The active microbial community more accurately reflects the anaerobic digestion process: 16S rRNA (gene) sequencing as a predictive tool

Background: Amplicon sequencing methods targeting the 16S rRNA gene have been used extensively to investigate microbial community composition and dynamics in anaerobic digestion. These methods successfully characterize amplicons but do not distinguish micro-organisms that are actually responsible for the process. In this research, the archaeal and bacterial community of 48 full-scale anaerobic digestion plants were evaluated on DNA (total community) and RNA (active community) level via 16S rRNA (gene) amplicon sequencing. Results: A significantly higher diversity on DNA compared with the RNA level was observed for archaea, but not for bacteria. Beta diversity analysis showed a significant difference in community composition between the DNA and RNA of both bacteria and archaea. This related with 25.5 and 42.3% of total OTUs for bacteria and archaea, respectively, that showed a significant difference in their DNA and RNA profiles. Similar operational parameters affected the bacterial and archaeal community, yet the differentiating effect between DNA and RNA was much stronger for archaea. Co-occurrence networks and functional prediction profiling confirmed the clear differentiation between DNA and RNA profiles. Conclusions: In conclusion, a clear difference in active (RNA) and total (DNA) community profiles was observed, implying the need for a combined approach to estimate community stability in anaerobic digestion

Metagenomic sequencing unravels gene fragments with phylogenetic signatures of O2-tolerant NiFe membrane-bound hydrogenases in lacustrine sediment

Many promising hydrogen technologies utilising hydrogenase enzymes have been slowed by the fact that most hydrogenases are extremely sensitive to O2. Within the group 1 membrane-bound NiFe hydrogenase, naturally occurring tolerant enzymes do exist, and O2 tolerance has been largely attributed to changes in iron–sulphur clusters coordinated by different numbers of cysteine residues in the enzyme’s small subunit. Indeed, previous work has provided a robust phylogenetic signature of O2 tolerance [1], which when combined with new sequencing technologies makes bio prospecting in nature a far more viable endeavour. However, making sense of such a vast diversity is still challenging and could be simplified if known species with O2-tolerant enzymes were annotated with information on metabolism and natural environments. Here, we utilised a bioinformatics approach to compare O2-tolerant and sensitive membrane-bound NiFe hydrogenases from 177 bacterial species with fully sequenced genomes for differences in their taxonomy, O2 requirements, and natural environment. Following this, we interrogated a metagenome from lacustrine surface sediment for novel hydrogenases via high-throughput shotgun DNA sequencing using the Illumina™ MiSeq platform. We found 44 new NiFe group 1 membrane-bound hydrogenase sequence fragments, five of which segregated with the tolerant group on the phylogenetic tree of the enzyme’s small subunit, and four with the large subunit, indicating de novo O2-tolerant protein sequences that could help engineer more efficient hydrogenases

Weathering of the Ethiopian volcanic province: a new weathering index to characterize and compare soils

