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PubMed2002

Substitution of Dmo1 with normal alleles results in decreased manifestation of diabetes in OLETF rats.

AIM: Dmo1 (Diabetes Mellitus OLETF type I) is a major quantitative trait locus for dyslipidaemia, obesity and diabetes phenotypes in the Otsuka Long Evans Tokushima Fatty (OLETF) rat strain. To evaluate possible metabolic and pathological improvements generated by correction of the Dmo1 genetic pathway, we produced congenic lines, in which both OLETF Dmo1 alleles are replaced by the F344-derived genome. METHODS: Congenic animals were produced by introgressing F344-derived Dmo1 alleles into the OLETF rat. Congenic animals of the fourth generation (BC4) were intercrossed to obtain F1 animals (BC4:F1). Animals of the next generation, BC4:F2, were used for this study. We used 23 BC4:F2 males harbouring homozygous replacement of the OLETF Dmo1 region with the F344-derived genome. Seven animals with OLETF-derived Dmo1 alleles were used as controls. RESULTS: Dmo1-F344/F344 congenic rats showed significant decreases in body weight, abdominal fat weight, serum triacylglycerols, total cholesterol, food consumption and blood glucose after glucose loading (13%, 39%, 45%, 27%, 18% and 27% respectively; p < 0.05) compared with Dmo1-OLETF/OLETF animals. Furthermore, histopathological analysis of the kidney showed that mesangial sclerosis, hyalin deposits and deposition of PAS-positive substance were significantly lower in Dmo1-F344/F344 animals (p < 0.05). CONCLUSION: Improvements in metabolic parameters and histopathological scores show that correction of the Dmo1 genetic pathway in the diabetic and mildly obese OLETF rat strain produces wide-ranging therapeutic effects. Thus, this pathway might represent a new drug target also applicable to humans.

Animals↗

PubMed2003

Haplotyping and mapping a large cluster of downy mildew resistance gene candidates in sunflower using multilocus intron fragment length polymorphisms.

Downy mildew (Plasmopara halstedii (Farl.) Berlese et de Toni) is a serious foliar pathogen of cultivated sunflower (Helianthus annuus L.). Genetic resistance is conditioned by several linked downy mildew resistance gene specificities in the HaRGC1 cluster of TIR-NBS-LRR resistance gene candidates (RGCs) on linkage group 8. The complexity and diversity of the HaRGC1 cluster was assessed by multilocus intron fragment length polymorphism (IFLP) genotyping using a single pair of primers flanking a hypervariable intron located between the TIR and NBS domains. Two to 23 bands were amplified per germplasm accession. The size of the included intron ranged from 89 to 858 nucleotides. Forty-eight unique markers were distinguished among 24 elite inbred lines, six partially isogenic inbred lines, nine open-pollinated populations, four Native American land races, and 20 wild H. annuus populations. Nine haplotypes (based on 24 RGCs) were identified among elite inbred lines and were correlated with known downy mildew resistance specificities. Sixteen out of 39 RGCs identified in wild H. annuus populations were not observed in elite germplasm. Five partially isogenic downy mildew resistant lines developed from wild H. annuus and H. praecox donors carried eight RGCs not found in other elite inbred lines. Twenty-four HaRGC1 loci were mapped to a 2-4 cM segment of linkage group 8. The multilocus IFLP marker and duplicated, hypervariable microsatellite markers tightly linked to the HaRGC1 cluster are powerful tools for distinguishing downy mildew resistance gene specificities and identifying and introgressing new downy mildew resistance gene specificities from wild sunflowers.

Journal Article↗

PubMed2003

There is more to tomato fruit colour than candidate carotenoid genes.

