Denaturing Gradient Gel Electrophoresis Fingerprinting Research Articles

Dysbiosis of bifidobacteria and Clostridium cluster XIVa in the cystic fibrosis fecal microbiota

BackgroundRecurrent antimicrobial interventions and disease-related intestinal dysfunction are suspected to contribute to the dysbiosis of the gastrointestinal microbial ecosystem in patients with cystic fibrosis (CF). The present study set out to detect and identify microbial discriminants in the gut microbiota composition that are associated with CF-related intestinal dysbiosis. MethodsAn in-depth description of CF-associated gut dysbiosis was obtained by screening denaturing gradient gel electrophoresis (DGGE) fingerprints for potentially discriminating bacterial species, and quantification by means of real-time PCR analyses using group-specific primers. ResultsA total of 8 DGGE band-classes assigned to the genus Bifidobacterium (n=3), and members of Clostridium clusters XIVa (n=3) and IV (n=2), were significantly (p<0.05) underrepresented in samples of patients with CF. Real-time PCR analyses confirmed a significantly lower abundance and temporal stability of bifidobacteria and Clostridium cluster XIVa in the fecal microbiota of patients with CF. ConclusionThis study is the first to report specific microbial determinants of dysbiosis in patients with CF.

Comparing Metabolic Functionalities, Community Structures, and Dynamics of Herbicide-Degrading Communities Cultivated with Different Substrate Concentrations

Two 4-chloro-2-methylphenoxyacetic acid (MCPA)-degrading enrichment cultures selected from an aquifer on low (0.1 mg liter(-1)) or high (25 mg liter(-1)) MCPA concentrations were compared in terms of metabolic activity, community composition, population growth, and single cell physiology. Different community compositions and major shifts in community structure following exposure to different MCPA concentrations were observed using both 16S rRNA gene denaturing gradient gel electrophoresis fingerprinting and pyrosequencing. The communities also differed in their MCPA-mineralizing activities. The enrichments selected on low concentrations mineralized MCPA with shorter lag phases than those selected on high concentrations. Flow cytometry measurements revealed that mineralization led to cell growth. The presence of low-nucleic acid-content bacteria (LNA bacteria) was correlated with mineralization activity in cultures selected on low herbicide concentrations. This suggests that LNA bacteria may play a role in degradation of low herbicide concentrations in aquifers impacted by agriculture. This study shows that subpopulations of herbicide-degrading bacteria that are adapted to different pesticide concentrations can coexist in the same environment and that using a low herbicide concentration enables enrichment of apparently oligotrophic subpopulations.

Effects of metal oxide nanoparticles on soil properties

In recent years the behavior and properties of nanoparticles released to the environment have been studied extensively to better assess the potential consequences of their broad use in commercial products. The fate, transport and mobility of nanoparticles in soil were shown to be strongly dependent on environmental conditions. However, little is known about the possible effects of nanoparticles on soil chemical, physical and biological properties. In this study, two types of metal oxide nanoparticles, CuO and Fe3O4 were mixed into two types of soil and the effects of the nanoparticles on various soil properties were assessed. Metal oxide nanoparticles were shown previously to catalyze the oxidation of organic pollutants in aqueous suspensions, and they were therefore expected to induce changes in the organic material in the soil, especially upon addition of an oxidant. It was found that the nanoparticles did not change the total amount of organic materials in the soil or the total organic carbon in the soil extract; however, three-dimensional fluorescence spectroscopy demonstrated changes in humic substances. The nanoparticles also affected the soil bacterial community composition, based on denaturing gradient gel electrophoresis (DGGE) fingerprinting, but had little impact on the macroscopic properties of the soil.

Evaluation of apple replant problems based on different soil disinfection treatments—links to soil microbial community structure?

