Enhancing Reniform Nematode Management in Sweetpotato by Complementing Host-Plant Resistance with Nonfumigant Nematicides.

The reniform nematode (Rotylenchulus reniformis Linford and Oliveira) adversely impacts the quality and quantity of sweetpotato storage roots. Management of R. reniformis in sweetpotato remains a challenge because host plant resistance is not available, fumigants are detrimental to the environment and health, and crop rotation is not effective. We screened a core set of 24 sweetpotato plant introductions (PIs) against R. reniformis. Four PIs were resistant, and 10 were moderately resistant to R. reniformis, suggesting these PIs can serve as sources of resistance for sweetpotato resistance breeding programs. PI 595869, PI 153907, and PI 599386 suppressed 83 to 89% egg production relative to the susceptible control 'Beauregard', and these PIs were employed in subsequent experiments to determine if their efficacy against R. reniformis can be further increased by applying nonfumigant nematicides oxamyl, fluopyram, and fluensulfone. A 34 to 93% suppression of nematode reproduction was achieved by the application of nonfumigant nematicides, with oxamyl providing the best suppression followed by fluopyram and fluensulfone. Although sweetpotato cultivars resistant to R. reniformis are currently not available and there is a need for the development of safer yet highly effective nonfumigant nematicides, results from the current study suggest that complementing host plant resistance with nonfumigant nematicides can serve as an important tool for effective and sustainable nematode management.

Identification of Sweet Potato Germplasm Resistant to Pathotypically Distinct Isolates of Meloidogyne enterolobii from the Carolinas.

Meloidogyne enterolobii (syn. mayaguensis) is an emergent species of root-knot nematode that has become a serious threat to sweet potato (Ipomoea batatas) production in the southeastern United States. The most popular sweet potato cultivars grown in this region are highly susceptible to M. enterolobii. As a result, this pest has spread across most of the sweet potato growing counties in the Carolinas, threatening the industry as well as other crops in the region. The development and release of new sweet potato cultivars with resistance to M. enterolobii would help to manage and slow the spread of this pest. To support sweet potato resistance breeding efforts, 93 accessions selected from the U.S. Department of Agriculture germplasm collection and breeding programs in the United States were screened to identify 19 lines with strong resistance to M. enterolobii. The resistance in these accessions was tested against two M. enterolobii isolates that were collected from sweet potato production fields in the Carolinas. These isolates were found to have distinct pathotypes, with galling and nematode reproduction differences observed on cotton as well as sweet potato. This study is the first report of intraspecific pathotypic variation in M. enterolobii, and it identifies sweet potato germplasm with resistance against both pathogenic variants of this nematode.

Low Genetic Diversity Suggests the Recent Introduction of Dogwood Powdery Mildew to North America.

Cornus florida (flowering dogwood) is a popular understory tree endemic to the eastern hardwood forests of the United States. In 1996, dogwood powdery mildew caused by Erysiphe pulchra, an obligate biotrophic fungus of large bracted dogwoods, reached epidemic levels throughout the C. florida growing region. In the late 1990s, both sexual and asexual stages of E. pulchra were regularly observed; thereafter, the sexual stage was found less frequently. We examined the genetic diversity and population structure of 167 E. pulchra samples on C. florida leaves using 15 microsatellite loci. Samples were organized into two separate collection zone data sets, separated as eight zones and two zones, for the subsequent analysis of microsatellite allele length data. Clone correction analysis reduced the sample size to 90 multilocus haplotypes. Our study indicated low genetic diversity, a lack of definitive population structure, low genetic distance among multilocus haplotypes, and significant linkage disequilibrium among zones. Evidence of a population bottleneck was also detected. The results of our study indicated a high probability that E. pulchra reproduces predominately via asexual conidia and lend support to the hypothesis that E. pulchra is an exotic pathogen to North America.[Formula: see text] Copyright © 2019 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Development of Genomic Resources for the Powdery Mildew, Erysiphe pulchra.

