AGP - 
 

Race Analysis

Knowledge relating to where stem rust is occurring is important, but information on which pathotypes (races) are present and at what frequency is vital for breeding and cultivar recommendation. The only way to determine virulence combinations and which specific pathotype (race) or pathotypes are present is to undertake bioassays under controlled conditions, using sets of differential testers (Race analysis). Race analyses of cereal rust pathogens have been undertaken in many countries for many years using more-or-less standardized methodologies of inoculation, incubation and then “reading” the sets (data recording).

With the identification of Ug99 and derivatives it is vital that information is obtained (1) on the current distribution of the various races, and (2) what variation exists in the stem rust pathogen. Advanced research institutes are providing technical support on race analysis; notably, the Agriculture and Agri-Food Lab, Winnipeg, Canada, Cereals Disease Lab, Minnesota, USA, University of the Free State, South Africa and PBI, University Sydney, Australia. In addition, in-country capacity to undertake race analysis is being strengthened in several at risk partner countries.

A schematic overview of the general protocol used for race analysis is given in the accompanying figure. A brief outline of the process is:

  • Following detection of stem rust infections in the field, samples of infected stems are collected. Core stem tissue is removed; samples kept in paper or glassine envelopes and dried.
  • For analysis in North America, samples are sent under import permit following strict phytosanitory protocols.
  • Rust spores are transferred from infected tissue to healthy leaves of a seedling of an extremely susceptible variety. This sub-culturing increases the amount of rust in a sample.
  • Infection is achieved using dew treatment for 12-16 hours under dark conditions at 21 C, followed by exposure to light for 3-4 hours. After 13-14 days of incubation at 18-25 C, infections will appear.
  • Spores from infected leaves can be collected and used to inoculate a differential set as a bulk or single pustule isolates. Use of single pustule isolates increases the chance of working with a single race.
  • For a single pustule isolate, spores are sub-cultured and multiplied again on the susceptible host. Following multiplication, a differential set is inoculated. The differential set consists of genotypes containing single major stem rust resistance genes. A standard international set of differentials is now being used to permit global comparison.
  • Seedling infection types on the differential set are evaluated using a standard 0-4 evaluation scale. Low infection types (0-2) are considered resistant and high infection types (3-4) susceptible. Races are identified based on the combination of virulence/avirulence patterns exhibited.

Generalized Protocol for Race Analysis - Seedling Assays

protocol
Schematic outline of protocol for seedling based race analysis (Click for larger view)