[Simultaneous Immobilization of Cadmium and Arsenic in Paddy Soils with Novel Fe-Mn Combined Graphene Oxide].

Novel Fe-Mn combined graphene oxide （GO-FM） material was produced and tested for its efficacy in remediating agricultural soil co-contaminated by Cd and As. In a 60-day soil incubation experiment， the remediation mechanism and immobilization effects of GO and GO-FM at different addition ratios （0.1%， 0.2%， and 0.3%） were investigated in Shangyu and Foshan soils， which had varying physicochemical properties and contamination degrees. The dynamic changes in pH， DOC concentration， bioavailable Cd and As content， and morphology of Cd and As were explored to determine the remediation efficacy of the materials. The results demonstrated that compared with that in the blank control， GO-FM increased the pH in Shangyu soil but decreased the pH in Foshan soil. After culture， both GO and GO-FM increased the soil DOC content. GO-FM decreased the soluble Cd concentration by 5.08%-19.19% and the bioavailability of Cd by 36.57%-42.8% in Foshan soil， and the main immobilization mechanism was electrostatic adsorption， complexation， and hydroxylated metal ion formation. The immobilization ability of GO-FM on Cd was lower than that of Foshan soil due to the influence of electrostatic repulsion in Shangyu acidic soil. However， with the increase in the amount of GO-FM， the trend of increasing the bioavailability of Cd by graphene oxide was inhibited. The addition of 0.2% and 0.3% GO-FM decreased the bioavailability of Cd by 6.45%-13.56% in Shangyu soil. Additionally， GO-FM decreased the bioavailability of As in Shangyu soil and Foshan soil by 4.34%-9.15% and 0.87%-5.71%， respectively. This was due to the immobilization mechanism of oxidation of As by manganese oxides and inner surface chelate between As and the surface hydroxyl group of iron oxides. In summary， the immobilization effect of GO-FM on Cd in Foshan soil was better than that in Shangyu soil， and the immobilization effect of GO-FM on As in Shangyu soil was better than that in Foshan soil， which can provide a theoretical basis and reference for the prevention and control of Cd and As co-contamination in different types of soil.

Safety and immunogenicity of a self-amplifying RNA vaccine against COVID-19: COVAC1, a phase I, dose-ranging trial.

BACKGROUND: Lipid nanoparticle (LNP) encapsulated self-amplifying RNA (saRNA) is a novel technology formulated as a low dose vaccine against COVID-19. METHODS: A phase I first-in-human dose-ranging trial of a saRNA COVID-19 vaccine candidate LNP-nCoVsaRNA, was conducted at Imperial Clinical Research Facility, and participating centres in London, UK, between 19th June to 28th October 2020. Participants received two intramuscular (IM) injections of LNP-nCoVsaRNA at six different dose levels, 0.1-10.0µg, given four weeks apart. An open-label dose escalation was followed by a dose evaluation. Solicited adverse events (AEs) were collected for one week from enrolment, with follow-up at regular intervals (1-8 weeks). The binding and neutralisation capacity of anti-SARS-CoV-2 antibody raised in participant sera was measured by means of an anti-Spike (S) IgG ELISA, immunoblot, SARS-CoV-2 pseudoneutralisation and wild type neutralisation assays. (The trial is registered: ISRCTN17072692, EudraCT 2020-001646-20). FINDINGS: 192 healthy individuals with no history or serological evidence of COVID-19, aged 18-45 years were enrolled. The vaccine was well tolerated with no serious adverse events related to vaccination. Seroconversion at week six whether measured by ELISA or immunoblot was related to dose (both p<0.001), ranging from 8% (3/39; 0.1µg) to 61% (14/23; 10.0µg) in ELISA and 46% (18/39; 0.3µg) to 87% (20/23; 5.0µg and 10.0µg) in a post-hoc immunoblot assay. Geometric mean (GM) anti-S IgG concentrations ranged from 74 (95% CI, 45-119) at 0.1µg to 1023 (468-2236) ng/mL at 5.0µg (p<0.001) and was not higher at 10.0µg. Neutralisation of SARS-CoV-2 by participant sera was measurable in 15% (6/39; 0.1µg) to 48% (11/23; 5.0µg) depending on dose level received. INTERPRETATION: Encapsulated saRNA is safe for clinical development, is immunogenic at low dose levels but failed to induce 100% seroconversion. Modifications to optimise humoral responses are required to realise its potential as an effective vaccine against SARS-CoV-2. FUNDING: This study was co-funded by grants and gifts from the Medical Research Council UKRI (MC_PC_19076), and the National Institute Health Research/Vaccine Task Force, Partners of Citadel and Citadel Securities, Sir Joseph Hotung Charitable Settlement, Jon Moulton Charity Trust, Pierre Andurand, Restore the Earth.

