Microbiome-Mediated Strategies to Manage Major Soil-Borne Diseases of Tomato.

The tomato (Solanum lycopersicum L.) is consumed globally as a fresh vegetable due to its high nutritional value and antioxidant properties. However, soil-borne diseases can severely limit tomato production. These diseases, such as bacterial wilt (BW), Fusarium wilt (FW), Verticillium wilt (VW), and root-knot nematodes (RKN), can significantly reduce the yield and quality of tomatoes. Using agrochemicals to combat these diseases can lead to chemical residues, pesticide resistance, and environmental pollution. Unfortunately, resistant varieties are not yet available. Therefore, we must find alternative strategies to protect tomatoes from these soil-borne diseases. One of the most promising solutions is harnessing microbial communities that can suppress disease and promote plant growth and immunity. Recent omics technologies and next-generation sequencing advances can help us develop microbiome-based strategies to mitigate tomato soil-borne diseases. This review emphasizes the importance of interdisciplinary approaches to understanding the utilization of beneficial microbiomes to mitigate soil-borne diseases and improve crop productivity.

Genotyping-by-Sequencing Reveals Population Differentiation and Linkage Disequilibrium in Alternaria linariae from Tomato.

Alternaria linariae is an economically important foliar pathogen that causes early blight disease in tomatoes. Understanding genetic diversity, population genetic structure, and evolutionary potential is crucial to contemplating effective disease management strategies. We leveraged genotyping-by-sequencing (GBS) technology to compare genome-wide variation in 124 isolates of Alternaria spp. (A. alternata, A. linariae, and A. solani) for comparative genome analysis and to test the hypotheses of genetic differentiation and linkage disequilibrium (LD) in A. linariae collected from tomatoes in western North Carolina. We performed a pangenome-aware variant calling and filtering with GBSapp and identified 53,238 variants conserved across the reference genomes of three Alternaria spp. The highest marker density was observed on chromosome 1 (7 Mb). Both discriminant analysis of principal components and Bayesian model-based STRUCTURE analysis of A. linariae isolates revealed three subpopulations with minimal admixture. The genetic differentiation coefficients (FST) within A. linariae subpopulations were similar and high (0.86), indicating that alleles in the subpopulations are fixed and the genetic structure is likely due to restricted recombination. Analysis of molecular variance indicated higher variation among populations (89%) than within the population (11%). We found long-range LD between pairs of loci in A. linariae, supporting the hypothesis of low recombination expected for a fungal pathogen with limited sexual reproduction. Our findings provide evidence of a high level of population genetic differentiation in A. linariae, which reinforces the importance of developing tomato varieties with broad-spectrum resistance to various isolates of A. linariae.

Molecular mapping of quantitative trait loci for resistance to early blight in tomatoes.

Early blight (EB), caused by Alternaria linariae (Neerg.) (syn. A. tomatophila) Simmons, is a disease that affects tomatoes (Solanum lycopersicum L.) throughout the world, with tremendous economic implications. The objective of the present study was to map the quantitative trait loci (QTL) associated with EB resistance in tomatoes. The F2 and F2:3 mapping populations consisting of 174 lines derived from NC 1CELBR (resistant) × Fla. 7775 (susceptible) were evaluated under natural conditions in the field in 2011 and in the greenhouse in 2015 by artificial inoculation. In all, 375 Kompetitive Allele Specific PCR (KASP) assays were used for genotyping parents and the F2 population. The broad-sense heritability estimate for phenotypic data was 28.3%, and 25.3% for 2011, and 2015 disease evaluations, respectively. QTL analysis revealed six QTLs associated with EB resistance on chromosomes 2, 8, and 11 (LOD 4.0 to 9.1), explaining phenotypic variation ranging from 3.8 to 21.0%. These results demonstrate that genetic control of EB resistance in NC 1CELBR is polygenic. This study may facilitate further fine mapping of the EB-resistant QTL and marker-assisted selection (MAS) to transfer EB resistance genes into elite tomato varieties, including broadening the genetic diversity of EB resistance in tomatoes.

