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July 7, 2019

Genomic analysis of Clavibacter michiganensis reveals insight into virulence strategies and genetic diversity of a gram-positive bacterial pathogen.

Clavibacter michiganensis subsp. michiganensis is a gram-positive bacterial pathogen that proliferates in the xylem vessels of tomato, causing bacterial canker disease. In this study, we sequenced and assembled genomes of 11 C. michiganensis subsp. michiganensis strains isolated from infected tomato fields in California as well as five Clavibacter strains that colonize tomato endophytically but are not pathogenic in this host. The analysis of the C. michiganensis subsp. michiganensis genomes supported the monophyletic nature of this pathogen but revealed genetic diversity among strains, consistent with multiple introduction events. Two tomato endophytes that clustered phylogenetically with C. michiganensis strains capable of infecting wheat and pepper and were also able to cause disease in these plants. Plasmid profiles of the California strains were variable and supported the essential role of the pCM1-like plasmid and the CelA cellulase in virulence, whereas the absence of the pCM2-like plasmid in some pathogenic C. michiganensis subsp. michiganensis strains revealed it is not essential. A large number of secreted C. michiganensis subsp. michiganensis proteins were carbohydrate-active enzymes (CAZymes). Glycome profiling revealed that C. michiganensis subsp. michiganensis but not endophytic Clavibacter strains is able to extensively alter tomato cell-wall composition. Two secreted CAZymes found in all C. michiganensis subsp. michiganensis strains, CelA and PelA1, enhanced pathogenicity on tomato. Collectively, these results provide a deeper understanding of C. michiganensis subsp. michiganensis diversity and virulence strategies.

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July 7, 2019

Recombination of virulence genes in divergent Acidovorax avenae strains that infect a common host.

Bacterial etiolation and decline (BED), caused by Acidovorax avenae, is an emerging disease of creeping bentgrass on golf courses in the United States. We performed the first comprehensive analysis of A. avenae on a nationwide collection of turfgrass- and maize-pathogenic A. avenae. Surprisingly, our results reveal that the turfgrass-pathogenic A. avenae in North America are not only highly divergent but also belong to two distinct phylogroups. Both phylogroups specifically infect turfgrass but are more closely related to maize pathogens than to each other. This suggests that, although the disease is only recently reported, it has likely been infecting turfgrass for a long time. To identify a genetic basis for the host specificity, we searched for genes closely related among turfgrass strains but distantly related to their homologs from maize strains. We found a cluster of 11 such genes generated by three ancient recombination events within the type III secretion system (T3SS) pathogenicity island. Ever since the recombination, the cluster has been conserved by strong purifying selection, hinting at its selective importance. Together our analyses suggest that BED is an ancient disease that may owe its host specificity to a highly conserved cluster of 11 T3SS genes.

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July 7, 2019

The genome of the cotton bacterial blight pathogen Xanthomonas citri pv. malvacearum strain MSCT1.

Xanthomonas citri pv. malvacearum is a major pathogen of cotton, Gossypium hirsutum L.. In this study we report the complete genome of the X. citri pv. malvacearum strain MSCT1 assembled from long read DNA sequencing technology. The MSCT1 genome is the first X. citri pv. malvacearum genome with complete coding regions for X. citri pv. malvacearum transcriptional activator-like effectors. In addition functional and structural annotations are presented in this study that will provide a foundation for future pathogenesis studies with MSCT1.

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July 7, 2019

Genome diversity and evolution in the budding yeasts (Saccharomycotina).

Considerable progress in our understanding of yeast genomes and their evolution has been made over the last decade with the sequencing, analysis, and comparisons of numerous species, strains, or isolates of diverse origins. The role played by yeasts in natural environments as well as in artificial manufactures, combined with the importance of some species as model experimental systems sustained this effort. At the same time, their enormous evolutionary diversity (there are yeast species in every subphylum of Dikarya) sparked curiosity but necessitated further efforts to obtain appropriate reference genomes. Today, yeast genomes have been very informative about basic mechanisms of evolution, speciation, hybridization, domestication, as well as about the molecular machineries underlying them. They are also irreplaceable to investigate in detail the complex relationship between genotypes and phenotypes with both theoretical and practical implications. This review examines these questions at two distinct levels offered by the broad evolutionary range of yeasts: inside the best-studied Saccharomyces species complex, and across the entire and diversified subphylum of Saccharomycotina. While obviously revealing evolutionary histories at different scales, data converge to a remarkably coherent picture in which one can estimate the relative importance of intrinsic genome dynamics, including gene birth and loss, vs. horizontal genetic accidents in the making of populations. The facility with which novel yeast genomes can now be studied, combined with the already numerous available reference genomes, offer privileged perspectives to further examine these fundamental biological questions using yeasts both as eukaryotic models and as fungi of practical importance. Copyright © 2017 by the Genetics Society of America.