Mapping Mechanostable Pulling Geometries of a Therapeutic Anticalin/CTLA-4 Protein Complex.

We used single-molecule AFM force spectroscopy (AFM-SMFS) in combination with click chemistry to mechanically dissociate anticalin, a non-antibody protein binding scaffold, from its target (CTLA-4), by pulling from eight different anchor residues. We found that pulling on the anticalin from residue 60 or 87 resulted in significantly higher rupture forces and a decrease in koff by 2-3 orders of magnitude over a force range of 50-200 pN. Five of the six internal anchor points gave rise to complexes significantly more stable than N- or C-terminal anchor points, rupturing at up to 250 pN at loading rates of 0.1-10 nN s-1. Anisotropic network modeling and molecular dynamics simulations helped to explain the geometric dependency of mechanostability. These results demonstrate that optimization of attachment residue position on therapeutic binding scaffolds can provide large improvements in binding strength, allowing for mechanical affinity maturation under shear stress without mutation of binding interface residues.

Postpartum depression in Covid-19 risk-stratified hospital zones: A cross-sectional study from India.

OBJECTIVE: To determine point prevalence of postpartum depression (PPD) and whether quarantine policies required in postpartum COVID-19 suspects increased the risk of depressive symptoms when compared COVID -19 non suspects in early postpartum period. METHODS: A cross-sectional study conducted in the postnatal ward of Lady Hardinge Medical College and Smt. Sucheta Kriplani Hospital, New Delhi from August 2020 to February 2021 using Edinburgh Postnatal Depression Scale (EPDS) to estimate point prevalence of postpartum depression (PPD) in the stratified zones (suspect and nonsuspect) of the hospital. RESULTS: Total of 408 postpartum women,204 each (COVID-19 suspects, and nonsuspects zone) were enrolled. Prevalence of Postpartum depression using an EDPS score of >9 was 11.9%. Prevalence of depression (17.9%; vs 4.85%, p <0.001) and level of depression (5.01±3.41 vs 4.14± 2.54, p 0.004) was significantly higher in mothers in isolated quarantine wards of COVID-19 suspect zone as compared to nonsuspect zones. Anhedonia was reported significantly higher (p <0.001) in suspect zone mothers. Multivariate analysis showed 'stay in COVID-19 suspect zone' and 'COVID-19 Suspect status' as a significant predictor of PPD. CONCLUSION: Covid -19 postpartum suspects are at increased risk of developing postpartum depressive symptoms due to isolation and quarantine measures required in suspect zones to mitigate viral transmission.Limitations - Due to the cross-sectional nature of the study long-term impacts on mental health due to isolation could not be assessed. Factors such as marital relationship, COVID -19 related risk factors weren't assessed which may have bearing on the risk of developing PPD.

Quantitative analysis of genome packaging in recombinant AAV vectors by charge detection mass spectrometry.

