James P. Stack

Using Pseudomonas spp. for Integrated Biological Control

ABSTRACT Pseudomonas spp. have been studied for decades as model organisms for biological control of plant disease. Currently, there are three commercial formulations of pseudomonads registered with the U.S. Environmental Protection Agency for plant disease suppression, Bio-Save 10 LP, Bio-Save 11 LP, and BlightBan A506. Bio-Save 10 LP and Bio-Save 11 LP, products of Jet Harvest Solutions, Longwood, FL, contain Pseudomonas syringae strains ESC-10 and ESC-11, respectively. These products are applied in packinghouses to prevent postharvest fungal diseases during storage of citrus, pome, stone fruits, and potatoes. BlightBan A506, produced by NuFarm Americas, Burr Ridge, IL, contains P. fluorescens strain A506. BlightBan A506 is applied primarily to pear and apple trees during bloom to suppress the bacterial disease fire blight. Combining BlightBan A506 with the antibiotic streptomycin improves control of fire blight, even in areas with streptomycin-resistant populations of the pathogen. BlightBan A506 also may reduce fruit russet and mild frost injury. These biocontrol products consisting of Pseudomonas spp. provide moderate to excellent efficacy against multiple production constraints, are relatively easy to apply, and they can be integrated with conventional products for disease control. These characteristics will contribute to the adoption of these products by growers and packinghouses.

Triticum mosaic virus: A New Virus Isolated from Wheat in Kansas

In 2006, a mechanically-transmissible and previously uncharacterized virus was isolated in Kansas from wheat plants with mosaic symptoms. The physiochemical properties of the virus were examined by purification on cesium chloride density gradients, electron microscopy, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), sequencing of the nucleotides and amino acids of the coat protein, and immunological reactivity. Purified preparations contained flexuous, rod-shaped particles that resembled potyviruses. The coat protein was estimated from SDS-PAGE to have a mass of approximately 35 kDa. Its amino acid sequence, as deduced from DNA sequencing of cloned, reverse-transcribed viral RNA and separately determined by time-of-flight mass spectrometry, was most closely related (49% similarity) to Sugarcane streak mosaic virus, a member of the Tritimovirus genus of the family Potyviridae. The virus gave strong positive reactions during enzyme-linked immunosorbent assays using polyclonal antibodies raised against purified preparations of the cognate virus but gave consistent negative reactions against antibodies to Wheat streak mosaic virus (WSMV), other wheat potyviruses, and the High Plains virus. When the virus was inoculated on the WSMV-resistant wheat cv. RonL, systemic symptoms appeared and plant growth was diminished significantly in contrast with WSMV-inoculated RonL. Taken together, the data support consideration of this virus as a new potyvirus, and the name Triticum mosaic virus (TriMV) is proposed.

The National Plant Diagnostic Network

Following September 11, Americas attention and resources were refocused on homeland security. While emphasizing the security of facilities, such as airports, tourist attraction sites, and major public buildings, etc. Congress also recognized the vulnerability of its agricultural systems. On June 12, 2002, the President signed into law the Agricultural Bio-terrorism Protection Act of 2002. The Act covers both animal and plant production and directed the Secretary of Agriculture to develop a network linking plant and animal disease diagnostic facilities across the country. The National Plant diagnostic Network (NPDN) focuses on the plant diseases and pest aspects of the program. Its mission is to enhance national agricultural security by quickly detecting introduced pests and pathogens. This will be achieved by creating a functional nationwide network of public agricultural institutions with a cohesive, distributed system to quickly detect deliberately introduced, high consequence, biological pests and pathogens into our agricultural and natural ecosystems by providing means for quick identifications and establishing protocols for immediate reporting to appropriate responders and decision makers. The network is comprised of Land Grant University plant disease and pest diagnostic facilities across the United States. Lead universities have been selected and designated as regional Centers to represent five regions across the country. These Centers are located at Cornell University, Michigan State University, Kansas State University, University of Florida at Gainesville, and University of California at Davis. The National Agricultural Pest Information System (NAPIS), located at Purdue University, has been designated as the central repository for archiving select data collected from the regions. NAPIS maintains information from the Cooperative Agricultural Pest Survey (CAPS), a network of state agricultural organizations and universities that survey for invasive species. As a part of the NPDN, NAPIS will expand its collection of data on plant diseases and other pests. The system will provide a national perspective on agricultural pests through dynamic maps and reports of plant pest distribution. Currently the pest information system houses 1.3 million records on more than 3,800 organisms, and that number will grow significantly as the plant diagnostic network centers start feeding information into the national database in the Spring, 2004. The establishment of the network will provide the means necessary for ensuring all participating Land Grant University diagnostic facilities are alerted of possible outbreaks and/or introductions and are technologically equipped to rapidly detect and identify pests and pathogens. This will be accomplished by establishing an effective communication network between regional expertise, developing harmonized reporting protocols with the national diagnostic network participants, and cataloging pest and disease occurrence to be included in the national database. The NPDN national database at Purdue University will provide summary reports, distribution maps, pattern analysis, and data sets for use in other studies.

