Evaluation of resistance reaction of maize germplasm to common foliar diseases in Kenya

Use of resistance varieties is the most practical method of managing crop diseases. There is a great variation in terms of resistance reaction to diseases among the various maize germplasm and with the liberization of seed market, the sector has witnessed proliferation of massive introduction of new varieties whose reaction to diseases cannot be ascertained. This study was conducted to evaluate the reaction of maize varieties to northern leaf blight (NLB), common maize rust, gray leaf spot (GLS) and maize streak disease (MSD). The experiment was conducted at Kabete Field Station, University of Nairobi for two seasons namely short rains and long rains. The germplasm was bought from the commercial seed stockists and the landraces obtained from KARI Katumani and from farmers. The diseases were assessed by monitoring and scoring for disease incidence and severity. The appropriate scoring keys were used for determination of disease severity. All the varieties showed symptoms of the four diseases in both seasons but the intensity of the diseases differed significantly among the different varieties. Disease incidence was highest for common rust with a mean of 14.29% for the variety DH04, while disease incidence was highest (19.21%) for northern leaf blight in season two for Kinyanya which is a landrace. Gray leaf spot and the common smuts had the lowest mean incidence ranging from 0 to 0.25% for common smut and 0 to 2.6% for gray leaf spot. Season two had comparatively higher disease incidence means compared to season one. Meteorological data showed that season two had more rains and this explains the reasons behind this. Though all the varieties screened were found to be affected by the diseases to various levels, the varieties displayed significant differences in the disease incidence and severity. This shows that use of resistance varieties should be considered in the management of maize diseases. Focus should also be on pyramiding genes for resistance in the breeding programmes to develop varieties with multiple resistance to different diseases. Key words: Disease incidence and severity, disease score, symptoms, varietie

Development Methodology of the Novel Endoscopic Stone Treatment Step 2/A Training/Assessment Curriculum and a Roadmap on Developing Hands-on Training Curriculums in Future: An International Collaborative Work by European Association of Urology Sections

Background: Basic simulation training in endourology was established with the endoscopic stone treatment step 1 (EST-s1), which is now recognized worldwide for training and examination. Following on from EST-s1, the endoscopic stone treatment step 2 (EST-s2) was started by the European Association of Urology (EAU) sections. Objective: We describe the methodology used in the development of EST-s2 assessment curriculum. Materials and Methods: The "full-life cycle curriculum development" template was followed for curriculum development, focusing on intermediate training of EST protocol with complex endourologic tasks. A cognitive task analysis (CTA) was run in accordance with EAU Urolithiasis guidelines. The protocol and its details underwent a first consensus by Delphi method with EAU Urolithiasis Section experts in March 2017. Once the outcome and metrics were decided, curriculum development was carried out. Purpose-built stones were developed, and simulator system requirement was defined. Preliminary testing was done in European Urology Residents Education Programme 2019 and in phase five the protocol was finalized with full tutor instruction sheet. Results: The EST-s2/A curriculum development took 38 months and involved EAU Uro-technology and urolithiasis sections with coordination from the European School of Urology training group. Starting from the initial CTA, a 1277-word revision with preliminary task description was produced. Nine intermediate skills were identified and included in the final training protocol. The training content and session evaluations were carried out by 26 experts and 16 final year trainees, respectively. Although the experts agreed that EST-s2/A protocol was well structured (96%), covered the complex endourologic maneuvers (92%), and was useful to optimize and improve hands-on-training (HoT) sessions (92%), the overall evaluation was scored 4.25/5 by trainees. Conclusion: We describe the development methodology for intermediate EST curriculum, which also provides a roadmap on developing other HoT protocols in future. Patients Summary: In this report we described the development of the novel intermediate training curriculum for EST, called EST-s2, which took 3 years of collaborative work inside the EAU. This article is aimed to strengthen the standards in curriculum development and clearly describe the background of this new EAU official endourology protocol

CRISPR-Cas9 Mediated Epitope Tagging Provides Accurate and Versatile Assessment of Myocardin

