Genetically engineered resistance to organophosphate herbicides provides a new scoreable and selectable marker system for transgenic plants

Organophosphate hydrolase (OPH, E.C. 3.1.8.1; encoded by the bacterial opd gene) provides a new scoreable and selectable genetic marker system for use in plant cell culture and regenerated plant tissue. OPH hydrolyzes a wide range of substrates that produce visually detectable products, which can be readily quantified in biological tissues. A variety of different OP compounds, both herbicides and pesticides, have been identified as acceptable enzymatic substrates, which can be used to generate transgenic markers for various types of plant tissues. For example, transgenic leaf tissue was easily differentiated from non-transgenic tissue by a simple fluorescent assay utilizing the OP insecticide coroxon. Transformed callus and intact whole seed could be easily distinguished from non-transformed tissue using novel non-destructive methods which allowed callus or seeds to grow and/or to germinate after phenotypic scoring with non-herbicidal OP insecticides such as paraoxon. In addition to being used as a scoreable phenotypic markers with various OP pesticides, the OP compounds Haloxon and Bensulide (Bensumec-4LF) were effective as positive selection agents for callus and germinating seeds.     

Bioassay determination of the distribution of imidacloprid in potato plants: implications to resistance development

Soil-applied imidacloprid exhibits exceptional efficacy as a systemic insecticide against the Colorado potato beetle, Leptinotarsa decemlineata (Say). An uneven distribution of the chemical within potato plants could result in differential concentrations, which may allow for discrimination between genotypes of varying susceptibility. In this study, susceptible and tolerant larvae were fed leaves from the lower, middle, and upper canopy of treated and untreated plants to characterize within-plant distribution of imidacloprid at 4, 6, 8, 10, 12, and 14 wk after planting. Significant differences in larval mortality and development indicated that the concentration of imidacloprid was unevenly distributed in the potato foliage during 6-14 wk after planting. The concentration of imidacloprid was lowest in the younger tissues of the upper leaves and highest in the older, lower leaves. At 6 wk, a time when the postdiapause beetles are colonizing potato fields, the lower concentration in upper leaves was toxic to susceptible larvae but did not kill a substantial portion of the tolerant larvae. Results suggest that higher concentrations of imidacloprid in the lower canopy leaves may act as a toxic barrier to colonizing susceptible beetles but may allow more tolerant individuals to reach the upper canopy with lower concentrations. Possible scenarios of how different concentrations of the systemic insecticide could influence the rate of resistance development are discussed.     

Possible implications of insulin resistance and glucose metabolism in Alzheimer's disease pathogenesis

Type 2 diabetes mellitus (DM) appears to be a significant risk factor for Alzheimer disease (AD). Insulin and insulin-like growth factor-1 (IGF-1) also have intense effects in the central nervous system (CNS), regulating key processes such as neuronal survival and longevity, as well as learning and memory. Hyperglycaemia induces increased peripheral utilization of insulin, resulting in reduced insulin transport into the brain. Whereas the density of brain insulin receptor decreases during age, IGF-1 receptor increases, suggesting that specific insulin-mediated signals is involved in aging and possibly in cognitive decline. Molecular mechanisms that protect CNS neurons against β-amyloid-derived-diffusible ligands (ADDL), responsible for synaptic deterioration underlying AD memory failure, have been identified. The protection mechanism does not involve simple competition between ADDLs and insulin, but rather it is signalling dependent down-regulation of ADDL-binding sites. Defective insulin signalling make neurons energy deficient and vulnerable to oxidizing or other metabolic insults and impairs synaptic plasticity. In fact, destruction of mitochondria, by oxidation of a dynamic-like transporter protein, may cause synapse loss in AD. Moreover, interaction between Aβ and τ proteins could be cause of neuronal loss. Hyperinsulinaemia as well as complete lack of insulin result in increased τ phosphorylation, leading to an imbalance of insulin-regulated τ kinases and phosphatates. However, amyloid peptides accumulation is currently seen as a key step in the pathogenesis of AD. Inflammation interacts with processing and deposit of β-amyloid. Chronic hyperinsulinemia may exacerbate inflammatory responses and increase markers of oxidative stress. In addition, insulin appears to act as 'neuromodulator', influencing release and reuptake of neurotransmitters, and improving learning and memory. Thus, experimental and clinical evidence show that insulin action influences cerebral functions. In this paper, we reviewed several mechanisms by which insulin may affect pathophysiology in AD.     

