Marie Jasieniuk

polysat : an R package for polyploid microsatellite analysis

We present an R package to help remedy the lack of software for manipulating and analysing autopolyploid and allopolyploid microsatellite data. polysat can handle genotype data of any ploidy, including populations of mixed ploidy, and assumes that allele copy number is always ambiguous in partial heterozygotes. It can import and export genotype data in eight different formats, calculate pairwise distances between individuals using a stepwise mutation and infinite alleles model, estimate ploidy based on allele counts and estimate allele frequencies and pairwise FST values. This software is freely available through the Comprehensive R Archive Network (http://cran.r‐project.org/) and includes a thorough tutorial.

Glyphosate-Resistant Italian Ryegrass (Lolium multiflorum) in California: Distribution, Response to Glyphosate, and Molecular Evidence for an Altered Target Enzyme

Abstract Selection by herbicides has resulted in widespread evolution of herbicide resistance in agricultural weeds. In California, resistance to glyphosate was first confirmed in rigid ryegrass in 1998. Objectives of this study were to determine the current distribution and level of glyphosate resistance in Italian ryegrass, and to assess whether resistance could be due to an altered target site. Seeds were sampled from 118 populations and seedlings were treated with glyphosate at 866 g ae ha−1. Percentage of survivors ranged from 5 to 95% in 54 populations. All plants from 64 populations died. One susceptible (S) population, four putatively resistant (R) populations, and one S accession from Oregon were used for pot dose–response experiments, shikimic acid analyses, and DNA sequencing. Seedlings were treated with glyphosate at eight rates, ranging from 108 to 13,856 g ae ha−1. Shoot biomass was evaluated 3 wk after treatment and fit to a log-logistic regression equation. On the basis of GR50 (herbicide rate required to reduce growth by 50%) values, seedlings from putatively R populations were roughly two to 15 times more resistant to glyphosate than S plants. Shikimic acid accumulation was similar in all plants before glyphosate treatment, but at 4 and 7 DAT, S plants from California and Oregon accumulated approximately two and three times more shikimic acid, respectively, than R plants. Sequencing of a cDNA fragment of the EPSPS coding region revealed two different codons, both of which encode proline at amino acid position 106 in S individuals. In contrast, all R plants sequenced exhibited missense mutations at site 106. Plants from one population revealed a mutation resulting in a proline to serine substitution. Plants from three R populations exhibited a mutation corresponding to replacement of proline with alanine. Our results indicate that glyphosate resistance is widespread in Italian ryegrass populations of California, and that resistance is likely due to an altered target enzyme. Nomenclature: Glyphosate; Italian ryegrass, Lolium multiflorum Lam. LOLMU; rigid ryegrass, Lolium rigidum Gaud. LOLRI.

Inheritance of Dicamba Resistance in Wild Mustard (Brassica kaber)

The inheritance of resistance to dicamba in wild mustard was deternined by making reciprocal crosses be- tween a resistant (R) population derived from a field treated repeatedly with auxin-type herbicides, and a known suscep- tible (S) population. The resulting F1 hybrids were selfed to produce F2 populations and backcrossed to the S parent. At the three- to four-leaf stage, parental, F1, F2, and backcross populations were screened for resistance to dicamba at three dosages (50, 200, and 400 g ai ha-1). F1 progeny survived all dosages and exhibited levels of injury similar to the R paren- tal population. F2 populations segregated in a 3:1 ratio of R to S phenotypes. Progeny of backcrosses segregated in a 1:1 (R:S) ratio. Responses of the F1, F2, and backcross popula- tions to treatment with dicamba indicate that resistance is determined by a single, completely dominant nuclear allele. Nomenclature: Dicamba, 3,6-dichloro-2-methoxybenzoic acid; wild mustard, Brassica kaber(DC.) L. C. Wheeler, Sinapis arvensis L. #3 SINAR. Additional index words. Genetics, herbicide resistance, auxin, Sinapis arvensis, SINAR.