Replant problems or soil sickness are known phenomena but still unsolved. The aims of this study were (i) to set up a test system for detecting replant problems using in vitro propagated apple rootstocks (M26) based on different soil disinfection treatments and (ii) to explore the treatment effects on root morphology and soil microbial community structure. The bio-test involved soil with apple replant problems (apple sick) and healthy soil from an adjacent plot, both either untreated, or submitted to treatments of 50 and 100 °C, or the chemical soil disinfectant Basamid. Histological analyses of roots and denaturing gradient gel electrophoresis (DGGE) fingerprints in rhizosphere soil collected at the final evaluation were performed. After 10 weeks, shoot dry mass on apple sick soil was 79, 108 and 124 % higher for soil treated at 50 °C, 100 °C and with Basamid, respectively, compared to the untreated soil. Roots in untreated apple sick soil showed destroyed epidermal and cortical layers. DGGE fingerprints revealed treatment dependent differences in community composition and relative abundance of total bacteria, Bacillus, Pseudomonas and total fungi. The clear differences detected in soil microbial communities are the first steps towards a better understanding of the causes for apple replant problems.

Effect of susceptibility to enterotoxigenic Escherichia coli F4 and of dietary tryptophan on gut microbiota diversity observed in healthy young pigs

Healthy weaned pigs susceptible to enterotoxigenic Escherichia coli F4 (ETEC) require more tryptophan (Trp) to maximize their performance. This may be related to an effect on intestinal microbiota. We studied the intestinal bacterial diversity of healthy pigs with different susceptibility to ETEC and fed different Trp levels. Thirty-six littermate weaned pigs were selected to obtain a set potentially formed of 50% ETEC-susceptible and 50% non-susceptible pigs, based on a Mucin 4 gene polymorphism. Pigs were fed a diet with 0.17 (TrpL) or 0.22 (TrpH) standardized ileal digestible Trp:Lys ratio for 21 days. Slaughtered pigs were classified into non-susceptible, mildly susceptible, and susceptible, by testing ETEC adhesion to intestinal villi. Bacterial diversity in jejunum content was assessed by the 16S rRNA gene-targeted denaturing gradient gel electrophoresis (DGGE) fingerprinting analysis and expressed by the Shannon index. Susceptible pigs had a reduced bacterial diversity, particularly with TrpL diet (p=0.003). The ETEC adhesion class affected the quantification of enterobacteria DNA (p=0.027). One DGGE band, which referred to Clostridium bartlettii, was not evidenced in all the susceptible pigs; less DNA from this microbe was quantified by RT-PCR in the jejunum from TrpH susceptible pigs (p=0.025) compared to TrpL. The gene expression for β-galactoside α-2,3-sialyltransferase 1 was higher in jejunal tissue of ETEC-susceptible pigs (p=0.019). In studies on pig gut microbiota, the presence of intestinal receptors for ETEC should be considered because of their contribution to a reduced bacterial diversity. This effect could be partially reversed by dietary Trp addition.

Spatial variability of cyanobacterial community composition in Sanya Bay as determined by DGGE fingerprinting and multivariate analysis

The cyanobacterial communities in the surface and bottom waters of Sanya Bay were investigated on April 24 and 25, 2010. Flow cytometry showed that the total cyanobacterial abundance in the surface and bottom layers ranged from 0.7×104 to 2.38×104 cells mL−1 and from 1×104 to 1.8×104 cells mL−1, respectively. Cyanobacterial diversity was analyzed using a molecular fingerprinting technique called denaturing gradient gel electrophoresis (DGGE), followed by DNA sequencing. The results were then interpreted through multivariate statistical analysis. Differences in the compositions of cyanobacterial communities were observed in the surface and bottom waters at the same station, with some bands obtained from both the surface and bottom layers, whereas some bands were present only in one layer. The predominant cyanobacterial species of the excised DGGE bands were related to Synechococcus or Synechococcus-like species (56.2%). Other phylogenetic groups identified included Chroococcidiopsis (6.3%), Cyanobium (6.3%) and some unclassified cyanobacteria (31.2%). A redundancy analysis (RDA) was conducted to reveal the relationships between the cyanobacterial community composition and environmental factors. Analysis results showed that the spatial variations in the cyanobacterial community composition in surface waters was significantly related to chlorophyll a (Chla), the biochemical oxygen demand (BOD), nitrate and phosphate (P<0.05). Meanwhile, the spatial variations in the bottom waters was significantly affected by nitrate, nitrite, and phosphate (P<0.05). Environmental parameters could explain 99.3% and 58.3% of the variations in the surface and bottom layers, respectively.

