Continuous monitoring of cellulase action on microcrystalline cellulose

Summary Cellobiose oxidase from Phanerochaete chrysosporium was used for continuous monitoring of cellulase action on microcrystalline cellulose (Avicel). Two protocols are described, the parameter monitored being either the decline in electrode potential as ferricyanide is reduced or consumption of dioxygen. Most experiments used a commercial cellulase preparation from Trichoderma reesei and ferricyanide as acceptor. Within 1 min of an addition of cellulase, ferricyanide reduction reached a steady rate. This was converted into a rate of production of substrate for celobiose oxidase, in mol·min^–1. Experiments were conducted either with a constant concentration of cellulase and increasing Avicel, or with constant Avicel and increasing cellulase. Kinetic analysis of the experiments with constant cellulase indicated a K _mof 4.8 ± 1.0 (g cellulose)·1^–1, which was close to the value predicted from binding studies. The specific activity of the cellulase was measured as 375±25 mol·(g cellulase)^–1·min^–1 in experiments with a high cellulose concentration, but was less than half this value when the cellulose was saturated with cellulase. The maximal rate of cellulose degradation was 9.6±1.3 mol·(g cellulose)^–1·min^–1.     

Nucleoside Diphosphate Levels in Taproots and Fibrous Roots of Beta vulgaris L. : Implifications for Control of Sucrose Allocation between Two Sinks

Sucrose synthase (EC 2.4.1.13) of sugarbeet (Beta vulgaris L.) fibrous roots has a distinctly different order of preference for nucleoside diphosphate substrates than that of the taproot enzyme (Silvius, Snyder 1979 Plant Physiol 64: 1070-1073). Separation and quantitation of UDP, ADP, and GDP in root extracts by high-pressure liquid chromatography revealed that UDP levels in taproot tissue were 5 to 10 times greater than those of fibrous root tissues. The lower fibrous root UDP levels were associated with significantly higher ADP and GDP levels in these roots as compared to the taproot. These differences are consistent with differences in the substrate affinity of sucrose synthase and suggest a regulatory role of the enzyme in the control of sucrose cleavage and utilization between the two root types.     

Effect of antioxidants on the genetic stability of cryopreserved mint shoot tips by encapsulation–dehydration

The effect of antioxidants applied in one step of a cryopreservation protocol by encapsulation–dehydration on recovery and genetic stability of mint shoot tips has been studied. Glutathione (0.16 or 0.24 mM), ascorbic acid (0.28 or 0.43 mM) and α-tocopherol (vitamin E) were added to the preculture medium (0.3 M sucrose). DNA was extracted from three different types of samples: leaves from shoots, callus at the base of shoots and callus. RAPD and AFLP markers were used to assess the genetic stability. The use of antioxidants did not improve recovery after cryopreservation. One of the genotypes, ‘MEN 198’, showed higher percentage of stable samples than the other one, ‘MEN 186’ (56 vs. 37?%; considering all treatments and types of explant). The use of vitamin E improved the percentage of stable samples with respect to control treatment (no antioxidant) in ‘MEN 186’. No differences in the percentages of stable samples were observed among cryopreserved and non-cryopreserved (treated similarly without immersion in liquid nitrogen) plant material. Recovered shoots of both genotypes showed higher stability (76–80?% stable samples) than callus samples (14–22?%).     

Reciprocal allopolyploid grasses (Festuca × Lolium) display stable patterns of genome dominance

Allopolyploidization entailing the merger of two distinct genomes in a single hybrid organism, is an important process in plant evolution and a valuable tool in breeding programs. Newly established hybrids often experience massive genomic perturbations, including karyotype reshuffling and gene expression modifications. These phenomena may be asymmetric with respect to the two progenitors, with one of the parental genomes being “dominant.” Such “genome dominance” can manifest in several ways, including biased homoeolog gene expression and expression level dominance. Here we employed a k-mer–based approach to study gene expression in reciprocal Festuca pratensis Huds. × Lolium multiflorum Lam. allopolyploid grasses. Our study revealed significantly more genes where expression mimicked that of the Lolium parent compared with the Festuca parent. This genome dominance was heritable to successive generation and its direction was only slightly modified by environmental conditions and plant age. Our results suggest that Lolium genome dominance was at least partially caused by its more efficient trans-acting gene expression regulatory factors. Unraveling the mechanisms responsible for propagation of parent-specific traits in hybrid crops contributes to our understanding of allopolyploid genome evolution and opens a way to targeted breeding strategies.     

