PCR-denaturing Gradient Gel Electrophoresis Research Articles

Diversity and screening of lactic acid bacteria in la-baicai made by Korean-Chinese in northeastern China

ABSTRACTThis study investigated the lactic acid bacteria (LAB) population in la-baicai (spicy cabbage), a traditional fermented food made by the Korean-Chinese community in northeastern China and screened for functional LAB. LAB diversity was analyzed using denaturing gradient gel electrophoresis (DGGE), and 81 LAB strains were isolated and identified based on 16S rDNA sequencing analysis. Polymerase chain reaction DGGE detected 21 LAB species, belonging to the genera Lactobacillus (Lb.), Leuconostoc (Leu.), Pediococcus and Weissella, in 45 la-baicai samples. Lb. plantarum and Lb. sakei were considered to be dominant in the bacterial community. Among 81 LAB isolated by traditional pure culture methods were Lb. plantarum (25 strains), Lb. brevis (two strains), Lb. casei, (four strains), Lb. pentosus (three strains), Enterococcus faecium (45 strains), and E. durans (two strains). The tolerances of these LAB were investigated. Six LAB with high salt (NaCl)-tolerance were screened from the isolates, and 16% (w/v) was the highest salt-tolerance reached. Among the isolates, strain N1, identified as Lb. pentosus, survived well in a simulated digestive environment. Traditional fermented la-baicai from northeastern China is a rich LAB resource. Further study is needed and would be worthwhile to advance the benefits of these LAB.

Microbial indication of soil health in apple orchards affected by replant disease

The aim of this study was to describe the interactions between soil physico-chemical parameters, plant growth, and soil microbial diversity under different fertilisation regimes in apple orchards affected by replant disease. The hypothesis was that fertiliser type and application, as well as tillage technique, could exert a shift in the soil chemical and biological parameters linked to soil sickness. Two apple orchards affected by ARD and located at different sites in South Tyrol, Italy, were selected as representatives of perennial crops affected by replant disease. The effects of application of six different amendments to the soil, at both sites, were investigated. Physico-chemical and ecophysiological properties, along with soil microbial communities (evaluated with PCR-Denaturing gradient gel electrophoresis and high throughput sequencing of the V1-V3 region of the bacterial 16S rRNA gene) were analysed. High-throughput sequencing resulted in 1,590,042 sequence reads, classified in 326,371 operational taxonomic units (OTUs). Considering a threshold of 1% relative abundance of OTUs for each sample, 39 bacterial phyla and 197 genera were detected. The phyla Proteobacteria, Actinobacteria, Acidobacteria and Bacteroidetes comprised between 24% and 71% of all sequences. Plant growth parameters were correlated with physico-chemical and ecophysiological parameters as well as with bacterial genus abundances. Findings revealed that in this study, Rhodanobacter, Blastocatella, Arenimonas, Variovorax, Ferruginobacter, Demequina and Schlesneria were the genera most affected by treatment type in terms of abundance. Furthermore, Ferruginobacter, Demequina and Schlesneria, genera not commonly described as involved in biocontrol, were strongly correlated with plant growth by a positive association. Treatments were found to exert contradictory effects on the bacterial communities and on the ecophysiological parameters at the different sites. These findings indicate not only a high complexity in the interaction between environmental and biological entities, but also that changes in environmental parameters result in changes in the interactions within bacterial communities. High-throughput sequencing confirmed the hypothesis that soil bacterial communities at the two sites were diverse and differed significantly, while they did not differ significantly according to treatment type. The dominant genera detected were not directly related to the treatment or to the growth of apple plants, suggesting that minor populations and unclassified sequences could be involved in replant disease. High-throughput sequencing can be proposed as a suitable method to deeply describe the soil microbiota found in ARD soils.

