Chain Reaction-denaturing Gradient Gel Electrophoresis Research Articles

Effects of moisture content on the performance of a two-stage thermophilic biofilter and choice of irrigation rate

Abstract The stabilization, innocuity, and reutilization of sludge can be obtained via the co-combustion of sewage sludge in cement producing kilns. Off-gases containing odours and volatile organic compounds (VOCs) are generated during the process of sewage sludge drying. A two-stage thermophilic biofilter was constructed in a cement plant to remove SO2, NH3, and total VOCs (TVOCs) from the plant’s exhaust. The average concentrations of TVOCs, NH3, and SO2 in the inlet stream were 164.33, 38.46, and 51.12 mg/m3, and in the outlet stream were 44.54, 9.18, and 3.91 mg/m3, respectively. During biofilter operation, nutrient solution was circularly pumped onto the packing material to ensure the packing material remained moist. The moisture content of the packing material increased from 40 to 85% as the irrigation rate varied from 0.05 to 0.42 m3/h. The removal efficiencies of NH3 and TVOCs changed accordingly. When the irrigation rate was 0.3 m3/h, removal efficiencies were 88.71% for NH3 and 81.38% for TVOCs. More than 90% SO2 removal occurred throughout the operation. The polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) method was used to assay the microbial population. Both microbial counts and diversity increased when the irrigation rate increased.

PCR-DGGE Analysis on Microbial Community Structure of Rural Household Biogas Digesters in Qinghai Plateau.

To investigate contribution of environmental factor(s) to microbial community structure(s) involved in rural household biogas fermentation at Qinghai Plateau, we collected slurry samples from 15 digesters, with low-temperature working conditions (11.1-15.7°C) and evenly distributed at three counties (Datong, Huangyuan, and Ledu) with cold plateau climate, to perform polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and further sequencing. The bacterial communities in the total 15 digesters were classified into 38 genera with Mangroviflexus (12.1%) as the first dominant, and the archaeal communities into ten genera with Methanogenium (38.5%) as the most dominant. For each county, the digesters with higher biogas production, designated as HP digesters, exclusively had 1.6-3.1°C higher fermentation temperature and the unique bacterial structure composition related, i.e., unclassified Clostridiales for all the HP digesters and unclassified Marinilabiliaceae and Proteiniclasticum for Ledu HP digesters. Regarding archaeal structure composition, Methanogenium exhibited significantly higher abundances at all the HP digesters and Thermogymnomonas was the unique species only identified at Ledu HP digesters with higher-temperature conditions. Redundancy analysis also confirmed the most important contribution of temperature to the microbial community structures investigated. This report emphasized the correlation between temperature and specific microbial community structure(s) that would benefit biogas production of rural household digesters at Qinghai Plateau.

Dynamic changes of bacterial communities and nitrite character during northeastern Chinese sauerkraut fermentation.

Northeastern Chinese sauerkraut is a well-known traditional fermented vegetable in China. Incomplete identification of the microorganisms' (bacteria in spontaneous fermentation) diversity and accumulation of nitrite make it difficult to normalize the fermentation process and product qualities of northeastern Chinese sauerkraut. Conventional culturing and polymerase chain reaction-denaturing gradient gel electrophoresis methods were combined to describe microbial structure and diversity. Lactobacillus, Leuconostoc, Enterobacter, Accumulibacter, Thermotoga, Pseudomonas, Clostridium, Rahnella and Citrobacter were predominant microorganisms in different fermentation periods. The pH value and nitrite concentration presentedacertainrelevancetothe amount of lacticacidbacteria. Lactobacillus and Leuconostoc had the ability to decrease nitrite by inhibiting nitrate-reducing bacteria such as Enterobacter. Therefore, Northeastern Chinese sauerkraut should not be eaten until 4weeks of fermentation for the safety and quality of fermented foods. Northeastern Chinese sauerkraut is rich in lacticacidbacteria, which demonstrate its ability as an excellent probiotic for applications in functional foods.

