Activity Of Acid Β-glucosidase Research Articles

Mechanisms activating trace heavy metals in the rhizosphere microenvironment of Amaranthus hypochondriacus L.

Soil heavy metal activation is closely related to rhizosphere soil pH, O2 and enzyme activity levels. In this study, we used laser ablation inductively coupled plasmamass spectrometry (ICPMS), planar optode (PO), soil in situ enzyme spectrum analysis, and the diffusive gradients in thin films (DGT) method to investigate heavy metal accumulation in Amaranthus hypochondriacus L. growing in polluted soil (Cd, Pb and Zn) and unpolluted soil, as well as dissolved oxygen (DO), pH, soil enzyme activity, and trace heavy metal dynamics in rhizosphere and nonrhizosphere soils. The results revealed that the growth of A. hypochondriacus was significantly inhibited under combined Cd, Pb, and Zn stress, with the most pronounced effects on root and branch biomass. Radial oxygen loss (ROL) and alkalization occurred in the roots of A. hypochondriacus, which were influenced primarily by root growth and environmental conditions. The enzyme activity patterns indicated that acid phosphatase (ACP), alkaline phosphatase (ALP), and β-glucosidase (BG) activities were primarily associated with the root system and that the activities of these enzymes were relatively low in nonrooted soil. Under heavy metal stress, the ACP, ALP and BG activities in the hotspots of soil increased. DO, pH, and soil enzyme activity were significantly correlated with the presence of heavy metals Cd, Pb, and Zn, influencing the mobilization mechanisms of trace metals in the rhizosphere. These findings lay the groundwork for understanding the impact of root-induced alterations in O2, pH, and soil enzyme activity on the mobility of trace heavy metals in soil.

Proteomics analysis of the brain from a Gaucher disease mouse identifies pathological pathways including a possible role for transglutaminase 1.

Gaucher disease (GD) is a lysosomal storage disorder (LSD) caused by the defective activity of acid β-glucosidase (GCase) which results from mutations in GBA1. Neurological forms of GD (nGD) can be generated in mice by intra-peritoneal injection of conduritol B-epoxide (CBE) which irreversibly inhibits GCase. Using this approach, a number of pathological pathways have been identified in mouse brain by RNAseq. However, unlike transcriptomics, proteomics gives direct information about protein expression which is more likely to provide insight into which cellular pathways are impacted in disease. We now perform non-targeted, mass spectrometry-based quantitative proteomics on brains from mice injected with 50 mg/kg body weight CBE for 13 days. Of the 5038 detected proteins, 472 were differentially expressed between control and CBE-injected mice of which 104 were selected for further analysis based on higher stringency criteria. We also compared these proteins with differentially expressed genes (DEGs) identified by RNAseq. Some lysosomal proteins were up-regulated as was interferon signaling, whereas levels of ion channel related proteins and some proteins associated with neurotransmitter signaling were reduced, as was cholesterol metabolism. One protein, transglutaminase 1 (TGM1), which is elevated in a number of neurodegenerative diseases, was absent from the control group but was found at high levels in CBE-injected mice, and located in the extracellular matrix (ECM) in layer V of the cortex and intracellularly in Purkinje cells in the cerebellum. Together, the proteomics data confirm previous RNAseq data and add additional mechanistic understanding about cellular pathways that may play a role in nGD pathology.

Ecoenzymatic Stoichiometry in the Rhizosphere and Bulk Soil of a Larix principis-rupprechtii Plantation in North China

Soil extracellular enzymes play an important role in ecosystem energy conversion and material cycling. Ecoenzymatic stoichiometry can reflect the relationship between the soil’s microbial nutrient cycle and nutrient limitation. However, there have been few studies on the differences in ecoenzymatic stoichiometry and nutrient limitation between rhizosphere soil and bulk soil. This study examined soil nutrients and enzyme activities in rhizosphere soil and bulk soil in a Larix principis-rupprechtii plantation in north China. The results showed that the levels of soil organic carbon (C), total nitrogen (N), and available nutrients in the rhizosphere soil were significantly higher than those in the bulk soil, whereas the total potassium (TK) level was significantly lower. The soil C:N, C:P, and N:P ratios of the rhizosphere soil also exceeded those of the bulk soil. The acid phosphatase (ACP), urease (UE), and β-glucosidase (β-GC) activities in the rhizosphere soil exceeded those in the bulk soil, whereas the activities of N-acetyl-β-D-glucosidase (NAG), aminopeptidase (LAP), and nitrogenase (NA) were lower. The ratios of C, N, and P acquisition activities changed from 1:1.7:1 in the rhizosphere soil to 1:2:1 in the bulk soil. Redundancy analysis showed that the available K and soil water content in the rhizosphere soil were the most important soil factors affecting soil enzyme activities and ecoenzymatic stoichiometry; those in the bulk soil were soil N:P and soil water content. These results suggest that not all soil enzyme activities present rhizosphere effects and that bulk soil is more susceptible to N limitation in Larix principis-rupprechtii plantations. Plant roots play an important role in regulating soil nutrients and soil activities, and future studies should examine the underlying mechanisms in more detail.

