Progress 09/15/07 to 08/31/12
Outputs
OUTPUTS: The first strategy was to initiate a disease epidemic in spring by planting mother plants infected with the systemic phase of Puccinia punctiformis (Pp). Observations indicated that these plants were stressed with disease, and had short longevity. Resulting disease was erratic and weak. In 2008, fall inoculations of rosettes of Canada thistle using ground-up leaves with teliospores of Pp resulted in these shoots being infected the following spring (2009) and numerous systemically diseased shoots in the spring of 2010. By spring 2011 there were no thistle plants surviving. Based on these results, a test plot was established at FDWSRU in Maryland in Fall 2009 with individual rosettes inoculated in September with about 1g per rosette of ground leaves bearing teliospores. The next spring (2010) systemically diseased shoots with Pp emerged, and by Spring 2011 an epidemic of disease was underway. In fall of 2010 in Maryland, rosettes were repeatedly inoculated with suspensions of 1 million teliospores, and probability of infection by Pp was correlated to number of times inoculated. Other experiments pointed to differential Pp susceptibility among thistle clones. To further enhance epidemics of Pp we needed additional knowledge of Pp epidemiology. We performed experiments to evaluate dispersal of various Pp spore types. Dispersal gradients were measured by releasing spores in windy field conditions and capturing spores at varying distances from the source. By all measures, aerial dispersal of the two major types of Pp spores is significantly different. Teliospores are much more restricted in dispersal distances than are urediniospores. The more limited distances traveled by teliospores likely has an important impact on the incidence of lethal systemic infections throughout the landscape. However the ability of both spore types to be dispersed in windborne leaf debris(see outcomes)reduces the impact of these differences in spore dispersal. From these observations and tests, the disease cycle of the rust fungus proceeds differently than previously reported. In the spring, systemically diseased shoots arise from shoots on roots that are previously infected with Pp. The first signs of the fungus on these shoots are orange haploid pycnia (spermagonia) that cross fertilize to form haploid dikaryotic aeciospores. These spores infect leaves of nearby plants and give rise to uredinia that produce haploid dikaryotic urediniospores that, in turn, infect other leaves. In the late summer the uredinia transform into telia which undergo nuclear fusion (karyogamy). Through mitotic division the telia give rise to two-celled diploid teliospores. In late summer and fall, the plants that emerged in the spring senesce and diseased leaves bearing telia dehisce and deposit teliospores onto newly emerging rosettes. Under conditions of adequate dew the teliospores undergo meiosis and germinate into haploid basidiospores that infect the rosettes. The fungus then develops hyphae that grow into the roots of the rosettes where it survives the winter. Systemically diseased shoots emerge from this rootstock the following spring. PARTICIPANTS: A key element of this project was our collaboration (as a sub-contract) with USDA's Agricultural Research Service, particularly, Dr. Dana Berner, and his assistant M. Smallwood at the Fort Detrick, Foreign Disease and Weed Science Research Unit. Their ability to test the principles developed in their own studies and with results from Penn State University, plus show that they would work in Greece, helped confirm the results generated in the U.S. in our laboratory and field locations. More importantly these trials indicated that results were practical and predictable. Numerous internships were provided to students from three Universities: Student interns: K. Thomas, Frederick Community College; T. Wiggins, James Madison University; R. Pongrance, Penn State University; R. Dabundo, Penn State University; M. Moschera, Penn State University; K. Nguyen, Penn State University; M. Sabol, Penn State University; and A. Harford, Penn State University. Also Graduate Students were wholly or partially trained as a result of this project, to include: S. Conaway, Ph.D. candidate, Penn State University and A. Testen, M.S. candidate, Penn State University. At least one thesis is presently being developed. Several publications are expected in the next two years. TARGET AUDIENCES: Target audiences are land owners who have canada thistle infected lands, and/or owners expected to manage conservation reserve lands with thistle infestations. Further, state highway departments, USDA NRCS personnel, State Depts of Conservation and Natural Resources who mandate land owner management practices, and state, local and federal park managers. All of these have been contacted and have responded with interest, and all are awaiting information on how to capitalize on these new management practices. Since much of these results were accumulated in the past year, we will continue to pass information to them over the next several years. PROJECT MODIFICATIONS: As previously reported, The PIs orignially thought that the use of systemically infected Canada thistle mother plants would provide the inoculum to develop a sustainable epidemic. These plants were too fragile, and provided only weak disease development. At he same time, it became clear that plant debris from locally sourced infected plants would provide inoculum to establish sustainable rust epidemics on Canada thistle. The project moved to that inoculum source to facilitate the epidemics. This inoculum provides the farmer with a virtually no-cost source of inoculum if he is diligent in finding the scattered infections in nearbye locations.