Bacterial Associates of Two Caribbean Coral Species Reveal Species-Specific Distribution and Geographic Variability

Scleractinian corals harbor microorganisms that form dynamic associations with the coral host and exhibit substantial genetic and ecological diversity. Microbial associates may provide defense against pathogens and serve as bioindicators of changing environmental conditions. Here we describe the bacterial assemblages associated with two of the most common and phylogenetically divergent reef-building corals in the Caribbean, Montastraea faveolata and Porites astreoides. Contrasting life history strategies and disease susceptibilities indicate potential differences in their microbiota and immune function that may in part drive changes in the composition of coral reef communities. The ribotype structure and diversity of coral-associated bacteria within the surface mucosal layer (SML) of healthy corals were assessed using denaturing gradient gel electrophoresis (DGGE) fingerprinting and 454 bar-coded pyrosequencing. Corals were sampled at disparate Caribbean locations representing various levels of anthropogenic impact. We demonstrate here that M. faveolata and P. astreoides harbor distinct, host-specific bacteria but that specificity varies by species and site. P. astreoides generally hosts a bacterial assemblage of low diversity that is largely dominated by one bacterial genus, Endozoicomonas, within the order Oceanospirillales. The bacterial assemblages associated with M. faveolata are significantly more diverse and exhibit higher specificity at the family level than P. astreoides assemblages. Both corals have more bacterial diversity and higher abundances of disease-related bacteria at sites closer to the mainland than at those furthest away. The most diverse bacterial taxa and highest relative abundance of disease-associated bacteria were seen for corals near St. Thomas, U.S. Virgin Islands (USVI) (2.5 km from shore), and the least diverse taxa and lowest relative abundance were seen for corals near our most pristine site in Belize (20 km from shore). We conclude that the two coral species studied harbor distinct bacterial assemblages within the SML, but the degree to which each species maintains specific microbial associations varies both within each site and across large spatial scales. The taxonomic scale (i.e., phylum versus genus) at which scientists examine coral-microbe associations, in addition to host-elicited factors and environmental fluctuations, must be considered carefully in future studies of the coral holobiont.

Comparative Analysis of Bacterial Diversity for Cutlassfish (&lt;i&gt;Trichiurus haumela&lt;/i&gt;) during Cold Storage by the Culture-Dependent and Culture-Independent Methods

The methods of culture-dependent and denaturing gradient gel electrophoresis (DGGE) based on the sequence of 16S rRNA V3 region gene were described to comparatively characterize the microbial population and community structure of cutlassfish (Trichiurus haumela) under the cold storage. The results showed that 13 kinds of bacteria were identified by the traditional culture-dependent methods, the dominant bacteria belonged to Shewanella putrefaciens and Pseudomonas fluorescens. To determine the community profiles of the samples on variable V3 region, the bacteria of 16S rRNA gene were amplified by PCR and 11 distinct PCR products were separated by DGGE fingerprinting technology. From the sequence analysis, Psychrobacter sp. was found to be the predominant bacteria in the initial stage of the storage. The proportion of Shewanella sp., Pseudomonas sp. increased gradually with the extension of storage time, and they took the place of Psychrobacter sp. to be the dominant bacteria. Thereinto, both Pseudomonas fluorescens and Vibrio sp. took high proportions in the process of storage due to the deterioration of cutlassfish (Trichiurus haumela).

Multitrophic interaction in the rhizosphere of maize: root feeding of Western corn rootworm larvae alters the microbial community composition.

BackgroundLarvae of the Western Corn Rootworm (WCR) feeding on maize roots cause heavy economical losses in the US and in Europe. New or adapted pest management strategies urgently require a better understanding of the multitrophic interaction in the rhizosphere. This study aimed to investigate the effect of WCR root feeding on the microbial communities colonizing the maize rhizosphere.Methodology/Principal FindingsIn a greenhouse experiment, maize lines KWS13, KWS14, KWS15 and MON88017 were grown in three different soil types in presence and in absence of WCR larvae. Bacterial and fungal community structures were analyzed by denaturing gradient gel electrophoresis (DGGE) of the16S rRNA gene and ITS fragments, PCR amplified from the total rhizosphere community DNA. DGGE bands with increased intensity were excised from the gel, cloned and sequenced in order to identify specific bacteria responding to WCR larval feeding. DGGE fingerprints showed that the soil type and the maize line influenced the fungal and bacterial communities inhabiting the maize rhizosphere. WCR larval feeding affected the rhiyosphere microbial populations in a soil type and maize line dependent manner. DGGE band sequencing revealed an increased abundance of Acinetobacter calcoaceticus in the rhizosphere of several maize lines in all soil types upon WCR larval feeding.Conclusion/SignificanceThe effects of both rhizosphere and WCR larval feeding seemed to be stronger on bacterial communities than on fungi. Bacterial and fungal community shifts in response to larval feeding were most likely due to changes of root exudation patterns. The increased abundance of A. calcoaceticus suggested that phenolic compounds were released upon WCR wounding.

