2010 Annual Report 1a.Objectives (from AD-416) The objective of this cooperative agreement is to survey and identify pathogenic and non-pathogenic Rhizoctonia spp. associated with turfgrasses with molecular pathological, cytological, ecological, and epidemiological criteria with a view to manage Rhizoctonia diseases. Another component of this objective is to molecularly characterize pathogenicity/virulence-associated factors at genome, gene or protein level. 1b.Approach (from AD-416) To identify Rhizoctonia infecting turfgrasses, a survey will be conducted to collect Rhizoctonia isolates from major golf clubs in Virginia, Maryland, and possibly a few North Eastern States. Rhizoctonia isolates will be classified to major groups (anastomosis groups (AGs), species, etc.), based on morphological, biochemical, and molecular biological characters. For molecular characterization, we may check the usefulness of rDNA PCR and sequencing, AFLP, RAPD, UP-PCR etc. We may also use non-turf Rhizoctonia isolates for comparative and disease control purposes. Molecular techniques like CHEF-gel electrophoresis, Northern, Southern and Western hybridizations, gene cloning, gene transformation may also be applied to study Rhizoctonia pathogenicity. For ecological and epidemiological studies, we may study climatic adaptation and prevalence of Rhizoctonia; susceptibility of various grass species and sensitivity of Rhizoctonia isolates to commonly used fungicides in turfgrasses. 3.Progress Report Under this new agreement, we are continuing a project on identification of brown patch-causing Rhizoctonia species in turfgrasses from Virginia and Maryland. Species of Rhizoctonia in general are not well identified due to morphological similarities and lack of mating relationships. The traditional method of grouping Rhizoctonia species based on hyphal anastomosis reactions is time-consuming and labor intensive. In addition, certain isolates do not self anastomose, or anastomose with more than one group. Knowledge of genetic diversity and evolutionary groups within different Rhizoctonia species could help in developing resistant turfgrass cultivars. Accurate identification of Rhizoctonia species and their anastomosis groups (AGs) is important since they have different levels of sensitivity to fungicides applied. More than 600 Rhizoctonia isolates were collected from diseased turfgrass leaves from five geographic areas in Virginia and Maryland. A random sample of 54 isolates was selected for further investigation. Grouping at the AG-level of the isolates was determined by hyphal fusion reactions with Rhizoctonia tester strains, where feasible. The sequences of ribosomal DNA internal transcriber spacer (ITS) region were determined, and DNA fingerprinting with the Universally Primed Polymerase chain reaction (UP-PCR) and cross-hybridization of UP-PCR products was carried out. UP-PCR is a DNA finger printing technique that can group isolates of Rhizoctonia into their infra-species level. Cross hybridization techniques can detect many unknown isolates simultaneously by using a known strain as the hybridizing probe. This research will result in knowledge about the genetic variations of Rhizoctonia solani pathogenic and non-pathogenic isolates from turfgrasses and the development of an identification method for the pathogen, allowing growers to utilize control measures more effectively for maintenance of healthy turf. This collaboration was monitored through regular phone and email conversations. A Ph.D. student conducting research in the ARS Beltsville lab interacted with the mentor at the University of Tennessee Knoxville on a weekly basis through emails or phone calls.