Soil enzymatic activity in Brazilian biomes under native vegetation and contrasting cropping and management

Enzymatic activity is a useful indicator for the evaluation of soil biological components. However, a detailed analysis of such indicators in Brazilian soils is lacking. In this study, the activity of soil enzymes was evaluated using data from 73 publications covering 57 locations across all six biomes and eight soil orders out of the 13 that occur in Brazil. The results indicated a wide variation of enzymatic activity in soils, where the means in μg p-nitrophenol g−1 soil h−1 were 569, 116, 112, and 80 for acid phosphatase, alkaline phosphatase, β-glucosidase, and arylsulfatase, respectively. Likewise, values of 148 μg reducing sugars g−1 soil day−1 for cellulase, 57 μg triphenyl tetrazolium formazan g−1 soil day−1 for dehydrogenase, and 38 μg N-NH4+ g−1 soil h−1 for urease were recorded. Soil organic matter and microbial biomass were the best attributes correlated with enzymatic activity, followed by clay content and pH. A more detailed analysis of the acid phosphatase, arylsulfatase, and β-glucosidase activities showed that land-use effects were more pronounced in the Atlantic Forest and Cerrado than in the Caatinga biome. Soils under non-tillage and integrated crop-livestock systems showed higher enzymatic activity than soils under conventional tillage. Soils housing higher biodiversity and delivering higher crop yields also presented higher enzymatic activity, especially in terms of arylsulfatase and β-glucosidase. Enzymatic activity has been shown to be a versatile indicator of soil quality, as it can elucidate how to achieve high-quality standards in tropical and subtropical soils.

Venglustat combined with imiglucerase for neurological disease in adults with Gaucher disease type 3: the LEAP trial.

Gaucher disease type 3 is a chronic neuronopathic disorder with wide-ranging effects, including hepatosplenomegaly, anaemia, thrombocytopenia, skeletal disease and diverse neurological manifestations. Biallelic mutations in GBA1 reduce lysosomal acid β-glucosidase activity, and its substrates, glucosylceramide and glucosylsphingosine, accumulate. Enzyme replacement therapy and substrate reduction therapy ameliorate systemic features of Gaucher disease, but no therapies are approved for neurological manifestations. Venglustat is an investigational, brain-penetrant, glucosylceramide synthase inhibitor with potential to improve the disease by rebalancing influx of glucosylceramide with impaired lysosomal recycling. The Phase 2, open-label LEAP trial (NCT02843035) evaluated orally administered venglustat 15 mg once-daily in combination with maintenance dose of imiglucerase enzyme replacement therapy during 1 year of treatment in 11 adults with Gaucher disease type 3. Primary endpoints were venglustat safety and tolerability and change in concentration of glucosylceramide and glucosylsphingosine in CSF from baseline to Weeks 26 and 52. Secondary endpoints included change in plasma concentrations of glucosylceramide and glucosylsphingosine, venglustat pharmacokinetics in plasma and CSF, neurologic function, infiltrative lung disease and systemic disease parameters. Exploratory endpoints included changes in brain volume assessed with volumetric MRI using tensor-based morphometry, and resting functional MRI analysis of regional brain activity and connectivity between resting state networks. Mean (SD) plasma venglustat AUC0-24 on Day 1 was 851 (282) ng•h/ml; Cmax of 58.1 (26.4) ng/ml was achieved at a median tmax 2.00 h. After once-daily venglustat, plasma concentrations (4 h post-dose) were higher compared with Day 1, indicating ∼2-fold accumulation. One participant (Patient 9) had low-to-undetectable venglustat exposure at Weeks 26 and 52. Based on mean plasma and CSF venglustat concentrations (excluding Patient 9), steady state appeared to be reached on or before Week 4. Mean (SD) venglustat concentration at Week 52 was 114 (65.8) ng/ml in plasma and 6.14 (3.44) ng/ml in CSF. After 1 year of treatment, median (inter-quartile range) glucosylceramide decreased 78% (72, 84) in plasma and 81% (77, 83) in CSF; median (inter-quartile range) glucosylsphingosine decreased 56% (41, 60) in plasma and 70% (46, 76) in CSF. Ataxia improved slightly in nine patients: mean (SD, range) total modified Scale for Assessment and Rating of Ataxia score decreased from 2.68 [1.54 (0.0 to 5.5)] at baseline to 1.55 [1.88 (0.0 to 5.0)] at Week 52 [mean change: -1.14 (95% CI: -2.06 to -0.21)]. Whole brain volume increased slightly in patients with venglustat exposure and biomarker reduction in CSF (306.7 ± 4253.3 mm3) and declined markedly in Patient 9 (-13894.8 mm3). Functional MRI indicated stronger connectivity at Weeks 26 and 52 relative to baseline between a broadly distributed set of brain regions in patients with venglustat exposure and biomarker reduction but not Patient 9, although neurocognition, assessed by Vineland II, deteriorated in all domains over time, which illustrates disease progression despite the intervention. There were no deaths, serious adverse events or discontinuations. In adults with Gaucher disease type 3 receiving imiglucerase, addition of once-daily venglustat showed acceptable safety and tolerability and preliminary evidence of clinical stability with intriguing but intrinsically inconsistent signals in selected biomarkers, which need to be validated and confirmed in future research.

