Zhian N. Kamvar

Poppr: an R package for genetic analysis of populations with clonal, partially clonal, and/or sexual reproduction.

Many microbial, fungal, or oomcyete populations violate assumptions for population genetic analysis because these populations are clonal, admixed, partially clonal, and/or sexual. Furthermore, few tools exist that are specifically designed for analyzing data from clonal populations, making analysis difficult and haphazard. We developed the R package poppr providing unique tools for analysis of data from admixed, clonal, mixed, and/or sexual populations. Currently, poppr can be used for dominant/codominant and haploid/diploid genetic data. Data can be imported from several formats including GenAlEx formatted text files and can be analyzed on a user-defined hierarchy that includes unlimited levels of subpopulation structure and clone censoring. New functions include calculation of Bruvo’s distance for microsatellites, batch-analysis of the index of association with several indices of genotypic diversity, and graphing including dendrograms with bootstrap support and minimum spanning networks. While functions for genotypic diversity and clone censoring are specific for clonal populations, several functions found in poppr are also valuable to analysis of any populations. A manual with documentation and examples is provided. Poppr is open source and major releases are available on CRAN: http://cran.r-project.org/package=poppr. More supporting documentation and tutorials can be found under ‘resources’ at: http://grunwaldlab.cgrb.oregonstate.edu/.

Novel R tools for analysis of genome-wide population genetic data with emphasis on clonality.

To gain a detailed understanding of how plant microbes evolve and adapt to hosts, pesticides, and other factors, knowledge of the population dynamics and evolutionary history of populations is crucial. Plant pathogen populations are often clonal or partially clonal which requires different analytical tools. With the advent of high throughput sequencing technologies, obtaining genome-wide population genetic data has become easier than ever before. We previously contributed the R package poppr specifically addressing issues with analysis of clonal populations. In this paper we provide several significant extensions to poppr with a focus on large, genome-wide SNP data. Specifically, we provide several new functionalities including the new function mlg.filter to define clone boundaries allowing for inspection and definition of what is a clonal lineage, minimum spanning networks with reticulation, a sliding-window analysis of the index of association, modular bootstrapping of any genetic distance, and analyses across any level of hierarchies.

Involving undergraduates in the annotation and analysis of global gene expression studies : Creation of a maize shoot apical meristem expression database

Through a multi-university and interdisciplinary project we have involved undergraduate biology and computer science research students in the functional annotation of maize genes and the analysis of their microarray expression patterns. We have created a database to house the results of our functional annotation of >4400 genes identified as being differentially regulated in the maize shoot apical meristem (SAM). This database is located at http://sam.truman.edu and is now available for public use. The undergraduate students involved in constructing this unique SAM database received hands-on training in an intellectually challenging environment, which has prepared them for graduate and professional careers in biological sciences. We describe our experiences with this project as a model for effective research-based teaching of undergraduate biology and computer science students, as well as for a rich professional development experience for faculty at predominantly undergraduate institutions.

Spatial and Temporal Analysis of Populations of the Sudden Oak Death Pathogen in Oregon Forests

Sudden oak death caused by the oomycete Phytophthora ramorum was first discovered in California toward the end of the 20th century and subsequently emerged on tanoak forests in Oregon before its first detection in 2001 by aerial surveys. The Oregon Department of Forestry has since monitored the epidemic and sampled symptomatic tanoak trees from 2001 to the present. Populations sampled over this period were genotyped using microsatellites and studied to infer the population genetic history. To date, only the NA1 clonal lineage is established in this region, although three lineages exist on the North American west coast. The original introduction into the Joe Hall area eventually spread to several regions: mostly north but also east and southwest. A new introduction into Hunter Creek appears to correspond to a second introduction not clustering with the early introduction. Our data are best explained by both introductions originating from nursery populations in California or Oregon and resulting from two distinct introduction events. Continued vigilance and eradication of nursery populations of P. ramorum are important to avoid further emergence and potential introduction of other clonal lineages.

apex: phylogenetics with multiple genes

Genetic sequences of multiple genes are becoming increasingly common for a wide range of organisms including viruses, bacteria and eukaryotes. While such data may sometimes be treated as a single locus, in practice, a number of biological and statistical phenomena can lead to phylogenetic incongruence. In such cases, different loci should, at least as a preliminary step, be examined and analysed separately. The r software has become a popular platform for phylogenetics, with several packages implementing distance‐based, parsimony and likelihood‐based phylogenetic reconstruction, and an even greater number of packages implementing phylogenetic comparative methods. Unfortunately, basic data structures and tools for analysing multiple genes have so far been lacking, thereby limiting potential for investigating phylogenetic incongruence. In this study, we introduce the new r package apex to fill this gap. apex implements new object classes, which extend existing standards for storing DNA and amino acid sequences, and provides a number of convenient tools for handling, visualizing and analysing these data. In this study, we introduce the main features of the package and illustrate its functionalities through the analysis of a simple data set.

Developing educational resources for population genetics in R: an open and collaborative approach.

The r computing and statistical language community has developed a myriad of resources for conducting population genetic analyses. However, resources for learning how to carry out population genetic analyses in r are scattered and often incomplete, which can make acquiring this skill unnecessarily difficult and time consuming. To address this gap, we developed an online community resource with guidance and working demonstrations for conducting population genetic analyses in r. The resource is freely available at http://popgen.nescent.org and includes material for both novices and advanced users of r for population genetics. To facilitate continued maintenance and growth of this resource, we developed a toolchain, process and conventions designed to (i) minimize financial and labour costs of upkeep; (ii) to provide a low barrier to contribution; and (iii) to ensure strong quality assurance. The toolchain includes automatic integration testing of every change and rebuilding of the website when new vignettes or edits are accepted. The process and conventions largely follow a common, distributed version control‐based contribution workflow, which is used to provide and manage open peer review by designated website editors. The online resources include detailed documentation of this process, including video tutorials. We invite the community of population geneticists working in r to contribute to this resource, whether for a new use case of their own, or as one of the vignettes from the ‘wish list’ we maintain, or by improving existing vignettes.

