Project description:The reniform nematode (Rotylenchulus reniformis Linford and Oliveira) is a semi-endoparasitic nematode that is a pathogen of numerous major crops such as cotton and soybean. Here, the authors present transcriptome assemblies of the egg, second-stage juvenile (J2), J3, vermiform adult, and sedentary female life stages of this important plant pathogen.The reniform nematode (Rotylenchulus reniformis Linford and Oliveira) is a semi-endoparasitic nematode that is a pathogen of numerous major crops such as cotton and soybean. Here, the authors present transcriptome assemblies of the egg, second-stage juvenile (J2), J3, vermiform adult, and sedentary female life stages of this important plant pathogen.
Project description:The reniform nematode (Rotylenchulus reniformis) is a sedentary semi-endoparasitic species that is pathogenic on many row crops, fruits, and vegetables. Here, the authors present a draft genome assembly of R. reniformis using small- and large-insert libraries sequenced on the Illumina GAIIx and MiSeq platforms.The reniform nematode (Rotylenchulus reniformis) is a sedentary semi-endoparasitic species that is pathogenic on many row crops, fruits, and vegetables. Here, the authors present a draft genome assembly of R. reniformis using small- and large-insert libraries sequenced on the Illumina GAIIx and MiSeq platforms.
Project description:Rotylenchulus reniformis is the predominant parasitic nematode of cotton in the Mid South area of the United States. Although variable levels of infection and morphological differences have been reported for this nematode, genetic variability has been more elusive. We developed microsatellite-enriched libraries for R. reniformis, produced 1152 clones, assembled 694 contigs, detected 783 simple sequence repeats (SSR) and designed 192 SSR-markers. The markers were tested on six R. reniformis cultures from four states, Texas, Louisiana, Mississippi and Georgia, in the USA. Based on performance we selected 156 SSR markers for R. reniformis from which 88 were polymorphic across the six reniform nematode populations, showing as the most frequent motif the dinucleotide AG. The polymorphic information content of the markers ranged from 0.00 to 0.82, and the percentage of multiallelic loci of the isolates was between 40.9 and 45.1%. An interesting finding in this study was the genetic variability detected among the three Mississippi isolates, for which 22 SSR markers were polymorphic. We also tested the level of infection of these isolates on six cotton genotypes, where significant differences were found between the Texas and Georgia isolates. Coincidentally, 62 polymorphic markers were able to distinguish these two populations. Further studies will be necessary to establish possible connections, if any, between markers and level of pathogenicity of the nematode. The SSR markers developed here will be useful in the assessment of the genetic diversity of this nematode, could assist in management practices for control of reniform nematode, be used in breeding programs for crop resistance, and help in detecting the origin and spread of this nematode in the United States.
Project description:Reniform nematode (Rotylenchulus reniformis) is a major pest of cotton in the southeastern United States. The objective of this study was to examine the variation of reniform nematode populations from cotton-growing locations in the United States where it is prevalent. Multivariate analysis of variance and discriminant analysis were used to determine the variability of morphology in males and immature females. Reproduction indices of populations were measured on selected soybean and cotton genotypes in the greenhouse. High variability in morphometrics and reproduction was observed within all the populations, and several differences were found among populations. DNA sequences of the nuclear ribosomal first internal transcribed spacer region (ITS1) were compared among populations from the United States and to sequences of populations from Brazil, Colombia, Honduras, and Japan. No polymorphic nucleotide sites were observed among the amphimictic populations. Only a parthenogenic population from Japan was distinct. The phenotypic polymorphism of the species in the United States could impact the effectiveness of management strategies based on host plant resistance.
