Search

Stable degrading 1,2-dichlorobenzene (1,2-DCB) enrichments were generated from original contaminated soil and groundwater via enrichment procedures using a mineral salt medium containing 1,2-DCB as the sole carbon and energy source. Four transferred enrichments showed stable 1,2-DCB-degrading ability and completely degraded 1,2-DCB within 32 h. PCR-denaturing gradient gel electrophoresis (DGGE) and 16S rRNA gene clone library analyses indicated that two bacterial strains, belonging to Acidovorax spp. and Ralstonia spp., respectively, were the predominant organisms in each enrichment. Moreover, these strains maintained a stable coexistence in the four transferred enrichments. These two bacteria were subsequently identified as Acidovorax sp. strain sk40 and Ralstonia sp. strain sk41. Strain sk40 was more tolerant to higher concentrations of 1,2-DCB than strain sk41, while strain sk41 maintained a shorter degradation time under lower concentrations of 1,2-DCB. Notably, however, both strains exhibited similar growth rates and degradation rates in media containing 40 mg/l 1,2-DCB, as well as complete degradation of the 1,2-DCB (40 mg/l) within 32 h. It is expected that these two strains will be used in future applications of bioremediation of 1,2-DCB contamination.

Mangroves constitute valuable coastal resources that are vulnerable to oil pollution. One of the major processes to remove oil from contaminated mangrove sediment is microbial degradation. A study on heavy oil- and hydrocarbon-degrading bacterial consortia from mangrove sediments in Okinawa, Japan was performed to evaluate their capacity to biodegrade and their microbial community composition. Surface sediment samples were obtained from mangrove sites in Okinawa (Teima, Oura, and Okukubi) and enriched with heavy oil as the sole carbon and energy source. The results revealed that all enriched microbial consortia degraded more than 20% of heavy oil in 21 days. The K1 consortium from Okukubi site showed the most extensive degradative capacity after 7 and 21 days. All consortia degraded more than 50% of hexadecane but had little ability to degrade polycyclic aromatic hydrocarbons (PAHs). The consortia were dominated by Pseudomonas or Burkholderia. When incubated in the presence of hydrocarbon compounds, the active bacterial community shifted to favor the dominance of Pseudomonas. The K1 consortium was a superior degrader, demonstrating the highest ability to degrade aliphatic and aromatic hydrocarbon compounds; it was even able to degrade heavy oil at a concentration of 15%(w/v). The dominance and turn-over of Pseudomonas and Burkholderia in the consortia suggest an important ecological role for and relationship between these two genera in the mangrove sediments of Okinawa.

The aims of this study were to identify the bacteria responsible for the rapid degradation of aromatic hydrocarbons, and to determine the roles of bacterial species in the microbial consortium. The population changes of the four major bacterial groups, namely Burkholderia spp., Achromobacter spp./Alcaligenes spp., Cupriavidus spp., and Rhodanobacter spp. in the Y consortium that preferentially degraded petroleum oil-derived aromatic hydrocarbons were assessed using quantitative real-time polymerase chain reaction (q-PCR). P-xylene was used as a representative aromatic compound, decane as a representative aliphatic compound, p-xylene-decane as a representative hydrocarbon mixture, and kerosene as a representative petroleum-based oil. The results showed that the remarkable increase of Burkholderia spp. coincided with the rapid degradation of aromatic hydrocarbons in kerosene, and p-xylene in single and dual-substrate cultures. Cupriavidus spp. significantly increased when decane was rapidly depleted. Achromobacter spp.,/Alcaligenes spp., and Rhodanobacter spp. were initially inhibited by the presence of aromatic hydrocarbons and seemed not to be active in hydrocarbon degradation, but utilized the metabolic products. This work provided an alternative way to explain the contribution of different microbial species in the degradation of hydrocarbon fractions. The behavior of the consortium is of interest in the context of the bioremediation of the more toxic aromatic hydrocarbons in environments contaminated by petroleum products.

