Transcriptomic Analysis of Degradative Pathways for Azo Dye Acid Blue 113 in Sphingomonas melonis B-2 from the Dye Wastewater Treatment Process.

BACKGROUND: Acid Blue 113 (AB113) is a typical azo dye, and the resulting wastewater is toxic and difficult to remove. METHODS: The experimental culture was set up for the biodegradation of the azo dye AB113, and the cell growth and dye decolorization were monitored. Transcriptome sequencing was performed in the presence and absence of AB113 treatment. The key pathways and enzymes involved in AB113 degradation were found through pathway analysis and enrichment software (GO, EggNog and KEGG). RESULTS: S. melonis B-2 achieved more than 80% decolorization within 24 h (50 and 100 mg/L dye). There was a positive relationship between cell growth and the azo dye degradation rate. The expression level of enzymes involved in benzoate and naphthalene degradation pathways (NADH quinone oxidoreductase, N-acetyltransferase and aromatic ring-hydroxylating dioxygenase) increased significantly after the treatment of AB113. CONCLUSIONS: Benzoate and naphthalene degradation pathways were the key pathways for AB113 degradation. NADH quinone oxidoreductase, N-acetyltransferase, aromatic ring-hydroxylating dioxygenase and CYP450 were the key enzymes for AB113 degradation. This study provides evidence for the process of AB113 biodegradation at the molecular and biochemical level that will be useful in monitoring the dye wastewater treatment process at the full-scale treatment.

Immune-enhancing effects of postbiotic produced by Bacillus velezensis Kh2-2 isolated from Korea Foods.

Postbiotics defined as soluble factors (products or metabolic byproducts) that are released after bacterial lysis or secreted by live bacteria, have attracted considerable attention because of their long shelf life, safety, and beneficial effects. In this study, we investigated the immune-enhancing activities of squid jeotgal (a traditional Korean fermented seafood)-derived Bacillus velezensis Kh2-2 (Kh2-2) postbiotics in vitro, ex vivo, and in vivo. Cell lysates of four Bacillus species were prepared by sonication. In particular, Kh2-2 lysates induced NO production by upregulating iNOS expression in RAW264.7 cells compared with the lysates of B. subtilis Kh2-1, B. vallismortis Kh8-3, and B. amyloliquefaciens Kh3-1. Furthermore, Kh2-2 lysates stimulated immune activation of macrophages by upregulating the NF-κB and MAPK signaling pathways and promoting immune-related cytokine secretion. In the ex vivo study, Kh2-2 lysates stimulated proliferation and polarized Th1 response by inducing the production of IL-2 and IFN-γ and inhibiting IL-10 expression in splenocytes. The in vivo immune-enhancing effects of Kh2-2 lysates and Kh2-2 were further evaluated using a cyclophosphamide (CTX)-induced immunosuppression mouse model. The results showed that oral administration of Kh2-2 lysates improved CTX-induced immunosuppression by enhancing innate and adaptive immunity, stimulating immune-related cytokine secretion, and modulating gut microbiota dysbiosis in mice. Thus, we concluded that Kh2-2 lysates have potential as a functional material for postbiotics with immune-enhancing effects.

Metagenomic Insight of a Full Scale Eco-Friendly Treatment System of Textile Dye Wastewater Using Bioaugmentation of the Composite Culture CES-1.

Effects of bioaugmentation of the composite microbial culture CES-1 on a full scale textile dye wastewater treatment process were investigated in terms of water quality, sludge reduction, dynamics of microbial community structures and their functional genes responsible for degradation of azo dye, and other chemicals. The removal efficiencies for Chemical Oxygen Demand (COD), Total Nitrogen (T-N), Total Phosphorus (T-P), Suspended Solids (SS), and color intensity (96.4%, 78.4, 83.1, 84.4, and 92.0, respectively) 300-531 days after the augmentation were generally improved after bioaugmentation. The denitrification linked to T-N removal appeared to contribute to the concomitant COD removal that triggered a reduction of sludge (up to 22%) in the same period of augmentation. Azo dye and aromatic compound degradation and other downstream pathways were highly metabolically interrelated. Augmentation of CES-1 increased microbial diversity in the later stages of augmentation when a strong microbial community selection of Acinetobacterparvus, Acinetobacterjohnsonii, Marinobacter manganoxydans, Verminephrobacter sp., and Arcobacter sp. occurred. Herein, there might be a possibility that the CES-1 augmentation could facilitate the indigenous microbial community successions so that the selected communities made the augmentation successful. The metagenomic analysis turned out to be a reasonable and powerful tool to provide with new insights and useful biomarkers for the complex environmental conditions, such as the full scale dye wastewater treatment system undergoing bioaugmentation.

