Progress 10/01/00 to 09/30/05
Outputs
Secretory proteins encoded by parasitism genes expressed in the esophageal gland cells of plant-parasitic nematodes play key roles in nematode infection and parasitism of host plants. Characterization of parasitism genes we have cloned from the soybean cyst nematode, Heterodera glycines, is enabling us to address the biology of how the soybean cyst nematode is able to attack the soybean root system to induce yield losses. The Hg-SYV46 parasitism gene encodes a secretory protein with function similar to CLAVATA3/ESR of Arabidopsis thaliana. New high yielding soybean breeding lines were identified with high levels of resistance to soybean cyst nematode, races 3 and 9, and the southern and peanut root-knot nematodes. During this year 22,164 soybean breeding lines were screened in the greenhouse for nematode resistance. Of these, 12,607 were screened for root-knot nematode resistance (7,364 to the southern root-knot nematode, 4,622 to the peanut root-knot nematode, and 6212
to the javanese root-knot nematode). The remaining 9,512 lines were screened for resistance to several races of the soybean cyst nematode (6,018 to race 3, 4,641 to race 9). In addition 3,206 lines were screened for major QTL for resistance to the southern root-knot nematode using molecular SSR (DNA) markers. The breeding lines identified in these screening tests were advanced to next level of testing in which the high yielding lines, also having other desirable agronomic characters, were selected for under field conditions.
Impacts
The nematode parasitism genes being discovered and characterized will provide potential targets for intervention in the parasitic process and lay the foundation for the development of novel strategies for controlling these economically important crop pathogens in the future. The availability of superior yielding soybean cultivars with multiple nematode resistance that are adapted to Georgia and the southeastern U.S. environments provides our farmers with a nematode management tactic that is efficient, economical, and environmentally sound. These benefits will result in an increase in grower
Publications
- Wang, Xiaohong. M. G. Mitchum, Bingli Gao, Chunying Li, Hanane Diab, Thomas J. Baum, Richard S. Hussey, and Eric L. Davis. 2005. A parasitism gene from a plant-parasitic nematode with function similar to CLAVATA3/ESR (CLE) of Arabidopsis thaliana. Mol. Plant Pathology 6:187-191.
- Wang, J., R. Hussey, E. Davis, T. Baum, and M. G. Mitchum. 2005. Identification and functional analysis of soybean cyst nematode parasitism proteins with a role in virulence. Phytopathology 95:S108.
- Tian, D., T. Baum, E. Davis, R. hussey, and X. Wang. 2005. Identification of putative parasitism genes of the potato cyst nematode, Globodera rostochiensis. Phytopathology 95:S103.
- Patel, N., H. Diab, B. Gao, X. Wang, R. S. Hussey, T. J. Baum, and E. L. Davis. 2004. Identification of putative parasitism genes of the beet cyst nematode (BCN), Heterodera schachtii. Phytopathology 94:S82.
- Hussey, R. S. and H. R. Boerma. 2005. Greenhouse ratings for resistance to three species of root-knot nematode and soybean cyst nematode, 2004. Pp. 59-61. In 2004 Field Crops Performance Tests. Research Report 676.
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Progress 01/01/04 to 12/31/04
Outputs
Secretory proteins encoded by parasitism genes expressed in the esophageal gland cells of plant-parasitic nematodes play key roles in nematode infection and parasitism of host plants. A cellulose-binding protein-encoding cDNA, designated Hg-cbp-1, was cloned from a Heterodera glycines oesophageal gland-cell long-distance PCR cDNA library. The cDNA hybridized to genomic DNA of H. glycines in Southern blots, and the genomic sequence of Hg-cbp-1 contained only one intron. The Hg-cbp-1 cDNA contained an open reading frame encoding 132 amino acids, with a predicted signal peptide sequence for secretion and a cellulose-binding domain. Bacterial expressed recombinant HG-CBP-1, minus the signal peptide sequence, had no hydrolytic activity on carboxymethyl-cellulose but was able to bind to cellulose. The developmental expression of Hg-cbp-1, determined by real-time reverse transcriptase PCR, showed that Hg-cbp-1 is expressed throughout the parasitic cycle of H. glycines, with
a relatively higher expression level in developing parasitic stages. Although a role for HG-CBP-1 in syncytial expansion has not been established, the fact that CBDs can modulate in vitro elongation of plant cells, suggests this might be a possible function of the secretory HG-CBP-1 in the parasitism of soybean by H. glycines. The expression of cell wall-modifying enzymes of plant origin is specifically upregulated in nematode feeding cells as most nematode celluloytic enzymes decrease. Future studies should address whether the secreted CBP from H. glycines acts as a cell wall loosening protein like expansins to aid in cell wall growth and remodeling by plant enzymes in nematode feeding cells. New high yielding soybean breeding lines were identified with high levels of resistance to soybean cyst nematode, races 3 and 9, and the southern and peanut root-knot nematodes. During this year 18,481 soybean breeding lines were screened in the greenhouse for nematode resistance. Of these,
10,255 were screened for root-knot nematode resistance (2,865 to the southern root-knot nematode, 6,758 to the peanut root-knot nematode, and 632 to the javanese root-knot nematode). The remaining 8,226 lines were screened for resistance to several races of the soybean cyst nematode (5,760 to race 3, 2,409 to race 9, and 57 to a mixture of races 1-5). The soybean breeding lines identified in these screening tests were advanced to next level of testing in which the high yielding lines, also having other desirable agronomic characters, were selected for under field conditions.