A dechlorinating consortium (designated as TES-1 culture) able to convert trichloroethene (TCE) to ethene was established from TCE-contaminated groundwater. This culture had the ability of complete dechlorination of TCE within about one month. From the clone library analysis of 16S rRNA gene, this culture was mainly composed of fermentation bacteria, such as Clostridium spp., and Desulfitobacterium spp. known as facultative dechlorinator. PCR using specific primers for Dehalococcoides spp. and the dehalogenase genes confirmed that the culture contained the Dehalococcoides spp. 16S rRNA gene and three dehalogenase genes, tceA, vcrA and bvcA. Dechlorination experiments using cis-dichloroethene (cis-DCE) at concentrations of 37-146 μM, revealed that the gene copy numbers of tceA, vcrA, and bvcA increased up to 10⁷ copy/mL, indicating that Dehalococcoides spp. containing these three dehalogenase genes were involved in cis-DCE dechlorination. However, in the culture to which 292 μM of cis-DCE was added, only the tceA gene and the Dehalococcoides spp. 16S rRNA gene increased up to 10⁷ copy/mL. The culture containing 292 μM of cis-DCE also exhibited about one tenth slower ethene production rate compared to the other cultures.

Numerous studies on the biodegradation of petroleum products using total petroleum hydrocarbons (TPH) have been carried out; however, the biodegradation of equivalent carbon number (EC) based hydrocarbon fractions in mineral oil has attracted little attention. This study investigated the ability of a microbial consortium to degrade the EC fractions in kerosene, which was used as a representative mineral oil. Based on the cloning and sequencing of the 16S rRNA gene, the microbial community was predominantly identified as Betaproteobacteria of the genera Achromobacter , Alcaligenes , and Cupriavidus . Degradation experiments in sealed 120-ml vials containing 1% (w v −1 ) kerosene revealed that aromatic fractions were degraded faster than aliphatic fractions. Aromatic fractions EC >7–8 and EC >8–10 were completely degraded after three days while aliphatic fractions EC >6–8 and EC >8–10 were only partially degraded. The aromatic EC >10–12 fraction was the third most degraded, and the aliphatic EC >10–12 and EC >12–16 fractions were the least degraded fractions. The first-order rate constants for the aromatic fractions ranged from 0.12 d −1 to 0.51 d −1 and from 0.06 d −1 to 0.32 d −1 for the aliphatic fractions. The microbial consortium preferentially utilized aromatic fractions, which are more toxic than aliphatic fractions. This finding is useful when considering risk-based bioremediation: A microbial community could potentially degrade the more toxic aromatic hydrocarbon components in petroleum-contaminated environments.

This paper describes hydrogen sulfide generation, isolation and characterization of sulfate-reducing bacteria in the activated sludge obtained from wastewater treatment plant. The sludge, collected from aerobic treatment system, was used as inoculums for hydrogen sulfide generation and for isolation of sulfate-reducing bacteria. Hydrogen sulfide was generated when the sludge was inoculated into a pH controlled medium. Two strains G2 and G3 of sulfate-reducing bacteria were isolated from the sludge. The characteristics and identification of these isolates was based on culture of the experiments and molecular analysis. Reduced sulfate rates in G2 and G3 is 0.39 and 0.41 mmol/dm3 respectively. Two isolates located in the genus Desulfovibrio in the phylogenetic tree based. First one was 16S rRNA gene analysis with the highest homology, 96.6 %, as Desulfovibrio carbinoliphilus (DSM 17524T) . However, the branch of the isolates was separated from the relatives and no strain was recognized in this branch. Also the dsr gene of isolate G3 had the highest homology with Desulfovibrio fructosovorans (DSM 3604T) , 92.7 %. The isolates G2 and G3 belonged to the genus Desulfovibrio as confirmed by the results of these analyses, however, these isolates oxidized organic compounds to carbon dioxide completely although relative strain oxidized it completely. Therefore, these isolates have high possibility that they should be recognized as a new strain.

