High-throughput Phenotyping of Wheat Seminal Root Traits in a Breeding Context

Water availability is a major limiting factor for wheat (Triticum aestivum L.) in rain-fed agricultural systems worldwide. Root architecture has important functional implications for the timing and extent of soil water extraction, yet selection for root traits in wheat breeding programs has been largely limited due to the lack of suitable phenotyping methods. The aim of this research was to develop a low-cost high-throughput phenotyping method to facilitate selection for desirable root traits. We developed a method to assess ‘seminal root angle’ and ‘seminal root number’ in seedlings – two proxy traits associated to root architecture of mature wheat plants (1). The method involves measuring the angle between the first pair of seminal roots and the number of roots of wheat seedlings grown in transparent pots (Figure 1). Images captured at 5 to 10 days after sowing are analyzed to calculate seminal root angle and number. Performing this technique under “speed breeding” conditions (plants grown at a density of 600 plants / m2, under controlled temperature and constant light) allows the selection based on the desired root traits of up to 5 consecutive generations within 12 months. Alternatively, when focusing only on germplasm screening, up to 52 successive phenotypic assays can be conducted within 12 months. This approach has been shown to be highly reproducible, it requires little resource (time, space, and labour) and can be used to rapidly enrich breeding populations with desirable alleles for narrow root angle and a high number of seminal roots to indirectly target the selection of deeper root system with higher branching at depth. Such root characteristics are highly desirable in wheat to cope with the climate model projections, especially in summer rainfall dominant regions including some Australian, Indian, South American and African cropping regions, where winter crops mainly rely on deep stored water.

Integrating Rapid Phenotyping and Speed Breeding to Improve Stay-Green and Root Adaptation of Wheat in Changing, Water-Limited, Australian Environments

Temperatures have increased and in-crop rainfall decreased over recent decades in many parts of the Australian wheat cropping region. With these trends set to continue or intensify, improving crop adaptation in the face of climate change is particularly urgent in this, already drought-prone, cropping region. Importantly, improved performance under water-limitation must be achieved while retaining yield potential during more favourable seasons. A multi-trait-based approach to improve wheat yield and yield stability in the face of water-limitation and heat has been instigated in northern Australia using novel phenotyping techniques and a nested association mapping (NAM) approach. An innovative laboratory technique allows rapid root trait screening of hundreds of lines. Using soil grown seedlings, the method offers significant advantages over many other lab-based techniques. Another recently developed method allows novel stay-green traits to be quantified objectively for hundreds of genotypes in standard field trial plots. Field trials in multiple locations and seasons allow evaluation of targeted trait values and identification of superior germplasm. Traits, including yield and yield components are measured for hundreds of NAM lines in rain fed environments under various levels of water-limitation. To rapidly generate lines of interest, the University of Queensland “speed breeding” method is being employed, allowing up to 7 plant generations per annum. A NAM population of over 1000 wheat recombinant inbred lines has been progressed to the F5 generation within 18 months. Genotyping the NAM lines with the genome-wide DArTseq molecular marker system provides up to 40,000 markers. They are now being used for association mapping to validate QTL previously identified in bi-parental populations and to identify novel QTL for stay-green and root traits. We believe that combining the latest techniques in physiology, phenotyping, genetics and breeding will increase genetic progress toward improved adaptation to water-limited environments.

Stem anatomy and adventitious root formation in cuttings of Angophora, Corymbia and Eucalyptus

Many plantation eucalypts are difficult to propagate from cuttings, and their rooted cuttings often possess very few adventitious roots. We microscopically examined the stem anatomy of cuttings from 12 species of eucalypts and we determined whether adventitious root formation in auxin-treated cuttings of four species was limited to particular positions around the vascular tissue. Most species contained a central pith that was arranged in a four-pointed stellate pattern. The surrounding vascular tissue was also arranged in a stellate pattern near the shoot apex but it developed a more rectangular shape at the outer phloem as the stems enlarged radially. Adventitious roots formed at, or slightly peripheral to, the vascular cambium, and they formed at both the corners and the sides of the rectangular-shaped vascular tissue. The study highlighted that auxin-treated eucalypt cuttings can produce roots at multiple positions around the vascular tissue and so propagation methods can aim to produce more than four adventitious roots per rooted cutting. Higher numbers of adventitious roots could improve the root system symmetry, stability, survival and growth rate of clonal eucalypt trees. © 2015 by the authors; licensee MDPI, Basel, Switzerland.