Powdery mildews (PMs) are important plant pathogens causing widespread damage. Here, we report the first draft genome of Erysiphe pulchra, the causative agent of PM of flowering dogwood, Cornus florida. The assembled genome was 63.5 Mbp and resulted in formation of 19,442 contigs (N50 = 11,686 bp) that contained an estimated 6,860 genes with a genome coverage of 62×. We found 102 candidate secreted effector proteins (CSEPs) in E. pulchra similar to E. necator genes that are potentially involved in disease development. This draft genome is an initial step for understanding the evolutionary history of the PMs and will also provide insight into evolutionary strategies that led to the wide host expansion and environmental adaptations so effectively employed by the PM lineages.

Genotypic and phenotypic evaluation of off-type grasses in hybrid Bermudagrass [Cynodon dactylon (L.) Pers. x C. transvaalensis Burtt-Davy] putting greens using genotyping-by-sequencing and morphological characterization.

BACKGROUND: Interspecific hybrid bermudagrass [Cynodon dactylon (L.) Pers. x C. transvaalensis Burtt-Davy] is one of the most widely used grasses on golf courses, with cultivars derived from 'Tifgreen' or 'Tifdwarf' particularly used for putting greens. Many bermudagrass cultivars established for putting greens can be genetically unstable and lead to the occurrence of undesirable off-type grasses that vary in phenotype. The objective of this research was to genetically and phenotypically differentiate off-type grasses and hybrid cultivars. Beginning in 2013, off-type and desirable hybrid bermudagrass samples were collected from golf course putting greens in the southeastern United States and genetically and phenotypically characterized using genotyping-by-sequencing and morphology. RESULTS: Genotyping-by-sequencing determined that 11% (5) of off-type and desirable samples from putting greens were genetically divergent from standard cultivars such as Champion, MiniVerde, Tifdwarf, TifEagle, and Tifgreen. In addition, genotyping-by-sequencing was unable to genetically distinguish all standard cultivars from one another due to their similar origin and clonal propagation; however, over 90,000 potentially informative nucleotide variants were identified among the triploid hybrid cultivars. CONCLUSIONS: Although few genetic differences were found in this research, samples harvested from golf course putting greens had variable morphology and were clustered into three distinct phenotypic groups. The majority of off-type grasses in hybrid bermudagrass putting greens were genetically similar with variable morphological traits. Off-type grasses within golf course putting greens have the potential to compromise putting surface functionality and aesthetics.

Population Structure and Genetic Diversity of Phytophthora nicotianae from Tobacco in Georgia.

Black shank, caused by Phytophthora nicotianae, occurs worldwide and is responsible for significant yield loss in tobacco production in Georgia. Management of the disease has primarily relied on utilization of tobacco cultivars with resistance to race 0 of the pathogen and application of the fungicide mefenoxam. Races of P. nicotianae currently prevalent in tobacco production in Georgia, their sensitivity to mefenoxam, and genetic diversity of the pathogen are largely unknown. To determine population structure and genetic diversity of the pathogen, simple sequence repeat (SSR) markers were used. Three races of P. nicotianae (races 0, 1, and 3) were isolated from infected tobacco plants, with race 3 identified in Georgia for the first time. The majority of isolates were identified as A2 mating type and all isolates were sensitive or intermediately sensitive to mefenoxam at 1 or 10 µg/ml, with effective concentration of mefenoxam for 50% mycelial growth reduction values ranging from <0.01 to 0.12 µg/ml. Bayesian and unweighted pair group method with arithmetic means analyses of 59 isolates using SSR markers grouped the isolates in two major groups. Group I contained 20 isolates, of which 19 isolates were collected from Berrien County. Group II contained 39 isolates collected from Bacon, Cook, Tift, and Toombs Counties as well as one sample from Berrien County. Genetic diversity of the isolates was associated with geographical location of collection, and isolates in group I were primarily (75%) race 1, whereas isolates in group II were primarily (69%) race 0. The presence of a single pathogen mating type at most of the locations implies low probability of sexual recombination that may have contributed to the low genetic diversity at a particular geographical location. Sensitivity of the isolates to mefenoxam indicates that the fungicide remains to be a potent tool for growers to combat the disease. Information generated in the study advances our knowledge about diversity and population structure of P. nicotianae, which facilitates development and implementation of effective disease management programs.