Genetic diversity and population structure of Alternaria species from tomato and potato in North Carolina and Wisconsin.

Early blight (EB) caused by Alternaria linariae or Alternaria solani and leaf blight (LB) caused by A. alternata are economically important diseases of tomato and potato. Little is known about the genetic diversity and population structure of these pathogens in the United States. A total of 214 isolates of A. alternata (n = 61), A. linariae (n = 96), and A. solani (n = 57) were collected from tomato and potato in North Carolina and Wisconsin and grouped into populations based on geographic locations and tomato varieties. We exploited 220 single nucleotide polymorphisms derived from DNA sequences of 10 microsatellite loci to analyse the population genetic structure between species and between populations within species and infer the mode of reproduction. High genetic variation and genotypic diversity were observed in all the populations analysed. The null hypothesis of the clonality test based on the index of association [Formula: see text] was rejected, and equal frequencies of mating types under random mating were detected in some studied populations of Alternaria spp., suggesting that recombination can play an important role in the evolution of these pathogens. Most genetic differences were found between species, and the results showed three distinct genetic clusters corresponding to the three Alternaria spp. We found no evidence for clustering of geographic location populations or tomato variety populations. Analyses of molecular variance revealed high (> 85%) genetic variation within individuals in a population, confirming a lack of population subdivision within species. Alternaria linariae populations harboured more multilocus genotypes (MLGs) than A. alternata and A. solani populations and shared the same MLG between populations within a species, which was suggestive of gene flow and population expansion. Although both A. linariae and A. solani can cause EB on tomatoes and potatoes, these two species are genetically differentiated. Our results provide new insights into the evolution and structure of Alternaria spp. and can lead to new directions in optimizing management strategies to mitigate the impact of these pathogens on tomato and potato production in North Carolina and Wisconsin.

RNA-Seq and Gene Regulatory Network Analyses Uncover Candidate Genes in the Early Defense to Two Hemibiotrophic Colletorichum spp. in Strawberry.

Two hemibiotrophic pathogens, Colletotrichum acutatum (Ca) and C. gloeosporioides (Cg), cause anthracnose fruit rot and anthracnose crown rot in strawberry (Fragaria × ananassa Duchesne), respectively. Both Ca and Cg can initially infect through a brief biotrophic phase, which is associated with the production of intracellular primary hyphae that can infect host cells without causing cell death and establishing hemibiotrophic infection (HBI) or quiescent (latent infections) in leaf tissues. The Ca and Cg HBI in nurseries and subsequent distribution of asymptomatic infected transplants to fruit production fields is the major source of anthracnose epidemics in North Carolina. In the absence of complete resistance, strawberry varieties with good fruit quality showing rate-reducing resistance have frequently been used as a source of resistance to Ca and Cg. However, the molecular mechanisms underlying the rate-reducing resistance or susceptibility to Ca and Cg are still unknown. We performed comparative transcriptome analyses to examine how rate-reducing resistant genotype NCS 10-147 and susceptible genotype 'Chandler' respond to Ca and Cg and identify molecular events between 0 and 48 h after the pathogen-inoculated and mock-inoculated leaf tissues. Although plant response to both Ca and Cg at the same timepoint was not similar, more genes in the resistant interaction were upregulated at 24 hpi with Ca compared with those at 48 hpi. In contrast, a few genes were upregulated in the resistant interaction at 48 hpi with Cg. Resistance response to both Ca and Cg was associated with upregulation of MLP-like protein 44, LRR receptor-like serine/threonine-protein kinase, and auxin signaling pathway, whereas susceptibility was linked to modulation of the phenylpropanoid pathway. Gene regulatory network inference analysis revealed candidate transcription factors (TFs) such as GATA5 and MYB-10, and their downstream targets were upregulated in resistant interactions. Our results provide valuable insights into transcriptional changes during resistant and susceptible interactions, which can further facilitate assessing candidate genes necessary for resistance to two hemibiotrophic Colletotrichum spp. in strawberry.

