Single-cell RNA-seq: advances and future challenges

Phenotypically identical cells can dramatically vary with respect to behavior during their lifespan and this variation is reflected in their molecular composition such as the transcriptomic landscape. Single-cell transcriptomics using next-generation transcript sequencing (RNA-seq) is now emerging as a powerful tool to profile cell-to-cell variability on a genomic scale. Its application has already greatly impacted our conceptual understanding of diverse biological processes with broad implications for both basic and clinical research. Different single-cell RNA-seq protocols have been introduced and are reviewed here-each one with its own strengths and current limitations. We further provide an overview of the biological questions single-cell RNA-seq has been used to address, the major findings obtained from such studies, and current challenges and expected future developments in this booming field

Sediment and phosphorus transport in irrigation furrows

Sediment and phosphorus (P) in agricultural runoff can impair water quality in streams, lakes, and rivers. We studied the factors affecting P transfer and transport in irrigated furrows in six freshly tilled fallow fields, 110 to 180 m long with 0.007 to 0.012 m m' slopes without the interference of raindrops or sheet flow that occur during natural or simulated rain. The soil on all fields was Portneuf silt loam (coarse-silty, mixed, superactive, mesic Durinodic Xeric Haplocalcids). Flow rate, sediment concentration, and P concentrations were monitored at four, equally spaced locations in each furrow. Flow rate decreased with distance down the furrow as water infiltrated. Sediment concentration varied with distance and time with no set pattern. Total P concentrations related directly to sediment concentrations (r2 = 0.75) because typically >90% of the transported P was particulate P, emphasizing the need to control erosion to reduce P loss. Dissolved reactive phosphorus (DRP) concentrations decreased with time at a specific furrow site but increased with distance down the furrow as contact time with soil and suspended sediment increased. The DRP concentration correlated better with sediment concentration than extractable furrow soil P concentration. However, suspended sediment concentration tended to not affect DRP concentration later in the irrigation (>2 h). These results indicate that the effects of soil P can be overshadowed by differences in flow hydraulics, suspended sediment loads, and non-equilibrium conditions

The <i>Castalia</i> mission to Main Belt Comet 133P/Elst-Pizarro

We describe Castalia, a proposed mission to rendezvous with a Main Belt Comet (MBC), 133P/Elst-Pizarro. MBCs are a recently discovered population of apparently icy bodies within the main asteroid belt between Mars and Jupiter, which may represent the remnants of the population which supplied the early Earth with water. Castalia will perform the first exploration of this population by characterising 133P in detail, solving the puzzle of the MBC’s activity, and making the first in situ measurements of water in the asteroid belt. In many ways a successor to ESA’s highly successful Rosetta mission, Castalia will allow direct comparison between very different classes of comet, including measuring critical isotope ratios, plasma and dust properties. It will also feature the first radar system to visit a minor body, mapping the ice in the interior. Castalia was proposed, in slightly different versions, to the ESA M4 and M5 calls within the Cosmic Vision programme. We describe the science motivation for the mission, the measurements required to achieve the scientific goals, and the proposed instrument payload and spacecraft to achieve these

Changes in meltwater chemistry over a 20-year period following a thermal regime switch from polythermal to cold-based glaciation at Austre Broggerbreen, Svalbard

Our long-term study gives a rare insight into meltwater hydrochemistry following the transition of Austre Brøggerbreen from polythermal to cold-based glaciation and its continued retreat. We find that the processes responsible for ion acquisition did not change throughout the period of records but became more productive. Two regimes before and after July/August 2000 were identified from changes in solute concentrations and pH. They resulted from increased chemical weathering occurring in ice-marginal and proglacial environments that have become progressively exposed by glacier retreat. Carbonate carbonation nearly doubled between 2000 and 2010, whilst increases in the weathering of silicate minerals were also marked. In addition, the end of ablation season chemistry was characterized by reactions in long residence time flow paths like those in subglacial environments, in spite of their absence in the watershed. Furthermore, the retreat of the glacier caused the sudden re-routing of meltwaters through its immediate forefield during 2009, which more than doubled crustal ion yields in this particular year and influenced chemical weathering in 2010 regardless of a low water flux. Such a “flush” of crustally derived ions can be meaningful for downstream terrestrial and marine ecosystems. We therefore find that, during glacier retreat, the recently exposed forefield is the most chemically active part of the watershed, making high rates of weathering possible, even when ice losses have caused a switch to cold-based conditions with no delayed subglacial drainage flowpaths. In addition, the drainage system reorganization events result in significant pCO2 depletion in an otherwise high pCO2 system

A randomized double blind placebo-controlled clinical trial of Ceterizine in horses affected by seasonal headshaking

