Micro- and macrodiversity in rbcL sequences in ambient phytoplankton populations from the southeastern Gulf of Mexico

Ribulose-1,5-diphosphate carboxylase/oxygenase (RuBisCO) large subunit genes (rbcL) were obtained by amplification and cloning of 554 or 614 bp sequences of indigenous phytoplankton populations at 2 stations in the southeastern Gulf of Mexico. One station (Stn 4) was located in a low salinity, high chlorophyll plume (the ŒGreen River¹) which has previously been shown to contain elevated levels of Form IA rbcL mRNA while the other (Stn 7) was in oligotrophic, oceanic water. A diversity of rbcL sequences was obtained, spanning 3 of the 4 evolutionary clades of Form I RuBisCOs. Six nucleotide sequences obtained from Stn 4 were closely related (92 to 96% similar) to the Form IA-containing Prochlorococcus GP2. Flow cytometry and pigment analysis indicated that Prochlorococcus was abundant at this site. Other sequences found included a Form IB rbcL closely related to prasinophytes, and Form ID sequences related to prymnesiophytes, diatoms, and pelagophytes. One sequence was nearly identical to the pelagophyte, Pelagomonas calceolata. At Stn 7, sequences were obtained that were more deeply rooted, and less similar to rbcLs in existing databases (77 to 83% similar), and no Form IA rbcLs were detected. HPLC pigment signatures and flow cytometry data were consistent with the forms obtained by cloning. The similarity of the 6 Prochlorococcus GP2-like sequences (93 to 98%) is consistent with the phenomenon of molecular microdiversity as found at other loci in marine (and other environmental) microorganisms

Application of COMPOCHIP Microarray to Investigate the Bacterial Communities of Different Composts

A microarray spotted with 369 different 16S rRNA gene probes specific to microorganisms involved in the degradation process of organic waste during composting was developed. The microarray was tested with pure cultures, and of the 30,258 individual probe-target hybridization reactions performed, there were only 188 false positive (0.62%) and 22 false negative signals (0.07%). Labeled target DNA was prepared by polymerase chain reaction amplification of 16S rRNA genes using a Cy5-labeled universal bacterial forward primer and a universal reverse primer. The COMPOCHIP microarray was applied to three different compost types (green compost, manure mix compost, and anaerobic digestate compost) of different maturity (2, 8, and 16 weeks), and differences in the microorganisms in the three compost types and maturity stages were observed. Multivariate analysis showed that the bacterial composition of the three composts was different at the beginning of the composting process and became more similar upon maturation. Certain probes (targeting Sphingobacterium, Actinomyces, Xylella/Xanthomonas/ Stenotrophomonas, Microbacterium, Verrucomicrobia, Planctomycetes, Low G + C and Alphaproteobacteria) were more influential in discriminating between different composts. Results from denaturing gradient gel electrophoresis supported those of microarray analysis. This study showed that the COMPOCHIP array is a suitable tool to study bacterial communities in composts

Degradation of a benzene–toluene mixture by hydrocarbon-adapted bacterial communities

We examined the rate of degradation of a benzene–toluene mixture in aerobic microcosms prepared with samples of an aquifer that lies below a petrochemical plant (SIReN, UK). Five samples exposed to different concentrations of benzene (from 0.6 to 317 mg l−1) were used. Fast degradation (approx. 1–6 mg l−1 day−1) of both contaminants was observed in all groundwater samples and complete degradation was recorded by the seventh day except for one sample. We also identified the microbial community in each of the samples by culture-independent techniques. Two of the less impacted samples harbour the aerobic benzene degrader Pseudomonas fluorescens, while Acidovorax and Arthrobacter spp. were found in the most polluted sample and are consistent with the population observed in situ. Hydrogenophaga was found in the deepest sample while Rhodoferax spp. were recovered in an alkaline sample (pH 8.4) and may also be implicated in benzene degradation. Time series analysis shows that each of the samples has a different community but they remain stable over the degradation period. This study provides new information on a well not previously studied (no. 309s) and confirms that adapted communities have the ability to degrade hydrocarbon mixtures and could be used in further bioaugmentation approaches in contaminated sites

Measurement of residual stresses in surrogate coated nuclear fuel particles using ring-core focussed ion beam digital image correlation

Coated fuel particles, most commonly tri-structural isotropic (TRISO), are intended for application in several designs of advanced nuclear reactors. A complete understanding of the residual stresses and local properties of these particles through their entire lifecycle is required to inform fuel element manufacturing, reactor operation, accident scenarios, and reprocessing. However, there is very little experimental data available in the literature on the magnitude of residual stresses in the individual coating layers of these particles. This work applies ring-core focussed ion beam milling combined with digital image correlation analysis (FIB-DIC) to cross-sections of TRISO and pyrolytic carbon coatings in surrogate coated fuel particles to evaluate the residual stresses. Tensile residual hoop stresses are identified in both pyrolytic carbon layers, while silicon carbide experiences a compressive residual hoop stress. Note that these residual stresses, which were not accounted for in the models reported in open literature, have magnitudes comparable to the stresses predicted to arise in real fuel particles during service. A 2D linear-elastic continuum-based finite element analysis has been conducted to investigate the stress relaxation phenomena caused by sectioning stressed coatings on spherical particles. The FIB-DIC method established here is independent of radiation defects and can be applied to irradiated TRISO particles to retrieve first-hand information regarding the residual stress evolution during service

Comparative evaluation of prokaryotic 16S rDNA clone libraries and SSCP in groundwater samples.

A comparison of ribosomal RNA sequence analysis methods based on clone libraries and single-strand conformational polymorphism technique (SSCP) was performed with groundwater samples obtained between 523-555 meters below surface. The coverage of analyzed clones by phylotype-richness estimates was between 88-100%, confirming that the clone libraries were adequately examined. Analysis of individual bands retrieved from SSCP gels identified 1-6 different taxonomic units per band, suggesting that a single SSCP band does often represent more than one single prokaryotic species. The prokaryotic diversity obtained by both methods showed an overall difference of 42-80%. In comparison to SSCP, clone libraries underestimated the phylogenetic diversity and only 36-66% of the phylotypes observed with SSCP were also detected with the clone libraries. An exception was a sample where the SSCP analysis of Archaea identified only half of the phylotypes retrieved by the clone library. Overall, this study suggests that the clone library and the SSCP approach do not provide an identical picture of the prokaryotic diversity in groundwater samples. The results clearly show that the SSCP method, although this approach is prone to generate methodological artifacts, was able to detect significantly more phylotypes than microbial community analysis based on clone libraries