Comparative Genome Analyses of 18 Verticillium dahliae Tomato Isolates Reveals Phylogenetic and Race Specific Signatures.

Host resistance is one of the few strategies available to combat the soil borne pathogenic fungus Verticillium dahliae. Understanding pathogen diversity in populations is key to successfully deploying host resistance. In this study the genomes of 18 V. dahliae isolates of races 1 (n = 2), 2 (n = 4), and 3 (n = 12) from Japan, California, and North Carolina were sequenced and mapped to the reference genome of JR2 (from tomato). The genomes were analyzed for phylogenetic and pathogen specific signatures to classify specific strains or genes for future research. Four highly clonal lineages/groups were discovered, including a lineage unique to North Carolina isolates, which had the rare MAT1-1 mating type. No evidence for recombination between isolates of different mating types was observed, even in isolates of different mating types discovered in the same field. By mapping these 18 isolates genomes to the JR2 reference genome, 193 unique candidate effectors were found using SignalP and EffectorP. Within these effectors, 144 highly conserved effectors, 42 mutable effectors (truncated or present in some isolates but absent in others), and 7 effectors present in highly variable regions of the chromosomes were discovered. Of the 144 core effectors, 21 were highly conserved in V. alfalfae and V. longisporum, 7 of which have no known function. Within the non-core effectors 30 contained large numbers of non-synonymous mutations, while 15 of them contained indels, frameshift mutations, or were present on highly variable regions of the chromosome. Two of these highly variable region effectors (HVREs) were only present in race 2 isolates, but not in race 3 isolates. The race 1 effector Ave1 was also present in a highly variable region. These data may suggest that these highly variable regions are enriched in race determinant genes, consistent with the two-speed genome hypothesis.

Meloidogyne enterolobii, a Major Threat to Tomato Production: Current Status and Future Prospects for Its Management.

The guava root-knot nematode, Meloidogyne enterolobii (Syn. M. mayaguensis), is an emerging pathogen to many crops in the world. This nematode can cause chlorosis, stunting, and reduce yields associated with the induction of many root galls on host plants. Recently, this pathogen has been considered as a global threat for tomato (Solanum lycopersicum L.) production due to the lack of known resistance in commercially accepted varieties and the aggressiveness of M. enterolobii. Both conventional morphological and molecular approaches have been used to identify M. enterolobii, an important first step in an integrated management. To combat root-knot nematodes, integrated disease management strategies such as crop rotation, field sanitation, biocontrol agents, fumigants, and resistant cultivars have been developed and successfully used in the past. However, the resistance in tomato varieties mediated by known Mi-genes does not control M. enterolobii. Here, we review the current knowledge on geographic distribution, host range, population biology, control measures, and proposed future strategies to improve M. enterolobii control in tomato.

Opportunities and Challenges in Studies of Host-Pathogen Interactions and Management of Verticillium dahliae in Tomatoes.

Tomatoes (Solanum lycopersicum L.) are a valuable horticultural crop that are grown and consumed worldwide. Optimal production is hindered by several factors, among which Verticillium dahliae, the cause of Verticillium wilt, is considered a major biological constraint in temperate production regions. V. dahliae is difficult to mitigate because it is a vascular pathogen, has a broad host range and worldwide distribution, and can persist in soil for years. Understanding pathogen virulence and genetic diversity, host resistance, and plant-pathogen interactions could ultimately inform the development of integrated strategies to manage the disease. In recent years, considerable research has focused on providing new insights into these processes, as well as the development and integration of environment-friendly management approaches. Here, we discuss the current knowledge on the race and population structure of V. dahliae, including pathogenicity factors, host genes, proteins, enzymes involved in defense, and the emergent management strategies and future research directions for managing Verticillium wilt in tomatoes.

Pathogenomics Characterization of an Emerging Fungal Pathogen, Fusarium oxysporum f. sp. lycopersici in Greenhouse Tomato Production Systems.

