Exploring the Diversity of the Human Blood Virome

Metagenomics is greatly improving our ability to discover new viruses, as well as their possible associations with disease. However, metagenomics has also changed our understanding of viruses in general. The vast expansion of currently known viral diversity has revealed a large fraction of non-pathogenic viruses, and offers a new perspective in which viruses function as important components of many ecosystems. In this vein, studies of the human blood virome are often motivated by the search for new viral diseases, especially those associated with blood transfusions. However, these studies have revealed the common presence of apparently non-pathogenic viruses in blood, particularly human anelloviruses and, to a lower extent, human pegiviruses (HPgV). To shed light on the diversity of the human blood virome, we subjected pooled plasma samples from 587 healthy donors in Spain to a viral enrichment protocol, followed by massive parallel sequencing. This showed that anelloviruses were clearly the major component of the blood virome and showed remarkable diversity. In total, we assembled 332 complete or near-complete anellovirus genomes, 50 of which could be considered new species. HPgV was much less frequent, but we, nevertheless, recovered 17 different isolates that we subsequently used for characterizing the diversity of this virus. In-depth investigation of the human blood virome should help to elucidate the ecology of these viruses, and to unveil potentially associated diseases.

Selection of a Nuclease-Resistant RNA Aptamer Targeting CD19

The transmembrane glycoprotein cluster of differentiation 19 (CD19) is a B cell–specific surface marker, expressed on the majority of neoplastic B cells, and has recently emerged as a very attractive biomarker and therapeutic target for B-cell malignancies. The development of safe and effective ligands for CD19 has become an important need for the development of targeted conventional and immunotherapies. In this regard, aptamers represent a very interesting class of molecules. Additionally referred to as ‘chemical antibodies’, they show many advantages as therapeutics, including low toxicity and immunogenicity. Here, we isolated a nuclease-resistant RNA aptamer binding to the human CD19 glycoprotein. In order to develop an aptamer also useful as a carrier for secondary reagents, we adopted a cell-based SELEX (Systematic Evolution of Ligands by EXponential Enrichment) protocol adapted to isolate aptamers able to internalise upon binding to their cell surface target. We describe a 2′-fluoro pyrimidine modified aptamer, named B85.T2, which specifically binds to CD19 and shows an exquisite stability in human serum. The aptamer showed an estimated dissociation constant (KD) of 49.9 ± 13 nM on purified human recombinant CD19 (rhCD19) glycoprotein, a good binding activity on human B-cell chronic lymphocytic leukaemia cells expressing CD19, and also an effective and rapid cell internalisation, thus representing a promising molecule for CD19 targeting, as well as for the development of new B-cell malignancy-targeted therapies.

Bacterial diversity dynamics in microbial consortia selected for lignin utilization

Lignin is nature’s largest source of phenolic compounds. Its recalcitrance to enzymatic conversion is still a limiting step to increase the value of lignin. Although bacteria are able to degrade lignin in nature, most studies have focused on lignin degradation by fungi. To understand which bacteria are able to use lignin as the sole carbon source, natural selection over time was used to obtain enriched microbial consortia over a 12-week period. The source of microorganisms to establish these microbial consortia were commercial and backyard compost soils. Cultivation occurred at two different temperatures, 30°C and 37°C, in defined culture media containing either Kraft lignin or alkaline-extracted lignin as carbon source. iTag DNA sequencing of bacterial 16S rDNA gene was performed for each of the consortia at six timepoints (passages). The initial bacterial richness and diversity of backyard compost soil consortia was greater than that of commercial soil consortia, and both parameters decreased after the enrichment protocol, corroborating that selection was occurring. Bacterial consortia composition tended to stabilize from the fourth passage on. After the enrichment protocol, Firmicutes phylum bacteria were predominant when lignin extracted by alkaline method was used as a carbon source, whereas Proteobacteria were predominant when Kraft lignin was used. Bray-Curtis dissimilarity calculations at genus level, visualized using NMDS plots, showed that the type of lignin used as a carbon source contributed more to differentiate the bacterial consortia than the variable temperature. The main known bacterial genera selected to use lignin as a carbon source were Altererythrobacter, Aminobacter, Bacillus, Burkholderia, Lysinibacillus, Microvirga, Mycobacterium, Ochrobactrum, Paenibacillus, Pseudomonas, Pseudoxanthomonas, Rhizobiales and Sphingobium. These selected bacterial genera can be of particular interest for studying lignin degradation and utilization, as well as for lignin-related biotechnology applications.

The Histone H1-like protein AlgP facilitates even spacing of polyphosphate granules in Pseudomonas aeruginosa

Synthesis of polyphosphate (polyP) is an ancient and universal stress and starvation response in bacteria. In many bacteria, polyP chains come together to form granular superstructures within cells. Some species appear to regulate polyP granule subcellular organization. Despite the critical role of polyP in starvation fitness, the composition of these structures, mechanism(s) underpinning their organization, and functional significance of such organization are poorly understood. We previously determined that granules become transiently evenly spaced on the cell’s long axis during nitrogen starvation in the opportunistic human pathogen Pseudomonas aeruginosa>. Here, we developed a granule-enrichment protocol to screen for polyP granule-localizing proteins. We identified AlgP as a protein that associates with polyP granules. We further discovered that AlgP is required for the even spacing of polyP granules. AlgP is a DNA-binding protein with a 154 amino acid C-terminal domain enriched in ‘KPAA’ repeats and variants of this repeat, with an overall sequence composition similar to the C-terminal tail of eukaryotic histone H1. Granule size, number, and spacing are significantly perturbed in the absence of AlgP, or when AlgP is truncated to remove the C-terminus. The ΔalgP and algPΔCTD mutants having fewer, larger granules. We speculate that AlgP may contribute to spacing by tethering polyP granules to the chromosome, thereby inhibiting fusion with neighboring granules. Our discovery that AlgP facilitates granule spacing allows us for the first time to directly uncouple granule biogenesis from even spacing, and will inform future efforts to explore the functional significance of granule organization on fitness during starvation.