Project description:The 18S rRNA gene is fundamental to cellular and organismal protein synthesis and because of its stable persistence through generations it is also used in phylogenetic analysis among taxa. Sequence variation in this gene within a single species is rare, but it has been observed in few metazoan organisms. More frequently it has mostly been reported in the non-transcribed spacer region. Here, we have identified two sequence variants within the near full coding region of 18S rRNA gene from a single reniform nematode (RN) Rotylenchulus reniformis labeled as reniform nematode variant 1 (RN_VAR1) and variant 2 (RN_VAR2). All sequences from three of the four isolates had both RN variants in their sequences; however, isolate 13B had only RN variant 2 sequence. Specific variable base sites (96 or 5.5%) were found within the 18S rRNA gene that can clearly distinguish the two 18S rDNA variants of RN, in 11 (25.0%) and 33 (75.0%) of the 44 RN clones, for RN_VAR1 and RN_VAR2, respectively. Neighbor-joining trees show that the RN_VAR1 is very similar to the previously existing R. reniformis sequence in GenBank, while the RN_VAR2 sequence is more divergent. This is the first report of the identification of two major variants of the 18S rRNA gene in the same single RN, and documents the specific base variation between the two variants, and hypothesizes on simultaneous co-existence of these two variants for this gene.
Project description:The reniform nematode, Rotylenchulus reniformis, is a sedentary semi-endoparasitic species with a host range that encompasses more than 77 plant families. Nematode effector proteins containing plant-ligand motifs similar to CLAVATA3/ESR (CLE) peptides have been identified in the Heterodera, Globodera, and Meloidogyne genera of sedentary endoparasites. Here, we describe the isolation, sequence analysis, and spatiotemporal expression of three R. reniformis genes encoding putative CLE motifs named Rr-cle-1, Rr-cle-2, and Rr-cle-3. The Rr-cle cDNAs showed >98% identity with each other and the predicted peptides were identical with the exception of a short stretch of residues at the carboxy(C)-terminus of the variable domain (VD). Each RrCLE peptide possessed an amino-terminal signal peptide for secretion and a single C-terminal CLE motif that was most similar to Heterodera CLE motifs. Aligning the Rr-cle cDNAs with their corresponding genomic sequences showed three exons with an intron separating the signal peptide from the VD and a second intron separating the VD from the CLE motif. An alignment of the RrCLE1 peptide with Heterodera glycines and Heterodera schachtii CLE proteins revealed a high level of homology within the VD region associated with regulating in planta trafficking of the processed CLE peptide. Quantitative RT-PCR (qRT-PCR) showed similar expression profiles for each Rr-cle transcript across the R. reniformis life-cycle with the greatest transcript abundance being in sedentary parasitic female nematodes. In situ hybridization showed specific Rr-cle expression within the dorsal esophageal gland cell of sedentary parasitic females.
Project description:<h4>Background</h4>Reniform nematode (Rotylenchulus reniformis) has emerged as one of the most destructive root pathogens of upland cotton (Gossypium hirsutum) in the United States. Management of R. reniformis has been hindered by the lack of resistant G. hirsutum cultivars; however, resistance has been frequently identified in germplasm accessions from the G. arboreum collection. To determine the genetic basis of reniform nematode resistance, a genome-wide association study (GWAS) was performed using 246 G. arboreum germplasm accessions that were genotyped with 7220 single nucleotide polymorphic (SNP) sequence markers generated from genotyping-by-sequencing.<h4>Results</h4>Fifteen SNPs representing 12 genomic loci distributed over eight chromosomes showed association with reniform nematode resistance. For 14 SNPs, major alleles were shown to be associated with resistance. From the 15 significantly associated SNPs, 146 genes containing or physically close to these loci were identified as putative reniform nematode resistance candidate genes. These genes are involved in a broad range of biological pathways, including plant innate immunity, transcriptional regulation, and redox reaction that may have a role in the expression of resistance. Eighteen of these genes corresponded to differentially expressed genes identified from G. hirsutum in response to reniform nematode infection.<h4>Conclusions</h4>The identification of multiple genomic loci associated with reniform nematode resistance would indicate that the G. arboreum collection is a significant resource of novel resistance genes. The significantly associated markers identified from this GWAS can be used for the development of molecular tools for breeding improved reniform nematode resistant upland cotton with resistance introgressed from G. arboreum. Additionally, a greater understanding of the molecular mechanisms of reniform nematode resistance can be determined through genetic structure and functional analyses of candidate genes, which will aid in the pyramiding of multiple resistance genes.
