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Karnal bunt (KB), caused by the pathogen Tilletia indica Mitra, infects bread wheat, (Triticum aestivum), durum wheat (T. durum), rye (Secale cereale), and triticale (Triticosecale sp.) (Sansford, 1998). Karnal bunt was first reported in the United States in 1996 in Arizona, California, New Mexico and Texas (Ykema et al., 1996). T. indica is a relatively weak plant pathogen that is highly sensitive to environmental conditions (Singh 1994) and few plant pathologists believe that the disease warrants the current level of regulation by the USDA.

Since 1996 Karnal bunt has occurred sporadically in Texas. In 2001 a grain elevator operator in Young County, Texas detected bunted kernels, and subsequent testing by APHIS detected the pathogen in the adjacent counties of Archer, Throckmorton and Baylor. If a field tests positive, it becomes quarantined and there is no mechanism to remove a field from quarantine. Furthermore, all land within a three-mile radius of the positive field becomes regulated. In 2002, because of the proximity of a Karnal bunt positive field to the county line, the regulated area expanded into a fifth county, Knox county. In 2003, 158,529 acres (2,972 fields) in these five counties of the Texas Rolling Plains are now regulated, which means that every wheat field must be checked for Karnal bunt.

Little can be done with wheat contaminated with T. indica bunted kernels except steaming and flaking for cattle consumption, a low value product. Also, wheat produced in regulated areas is difficult to sell, resulting in a plummet in land value. The sustainability of farming operations in Karnal Bunt regulated areas is questionable. Additionally, farmers in the portion of N. Texas where Karnal Bunt occurs have few alternative crop options.

At present, there are no regionally adapted cultivars with specific resistance to Karnal bunt or fungicides labeled for the disease. It is unknown how teliospores, deposited in the soil during previous harvests, will impact disease development in future crops or how soil type or microclimatic conditions will impact spore viability. It is imperative that research be conducted on the ecology and epidemiology of this disease in order to provide regulatory agencies with the type of scientific data needed to deregulate the pathogen, and producers with better management options.

The Texas Agriculture Experiment Station in Bushland, TX has received a permit from the USDA to work on Karnal Bunt. In accordance with the permit, we have modified a lab with containment equipment to eliminate the risk of pathogen escape. Thus includes the installation of a HEPA filtration system to create negative air pressure in the laboratory, a waste water decontamination system, and other preventative measures. We currently have the following research objectives:

Objective 1. Develop improved methodology to detect T. indica in infected wheat spikes and grain samples. PCR techniques have been developed to differentiate between T. indica and other smut fungi. However, all published techniques to date were developed for diagnostic purposes and not for disease management. It is unknown whether any of these are capable of detecting the pathogen in planta. We will test and compare published PCR techniques for efficacy in detecting T. indica in infected wheat spikes and immature kernels before teliospore formation. If existing primers do not work, a PCR protocol that detects the pathogen in infected plant tissue will be developed.

Tests for detection of T. indica teliospores and bunted kernels in grain samples are required of producers seeking a phytosanitary certificate for shipment and sell of their wheat as seed. The potential to detect T. indica in infested grain samples will be tested with a novel solid phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS). Healthy and infested grains release volatile organic compounds (VOCs) to ambient air that are specific to biologic activity. The SPME samplers are very sensitive in extracting VOCs from air and the GC-MS is capable of separating, identifying, and quantifying specific VOCs that are released and extracted with SPME. Wheat infested with T. indica or T. caries, the casual agent of common bunt of wheat, will be used in studies to determine whether SPME-GC-MS can distinguish between the two pathogens. Studies will also be initiated to compare the limits of detection using SPME-GC-MS and traditional sieving and spore washing techniques. Initially, wheat samples artificially contaminated with either T. indica or T. caries will be analyzed and the specific VOCs from each sample will be recorded. This data will be used to develop a specific "volatile signature" for each pathogen. Secondly, specific numbers of spores will be added to grain samples that will then be tested for KB contamination using traditional methods or GC detection.

Objective 2. Develop methodology to detect and quantify genetic resistance to T. indica using real-time fluorescent PCR. Fluorescent PCR is a technique that allows quantification of target DNA. Wheat heads, from lines varying in their susceptibility to T. indica, will be inoculated and subsequently harvested every 3-4 days, over a three-week period. Fungal DNA will be extracted and quantified from each line at each harvest date to determine whether fluorescent PCR can be used to quantify resistance in each line. This method of quantifying pathogen development in planta will benefit breeders in their selection and screening processes, and also could be useful in quantifying efficacy of new fungicides.

Objective 3. Investigate the relationship among soil type, time of spore deposition, and teliospore viability. Soils were collected in April 2002 from wheat fields in San Saba County that tested positive for KB in 1997 only, 2001 only, 1997 and 2001, or never tested positive. Samples were collected from at least two fields in each category. Soil type will be determined, with special emphasis on texture, and percent spore survival will be determined by sieving teliospores from soil and plating on acidified water agar. Data will be analyzed to determine the correlation between spore viability and time of spore deposition, or spore viability and soil texture.

Images of Karnal Bunt:


Bunted Wheat Head	Bunted Kernels	Tilletia indica teliospores
All images reproduced with permission from the UDSA. Click on a thumbnail for a larger image (warning large).

Maps of the regulated counties in N. Texas (~150kb). Reproduced with permission by the USDA.

For more information about Karnal Bunt of Wheat visit:

References:

Sansford, C. E. 1998. Karnal bunt (Tilletia indica): An assessment of the significance of the initial detection of Tilletia indica Mitra in the USA in early 1996 and the potential risk to the United Kingdom (and the European Union). at Bunts and Smuts of Wheat: An International Symposium, at North Carolina, USA.

Singh, A., 1994. Epidemiology and management of Karnal bunt disease of wheat. University of Agriculture and Technology Research Bulletin No. 127, Pantnagar, U.P., India.

Ykema, R. E., Palm, M. E. and Peterson, G. L., 1996. First report of Karnal bunt of wheat in the United States. Plant Dis 80, 1207-1207.