Effect of Stock Plant Growing Medium and Density upon a Cutting Propagation System for Tea Tree, Melaleuca alternifolia.

To offer a viable alternative to seedling deployment of tea tree, clones will require the development of an efficient, robust, and vegetative propagation system for the large number of plants needed for plantations (i.e., typically 33,000 plants/ha). This study investigated the productivity of an intensive management system for tea tree stock plants and rooted cuttings grown in a subtropical environment (Lismore, NSW, Australia). Three stock plant densities (30, 100, and 200 plants/m2) were tested in coir and potting mix media (consisting of peat+perlite+vermiculite), with 11 settings of cuttings undertaken between April 2019 and March 2020. All stock plants in each media type survived 11 harvests and remained productive; however after 13 months, many plants in the coir media, appeared chlorotic and showed symptoms of iron deficiency. Rooting and cutting survival rates using the mini cutting technique were high, ranging from a maximum mean monthly setting value of 87.7% ± 4 at 84 days post-setting in potting mix, to a minimum of 80.4% ± 3.7 in coir. The most productive treatment was at high stock plant density in potting mix which had the potential to produce 13,440 plants/year/m2. Overall coir appeared less productive, but the pattern of difference among treatments was similar. For the highest system productivity, it is recommended to grow stock plants in potting mix at high densities and modulate temperatures to between 18 °C and 28 °C. Late spring and early summer were the best time for harvesting and setting tea tree mini cuttings in the subtropics.

Development of Selection Indices for Improvement of Seed Yield and Lipid Composition in Bambara Groundnut (Vigna subterranea (L.) Verdc.).

The underutilised grain legume bambara groundnut (Vigna subterranea) has the potential to contribute significantly to nutritional security. However, the lack of commercial cultivars has hindered its wider adoption and utilisation as a food source. The development of competitive cultivars is impeded by (1) lack of systematic data describing variation in nutritional composition within the gene pool, and (2) a poor understanding of how concentrations of different nutritional components interact. In this study, we analysed seed lipid and protein concentration and lipid composition within a collection of 100 lines representing the global gene pool. Seed protein and lipid varied over twofold with a normal distribution, but no significant statistical correlation was detected between the two components. Seed lipid concentration (4.2-8.8 g/100 g) is primarily determined by the proportion of oleic acid (r2 = 0.45). Yield and composition data for a subset of 40 lines were then used to test selection parameters for high yielding, high lipid breeding lines. From five selection indices tested using 15 scenarios, an index based on the seed number, seed weight, and oleic acid yielded a >50% expected increase in each of the mean values of seed number, pod dry weight, seed dry weight, and seed size, as well as an expected 7% increase in seed lipid concentration.

Complex Patterns of Cannabinoid Alkyl Side-Chain Inheritance in Cannabis.

The cannabinoid alkyl side-chain represents an important pharmacophore, where genetic targeting of alkyl homologs has the potential to provide enhanced forms of Cannabis for biopharmaceutical manufacture. Delta(9)-tetrahydrocannabinolic acid (THCA) and cannabidiolic acid (CBDA) synthase genes govern dicyclic (CBDA) and tricyclic (THCA) cannabinoid composition. However, the inheritance of alkyl side-chain length has not been resolved, and few studies have investigated the contributions and interactions between cannabinoid synthesis pathway loci. To examine the inheritance of chemical phenotype (chemotype), THCAS and CBDAS genotypes were scored and alkyl cannabinoid segregation analysed in 210 F2 progeny derived from a cross between two Cannabis chemotypes divergent for alkyl and cyclic cannabinoids. Inheritance patterns of F2 progeny were non-Gaussian and deviated from Mendelian expectations. However, discrete alkyl cannabinoid segregation patterns consistent with digenic as well as epistatic modes of inheritance were observed among F2 THCAS and CBDAS genotypes. These results suggest linkage between cannabinoid pathway loci and highlight the need for further detailed characterisation of cannabinoid inheritance to facilitate metabolic engineering of chemically elite germplasm.

Remobilization and fate of sulphur in mustard.

BACKGROUND AND AIMS: Sulphur (S) is an essential macronutrient involved in numerous metabolic pathways required for plant growth. Crops of the plant family Brassicaceae require more S compared with other crops for optimum growth and yield, with most S ultimately sequestered in the mature seeds as the storage proteins cruciferin and napin, along with the unique S-rich secondary metabolite glucosinolate (GSL). It is well established that S assimilation primarily takes place in the shoots rather than roots, and that sulphate is the major form in which S is transported and stored in plants. We carried out a developmental S audit to establish the net fluxes of S in two lines of Brassica juncea mustard where seed GSL content differed but resulted in no yield penalty. METHODS: We quantified S pools (sulphate, GSL and total S) in different organs at multiple growth stages until maturity, which also allowed us to test the hypothesis that leaf S, accumulated as a primary S sink, becomes remobilized as a secondary source to meet the requirements of GSL as the dominant seed S sink. KEY RESULTS: Maximum plant sulphate accumulation had occurred by floral initiation in both lines, at which time most of the sulphate was found in the leaves, confirming its role as the primary S sink. Up to 52 % of total sulphate accumulated by the low-GSL plants was lost through senesced leaves. In contrast, S from senescing leaves of the high-GSL line was remobilized to other tissues, with GSL accumulating in the seed from commencement of silique filling until maturity. CONCLUSION: We have established that leaf S compounds that accumulated as primary S sinks at early developmental stages in condiment type B. juncea become remobilized as a secondary S source to meet the demand for GSL as the dominant seed S sink at maturity.

