Gene cloning, expression and performance validation of nitric oxide dismutase.

Nitrous oxide (N2O) is a significant contributor to global warming and possesses an ozone-depleting impact nearly 298 times that of CO2. To reduce N2O emissions, the newly-discovered nod gene which can directly convert NO into N2 and O2 was successfully cloned from the anaerobic denitrification sludge. The recombinant plasmid containing the nod gene was built, and the expression of nod gene in Escherichia coli was determined, leading to the construction of recombinant engineering bacteria. Results showed that the recombinant engineering bacteria E. coli BL21 (DE3)-pET28a-nod could autonomously degrade NO, with a degradation rate of 72 % within 48 h, and could produce 2479.72 ppm of N2 and 75.12 mL of O2. The cumulative O2 production of the sludge sample and recombinant E. coli within 8 h was 1.75 mL and 8.45 mL, respectively. The cumulative O2 production of recombinant E. coli was at least 4.82 times higher than that of the sludge sample. The investigation proposed a new biodegradation pathway for nitrogen pollution.

Effects of bamboo-charcoal modified by bimetallic Fe/Pd nanoparticles on n-hexane biodegradation by bacteria Pseudomonas mendocina NX-1.

The high hydrophobicity of n-hexane is the main reason why it is difficult to be removed biologically. In this study, the effects of bamboo-charcoal modified by bimetallic Fe/Pd (BBC) on n-hexane biodegradation by Pseudomonas mendocina NX-1 (PM) was investigated. The n-hexane removal efficiency was increased in the presence of BC. The highest n-hexane removal efficiency at 90.0% was achieved at 0.05 g L-1 BCE and 3 g L-1 NH4+ under pH 7.7 and 35 °C. Additionally, protein content (45.9 µg mL-1) and negative cell surface zeta potential (-26.4 mV) were increased during biodegradation process, with PM-BBC being 43.1 µg mL-1 and 19.1 mV. Bacterial growth was improved and maximum cell surface hydrophobicity was obtained after 20 h, which was 59.4% higher than the control with PM-BBC (37.7%) or PM (16.1%), showing biodegradation products of 1-butanol and acetic acid. The results indicate that BBC improved n-hexane biodegradation efficiency by promoting bacterial growth, reducing cell zeta potential, exposing hydrophobic proteins, and increasing cell surface hydrophobicity of bacterial strain NX-1. This investigation suggests that BBC-enhanced biodegradation can be promising to treat n-hexane-containing gas.

Co-culturing fungus Penicillium citrinum and strain Citrobacter freundii improved nitrate removal and carbon utilization by promoting glyceride metabolism.

Exploring the interaction between denitrifying microbial species is significant for improving denitrification performance. In this study, the effects of co-culturing fungus Penicillium citrinum and strain Citrobacter freundii on denitrification were investigated. Results showed that the maximum nitrate removal and carbon utilization in co-culture were 68.0 and 14.1 mg·L-1·d-1, respectively. The total adenosine triphosphatase activity was increased to 9.87 U‧mg-1 protein in co-culture, and nicotinamide adenine dinucleotide production was 1.7-2.3 times that of monoculture, attributing to increased carbon utilization. Further metabolomics and membrane permeability assay revealed that co-culture increased the metabolism of glycerides, thereby enhancing the membrane permeability of strain Citrobacter freundii and promoting the transmembrane transport of nitrate and glucose, which boosted nitrate reductase activity and nicotinamide adenine dinucleotide production in turn. In summary, co-culturing promoted carbon utilization and enhanced substrate removal efficiency through the metabolism of glycerides, which provided a strategy to enhance denitrification performance in wastewater treatment.

Characterization of the complete chloroplast genome of Elaeagnus pungens (elaeagnaceae) and phylogeny within elaeagnaceae.

Elaeagnus pungens is an evergreen shrub with high medicinal values. In this study, the complete chloroplast (cp) genome of E. pungens was characterized. The chloroplast genome of E. pungens is 152,218 bp in length, consisting of two 25,876 bp inverted repeats, 18,231 bp small single copy region, and 82,235 bp large single copy region. The chloroplast genome contains 113 unique genes, including 79 protein-coding genes, 30 tRNA genes, and 4 rRNA genes. Phylogenomic analysis revealed that E. pungens and species from Elaeagnus formed a monophyletic clade sister to the clade consisting of species from Hippophae.

NosZ gene cloning, reduction performance and structure of Pseudomonas citronellolis WXP-4 nitrous oxide reductase.

