Degradation of DDT by γ-hexachlorocyclohexane dehydrochlorinase LinA.

1,1,1-Trichloro-2,2-bis(4-chlorophenyl)-ethane (DDT) is the first synthetic insecticide and one of the most widely used pesticides. The use of DDT has been banned, but it remains one of the most notorious environmental pollutants around the world. In this study, we found that γ-hexachlorocyclohexane (γ-HCH) dehydrochlorinase LinA from a γ-HCH-degrading bacterium, Sphingobium japonicum UT26, converts DDT to 1,1-dichloro-2,2-bis(4-chlorophenyl)-ethylene (DDE). Because of the weak DDT degradation activity of LinA, we could not detect such activity in UT26 cells expressing LinA constitutively. However, the linA-deletion mutant of UT26 harboring a plasmid for the expression of LinA, in which LinA was expressed at a higher level than UT26, showed the DDT degradation activity. This outcome highlights the potential for constructing DDT-degrading sphingomonad cells through elevated LinA expression.

Whole-Genome Sequence of an Arsenite-Oxidizing Bacterium, Pandoraea sp. Strain NE5, Isolated from the Rhizosphere of the Arsenic Hyperaccumulator Pteris vittata.

Pandoraea sp. strain NE5, an arsenite-oxidizing bacterium, was isolated from the rhizosphere of an arsenic hyperaccumulator fern (Pteris vittate). Here, the genome sequence of Pandoraea sp. strain NE5 is announced.

Arsenic uptake by Pteris vittata in a subarctic arsenic-contaminated agricultural field in Japan: An 8-year study.

In this study, the phytoremediation potential of tropical and subtropical arsenic (As) hyperaccumulating fern Pteris vittata in an As contaminated farmland field near an abandoned goldmine was investigated. The tested field is located in a subarctic area of northeast Japan. This study was aimed at decreasing the risk of As in the soil (water-soluble As) with nurturing the soil and respecting the plant life cycle for the sustainable phytoremediation for 8 years. The field was tilled and planted with new seedlings of the fern every spring and the grown fern was harvested every autumn. The biomass and As concentration in fronds, rhizomes and roots of the fern were analyzed separately after harvesting each year. The biomass of the fronds of P. vittata was significantly affected by the yearly change of the weather condition, but As concentration in fronds was kept at 100-150 mg/kg dry weight. The accumulated As in P. vittata was higher than that of As-hyperaccumulator fern Pteris cretica, the native fern in the field trial area. Harvested biomass of P. vittata per plant was also higher than that of P. cretica. More than 43.5 g As/154 m2 (convertible to 2.82 kg of As per hectare) was removed from the farmland field by P. vittata phytoremediation at the end of the 8-year experiment. Because of the short-term plant growth period and soil tilling process, total As in soil did not show significant depletion. However, the water-soluble As in the surface and deeper soil, which is phytoavailable and easily taken in cultivated plants, decreased to 10 µg/L (Japan Environmental Quality Standard for water-soluble As in soil) by the 8-year phytoremediation using P. vittata. These research data elucidate that the tropical and subtropical As hyperaccumulating fern, P. vittata, is applicable for As phytoremediation in the subarctic climate area.

New evidence of arsenic translocation and accumulation in Pteris vittata from real-time imaging using positron-emitting 74As tracer.

Pteris vittata is an arsenic (As) hyperaccumulator plant that accumulates a large amount of As into fronds and rhizomes (around 16,000 mg/kg in both after 16 weeks hydroponic cultivation with 30 mg/L arsenate). However, the sequence of long-distance transport of As in this hyperaccumulator plant is unclear. In this study, we used a positron-emitting tracer imaging system (PETIS) for the first time to obtain noninvasive serial images of As behavior in living plants with positron-emitting 74As-labeled tracer. We found that As kept accumulating in rhizomes as in fronds of P. vittata, whereas As was retained in roots of a non-accumulator plant Arabidopsis thaliana. Autoradiograph results of As distribution in P. vittata showed that with low As exposure, As was predominantly accumulated in young fronds and the midrib and rachis of mature fronds. Under high As exposure, As accumulation shifted from young fronds to mature fronds, especially in the margin of pinna, which resulted in necrotic symptoms, turning the marginal color to gray and then brown. Our results indicated that the function of rhizomes in P. vittata was As accumulation and the regulation of As translocation to the mature fronds to protect the young fronds under high As exposure.