Rice straw decomposition affects diversity and dynamics of soil fungal community, but not bacteria

Straw decomposition increased amounts of soil organic carbon and changed microbial biomass. But the different impacts of rice straw decomposition on the succession of bacterial and fungal community composition in field conditions were poorly understood. The objective of this study was to investigate the development of soil bacterial and fungal communities during rice straw decomposition in field conditions, and the relationship between soil chemical/physical properties and the evolution of microbial communities. A 1-year field study (90, 180, 270, and 360 days) was conducted, including the straw decomposition soil and control (no straw decomposition soil). The bulk soil samples (0–15-cm depths) from three replicate plots per treatment were collected for the analysis of soil properties and of microbial diversity parameters. Soil bacterial and fungal community structures and population sizes were determined by applying PCR-denaturing gradient gel electrophoresis (DGGE) and quantitative PCR (qPCR). The bacterial and fungal community diversity was evaluated using the following parameters: Shannon–Wiener diversity index, richness, and evenness. Moreover, the relationship between soil properties and the changes of microbial communities was analyzed using redundancy analysis (RDA). The results showed that, in contrast to bacteria, the soil fungal population size and diversity indices were significantly increased during different time points of rice straw decomposition, and reached to the highest value at 360 days. When compared with the bacterial DGGE profiles, the fungal DGGE profiles significantly changed between the straw decomposition soil and control, and the dominant soil fungal genera varied apparently. Bacterial 16S ribosomal RNA (rRNA) and fungal 18S rRNA gene sequences obtained from the main DGGE bands were further sequenced, among which Penicillium sp., Aspergillus sp., and Acremonium sp. have the ability to degrade cellulose. RDA suggested that the soil available P, organic C, pH, and bulk density were the main factors influencing the variation in the fungal community structures and diversities. The fungal community structures displayed distinct successions during rice straw decomposition. But such finding was not observed in bacterial communities. The ratio of fungi to bacteria measured by qPCR was higher in the straw decomposition soil compared with the findings in control, indicating that fungi predominated in straw decomposition. Especially, our study deduced that the diversified cellulolytic fungal genera highly enriched in rice straw decomposition soils had great potential of mining novel cellulose-decomposing enzymes.

English

Amorphophallus konjac cannot be cultivated on the same field consecutively for extended periods, but can be cultivated on the same grove. The underlying mechanism for this is unknown and may involve the rhizosphere microbial community. Therefore, the different duration and mode for konjac were researched. Polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE) showed that microbial communities varied by cultivation duration and mode. Seven bacterial phyla were detected in the soil and Proteobacteria was the most abundant of these phyla. The microbial metabolic diversity following three years of continuous cultivation (sample QXFH3) was the highest, and this sample may have contained pathogenic and antagonistic microorganisms. There was almost no disease in the soil sample from grove conditions (QXN0), which was readily distinguishable from soils from field conditions (QXF0). Amino acids, carboxylic acids, and miscellaneous carbohydrates were the main carbon sources utilized by microbes in these soils. The microbial diversity index and multivariate analyses revealed that bacterial diversity increased with cultivation duration.   Key words: Discontinuous cultivation, metabolic function, microbial composition, Amorphophallus konjac.

Ecotoxicological assessment of pesticides and their combination on rhizospheric microbial community structure and function of Vigna radiata.

India is one of the leading countries in production and indiscriminate consumption of pesticides. Owing to their xenobiotic nature, pesticides affect soil microorganisms that serve as mediators in plant growth promotion. Our study aimed to deliver a comprehensive picture, by comparing the effects of synthetic pesticides (chlorpyriphos, cypermethrin, and a combination of both) with a biopesticide (azadirachtin) at their recommended field application level (L), and three times the recommended dosage (H) on structure and function of microbial community in rhizosphere of Vigna radiata. Effect on culturable fraction was assessed by enumeration on selective media, while PCR-denaturing gradient gel electrophoresis (DGGE) was employed to capture total bacterial community diversity. This was followed by a metabolic sketch using community-level physiological profiling (CLPP), to obtain a broader picture of the non-target effects on rhizospheric microbial community. Although plant parameters were not significantly affected by pesticide application, the microbial community structure experienced an undesirable impact as compared to control devoid of pesticide treatment. Examination of DGGE banding patterns through cluster analysis revealed that microbial community structure of pesticide-treated soils had only 70% resemblance to control rhizospheric soil even at 45days post application. Drastic changes in the metabolic profiles of pesticide-treated soils were also detected in terms of substrate utilization, rhizospheric diversity, and evenness. It is noteworthy that the effects exacerbated by biopesticide were comparable to that of synthetic pesticides, thus emphasizing the significance of ecotoxicological assessments before tagging biopesticides as "safe alternatives."

