PCR-denaturing Gradient Gel Electrophoresis Research Articles

Decreased microbial diversity and Lactobacillus group in the intestine of geriatric giant pandas (Ailuropoda melanoleuca)

It has been established beyond doubt that giant panda genome lacks lignin-degrading related enzyme, gastrointestinal microbes may play a vital role in digestion of highly fibrous bamboo diet. However, there is not much information available about the intestinal bacteria composition in captive giant pandas with different ages. In this study, we compared the intestinal bacterial community of 12 captive giant pandas from three different age groups (subadults, adults, and geriatrics) through PCR-denaturing gradient gel electrophoresis (DGGE) and real-time PCR analysis. Results indicated that microbial diversity in the intestine of adults was significantly higher than that of the geriatrics (p < 0.05), but not significant compared to the subadults (p > 0.05). The predominant bands in DGGE patterns shared by the twelve pandas were related to Firmicutes and Proteobacteria. Additionally, in comparison to healthy individuals, antibiotic-treated animals showed partial microbial dysbiosis. Real-time PCR analyses confirmed a significantly higher abundance of the Lactobacillus in the fecal microbiota of adults (p < 0.05), while other bacterial groups and species detected did not significantly differ among the three age groups (p > 0.05). This study revealed that captive giant pandas with different ages showed different intestinal bacteria composition.

Enhanced biodegradation of antibiotic combinations via the sequential treatment of the sludge resulting from pharmaceutical wastewater treatment using white-rot fungi Trametes versicolor and Bjerkandera adusta.

While anaerobic treatment is capable of treating pharmaceutical wastewater and removing antibiotics in liquid phases, solid phases may still contain significant amounts of antibiotics following this treatment. The main goal of this study was to evaluate the use of white-rot fungi to remove erythromycin, sulfamethoxazole, and tetracycline combinations from biosolids. The degradation potential of Trametes versicolor and Bjerkandera adusta was evaluated via the sequential treatment of anaerobic sludge. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analyses were used to identify competition between the autochthonous microbial communities and white-rot fungi. Solid-phase treatment using white-rot fungi substantially reduced antibiotic concentrations and toxicity in sludge. According to PCR-DGGE results, there is an association between species of fungus and antibiotic type as a result of the different transformation pathways of fungal strains. Fungal post-treatment of sludge represents a promising method of removing antibiotic combinations, therefore holding a significant promise as an environmentally friendly means of degrading the antibiotics present in sludge.

The duodenal microbiota composition in children with active coeliac disease is influenced by the degree of enteropathy

Abstract Objectives To establish whether the duodenal mucosa microbiota of children with active coelic disease (CD) and healthy controls (HC) differ in composition and biodiversity. Material and methods Samples of duodenal biopsies in 11 CD patients were obtained at diagnosis, and in 6 HC who were investigated for functional intestinal disorders of non-CD origin. Total duodenal microbiota and that belonging to the genus Lactobacillus using PCR-denaturing gradient gel electrophoresis (DGGE) were analyzed. The banding patterns obtained in the resulting gels were analyzed to determine the differences between the microbiota of CD patients and HC (FPQuest 4.5) while environmental indexes (richness, diversity and habitability) were calculated with the past version 2.17 program. Results The intestinal microbiota of patients with Marsh 3c lesion showed similarity of 98% and differs from other CD patients with other type of histologic lesion as Marsh 3a, Marsh 3b and Marsh 2. The main differences were obtained in ecological indexes belonging to the genus Lactobacillus, with significant richness, diversity and habitability reduction in CD patients. In CD bands were categorized primarily with Streptococcus, Bacteroides and Escherichia coli species. In HC the predominant bands were Bifidobacterium, Lactobacillus and Acinetobacter, though the Streptococcus and Bacteroides were lower. Conclusions The celiac patients with major histological affectation presented a similar microbiota duodenal. The ecological indexes applied to the genus Lactobacillus were significantly reduced in CD.