Identification of A Novel Arsenic Resistance Transposon Nested in A Mercury Resistance Transposon of Bacillus sp. MB24.

A novel TnMERI1-like transposon designated as TnMARS1 was identified from mercury resistant Bacilli isolated from Minamata Bay sediment. Two adjacent ars operon-like gene clusters, ars1 and ars2, flanked by a pair of 78-bp inverted repeat sequences, which resulted in a 13.8-kbp transposon-like fragment, were found to be sandwiched between two transposable genes of the TnMERI1-like transposon of a mercury resistant bacterium, Bacillus sp. MB24. The presence of a single transcription start site in each cluster determined by 5'-RACE suggested that both are operons. Quantitative real time RT-PCR showed that the transcription of the arsR genes contained in each operon was induced by arsenite, while arsR2 responded to arsenite more sensitively and strikingly than arsR1 did. Further, arsenic resistance complementary experiments showed that the ars2 operon conferred arsenate and arsenite resistance to an arsB-knocked out Bacillus host, while the ars1 operon only raised arsenite resistance slightly. This transposon nested in TnMARS1 was designated as TnARS1. Multi-gene cluster blast against bacteria and Bacilli whole genome sequence databases suggested that TnMARS1 is the first case of a TnMERI1-like transposon combined with an arsenic resistance transposon. The findings of this study suggested that TnMERI1-like transposons could recruit other mobile elements into its genetic structure, and subsequently cause horizontal dissemination of both mercury and arsenic resistances among Bacilli in Minamata Bay.

Comparative geochemical evaluation of toxic metals pollution and bacterial communities of industrial effluent tributary and a receiving estuary in Nigeria.

Toxic metals/metalloid contaminations of estuarine sediments due to compromised tributaries arouse significant interest in studying bacterial community that triggers natural attenuation processes. Geo-accumulation index (Igeo), contamination factor (CF), pollution load index (PLI), and Hakanson potential ecological risk index (RI) as a sum of risk factors (Er) were used to quantify toxic metal/metalloid-pollution status of Lagos Lagoon (2W) and 'Iya-Alaro' tributary (4W) sediments in comparison with pristine 'Lekki Conservation Centre' sediment (L1-B). Bacteriology of the ecosystems was based on culture-independent analyses using pyrosequencing. 2W and 4W were extremely contaminated with mercury (Igeo > 7), whereas, cadmium contamination was only observed in 4W. The two ecosystems were polluted with toxic metal based on PLI, where mercury (Er = 2900 and 1900 for 4W and 2W, respectively) posed very high ecological risks. Molecular fingerprinting revealed that Proteobacteria, Firmicutes, and Acidobacteria predominately contributed the 20 most abundant genera in the two ecosystems. The 240 and 310 species present in 2W and 4W, respectively, but absent in L1-B, thrive under the metal concentrations in the polluted hydrosphere. Whereas, the 58,000 species missing in 2W and 4W but found in L1-B would serve as indicators for systems impacted with metal eco-toxicity. Despite toxic metal pollution of the ecosystems understudied, bacterial communities play vital roles in self-recovery processes occurring in the hydrosphere.

Bio-oxidation of elemental mercury during growth of mercury resistant yeasts in simulated hydrosphere.

Transformation of metallic mercury (Hg°) to mercuric ion (Hg2+) in hydrosphere is the entrance of mercury cycle in water environments and leads to toxicological impact of serious global concern. Two yeast strains of Yarrowia (Idd1 and Idd2) isolated from Hg-contaminated sediments were studied for their mediating role in Hg° dissolution and oxidation. Growth of the Yarrowia cells in Hg-free liquid medium, incubated for 5 d in closed air-tight systems containing Hg°, produced extracellular polymeric substances (EPS). Approximately 230 (±5.7) ng and 120 (±6.8) ng of the dissolved Hg° were oxidized to Hg2+ by the cultures of Idd1 and Idd2, respectively, 5 day post-inoculation. Transmission electron microscopy (TEM) and X-ray energy dispersive spectrophotometry (XEDS) analysis of the EPS and cell mass revealed the presence of extracellular Hg nanoparticles, presumably HgS, as an indication of EPS-Hg complexation that is useful for Hg° dissolution and its eventual oxidation to Hg2+ by the cells. Fourier transmission infra-red (FTIR) analyses of the EPS and cell-mass during Hg-oxidation revealed that amine and carbonyl groups were used by EPS for Hg complexation. Our findings provided information about mediatory role played by Yarrowia (Idd1 and Idd2) in hydrosphere in biogeochemical cycling of Hg.

