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In order to study the genetic variation and phylogenetic relationship in Elymus repens, amplified fragment length polymorphism (AFLP) were used, together with sequence data for the nuclear gene phytochrome B, phyB, and the chloroplast ribosomal protein encoding gene rps4. A total of 83 collected E. repens samples, 3 E. repens reference samples and 18 related species accessions were analysed and compared with 13 GenBank sequences. AMOVA analysis revealed a moderate genetic differentiation between the populations of E. repens in the three Swedish provinces investigated, while no differentiation was observed due to landscape type. A moderate genetic differentiation was also found when samples from different fields in one province were compared to samples from a selected field. A common female origin was found in E. repens and seven other Elymus species, Pseudoroegneria spicata, Thinopyrum intermedium, T. junceum, Hordeum bogdanii and H. stenostachys. The latter two both harbour the H genome. Taken together, the data suggest that the Swedish E. repens population is slightly heterogeneous and comprises multiple origins of genome donors; a nuclear genome with contributions from Pseudoroegneria (St), Hordeum (H), Thinopyrum (E) and Y with an unknown donor together with a maternal genome donated from Pseudoroegneria. [ABSTRACT FROM AUTHOR]

The inheritance of resistance to Leptosphaeria maculans was studied in near-isogenic lines derived from asymmetric somatic hybrids between Brassica napus + Brassica nigra and Brassica napus + Brassica juncea, respectively. The hybrids had been backcrossed to B. napus for seven generations before the genetic segregation of the blackleg resistance was determined. The results of the inheritance studies suggested that one single dominant allele controls the resistance in the Brassica napojuncea line, whereas two independent dominant loci were found in the Brassica naponigra line. Total leaf DNA from the near-isogenic lines was isolated and 89 loci were detected by hybridization to 66 restriction fragment length polymorphism (RFLP) markers previously mapped in the B. nigra genome. Out of the 89 loci, eight loci were detected in the B. naponigra line and six were found in the B. napojuncea line. RFLP markers co-segregating with blackleg resistance in adult leaves were also found. Two markers associated with linkage group 5 and 8, respectively, of the B genome were found in the B. naponigra line and one marker was associated with linkage group 2 in the B. napojuncea line. [ABSTRACT FROM AUTHOR]

Out of 400 reciprocal crosses made between Brassica napus and Arabidopsis thaliana, 200 maternal B. napus flowers produced 31 seeds and 10 seeds developed when A. thaliana was used as the female parent. Of the latter, three seeds successfully germinated but did not develop into mature plants. In contrast, plants from all 31 seeds generated from B. napus were established. DNA from these 31 plants was analysed by 18 probes mapped to the A. thaliana genome, two selectable marker genes and one A. thaliana species-specific repetitive probe. It was found that one plant had identical restriction fragments to A. thaliana with two of the 21 probes used. The gene flow within Brassicaceae is discussed in light of this result. [ABSTRACT FROM AUTHOR]

Abstract Background The fungal pathogen Setosphaeria turcica causes turcicum or northern leaf blight disease on maize, sorghum and related grasses. A prevalent foliar disease found worldwide where the two host crops, maize and sorghum are grown. The aim of the present study was to find genes controlling the host defense response to this devastating plant pathogen. A cDNA-AFLP approach was taken to identify candidate sequences, which functions were further validated via virus induced gene silencing (VIGS), and real-time PCR analysis. Phylogenetic analysis was performed to address evolutionary events. Results cDNA-AFLP analysis was run on susceptible and resistant sorghum and maize genotypes to identify resistance-related sequences. One CC-NB-LRR encoding gene GRMZM2G005347 was found among the up-regulated maize transcripts after fungal challenge. The new plant resistance gene was designated as St referring to S. turcica. Genome sequence comparison revealed that the CC-NB-LRR encoding St genes are located on chromosome 2 in maize, and on chromosome 5 in sorghum. The six St sorghum genes reside in three pairs in one locus. When the sorghum St genes were silenced via VIGS, the resistance was clearly compromised, an observation that was supported by real-time PCR. Database searches and phylogenetic analysis suggest that the St genes have a common ancestor present before the grass subfamily split 50-70 million years ago. Today, 6 genes are present in sorghum, 9 in rice and foxtail millet, respectively, 3 in maize and 4 in Brachypodium distachyon. The St gene homologs have all highly conserved sequences, and commonly reside as gene pairs in the grass genomes. Conclusions Resistance genes to S. turcica, with a CC-NB-LRR protein domain architecture, have been found in maize and sorghum. VIGS analysis revealed their importance in the surveillance to S. turcica in sorghum. The St genes are highly conserved in sorghum, rice, foxtail millet, maize and Brachypodium, suggesting an essential evolutionary function.