Fish-mediated changes in bacterioplankton community composition: an in situ mesocosm experiment

We characterized variations in bacterioplankton community composition (BCC) in mesocosms subject to three different treatments. Two groups contained fish (group one: Cyprinus carpio; group two: Hypophthalmichthys molitrix); and group three, the untreated mesocosm, was the control. Samples were taken seven times over a 49-d period, and BCC was analyzed by PCR-denaturing gradient gel electrophoresis (DGGE) and real-time quantitative PCR (qPCR). Results revealed that introduction of C. carpio and H. molitrix had a remarkable impact on the composition of bacterioplankton communities, and the BCC was significantly different between each treatment. Sequencing of DGGE bands revealed that the bacterioplankton community in the different treatment groups was consistent at a taxonomic level, but differed in its abundance. H. molitrix promoted the richness of Alphaproteobacteria and Actinobacteria, while more bands affiliated to Cyanobacteria were detected in C. carpio mesocosms. The redundancy analysis (RDA) result demonstrated that the BCC was closely related to the bottom-up (total phosphorus, chlorophyll a, phytoplankton biomass) and top-down forces (biomass of copepods and cladocera) in C. carpio and control mesocosms, respectively. We found no evidence for top-down regulation of BCC by zooplankton in H. molitrix mesocosms, while grazing by protozoa (heterotrophic nanoflagellates, ciliates) became the major way to regulate BCC. Total bacterioplankton abundances were significantly higher in C. carpio mesocosms because of high nutrient concentration and suspended solids. Our study provided insights into the relationship between fish and bacterioplankton at species level, leading to a deep understanding of the function of the microbial loop and the aquatic ecosystem.

English

Bacterial populations within hypersaline lakes often exhibit uncultured unique species. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), followed by sequencing of 16S rRNA gene approach was applied in order to explore, for the first time, the bacterial communities within the hypersaline Lakes, Aghormy, Zeiton and Maraqi, Siwa Oasis, Egypt. The DGGE profile displayed 12 phylotypes of 16S rRNA gene, representing the total species richness within the three studied lakes. The phylotypes were varied among lakes, but were restricted to two phylogenetic groups, Bacteroidetes, which occurred in all lakes, and Alphaproteobacetria, which showed abundance in only Lakes Zeiton and Maraqi. Single spirochete-like phylotype characterized the Lake Aghormy. A phylotype, which was recorded in both of Aghormy and Zeiton, represented a first recorded of Lewinella agarilytica in hypersaline inland lakes. Sequence homology results suggested novel indigenous bacterial phylotypes. Key words: Siwa lakes, halophilic bacteria, 16S rRNA gene, molecular diversity.

Microbial Diversity during Fermentation of Sweet Paste, a Chinese Traditional Seasoning, Using PCR-Denaturing Gradient Gel Electrophoresis.

The aim of this study was to elucidate the changes in the microbial community and biochemical properties of a traditional sweet paste during fermentation. PCR-denaturing gradient gel electrophoresis (DGGE) analysis showed that Aspergillus oryzae was the predominant species in the koji (the fungal mixture), and the majority of the fungi isolated belonged to two Zygosaccharomyces species in the mash. The bacterial DGGE profiles revealed the presence of Bacillus subtilis during fermentation, and Lactobacillus acidipiscis, Lactobacillus pubuzihii, Lactobacillus sp., Staphylococcus kloosi, and several uncultured bacteria were also detected in the mash after 14 days of main fermentation. Additionally, during main fermentation, amino-type nitrogen and total acid increased gradually to a maximum of 6.77 ± 0.25 g/kg and 19.10 ± 0.58 g/kg (30 days) respectively, and the concentration of reducing sugar increased to 337.41 ± 3.99 g/kg (7 days). The 180-day fermented sweet paste contained 261.46 ± 19.49 g/kg reducing sugar and its pH value remained at around 4.65. This study has used the PCR-DGGE technique to demonstrate the microbial community (including bacteria and fungi) in sweet paste and provides useful information (biochemical properties) about the assessment of the quality of sweet paste throughout fermentation.