Intracellular synthesis of gold nanoparticles by Gluconacetobacter liquefaciens for delivery of peptide CopA3 and ginsenoside and anti-inflammatory effect on lipopolysaccharide-activated macrophages.

Probiotic Gluconacetobacter strains are intestinal microbes with beneficial effects on human health. Recently, researchers have used these strains to biosynthesize metal and non-metal nanoparticles for treating various chronic diseases. Despite their importance in nanotechnology, gold nanoparticles (AuNPs) biosynthesized by Gluconacetobacter species have not been clearly identified for treating inflammation and inflammation-associated diseases. While ginsenoside CK has strong pharmaceutical activity, it also has strong cytotoxicity and hydrophobicity which is hurdle to make formulation. Peptide-nanoparticle hybrids are gaining increasing attention for their potential biomedical applications, including human inflammatory diseases. Herein, we developed peptide CopA3 surface conjugated and ginsenoside compound K (CK) loaded gold nanoparticles (GNP-CK-CopA3), which intracellularly synthesised by the probiotic Gluconacetobacter liquefaciens kh-1, to target lipopolysaccharide (LPS)-activated RAW264.7 macrophages. The synthetic GNP-CK-CopA3 was characterised by various instrumental techniques. The results of our cellular uptake and MTT assays exhibited obvious drug intracellular delivery without significant cytotoxicity. In addition, pre-treatment with GNP-CK-CopA3 significantly ameliorated LPS-induced nitric oxide (NO) and reactive oxygen species (ROS) production and suppressed the mRNA and protein expression of pro-inflammatory cytokines in macrophages. Furthermore, GNP-CK-CopA3 efficiently inhibited the activation of the nuclear factor-κB (NF-κB) and mitogen-activating protein kinase (MAPK) signalling pathways. Taken together, our findings highlight the potential of using peptide-nanoparticle hybrids in the development of anti-inflammatory approaches and providing the experimental foundation for further application.

Metagenomic analysis of relationships between the denitrification process and carbon metabolism in a bioaugmented full-scale tannery wastewater treatment plant.

The goal of this study was to investigate the relationship between the denitrification process and carbon metabolism in a full-scale tannery wastewater treatment plant bioaugmented with the microbial consortium BM-S-1. The metagenomic analysis of the microbial community showed that Brachymonas denitrificans, a known denitrifier, was present at a high level in the treatment stages of buffering (B), primary aeration (PA), and sludge digestion (SD). The occurrences of the amino acid-degrading enzymes alpha ketoglutarate dehydrogenase (α-KGDH) and tryptophan synthase were highly correlated with the presence of denitrification genes, such as napA, narG, nosZ and norB. The occurrence of glutamate dehydrogenase (GDH) was also highly paralleled with the occurrence of denitrification genes such as napA, narG, and norZ. The denitrification genes (nosZ, narG, napA, norB and nrfA) and amino acid degradation enzymes (tryptophan synthase, α-KGDH and pyridoxal phosphate dependent enzymes) were observed at high levels in B. This indicates that degradation of amino acids and denitrification of nitrate may potentially occur in B. The high concentrations of the fatty acid degradation enzyme groups (enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase and ß-ketothiolase) were observed together with the denitrification genes, such as napA, narG and nosZ. Phospholipase/carboxylesterase, enoyl-CoA hydratase/isomerase, acyl-CoA dehydrogenase, phenylacetate degradation enzyme and 3-hydroxyacyl-CoA dehydrogenase 2 were also dominant in B. All these results clearly indicate that the denitrification pathways are potentially linked to the degradation pathways of amino acids and fatty acids whose degradation products go through the TCA cycle, generating the NADH that is used as electron donors for denitrification.

