Cultivation-dependent and cultivation-independent characterization of the microbial community in acid mine drainage associated with acidic Pb/Zn mine tailings at Lechang, Guangdong, China.

Cultivation-based and molecular approaches were used to characterize the phylogenetic composition and structure of the microbial community in an extremely acidic (pH 2.0) acid mine drainage (AMD) associated with Pb/Zn mine tailings that were undergoing vigorous acid generation. Acidophilic bacteria were isolated and enumerated on solid media, and were found to be restricted to isolates related to Acidithiobacillus ferrooxidans and Acidiphilium cryptum. By contrast, cloning and phylogenetic analysis of 16S rRNA genes revealed that, although low in total taxonomically distinct groups, the tailings AMD ecosystem harbored a wide range of phylogenetically diverse microbes. Of the 141 clones examined, 104 were phylogenetically affiliated with the recently discovered, iron-oxidizing Leptospirillum group III within the Nitrospira. It thus appears that iron serves as the major electron donor in this habitat. Thirty clones were affiliated with the Proteobacteria, half of which belonged to organisms related to Alphaproteobacteria species capable of ferric iron reduction. Other clones were grouped with Betaproteobacteria and Gammaproteobacteria (six clones each), and even with Deltaproteobacteria (three clones), a subdivision with anaerobic sulfate or metal (iron) reduction as the predominant physiological trait of its members. Finally, four clones were clustered within the Firmicutes and the Acidobacteria. Approximately half of the sequence types representing the majority of the total clones fell into lineages that are poorly represented by cultured organisms or have thus far been represented by only a few environmental sequences. Thus, the present study extends our knowledge of the biodiversity of microorganisms populating highly acidic AMD environments.

Distribution patterns of octylphenol and nonylphenol in the aquatic system at mai po marshes nature reserve, a subtropical estuarine wetland in Hong Kong.

Spatial and temporal distribution of octylphenol (OP) and nonylphenol (NP) in Mai Po Marshes, a subtropical estuarine wetland in Hong Kong, were investigated. Surface water samples were collected every month from 11 sites during the period of September-December 2004. Concentrations of OP and NP ranged from 11.3 to 348 ng/L and from 29 to 2591 ng/L, respectively. The high levels of NP and OP were found in November and December than in September and October. The levels of OP and NP have no significant spatial differences except September. Total organic matter in the sediments appeared to be an important factor in controlling the fate of these compounds in the aquatic environment.

Field released transgenic papaya effect on soil microbial communities and enzyme activities.

Soil properties, microbial communities and enzyme activities were studied in soil amended with replicase (RP)-transgenic or non-transgenic papaya under field conditions. Compared with non-transgenic papaya, significant differences (P<0.05) were observed in total nitrogen in soils grown with transgenic papaya. There were also significant differences (P<0.05) in the total number of colony forming units (CFUs) of bacteria, actinomycetes and fungi between soils amended with RP-transgenic plants and non-transgenic plants. Compared with non-transgenic papaya, the total CFUs of bacteria, actinomycetes and fungi in soil with transgenic papaya increased by 0.43-1.1, 0.21-0.80 and 0.46-0.73 times respectively. Significantly higher (P<0.05) CFUs of bacteria, actinomycetes and fungi resistant to kanamycin (Km) were obtained in soils with RP-transgenic papaya than those with non-transgenic papaya in all concentrations of Km. Higher resistance quotients for Kmr (kanamycin resistant) bacteria, actinomycetes and fungi were found in soil planted with RP-transgenic papaya, and the resistance quotients for Kmr bacteria, actinomycetes and fungi in soils with transgenic papaya increased 1.6-4.46, 0.63-2.5 and 0.75-2.30 times. RP-transgenic papaya and non-transgenic papaya produced significantly different enzyme activities in arylsulfatase (5.4-5.9x), polyphenol oxidase (0.7-1.4x), invertase (0.5-0.79x), cellulase (0.23-0.35x) and phosphodiesterase (0.16-0.2x). The former three soil enzymes appeared to be more sensitive to the transgenic papaya than the others, and could be useful parameters in assessing the effects of transgenic papaya. Transgenic papaya could alter soil chemical properties, enzyme activities and microbial communities.