Serum lipid and bone metabolism effects of Toremifene vs. Letrozole as adjuvant therapy for postmenopausal early breast cancer patients: results of a multicenter open randomized study.

A prospective randomized phase II trial was conducted to evaluate the time course effects of toremifene (TOR) and letrozole (LET), as adjuvant hormone therapy, on serum lipid profiles and bone metabolism in estrogen receptor (ER)-positive, postmenopausal breast cancer patients.Fifty-four postmenopausal breast cancer patients [ER positive, HER2 negative, T1-2, node metastases (n = 0-3), M0] who had undergone curative resection were enrolled. They were randomized to receive either TOR 40 mg/day or LET 2.5 mg/day as adjuvant hormone therapy. Serum lipids and bone markers were measured prior to, and again at 6, 12, and 24 months after initiation of treatment. Changes in serum lipids and bone markers were compared. Serum levels of total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) were decreased compared with the baseline values at 6 months in 6.5 and 14.0% of patients, respectively, receiving TOR. Lipid levels did not change in patients administered LET. Significant differences were observed in TC and LDL-C between the two groups at 12 and 24 months. In the TOR group, serum bone-specific alkaline phosphatase (BAP) was decreased by 25.0% at 12 months, and serum cross-linked N-telopeptide of type-I collagen (NTx) was decreased by 13.6% at 6 months, and these reductions were maintained for at least 24 months. In contrast, in the LET group, serum BAP did not change and NTx was increased by 16.0% at 6 months and by 18.6% at 24 months, as compared with the baseline.TOR and LET exert different effects on serum lipid profiles and bone metabolism markers. The effects of TOR, as adjuvant hormone therapy, on both lipids and bone metabolism in postmenopausal breast cancer patients are superior to those of LET.

Body mass index and menopausal disorders during menopause affect vasomotor symptoms of postmenopausal Japanese breast cancer patients treated with anastrozole: a prospective multicenter cohort study of patient-reported outcomes.

BACKGROUND: Adverse events related to endocrine therapies have a major impact not only on patients' quality of life but also on treatment discontinuation. Although vasomotor symptoms induced by aromatase inhibitors are frequently recognized, risk factors, especially for Japanese women, are not well reported. To identify risk factors for vasomotor symptoms of Japanese breast cancer patients treated with adjuvant anastrozole, we conducted a prospective cohort study based on patient-reported outcomes (PROs). PATIENTS AND METHODS: For this prospective cohort study (SAVS-JP, UMIN000002455), 391 postmenopausal Japanese estrogen receptor-positive breast cancer patients who were treated with adjuvant anastrozole were recruited from 28 centers. The PRO assessment was obtained from a self-reported questionnaire at baseline, 3, 6, 9 and 12 months between August 2009 and April 2012. Vasomotor symptoms, comprising hot flashes, night sweats, and cold sweats, were categorized into four grades (none, Grade 1: mild, Grade 2: moderate, Grade 3: severe). Pre-existing symptoms were only included if they had become worse than at baseline. RESULTS: Hot flashes, night sweats, and cold sweats at baseline were reported by 20.5, 15.1, and 8.2 % of the patients, respectively, and new appearance or worsening of symptoms in comparison with baseline by 38.4, 29.3, and 28.7 %, respectively. About 80 % of newly occurring symptoms were Grade 1, and less than 5 % were Grade 3. Vasomotor symptoms were reported by 201 out of 362 patients (55.5 %) during the first year and the mean time to onset was 5.6 months. Patients with vasomotor symptoms were significantly younger (mean 62.8 years, range 38-86 vs 64.7 years, range 37-84; p = 0.02), had higher body mass index (BMI) (23.4 kg/m2, range 15.8-39.9 vs 22.4 kg/m2, range 15.8-34.9; p = 0.01), had vasomotor symptoms sooner after menopause (12.4 years, range 0-51 vs 15.1 years, range 1-37; p = 0.002), and had more menopausal disorders during menopause (63.3 vs 36.7 %; p = 0.002). Multivariate analysis showed that BMI [odds ratio (OR) 1.09 per unit of increase, 95 % confidence interval (CI) 1.02-1.16; p = 0.009] and experiencing menopausal disorders (OR 2.11, 95 % CI 1.35-3.30; p = 0.001) were significantly associated with vasomotor symptoms. CONCLUSION: High BMI and experiencing menopausal disorders at menopause were found to be significantly associated with the occurrence of vasomotor symptoms. These findings are expected to prove useful for the management of postmenopausal Japanese women treated with aromatase inhibitors.