Impacts
These newly discovered nematode parasitism genes will provide potential targets for intervention in the parasitic process and lay the foundation for the development of novel strategies for controlling these economically important crop pathogens in the future. The direct cloning of soybean cyst nematode parasitism gene candidates is enabling us to address the biology of how the soybean cyst nematode is able to attack the soybean root system to induce yield losses. The availability of superior yielding soybean cultivars with multiple nematode resistance that are adapted to Georgia and the southeastern U.S. environments provides our farmers with a nematode management tactic that is efficient, economical, and environmentally sound. These benefits will result in an increase in grower profits from soybean production in Georgia and throughout the southeastern U.S.
Publications
- Ha, B-K, B. Bennett, R. S. Hussey, and H. R. Boerma. 2004. Pedigree analysis of a major QTL for soybean resistance to southern root-knot nematode. Crop Sci. 44:758-763.
- Gao, B., R. Allen, E. L. Davis, T. J. Baum, and R. S. Hussey. 2004. Molecular characterisation and developmental expression of a cellulose-binding protein gene in the soybean cyst nematode Heterodera glycines. Int. J. Parasitol. 34:1377-1383.
- Hussey, R. S. 2004. Feeding strategies. Pp. 784-787. In R. M. Goodman (ed.) Encyclopedia of Plant and Crop Science. Marcel Dekker, New York.
- Baum, T. J., B. Gao, T. Maier, R. S. Hussey, and E. L. Davis. 2004. Secrets in secretions: analysis of the parasitome of Heterodera glycines. Pp. 289-292. In Biology of Plant-Microbe Interactions, Vol 4. I. Tikhonovich, B. Lugtenberg and N. Provorov (eds).
- Smant, T., E. L. Davis, R. S. Hussey, T. J. Baum, M-N. Rosso, J. Helder, and J. Bakker. 2004. On the evolution of parasitism genes. Proceedings of the Fourth International Congress of Nematology. Nematology Monographs & Perspectives 2:573-579.
- Gao, B., T. Davis, T. J. Baum, and R. S. Hussey. 2004. A novel secretory ubiquitin extension protein from the soybean cyst nematode. Phytopathology 94:S33.
- Diab, H. G., B. Gao, T. J. Baum, R. S. Hussey, X. Wang, and E. L. Davis. 2004. Functional analyses of a novel esophageal parasitism gene from Heterodera glycines. Phytopathology 94:S25
- Patel, N., H. Diab, B. Gao, X. Wang, R. S. Hussey, T. J. Baum, and E. L. Davis. 2004. Identification of putative parasitism genes of the beet cyst nematode (BCN), Heterodera schachtii. Phytopathology 94:S82.
- Sunko, S., J. McCuiston, X. Wang, R. S. Hussey, T. J. Baum, and E. L. Davis. 2004. Towards functional analysis of putative parasitism genes in Heterodera glycines using RNA interference. Phytopathology 94:S100.
- Hussey, R. S. and H. R. Boerma. 2004. Greenhouse ratings for resistance to three species of root-knot nematode and soybean cyst nematode, 2003. Pp. 59-61. In 2003 Field Crops Performance Tests. Research Report 676.
- Gao, B., R. Allen, E. L. Davis, T. J. Baum, and R. S. Hussey. 2004. Developmental expression and biochemical properties of a beta-1,4-endoglucanase family in the soybean cyst nematode Heterodera glycines. Mol. Plant Pathol. 5:93-104.