Tracking Sweet Potato Leaf Curl Virus through Field Production: Implications for Sustainable Sweetpotato Production and Breeding Practices.

Sweet potato leaf curl virus (SPLCV) is a whitefly-transmitted begomovirus infecting sweetpotato and other morning glory (Convolvulaceae) species worldwide. The virus is widespread at the USDA, ARS, U.S. Vegetable Laboratory (USVL), and testing of germplasm maintained in the breeding program indicates nearly 100% infection in storage roots of materials propagated for at least four years. Prior to the public release of new germplasm, viruses must be eliminated via laborious and time-consuming meristem-tip culture. The identification of virus-free seedlings early in the selection process can offer an alternative to meristem-tip culture. In this study, we investigated the transmission of SPLCV over two years of consecutive field plantings (early and late) of sweetpotato. While SPLCV is endemic at the USVL, virus transmission pressure over the typical cultivation season is unknown, and avoidance of virus transmission paired with the selection and maintenance of clean material may be a viable alternative to virus elimination. In 2022, the storage roots of 39 first-year seedling (FYS) selections were tested for SPLCV after early-season cultivation, revealing a single selection (2.6%) with a positive test. Similar testing was conducted in 2023 with no SPLCV-positive FYS selections detected. To further assess SPLCV acquisition in the field, replicated late-season plantings of each selected FYS (n = 37) were monitored from planting to harvest. Testing was conducted at 60 and 120 days after planting (DAP). Approximately 35% of the bulk samples were infected at 60 DAP, and infection increased to 52.3% by 120 DAP. Testing of individuals within selected positive bulked samples did not support 100% infection at harvest. Altogether, these results demonstrate that SPLCV transmission during early planting is sufficiently low to facilitate the maintenance of virus-free selections, offering an alternative to virus cleaning and a cultivation strategy that may be leveraged for production.

A Public Mid-Density Genotyping Platform for Hexaploid Sweetpotato (Ipomoea batatas [L.] Lam).

Small public breeding programs focused on specialty crops have many barriers to adopting technology, particularly creating and using genetic marker panels for genomic-based decisions in selection. Here, we report the creation of a DArTag panel of 3120 loci distributed across the sweetpotato (Ipomoea batatas [L.] Lam) genome for molecular-marker-assisted breeding and genomic prediction. The creation of this marker panel has the potential to bring cost-effective and rapid genotyping capabilities to sweetpotato breeding programs worldwide. The open access provided by this platform will allow the genetic datasets generated on the marker panel to be compared and joined across projects, institutions, and countries. This genotyping resource has the power to make routine genotyping a reality for any breeder of sweetpotato.

Metagenome-enabled models improve genomic predictive ability and identification of herbivory-limiting genes in sweetpotato.

Plant-insect interactions are often influenced by host- or insect-associated metagenomic community members. The relative abundance of insects and the microbes that modulate their interactions were obtained from sweetpotato (Ipomoea batatas) leaf-associated metagenomes using quantitative reduced representation sequencing and strain/species-level profiling with the Qmatey software. Positive correlations were found between whitefly (Bemisia tabaci) and its endosymbionts (Candidatus Hamiltonella defensa, Candidatus Portiera aleyrodidarum, and Rickettsia spp.) and negative correlations with nitrogen-fixing bacteria that implicate nitric oxide in sweetpotato-whitefly interaction. Genome-wide associations using 252 975 dosage-based markers, and metagenomes as a covariate to reduce false positive rates, implicated ethylene and cell wall modification in sweetpotato-whitefly interaction. The predictive abilities (PA) for whitefly and Ocypus olens abundance were high in both populations (68%-69% and 33.3%-35.8%, respectively) and 69.9% for Frankliniella occidentalis. The metagBLUP (gBLUP) prediction model, which fits the background metagenome-based Cao dissimilarity matrix instead of the marker-based relationship matrix (G-matrix), revealed moderate PA (35.3%-49.1%) except for O. olens (3%-10.1%). A significant gain in PA after modeling the metagenome as a covariate (gGBLUP, ≤11%) confirms quantification accuracy and that the metagenome modulates phenotypic expression and might account for the missing heritability problem. Significant gains in PA were also revealed after fitting allele dosage (≤17.4%) and dominance effects (≤4.6%). Pseudo-diploidized genotype data underperformed for dominance models. Including segregation-distorted loci (SDL) increased PA by 6%-17.1%, suggesting that traits associated with fitness cost might benefit from the inclusion of SDL. Our findings confirm the holobiont theory of host-metagenome co-evolution and underscore its potential for breeding within the context of G × G × E interactions.