Comparison and application of the extraction method for the determination of enzymatic profiles in matured soybean koji

In this study, two methods of extracting crude enzymes from soy sauce koji were compared to investigate the effect of replacing the buffer extraction with distilled water extraction by measuring the enzyme activity. Results showed the enzyme activities of the two extracts in each koji sample were not significant differences. The stability of the different enzymes in the extracts varied. Acid protease, β-glucosidase and cellulase activities showed the best stability during the storage. Neutral protease, glucoamylase and aminopeptidase were moderately stable and their activities needed to be determined within 3 d of extract preparation, while xylanase activity was the least stable, decreasing by 40% on the second day of storage. Therefore, the recommended procedure for the analysis of koji enzyme profiles was as follows, xylanase was determined immediately after distilled water extraction, and then all other enzyme activity analyses need to be completed within 3 d to ensure the validity of enzyme analysis. This method was applied to the enzymatic analysis of 7 kojis made by different strains of Aspergillus oryzae, and to explore the association between the enzyme profile and the taste characteristics of seven fermented soy sauce. The results showed proteases were correlated with sour and richness, while carbohydrate-degrading enzymes were relatively more relevant to umami taste.

Acid β-glucosidase insufficiency activates the C-X-C Motif Chemokine Ligand 9/CXCR3 axis leading to T cell mediated inflammation in Gaucher disease

Abstract Gaucher disease (GD) is caused by GBA1 mutations that lead to decreased activity of lysosomal acid β-glucosidase and abnormal tissue accumulation of its substrate, glucosylceramide (GC). Monocyte lineage cells, e.g., macrophages (Mφs) and dendritic cells (DCs) are prominent disease effectors due to their massive accumulation of GC resulting in “Gaucher cells.” Interaction of Gaucher cells and T lymphocytes trigger massive secretion of IFNγ, TNFα, IL6, IL12p40, IL12p70, IL23, and IL17A/F, that leads to the tissue destruction in GD. The exact mechanisms that trigger excess tissue T cells recruitment in GD are still unclear. CXCR3+ T cells are enriched at inflammatory sites in patients with rheumatoid arthritis and multiple sclerosis. Using D409V/null (Gba19V/−) GD mouse model, this study uncovered the role of CXCR3 receptor and its C-X-C Motif Chemokine Ligand 9 (CXCL9) in induction of excess tissue recruitment of T cells in GD. Intracellular FACS staining of Mφs and DCs from Gba19V/− mice showed elevated amounts of CXCL9. CD4+ T and the CD8+ T cells purified from Gba19V/− mice showed increased amounts of CXCR3. Ex vivo and in vivo chemotaxis experiments, evaluated CXCL9’s recruitment of Gba19V/− T cells. Blockade of T-cell CXCL9 receptor using antibodies to CXCR3 caused marked reduction in their CXCL9-mediated chemotaxis in Gba19V/− mice. These data suggest that Gba1 defects and the resultant deficiency of acid β-glucosidase activate CXCL9-CXCR3 axis for enhanced tissue Tells recruitment of T cells in GD. Collectively, our results indicate a critical role for CXCR3 mediated T cell transmigration to sites of inflammation. Molecular targeting of CXCL9/CXCR3 axis may provide needed anti-inflammatory therapies in human patients with GD. Supported by Cincinnati Children’s Hospital Medical Center, 3333 Burnet Avenue, Cincinnati, Ohio 45229, USA

Mechanistic Insight into the Mode of Action of Acid β-Glucosidase Enhancer Ambroxol.