An introduction to the new degradation of polyvinyl chloride (PVC) shower hose

ABSTRACTRecently, polyvinyl chloride (PVC) shower hoses have hardened throughout the eastern Shizuoka Prefecture, Japan. According to the elemental analysis results, the carbon and oxygen concentrations were much lower in the damaged hoses. These findings reveal that oxygen-containing, carbon-based plasticisers may leach from the damaged hose. As a result, the hoses lost flexibility after one year of use. The highest number of heterotrophic bacteria was detected by PCR-denaturing gradient gel electrophoresis in the shower water, and the bacterial DNA concentration was higher when hot water contacted the hose surface. I conclude that the plasticiser was leached from the stiffened hose through a bioaccumulation process.

Factors influencing the distribution of lactic acid bacteria on Pennisetum grasses

AbstractSuccessful silage production depends on the promotion of lactic acid fermentation and one of the key factors influencing the fermentation quality of silage is lactic acid bacteria (LAB). Therefore, epiphytic LAB on grasses play an important role in silage production. The present research investigated the distribution of LAB on Pennisetum spp. and factors influencing its presence. Lactic acid bacteria on four grass cultivars, cv. MT‐1, cv. Mott, cv. Reyan No. 4 (Reyan) and cv. Guimu No. 1 (Guimu), were investigated by both culture‐dependent methods and 16S rRNA amplicon sequence analysis and polymerase chain reaction‐denaturing gradient gel electrophoresis (PCR‐DGGE). Mott grass had significantly lower contents of dry matter and lower pH and higher water‐soluble carbohydrates (WSC) content than the other three cultivars (P < 0.05). Mott also epiphyted the largest number of LAB (2.56 × 103 cfu g−1), while MT‐1 supported the least (41 cfu g−1). A diversity of LAB were recovered from the species including the following proportional representation: 33.4% Lactobacillus plantarum, 8.3% Lactobacillus pentosus, 8.3% Lactococcus lactis, 8.3% Leuconostoc citreum, 25.0% Weissella confusa and 16.7% Weissella cibaria on Mott, 20.0% L. plantarum, 20.0% L. pentosus and 60.0% Lc. lactis on MT‐1, 83.3% L. plantarum and 16.7% Lc. lactis on Reyan, 16.7% L. plantarum and 83.3% Lc. lactis on Guimu. The DGGE analyses of rRNA amplicons also revealed a similar distribution of these species. While the species composition and abundance of LAB differed among the four grasses. The number of LAB on grasses was positively correlated with the WSC content in the grasses (P < 0.01), and negatively correlated with the pH of the grasses (P < 0.05).

Evidence of Campylobacter jejuni reduction in broilers with early synbiotic administration

C. jejuni is considered a food safety concern to both public health authorities and consumers since it is the leading bacterial cause of food-borne gastroenteritis in humans. A high incidence of C. jejuni in broiler flocks is often correlated to pathogen recovery in retail poultry meat, which is the main source of human infection. In this work broiler chickens were fed with a synbiotic product mixed with conventional feed using two different administration strategies. The synbiotic was formulated with the microencapsulated probiotic Bifidobacterium longum PCB133 and a xylo-oligosaccharide (XOS). 1-day old chicks were infected with C. jejuni strain M1 (105 cells) and the synbiotic mixture was then administered starting from the first and the 14th day of chicken life (for animal groups GrpC and GrpB respectively). The goal of this study was to monitor C. jejuni load at caecum level at different sampling time by real-time PCR, identifying the best administration strategy. The microbiological analysis of the caecal content also considered the quantification of Campylobacter spp., Bifidobacterium spp. and B. longum.The supplemented synbiotic was more successful in reducing C. jejuni and Campylobacter spp. when administered lifelong, compared to the shorter supplementation (GrpB). Bifidobacterium spp. quantification did not show significant differences among treatments and B. longum PCB133 was detected in both supplemented groups evidencing the successful colonization of the strain. Moreover, the samples of the control group (GrpA) and GrpC were analysed with PCR-denaturing gradient gel electrophoresis (PCR-DGGE) to compare the caecal microbial community profiles at the beginning and at the end of the trial. Pattern analysis evidenced the strong influence of the early synbiotic supplementation, although a physiological change in the microbial community, occurring during growth, could be observed. Experimental results demonstrate that the synbiotic approach at farm level can be an effective strategy, combined with biosecurity measures, to improve the safety of poultry meat.