Dependence of bisphenol A photodegradation on the initial concentration of oxalate in the lepidocrocite-oxalate complex system.

To understand the degradation of endocrine disrupting chemicals (EDCs) in natural environment with existence of iron oxides and carboxylic acids, the dependence of bisphenol A (BPA) photodegradation on the initial concentration of oxalate (COX) in lepidocrocite (gamma-FeOOH) aqueous suspension was investigated under both UV and visible lights in this study. Lepidocrocite powder was home-prepared by a hydrothermal process. It was found that BPA degradation was promoted greatly in the presence of oxalate owing to the formation of lepidocrocite-oxalate complex. And there was an optimal COX, which was 2.0 and 2.4 mmol/L, under UV and visible lights, respectively. The first-order kinetic constant, k value increased 38 times from 0.17 x 10(-2) min(-1) in the absence of oxalate to 6.39 x 10(-2) min(-1) in the presence of oxalate with an optimal COX (2.0 mmol/L) under UV irradiation, and almost 306 times from 0.02 x 10(-2) min(-1) in the absence of oxalate to 6.11 x 10(-2) min(-1) in the presence of oxalate with an optimal COX (2.4 mmol/L) under visible irradiation. The BPA degradation rate increased and the first-order kinetic constants decreased with the increase in BPA initial concentration. The dependence of the variation of pH value, total-Fe and Fe2+ during the photoreaction on COX was also investigated. The pH value increased obviously with the reaction time. Total-Fe increased dramatically at the first 5 min and then decreased quickly under UV irradiation and slowly under visible irradiation. The initial concentration of oxalate is a main factor to affect BPA photodegradation in aqueous suspension under both UV and visible lights.

Diversity and structure of the archaeal community in the leachate of a full-scale recirculating landfill as examined by direct 16S rRNA gene sequence retrieval.

The diversity and structure of the archaeal community in the effluent leachate from a full-scale recirculating landfill was characterized by direct 16S rRNA gene (16S rDNA) retrieval. Total-community DNA was extracted from the microbial assemblages in the landfill leachate, and archaeal 16S rDNAs were amplified with a universally conserved primer and an Archaea-specific primer. The amplification product was then used to construct a 16S rDNA clone library, and 70 randomly selected archaeal clones in the library were grouped by restriction fragment length polymorphism (RFLP) analysis. Sequencing and phylogenetic analysis of representatives from each unique RFLP type showed that the archaeal library was dominated by methanogen-like rDNAs. Represented in the kingdom of Euryarchaeota were phylotypes highly similar to the methanogenic genera Methanoculleus, Methanosarcina, Methanocorpusculum, Methanospirillum and Methanogenium, where the clone distribution was 48, 11, 3, 1 and 1, respectively. No sequences related to known Methanosaeta spp. were retrieved. Four rDNA clones were not affiliated with the known methanogenic Archaea, but instead, they were clustered with the uncultured archaeal sequences recently recovered from anaerobic habitats. Two chimeric sequences were identified among the clones analyzed.

Evaluation of leachate recirculation on nitrous oxide production in the Likang Landfill, China.

