The taxonomy of rhizobia: an overview

The taxonomy of rhizobia, bacteria capable of nodulating leguminous plants, has changed considerably over the last 20 years, with the original genus Rhizobium, a member of the alpha-Proteobacteria, now divided into several genera. The study of new geographically dispersed host plants, has been a source of many new species and is expected to yield many more. Here we provide an overview of the history of the rhizobia, but focus on the Rhizobium–Allorhizobium–Agrobacterium relationship. Finally, we review recent reports of nodulation and nitrogen fixation with legume hosts by bacteria that are outside the traditional rhizobial phylogenetic lineages. They include species of Methylobacterium and Devosia in the alpha- Proteobacteria and of Burkholderia and Ralstonia in the beta-Proteobacteria.     

Osmoregulation in the model organismEscherichia coli: genes governing the synthesis of glycine betaine and trehalose and their use in metabolic engineering of stress tolerance

Glycine betaine is known to be the preferred osmoprotectant in many bacteria, and glycine betaine accumulation has also been correlated with increased cold tolerance. Trehalose is often a minor osmoprotectant in bacteria and it is a major determinant for desiccation tolerance in many so-called anhydrobiotic organisms such as baker's yeast(Saccharomyces cerevisiae). Escherichia coli has two pathways for synthesis of these protective molecules; i.e., a two-step conversion of UDP-glucose and glucose-6-phosphate to trehalose and a two-step oxidation of externally-supplied choline to glycine betaine. The genes governing the choline-to-glycine betaine pathway have been studied inE. coli and several other bacteria and higher plants. The genes governing UDP-glucose-dependent trehalose synthesis have been studied inE. coli andS. cerevisiae. Because of their well-documented function in stress protection, glycine betaine and trehalose have been identified as targets for metabolic engineering of stress tolerance. Examples of this experimental approach include the expression of theE. coli betA andArthrobacter globiformis codA genes for glycine betaine synthesis in plants and distantly related bacteria, and the expression of theE. coli otsA and yeastTPS1 genes for trehalose synthesis in plants. The published data show that glycine betaine synthesis protects transgenic plants and phototrophic bacteria against stress caused by salt and cold. Trehalose synthesis has been reported to confer increased drought tolerance in transgenic plants, but it causes negative side effects which is of concern. Thus, the much-used model organismE. coli has now become a gene resource for metabolic engineering of stress tolerance.     

Constitutive expression of a meiotic recombination protein gene homolog, OsTOP6A1, from rice confers abiotic stress tolerance in transgenic Arabidopsis plants

Plant productivity is greatly influenced by various environmental stresses, such as high salinity and drought. Earlier, we reported the isolation of topoisomerase 6 homologs from rice and showed that over expression of OsTOP6A3 and OsTOP6B confers abiotic stress tolerance in transgenic Arabidopsis plants. In this study, we have assessed the function of nuclear-localized topoisomerase 6 subunit A homolog, OsTOP6A1, in transgenic Arabidopsis plants. The over expression of OsTOP6A1 in transgenic Arabidopsis plants driven by cauliflower mosaic virus-35S promoter resulted in pleiotropic effects on plant growth and development. The transgenic Arabidopsis plants showed reduced sensitivity to stress hormone, abscisic acid (ABA), and tolerance to high salinity and dehydration at the seed germination; seedling and adult stages as reflected by the percentage of germination, fresh weight of seedlings and leaf senescence assay, respectively. Concomitantly, the expression of many stress-responsive genes was enhanced under various stress conditions in transgenic Arabidopsis plants. Moreover, microarray analysis revealed that the expression of a large number of genes involved in various processes of plant growth and development and stress responses was altered in transgenic plants. Although AtSPO11-1, the homolog of OsTOP6A1 in Arabidopsis, has been implicated in meiotic recombination; the present study demonstrates possible additional role of OsTOP6A1 and provides an effective tool for engineering crop plants for tolerance to different environmental stresses. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Micropropagation of Ranunculus Asiaticus: A Review and Perspectives

Ranunculus asiaticus is an important ornamental species mainly cultivated in the countries surrounding the Mediterranean sea. So far the multiplication of this plant has been mainly carried out by seed and rhizome division; however, these systems present many drawbacks. Tissue culture is an attractive alternative for accelerated propagation of selected and indexed genotypes. In this paper we review the work carried out on in vitro culture of R. asiaticus and present a flow chart for its commercial production, using axillary budding and embryogenesis. Although the price of micropropagated plants is higher compared to traditional material (seedlings and rhizomes from seed populations), it should be considered that tissue cultured plants have a better rhizome yield per plant; moreover, the tissue culture approach allows to offer clonal material of selected lines.     

