Polymerase Chain Reaction-denaturing Gradient Gel Electrophoresis Analysis Research Articles

Removal of Hydrogen Sulfide from Swine-Waste Biogas on a Pilot Scale Using Immobilized Paracoccus versutus CM1.

Hydrogen sulfide (H2S) is a toxic and corrosive component that commonly occurs in biogas. In this study, H2S removal from swine-waste biogas using sulfur-oxidizing Paracoccus versutus CM1 immobilized in porous glass (PG) and polyurethane foam (PUF) biofilters was investigated. Bacterial compositions in the biofilters were also determined using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). The biofilters were first tested on a laboratory scale under three space velocities (SV): 20, 30, and 40 h-1. Within 24 h, at an SV of 20 h-1, PG and PUF biofilters immobilized with P. versutus CM1 removed 99.5% and 99.7% of H2S, respectively, corresponding to the elimination capacities (EC) of 83.5 and 86.2 gm-3 h-1. On a pilot scale, with the horizontal PG-P. versutus CM1 biofilter operated at an SV of 30 h-1, a removal efficiency of 99.7% and a maximum EC of 113.7 gm-3 h-1 were achieved. No reduction in methane content in the outlet biogas was observed under these conditions. The PCR-DGGE analysis revealed that Paracoccus, Acidithiobacillus, and Thiomonas were the predominant bacterial genera in the biofilters, which might play important roles in H2S removal. This PG-P. versutus CM1 biofiltration system is highly efficient for H2S removal from swine-waste biogas.

Mapping Archaeal Diversity in Soda Lakes by Coupling 16S rRNA PCR-DGGE Analysis with Remote Sensing and GIS Technology

The haloarchaeal diversity of four hypersaline alkaline lakes from the Wadi El-Natrun depression (Northern Egypt) was investigated using culture-independent polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) of 16S rRNA gene phylotypes, which was combined with remote sensing and geographic information system (GIS) data to highlight the distribution pattern of the microbial diversity in water and sediment samples. The majority of archaeal sequences identified in all four lakes belonged to the phyla Euryarchaeota and Crenarchaeota. Sediment samples from Beida Lake and water samples from El-Hamra Lake showed the highest levels of archaeal diversity. Sequence similarities ≥ 95% were found between six of the acquired clones and uncultured Halorhabdus, Euryarchaeota, and archaeon clones. In addition, two clones shared a high level of sequence similarity (97%) with unclassified archaea, while other nine clones exhibited 96% to 99% sequence similarity with uncultured archaeon clones, and only one clone showed 97% identity with an uncultured Crenarchaeota. Likewise, 7 DGGE bands presented a sequence similarity of 90 to 98% to Halogranum sp., Halalkalicoccus tibetensis, Halalkalicoccus jeotgali, uncultured Halorubrum, Halobacteriaceae sp., or uncultured haloarchaeon. In conclusion, while the variety of alkaliphilic haloarchaea in the examined soda lakes was restricted, the possibility of uncovering novel species for biotechnological applications from these extreme habitats remains promising.

PCR-DGGE Analysis Proves the Suppression of Rhizoctonia and Sclerotium Root Rot Due to Successive Inoculations

The soil-borne pathogens Rhizoctonia solani and Sclerotium rolfsii have emerged as major pathogens of radish (Raphanus sativus) worldwide. The induction of soil suppressive of radish root rot disease was evaluated in soil repeatedly inoculated with R. solani, nonpathogenic binucleate Rhizoctonia sp. AG-A W1 (BNR) and S. rolfsii. The repeated inoculations of soil with R. solani and BNR significantly suppressed the disease severity of R. solani and S. rolfsii compared to the control. In contrast, the repeated inoculation of soil with S. rolfsii significantly suppressed only the pathogen, S. rolfsii. The community structure was examined using PCR-DGGE (polymerase chain reaction denaturing gradient gel electrophoresis) method. The bands of Trichoderma sp. were observed in the first, second and third inoculations of the soil with BNR. Similarly, bands of Trichoderma sp. were observed in the second and third inoculations of the soil with S. rolfsii and R. solani. Compared to the control, disease severity was significantly reduced in the soil repeatedly inoculated with S. rolfsii and R. solani . In conclusion, Trichoderma species were accumulated in specific patterns depending on the applied fungal inoculum in the suppressive soil.