Determining gene sequences responsible for complex phenotypes has remained a major objective in modern biology. The candidate gene approach is attempting to link, through mapping analysis, sequences that have a known functional role in the measured phenotype with quantitative trait loci (QTL) that are responsible for the studied variation. To explore the potential of the candidate approach for complex traits we conducted a mapping analysis of QTL for the intensity of the red colour of the tomato fruit (mainly lycopene) and for probes associated with the well-characterized carotenoid biosynthesis pathway. Seventy-five tomato introgression lines (ILs), each containing a single homozygous RFLP-defined chromosome segment from the green-fruited species Lycopersicon pennellii delimited 107 marker-defined mapping bins. Three of the bins resolved known qualitative colour mutations for yellow (r) and orange (B and Del) fruits resulting from variation in specific carotenoid biosynthesis genes. Based on trials in different environments, 16 QTL that modified the intensity of the red colour of ripe fruit were assigned to bins. Candidate sequences associated with the carotenoid biosynthesis pathway were mapped to 23 loci. Only five of the QTL co-segregated with the same bins that contained candidate genes - a number that is expected by chance alone. Furthermore, similar map location of a QTL and a candidate is far from a direct causative relationship between a gene and a phenotype. This study highlights the wealth and complexity of the variation present in the genus Lycopersicon that could be employed for basic research and genetic improvement of fruit colour in tomato.

Journal Article↗

PubMedDate not supplied

Mimicry: developmental genes that contribute to speciation.

Despite renewed interest in the role of natural selection as a catalyst for the origin of species, the developmental and genetic basis of speciation remains poorly understood. Here we describe the genetics of Müllerian mimicry in Heliconius cydno and H. melpomene (Lepidoptera: Nymphalidae), sister species that recently diverged to mimic other Heliconius. This mimetic shift was a key step in their speciation, leading to pre- and postmating isolation. We identify 10 autosomal loci, half of which have major effects. At least eight appear to be homologous with genes known to control pattern differences within each species. Dominance has evolved under the influence of identifiable "modifier" loci rather than being a fixed characteristic of each locus. Epistasis is found at many levels: phenotypic interaction between specific pairs of genes, developmental canalization due to polygenic modifiers so that patterns are less sharply defined in hybrids, and overall fitness through ecological selection against nonmimetic hybrid genotypes. Most of the loci are clustered into two genomic regions or "supergenes," suggesting color pattern evolution is constrained by preexisting linked elements that may have arisen via tandem duplication rather than having been assembled by natural selection. Linkage, modifiers, and epistasis affect the strength of mimicry as a barrier to gene flow between these naturally hybridizing species and may permit introgression in genomic regions unlinked to those under disruptive selection. Müllerian mimics in Heliconius use different genetic architectures to achieve the same mimetic patterns, implying few developmental constraints. Therefore, although developmental and genomic constraints undoubtedly influence the evolutionary process, their effects are probably not strong in comparison with natural selection.

Animals↗

PubMed2002

Fitness effects of Alternaria dauci on wild carrot in The Netherlands.

In a field experiment with a susceptible population of annual wild carrot (Daucus carota) from Iran, artificial inoculations with the fungal pathogen A. dauci led to a strong and very significant increase of the diseased leaf area. The pathogen caused a very significant decrease in fecundity and seed survival of the host. This considerable fitness reduction by A. dauci would suggest that introgression of disease resistance from cultivated (transgenic) carrot cultivars into wild carrot populations could strongly increase the fitness of wild carrot. In spite of the potential ability of A. dauci to lower the fitness of the host considerably, wild carrot is very common in The Netherlands. Disease levels of wild carrot were estimated in 26 natural populations in 1998. No A. dauci could be detected on the leaves, and only 0.4% of the seeds were contaminated with A. dauci, in spite of the conducive weather for A. dauci. Resistance tests showed that all 26 monitored populations were highly resistant to A. dauci strains from The Netherlands. It is probable that the strong potential fitness reduction by A. dauci led to a high selection pressure towards resistance in The Netherlands. In conclusion, our results suggest that transgenic resistance to A. dauci would not be beneficial to wild carrot populations in The Netherlands because they are already resistant to this pathogen.

Alternaria↗

PubMedDate not supplied

Comparative fitness of a wild squash species and three generations of hybrids between wild x virus-resistant transgenic squash.