This paper described that relationship between Acidithiobacillus feroooxidans T23-3 and fungus TEF1-1 in an enrichment culture. In order to evaluate the relationship, some culture experiments using cell-free extracts from T23-3 and TEF1-1 were carried out. Chemolitotrophic iron-oxidizing bacterium, A. ferrooxidans T23-3, was able to grow using inorganic compounds in cell-free extract of fungus TEF1-1 or A. ferrooxidans T23-3. Since the fungus TEF1-1 was able to grow using cell-free extract of the both microorganisms at pH 1.5, the fungus used some organic compounds in the cell-free extract as energy and carbon sources. When the cell-free extract of TEF1-1 used for the cultivation of A. ferrooxidans T23-3, lag-phase was observed because of lack of magnesium ion. Minimum concentration of magnesium ion for complete growth of A. ferrooxidans T23-3 was found to be 2.0 x 10-5 mol/dm3 from the experiments using synthetic medium. Also the cell-free extract contained enough nutritious components except magnesium ion for the growth of A. ferrooxidans T23-3. It was suggested that fungi grew using organics and inorganic ions released from the cell of chemolithotrophic iron-oxidizing bacteria by bacteriolysis and/or metabolisum and that the bacteria grew using inorganic ions from the lysis of fungi cells in the enrichment culture of iron-oxidizing bacteria. In addition, chemolithotrophic iron-oxidizing bacterium fixes carbon dioxide to convert some organic compounds in the culture. Therefore, it was evaluated that the relationship between chemolithotrophic iron-oxidizing bacteria and fungi was mutualistic symbiosis in the enrichment culture.

The copper extraction yield from thermophilic bioleaching of chalcopyrite depends on temperature, pH, and the oxidation-reduction potential (ORP), as well as on the activity of the thermophile used. The copper extraction yields obtained with three thermophiles under various pH and temperature conditions and with different initial amounts of Fe 3+ were studied. The results indicated that because of the low ability of Acidianus brierleyi to leach iron as Fe 3+ , high biomass concentrations were reflected by ORP close to a critical value (450 mV, Ag 0 /AgCl reference), at which copper extractions were highest. By contrast, because of the higher ability of Sulfolobus metallicus and Metallosphaera sedula to leach iron as Fe 3+ , high biomass concentrations were reflected by high ORP which in combination with the precipitation of Fe 3+ as jarosite attenuated the leaching rates. Therefore, optimum temperatures for the growth of thermophiles did not always mean high copper extraction yields. Generally, highest copper extractions were obtained at initial pH 1.5. However, higher copper extractions were observed at initial pH 2.5 than at pH 2.0, suggesting that at high pH the bioleaching of chalcopyrite is controlled by the ORP rather than by the pH or temperature. The bioleaching capacity of A. brierleyi was reduced or suppressed when insufficient initial Fe 3+ was provided to trigger the leaching reaction, whereas S. metallicus and M. sedula were less sensitive to the initial availability of Fe 3+ . This result confirmed that a direct enzymatic attack on the mineral surface could initiate the leaching reaction, but later ORP governed the leaching rate of chalcopyrite.

This paper describes about microbial diversity in an iron oxidation tank of an AMD treatment plant established at an abandoned sulphur mine in Japan. Since the mining operation was stopped, this mine has produced strong acidic mine drainage, pH 1.8, including a high concentration of ferrous iron, 301mg/L, and the flow rate was about 4m3/min. In 2006, a pilot scale microbial iron oxidizing system was installed to remove total iron more easily from the AMD by oxidizing ferrous iron to ferric iron. From the start of this pilot operation, microbial diversity in the iron oxidation tank was investigated using a PCR-DGGE method for about two months. In the PCR, V3 region of 16S rRNA gene for Bacteria was amplified. The profiles of DGGE showed that there were three dominant species in the iron oxidation tank through the experimental period. Number of bands on DGGE profiles decreased with dates of sampling so that the microbial population became less diverse because of the iron oxidizing operation. There was a wide variety of bacterial species of even though conditions were strongly acidic.

This paper describes that evaluation of soil remediation contaminated with oil on quicklime process and the effect of the treatment for oil degradable bacteria in the soil. In this paper, two different contaminated sites were used. When quicklime method was applied for a contaminated site, the temperature of soil increased over 60 °C by reaction heat and the pH value also rose to about 12. Although the density of bacteria in the contaminated soil decreased from 40 to 5 - 10 x 1010 cells/g-dry soil, they could survive against the increasing of heat and alkalinity caused by the treatment. Some oil degradable bacteria were isolated from original contaminated and treated soil and the abilities of oil degradation were evaluated. These bacteria might contribute the degradation of remained oil in the treated soil. On another site, the oil concentration in soil decreased from 3,555 to 1,430 mg-oil/kg-soil by the treatment. In this case, some light parts of oil, C6 to C10 and C10 to C28, were almost volatized to below 5 and 35% respectively, but the heavy part, C28 to C44 was still remained. However the remained oil decreased gradually after the treatment to 250 mg-oil/kg-soil within 6 month. This shows the high possibility of oil remediation by oil degradable bacteria which survive in contaminated soil against the treatment.