Inter- and Intra-Species Diversity of Lactic Acid Bacteria in Apis mellifera ligustica Colonies.

Lactic acid bacteria could positively affect the health of honey bees, including nutritional supplementation, immune system development and pathogen colonization resistance. Based on these considerations the present study evaluated predominant Lactic Acid Bacteria (LAB) species from beebread as well as from the social stomach and midgut of Apis mellifera ligustica honey bee foragers. In detail, for each compartment, the diversity in species and biotypes was ascertained through multiple culture-dependent approaches, consisting of Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE), 16S rRNA gene sequencing and Randomly Amplified Polymorphic DNA-Polymerase Chain Reaction (RAPD-PCR). The study of a lactic acid bacteria community, performed with PCR-DGGE and sequence analysis targeting the V1–V3 region of the 16S rRNA gene (rDNA), highlighted the presence of a few species, including Apilactobacillus kunkeei, Lactiplantibacillus plantarum, Fructobacillus fructosus, Levilactobacillus brevis and Lactobacillus delbrueckii subsp. lactis. Depending on the different compartments, diverse levels of biodiversity in species were found. Particularly, a very low inter-species biodiversity was detected in the midgut that was prevalently dominated by the presence of Apilactobacillus kunkeei. On the other hand, the beebread was characterized by a reasonable biodiversity showing the presence of five species and the predominance of Apilactobacillus kunkeei, Lactiplantibacillus plantarum and Fructobacillus fructosus. The RAPD-PCR analysis performed on the three predominant species allowed the differentiation into several biotypes for each species. Moreover, a relationship between biotypes and compartments has been detected and each biotype was able to express a specific biochemical profile. The biotypes that populated the social stomach and midgut were able to metabolize sugars considered toxic for bees while those isolated from beebread could contribute to release useful compounds with functional properties. Based on this knowledge, new biotechnological approaches could be developed to improve the health of honey bees and the quality of bee products.

Plant Species-Dependent Effects of Liming and Plant Residue Incorporation on Soil Bacterial Community and Activity in an Acidic Orchard Soil

Both liming and plant residue incorporation are widely used practices for the amelioration of acidic soils—however, the difference in their effects is still not fully understood, especially regarding the microbial community. In this study, we took the acidic soils from a subtropical orchard as target soils, and implemented liming and plant residue incorporation with a leguminous and a gramineous cover crop as test plants. After six months of growth, soil pH, total organic carbon (TOC), dissolved organic carbon (DOC) and nutrient contents were determined, soil enzymes involving C, N, P cycling were assayed, and microbial communities were also analyzed using Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE). Results showed that liming was more effective in elevating soil pH, while plant residue incorporation exerted a more comprehensive influence—not only on soil pH, but also on soil enzyme activity and microbial community. PCR-DGGE analysis revealed that liming changed the microbial community structure more greatly than plant residue incorporation, while plant residue incorporation altered the microbial community composition much more than liming. The growth responses of test plants to liming and plant residue incorporation depended on plant species, indicating the necessity to select appropriate practice for a particular crop. A further, detailed investigation into the microbial community composition, and the respective functions using metagenomic approach, is also suggested.

Assessing the Microbial Communities in Four Different Daqus by Using PCR-DGGE, PLFA, and Biolog Analyses.

Daqu made from raw wheat, barley or pea is used as an inoculum for the fermentation of Chinese Baijiu. In this study, the microbial communities of four different types of Daqus (sauce-flavor Wuling Daqu, sauce and strong-flavor Baisha Daqu, strong-flavor Deshan Daqu, and light-flavor Niulanshan Daqu) were analyzed by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), phospholipid fatty acid (PLFA) analysis, and Biolog EcoPlates analysis (Biolog). Clear differences were seen between the microbial communities of the four Daqus. PCR-DGGE showed differences in the number and brightness of bands between the Daqus, indicating the presence of unique bacterial species in Deshan Daqu, Wuling Daqu, and Niulanshan Daqu. Lactobacillus sanfranciscensis, Bacillus thermoamylovorans, and some unclassified bacteria were unique to Wuling Daqu, Deshan Daqu, and Niulanshan Daqu, respectively. Moreover, some bacterial species were observed in all four Daqus. A total of 26 PLFAs between C12 to C20 were detected from the four Daqus by PLFA analysis. Wuling Daqu had the highest total and fungal biomasses, Baisha Daqu had the highest bacterial biomass, and Niulanshan Daqu had the highest ratio of fungal biomass to bacterial biomass. The Biolog results indicated differences in the carbon source use and mode of the four Daqus, and also demonstrated that each Daqu had varying abilities to utilize different types of carbon sources. The cluster analysis of the three methods showed that the microbial communities of the four Daqus were different. This study also demonstrates the applicability of the three analytical methods in the evaluating of the microbial communities of Daqus.