Ambroxol (ABX) is a mucolytic agent used for the treatment of respiratory diseases. Bioactivity has been demonstrated as an enhancement effect on lysosomal acid β-glucosidase (β-Glu) activity in Gaucher disease (GD). The positive effects observed have been attributed to a mechanism of action similar to pharmacological chaperones (PCs), but an exact mechanistic description is still pending. The current study uses cell culture and in vitro assays to study the effects of ABX on β-Glu activity, processing, and stability upon ligand binding. Structural analogues bromohexine, 4-hydroxybromohexine, and norbromohexine were screened for chaperone efficacy, and in silico docking was performed. The sugar mimetic isofagomine (IFG) strongly inhibits β-Glu, while ABX exerts its inhibitory effect in the micromolar range. In GD patient fibroblasts, IFG and ABX increase mutant β-Glu activity to identical levels. However, the characteristics of the banding patterns of Endoglycosidase-H (Endo-H)-digested enzyme and a substantially lower half-life of ABX-treated β-Glu suggest different intracellular processing. In line with this observation, IFG efficiently stabilizes recombinant β-Glu against thermal denaturation in vitro, whereas ABX exerts no significant effect. Additional β-Glu enzyme activity testing using Bromohexine (BHX) and two related structures unexpectedly revealed that ABX alone can refunctionalize β-Glu in cellula. Taken together, our data indicate that ABX has little in vitro ability to act as PC, so the mode of action requires further clarification.

Effect of Lactobacillus plantarum or Enterococcus faecalis as co-inoculants with Aspergillus oryzae in koji making on the physicochemical properties of soy sauce.

Soy sauce fermentation is largely mediated by bacteria, including several non-salt-tolerant bacterial species. The present study aimed to compare the use of Aspergillus oryzae alone or in co-inoculation with Lactobacillus plantarum or Enterococcus faecalis on the resulting enzymatic activity, content of phenolic compounds, free amino acids, organic acids, and antioxidant capacity of soy sauce. Co-inoculation significantly increased acid protease, β-glucosidase, and pectinase activities during koji making. Additionally, the contents of total phenols, total flavonoids, organic acids, amino acids, isoflavones, as well as the antioxidant capacity were higher during moromi fermentation using co-inoculants compared to those obtained when A. oryzae was used as single inoculant. Overall, co-inoculation of A. oryzae with L. plantarum or E. faecalis during koji making was shown to effectively promote the production of active substances in soy sauce. PRACTICAL APPLICATION: In this study, Aspergillus oryzae in mixed inoculation with Lactobacillus plantarum or Enterococcus faecalis could be effectively used in koji making for soy sauce. The use of co-inoculants in koji making effectively promoted the production of active substances in soy sauce, hence it is improving health benefits, whilst diversifying the use of raw materials and the application value of soybean meal.

A first-year melon/cowpea intercropping system improves soil nutrients and changes the soil microbial community

The melon/cowpea intercropping system can be a specific and efficient cropping pattern in a horticultural field. Intercropping systems contribute to the optimization of land use, fostering sustainable and efficient agriculture. This study entails a first-year comparative intercropping assay using cowpea (Vigna unguiculata) and melon (Cucumis melo) under organic management with different patterns and 30% less organic fertilization than usual in monocrops. We determined the soil nutrients, physicochemical properties, enzyme activities and microbes by high-throughput sequencing. We found that the intercropping system changed the bacterial community structure independently of the intercropping pattern. The bacterial community was characterized by a higher abundance of the phyla Proteobacteria and Bacteroidetes phyla and of the genus Pseudomonas, which are related to nutrient cycling, and by greater amounts of other beneficial microorganisms like Bacillus, Streptomyces and Sphingomonas. The intercropped systems significantly boosted the total nitrogen, available phosphorus and total organic carbon levels in addition to the melon yield. They also enhanced the acid phosphatase and β-glucosidase activity compared to the melon monocrop. Results from this study suggest that melon/cowpea intercropping, starting from the first year, not only provides a stable supply of food and income due to the diversified cropping systems, but is also beneficial for the soil microbial community and environment.

Gaucher disease and SARS-CoV-2 infection: Experience from 181 patients in New York

SARS-CoV-2 infection carries high morbidity and mortality in individuals with chronic disorders. Its impact in rare disease populations such as Gaucher disease (GD) is unknown. In GD, decreased acid β-glucosidase activity leads to the accumulation of inflammatory glycosphingolipids and chronic myeloid cell immune activation which a priori could predispose to the most severe effects of SARS-CoV-2. To evaluate the determinants of SARS-CoV-2 infection in GD, we conducted a cross-sectional study in a large cohort. 181 patients were enrolled, including 150 adults and 31 children, with a majority of patients on treatment (78%). Information on COVID-19 exposure, symptoms, and SARS-CoV-2 nucleic acid and/or antibody testing was obtained during the peak of the pandemic in the New York City metropolitan area. Forty-five adults reported a primary exposure to someone with COVID-19 and 17 (38%) of these patients reported at least one COVID-19 symptom. A subset of adults was tested (n = 88) and in this group 18% (16/88) were positive. Patients testing positive for SARS-CoV-2 had significantly more symptoms (4.4 vs 0.3, p < 0.001) than patients testing negative. Among patients who were antibody-positive, quantitative titers indicated moderate to high antibody response. In GD adults, male gender, older age, increased BMI, comorbidities, GBA genotype, prior splenectomy and treatment status were not associated with the probability of reporting symptoms or testing positive. No patient required COVID-19-specific treatments and there were no deaths. Our data suggests that GD does not confer a heightened risk for severe effects of SARS-CoV-2 infection feared based on the known chronic inflammatory state in these patients.