Preoperative portal embolization (PE) is useful for the prevention of postoperative liver failure after extended hepatectomy. However, clinical evaluation of liver function in the hypertrophying lobe after PE has not been studied. Here we report functional changes in the hypertrophying lobe using a 80% portal-branch-ligation rabbit model. Liver function was evaluated by the expression of liver-specific genes detected by Northern blot analysis and plasma disappearance rate of indocyanine green (ICG). The weight of the unligated lobe after portal ligation increased about twofold on the 7th postoperative day (POD) and about threefold on the 14th POD. The mRNA levels of the liver-specific genes (albumin, aldolase B, and tyrosine aminotransferase) in the unligated lobe decreased to about 50% on the 1st POD and returned to the preoperative levels on the 7-14th POD. In contrast, the expression of histone H2B mRNA increased on the 3rd-7th POD. The plasma disappearance rate of ICG (K-ICG) in the rabbit that has only the unligated lobe did not significantly change during the first 7 days, but then improved and recovered to 80% of that in the rabbit that has whole liver on the 14th POD. These results indicate that liver function of the hypertrophying lobe after portal branch ligation does not increase during the first 7 days despite an increase in liver weight. This finding suggests that the compensatory hypertrophying liver is enlarging without functional augmentation in the early period after PE.

In the enrichment culture of the sulfur-oxidizing bacteria collected from an acid mine drainage, we have confirmed protozoa that belongs to Bodo edax. This paper describes an interaction between the bacteria and the protozoa in the presence of cupric ion. The SO42- produced by the bacteria from S0 and the cell numbers of the bacteria and the protozoa were measured as a function of time in the culture containing Cu2+ of 0, 1, 10, 100 and 1, 000mg dm-3. The results obtained were as follow;(1) the bacterial oxidation rate and the yield factor of the bacteria growth became small due to the protozoa;(2) in the cultivation of the bacteria alone, the lag phase of the bacteria growth became long over 1 mg dm-3 of Cu2+ concentration but the growth rate was not affected;(3) in the culture coexisting the bacteria and the protozoa the bacteria grew, but the protozoa did not grow at Cu2+ concentration of 100mg dm-3;(4) preincupation in the culture containing Cu2+ gave the ability to resist Cu2+ to the bacteria, but did not to the protozoa.

Sulfur is almost insoluble in water at ambient temperatures, and therefore polysulfide (S(n)(2-)) has been considered as a possible intermediate that is used directly by bacteria in sulfur respiration. Sulfur-reducing reductases have been purified and characterized from a few sulfur reducers. However, polysulfide reduction has only been confirmed in Wolinella succinogenes. In our previous study, the direct production of hydrogen sulfide from polysulfide was confirmed by an enrichment culture obtained from natural samples under sulfate-reducing conditions. The present study attempted to isolate and identify polysulfide-reducing bacteria from the enrichment cultures. Almost all the isolated strains were classified into the genus Clostridium, based on 16S rRNA gene sequence analysis. The isolates, and some closely related strains, were able to reduce polysulfide to hydrogen sulfide. During production of 1 mol of hydrogen sulfide, approximately 2 mol of lactate was converted to acetate. Thus, dissimilatory polysulfide reduction occurred using lactate as an electron donor. The ability to reduce elemental sulfur was also examined with the isolates and the related strains. Although elemental sulfur reducing strains can reduce polysulfides, not all polysulfide-reducing strains can reduce elemental sulfur. These results demonstrate that the conversion of elemental sulfur to polysulfide seems to be important in the reduction process of sulfur.