The effect of Jatropha curcas L. leaf litter decomposition on soil carbon and nitrogen status and bacterial community structure (Senegal)

The cultivation of Jatropha curcas L. as a biodiesel feedstock has been encouraged in Senegal to reduce dependence on fossil fuels and mitigate the effects of climate change. J. curcasis is a poisonous plant which sheds its leaves during the dry season. Although the leaves are toxic for animals, they can help to recycle soil organic matter. This study set out to determine the effect of the decomposition dynamics of green and senescent J. curcas leaves on the soil C and N contents and on the structure of the bacterial community. Leaf litter decomposition was studied for 4 months by laboratory incubation and samples were taken at the start of incubation and at 3, 28, 56, 90 and 120 days. Green leaves had a higher N content, higher concentrations of water soluble compounds and hemicelluloses, but a lower C:N ratio and lignin content than senescent leaves regardless of the cultivar. The cultivar, the type of litter and the interaction between them, all had a significant effect on the soil N content (p<0.0001, R2=0.995) and C:N ratio (p<0.0001, R2=0.998). However, the cultivar was the only factor that affected the leaf C content (p<0.05, R2=0.624). The initial N content explained the N-NH4+ mineralization at the start of decomposition and the initial lignin content explained the N-NH4+ mineralization at later stages of decomposition. The recalcitrant C content in the green leaves was estimated as being between 70.01 and 73.33% of the total C content and between 72 and 77.33% in senescent leaves. This suggests that Jatropha litter may contribute significantly to soil C sequestration. The results indicate that the soil had higher bacterial diversity in the later stages of litter decomposition, for both types of litter and all cultivars. Key words: Jatropha curcas L., leaf litter, nitrogen mineralization, carbon mineralization, bacterial community, polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE), Senegal.

Biodiversity and technological-functional potential of lactic acid bacteria isolated from spontaneously fermented quinoa sourdoughs.

To analyse lactic acid bacteria (LAB) diversity and technological-functional and safety properties of strains present during spontaneous fermented quinoa sourdoughs. Fermentation was performed by daily backslopping at 30°C for 10days. Autochthonous LAB microbiota was monitored by a biphasic approach combining random amplified polymorphic DNA (RAPD)-PCR and rRNA gene sequencing with PCR-denaturing gradient gel electrophoresis (DGGE) analysis. Identification and intraspecies differentiation allowed to group isolates within nine LAB species belonging to four genera. A succession of LAB species occurred during 10-days backslopping; Lactobacillus plantarum and Lactobacillus brevis were detected as dominant species in the consortium. The characterization of 15 representative LAB strains was performed based on the acidifying capacity, starch and protein hydrolysis, γ-aminobutyric acid and exopolysaccharides production, antimicrobial activity and antibiotic resistance. Strains characterization led to the selection of Lact.plantarum CRL1905 and Leuconostoc mesenteroides CRL1907 as candidates to be assayed as functional starter culture for the gluten-free (GF) quinoa fermented products. Results on native LAB microbiota present during quinoa sourdough fermentation will allow the selection of strains with appropriate technological properties to be used as a novel functional starter culture for GF-fermented products.

Diversity of parasitic fungi from soybean cyst nematode associated with long-term continuous cropping of soybean in black soil

ABSTRACTThe soybean cyst nematode (SCN) is a major yield-limiting pest of soybean. In this study, experiments were conducted to examine the diversity of parasitic fungi from SCN associated with disease-suppressive soil fields in Northeast China. Soil samples were collected from three fields under different rotation systems that were established in 1991: (1) a continuous long-term cropping field with soybean (SSSS) that had been shown to be SCN-suppressive, (2) cycles of three-year rotation with corn, soybean, and wheat (WCS), and (3) continuous cropping field with three-year cycles of two years soybean and one-year corn (SSC). In the traditional method result, cyst densities of SCN declined as increase of parasitic fungi, and the percentage of parasitic fungi associated with cyst of SCN was higher in SSSS field than other two fields. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) also showed that parasitic fungi of SCN were also increased in SSSS field, compared with the other two fields. Principal component analysis based on PCR-DGGE data revealed that fungal communities on cysts could be divided into three groups: one group occurred in SSSS, and the other two groups were in WCS and SSC fields, respectively. Long-term cropping with soybean monoculture in the black soil field might increase parasitic fungi of SCN. These fungal communities may play an important role in the ecological suppression of SCN in disease-suppressive soil.