Development and characterization of simple sequence repeats for Bipolaris sorokiniana and cross transferability to related species.

Simple sequence repeats (SSR) markers were developed from a small insert genomic library for Bipolaris sorokiniana, a mitosporic fungal pathogen that causes spot blotch and root rot in switchgrass. About 59% of sequenced clones (n = 384) harbored SSR motifs. After eliminating redundant sequences, 196 SSR loci were identified, of which 84.7% were dinucleotide repeats and 9.7% and 5.6% were tri- and tetra-nucleotide repeats, respectively. Primer pairs were designed for 105 loci and 85 successfully amplified loci. Sixteen polymorphic loci were characterized with 15 B. sorokiniana isolates obtained from infected switchgrass plant materials collected from five states in USA. These loci successfully cross-amplified isolates from at least one related species, including Bipolaris oryzae, Bipolaris spicifera and Bipolaris victoriae, that causes leaf spot on switchgrass. Haploid gene diversity per locus across all isolates studied varied 0.633-0.861. Principal component analysis of SSR data clustered isolates according to their respective species. These SSR markers will be a valuable tool for genetic variability and population studies of B. sorokiniana and related species that are pathogenic on switchgrass and other host plants. In addition, these markers are potential diagnostic tools for species in the genus Bipolaris.

Pityopsis ruthii: An Updated Review of Conservation Efforts for an Endangered Plant.

Pityopsis ruthii (Small) Small, Ruth's golden aster, is an endangered Asteraceae species that grows in the riparian zone along small sections of two rivers in the Southern Appalachian Mountains of the United States of America (USA). Since 1985, the species has been listed under the Endangered Species Act by the United States Fish and Wildlife Service (USFWS). The mission of the USFWS is to conserve, protect, and enhance fish, wildlife, and plants and their habitats for the continued benefit of the American people. The agency provides national leadership in the recovery and conservation of imperiled plant species by working with the scientific community to protect important habitats, increase species' populations, and identify and reduce threats to species survival with the goal of removal from federal protection. Over the past 35 years, research efforts have focused on studies designed to delineate the range and size of populations, determine habitat requirements, reproductive and propagation potential, and understand the demographic, ecological, and genetic factors that may increase vulnerability to extinction for P. ruthii. Cooperative partnerships have driven the completion of actions called for in the strategy to recover P. ruthii, and in this review, we highlight these efforts within the context of species conservation.

Mitochondrial genome datasets for the sweetpotato weevil, Cylas formicarius elegantulus (Coleoptera: Brentidae), collected in the United States.

The sweetpotato weevil, Cylas formicarius elegantulus (Summers) (Coleoptera: Brentidae), is one of the most destructive pests of sweetpotato worldwide. Genomic analyses of sweetpotato weevils can provide insights into their genetic diversity, population structure, and dispersal as well as provide information to support management strategies. Adult sweetpotato weevils were collected by various methods from Ipomoea batatas L. (sweetpotato) or I. coccinea L. (red morning glory) in the U.S. states of Georgia, Hawaii, South Carolina, and Texas. Genomic DNA was extracted from individual weevil specimens and sequenced using Illumina NovaSeq. A total of 181 GB of 150 base pair (bp) paired-end reads were generated for 40 specimens. Mitochondrial genomes were assembled for each specimen via reference mapping and annotated using Geneious Prime. Full mitochondrial genome sequences range from 17,141 to 17,152 bp with an average GC content of 21.8% and average coverage of 3307 × . A maximum likelihood phylogenetic analysis considering the mitochondrial protein coding genes is provided. Mitochondrial genomes and assembled reads are deposited in NCBI GenBank, providing 40 mitogenomes of C. formicarius elegantulus collected in the U.S.