Recombinant adeno-associated virus (rAAV) has emerged as an important gene therapy vector with many clinical trials currently in progress. Analytical characterization and quantitation of particle content remain challenges in both the development and production of rAAV vectors. In this study, charge detection mass spectrometry (CDMS) and gel electrophoresis are used to characterize the DNA content of recombinant AAV8 (rAAV8) vectors with a wide range of target genome sizes. We show that the differences between the masses of empty particles and particles with the genome of interest (GOI) are correlated with the expected genome mass. A small systematic deviation (around 2%) is attributed to the packaging of counterions along with the DNA. In addition to the GOI, a broad distribution of heterogeneous DNA is packaged. The distribution peaks are close to the packaging capacity of the rAAV8 vectors. There is also evidence for the co-packaging of small DNA fragments along with the GOI. Finally, we present evidence that incubation at an elevated temperature can reduce the heterogeneity of the packaged DNA. Taken together, these results show that CDMS is a viable tool for characterization of the packaged genome.

High Performance p-i-n Photodetectors on Ge-on-Insulator Platform.

In this article, we demonstrated novel methods to improve the performance of p-i-n photodetectors (PDs) on a germanium-on-insulator (GOI). For GOI photodetectors with a mesa diameter of 10 µm, the dark current at -1 V is 2.5 nA, which is 2.6-fold lower than that of the Ge PD processed on Si substrates. This improvement in dark current is due to the careful removal of the defected Ge layer, which is formed with the initial growth of Ge on Si. The bulk leakage current density and surface leakage density of the GOI detector at -1 V are as low as 1.79 mA/cm2 and 0.34 µA/cm, respectively. GOI photodetectors with responsivity of 0.5 and 0.9 A/W at 1550 and 1310 nm wavelength are demonstrated. The optical performance of the GOI photodetector could be remarkably improved by integrating a tetraethylorthosilicate (TEOS) layer on the oxide side due to the better optical confinement and resonant cavity effect. These PDs with high performances and full compatibility with Si CMOS processes are attractive for applications in both telecommunications and monolithic optoelectronics integration on the same chip.

Another insight into the contamination levels at Ogoniland in Niger Delta, Nigeria, with focus on Goi Creek.

Environmental pollution arising from crude oil exploratory activities has adversely impacted both the living and non-living components of the ecosystem. An insight into the impact of such activities with focus on Goi Creek in Ogoniland was provided in this study. We generated reliable data for physicochemical, heavy metal, and total hydrocarbon levels in the creek via a total of 40 samples collected across 4 stations designated within the creek in a duration of 10 months. With the exception of temperature, total dissolve solid, dissolved oxygen, chloride, sulfate, nitrate, zinc, and vanadium, the levels of other regulated parameters in the samples were not compatible with the national and international guidelines and standards for surface water quality including Federal Ministry of Environment Guidelines and standards for water quality in Nigeria and Environmental Protection Agency National recommended water quality criteria, respectively. The principal components (PCs) 1 and 2 of principal component analysis biplot revealed that the concentrations of the physicochemical parameters showed a mixed distribution among the stations; however, the heavy metals were more localized to station 4. Moreover, the dataset was subjected to water quality index (WQI), contamination index (Cd), heavy metal evaluation index (HEI), trace element toxicity index (TETI), and environmental water quality index (EWQI) to determine the extent of contamination. The result showed poor WQI, high contamination for Cd, and low-to-medium contamination for HEI. Although, there were variations among the stations on the contamination level, uniform remediation method should be adopted due to the similarity of the contaminants in all the stations.

Viability of power distribution in India - Challenges and Way Forward.

Energy is a key component for economic growth as well as for human development. India is the third-ranking electricity generator in the world but ranks 106th in terms of per capita consumption. Specifically, the distribution of power is the most important link in the value chain of the power sector since it is the cash register for the entire sector. In India, electricity is a concurrent subject shared between the Central and State Governments. While the generation of power has been deregulated since 2003, the supply of power to the consumers is generally carried out by Government-owned power Distribution Companies (DISCOMs). In this paper, the authors analyze the financial distress of DISCOMs in India, and explain how the supply-demand mismatch due to over-ambitious demand projections, the fixed and energy charges of thermal power plants, and the excess procurement of "must-run" renewable sources together contribute to the high cost of power procurement that drives the financial stress faced by DISCOMs. The authors support their assertion with a study of nine DISCOMs supplying electricity to 155 million people in three States and propose policy recommendations for a turnaround of such DISCOMs which can be rolled out with suitable modifications across India.