Project description:Reniform nematodes of the genus <i>Rotylenchulus</i> are semi-endoparasites of numerous herbaceous and woody plant roots that occur largely in regions with temperate, subtropical, and tropical climates. In this study, we compared 12 populations of <i>Rotylenchulus</i><i>borealis</i> and 16 populations of <i>Rotylenchulus</i><i>macrosoma</i>, including paratypes deposited in nematode collections, confirming that morphological characters between both nematode species do not support their separation. In addition, analysis of molecular markers using nuclear ribosomal DNA (<i>28S</i>, <i>ITS1</i>) and mitochondrial DNA (<i>coxI</i>) genes, as well as phylogenetic approaches, confirmed the synonymy of <i>R. macrosoma</i> with <i>R. borealis</i>. This study also demonstrated that <i>R. borealis</i> (= <i>macrosoma</i>) from Israel has two distinct rRNA gene types in the genome, specifically the two types of <i>D2-D3</i> (A and B). We provide a global geographical distribution of the genus <i>Rotylenchulus</i>. The two major pathogenic species (<i>Rotylenchulus</i><i>reniformis</i> and <i>Rotylenchulus</i><i>parvus</i>) showed their close relationship with warmer areas with high annual mean temperature, maximum temperature of the warmest month, and minimum temperature of the coldest month. The present study confirms the extraordinary morphological and molecular diversity of <i>R. borealis</i> in Europe, Africa, and the Middle East and comprises a paradigmatic example of remarkable flexibility of ecological requirements within reniform nematodes.
Project description:Soybean (<i>Glycine max</i>) is a major plant protein source and oilseed crop. However, plant-parasitic nematodes (PPNs) affect its annual yield. In the current study, in order to better understand the regulation of defense mechanism against PPNs in soybean, we investigated the role of long non-coding RNAs (lncRNAs) in response to two nematode species, <i>Heterodera glycines</i> (SCN: soybean cyst nematode) and <i>Rotylenchulus reniformis</i> (reniform). To this end, two publicly available RNA-seq data sets (SCN data set and RAD: reniform-associated data set) were employed to discover the lncRNAome profile of soybean under SCN and reniform infection, respectively. Upon identification of unannotated transcripts in these data sets, a seven-step pipeline was utilized to sieve these transcripts, which ended up in 384 and 283 potential lncRNAs in SCN data set and RAD, respectively. These transcripts were then used to predict cis and trans nematode-related targets in soybean genome. Computational prediction of target genes function, some of which were also among differentially expressed genes, revealed the involvement of putative nematode-responsive genes as well as enrichment of multiple stress responses in both data sets. Finally, 15 and six lncRNAs were proposed to be involved in microRNA-mediated regulation of gene expression in soybean in response to SNC and reniform infection, respectively. Collectively, this study provides a novel insight into the signaling and regulatory network of soybean-pathogen interactions and opens a new window for further research.
Project description:Sedentary plant-parasitic nematodes (PPNs) induce and maintain an intimate relationship with their host, stimulating cells adjacent to root vascular tissue to re-differentiate into unique and metabolically active 'feeding sites'. The interaction between PPNs and their host is mediated by nematode effectors. We describe the discovery of a large and diverse family of effector genes, encoding C-TERMINALLY ENCODED PEPTIDE (CEP) plant hormone mimics (RrCEPs), in the syncytia-forming plant parasite Rotylenchulus reniformis. The particular attributes of RrCEPs distinguish them from all other CEPs, regardless of origin. Together with the distant phylogenetic relationship of R. reniformis to the only other CEP-encoding nematode genus identified to date (Meloidogyne), this suggests that CEPs probably evolved de novo in R. reniformis. We have characterized the first member of this large gene family (RrCEP1), demonstrating its significant up-regulation during the plant-nematode interaction and expression in the effector-producing pharyngeal gland cell. All internal CEP domains of multi-domain RrCEPs are followed by di-basic residues, suggesting a mechanism for cleavage. A synthetic peptide corresponding to RrCEP1 domain 1 is biologically active and capable of up-regulating plant nitrate transporter (AtNRT2.1) expression, whilst simultaneously reducing primary root elongation. When a non-CEP-containing, syncytia-forming PPN species (Heterodera schachtii) infects Arabidopsis in a CEP-rich environment, a smaller feeding site is produced. We hypothesize that CEPs of R. reniformis represent a two-fold adaptation to sustained biotrophy in this species: (i) increasing host nitrate uptake, whilst (ii) limiting the size of the syncytial feeding site produced.