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A pilot-scale biocover was constructed at a sanitary landfill and the mitigation of methane and odor compounds was compared between the summer and non-summer seasons. The average inlet methane concentrations were 22.0%, 16.3%, and 31.3%, and the outlet concentrations were 0.1%, 0.1%, and 0.2% during winter, spring, and summer, respectively. The odor removal efficiency was 98.0% during summer, compared to 96.6% and 99.6% during winter and spring, respectively. No deterioration in methane and odor removal performance was observed even when the internal temperature of the biocover increased to more than 40°C at midday during summer. During summer, the packing material simultaneously degraded methane and dimethyl sulfide (DMS) under both moderately thermophilic (40-50°C) and mesophilic conditions (30°C). Hyphomicrobium and Brevibacillus, which can degrade methane and DMS at 40°C and 50°C, were isolated. The diversity of the bacterial community in the biocover during summer did not decrease significantly compared to other seasons. The thermophilic environment of the biocover during summer promoted the growth of thermotolerant and thermophilic bacterial populations. In particular, the major methane-oxidizing species were Methylocaldum spp. during summer and Methylobacter spp. during the nonsummer seasons. The performance of the biocover remained stable under moderately thermophilic conditions due to the replacement of the main species and the maintenance of bacterial diversity. The information obtained in this study could be used to design biological processes for methane and odor removal during summer and/or in subtropical countries.

Petroleum-contaminated soil is considered among the most important potential anthropogenic atmospheric methane sources. Additionally, various rhizoremediation factors can affect methane emissions by altering soil ecosystem carbon cycles. Nonetheless, greenhouse gas emissions from soil have not been given due importance as a potentially relevant parameter in rhizoremediation techniques. Therefore, in this study we sought to investigate the effects of different plant and soil amendments on both remediation efficiencies and methane emission characteristics in dieselcontaminated soil. An indoor pot experiment consisting of three plant treatments (control, maize, tall fescue) and two soil amendments (chemical nutrient, compost) was performed for 95 days. Total petroleum hydrocarbon (TPH) removal efficiency, dehydrogenase activity, and alkB (i.e., an alkane compound-degrading enzyme) gene abundance were the highest in the tall fescue and maize soil system amended with compost. Compost addition enhanced both the overall remediation efficiencies, as well as pmoA (i.e., a methane-oxidizing enzyme) gene abundance in soils. Moreover, the potential methane emission of diesel-contaminated soil was relatively low when maize was introduced to the soil system. After microbial community analysis, various TPH-degrading microorganisms (Nocardioides, Marinobacter, Immitisolibacter, Acinetobacter, Kocuria, Mycobacterium, Pseudomonas, Alcanivorax) and methane-oxidizing microorganisms (Methylocapsa, Methylosarcina) were observed in the rhizosphere soil. The effects of major rhizoremediation factors on soil remediation efficiency and greenhouse gas emissions discussed herein are expected to contribute to the development of sustainable biological remediation technologies in response to global climate change.

Several non-methylotrophic bacteria have been reported to improve the growth and activity of methanotrophs; however, their interactions remain to be elucidated. We investigated the interaction between Methylocystis sp. M6 and Microbacterium sp. NM2. A batch co-culture experiment showed that NM2 markedly increased the biomass and methane removal of M6. qPCR analysis revealed that NM2 enhanced both the growth and methane-monooxygenase gene expression of M6. A fed-batch experiment showed that co-culture was more efficient in removing methane than M6 alone (28.4 vs. 18.8 μmol·l-1·d-1), although the biomass levels were similar. A starvation experiment for 21 days showed that M6 population remained stable while NM2 population decreased by 66% in co-culture, but the results were opposite in pure cultures, indicating that M6 may cross-feed growth substrates from NM2. These results indicate that M6 apparently had no negative effect on NM2 when M6 actively proliferated with methane. Interestingly, a batch experiment involving a dialysis membrane indicates that physical proximity between NM2 and M6 is required for such biomass and methane removal enhancement. Collectively, the observed interaction is beneficial to the methanotroph but adversely affects the non-methylotroph; moreover, it requires physical proximity, suggesting a tight association between methanotrophs and non-methylotrophs in natural environments.