Genomic regions influencing seminal root traits in barley

Water availability is a major limiting factor for crop production, making drought adaptation and its many component traits a desirable attribute of plant cultivars. Previous studies in cereal crops indicate that root traits expressed at early plant developmental stages, such as seminal root angle and root number, are associated with water extraction at different depths. Here, we conducted the first study to map seminal root traits in barley (Hordeum vulgare L.). Using a recently developed high-throughput phenotyping method, a panel of 30 barley genotypes and a doubled-haploid (DH) population (ND24260 × 'Flagship') comprising 330 lines genotyped with diversity array technology (DArT) markers were evaluated for seminal root angle (deviation from vertical) and root number under controlled environmental conditions. A high degree of phenotypic variation was observed in the panel of 30 genotypes: 13.5 to 82.2 and 3.6 to 6.9° for root angle and root number, respectively. A similar range was observed in the DH population: 16.4 to 70.5 and 3.6 to 6.5° for root angle and number, respectively. Seven quantitative trait loci (QTL) for seminal root traits (root angle, two QTL; root number, five QTL) were detected in the DH population. A major QTL influencing both root angle and root number (RAQ2/RNQ4) was positioned on chromosome 5HL. Across-species analysis identified 10 common genes underlying root trait QTL in barley, wheat (Triticum aestivum L.), and sorghum [Sorghum bicolor (L.) Moench]. Here, we provide insight into seminal root phenotypes and provide a first look at the genetics controlling these traits in barley.

Fine root dry matter relative to mango (Mangifera indica) tree scion size grafted on size-controlling rootstocks, is negatively related to scion growth rate

Knowledge of root dry matter (DM) allocation, in relation to differing vigour conferred by rootstock cultivars, is required to understand the structural relationships between rootstock and scion. We investigated the mass of roots (four size classes up to 23 mm diameter) by coring proximal to five polyembryonic mango rootstock cultivars known to differ in their effects on the vigour and productivity of scion cultivar ‘Kensington Pride’, in a field trial of 13-year-old trees. Significant differences in fine (<0.64 and 0.64–1.88 mm diameter) and small (1.88–7.50 mm) root DM contents were observed between rootstock cultivars. There was a complex relationship between the amount of feeder (fine and small size classes) roots and scion size (trunk cross sectional area, TCSA), with intermediate size trees on rootstock MYP having the most feeder roots, while the smallest trees, on the rootstock Vellaikulamban had the least of these roots. Across rootstock cultivars, tree vigour (TCSA growth rate) was negatively and significantly related to the ratio of fine root DM/scion TCSA, suggesting this may be a useful indicator of the vigour that different rootstocks confer on the scion. In contrast non-ratio root DM and scion TCSA results had no significant relationships. The significant rootstock effects on orchard root growth and tree size could not be predicted from earlier differences in nursery seedling vigour, nor did seedling vigour predict root DM allocation.

Highly heritable resistance to root-lesion nematode (Pratylenchus thornei) in Australian chickpea germplasm observed using an optimised glasshouse method and multi-environment trial analysis

Pratylenchus thornei is a root-lesion nematode (RLN) of economic significance in the grain growing regions of Australia. Chickpea (Cicer arietinum) is a significant legume crop grown throughout these regions, but previous testing found most cultivars were susceptible to P. thornei. Therefore, improved resistance to P. thornei is an important objective of the Australian chickpea breeding program. A glasshouse method was developed to assess resistance of chickpea lines to P. thornei, which requires relatively low labour and resource input, and hence is suited to routine adoption within a breeding program. Using this method, good differentiation of chickpea cultivars for P. thornei resistance was measured after 12 weeks. Nematode multiplication was higher for all genotypes than the unplanted control, but of the 47 cultivars and breeding lines tested, 17 exhibited partial resistance, allowing less than two fold multiplication. The relative differences in resistance identified using this method were highly heritable (0.69) and were validated against P. thornei data from seven field trials using a multi-environment trial analysis. Genetic correlations for cultivar resistance between the glasshouse and six of the field trials were high (>0.73). These results demonstrate that resistance to P. thornei in chickpea is highly heritable and can be effectively selected in a limited set of environments. The improved resistance found in a number of the newer chickpea cultivars tested shows that some advances have been made in the P. thornei resistance of Australian chickpea cultivars, and that further targeted breeding and selection should provide incremental improvements.