Bacterial and Fungal Communities of Gioddu as Revealed by PCR-DGGE Analysis.

Gioddu is the sole variety of fermented milk originating in Italy. Despite the long history of consumption, Gioddu still represents an undisclosed source of microbial diversity. The present study was aimed to get an insight into the bacterial and fungal diversity of Gioddu samples collected from two artisan producers located in Sardinia. To this end 3 batches of Gioddu were collected from each producer and subjected to Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) analyses. Gioddu was produced with sheep milk in accordance with the local tradition. Regarding the bacterial population, a low biodiversity emerged. In more detail, the sole species Lactobacillus delbrueckii subsp. bulgaricus was detected in all the samples, irrespective of the producer or the batch. A more ample microbial diversity was highlighted for the fungal population that included closest relatives to Pichia cactophila, Kluyveromyces marxianus and Galactomyces candidum. Based on the results, the detected bacterial and fungal species generally clustered in accordance with the producer, irrespective of the batch considered. It is noteworthy that, Gioddu revealed several microbiological similarities with kefir beverage, thus suggesting, by analogy, potential health benefits related to its consumption. More research is needed to better clarify the microbiota composition of Gioddu by using more powerful metagenomic techniques.

Unveiling hákarl: A study of the microbiota of the traditional Icelandic fermented fish

Hákarl is produced by curing of the Greenland shark (Somniosus microcephalus) flesh, which before fermentation is toxic due to the high content of trimethylamine (TMA) or trimethylamine N-oxide (TMAO). Despite its long history of consumption, little knowledge is available on the microbial consortia involved in the fermentation of this fish. In the present study, a polyphasic approach based on both culturing and DNA-based techniques was adopted to gain insight into the microbial species present in ready-to-eat hákarl. To this aim, samples of ready-to-eat hákarl were subjected to viable counting on different selective growth media. The DNA directly extracted from the samples was further subjected to Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) and 16S amplicon-based sequencing. Moreover, the presence of Shiga toxin-producing Escherichia coli (STEC) and Pseudomonas aeruginosa was assessed via qualitative real-time PCR assays. pH values measured in the analyzed samples ranged from between 8.07 ± 0.06 and 8.76 ± 0.00. Viable counts revealed the presence of total mesophilic aerobes, lactic acid bacteria and Pseudomonadaceae. Regarding bacteria, PCR-DGGE analysis highlighted the dominance of close relatives of Tissierella creatinophila. For amplicon sequencing, the main operational taxonomic units (OTUs) shared among the data set were Tissierella, Pseudomonas, Oceanobacillus, Abyssivirga and Lactococcus. The presence of Pseudomonas in the analyzed samples supports the hypothesis of a possible role of this microorganism on the detoxification of shark meat from TMAO or TMA during fermentation. Several minor OTUs (<1%) were also detected, including Alkalibacterium, Staphylococcus, Proteiniclasticum, Acinetobacter, Erysipelothrix, Anaerobacillus, Ochrobactrum, Listeria and Photobacterium. Analysis of the yeast and filamentous fungi community composition by PCR-DGGE revealed the presence of close relatives of Candida tropicalis, Candida glabrata, Candida parapsilosis, Candida zeylanoides, Saccharomyces cerevisiae, Debaryomyces, Torulaspora, Yamadazyma, Sporobolomyces, Alternaria, Cladosporium tenuissimum, Moristroma quercinum and Phoma/Epicoccum, and some of these species probably play key roles in the development of the sensory qualities of the end product. Finally, qualitative real-time PCR assays revealed the absence of STEC and Pseudomonas aeruginosa in all of the analyzed samples.

Adhesion Behavior of Microorganisms Isolated from Soil on Hydroxyapatite and Other Materials.