Mercury removal during growth of mercury tolerant and self-aggregating Yarrowia spp.

Ecotoxicological implications of mercury (Hg) pollution of hydrosphere require effective Hg-removal strategies as antidote to the environmental problems. Mercury-tolerant yeasts, Yarrowia spp. Idd1 and Idd2 strains, were studied for intracellular accumulation and extracellular micro-precipitation of Hg during growth stage of the yeast strains. In a liquid medium containing 870 (±23.6) µg of bioavailable Hg2+, 419.0 µg Hg2+ (approx.) was taken up by the wet biomasses of the yeast strains after 48 h post-inoculation. Large portion of the adsorbed Hg was found in cell wall (approx. 49-83 %) and spheroplast (approx. 62-89 %). Negligible quantities of Hg were present in the mitochondria (0.02-0.02 %), and appreciable amount of Hg was observed in nuclei and cell debris (15.2-65.3 %) as evidence of bioaccumulation. Extracellular polymeric substances (EPS) produced by the growing Yarrowia cells was a complex of protein, carbohydrates and other substances, immobilizing 43.8 (±0.7)-58.7 (±1.0) % of initial Hg in medium as micro-precipitates, while 10.13 ± 0.4-39.2 ± 4.3 % Hg content was volatilized. Transmission electron microscopy coupled with X-ray energy dispersive spectrophotometry confirmed the cellular removal of Hg and formation of EPS-Hg complex colloids in the surrounding bulk solution as micro-precipitates in form of extracellular Hg-nanoparticles. Hg mass balance in the bio-sequestration experiment revealed excellent Hg removal (>97 %) from the medium (containing ≤16 µg ml-1 Hg2+) by the yeast strains via bioaccumulation, volatilization and micro-precipitation. The yeast strains are also effectively applicable in biological purification technology for Hg contaminated water because of their high self-aggregation activity and separatability from the aquatic environments. Graphical abstract Yarrowia species are oligotrophic marine yeasts that exhibited great potentials for mercuric ion remediation technologies, which are classified into four categories based on the process acting on the metal. These include immobilization through biosorption, compartmentation via bioaccumulation, separation from bulk solution via micro-precipitation upon EPS-Hg complex formation, and destruction that is a process to reduce the mercuric ion to metallic mercury.

Extracellular mercury sequestration by exopolymeric substances produced by Yarrowia spp.: Thermodynamics, equilibria, and kinetics studies.

Exopolymeric substances (EPS) produced by highly mercury-resistant strains of the yeast Yarrowia spp. (Idd1 and Idd2) were isolated and studied for their mercury binding potential. Excellent yield (approximately 0.3 g EPS per gram biomass) of soluble EPS in medium with 3% glucose was observed in the Yarrowia cultures 7 day post-inoculation. A gram dry weight of the EPS consists mainly of carbohydrates (0.4 g), protein (0.3-0.4 g), uronic acid (0.02 g), and nucleic acids (0.002 g). Mercury interactions with the biopolymer were measured as uptake kinetics from a simulated aquatic system and modelled with thermodynamics and calculated mass action equilibria. The EPS forms a complex with Hg2+ in water with small activation energy (≤2 kJ mol-1), achieving about 30 mg Hg2+ adsorption per gram dry weight of EPS. The adsorption models confirmed complexation of Hg2+ by the EPS via heterogeneous multilayer adsorption that obey second-order kinetics at constant rate of 4.0 and 8.1 mg g-1 min-1. The EPS used chemisorption as rate-limiting step that controls the uptake of Hg2+ from aquatic systems during micro-precipitation as bio-removal strategy. The EPS are promising biotechnological tools to design bioreactors for treatment of mercury-rich industrial wastewater.