Impacts
The above outputs were reduced to practice in a project developed in summer of 2010. A collaborative project on establishing in-field epiphytotics of P. punctiformis on Canada thistle was established in Greece. A set of protocols on the collection of diseased leaves bearing teliospores and on repeated inoculations with these leaves was sent to the Greek collaborators who inoculated rosettes in early October of 2010. In the spring of 2011, rosettes receiving multiple inoculations had produced systemically diseased shoots. It was also noticed that systemically diseased plants had emerged on the windward side of a small hill near the inoculation area. Evidently, teliospore-bearing leaf material from the inoculation area had blown onto this hill. This observation is consistent with additional observations in MD, in which natural epiphytotics of systemic rust disease may follow dehiscing and wind-blown leaves bearing high concentrations of teliospores. These data also point to the potential for farmers to locate systemically infected Canada thistle plants, displaying black telia, air dry these plants, maintain them dry till fall, grind the plants, and then inoculate rosettes with pinches of the plant debris in multiple patches to achieve multiple infections the next spring. After these initial efforts, disease should persist to continue the decline of the thistle patch. Student interns/trainees: K. Thomas, Frederick Community College; T. Wiggins, James Madison University; R. Pongrance, Penn State University; R. Dabundo, Penn State University; M. Moschera, Penn State University; K. Nguyen, Penn State University; M. Sabol, Penn State University; A. Harford, Penn State University. Graduate Students: S. Conaway, Ph.D. candidate, Penn State University; A. Testen, M.S. candidate, Penn State University.
Publications
- Berner, D. K., E. L. Smallwood, C. A. Cavin, A. L. Lagopodi, and J. N. Kashefi. 2012. Routine establishment of epidemics of systemic disease of Canada thistle (Cirsium arvense) caused by the rust fungus Puccinia punctiformis. APS Potomac Division Meetings March 14-16, 2012. Phytopathology 102:S3.1.
- Berner, D. K. and P. A. Backman. 2007. Biological control of Canada thistle in pastures and parks: a call for collaborators on simple field tests. Proceedings Northeastern Weed Science Society Annual Meeting. Baltimore, MD. Jan. 2-7, 2007. 61:1 http://www.newss.org/proceedings/proceedings_2007.pdf.
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Progress 09/15/10 to 09/14/11
Outputs
OUTPUTS: In 2008, fall inoculations of rosettes of Canada thistle with ground leaves bearing teliospores of Puccinia punctiformis resulted in systemically diseased shoots the following spring and systemically diseased shoots in the spring of 2010. By the spring of 2011 there were no Canada thistle plants in the inoculated area. In greenhouse tests, inoculation of rosettes with 10 million teliospores per plant produced significantly more systemically diseased shoots than lower inoculum levels. Additionally, a Canada thistle test plot was established in Maryland in the summer of 2009 where individual rosettes were inoculated one time in September of that year with about 1 gram per rosette of ground leaves bearing teliospores of Puccinia punctiformis. The following spring (2010), systemically diseased shoots with P. punctiformis emerged, and by the spring of 2011, 140 systemically diseased shoots emerged in this plot. A subsequent greenhouse test on repeated inoculations of rosettes with 1 million teliospores per rosette each time, resulted in a linear and significant increase in proportion of systemically diseased plants with increasing numbers of inoculations. In the fall of 2010, rosettes of Canada thistle in a farmer's field in Maryland were repeatedly inoculated with suspensions of 1 million teliospores. Repeated inoculations were done on each rosette. In the spring of 2011, systemically diseased shoots emerged in this previously disease-free field. Additional inoculation methods are also being developed and evaluated. Healthy Canada thistle transplants that were subjected to root-dip inoculations into P. punctiformis spore suspensions of either one million or 100,000 per ml in the Fall, yielded >90% systemically diseased shoots regardless of inoculum level. This technique would allow for onsite production of inoculum with minimal inputs and resources. Recently, it was deployed into established patches of Canada thistle to investigate