Development of functional composts using spent coffee grounds, poultry manure and biochar through microbial bioaugmentation.

Spent coffee grounds (SCG), poultry manure, and agricultural waste-derived biochar were used to manufacture functional composts through microbial bioaugmentation. The highest yield of tomato stalk-based biochar (40.7%) was obtained at 450°C with a surface area of 2.35 m2 g-1. Four pilot-scale composting reactors were established to perform composting for 45 days. The ratios of NH4+-N/NO3--N, which served as an indicator of compost maturity, indicate rapid, and successful composting via microbial bioaugmentation and biochar amendment. Moreover, germination indices for radish also increased by 14-34% through augmentation and biochar amendment. Microbial diversity was also enhanced in the augmented and biochar-amended composts by 7.1-8.9%, where two species of Sphingobacteriaceae were dominant (29-43%). The scavenging activities of 2,2-diphenyl-1-picrylhydrazyl (DPPH) were enhanced by 14.1% and 8.6% in the fruits of pepper plants grown in the presence of the TR-2 (augmentation applied only) and TR-3 (both augmentation and biochar amendment applied) composts, respectively. Total phenolic content was also enhanced by 68% in the fruits of the crops grown in TR-3. Moreover, the other compost, TR-L (augmentation applied only), boosted DPPH scavenging activity by 111% in leeks compared with commercial organic fertilizer, while TR-3 increased the phenolic content by 44.8%. Composting facilitated by microbial augmentation and biochar amendment shortened the composting time and enhanced the quality of the functional compost. These results indicate that functional compost has great potential to compete with commercially available organic fertilizers and that the novel composting technology could significantly contribute to the eco-friendly recycling of organic wastes such as spent coffee grounds, poultry manure, and agricultural wastes.

Metagenomic insight of nitrogen metabolism in a tannery wastewater treatment plant bioaugmented with the microbial consortium BM-S-1.

Nitrogen (N) removal in a tannery wastewater treatment plant was significantly enhanced by the bioaugmentation of the novel consortium BM-S-1. In order to identify dominant taxa responsible for N metabolisms in the different stages of the treatment process, Illumina MiSeq Sequencer was used to conduct metagenome sequencing of the microbial communities in the different stages of treatment system, including influent (I), buffering (B), primary aeration (PA), secondary aeration (SA) and sludge digestion (SD). Based on MG-RAST analysis, the dominant phyla were Proteobacteria, Bacteroidetes and Firmicutes in B, PA, SA and SD, whereas Firmicutes was the most dominant in I before augmentation. The augmentation increased the abundance of the denitrification genes found in the genera such as Ralstonia (nirS, norB and nosZ), Pseudomonas (narG, nirS and norB) and Escherichia (narG) in B and PA. In addition, Bacteroides, Geobacter, Porphyromonasand Wolinella carrying nrfA gene encoding dissimilatory nitrate reduction to ammonium were abundantly present in B and PA. This was corroborated with the higher total N removal in these two stages. Thus, metagenomic analysis was able to identify the dominant taxa responsible for dissimilatory N metabolisms in the tannery wastewater treatment system undergoing bioaugmentation. This metagenomic insight into the nitrogen metabolism will contribute to a successful monitoring and operation of the eco-friendly tannery wastewater treatment system.

Denitratimonas tolerans gen. nov., sp. nov., a denitrifying bacterium isolated from a bioreactor for tannery wastewater treatment.