Plant defense responses to the soil-borne fungus Verticillium longisporum causing stem stripe disease on oilseed rape (Brassica napus) are poorly understood. In this study, a population of recombinant inbred lines (RILs) using the Arabidopsis accessions Sei-0 and Can-0 was established. Composite interval mapping, transcriptome data, and T-DNA mutant screening identified the NITRATE/PEPTIDE TRANSPORTER FAMILY 5.12 (AtNPF5.12) gene as being associated with disease susceptibility in Can-0. Co-immunoprecipitation revealed interaction between AtNPF5.12 and the MAJOR LATEX PROTEIN family member AtMLP6, and fluorescence microscopy confirmed this interaction in the plasma membrane and endoplasmic reticulum. CRISPR/Cas9 technology was applied to mutate the NPF5.12 and MLP6 genes in B. napus. Elevated fungal growth in the npf5.12 mlp6 double mutant of both oilseed rape and Arabidopsis demonstrated the importance of these genes in defense against V. longisporum. Colonization of this fungus depends also on available nitrates in the host root. Accordingly, the negative effect of nitrate depletion on fungal growth was less pronounced in Atnpf5.12 plants with impaired nitrate transport. In addition, suberin staining revealed involvement of the NPF5.12 and MLP6 genes in suberin barrier formation. Together, these results demonstrate a dependency on multiple plant factors that leads to successful V. longisporum root infection., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Advances in sequencing technologies and bioinformatic analyses are accelerating the quantity and quality of data from all domains of life. This rich resource has the potential to reveal a number of important incidences with respect to possible exchange of nucleic acids. Ancient events have impacted species evolution and adaptation to new ecological niches. However, we still lack a full picture of processes ongoing within and between somatic cells, gametes, and different organisms. We propose that events linked to acceptance of alien nucleic acids grossly could be divided into 2 main routes in plants: one, when plants are exposed to extreme challenges and, the second level, a more everyday or season-related stress incited by biotic or abiotic factors. Here, many events seem to comprise somatic cells. Are the transport and acceptance processes of alien sequences random or are there specific regulatory systems not yet fully understood? Following entrance into a new cell, a number of intracellular processes leading to chromosomal integration and function are required. Modification of nucleic acids and possibly exchange of sequences within a cell may also occur. Such fine-tune events are most likely very common. There are multiple questions that we will discuss concerning different types of vesicles and their roles in nucleic acid transport and possible intracellular sequence exchange between species., Competing Interests: Conflicts of interest statement: The authors declare no conflict of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of The Genetics Society of America.)

Aims: This study evaluated the red clover (Trifolium pratense) root-associated microbiota to clarify the presence of pathogenic and beneficial microorganisms in 89 Swedish field sites., Methods and Results: 16S rRNA and ITS amplicon sequencing analysis were performed on DNA extracted from the red clover root samples collected to determine the composition of the prokaryotic and eukaryotic root-associated microbe communities. Alpha and beta diversities were calculated and relative abundance of various microbial taxa and their co-occurrence were analyzed. Rhizobium was the most prevalent bacterial genus, followed by Sphingomonas, Mucilaginibacter, Flavobacterium, and the unclassified Chloroflexi group KD4-96. The Leptodontidium, Cladosporium, Clonostachys, and Tetracladium fungal genera known for endophytic, saprotrophic, and mycoparasitic lifestyles were also frequently observed in all samples. Sixty-two potential pathogenic fungi were identified with a bias toward grass pathogens and a higher abundance in samples from conventional farms., Conclusions: We showed that the microbial community was mainly shaped by geographic location and management procedures. Co-occurrence networks revealed that the Rhizobiumleguminosarum bv. trifolii was negatively associated with all fungal pathogenic taxa recognized in this study., (© The Author(s) 2023. Published by Oxford University Press on behalf of Applied Microbiology International.)