Headshaking is a threat to the welfare of affected horses. As seasonal headshaking may be triggered by allergic conditions, this clinical trial investigated whether the second-generation antihistamine cetirizine decreased headshaking. The objective was to assess the clinical effect of cetirizine versus placebo on seasonal headshaking. The hypothesis was that it would reduce headshaking by 50% in 50% of the horses. Thirty client-owned horses with seasonal headshaking were selected on the basis of information from the owner and a general clinical examination. In this crossover randomised double-blind placebo-controlled clinical trial, horses were given cetirizine (0.4mg/kg twice daily PO for 7 days) or placebo (same number of tablets twice daily PO for 7 days) in a randomised order, with a washout period of 1 week in between. A 9-minute lunge protocol was recorded on video at the start and after both treatment weeks, and the number of headshakes in this 9-minute period was scored by two assessors in a blinded manner. Data of 29 horses were analysed. The number of headshakes decreased by more than 50% in 10 horses when they were given cetirizine and in 8 horses when they were given placebo. This difference was not significant (p=0.73). In a mixed linear model incorporating weather conditions no significant treatment effect was found either. In conclusion: no significant effect of cetirizine on seasonal headshaking was found in the group of horses included in this study

Dual RNA-seq of pathogen and host

A comprehensive understanding of host–pathogen interactions requires a knowledge of the associated gene expression changes in both the pathogen and the host. Traditional, probe-dependent approaches using microarrays or reverse transcription PCR typically require the pathogen and host cells to be physically separated before gene expression analysis. However, the development of the probe-independent RNA sequencing (RNA-seq) approach has begun to revolutionize transcriptomics. Here, we assess the feasibility of taking transcriptomics one step further by performing 'dual RNA-seq', in which gene expression changes in both the pathogen and the host are analysed simultaneously

A randomized double blind placebo-controlled clinical trial of Ceterizine in horses affected by seasonal headshaking

Headshaking is a threat to the welfare of affected horses. As seasonal headshaking may be triggered by allergic conditions, this clinical trial investigated whether the second-generation antihistamine cetirizine decreased headshaking. The objective was to assess the clinical effect of cetirizine versus placebo on seasonal headshaking. The hypothesis was that it would reduce headshaking by 50% in 50% of the horses. Thirty client-owned horses with seasonal headshaking were selected on the basis of information from the owner and a general clinical examination. In this crossover randomised double-blind placebo-controlled clinical trial, horses were given cetirizine (0.4mg/kg twice daily PO for 7 days) or placebo (same number of tablets twice daily PO for 7 days) in a randomised order, with a washout period of 1 week in between. A 9-minute lunge protocol was recorded on video at the start and after both treatment weeks, and the number of headshakes in this 9-minute period was scored by two assessors in a blinded manner. Data of 29 horses were analysed. The number of headshakes decreased by more than 50% in 10 horses when they were given cetirizine and in 8 horses when they were given placebo. This difference was not significant (p=0.73). In a mixed linear model incorporating weather conditions no significant treatment effect was found either. In conclusion: no significant effect of cetirizine on seasonal headshaking was found in the group of horses included in this study

A placebo controlled clinical trial on cetirizine in seasonal headshakers

Trigeminal mediated headshaking is a syndrome that may have several causes and is a potential threat to the wellbeing of the horse. Because one of the possible etiologies is an allergic rhinitis, a randomized double-blinded placebo controlled clinical trial on the effectiveness of cetirizine, an antihistamine, in 30 seasonal headshakers was performed. A lunge protocol of 9 min, consisting of rest, walk and trot, was recorded at the start, after a week of treatment with cetirizine and after a week of placebo. Weather conditions were also recorded and owners kept a diary.The videos were scored for the amount of shakes. The overall amount of shakes was 67 ± 94 at the start and 47 ± 72 on cetirizine. This difference was significant (P = 0.01). Placebo compared to start was not significantly different (P = 0.18) as was cetirizine vs placebo (P = 0.74). Evaluating results of individual horses 10 horses showed >50% improvement on cetirizine and 11 horses showed a >50% improvement on placebo (compared to t = 0). Factors like weather circumstances will be taken into account.This study shows the difficulty of using a clinical trial for a syndrome influenced by weather and many other factors. It does suggest however that even if cetirizine will not be beneficial in all (seasonal) headshaking horses, it may be in some, both diagnostic for proving an allergic etiology, as well as therapeutic which is also suggested by follow-up of these horses and other clinical cases

Sediment and phosphorus transport in irrigation furrows

Sediment and phosphorus (P) in agricultural runoff can impair water quality in streams, lakes, and rivers. We studied the factors affecting P transfer and transport in irrigated furrows in six freshly tilled fallow fields, 110 to 180 m long with 0.007 to 0.012 m m' slopes without the interference of raindrops or sheet flow that occur during natural or simulated rain. The soil on all fields was Portneuf silt loam (coarse-silty, mixed, superactive, mesic Durinodic Xeric Haplocalcids). Flow rate, sediment concentration, and P concentrations were monitored at four, equally spaced locations in each furrow. Flow rate decreased with distance down the furrow as water infiltrated. Sediment concentration varied with distance and time with no set pattern. Total P concentrations related directly to sediment concentrations (r2 = 0.75) because typically >90% of the transported P was particulate P, emphasizing the need to control erosion to reduce P loss. Dissolved reactive phosphorus (DRP) concentrations decreased with time at a specific furrow site but increased with distance down the furrow as contact time with soil and suspended sediment increased. The DRP concentration correlated better with sediment concentration than extractable furrow soil P concentration. However, suspended sediment concentration tended to not affect DRP concentration later in the irrigation (>2 h). These results indicate that the effects of soil P can be overshadowed by differences in flow hydraulics, suspended sediment loads, and non-equilibrium conditions