Chemical profile and analysis of biosynthetic pathways and genes of volatile terpenes in Pityopsis ruthii, a rare and endangered flowering plant.

It is critical to gather biological information about rare and endangered plants to incorporate into conservation efforts. The secondary metabolism of Pityopsis ruthii, an endangered flowering plant that only occurs along limited sections of two rivers (Ocoee and Hiwassee) in Tennessee, USA was studied. Our long-term goal is to understand the mechanisms behind P. ruthii's adaptation to restricted areas in Tennessee. Here, we profiled the secondary metabolites, specifically in flowers, with a focus on terpenes, aiming to uncover the genomic and molecular basis of terpene biosynthesis in P. ruthii flowers using transcriptomic and biochemical approaches. By comparative profiling of the nonpolar portion of metabolites from various tissues, P. ruthii flowers were rich in terpenes, which included 4 monoterpenes and 10 sesquiterpenes. These terpenes were emitted from flowers as volatiles with monoterpenes and sesquiterpenes accounting for almost 68% and 32% of total emission of terpenes, respectively. These findings suggested that floral terpenes play important roles for the biology and adaptation of P. ruthii to its limited range. To investigate the biosynthesis of floral terpenes, transcriptome data for flowers were produced and analyzed. Genes involved in the terpene biosynthetic pathway were identified and their relative expressions determined. Using this approach, 67 putative terpene synthase (TPS) contigs were detected. TPSs in general are critical for terpene biosynthesis. Seven full-length TPS genes encoding putative monoterpene and sesquiterpene synthases were cloned and functionally characterized. Three catalyzed the biosynthesis of sesquiterpenes and four catalyzed the biosynthesis of monoterpenes. In conclusion, P. ruthii plants employ multiple TPS genes for the biosynthesis of a mixture of floral monoterpenes and sesquiterpenes, which probably play roles in chemical defense and attracting insect pollinators alike.

Ten polymorphic microsatellite loci identified from a small insert genomic library for Peronospora tabacina.

Ten polymorphic microsatellite loci for the obligate biotrophic, oomycete pathogen of tobacco, Peronospora tabacina, were identified from a small insert genomic library enriched for GT motifs. Eighty-five percent of the 162 loci identified were composed of dinucleotide repeats, whereas only 4% and 11% were tri-and tetra-nucleotide repeats respectively. About 82% of all the microsatellites were perfect and within the library; only about 7% of the loci were duplicated. Primers were designed for 63 loci; 10 loci were polymorphic, 19 were monomorphic and 34 either failed to amplify or produced ambiguous/inconsistent results. The 10 polymorphic loci were characterized with 44 isolates of P. tabacina collected from tobacco plants growing in Europe, the Near East and North and South America. The number of alleles per locus was either three or four with a mean of 3.2, and the mean number of genotypes per locus was 3.6. Observed heterozygosity was 0.32-0.95, whereas expected heterozygosity was 0.44-0.69 for these loci. All loci except PT054 did not conform to the Hardy-Weinberg distribution. Polymorphic information content (PIC) for the loci was 0.35-0.69 with a mean of 0.50. These microsatellite loci provide a set of markers sufficient to perform genetic diversity and population studies of P. tabacina, and possibly other species of Peronospora.

Characterization of expressed sequence tag-derived simple sequence repeat markers for Aspergillus flavus: emphasis on variability of isolates from the southern United States.