Nitrification of archaeal ammonia oxidizers in acid soils is supported by hydrolysis of urea

The hydrolysis of urea as a source of ammonia has been proposed as a mechanism for the nitrification of ammonia-oxidizing bacteria (AOB) in acidic soil. The growth of Nitrososphaera viennensis on urea suggests that the ureolysis of ammonia-oxidizing archaea (AOA) might occur in natural environments. In this study, (15)N isotope tracing indicates that ammonia oxidation occurred upon the addition of urea at a concentration similar to the in situ ammonium content of tea orchard soil (pH 3.75) and forest soil (pH 5.4) and was inhibited by acetylene. Nitrification activity was significantly stimulated by urea fertilization and coupled well with abundance changes in archaeal amoA genes in acidic soils. Pyrosequencing of 16S rRNA genes at whole microbial community level demonstrates the active growth of AOA in urea-amended soils. Molecular fingerprinting further shows that changes in denaturing gradient gel electrophoresis fingerprint patterns of archaeal amoA genes are paralleled by nitrification activity changes. However, bacterial amoA and 16S rRNA genes of AOB were not detected. The results strongly suggest that archaeal ammonia oxidation is supported by hydrolysis of urea and that AOA, from the marine Group 1.1a-associated lineage, dominate nitrification in two acidic soils tested.

A Proposed Taxonomy of Anaerobic Fungi (Class Neocallimastigomycetes) Suitable for Large-Scale Sequence-Based Community Structure Analysis

Anaerobic fungi are key players in the breakdown of fibrous plant material in the rumen, but not much is known about the composition and stability of fungal communities in ruminants. We analyzed anaerobic fungi in 53 rumen samples from farmed sheep (4 different flocks), cattle, and deer feeding on a variety of diets. Denaturing gradient gel electrophoresis fingerprinting of the internal transcribed spacer 1 (ITS1) region of the rrn operon revealed a high diversity of anaerobic fungal phylotypes across all samples. Clone libraries of the ITS1 region were constructed from DNA from 11 rumen samples that had distinctly different fungal communities. A total of 417 new sequences were generated to expand the number and diversity of ITS1 sequences available. Major phylogenetic groups of anaerobic fungi in New Zealand ruminants belonged to the genera Piromyces, Neocallimastix, Caecomyces and Orpinomyces. In addition, sequences forming four novel clades were obtained, which may represent so far undetected genera or species of anaerobic fungi. We propose a revised phylogeny and pragmatic taxonomy for anaerobic fungi, which was tested and proved suitable for analysis of datasets stemming from high-throughput next-generation sequencing methods. Comparing our revised taxonomy to the taxonomic assignment of sequences deposited in the GenBank database, we believe that >29% of ITS1 sequences derived from anaerobic fungal isolates or clones are misnamed at the genus level.

Effects of two different soil amendments on the biocontrol efficacy of biological control agents (BCA) against Ralstonia wilt on ginger