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Progress 01/01/03 to 12/31/03
Outputs
Parasitism genes expressed in the esophageal gland cells of plant-parasitic nematodes encode the secretions that control the complex process of plant parasitism. We developed a novel and direct method to identify these parasitism genes and have discovered some of the secrets of the secretions that the nematode uses to attack crops. The emerging model of plant parasitism by nematodes is much more complex than originally hypothesized. One of the secrets of the disease-inducing secretions that we have discovered suggests that certain plant-parasitic nematodes may have co-opted plant signaling molecules for parasitic modification of host plant cells. This extraordinary form of molecular mimicry, where a nematode molecule mimics a host molecule for a biological reason, may have evolved in plant nematodes so that the nematode could modulate host cellular processes to its benefit. The soybean cyst nematode (SCN) produces six beta-1,4-endoglucanases (cellulases) that are
secreted during infection of soybean. Two of the endoglucanases, HG-ENG-5 and HG-ENG-6, differed significantly in their amino acid sequence of the catalytic domains and their gene structure from that of the other four beta-1,4-endoglucanases. Quantitative real-time RT-PCR revealed distinct developmental expression differences among the hg-eng family members during the early stages of parasitism and relatively low expression levels in late parasitic stages, with the exception of the adult male stage for some eng genes. Recombinant HG-ENGs degraded carboxymethylcellulose and optimum enzyme activity ranged from pH 5.5 for HG-ENG-5 to pH 8 for HG-ENG-6. EDTA, Ca2+, Co2+, Mg2+, and Fe2+ did not affect enzyme activity of any ENG protein, while Zn2+, Cu2+ and Mn2+ inhibited enzyme activity from 23% to 73% in some cases. In tests with 12 different polysaccharide substrates, enzyme activity was restricted to beta-1,4 linkages with all ENG proteins tested. Only HG-ENG-5 and HG-ENG-6 had
relatively high activity on xylan and slightly degraded microcrystalline cellulose. Together, these data reveal distinct differences in expression and biochemistry of cyst nematode parasitism genes and proteins, respectively, and cast light on the intricate interactions between a parasitic animal and its plant host. New high yielding soybean breeding lines were identified with high levels of resistance to soybean cyst nematode, races 3 and 9, and the southern and peanut root-knot nematodes. During this year 18,481 soybean breeding lines were screened in the greenhouse for nematode resistance. Of these, 10,255 were screened for root-knot nematode resistance (2,865 to the southern root-knot nematode, 6,758 to the peanut root-knot nematode, and 632 to the javanese root-knot nematode). The remaining 8,226 lines were screened for resistance to several races of the soybean cyst nematode (5,760 to race 3, 2,409 to race 9, and 57 to a mixture of races 1-5). The soybean breeding lines
identified in these screening tests were advanced to next level of testing in which the high yielding lines, also having other desirable agronomic characters, were selected for under field conditions.
Impacts
These newly discovered nematode parasitism genes will provide potential targets for intervention in the parasitic process and lay the foundation for the development of novel strategies for controlling these economically important crop pathogens in the future. The direct cloning of soybean cyst nematode parasitism gene candidates is enabling us to address the biology of how the soybean cyst nematode is able to attack the soybean root system to induce yield losses. The availability of superior yielding soybean cultivars with multiple nematode resistance that are adapted to Georgia and the southeastern U.S. environments provides our farmers with a nematode management tactic that is efficient, economical, and environmentally sound. These benefits will result in an increase in grower profits from soybean production in Georgia and throughout the southeastern U.S.
Publications
- Hussey, R. S. and E. L. Davis. 2003. Chapter 5. Nematode esophageal glands and plant parasitism. Pp. 257-292. In Nematology, Advances and Perspectives Vol. 1. Z. X. Chen, S. Y. Chen, and D. W. Dickson, eds. Tsinghua University Press, China.
- Gao, B., T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2003. Developmental expression and biochemical characterization of cellulases from Heterodera glycines. Phytopathology 93:S28.
- Gao, B., R. Allen, T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2003. The parasitome of the phytonematode Heterodera glycines. Mol. Plant Microbe Interact. 16:720-726.
- Harris, D. K., H. R. Boerma, R. S. Hussey, and S. L. Finnerty. 2003. Additional sources of soybean germplasm resistant to two species of root knot nematode. Crop Sci. 43:1848-1851.
- Beuchat, L. R., A. J. Scouten, R. I. Allen, and R. S. Hussey. 2003. Potential of a plant parasitic nematode to facilitate internal contamination of tomato plants by Salmonella. J. Food Protect. 66:1459-1461.