Rhizoctonia solani is a soil-borne fungus, which rarely produces any spores in culture. Hence, all inoculation procedures are based on mycelia, often as a coat on cereal kernels, placed in close vicinity to the plant to be infected. In this protocol, an inoculation method is described where the fungus is first allowed to infest a perlite-maize flour substrate for 10 days, followed by thorough soil mixing to generate uniform fungal distribution. Pre-grown seedlings are then replanted in the infested soil. Plant materials can be harvested, five (sugar beet) and ten days ( Arabidopsis ) post infection, followed by a rapid cleaning step ahead of any nucleic acid preparation. Commercial DNA or RNA extraction kits can be used or, if higher DNA yield is required, a CTAB extraction method. Our purpose was to develop a reliable and reproducible protocol to determine the infection levels in planta upon infection with R. solani . This protocol is less laborious compared to previous ones, improves the consistency of plant infection, reproducibility between experiments, and suits both a root crop and Arabidopsis . Graphic abstract: Plant infection Inoculation of R. solani Preparation of inoculumPCR analysis Overview of the R. solani infection procedure., Competing Interests: Competing interestsThe authors declare no conflict of interest., (Copyright © 2022 The Authors; exclusive licensee Bio-protocol LLC.)

Phytophthora spp. cause serious damage to plants by exploiting a large number of effector proteins and small RNAs (sRNAs). Several reports have described modulation of host RNA biogenesis and defence gene expression. Here, we analysed Phytophthora infestans Argonaute (Ago) 1 associated small RNAs during potato leaf infection. Small RNAs were co-immunoprecipitated, deep sequenced and analysed against the P. infestans and potato genomes, followed by transcript analyses and transgenic assays on a predicted target. Extensive targeting of potato and pathogen-derived sRNAs to a range of mRNAs was observed, including 638 sequences coding for resistance (R) proteins in the host genome. The single miRNA encoded by P. infestans (miR8788) was found to target a potato alpha/beta hydrolase-type encoding gene (StABH1), a protein localized to the plasma membrane. Analyses of stable transgenic potato lines harbouring overexpressed StABH1 or artificial miRNA gene constructs demonstrated the importance of StABH1 during infection by P. infestans. miR8788 knock-down strains showed reduced growth on potato, and elevated StABH1 expression levels were observed when plants were inoculated with the two knock-down strains compared to the wild-type strain 88069. The findings of our study suggest that sRNA encoded by P. infestans can affect potato mRNA, thereby expanding our knowledge of the multifaceted strategies this species uses to facilitate infection., (© 2021 The Authors. New Phytologist © 2021 New Phytologist Foundation.)

Degradome sequencing is commonly used to generate high-throughput information on mRNA cleavage sites mediated by small RNAs (sRNA). In our datasets of potato ( Solanum tuberosum , St) and Phytophthora infestans (Pi), initial predictions generated high numbers of cleavage site predictions, which highlighted the need of improved analytic tools. Here, we present an R package based on a deep learning convolutional neural network (CNN) in a machine learning environment to optimize discrimination of false from true cleavage sites. When applying smartPARE to our datasets on potato during the infection process by the late blight pathogen, 7.3% of all cleavage windows represented true cleavages distributed on 214 sites in P. infestans and 444 sites in potato. The sRNA landscape of the two organisms is complex with uneven sRNA production and cleavage regions widespread in the two genomes. Multiple targets and several cases of complex regulatory cascades, particularly in potato, was revealed. We conclude that our new analytic approach is useful for anyone working on complex biological systems and with the interest of identifying cleavage sites particularly inferred by sRNA classes beyond miRNAs.