if secondary spread from infected transplants is a possibility with results pending Spring 2012. It is hypothesized that Canada thistle plants can have genetic resistance to some P. punctiformis lines. To investigate this possibility, we have compared resistance of thistle genotypes across Pennsylvania, which showed differential responses. Systematic sampling of thistle patches was performed to assess host diversity within and between patches with ssr genotyping techniques. Unique genets are being used as the basis of a study into the spatial distribution of host resistance. Three weeks of continuous spore trapping were used to sample spore spore dispersal dynamics in a 6m diameter natural disease focus. P. punctiformis spore counts were aligned with local weather data. Preliminary analysis suggested that peak spore release events are correlated with leaf dry-off conditions and ground turbulence occuring near mid-day when temperatures have risen and relative humidity has decreased. These data will contribute to future modeling of P. punctiformis dispersal and epidemiology. PARTICIPANTS: Graduate student, S. Conaway spent a week visiting the USDA-ARS Cereal Disease Laboratory in March 2011 presenting Canada thistle rust research to staff and collecting applicable techniques from cereal rust studies. Experimental methods for elucidating differential host resistance in Canada thistle was discussed with CDL researchers to the benefit of the project. Mr. Conaway also collaborated with biological control research scientists and extension personnel at the USDA-ARS Pacific Basin Agricultural Research Center on a variety of classical biocontrol projects. Further thistle genotyping work was done at PBARC along with developing automated machine counting techniques for spore dispersal studies. While in Hawaii, research was presented at the American Phytopathological Society 2011 annual meeting and at the 13th International Symposium on Biological Control of Weeds. Undergraduate student R. Dabundo was trained in field data collection, and genotyping of thistle field accessions. He worked on a project to develop an automated disease severity assessment program based on scans of infected Canada thistle leaves. TARGET AUDIENCES: Twenty high school students visiting the Penn State University campus for the Agroecology field day, toured plots, heard a short presentation, and examined exhibits to learn about biological control of weeds based on the Canada thistle x Puccinia punctiformis system. Three undergraduate students at Penn State University, received hands on training working on this project both on-campus and at the experimental farm in Rock Springs. An undergraduate student from Hood College (MD) worked on the project as an internship at the Fort Detrick Weed Science and Research Unit of USDA/ARS in Frederick MD. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
In the summer of 2010, a collaborative project on establishing in-field epidemics of P. punctiformis on Canada thistle was established in Greece. A set of protocols on collection of diseased leaves bearing teliospores and on repeated inoculations with dried and ground leaves was sent to the Greek collaborators who inoculated rosettes in early October of 2010. In the spring of 2011, rosettes receiving multiple inoculations had produced systemically diseased shoots. While searching for the diseased shoots it was noticed that systemically diseased plants had emerged on the windward side of a small hill near the inoculation area. Evidently, teliospore-bearing leaf material from the inoculation area had blown onto this hill which acted like a wind-break and blocked further movement of the leaf material which was deposited on rosettes on the hill. This observation is consistent with what has been observed in the U.S. and, in conjunction with results of inoculations with teliospore-bearing leaves, indicates that the form of inoculum responsible for natural epidemics of systemic rust disease is probably dehiscing and wind-blown leaves bearing high concentrations of teliospores. Presentations of research results were made to two USDA/ARS laboratories, and to Southern University in Louisiana.
Publications
- S. Conaway, K. Shea, D. K. Berner, and P. A. Backman. 2011. Using pathogen dispersal characteristics to improve biological control of Canada thistle with the rust fungus Puccinia punctiformis. Proceedings of APS-IPPC Joint Meeting, Aug. 6-10, 2011. Phytopathology 101(6S):S1-S206.