We demonstrated the quantitative detection of a toxin-producing Microcystis aeruginosa (M. aeruginosa) strain with the laboratory protocol of the NanoGene assay. The NanoGene assay was selected because its laboratory protocol is in the process of being transplanted into a portable system. The mcyD gene of M. aeruginosa was targeted and, as expected, its corresponding fluorescence signal was linearly proportional to the mcyD gene copy number. The sensitivity of the NanoGene assay for this purpose was validated using both dsDNA mcyD gene amplicons and genomic DNAs (gDNA). The limit of detection was determined to be 38 mcyD gene copies per reaction and 9 algal cells/ml water. The specificity of the assay was also demonstrated by the addition of gDNA extracted from environmental algae into the hybridization reaction. Detection of M. aeruginosa was performed in the environmental samples with environmentally relevant sensitivity (~105 algal cells/ml) and specificity. As expected, M. aeruginosa were not detected in nonspecific environmental algal gDNA over the range of 2 × 100 to 2 × 107 algal cells/ml.

The dye decolorization rate in a cell-free culture broth of the white-rot fungus Trametes versicolor CBR43 was studied, including the effects of inhibitors of NaCl, Zn(II), and Cd(II) on dye decolorization activity. The maximum rates of dye decolorization in cell-free culture broth were 1,410, 44.7, 41.2, and 0.19 μmol·l-1·min-1 for Acid Blue 62, Acid Black 175, Reactive Blue 4, and Acid Red 114, respectively. The inhibition effects of NaCl, Zn(II), and Cd(II) on dye decolorization were quantitatively compared using the half maximal inhibition concentration (IC50), which indicates the concentration of an inhibitor required for 50% inhibition. Based on IC50 values, dye decolorization in the cell-free culture broth of CBR43 was most potently inhibited by Cd(II), whereas the inhibitory effect of NaCl was relatively low. The dye decolorization rates and IC50 data can be used in the design and development of a dyewastewater treatment process using T. versicolor CBR43 and its operating factors.

Several non-methylotrophic bacteria have been reported to improve the growth and activity of methanotrophs; however, their interactions remain to be elucidated. We investigated the interaction between

Soil burial is the most widely used disposal method for infected pig carcasses, but composting has gained attention as an alternative disposal method because pig carcasses can be decomposed rapidly and safely by composting. To understand the pig carcass decomposition process in soil burial and by composting, pilot-scale test systems that simulated soil burial and composting were designed and constructed in the field. The envelope material samples were collected using special sampling devices without disturbance, and bacterial community dynamics were analyzed by high-throughput pyrosequencing for 340 days. Based on the odor gas intensity profiles, it was estimated that the active and advanced decay stages were reached earlier by composting than by soil burial. The dominant bacterial communities in the soil were aerobic and/or facultatively anaerobic gram-negative bacteria such as Pseudomonas, Gelidibacter, Mucilaginibacter, and Brevundimonas. However, the dominant bacteria in the composting system were anaerobic, thermophilic, endospore-forming, and/or halophilic gram-positive bacteria such as Pelotomaculum, Lentibacillus, Clostridium, and Caldicoprobacter. Different dominant bacteria played important roles in the decomposition of pig carcasses in the soil and compost. This study provides useful comparative date for the degradation of pig carcasses in the soil burial and composting systems.

We demonstrated the quantitative detection of a toxin-producing

Co-digesting molasses wastewater and sewage sludge was evaluated for hydrogen production by response surface methodology (RSM). Batch experiments in accordance with various dilution ratios (40- to 5-fold) and waste mixing composition ratios (100:0, 80:20, 60:40, 40:60, 20:80, and 0:100, on a volume basis) were conducted. Volatile solid (VS) concentration strongly affected the hydrogen production rate and yield compared with the waste mixing ratio. The specific hydrogen production rate was predicted to be optimal when the VS concentration ranged from 10 to 12 g/l at all the mixing ratios of molasses wastewater and sewage sludge. A hydrogen yield of over 50 ml H2/g VS(removed) was obtained from mixed waste of 10% sewage sludge and 10 g/l VS (about 10-fold dilution ratio). The optimal chemical oxygen demand/ total nitrogen ratio for co-digesting molasses wastewater and sewage sludge was between 250 and 300 with a hydrogen yield above 20 ml H2/g VS(removed).