Landfill leachate recirculation is efficient in reducing the leachate quantity handled by a leachate treatment plant. However, after land application of leachate, nitrification and denitrification of the ammoniacal N becomes possible and the greenhouse gas nitrous oxide (N2O) is produced. Lack of information on the effects of leachate recirculation on N2O production led to a field study being conducted in the Likang Landfill (Guangzhou, China) where leachate recirculation had been practiced for 8 yr. Monthly productions and fluxes of N2O from leachate and soil were studied from June to November 2000. Environmental and chemical factors regulating N2O production were also accessed. An impermeable top liner was not used at this site; municipal solid waste was simply covered by inert soil and compacted by bulldozers. A high N2O emission rate (113 mg m-2 h-1) was detected from a leachate pond purposely formed on topsoil within the landfill boundary after leachate irrigation. A high N2O level (1.09 micrograms L-1) was detected in a gas sample emitted from topsoil 1 m from the leachate pond. Nitrous oxide production from denitrification in leachate-contaminated soil was at least 20 times higher than that from nitrification based on laboratory incubation studies. The N2O levels emitted from leachate ponds were compared with figures reported for different ecosystems and showed that the results of the present study were 68.7 to 88.6 times higher. Leachate recirculation can be a cost-effective operation in reducing the volume of leachate to be treated in landfill. However, to reduce N2O flux, leachate should be applied to underground soil rather than being irrigated and allowed to flow on topsoil.

[Analyses of virus resistance and transgenes for transgenic papaya].

Virus resistance in field and molecular biological characterizations of the transgenes were analyzed for two lines of T(1) generation of transgenic papaya with the replicase mutant gene from papaya ringspot virus (PRV). The transgenic plants showed highly resistant or immune against PRV. Results indicated that the transgenes inherited to and expressed at RNA level in the progenies.

Accumulation of heavy metals in four grasses grown on lead and zinc mine tailings.

A field experiment was conducted to compare the growth and metal accumulation of Vetiveria zizanioides, Paspalum notatum, Cynodon dactylon and Imperata cylindraca var. major on the tailings, amended with 10 cm domestic refuse + complex NPK fertilizer(Treatment A), 10 cm domestic refuse(Treatment B) and complex NPK fertilizer (Treatment C) respectively, and without any amendment used as control (Treatment D). The results indicated that V. zizanioides was a typical heavy metal excluder, because the concentrations in shoots of the plants were the lowest among the four plants tested. The most of metal accumulated in V. zizanioides distributed in its root, and transportation of metal in this plant from root to shoot was restricted. Therefore, V. zizanioides was more suitable for phytostabilization of toxic mined lands than P. notatum and C. dactylon, which accumulated a relatively high level of metals in their shoots and roots. It was also found that I. cylindraca var. major accumulated lower amounts of Pb, Zn and Cu than C. dactylon and P. notatum, and could also be considered for phytostalilisaton of tailings. Although the metal(Pb, Zn and Cu) concentrations in shoots and roots of V. zizanioides were the lowest, the total amounts of heavy metals accumulated in shoots of V. zizanioides were the highest among the four tested plants due to the highest dry weight yield of it. The results indicated that V. zizanioides was the best choice among the four species used for phytoremediation (for both phytostabilization and phytoextraction) of metal contaminated soils.

Seasonal and spatial variations in microbial community structure and diversity in the acid stream draining across an ongoing surface mining site.

This study examined the microbial community in an acidic stream draining across the Yun-Fu pyrite mine (Guangdong, China), where extremely acidic mine water is a persistent feature due to the intensive surface mining activities. Analysis of terminal restriction fragment length polymorphism (TRFLP) of 16S rRNA gene sequences showed that microbial populations varied spatially and seasonally and correlated with geochemical and physical conditions. After the stream moves from underground to the surface, the microbial community in the acidic water rapidly evolves into a distinct community close to that in the downstream storage pond. Comparisons of TRFLP peaks with sequenced clone libraries indicated that bacteria related to the recently isolated iron-oxidizer Ferrovum myxofaciens dominated the acidophilic community throughout the year except for the samples collected in spring from the storage pond, where Ferroplasma acidiphilum-like archaea represented the most abundant group. Acidithiobacillus ferrooxidans-affiliated organisms increased along the acid stream and remained common over the year, whereas Leptospirillum ferrooxidans-like bacteria were negligible or even not detected in the analyzed samples. The data indicate that changes in environmental conditions are accompanied by significant shifts in community structure of the prokaryotic assemblages at this opencast mining site.

