Managing nematodes in gardens

Revised by Robert Heinz (Coordinator, MU Extension Plant Nematology Lab, Division of Plant Sciences), Melissa Goellner Mitchum (Assistant Professor, Division of Plant Sciences and Bond Life Sciences Center)"Most plants, whether naturally occurring or cultivated, have nematodes associated with them. Some nematodes are endoparasitic, spending their lives within plants (Figure 1). Others are ectoparasitic, and only their stylets — hollow, protrusible spears used to puncture plants — enter the plant to extract plant nutrients (Figure 2)."--Page 1.Robert Heinz (Coordinator, MU Extension Plant Nematology Lab, Division of Plant Sciences), Melissa Goellner Mitchum (Assistant Professor, Division of Plant Sciences and Bond Life Sciences Center)Revised 4/10; Reprinted 1/12/1.5M

Managing nematodes in gardens (2012)

Revised 4/10; Reprinted 1/12/1.5M

Identification of cyst nematode B-type CLE peptides and modulation of the vascular stem cell pathway for feeding cell formation.

Stem cell pools in the SAM (shoot apical meristem), RAM (root apical meristem) and vascular procambium/cambium are regulated by CLE-receptor kinase-WOX signaling modules. Previous data showed that cyst nematode CLE-like effector proteins delivered into host cells through a stylet, act as ligand mimics of plant A-type CLE peptides and are pivotal for successful parasitism. Here we report the identification of a new class of CLE peptides from cyst nematodes with functional similarity to the B-type CLE peptide TDIF (tracheary element differentiation inhibitory factor) encoded by the CLE41 and CLE44 genes in Arabidopsis. We further demonstrate that the TDIF-TDR (TDIF receptor)-WOX4 pathway, which promotes procambial meristem cell proliferation, is involved in beet cyst nematode Heterodera schachtii parasitism. We observed activation of the TDIF pathway in developing feeding sites, reduced nematode infection in cle41 and tdr-1 wox4-1 mutants, and compromised syncytium size in cle41, tdr-1, wox4-1 and tdr-1 wox4-1 mutants. By qRT-PCR and promoter:GUS analyses, we showed that the expression of WOX4 is decreased in a clv1-101 clv2-101 rpk2-5 mutant, suggesting that WOX4 is a potential downstream target of nematode CLEs. Exogenous treatment with both nematode A-type and B-type CLE peptides induced massive cell proliferation in wild type roots, suggesting that the two types of CLEs may regulate cell proliferation during feeding site formation. These findings highlight an important role of the procambial cell proliferation pathway in cyst nematode feeding site formation

Signal Peptide-Selection of cDNA Cloned Directly from the Esophageal Gland Cells of the Soybean Cyst Nematode Heterodera glycines

Secretions from the esophageal gland cells of plantparasitic nematodes play critical roles in the nematodeparasitic cycle. A novel method to isolate cDNA encoding putative nematode secretory proteins was developed that utilizes mRNA for reverse transcription-polymerase chain reaction derived from microaspiration of the esophageal gland cell contents of parasitic stages of the soybean cyst nematode Heterodera glycines. The resulting H. glycines gland cell cDNA was cloned into the pRK18 vector, and plasmid DNA was transformed into a mutated yeast host for specific selection of cDNA inserts that encode proteins with functional signal peptides. Of the 223 cDNA clones recovered from selection in yeast, 97% of the clones encoded a predicted signal peptide. Fourteen unique cDNA clones hybridized to genomic DNA of H. glycines on Southern blots and, among them, nine cDNA clones encoded putative extracellular proteins, as predicted by PSORT II computer analysis. Four cDNA clones hybridized to transcripts within the dorsal esophageal gland cell of parasitic stages of H. glycines, and in situ hybridization within H. glycines was not detected for eight cDNA clones. The protocol provides a direct means to isolate potential plant-parasitic nematode esophageal gland secretory protein genes