© Walter de Gruyter Berlin/Boston 2015.Soil formation occurs through numerous physical and chemical weathering processes acting to alter the parent rock on the Earths surface. Samples of surface soils were collected over a range of elevations (2000-3600 m) from profiles directly overlying basaltic to more felsic parent rocks, over a region in NW Ethiopia. The soils were investigated to determine their chemical composition and X-ray diffraction was used to identify and quantify individual mineral phases. The data set was analyzed using non-parametric statistics (Spearmans Rank and Mann-Whitney U tests) to compare the soils forming over the two parent rocks. Principal component analysis (PCA) was used to identify the mineral alteration assemblage and formation during pedogenesis. The extent of alteration was quantified using several chemical weathering indices (Chemical Index of Alteration = CIA; Chemical Index of Weathering = CIW), including an index calculated by multivariate analyses of the soil chemical composition data (weathering W index). Further to this we devised and tested a new weathering index (Wmin) using multivariate analysis of the soil mineralogy, to estimate the extent of weathering and physico-chemical proprieties of the parent rock from which the soil formed. The soils present a fair to advanced stage of alteration, with abundant iron (Fe) oxides (up to 40 wt%) and phyllosilicates (up to 57 wt%), including kaolinite-smectite (K-S) mixed-layer phases. The K-S was composed of either 30-50% kaolinite or 94-98% kaolinite layers. Discrete kaolinite was also present. The bimodal K-S mineralogical composition is likely due to two precursor phases: feldspar for the kaolinite-rich K-S and volcanic glass for the smectite-rich K-S. K-S with intermediate composition (50-94% kaolinite) was rare, due to its instability. Statistical analysis showed significant differences between the chemical compositions of the soils developed on the two different parent volcanic compositions. The soils overlying the more felsic parent rocks were less altered than those overlying the flood basalt. When comparing the weathering indices calculated in this study, we conclude that while the CIA and CIW may be more readily determined, the W and Wmin indices can elucidate information on the composition of the original rock from which they formed. The W index is more sensitive to certain variables when compared with the newly derived mineralogical Wmin index; however the Wmin index takes into account mineral phases within the sample, which provides a more detailed interpretation of weathering rates than chemistry alone. In addition the Wmin index correlated with meteorological variables, such as elevation (and consequently temperature and precipitation), known to influence the degree of pedogenesis. The Wmin index can be used to enhance our understanding of the processes that occur during weathering processes to supplement information gained from traditional chemical weathering indices

Information and resource management systems for Internet of Things: Energy management, communication protocols and future applications

The idea of the Internet of Things (IoT) has enabled the objects of our surroundings to intercommunicate with each other in diverse working environments by utilizing their embedded architectural and communication technologies. IoT has provided humans the capability to manipulate the operations and data available from different information systems using these intelligent objects available in the surroundings. The scope of IoT is to serve humanity across different domains of life covering industrial, health, home and day-to-day operations of Information Systems (IS). Due to the huge number of heterogeneous network elements interacting and working under IoT based information systems, there is an enormous need for resource management for the smooth running of IoT operations. The key aspect in IoT implementations is to have resource-constrained embedded devices and objects participating in IoT operations. It is important to meet the challenges raised during management and sharing of resources in IoT based information systems. Managing resources by implementing protocols, algorithms and techniques are required to enhance the scalability, reliability and stability in IoT operations across different fields of technology. This special issue opens the new areas of interest for the researchers in the domain of resource management in IoT operations

NanoAmpli-Seq: a workflow for amplicon sequencing for mixed microbial communities on the nanopore sequencing platform

Background: Amplicon sequencing on Illumina sequencing platforms leverages their deep sequencing and multiplexing capacity but is limited in genetic resolution due to short read lengths. While Oxford Nanopore or Pacific Biosciences sequencing platforms overcome this limitation, their application has been limited due to higher error rates or lower data output. Results: In this study, we introduce an amplicon sequencing workflow, i.e., NanoAmpli-Seq, that builds on the intramolecular-ligated nanopore consensus sequencing (INC-Seq) approach and demonstrate its application for full-length 16S rRNA gene sequencing. NanoAmpli-Seq includes vital improvements to the INC-Seq protocol that reduces sample processing time while significantly improving sequence accuracy. The developed protocol includes chopSeq software for fragmentation and read orientation correction of INC-Seq consensus reads while nanoClust algorithm was designed for read partitioning-based de novo clustering and within cluster consensus calling to obtain accurate full-length 16S rRNA gene sequences. Conclusions: NanoAmpli-Seq accurately estimates the diversity of tested mock communities with average consensus sequence accuracy of 99.5% for 2D and 1D2 sequencing on the nanopore sequencing platform. Nearly all residual errors in NanoAmpli-Seq sequences originate from deletions in homopolymer regions, indicating that homopolymer aware base calling or error correction may allow for sequencing accuracy comparable to short-read sequencing platforms