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Progress 09/15/09 to 09/14/10
Outputs
OUTPUTS: The rust pathogen Puccinia punctiformis is a promising biological control agent that reduces Canada thistle infestations through fatal, systemic infections. Inoculum for P. punctiformis can only be grown on living Canada thistle plants and is considerably more labor intensive than growing it in culture. Estimating how P. punctiformis moves throughout the landscape will help in utilizing the pathogen's natural spread mechanisms for more efficient deployment of limited initial inoculum. The dispersal gradients for both urediniospores and teliospores were measured by releasing spores in windy field conditions and capturing spores at varying distances (up to 200 meters) from the source with rotorod air samplers. Field releases conducted in 2009 and 2010, indicated that teliospores had a significantly steeper dispersal gradient than did urediniospores. Terminal velocities of spores were also compared in a particle settling tower in an effort to explain the dispersal differences between the 2 spore types. Mean terminal velocities were approximately 3 times faster for teliospores compared to urediniospores. In the 2010 field season volumetric air samplers were deployed in naturally diseased Canada thistle patches. Spore counts obtained after 3 weeks of collections were compared to local temperature, relative humidity, light intensity, wind, and precipitation data. Priliminary analysis suggests that peak release of both urediniospores and teliospores occurs in mid-day during which conditions of highest temperatures and lowest humidity occur. These conditions result in leaf dry-off and high turbulence. We compared resistance of thistle genotypes across Pennsylvania from distinct patches within seven different locations. A greenhouse experiment conducted in 2010 tested resistance of 37 clones of thistle to two lines of rust. Systemic and local infection susceptibility was tested with separate foliar inoculations and root dip inoculations. The preliminary analysis of systemic disease results showed differential responses between thistle clones and rust lines. To assess optimum timing and host tissue infection court, we evaluated the effects of season and placement of inoculum on disease incidence. To clarify this issue, a field experiment in 2009 and 2010 looked at inoculations performed in spring and fall and on emerging shoot tissue versus root tissue. The inoculations were performed with liquid spore suspensions on Canada thistle clones transplanted into a thistle-free field. Systemic disease was only observed on the fall treatments. Root-shoot inoculations produced higher incidence than emerged shoot inoculations. The experiment will be repeated in 2011 with physical barriers to exclude run-off from reaching root tissue in order to determine whether the shoot treatments actually infected aboveground host tissue. PARTICIPANTS: This project is the primary source of funding for a Ph.D. student's (Steven Conaway)research project on the development of rust epidemics that would facilitate the biological control of Canada thistle. This project facilitated the professional development of Misha Moschera Masters student in Agricultural Extension and Education during the Summer 2010, during which she trained in field sampling techniques; greenhouse propagation and maintenance; rust inoculation and harvesting; DNA extraction and PCR, and she helped the project with the development of educational products to help with the extension component of this project. This project facilitated the training of Charlene Gray, Undergraduate in Ecology. Summer 2010 trained in field sampling technique; greenhouse propagation; rust inoculation and analysis of spores; DNA extraction and PCR. This project facilitated the development of Alicia Spangler, who worked during 2009-10 on factors responsible for rust epidemics in thistle. Her work on the project was partially responsible for her selection as the Outstanding undergraduate student for 2010 of Penn State University College of Agricultural Sciences. She metriculated to an M.S. program in Weed Science at Michigan State University. TARGET AUDIENCES: Key groups of target audiences are land owners that may or may not be farming. Research sites have been placed in conservation reserve land, raparian strips, grazing land, and hiway medians. These research trials frequently become points of interest to these owners and we educate as we learn more about the pathosystem. PROJECT MODIFICATIONS: Results in the first two years indicated that infected plants as inoculum sources have issues that can result in erratic outcomes. One is that the appropriate environment for dispersal and infection may not occur at the time for optimal infection. The second is that not all clones of thistle have the same level of susceptibility to the inoculum provided as infected plants. This has resulted in additional studies involving release of inoculum at later periods in the year (coincidental with the most probable infection period), and has also caused us to investigate mixed inoculum sources to make epidemics more predictably severe.
Impacts
Training of an undergraduate student in the area of weed ecology and biological control of weeds. Training and technical support of a M.S. candidate in weed ecology and biological control, and her assistance in developing educational products supporting the projects goals and outreach. Award of 2009 undergraduate student who worked on development of rust epidemics in Canada thistle, and who received the undergraduate student of the year award from Penn State University's College of Agricultural Sciences.