The fungal species Rhizoctonia solani belongs to the Basidiomycota division and is a ubiquitous soil-borne pathogen. It is the main agent of the damping-off disease in seedlings and causes the root and crown rot disease in sugar beets. Plant pathogens deploy small secreted proteins, called effectors, to manipulate plant immunity in order to infect the host. Here, a gene (RsCRP1) encoded a putative effector cysteine-rich protein was cloned, expressed in Cercospora beticola and used for virulence assays. The RsCRP1 gene was highly induced upon the early-infection stage of sugar beet seedlings and disease was promoted. Confocal microscopy demonstrated localization to the chloroplasts and mitochondria upon transient expression of RsCRP1 in leaves of Nicotiana benthamiana. Further, this effector was unable to induce necrosis or to suppress hypersensitive response induced by the Avr4/Cf4 complex in N. benthamiana. Overall, these data indicate that RsCRP1 is a novel effector targeting distinct plant cell organelles in order to facilitate a successful infection at the early stages of the disease development., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Sugar beets are attacked by several pathogens that cause root damages. Rhizoctonia (Greek for "root killer") is one of them. Rhizoctonia root rot has become an increasing problem for sugar beet production and to decrease yield losses agronomical measures are adopted. Here, two partially resistant and two susceptible sugar beet genotypes were used for transcriptome analysis to discover new defense genes to this fungal disease, information to be implemented in molecular resistance breeding. Among 217 transcripts with increased expression at 2 days post-infection (dpi), three resistance-like genes were found. These genes were not significantly elevated at 5 dpi, a time point when increased expression of three Bet v I/Major latex protein (MLP) homologous genes BvMLP1, BvMLP2 and BvML3 was observed in the partially resistant genotypes. Quantitative RT-PCR analysis on diseased sugar beet seedlings validated the activity of BvMLP1 and BvMLP3 observed in the transcriptome during challenge by R. solani. The three BvMLP genes were cloned and overexpressed in Arabidopsis thaliana to further dissect their individual contribution. Transgenic plants were also compared to T-DNA mutants of orthologous MLP genes. Plants overexpressing BvMLP1 and BvMLP3 showed significantly less infection whereas additive effects were seen on Atmlp1/Atmlp3 double mutants. The data suggest that BvMLP1 and BvMLP3 may contribute to the reduction of the Rhizoctonia root rot disease in sugar beet. Impact on the defense reaction from other differential expressed genes observed in the study is discussed.

Regulatory small RNAs (sRNAs) play important roles in many fundamental processes in plant biology such as development, fertilization and stress responses. The AGO protein family has here a central importance in gene regulation based on their capacity to associate with sRNAs followed by mRNA targeting in a sequence-complementary manner. The present study explored Argonautes (AGOs) in the Solanaceae family, with emphasis on potato, Solanum tuberosum (St). A genome-wide monitoring was performed to provide a deeper insight into gene families, genomic localization, gene structure and expression profile against the potato late blight pathogen Phytophthora infestans. Among 15 species in the Solanaceae family we found a variation from ten AGOs in Nicotiana obtusifolia to 17 in N. tabacum. Comprehensive analyses of AGO phylogeny revealed duplication of AGO1, AGO10 and AGO4 paralogs during early radiation of Solanaceae. Fourteen AGOs were identified in potato. Orthologs of AGO8 and AGO9 were missing in the potato genome. However, AGO15 earlier annotated in tomato was identified. StAGO15 differs from the other paralogs having residues of different physico-chemical properties at functionally important amino acid positions. Upon pathogen challenge StAGO15 was significantly activated and hence may play a prominent role in sRNA-based regulation of potato defense.