Diagnosis and principles of management of gestational diabetes mellitus in the prevailing COVID-19 pandemic.

BACKGROUND: Limited medical facilities are available due to Covid-19 pandemic. Nevertheless, all efforts should be made in planning judicial and possible methods of delivering health care, particularly to pregnant woman with GDM. GDM may play a crucial role in the increasing prevalence of diabetes and obesity and also may be the origin of cardiometabolic diseases. METHODS: It is mandatary to diagnose and care pregnant woman with GDM. The test suggested to diagnose GDM has to be evidence based and in this regard "a single test procedure" evaluated meets this requirement. This doable test has been accepted by the Diabetes in Pregnancy Study Group India (DIPSI) and approved by MHFW-GOI, WHO, International Diabetes Federation, and International Federation of Obstetricians and Gynecologists. MHFW-GOI also recommends testing at first antenatal visit and then at 24-28 weeks of gestation. This opportunity can also be utilized for performing ultrasonography for assessing fetal development. RESULT: The first-line management is MNT and life style modifications. Non-responders may require insulin or OHA. The target glycemic control is FPG ~ 5.0 mmol/dl (90 mg/dl) and 2 h PPPG ~ 6.7 mmol/dl (120 mg/dl). The goal is to obtain newborns birth weight appropriate for gestational age between 2.5 and 3.5 kg, a step to prevent offspring developing diabetes. CONCLUSION: The essential precaution required during COVID pandemic is to wear face mask, avoid crowded places, and maintain social distancing. Finally, the economical and evidence based "single test procedure" of DIPSI is most appropriate for screening during the COVID pandemic.

Greater Omentum Imaging-Reporting and Data System: establishing the grade of benign and malignant lesions of the greater omentum using ultrasonography.

OBJECTIVE: To establish Greater Omentum Imaging-Reporting and Data System (GOI-RADS) to evaluate the possibility of omental diseases being malignant. METHOD: A retrospective analysis was made of 883 patients who had undergone biopsy of the greater omentum in our center from October 2009 to October 2019. Twelve parameters of ultrasonographic images were evaluated, and the odds ratio of each group calculated. We assigned scores for the direct signs (omental echo, omental structure, and omental nodules) and indirect signs (separation of ascites, echo of ascites, mesenteric lymph nodes, and thickening of parietal peritoneum) of omental lesions. We created an omental score (OS) for each patient and receiver operating characteristic (ROC) curve to analyze its effectiveness in the differential diagnosis of benign and malignant omental diseases. RESULTS: The OS was divided into ≤5, 6, 7, 8, 9, 10, 11, 12, 13, and ≥ 14 points, and the malignant rate was 0, 1.85, 5.56, 30.36, 37.25, 87.72, 96.72, 98.28, 99.08, and 100%, respectively. The area under the ROC curve (AUC) was 0.976. When taking 10 points as the cutoff value to diagnose benign and malignant omental diseases, the sensitivity and specificity was 93.85 and 98.21%, respectively. A grading system was established: grade 1: omental score ≤ 5, malignant rate 0%; grade 2: omental score 6-7, malignant rate ≤ 5.56%; grade 3: omental score 8--9, malignant rate ≤ 37.25%; grade 4: omental score ≥ 10, malignant rate ≥ 87.72. CONCLUSION: GOI-RADS had high sensitivity and specificity in the differential diagnosis of benign and malignant omental lesions. We believe that GOI-RADS will aid the diagnosis of omental diseases based on objective and accurate interpretation of ultrasound features, and also to promote the ultrasonography of omental diseases in clinical application.

Temporary reduction in fine particulate matter due to 'anthropogenic emissions switch-off' during COVID-19 lockdown in Indian cities.