Publications
- No publications reported this period
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Progress 09/15/08 to 09/14/09
Outputs
OUTPUTS: Successful, large-scale infections of potted Canada thistle were achieved with Puccinia punctiformis in the greenhouse. Seven Pennsylvania Canada thistle locations were established in varying agroecologies. The first year results showed no disease spread during the summer from transplants that had produced only teliospores. However, latent systemic infections or overwintering teliospores may produce systemic disease spread in 2010. These results indicate that initial transplanted seedlings should bear urediniospores to produce local lesions on surrounding foliage that subsequently produces teliospores in the late summer and fall. In Maryland, five Canada thistle sites were identified in the fall of 2007 that had no signs of P. punctiformis, and were selected to receive transplants systemically diseased with this fungus in early spring of 2008. Establishment of systemic disease was successful at one site and local lesions bearing telia developed on surrounding plants at this site. These plants, in turn, produced teliospores that generated an epidemic of systemic disease visible in the spring of 2009. Monitoring of a naturally diseased site in Maryland over two years showed that systemically diseased shoots lead to local lesion production on surrounding foliage at a rate of about 3 locally diseased shoots (LDS) for each systemically diseased shoot (SDS); LDS=3.51 + 3.37 X SDS; Rsq=0.95. Healthy thistle shoots in heavily diseased patches at this site decreased over 50% from spring 2008 to spring 2009, while systemically diseased shoots doubled. Evaluations of the aerobiology of spore flight indicate that both teliospores and urediniospores are wind-borne, though to differing degrees: teliospores have a steeper dispersal gradient than urediniospores in windy field conditions and a mean terminal velocity almost double that of urediniospores, which are over twice as heavy as teliospores. Greenhouse and field inoculations in MD and PA show that systemic disease develops from teliospore inoculations of young shoots. This has led us to revise the natural disease development cycle of the rust which is now thought to be initiated by basidiospore (from teliospores) infection of secondary shoots in the fall. The fungus then grows into the roots and overwinters in the root system, also reported from older literature sources, which produce systemically diseased shoots the following spring. Failures in past greenhouse inoculations of some thistle isolates have led to an hypothesis of genetic resistance to systemic rust development. In MD, susceptible thistle isolates showing systemic disease were cured of disease with systemic fungicides, and these susceptible clones remain significantly more susceptible than plants from clones that were not cured. Roots from thistle populations in PA were collected and propagated for further tests of resistance. Three field experiments were established to pin-point the susceptible tissue and seasonal window for systemic infection in the field. Spring results of field experiments with varied inoculation sites and timing could further this hypothesis and lead to modified methods to establish epidemics. PARTICIPANTS: Partnering with USDA Natural Resources Conservation Service to manage state lands in Conservation Reserve and raparian zones. Also with USDA Farm Service Agency on Rules and Regulations required for Federal oversight of managed lands. Collaborators also include MD and PA Departments of Transportation for management of highway right-of-way zones. In all cases,the appropriate authorities are kept aware of progress and difficulties. TARGET AUDIENCES: State and Federal regulatory and transportation agencies. Also land owners who need to manage canada thistle in their production fields or pastures. PROJECT MODIFICATIONS: Unexpected and previously unreported resistance to this pathogen indicate that successful inonoculum in the future may require a blend of several genotypes of Puccinia punctiformis to be successful. This occurs as differential responses are observed in the field and are being verified in the greenhouse.
Impacts
Three summer undergraduate interns were trained in field techniques and weed biocontrol biology. One intern has since entered a masters program in the Penn State department of Agricultural Extension Education. A former undergraduate intern has developed a poster detailing her work on transplanting diseased thistle and optimizing transplant conditions. Her poster will be presented at the annual meeting of the American Society of Agronomy this November. She is also developing a presentation for the Northeastern Weed Science Society annual meeting in January. A second intern joined the USDA as a summer technician at the ARS foreign disease and weed science laboratory. The project graduate research assistant introduced a number of different audiences to concepts and methods of Canada thistle biocontrol by speaking with CREP participants volunteering land for biocontrol field work; giving a departmental seminar on research progress; and teaching four sessions on biocontrol of invasive weeds to a group of high school agriculture and science students.
Publications
- No publications reported this period
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