Nusinersen treatment and cerebrospinal fluid neurofilaments: An explorative study on Spinal Muscular Atrophy type 3 patients

The antisense oligonucleotide Nusinersen has been recently licensed to treat spinal muscular atrophy (SMA). Since SMA type 3 is characterized by variable phenotype and milder progression, biomarkers of early treatment response are urgently needed. We investigated the cerebrospinal fluid (CSF) concentration of neurofilaments in SMA type 3 patients treated with Nusinersen as a potential biomarker of treatment efficacy. The concentration of phosphorylated neurofilaments heavy chain (pNfH) and light chain (NfL) in the CSF of SMA type 3 patients was evaluated before and after six months since the first Nusinersen administration, performed with commercially available enzyme-linked immunosorbent assay (ELISA) kits. Clinical evaluation of SMA patients was performed with standardized motor function scales. Baseline neurofilament levels in patients were comparable to controls, but significantly decreased after six months of treatment, while motor functions were only marginally ameliorated. No significant correlation was observed between the change in motor functions and that of neurofilaments over time. The reduction of neurofilament levels suggests a possible early biochemical effect of treatment on axonal degeneration, which may precede changes in motor performance. Our study mandates further investigations to assess neurofilaments as a marker of treatment response.     

Monitoring tropical forests under a functional perspective with satellite‐based vegetation optical depth

Monitoring ecosystem functions in forests is a priority in a climate change scenario, as climate‐induced events may initially alter the functions more than slow‐changing attributes, such as biomass. The ecosystem functional properties (EFPs) are quantities that characterize key ecosystem processes. They can be derived by point observations of gas and energy exchanges between the ecosystems and the atmosphere that are collected globally at FLUXNET flux tower sites and upscaled at ecosystem level. The properties here considered describe the ability of ecosystems to optimize the use of resources for carbon uptake. They represent functional forest information, are dependent on environmental drivers, linked to leaf traits and forest structure, and influenced by climate change effects. The ability of vegetation optical depth (VOD) to provide forest functional information is investigated using 2011–2014 satellite data collected by the Soil Moisture and Ocean Salinity mission and using the EFPs as reference dataset. Tropical forests in Africa and South America were analyzed, also according to ecological homogeneous units. VOD jointly with water deficit information explained 93% and 87% of the yearly variability in both flux upscaled maximum gross primary productivity and light use efficiency functional properties, in Africa and South America forests respectively. Maps of the retrieved properties evidenced changes in forest functional responses linked to anomalous climate‐induced events during the study period. The findings indicate that VOD can support the flux upscaling process in the tropical range, affected by high uncertainty, and the detection of forest anomalous functional responses. Preliminary temporal analysis of VOD and EFP signals showed fine‐grained variability in periodicity, in signal dephasing, and in the strength of the relationships. In selected drier forest types, these satellite data could also support the monitoring of functional dynamics.     

How to measure,report and verify soil carbon change to realize the potential of soil carbon sequestration for atmospheric greenhouse gas removal

There is growing international interest in better managing soils to increase soil organic carbon (SOC) content to contribute to climate change mitigation, to enhance resilience to climate change and to underpin food security, through initiatives such as international ‘4p1000’ initiative and the FAO's Global assessment of SOC sequestration potential (GSOCseq) programme. Since SOC content of soils cannot be easily measured, a key barrier to implementing programmes to increase SOC at large scale, is the need for credible and reliable measurement/monitoring, reporting and verification (MRV) platforms, both for national reporting and for emissions trading. Without such platforms, investments could be considered risky. In this paper, we review methods and challenges of measuring SOC change directly in soils, before examining some recent novel developments that show promise for quantifying SOC. We describe how repeat soil surveys are used to estimate changes in SOC over time, and how long‐term experiments and space‐for‐time substitution sites can serve as sources of knowledge and can be used to test models, and as potential benchmark sites in global frameworks to estimate SOC change. We briefly consider models that can be used to simulate and project change in SOC and examine the MRV platforms for SOC change already in use in various countries/regions. In the final section, we bring together the various components described in this review, to describe a new vision for a global framework for MRV of SOC change, to support national and international initiatives seeking to effect change in the way we manage our soils.