In this research, we developed an integrated approach to control Ralstonia wilt on ginger. Two organic composts (maize powder and soybean residue) were combined with one combination (the aqueous solution of Bacillus subtilis and Bacillus cereus ) and two single ( B. subtilis 1JN2 and Bacillus megaterium B1301) biological control agents (BCA) which together showed significant biocontrol efficacies in our previous study. Biocontrol efficacy of each treatment was measured. According to the results, a 3 - 30% biocontrol efficacy improvement was obtained by addition of the two organic composts compared to the BCA treatment. The best biocontrol activity (73.7%) was obtained from the combination of B. megaterium B1301 and maize powder. Colonization ability and the impacts on the bacterial community structure of each BCA were determined by denaturing gradient gel electrophoresis (DGGE). The DGGE fingerprints gave us an insight into the rhizosphere of the plant. And there is no noticeable bacterial structure shift, which means the exotic BCAs by artificial inoculation have little impacts on the local organism community. Additionally, the colonization ability of the tested BCAs was excellent and they could be detected at 120 days post inoculation. Furthermore, the available N, P, K and organic matter of rhizosphere soil from each treatment was measured. The organic matter did not show significant change among each treatment, while maize powder, soybean residue and BCA 1JN2 enhanced the available N, P and K significantly ranging from 79 - 133%, 166 - 251% and 252 - 368%, respectively compared to the blank control. Key words: Integrated management, biological control agents (BCA), organic compost, ginger wilt.

A protective coat of microorganisms on macroalgae: inhibitory effects of bacterial biofilms and epibiotic microbial assemblages on barnacle attachment.

Effects of epibiotic bacteria associated with macroalgae on barnacle larval attachment were investigated. Eight bacterial isolates obtained from samples of three macroalga species were cultured as monospecies bacterial films and tested for their activity against barnacle (Amphibalanus improvisus) attachment in field experiments (Western Baltic Sea). Furthermore, natural biofilm communities associated with the surface of the local brown alga, Fucus vesiculosus, which were exposed to different temperatures (5, 15 and 20°C), were harvested and subsequently tested. Generally, monospecies bacterial biofilms, as well as natural microbial assemblages, inhibited barnacle attachment by 20-67%. denaturing gradient gel electrophoresis fingerprints showed that temperature treatment shifted the bacterial community composition and weakened the repellent effects at 20°C. Repellent effects were absent when settlement pressure of cyprids was high. Nonviable bacteria tended to repel cyprids when compared to the unfilmed surfaces. We conclude that biofilms can have a repellent effect benefiting the host by preventing heavy fouling on its surface. However, severe settlement pressure, as well as stressful temperature, may reduce the protective effects of the alga's biofilm. Our results add to the notion that the performance of F.vesiculosus may be reduced by multiple stressors in the course of global warming.

The effect of maize silage as co-substrate for swine manure on the bacterial community structure in biogas plants

The qualitative and quantitative changes in the bacterial community composition in two mesophilic, commercially used biogas plants were monitored by denaturing gradient gel electrophoresis (DGGE) and real-time PCR. The main objective was to evaluate the influence of the co-substrate maize silage on total bacteria and some selected bacterial groups by comparing full-scale reactors fed solely with pig manure or additionally with maize silage. DGGE fingerprints reflected shifts in the bacterial community structure associated with maize silage as co-substrate and the real-time PCR results showed clear changes in the quantitative composition of the bacterial consortia of each fermenter. A clear dominance of Clostridia in all surveyed fermenters and considerably lower abundance of Bacteroidetes in the biogas plant fed with maize silage was shown.

Benthic bacterial diversity from freshwater tufas of the Iberian Range (Spain)

Aiming to characterize the bacterial diversity of modern tufa systems of the Iberian Range (Spain), we surveyed the 16S rRNA gene sequence diversity from 24 sites within three rivers (Añamaza, Mesa and Piedra). These tufas record substantial calcareous growth under different physicochemical conditions and are part of an important, regional landscape-building system. The bacterial community structure and composition, richness and diversity were quantified from denaturing gradient gel electrophoresis fingerprints. Retrieved DNA sequences could be assigned to 10 bacterial phyla and included a variety of phototrophic and heterotrophic groups. Cyanobacteria, mainly filamentous taxa, constituted 43% of all the retrieved sequences, followed by Firmicutes (11%), Gammaproteobacteria (10%), Alphaproteobacteria (7%), Acidobacteria (6%), Bacteroidetes (5%), Betaproteobacteria (4%), Planctomycetes (4%), Actinobacteria (3%) and Deltaproteobacteria (2%). Diatom and Xanthophyceae chloroplast sequences were also detected. Physicochemical variables measured at each site were modelled with multivariate statistics. Principal component analyses yielded the highest variance for salinity-related variables (conductivity; Na(+) , Cl(-) and SO4(2-) concentrations), which correlated negatively and significantly with diversity indices. However, the highest variance explained by individual principal components was relatively low (< 34%). Overall, we show that these young fluvial tufas are inhabited by a large variety of bacteria in diverse and widespread communities.