Reducing bias in complex microbial community analysis in shrimp based on propidium monoazide combined with PCR-DGGE

Microbial community plays a prominent role in food quality and safety, which was regarded as the major cause for food spoilage and foodborne disease. Pre-PCR propidium monoazide (PMA) treatment is an effective approach to exclude DNA amplification signal from dead cells, and application of this method in the analysis of bacterial community might help us better understand the diversity and distribution of viable bacteria in food. In this study, the effects of PMA treatment on bacterial community characteristics in shrimp were assessed by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) fingerprinting technique. The results showed that the microbial diversity was dramatically influenced by PMA treatment. During storage at 4 °C, Shannon–Wiener index and PCoA analysis indicated there were significant differences between bacterial diversity in samples treated with and without PMA. At the level of species, compared with samples without PMA treatment, Vibrio cholera, Exiguobacterium acetylicum, Aeromonas enteropelogenes, Eubacterium multiforme, Vibrio mimicus in PMA-treated samples disappeared during storage. When samples stored at 25 °C, PMA treatment has little influence on banding patterns in DGGE profiles. In conclusion, this study provides an effective tool for precisely monitoring the diversity of viable bacteria in food samples, especially at low temperature.

Differences in the Biodiversity of the Fecal Microbiota of Infants With Rotaviral Diarrhea and Healthy Infants.

BackgroundRotaviral diarrhea (RD) has been associated with the biodiversity of the fecal microbiota in infants; however, the differences in the biodiversity of the fecal microbiota between infants with RD and healthy (H) infants have not been clearly elucidated.ObjectivesThis study aimed to reveal the changes in the biodiversity of the fecal microbiota of infants with RD.Patients and MethodsFor this study, 30 fecal samples from 15 RD infants and 15 H infants were collected. The biodiversity of the fecal microbiota from the two groups was compared via polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and gene sequencing.ResultsThe Shannon-Weaver index showed that the biodiversity of the fecal microbiota from the RD infants was significantly lower (P < 0.05) than that from the H infants. All fifteen RD infants were grouped into one cluster and were separated from the H infants by the un weighted-pair group method, with the arithmetic average (UPGMA) clustering algorithm. In addition, when compared with the healthy infants, the communities of the dominant microbes, Lactobacillus and Bifidobacterium, in the fecal microbiota from the RD infants have obviously changed.ConclusionsWith regard to improving the understanding of the differences in the biodiversity of the fecal microbiota between RD infants and H infants, the findings of this study can provide a possible basis to reveal the relationship between RD and intestinal microbiota.

Fiber degradation potential of natural co-cultures of Neocallimastix frontalis and Methanobrevibacter ruminantium isolated from yaks (Bos grunniens) grazing on the Qinghai Tibetan Plateau

Several natural anaerobic fungus-methanogen co-cultures have been isolated from rumen and feces source of herbivores with strong fiber degrading ability. In this study, we isolated 7 Neocallimastix with methanogen co-cultures from the rumen of yaks grazing on the Qinghai Tibetan Plateau. Based on morphological characteristics and internal transcribed spacer 1 sequences (ITS1), all the fungi were identified as Neocallimastix frontalis. The co-cultures were confirmed as the one fungus – one methanogen pattern by the PCR-denatured gradient gel electrophoresis (DGGE) assay. All the methanogens were identified as Methanobrevibacter ruminantium by 16s rRNA gene sequencing. We investigated the biodegrading capacity of the co-culture (N. frontalis + M. ruminantium) Yaktz1 on wheat straw, corn stalk and rice straw in a 7 days-incubation. The in vitro dry matter digestibility (IVDMD), acid detergent fiber digestibility (ADFD) and neural detergent fiber digestibility (NDFD) values of the substrates in the co-culture were significantly higher than those in the mono-culture N. frontalis Yaktz1. The co-culture exhibited high polysaccharide hydrolase (xylanase and FPase) and esterase activities. The xylanase in the co-culture reached the highest activity of 12500 mU/ml on wheat straw at the day 3 of the incubation. At the end of the incubation, 3.00 mmol–3.29 mmol/g dry matter of methane were produced by the co-culture. The co-culture also produced high level of acetate (40.00 mM–45.98 mM) as the end-product during the biodegradation. Interestingly, the N. frontalis Yaktz1 mono-culture produced large amount of lactate (8.27 mM–11.60 mM) and ethanol (163.11 mM–242.14 mM), many times more than those recorded in the previously reported anaerobic fungi. Our data suggests that the (N. frontalis + M. ruminantium) Yaktz1 co-culture and the N. frontalis Yaktz1 mono-culture both have great potentials for different industrial use.