We compared some fitness components of the wild squash species Cucurbita pepo spp. ovifera var. texana (C. texana) and three generations of hybrids (F1, BC1, and BC2) between C. texana and commercial transgenic squash CZW-3 over three consecutive years under field conditions of low (LDP) and high disease pressure (HDP) by Cucumber mosaic virus (CMV), Zucchini yellow mosaic virus (ZYMV) and Watermelon mosaic virus (WMV). Transgenic squash CZW-3 expresses the coat protein (CP) genes of CMV, ZYMV, and WMV, and is resistant to these three aphid-borne viruses. Across all HDP trials, transgenic BC1 and BC2 hybrids expressing the three CP genes grew more vigorously, displayed resistance to CMV, ZYMV, and WMV, and produced a greater number of mature fruits and viable seeds than nontransgenic hybrid segregants and C. texana. Transgenic F1 hybrids behaved similarly to BC1 and BC2 hybrids but grew less vigorously than C. texana. In contrast, across all LDP trials, C. texana outperformed the transgenic and nontransgenic hybrid segregants. Further, only one back cross was necessary to recover individuals with most of the C. texana characteristics and yet maintain virus resistance. Our data suggest that C. texana acquiring CP transgenes upon hybridization and introgression could have a selective advantage if CMV, ZYMV, and WMV are severely limiting the growth and reproductibility of wild squash populations.

Animals↗

PubMedDate not supplied

Transgene expression and fitness of hybrids between GM oilseed rape and Brassica rapa.

Oilseed rape (Brassica napus) is sexually compatible with its wild and weedy relative B. rapa, and introgression of genes from B. napus has been found to occur over a few generations. We simulated the early stages of transgene escape by producing F1 hybrids and the first backcross generation between two lines of transgenic B. napus and two populations of weedy B. rapa. Transgene expression and the fitness of the hybrids were examined under different environmental conditions. Expression of the transgenes was analyzed at the mRNA level by quantitative PCR and found to be stable in the hybrids, regardless of the genetic background and the environment, and equal to the level of transcription in the parental B. napus lines. Vigor of the hybrids was measured as the photosynthetic capability; pollen viability and seed set per silique. Photosynthetic capability of first generation hybrids was found to be at the same level, or higher, than that of the parental species, whereas the reproductive fitness was significantly lower. The first backcross generation had a significantly lower photosynthetic capability and reproductive fitness compared to the parental species. This is the first study that examines transgene expression at the mRNA level in transgenic hybrids of B. napus of different genetic background exposed to different environmental conditions. The data presented clarify important details of the overall risk assessment of growing transgenic oilseed rape.

Brassica napus↗

PubMedDate not supplied

Tiered tests to assess the environmental risk of fitness changes in hybrids between transgenic crops and wild relatives: the example of virus resistant Brassica napus.

Over the last 20 years, there has been much research aimed at improving environmental risk assessment of transgenic crops. Despite large amounts of data, decisions to allow or prohibit the release of transgenic crops remain confused and controversial. We argue that part of the reason for confusion is the lack of clear definitions of components of the environment that should be protected, and, as a consequence, there is no way to judge the relevance of data collected under the auspices of 'environmental risk assessment'. Although this criticism applies to most aspects of environmental risk assessment of transgenic crops, it is most pertinent to effects that might result from an increase in plant fitness, often referred to as increased weediness. Environmental risk assessment of weediness is regarded as complicated: an increase in the fitness of a transgenic plant compared with non-transgenic counterparts will be the result of an interaction between the altered plant phenotype and an enormous number of environmental variables. This has led to the idea that risk assessment of weediness needs to "understand" these interactions, with the implication that exhaustive data are required. Here we argue that environmental risk assessment of the weediness of transgenic plants need not be complicated. Analysis of the conditions that must be met for increased weediness to occur suggests a series of studies that starts with simple tests in the laboratory under "worst case" assumptions, and becomes increasingly complex and realistic should the simpler studies not indicate negligible risk with sufficient certainty. We illustrate how the approach might work for assessing the risks of increased weediness using the example of possible introgression of a gene for Turnip mosaic virus (TuMV) resistance from oilseed rape to certain wild Brassica species.

Brassica↗

PubMed2006

Monitoring the escape of transgenic oilseed rape around Japanese ports and roadsides.