Population structure and phenotypic variation of Sclerotinia sclerotiorum from dry bean (Phaseolus vulgaris) in the United States

The ascomycete pathogen Sclerotinia sclerotiorum is a necrotrophic pathogen on over 400 known host plants, and is the causal agent of white mold on dry bean. Currently, there are no known cultivars of dry bean with complete resistance to white mold. For more than 20 years, bean breeders have been using white mold screening nurseries (wmn) with natural populations of S. sclerotiorum to screen new cultivars for resistance. It is thus important to know if the genetic diversity in populations of S. sclerotiorum within these nurseries (a) reflect the genetic diversity of the populations in the surrounding region and (b) are stable over time. Furthermore, previous studies have investigated the correlation between mycelial compatibility groups (MCG) and multilocus haplotypes (MLH), but none have formally tested these patterns. We genotyped 366 isolates of S. sclerotiorum from producer fields and wmn surveyed over 10 years in 2003–2012 representing 11 states in the United States of America, Australia, France, and Mexico at 11 microsatellite loci resulting in 165 MLHs. Populations were loosely structured over space and time based on analysis of molecular variance and discriminant analysis of principal components, but not by cultivar, aggressiveness, or field source. Of all the regions tested, only Mexico (n = 18) shared no MLHs with any other region. Using a bipartite network-based approach, we found no evidence that the MCGs accurately represent MLHs. Our study suggests that breeders should continue to test dry bean lines in several wmn across the United States to account for both the phenotypic and genotypic variation that exists across regions.

First Report of the EU1 Clonal Lineage of Phytophthora ramorum on Tanoak in an Oregon Forest

Initially reported in California as the causal agent of sudden oak death (SOD), efforts to limit spread of Phytophthora ramorum in Oregon natural forests have concentrated on quarantine regulations and eradication of the pathogen from infested areas. P. ramorum has four clonal lineages: NA1; NA2; EU1; and EU2 (Grünwald et al. 2012; Van Poucke et al. 2012). Forest infestations in Oregon have been limited to the NA1 clonal lineage, whereas EU1, NA1, and NA2 clonal lineages have all been found in U.S. nurseries (Kamvar et al. 2015; Prospero et al. 2007). In February 2015, in response to an aerial survey, P. ramorum was isolated from a dying Notholithocarpus densiflorus tree in the South Fork Pistol River drainage of Curry Co., Oregon. The isolated strain was identified as P. ramorum based on presence of chlamydospores, characteristic hyphae, and sporangial morphology. Microsatellite genotyping at 14 loci (Vercauteren et al. 2011) and comparison with reference cultures revealed that these isolates belonged to the EU1 clonal lineage. Subsequently, five more isolates were obtained from the original tree stump and the EU1 lineage was confirmed. Microsatellite alleles of the forest EU1 isolates were nearly identical to EU1 isolates collected in 2012 from a nearby nursery during routine P. ramorum nursery monitoring, except for one allele at locus PrMS145a. Interestingly, several isolates differed at locus ILVOPrMS131a within both the 2015 forest and the 2012 nursery findings with identical allele frequencies in each population for this locus. These data provide inconclusive support for the introduction of EU1 into Curry Co. from the 2012 populations found in nurseries, given that no direct match was found probably owing to the paucity of EU1 samples from nurseries. These results provide further evidence that multiple distinct P. Quick Links Add to favorites

Best Practices for Population Genetic Analyses

Population genetic analysis is a powerful tool to understand how pathogens emerge and adapt. However, determining the genetic structure of populations requires complex knowledge on a range of subtle skills that are often not explicitly stated in book chapters or review articles on population genetics. What is a good sampling strategy? How many isolates should I sample? How do I include positive and negative controls in my molecular assays? What marker system should I use? This review will attempt to address many of these practical questions that are often not readily answered from reading books or reviews on the topic, but emerge from discussions with colleagues and from practical experience. A further complication for microbial or pathogen populations is the frequent observation of clonality or partial clonality. Clonality invariably makes analyses of population data difficult because many assumptions underlying the theory from which analysis methods were derived are often violated. This review provides practical guidance on how to navigate through the complex web of data analyses of pathogens that may violate typical population genetics assumptions. We also provide resources and examples for analysis in the R programming environment.

Population Structure of Pythium irregulare, P. ultimum, and P. sylvaticum in Forest Nursery Soils of Oregon and Washington

Pythium species are important soilborne pathogens occurring in the forest nursery industry of the Pacific Northwest. However, little is known about their genetic diversity or population structure and it is suspected that isolates are moved among forest nurseries on seedling stock and shared field equipment. In order to address these concerns, a total of 115 isolates of three Pythium species (P. irregulare, P. sylvaticum, and P. ultimum) were examined at three forest nurseries using simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) markers. Analyses revealed distinct patterns of intraspecific variation for the three species. P. sylvaticum exhibited the most diversity, followed by P. irregulare, while substantial clonality was found in P. ultimum. For both P. irregulare and P. sylvaticum, but not P. ultimum, there was evidence for significant variation among nurseries. However, all three species also exhibited at least two distinct lineages not associated with the nursery of origin. Finally, evidence was found that certain lineages and clonal genotypes, including fungicide-resistant isolates, are shared among nurseries, indicating that pathogen movement has occurred.