Nematode effector proteins: an emerging paradigm of parasitism

Phytonematodes use a stylet and secreted effectors to modify host cells and ingest nutrients to support their growth and development. The molecular function of nematode effectors is currently the subject of intense investigation. In this review, we summarize our current understanding of nematode effectors, with a particular focus on proteinaceous stylet-secreted effectors of sedentary endoparasitic phytonematodes, for which a wealth of information has surfaced in the past 10 yr. We provide an update on the effector repertoires of several of the most economically important genera of phytonematodes and discuss current approaches to dissecting their function. Lastly, we highlight the latest breakthroughs in effector discovery that promise to shed new light on effector diversity and function across the phylum Nematoda

Endo-β-1,4-Glucanase Expression in Compatible Plant–Nematode Interactions

Cyst nematodes and root-knot nematodes elaborately transform cells within the vascular cylinders of plant roots into enlarged, multinucleate, and metabolically active feeding cells. The giant cells of root-knot nematodes are formed by repeated karyokinesis uncoupled from cytokinesis, whereas the syncytia formed by cyst nematodes arise from coordinated cell wall dissolution and the coalescing of cell cytoplasm of adjacent cells. Both giant cells and syncytia undergo extensive cell wall architectural modifications, including thickening and the formation of numerous ingrowths that increase the plasmalemma surface area for solute uptake. The origin of enzymes involved in these cell wall modifications has been the subject of debate for several decades. Immunolocalization of endo-β-1,4-glucanases (EGases) secreted from cyst nematodes was observed in root cortical tissue during the intracellular migration of the nematodes, but secretion of cyst nematode EGases into developing syncytia was not detected. We have identified five EGase genes from tobacco that are upregulated within plant roots upon infection by both root-knot and cyst nematodes. In situ localization of tobacco EGase transcripts demonstrated that their expression was specifically and developmentally upregulated within giant cells, syncytia, root tips, and lateral root primordia. These data confirm that cell wall modifications within plant-parasitic-nematode feeding cells arise from cell wall–modifying enzymes of plant, rather than nematode, origin

Cellulose Binding Protein from the Parasitic Nematode Heterodera schachtii Interacts with Arabidopsis Pectin Methylesterase: Cooperative Cell Wall Modification during Parasitism[W]

Plant–parasitic cyst nematodes secrete a complex of cell wall–digesting enzymes, which aid in root penetration and migration. The soybean cyst nematode Heterodera glycines also produces a cellulose binding protein (Hg CBP) secretory protein. To determine the function of CBP, an orthologous cDNA clone (Hs CBP) was isolated from the sugar beet cyst nematode Heterodera schachtii, which is able to infect Arabidopsis thaliana. CBP is expressed only in the early phases of feeding cell formation and not during the migratory phase. Transgenic Arabidopsis expressing Hs CBP developed longer roots and exhibited enhanced susceptibility to H. schachtii. A yeast two-hybrid screen identified Arabidopsis pectin methylesterase protein 3 (PME3) as strongly and specifically interacting with Hs CBP. Transgenic plants overexpressing PME3 also produced longer roots and exhibited increased susceptibility to H. schachtii, while a pme3 knockout mutant showed opposite phenotypes. Moreover, CBP overexpression increases PME3 activity in planta. Localization studies support the mode of action of PME3 as a cell wall–modifying enzyme. Expression of CBP in the pme3 knockout mutant revealed that PME3 is required but not the sole mechanism for CBP overexpression phenotype. These data indicate that CBP directly interacts with PME3 thereby activating and potentially targeting this enzyme to aid cyst nematode parasitism

Cellulose Binding Protein from the Parasitic Nematode Heterodera schachtii Interacts with Arabidopsis Pectin Methylesterase: Cooperative Cell Wall Modification during Parasitism