A denitrifying bacterium, designated strain E4-1(T), was isolated from a bioreactor for tannery wastewater treatment, and its taxonomic position was investigated using a polyphasic approach. Strain E4-1(T), a facultative anaerobic bacterium, was observed to grow between 0 and 12 % (w/v) NaCl, between pH 3.0 and 12.0. Cells were found to be oxidase-positive and catalase-negative. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain E4-1(T) forms a distinct lineage with respect to closely related genera in the family Xanthomonadaceae, and is closely related to Chiayiivirga, Aquimonas and Dokdonella, and the levels of 16S rRNA gene sequence similarity with respect to the type species of related genera are less than 93.9 %. The predominant respiratory quinone was determined to be ubiquinone-8 (Q-8) and the major cellular fatty acids were determined to be iso-C15:0, iso-C17:1 ω9c, iso-C11:0 and iso-C11:0 3OH. Based on physiological, biochemical and chemotaxonomic properties together with results of comparative 16S rRNA gene sequence analysis, strain E4-1(T) is considered to represent a novel species in a new genus, for which the name Denitratimonas tolerans gen. nov., sp. nov. is proposed. The type strain is E4-1(T) (=KACC 17565(T) = NCAIM B 025327(T)).

Paenibacillus yonginensis DCY84(T) induces changes in Arabidopsis thaliana gene expression against aluminum, drought, and salt stress.

Current agricultural production methods, for example the improper use of chemical fertilizers and pesticides, create many health and environmental problems. Use of plant growth-promoting bacteria (PGPB) for agricultural benefits is increasing worldwide and also appears to be a trend for the future. There is possibility to develop microbial inoculants for use in agricultural biotechnology, based on these beneficial plant-microbe interactions. For this study, ten bacterial strains were isolated from Yongin forest soil for which in vitro plant-growth promoting trait screenings, such as indole acetic acid (IAA) production, a phosphate solubilization test, and a siderophore production test were used to select two PGPB candidates. Arabidopsis thaliana plants were inoculated with Paenibacillus yonginensis DCY84(T) and Micrococcus yunnanensis PGPB7. Salt stress, drought stress and heavy metal (aluminum) stress challenges indicated that P. yonginensis DCY84(T)-inoculated plants were more resistant than control plants. AtRSA1, AtVQ9 and AtWRKY8 were used as the salinity responsive genes. The AtERD15, AtRAB18, and AtLT178 were selected to check A. thaliana responses to drought stress. Aluminum stress response was checked using AtAIP, AtALS3 and AtALMT1. The qRT-PCR results indicated that P. yonginensis DCY84(T) can promote plant tolerance against salt, drought, and aluminum stress. P. yonginensis DCY84(T) also showed positive results during in vitro compatibility testing and virulence assay against X. oryzae pv. oryzae Philippine race 6 (PXO99). Better germination rates and growth parameters were also recorded for the P. yonginensis DCY84(T) Chuchung cultivar rice seed which was grown on coastal soil collected from Suncheon. Based on these results, P. yonginensis DCY84(T) can be used as a promising PGPB isolate for crop improvement.

Paenibacillus yonginensis sp. nov., a potential plant growth promoting bacterium isolated from humus soil of Yongin forest.

Strain DCY84(T), a Gram-stain positive, rod-shaped, aerobic, spore-forming bacterium, motile by means of peritrichous flagella, was isolated from humus soil from Yongin forest in Gyeonggi province, South Korea. Strain DCY84(T) shared the highest sequence similarity with Paenibacillus barengoltzii KACC 15270(T) (96.86 %), followed by Paenibacillus timonensis KACC 11491(T) (96.49 %) and Paenibacillus phoenicis NBRC 106274(T) (95.77 %). Strain DCY84(T) was found to able to grow best in TSA at temperature 30 °C, at pH 8 and at 0.5 % NaCl. MK-7 menaquinone was identified as the isoprenoid quinone. The major polar lipids were identified as phosphatidylethanolamine, an unidentified aminophospholipid, two unidentified aminolipids and an unidentified polar lipid. The peptidoglycan was found to contain the amino acids meso-diaminopimelic acid, alanine and D-glutamic acid. The major fatty acids of strain DCY84(T) were identified as branched chain anteiso-C15:0, saturated C16:0 and branched chain anteiso-C17:0. The cell wall sugars of strain DCY84(T) were found to comprise of ribose, galactose and xylose. The major polyamine was identified as spermidine. The DNA G+C content was determined to be 62.6 mol%. After 6 days of incubation, strain DCY84(T) produced 52.96 ± 1.85 and 72.83 ± 2.86 µg/ml L-indole-3-acetic acid, using media without L-tryptophan and supplemented with L-tryptophan, respectively. Strain DCY84(T) was also found to be able to solubilize phosphate and produce siderophores. On the basis of the phenotypic characteristics, genotypic analysis and chemotaxonomic characteristics, strain DCY84(T) is considered to represent a novel species of the genus Paenibacillus, for which the name Paenibacillus yonginensis sp. nov. is proposed. The type strain is DCY84(T) (=KCTC 33428(T) = JCM 19885(T)).