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July 7, 2019

Morphological and genetic analyses of the invasive forest pathogen Phytophthora austrocedri reveal two clonal lineages colonised Britain and Argentina from a common ancestral population.

Phytophthora austrocedri is causing widespread mortality of Austrocedrus chilensis in Argentina and Juniperus communis in Britain. The pathogen has also been isolated from J. horizontalis in Germany. Isolates from Britain, Argentina and Germany are homothallic with no clear differences in the dimensions of sporangia, oogonia or oospores. Argentinian and German isolates grew faster than British isolates across a range of media and had a higher temperature tolerance although most isolates regardless of origin grew best at 15°C and all isolates were killed at 25°C. Argentinian and British isolates caused lesions on both hosts when inoculated onto A. chilensis and J. communis; however the Argentinian isolate caused longer lesions on A. chilensis than on J. communis and vice versa for the British isolate. Genetic analyses of nuclear and mitochondrial loci showed that all British isolates are identical. Argentinian isolates and the German isolate are also identical but differ from the British isolates. Single nucleotide polymorphisms are shared between the British and Argentinian isolates. It is concluded that British isolates and Argentinian isolates conform to two distinct clonal lineages of P. austrocedri founded from the same as-yet unidentified source population. These lineages should be recognised and treated as separate risks by international plant health legislation.

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July 7, 2019

Improved annotation of the insect vector of citrus greening disease: biocuration by a diverse genomics community.

The Asian citrus psyllid (Diaphorina citri Kuwayama) is the insect vector of the bacterium Candidatus Liberibacter asiaticus (CLas), the pathogen associated with citrus Huanglongbing (HLB, citrus greening). HLB threatens citrus production worldwide. Suppression or reduction of the insect vector using chemical insecticides has been the primary method to inhibit the spread of citrus greening disease. Accurate structural and functional annotation of the Asian citrus psyllid genome, as well as a clear understanding of the interactions between the insect and CLas, are required for development of new molecular-based HLB control methods. A draft assembly of the D. citri genome has been generated and annotated with automated pipelines. However, knowledge transfer from well-curated reference genomes such as that of Drosophila melanogaster to newly sequenced ones is challenging due to the complexity and diversity of insect genomes. To identify and improve gene models as potential targets for pest control, we manually curated several gene families with a focus on genes that have key functional roles in D. citri biology and CLas interactions. This community effort produced 530 manually curated gene models across developmental, physiological, RNAi regulatory and immunity-related pathways. As previously shown in the pea aphid, RNAi machinery genes putatively involved in the microRNA pathway have been specifically duplicated. A comprehensive transcriptome enabled us to identify a number of gene families that are either missing or misassembled in the draft genome. In order to develop biocuration as a training experience, we included undergraduate and graduate students from multiple institutions, as well as experienced annotators from the insect genomics research community. The resulting gene set (OGS v1.0) combines both automatically predicted and manually curated gene models.

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July 7, 2019

Comparative genome analysis of Rathayibacter tritici NCPPB 1953 with Rathayibacter toxicus strains can facilitate studies on mechanisms of nematode association and host infection.

Rathayibacter tritici, which is a Gram positive, plant pathogenic, non-motile, and rod-shaped bacterium, causes spike blight in wheat and barley. For successful pathogenesis, R. tritici is associated with Anguina tritici, a nematode, which produces seed galls (ear cockles) in certain plant varieties and facilitates spread of infection. Despite significant efforts, little research is available on the mechanism of disease or bacteria-nematode association of this bacterium due to lack of genomic information. Here, we report the first complete genome sequence of R. tritici NCPPB 1953 with diverse features of this strain. The whole genome consists of one circular chromosome of 3,354,681 bp with a GC content of 69.48%. A total of 2,979 genes were predicted, comprising 2,866 protein coding genes and 49 RNA genes. The comparative genomic analyses between R. tritici NCPPB 1953 and R. toxicus strains identified 1,052 specific genes in R. tritici NCPPB 1953. Using the BlastKOALA database, we revealed that the flexible genome of R. tritici NCPPB 1953 is highly enriched in ‘Environmental Information Processing’ system and metabolic processes for diverse substrates. Furthermore, many specific genes of R. tritici NCPPB 1953 are distributed in substrate-binding proteins for extracellular signals including saccharides, lipids, phosphates, amino acids and metallic cations. These data provides clues on rapid and stable colonization of R. tritici for disease mechanism and nematode association.