Winter cover crops alter methanotrophs community structure in a double-rice paddy soil

Methanotrophs play a vital role in the mitigation of methane emission from soils. However, the influences of cover crops incorporation on paddy soil methanotrophic community structure have not been fully understood. In this study, the impacts of two winter cover crops (Chinese milk vetch (Astragalus sinicus L.) and ryegrass (Lolium multiflorum Lam.), representing leguminous and non-leguminous cover crops, respectively) on community structure and abundance of methanotrophs were evaluated by using PCR-DGGE (polymerase chain reaction-denaturing gradient gel electrophoresis) and real-time PCR technology in a double-rice cropping system from South China. Four treatments were established in a completely randomized block design: 1) double-rice cropping without nitrogen fertilizer application, CK; 2) double-rice cropping with chemical nitrogen fertilizer application (200 kg ha–1 urea for entire double-rice season), CF; 3) Chinese milk vetch cropping followed by double-rice cultivation with Chinese milk vetch incorporation, MV; 4) ryegrass cropping followed by double-rice cultivation with ryegrass incorporation, RG. Results showed that cultivating Chinese milk vetch and ryegrass in fallow season decreased soil bulk density and increased rice yield in different extents by comparison with CK. Additionally, methanotrophic bacterial abundance and community structure changed significantly with rice growth. Methanotrophic bacterial pmoA gene copies in four treatments were higher during late-rice season (3.18×107 to 10.28×107 copies g–1 dry soil) by comparison with early-rice season (2.1×107 to 9.62×107 copies g–1 dry soil). Type I methanotrophs absolutely predominated during early-rice season. However, the advantage of type I methanotrophs kept narrowing during entire double-rice season and both types I and II methanotrophs dominated at later stage of late-rice.

Microbial Community Structure of Korean Cabbage Kimchi and Ingredients with Denaturing Gradient Gel Electrophoresis.

Kimchi is a traditional Korean fermented vegetable food, the production of which involves brining of Korean cabbage, blending with various other ingredients (red pepper powder, garlic, ginger, salt-pickled seafood, etc.), and fermentation. Recently, kimchi has also become popular in the Western world because of its unique taste and beneficial properties such as antioxidant and antimutagenic activities, which are derived from the various raw materials and secondary metabolites of the fermentative microorganisms used during production. Despite these useful activities, analysis of the microbial community present in kimchi has received relatively little attention. The objective of this study was to evaluate the bacterial community structure from the raw materials, additives, and final kimchi product using the culture-independent method. Specifically, polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) was used to analyze the 16S rRNA partial sequences of the microflora. One primer set for bacteria, 341F(GC)-518R, reliably produced amplicons from kimchi and its raw materials, and these bands were clearly separated on a 35-65% denaturing gradient gel. Overall, 117 16S rRNA fragments were identified by PCR-DGGE analysis. Pediococcus pentosaceus, Leuconostoc citreum, Leuconostoc gelidum, and Leuconostoc mesenteroides were the dominant bacteria in kimchi. The other strains identified were Tetragenococcus, Pseudomonas, Weissella, and uncultured bacterium. Comprehensive analysis of these microorganisms could provide a more detailed understanding of the biologically active components of kimchi and help improve its quality. PCR-DGGE analysis can be successfully applied to a fermented food to detect unculturable or other species.

Effects of combined organic acid treatments during the cutting process on the natural microflora and quality of chicken drumsticks

Reducing the microbial load on broiler chicken carcasses at each stage of poultry meat processing, is highly important for hygienic meat production. This study was conducted to assess the efficacy of various combined organic acid treatments during the cutting process on the microbial decontamination of chicken drumsticks. Changes in naturally occurring microflora before and after treatment were analyzed through microbiological counting and polymerase chain reaction–denaturing gradient gel electrophoresis (PCR-DGGE). Results revealed that the most effective treatments obtained through the combination of orthogonal design and sensory evaluation were as follows: 0.5% lactic acid (w/v), 1% citric acid (w/v), and spray-washing for 30 s. Microbiological counting results and PCR-DGGE analysis indicated that the microbial load on the chicken drumsticks decreased significantly after the treatment was administered. The treatment did not affect the physicochemical properties and sensory attributes of the quick-frozen chicken drumsticks during storage. Therefore, the proposed technique could be used to improve the microbial quality of chicken drumsticks. The technique could also be employed in poultry meat production chains.