Biocontrol of a perennial legume,Sesbania punicea,using a florivorous weevil,Trichapion lativentre: weed population dynamics with a scarcity of seeds

Summary The establishment in South Africa of a florivorous, apionid weevil, Trichapion lativentre, on Sesbania punicea, a leguminous weed of South American origin, has reduced seed production of the plants by >98%. Surveys of the age structure and density of plants in infestations of S. punicea throughout South Africa have shown that the rate of recruitment of seedlings has drastically declined within a few years in many areas, due to the weevils. However, there has unexpectedly not been a corresponding decline in the density of mature plants in extant infestations of S. punicea. In spite of this, T. lativentre has curtailed the rate of spread of the weed into uninvaded habitats and has impeded reinvasion into areas cleared of infestations by mechanical means or by another complimentary biocontrol agent.     

Nickel hyperaccumulation as an anti-herbivore trait: considering the role of tolerance to damage

Metal hyperaccumulation is a striking trait exhibited by many plant species, but the evolutionary ecology of metal hyperaccumulation is poorly understood. It has been widely hypothesized that metal hyperaccumulation evolved to protect plants from herbivory. However, there is currently little evidence that metal hyperaccumulation enhances the fitness of plants in the presence of herbivory. In this study, we conducted a multi-factor greenhouse experiment to examine the effects of two soil nickel concentrations (unamended (0 μg/g) and nickel amended (600 μg/g)), and three levels of artificial damage (0, 10 and 50%) on the growth of plants from two populations of Thlaspi montanum var. montanum. We observed a significant interaction between soil nickel and artificial damage. An a posteriori analysis of this interaction revealed that the presence of nickel significantly improved the ability of T. montanum to tolerate the negative effects of intense damage. Our results indicate that metal hyperaccumulation could benefit T. montanum by increasing its tolerance to damage. This study suggests that there is a potential for the evolution of metal hyperaccumulation in response to intense herbivory on T. montanum.     

The role of the novel adenosine 5′-phosphosulfate reductase in regulation of sulfate assimilation of <Emphasis Type="Italic">Physcomitrella patens</Emphasis>

Sulfate assimilation provides reduced sulfur for the synthesis of the amino acids cysteine and methionine and for a range of other metabolites. The key step in control of plant sulfate assimilation is the reduction of adenosine 5′-phosphosulfate to sulfite. The enzyme catalyzing this reaction, adenosine 5′phosphosulfate reductase (APR), is found as an iron sulfur protein in plants, algae, and many bacteria. In the moss Physcomitrella patens, however, a novel isoform of the enzyme, APR-B, has recently been discovered lacking the co-factor. To assess the function of the novel APR-B we used homologous recombination to disrupt the corresponding gene in P. patens. The knock-out plants were able to grow on sulfate as a sole sulfur source and the content of low molecular weight thiols was not different from wild type plants or plants where APR was disrupted. However, when treated with low concentrations of cadmium the APR-B knockout plants were more sensitive than both wild type and APR knockouts. In wild type P. patens, the two APR isoforms were not affected by treatments that strongly regulate this enzyme in flowering plants. The data thus suggest that in P. patens APS reduction is not the major control step of sulfate assimilation.     

<Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> plants expressing human beta-defensin-2 are more resistant to fungal attack: functional homology between plant and human defensins

Human beta-defensin-2 (hBD-2) is a small antimicrobial peptide with potent activity against different Gram-negative bacteria and fungal/yeast species. Since human beta-defensins and plant defensins share structural homology, we set out to analyse whether there also exists a functional homology between these defensins of different eukaryotic kingdoms. To this end, we constructed a plant transformation vector harbouring the hBD-2 coding sequence, which we transformed to Arabidopsis thaliana plants, giving rise to A. thaliana plants indeed expressing hBD-2. Furthermore, we could demonstrate that this heterologously produced hBD-2 possesses antifungal activity in vitro. Finally, we could show that hBD-2 expressing A. thaliana plants are more resistant against the broad-spectrum fungal pathogen Botrytis cinerea as compared to untransformed A. thaliana plants, and that this resistance is correlated with the level of active hBD-2 produced in these transgenic plants. Hence, we demonstrated a functional homology, next to the already known structural homology, between defensins originating from different eukaryotic kingdoms. To our knowledge, this is the first time that this is specifically demonstrated for plant and mammalian defensins.     