Growing Medium Type Affects Organic Fertilizer Mineralization and CNPS Microbial Enzyme Activities

Managing plant fertilization is a major concern of greenhouse growers to achieve sustainable production with growing media (GM). Organic fertilization is popular but is more difficult to control, since organic compounds need first to be mineralized by microbes. After 7, 14, 28, and 56 days of incubation, we investigated the response of microbial activities and nutrient releases from three frequently used organic fertilizers (horn and two plant-based fertilizers) in three frequently employed GM types (peat, coir, and bark). We measured pH, electrical conductivity, nutrient contents (NH4+-N, NO3−-N, PO43−-P, SO42−-S), and enzyme activities (β-1.4-glucosidase, urease, acid phosphatase, arylsulfatase). After fertilization, microbes in coir expressed all the C, N, P, and S functions studied, making related nutrients available. In peat and bark, some C, N, P, and S-related pathways were locked. Peat presented high NH4+-N and PO43−-P releases linked to high acid phosphatase and β-glucosidase activities, while bark showed high nitrification rates but weak enzyme activities. Fertilizer types modulated these responses with lower activities and nutrient releases with horn. Our results contributed to better understanding mineralization processes in GM, showing different microbial responses to fertilization. This study pointed out the necessity to look deeper into microbial functions in GM optimizing biological and physicochemical properties.

In vitro and in vivo effects of Ambroxol chaperone therapy in two Italian patients affected by neuronopathic Gaucher disease and epilepsy

Gaucher disease (GD) is an autosomal recessive lysosomal storage disorder caused by mutations in the acid β-glucosidase encoding gene (GBA1), resulting in the deficient activity of acid β-glucosidase (GCase). To date, there is no approved treatment for the neurological manifestations of the disease. The role of Ambroxol as a chaperone for mutant GCase has been extensively demonstrated in vitro. Furthermore, different authors have reported beneficial effects of high doses of Ambroxol on neurological manifestations in patients affected by GD.In this report, we describe the in vitro and in vivo effects of Ambroxol in two patients (P1 and P2) affected by the neurological form of GD and epilepsy, carrying mutations already reported as responsive to the chaperone. Indeed, P1 presented the N188S mutation in compound heterozygous with a null allele (IVS2 + 1G > A) and P2 was homozygous for the L444P mutation. As expected, a beneficial effect of Ambroxol was observed in cultured fibroblasts as well as in vivo, both on epilepsy and on biomarkers of GD, in P1. However, Ambroxol was completely undefective in P2, suggesting that other factors besides the GBA1 mutation itself would be involved in the response therapy which would be difficult to predict based on the patient genotype. The present report expands the experience of Ambroxol treatment in neurological GD patients and highlights the need to in vitro test the individual response to Ambroxol even in patients carrying mutations already classified as responsive to the chaperone.

Prevalence of Gaucher Disease in Patients of Unknown Cause of Splenomegaly and/or Thrombocytopenia in Saudi Arabia

Rationale: Gaucher disease (GD) is the most common amongst the lysosomal storage disorders. Prevalence of GD in Saudi Arabia is not available in published literature and it is expected to be high and remains undiagnosed. In 2004, a Saudi study reported that GD accounts for 6% of all genetic metabolic disorders. While acid sphingomyelinase deficiency (ASMD) is a rare progressive genetic disorder with no clear data about its incidence in Saudi Arabia. Consequently, this study proposes to determine the prevalence of GD and ASMD in outpatient settings in Saudi Arabia by screening patients with unknown causes of splenomegaly and/or thrombocytopenia. These data will change the local practice and increase the awareness towards GD and ASMD in Saudi Arabia. Objectives: To determine the prevalence of GD in high risk group (defined as patients with splenomegaly and/or thrombocytopenia of unknown cause) and describe their characteristics. We also aim to determine the prevalence of ASMD in high risk group. Methods: A multi-center, observational, study will be conducted in 25 specialty care centres across Saudi Arabia, these centres are mainly focusing on pediatric hematology, adult hematology and hemato-pathology. Female or male patients (aged 2-75 years) will deemed eligible if they have clinical, instrumental or laboratory signs of splenomegaly or thrombocytopenia over a period of 12 months without definitive cause. These patients will be tested for acid β-glucosidase and acid sphingomyelinase enzymes activity on dried blood spot (DBS) samples. Patients with hematological malignancies, hemolytic anemia, and/or thalassemia (except sickle cell disease) will be excluded. A total of 400 patients from Saudi Arabia who fulfill the eligibility criteria will be enrolled in the study. All patient data will be collected in a single visit. Each enrolled patient will visit the investigator for a baseline visit. The investigator will contact them later for sharing the investigation blood test results. During the baseline visit, data and blood sample for enzyme tests and genotyping will be collected by the investigator/designated person at the site. Discussion: Arab world represents one of the leading regions in terms of the incidence of congenital and genetic disorders; a growing body of published literature reported a notable trends towards higher incidence of congenital and genetic diseases, compared to other parts of the world(1). High consanguinity rates which reach up to 60% in some regions, high prevalence of haemoglobinopathies and metabolic disorders, relatively high maternal and parental age, and lack of proper genetic screening were reported as contributing factors for this high prevalence of genetic disorders in the Arab world(1-3). In Saudi Arabia, the situation appears to be no different as previous retrospective studies showed a relatively high incidence of genetic diseases such as inborn error of metabolism. The data of the present study will change the local practice and increase the awareness towards GD and ASMD in Saudi Arabia.