Plant–parasitic cyst nematodes secrete a complex of cell wall–digesting enzymes, which aid in root penetration and migration. The soybean cyst nematode Heterodera glycines also produces a cellulose binding protein (Hg CBP) secretory protein. To determine the function of CBP, an orthologous cDNA clone (Hs CBP) was isolated from the sugar beet cyst nematode Heterodera schachtii, which is able to infect Arabidopsis thaliana. CBP is expressed only in the early phases of feeding cell formation and not during the migratory phase. Transgenic Arabidopsis expressing Hs CBP developed longer roots and exhibited enhanced susceptibility to H. schachtii. A yeast two-hybrid screen identified Arabidopsis pectin methylesterase protein 3 (PME3) as strongly and specifically interacting with Hs CBP. Transgenic plants overexpressing PME3 also produced longer roots and exhibited increased susceptibility to H. schachtii, while a pme3 knockout mutant showed opposite phenotypes. Moreover, CBP overexpression increases PME3 activity in planta. Localization studies support the mode of action of PME3 as a cell wall–modifying enzyme. Expression of CBP in the pme3 knockout mutant revealed that PME3 is required but not the sole mechanism for CBP overexpression phenotype. These data indicate that CBP directly interacts with PME3 thereby activating and potentially targeting this enzyme to aid cyst nematode parasitism.This article is published as Hewezi, Tarek, Peter Howe, Tom R. Maier, Richard S. Hussey, Melissa Goellner Mitchum, Eric L. Davis, and Thomas J. Baum. "Cellulose binding protein from the parasitic nematode Heterodera schachtii interacts with Arabidopsis pectin methylesterase: cooperative cell wall modification during parasitism." The Plant Cell 20, no. 11 (2008): 3080-3093, doi: 10.1105/tpc.108.063065. Copyright American Society of Plant Biologists</p

Signal Peptide-Selection of cDNA Cloned Directly from the Esophageal Gland Cells of the Soybean Cyst Nematode Heterodera glycines

Secretions from the esophageal gland cells of plantparasitic nematodes play critical roles in the nematodeparasitic cycle. A novel method to isolate cDNA encoding putative nematode secretory proteins was developed that utilizes mRNA for reverse transcription-polymerase chain reaction derived from microaspiration of the esophageal gland cell contents of parasitic stages of the soybean cyst nematode Heterodera glycines. The resulting H. glycines gland cell cDNA was cloned into the pRK18 vector, and plasmid DNA was transformed into a mutated yeast host for specific selection of cDNA inserts that encode proteins with functional signal peptides. Of the 223 cDNA clones recovered from selection in yeast, 97% of the clones encoded a predicted signal peptide. Fourteen unique cDNA clones hybridized to genomic DNA of H. glycines on Southern blots and, among them, nine cDNA clones encoded putative extracellular proteins, as predicted by PSORT II computer analysis. Four cDNA clones hybridized to transcripts within the dorsal esophageal gland cell of parasitic stages of H. glycines, and in situ hybridization within H. glycines was not detected for eight cDNA clones. The protocol provides a direct means to isolate potential plant-parasitic nematode esophageal gland secretory protein genes.This article is published as Wang, Xiaohong, Rex Allen, Xiongfei Ding, Melissa Goellner, Tom Maier, Jan M. de Boer, Thomas J. Baum, Richard S. Hussey, and Eric L. Davis. "Signal peptide-selection of cDNA cloned directly from the esophageal gland cells of the soybean cyst nematode Heterodera glycines." Molecular Plant-Microbe Interactions 14, no. 4 (2001): 536-544, doi: 10.1094/MPMI.2001.14.4.536. Posted with permission.</p

Expression pattern of TDIF-TDR-WOX4 pathway upon nematode infection.

<p>Three Arabidopsis promoter:GUS lines including (A) TDRp:GUS. (B) WOX4p:GUS. (C) CLE41p:GUS were infected with <i>H</i>. <i>schachtii</i>. GUS staining was performed at different time points (days) after inoculation (dpi). For cross-sectioning, roots at 3 dpi were used. N = nematode; S = syncytium.</p