An investigation was carried out to monitor the escape and spread of oilseed rape (Brassica napus) transgenic plants and the introgression of transgenes to its closely related feral species in Japan. We screened a total of about 7500 feral B. napus, 300 B. rapa, and 5800 B. juncea seedlings from maternal plants in 143 locations at several ports, roadsides, and riverbanks. The presence of glufosinate-resistance or glyphosate-resistance transgenes in these seedlings was confirmed by means of herbicide treatments and also immunochemical and DNA analyses. B. napus plants with herbicide-resistant transgenic seeds were found at five of six major ports and along two of four sampled roadsides in the Kanto District. Transgenic oilseed rape plants have not been commercially cultivated in Japan, suggesting that the transgenes would probably have come from imported transgenic seeds that were spilled during transportation to oilseed processing facilities. No transgenes were detected in seeds collected from B. napus plants growing along riverbanks in the Kanto District or in seeds from closely related species (B. rapa and B. juncea). To our knowledge, this is the first published example of feral, transgenic populations occurring in a nation where the transgenic crop has not been cultivated commercially.

Aminobutyrates↗

PubMed2006

Detecting (trans)gene flow to landraces in centers of crop origin: lessons from the case of maize in Mexico.

There is much discussion of the probability of transgene flow from transgenic crop varieties to landraces and wild relatives in centers of origin or diversity, and its genetic, ecological, and social consequences. Without costly research on the variables determining gene flow, research on transgene frequencies in landrace (or wild relative) populations can be valuable for understanding transgene flow and its effects. Minimal research requirements include (1) understanding how farmer practices and seed systems affect landrace populations, (2) sampling to optimize Ne/n (effective/census population size), (3) minimizing variance at all levels sampled, and (4) using Ne to calculate binomial probabilities for transgene frequencies. A key case is maize in Mexico. Two peer-reviewed papers, based on landrace samples from the Sierra Juárez region of Oaxaca, Mexico, reached seemingly conflicting conclusions: transgenes are present (Quist and Chapela, 2001, Nature 414: 541-543; 2002, Nature 416: 602) or "detectable transgenes" are absent (Ortiz-García et al., 2005, Proc. Natl. Acad. Sci. USA 102: 12338-12343 and 18242). We analyzed these papers using information on Oaxacan maize seed systems and estimates of Ne. We conclude that if Quist and Chapela's results showing presence are accepted, Ortiz-García et al.'s conclusions of no evidence of transgenes at detectable levels or for their introgression into maize landraces in the Sierra de Juárez of Oaxaca are not scientifically justified. This is because their samples are not representative, and their statistical analysis is inconclusive due to using n instead of Ne. Using estimates of Ne based on Ortiz-García et al.'s n, we estimate that transgenes could be present in maize landraces in the Sierra Juárez region at frequencies of approximately 1-4%, and are more likely to be present in the 90% of Oaxacan landrace area that is not mountainous. Thus, we have no scientific evidence of maize transgene presence or absence in recent years in Mexico, Oaxaca State, or the Sierra Juárez region.

Crops, Agricultural↗

PubMed2005

Postglacial dispersal of Phlebotomus perniciosus into France.

Phlebotomus perniciosus was identified morphologically in samples from France and northeast Spain, and individuals were then characterized at three polymorphic isoenzyme loci (by isoelectrofocusing) and at the mitochondrial DNA locus (by comparative DNA sequence analysis of a fragment of the Cytochrome b gene). The four polymorphic loci gave conflicting patterns of population relationships, which can be explained by hypothesizing different amounts of gene introgression at each locus when two distinctive lineages met in southern France or northeast Spain after isolation in southern Italy and Spain during the Pleistocene Ice Ages. P. perniciosus is an important vector of leishmania infantum and so these population differentiation studies are relevant for predicting the emergence and spread of leishmaniasis in relation to environmental changes, including climate.

Animals↗

PubMed2006

Natural hybridization in Saxifraga callosa Sm.

Saxifraga callosa Sm. is an evergreen perennial species distributed from Eastern Spain, through the Western Alps and the Apennines, to southern Italy. The existence of high morphological variation within different subspecies indicates that phenotypic characters are useful but not sufficient taxonomic tools. Indeed, available morphological data already suggested that S. callosa subentity lantoscana may be an outcross between S. callosa and S. cochlearis. In this work, by analyzing ITS (Internal Transcribed Sequences), AFLP (Amplified Fragment Length Polymorphisms), and cpDNA (chloroplast DNA) markers, a comprehensive study of the genomic relationships among S. callosa and related species has been carried out. The sequence of the ITS region of S. callosa subentity lantoscana gave no conclusive results on the taxonomy status of S. callosa subentity lantoscana. On the other hand, the use of the "NewHybrids" software to analyze an AFLP data-set (208 polymorphic amplified fragments) supported a significant posterior probability that S. callosa subentity lantoscana individuals are natural hybrids between S. callosa and S. cochlearis. The level of introgression of genes from alien genomes was confirmed by a simpler and quick methodology that analyze length variation in cpDNA sequences.