Simple sequence repeat (SSR) markers were developed from Aspergillus flavus expressed sequence tag (EST) database to conduct an analysis of genetic relationships of Aspergillus isolates from numerous host species and geographical regions, but primarily from the United States. Twenty-nine primers were designed from 362 tri-nucleotide EST-SSR sequences. Eighteen polymorphic loci were used to genotype 96 Aspergillus species isolates. The number of alleles detected per locus ranged from 2 to 24 with a mean of 8.2 alleles. Haploid diversity ranged from 0.28 to 0.91. Genetic distance matrix was used to perform principal coordinates analysis (PCA) and to generate dendrograms using unweighted pair group method with arithmetic mean (UPGMA). Two principal coordinates explained more than 75 % of the total variation among the isolates. One clade was identified for A. flavus isolates (n = 87) with the other Aspergillus species (n = 7) using PCA, but five distinct clusters were present when the others taxa were excluded from the analysis. Six groups were noted when the EST-SSR data were compared using UPGMA. However, the latter PCA or UPGMA comparison resulted in no direct associations with host species, geographical region or aflatoxin production. Furthermore, there was no direct correlation to visible morphological features such as sclerotial types. The isolates from Mississippi Delta region, which contained the largest percentage of isolates, did not show any unusual clustering except for isolates K32, K55, and 199. Further studies of these three isolates are warranted to evaluate their pathogenicity, aflatoxin production potential, additional gene sequences (e.g., RPB2), and morphological comparisons.

Genetic diversity, population structure, and selection of breeder germplasm subsets from the USDA sweetpotato (Ipomoea batatas) collection.

Sweetpotato (Ipomoea batatas) is the sixth most important food crop and plays a critical role in maintaining food security worldwide. Support for sweetpotato improvement research in breeding and genetics programs, and maintenance of sweetpotato germplasm collections is essential for preserving food security for future generations. Germplasm collections seek to preserve phenotypic and genotypic diversity through accession characterization. However, due to its genetic complexity, high heterogeneity, polyploid genome, phenotypic plasticity, and high flower production variability, sweetpotato genetic characterization is challenging. Here, we characterize the genetic diversity and population structure of 604 accessions from the sweetpotato germplasm collection maintained by the United States Department of Agriculture (USDA), Agricultural Research Service (ARS), Plant Genetic Resources Conservation Unit (PGRCU) in Griffin, Georgia, United States. Using the genotyping-by-sequencing platform (GBSpoly) and bioinformatic pipelines (ngsComposer and GBSapp), a total of 102,870 polymorphic SNPs with hexaploid dosage calls were identified from the 604 accessions. Discriminant analysis of principal components (DAPC) and Bayesian clustering identified six unique genetic groupings across seven broad geographic regions. Genetic diversity analyses using the hexaploid data set revealed ample genetic diversity among the analyzed collection in concordance with previous analyses. Following population structure and diversity analyses, breeder germplasm subsets of 24, 48, 96, and 384 accessions were established using K-means clustering with manual selection to maintain phenotypic and genotypic diversity. The genetic characterization of the PGRCU sweetpotato germplasm collection and breeder germplasm subsets developed in this study provide the foundation for future association studies and serve as precursors toward phenotyping studies aimed at linking genotype with phenotype.

Development of microsatellite loci for the endangered species Pityopsis ruthii (Asteraceae).

PREMISE OF THE STUDY: Microsatellite loci were developed for the endangered species Pityopsis ruthii and will permit genetic and conservation studies of the species. METHODS AND RESULTS: A microsatellite-enriched library was used to develop 12 polymorphic microsatellite loci for P. ruthii. The loci amplified perfect and imperfect repeats with three to seven alleles per locus. Observed heterozygosity ranged from 0.05 to 0.80 and expected heterozygosity ranged from 0.23 to 0.75. CONCLUSIONS: These microsatellite loci provide a sufficient set of markers for further investigation of population genetics of P. ruthii.

Large-Scale Seedling Grow-Out Experiments Do Not Support Seed Transmission of Sweet Potato Leaf Curl Virus in Sweet Potato.