De Novo Genome Assembly of the Economically Important Weed Horseweed Using Integrated Data from Multiple Sequencing Platforms

De novo genome assembly and genomic resources of horseweed will be useful to understand the genetic and molecular bases of weediness. Horseweed (Conyza canadensis), a member of the Compositae (Asteraceae) family, was the first broadleaf weed to evolve resistance to glyphosate. Horseweed, one of the most problematic weeds in the world, is a true diploid (2n = 2x = 18), with the smallest genome of any known agricultural weed (335 Mb). Thus, it is an appropriate candidate to help us understand the genetic and genomic bases of weediness. We undertook a draft de novo genome assembly of horseweed by combining data from multiple sequencing platforms (454 GS-FLX, Illumina HiSeq 2000, and PacBio RS) using various libraries with different insertion sizes (approximately 350 bp, 600 bp, 3 kb, and 10 kb) of a Tennessee-accessed, glyphosate-resistant horseweed biotype. From 116.3 Gb (approximately 350× coverage) of data, the genome was assembled into 13,966 scaffolds with 50% of the assembly = 33,561 bp. The assembly covered 92.3% of the genome, including the complete chloroplast genome (approximately 153 kb) and a nearly complete mitochondrial genome (approximately 450 kb in 120 scaffolds). The nuclear genome is composed of 44,592 protein-coding genes. Genome resequencing of seven additional horseweed biotypes was performed. These sequence data were assembled and used to analyze genome variation. Simple sequence repeat and single-nucleotide polymorphisms were surveyed. Genomic patterns were detected that associated with glyphosate-resistant or -susceptible biotypes. The draft genome will be useful to better understand weediness and the evolution of herbicide resistance and to devise new management strategies. The genome will also be useful as another reference genome in the Compositae. To our knowledge, this article represents the first published draft genome of an agricultural weed.

Evolution and spread of glyphosate resistance in Conyza canadensis in California.

Recent increases in glyphosate use in perennial crops of California, USA, are hypothesized to have led to an increase in selection and evolution of resistance to the herbicide in Conyza canadensis populations. To gain insight into the evolutionary origins and spread of resistance and to inform glyphosate resistance management strategies, we investigated the geographical distribution of glyphosate resistance in C. canadensis across and surrounding the Central Valley, its spatial relationship to groundwater protection areas (GWPA), and the genetic diversity and population structure and history using microsatellite markers. Frequencies of resistant individuals in 42 sampled populations were positively correlated with the size of GWPA within counties. Analyses of population genetic structure also supported spread of resistance in these areas. Bayesian clustering and approximate Bayesian computation (ABC) analyses revealed multiple independent origins of resistance within the Central Valley. Based on parameter estimation in the ABC analyses, resistant genotypes underwent expansion after glyphosate use began in agriculture, but many years before it was detected. Thus, diversity in weed control practices prior to herbicide regulation in GWPA probably kept resistance frequencies low. Regionally coordinated efforts to reduce seed dispersal and selection pressure are needed to manage glyphosate resistance in C. canadensis.

De novo genome assembly of the economically-important weed Conyza canadensis using integrated data from multiple sequencing platforms

De novo genome assembly and genomic resources of horseweed will be useful to understand the genetic and molecular bases of weediness. Horseweed (Conyza canadensis), a member of the Compositae (Asteraceae) family, was the first broadleaf weed to evolve resistance to glyphosate. Horseweed, one of the most problematic weeds in the world, is a true diploid (2n = 2x = 18), with the smallest genome of any known agricultural weed (335 Mb). Thus, it is an appropriate candidate to help us understand the genetic and genomic bases of weediness. We undertook a draft de novo genome assembly of horseweed by combining data from multiple sequencing platforms (454 GS-FLX, Illumina HiSeq 2000, and PacBio RS) using various libraries with different insertion sizes (approximately 350 bp, 600 bp, 3 kb, and 10 kb) of a Tennessee-accessed, glyphosate-resistant horseweed biotype. From 116.3 Gb (approximately 350× coverage) of data, the genome was assembled into 13,966 scaffolds with 50% of the assembly = 33,561 bp. The assembly covered 92.3% of the genome, including the complete chloroplast genome (approximately 153 kb) and a nearly complete mitochondrial genome (approximately 450 kb in 120 scaffolds). The nuclear genome is composed of 44,592 protein-coding genes. Genome resequencing of seven additional horseweed biotypes was performed. These sequence data were assembled and used to analyze genome variation. Simple sequence repeat and single-nucleotide polymorphisms were surveyed. Genomic patterns were detected that associated with glyphosate-resistant or -susceptible biotypes. The draft genome will be useful to better understand weediness and the evolution of herbicide resistance and to devise new management strategies. The genome will also be useful as another reference genome in the Compositae. To our knowledge, this article represents the first published draft genome of an agricultural weed.

Cross-Resistance to Herbicides of Five ALS-Inhibiting Groups and Sequencing of the ALS Gene in Cyperus difformis L.

