Composition and Distribution of Free Amino Acids in Flatpea (Lathyrus sylvestris L.) as Influenced by Water Deficit and Plant Age

Drought-stressed flatpea (Lathyrus sylvestris L.) plants from8 to 22 weeks old were analysed for nitrogen, soluble proteinand free amino acids. An increase in nitrogen and free aminoacid concentrations and a decrease in soluble protein levelwere observed in roots of plants up to 16 weeks old. The cumulativeconcentration of free amino acids increased with drought stress.Tissue concentrations of 2, 4-diaminobutyric acid (1.6–2.6%of the dry weight) were highest in leaves. Levels increasedsteadily, nearly doubling, in leaves and stems between weeks10 and 16. Levels in drought-stressed leaves were, on average,11.9% higher than those of controls. Estimated concentrationsof a mixture of 4-aminobutyric acid and an unknown amino acidwere highest in stems, increased in this tissue with age andtended to increase in stems and leaves and decrease in rootsin response to water deficit. Levels of the mixture of homoserineand another unidentified amino acid were not influenced by ageor water status of the plants. Root concentrations of asparagine,arginine, glutamine, aspartate, and another prominent, unidentifiedamino acid increased with plant age and reached a peak at thetime of flowering (14 to 18 weeks). Only the concentration ofthe unknown compound was elevated following drought stress.Concentrations of valine, isoleucine, leucine, phenylalanine,and methionine also increased during this period and were elevatedin drought-stressed plants. Proline levels increased with plantage and drought stress, but proline accounted for only about10% of the total free amino acids in the drought-stressed plants. Key words: 2, 4-Diaminobutyric acid, drought, flatpea     

Crossing the Tasman Sea: Inferring the introduction history of Litoria aurea and Litoria raniformis (Anura: Hylidae) from Australia into New Zealand

Abstract The amphibian fauna of New Zealand consists of three native species (Leiopelma spp.), and three Litoria species introduced from Australia in the last 140 years. We conducted a molecular phylogeographical study that aimed to identify the Australian origins of two species, Litoria aurea and Litoria raniformis. We used partial sequences of the mitochondrial cytochrome oxidase I (cox1) gene from 59 specimens sampled from across the range of both species to identify the probable source populations for the New Zealand introductions, and to describe the current genetic diversity among New Zealand Litoria populations. Our genetic data suggest that L. aurea was introduced into the North Island of New Zealand from two regions in Australia, once from the northern part of coastal New South Wales and once from the southern part of coastal New South Wales. Our data indicate that L. raniformis introductions originated from the Melbourne region of southern Victoria and once established in the South Island of New Zealand, the species subsequently spread throughout both islands. In addition, we found a distinct haplotype in L. raniformis from Tasmania that strongly suggests, contrary to earlier reports, that this species was not introduced into New Zealand from Tasmania. Finally, we identified two very distinctive mitochondrial lineages of L. raniformis within the mainland Australia distribution, which may be previously unrecognized species.     

Ferricrocin synthesis in Magnaporthe grisea and its role in pathogenicity in rice

Iron is an essential element for the growth of nearly all organisms. In order to overcome the problem of its low bioavailability, microorganisms (including fungi) secrete siderophores, high-affinity iron chelators. As the acquisition of iron is also a key step in infection processes, siderophores have been considered as potential virulence factors in several host–pathogen interactions. Most fungi produce siderophores of the hydroxamate-type, which are synthesized by non-ribosomal peptide synthetases (NRPSs). Magnaporthe grisea , the causal agent of rice blast disease, produces ferricrocin as intracellular storage siderophore and excretes coprogens. In the M. grisea genome we identified SSM1 , an NRPS gene, and a gene encoding an l -ornithine N5-monooxygenase ( OMO1 ) that is clustered with SSM1 and responsible for catalysing the first step in siderophore biosynthesis, the N⁵ hydroxylation of ornithine. Disruption of SSM1 confirmed that the gene encodes ferricrocin synthetase. Pathogenicity of these mutants towards rice was reduced, suggesting a role of this siderophore in pathogenicity of M. grisea .     

Inadvertent gene silencing of argininosuccinate synthase (bcass1) in Botrytis cinerea by the pLOB1 vector system

For several years, researchers working on the plant pathogen Botrytis cinerea and a number of other related fungi have routinely used the pLOB1 vector system, based on hygromycin resistance, under the control of the Aspergillus nidulans oliC promoter and what was reported to be the β‐tubulin (tubA) terminator. Recently, it has been demonstrated that this vector contains a 446‐bp portion of the B. cinerea argininosuccinate synthase gene (bcass1) rather than the tubA terminator. As argininosuccinate synthase is essential for the production of l ‐arginine, inadvertent gene silencing of bcass1 may result in partial l ‐arginine auxotrophy and, indeed, may lead to altered phenotypes in planta. In this article, we report our findings relating to possible problems arising from this incorrect plasmid construction. As an absolute baseline, gene disruption of bcass1 was carried out and generated a strict auxotroph, unable to grow without exogenous arginine supplementation. The knockout displayed an alteration in host range in planta, showing a reduction in pathogenicity on strawberries, French bean leaves and tomatoes, but maintained wild‐type growth on grape, which is in accordance with the reported arginine availability in such tissues. Deliberate gene silencing of bcass1 mirrored these effects, with strongly silenced lines showing reduced virulence. The degree of silencing as seen by partial auxotrophy was correlated with an observed reduction in virulence. We also showed that inadvertent silencing of bcass1 is possible when using the pLOB1 vector or derivatives thereof. Partial arginine auxotrophy and concomitant reductions in virulence were triggered in approximately 6% of transformants obtained when expressing enhanced green fluorescent protein, luciferase, monomeric red fluorescent protein or β‐glucuronidase using the pLOB1‐based expression system, which inadvertently contains 446 bp of the bcass1 coding sequence. We recommend the testing of transformants obtained using this vector system for arginine auxotrophy in order to provide assurance that any observed effects on the development or virulence are a result of the desired genetic alteration rather than accidental bcass1 silencing.