Probiotic strain Stenotrophomonas acidaminiphila BJ1 degrades and reduces chlorothalonil toxicity to soil enzymes, microbial communities and plant roots

Chlorothalonil, a non-systemic and broad-spectrum fungicide, is widely used to control the pathogens of agricultural plants. Although microbial degradation of chlorothalonil is known, we know little about the colonization and degradation capacity of these microbes in the natural and semi-natural soil environments. Therefore, we studied the colonization and detoxification potential of a chlorothalonil degrading Stenotrophomonas acidaminiphila probiotic strain BJ1 in the soil under green conditions. The results from polymerase chain reaction-denaturing gradient gel electrophoresis demonstrated that probiotic strain BJ1 successfully colonized the soil by competing with the native biota. Moreover, the bacterial inoculation stimulated some members of indigenous soil microbial communities. Meantime, the degradation half-life of chlorothalonil decreased from 9.0 to 4.9 days in the soil environment. Moreover, the results from enzymatic activities and micronucleus test of Vicia faba root tips showed that the probiotic strain BJ1 reduced the ecotoxicity and genotoxicity of chlorothalonil in the soil. We suggest that probiotic strains like BJ1 could potentially alleviate the toxic effects of pesticides on soil microbes and plant roots under greenhouse conditions.

Profiling of heavy metal(loid)-resistant bacterial community structure by metagenomic-DNA fingerprinting using PCR–DGGE for monitoring and bioremediation of contaminated environment

Frequent exposure of microbes to hazardous metalloids/heavy metals in contaminated environment results in the development of heavy metal(loid)-resistance properties. The study attempted to assess the profile of elevated arsenic (As), cadmium (Cd) and mercury (Hg)—resistant bacterial community structures of sludge (S1, India), sludge and sediment (S2 and S3, Japan) and sediment (S4, Vietnam) samples by metagenomic-DNA fingerprinting using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR–DGGE) for monitoring and bioremediation of hazardous metal(loid) contamination in environment. The results revealed that As-resistant bacteria were dominant compared to Cd- and Hg-resistant bacteria with higher species diversity (Lysinibacillus sp., Uncultured soil bacterium clone, Staphylococcus sciuri, Bacillus fastidiosus, Bacillus niacini, Clostridium sp. and Bacillus sp.) in S1 and S4 than that of S2 and S3 samples. The occurrence of dominant As-resistant bacteria may indicate arsenic contamination in the investigated coastal habitats of India, Japan and Vietnam. The As-, Cd- and Hg-resistant bacteria/bacterial consortiums showed appreciable uptake ability of respective metal(loid) (0.042–0.125 mg As/l, 0.696–0.726 mg Cd/l and 0.34–0.412 mg Hg/l). Therefore, it might be concluded that the profiling of metalloids/heavy metal-resistant bacterial community structure by metagenomic-DNA fingerprinting using PCR–DGGE could be used to explore high metal(loid)-resistant bacteria for applying in metal(loid) bioremediation and as an indicator for monitoring hazardous metal(loid) contamination in environment.

Bio-protective potential of lactic acid bacteria: Effect of Lactobacillus sakei and Lactobacillus curvatus on changes of the microbial community in vacuum-packaged chilled beef

ObjectiveThis study was to determine the bacterial diversity and monitor the community dynamic changes during storage of vacuum-packaged sliced raw beef as affected by Lactobacillus sakei and Lactobacillus curvatus.MethodsL. sakei and L. curvatus were separately incubated in vacuumed-packaged raw beef as bio-protective cultures to inhibit the naturally contaminating microbial load. Dynamic changes of the microbial diversity of inoculated or non-inoculated (control) samples were monitored at 4°C for 0 to 38 days, using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE).ResultsThe DGGE profiles of DNA directly extracted from non-inoculated control samples highlighted the order of appearance of spoilage bacteria during storage, showing that Enterbacteriaceae and Pseudomonas fragi emerged early, then Brochothrix thermosphacta shared the dominant position, and finally, Pseudomonas putida showed up became predominant. Compared with control, the inoculation of either L. sakei or L. curvatus significantly lowered the complexity of microbial diversity and inhibited the growth of spoilage bacteria (p<0.05). Interestingly, we also found that the dominant position of L. curvatus was replaced by indigenous L. sakei after 13 d for L. curvatus-inoculated samples. Plate counts on selective agars further showed that inoculation with L. sakei or L. curvatus obviously reduced the viable counts of Enterbacteraceae, Pseudomonas spp. and B. thermosphacta during later storage (p< 0.05), with L. sakei exerting greater inhibitory effect. Inoculation with both bio-protective cultures also significantly decreased the total volatile basic nitrogen values of stored samples (p<0.05).ConclusionTaken together, the results proved the benefits of inoculation with lactic acid bacteria especially L. sakei as a potential way to inhibit growth of spoilage-related bacteria and improve the shelf life of vacuum-packaged raw beef.