Impact of salinity on organic matter and nitrogen removal from a municipal wastewater RO concentrate using biologically activated carbon coupled with UV/H2O2

The concentrate streams generated from reverse osmosis (RO)-based municipal wastewater reclamation processes contain organic substances and nutrients at elevated concentrations, posing environmental and health risks on their disposal to confined receiving environments such as bays. The impact of salinity (TDS at 7, 10 and 16 g/L) of a RO concentrate (ROC) on the treatment efficiency of a biological activated carbon (BAC) system after pre-oxidation with UV/H2O2 was characterised in terms of removal of organic matter and nitrogen species, and the bacterial communities. Organic matter removal was comparable for the ROC over the tested salinity range, with 45–49% of DOC and 70–74% of UVA254 removed by the combined treatment. However, removal in total nitrogen (TN) was considerably higher for the ROC at the high salinity (TDS ∼ 16 mg/L) compared with the low (∼7 g/L) and medium salinity (∼10 g/L). Effective nitrification with high ammonium removal (>90%) was achieved at all salinity levels, whereas greater denitrification (39%) was obtained at high salinity than low (23%) and medium salinity (27%) which might suggest that the bacterial communities contributing to the greater denitrification were more halotolerant. Microbiological characterisation using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and culture based techniques showed that diversified bacterial communities were present in the BAC system as evident from different 16S rDNA. The major bacterial groups residing on the BAC media belonged to Bacillus (Firmicutes), Pseudomonas (γ-Proteobacteria), and Rhodococcus (Actinobacteria) for all salinity levels, confirming that these microbial communities could be responsible for carbon and nitrogen removal at the different salinity levels. This has implications in understanding the effectiveness and robustness of the BAC system over the salinity range of the ROC and so would be useful for optimising the treatment efficiency of the BAC system.

Nitrogen removal properties in a continuous marine anammox bacteria reactor under rapid and extensive salinity changes

Salinity tolerance is one of the most important factors for the application of bioreactors to high-salinity wastewater. Although marine anammox bacteria (MAB) might be expected to tolerate higher salinities than freshwater anammox bacteria, there is little information on the effects of salinity on MAB activity. This study aimed to reveal the nitrogen removal properties in a continuous MAB reactor under conditions of rapid and extensive salinity changes. The reactor demonstrated stable nitrogen removal performance with a removal efficiency of over 85% under salinity conditions ranging from 0 to 50 g/L NaCl. The reactor performance was also well maintained, even though the salinity was rapidly changed from 30 to 50 g/L and from 30 to 0 g/L. Other evidence suggested that the seawater medium used contained components essential for effective MAB performance. Bacterial community analysis using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) showed that planctomycete UKU-1, the dominant MAB species in the inoculum, was the main contributor to anammox activity under all conditions. The PCR-DGGE using a universal bacterial primer set showed different DNA band patterns between the reactor biomass sample collected under conditions of 75 g/L NaCl and all other conditions (0, 30, 50 and freshwater-medium). All DNA sequences determined were very similar to those of bacterial species from marine environments, anaerobic environments, or wastewater-treatment facilities.

Shifts of microbial communities of wheat (Triticum aestivum L.) cultivation in a closed artificial ecosystem.

The microbial communities of plant ecosystems are in relation to plant growing environment, but the alteration in biodiversity of rhizosphere and phyllosphere microbial communities in closed and controlled environments is unknown. The purpose of this study is to analyze the change regularity of microbial communities with wheat plants dependent-cultivated in a closed artificial ecosystem. The microbial community structures in closed-environment treatment plants were investigated by a culture-dependent approach, polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), and Illumina Miseq high-throughput sequencing. The results indicated that the number of microbes decreased along with time, and the magnitude of bacteria, fungi, and actinomycetes were 10(7)-10(8), 10(5), and 10(3)-10(4)CFU/g (dry weight), respectively. The analysis of PCR-DGGE and Illumina Miseq revealed that the wheat leaf surface and near-root substrate had different microbial communities at different periods of wheat ecosystem development and showed that the relative highest diversity of microbial communities appeared at late and middle periods of the plant ecosystem, respectively. The results also indicated that the wheat leaf and substrate had different microbial community compositions, and the wheat substrate had higher richness of microbial community than the leaf. Flavobacterium, Pseudomonas, Paenibacillus, Enterobacter, Penicillium, Rhodotorula, Acremonium, and Alternaria were dominant in the wheat leaf samples, and Pedobacter, Flavobacterium, Halomonas, Marinobacter, Salinimicrobium, Lysobacter, Pseudomonas, Halobacillus, Xanthomonas, Acremonium, Monographella, and Penicillium were dominant populations in the wheat near-root substrate samples.