Conservation In vitro of threatened plants—Progress in the past decade

Summary In vitro techniques have found increasing use in the conservation of threatened plants in recent years and this trend is likely to continue as more species face risk of extinction. The Micropropagation Unit at Royal Botanic Gardens, Kew, UK (RBG Kew) has an extensive collection of in vitro plants including many threatened species from throughout the world. The long history of the unit and the range of plants cultured have enabled considerable expertise to be amassed in identifying the problems and developing experimental strategies for propagation and conservation of threatened plants. While a large body of knowledge is available on the in vitro culture of plants, there are limited publications relating to threatened plant conservation. This review highlights the progress in in vitro culture and conservation of threatened plants in the past decade (1995–2005) and suggests future research directions. Works on non-threatened plants are also included wherever methods have applications in rare plant conservation. Recalcitrant plant materials collected from the wild or ex situ collections are difficult to grow in culture. Different methods of sterilization and other treatments to establish clean material for culture initiation are reviewed. Application of different culture methods for multiplication, and use of unconventional materials for rooting and transplantation are reviewed. As the available plant material for culture initiation is scarce and in many cases associated with inherent problems such as low viability and endogenous contamination, reliable protocols on multiplication, rooting, and storage methods are very important. In this context, photoautotrophic micropropagation has the potential for development as a routine method for the in vitro conservation of endangered plants. Long-term storage of material in culture is challenging and the potential applications of cryopreservation are significant in this area. Future conservation biotechnology research and its applications must be aimed at conserving highly threatened, mainly endemic, plants from conservation hotspots.     

Growth retardation of plants transformed by overexpression of NtFtsZ1-2 in Tobacco

We transformed tobacco plants (Mcotiana tabacum L, Xanthi) by introducing a sense construct ofNtFtsZ1-2. This tobacco nuclear gene encodes a chloroplast-localized homologue of FtsZ, the bacterial cell-division protein. The overexpressing plants contained enlarged chloroplasts in their leaf mesophyll cells. In the T₁ progeny, we observed three different phenotypes: 1 ) plants with cells containing many small chloroplasts, which was the same as for wild-type plants; 2) plants in which the celts contained one to three enlarged chloroplasts (severe type); and 3) plants whose cells contained a combination of many small chloroplasts and one to three enlarged chloroplasts (intermediate type). The outward appearance of the severe and intermediate types of transgenic plants did not differ noticeably from the wild-types. However, the severe-type plants were most retarded in their growth under both high- and low-light conditions, followed by the intermediate-types. Under medium levels of light, the two types of transgenic plants exhibited growth rates comparable to that of the wild types. Based on the overall results, we suggest that many small chloroplasts, rather than a few large chloroplasts, are required for efficient use of light energy in the mesophyll cells.     

Genetic determination of pappus part number in the annual hybridMicroseris B 87 (Asteraceae—Lactuceae)

The ChileanMicroseris pygmaea has a ten-part paleaceous pappus while the CalifornianM. bigelovii has five pappus parts on each achene. Hybrids between the two species have between five and ten pappus parts with averages below 7.5. Hybrid B 87 has an F 1 average value of 6.7 pappus parts. 140 F 2 plants were raised from this hybrid, and 12 F 3 families were obtained by selfing from F 2 plants. One larger F 4 family has been raised. Pappus part number in all of these is still canalized between 5 and 10. Variation within these limits is genetically determined by a quantitatively acting polygenic system. Modeling of this system suggests that a minimum of four, but probably not many more, genes are involved. This opens the possibility of a complete genetic analysis of the system.     

Polyphosphate kinase genes from full-scale activated sludge plants

The performance of enhanced biological phosphorus removal (EBPR) wastewater treatment processes depends on the presence of bacteria that accumulate large quantities of polyphosphate. One such group of bacteria has been identified and named Candidatus Accumulibacter phosphatis. Accumulibacter-like bacteria are abundant in many EBPR plants, but not much is known about their community or population ecology. In this study, we used the polyphosphate kinase gene (ppk1) as a high-resolution genetic marker to study population structure in activated sludge. Ppk1 genes were amplified from samples collected from full-scale wastewater treatment plants of different configurations. Clone libraries were constructed using primers targeting highly conserved regions of ppk1, to retrieve these genes from activated sludge plants that did, and did not, perform EBPR. Comparative sequence analysis revealed that ppk1 fragments were retrieved from organisms affiliated with the Accumulibacter cluster from EBPR plants but not from a plant that did not perform EBPR. A new set of more specific primers was designed and validated to amplify a 1,100 bp ppk1 fragment from Accumulibacter-like bacteria. Our results suggest that the Accumulibacter cluster has finer-scale architecture than previously revealed by 16S ribosomal RNA-based analyses. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Short term temperature drops do not enhance cold tolerance