Rhizoctonia solani (Rs) is a soil-borne pathogen with a broad host range. This pathogen incites a wide range of disease symptoms. Knowledge regarding its infection process is fragmented, a typical feature for basidiomycetes. In this study, we aimed at identifying potential fungal effectors and their function. From a group of 11 predicted single gene effectors, a rare lipoprotein A (RsRlpA), from a strain attacking sugar beet was analyzed. The RsRlpA gene was highly induced upon early-stage infection of sugar beet seedlings, and heterologous expression in Cercospora beticola demonstrated involvement in virulence. It was also able to suppress the hypersensitive response (HR) induced by the Avr4/Cf4 complex in transgenic Nicotiana benthamiana plants and functioned as an active protease inhibitor able to suppress Reactive Oxygen Species (ROS) burst. This effector contains a double-psi beta-barrel (DPBB) fold domain, and a conserved serine at position 120 in the DPBB fold domain was found to be crucial for HR suppression. Overall, R. solani seems to be capable of inducing an initial biotrophic stage upon infection, suppressing basal immune responses, followed by a switch to necrotrophic growth. However, regulatory mechanisms between the different lifestyles are still unknown.

The potato late blight pathogen Phytophthora infestans has both an asexual and a sexual mode of reproduction. In Scandinavia, the pathogen is reproducing sexually on a regular basis, whereas clonal lineages dominate in other geographical regions. This study aimed at elucidating events or key genes underlying this difference in sexual behavior. First, the transcriptomes of eight strains, known as either clonal or sexual, were compared during early stages of mating. Principal component analysis (PCA) divided the samples in two clusters A and B and a clear grouping of the mating samples together with the A1 mating type parents was observed. Induction of genes encoding DNA adenine N6-methylation (6mA) methyl-transferases clearly showed a bias toward the cluster A. In contrast, the Avrblb2 effector gene family was highly induced in most of the mating samples and was associated with cluster B in the PCA, similarly to genes coding for acetyl-transferases, which play an important role in RXLR modification prior to secretion. Avrblb2 knock-down strains displayed a reduction in virulence and oospore formation, suggesting a role during the mating process. In conclusion, a number of gene candidates important for the reproductive processes were revealed. The results suggest a possible epigenetic influence and involvement of specific RXLR effectors in mating-related processes., (Copyright © 2020 Tzelepis, Hodén, Fogelqvist, Åsman, Vetukuri and Dixelius.)

Plasmodiophora brassicae is a soil-borne pathogen that attacks roots of cruciferous plants causing clubroot disease. The pathogen belongs to the Plasmodiophorida order in Phytomyxea. Here we used long-read SMRT technology to clarify the P. brassicae e3 genomic constituents along with comparative and phylogenetic analyses. Twenty contigs representing the nuclear genome and one mitochondrial (mt) contig were generated, together comprising 25.1 Mbp. Thirteen of the 20 nuclear contigs represented chromosomes from telomere to telomere characterized by [TTTTAGGG] sequences. Seven active gene candidates encoding synaptonemal complex-associated and meiotic-related protein homologs were identified, a finding that argues for possible genetic recombination events. The circular mt genome is large (114,663 bp), gene dense and intron rich. It shares high synteny with the mt genome of Spongospora subterranea, except in a unique 12 kb region delimited by shifts in GC content and containing tandem minisatellite- and microsatellite repeats with partially palindromic sequences. De novo annotation identified 32 protein-coding genes, 28 structural RNA genes and 19 ORFs. ORFs predicted in the repeat-rich region showed similarities to diverse organisms suggesting possible evolutionary connections. The data generated here form a refined platform for the next step involving functional analysis, all to clarify the complex biology of P. brassicae.

Rhizoctonia solani is a fungal species that belongs to the fungal division Basidiomycota. It is a soil-borne pathogen that attacks a broad range of plant species and crops. Disease symptoms are commonly seen as damping off of seedlings and root rot, although it can infect plants at any developmental stage. Despite the severity of this disease, many aspects in R. solani infection biology remain unclear. Here we investigated the role of a LysM effector, previously predicted from the genome of a R. solani AG2-2IIIB strain that has sugar beet as a host. Gene expression analysis showed that RsLysM was highly induced upon sugar beet infection. When RsLysM was heterologously expressed in Cercospora beticola, necrotic lesion size and fungal colonization ability were increased, indicating a role in virulence. RsLysM displayed chitin-binding affinity and suppression of chitin-triggered immunity but could not protect hyphae from hydrolysis. Overall, this study is the first characterization of a LysM effector from Basidiomycota, suggesting that this necrotrophic fungal species relies on perturbation of chitin-triggered immunity to establish a successful infection.