The adhesion behavior of microorganisms on different materials was examined to obtain basic knowledge for designing support materials for microorganisms. The microorganisms were isolated from soil, and their adhesion behavior on hydroxyapatite (HA), carbon-coated HA (Carbon), poly (vinyl chloride) (PVC), and polyurethane (PU) pellets was investigated. The total metabolic activity on and adherence of microorganisms to the tested materials were in the following order: HA > Carbon > PVC > PU. This order was consistent with the extent of hydrophilicity of the materials. Morphological examination and polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE) analysis of microorganisms adhered to the materials revealed that the activities and states of microorganisms were affected by the composition of the pellets. PCR-DGGE analysis revealed various species of microorganisms adhered to the HA pellet. HA ceramics are expected to be one of the most suitable materials for supporting microorganisms.

Effects of C/N ratio on nitrous oxide production from nitrification in a laboratory-scale biological aerated filter reactor.

Emission of nitrous oxide (N2O) during biological wastewater treatment is of growing concern. This paper reports findings of the effects of carbon/nitrogen (C/N) ratio on N2O production rates in a laboratory-scale biological aerated filter (BAF) reactor, focusing on the biofilm during nitrification. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and microelectrode technology were utilized to evaluate the mechanisms associated with N2O production during wastewater treatment using BAF. Results indicated that the ability of N2O emission in biofilm at C/N ratio of 2 was much stronger than at C/N ratios of 5 and 8. PCR-DGGE analysis showed that the microbial community structures differed completely after the acclimatization at tested C/N ratios (i.e., 2, 5, and 8). Measurements of critical parameters including dissolved oxygen, oxidation reduction potential, NH4+-N, NO3--N, and NO2--N also demonstrated that the internal micro-environment of the biofilm benefit N2O production. DNA analysis showed that Proteobacteria comprised the majority of the bacteria, which might mainly result in N2O emission. Based on these results, C/N ratio is one of the parameters that play an important role in the N2O emission from the BAF reactors during nitrification.

The diversity and proteolytic properties of psychrotrophic bacteria in raw cows' milk from North China

Most psychrotrophic bacteria have the ability to produce thermoresistant proteases that can destroy the quality of milk and dairy products. To investigate the population dynamics of psychrotrophic bacteria during refrigeration, three raw cows' milk samples (sample A comprising milk from 10 farms in Beijing, sample B comprising milk from 5 farms in Heihe, and sample C comprising milk from 7 farms in Harbin) were refrigerated at 0–5 °C and 5–10 °C. PCR-DGGE (Polymerase chain reaction-denaturing gradient gel electrophoresis) analysis revealed that the bacterial community profiles varied from geographical site to site, and with refrigeration temperature. The dominant psychrotrophic bacteria among the samples after storage were affiliated with the order Pseudomonadales. Following isolation and identification, 8 psychrotrophic isolates were selected as stronger protease producers and their growth and proteolytic activities were assessed. The results indicate that the composition of psychrotrophic bacteria play an important role in the determination of the quality of milk and dairy products.

PCR-DGGE analysis of population dynamics of lactic acid bacteria in kimchi by addition of bacteriocins.

The effects of two crude bacteriocins (DF01 and K10) on lactic acid bacteria (LAB) communities and pH during kimchi fermentation were analyzed by polymerase chain reactiondenaturing gradient gel electrophoresis (PCR-DGGE). Crude LAB bacteriocins, prepared by ammonium sulfate precipitation, were added at 5 AU/mL, and kimchi was incubated at 20°C for 7 days. The pH and titratable acidity of the kimchies were determined daily, and the amplified 16S rRNA products were analyzed by PCR-DGGE. The common and main LAB were Weissella spp., Leuconostoc spp., and Lactobacillus spp. from both control and bacteriocin-treated samples. Among them, W. koreensis, W. confusa, and Lb. sakei were the predominant microorganisms throughout the fermentation period. Some obligate and facultatively heterofermentative LAB were detected from the bacteriocin-treated samples. The pH of the kimchi samples treated with each bacteriocin was higher (ca. 0.8 unit) than that of the control at day 4 and 5.

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.