In recent years, greenhouse-grown tomato (Solanum lycopersicum) plants showing vascular wilt and yellowing symptoms have been observed between 2015 and 2018 in North Carolina (NC) and considered as an emerging threat to profitability. In total, 38 putative isolates were collected from symptomatic tomatoes in 12 grower greenhouses and characterized to infer pathogenic and genomic diversity, and mating-type (MAT) idiomorphs distribution. Morphology and polymerase chain reaction (PCR) markers confirmed that all isolates were Fusarium oxysporum f. sp. lycopersici (FOL) and most of them were race 3. Virulence analysis on four different tomato cultivars revealed that virulence among isolates, resistance in tomato cultivars, and the interaction between the isolates and cultivars differed significantly (P < 0.001). Cultivar 'Happy Root' (I-1, I-2, and I-3 genes for resistance) was highly resistant to FOL isolates tested. We sequenced and examined for the presence of 15 pathogenicity genes from different classes (Fmk1, Fow1, Ftf1, Orx1, Pda1, PelA, PelD, Pep1, Pep2, eIF-3, Rho1, Scd1, Snf1, Ste12, and Sge1), and 14 Secreted In Xylem (SIX) genes to use as genetic markers to identify and differentiate pathogenic isolates of FOL. Sequence data analysis showed that five pathogenicity genes, Fmk1, PelA, Rho1, Sge1, and Ste12 were present in all isolates while Fow1, Ftf1, Orx1, Peda1, Pep1, eIF-3, Scd1, and Snf1 genes were dispersed among isolates. Two genes, Pep2 and PelD, were absent in all isolates. Of the 14 SIX genes assessed, SIX1, SIX3, SIX5, SIX6, SIX7, SIX8, SIX12, and SIX14 were identified in most isolates while the remaining SIX genes varied among isolates. All isolates harbored one of the two mating-type (MAT-1 or MAT-2) idiomorphs, but not both. The SIX4 gene was present only in race 1 isolates. Diversity assessments based on sequences of the effector SIX3- and the translation elongation factor 1-α encoding genes SIX3 and tef1-α, respectively were the most informative to differentiate pathogenic races of FOL and resulted in race 1, forming a monophyletic clade while race 3 comprised multiple clades. Furthermore, phylogeny-based on SIX3- and tef1-α gene sequences showed that the predominant race 3 from greenhouse production systems significantly overlapped with previously designated race 3 isolates from various regions of the globe.

Gene Genealogies Reveal High Nucleotide Diversity and Admixture Haplotypes Within Three Alternaria Species Associated with Tomato and Potato.

Early blight (EB) and leaf blight are two destructive diseases of tomato in North Carolina (NC), caused by Alternaria linariae and A. alternata, respectively. During the last decade, EB caused by A. solani has increased in potato-producing areas in Wisconsin (WI). We collected 152 isolates of three Alternaria spp. associated with tomato and potato in NC and WI and used the gene genealogical approach to compare the genetic relationships among them. Two nuclear genes: the glyceraldehyde-3-phosphate dehydrogenase (GPDH), RNA polymerase second largest subunit (RPB2), and the rDNA internal transcribed spacer (ITS) region of these isolates were sequenced. Besides, sequences of the GPDH locus from international isolates described in previous studies were included for comparison purposes. A set of single nucleotide polymorphisms was assembled to identify locus-specific and species-specific haplotypes. Nucleotide diversity varied among gene sequences and species analyzed. For example, the estimates of nucleotide diversity and Watterson's theta were higher in A. alternata than in A. linariae and A. solani. There was little or no polymorphisms in the ITS sequences and thus restricted haplotype placement. The RPB2 sequences were less informative to detect haplotype diversity in A. linariae and A. solani, yet six haplotypes were detected in A. alternata. The GPDH sequences enabled strongly supported phylogenetic inferences with the highest haplotype diversity and belonged to five haplotypes (AaH1 to AaH5), which consisted of only A. alternata from NC. However, 13 haplotypes were identified within and among A. linariae and A. solani sequences. Among them, six (AsAlH1 to AsAlH6) were identical to previously reported haplotypes in global samples and the remaining were new haplotypes. The most divergent haplotypes were AaH1, AsAlH2/AsAlH3, and AsAlH4 and consisted exclusively of A. alternata, A. linariae, and A. solani, respectively. Neutrality tests suggested an excess of mutations and population expansion, and selection may play an important role in nucleotide diversity of Alternaria spp.