Bioreactor scalability: laboratory-scale bioreactor design influences performance, ecology, and community physiology in expanded granular sludge bed bioreactors

Studies investigating the feasibility of new, or improved, biotechnologies, such as wastewater treatment digesters, inevitably start with laboratory-scale trials. However, it is rarely determined whether laboratory-scale results reflect full-scale performance or microbial ecology. The Expanded Granular Sludge Bed (EGSB) bioreactor, which is a high-rate anaerobic digester configuration, was used as a model to address that knowledge gap in this study. Two laboratory-scale idealizations of the EGSB—a one-dimensional and a three- dimensional scale-down of a full-scale design—were built and operated in triplicate under near-identical conditions to a full-scale EGSB. The laboratory-scale bioreactors were seeded using biomass obtained from the full-scale bioreactor, and, spent water from the distillation of whisky from maize was applied as substrate at both scales. Over 70 days, bioreactor performance, microbial ecology, and microbial community physiology were monitored at various depths in the sludge-beds using 16S rRNA gene sequencing (V4 region), specific methanogenic activity (SMA) assays, and a range of physical and chemical monitoring methods. SMA assays indicated dominance of the hydrogenotrophic pathway at full-scale whilst a more balanced activity profile developed during the laboratory-scale trials. At each scale, Methanobacterium was the dominant methanogenic genus present. Bioreactor performance overall was better at laboratory-scale than full-scale. We observed that bioreactor design at laboratory-scale significantly influenced spatial distribution of microbial community physiology and taxonomy in the bioreactor sludge-bed, with 1-D bioreactor types promoting stratification of each. In the 1-D laboratory bioreactors, increased abundance of Firmicutes was associated with both granule position in the sludge bed and increased activity against acetate and ethanol as substrates. We further observed that stratification in the sludge-bed in 1-D laboratory-scale bioreactors was associated with increased richness in the underlying microbial community at species (OTU) level and improved overall performance

Biodiesel production from Cannabis sativa oil from Pakistan

The present study was appraised using response surface methodology for process optimization owing to strong interaction of reaction variables: NaOCH3 catalyst concentration (0.25–1.50%), methanol/oil molar ratio (3:1–9:1), reaction time (30–90 min), and reaction temperature (45–65°C). The quadratic polynomial equation was determined using response surface methodology for predicting optimum methyl esters yield from Cannabis sativa oil. The analysis of variance results indicated that molar ratio and reaction temperature were the key factors that appreciably influence the yield of Cannabis sativa oil methyl esters. The significant (p < 0.0001) variable interaction between molar ratio × catalyst concentration and reaction time × molar ratio was observed, which mostly affect the Cannabis sativa oil methyl esters yield. The optimum Cannabis sativa oil methyl esters yield, i.e., 86.01% was gained at 53°C reaction temperature, 7.5:1 methanol/oil molar ratio, 65 min reaction time, and 0.80% catalyst concentration. The results depicted a linear relationship between observed and predicted values. The residual analysis predicted the appropriateness of the central composite design. The Cannabis sativa oil methyl esters, analyzed by gas chromatography, elucidated six fatty acid methyl esters (linoleic, α-linolenic, oleic, palmitic, stearic, and γ-linolenic acids). In addition, the fuel properties, such as kinematic viscosity at 40°C; cetane number; acid value; flash point; cloud, pour, and cold filter plugging points; ash content; density; and sulphur content, of Cannabis sativa oil methyl esters were evaluated and discussed with reference to ASTM D 6751 and EU 14214 biodiesel specifications

Lower-limb alignment and posterior tibial slope in Pakistanis: a radiographic study