ANAMMOX-like performances for nitrogen removal from ammonium-sulfate-rich wastewater in an anaerobic sequencing batch reactor

ABSTRACTAmmonium removal by the ANaerobic AMonium OXidation (ANAMMOX) process was observed through the Sulfate-Reducing Ammonium Oxidation (SRAO) process. The same concentration of ammonium (100 mg N L−1) was applied to two anaerobic sequencing batch reactors (AnSBRs) that were inoculated with the same activated sludge from the Vermicelli wastewater treatment process, while nitrite was fed in ANAMMOX and sulfate in SRAO reactors. In SRAO-AnSBR, in substrates that were fed with a ratio of NH4+/SO42− at 1:0.4 ± 0.03, a hydraulic retention time (HRT) of 48 h and without sludge draining, the Ammonium Removal Rate (ARR) was 0.02 ± 0.01 kg N m−3.d−1. Adding specific ANAMMOX substrates to SRAO-AnSBR sludge in batch tests results in specific ammonium and nitrite removal rates of 0.198 and 0.139 g N g−1 VSS.d, respectively, indicating that the ANAMMOX activity contributes to the removal of ammonium in the SRAO process using the nitrite that is produced from SRAO. Nevertheless, the inability of ANAMMOX to utilize sulfate to oxidize ammonium was also investigated in batch tests by augmenting enriched ANAMMOX culture in SRAO-AnSBR sludge and without nitrite supply. The time course of sulfate in a 24-hour cycle of SRAO-AnSBR showed an increase in sulfate after 6 h. For enriched SRAO culture, the uptake molar ratio of NH4+/SO42− at 8 hours in a batch test was 1:0.82 lower than the value of 1:0.20 ± 0.09 as obtained in an SRAO-AnSBR effluent, while the stoichiometric ratio of 1:0.5 that includes the ANAMMOX reaction was in this range. After a longer operation of more than 2 years without sludge draining, the accumulation of sulfate and the reduction of ammonium removal were observed, probably due to the gradual increase in the sulfur denitrification rate and the competitive use of nitrite with ANAMMOX. The 16S rRNA gene PCR-DGGE (polymerase chain reaction-denaturing gradient gel electrophoresis) and PCR cloning analyses resulted in the detection of the ANAMMOX bacterium (Candidatus Brocadia sinica JPN1) Desulfacinum subterraneum belonging to the genus Desulfacinum and bacteria that are involved in sulfur metabolism (Pseudomonas aeruginosa strain SBTPe-001 and Paracoccus denitrificans strain IAM12479) in SRAO-AnSBR.

Fungal Community Successions in Rhizosphere Sediment of Seagrasses Enhalus acoroides under PAHs Stress

Seagrass meadows represent one of the highest productive marine ecosystems and are of great ecological and economic values. Recently, they have been confronted with worldwide decline. Fungi play important roles in sustaining the ecosystem health as degraders of polycyclic aromatic hydrocarbons (PAHs), but fewer studies have been conducted in seagrass ecosystems. Hence, we investigated the dynamic variations of the fungal community succession under PAH stress in rhizosphere sediment of seagrasses Enhalus acoroides in this study. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), quantitative PCR (qPCR) and a clone library have been employed to analyze the fungal community’s shifts. Sequencing results of DGGE and the clone library showed that the predominant species belong to phyla Ascomycota and Basidiomycota. The abundance of three groups decreased sharply over the incubation period, whereas they demonstrated different fungal diversity patterns. Both the exposure time and the PAH concentrations affected the microbial diversity as assessed by PCR-DGGE analysis. Redundancy analysis (RDA) indicated that significant factors driving community shifts were ammonium and pH (p < 0.05). Significant amounts of the variations (31.1%) were explained by pH and ammonium, illustrating that those two parameters were the most likely ones to influence or be influenced by the fungal communities’ changes. Investigation results also indicated that fungal communities in seagrass meadow were very sensitive to PAH-induced stress and may be used as potential indicators for the PAH contamination.

Toxicological effects of dietary nickel chloride on intestinal microbiota

This study was designed to evaluate the toxicological effect of dietary nickel chloride (NiCl2) on the counts of intestinal bacteria and diversity of microorganisms in broilers. Plate counting and polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) assays were used. A total of 240 one-day-old avian broilers chicks were divided into four equal groups and kept on corn-soybean basal diet along with supplementation of 0, 300, 600 and 900mg/kg NiCl2 for 42 days. Samples were taken at 21 and 42 days of age during the experiment. The bacterial count results showed that dietary NiCl2 in the range of 300 to 900mg/kg decreased the counts of Bifidobacterium spp. and Lactobacillus, increased Escherichia coli (E. coli) and Enterococcus spp. in the ileum and cecum. PCR-DGGE analysis showed that bacterial band numbers, profile similarity, and the Shannon index of the ileum and cecum were all decreased in the 300, 600, and 900mg/kg groups at 21 and 42 days of age. In conclusion, dietary NiCl2 affected the amount and diversity of intestinal microbiota in the ileum and cecum of broilers. This finding implies that NiCl2 has toxicological effect on the intestinal ecosystem and, possibly functions.