The fitness costs to plants of resistance to pathogens

It has long been predicted that genes giving resistance to pathogens impose a cost on the fitness of plants. A new study has shown this to be true for one resistance gene in Arabidopsis. This raises intriguing theoretical and practical questions about how generally the results apply and how such costs are controlled in plants carrying resistance genes to several different pathogens.     

The genetic architecture of disease resistance in plants and the maintenance of recombination by parasites

Parasites represent strong selection on host populations because they are ubiquitous and can drastically reduce host fitness. It has been hypothesized that parasite selection could explain the widespread occurrence of recombination because it is a coevolving force that favours new genetic combinations in the host. A review of deterministic models for the maintenance of recombination reveals that for recombination to be favoured, multiple genes that interact with each other must be under selection. To evaluate whether parasite selection can explain the maintenance of recombination, we review 85 studies that investigated the genetic architecture of plant disease resistance and discuss whether they conform to the requirements that emerge from theoretical models. General characteristics of disease resistance in plants and problems in evaluating resistance experimentally are also discussed. We found strong evidence that disease resistance in plants is determined by multiple loci. Furthermore, in most cases where loci were tested for interactions, epistasis between loci that affect resistance was found. However, we found weak support for the idea that specific allelic combinations determine resistance to different host genotypes and there was little data on whether epistasis between resistance genes is negative or positive. Thus, the current data indicate that it is possible that parasite selection can favour recombination, but more studies in natural populations that specifically address the nature of the interactions between resistance genes are necessary. The data summarized here suggest that disease resistance is a complex trait and that environmental effects and fitness trade-offs should be considered in future models of the coevolutionary dynamics of host and parasites.     

The evolving genetic landscape of congenital disorders of glycosylation

Congenital Disorders of Glycosylation (CDG) are an expanding and complex group of rare genetic disorders caused by defects in the glycosylation of proteins and lipids. The genetic spectrum of CDG is extremely broad with mutations in over 140 genes leading to a wide variety of symptoms ranging from mild to severe and life-threatening. There has been an expansion in the genetic complexity of CDG in recent years. More specifically several examples of alternate phenotypes in recessive forms of CDG and new types of CDG following an autosomal dominant inheritance pattern have been identified. In addition, novel genetic mechanisms such as expansion repeats have been reported and several already known disorders have been classified as CDG as their pathophysiology was better elucidated. Furthermore, we consider the future and outlook of CDG genetics, with a focus on exploration of the non-coding genome using whole genome sequencing, RNA-seq and multi-omics technology.     

The effect of herbicides on the root system of wheat plants

Three commercial herbicides, at the rates normally recommended for selective weed control in cereal crops, deformed the roots of spring wheat plants grown in sand culture. Affected roots produced large numbers of short swollen lateral branches which later grew on to form tassel-like bundles of side roots. Only the mecoprop and MCPA components of the mixtures were responsible for the deformities; ioxynil and dicamba had no effect. Mecoprop applied to the rooting medium severely deformed the roots but foliar application also resulted in some abnormalities. The effects of the herbicides on root and shoot dry weights were not correlated with their deformative effects.     

The costs of evolving resistance in heterogeneous parasite environments

The evolution of host resistance to parasites, shaped by associated fitness costs, is crucial for epidemiology and maintenance of genetic diversity. Selection imposed by multiple parasites could be a particularly strong constraint, as hosts either accumulate costs of multiple specific resistances or evolve a more costly general resistance mechanism. We used experimental evolution to test how parasite heterogeneity influences the evolution of host resistance. We show that bacterial host populations evolved specific resistance to local bacteriophage parasites, regardless of whether they were in single or multiple-phage environments, and that hosts evolving with multiple phages were no more resistant to novel phages than those evolving with single phages. However, hosts from multiple-phage environments paid a higher cost, in terms of population growth in the absence of phage, for their evolved specific resistances than those from single-phage environments. Given that in nature host populations face selection pressures from multiple parasite strains and species, our results suggest that costs may be even more critical in shaping the evolution of resistance than previously thought. Furthermore, our results highlight that a better understanding of resistance costs under combined control strategies could lead to a more 'evolution-resistant' treatment of disease.     