Advances and Challenges in Bacterial Spot Resistance Breeding in Tomato (Solanum lycopersicum L.).

Bacterial spot is a serious disease of tomato caused by at least four species of Xanthomonas. These include X. euvesicatoria (race T1), X. vesicatoria (race T2), X. perforans (races T3 and T4), and X. gardneri, with the distinct geographical distribution of each group. Currently, X. gardneri and X. perforans are two major bacterial pathogens of tomato in North America, with X. perforans (race T4) dominating in east-coast while X. gardneri dominating in the Midwest. The disease causes up to 66% yield loss. Management of this disease is challenging due to the lack of useful chemical control measures and commercial resistant cultivars. Although major genes for resistance (R) and quantitative resistance have been identified, breeding tomato for resistance to bacterial spot has been impeded by multiple factors including the emergence of new races of the pathogen that overcome the resistance, multigenic control of the resistance, linkage drag, non-additive components of the resistance and a low correlation between seedling assays and field resistance. Transgenic tomato with Bs2 and EFR genes was effective against multiple races of Xanthomonas. However, it has not been commercialized because of public concerns and complex regulatory processes. The genomics-assisted breeding, effectors-based genomics breeding, and genome editing technology could be novel approaches to achieve durable resistance to bacterial spot in tomato. The main goal of this paper is to understand the current status of bacterial spot of tomato including its distribution and pathogen diversity, challenges in disease management, disease resistance sources, resistance genetics and breeding, and future prospectives with novel breeding approaches.

Assessing Rate-Reducing Foliar Resistance to Anthracnose Crown Rot and Fruit Rot in Strawberry.

Anthracnose fruit rot and anthracnose crown rot (ACR) caused by two species complexes of the fungus referred to as Colletotrichum acutatum and Colletotrichum gloeosporioides, respectively, are major pathogens of strawberry in North Carolina. Anthracnose epidemics are common when susceptible cultivars and asymptomatic planting stocks carrying quiescent Colletotrichum infection or hemibiotrophic infection (HBI) are planted. The main objective of this study was to assess resistance to HBI and ACR in strawberry. Strawberry cultivars and breeding lines were spray inoculated with isolates of C. acutatum or C. gloeosporioides. Four epidemiological parameters providing estimates of rate-reducing resistance to HBI and ACR in strawberry cultivars and lines were evaluated in repeated experiments in controlled environments in a greenhouse. HBI severity, measured as the percentage of total leaf area covered by acervuli, was estimated visually and by image analysis. ACR severity was rated weekly for wilt symptoms, and relative area under disease progress curve scores were calculated for comparing strawberry cultivars and lines. Significant differences (P ≤ 0.005) in HBI severity were found among strawberry genotypes; however, the correlations were not remarkable between Colletotrichum species (r = 0.4251). Although significant variation in resistance was observed for ACR, this was also weakly correlated (r = 0.2430) with resistance to C. gloeosporioides HBI. Overall, rate-reducing resistance to HBI and ACR in strawberry identified in this study could be utilized in breeding programs to develop durable resistance to anthracnose in North Carolina.

Phenotypic and Genetic Diversity of Xanthomonas perforans Populations from Tomato in North Carolina.