Impact of tree species on barley rhizosphere-associated fungi in an agroforestry ecosystem as revealed by 18S rDNA PCR-DGGE

Agroforestry systems have been considered a form of sustainable land use. Woody species in agroforestry systems can improve soil physicochemical properties by supplying leaf or stem litter. However, little is known about fungal community structure and diversity in agroforestry systems. In the present study, the culture-independent 18S rDNA-based polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) method was used to investigate fungal community structure in rhizosphere and bulk soil in Populus euramevicana-barley and Taxodium distichum-barley agroforestry systems. DGGE profiling and cluster analysis revealed that the fungal community structure in the rhizosphere was more complex than that of bulk soil. Our results also indicated that the rhizosphere fungal community in barley was less affected by T. distichum than by P. euramevicana. In addition, an increase in the relative abundance of certain rhizosphere fungal populations was detected in this agroforestry system. Sequencing of prominent DGGE bands revealed an increase in the rhizosphere of a fungal species belonging to the genera Chaetomium, which includes potential biocontrol agents. A rare cellulolytic fungus, Acremonium alcalophilum, was found in the bulk soil from P. euramevicana and barley grown under P. euramevicana. Taken together, our findings may provide new insights into agroforestry practices.

Operational and biological analyses of branched water-adjustment and combined treatment of wastewater from a chemical industrial park

ABSTRACTThe combined biological processes of branched water-adjustment, chemical precipitation, hydrolysis acidification, secondary sedimentation, Anoxic/Oxic and activated carbon treatment were used for chemical industrial wastewater treatment in the Taihu Lake Basin. Full-scale treatment resulted in effluent chemical oxygen demand, total nitrogen, NH3–N and total phosphorus of 35.1, 5.20, 3.10 and 0.15 mg/L, respectively, with a total removal efficiency of 91.1%, 67.1%, 70.5% and 89.3%, respectively. In this process, short-circuited organic carbon from brewery wastewater was beneficial for denitrification and second-sulfate reduction. The concentration of effluent fluoride was 6.22 mg/L, which also met the primary standard. Gas Chromatography-Mass Spectrometry analysis revealed that many types of refractory compounds were present in the inflow. Microbial community analysis performed in the summer by PCR-denaturing gradient gel electrophoresis and MiSeq demonstrated that certain special functional bacteria, such as denitrificans, phosphorus-accumulating bacteria, sulfate- and perhafnate-reducing bacteria, aromatic compound-degrading bacteria and organic fluoride-degrading bacteria, present in the bio-tanks were responsible for the acceptable specific biological pollutant reduction achieved.

Study of the treatment of domestic sewage using PVA gel beads as a biomass carrier

Abstract Technologies for domestic sewage treatment with low excess sludge yield rate and highly efficient biological treatment methods are needed, and the moving-bed biofilm reactor has great promise for meeting this need. To provide technical support for the treatment of typical domestic sewage, this paper provides an estimate of the bacterial diversity in polyvinyl alcohol (PVA) gel beads determined by a 16S-rRNA gene-based polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) approach, proposes a method for reducing the excess sludge yield rate, and describes the startup and continuous operation of the PVA technology. Domestic sewage was treated by a moving-bed biofilm reactor system using PVA gel beads as a biomass carrier. A significant amount of sphalerite, filamentous bacteria and bacillus were observed on the surface and internal structure of the PVA-gel beads by scanning electron microscope. Clostridiaceae bacterium, Alpha proteobacterium, Phenylobacterium haematophilum and Rhodobacter were identified as dominant bacteria strains using 16S-rRNA PCR-DGGE. The active sludge tanks were found to play a significant role in the reduction of excess sludge.

Comparison of Prokaryotic Diversity in Cold, Oligotrophic Remote Lakes of Chilean Patagonia