The COVID-19 pandemic elicited a global response to limit associated mortality, with social distancing and lockdowns being imposed. In India, human activities were restricted from late March 2020. This 'anthropogenic emissions switch-off' presented an opportunity to investigate impacts of COVID-19 mitigation measures on ambient air quality in five Indian cities (Chennai, Delhi, Hyderabad, Kolkata, and Mumbai), using in-situ measurements from 2015 to 2020. For each year, we isolated, analysed and compared fine particulate matter (PM2.5) concentration data from 25 March to 11 May, to elucidate the effects of the lockdown. Like other global cities, we observed substantial reductions in PM2.5 concentrations, from 19 to 43% (Chennai), 41-53% (Delhi), 26-54% (Hyderabad), 24-36% (Kolkata), and 10-39% (Mumbai). Generally, cities with larger traffic volumes showed greater reductions. Aerosol loading decreased by 29% (Chennai), 11% (Delhi), 4% (Kolkata), and 1% (Mumbai) against 2019 data. Health and related economic impact assessments indicated 630 prevented premature deaths during lockdown across all five cities, valued at 0.69 billion USD. Improvements in air quality may be considered a temporary lockdown benefit as revitalising the economy could reverse this trend. Regulatory bodies must closely monitor air quality levels, which currently offer a baseline for future mitigation plans.

Psychometric Properties of the General Organizational Index (GOI): A Measure of Individualization and Quality Improvement to Complement Program Fidelity.

To assess the implementation of effective practices, mental health programs need standardized measures. The General Organizational Index (GOI), although widely used for this purpose, has received minimal psychometric research. For this study, we assessed psychometric properties of the GOI scale administered four times over 18 months during the implementation of a new program in 11 sites. The GOI scale demonstrated high levels of interrater reliability (.97), agreement between assessors on item ratings (86% overall), internal consistency (.77-.80 at three time points), sensitivity to change, and feasibility. We conclude that the GOI scale has acceptable psychometric properties, and its use may enhance implementation and research on evidence-based mental health practices.Trial registration: REK2015/2169. ClinicalTrials.gov Identifier: NCT03271242.

Correlation Between Allosteric and Orthosteric Sites.

Correlation between an allosteric site and its orthosteric site refers to the phenomenon that perturbations like ligand binding, mutation, or posttranslational modifications at the allosteric site leverage variation in the orthosteric site. Understanding this kind of correlation not only helps to disclose how information is transmitted in allosteric regulation but also provides clues for allosteric drug discovery. This chapter starts with an overview of correlation studies on allosteric and orthosteric sites and then introduces recent progress in evolutionary and simulation-based dynamic studies. Discussions and perspectives on future directions are also given.

Knowledge and Attitude Regarding Hepatitis B and C Among Blood Donors and Nondonors in North India.

BACKGROUND: The selection of a low-risk blood donor involves a dialogue between the trained medical staff and the volunteer blood donor, and this is where the knowledge of the prospective blood donor with regard to the risk factors for acquiring hepatitis B and C and the mode of spread through a blood transfusion is of utmost importance. Therefore, the study was conducted to assess the knowledge and attitude on hepatitis B and C with regard to blood donation, in the existing and the potential donor base. MATERIALS AND METHODS: This is a cross-sectional study conducted on 4000 participants, including 2000 blood donors and 2000 nondonors. The study tool was a pilot-tested, self-administered questionnaire, content and construct validated using Delphi methodology. RESULTS: The mean age of the study participants was 25.12 ± 8.43 years ranging from 18 to 60 years; 24.64 ± 8.31 years in donors and 25.61 ± 8.55 years in non-donors. The study included 69.8% males and 30.2% females, with 87.5% males and 12.6% females in donors and 52.1% males and 47.9% females in non-donors. Overall knowledge score was 51.02%, being 51.21% in donors and 50.84% in non-donors. Overall attitude score was 47.93%, being 47.09% in donors and 48.77% in non-donors. There was a low degree of significant linear correlation between knowledge and attitude in the study participants. CONCLUSION: Based on the results obtained in the study, it is evident that neither the existing level of knowledge nor the attitude of both donors and nondonors towards hepatitis B and C is adequate for being able to select a low-risk blood donor.