Crosses, Genetic↗

PubMed2004

Distinguishing human ethnic groups by means of sequences from Helicobacter pylori: lessons from Ladakh.

The history of mankind remains one of the most challenging fields of study. However, the emergence of anatomically modern humans has been so recent that only a few genetically informative polymorphisms have accumulated. Here, we show that DNA sequences from Helicobacter pylori, a bacterium that colonizes the stomachs of most humans and is usually transmitted within families, can distinguish between closely related human populations and are superior in this respect to classical human genetic markers. H. pylori from Buddhists and Muslims, the two major ethnic communities in Ladakh (India), differ in their population-genetic structure. Moreover, the prokaryotic diversity is consistent with the Buddhists having arisen from an introgression of Tibetan speakers into an ancient Ladakhi population. H. pylori from Muslims contain a much stronger ancestral Ladakhi component, except for several isolates with an Indo-European signature, probably reflecting genetic flux from the Near East. These signatures in H. pylori sequences are congruent with the recent history of population movements in Ladakh, whereas similar signatures in human microsatellites or mtDNA were only marginally significant. H. pylori sequence analysis has the potential to become an important tool for unraveling short-term genetic changes in human populations.

DNA, Bacterial↗

PubMed2004

Evidence for landscape-level, pollen-mediated gene flow from genetically modified creeping bentgrass with CP4 EPSPS as a marker.

Sampling methods and results of a gene flow study are described that will be of interest to plant scientists, evolutionary biologists, ecologists, and stakeholders assessing the environmental safety of transgenic crops. This study documents gene flow on a landscape level from creeping bentgrass (Agrostis stolonifera L.), one of the first wind-pollinated, perennial, and highly outcrossing transgenic crops being developed for commercial use. Most of the gene flow occurred within 2 km in the direction of prevailing winds. The maximal gene flow distances observed were 21 km and 14 km in sentinel and resident plants, respectively, that were located in primarily nonagronomic habitats. The selectable marker used in these studies was the CP4 EPSPS gene derived from Agrobacterium spp. strain CP4 that encodes 5-enol-pyruvylshikimate-3-phosphate synthase and confers resistance to glyphosate herbicide. Evidence for gene flow to 75 of 138 sentinel plants of A. stolonifera and to 29 of 69 resident Agrostis plants was based on seedling progeny survival after spraying with glyphosate in greenhouse assays and positive TraitChek, PCR, and sequencing results. Additional studies are needed to determine whether introgression will occur and whether it will affect the ecological fitness of progeny or the structure of plant communities in which transgenic progeny may become established.

3-Phosphoshikimate 1-Carboxyvinyltransferase↗

PubMed2002

Integration of foreign DNA during natural transformation of Acinetobacter sp. by homology-facilitated illegitimate recombination.

The active uptake of extracellular DNA and its genomic integration is termed natural transformation and constitutes a major horizontal gene-transfer mechanism in prokaryotes. Chromosomal DNA transferred within a species can be integrated effectively by homologous recombination, whereas foreign DNA with low or no sequence homology would rely on illegitimate recombination events, which are rare. By using the nptII(+) gene (kanamycin resistance) as selectable marker, we found that the integration of foreign DNA into the genome of the Gram-negative Acinetobacter sp. BD413 during transformation indeed was at least 10(9)-fold lower than that of homologous DNA. However, integration of foreign DNA increased at least 10(5)-fold when it was linked on one side to a piece of DNA homologous to the recipient genome. Analysis of foreign DNA integration sites revealed short stretches of sequence identity (3-8 bp) between donor and recipient DNA, indicating illegitimate recombination events. These findings suggest that homologous DNA served as a recombinational anchor facilitating illegitimate recombination acting on the same molecule. Homologous stretches down to 183 nucleotides served as anchors. Transformation with heteroduplex DNA having different nucleotide sequence tags in the strands indicated that strands entered the cytoplasm 3' to 5' and that strands with either polarity were integrated by homologous recombination. The process led to the genomic integration of thousands of foreign nucleotides and often was accompanied by deletion of a roughly corresponding length of recipient DNA. Homology-facilitated illegitimate recombination would explain the introgression of DNA in prokaryotic genomes without the help of mobile genetic elements.