An anaerobic mixed microbial culture was enriched from soil and groundwater taken from a site contaminated with trichloroethene (TCE). This enrichment culture could dechlorinate TCE sequentially to cis‐dichloroethene (cis‐DCE), vinyl chloride (VC), and ethene rapidly within 2 weeks. This enrichment culture could utilize various organic compounds, such as methanol, ethanol, and sodium acetate and so on, as electron donor. This culture had maintained high ability of TCE dechlorination for about 3 years since the start of enrichment cultivation. The optimum pH value for the dechlorination activity of this culture, which reacts from TCE to ethene, was 6.7. However above the pH value 7.1, it lost the dechlorination ability of cis‐DCE and VC. So cis‐DCE was remained at that pH conditions. From the DNA sequencing analysis of 16SrRNA gene, this enrichment culture includes Dehalococcoides sp. which has the ability to dechlorinate TCE to VC completely with hydrogen. It suggested that this Dehalococcoides sp. takes part in the dechlorination of chloroethenes.

An acidophilic heterotrophic bacterium, designated as HIB4, having the ability to oxidize ferrous ion was newly isolated from a sample of an enrichment culture for iron-oxidizing bacteria, using the modified washed agarose/yeast extract (WAYE) medium with ferrous sulphate. The isolate HIB4 was an acidophilic, heterotrophic, mesophilic and gram-positive bacterium. Phylogenetically, it was classified under the genus Alicyclobacillus and was the closest to Alicyclobacillus disulfidooxidans SD-11 with 99.7% 16S rDNA homology. It grew and oxidized ferrous ion in the medium containing 0.02% (w/v) yeast extract. Yeast extract was an essential substrate for this bacterium because it could not grow or oxidize ferrous ion without yeast extract. However, a higher concentration of yeast extract inhibited the growth of HIB4, so that the optimum concentration of yeast extract for this bacterium to grow was 0.02% (w/v) at 0.08 mol/l of ferrous ion. On the other hand, ferrous ion oxidation occurred almost at the end of the bacterium's logarithmic growth phase and the isolate was able to grow without ferrous ion. These results denote that HIB4 did not obtain any energy from the ferrous ion oxidation and that HIB4 is an obligate heterotrophic and aerobic bacterium even though it oxidized ferrous ion. Also, HIB4 could not utilize any organic compounds, among the several organic chemicals used in this study, as a carbon source except yeast extract. These characteristics were completely different from these of A. disulfidooxidans SD-11 so that HIB4 might be a different species.

To find an efficient bacterium, which has the strong capacity to produce hydrogen sulfide from polysulfide as the waste of process generating hydrogen from hydrogen sulfide by photocatalytic reaction using sun light, is very important for constructing hydrogen producing system. 10 strains of sulfate‐reducing bacteria (SRB), which can reduce polysulfide directly, have been isolated from various natural samples such as TCE contaminated soil, soil and sludge around hot spring environment, and the cooling tower of a geothermal plant. This study describes physiological and phylogenetic characterization of SRB which can reduce polysulfide. All of isolates had the ability to reduce polusulfide but these reduction rates were difference depend on isolates. Phylogetetically, all of isolates located difference position for general SRB including Desulfovibrio desulfuricans, which is used standard strain in this study, so they do not belong to Proteobacteria. These have close relation to the genus Desulfotomaculum which can reduce elemental sulfur. It suggests that the ability of reducing elemental sulfur is important for reducing polysulfide to hydrogen sulfide.

A heterotrophic bacterium, designated as HIB4, having an ability to oxidize ferrous iron was isolated from the sample of an enrichment culture with 9K medium, by using the modified WAYE (washed agarose/yeast extract) medium with ferrous sulphate. This isolate was identified as Alicyclobacillus disulfidooxidans from 16S rDNA sequence analysis. The isolate grew and oxidized ferrous iron in an inorganic medium containing 0.02 % (w/v) of yeast extract and Ferrous iron oxidation occurred at the almost end of its logarithmic phase. Yeast extract was an essential substrate for the isolate because the isolate could not grow or oxidize ferrous iron without yeast extract. However, higher concentration of yeast extract inhibited the growth of the isolate. On the other hand, it was confirmed that the isolate was able to grow without ferrous ion so that it did not get any energy by ferrous ion oxidation. Its optimum concentration of yeast extract was 0.02% (w/v) at the concentration of ferrous ion 0.08mol/liter. Its o...