The Relationship between Green Organization Identity and Corporate Environmental Performance: The Mediating Role of Sustainability Exploration and Exploitation Innovation.

The link between green organizational identity (GOI) and corporate environmental performance (CEP) has been investigated, but existing studies have no consistent conclusion. A significant research gap remains regarding the mediating role of sustainability exploration innovation (SER), sustainability exploitation innovation (SEI), and the moderating role of government environmental regulation (GER). This study explored the relationship between GOI and CEP in a moderated meditation model which includes SER, SEI, and GER. Using structural equation modelling and bootstrap method based on data sets from of 380 Chinese companies, the results show that: (1) GOI promotes SER, thereby enhancing CEP; (2) GOI promotes SEI, thereby enhancing CEP; (3) GER can positively moderate the indirect effect of GOI on CEP via SER; (4) GER negatively moderate the indirect effect of GOI on CEP via SEI. These findings suggest that firms choose different innovative ways between SER and SEI to improve CEP which depends on different levels of GER in China.

Expected Number and Height Distribution of Critical Points of Smooth Isotropic Gaussian Random Fields.

We obtain formulae for the expected number and height distribution of critical points of smooth isotropic Gaussian random fields parameterized on Euclidean space or spheres of arbitrary dimension. The results hold in general in the sense that there are no restrictions on the covariance function of the field except for smoothness and isotropy. The results are based on a characterization of the distribution of the Hessian of the Gaussian field by means of the family of Gaussian orthogonally invariant (GOI) matrices, of which the Gaussian orthogonal ensemble (GOE) is a special case. The obtained formulae depend on the covariance function only through a single parameter (Euclidean space) or two parameters (spheres), and include the special boundary case of random Laplacian eigenfunctions.

Glioblastomas with copy number gains in EGFR and RNF139 show increased expressions of carbonic anhydrase genes transformed by ENO1.

BACKGROUND: Prominence of glycolysis in glioblastomas may be non-specific or a feature of oncogene-related subgroups (i.e. amplified EGFR, etc.). Relationships between amplified oncogenes and expressions of metabolic genes associated with glycolysis, directly or indirectly via pH, were therefore investigated. METHODS: Using multiplex ligation-dependent probe amplification, copy numbers (CN) of 78 oncogenes were quantified in 24 glioblastomas. Related expressions of metabolic genes encoding lactate dehydrogenases (LDHA, LDHC), carbonic anhydrases (CA3, CA12), monocarboxylate transporters (SLC16A3 or MCT4, SLC16A4 or MCT5), ATP citrate lyase (ACLY), glycogen synthase1 (GYS1), hypoxia inducible factor-1A (HIF1A), and enolase1 (ENO1) were determined in 22 by RT-qPCR. To obtain supra-glycolytic levels and adjust for heterogeneity, concurrent ENO1 expression was used to mathematically transform the expression levels of metabolic genes already normalized with delta-delta crossing threshold methodology. RESULTS: Positive correlations with EGFR occurred for all metabolic genes. Significant differences (Wilcoxon Rank Sum) for oncogene CN gains in tumors of at least 2.00-fold versus less than 2.00-fold occurred for EGFR with CA3's expression (p < 0.03) and for RNF139 with CA12 (p < 0.004). Increased CN of XIAP associated negatively. Tumors with less than 2.00-fold CN gains differed from those with gains for XIAP with CA12 (p < 0.05). Male gender associated with CA12 (p < 0.05). CONCLUSIONS: Glioblastomas with CN increases in EGFR had elevated CA3 expression. Similarly, tumors with RNF149 CN gains had elevated CA12 expression. GENERAL SIGNIFICANCE: In larger studies, subgroups of glioblastomas may emerge according to oncogene-related effects on glycolysis, such as control of pH via effects on carbonic anhydrases, with prognostic and treatment implications.

The syntaxin 31-induced gene, LESION SIMULATING DISEASE1 (LSD1), functions in Glycine max defense to the root parasite Heterodera glycines.