Optimal cultivation and characteristics of aerobic granules with typical domestic sewage in an alternating anaerobic/aerobic sequencing batch reactor

The present work investigated some important factors for optimal aerobic granulation using typical domestic sewage as a substrate in a pilot-scale alternating anaerobic/aerobic sequencing batch reactor. High sludge concentration and low sludge loading, for the first time, were used for the reactor start-up. A vast number of small particles appeared on day8. Subsequently, several measures for controlling sludge concentration and sludge loading within an appropriate range were applied to optimize the granulation process. On day 45, complete sludge granulation was achieved. After 60days of operation, the aerobic granules always kept in stable state, with an average diameter of 750μm and the SVI30 of 20–35ml/g. The COD, TN, and TP removal ratios were 92%, 81%, and 85%, respectively. The results demonstrated that it was feasible to form aerobic granules quickly using typical domestic sewage under optimal operation strategies, which was further proved by the results from denaturing gradient gel electrophoresis fingerprint.

Effects of disodium fumarate on ruminal fermentation and microbial communities in sheep fed on high-forage diets

This study was conducted to investigate effects of disodium fumarate (DF) on fermentation characteristics and microbial populations in the rumen of Hu sheep fed on high-forage diets. Two complementary feeding trials were conducted. In Trial 1, six Hu sheep fitted with ruminal cannulae were randomly allocated to a 2 × 2 cross-over design involving dietary treatments of either 0 or 20 g DF daily. Total DNA was extracted from the fluid- and solid-associated rumen microbes, respectively. Numbers of 16S rDNA gene copies associated with rumen methanogens and bacteria, and 18S rDNA gene copies associated with rumen protozoa and fungi were measured using real-time PCR, and expressed as proportion of total rumen bacteria 16S rDNA. Ruminal pH decreased in the DF group compared with the control (P < 0.05). Total volatile fatty acids increased (P < 0.001), but butyrate decreased (P < 0.01). Addition of DF inhibited the growth of methanogens, protozoa, fungi and Ruminococcus flavefaciens in fluid samples. Both Ruminococcus albus and Butyrivibrio fibrisolvens populations increased (P < 0.001) in particle-associated samples. Trial 2 was conducted to investigate the adaptive response of rumen microbes to DF. Three cannulated sheep were fed on basal diet for 2 weeks and continuously for 4 weeks with supplementation of DF at a level of 20 g/day. Ruminal samples were collected every week to analyze fermentation parameters and microbial populations. No effects of DF were observed on pH, acetate and butyrate (P > 0.05). Populations of methanogens and R. flavefaciens decreased in the fluid samples (P < 0.001), whereas addition of DF stimulated the population of solid-associated Fibrobacter succinogenes. Population of R. albus increased in the 2nd to 4th week in fluid-associated samples and was threefold higher in the 4th week than control week in solid samples. Analysis of denaturing gradient gel electrophoresis fingerprints revealed that there were significant changes in rumen microbiota after adding DF. Ten of 15 clone sequences from cut-out bands appearing in both the 2nd and the 4th week were 94% to 100% similar to Prevotella-like bacteria, and four sequences showed 95% to 98% similarity to Selenomonas dianae. Another 15 sequences were obtained from bands, which appeared in the 4th week only. Thirteen of these 15 sequences showed 95% to 99% similarity to Clostridium sp., and the other two showed 95% and 100% similarity to Ruminococcus sp. In summary, the microorganisms positively responding to DF addition were the cellulolytic bacteria, R. albus, F. succinogenes and B. fibrisolvens as well as proteolytic bacteria, B. fibrisolvens, P. ruminicola and Clostridium sp.