Pregnancy outcome in women with Gaucher disease type 1 who had unplanned pregnancies during eliglustat clinical trials.

Gaucher disease type 1 (GD1) is an inherited lysosomal storage disorder caused by deficient enzymatic activity of acid β‐glucosidase, resulting in accumulation of its substrate glucosylceramide, leading to debilitating visceral, hematologic, and skeletal manifestations. Women with GD1 are at increased risk for complications during pregnancy, delivery, and postpartum. Treatment with enzyme replacement therapy is generally recommended before and during pregnancy to reduce risks. Eliglustat, an oral substrate‐reduction therapy, is a first‐line treatment for adults with GD1 adults who have extensive, intermediate, or poor CYP2D6‐metabolizer phenotypes (>90% of patients). We report on pregnancy outcomes among women in eliglustat trials who had unplanned pregnancies and female partners of men in the trials. In four phase 2 and 3 eliglustat trials of 393 adults with GD1, women of childbearing potential were required to use contraception, have monthly pregnancy tests, and discontinue eliglustat promptly if pregnant. In phase 2 and 3 trials, 18 women had 19 pregnancies, resulting in 14 healthy infants from 13 pregnancies (one set of twins), three elective terminations, one ectopic pregnancy, one spontaneous abortion, and one in utero death. Median estimated eliglustat exposure duration during pregnancy was 38 days. In phase 1 trials (non‐GD1 subjects), one woman had a spontaneous abortion. Partners of 16 eliglustat‐treated men with GD1 had 18 pregnancies, all resulting in healthy infants. Eliglustat is not approved during pregnancy due to limited data. Guidelines for clinicians and patients with GD that address use of eliglustat in women of childbearing potential are needed.

Acid phosphomonoesterase and β-glucosidase activities in volcanic soils under permanent fertilized pastures: distribution profile and microbial effort toward P acquisition

ABSTRACT This work focused on the amounts and distribution of organic C (OC), organic phosphorus (Po), and acid phosphomonoesterase (AcP) and β-glucosidase (βG) activities in volcanic soils, as well as on the effect of inorganic P (Pi) on potential P-acquiring effort. The OC and Po contents are correlated with the active aluminum (Alox) and iron (Feox) and with the stratified gradients of the enzyme activities. The AcP activity does not seem to be suppressed by soil Pi content, showing a positive linear relationship with Pi. High values for the AcP:βG activity ratio are observed, which suggests a high energy allocation (C consumption) to ensure P demand. With the increase of Alox and Feox, the AcP:βG ratios increase and the potential C:P acquisition (ln(βG):ln(AcP)) values decrease. Nevertheless, the average C:P molar ratio of the six soils (115) are below the critical value, above which soils may present low P bioavailability. The results suggest that the effort for P acquisition (AcP:βG) and the potential C:P acquisition are both affected by the size of the Pi pools and by Alox and Feox contents. If Pi retention increases due to high contents of Alox and Feox, energy allocation shifts toward AcP production, in detriment of microbial population and plant nutrition. Considering the OC accumulation in the studied soils, several hypotheses may be considered such as the energy demand toward P acquisition, and the ability of Alox and Feox metal complexes to protect organic molecules to depress βG activity.