Density of catalytic organisms can determine the biodegradation capacity and specific biodegradation rate (SBR). A new index, biodegradation capacity utilization (BCU, %), was developed for estimating the extent of actual biodegradation of a gas compound over the full capacity. Three methanotrophic cultures were serially diluted (1-1/25), and methane SBR and BCU were measured. Consistently, biomass reduction increased the SBR and decreased the BCU. Linearity (p0.05, r0.97) between the BCU and cell density indicated the reflection of biodegradation capacity by BCU. Therefore, BCU is indicative of whether the density of catalytic organisms is pertinent for SBR evaluation of low-soluble gaseous compounds.

The dye decolorization rate in a cell-free culture broth of the white-rot fungus

The anaerobic oxidation of methane (AOM) in anaerobic sewage sludge was characterized. The net methane oxidation was observed in samples amended with methane plus sulfate or with methane alone, whereas methane formation was observed in the samples without methane, indicating that methane oxidation and formation occurred simultaneously. The ratio of the net methane oxidation rate to H2S formation was 100:1, suggesting that the AOM was not closely associated with sulfate reduction in the anaerobic sludge. The net AOM was positively associated with the methane concentration and sludge dilution ratio. However, the rate of AOM was negatively correlated with organic substrate (acetate) concentration. Therefore, the production and oxidation of methane could be controlled by environmental conditions and dissolved organic compounds in the bulk solution.

Methylotrophs within biological activated carbon (BAC) systems have not received attention although they are a valuable biological resource for degradation of organic pollutants. In this study, methylotrophic populations were monitored for four consecutive seasons in BAC of an actual drinking water plant, using ribosomal tag pyrosequencing. Methylotrophs constituted up to 5.6% of the bacterial community, and the methanotrophs Methylosoma and Methylobacter were most abundant. Community comparison showed that the temperature was an important factor affecting community composition, since it had an impact on the growth of particular methylotrophic genera. These results demonstrated that BAC possesses a substantial methylotrophic activity and harbors the relevant microbes.

Current ecological knowledge of methanotrophic biofilms is incomplete, although they have been broadly studied in biotechnological processes. Four individual DNA samples were prepared from a methanotrophic biofilm, and a multiplex 16S rDNA pyrosequencing was performed. A complete library (before being de-multiplexed) contained 33,639 sequences (average length, 415 nt). Interestingly, methanotrophs were not dominant, only making up 23% of the community. Methylosinus, Methylomonas, and Methylosarcina were the dominant methanotrophs. Type II methanotrophs were more abundant than type I (56 vs. 44%), but less richer and diverse. Dominant non-methanotrophic genera included Hydrogenophaga, Flavobacterium, and Hyphomicrobium. The library was de-multiplexed into four libraries, with different sequencing efforts (3,915-20,133 sequences). Sorrenson abundance similarity results showed that the four libraries were almost identical (indices > 0.97), and phylogenetic comparisons using UniFrac test and P-test revealed the same results. It was demonstrated that the pyrosequencing was highly reproducible. These survey results can provide an insight into the management and/or manipulation of methanotrophic biofilms.

A methane-oxidizing bacterium was isolated from the enriched culture of a landfill cover soil. The closest relative of the isolate, designated M6, is Methylocystis sp. Based on a kinetic analysis, the maximum specific methane oxidation rate and saturation constant were 4.93 mmol·g--dry cell weight--1·h⁻¹ and 23 microM, respectively. This was the first time a kinetic analysis was performed using pure methanotrophic culture. The methane oxidation by M6 was investigated in the presence of aromatic (m- and p-xylene and ethylbenzene) or sulfur (hydrogen sulfide, dimethyl sulfide, methanthiol) compounds. The methane oxidation was inhibited by the presence of aromatic or sulfur compounds.