Seed glucosinolate yield is maximized by higher rates of sulfur nutrition than required for seed yield in condiment mustard (Brassica juncea L.).

Brassica crops require high amounts of inorganic sulfur (S) for optimum yield, and are characterized by the synthesis of S-rich glucosinolates (GSL). Although it is well established that seed and GSL yield can be increased by S fertilizer, the detailed relationship between S supply as primary source and the harvestable sinks of seed GSL and storage proteins is poorly understood. We tested the hypothesis that Brassica juncea mustard seed acts as a secondary S sink, and so require a higher rate of S to achieve maximum seed GSL compared to rates required to attain maximum seed biomass. Our experimental strategy involved comparing responses to available S for seed biomass, GSL, and protein. This was carried out in a protected environment using sand culture for a high-GSL condiment-type homozygous B. juncea genotype. A low-GSL canola-type was used as a control, in order to establish a base-line of response. Significantly more S was required to achieve maximum seed GSL than was required to achieve maximum seed mass. Total seed protein content was not significantly affected by increased S. The high-GSL line appeared to have an efficient mechanism of S supply to the secondary S sink, given the observed increase in seed S with increased S availability. From a practical point of view, increases in seed GSL with S availability suggests that S fertilizer rates should be optimized for maximum seed GSL yield, rather that optimizing for seed yield, as occurs for most other crops.

Developmental Plasticity of the Major Alkyl Cannabinoid Chemotypes in a Diverse Cannabis Genetic Resource Collection.

Cannabis is a chemically diverse domesticated plant genus which produces a unique class of biologically active secondary metabolites referred to as cannabinoids. The affinity and selectivity of cannabinoids to targets of the human endocannabinoid system depend on alkyl side chain length, and these structural-activity relationships can be utilized for the development of novel therapeutics. Accurate early screening of germplasm has the potential to accelerate selection of chemical phenotypes (chemotypes) for pharmacological exploitation. However, limited attempts have been made to characterize the plasticity of alkyl cannabinoid composition in different plant tissues and throughout development. A chemotypic diversity panel comprised of 99 individuals from 20 Cannabis populations sourced from the Ecofibre Global Germplasm Collection (ecofibre.com.au and anandahemp.com) was used to examine alkyl cannabinoid variation across vegetative, flowering and maturation stages. A wide range of di-/tri-cyclic as well as C3-/C5-alkyl cannabinoid composition was observed between plants. Chemotype at the vegetative and flowering stages was found to be predictive of chemotype at maturation, indicating a low level of plasticity in cannabinoid composition. Chemometric cluster analysis based on composition data from all three developmental stages categorized alkyl cannabinoid chemotypes into three classes. Our results suggest that more extensive chemical and genetic characterization of the Cannabis genepool could facilitate the metabolic engineering of alkyl cannabinoid chemotypes.

Genotypic Variation in Wheat Flour Lysophospholipids.

Lysophospholipids (LPLs) are the most abundant polar lipids in wheat endosperm and naturally complex with amylose, affecting starch physicochemical properties. We analyzed LPLs in wheat flour from 58 cultivars which differ by grain hardness using liquid chromatography mass spectrometry (LCMS). There were significant differences in LPL content between cultivars, demonstrating that genotype rather than environment contributes most to the total variance in wheat endosperm LPLs. Polar lipids such as LPLs may play a role in grain hardness through their interaction with puroindoline proteins, however, no strong correlation between kernel hardness and LPLs was detected. This may reflect the location of LPLs within the starch granule as opposed to the puroindoline proteins outside starch granules. LPLs may have an indirect relationship with kernel hardness as they could share the same origin as polar lipids that interact with puroindoline on the starch granule surface.

GC-MS method validation and levels of methyl eugenol in a diverse range of tea tree (Melaleuca alternifolia) oils.