Development of suitable hydroponics system for phytoremediation of arsenic-contaminated water using an arsenic hyperaccumulator plant Pteris vittata.

In this study, we found that high-performance hydroponics of arsenic hyperaccumulator fern Pteris vittata is possible without any mechanical aeration system, if rhizomes of the ferns are kept over the water surface level. It was also found that very low-nutrition condition is better for root elongation of P. vittata that is an important factor of the arsenic removal from contaminated water. By the non-aeration and low-nutrition hydroponics for four months, roots of P. vittata were elongated more than 500 mm. The results of arsenate phytofiltration experiments showed that arsenic concentrations in water declined from the initial concentrations (50 µg/L, 500 µg/L, and 1000 µg/L) to lower than the detection limit (0.1 µg/L) and about 80% of arsenic removed was accumulated in the fern fronds. The improved hydroponics method for P. vittata developed in this study enables low-cost phytoremediation of arsenic-contaminated water and high-affinity removal of arsenic from water.

Risk factors for joint symptoms in postmenopausal Japanese breast cancer patients treated with anastrozole: a prospective multicenter cohort study of patient-reported outcomes.

BACKGROUND: Endocrine treatment-related adverse events have a strong impact on patients' quality of life and sometimes result in treatment discontinuation. Since joint symptoms are the most frequently recognized side effect of aromatase inhibitors, evaluation of associated risk factors may yield significant findings. PATIENTS AND METHODS: A total of 391 postmenopausal Japanese women with estrogen receptor-positive breast cancer and treated with adjuvant anastrozole were enrolled from 28 centers for assessment of patient-reported outcomes (PROs) in this prospective cohort study (SAVS-JP, UMIN000002455). Patients completed the self-report questionnaire at baseline and after 3, 6, 9, and 12 months of treatment for evaluation of frequency of treatment-related joint symptoms (arthralgia, decrease in range of joint motion, and joint stiffness). RESULTS: We obtained PROs from 362 patients (92.6 %) at baseline and at one or more subsequent points. New or worsening from baseline of joint symptoms were reported by 260 patients (71.8 %). More than 90 % of the symptoms were mild or moderate and nearly 80 % had occurred by 6 months. Multivariate analysis showed that a short time span after menopause [odds ratio (OR) 0.95, 95 % confidence interval (CI) 0.90-0.99; P = 0.02] and adjuvant chemotherapy (OR 2.29, 95 % CI 1.06-4.95; P = 0.03) were significant independent risk factors for joint symptoms. No significant relationships between body mass index (BMI) and joint symptoms were identified. Eighteen patients discontinued treatment during the 1st year and eight of them reported joint symptoms. CONCLUSION: Taking into consideration that PROs may yield higher prevalence rates than physician ratings for symptoms published in pivotal clinical trials, we found that a short time span after menopause and use of adjuvant chemotherapy, but not high BMI, were significantly associated with joint symptoms. These findings might prove useful for counseling before initiating treatment with adjuvant aromatase inhibitors in postmenopausal Japanese women.

Association of body mass index with risk of luminal A but not luminal B estrogen receptor-positive and HER2-negative breast cancer for postmenopausal Japanese women.

BACKGROUND: The impact of body mass index (BMI) on the risk of postmenopausal estrogen receptor (ER)-positive breast cancers has been well documented. However, the mechanism for the impact of BMI on the etiology of luminal A and luminal B subtypes has not yet been identified. METHODS: We analyzed associations between BMI and breast cancers stratified by immunohistochemically defined intrinsic subtypes, and 1,297 Japanese women (615 breast cancer patients and 682 healthy women from a breast cancer screening program) were enrolled in a case-control study. ER-positive/human epidermal growth factor receptor 2 (HER2)-negative breast cancers were classified into luminal A and B subtypes according to Ki67 expression levels. RESULTS: Higher BMI was significantly positively associated with postmenopausal breast cancer risk for one-unit increase in BMI (adjusted odds ratio (aOR) 1.09, 95 % confidence interval (CI) 1.04-1.15; P = 0.0008). Analyses of postmenopausal women revealed that BMI was consistently and exclusively associated with luminal A incidence (aOR 1.18, 95 % CI 1.10-1.26; P < 0.0001). When BMI was divided into three categories corresponding to those of controls, among postmenopausal women, the observed positive association was confined to luminal A (high vs low, aOR 2.98, 95 % CI 1.53-5.80; P < 0.005), but not luminal B (aOR 0.95, 95 % CI 0.47-1.91) subtypes. CONCLUSIONS: We observed that BMI was significantly positively associated with increased risk of postmenopausal breast cancer for Japanese women with luminal A, but not with luminal B tumor subtype.

