The development of powerful “omics” technologies has enabled researchers to identify many genes of interest for which comprehensive functional analyses are highly desirable. However, the production ...of lines which ectopically express recombinant genes, or those in which endogenous genes are knocked down via stable transformation, remains a major bottleneck for the association between genetics and gene function in monocotyledonous crops. Methods of effective DNA transfer into regenerable cells of immature embryos from cereals by means of Agrobacterium tumefaciens have been modified in a stepwise manner. The effect of particular improvement measures has often not been significantly evident, whereas their combined implementation has resulted in meaningful advances. Here, we provide updated protocols for the Agrobacterium-mediated generation of stably transgenic barley, wheat, triticale and maize. Based upon these methods, several hundred independent transgenic lines have been delivered, with efficiencies of inoculated embryos leading to stably transgenic plants reaching 86% in barley, 10% in wheat, 4% in triticale, and 24% in maize.
Plants producing antisense or double-stranded RNA molecules that target specific genes of eukaryotic pests or pathogens can become protected from their attack. This beneficial effect was also ...reported for plant–fungus interactions and is believed to reflect uptake of the RNAs by the fungus via an as yet unknown mechanism, followed by target gene silencing. Here we report that wheat plants pre-infected with Barley stripe mosaic virus (BSMV) strains containing antisense sequences against target genes of the Fusarium head blight (FHB) fungus F. culmorum caused a reduction of corresponding transcript levels in the pathogen and reduced disease symptoms. Stable transgenic wheat plants carrying an RNAi hairpin construct against the β-1, 3-glucan synthase gene FcGls1 of F. culmorum or a triple combination of FcGls1 with two additional, pre-tested target genes also showed enhanced FHB resistance in leaf and spike inoculation assays under greenhouse and near-field conditions, respectively. Microscopic evaluation of F. culmorum development in plants transiently or stably expressing FcGls1 silencing constructs revealed aberrant, swollen fungal hyphae, indicating severe hyphal cell wall defects. The results lead us to propose host-induced gene silencing (HIGS) as a plant protection approach that may also be applicable to highly FHB-susceptible wheat genotypes.
Summary
Maize (corn) is one of the most widely grown cereal crops globally. Fungal diseases of maize cause significant economic damage by reducing maize yields and by increasing input costs for ...disease management. The most sustainable control of maize diseases is through the release and planting of maize cultivars with durable disease resistance. The wheat gene Lr34 provides durable and partial field resistance against multiple fungal diseases of wheat, including three wheat rust pathogens and wheat powdery mildew. Because of its unique qualities, Lr34 became a cornerstone in many wheat disease resistance programmes. The Lr34 resistance is encoded by a rare variant of an ATP‐binding cassette (ABC) transporter that evolved after wheat domestication. An Lr34‐like disease resistance phenotype has not been reported in other cereal species, including maize. Here, we transformed the Lr34 resistance gene into the maize hybrid Hi‐II. Lr34‐expressing maize plants showed increased resistance against the biotrophic fungal disease common rust and the hemi‐biotrophic disease northern corn leaf blight. Furthermore, the Lr34‐expressing maize plants developed a late leaf tip necrosis phenotype, without negative impact on plant growth. With this and previous reports, it could be shown that Lr34 is effective against various biotrophic and hemi‐biotrophic diseases that collectively parasitize all major cereal crop species.
The majority of endometrial and cervical cancers present with abnormal vaginal bleeding but only a small proportion of women suffering from vaginal bleeding actually have such a cancer. A simple, ...operator‐independent and accurate test to correctly identify women presenting with abnormal bleeding as a consequence of endometrial or cervical cancer is urgently required. We have recently developed and validated the WID‐qEC test, which assesses DNA methylation of ZSCAN12 and GYPC via real‐time PCR, to triage women with symptoms suggestive of endometrial cancer using ThinPrep‐based liquid cytology samples. Here, we investigated whether the WID‐qEC test can additionally identify women with cervical cancer. Moreover, we evaluate the test's applicability in a SurePath‐based hospital‐cohort by comparing its ability to detect endometrial and cervical cancer to cytology. In a set of 23 cervical cancer cases and 28 matched controls the receiver operating characteristic (ROC) area under the curve (AUC) is 0.99 (95% confidence interval CI: 0.97‐1.00) with a sensitivity and specificity of 100% and 92.9%, respectively. Amongst the hospital‐cohort (n = 330), the ROC AUC is 0.99 (95% CI: 0.98‐1) with a sensitivity and specificity of 100% and 82.5% for the WID‐qEC test, respectively, and 33.3% and 96.9% for cytology (considering PAP IV/V as positive). Our data suggest that the WID‐qEC test detects both endometrial and cervical cancer with high accuracy.
