Mercury Tolerance of Thermophilic <Emphasis Type="Italic">Bacillus</Emphasis> sp. and <Emphasis Type="Italic">Ureibacillus</Emphasis> sp

Although resistance of microorganisms to Hg(II) salts has been widely investigated and resistant strains have been reported from many eubacterial genera, there are few reports of mercuric ion resistance in extremophilic microorganisms. Moderately thermophilic mercury resistant bacteria were selected by growth at 62 °C on Luria agar containing HgCl₂. Sequence analysis of 16S rRNA genes of two isolates showed the closest matches to be with Bacillus pallidus and Ureibacillus thermosphaericus. Minimum inhibitory concentration (MIC) values for HgCl₂ were 80 μg/ml and 30 μg/ml for these isolates, respectively, compared to 10 μg/ml for B. pallidus H12 DSM3670, a mercury-sensitive control. The best-characterised mercury-resistant Bacillus strain, B. cereus RC607, had an MIC of 60 μg/ml. The new isolates had negligible mercuric reductase activity but removed Hg from the medium by the formation of a black precipitate, identified as HgS by X-ray powder diffraction analysis. No volatile H₂S was detected in the headspace of cultures in the absence or presence of Hg²⁺, and it is suggested that a new mechanism of Hg tolerance, based on the production of non-volatile thiol species, may have potential for decontamination of solutions containing Hg²⁺ without production of toxic volatile H₂S.     

Biodegradation of Ethametsulfuron-Methyl by <Emphasis Type="Italic">Pseudomonas</Emphasis> sp. SW4 Isolated from Contaminated Soil

A soil bacterium SW4, capable of degrading the sulfonylurea herbicide ethametsulfuron-methyl (ESM), was isolated from the bottom soil of a herbicide factory. Based on physiological characteristics, biochemical tests and phylogenetic analysis of the 16S rRNA gene sequence, the strain was identified as a Pseudomonas sp. The total degradation of ESM in the medium containing glucose was up to 84.6% after 6 days of inoculation with SW4 strain. The inoculation of strain SW4 to soil treated with ESM resulted in a higher degradation rate than in noninoculated soil regardless of the soil sterilized or nonsterilized. Five metabolites of ESM degradation were analyzed by liquid chromatography/mass spectrometry. Based on the identified products, strain SW4 seemed to degrade ESM after two separate and different pathways: one leads to the cleavage of the sulfonylurea bridge, whereas the other to the dealkylation and opening of the triazine ring of ESM.     

Comparative effectiveness of <Emphasis Type="Italic">Pseudomonas</Emphasis> and <Emphasis Type="Italic">Serratia</Emphasis> sp. containing ACC-deaminase for improving growth and yield of wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) under salt-stressed conditions

Ethylene synthesis is accelerated in response to various environmental stresses like salinity. Ten rhizobacterial strains isolated from wheat rhizosphere taken from different salt affected areas were screened for growth promotion of wheat under axenic conditions at 1, 5, 10 and 15 dS m⁻¹. Three strains, i.e., Pseudomonas putida (N21), Pseudomonas aeruginosa (N39) and Serratia proteamaculans (M35) showing promising performance under axenic conditions were selected for a pot trial at 1.63 (original), 5, 10 and 15 dS m⁻¹. Results showed that inoculation was effective even in the presence of higher salinity levels. P. putida was the most efficient strain compared to the other strains and significantly increased the plant height, root length, grain yield, 100-grain weight and straw yield up to 52, 60, 76, 19 and 67%, respectively, over uninoculated control at 15 dS m⁻¹. Similarly, chlorophyll content and K⁺/Na⁺ of leaves also increased by P. putida over control. It is highly likely that under salinity stress, 1-aminocyclopropane-1-carboxylic acid-deaminase activity of these microbial strains might have caused reduction in the synthesis of stress (salt)-induced inhibitory levels of ethylene. The results suggested that these strains could be employed for salinity tolerance in wheat; however, P. putida may have better prospects in stress alleviation/reduction.     