Cover crop biomass and species composition affect soil microbial community structure and enzyme activities in semiarid cropping systems

Cover crops are promoted to increase soil organic carbon (SOC) storage and improve soil biological health in agricultural systems. However, cover crop effects on soil microbial communities – key regulators of SOC and nutrients – and functioning are not clear in semiarid environments. This study investigated the response of soil microbial community structure and enzyme activities to cover crop integration under limited-irrigation winter wheat (Triticum aestivum)-sorghum (Sorghum bicolor)-fallow rotation. The study had a randomized complete block design with eight treatments and three replications. Treatments were pea (Pisum sativum); oat (Avena sativa); canola (Brassica napus L.); and mixtures of pea + oat (POmix), pea + canola (PCmix), pea + oat + canola (POCmix), and pea + oat + canola + hairy vetch (Vicia villosa) + forage radish (Raphanus sativus L.) + barley (Hordeum vulgare L.) (diverse-mix) as cover crops; and a fallow. Soil samples were collected in the summer of 2017 and 2018 from 0 to 15 cm depth of each plot established in fall 2015. Microbial community size and structure were evaluated via ester-linked fatty acid methyl ester (EL-FAME) analysis, which showed that total microbial community size and fungal community were similar between soils under oats and diverse-mix, and were 31% and 41% greater than fallow, respectively. The FAME marker for arbuscular mycorrhizal fungi (AMF) was 84% greater under oats than fallow. The combined enzyme activity of acid phosphatase, β-glucosidase, and β-glucosaminidase was 294 mg p-nitrophenol (PNP) kg−1 soil h−1 under diverse-mix, which was greater than fallow (204 mg PNP kg−1 soil h−1, p = 0.021) in 2018. Cover cropping during fallow period in a crop-fallow rotation could increase total microbial community size, fungal abundance, and enzyme activities associated with carbon (C) and nutrient cycling. Among cover crops, oat and its mixture with legumes (pea and hairy vetch) and brassicas (canola and forage radish) were most effective in improving soil health and biogeochemical cycling in a hot and dry semiarid climate.

Bioremediation of Cd-contaminated soil by earthworms (Eisenia fetida): Enhancement with EDTA and bean dregs

The remediation of cadmium (Cd) contaminated soil has become a global problem due to its toxicity to living organisms. In this study, earthworm (Eisenia fetida) alone or combined with EDTA or bean dregs were used for Cd removal from soils. The total and available Cd in soils, soil physicochemical and biological (soil enzyme) properties, Cd accumulation in the earthworm and its antioxidant responses towards Cd, were determined during the 35 days of soil incubation experiment. Our results showed that earthworms were capable of removing Cd from soils, and the remediation process was accelerated by both EDTA and bean dregs. By translocation of Cd from soils, the content of Cd in earthworm steadily increased with the exposure time to 8.11, 12.80, and 9.26 mg kg−1 on day 35 for T2 (earthworm alone), T3 (EDTA enhancement), and T4 (bean dregs enhancement), respectively. Consequently, a great reduction in the Cd contents in soils was achieved in T3 (36.53%) and T4 (30.8%) compared with T2 (28.95%). The concentrations of water/DTPA extractable Cd were also reduced, indicating the low Cd mobility after amendment. Finally, the soil became more fertile and active after wermi-remediation. The soil pH, EC, NO3−-N, available P, and K contents increased, while soil SOM, DOC, and NH4+-N contents were decreased. There were higher soil enzyme activities (including acid phosphatase activity, β-glucosidase activity, and urease activity) among treatments with earthworms. Additionally, the operational taxonomic units (OTUs) increased by 100–150 units, and the higher chao1 and Shannon indexes indicated the enhanced microbial community after wermi-remediation, especially among treatment with EDTA and bean dregs. Therefore, we concluded that earthworms, alone or combined with EDTA and bean dregs, are feasible for the remediation of Cd-contaminated soil.

Biomarker Response to Oral Eliglustat in Adults with Gaucher Disease Type 1: Results from 4 Completed Clinical Trials