Evolving strategies for the genetic engineering of herbicide resistance in plants

Developments in plant genetic engineering technology will shortly permit the commercial introduction of transgenic crop varieties resistant to a number of non-selective herbicides. High levels of tolerance have been achieved both by overexpression of a target protein and by modification of that target to an insensitive form. However the results of preliminary trials suggest that in some instances the yield penalty for such genetic alterations will be prohibitive. An alternative strategy, based on the transfer and expression of a gene encoding a herbicide-detoxifying enzyme, appears to offer high resistance levels at low metabolic cost and is expected to assume increasing importance, although it may not prove suitable for all herbicides.     

Phylogenetic perspectives on nodulation: evolving views of plants and symbiotic bacteria

Phylogenetic studies are contributing greatly to our knowledge of relationships on both sides of the plant–bacteria nodulation symbiosis. Multiple origins of nodulation (perhaps even within the legume family) appear likely. However, all nodulating flowering plants are more closely related than previously suspected, suggesting that the predisposition to nodulate might have arisen only once. Phylogenies of 16S rRNA genes highlight the evolutionary diversity of symbiotic bacteria and appear to rule out any broad coevolution with their plant hosts, but high levels of gene transfer might obscure the relevant pattern. The origins of nodulation, and the extent to which developmental programs are conserved in nodules remain unclear, but an improved understanding of the relationships between nodulin genes is providing some clues.     

The problem of durable resistance of plants to different pests

Different hypotheses concerning durable plant resistance against different pest were tested: 1) resistance is weak and polygenetically controlled; 2) resistance depends on "residual effect" of oligogenes that were overcome by pests. Contrast pair of plants and pests were used in experiments: wheat, barley--facultative parasite Bipolaris sorokiniana Shoem., wheat--obligate parasite Puccinia recondita Rob. ex Desm., sorghum--greenbug Schizaphis graminum Rond. Differential interaction between parasite and host plant resulted in their increased compatibility under long reproduction of parasite on resistant varieties were regarded as criteria of quick overcoming of resistance. The results did not support any hypothesis. The rate of adaptation of B. sorokiniana to the resistant varieties of wheat and barley did not depend on the level of resistance expression (weak, moderate or strong) and genetic control (oligogenic, polygenic or cytoplasmatic). It was shown by hybridological analysis that "residual effect" of oligogenes of sorghum resistance against greenbug depended on small resistance genes, that can be independent or weakly connected with marker oligogene. These data allows to doubt in phenomenon of "residual effect" of oligogenes. It was shown that non specific pathogenicity of parasitic fungi increased during their reproductions on sensitive varieties of plants. Thus, cultivation of sensitive varieties causes damage of crop culture non only because of their own losses, but also by increasing the infection of moderately resistant varieties.     

The genetic basis of resistance to anticoagulants in rodents

Anticoagulant compounds, i.e., derivatives of either 4-hydroxycoumarin (e.g., warfarin, bromadiolone) or indane-1,3-dione (e.g., diphacinone, chlorophacinone), have been in worldwide use as rodenticides for >50 years. These compounds inhibit blood coagulation by repression of the vitamin K reductase reaction (VKOR). Anticoagulant-resistant rodent populations have been reported from many countries and pose a considerable problem for pest control. Resistance is transmitted as an autosomal dominant trait although, until recently, the basic genetic mutation was unknown. Here, we report on the identification of eight different mutations in the VKORC1 gene in resistant laboratory strains of brown rats and house mice and in wild-caught brown rats from various locations in Europe with five of these mutations affecting only two amino acids (Tyr139Cys, Tyr139Ser, Tyr139Phe and Leu128Gln, Leu128Ser). By recombinant expression of VKORC1 constructs in HEK293 cells we demonstrate that mutations at Tyr139 confer resistance to warfarin at variable degrees while the other mutations, in addition, dramatically reduce VKOR activity. Our data strongly argue for at least seven independent mutation events in brown rats and two in mice. They suggest that mutations in VKORC1 are the genetic basis of anticoagulant resistance in wild populations of rodents, although the mutations alone do not explain all aspects of resistance that have been reported. We hypothesize that these mutations, apart from generating structural changes in the VKORC1 protein, may induce compensatory mechanisms to maintain blood clotting. Our findings provide the basis for a DNA-based field monitoring of anticoagulant resistance in rodents.     