Exploring the Diversity of the Human Blood Virome

Metagenomics is greatly improving our ability to discover new viruses, as well as their possible associations with disease. However, metagenomics has also changed our understanding of viruses in general. The vast expansion of currently known viral diversity has revealed a large fraction of non-pathogenic viruses, and offers a new perspective in which viruses function as important components of many ecosystems. In this vein, studies of the human blood virome are often motivated by the search for new viral diseases, especially those associated with blood transfusions. However, these studies have revealed the common presence of apparently non-pathogenic viruses in blood, in particular human anelloviruses and, to a lower extent, human pegiviruses (HPgV). To shed light on the diversity of the human blood virome, we subjected pooled plasma samples from 587 healthy donors in Spain to a viral enrichment protocol followed by massive parallel sequencing. This showed that anelloviruses were clearly the major component of the blood virome and showed remarkable diversity. In total, we assembled 332 complete or near-complete anellovirus genomes, 50 of which could be considered new species. HPgV was much less frequent, but we nevertheless recovered 17 different isolates that we subsequently used for characterizing the diversity of this virus. In-depth investigation of the human blood virome should help to elucidate the ecology of these viruses, and to unveil potentially associated diseases.

Implementing the Soil Enrichment Protocol at Scale: Opportunities for an Agricultural Carbon Market

High-quality agricultural carbon credits that incentivize regenerative practices can help address climate change through greenhouse gas (GHG) abatement and CO2 sequestration. Generating large volumes of such credits requires rigorous crediting methodologies. The Soil Enrichment Protocol (SEP) by the Climate Action Reserve (CAR) aims to unlock this type of crediting potential. The SEP includes new expert-driven standards for validating the use of soil biogeochemical modeling to generate credits. Technical experts at Indigo Ag participated in the SEP working group and are supporting implementation of the first project, CAR 1459_RP1, on hundreds of thousands of acres in the US. The authors share their thoughts on new approaches enabled by the SEP as both contributors to the theory behind and practitioners of these approaches. The SEP enables scalable, high-quality credits through four main advances: (1) allowing flexibility in the use of biogeochemical models that meet explicit performance requirements, (2) enabling a new approach to field-level, modeled baselines, (3) supporting a hybrid approach of credit generation using both soil measurement and modeling, and (4) requiring a new type of credit uncertainty quantification that accounts for multiple sources of uncertainty. Together these advances support agricultural credit quantification that enables payments to offset transitional costs for growers, at large enough scales to create a robust market, with a level of rigor that ensures any credited emission reductions have real climate impact. Innovations in soil analyses, advances in research, and improvements in data collection could further improve the potential for agricultural carbon credits to scale.

n-3 LC-PUFA Enrichment Protocol for Red Earthworm, Eisenia fetida: A Cheap and Sustainable Method

This study assessed the potential of omega-3 long chain polyunsaturated fatty acids (n-3 LC-PUFA) enrichment in red earthworm (Eisenia fetida, REW), via the addition of fish oil (FO, anchovy oil) to the culture-compost. FO addition to compost was tested at graded inclusion doses (0, 2.5, 5 and 10 g/100 g compost) and for different timeperiods, up to 96h. Dose, time, as well as dose and time interactions of FO compost enrichment had significant effects on all fatty acids of earthworm bodies. The enrichment with 10 g FO/100 g compost sharply increased DHA levels of the worms (14.01 mol%) within just 24 h, compared to the control group (0.92 mol%). Similar increases (ranging between 10.99 and 15.55 mol%) occurred only after 48 and 96 h in lower FO enrichment levels (2.5 and 5.0 g/100 g compost, respectively). Therefore, it was concluded that, to obtain maximum n-3 LC-PUFA enrichment efficiency in REW, an enrichment period of 24 h is ideal and recommended for the 10 g FO/ 100 g compost, whereas longer enrichment periods (48-96 h) are more suitable for lower levels of FO inclusions (2.5 or 5 g/100 g compost).

Ultra-rapid and highly efficient enrichment of organic pollutants via magnetic nanoparticles/mesoporous nanosponge compounds for ultrasensitive nanosensors

Developing advanced sensing and detection technologies to effectively monitor organic micropollutants in water is under urgent demand in both scientific and industrial communities. Currently, owing to the ultrahigh sensitivity on the single-molecule level with highly informative spectra characteristics, SERS technique is regarded as the most direct and effective detection technique. However, some weakly adsorbed molecules, such as most of persistent organic pollutants, cannot exhibit strong SERS signals, which is a long-standing key challenge that has not been solved. Here, we show an enrichment-typed sensing strategy based on a powerful porous composite material, call mesoporous nanosponge. The nanosponge consists of magnetic nanoparticles immobilized porous β-cyclodextrin polymers, demonstrating remarkable capability of effective and fast removal of organic micropollutants, e.g. ~90% removal efficiency within ~1 min. With the anchoring of magnetic nanoparticles, the current new polymer adsorbent can be easily recycled from water and re-dispersed in ethanol so that the target molecules in the cavity of adsorbent is concentrated, with an enrichment factor up to ~103. By means of the current enrichment strategy, the limit of detection (LOD) of the typical organic pollutants can be significantly improved, i.e. increasing 2~3 orders of magnitude, compared with the detection without molecule enrichment protocol. Consequently, the current enrichment strategy is proved to be applicable in a variety of fields for portable and fast detection, such as Raman and fluorescent.