Three hundred and seventy-four rhizobacteria were isolated from the rhizosphere soil (RS) or rhizoplane (RP) of Echinochloa crus-galli, Carex leiorhyncha, Commelina communis, Persicaria lapathifolia, Carex kobomugi, and Equisetum arvense, grown in contaminated soil with petroleum and heavy metals. The isolates were screened for plant growth-promoting potential (PGPP), including indole acetic acid (IAA) productivity, 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase activity, and siderophore(s) synthesis ability. IAA production was detected in 86 isolates (23.0%), ACC deaminase activity in 168 isolates (44.9%), and siderophore(s) synthesis in 213 isolates (57.0%). Among the rhizobateria showing PGPP, 162 rhizobacteria had multiple traits showing more than two types of PGPP. The PGPP-having rhizobateria were more abundant in the RP (82%) samples than the RS (75%). There was a negative correlation (-0.656, p0.05) between the IAA-producers and the ACC deaminase producers. Clustering analysis by principal component analysis showed that RP was the most important factor influencing ecological distribution and physiological characterization of PGPP-possesing rhizobateria.

The role of plant growth-promoting rhizobacteria (PGPR) in the phytoremediation of heavy-metal-contaminated soils is important in overcoming its limitations for field application. A plant growth-promoting rhizobacterium, Serratia sp. SY5, was isolated from the rhizoplane of barnyard grass (Echinochloa crus-galli) grown in petroleum and heavy-metal-contaminated soil. This isolate has shown capacities for indole acetic acid production and siderophores synthesis. Compared with a non-inoculated control, the radicular root growth of Zea mays seedlings inoculated with SY5 can be increased by 27- or 15.4-fold in the presence of 15 mg-Cd/l or 15 mg-Cu/l, respectively. The results from hydroponic cultures showed that inoculation of Serratia sp. SY5 had a favorable influence on the initial shoot growth and biomass of Zea mays under noncontaminated conditions. However, under Cd-contaminated conditions, the inoculation of SY5 significantly increased the root biomass of Zea mays. These results indicate that Serratia sp. SY5 can serve as a promising microbial inoculant for increased plant growth in heavy-metal-contaminated soils to improve the phytoremediation efficiency.

In this study, we investigated the inhibition effects of single and mixed heavy metal ions (Zn2+, Ni2+, Cu2+, and Cd2+) on iron oxidation by Acidithiobacillus ferrooxidans. Effects of metals on the iron oxidation activity of A. ferrooxidans are categorized into four types of patterns according to its oxidation behavior. The results indicated that the inhibition effects of the metals on the iron oxidation activity were noncompetitive inhibitions. We proposed a reduced inhibition model, along with the reduced inhibition constant (alpha i), which was derived from the inhibition constant (KI) of individual metals and represented the tolerance of a given inhibitor relative to that of a reference inhibitor. This model was used to evaluate the toxicity effect (inhibition effect) of metals on the iron oxidation activity of A. ferrooxidans. The model revealed that the iron oxidation behavior of the metals, regardless of metal systems (single, binary, ternary, or quaternary), is closely matched to that of any reference inhibitor at the same reduced inhibition concentration, [I]reduced, which defines the ratio of the inhibitor concentration to the reduced inhibition constant. The model demonstrated that single metal systems and mixed metal systems with the same reduced inhibitor concentrations have similar toxic effects on microbial activity.

In the current studies, we characterized the degradation of a hot mixture of benzene and toluene (BT) gases by a thermophilic biofilter using polyurethane as packing material and high-temperature compost as a microbial source. We also examined the effect of supplementing the biofilter with yeast extract (YE). We found that YE substantially enhanced microbial activity in the thermophilic biofilter. The degrading activity of the biofilter supplied with YE was stable during long-term operation (approximately 100 d) without accumulating excess biomass. The maximum elimination capacity (1,650 g x m(-3) h(-1)) in the biofilter supplemented with YE was 3.5 times higher than that in the biofilter without YE (470 g g x m(-3) h(-1)). At similar retention times, the capacity to eliminate BT for the YE-supplemented biofilter was higher than for previously reported mesophilic biofilters. Thus, thermophilic biofiltration can be used to degrade hydrophobic compounds such as a BT mixture. Finally, 16S rDNA polymerase chain reaction-DGGE (PCR-DGGE) fingerprinting revealed that the thermophilic bacteria in the biofilter included Rubrobacter sp. and Mycobacterium sp.