The genetic manipulation of medicinal and aromatic plants

Medicinal and aromatic plants have always been intimately linked with human health and culture. Plant-derived medicines constitute a substantial component of present day human healthcare systems in industrialized as well as developing countries. They are products of plant secondary metabolism and are involved in many other aspects of a plant's interaction with its immediate environment. The genetic manipulation of plants together with the establishment of in vitro plant regeneration systems facilitates efforts to engineer secondary product metabolic pathways. Advances in the cloning of genes involved in relevant pathways, the development of high throughput screening systems for chemical and biological activity, genomics tools and resources, and the recognition of a higher order of regulation of secondary plant metabolism operating at the whole plant level facilitate strategies for the effective manipulation of secondary products in plants. Here, we discuss advances in engineering metabolic pathways for specific classes of compounds in medicinal and aromatic plants and we identify remaining constraints and future prospects in the field. In particular we focus on indole, tropane, nicotine, isoquinoline alcaloids, monoterpenoids such as menthol and related compounds, diterpenoids such as taxol, sequiterpenoids such as artemisinin and aromatic amino acids.     

Agrobacterium-mediated genetic transformation of plants: The role of host

Agrobacterium-mediated genetic transformation is the most widely used technology to obtain overexpression of recombinant proteins in plants. Molecular events that occur within Agrobacterium during interactions with host plants have been studied extensively, and now we have a reasonable understanding the key factors involved in the regulation of T-DNA nuclear import and genomic integration. By contrast, very little is known about the events that take place in the host cells during genetic transformation by Agrobacterium. Here, we describe the plant-related factors including genotype, genes, proteins, competency of target tissues and phenolic compounds that participate in Agrobacterium-mediated genetic transformation and discuss their possible roles in this process. Because Agrobacterium probably adapts existing cellular processes for its life cycle, identifying the processes in host cells during Agrobacterium infection might contribute to better understanding of basic biological processes as cell communication, intracellular transport and DNA repair and recombination as well as to expanding the host range of Agrobacterium as a genetic engineering tool.     

Understanding invasion history: genetic structure and diversity of two globally invasive plants and implications for their management

Aim Resolving the origin of invasive plant species is important for understanding the introduction histories of successful invaders and aiding strategies aimed at their management. This study aimed to infer the number and origin(s) of introduction for the globally invasive species, Macfadyena unguis‐cati and Jatropha gossypiifolia using molecular data. Location Native range: Neotropics; Invaded range: North America, Africa, Europe, Asia, Pacific Islands and Australia. Methods We used chloroplast microsatellites (cpSSRs) to elucidate the origin(s) of introduced populations and calculated the genetic diversity in native and introduced regions. Results Strong genetic structure was found within the native range of M. unguis‐cati, but no genetic structuring was evident in the native range of J. gossypiifolia. Overall, 27 haplotypes were found in the native range of M. unguis‐cati. Only four haplotypes were found in the introduced range, with more than 96% of introduced specimens matching a haplotype from Paraguay. In contrast, 15 haplotypes were found in the introduced range of J. gossypiifolia, with all invasive populations, except New Caledonia, comprising multiple haplotypes. Main conclusions These data show that two invasive plant species from the same native range have had vastly different introduction histories in their non‐native ranges. Invasive populations of M. unguis‐cati probably came from a single or few independent introductions, whereas most invasive J. gossypiifolia populations arose from multiple introductions or alternatively from a representative sample of genetic diversity from a panmictic native range. As introduced M. unguis‐cati populations are dominated by a single haplotype, locally adapted natural enemies should make the best control agents. However, invasive populations of J. gossypiifolia are genetically diverse and the selection of bio‐control agents will be considerably more complex.