Bacterial spot caused by Xanthomonas spp. is one of the most devastating diseases of tomato in North Carolina (NC). In total, 290 strains of Xanthomonas spp. from tomato in NC collected over 2 years (2015 and 2016) were analyzed for phenotypic and genetic diversity. In vitro copper and streptomycin sensitivity assays revealed that >95% (n = 290) of the strains were copper tolerant in both years, whereas 25% (n = 127) and 46% (n = 163) were streptomycin tolerant in 2016 and 2015, respectively. Using BOX repetitive element PCR assay, fingerprint patterns showed four haplotypes (H1, H2, H3, and H4) among the strains analyzed. The multiplex real-time quantitative PCR on a subset of representative strains (n = 45) targeting the highly conserved hrcN gene identified Xanthomonas strains from tomato in NC that belonged to X. perforans. Race profiling of the representative strains (n = 45) on tomato and pepper differentials confirmed that ∼9 and 91% of strains are tomato races T3 and T4, respectively. Additionally, PCR assays and sequence alignments confirmed that the copL, copA, copB (copLAB copper tolerance gene cluster), and avrXv4 genes are present in the strains analyzed. Phylogenetic and comparative sequence analyses of six genomic regions (elongation factor G [fusA], glyceraldehyde-3-phosphate dehydrogenase A [gapA], citrate synthase [gltA], gyrase subunit B [gyrB], ABC transporter sugar permease [lacF], and GTP binding protein [lepA]) suggested that 13 and 74% of X. perforans strains from NC were genetically similar to races T3 and T4 from Florida, respectively. Our results provide insights that bacterial spot management practices in tomato should focus on deploying resistance genes to combat emerging pathogenic races of X. perforans and overcome the challenges currently posed by intense use of copper-based bactericides.

Inheritance of Resistance to Colletotrichum gloeosporioides and C. acutatum in Strawberry.

Information on the inheritance of resistance to Colletotrichum gloeosporioides and C. acutatum hemibiotrophic infections (HBI) in strawberry leaf tissue and the genetic control of anthracnose crown rot (ACR) in crown tissue are relatively unknown. Six parental genotypes were crossed in a half-diallel mating design to generate 15 full-sib families. HBI and ACR experiments were conducted concurrently. Both seedlings and parental clones were inoculated with 1 × 106 conidia/ml of C. gloeosporioides or C. acutatum. Percent sporulating leaf area, wilt symptoms, and relative area under the disease progress curve were calculated to characterize resistance among genotypes and full-sib families. Low dominance/additive variance ratios for C. acutatum HBI (0.13) and C. gloeosporioides ACR (0.20) were observed, indicating additive genetic control of resistance to these traits. Heritability estimates were low for C. acutatum HBI (0.25) and C. gloeosporioides HBI (0.16) but moderate for C. gloeosporioides ACR (0.61). A high genetic correlation (rA = 0.98) between resistance to C. acutatum HBI and C. gloeosporioides HBI was observed, suggesting that resistance to these two Colletotrichum spp. may be controlled by common genes in strawberry leaf tissue. In contrast, negative genetic correlations between ACR and both HBI traits (rA = -0.85 and -0.61) suggest that resistance in crown tissue is inherited independently of resistance in leaf tissue in the populations tested. Overall, these findings provide valuable insight into the genetic basis of resistance, and the evaluation and deployment of resistance to HBIs and ACR in strawberry breeding programs.

A New Map Location of Gene Stb3 for Resistance to Septoria Tritici Blotch in Wheat.

Septoria tritici blotch (STB), caused by Mycosphaerella graminicola (synonym: Zymoseptoria tritici; asexual stage: Septoria tritici), is an important disease of wheat worldwide. Management of the disease usually is by host resistance or fungicides. However, M. graminicola has developed insensitivity to most commonly applied fungicides so there is a continuing need for well-characterized sources of host resistance to accelerate the development of improved wheat cultivars. Gene Stb3 has been a useful source of major resistance, but its mapping location has not been well characterized. Based on linkage to a single marker, a previous study assigned Stb3 to a location on the short arm of chromosome 6D. However, the results from the present study show that this reported location is incorrect. Instead, linkage analysis revealed that Stb3 is located on the short arm of wheat chromosome 7A, completely linked to microsatellite (SSR) locus Xwmc83 and flanked by loci Xcfa2028 (12.4 cM distal) and Xbarc222 (2.1 cM proximal). Linkage between Stb3 and Xwmc83 was validated in BC1F3 progeny of other crosses, and analyses of the flanking markers with deletion stocks showed that the gene is located on 7AS between fraction lengths 0.73 and 0.83. This revised location of Stb3 is different from those for other STB resistance genes previously mapped in hexaploid wheat but is approximately 20 cM proximal to an STB resistance gene mapped on the short arm of chromosome 7Am in Triticum monococcum. The markers described in this study are useful for accelerating the deployment of Stb3 in wheat breeding programs.