The prokaryotic abundance and diversity in three cold, oligotrophic Patagonian lakes (Témpanos, Las Torres and Mercedes) in the northern region Aysén (Chile) were compared in winter and summer using 16S rRNA fluorescence in situ hybridization and PCR-denaturing gradient gel electrophoresis technique. Prokaryotic abundances, numerically dominated by Bacteria, were quite similar in the three lakes, but higher in sediments than in waters, and they were also higher in summer than in winter. The relative contribution of Archaea was greater in waters than in sediments, and in winter rather than in summer. Despite the phylogenetic analysis indicated that most sequences were affiliated to a few taxonomic groups, mainly referred to Proteobacteria (consisting of Beta-, Alpha- and Gammaproteobacteria) and Euryarchaeota (mainly related to uncultured methanogens), their relative abundances differed in each sample, resulting in different bacterial and archaeal assemblages. In winter, the abundance of the dominant bacterial phylotypes were mainly regulated by the increasing levels of total organic carbon in waters. Archaeal abundance and richness appeared mostly influenced by pH in winter and total nitrogen content in summer. The prokaryotic community composition at Témpanos lake, located most northerly and closer to a glacier, greatly differed in respect to the other two lakes. In this lake was detected the highest bacterial diversity, being Betaproteobacteria the most abundant group, whereas Alphaproteobacteria were distinctive of Mercedes. Archaeal community associated with sediments was mainly represent by members related to the order of Methanosarcinales at Mercedes and Las Torres lakes, and by Crenarchaeota at Témpanos lake. Our results indicate that the proximity to the glacier and the seasonality shape the composition of the prokaryotic communities in these remote lakes. These results may be used as baseline information to follow the microbial community responses to potential global changes and to anthropogenic impacts.

A preliminary analysis of changes in composition of intestinal microbiota during infancy using polymerase chain reaction-denaturing gradient gel electrophoresis

To investigate the composition of bacteria in the stools of infants and the colonization of intestinal microbiota during infancy. Fresh stools were collected from 15 healthy infants at 0, 2, 4, 7, 10, 14, and 28 days and 3, 6, and 12 months after birth. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) was used to analyze the composition of intestinal microbiota, perform sequencing of dominant bacteria, and to analyze the changes in the composition of intestinal microbiota during infancy. DGGE fingerprint showed that the composition of intestinal microbiota during infancy changed significantly over time after birth. The cloning and sequencing results indicated that Proteobacteria colonized the earliest, mainly the obligate aerobes Enterobacter and Pseudomonas, followed by the obligate anaerobes (Clostridium hathewayi and Veillonella parvula) and the facultative anaerobe Clostridium ramosum in Firmicutes, and Verrucomicrobia. Actinobacteria colonized the latest, mainly Bifidobacterium, and gradually became dominant bacteria. During infancy, obligate aerobes colonize the intestinal tract the earliest, followed by obligate anaerobes and facultative anaerobes. Proteobacteria colonizes the earliest, followed by Firmicutes and Verrucomicrobia, and Actinobacteria, mainly Bifidobacterium, colonizes the latest.

Identification of microbial populations contributing to nitrification-associated nitrous oxide emission during cattle manure composting process with forced aeration

This study was conducted to clarify the fates of whole nitrogen cycle-associated microorganisms during laboratory-scale composting of cattle manure with continuous forced aeration, with the primary goal of identifying microbial populations that contributed greatly to high nitrous oxide (N2O) emissions triggered by nitrification. DNA microarray analysis targeting functional genes associated with the entire microbial nitrogen cycle was applied for the first time to investigate the fates of entire nitrogen cycle-associated microbial community during composting, and revealed increased diversity of nitrogen cycle-associated microorganisms, including ammonia-oxidizing populations, during active nitrification and N2O emission periods. Further analyses of the compositions of ammonia-oxidizing populations by DNA microarray, PCR-denaturing gradient gel electrophoresis and clone library methods revealed that Nitrosomonas europaea, Nitrosomonas eutropha, and their relatives were the dominant ammonia-oxidizing populations. Overall, the analyses of entire nitrogen cycle-associated microbial community and specific functional groups by a combination of different molecular biological methods in this study could confirm that N. europaea/eutropha and their relatives contributed greatly to high N2O emissions via active nitrification during cattle manure composting with forced aeration.