Acinetobacter↗

PubMed2005

An ancestral oomycete locus contains late blight avirulence gene Avr3a, encoding a protein that is recognized in the host cytoplasm.

The oomycete Phytophthora infestans causes late blight, the potato disease that precipitated the Irish famines in 1846 and 1847. It represents a reemerging threat to potato production and is one of >70 species that are arguably the most devastating pathogens of dicotyledonous plants. Nevertheless, little is known about the molecular bases of pathogenicity in these algae-like organisms or of avirulence molecules that are perceived by host defenses. Disease resistance alleles, products of which recognize corresponding avirulence molecules in the pathogen, have been introgressed into the cultivated potato from a wild species, Solanum demissum, and R1 and R3a have been identified. We used association genetics to identify Avr3a and show that it encodes a protein that is recognized in the host cytoplasm, where it triggers R3a-dependent cell death. Avr3a resides in a region of the P. infestans genome that is colinear with the locus containing avirulence gene ATR1(NdWsB) in Hyaloperonospora parasitica, an oomycete pathogen of Arabidopsis. Remarkably, distances between conserved genes in these avirulence loci were often similar, despite intervening genomic variation. We suggest that Avr3a has undergone gene duplication and that an allele evading recognition by R3a arose under positive selection.

Agrobacterium tumefaciens↗

PubMed2006

Experimental evidence for interspecific directional selection on moth pheromone communication.

The chemical composition of the sexual communication signals of female moths is thought to be under strong stabilizing selection, because females that produce atypical pheromone blends suffer lower success in finding mates. This intraspecific selection pressure cannot explain the high diversity of moth pheromone blends found in nature. We conducted experiments to determine whether communication interference from males of closely related species could exert strong enough directional selection to cause evolution of these signals. Attraction and mating success of Heliothis subflexa (Hs) females with a normal quantitative trait locus for production of acetate pheromone components (norm-OAc) were compared with Hs females with an introgressed quantitative trait locus from Heliothis virescens (Hv) that dramatically decreased the amount of acetate esters in their pheromone glands (low-OAc). In field experiments with natural Hv and Hs populations, 10 times more Hv males were captured in traps baited with live low-OAc Hs females than in traps with norm-OAc Hs females. This pattern was confirmed in mate-choice assays in cages. Hybrids resulting from Hv-Hs matings have effectively zero fitness in the field. Combining our results with the extensive data set gathered in the past 40 years on the reproductive biology of Hv, we can quantitatively estimate that the directional selection exerted by Hv males on Hs females to produce relatively high amounts (>5%) of acetates can range from 0.135 to 0.231. Such intense interspecific selection may counteract intraspecific stabilizing selection that impedes evolutionary changes in pheromone blends and could lead to diversification of sexual signals.

Animals↗

PubMed2006

The origin of European cattle: evidence from modern and ancient DNA.

Cattle domestication from wild aurochsen was among the most important innovations during the Neolithic agricultural revolution. The available genetic and archaeological evidence points to at least two major sites of domestication in India and in the Near East, where zebu and the taurine breeds would have emerged independently. Under this hypothesis, all present-day European breeds would be descended from cattle domesticated in the Near East and subsequently spread during the diffusion of herding and farming lifestyles. We present here previously undescribed genetic evidence in contrast with this view, based on mtDNA sequences from five Italian aurochsen dated between 7,000 and 17,000 years B.P. and >1,000 modern cattle from 51 breeds. Our data are compatible with local domestication events in Europe and support at least some levels of introgression from the aurochs in Italy. The distribution of genetic variation in modern cattle suggest also that different south European breeds were affected by introductions from northern Africa. If so, the European cattle may represent a more variable and valuable genetic resource than previously realized, and previous simple hypotheses regarding the domestication process and the diffusion of selected breeds should be revised.

Animals↗