Evaluation of biochars from different stock materials as carriers of bacterial strain for remediation of heavy metal-contaminated soil

Two kinds of biochars, one derived from corn straw and one from pig manure, were studied as carriers of a mutant genotype from Bacillus subtilis (B38) for heavy metal contaminated soil remediation. After amendment with biochar, the heavy metal bioavailability decreased. Moreover, the heavy metal immobilization ability of the biochar was enhanced by combining it with B38. The simultaneous application of B38 and pig manure-derived biochar exhibited a superior effect on the promotion of plant growth and the immobilization of heavy metals in soil. The plant biomass increased by 37.9% and heavy metal concentrations in the edible part of lettuce decreased by 69.9–96.1%. The polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) profiles revealed that pig manure-derived biochar could enhance the proliferation of both exotic B38 and native microbes. These results suggest that B38 carried by pig manure-derived biochar may be a promising candidate for the remediation of soils contaminated by multiple heavy metals.

Microbial community response and SDS-PAGE reveal possible mechanism of waste activated sludge acidification enhanced by microaeration coupled thermophilic pretreatment

Aiming to strengthen the performance of waste activated sludge digestion, pretreatment is a prerequisite for keeping operations within an industrially acceptable time- frame. In this study, the performance of microaeration coupled with thermophilic (MT) pretreatment on waste activated sludge solubilization and acidification was investigated. The results showed that the maximum soluble organics concentrations, including soluble proteins and carbohydrates, reached 2290 mg COD/L in 24 h when the ventilation rate and temperature were 0.05 vvm and 70 °C. SDS-polyacrylamide gel electrophoresis (SDS-PAGE) test on pure-culture (E.coli) and scanning electron microscopy analysis indicated that MT pretreatment effectively destroyed microbial cell wall and resulted in an increase in soluble proteins. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis revealed that MT pretreatment reduced the diversity of the bacterial community during pretreatment. Along with the dissolution of a large quantity of organic matter, microbial species such as Lactococcus and Methyloversatilis bloomed at the end of acidification period, which contributed to SCFAs production. This study revealed that MT pretreatment facilitated the WAS hydrolysis efficiency and enhanced SCFAs (3149 mg COD/L with fermentation for 96 h) production, especially for acetic acid (50%) accumulation, which provides a new perspective for the application and recovery of sludge resources.

Biodegradation of aniline by a novel bacterial mixed culture AC

In this study, a novel bacterial mixed culture AC, capable of degrading aniline, was developed by us. The mixed culture presented better aniline degradation abilities comparing with different pure strains. Aniline at high concentrations could be tolerated and degraded by the mixed culture. An almost complete degradation of aniline at the concentration of 1500 mg L−1 was observed within 60 h. To provide further insight into the microbial diversity of the bacterial mixed culture, polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and high-throughput sequencing analyses were performed. PCR-DGGE profiles revealed that the microbial community had changed with varying initial concentrations of aniline. The taxonomic classification of the bacterial communities showed that the acclimated mixed culture was mainly composed of Serratia, Escherichia/Shigella, Bacillus and Acinetobacter. The mixed culture was immobilized by entrapment in different polumeric matrices. The immobilized bead exhibited good activity for aniline degradation and kept stable during successive repeated experiments. The effects of physico-chemical parameters on the aniline degradation were determined. Comparing with the free cells, the immobilized bead exhibited better degradation ability in wide range of pH, temperature and salinity. HPLC analysis showed that the mixed culture could convert aniline to catechol, and then catechol was further biodegraded to cis, cis-muconic acid. Ames test showed that the metabolic transformation products of aniline had no inherent toxicity. All these results showed that the mixed culture had a great potential to be used in the treatment of wastewater containing aniline.