Nitrous oxide reductase (N2OR) is the only known enzyme that can reduce the powerful greenhouse gas nitrous oxide (N2O) to harmless nitrogen at the final step of bacterial denitrification. To alleviate the N2O emission, emerging approaches aim at microbiome biotechnology. In this study, the genome sequence of facultative anaerobic bacteria Pseudomonas citronellolis WXP-4, which efficiently degrades N2O, was obtained by de novo sequencing for the first time, and then, four key reductase structure coding genes related to complete denitrification were identified. The single structural encoding gene nosZ with a length of 1914 bp from strain WXP-4 was cloned in Escherichia coli BL21(DE3), and the N2OR protein (76 kDa) was relatively highly efficiently expressed under the optimal inducing conditions of 1.0 mM IPTG, 5 h, and 30 °C. Denitrification experiment results confirmed that recombinant E. coli had strong denitrification ability and reduced 10 mg L-1 of N2O to N2 within 15 h under the optimal conditions of pH 7.0 and 40 °C, its corresponding N2O reduction rate was almost 2.3 times that of Alcaligenes denitrificans strain TB, but only 80% of that of wild strain WXP-4, meaning that nos gene cluster auxiliary gene deletion decreased the activity of N2OR. The 3D structure of N2OR predicted on the basis of sequence homology found that electron transfer center CuA had only five amino acid ligands, and the S2 of the catalytically active center CuZ only bound one CuI atom. The unique 3D structure was different from previous reports and may be closely related to the strong N2O reduction ability of strain WXP-4 and recombinant E. coli. The findings show a potential application of recombinant E. coli in alleviating the greenhouse effect and provide a new perspective for researching the relationship between structure and function of N2OR.

Chemical Composition, In Vitro and In Situ Antimicrobial and Antibiofilm Activities of Syzygium aromaticum (Clove) Essential Oil.

The essential oil of Syzygium (S.) aromaticum (CEO) is known for its good biological activity. The aim of the research was to evaluate in vitro and in situ antimicrobial and antibiofilm activity of the essential oil produced in Slovakia. The main components of CEO were eugenol 82.4% and (E)-caryophyllene 14.0%. The antimicrobial activity was either weak or very strong with inhibition zones ranging from 4.67 to 15.78 mm in gram-positive and gram-negative bacteria and from 8.22 to 18.56 mm in yeasts and fungi. Among the tested bacteria and fungi, the lowest values of MIC were determined for Staphylococcus (S.) aureus and Penicillium (P.) expansum, respectively. The vapor phase of CEO inhibited the growth of the microscopic filamentous fungi of the genus Penicillium when tested in situ on bread. The strongest effect of mycelia inhibition in a bread model was observed against P. expansum at concentrations of 250 and 500 µL/mL. The best antimicrobial activity of CEO in the carrot model was found against P. chrysosenum. Differences between the mass spectra of Bacillus (B.) subtilis biofilms on the tested surfaces (wood, glass) and the control sample were noted from the seventh day of culture. There were some changes in mass spectra of Stenotrophomonas (S.) maltophilia, which were observed in both experimental groups from the fifth day of culture. These findings confirmed the impact of CEO on the protein structure of older biofilms. The findings indicate that, besides being safe and sensorially attractive, S. aromaticum has antimicrobial activity, which makes it a potential substitute for chemical food preservatives.

A fungus-bacterium co-culture synergistically promoted nitrogen removal by enhancing enzyme activity and electron transfer.

The fungus Penicillium citrinum WXP-2 and the bacterium Citrobacter freundii WXP-9 were isolated and found to have poor denitrification performance. Surprisingly, co-culture of the two strains which formed fungus-bacterium pellets (FBPs) promoted the removal efficiency of nitrate (NO3--N; 95.78%) and total nitrogen (TN; 81.73%). Nitrogen balance analysis showed that excess degraded NO3--N was primarily converted to N2 (77.53%). Moreover, co-culture increased the dry weight to 0.74 g/L. The diameter of pellets and cell viability also increased by 1.49 and 1.78 times, respectively, indicating that the co-culture exerted a synergistic effect to promote growth. The increase in electron-transmission system activity [99.01 mg iodonitrotetrazolium formazan/(g·L)] and nitrate reductase activity [8.65 mg N/(min·mg protein)] were responsible for denitrification promotion. The FBPs also exhibited the highest degradation rate at 2:1 inoculation ratio and 36 h delayed inoculation of strain WXP-9. Finally, recycling experiments of FBP demonstrated that the high steady TN removal rate could be maintained for five cycles.

Chemical Composition and Antimicrobial Activity of Selected Essential Oils against Staphylococcus spp. Isolated from Human Semen.

Staphylococcus spp. is not only a commensal bacteria but also a major human pathogen that causes a wide range of clinical infections. Recent evidence suggests that Staphylococcus has the ability to colonize the reproductive system and to affect its structure and functions. The objective of this study was to determine the chemical properties and antibacterial effects of select essential oils (EOs): Amyris balsamifera L., Boswellia carterii Birdw., Canarium luzonicum (Blume) A. Gray, Cinnamomum camphora (L.) J. Presl., Cinnamomum camphora var. linaloolifera Y. Fuita, Citrus x aurantium L., Gaultheria procumbens L., Litsea cubeba (Lour.) Pers., Melaleuca ericifolia Smith., Melaleuca leucadendra L., Pogostemon cablin (Blanco) Benth., Citrus limon (L.) Osbeck, Santalum album L., and Vetiveria zizanoides (L.) Roberty against 50 Staphylococcus spp. cultures isolated from human semen, specifically Staphylococcus aureus, S. capiti, S. epidermidis, S. haemoliticus, and S. hominis. The disc diffusion and broth microdilution methods were used to assess the antimicrobial potential and to determine the minimum inhibitory concentration (MIC) of the selected EOs. The best anti-Staphylococcus activities were found with both methods for the essential oils of C. luzonicum (Blume) A. Gray, A. balsamifera, C. camphora, and P. cabli.