Full-scale biological treatment of tannery wastewater using the novel microbial consortium BM-S-1.

In order to develop a more effective and eco-friendly treatment technology, a full-scale tannery wastewater treatment plant with a sludge digestion system was augmented with a novel microbial consortium (BM-S-1). The aim of this study was to determine if the BM-S-1 could successfully treat the tannery wastewater in a full-scale treatment system without chemical pretreatment and to investigate effect of the augmentation on sludge production. Chemical oxygen demand (COD), total nitrogen (TN), total phosphorus (TP), chromium (Cr) and mixed liquor suspended solids (MLSS) were measured to monitor treated water quality and treatment efficiency. Microbial community structures in the treatment were also examined using pyrosequencing analysis of 16S rRNA gene and quantitative PCR (qPCR) of the nitrous oxide reductase gene (nosZ). The removal efficiencies of COD, TN, TP, and Cr were estimated to be 98.3%, 98.6%, 93.6%, and 88.5%, respectively, while the system without a continuous augmentation was broken down. The pyrosequencing analysis showed Brachymonas denitrificans to be the most dominant microbial population in the buffering tank (B; 37.5%). Potential polymeric substance degraders (Clostridia), sulfate reducers (Desulfuromonas palmitatis), and sulfur oxidizers (uncultured Thiobacillus) were dominant in the sludge digestion (SD) tank. The denitrifiers assayed by nosZ qPCR were dominant in B and SD. These microbial communities appeared to play important roles in removing nutrients and odor, and reducing sludge in the wastewater treatment plant without chemical pretreatment.

An eco-friendly treatment of tannery wastewater using bioaugmentation with a novel microbial consortium.

A novel microbial consortium (BM-S-1) enriched from natural soils was successfully used to treat tannery wastewater from leather manufacturing industries in Korea on a pilot scale. The objective of this study was to determine whether augmentation with a novel microbial consortium BM-S-1could successfully treat the recalcitrant wastewater without chemical pre-treatment in a tannery wastewater treatment system. Chemical oxygen demand (COD), total nitrogen (TN) and total phosphorus (TP) were monitored for water quality. The microbial population dynamics were analyzed using pyrosequencing, and denitrifying bacteria were quantified using real-time PCR (RT-PCR). The removal efficiencies for COD, TN and TP were greater than 91%, 79%, and 90%, respectively. The dominant phyla in the buffering tank (B), primary aeration (PA), secondary aeration (SA) and sludge digestion tank (SD) were Proteobacteria, Firmicutes, Bacteroidetes, Planctomycetes and Deinococcus-Thermus. Cluster analysis based on the UniFrac distance of the species in the different stages showed that the PA is similar to the SA, whereas the B is similar to the SD. qPCR of the nosZ genes showed the highest abundance of denitrifiers in B, which was increased 734-fold compared to the influent (I). It was hypothesized that anaerobic denitrifiers and the diverse microbial community may play important roles in the biological treatment of tannery wastewater. This technology may also contribute to the full-scale treatment of industrial wastewater containing food processing wastewater and marine sediment with high organic content.

Functions of effective microorganisms in bioremediation of the contaminated harbor sediments.