- Baum, T. J., R. S. Hussey, and E. L. Davis. 2003. Parasitism gene discovery in sedentary phytonematodes. Nematology Monographs and Perspectives 2:1-8.
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Progress 01/01/02 to 12/31/02
Outputs
Parasitism genes expressed in the esophageal gland cells of plant-parasitic nematodes encode stylet secretions that control the process of plant parasitism. In the soybean cyst nematode, the parasitome, i.e. the secreted products of parasitism genes, facilitate juvenile migration in soybean roots and mediate the modification of root cells into elaborate feeding cells required to support the growth and development of the nematode. The most direct and efficient approach for cloning parasitism genes and rapidly advancing our understanding of the molecular interactions during nematode parasitism of plants is to create gland-cell cDNA libraries using cytoplasm microaspirated from the esophageal gland cells of parasitic stages. By combining expressed sequence tag analysis of a gland-cell cDNA library with high-throughput in situ expression localization of clones encoding secretory proteins we obtained the first comprehensive parasitome profile for any parasitic nematode. We
identified 54 new soybean cyst nematode gland-expressed candidate parasitism genes of which 42 genes constitute completely novel sequences. The few parasitism gene products with similarity to proteins of known functions are either involved in cell-wall digestion or suggest involvement in the regulatory machinery of plant cells. Individual parasitome members showed distinct expression patterns throughout the parasitic cycle and the discovered parasitome complexity paints a complicated picture of host cellular events under specific control by the nematode parasite. The parasitome discovered provides a comprehensive template to target functional analyses for dissecting the molecular interactions of soybean cyst nematode with soybean.
Impacts
Coupling expressed sequence tag analyses with techniques designed to isolate and characterize genes expressed exclusively within the esophageal gland cells of a plant-parasitic nematode has provided for the first time direct and surprising information on the complexity and dynamics of the parasitome of a multicellular parasite. The direct cloning of candidate soybean cyst nematode parasitism genes is enabling us to address the biology of how the soybean cyst nematode is able to attack the soybean root system to induce yield losses. Our progress has provided many candidate parasitism genes from soybean cyst nematode to study as potential new targets to inhibit soybean cyst nematode parasitism of soybean.
Publications
- Gao, B., R. Allen, T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2002. Identification of a new beta-1,4-endoglucanase gene expressed in the subventral gland cells of Heterodera glycines. J. Nematol. 33:12-15.
- Gao, B., R. Allen, T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2002. Characterisation and developmental expression of a chitinase gene in Heterodera glycines. Int. J. Parasitol. 32:1293-1300.
- De Boer, J. M., J. P. McDermott, E. L. Davis, R. S. Hussey, G. Smant, and T. J. Baum. 2002. Cloning of a putative pectate lyase gene expressed in the subventral esophageal glands of Heterodera glycines. J. Nematol. 33:9-11.
- De Boer, J. M., J. P. McDermott, X. Wang, T. Maier, F. Qui, R. S. Hussey, E. L. Davis, and T. J. Baum. 2002. The use of DNA microarrays for the developmental expression analysis of cDNAs from the oesophageal gland cell region of Heterodera glycines. Mol. Plant Pathol. 3:261-270.
- Gao, B., R. Allen, T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2002. Large-scale identification of parasitism genes of the soybean cyst nematode. Pp. 212-216. In S. A. Leong, C. Allen, E. Tripplet, (eds). Biology of Plant-Microbe Interactions, Volume 3. APS Press, St. Paul, MN.
- Gao, B., T. Maier, E. Davis, T. Baum and R. Hussey. 2002. Defining a plant-parasitic nematode: a profile of putative parasitism genes expressed in the esophageal gland cells of Heterodera glycines. Nematology 4:218.
- Gao, B., T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2002. A profile of putative parasitism genes expressed in the esophageal glands of Heterodera glycines. Phytopathology 92:S27
- Du, W., R. Hussey, T. Baum, and E. Davis. 2002. Peptides that bind to and inhibit cellulase of Heterodera glycines identified by combinatorial library screening. Nematology 4:218.
- Wang, X., T. Maier, R. Hussey, T. Baum, and E. Davis. 2002. Cloning parasitism genes expressed in the esophageal gland cells of Heterodera glycines using an amplified RNA technique. Nematology 4:224.