Inhibition of Bacterial Ammonia Oxidation by Organohydrazines in Soil Microcosms

Hydroxylamine oxidation by hydroxylamine oxidoreductase (HAO) is a key step for energy-yielding in support of the growth of ammonia-oxidizing bacteria (AOB). Organohydrazines have been shown to inactivate HAO from Nitrosomonas europaea, and may serve as selective inhibitors to differentiate bacterial from archaeal ammonia oxidation due to the absence of bacterial HAO gene homolog in known ammonia-oxidizing archaea (AOA). In this study, the effects of three organohydrazines on activity, abundance, and composition of AOB and AOA were evaluated in soil microcosms. The results indicate that phenylhydrazine and methylhydrazine at the concentration of 100 μmol g−1 dry weight soil completely suppressed the activity of soil nitrification. Denaturing gradient gel electrophoresis fingerprinting and sequencing analysis of bacterial ammonia monooxygenase subunit A gene (amoA) clearly demonstrated that nitrification activity change is well paralleled with the growth of Nitrosomonas europaea-like AOB in soil microcosms. No significant correlation between AOA community structure and nitrification activity was observed among all treatments during the incubation period, although incomplete inhibition of nitrification activity occurred in 2-hydroxyethylhydrazine-amended soil microcosms. These findings show that the HAO-targeted organohydrazines can effectively inhibit bacterial nitrification in soil, and the mechanism of organohydrazine affecting AOA remains unclear.

Specificity of Salt Marsh Diazotrophs for Vegetation Zones and Plant Hosts: Results from a North American marsh

Salt marshes located on the east coast of temperate North America are highly productive, typically nitrogen-limited, and support diverse assemblages of free-living nitrogen fixing (diazotrophic) bacteria. This article reviews and analyzes data from North Inlet estuary (SC, USA), addressing diazotroph assemblage structure and the influence of plant host and environmental conditions on the assemblage. The North Inlet estuary is a salt marsh ecosystem in which anthropogenic influences are minimal and the distributions of diazotrophs are governed by the natural biota and dynamics of the system. Denaturing gradient gel electrophoresis fingerprinting and phylogenetic analyses of recovered sequences demonstrated that the distributions of some diazotrophs reflect plant host specificity and that diazotroph assemblages distributed across marsh gradients are also heavily influenced by edaphic conditions. Broadly distributed diazotrophs that are capable of maintaining populations under all environmental conditions spanning such gradients are also present in these assemblages. Statistical analyses indicate that the structures of diazotroph assemblages in different vegetation zones are significantly (p < 0.01) different. New data presented here demonstrate the heterogeneity of salt marsh rhizosphere microenvironments, and corroborate previous findings from different plant hosts growing at several locations within this estuary. The data from these collected works support the hypothesis that the biogeography of microorganisms is non-random and these biogeographic patterns are predictable.

Changes in Human Fecal Microbiota Due to Chemotherapy Analyzed by TaqMan-PCR, 454 Sequencing and PCR-DGGE Fingerprinting

BackgroundWe investigated whether chemotherapy with the presence or absence of antibiotics against different kinds of cancer changed the gastrointestinal microbiota.Methodology/Principal FindingsFeces of 17 ambulant patients receiving chemotherapy with or without concomitant antibiotics were analyzed before and after the chemotherapy cycle at four time points in comparison to 17 gender-, age- and lifestyle-matched healthy controls. We targeted 16S rRNA genes of all bacteria, Bacteroides, bifidobacteria, Clostridium cluster IV and XIVa as well as C. difficile with TaqMan qPCR, denaturing gradient gel electrophoresis (DGGE) fingerprinting and high-throughput sequencing. After a significant drop in the abundance of microbiota (p = 0.037) following a single treatment the microbiota recovered within a few days. The chemotherapeutical treatment marginally affected the Bacteroides while the Clostridium cluster IV and XIVa were significantly more sensitive to chemotherapy and antibiotic treatment. DGGE fingerprinting showed decreased diversity of Clostridium cluster IV and XIVa in response to chemotherapy with cluster IV diversity being particularly affected by antibiotics. The occurrence of C. difficile in three out of seventeen subjects was accompanied by a decrease in the genera Bifidobacterium, Lactobacillus, Veillonella and Faecalibacterium prausnitzii. Enterococcus faecium increased following chemotherapy.Conclusions/SignificanceDespite high individual variations, these results suggest that the observed changes in the human gut microbiota may favor colonization with C.difficile and Enterococcus faecium. Perturbed microbiota may be a target for specific mitigation with safe pre- and probiotics.