Tea tree oil distilled from Melaleuca alternifolia has widespread use in the cosmetic industry as an antimicrobial as well as for other functions in topical products. Concerns were first raised by the European Commission's Scientific Committee on Consumer Products in 2004 about the level of the potentially carcinogenic phenylpropanoid compound methyl eugenol in tea tree oil. Limits on oil content in different types of cosmetic products were set based on a reported upper level of 0.9% methyl eugenol in the oil. A previous publication indicated that these levels were based on oil from a Melaleuca species not used in the commercial production of oil. Even the highest recorded levels in Melaleuca alternifolia, the overwhelmingly most common species used, were ∼15 times less than this, meaning that more oil could be safely used in the products. The current study, including details on methodology and reproducibility, extends that work across a suite of 57 plantation-sourced oils from a range of geographical locations and production years, as well as many Australian and international commercial oils. Lower levels of methyl eugenol in oils of known provenance were confirmed, with a recorded range of 160-552 ppm and a mean of 337 ppm. Analysis of variance showed methyl eugenol levels in Australian plantation oils to be correlated to the geographical region but not to the year of production. Average methyl eugenol levels in commercial oils were significantly lower, and these samples were divided into an authentic group and a group that were suspected of being adulterated based on an independent test. Authentic commercial oils had similar levels of methyl eugenol to Australian provenance material, whilst the oils classed as suspect had significantly lower levels.

Single-cell-type quantitative proteomic and ionomic analysis of epidermal bladder cells from the halophyte model plant Mesembryanthemum crystallinum to identify salt-responsive proteins.

BACKGROUND: Epidermal bladder cells (EBC) are large single-celled, specialized, and modified trichomes found on the aerial parts of the halophyte Mesembryanthemum crystallinum. Recent development of a simple but high throughput technique to extract the contents from these cells has provided an opportunity to conduct detailed single-cell-type analyses of their molecular characteristics at high resolution to gain insight into the role of these cells in the salt tolerance of the plant. RESULTS: In this study, we carry out large-scale complementary quantitative proteomic studies using both a label (DIGE) and label-free (GeLC-MS) approach to identify salt-responsive proteins in the EBC extract. Additionally we perform an ionomics analysis (ICP-MS) to follow changes in the amounts of 27 different elements. Using these methods, we were able to identify 54 proteins and nine elements that showed statistically significant changes in the EBC from salt-treated plants. GO enrichment analysis identified a large number of transport proteins but also proteins involved in photosynthesis, primary metabolism and Crassulacean acid metabolism (CAM). Validation of results by western blot, confocal microscopy and enzyme analysis helped to strengthen findings and further our understanding into the role of these specialized cells. As expected EBC accumulated large quantities of sodium, however, the most abundant element was chloride suggesting the sequestration of this ion into the EBC vacuole is just as important for salt tolerance. CONCLUSIONS: This single-cell type omics approach shows that epidermal bladder cells of M. crystallinum are metabolically active modified trichomes, with primary metabolism supporting cell growth, ion accumulation, compatible solute synthesis and CAM. Data are available via ProteomeXchange with identifier PXD004045.

Novel Applications for Oxalate-Phosphate-Amine Metal-Organic-Frameworks (OPA-MOFs): Can an Iron-Based OPA-MOF Be Used as Slow-Release Fertilizer?

A porous iron-based oxalate-phosphate-amine metal-organic framework material (OPA-MOF) was investigated as a microbially-induced slow-release nitrogen (N) and phosphorus (P) fertilizer. Seedling growth, grain yields, nutrient uptake of wheat plants, and soil dynamics in incubated soil, were investigated using OPA-MOF vs standard P (triple-superphosphate) and N (urea) fertilizers in an acidic Ferralsol at two application rates (equivalent 120 and 40 kg N ha(-1)). While urea hydrolysis in the OPA-MOF treatment was rapid, conversion of ammonium to nitrate was significantly inhibited compared to urea treatment. Reduced wheat growth in OPA-MOF treatments was not caused by N-deficiency, but by limited P-bioavailability. Two likely reasons were slow P-mobilisation from the OPA-MOF or rapid P-binding in the acid soil. P-uptake and yield in OPA-MOF treatments were significantly higher than in nil-P controls, but significantly lower than in conventionally-fertilised plants. OPA-MOF showed potential as enhanced efficiency N fertilizer. However, as P-bioavailability was insufficient to meet plant demands, further work should determine if P-availability may be enhanced in alkaline soils, or whether central ions other than Fe, forming the inorganic metal-P framework in the MOF, may act as a more effective P-source in acid soils.

Pectin Methylesterase genes influence solid wood properties of Eucalyptus pilularis.

This association study of Eucalyptus pilularis populations provides empirical evidence for the role of Pectin Methylesterase (PME) in influencing solid wood characteristics of Eucalyptus. PME6 was primarily associated with the shrinkage and collapse of drying timber, which are phenotypic traits consistent with the role of pectin as a hydrophilic polysaccharide. PME7 was primarily associated with cellulose and pulp yield traits and had an inverse correlation with lignin content. Selection of specific alleles in these genes may be important for improving trees as sources of high-quality wood products. A heterozygote advantage was postulated for the PME7 loci and, in combination with haplotype blocks, may explain the absence of a homozygous class at all single-nucleotide polymorphisms investigated in this gene.