Preliminary Assessment of Resistance Among U.S. Wheat Cultivars to the Triticum Pathotype of Magnaporthe oryzae

Magnaporthe oryzae is the causal agent of blast disease on several graminaceous plants. The M. oryzae population causing wheat blast has not been officially reported outside South America. Wheat production in the United States is at risk to this pathogen if it is introduced and established. Proactive testing of U.S. wheat cultivars for their reaction to blast and identification of resistance resources is crucial due to the national and global importance of the U.S. wheat industry. In this preliminary study, the phenotypic reaction of 85 U.S. wheat cultivars to M. oryzae (Triticum pathotype) was determined. Although there was a significant correlation in the reaction to blast at the seedling and adult plant stages, only 57% of the head reaction was explained by the seedling reaction. Because of the importance of disease development at the head stage in the field, assessment of all 85 cultivars occurred at the head stage. Among cultivars tested, a continuum in severity to head blast was observed; cultivars Everest and Karl 92 were highly susceptible with more than 90% disease severity, while cultivars Postrock, JackPot, Overley, Jagalene, Jagger, and Santa Fe showed less than 3% infection. No evidence of the presence of physiological races among isolates T-7, T-12, T-22, and T-25 was found.

Number of Experiments Needed to Determine Wheat Disease Phenotypes for Four Wheat Diseases

Disease phenotypes for winter wheat cultivars were determined in numerous inoculated greenhouse and field experiments over many years. For four diseases, Fusarium head blight, tan spot, Septoria leaf blotch, and Stagonospora leaf blotch, at least 20 cultivars each had been evaluated in a minimum of five experiments. Reference cultivars of known disease reaction were included in each experiment, which allowed transformation of the percent disease severity data to a 1-to-9 scale for comparisons between experiments. Variations in scale values obtained for individual cultivars among the different experiments were used to calculate standard deviations for disease phenotype data. Standard deviations were used to calculate the number of experiment repetitions needed within each disease to achieve different levels of accuracy (margins of error). A margin of error of ±1.5 for the 1-to-9 scale was chosen as the best level of accuracy. Rounding values within this range would put the estimated disease phenotype within ±1 unit of the actual phenotype. To achieve a margin of error of ±1.5 for Fusarium head blight, tan spot, Septoria leaf blotch, and Stagonospora leaf blotch would require a mean that was calculated from a minimum of five, five, seven, and eight experiments, respectively. Personnel who report disease phenotype data to wheat producers or breeders should be aware of the number of experiments upon which they are basing their reports and adjust any disclaimers accordingly. Similarly, wheat breeders should be aware of the inherent variability in phenotyping these four wheat diseases and make appropriate adjustments to their selection protocols. With a minimum of five experimental repetitions, disease phenotype values obtained from inoculated greenhouse and field experiments had very high correlations (r = 0.81 to 0.92, P < 0.0001) with published Kansas State University Research and Extension ratings obtained from commercial fields.

Expression of Susceptibility to Fusarium Head Blight and Grain Mold in A1 and A2 Cytoplasms of Sorghum bicolor