Sweet potato leaf curl virus (SPLCV) threatens global sweet potato production. SPLCV is transmitted by Bemisia tabaci or via infected vegetative planting materials; however, SPLCV was suggested to be seed transmissible, which is a characteristic that is disputed for geminiviruses. The objective of this study was to revisit the validity of seed transmission of SPLCV in sweet potato. Using large-scale grow-out of sweet potato seedlings from SPLCV-contaminated seeds over 4 consecutive years, approximately 23,034 sweet potato seedlings of 118 genotype entries were evaluated. All seedlings germinating in a greenhouse under insect-proof conditions or in a growth chamber were free of SPLCV; however, a few seedlings grown in an open bench greenhouse lacking insect exclusion tested positive for SPLCV. Inspection of these seedlings revealed that B. tabaci had infiltrated the greenhouse. Therefore, transmission experiments were conducted using B. tabaci MEAM1, demonstrating successful vector transmission of SPLCV to sweet potato. Additionally, tests on contaminated seed coats and germinating cotyledons demonstrated that SPLCV contaminated a high percentage of seed coats collected from infected maternal plants, but SPLCV was never detected in emerging cotyledons. Based on the results of grow-out experiments, seed coat and cotyledon tests, and vector transmission experiments, we conclude that SPLCV is not seed transmitted in sweet potato.

"Jumping Jack": Genomic Microsatellites Underscore the Distinctiveness of Closely Related Pseudoperonospora cubensis and Pseudoperonospora humuli and Provide New Insights Into Their Evolutionary Past.

Downy mildews caused by obligate biotrophic oomycetes result in severe crop losses worldwide. Among these pathogens, Pseudoperonospora cubensis and P. humuli, two closely related oomycetes, adversely affect cucurbits and hop, respectively. Discordant hypotheses concerning their taxonomic relationships have been proposed based on host-pathogen interactions and specificity evidence and gene sequences of a few individuals, but population genetics evidence supporting these scenarios is missing. Furthermore, nuclear and mitochondrial regions of both pathogens have been analyzed using microsatellites and phylogenetically informative molecular markers, but extensive comparative population genetics research has not been done. Here, we genotyped 138 current and historical herbarium specimens of those two taxa using microsatellites (SSRs). Our goals were to assess genetic diversity and spatial distribution, to infer the evolutionary history of P. cubensis and P. humuli, and to visualize genome-scale organizational relationship between both pathogens. High genetic diversity, modest gene flow, and presence of population structure, particularly in P. cubensis, were observed. When tested for cross-amplification, 20 out of 27 P. cubensis-derived gSSRs cross-amplified DNA of P. humuli individuals, but few amplified DNA of downy mildew pathogens from related genera. Collectively, our analyses provided a definite argument for the hypothesis that both pathogens are distinct species, and suggested further speciation in the P. cubensis complex.

Variability of United States isolates of Macrophomina phaseolina based on simple sequence repeats and cross genus transferability to related genera within botryosphaeriaceae.

Twelve simple sequence repeat (SSRs) loci were used to evaluate genetic diversity of 109 isolates of Macrophomina phaseolina collected from different geographical regions and host species throughout the United States (US). Genetic diversity was assessed using Nei's minimum genetic distance, and the usefulness of each locus was determined by calculating the polymorphism information content (PIC). A total of 98 alleles were detected and of these 31 were unique to individual genotypes. Eight of twelve loci were highly informative with PIC values greater than 0.50. The majority of pairwise comparisons of genetic distance were greater than 0.60 indicating moderate to high genetic diversity. Dendrograms based on the genetic dissimilarities were created for the 109 isolates of which 79 were from soybean. Some clustering by host and geography was noted, but, the dendrograms generally grouped isolates independent of host or geography. Additionally, sequencing of the internal transcribed spacer region (ITS) for 10 isolates revealed that all of these isolates were 99% similar. Three SSR loci from M. phaseolina were cross amplified in other genera in the Botryosphaeriaceae. This was the first study of genotyping and assessing genetic diversity of M. phaseolina isolates collected from a widespread host and geographic range across the US with SSRs. With an additional 34 loci publically available for M. phaseolina, the results indicate that previously developed SSRs from one species can be used in future population, ecological, and genetic studies of M. phaseolina and other genera within the Botryosphaeriaceae.