Decontamination of heavy metal laden sewage sludge with simultaneous solids reduction using thermophilic sulfur and ferrous oxidizing species

A possibility of using simultaneous sewage sludge digestion and metal leaching (SSDML) process at the thermophilic temperature to remove heavy metals and suspended solids from sewage sludge is explored in this study. Though thermophilic sludge digestion efficiently produces a stable sludge, its inability to remove heavy metals requires it to be used in tandem with another process like bioleaching for metal reduction. Previously, different temperature optima were known for the heterotrophs (thermophilic) responsible for the sludge digestion and the autotrophs involved in bioleaching (mesophilic), because of which the metal concentration was brought down separately in a different reactor. In our study, SSDML process was carried out at 50 °C (thermophilic) by using ferrous sulfate (batch-1) and sulfur (batch-2) as the energy source in two reactors. The concentration of volatile suspended solids reduced by >40% in both batches, while that of heavy metals zinc, copper, chromium, cadmium and nickel decreased by >50% in both batch-1 and batch-2. Lead got leached out only in batch-1. Using 16S rRNA gene-based PCR-denaturing gradient gel electrophoresis analysis, Alicyclobacillus tolerans was found to be the microorganism responsible for lowering the pH in both the reactors at thermophilic temperature. The indicator organism count was also below the maximum permissible limit making sludge suitable for agricultural use. Our results indicate that SSDML at thermophilic temperature can be effectively used for reduction of heavy metals and suspended solids from sewage sludge.

Diversity of ammonia-oxidizing bacteria in relation to soil environment in Ebinur Lake Wetland

ABSTRACTAmmonia oxidation is the first and rate-limiting step of nitrification and is carried out by ammonia-oxidizing bacteria (AOB). Ebinur Lake Wetland, the most representative temperate arid zone wetland ecosystem in China, is the centre of oasis and desertification of the northern slope of Tianshan conjugate. Soil samples were collected from three sites (Tamarix ramosissima, Halocnemum strobilaceum and Phragmites australis) and different soil layers (0–5, 5–15, 15–25 and 25–35 cm) in this wetland in spring, summer and autumn and were used to characterize the diversity of AOB based on the ammonia monooxygenase (amoA) gene. Polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE) and bivariate correlation analysis were used to analyse the relationship between the diversity of AOB and soil environment factors. The PCR-DGGE indicated that the diversity of AOB was high in the entire sample and the Shannon diversity index varied from 1.369 to 2.471. The phylogenetic analysis showed that the amoA fragments were grouped into Nitrosospira sp. and Nitrosomonas sp. Most amoA gene sequences fell within the Nitrosospira sp. cluster, and only a few sequences were clustered with Nitrosomonas sp., indicating that Nitrosospira sp. may be more adaptable than Nitrosomonas sp. in this area. Bivariate correlation analysis showed that the diversity of AOB was significantly correlated with soil organic matter, conductivity, total phosphorus and nitrate in the Ebinur Lake Wetland in Xinjiang.