Drought stress often leads to reduced yields and is a perilous delimiter for expanded cultivation and increased productivity of sweet potato. Cell wall stabilization proteins have been identified to play a pivotal role in mechanical stabilization during desiccation stress mitigation in plants. They are involved in numerous cellular processes that modify cell wall properties to tolerate the mechanical stress during dehydration. This provides a plausible approach to engineer crops for enhanced stable yields under adverse climatic conditions. In this study, we genetically engineered sweet potato cv. Jewel with XvSap1 gene encoding a protein related to cell wall stabilization, isolated from the resurrection plant Xerophyta viscosa , under stress-inducible XvPSap1 promoter via Agrobacterium -mediated transformation. Detection of the transgene by PCR, Southern blot, and quantitative real-time PCR (qRT-PCR) analyses revealed the integration of XvSap1 in the three independent events. Phenotypic evaluation of shoot length, number of leaves, and yield revealed that the transgenic plants grew better than the wild-type plants under drought stress. Assessment of biochemical indices during drought stress showed higher levels of chlorophyll, free proline, and relative water content and decreased lipid peroxidation in transgenic plants than in wild types. Our findings demonstrate that XvSap1 enhances drought tolerance in transgenic sweet potato without causing deleterious phenotypic and yield changes. The transgenic drought-tolerant sweet potato lines provide a valuable resource as a drought-tolerant crop on arid lands of the world., (Copyright © 2019 Mbinda, Dixelius and Oduor.)

Plasmodiophora brassicae is a soil-borne pathogen that belongs to Rhizaria, an almost unexplored eukaryotic organism group. This pathogen requires a living host for growth and multiplication, which makes molecular analysis further complicated. To broaden our understanding of a plasmodiophorid such as P. brassicae, we here chose to study immunophilins, a group of proteins known to have various cellular functions, including involvement in plant defense and pathogen virulence. Searches in the P. brassicae genome resulted in 20 putative immunophilins comprising of 11 cyclophilins (CYPs), 7 FK506-binding proteins (FKBPs) and 2 parvulin-like proteins. RNAseq data showed that immunophilins were differentially regulated in enriched life stages such as germinating spores, maturing spores, and plasmodia, and infected Brassica hosts (B. rapa, B. napus and B. oleracea). PbCYP3 was highly induced in all studied life stages and during infection of all three Brassica hosts, and hence was selected for further analysis. PbCYP3 was heterologously expressed in Magnaporthe oryzae gene-inactivated ΔCyp1 strain. The new strain ΔCyp1+ overexpressing PbCYP3 showed increased virulence on rice compared to the ΔCyp1 strain. These results suggest that the predicted immunophilins and particularly PbCYP3 are activated during plant infection. M. oryzae is a well-studied fungal pathogen and could be a valuable tool for future functional studies of P. brassicae genes, particularly elucidating their role during various infection phases.

Sweetpotato is a significant crop which is widely cultivated particularly in the developing countries with high and stable yield. However, drought stress is a major limiting factor that antagonistically influences the crop's productivity. Dehydration stress caused by drought causes aggregation of reactive oxygen species (ROS) in plants, and aldose reductases are first-line safeguards against ROS caused by oxidative stress. In the present study, we generated transgenic sweetpotato plants expressing aldose reductase, XvAld1 isolated from Xerophyta viscosa under the control of a stress-inducible promoter via Agrobacterium-mediated transformation. Our results demonstrated that the transgenic sweetpotato lines displayed significant enhanced tolerance to simulated drought stress and enhanced recuperation after rehydration contrasted with wild-type plants. In addition, the transgenic plants exhibited improved photosynthetic efficiency, higher water content and more proline accumulation under dehydration stress conditions compared with wild-type plants. These results demonstrate that exploiting the XvAld1 gene is not only a compelling and attainable way to improve sweetpotato tolerance to drought stresses without causing any phenotypic imperfections but also a promising gene candidate for more extensive crop improvement.