Substrate specificity of a long-chain alkylamine-degrading <Emphasis Type="Italic">Pseudomonas</Emphasis> sp isolated from activated sludge

A bacterium strain BERT, which utilizes primary long-chain alkylamines as nitrogen, carbon and energy source, was isolated from activated sludge. This rod-shaped motile, Gram-negative strain was identified as a Pseudomonas sp. The substrate spectrum of this Pseudomonas strain BERT includes primary alkylamines with alkyl chains ranging from C₃ to C₁₈, and dodecyl-1,3-diaminopropane. Amines with alkyl chains ranging from 8 to 14 carbons were the preferred substrates. Growth on dodecanal, dodecanoic acid and acetic acid and simultaneous adaptation studies indicated that this bacterium initiates degradation through a C_alkyl–N cleavage. The cleavage of alkylamines to the respective alkanals in Pseudomonas strain BERT is mediated by a PMS-dependent alkylamine dehydrogenase. This alkylamine dehydrogenase produces stoichiometric amounts of ammonium from octylamine. The PMS-dependent alkylamine was found to oxidize a broad range of long-chain alkylamines. PMS-dependent long-chain aldehyde dehydrogenase activity was also detected in cell-free extract of Pseudomonas strain BERT grown on octylamine. The proposed pathway for the oxidation of alkylamine in strain BERT proceeds from alkylamine to alkanal, and then to the fatty acid.     

Quantitative studies on phosphorus transference occuring between <Emphasis Type="Italic">Microcystis aeruginosa</Emphasis> and its attached bacterium (<Emphasis Type="Italic">Pseudomonas</Emphasis> sp.)

Phosphorus release from Microcystis aeruginosa and attached bacterium (Pseudomonas sp.) isolated from Lake Taihu was examined using a phosphorus isotope tracer in order to investigate the phosphorus transference between the two species. Our results reveal that the amount of phosphorus released form ³²P-saturated M. aeruginosa is determined by its growth phase and most of phosphorus is assimilated by Pseudomonas finally while the amount of phosphorus released from ³²P-saturated Pseudomonas is also determined by the growth phase of M. aeruginosa and most of them are assimilated by M. aeruginosa. The results suggest that phosphorus transference occurs between M. aeruginosa and its attached Pseudomonas . This process makes microenvironment of mucilage of M. aeruginosa attached bacteria maintain relative high amounts of phosphorus. Attached bacteria may be a temporary phosphorus bank to the growth of M. aeruginosa, and assimilation of phosphorus by M. aeruginosa becomes easy when M. aeruginosa is in lag growth phase. Thus, the phosphorus exchange between M. aeruginosa and attached Pseudomonas in microenvironment may be important to microfood web and cyanobacteria bloom.     

Assessing the toxic effect of 2,4,6-trinitrotoluene on cells of <Emphasis Type="Italic">Escherichia coli</Emphasis> K12 by flow cytofluorometry

The magnitude of transmembrane potential Δψ in cells of Escherichia coli K12 was determined by the method of flow cytofluorometry for different phases of growth. It was large in the log phase, whereas in the lag and stationary phases, the population was shown to consist of two subpopulations with low and large values of Δψ in cells. In the presence of 200 mg/l of 2,4,6-trinitrotoluene (TNT), this bimodal distribution of Δψ over the population was observed during the entire growth period until TNT was almost completely eliminated from the cultivation medium (to a concentration of 18–20 mg/l). The mean value of Δψ in cells of the population grown in the presence of TNT was substantially smaller than that in controls due to the larger fraction of the subpopulation with a low value of Δψ. Upon elimination of TNT, the distribution of Δψ in cells of the culture became unimodal and close to that in the control culture in the early log phase of growth. These findings are discussed from the standpoint that considers heterogeneity of the culture of Escherichia coli K12 as a mechanism of its adaptation to the presence of xenobiotics.     

Induction of the unculturable state in <Emphasis Type="Italic">Escherichia coli</Emphasis> K12 with 2,4,6-trinitrotoluene

The toxic effect of high (200 mg/l) 2,4,6-trinitrotoluene (TNT) concentrations on Escherichia coli K12 cells in the absence of exogenous nutrient sources (incubation in 0.5% NaCl) was shown to manifest itself in the transfer of the culture to an unculturable but viable state; its reversal depends on the duration of culture contact with the xenobiotic and the conditions of cell recultivation. The likelihood that cell succession to death forms the basis of the physiologo-biochemical mechanism of the unculturable state in Escherichia coli K12 population under conditions of combined toxic and starvation stress is discussed.     