INTRODUCTION: Gaucher disease type 1 (GD1) is caused by hereditary deficiency in activity of lysosomal acid β-glucosidase and lysosomal dysfunction due to abnormal recycling and storage of glycosphingolipids especially in tissue macrophages. Clinical manifestations can include anemia, thrombocytopenia, hepatosplenomegaly, and debilitating bone disease. Hematologists care for most patients with GD1 but may be unaware of the importance of plasma biomarkers not only as tissue response surrogates that are useful for monitoring treatment efficacy but also as pathogenic molecules that can directly contribute to disease manifestations. These biomarkers include chitotriosidase, macrophage inflammatory protein 1β (MIP-1β), glucosylceramide, glucosylsphingosine, and monosialodihexosyl ganglioside (GM3). Here we show differing changes in biomarker levels after inception of eliglustat therapy in treatment-naïve GD1 patients and in patients previously treated with enzyme replacement therapy (ERT). Eliglustat is an oral inhibitor of glucosylceramide biosynthesis approved by FDA as a first-line treatment of adults with GD1 who have extensive, intermediate, or poor CYP2D6 metabolizer phenotypes (&gt;90% of patients). METHODS: We evaluated percent change from baseline in biomarker levels during time on eliglustat in the 4 completed clinical trials of oral eliglustat for GD1 adults (Sanofi Genzyme): Phase 2/NCT00358150 (treatment-naïve patients with moderate to severe baseline disease, N=26); Phase 3 ENGAGE/NCT00891202 (treatment-naïve patients with mild to moderate baseline disease, N=40); Phase 3 ENCORE/NCT00943111 (switch patients at prespecified therapeutic goals stabilized after a mean of 10 years of ERT], N=159); Phase 3 EDGE/NCT01074944 (mixture of treatment-naïve [N=22] and ERT-switch patients [N=148] with mild to moderate baseline disease and variable prior time on ERT). All percentages were calculated in the subset of patients with biomarkers measured at baseline and the reported timepoint. The attrition of patients with data at the longer-term time points is attributable primarily to a combination of 2-year enrollment periods, trial design, and protocol-mandated switches to commercial therapy for US patients. Most patients (81%) continued until eliglustat became commercially available or completion of their trial; 9 patients (2.3% overall) withdrew due to adverse events reported as drug-related. For Phase 2, ENGAGE, and ENCORE, chitotriosidase genotypes were obtained and values were normalized; patients homozygous for the CHIT null mutation were excluded and values for heterozygous patients were doubled. For EDGE patients, chitotriosidase genotype data were not obtained but patients with ≥4 zero values (n=12) were excluded. RESULTS: In treatment-naïve patients, all biomarkers were elevated at baseline. Median levels of glucosylceramide, GM3, and MIP-1β normalized within 1 year of starting eliglustat. Chitotriosidase and glucosylsphingosine levels markedly decreased but did not normalize. After 3-8 years on treatment, median percent reduction in chitotriosidase and glucosylsphingosine (Phase 2 and ENGAGE) was 85-92% (Table) but values did not normalize. In patients switching from ERT, median chitotriosidase was still modestly elevated at baseline. Median values for other biomarkers were normal or near normal. After 1 year of treatment, chitotriosidase decreased (Table). Median glucosylceramide, GM3, and MIP-1β remained normal or normalized. After 3-4 years, GM3 and MIP-1β remained normal. Median percent reduction in chitotriosidase from baseline was 60-63% but values did not normalize. Eliglustat was generally well-tolerated in all 4 trials; 97% of adverse events were mild/moderate and 86% considered unrelated to eliglustat by the investigator. CONCLUSIONS: Gaucher disease biomarkers decreased substantially in treatment-naïve patients after eliglustat treatment. In switch patients, including very stable patients after a mean of 10 years on ERT, chitotriosidase and MIP-1β also decreased substantively after eliglustat treatment. Future longitudinal follow-up of eliglustat-treated patients should indicate whether the biomarker and plasma glycosphingolipid responses reported here in stable patients at long-term therapeutic goals are associated with immunological and clinical improvements. Disclosures Weinreb: Shire HGT: Consultancy, Honoraria, Speakers Bureau; Sanofi Genzyme: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Pfizer Corporation: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Cox:Sanofi Genzyme: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Reimbursement, Research Funding. Mistry:Sanofi Genzyme: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Reimbursement, Research Funding; NIH NIMS: Research Funding; Shire: Honoraria, Other: Travel Reimbursement, Research Funding; Pfizer: Honoraria, Other: Travel Reimbursement; Synageva: Honoraria, Other: Travel Reimbursement. Charrow:BioMarin: Research Funding; Shire: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi Genzyme: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amicus: Research Funding. Lukina:JSC GENERIUM: Research Funding; Sanofi Genzyme: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Reimbursement, Research Funding; Shire: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Reimbursement. Foster:Sanofi Genzyme: Employment. Peterschmitt:Sanofi Genzyme: Employment.

Evaluation of Organic Substrates and Microorganisms as Bio-Fertilisation Tool in Container Crop Production

Microorganisms are only effective when adequate conditions for their survival and development are provided. Among the factors that influence its effectiveness includes the type of soil or culture substrate, which works as an energy source reserve. Therefore, a tomato and a melon crop were established in different cycles to assess the effect of the physicochemical properties of organic substrates based on coconut fibre and vermicompost in three proportions, 0:100, 40:60 and 60:40 (% v:v), on the microbial activity in the rhizosphere when the bacteria Azotobacter vinelandii, Bacillus megaterium and Frateuria aurantia were applied. Concentrations of NO3−, H2PO4−, K+ and Ca2+ in the petiole cellular extract (PCE) were quantified at 60, 90 and 120 days after transplantation (DAT) for tomato and 45 and 65 DAT for melon. We analysed dehydrogenase activity (DHA), acid phosphatase activity (FTA) and β-glucosidase activity (β-GLU). In order to maintain optimal volumetric moisture for the survival of microorganisms, automatic control was used to manage the irrigation frequency between 22%–28%. The results showed that physicochemical substrate properties, by incorporating 40% vermicompost into the coconut fibre mixture, increased enzymatic activity. Plants that were inoculated with Azotobacter vinelandii and Frateuria aurantia showed an improvement in NO3− and K+ assimilation achieving highest yields.