What's new?
While abnormal vaginal bleeding is a presenting symptom of endometrial and cervical cancers, only a small proportion of women who present with vaginal bleeding have endometrial or cervical cancer. Currently, the tests used to triage women with abnormal bleeding, such as ultrasound or cytology, are subjective and have modest accuracy. Here, the authors demonstrate that a real‐time PCR‐based test, which assesses DNA methylation at three gene regions using a cervical or vaginal sample, is able to identify 100% of women who have cervical or endometrial cancer with a high specificity (>80%), irrespective of the sample collection system.
Introduction
Green tea is associated with decreased risk for cardiovascular disease and stroke. Matcha is a special kind of powdered green tea known for its use in the Japanese tea ceremony. Due to ...its influence on lipoprotein parameters, it has been postulated to exert antiatherogenic effects. This study investigates whether it modulates the high‐density lipoprotein (HDL) function and thereby influences the atherogenic process in an animal model with a strong influence on humans' situation.
Methods and Results
After a pretreatment phase based on a standard diet, 10 female New Zealand White (NZW) rabbits are fed a high‐fat diet for 20 weeks. The treatment group is additionally administered 1% matcha during the whole experiment. Long‐term matcha treatment leads to lowered HDL cholesterol, impaired cholesterol transport manifested by reduced in vitro cholesterol efflux capacity, reduced cholesteryl ester transfer protein (CETP)‐mediated cholesterol ester (CE) transfer between HDL and triglyceride‐rich particles, and reduced macrophage‐specific in vivo transfer, where ian increased absorption of cholesterol in the liver but a decreased secretion into bile is observed. Pulse wave velocity, assessed by nuclear magnetic resonance, is increased in matcha‐treated animals, and a similar trend is observed for atherosclerotic lesion formation.
Conclusion
Long‐term matcha green tea treatment of hypercholesterolemic rabbits cause impaired reverse cholesterol transport and increased vascular stiffness, and susceptibility for atherosclerotic lesion development.
The reverse transport of cholesterol is responsible for the body's regulation of excess cholesterol. In rabbits, an animal model closest to the human situation in terms of lipoprotein metabolism, long‐term adjuvant therapy with matcha green tea alters this mechanism. The resulting metabolic changes are associated with atherosclerosis and exacerbate the clinical symptoms of cardiovascular disease.
Infection of maize (Zea mays) plants with the corn smut fungus Ustilago maydis leads to the formation of large tumors on the stem, leaves and inflorescences. In this biotrophic interaction, plant ...defense responses are actively suppressed by the pathogen, and previous transcriptome analyses of infected maize plants showed massive and stage-specific changes in host gene expression during disease progression. To identify maize genes that are functionally involved in the interaction with U. maydis, we adapted a virus-induced gene silencing (VIGS) system based on the brome mosaic virus (BMV) for maize. Conditions were established that allowed successful U. maydis infection of BMV-preinfected maize plants. This set-up enabled quantification of VIGS and its impact on U. maydis infection using a quantitative real-time PCR (qRT-PCR)-based readout. In proof-of-principle experiments, an U. maydis-induced terpene synthase was shown to negatively regulate disease development while a protein involved in cell death inhibition was required for full virulence of U. maydis. The results suggest that this system is a versatile tool for the rapid identification of maize genes that determine compatibility with U. maydis.
Ustilago maydis is a biotrophic pathogen causing maize (Zea mays) smut disease. Transcriptome profiling of infected maize plants indicated that a gene encoding a putative cystatin (CC9) is induced ...upon penetration by U. maydis wild type. By contrast, cc9 is not induced after infection with the U. maydis effector mutant ∆pep1, which elicits massive plant defenses. Silencing of cc9 resulted in a strongly induced maize defense gene expression and a hypersensitive response to U. maydis wild-type infection. Consequently, fungal colonization was strongly reduced in cc9-silenced plants, while recombinant CC9 prevented salicylic acid (SA)-induced defenses. Protease activity profiling revealed a strong induction of maize Cys proteases in SA-treated leaves, which could be inhibited by addition of CC9. Transgenic maize plants overexpressing cc9-mCherry showed an apoplastic localization of CC9. The transgenic plants showed a block in Cys protease activity and SA-dependent gene expression. Moreover, activated apoplastic Cys proteases induced SA-associated defense gene expression in naïve plants, which could be suppressed by CC9. We show that apoplastic Cys proteases play a pivotal role in maize defense signaling. Moreover, we identified cystatin CC9 as a novel compatibility factor that suppresses Cys protease activity to allow biotrophic interaction of maize with the fungal pathogen U. maydis.