Intestinal Microbiota in Large Yellow Croaker, Larimichthys crocea, at Different Ages

AbstractThe polymerase chain reaction–denaturing gradient gel electrophoresis (PCR‐DGGE) of 16S ribosomal RNA gene was used to investigate bacterial communities in the intestines of large yellow croaker at six different ages (12 d, 18 d, 26 d, 40 d, 3 mo, and 1 yr old) as well as within the corresponding feed and culture water. In addition, Illumina Miseq sequencing was utilized to compare intestinal microbiota between 12‐d‐old and 1‐yr‐old individuals. PCR‐DGGE results revealed that the culture water had the highest bacterial diversity, followed by the feed, while the intestines had the lowest diversity. The intestinal microbiota at six ages changed severely; however, the change did not follow any trend. The large yellow croaker intestines harbored specific bacterial communities that differed from those in both feed and water. Illumina Miseq sequencing results revealed that the diversity of intestinal bacteria in 12‐d‐old fish was higher than that in 1‐yr‐old fish, and the bacterial composition differed significantly between them. γ‐Proteobacteria and Pseudoalteromonas supplied the most abundant phylum and genus in the 12‐d‐old fish intestine. However, in the 1‐yr‐old fish intestine, Firmicutes and Clostridium were the most dominant, respectively. The study may contribute to a better understanding of gut microbiota and dynamics of the large yellow croaker and the relationship with their surrounding environment.

Effects of tourmaline on nitrogen removal performance and biofilm structures in the sequencing batch biofilm reactor

The effects of tourmaline on nitrogen removal performance and biofilm structures were comparatively investigated in two identical laboratory-scale sequencing batch biofilm reactors (SBBRs) (denoted SBBR1 and SBBR2) at different nitrogen loading rates (NLRs) varying from (0.24±0.01) to (1.26±0.02) g N/(L·day). SBBR1 was operated in parallel with SBBR2, but SBBR1 was filled with polyurethane foam loaded tourmaline (TPU) carriers and another (SBBR2) filled with polyurethane foam (PU) carriers. Results obtained from this study showed that the excellent and stable performance of SBBR1 was obtained. Ammonia nitrogen removal and total nitrogen removal were higher in SBBR1 than that in SBBR2 with increase of NLR. At an NLR of (0.24±0.01) g N/(L·day), the majority of the spherical and elliptical bacteria were surrounded by the extracellular polymeric substance (EPS) and bacillus or filamentous bacteria in two SBBRs biofilms. When NLR increased to (1.26±0.02) g N/(L·day), the clusters were more obvious in the SBBR1 biofilm than that in the SBBR2 biofilm. Bacteria in SBBR1 were inclined to synthesis more EPS, and the formed EPS could protect the bacteria from free ammonia (FA) under extreme condition NLR (1.26±0.02) g N/(L·day). The results of polymerase chain reaction-denaturing gradient gel electrophoresis analysis showed that the microbial community similarity in SBBR2 decreased more obviously than that in SBBR1 with the increase of NLR, which the microbial community in SBBR1 was relatively stable.

Microeukaryotic Community Structure and Dynamics During Macrobenthos Bioremediation in Intertidal Mudflat Sediments of Sansha Bay, China

Background: Microorganisms play important roles in the macrobenthic bioremediation process. At present, studies regarding microeukaryotes in intertidal sediments are not enough, and studies of the microeukaryotic dynamics in the bioremediation process using macrobenthos have received much less attention. Objectives: This study aimed to reveal the microeukaryotic community structure and dynamics during the bioremediation process using macrobenthos in the intertidal sediments of Sansha Bay of China. Methods: Twenty sediment samples were collected before and after bioremediation respectively. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) as well as 18S rDNA clone libraries analyses were used. Results: The microeukaryotic communitywere divided into 10 phyla, in which Annelida and Arthropoda were the most dominant, and Gastrotricha and Nematoda were the second dominant groups. A small amount of Cercozoa, Bacillariophyta, Chlorophyta, Alveolata, Amoebazoa, Chytridiomycota, and some unknown eukaryotes were also present. The microeukaryotic community structure presented a clear variation with time. In June, Annelida was the most dominant, and Arthropoda, Nematoda, and Cercozoa were the subdominant phyla. While in September, phylum Annelida decreased dramatically, and Arthropoda and Gastrotricha increased greatly. December harbored a quite different and highest diversity of microeukaryotic community: the phylum Annelida, which was dominant in June and September, was not detected, while the phyla Bacillariophyta, Nematoda, and Alveolata increased greatly, and Amoebozoa and Chytridiomycota were only detected in December. The bioremediation using macrobenthos changed the microeukartotic community structure and increased the diversity. The phyla Annelida and Arthropoda increased, while Gastrotricha, Cercozoa, and Alveolata decreased during the bioremediation. Conclusions: The microeukaryotic community in the intertidal sediment of Sansha Bay of China, were divided into 10 phyla, and presented a clear variation with time. The bioremediation using macrobenthos changed the microeukartotic community structure and increased the diversity.