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July 7, 2019

Whole genome sequence of two Rathayibacter toxicus strains reveals a tunicamycin biosynthetic cluster similar to Streptomyces chartreusis.

Rathayibacter toxicus is a forage grass associated Gram-positive bacterium of major concern to food safety and agriculture. This species is listed by USDA-APHIS as a plant pathogen select agent because it produces a tunicamycin-like toxin that is lethal to livestock and may be vectored by nematode species native to the U.S. The complete genomes of two strains of R. toxicus, including the type strain FH-79, were sequenced and analyzed in comparison with all available, complete R. toxicus genomes. Genome sizes ranged from 2,343,780 to 2,394,755 nucleotides, with 2079 to 2137 predicted open reading frames; all four strains showed remarkable synteny over nearly the entire genome, with only a small transposed region. A cluster of genes with similarity to the tunicamycin biosynthetic cluster from Streptomyces chartreusis was identified. The tunicamycin gene cluster (TGC) in R. toxicus contained 14 genes in two transcriptional units, with all of the functional elements for tunicamycin biosynthesis present. The TGC had a significantly lower GC content (52%) than the rest of the genome (61.5%), suggesting that the TGC may have originated from a horizontal transfer event. Further analysis indicated numerous remnants of other potential horizontal transfer events are present in the genome. In addition to the TGC, genes potentially associated with carotenoid and exopolysaccharide production, bacteriocins and secondary metabolites were identified. A CRISPR array is evident. There were relatively few plant-associated cell-wall hydrolyzing enzymes, but there were numerous secreted serine proteases that share sequence homology to the pathogenicity-associated protein Pat-1 of Clavibacter michiganensis. Overall, the genome provides clear insight into the possible mechanisms for toxin production in R. toxicus, providing a basis for future genetic approaches.

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July 7, 2019

Xanthomonas adaptation to common bean is associated with horizontal transfers of genes encoding TAL effectors.

Common bacterial blight is a devastating bacterial disease of common bean (Phaseolus vulgaris) caused by Xanthomonas citri pv. fuscans and Xanthomonas phaseoli pv. phaseoli. These phylogenetically distant strains are able to cause similar symptoms on common bean, suggesting that they have acquired common genetic determinants of adaptation to common bean. Transcription Activator-Like (TAL) effectors are bacterial type III effectors that are able to induce the expression of host genes to promote infection or resistance. Their capacity to bind to a specific host DNA sequence suggests that they are potential candidates for host adaption.To study the diversity of tal genes from Xanthomonas strains responsible for common bacterial blight of bean, whole genome sequences of 17 strains representing the diversity of X. citri pv. fuscans and X. phaseoli pv. phaseoli were obtained by single molecule real time sequencing. Analysis of these genomes revealed the existence of four tal genes named tal23A, tal20F, tal18G and tal18H, respectively. While tal20F and tal18G were chromosomic, tal23A and tal18H were carried on plasmids and shared between phylogenetically distant strains, therefore suggesting recent horizontal transfers of these genes between X. citri pv. fuscans and X. phaseoli pv. phaseoli strains. Strikingly, tal23A was present in all strains studied, suggesting that it played an important role in adaptation to common bean. In silico predictions of TAL effectors targets in the common bean genome suggested that TAL effectors shared by X. citri pv. fuscans and X. phaseoli pv. phaseoli strains target the promoters of genes of similar functions. This could be a trace of convergent evolution among TAL effectors from different phylogenetic groups, and comforts the hypothesis that TAL effectors have been implied in the adaptation to common bean.Altogether, our results favour a model where plasmidic TAL effectors are able to contribute to host adaptation by being horizontally transferred between distant lineages.

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July 7, 2019

Centrochromatin of fungi.