Degradation of ethylbenzene by free and immobilized <Emphasis Type="Italic">Pseudomonas</Emphasis><Emphasis Type="Italic">fluorescens-</Emphasis>CS2

Pseudomonas fluorescens-CS2 metabolized ethylbenzene as the sole source of carbon and energy. The involvement of catechol as the hydroxylated intermediate during the biodegradation of ethylbenzene was established by TLC, HPLC and enzyme analysis. The specific activity of Catechol 2,3-dioxygenase in the cell free extracts of P. fluorescens-CS2 was determined to be 0.428 μmoles min⁻¹ mg⁻¹ protein. An aqueous-organic, Two-Phase Batch Culture System (TPBCS) was developed to overcome inhibition due to higher substrate concentrations. In TPBCS, P. fluorescens-CS2 demonstrated ethylbenzene utilization up to 50 mM without substrate inhibition on inclusion of n-decanol as the second phase. The rate of ethylbenzene metabolism in TPBCS was found enhance by fivefold in comparison with single phase system. Alternatively the alginate, agar and polyacrylamide matrix immobilized P. fluorescens-CS2 cells efficiently degraded ethylebenzene with enhanced efficiency compared to free cell cultures in single and two-phase systems. The cells entrapped in ployacrylamide and alginate were found to be stable and degradation efficient for a period of 42 days where as agar-entrapped P. fluorescens was stable and efficient a period of 36 days. This demonstrates that alginate and polyacrylamide matrices are more promising as compared to agar for cell immobilization.     

Four novel <Emphasis Type="Italic">Candida</Emphasis> species in the <Emphasis Type="Italic">Candida albicans</Emphasis>/<Emphasis Type="Italic">Lodderomyces elongisporus</Emphasis> clade isolated from the gut of flower beetles

Flower-visiting beetles belonging to three species of Cetoniidae were collected on three mountains near Beijing, China, and yeasts were isolated from the gut of the insects collected. Based on the 26S rDNA D1/D2 domain and internal transcribed spacer (ITS) region sequence analysis and phenotypic characterization, four novel anamorphic yeast species located in the Candida albicans/Lodderomyces elongisporus clade were identified from 18 of the strains isolated. The new species and type strains are designated as Candida blackwellae AS 2.3639^T (=CBS 10843^T), Candida jiufengensis AS 2.3688^T (=CBS 10846^T), Candida oxycetoniae AS 2.3656^T (=CBS 10844^T), and Candida pseudojiufengensis AS 2.3693^T (=CBS 10847^T). C. blackwellae sp. nov. was basal to the branch formed by C. albicans and C. dubliniensis with moderately strong bootstrap support. The closest relative of C. oxycetoniae was L. elongisporus. C. jiufengensis sp. nov. and C. pseudojiufengensis sp. nov. were closely related with each other and formed a branch in a subclade represented by C. parapsilosis and L. elongisporus.     

Phenanthrene degradation by bacteria of the genera <Emphasis Type="Italic">Pseudomonas</Emphasis> and <Emphasis Type="Italic">Burkholderia</Emphasis> in model soil systems

Degradation of phenanthrene by strains Pseudomona, Moscow, KMK, 2004simova, I.A. and Chernov, I.s putida BS3701 (pBS1141, pBS1142), Pseudomonas putida BS3745 (pBS216), and Burkholderia sp. BS3702 (pBS1143) were studied in model soil systems. The differences in accumulation and uptake rate of phenanthrene intermediates between the strains under study have been shown. Accumulation of 1-hydroxy-2-naphthoic acid in soil in the course of phenanthrene degradation by strain BS3702 (pBS1143) in a model system has been revealed. The efficiency of phenanthrene biodegradation was assessed using the mathematical model proposed previously for assessment of naphthalene degradation efficiency. The efficiency of degradation of both phenanthrene and the intermediate products of its degradation in phenanthrene-contaminated soil is expected to increase with the joint use of strains P. Putida BS3701 (pBS1141, pBS1142) and Burkholderia sp. BS3702 (pBS1143).     

Diverse mechanisms adopted by fluorescent <Emphasis Type="Italic">Pseudomonas</Emphasis> PGC2 during the inhibition of <Emphasis Type="Italic">Rhizoctonia solani</Emphasis> and <Emphasis Type="Italic">Phytophthora capsici</Emphasis>

Rhizoctonia solani and Phytophthora capsici are two of the most destructive phytopathogens occurring worldwide and are only partly being managed by traditional control strategies. Fluorescent Pseudomonas isolates PGC1 and PGC2 were checked for the antifungal potential against R. solani and P. capsici. Both the isolates were screened for the ability to produce a range of antifungal compounds. The results of this study indicated the role of chitinase and β-1,3-glucanase in the inhibition of R. solani, however, antifungal metabolites of a non-enzymatic nature were responsible for inhibition of P. capsici. The study confirmed that multiple and diverse mechanisms are adopted by the same antagonist to suppress different phytopathogens, as evidenced in case of R. solani and P. capsici.     

Extracellular phycoerythrin-like protein released by freshwater cyanobacteria <Emphasis Type="Italic">Oscillatoria</Emphasis> and <Emphasis Type="Italic">Scytonema</Emphasis> sp.