Mercury bioremoval by Yarrowia strains isolated from sediments of mercury-polluted estuarine water.

Difference in mercuric ion removal by resting and growing cells of two mercury-resistant yeast strains, identified as Yarrowia spp. (strains Idd1 and Idd2), were studied. Resting cells of strain Idd2 exhibited high maximum Hg(2+) removal capacity (59 mg mercury per g dry cell weight [gdw(-1)]) by adsorption than those of resting cells of strain Idd1 (32 mg gdw(-1)). The resting cells of strain Idd2 exhibited a higher Hg(2+) desorption capacity using CaCl2 (68 %) and EDTA (48 %) than strain Idd1, depicting weaker binding of Hg(2+) onto strain Idd2 unlike strain Idd1. The actively growing yeast cells showed opposite Hg removal characteristics to those of the resting cells. Strain Idd1 adsorbed less Hg(2+) from culture medium supplemented with Hg(2+) than strain Idd2. However, the growing strain Idd1 reduced and vaporized 27 % of supplemented Hg(2+) as metallic mercury (Hg(0)), while the growing strains Idd2 vaporized 15 % of the supplemented Hg(2+). These two yeast strains are potential biotechnological tools for the eventual bioremediation of polluted aquatic systems.

Phase II study of S-1 in combination with trastuzumab for HER2-positive metastatic breast cancer.

AIM: We undertook a prospective phase II study to evaluate the efficacy of S-1 plus trastuzumab combination regimen for human epidermal-growth factor receptor-2 (HER2)-positive metastatic breast cancer (MBC). PATIENTS AND METHODS: HER2-positive MBC patients received oral administration of S-1 (80 mg/m2/day, days 1 to 28, every 6 weeks) and intravenous weekly trastuzumab (2 mg/kg), according to the results of a prior Phase I trial of our group. RESULTS: A total of 28 patients were enrolled and received a median of 3.5 (range 1-10) cycles of treatment. Overall response rate and clinical benefit rate were 53.6% and 75.0%, respectively. Progression-free survival was 30 weeks. With regard to grade 3 and 4 adverse effects, leucopenia, neutropenia, increase in serum alanine aminotransferase, and diarrhea were observed. CONCLUSION: Combination of S-1 and trastuzumab was tolerable and had excellent efficacy with good response and disease control in this trial.

Effectiveness of heat treatment to protect introduced denitrifying bacteria from eukaryotic predatory microorganisms in a pilot-scale bioreactor.

Bioaugmentation of bioreactor systems with pre-cultured bacteria has proven difficult because inoculated bacteria are easily eliminated by predatory eukaryotic-microorganisms. Here, we demonstrated an intermediate thermal treatment was effective for protecting introduced denitrifying bacteria from eukaryotic predators and consequently allowed the inoculated bacteria to survive longer in a denitrification reactor.

Community structure and population dynamics of ammonia oxidizers in composting processes of ammonia-rich livestock waste.