Objective- Unreliable antibodies often hinder the accurate detection of an endogenous protein, and this is particularly true for the cardiac and smooth muscle cofactor, MYOCD (myocardin). Accordingly, the mouse Myocd locus was targeted with 2 independent epitope tags for the unambiguous expression, localization, and activity of MYOCD protein. Approach and Results- 3cCRISPR (3-component clustered regularly interspaced short palindromic repeat) was used to engineer a carboxyl-terminal 3×FLAG or 3×HA epitope tag in mouse embryos. Western blotting with antibodies to each tag revealed a MYOCD protein product of ≈150 kDa, a size considerably larger than that reported in virtually all publications. MYOCD protein was most abundant in some adult smooth muscle-containing tissues with surprisingly low-level expression in the heart. Both alleles of Myocd are active in aorta because a 2-fold increase in protein was seen in mice homozygous versus heterozygous for FLAG-tagged Myocd. ChIP (chromatin immunoprecipitation)-quantitative polymerase chain reaction studies provide proof-of-principle data demonstrating the utility of this mouse line in conducting genome-wide ChIP-seq studies to ascertain the full complement of MYOCD-dependent target genes in vivo. Although FLAG-tagged MYOCD protein was undetectable in sections of adult mouse tissues, low-passaged vascular smooth muscle cells exhibited expected nuclear localization. Conclusions- This report validates new mouse models for analyzing MYOCD protein expression, localization, and binding activity in vivo and highlights the need for rigorous authentication of antibodies in biomedical research

CRISPR-Cas9 Mediated Epitope Tagging Provides Accurate and Versatile Assessment of Myocardin--Brief Report

OBJECTIVE: Unreliable antibodies often hinder the accurate detection of an endogenous protein, and this is particularly true for the cardiac and smooth muscle cofactor, MYOCD (myocardin). Accordingly, the mouse Myocd locus was targeted with 2 independent epitope tags for the unambiguous expression, localization, and activity of MYOCD protein. APPROACH AND RESULTS: 3cCRISPR (3-component clustered regularly interspaced short palindromic repeat) was used to engineer a carboxyl-terminal 3xFLAG or 3xHA epitope tag in mouse embryos. Western blotting with antibodies to each tag revealed a protein product of approximately 150 kDa, a size considerably larger than that reported in virtually all publications. MYOCD protein was most abundant in some adult smooth muscle-containing tissues with surprisingly low-level expression in the heart. Both alleles of Myocd are active in aorta because a 2-fold increase in protein was seen in mice homozygous versus heterozygous for FLAG-tagged Myocd. ChIP-quantitative polymerase chain reaction studies provide proof-of-principle data demonstrating the utility of this mouse line in conducting genome-wide ChIP-seq studies to ascertain the full complement of MYOCD-dependent target genes in vivo. Although FLAG-tagged MYOCD protein was undetectable in sections of adult mouse tissues, low-passaged vascular smooth muscle cells exhibited expected nuclear localization. CONCLUSIONS: This report validates new mouse models for analyzing MYOCD protein expression, localization, and binding activity in vivo and highlights the need for rigorous authentication of antibodies in biomedical research

Retinoid-Induced Expression and Activity of an Immediate Early Tumor Suppressor Gene in Vascular Smooth Muscle Cells

Retinoids are used clinically to treat a number of hyper-proliferative disorders and have been shown in experimental animals to attenuate vascular occlusive diseases, presumably through nuclear receptors bound to retinoic acid response elements (RARE) located in target genes. Here, we show that natural or synthetic retinoids rapidly induce mRNA and protein expression of a specific isoform of A-Kinase Anchoring Protein 12 (AKAP12β) in cultured smooth muscle cells (SMC) as well as the intact vessel wall. Expression kinetics and actinomycin D studies indicate Akap12β is a retinoid-induced, immediate-early gene. Akap12β promoter analyses reveal a conserved RARE mildly induced with atRA in a region that exhibits hyper-acetylation. Immunofluorescence microscopy and protein kinase A (PKA) regulatory subunit overlay assays in SMC suggest a physical association between AKAP12β and PKA following retinoid treatment. Consistent with its designation as a tumor suppressor, inducible expression of AKAP12β attenuates SMC growth in vitro. Further, immunohistochemistry studies establish marked decreases in AKAP12 expression in experimentally-injured vessels of mice as well as atheromatous lesions in humans. Collectively, these results demonstrate a novel role for retinoids in the induction of an AKAP tumor suppressor that blocks vascular SMC growth thus providing new molecular insight into how retiniods may exert their anti-proliferative effects in the injured vessel wall

Biocontrol-based strategies for improving soil health and managing plant-parasitic nematodes in coffee production

