Lime pretreatment of sugar beet pulp and evaluation of synergy between ArfA, ManA and XynA from Clostridium cellulovorans on the pretreated substrate

Sugar beet pulp (SBP) is a waste product from the sugar beet industry and could be used as a potential biomass feedstock for second generation biofuel technology. Pretreatment of SBP with ‘slake lime’ (calcium hydroxide) was investigated using a 23 factorial design and the factors examined included lime loading, temperature and time. The pretreatment was evaluated for its ability to enhance enzymatic degradation using a combination of three hemicellulases, namely ArfA (an arabinofuranosidase), ManA (an endo-mannanase) and XynA (an endo-xylanase) from C. cellulovorans to determine the conditions under which optimal activity was facilitated. Optimal pretreatment conditions were found to be 0.4 g lime/g SBP, with 36 h digestion at 40 °C. The synergistic interactions between ArfA, ManA and XynA from C. cellulovorans were subsequently investigated on the pretreated SBP. The highest degree of synergy was observed at a protein ratio of 75% ArfA to 25% ManA, with a specific activity of 2.9 U/g protein. However, the highest activity was observed at 4.2 U/g protein at 100% ArfA. This study demonstrated that lime treatment enhanced enzymatic hydrolysis of SBP. The ArfA was the most effective hemicellulase for release of sugars from pretreated SBP, but the synergy with the ManA indicated that low levels of mannan in SBP were probably masking the access of the ArfA to its substrate. XynA displayed no synergy with the other two hemicellulases, indicating that the xylan in the SBP was not hampering the access of ArfA or ManA to their substrates and was not closely associated with the mannan and arabinan in the SBP

The role of multiple marks in epigenetic silencing and the emergence of a stable bivalent chromatin state

We introduce and analyze a minimal model of epigenetic silencing in budding yeast, built upon known biomolecular interactions in the system. Doing so, we identify the epigenetic marks essential for the bistability of epigenetic states. The model explicitly incorporates two key chromatin marks, namely H4K16 acetylation and H3K79 methylation, and explores whether the presence of multiple marks lead to a qualitatively different systems behavior. We find that having both modifications is important for the robustness of epigenetic silencing. Besides the silenced and transcriptionally active fate of chromatin, our model leads to a novel state with bivalent (i.e., both active and silencing) marks under certain perturbations (knock-out mutations, inhibition or enhancement of enzymatic activity). The bivalent state appears under several perturbations and is shown to result in patchy silencing. We also show that the titration effect, owing to a limited supply of silencing proteins, can result in counter-intuitive responses. The design principles of the silencing system is systematically investigated and disparate experimental observations are assessed within a single theoretical framework. Specifically, we discuss the behavior of Sir protein recruitment, spreading and stability of silenced regions in commonly-studied mutants (e.g., sas2, dot1) illuminating the controversial role of Dot1 in the systems biology of yeast silencing.Comment: Supplementary Material, 14 page

Synergy between EngE, XynA and ManA from Clostridium cellulovorans on corn stalk, grass and pineapple pulp substrates

The synergistic interaction between various hemi/cellulolytic enzymes has become more important in order to achieve effective and optimal degradation of complex lignocellulose substrates for biofuel production. This study investigated the synergistic effect of three enzymes endoglucanase (EngE), mannanase (ManA) and xylanase (XynA) on the degradation of corn stalk, grass, and pineapple fruit pulp and determined the optimal degree of synergy between combinations of these enzymes. It was established that EngE was essential for degradation of all of the substrates, while the hemicellulases were able to contribute in a synergistic fashion to increase the activity on these substrates. Maximum specific activity and degree of synergy on the corn stalk and grass was found with EngE:XynA in a ratio of 75:25%, with a specific activity of 41.1 U/mg protein and a degree of synergy of 6.3 for corn stalk, and 44.1 U/mg protein and 3.4 for grass, respectively. The pineapple fruit pulp was optimally digested using a ManA:EngE combination in a 50:50% ratio; the specific activity and degree of synergy achieved were 52.4 U/mg protein and 2.7, respectively. This study highlights the importance of hemicellulases for the synergistic degradation of complex lignocellulose. The inclusion of a mannanase in an enzyme consortium for biomass degradation should be examined further as this study suggests that it may play an important, although mostly overlooked, role in the synergistic saccharification of lignocellulose

Effectiveness and costs of phototest in dementia and cognitive impairment screening