Expression of hyoscyamine 6 beta-hydroxylase in the root pericycle cells and accumulation of its product scopolamine in leaf and stem tissues of Datura metel L.

Hyoscyamine 60-hydroxylase (H6H: EC 1.14.11.11), a key enzyme at the terminal step of tropane alkaloid biosynthesis, converts hyoscyamine to scopolamine. The accumulation of scopolamine in different organs, in particular the aerial parts for storage, is subject to the expression of hyoscyamine 6-phydroxylase as well as its transport from the site of synthesis. To understand the molecular basis of this regulation, we have analyzed, in parallel, the relative levels of hyoscyamine and scopolamine, and the accumulation of H6H (both protein and transcript) in leaves, stems and roots of D. metel. The root, stem and leaf tissues all contain about 0.51-0.65 mg g(-1) dry weight of scopolamine. Hyoscyamine content was extremely low in leaf and stem tissues and was about 0.28 mg g(-1) dry weight in the root tissue. H6H protein and its transcript were found only in roots but not in the aerial parts viz. stems and leaves. The immunolocalization studies performed on leaf, stem, root as well as hairy root tissues showed that H6H was present only in the pericycle cells of young lateral and hairy roots. These studies suggest that the conversion of hyoscyamine to scopolamine takes place in the root pericycle cells, and the alkaloid biosynthesized in the roots gets translocated to the aerial parts in D. metel. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Lung cancer stem cells: The root of resistance

In the absence of specific treatable mutations, platinum-based chemotherapy remains the gold standard of treatment for lung cancer patients. However, 5-year survival rates remain poor due to the development of resistance and eventual relapse. Resistance to conventional cytotoxic therapies presents a significant clinical challenge in the treatment of this disease. The cancer stem cell (CSC) hypothesis suggests that tumors are arranged in a hierarchical structure, with the presence of a small subset of stem-like cells that are responsible for tumor initiation and growth. This CSC population has a number of key properties such as the ability to asymmetrically divide, differentiate and self-renew, in addition to having increased intrinsic resistance to therapy. While cytotoxic chemotherapy kills the bulk of tumor cells, CSCs are spared and have the ability to recapitulate the heterogenic tumor mass. The identification of lung CSCs and their role in tumor biology and treatment resistance may lead to innovative targeted therapies that may ultimately improve clinical outcomes in lung cancer patients. This review will focus on lung CSC markers, their role in resistance and their relevance as targets for future therapies.

Guided respiration mindfulness therapy: Development and evaluation of a brief therapist training program

The present paper describes the development and evaluation of a standardized multi-component therapist training program in guided respiration mindfulness therapy (GRMT). GMRT is a manual-based, experimental clinical intervention involving concentrated focus on sustained self-regulation of breathing, application of mindfulness to emergent somatic experience and relaxation. Therapists (n = 61) new to the approach attended a 2-day experiential workshop and were evaluated pre-post workshop for change in intervention knowledge, as well as change in mindfulness. These trainees also participated in post-workshop focus group sessions to explore perception of the intervention. A subset of 40 therapists participated in a second training component, and 14 of these were rated for competent delivery of the intervention during participation in a clinical trial. During training, therapists personally received the treatment giving the opportunity to assess treatment session (n = 283) impact on sense of wellbeing. Results indicated a brief focused training program can equip therapists with basic knowledge and skills required to deliver the standardized manual-based treatment. Qualitative analysis of focus group sessions showed that therapists endorsed the intervention for clinical use and found it personally beneficial. This research provides a foundation for further evaluation of clinical effectiveness of the intervention.

Interrelations of micro-organisms and mulberry II. Phyllosphere microflora and nitrogen fixation in leaf and root surfaces

The mulberry leaves were shown to harbour substantial populations of bacteria, streptomycetes, yeasts, and moulds. Azotobacter and Beijerinckia were observed to contribute to nearly 5 to 10 per cent of the bacterial population. When grown in water culture under sterile conditions, Azotobacter inoculation on the leaf or root surface was found to increase plant growth, dry wt, and nitrogen content of the mulberry. The beneficial effect of Azotobacter was largely influenced by the presence of a carbon source in the plant nutrient solution. The root inoculation in comparison to leaf application was found to confer greater benefits to the growing plant. The presence of carbohydrates and amino acids in the leaf leachates of mulberry was shown. The mutual beneficial nature of the association of the plant and Azotobacter has been brought to light.