Analysis of intestinal bacterial community diversity of adult Dastarcus helophoroides.

Polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE), and a culturedependent technique were used to study the diversity of the intestinal bacterial community in adult Dastarcus helophoroides (Fairmaire) (Coleoptera: Bothrideridae). Universal bacterial primers targeting 200 bp regions of the 16S rDNA gene were used in the PCR-DGGE assay, and 14 bright bands were obtained. The intestinal bacteria detected by PCR-DGGE were classified to Enterococcus (Lactobacillales: Enterococcaceae), Bacillus (Bacillales: Bacillaceae), Cellvibrio (Pseudomonadales: Pseudomonadaceae), Caulobacter (Caulobacterales: Caulobacteraceae), and uncultured bacteria, whereas those isolated by the culture-dependent technique belonged to Staphylococcus (Bacillales: Staphylococcaceae), Pectobacterium Enterobacteriales: Enterobacteriaceae), and Enterobacter (Enterobacteriales: Enterobacteriaceae). These intestinal bacteria represented the groups Lactobacillales (Enterococcus), Pseudomonadales (Cellvibrio), Caulobacterales (Caulobacter), Bacilli (Bacillus and Staphylococcus), and Gammaproteobacteria (Pectobacterium and Enterobacter). Our results demonstrated that PCR-DGGE analysis and the culture-dependent technique were useful in determining the intestinal bacteria of D. helophoroides and the two methods should be integrated to characterize the microbial community and diversity.

Microbial and metabolic characterization of a denitrifying phosphorus-uptake/side stream phosphorus removal system for treating domestic sewage.

In this study, an advanced wastewater treatment process, the denitrifying phosphorus/side stream phosphorus removal system (DPR-Phostrip), was developed for the purpose of enhancing denitrifying phosphorus removal. The enrichment of denitrifying phosphorus-accumulating organisms (DPAOs) and the microbial community structure of DPR-Phostrip were evaluated by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), and the metabolic activity of seed sludge and activated sludge collected after 55 days of operation were evaluated by Biolog™ analysis. This experimental study of DPR-Phostrip operation showed that nutrients were removed effectively, and denitrifying phosphorus removal was observed during the pre-anoxic period. PCR-DGGE analysis indicated that DPR-Phostrip supported DPAO growth while inhibiting PAOs and GAOs. The major dominant species in DPR-Phostrip were Bacteroidetes bacterium, Saprospiraceae bacterium, and Chloroflexi bacterium. Moreover, the functional diversity indices calculated on the basis of Biolog analysis indicated that DPR-Phostrip had almost no effect on microbial community diversity but was associated with a shift in the dominant species, which confirms the results of the PCR-DGGE analysis. The results for average well color development, calculated via Biolog analysis, showed that DPR-Phostrip had a little impact on the metabolic activity of sludge. Further principal component analysis suggested that the ability to utilize low-molecular-weight organic compounds was reduced in DPR-Phostrip.

Clinical signs and bacterial communities of deciduous necrotic root canals detected by PCR-DGGE analysis: Research association

ObjectiveThis study sought to investigate the possible association between clinical and radiographic data of the patients with the bacterial community profiles involved in cases of necrosis in primary root canals. MethodsMicrobial community profiles for 25 samples from necrotic deciduous root canals were analyzed using the polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) fingerprinting approach. These results were related to the clinical and radiographic data of these patients. ResultsThe analysis showed a large diversity of microbial communities in necrotic deciduous root canals. The statistical results pointed out that posterior and anterior teeth were associated with <20 bands and >20 bands in PCR-DGGE method, respectively. A relationship was verified between ages >4 years old and posterior teeth and, ages ≤4 years old and anterior teeth. ConclusionsThe data showed a polymicrobial community and pointed out the association of age with necrosis in anterior and posterior teeth.