<p>Abstract</p> <p>Background</p> <p>To assess and compare the effectiveness and costs of Phototest, Mini Mental State Examination (MMSE), and Memory Impairment Screen (MIS) to screen for dementia (DEM) and cognitive impairment (CI).</p> <p>Methods</p> <p>A phase III study was conducted over one year in consecutive patients with suspicion of CI or DEM at four Primary Care (PC) centers. After undergoing all screening tests at the PC center, participants were extensively evaluated by researchers blinded to screening test results in a Cognitive-Behavioral Neurology Unit (CBNU). The gold standard diagnosis was established by consensus of expert neurologists. Effectiveness was assessed by the proportion of correct diagnoses (diagnostic accuracy [DA]) and by the kappa index of concordance between test results and gold standard diagnoses. Costs were based on public prices and hospital accounts.</p> <p>Results</p> <p>The study included 140 subjects (48 with DEM, 37 with CI without DEM, and 55 without CI). The MIS could not be applied to 23 illiterate subjects (16.4%). For DEM, the maximum effectiveness of the MMSE was obtained with different cutoff points as a function of educational level [k = 0.31 (95% Confidence interval [95%CI], 0.19-0.43), DA = 0.60 (95%CI, 0.52-0.68)], and that of the MIS with a cutoff of 3/4 [k = 0.63 (95%CI, 0.48-0.78), DA = 0.83 (95%CI, 0.80-0.92)]. Effectiveness of the Phototest [k = 0.71 (95%CI, 0.59-0.83), DA = 0.87 (95%CI, 0.80-0.92)] was similar to that of the MIS and higher than that of the MMSE. Costs were higher with MMSE (275.9 ± 193.3€ [mean ± sd euros]) than with Phototest (208.2 ± 196.8€) or MIS (201.3 ± 193.4€), whose costs did not significantly differ. For CI, the effectiveness did not significantly differ between MIS [k = 0.59 (95%CI, 0.45-0.74), DA = 0.79 (95%CI, 0.64-0.97)] and Phototest [k = 0.58 (95%CI, 0.45-0.74), DA = 0.78 (95%CI, 0.64-0.95)] and was lowest for the MMSE [k = 0.27 (95%CI, 0.09-0.45), DA = 0.69 (95%CI, 0.56-0.84)]. Costs were higher for MMSE (393.4 ± 121.8€) than for Phototest (287.0 ± 197.4€) or MIS (300.1 ± 165.6€), whose costs did not significantly differ.</p> <p>Conclusion</p> <p>MMSE is not an effective instrument in our setting. For both DEM and CI, the Phototest and MIS are more effective and less costly, with no difference between them. However, MIS could not be applied to the appreciable percentage of our population who were illiterate.</p

Familial hypercholesterolaemia in children and adolescents from 48 countries: a cross-sectional study

Background Approximately 450 000 children are born with familial hypercholesterolaemia worldwide every year, yet only 2·1% of adults with familial hypercholesterolaemia were diagnosed before age 18 years via current diagnostic approaches, which are derived from observations in adults. We aimed to characterise children and adolescents with heterozygous familial hypercholesterolaemia (HeFH) and understand current approaches to the identification and management of familial hypercholesterolaemia to inform future public health strategies. Methods For this cross-sectional study, we assessed children and adolescents younger than 18 years with a clinical or genetic diagnosis of HeFH at the time of entry into the Familial Hypercholesterolaemia Studies Collaboration (FHSC) registry between Oct 1, 2015, and Jan 31, 2021. Data in the registry were collected from 55 regional or national registries in 48 countries. Diagnoses relying on self-reported history of familial hypercholesterolaemia and suspected secondary hypercholesterolaemia were excluded from the registry; people with untreated LDL cholesterol (LDL-C) of at least 13·0 mmol/L were excluded from this study. Data were assessed overall and by WHO region, World Bank country income status, age, diagnostic criteria, and index-case status. The main outcome of this study was to assess current identification and management of children and adolescents with familial hypercholesterolaemia. Findings Of 63 093 individuals in the FHSC registry, 11 848 (18·8%) were children or adolescents younger than 18 years with HeFH and were included in this study; 5756 (50·2%) of 11 476 included individuals were female and 5720 (49·8%) were male. Sex data were missing for 372 (3·1%) of 11 848 individuals. Median age at registry entry was 9·6 years (IQR 5·8–13·2). 10 099 (89·9%) of 11 235 included individuals had a final genetically confirmed diagnosis of familial hypercholesterolaemia and 1136 (10·1%) had a clinical diagnosis. Genetically confirmed diagnosis data or clinical diagnosis data were missing for 613 (5·2%) of 11 848 individuals. Genetic diagnosis was more common in children and adolescents from high-income countries (9427 [92·4%] of 10 202) than in children and adolescents from non-high-income countries (199 [48·0%] of 415). 3414 (31·6%) of 10 804 children or adolescents were index cases. Familial-hypercholesterolaemia-related physical signs, cardiovascular risk factors, and cardiovascular disease were uncommon, but were more common in non-high-income countries. 7557 (72·4%) of 10 428 included children or adolescents were not taking lipid-lowering medication (LLM) and had a median LDL-C of 5·00 mmol/L (IQR 4·05–6·08). Compared with genetic diagnosis, the use of unadapted clinical criteria intended for use in adults and reliant on more extreme phenotypes could result in 50–75% of children and adolescents with familial hypercholesterolaemia not being identified. Interpretation Clinical characteristics observed in adults with familial hypercholesterolaemia are uncommon in children and adolescents with familial hypercholesterolaemia, hence detection in this age group relies on measurement of LDL-C and genetic confirmation. Where genetic testing is unavailable, increased availability and use of LDL-C measurements in the first few years of life could help reduce the current gap between prevalence and detection, enabling increased use of combination LLM to reach recommended LDL-C targets early in life. Funding Pfizer, Amgen, Merck Sharp & Dohme, Sanofi–Aventis, Daiichi Sankyo, and Regeneron