Enhanced denitrification performance of Alcaligenes sp. TB by Pd stimulating to produce membrane adaptation mechanism coupled with nanoscale zero-valent iron.

The microbial reduction of nitrate in the presence of nanoscale zero-valent iron (nZVI) was evaluated to assess the feasibility of employing nZVI for biological denitrification treatment. The effect of modified nZVI on the growth, metabolism, and denitrification performance of Alcaligenes sp. TB under aerobic conditions was studied. Results showed that Alcaligenes sp. TB with nZVI/Pd had 31.5% increase in nitrate removal and 18.1% decrease in nitrite accumulation within 28 h. nZVI/Pd exhibited less inhibition on the cell growth (OD600 = 0.725), NADH/NAD+ ratio (86% of control), and electron transfer system activity (68.5% of control). In addition, nZVI/Pd decreased the membrane fluidity by increasing the trans/cis isomerization ratio (317.7% of control) to enhance the resistance of nZVI. This study underlines the effects of nZVI/Pd on membrane susceptibility via membrane fatty acid transformation during denitrification and suggests the influence of engineered nanomaterials on denitrification.

Effects of carbon nanotube on denitrification performance of Alcaligenes sp. TB: Promotion of electron generation, transportation and consumption.

Multi-walled carbon nanotubes (MWCNTs) promote biodegradation in water treatment, but the effect of MWCNT on denitrification under aerobic conditions is still unclear. This investigation focused on the denitrification performance of MWCNT and its toxic effects on Alcaligenes sp. TB which showed that 30 mg/L MWCNTs increased NO3- removal efficiency from 84% to 100% and decreased the NO2-and N2O accumulation rates by 36% and 17.5%, respectively. Nitrite reductase and nitrous oxide reductase activities were further increased by 19.5% and 7.5%, respectively. The mechanism demonstrated that electron generation (NADH yield) and electron transportation system activity increased by 14.5% and 104%, respectively. Cell membrane analysis found that MWCNT caused an increase in polyunsaturated fatty acids, which had positive effects on electron transportation and membrane fluidity at a low concentration of 96 mg/kg but caused membrane lipid peroxidation and impaired membrane integrity at a high concentration of 115 mg/L. These findings confirmed that MWCNT affects the activity of Alcaligenes sp. TB and consequently enhances denitrification performance.

The mRNA expression of various angiogenesis-related genes in pediatric sarcomas and nonmalignant lesions of tissue.

For better understanding cancer pathogenesis and searching a potential target for antineoplastic therapy, the authors have studied mRNA expression profile in tissues from 39 children with histological confirmed malignant sarcomas and from 23 patients with bone and soft tissue nonmalignant lesions. mRNA levels of Angiogenesis-related genes VEGFA (including isoforms of 121, 165, 189), VEGFC, VEGFR-1, VEGFR-2, VEGFR-3, HIF-1α, TF, TFPI-1, TFPI-2, uPA, PAI-1 in pediatric specimens were examined using quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR). uPA, HIF-1α, VEGFR-1, VEGFR-2, VEGFR-3, and VEGFC mRNA levels from nonmalignant tissue were significantly higher than those from cancer tissue. On the other hand, isoform VEGFA121 and VEGFA165 and ratio VEGFA165/189 mRNA levels in cancer were higher in comparison with nonmalignant tissue. There was a strong correlation between VEGFA165 and VEGFA189 mRNA expression levels both in cancer tissue and in nonmalignant tissue. In grade 4 tumors in comparison with grade 2 tumors, there was a reduced VEGFA165/189 ratio. Moreover, TFPI-1 and TFPI-2 mRNA levels were significantly lower in sarcomas than in nonmalignant lesions and TFPI-2 was significantly lower in grade 4 tumors than in grade 2. The present data suggested that mRNA overexpression of angiogenesis-related genes is not a prerogative of malignant tissues. The authors supposed that in pediatric bone and soft tissue pathology, high expression of mRNAs of some angiogenesis-related genes may be associated with inflammation and physiological angiogenesis rather than with the development of a malignant tumor. The authors showed the importance of VEGFA121 and/or VEGFA165 and VEGFA165/189 isoform ratio in pediatric sarcomas neoangiogenesis and TFPI-2 for tumors grade 4.

Translocation (10;11)(p12;q23) in childhood acute myeloid leukemia: incidence and complex mechanism.

Using both conventional and molecular cytogenetic methods, we found five new cases of t(10;11)(p12;q23). This translocation represented 28% of all cases of childhood AML treated at our center in 2004, and 63% of AML with rearrangements of 11q23. We describe three mechanisms for the translocation. Different fragments of 11q were involved in four of the five cases. One patient showed a cytogenetically cryptic insertion of 5' part of MLL into the 3' part of MLLT10 in 10p12. The median event-free survival of patients was 8.1 months, and we conclude that the t(10;11)(p12;q23) is associated with unfavorable prognosis in childhood acute myeloid leukemia.