Nitritation via heat shock using immobilized active sludge aggregates

The nitritation process is considered to be one of the most energy-saving and efficient methods for treating polluted water. In this study, active sludge was immobilized with waterborne polyurethane (WPU), and a heat-shock method was employed to treat the immobilized aggregates. When environmental factors that adversely affect nitritation (pH, dissolved oxygen, temperature, etc.) were controlled, steady nitrite nitrogen accumulation was also successfully achieved. We investigated the effect of temperature and timing of the heat-shock method on the activity of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). Subsequently, temperatures of 60 and 70°C were selected to evaluate the nitritation stability of the heat-shocked immobilized aggregates, and continuous-flow experiments were conducted. The results of preliminary experiments indicated that NOB can be completely deactivated at temperatures above 60°C after 10 min, whereas AOB continued to exhibit some activity. In addition, the effect of temperature was more significant than the effect of heating time for NOB. The results of the nitritation stability assay indicated that the heat-shocked immobilized aggregates that formed at two temperatures remained in stable nitritation; however, the performance of the AOB was decreased at the highest temperature tested. A high temperature and long duration were required for stable nitritation. In the continuous-flow experiments, we discovered that nitrate nitrogen accumulation occurred after 65 d of operation. Therefore, the immobilized aggregates were heat shocked again, and nitrite nitrogen accumulation recurred in the reactor (results not shown). Heat shock is an effective method for stabilizing the nitritation of immobilized nitrifying sludge; this method also provides new ideas for a nitritation-based nitrogen removal process. Key innovations are: (1) a new nitration method based on heat shock has been proposed to generate steady nitritation using immobilized bacterial aggregates; (2) polymerase chain reaction (PCR) has been applied to analyze variations in the AOB and NOB populations of immobilized aggregates before and after heat shock; (3) optimal heat-shock conditions (including duration and temperature) have been identified.

The performance of the microbial fuel cell-coupled constructed wetland system and the influence of the anode bacterial community

ABSTRACTIn order to analyse the influences of substrate and electrode on the performance of microbial fuel cell-coupled constructed wetland (CW-MFC), the electrical generation efficiencies, the decolourization mechanism of reactive brilliant red X-3B, and the microbial communities in the anode were investigated. The closed circuit reactor fed with a mixture of X-3B and glucose (166.7 mg/L X-3B and 140 mg/L glucose) (the mixture CC reactor) got a decolourization rate of 92.79%, which was higher than the open circuit reactor (the mixture OC reactor) and the reactor fed with X-3B (the X-3B reactor). The mixture CC reactor got a maximum power density of 0.200 W/m3, which was much higher than the X-3B reactor. The intermediates produced by X-3B decolourization were further degraded in CW-MCs. The PCR-denatured gradient gel electrophoresis analysis indicated the dominance of Proteobacteria-like 16S rRNA gnen sequences. The brightest band was detected to be dominant by a Lactobacillus kefiranofaciens-like sequence. The electrogenic bacteria-associated sequences, such as Geobacter metallireducens and Desulfobulbaceae, both existed in the closed circuit and the open circuit reactors, accompanied with Desulfobacterium sp., Klebsiella sp., Aminobacter sp., Flavobacterium sp., Thauera aromatic, and Sphingomonas sp. The abundances of Geobacter sulfurreducens and Betaproteobacteria in the mixture CC reactor were 32.2% and 7.2%, respectively, and were higher than those in the mixture OC reactor. In summary, substrate and electrode can promote the performance of the CW-MFC and have effects on the microbial community in the anode of the CW-MFC.

Bacterial dynamics and metabolite changes in solid-state acetic acid fermentation of Shanxi aged vinegar.

Solid-state acetic acid fermentation (AAF), a natural or semi-controlled fermentation process driven by reproducible microbial communities, is an important technique to produce traditional Chinese cereal vinegars. Highly complex microbial communities and metabolites are involved in traditional Chinese solid-state AAF, but the association between microbiota and metabolites during this process are still poorly understood. In this study, we performed amplicon 16S rRNA gene sequencing on the Illumina MiSeq platform, PCR-denaturing gradient gel electrophoresis, and metabolite analysis to trace the bacterial dynamics and metabolite changes under AAF process. A succession of bacterial assemblages was observed during the AAF process. Lactobacillales dominated all the stages. However, Acetobacter species in Rhodospirillales were considerably accelerated during AAF until the end of fermentation. Quantitative PCR results indicated that the biomass of total bacteria showed a "system microbe self-domestication" process in the first 3days, and then peaked at the seventh day before gradually decreasing until the end of AAF. Moreover, a total of 88 metabolites, including 8 organic acids, 16 free amino acids, and 66 aroma compounds were detected during AAF. Principal component analysis and cluster analyses revealed the high correlation between the dynamics of bacterial community and metabolites.