Objective
To explore ongoing symptoms, unmet needs, psychological wellbeing, self‐efficacy and overall health status in survivors of prostate cancer.
Patients and Methods
An invitation to participate ...in a postal questionnaire survey was sent to 546 men, diagnosed with prostate cancer 9–24 months previously at two UK cancer centres. The study group comprised men who had been subject to a range of treatments: surgery, radiotherapy, hormone therapy and active surveillance. The questionnaire included measures of prostate‐related quality of life (Expanded Prostate cancer Index Composite 26‐item version, EPIC‐26); unmet needs (Supportive Care Needs Survey 34‐item version, SCNS‐SF34); anxiety and depression (Hospital Anxiety and Depression Scale, HADS), self‐efficacy (modified Self‐efficacy Scale), health status (EuroQol 5D, EQ‐5D) and satisfaction with care (questions developed for this study). A single reminder was sent to non‐responders after 3 weeks. Data were analysed by age, co‐morbidities, and treatment group.
Results
In all, 316 men completed questionnaires (64.1% response rate). Overall satisfaction with follow‐up care was high, but was lower for psychosocial than physical aspects of care. Urinary, bowel, and sexual functioning was reported as a moderate/big problem in the last month for 15.2% (n = 48), 5.1% (n = 16), and 36.5% (n = 105) men, respectively. The most commonly reported moderate/high unmet needs related to changes in sexual feelings/relationships, managing fear of recurrence/uncertainty, and concerns about the worries of significant others. It was found that 17% of men (51/307) reported potentially moderate‐to‐severe levels of anxiety and 10.2% (32/308) reported moderate‐to‐severe levels of depression. The presence of problematic side‐effects was associated with higher psychological morbidity, poorer self‐efficacy, greater unmet needs, and poorer overall health status.
Conclusion
While some men report relatively few problems after prostate cancer treatment, this study highlights important physical and psycho‐social issues for a significant minority of survivors of prostate cancer. Strategies for identifying those men with on‐going problems, alongside new interventions and models of care, tailored to individual needs, are needed to improve quality of life.
Screening for ovarian cancer (OC) in women at high risk consists of a combination of carbohydrate antigen 125 (CA125) and transvaginal ultrasound, despite their low sensitivity and specificity. This ...could be improved by the combination of several biomarkers, which has been shown in average risk patients but has not been investigated until now in female BRCA mutation carriers.
Using a multiplex, bead-based, immunoassay system, we analyzed the concentrations of leptin, prolactin, osteopontin, insulin-like growth factor II, macrophage inhibitory factor, CA125 and human epididymis antigen 4 in 26 healthy wild type women, 26 healthy BRCA1 mutation carriers, 28 wildtype OC patients and 26 OC patients with BRCA1 mutation.
Using the ROC analysis, we found a high overall sensitivity of 94.3% in differentiating healthy controls from OC patients with comparable results in the wildtype subgroup (sensitivity 92.8%, AUC = 0.988; p = 5.2e-14) as well as in BRCA1 mutation carriers (sensitivity 95.2%, AUC = 0.978; p = 1.7e-15) at an overall specificity of 92.3%. The used algorithm also allowed to identify healthy BRCA1 mutation carriers when compared to healthy wildtype women (sensitivity 88.4%, specificity 80.7%, AUC = 0.895; p = 6e-08), while this was less pronounced in patients with OC (sensitivity 66.7%, specificity 67.8%, AUC = 0.724; p = 0.00065).
We have developed an algorithm, which can differentiate between healthy women and OC patients and have for the first time shown, that such an algorithm can also be used in BRCA mutation carriers. To clarify a suggested benefit to the existing early detection program, large prospective trials with mainly early stage OC cases are warranted.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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