Experiments show the membrane fusion genes α soluble NSF attachment protein (α-SNAP) and syntaxin 31 (Gm-SYP38) contribute to the ability of Glycine max to defend itself from infection by the plant parasitic nematode Heterodera glycines. Accompanying their expression is the transcriptional activation of the defense genes ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1) and NONEXPRESSOR OF PR1 (NPR1) that function in salicylic acid (SA) signaling. These results implicate the added involvement of the antiapoptotic, environmental response gene LESION SIMULATING DISEASE1 (LSD1) in defense. Roots engineered to overexpress the G. max defense genes Gm-α-SNAP, SYP38, EDS1, NPR1, BOTRYTIS INDUCED KINASE1 (BIK1) and xyloglucan endotransglycosylase/hydrolase (XTH) in the susceptible genotype G. max[Williams 82/PI 518671] have induced Gm-LSD1 (Gm-LSD1-2) transcriptional activity. In reciprocal experiments, roots engineered to overexpress Gm-LSD1-2 in the susceptible genotype G. max[Williams 82/PI 518671] have induced levels of SYP38, EDS1, NPR1, BIK1 and XTH, but not α-SNAP prior to infection. In tests examining the role of Gm-LSD1-2 in defense, its overexpression results in ∼52 to 68% reduction in nematode parasitism. In contrast, RNA interference (RNAi) of Gm-LSD1-2 in the resistant genotype G. max[Peking/PI 548402] results in an 3.24-10.42 fold increased ability of H. glycines to parasitize. The results identify that Gm-LSD1-2 functions in the defense response of G. max to H. glycines parasitism. It is proposed that LSD1, as an antiapoptotic protein, may establish an environment whereby the protected, living plant cell could secrete materials in the vicinity of the parasitizing nematode to disarm it. After the targeted incapacitation of the nematode the parasitized cell succumbs to its targeted demise as the infected root region is becoming fortified.

CRISPR-Cas system enables fast and simple genome editing of industrial Saccharomyces cerevisiae strains.

There is a demand to develop 3rd generation biorefineries that integrate energy production with the production of higher value chemicals from renewable feedstocks. Here, robust and stress-tolerant industrial strains of Saccharomyces cerevisiae will be suitable production organisms. However, their genetic manipulation is challenging, as they are usually diploid or polyploid. Therefore, there is a need to develop more efficient genetic engineering tools. We applied a CRISPR-Cas9 system for genome editing of different industrial strains, and show simultaneous disruption of two alleles of a gene in several unrelated strains with the efficiency ranging between 65% and 78%. We also achieved simultaneous disruption and knock-in of a reporter gene, and demonstrate the applicability of the method by designing lactic acid-producing strains in a single transformation event, where insertion of a heterologous gene and disruption of two endogenous genes occurred simultaneously. Our study provides a foundation for efficient engineering of industrial yeast cell factories.

Polishing the craft of genetic diversity creation in directed evolution.

Genetic diversity creation is a core technology in directed evolution where a high quality mutant library is crucial to its success. Owing to its importance, the technology in genetic diversity creation has seen rapid development over the years and its application has diversified into other fields of scientific research. The advances in molecular cloning and mutagenesis since 2008 were reviewed. Specifically, new cloning techniques were classified based on their principles of complementary overhangs, homologous sequences, overlapping PCR and megaprimers and the advantages, drawbacks and performances of these methods were highlighted. New mutagenesis methods developed for random mutagenesis, focused mutagenesis and DNA recombination were surveyed. The technical requirements of these methods and the mutational spectra were compared and discussed with references to commonly used techniques. The trends of mutant library preparation were summarised. Challenges in genetic diversity creation were discussed with emphases on creating "smart" libraries, controlling the mutagenesis spectrum and specific challenges in each group of mutagenesis methods. An outline of the wider applications of genetic diversity creation includes genome engineering, viral evolution, metagenomics and a study of protein functions. The review ends with an outlook for genetic diversity creation and the prospective developments that can have future impact in this field.