Panicle diseases are among the major constraints to sorghum (Sorghum bicolor) production in the northern Great Plains; host plant resistance is the primary management option. However, essentially all commercial sorghum hybrids contain A1 cytoplasm, which raises the concern about increased disease risk as a result of cytoplasmic genetic uniformity. To determine the influence of cytoplasmic background on the expression of susceptibility to panicle diseases, F1 hybrids with four nuclear genotypes in each of two cytoplasms (A1 and A2) were planted in three environmentally diverse geographic locations in Nebraska. Fusarium head blight ranged in incidence from 13 to 100% across locations. Grain mold, caused primarily by species of Alternaria, Fusarium, and Cladosporium, ranged in incidence from 5 to 100% across locations. There was a significant effect of nuclear genotype on the incidence and severity of both head blight and grain mold across the three locations. Cytoplasm had no effect on head blight incidence or severity, or on grain mold severity. Cytoplasm had a significant effect on grain mold incidence, with A1 exhibiting slightly lower incidence than A2 (64 versus 70%). Although the cytoplasm effect for grain mold incidence was statistically significant, most of the variation in grain mold incidence was attributable to nuclear genotype. The slight increase in grain mold incidence attributable to A2 cytoplasm should be overcome easily by selection of nuclear genotypes with grain mold resistance. The use of A2 cytoplasm to incorporate genetic diversity into grain sorghum hybrids should not increase the risk of head blight or grain mold in commercial grain production.

The National Plant Diagnostic Network: partnering to protect plant systems

The National Plant Diagnostic Network (NPDN) has developed into a critical component of the plant biosecurity infrastructure of the United States. The vision set forth in 2002 for a distributed but coordinated system of plant diagnostic laboratories at land grant universities and state departments of agriculture has been realized. NPDN, in concept and in practice, has become a model for cooperation among the public and private entities necessary to protect our natural and agricultural plant resources. Aggregated into five regional networks, NPDN laboratories upload diagnostic data records into a National Data Repository at Purdue University. By facilitating early detection and providing triage and surge support during plant disease outbreaks and arthropod pest infestations, NPDN has become an important partner among federal, state, and local plant protection agencies and with the industries that support plant protection.

Genomics-Based Marker Discovery and Diagnostic Assay Development for Wheat Blast

Wheat blast has emerged as a major threat to wheat production in South America. Although originally restricted to Brazil, the disease has since been observed in the neighboring countries of Argentina, Bolivia, and Paraguay and recently the pathogen, Magnaporthe oryzae Triticum pathotype, was isolated from infected wheat in Bangladesh. There is growing concern that the pathogen may continue to spread to other parts of the world, including the United States, where several M. oryzae pathotypes are endemic. M. oryzae pathotypes are morphologically indistinguishable and, therefore, must be characterized genotypically. Symptoms of wheat blast include bleaching of the head, which closely resembles the symptoms of Fusarium head blight, further complicating efforts to monitor for the presence of the pathogen in the field. We used a genomics-based approach to identify molecular markers unique to the Triticum pathotype of M. oryzae. One of these markers, MoT3, was selected for the development of a polymerase chain reaction (PCR)-based diagnostic assay that was evaluated for specificity using DNA from 284 M. oryzae isolates collected from a diverse array of host species. Conventional PCR primers were designed to amplify a 361-bp product, and the protocol consistently amplified from as little as 0.1 ng of purified DNA. The specificity of the MoT3-based assay was also evaluated using Fusarium spp. DNA, from which no amplicons were detected.

Bioterrorism: A Threat to Plant Biosecurity?

The food systems that provide the caloric requirements for most of the world’s population are plant-based, including, rice, wheat and maize. The health and productivity of plant systems is a prerequisite of food security and human health. There are many general threats to plant systems that put plant biosecurity at risk, including global trade of plants and plant products, climate change, population growth and landscape exploitation. Bioterrorism is one more threat to consider when developing a strategy for plant biosecurity. Plant systems are vulnerable to biocrime and bioterrorism. Various lists of threat agents and different approaches to risk assessment have been developed to guide research and policy decisions. Yet an integrated strategy for global plant biosecurity is lacking.

Climate Suitability for Magnaporthe oryzae Triticum Pathotype in the United States

Wheat blast, caused by the Triticum pathotype of Magnaporthe oryzae, is an emerging disease considered to be a limiting factor to wheat production in various countries. Given the importance of wheat blast as a high-consequence plant disease, weather-based infection models were used to estimate the probabilities of M. oryzae Triticum establishment and wheat blast outbreaks in the United States. The models identified significant disease risk in some areas. With the threshold levels used, the models predicted that the climate was adequate for maintaining M. oryzae Triticum populations in 40% of winter wheat production areas of the United States. Disease outbreak threshold levels were only reached in 25% of the country. In Louisiana, Mississippi, and Florida, the probability of years suitable for outbreaks was greater than 70%. The models generated in this study should provide the foundation for more advanced models in the future, and the results reported could be used to prioritize research efforts regarding the biology of M. oryzae Triticum and the epidemiology of the wheat blast disease.