During growth of the freshwater cyanobacteria, Oscillatoria sp. BTCC/A0004, and Scytonema sp. TISTR 8208, a pink pigment is released into the growth medium. The pigment from each source had a molecular weight of approximately 250 kDa and had adsorption maxima at 560 and 620 nm. These results suggest that pink pigment is a phycoerythrin-like protein. It inhibited the growth of green algae, Chlorella fusca and Chlamydomonas reinhardtii, but not other cyanobacteria or true bacteria. The concentration at which growth inhibition 50% occurred was 0.5, 6 and more than 10 mg ml⁻¹, respectively.     

Effect of temperature on the life history of <Emphasis Type="Italic">Eretmocerus</Emphasis> sp. nr. <Emphasis Type="Italic">furuhashii</Emphasis>, a parasitoid of <Emphasis Type="Italic">Bemisia tabaci</Emphasis>

Eretmocerus sp. nr. furuhashii (Hymenoptera: Aphelinidae) is an indigenous parasitoid of Bemisia tabaci (Gennadius)(Hemiptera: Aleyrodidae) from southern China; the effects of constant temperatures on the life history of E. sp. nr. furuhashii were examined in the laboratory. The developmental period ranged from 39.2 days at 20°C to 12.40 days at 32°C. A total of 263.4 degree-days were required to complete development with a lower developmental threshold temperature of 11.1°C. Of the eggs produced, 59.3% completed development at 20°C with completion increasing to 71.5% at 26°C. Adult female longevity was 10.8 days at 20°C and 5.2 days at 32°C while the mean daily offspring reproduced per female was highest at 29°C with 5.9 offspring. Adult oviposition peaked three days after emergence at 26, 29 and 32°C, and four days post-emergence at 20°C and 23°C. The total numbers of offspring produced per female ranged from 25.7 individuals at 32°C to 41.1 individuals at 20°C. The sex ratio had a female bias and ranged from 0.72 at 17°C to 0.51 at 35°C. The intrinsic rate of increase was 0.1727 at 29°C followed with 0.1606 at 32°C. Results indicated that E. sp. nr. furuhashii reaches its maximum biological potential at temperatures ranging from 26°C to 32°C.     

Reduction of Fe(II)EDTA-NO by a newly isolated <Emphasis Type="Italic">Pseudomonas</Emphasis> sp. strain DN-2 in NO<Subscript><Emphasis Type="Italic">x</Emphasis></Subscript> scrubber solution

Biological reduction of nitric oxide (NO) chelated by ferrous ethylenediaminetetraacetate (Fe(II)EDTA) to N₂ is one of the core processes in a chemical absorption–biological reduction integrated technique for nitrogen oxide (NO _x ) removal from flue gases. A new isolate, identified as Pseudomonas sp. DN-2 by 16S rRNA sequence analysis, was able to reduce Fe(II)EDTA-NO. The specific reduction capacity as measured by NO was up to 4.17 mmol g DCW⁻¹ h⁻¹. Strain DN-2 can simultaneously use glucose and Fe(II)EDTA as electron donors for Fe(II)EDTA-NO reduction. Fe(III)EDTA, the oxidation of Fe(II)EDTA by oxygen, can also serve as electron acceptor by strain DN-2. The interdependency between various chemical species, e.g., Fe(II)EDTA-NO, Fe(II)EDTA, or Fe (III)EDTA, was investigated. Though each complex, e.g., Fe(II)EDTA-NO or Fe(III)EDTA, can be reduced by its own dedicated bacterial strain, strain DN-2 capable of reducing Fe(III)EDTA can enhance the regeneration of Fe(II)EDTA, hence can enlarge NO elimination capacity. Additionally, the inhibition of Fe(II)EDTA-NO on the Fe(III)EDTA reduction has been explored previously. Strain DN-2 is probably one of the major contributors for the continual removal of NO _x due to the high Fe(II)EDTA-NO reduction rate and the ability of Fe(III)EDTA reduction.     

In planta transformation of <Emphasis Type="Italic">Notocactus scopa</Emphasis> cv. <Emphasis Type="Italic">Soonjung</Emphasis> by <Emphasis Type="Italic">Agrobacterium </Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>

Notocactus scopa cv. Soonjung was subjected to in planta Agrobacterium tumefaciens-mediated transformation with vacuum infiltration, pin-pricking, and a combination of the two methods. The pin-pricking combined with vacuum infiltration (20-30 cmHg for 15 min) resulted in a transformation efficiency of 67-100%, and the expression of the uidA and nptII genes was detected in transformed cactus. The established in planta transformation technique generated a transgenic cactus with higher transformation efficiency, shortened selection process, and stable gene expression via asexual reproduction. All of the results showed that the in planta transformation method utilized in the current study provided an efficient and time-saving procedure for the delivery of genes into the cactus genome, and that this technique can be applied to other asexually reproducing succulent plant species.     