Background: Brassica plant species are attacked by a number of pathogens; among them, the ones with a soil-borne lifestyle have become increasingly important. Verticillium stem stripe caused by Verticillium longisporum is one example. This fungal species is thought to be of a hybrid origin, having a genome composed of combinations of lineages denominated A and D. In this study we report the draft genomes of 2 V. longisporum field isolates sequenced using the Illumina technology. Genomic characterization and lineage composition, followed by selected gene analysis to facilitate the comprehension of its genomic features and potential effector categories were performed., Results: The draft genomes of 2 Verticillium longisporum single spore isolates (VL1 and VL2) have an estimated ungapped size of about 70 Mb. The total number of protein encoding genes identified in VL1 was 20,793, whereas 21,072 gene models were predicted in VL2. The predicted genome size, gene contents, including the gene families coding for carbohydrate active enzymes were almost double the numbers found in V. dahliae and V. albo-atrum. Single nucleotide polymorphisms (SNPs) were frequently distributed in the two genomes but the distribution of heterozygosity and depth was not independent. Further analysis of potential parental lineages suggests that the V. longisporum genome is composed of two parts, A1 and D1, where A1 is more ancient than the parental lineage genome D1, the latter being more closer related to V. dahliae. Presence of the mating-type genes MAT1-1-1 and MAT1-2-1 in the V. longisporum genomes were confirmed. However, the MAT genes in V. dahliae, V. albo-atrum and V. longisporum have experienced extensive nucleotide changes at least partly explaining the present asexual nature of these fungal species., Conclusions: The established draft genome of V. longisporum is comparatively large compared to other studied ascomycete fungi. Consequently, high numbers of genes were predicted in the two V. longisporum genomes, among them many secreted proteins and carbohydrate active enzyme (CAZy) encoding genes. The genome is composed of two parts, where one lineage is more ancient than the part being more closely related to V. dahliae. Dissimilar mating-type sequences were identified indicating possible ancient hybridization events.

Verticillium species are soilborne plant pathogens, responsible for big yield losses worldwide. Here, we report improved procedures to generate DNA from Verticillium species imbedded in farm soils. Using new genomic sequence information, primers for V. dahliae, V. albo-atrum, V. tricorpus, and V. longisporum were designed. In a survey of 429 samples from intensively farmed soil of two Swedish regions, only V. dahliae and V. longisporum were identified. A bias towards V. longisporum (40%) was seen in the south, whereas V. dahliae was more frequent in the western region (19%). Analyses of soil and leaf samples from 20 sugar beet fields, where foliar wilting had been observed, revealed V. dahliae DNA in all leaf and soil samples and V. longisporum in 18 soil samples, illustrating host choice and longevity of the V. longisporum microsclerotia. This study demonstrates the applicability of new molecular diagnostic tools that are important for growers of variable crops.

Clubroot disease caused by Plasmodiophora brassicae is one of the most important diseases of cultivated brassicas. P. brassicae occurs in pathotypes which differ in the aggressiveness towards their Brassica host plants. To date no DNA based method to distinguish these pathotypes has been described. In 2011 polymorphism within the 28S rDNA of P. brassicae was reported which potentially could allow to distinguish pathotypes without the need of time-consuming bioassays. However, isolates of P. brassicae from around the world analysed in this study do not show polymorphism in their LSU rDNA sequences. The previously described polymorphism most likely derived from soil inhabiting Cercozoa more specifically Neoheteromita-like glissomonads. Here we correct the LSU rDNA sequence of P. brassicae. By using FISH we demonstrate that our newly generated sequence belongs to the causal agent of clubroot disease., (Copyright © 2016 The Authors. Published by Elsevier GmbH.. All rights reserved.)