Age-Related Variations in Intestinal Microflora of Free-Range and Caged Hens.

Free range feeding pattern puts the chicken in a mixture of growth materials and enteric bacteria excreted by nature, while it is typically unique condition materials and enteric bacteria in commercial caged hens production. Thus, the gastrointestinal microflora in two feeding patterns could be various. However, it remains poorly understood how feeding patterns affect development and composition of layer hens’ intestinal microflora. In this study, the effect of feeding patterns on the bacteria community in layer hens’ gut was investigated using free range and caged feeding form. Samples of whole small intestines and cecal digesta were collected from young hens (8-weeks) and mature laying hens (30-weeks). Based on analysis using polymerase chain reaction-denaturing gradient gel electrophoresis and sequencing of bacterial 16S rDNA gene amplicons, the microflora of all intestinal contents were affected by both feeding patterns and age of hens. Firmicutes, Bacteroidetes, Actinobacteria, Proteobacteria, and Fusobacteria were the main components. Additionally, uncultured environmental samples were found too. There were large differences between young hens and adult laying hens, the latter had more Firmicutes and Bacteroidetes, and bacterial community is more abundant in 30-weeks laying hens of all six phyla than 8-weeks young hens of only two phyla. In addition, the differences were also observed between free range and caged hens. Free range hens had richer Actinobacteria, Bacteroidetes, and Proteobacteria. Most of strains found were detected more abundant in small intestines than in cecum. Also the selected Lactic acid bacteria from hens gut were applied in feed and they had beneficial effects on growth performance and jejunal villus growth of young broilers. This study suggested that feeding patterns have an importance effect on the microflora composition of hens, which may impact the host nutritional status and intestinal health.

Impact of dissolved oxygen on the production of nitrous oxide in biological aerated filters

Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and microelectrode technology were employed to evaluate the Nitrous oxide (N2O) production in biological aerated filters (BAFs) under varied dissolved oxygen (DO) concentrations during treating wastewater under laboratory scale. The average yield of gasous N2O showed more than 4-fold increase when the DO levels were reduced from 6.0 to 2.0 mg∙L–1, indicating that low DO may drive N2O generation. PCR-DGGE results revealed that Nitratifractor salsuginis were dominant and may be responsible for N2O emission from the BAFs system. While at a low DO concentration (2.0 mg∙L–1), Flavobacterium urocaniciphilum might play a role. When DO concentration was the limiting factor (reduced from 6.0 to 2.0 mg∙L–1) for nitrification, it reduced NO 2 - -N oxidation as well as the total nitrification. The data from this study contribute to explain how N2O production changes in response to DO concentration, and may be helpful for reduction of N2O through regulation of DO levels.

Plant-fed versus chemicals-fed rhizobacteria of Lucerne: Plant-only teabags culture media not only increase culturability of rhizobacteria but also recover a previously uncultured Lysobacter sp., Novosphingobium sp. and Pedobacter sp.

In an effort to axenically culture the previously uncultivable populations of the rhizobacteria of Lucerne (Medicago sativa L.), we propose plant-only teabags culture media to mimic the nutritional matrix available in the rhizosphere. Here, we show that culture media prepared from Lucerne powder teabags substantially increased the cultivability of Lucerne rhizobacteria compared with a standard nutrient agar, where we found that the cultivable populations significantly increased by up to 60% of the total bacterial numbers as estimated by Quantitative Real-time Polymerase Chain Reaction (qRT-PCR). Cluster analysis of 16S rDNA Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) of cultivable Colony-Forming Units (CFUs) revealed a more distinct composition and separation of bacterial populations recovered on the plant-only teabags culture media than those developed on a standard nutrient agar. Further, the new plant medium gave preference to the micro-symbiont Sinorhizobium meliloti, and succeeded in isolating a number of not-yet-cultured bacteria, most closely matched to Novosphingobium sp., Lysobacter sp. and Pedobacter sp. The present study may encourage other researchers to consider moving from the well-established standard culture media to the challenging new plant-only culture media. Such a move may reveal previously hidden members of rhizobacteria, and help to further explore their potential environmental impacts.

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.