The aim of this study was to apply loess balls containing effective microorganisms (EM) to the remediation of contaminated harbor sediments, and to thereby elucidate the functions of EM in remediation. Changes in physicochemical, biochemical, and microbiological parameters were measured to monitor the remediation process at a laboratory scale. Treatment with high concentrations of EM stock culture and EM loess balls (4%), and a low concentration of EM loess balls (0.1%) that contained molasses (0.05%) contributed to more rapid removal of malodor. Acetic acid, propionic acid, valeric acid, caponic acid, and lactic acid were rapidly removed in the presence of molasses (0.05% w/w) as a carbon nutrient source, indicating enhanced EM activity by amendment with molasses. Fermentation of molasses by EM showed that more acetic acid was produced compared with other organic acids, and that the majority of organic acids were eventually converted to acetate via intermediate metabolites. Sediment bioremediation tests showed there was no significant difference in eubacterial density with the control and the treatments. However, the density of a Lactobacillus sp. in sediments treated with 0.1% and 4.0% EM loess balls was significantly higher than the control, which indicated the bioaugmentation effect of EM loess balls in the polluted sediments. Treatment with EM loess balls and an appropriate amount of molasses, or other nutrients, will facilitate the remediation of polluted marine sediments by malodor removal, via EM degradation or utilization of offensive organic acids. To our knowledge, this is the first study to remediate contaminated marine (harbor) sediments using EM loess balls and to understand EM function during the bioaugmentation process, both in terms of organic acid metabolism and the dynamics of the engineered microbial community.

Complete reductive dechlorination of tetrachloroethene to ethene by anaerobic microbial enrichment culture developed from sediment.

A mixed, anaerobic microbial enrichment culture, AMEC-4P, was developed that uses lactate as the electron donor for the reductive dechlorination of tetrachloroethene (PCE) to ethene. AMEC-4P consistently and completely converted 2 mM PCE to cis-1,2-dichloroethene (cis-DCE) within 13 days, and the intermediate, cis-DCE, was then completely dechlorinated to ethene after 130 days. Dechlorination rates for PCE to cis-DCE, cis-DCE to VC, and VC to ethene were 243, 27, and 41 µmol/l/day, respectively. Geobacter lovleyi and a Dehalococcoides sp. were identified from their 16S rRNA sequences to be the dominant phylotypes in AMEC-4P.

Characterization of the Bradyrhizobium japonicum galE gene: its impact on lipopolysaccharide profile and nodulation of soybean.

The galE gene from Bradyrhizobium japonicum 61A101C, a soybean endosymbiont, was cloned and characterized. Its deduced amino-acid sequence showed a high similarity with that of other rhizobia. Functional identification of the galE gene was achieved by complementation of a galE mutant strain, PL2, with a series of pKM subclones. Disruption of the B. japonicum galE gene affects the lipopolysaccharide profile compared with that of the wild type, suggesting that galE is responsible for alteration of lipopolysaccharide structure. Examination of nodule formation by the wild-type and galE mutant revealed that the former displayed normal nodule development on soybean roots, whereas the latter showed no nodule formation at all time points examined except for 20 days after inoculation when <10% of soybean formed pseudo-nodules.

Implementation of artificial neural networks (ANNs) to analysis of inter-taxa communities of benthic microorganisms and macroinvertebrates in a polluted stream.

This study was performed to gain an understanding of the structural and functional relationships between inter-taxa communities (macroinvertebrates as consumers, and microbes as decomposers or preys for the invertebrates) in a polluted stream using artificial neural networks techniques. Sediment samples, carrying microorganisms (eubacteria) and macroinvertebrates, were seasonally collected from similar habitats in streams with different levels of pollution. Microbial community taxa and densities were determined using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and 16S rDNA sequence analysis techniques. The identity and density of macroinvertebrates were concurrently determined. In general, differences were observed on grouping by self-organizing map (SOM) in polluted, clean and recovering sites based on the microbial densities, while the community patterns were partly dependent on the sampling period. A Spearman rank order correlation analysis revealed correlations of several eubacterial species with those of macroinvertebrates: a negative correlation was observed between Acidovorax sp. (from polluted sites) and Gammaridae (mostly from the clean site), while Herbaspirillum sp. and Janthinobacterium sp. appeared to have positive correlations with some macroinvertebrate species. The population dynamics of the tolerant texa, Tubificidae and Chironomidae, appeared to be related with changes in the densities of Acidovorax sp. This study revealed community relationships between macroinvertebrates and microorganisms, reflecting the connectivity between the two communities via the food chain. A further physio-ecological and symbiological study on the invertebrate-microorganism relationships will be required to understand the degradation and utilization of detritus in aquatic ecosystems as well as to elucidate the roles of the inter-taxa in the recovery of polluted aquatic environments.