- Du, W., R. Hussey, T. Baum, and E. Davis. 2002. Identifying peptides that bind to and inhibit cellulase of Heterodera glycines identified by combinatorial library screening. Phytopathology 92:S20
- Wang, X., T. Maier, R. Hussey, T. Baum, and E. Davis. 2002. Construction of a secretory gland cell-specific cDNA library of Heterodera glycines through an amplified RNA approach. Phytopathology 92:S20
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Progress 01/01/01 to 12/31/01
Outputs
Our central hypothesis is that stylet secretions originating in the esophageal gland cells of the soybean cyst nematode (SCN) play essential roles in pathogenesis of soybean. A gland-cell long distance PCR cDNA library was constructed from cytoplasm microaspirated from 10 parasitic SCN representing different time points in the parasitic cycle. Of 1,441 cDNA clones sequenced from the LD PCR cDNA library, deduced protein sequences of 150 cDNAs were preceded by a putative signal peptide for secretion and could have roles in SCN parasitism of soybean. These clones were further characterized by in situ hybridizations with digoxigenin-labeled antisense cDNA probes to determine their developmental expression pattern in the life cycle of SCN. Ninety-eight cDNA clones encoding putative signal peptides have been tested by in situ hybridizations with probes of 32 unique clones specifically hybridizing with transcripts within the subventral (8 clones or 25%) or dorsal (24 clones
or 75%) esophageal gland cells of SCN. These clones are considered prime candidates for further functional analyses to determine their role in the soybean-nematode interaction. PSORT II predicted 24 of the deduced proteins to be extracellular and 8 proteins as nuclear localized. The predicted nuclear localized proteins are additional candidates for SCN secretory proteins that might enter the plant nucleus to modulate host gene expression if secreted into a parasitized root cell. In BLASTp analysis 23 (75%) of the predicted proteins had no similarity with known proteins in the databases. The predicted proteins that had similarities with known proteins included a venom allergen like protein, two beta-1,4- endoglucanases, a pectate lyase, a chitinase, RanBPM-like proteins, and a unique cellulose binding protein. Only two of the gland in situ positive clones had homologues in Caenorhabditis elegans. Also the developmental expression for many of the putative parasitism genes differed among
the SCN parasitism stages, suggesting that the regulation of some parasitism genes requires stimulation factors within soybean root tissue.
Impacts
The direct cloning of putative soybean cyst nematode parasitism genes using the new techniques we have developed is enabling us to address the biology of how the soybean cyst nematode is able to attack the soybean root system to induce yield losses. The progress we have made has provided many candidate parasitism genes from soybean cyst nematode to study as potential new targets to inhibit soybean cyst nematode parasitism of soybean.
Publications
- Gao, B., R. Allen, T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2001. Identification of putative parasitism genes expressed in the esophageal gland cells of the soybean cyst nematode, Heterodera glycines. Mol. Plant-Microbe Interact. 14:1247-1254.
- Gao, B., R. Allen, T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2001. Molecular characterization and expression of two venom allergen-like secretory proteins from the soybean cyst nematode, Heterodera glycines. Int. J. Parasitology 31:1617-1625.
- Wang, X., R. Allen, X. Ding, M. Goellner, T. Maier, J. M. DeBoer, T. J. Baum, R. S. Hussey, and E. L. Davis. 2001. Signal peptide-selection of cDNA cloned directly from the esophageal gland cells of Heterodera glycines. Mol. Plant-Microbe Interact. 14:536-544.
- Davis, E. L., Y. Yan, M. N. Rosso, R. S. Hussey, T. J. Baum, and G. Smant. 2001. Endoglucanase genes active in plant-nematode interactions. Poster, Gordon Research Conference on Cellulases and Cellulosomes. Proctor Academy, Andover, NH. July 30, 2001.
- Gao, B., R. Allen, T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2001. Cloning putative parasitism genes expressed in the esophageal gland cells of the soybean cyst nematode. Phytopathology 91: S134.
- Gao, B., R. Allen, T. Maier, E. L. Davis, T. J. Baum, and R. S. Hussey. 2001. Large-scale identification of parasitism genes of the soybean cyst nematode. 10th International Congress of Molecular Plant- Microbe Interactions abstract No. 464.
- De Boer, J., X. Wang, R. Allen, R. S. Hussey, E. L. Davis, and T. J. Baum. 2001. Microarray screening of a cDNA library from the esophageal glands of the soybean cyst nematode. 10th International Congress of Molecular Plant-Microbe Interactions abstract No. 461.
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Progress 01/01/00 to 12/31/00
Outputs
There is no progress to report. This is a new project.
Impacts
(N/A)
Publications
- No publications reported this period
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