Open Access JournalCoffee is an important commodity for Kenya, where production is steadily declining, despite a global rise in demand. Of the various constraints affecting production, plant-parasitic nematodes are a significant, but often overlooked, threat. As a perennial crop, treating plantations once infected with nematodes becomes difficult. The current study evaluated the drenching application of two biocontrol agents, Trichoderma asperellum and Purpureocillium lilacinum, for their nematode control efficacy, as well as their impact on the soil nematode community structure on mature, established coffee trees in Kenya. Seven Arabica coffee field trials were conducted over two years on trees of various ages. All the fields were heavily infested with Meloidogyne hapla, the first report of the species on coffee in Kenya. Both fungal biocontrol agents were detected endophytically infecting roots and recovered from soil but not until six months after initial applications. The population densities of M. hapla had significantly declined in roots of treated trees 12 months after the initial application, although soil nematode density data were similar across treatments. Based upon the maturity index and the Shannon index, treatment with T. asperellum led to improved soil health conditions and enrichment of diversity in the microbial community. Application of P. lilacinum, in particular, led to an increased abundance of fungivorous nematodes, especially Aphelenchus spp., for which P. lilacinum would appear to be a preferred food source. The soils in the trials were all stressed and denuded, however, which likely delayed the impact of such treatments or detection of any differences between treatments using indices, such as the functional metabolic footprint, over the period of study. A longer period of study would therefore likely provide a better indication of treatment benefits. The current study positively demonstrates, however, the potential for using biologically based options for the environmentally and climate-smart management of nematode threats in a sustainable manner on established, mature coffee plantations

A quantitatively-modeled homozygosity mapping algorithm, qHomozygosityMapping, utilizing whole genome single nucleotide polymorphism genotyping data

Homozygosity mapping is a powerful procedure that is capable of detecting recessive disease-causing genes in a few patients from families with a history of inbreeding. We report here a homozygosity mapping algorithm for high-density single nucleotide polymorphism arrays that is able to (i) correct genotyping errors, (ii) search for autozygous segments genome-wide through regions with runs of homozygous SNPs, (iii) check the validity of the inbreeding history, and (iv) calculate the probability of the disease-causing gene being located in the regions identified. The genotyping error correction restored an average of 94.2% of the total length of all regions with run of homozygous SNPs, and 99.9% of the total length of them that were longer than 2 cM. At the end of the analysis, we would know the probability that regions identified contain a disease-causing gene, and we would be able to determine how much effort should be devoted to scrutinizing the regions. We confirmed the power of this algorithm using 6 patients with Siiyama-type α1-antitrypsin deficiency, a rare autosomal recessive disease in Japan. Our procedure will accelerate the identification of disease-causing genes using high-density SNP array data

Multi-institutional Evaluation of Producing and Testing a Novel 3D-Printed Laparoscopic Trainer

To create, distribute, and evaluate the efficacy of a portable, cost effective 3D-printed laparoscopic trainer for surgical skills development.Objective: To create, distribute, and evaluate the efficacy of a portable, cost-effective 3D-printed laparoscopic trainer for surgical skills development. Methods: The UCI Trainer (UCiT) laparoscopic simulator was developed via computer-aided designs (CAD), which were used to 3D-print the UCiT. Once assembled, a tablet computer with a rear-facing camera was attached for video and optics. Four institutions were sent the UCiT CAD files with a 3D-printer and instructions for UCiT assembly. For a comparison of the UCiT to a standard trainer, peg transfer and intracorporeal knot tying skills were accessed. These tasks were scored, and participants were asked to rate their experience with the trainers. Lastly, a questionnaire was given to individuals who 3D-printed and assembled the UCiT. Results: We recruited 25 urologists; none had any 3D-printing experience. The cost of printing each trainer was $26.50 USD. Each institution used the Apple iPad for optics. Six of eight participants assembled the UCiT in < 45 minutes, and rated assembly as somewhat easy. On objective scoring, participants performed tasks equally well on the UCiT vs the conventional trainer. On subjective scoring, the conventional trainer provided a significantly better experience vs the UCiT; however, all reported that the UCiT was useful for surgical education. Conclusion: The UCiT is a low cost, portable training tool that is easy to assemble and use. UCiT provided a platform whereby participants performed laparoscopic tasks equal to performing the same tasks on the more expensive, nonportable standard trainer