Genome-wide association study reveals novel quantitative trait Loci associated with resistance to multiple leaf spot diseases of spring wheat.

Accelerated wheat development and deployment of high-yielding, climate resilient, and disease resistant cultivars can contribute to enhanced food security and sustainable intensification. To facilitate gene discovery, we assembled an association mapping panel of 528 spring wheat landraces of diverse geographic origin for a genome-wide association study (GWAS). All accessions were genotyped using an Illumina Infinium 9K wheat single nucleotide polymorphism (SNP) chip and 4781 polymorphic SNPs were used for analysis. To identify loci underlying resistance to the major leaf spot diseases and to better understand the genomic patterns, we quantified population structure, allelic diversity, and linkage disequilibrium. Our results showed 32 loci were significantly associated with resistance to the major leaf spot diseases. Further analysis identified QTL effective against major leaf spot diseases of wheat which appeared to be novel and others that were previously identified by association analysis using Diversity Arrays Technology (DArT) and bi-parental mapping. In addition, several identified SNPs co-localized with genes that have been implicated in plant disease resistance. Future work could aim to select the putative novel loci and pyramid them in locally adapted wheat cultivars to develop broad-spectrum resistance to multiple leaf spot diseases of wheat via marker-assisted selection (MAS).

Phenotypic and Molecular Diversity of Cochliobolus sativus Populations from Wheat.

Spot blotch, caused by Cochliobolus sativus, is a devastating foliar disease of wheat in Nepal and in the Northern Great Plains of the United States. However, limited information on variation in virulence and genetic structure of C. sativus from wheat is available. In this study, pathogenic variation of 96 isolates of C. sativus from the Hill and Plain areas in Nepal (n = 48) and in the Central and Northern areas in North Dakota (n = 48) were evaluated on 12 differential wheat lines. DNA polymorphisms in all isolates were analyzed using eight selected amplified fragment length polymorphism primer combinations. Phenotypic data analysis showed the isolates varied greatly and were classified into 47 pathotypes. Cluster analysis indicated the isolates fell into three distinct groups with low, intermediate, and high virulence. Population genetic analysis revealed significant linkage disequilibrium ( = 0.066 to 0.292), indicating that sexual reproduction plays little or no role in evolution and disease epidemiology in wheat fields. Furthermore, the corrected standardized fixation index (G″ST = 0.05 and 0.02) showed no evidence of genetic differentiation in C. sativus populations. Collectively, these results confirmed high pathogenic and molecular diversity in the C. sativus populations collected from wheat foliar infections and will be useful to assist in developing resistant cultivars to manage this disease.

Pathogenic and genetic diversity of Xanthomonas translucens pv. undulosa in North Dakota.

Bacterial leaf streak (BLS), caused by Xanthomonas translucens pv. undulosa, has become more prevalent recently in North Dakota and neighboring states. From five locations in North Dakota, 226 strains of X. translucens pv. undulosa were collected and evaluated for pathogenicity and then selected strains were inoculated on a set of 12 wheat cultivars and other cereal hosts. The genetic diversity of all strains was determined using repetitive sequence-based polymerase chain reaction (rep-PCR) and insertion sequence-based (IS)-PCR. Bacterial strains were pathogenic on wheat and barley but symptom severity was greatest on wheat. Strains varied greatly in aggressiveness, and wheat cultivars also showed differential responses to several strains. The 16S ribosomal DNA sequences of the strains were identical, and distinct from those of the other Xanthomonas pathovars. Combined rep-PCR and IS-PCR data produced 213 haplotypes. Similar haplotypes were detected in more than one location. Although diversity was greatest (≈92%) among individuals within a location, statistically significant (P ≤ 0.001 or 0.05) genetic differentiation among locations was estimated, indicating geographic differentiation between pathogen populations. The results of this study provide information on the pathogen diversity in North Dakota, which will be useful to better identify and characterize resistant germplasm.