Identification of potential biomarkers for diazinon exposure to Japanese Medaka (Oryzias latipes) using annealing control primers.

A new differential display-polymerase chain reaction (PCR) method based on annealing control primers was used to screen and identify potential biomarkers from differentially expressed genes (DEGs) in medaka exposed to sub-lethal concentration of diazinon (100 ppb). Among the differentially expressed genes identified, the majority were in functional categories of protein biosynthesis, transport and metabolism according to the gene ontology classification. The differential expression of ribosomal protein genes was quantified by real time PCR. The genes encoding ribosomal proteins including L3 and S17 were selected as potential biomarkers for diazinon exposure in medaka fish.

Functional identification of p-cumate operons in the terpene-degrading Rhodococcus sp. strain T104.

We identified a p-cumate degrading gene cluster (designated cmt) with a novel organization in our genomic studies of the terpene-degrading Rhodococcus sp. strain T104. The mutant strain SN140, isolated for the inability to grow on p-cumate, accumulates 2,3-dihydroxy-p-cumate as identified by liquid chromatography-mass spectrometry and 300 MHz proton nuclear magnetic resonance spectroscopy. Reverse transcriptase polymerase chain reaction experiments showed that the cmt genes are operonic and induced specifically by growth on p-cumate. This report is the first example of identifying the genes for p-cumate degradation in a gram-positive organism based on functional data.

Reductive dechlorination of polychlorinated biphenyls as affected by natural halogenated aromatic compounds.

We investigated the effects of halogenated aromatic compounds (HACs) including naturally occurring ones (L-thyroxine, 3-chloro-L-tyrosine, 5-chloroindole, 2-chlorophenol, 4-chlorophenol and chlorobenzene) on polychlorinated biphenyl (PCB) dechlorination in sediment cultures. A PCB-dechlorinating enrichment culture of sediment microorganisms from the St. Lawrence River was used as an initial inoculum. When the culture was inoculated into Aroclor 1248 sediments amended with each of the six HACs, the extent of dechlorination was not enhanced by amendment with HACs. The dechlorination patterns in the HAC-amended sediments were nearly identical to that of the HAC-free sediments except the 3-chloro-L-tyrosine-amended ones where no dechlorination activity was observed. When these sediment cultures were transferred into fresh sediments with the same HACs, the dechlorination specificities remained the same as those of the initial inoculations. Thus, in the present study, the substrate range of the highly selected enrichment culture could not be broadened by the HACs. It appears that HACs affect PCB dechlorination mainly through population selection rather than enzyme induction of single population.

Movement behaviour of Medaka (Oryzias latipes) in response to sublethal treatments of diazinon and cholinesterase activity in semi-natural conditions.

Behavioural changes of medaka (Oryzias latipes) treated with an anticholinesterase insecticide, diazinon (0.1 mg L(-1)), were continuously observed for 4 days in semi-natural conditions. Although variations occurred in individual specimens, the movement tracks appeared differently with typical short-range movement with irregular turns and shaking after the treatments. Eight movement patterns frequently observed before and after the treatments were selected, and the variables characterising the movement patterns were compared quantitatively. The variables were clearly differentiated when the movement patterns were correspondingly matched before and after the treatments (e.g., vertical movements, horizontal movements, etc). Meander and stop duration were highly different among the selected movement patterns. Additionally, different degree of toxic response behaviours could also be detected by quantitative characterisation of the variables. Response behaviour was confirmed with toxicological experiments that show the decrease in the acetylcholine esterase activity in the head and body of specimens. Quantitative investigations on the variables of the movement tracks suggested the usefulness of response behaviour as a monitoring tool for environmental assessment.