screening of demulsifying bacteria. After the comparison to the screening results obtained in demulsification test, 50 mN/m of surface tension of culture was proposed as a preliminary screening standard for potential demulsifying bacteria. For the identification of efficient demulsifying strains, surface tension level was set at 40 mN/m. The detected strains were further verified in demulsification test. Compared to using demulsification test alone as screening method, the proposed screening protocol would be more efficient. From the screening, a highly efficient demulsifying stain, S-XJ-1, was isolated from petroleum-contaminated soil and identified as Alcaligenes sp. by 16S rRNA gene and physiological test. It achieved 96.5% and 49.8% of emulsion breaking ratio in W/O and O/W kerosene emulsion within 24h, respectively, and also showed 95% of water separation ratio in oilfield petroleum emulsion within 2h. The bio-demulsifier was found to be cell-wall combined. After soxhlet extraction and purification through silicon-gel column, the bio-demulsifier was then identified as lipopeptide biosurfactant by TLC and FT-IR.

Arsenite oxidation by Alcaligenes sp. strain RS-19 isolated from arsenic-contaminated mines in the Republic of Korea.

Arsenite [As(III)]-oxidizing bacteria play important roles in reducing arsenic [As] toxicity and mobility in As-contaminated areas. As-resistant bacteria were isolated from the soils of two abandoned mines in the Republic of Korea. The isolated bacteria showed relatively high resistances to As(III) up to 26 mM. The PCR-based 16S rRNA analysis revealed that the isolated As-resistant bacteria were close relatives to Serratia marcescensa, Pseudomonas putida, Pantoea agglomerans, and Alcaligenes sp. Among the five As-resistant bacterial isolates, Alcaligenes sp. strain RS-19 showed the highest As(III)-oxidizing activity in batch tests, completely oxidizing 1 mM of As(III) to As(V) within 40 h during heterotrophic growth. This study suggests that the indigenous bacteria have evolved to retain the ability to resist toxic As in the As-contaminated environments and moreover to convert the species to a less toxic form [e.g., from As(III) to As(V)] and also contribute the biogeochemical cycling of As by being involved in speciation of As.

[The spectrum of microbiological agents causing pulmonary MALT-type lymphomas. A 16S rRNA-based analysis of microbial diversity]. / Das Erregerspektrum pulmonaler MALT-Typ-Lymphome. Eine 16S-rRNA-basierte Analyse der mikrobiellen Diversität.

For several anatomical localisations of extranodal marginal zone B-cell lymphoma of MALT type (eMZBCL), an association with chronic inflammation caused by microbiological agents (e.g. Helicobacter pylori in the stomach) has been described. In the lung, a link between lymphomagenesis and a defined causative organism is still missing. A comprehensive diversity survey using 16S-rDNA library construction followed by restriction fragment length polymorphism (RFLP) analysis, sequencing, and phylogenetic tree construction was employed for nine cases each of BALT lymphoma and control lung tissues (normal foetal lung, pneumonitis, carcinoid). This highly sensitive method, hereafter termed SHARP screening allowed for identification of the entire bacterial population in the tissue in a cultivation-independent manner. It was noteworthy that in eight of the nine cases of BALT lymphoma, bacteria of the Alcaligenaceae family (Alcaligenes, Achromobacter, AKIW733), were detected, whereas none of the control cases showed the presence of these clades. 16S-rDNA library construction in combination with RFLP screening and phylogenetic analyses, hereafter described as SHARP screening, is a cultivation-independent tool for analysing the microbial environment in chronic inflammation processes giving rise to extranodal marginal zone B-cell lymphomas of MALT-type. Betaproteobacteria of the Alcaligenaceae family may be affiliated with and possibly involved in the lymphomagenesis of BALT lymphomas.

Preparation of (R)-(-)-mandelic acid and its derivatives from racemates by enantioselective degradation with a newly isolated bacterial strain Alcaligenes sp. ECU0401.

An enantioselective mandelate-degrading bacterium, Alcaligenes sp. ECU0401, was newly isolated from soil. By fed-batch culture, (R)-(-)-mandelic acid was successfully prepared in a 5-L fermenter with 32.8% isolated yield and >99.9% enantiomeric excesses (e.e.) from totally 3.04% (w/v) of racemic mandelic acid after 99 h of biotransformation. The optimal reaction pH and temperature were 6.5 and 30 degrees C, respectively. Using the resting cell as a biocatalyst for asymmetric degradation of racemic mandelic acid and chloro-substituted derivatives thereof, (R)-(-)-mandelic acid, (R)-(-)-o-chloromandelic acid, (S)-(+)-m-chloromandelic acid and (S)-(+)-p-chloromandelic acid were recovered with high analytic yields and excellent enantiomeric excesses (e.e. > 99.9%). (R)-(-)-Mandelic acid could also be obtained after 12 h of biotransformation with 41.5% isolated yield and >99.9% e.e.

Hydrogen cyanide-producing rhizobacteria kill subterranean termite Odontotermes obesus (Rambur) by cyanide poisoning under in vitro conditions.

The subterranean termite Odontotermes obesus is an important pest of the Indian subcontinent, causing extensive damage to major agricultural crops and forest plantation trees. Control of termites by strategies employing their parasites has limitations because they have evolved a complex social structure, immune responses, and adaptive behavior toward pathogen-infected individuals. Nonparasitic rhizobacteria that produce harmful metabolites might facilitate the biocontrol of termites. In the present investigation, three different species of hydrogen cyanide-producing rhizobacteria were tested for their potential to kill O. obesus. The three bacterial species were found to be effective in killing the termites under in vitro conditions.

Castellaniella gen. nov., to accommodate the phylogenetic lineage of Alcaligenes defragrans, and proposal of Castellaniella defragrans gen. nov., comb. nov. and Castellaniella denitrificans sp. nov.

Comparative 16S rRNA gene sequence analysis indicates that two distinct sublineages exist within the genus Alcaligenes: the Alcaligenes faecalis lineage, comprising Alcaligenes aquatilis and A. faecalis (with the three subspecies A. faecalis subsp. faecalis, A. faecalis subsp. parafaecalis and A. faecalis subsp. phenolicus), and the Alcaligenes defragrans lineage, comprising A. defragrans. This phylogenetic discrimination is supported by phenotypic and chemotaxonomic differences. It is proposed that the A. defragrans lineage constitutes a distinct genus, for which the name Castellaniella gen. nov. is proposed. The type strain for Castellaniella defragrans gen. nov., comb. nov. is 54PinT (=CCUG 39790T = CIP 105602T = DSM 12141T). Finally, on the basis of data from the literature and new DNA-DNA hybridization and phenotypic data, the novel species Castellaniella denitrificans sp. nov. (type strain NKNTAUT = DSM 11046T = CCUG 39541T) is proposed for two strains previously identified as strains of A. defragrans.

Neumonía adquirida en la comunidad y bacteriemia por Alcaligenes xylososidans / Community-acquired pneumonia and bacteriemia due to Alcaligenes xylososidans

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