The centromere is an essential chromosomal locus that dictates the nucleation point for assembly of the kinetochore and subsequent attachment of spindle microtubules during chromosome segregation. Research over the last decades demonstrated that centromeres are defined by a combination of genetic and epigenetic factors. Recent work showed that centromeres are quite diverse and flexible and that many types of centromere sequences and centromeric chromatin (“centrochromatin”) have evolved. The kingdom of the fungi serves as an outstanding example of centromere plasticity, including organisms with centromeres as diverse as 0.15-300 kb in length, and with different types of chromatin states for most species examined thus far. Some of the species in the less familiar taxa provide excellent opportunities to help us better understand centromere biology in all eukaryotes, which may improve treatment options against fungal infection, and biotechnologies based on fungi. This review summarizes the current knowledge of fungal centromeres and centrochromatin, including an outlook for future research.

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July 7, 2019

Complete genome sequences of the plant pathogens Ralstonia solanacearum type strain K60 and R. solanacearum race 3 biovar 2 strain UW551.

Ralstonia solanacearum is a globally distributed plant pathogen that causes bacterial wilt diseases of many crop hosts, threatening both sustenance farming and industrial agriculture. Here, we present closed genome sequences for the R. solanacearum type strain, K60, and the cool-tolerant potato brown rot strain R. solanacearum UW551, a highly regulated U.S. select agent pathogen. Copyright © 2017 Hayes et al.

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July 7, 2019

Genome sequencing and comparative genomics reveal the potential pathogenic mechanism of Cercospora sojina Hara on soybean.

Frogeye leaf spot, caused by Cercospora sojina Hara, is a common disease of soybean in most soybean-growing countries of the world. In this study, we report a high-quality genome sequence of C. sojina by Single Molecule Real-Time sequencing method. The 40.8-Mb genome encodes 11,655 predicated genes, and 8,474 genes are revealed by RNA sequencing. Cercospora sojina genome contains large numbers of gene clusters that are involved in synthesis of secondary metabolites, including mycotoxins and pigments. However, much less carbohydrate-binding module protein encoding genes are identified in C. sojina genome, when compared with other phytopathogenic fungi. Bioinformatics analysis reveals that C. sojina harbours about 752 secreted proteins, and 233 of them are effectors. During early infection, the genes for metabolite biosynthesis and effectors are significantly enriched, suggesting that they may play essential roles in pathogenicity. We further identify 13 effectors that can inhibit BAX-induced cell death. Taken together, our results provide insights into the infection mechanisms of C. sojina on soybean.© The Author 2017. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

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July 7, 2019

Genome sequence of the white-rot fungus Irpex lacteus F17, a type strain of lignin degrader fungus.

Irpex lacteus, a cosmopolitan white-rot fungus, degrades lignin and lignin-derived aromatic compounds. In this study, we report the high-quality draft genome sequence of I. lacteus F17, isolated from a decaying hardwood tree in the vicinity of Hefei, China. The genome is 44,362,654 bp, with a GC content of 49.64% and a total of 10,391 predicted protein-coding genes. In addition, a total of 18 snRNA, 842 tRNA, 15 rRNA operons and 11,710 repetitive sequences were also identified. The genomic data provides insights into the mechanisms of the efficient lignin decomposition of this strain.

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July 7, 2019

Root-associated fungal microbiota of nonmycorrhizal Arabis alpina and its contribution to plant phosphorus nutrition.

Most land plants live in association with arbuscular mycorrhizal (AM) fungi and rely on this symbiosis to scavenge phosphorus (P) from soil. The ability to establish this partnership has been lost in some plant lineages like the Brassicaceae, which raises the question of what alternative nutrition strategies such plants have to grow in P-impoverished soils. To understand the contribution of plant-microbiota interactions, we studied the root-associated fungal microbiome of Arabis alpina (Brassicaceae) with the hypothesis that some of its components can promote plant P acquisition. Using amplicon sequencing of the fungal internal transcribed spacer 2, we studied the root and rhizosphere fungal communities of A. alpina growing under natural and controlled conditions including low-P soils and identified a set of 15 fungal taxa consistently detected in its roots. This cohort included a Helotiales taxon exhibiting high abundance in roots of wild A. alpina growing in an extremely P-limited soil. Consequently, we isolated and subsequently reintroduced a specimen from this taxon into its native P-poor soil in which it improved plant growth and P uptake. The fungus exhibited mycorrhiza-like traits including colonization of the root endosphere and P transfer to the plant. Genome analysis revealed a link between its endophytic lifestyle and the expansion of its repertoire of carbohydrate-active enzymes. We report the discovery of a plant-fungus interaction facilitating the growth of a nonmycorrhizal plant under native P-limited conditions, thus uncovering a previously underestimated role of root fungal microbiota in P cycling.

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