Genetic differentiation at microsatellite loci among populations of Mycosphaerella graminicola from California, Indiana, Kansas, and North Dakota.

Mycosphaerella graminicola causes Septoria tritici blotch (STB) in wheat (Triticum aestivum) and is considered one of the most devastating pathogens of that crop in the United States. Although the genetic structures of M. graminicola populations from different countries have been analyzed using various molecular markers, relatively little is known about M. graminicola populations from geographically distinct areas of the United States and, in particular, of those from spring versus winter wheat. These are exposed to great differences in environmental conditions, length and season of host-free periods, and resistance sources used in geographically separated wheat breeding programs. Thus, there is more likely to be genetic differentiation between populations from spring versus winter wheat than there is among those within each region. To test this hypothesis, 330 single-spore isolates of M. graminicola representing 11 populations (1 from facultative winter wheat in California, 2 from spring wheat in North Dakota, and 8 from winter wheat in Indiana and Kansas) were analyzed for mating type frequency and for genetic variation at 17 microsatellite or simple-sequence repeat (SSR) loci. Analysis of clone-corrected data revealed an equal distribution of both mating types in the populations from Kansas, Indiana, and North Dakota, but a deviation from a 1:1 ratio in the California population. In total, 306 haplotypes were detected, almost all of which were unique in all 11 populations. High levels of gene diversity (H = 0.31 to 0.56) were observed within the 11 populations. Significant (P ≤ 0.05) gametic disequilibrium, as measured by the index of association (rBarD), was observed in California, one Indiana population (IN1), and three populations (KS1, KS2, and KS3) in Kansas that could not be explained by linkage. Corrected standardized fixation index (G″(ST)) values were 0.000 to 0.621 between the 11 populations and the majority of pairwise comparisons were statistically significant (P ≤ 0.001), suggesting some differentiation between populations. Analysis of molecular variance showed that there was a small but statistically significant level of genetic differentiation between populations from spring versus winter wheat. However, most of the total genetic variation (>98%) occurred within spring and winter wheat regions while <2% was due to genetic differentiation between these regions. Taken together, these results provide evidence that sexual recombination occurs frequently in the M. graminicola populations sampled and that most populations are genetically differentiated over the major spring- and winter-wheat-growing regions of the United States.

Association mapping of quantitative resistance to Phaeosphaeria nodorum in spring wheat landraces from the USDA National Small Grains Collection.

Stagonospora nodorum blotch (SNB), caused by Phaeosphaeria nodorum, is a destructive disease of wheat (Triticum aestivum) found throughout the United States. Host resistance is the only economically feasible option for managing the disease; however, few SNB-resistant wheat cultivars are known to exist. In this study, we report findings from an association mapping (AM) of resistance to P. nodorum in 567 spring wheat landraces of diverse geographic origin. The accessions were evaluated for seedling resistance to P. nodorum in a greenhouse. Phenotypic data and 625 polymorphic diversity array technology (DArT) markers have been used for linkage disequilibrium (LD) and association analyses. The results showed that seven DArT markers on five chromosomes (2D, 3B, 5B, 6A, and 7A) were significantly associated with resistance to P. nodorum. Genetic regions on 2D, 3B, and 5B correspond to previously mapped quantitative trait loci (QTL) conferring resistance to P. nodorum whereas the remaining QTL appeared to be novel. These results demonstrate that the use of AM is an effective method for identifying new genomic regions associated with resistance to P. nodorum in spring wheat landraces. Additionally, the novel resistance found in this study could be useful in wheat breeding aimed at controlling SNB.