Resistance to ALS-inhibiting herbicides in Cyperus difformis has evolved rapidly in many rice areas worldwide. This study identified the mechanism of resistance, assessed cross-resistance patterns to all five chemical groups of ALS-inhibiting herbicides in four C. difformis biotypes, and attempted to sequence the ALS gene. Whole-plant and ALS enzyme activity dose-response assays indicated that the WA biotype was resistant to all ALS-inhibiting herbicides evaluated. The IR biotype was resistant to bensulfuron-methyl, orthosulfamuron, imazethapyr, and propoxycarbazone-sodium and less resistant to bispyribac-sodium and halosulfuron-methyl, and susceptible to penoxsulam. ALS enzyme activity assays indicated that resistance is due to an altered target site yet mutations previously found to endow target-site resistance in weeds were not detected in the sequences obtained. The inability to detect resistance mutations in C. difformis may result from the presence of additional ALS genes, which were not amplified by the primers used. This study reports the first ALS gene sequence from Cyperus difformis. Certain ALS-inhibiting herbicides can still be used to control some resistant C. difformis biotypes. However, because cross-resistance to all five classes of ALS-inhibitors was detected in other resistant biotypes, these herbicides should only be used within an integrated weed management program designed to delay the evolution of herbicide resistance.

Microsatellite variation points to local landscape plantings as sources of invasive pampas grass (Cortaderia selloana) in California

International trade in horticultural plants is a major pathway of introduction of invasive species. Pampas grass (Cortaderia selloana) is an invasive species of horticultural origin that is native to South America but cultivated as an ornamental in regions with Mediterranean climates worldwide. To gain insight into the introduction history of invasive populations in California, we analysed microsatellite marker variation in cultivated and invasive C. selloana. We sampled 275 cultivated plants from diverse sources and 698 invasive plants from 33 populations in four geographical regions of California. A model‐based Bayesian clustering analysis identified seven distinct gene pools in cultivated C. selloana. Probabilities of assignment of invasive individuals to cultivated gene pools indicated that two gene pools accounted for the genomic origin of 78% of the invasive C. selloana sampled. Extensive admixture between cultivated source gene pools was detected within invasive individuals. Sources of admixed invasive individuals are probably landscape plantings. Consistent with the Bayesian assignment results indicating that multiple cultivated gene pools and landscape plantings are probable sources of invasive populations, FST and neighbour‐joining clustering analyses indicated multiple escapes from shared sources in each geographical region. No isolation by distance or geographical trend in reduction of genetic diversity was evident. Furthermore, a generally random and discontinuous distribution of proportional assignments of invasive populations to cultivated gene pools suggests that introductions occurred recurrently within each geographical region. Our results strongly suggest that dispersal through local landscape plantings has contributed to the range expansion of invasive C. selloana in California.

Review: Sampling Weedy and Invasive Plant Populations for Genetic Diversity Analysis

Abstract Recent advances in molecular methods and statistical analyses provide weed scientists with powerful tools for examining the genetic structure of weedy plant populations. The value of these studies depends on effective sampling protocols; however, there is little consensus on how to sample plant populations for genetic diversity analyses. In this review, we draw on published literature that incorporates sampling theory and spatial statistics in population genetic analyses to identify key factors to consider when designing a sampling strategy. We discuss how sampling design is affected by research objectives, biology of the study species, population structure, marker choice, and the genetic parameters to be investigated, and we offer suggestions on defining sampling units and developing sampling protocols.

Weed science research and funding: A call to action

Abstract Weed science has contributed much to agriculture, forestry and natural resource management during its history. However, if it is to remain relevant as a scientific discipline, it is long past time for weed scientists to move beyond a dominating focus on herbicide efficacy testing and address the basic science underlying complex issues in vegetation management at many levels of biological organization currently being solved by others, such as invasion ecologists and molecular biologists. Weed science must not be circumscribed by a narrowly-defined set of tools but rather be seen as an integrating discipline. As a means of assessing current and future research interests and funding trends among weed scientists, the Weed Science Society of America conducted an online survey of its members in summer of 2007. There were 304 respondents out of a membership of 1330 at the time of the survey, a response rate of 23%. The largest group of respondents (41%) reported working on research problems primarily focused on herbicide efficacy and maintenance, funded mainly by private industry sources. Another smaller group of respondents (22%) reported focusing on research topics with a complex systems focus (such as invasion biology, ecosystem restoration, ecological weed management, and the genetics, molecular biology, and physiology of weedy traits), funded primarily by public sources. Increased cooperation between these complementary groups of scientists will be an essential step in making weed science increasingly relevant to the complex vegetation management issues of the 21st century.