To successfully transplant agricultural species in the spring, prior hardening is of great significance. Low, non-freezing temperature increases cold tolerance in many species. Also, diurnal temperature drops have been suggested to improve cold tolerance, as assessed by ultrastructural studies after short term freezing of leaf discs. Pre-treatment with lower day than night temperature prior to hardening has also been reported to enhance cold resistance in winter rape. This study investigated the effect of temperature drops on cold resistance of different species. In contrast to a period of continuous low temperature, short diurnal temperature drops did not enhance cold tolerance in Arabidopsis, swede, white cabbage or pea, compared to control plants. Exposure to low temperature of 6°C for 6 days increased cold tolerance by 2–5°C compared to plants exposed to diurnal temperature drops or control plants. Pre-treatment with diurnal temperature drops in the entire growth period prior to hardening with constant low temperature did not give any additional hardening in swede and pea. In conclusion, by freeze testing of whole plants under controlled conditions we have found no evidence supporting the hypothesis that diurnal temperature drops improve cold tolerance. However, temperature drops reduce plants size like shown earlier for a number of other species, and thus is a tool to produce compact, robust plants.     

Involvement of AtLAC15 in lignin synthesis in seeds and in root elongation of Arabidopsis

Laccase, EC 1.10.3.2 or p-diphenol:dioxygen oxidoreductase, has been proposed to be involved in lignin synthesis in plants based on its in vitro enzymatic activity and a close correlation with the lignification process in plants. Despite many years of research, genetic evidence for the role of laccase in lignin synthesis is still missing. By screening mutants available for the annotated laccase gene family in Arabidopsis, we identified two mutants for a single laccase gene, AtLAC15 (At5g48100) with a pale brown or yellow seed coat which resembled the transparent testa (tt) mutant phenotype. A chemical component analysis revealed that the mutant seeds had nearly a 30% decrease in extractable lignin content and a 59% increase in soluble proanthocyanidin or condensed tannin compared with wild-type seeds. In an in vitro enzyme assay, the developing mutant seeds showed a significant reduction in polymerization activity of coniferyl alcohol in the absence of H₂O₂. Among the dimers formed in the in vitro assay using developing wild-type seeds, 23% of the linkages were β-O-4 which resembles the major linkages formed in native lignin. The evidence strongly supports that AtLAC15 is involved in lignin synthesis in plants. To our knowledge, this is the first genetic evidence for the role of laccase in lignin synthesis. Changes in seed coat permeability, seed germination and root elongation were also observed in the mutant.Electronic Supplementary Material Supplementary material is available to authorised users in the online version of this article at .     

Pappus part number in annual species ofMicroseris (Compositae,Cichoriaceae)

All North American annual species of the genusMicroseris have a five-part pappus, the one South American annual,M. pygmaea, has ten pappus parts. The pappus develops over a constant number of ten provascular bundles with or without inhibition between alternate sites of pappus development. Each natural population contains a predictable proportion of achenes with aberrant pappus part numbers. Hybridization betweenM. bigelovii (5 parts) andM. pygmaea results in F 1 and F 2 plants with many aberrant achenes. In each plant either five or ten can be shown to be the basic number with aberrant numbers following a Poisson distribution for numbers added to 5 or deleted from 10. Occasional plants show no basic number but have a random distribution of numbers about an intermediate mean. The evolutionary genetics of this character is discussed.     

Morphological and anatomical evaluation of adult and juvenile leaves of olive plants

The olive tree (Olea europaea L.), like many other woody plants, has a long juvenile period in which the plant is not able to produce flowers. Knowledge of the moment when the plant is capable of flowering is important for breeding programs and also for determining the physiological basis for sexual reproductive behavior, but currently the only indicator of that moment is the actual flowering. In many species, the juvenile-to-adult phase shift includes changes in leaf structure known as heteroblasty, that is, varied form of successive leaves on the same plant. Some differences have been observed between juvenile and adult olive leaves, particularly in size and form, but to our knowledge, no complete systematic study has been carried out. In this research, we measured size, morphology and anatomy for juvenile and adult leaves of olive plants grown from seeds. Differences were found in most of the parameters studied, including leaf size, form, mesophyll thickness, layers of palisade parenchyma and quantity of peltate trichomes, which were generally significant but overlapping between the two leaf types. The most consistent and striking difference was the presence of an organized layer of subepidermal cells only in the abaxial mesophyll of adult leaves. This characteristic could be a simple and effective criterion of phase change in the olive tree.