Analysis of Microbial Diversity and Greenhouse Gas Production of Decaying Pine Logs

In Sustainable Forest Management, decaying wood plays an important role in forest biodiversity, carbon balance and nutrient cycling. The management of this important component of forest ecosystems is limited by the fact that little is known about relationships between substrate quality and community structure of wood-inhabiting microorganisms. During decomposition, carbon stored in deadwood is lost either in the atmosphere or in the soil, but to our knowledge, limited information on the quantities of CO2 and other greenhouse gases (GHG) emitted is available. In the present research we investigated the correlation between the decay of logs, the decomposer microorganisms and their activities, in terms of GHG production and enzymes, in a black pine (Pinus nigra Arnold ssp. nigra) degraded forest. The decomposition of deadwood was visually assessed using a five-class system, and for each decay class four wood samples were collected. CO2, CH4 and N2O potential production from each decay class was measured in closed systems by means of gas chromatography. Enzyme activities related to carbon, nitrogen, sulphur and phosphorus cycling were measured fluorometrically. The composition of decomposer microbial communities (fungi, bacteria and actinobacteria) was assessed by using polymerase chain reaction-denaturing gradient gel electrophoresis fingerprinting. CO2 production and enzyme activities were significantly higher in the last decay classes of deadwood. The molecular approach highlighted differences in microbial community structure both at species and abundance levels, depending on the rate of decay.

Fermentation of hydrogen, 1,3-propanediol and ethanol from glycerol as affected by organic loading rate using up-flow anaerobic sludge blanket (UASB) reactor

The influences of organic loading rate (OLR) on the production of biohydrogen, 1,3-propanediol (1,3-PD) and ethanol from glycerol in an up-flow anaerobic sludge blanket (UASB) by anaerobic mixed cultures were investigated. UASB reactor was firstly operated using pure glycerol as the substrate at various OLRs of 25, 37.5, 50, 62.5 g/L d. Results showed that a change in OLR led to a change in hydrogen production, hydrogen production rate (HPR), hydrogen yield (HY), 1,3-PD and ethanol production as well as glycerol consumption, and microbial community. The optimal OLR for hydrogen and ethanol production was found to be 50 g/L d when the UASB was fed with pure glycerol, while the optimal OLR for 1,3-PD production was 62.5 g/L d. After the optimum OLR was obtained, the reactor was fed with crude glycerol. Maximum hydrogen, ethanol and 1,3-PD production from crude glycerol was obtained at the same OLR as of pure glycerol. A maximum hydrogen production, HY, ethanol and 1,3-PD production of 134.2 mmol/L, 579.8 mmol H2/mol glycerol, 78.9 mmol/L and 60.9 mmol/L, respectively, were obtained when pure glycerol was used as the substrate. However, when crude glycerol was used as the substrate, a maximum hydrogen production, HY, ethanol and 1,3-PD production were lower than that of using pure glycerol as the substrate. Analysis of microbial community by polymerase chain reaction-denaturing gradient gel electrophoresis indicated that the predominant hydrogen producers at the optimum OLR of 50 g/L d, when crude glycerol was used as the substrate, were Clostridium sp., Enterobacter sp., uncultured Firmicutes bacterium, and uncultured Actinobacterium while Enterobacter sp., Klebsiella sp., were the predominant 1,3-PD producers and Clostridium sp., as well as Enterobacter sp., were the predominant ethanol producers.

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.

Isolation of Viable but Non-culturable Bacteria from Printing and Dyeing Wastewater Bioreactor Based on Resuscitation Promoting Factor.

Printing and dyeing wastewater with high content of organic matters, high colority, and poor biochemical performance is hard to be degraded. In this study, we isolated viable but non-culturable (VBNC) bacteria from printing and dyeing wastewater with the culture media contained resuscitation promoting factor (Rpf) protein secreted by Micrococcus luteus, counted the culturable cells number with the most probable number, sequenced 16S rRNA genes, and performed polymerase chain reaction-denaturing gradient gel electrophoresis. It is obviously that the addition of Rpf in the enrichment culture could promote growth and resuscitation of bacteria in VBNC state to obtain more fastidious bacteria significantly. The identified bacteria were assigned to nine genera in the treatment group, while the two strains of Ochrobactrum anthropi and Microbacterium sp. could not be isolated from the control group. The function of isolated strains was explored and these strains could degrade the dye of Congo red. This study provides a new sight into the further study including the present state, composition, formation mechanism, and recovery mechanism about VBNC bacteria in printing and dyeing wastewater, which would promote to understand bacterial community in printing and dyeing wastewater, and to obtain VBNC bacteria from ecological environment.