This study investigated the relationship between the population dynamics of ammonia-oxidizing bacteria (AOB) and archaea (AOA), and changes in the concentrations of nitrogenous compounds during ammonia-rich livestock waste-composting processes. The data showed that ammonia in beef and dairy cow livestock waste-composting piles was slowly oxidized to nitrite and nitrate after approximately 21-35 days under thermophilic or moderately thermophilic and mesophilic conditions. Real-time quantitative PCR (qPCR) assays showed a relative abundance of betaproteobacterial AOB during ammonia oxidation but did not detect AOA in any composting stage. Furthermore, real-time qPCR and terminal-restriction fragment length polymorphism (T-RFLP) analyses for the AOB in two composting processes (beef and dairy cow livestock waste) out of the three studied found that thermophilic or moderately thermophilic uncultured betaproteobacterial AOB from the "compost AOB cluster" contributed to ammonia oxidation during hot composting stages. Non-metric multidimensional scaling analyses of the data from T-RFLP showed that only a few analogous species predominated during composting of beef, dairy cow and pig livestock wastes, and thus, the AOB community structures in the three composting piles operating under different conditions were similar. AOB-targeted clone library analyses revealed that uncultured members of the "compost AOB cluster", which could be clearly distinguished from the authentic species of the genus Nitrosomonas, were the major constituents of the AOB populations. These results suggested that a limited and unique species of AOB played a role in ammonia oxidation during the composting of ammonia-rich livestock waste.

Gene cluster and regulation system for 1,1-dichloro-2,2-bis(4-chlorophenyl)ethylene (DDE) degradation in Janibacter sp. TYM3221.

A DDE-degrading bacterium, Janibacter sp. TYM3221, is able to grow on biphenyl and degrades 1,1-dichloro-2,2-bis(4-chlorophenyl)ethylene (DDE) via a meta-ring cleavage pathway. The bphAa gene, encoding a biphenyl dioxygenase large subunit, was previously demonstrated to be involved in the degradation of DDE in TYM3221. The bph gene cluster, containing orf2 and bphDAaAbAcAdBCST was cloned and characterized. Reverse transcription-PCR (RT-PCR) analysis indicated that these genes were transcribed as an operon. The real-time RT-PCR on orf2, bphAa, bphC, and bphS suggest the presence of the inducible orf2 promoter (orf2p) and constitutive bphAa promoter (bphAap). The TYM3221 bphST conducted biphenyl-dependent inducible activation plus constitutive basal activation of orf2p and constitutive activation of bphAap in a rhodococcal host strain, Rhodococcus erythropolis IAM1399, suggesting that expression of the TYM3221 bph operon depends on the bphST-coded two-component regulatory system. Both of these promoters were also induced by the bphS1T1 of a biphenyl degrader, Rhodococcus jostii RHA1, and contained the 24-bp consensus sequences of RHA1 bphS1T1-dependent promoters. The replacement of RHA1 bphS1 with TYM3221 bphS in combination with RHA1 bphT1 suggests that TYM3221 bphS is responsible for low inducible and high constitutive activation of orf2p in IAM1399 by the TYM3221 bphST-system. Expression of bphAaAbAcAdBC in IAM1399 resulted in the transformation of DDE to the meta-ring cleavage product via 2,3-hydroxylation, suggesting that these genes are involved in DDE degradation.

Characterization of nitrate and nitrite utilization system in Rhodococcus jostii RHA1.

A polychlorinated-biphenyl degrader, Rhodococcus jostii RHA1, has the potential to be used in soil for the remediation of environmental contamination. It has been found that RHA1 genes, ro06365 (narK) and ro06366, encoding a nitrate/nitrite transporter and nitrite reductase, respectively, were highly upregulated during the growth in sterile soil. In this study, these genes and ro00862, a paralog of ro06366 were characterized to reveal the nitrate and nitrite utilization systems of RHA1. The transcriptional induction of ro06366 (nirB1) and ro00862 (nirB2) by either nitrate or nitrite was revealed by qRT-PCR. Deletion mutants for each gene exhibited retarded growth on either nitrate or nitrite as a sole nitrogen source. Furthermore, their double mutant, Dnit, grew on and consumed neither nitrate nor nitrite as a sole nitrogen source, suggesting that both nirB1 and nirB2 are involved in the utilization of nitrite and nitrate. A narK mutant, DnarK, exhibited no growth on nitrate and retarded growth on nitrite as the sole nitrogen source. DnarK showed no consumption of nitrate and reduced consumption of nitrite, suggesting that narK is essential for nitrate uptake and is partially involved in nitrite uptake. The induced transcription of nirB1, nirB2, and narK was repressed in the presence of 3 mM ammonium or more. The upregulation of nirB1 and narK in sterilized soil containing ammonium and nitrate suggests that the ammonium concentration of the sterilized soil is equivalent to less than 3 mM. The unique nitrogen metabolism system of RHA1 and its importance for the growth in soil are discussed.