Abstract PURPOSE: To assess the lower-limb alignment and posterior tibial slope in Pakistanis. METHODS: 40 male and 19 female healthy Pakistanis aged 20 to 45 years were recruited. A full weight-bearing anteroposterior radiograph of the entire lower limb of each subject was obtained. The axial alignment was measured based on the centres of the femoral head, knee, and ankle. The tibiofemoral (TF) angle, knee joint line obliquity angle (angle J), and posterior tibial slope were determined. RESULTS: The mean TF angle was more varus in men than women (178.4 degrees vs. 180.0 degrees, p\u3c0.001). The mean angle J was more medially inclined in men than women (93.4 degrees vs. 91.4 degrees, p=0.007). The mean medial tibial slope was greater in women than men (16.0 degrees vs. 12.5 degrees, p\u3c0.001). The posterior tibial slope was greater in women than men (14.1 degrees vs. 12.5 degrees, p=0.02), and was greater than the 5 degrees to 10 degrees commonly reported. CONCLUSIONS: Knee alignment and geometry vary in different population subsets. With regard to total knee arthroplasty, the more medially inclined angle J in Pakistani men suggests that an anteroposterior cut of the distal femur should be in increased external rotation, compared with Pakistani women. Whereas the greater posterior tibial slope in Pakistanis suggests that a proximal tibial cut with a greater posterior tibial slope may reduce the chance of tibial loosening and increase postoperative knee range of motion, especially when using posterior cruciate ligament-retaining designs

The distinct features of microbial 'dysbiosis' of Crohn's disease do not occur to the same extent in their unaffected, genetically linked kindred

Background/Aims: Studying the gut microbiota in unaffected relatives of people with Crohn’s disease (CD) may advance our understanding of the role of bacteria in disease aetiology. Methods: Faecal microbiota composition (16S rRNA gene sequencing), genetic functional capacity (shotgun metagenomics) and faecal short chain fatty acids (SCFA) were compared in unaffected adult relatives of CD children (CDR, n = 17) and adult healthy controls, unrelated to CD patients (HUC, n = 14). The microbiota characteristics of 19 CD children were used as a benchmark of CD ‘dysbiosis’. Results: The CDR microbiota was less diverse (p = 0.044) than that of the HUC group. Local contribution of β-diversity analysis showed no difference in community structure between the CDR and HUC groups. Twenty one of 1,243 (1.8%) operational taxonomic units discriminated CDR from HUC. The metagenomic functional capacity (p = 0.207) and SCFA concentration or pattern were similar between CDR and HUC (p&#62;0.05 for all SCFA). None of the KEGG metabolic pathways were different between these two groups. Both of these groups (HUC and CDR) had a higher microbiota α-diversity (CDR, p = 0.026 and HUC, p&#60;0.001) with a community structure (β-diversity) distinct from that of children with CD. Conclusions: While some alterations were observed, a distinct microbial ‘dysbiosis’, characteristic of CD patients, was not observed in their unaffected, genetically linked kindred

The active microbial community more accurately reflects the anaerobic digestion process : 16S rRNA (gene) sequencing as a predictive tool

Amplicon sequencing methods targeting the 16S rRNA gene have been used extensively to investigate microbial community composition and dynamics in anaerobic digestion. These methods successfully characterise amplicons, but do not distinguish micro-organisms that are actually responsible for the process. In this research, the archaeal and bacterial community of 48 full-scale anaerobic digestion plants was evaluated on DNA (total community) and RNA (active community) level via 16S rRNA (gene) amplicon sequencing. A significantly higher diversity on DNA compared with the RNA level was observed for archaea, but not for bacteria. Beta diversity analysis showed a significant difference in community composition between the DNA and RNA of both bacteria and archaea. This related with 25.5 and 42.3% of total OTUs for bacteria and archaea, respectively, that showed a significant difference in their DNA and RNA profiles. Similar operational parameters affected the bacterial and archaeal community, yet, the differentiating effect between DNA and RNA was much stronger for archaea. In conclusion, a clear difference in active (RNA) and total (DNA) community profiles was observed, implying the need for a combined approach to estimate community stability in anaerobic digestion