The <Emphasis Type="Italic">Caenorhabditis</Emphasis><Emphasis Type="Italic">briggsae</Emphasis> genome contains active <Emphasis Type="Italic">CbmaT1</Emphasis> and <Emphasis Type="Italic">Tcb1</Emphasis> transposons

The maT clade of transposons is a group of transposable elements intermediate in sequence and predicted protein structure to mariner and Tc transposons, with a distribution thus far limited to a few invertebrate species. We present evidence, based on searches of publicly available databases, that the nematode Caenorhabditis briggsae has several maT-like transposons, which we have designated as CbmaT elements, dispersed throughout its genome. We also describe two additional transposon sequences that probably share their evolutionary history with the CbmaT transposons. One resembles a fold back variant of a CbmaT element, with long (380-bp) inverted terminal repeats (ITRs) that show a high degree (71%) of identity to CbmaT1. The other, which shares only the 26-bp ITR sequences with one of the CbmaT variants, is present in eight nearly identical copies, but does not have a transposase gene and may therefore be cross mobilised by a CbmaT transposase. Using PCR-based mobility assays, we show that CbmaT1 transposons are capable of excising from the C. briggsae genome. CbmaT1 excised approximately 500 times less frequently than Tcb1 in the reference strain AF16, but both CbmaT1 and Tcb1 excised at extremely high frequencies in the HK105 strain. The HK105 strain also exhibited a high frequency of spontaneous induction of unc-22 mutants, suggesting that it may be a mutator strain of C. briggsae.     

Characterization of <Emphasis Type="Italic">twist</Emphasis> and <Emphasis Type="Italic">snail</Emphasis> gene expression during mesoderm and nervous system development in the polychaete annelid <Emphasis Type="Italic">Capitella</Emphasis> sp. I

To investigate the evolutionary history of mesoderm in the bilaterian lineage, we are studying mesoderm development in the polychaete annelid, Capitella sp. I, a representative lophotrochozoan. In this study, we focus on the Twist and Snail families as candidate mesodermal patterning genes and report the isolation and in situ expression patterns of two twist homologs (CapI-twt1 and CapI-twt2) and two snail homologs (CapI-sna1 and CapI-sna2) in Capitella sp. I. CapI-twt1 is expressed in a subset of mesoderm derivatives during larval development, while CapI-twt2 shows more general mesoderm expression at the same stages. Neither twist gene is detected before the completion of gastrulation. The two snail genes have very distinct expression patterns. At cleavage and early gastrula stages, CapI-sna1 is broadly expressed in precursors of all three germ layers and becomes restricted to cells around the closing blastopore during late gastrulation; CapI-sna2 expression is not detected at these stages. After gastrulation, both snail genes are expressed in the developing central nervous system (CNS) at stages when neural precursor cells are internalized, and CapI-sna1 is also expressed laterally within the segmental mesoderm. Based on the expression patterns in this study, we suggest a putative function for Capitella sp. I twist genes in mesoderm differentiation and for snail genes in regulating CNS development and general cell migration during gastrulation. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Diversification of the barley and wheat <Emphasis Type="Italic">blt101</Emphasis>/<Emphasis Type="Italic">wpi6</Emphasis> promoters by the <Emphasis Type="Italic">Xumet</Emphasis> element without affecting stress responsiveness

BLT101-family plasma membrane proteins are found in a wide range of organisms from bacteria to nematodes and are involved in the regulation of cellular cation concentration under stress conditions. A comparison of the promoter regions of barley blt101 and its wheat ortholog, wpi6, revealed highly conserved nucleotide sequences between both genes and a unique insertion of a Xumet element in the blt101 promoter. The Xumet insertion occurred between a putative abscisic acid-responsive element (ABRE) and the dehydration-responsive element/c-repeat (DRE/CRT) within the blt101 promoter. However, blt101 and wpi6 were induced similarly in response to ABA, drought and low temperature, suggesting that the insertion does not affect promoter functions. The Xumet insertion in the blt101/wpi6 promoter region was detected in five barley cultivars, but absent in two wheat cultivars tested, suggesting that the insertion is barley-specific. Genomic Southern blot analysis revealed a large number of Xumet sequences interspersed in the barley genome, whereas only one or very few copies are present in the wheat genome. The data suggested that an expansion in copy number of Xumet elements occurred in the barley genome through evolution.