Copper and Zinc in a paddy field and their potential ecological impacts affected by wastewater from a lead/zinc mine, P. R. China.

As well known, at normal levels, copper and zinc are essential micronutrients for plants, animals, and humans. However, excessive Cu and Zn are toxic and disturb a wide range of biochemical and physiological processes. Using Atomic Absorption Spectrophotometer (AAS; Perkin-Elmer 3030, USA), soil and rice plant (Oryza sativa L.) samples collected from a paddy field in Lechang lead-zinc mine area, Guangdong Province, China were analyzed and their potential ecological impacts to local human and livestock were evaluated. The results showed that the paddy soils were contaminated with Cu and Zn. Both metals in soils had low bio-available fractions for paddy plants, animal and human by three chemically analytical techniques. Generally, were concentrations of copper and zinc root >> straw > stalk > grain with hull > grain without hull (i.e. unpolished rice) and in the normal ranges indicating no ecological risk for local livestock and residents. All positive correlation coefficients, however, between heavy metals in rice plant and total, exchangeable (step 1 in Tessier's method established in 1979) and DTPA-extractable fractions in soils were found in this study indicating that elevated heavy metal in soils would increase long-term exposition and possible consequence of ecological hazard through food chains.

Culturable and molecular phylogenetic diversity of microorganisms in an open-dumped, extremely acidic Pb/Zn mine tailings.

A combination of cultivation-based and molecular-based approaches was used to reveal the culturable and molecular diversity of the microbes inhabiting an open-dumped Pb/Zn mine tailings that was undergoing intensive acid generation (pH 1.9). Culturable bacteria found in the extremely acidic mine tailings were Acidithiobacillus ferrooxidans, Leptospirillum ferriphilum, Sulfobacillus thermotolerans and Acidiphilium cryptum, where the number of acidophilic heterotrophs was ten times higher than that of the iron- and sulfur-oxidizing bacteria. Cloning and phylogenetic analysis revealed that, in contrast to the adjacent AMD, the mine tailings possessed a low microbial diversity with archaeal sequence types dominating the 16S rRNA gene library. Of the 141 clones examined, 132 were represented by two sequence types phylogenetically affiliated with the iron-oxidizing archaea Ferroplasma acidiphilum and three belonged to two tentative groups within the Thermoplasma lineage so far represented by only a few environmental sequences. Six clones in the library were represented by the only bacterial sequence type and were closely related to the well-described iron-oxidizer L. ferriphilum. The significant differences in the prokaryotic community structures of the extremely acidic mine tailings and the AMD associated with it highlights the importance of studying the microbial communities that are more directly involved in the iron and sulfur cycles of mine tailings.

Characterization of methanogenic Archaea in the leachate of a closed municipal solid waste landfill.

Cultivation-independent molecular approaches were used to investigate the phylogenetic composition of Archaea and the relative abundance of phylogenetically defined groups of methanogens in the leachate of a closed municipal solid waste landfill. Cloning and phylogenetic analysis of archaeal 16S rRNA gene sequences (16S rDNA) revealed that the landfill leachate harbored a diverse Archaea community, with sequence types distributed within the two archaeal kingdoms of the Euryarchaeota and the Crenarchaeota. Of the 80 clones examined, 51 were phylogenetically associated with well-defined methanogen lineages covering two major methanogenic phenotypes; 20 were related to Thermoplasma and were grouped with some novel archaeal rRNA gene sequences recently recovered from various anaerobic habitats; finally, five belonged to Crenarchaeota and were not closely related to any hitherto cultivated species. Most of the methanogen-like clones were affiliated with the hydrogenotrophic Methanomicrobiales and the methylotrophic and acetoclastic Methanosarcinales. Quantitative oligonucleotide hybridization experiments showed that methanogens in the leachate accounted for only a very small fraction of the total community (approximately 2%) and that Methanomicrobiales and Methanosarcinales constituted the majority of the total methanogenic population.