The Influence of Micro-Oxygen Addition on Desulfurization Performance and Microbial Communities during Waste-Activated Sludge Digestion in a Rusty Scrap Iron-Loaded Anaerobic Digester

In this study, micro-oxygen was integrated into a rusty scrap iron (RSI)-loaded anaerobic digester. Under an optimal RSI dosage of 20 g/L, increasing O2 levels were added stepwise in seven stages in a semi-continuous experiment. Results showed the average methane yield was 306 mL/g COD (chemical oxygen demand), and the hydrogen sulphide (H2S) concentration was 1933 ppmv with RSI addition. O2 addition induced the microbial oxidation of sulphide by stimulating sulfur-oxidizing bacteria and chemical corrosion of iron, which promoted the generation of FeS and Fe2S3. In the 6th phase of the semi-continuous test, deep desulfurization was achieved without negatively impacting system performance. Average methane yield was 301.1 mL/g COD, and H2S concentration was 75 ppmv. Sulfur mass balance was described, with 84.0%, 11.90% and 0.21% of sulfur present in solid, liquid and gaseous phases, respectively. The Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) analysis revealed that RSI addition could enrich the diversity of hydrogenotrophic methanogens and iron-reducing bacteria to benefit methanogenesis and organic mineralization, and impoverish the methanotroph (Methylocella silvestris) to reduce the consumption of methane. Micro-oxygen supplementation could enhance the diversity of iron-oxidizing bacteria arising from the improvement of Fe(II) release rate and enrich the sulphur-oxidising bacteria to achieved desulfurization. These results demonstrated that RSI addition in combination with micro-oxygenation represents a promising method for simultaneously controlling biogas H2S concentration and improving digestion performance.

Effects of Different Vegetation Types and Reclamation Years on Soil Bacterial Community Structure in Reclaimed Mine Areas

Effects of different vegetation types (Ulmus pumila, Larix gmelinii, Armeniaca vulgaris, Picea asperata and Robinia pseudoacacia) and reclamation years (15 and 20 years) on soil bacterial community structure in reclaimed Antaibao opencast mine areas were investigated using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and clone sequencing. For 20-year reclaimed soils, the significantly highest and lowest bacterial diversity were found in U. pumila and A. vulgaris stand, respectively, whereas no significant differences were found between the other three vegetation types. Under 15-year plantations, soil bacterial diversity index of P. asperata was significantly higher than that of R. pseudoacacia. Soil bacterial diversity index significantly increased in R. pseudoacacia planted soils but decreased in P. asperata treatment with the increase of reclaimed years. No significant change of soil bacterial community structure was observed in the same reclamation years based on the similarity coefficient analysis, cluster analysis and principal component analysis (PCA). Pearson correlation analysis demonstrated that bacterial diversity index was significantly positively correlated with soil pH. Nitrospira, Sphingomonas, Arthrobacter, Brachybacterium, Rhizobium as well as Mesorhizobium, which play important roles in the nitrogen cycle, degradation of polycyclic aromatic hydrocarbons and other organic matter, were identified by clone sequencing of the DGGE bands. Our results indicated that U. pumila and P. asperata were conducive to the recovery of soil bacterial diversity. The most dominant bacterial community from reclaimed mine soil would be beneficial for restoring wasteland contaminated soil and improving soil fertility.

Effect of fermentation temperature on hydrogen production from xylose and the succession of hydrogen‐producing microflora

AbstractBACKGROUNDHydrogen production through anaerobic dark fermentation is considered to be a potential biological process for xylose utilization. Temperature is one of the most important environmental factors, however, most studies have been carried out over a small temperature range. Batch tests were carried out to investigate the temperature effect on hydrogen production from xylose using a mixed culture over a wide temperature range (35–65°C). Hydrogen production, metabolite distribution and dynamics of microbial communities were investigated.RESULTSHydrogen‐producing cultures were successfully enriched at each tested temperature. Two peaks of fermentation temperature for hydrogen production were observed at 35 and 55°C (1.11 and 1.30 mol‐H2 mol−1‐xyloseconsumed, respectively). Butyrate and acetate were the major liquid metabolites at 35–60°C. While at 65°C the main by‐product was ethanol. Polymerase chain reaction‐denaturing gradient gel electrophoresis (PCR‐DGGE) analysis indicated that Clostridium species were dominant at 35–40°C, while Thermoanaerobacterium dominated at 45–60°C. Both species were found at 65°C, but with lowest microbial community diversity.CONCLUSIONHydrogen production efficiency was mainly affected by the liquid metabolite distributions, which depended mainly upon the temperature. Several microbial community structures were formed at mesophilic, transition, thermophilic and extreme‐thermophilic conditions, resulting in different metabolic pathways of xylose and hydrogen production capacity. © 2017 Society of Chemical Industry