Reactions of Soybean Plant Introductions
(PI273483 to PI427107)
Following Inoculation with Phytophthora sojae

Research Bulletin 1193-01

Materials and Methods

The soybean germplasm examined in this study included accession numbers PI273483 to PI427107, in maturity groups 000 to IV, from the USDA Soybean Germplasm Collection in Urbana, Illinois.

Laviolette and Athow evaluated many of the accessions in this collection previously for resistance to frogeye leafspot race 2 and P.sojae races 1, 2, 3, and 4 (2). Some lines were also evaluated for reaction to P.sojae races 5, 6, 7, 8, and 9 (2). The 1,015 accessions selected for the current study were resistant or not tested in Laviolette and Athow's previous evaluation.

Detection of Sources of Rps Genes

The isolates that were used in inoculations and their virulence pathotype are listed in Table 1. All of the isolates originated in Ohio. Races 7, 17, and 25 collectively have susceptible interactions with all of the designated Rps genes and many Rps gene combinations.

Table 1. Differential Reaction and Race Designations of Isolates of Phytophthora sojae Used to Detect New Sources or Resistance in Soybean Plant Introductions (PI273483-PI427107). P. sojaea Race Virulence Pathotypeb 7 1a, 2, 3a, 3c, 4, 5, 6, 7 17 1b, 1d, 2, 3a, 3b, 3c, 4, 5, 6, 7 25 1a, 1b, 1c, 1k, 7 31 1b, 1c, 1k, 2, 3c, 4, 5, 6, 7 30-T 1a, 1b, 1k, 2, 3a, 3c, 4, 5, 6, 7 30 1a, 1b, 1k, 2, 3a, 3b, 3c, 4, 5, 6, 7 33 1a, 1b, 1c, 1d, 1k, 3b, 5, 7 38 1a, 1b, 1c, 1d, 1k, 3a, 3b, 3c, 5, 6, 7 aIsolates are listed as the race designation; T is used to separate two isolates which would be designated race 30 based on the reaction of the isolate on differentials Rps1a, Rps1b, Rps1c, Rps1d, Rps1k, Rps3a, Rps6, and Rps7. bThe pathotype indicates which Rps genes in the host the P.sojae isolates have a susceptible interaction. P.sojae races were determined on the following differentials: Williams (universal suscept); Harlon (Rps1a); Harosoy 13XX (Rps1b); Williams 79 (Rps1c); PI103091 (Rps1d); Williams 82 (Rps1k); L76-1988 (Rps2); L83-570 (Rps3); PRX 146-36 (Rps3b); PRX 145-48 (Rps3c); L85-2352 (Rps4); L85-3059 (Rps5); Harosoy 62XX (Rps6); and Harosoy (Rps7).

Additional isolates were then selected that elicit a susceptible interaction with specific 2-Rps gene combinations (Table 2). For example, P.sojae race 31 has a susceptible interaction with Rps gene combination of Rps1k and Rps6, but races 7, 17, or 25 have a resistant reaction.

Table 2. Selected 2 Rps-Gene Combinations That Are Susceptible to Specific P.Sojae Races. P. sojaerace Selected susceptible Rps gene combinations 7 1a,2; 1a,3a; 1a,3c; 1a,4; 1a,5; 1a,6; 1a,7; 2,3a; 2,3c; 2,4; 2,5; 2,6; 2,7; 3a,4; 3a,5; 3a,6; 3a,7; 3c,4; 3c,5; 3c,6; 3c,7; 4,5; 4,6; 4,7; 5,6; 5,7; 6,7 17 1b,2; 1b,3a; 1b,3c; 1b, 4; 1b,5; 1b,6; 1b,7; 2,3b; 3b,4; 3b,5, 3b,6, 3b,7 25 1c,7; 1k,7 31 1c,2; 1c,3c; 1c,4; 1c,5; 1c,6; 1k,2; 1k,3c; 1k,4; 1k, 5; 1k,6 30 1k,3a 30T 1a,3b; 1k,3b 33 1c,3b 38 1c,3a

The race designations for P.sojae are based on the susceptible reaction of eight of the 13 differentials Rps1a, Rps1b, Rps1c, Rps1d, Rps1k, Rps3a, Rps6, and Rps7 (1, 5). The authors used the combination of race-isolate code for 30-T to indicate that there is a difference in the pathotype among two isolates that are designated P.sojae race 30 (Table 1).

The hypocotyl inoculation technique (6) was used to evaluate the accessions for Rps genes. Inoculum was prepared by growing P.sojae for approximately seven days on lima bean-soft agar (12g agar per L). Colonized areas of the plate were removed and placed in a hypodermic syringe; this was mixed to make a slurry by forcing the mycelium and agar through the syringe two times.

Eight to 10 seeds of each accession were planted in vermiculite in the greenhouse. After seven to 10 days, an incision was made in the hypocotyl, and approximately 0.2 to 0.4 mls of a mycelium agar slurry was placed in the wound. The plants were covered overnight with plastic to prevent the agar from drying out.

Seedlings were evaluated for resistant (hypersensitive) and susceptible reactions four to six days after inoculations. Inoculations of accessions that were resistant to P.sojae races 7, 17, and 25 were repeated five times. Accessions that were inoculated with P.sojae races 31, 30-T, 30, 33, and 38 were evaluated a minimum of three times with each race.

Detection of Partial Resistance

Once a P.sojae isolate was identified that had a susceptible interaction with an accession, that isolate was then chosen to evaluate partial resistance. P.sojae races 7, 17, 25, 30, and 31 were used to evaluate for partial resistance with the inoculum-layer test (6) in 14 separate screenings.

Each inoculum-layer test was arranged in a randomized complete block design with three replications which included five soybean cultivars with known resistance response. For example, the controls for P.sojae race 25 were: Chapman (Rps-3a, resistant), Conrad (high partial resistance), Resnik (moderate partial resistance), Williams (moderately susceptible), and Sloan (susceptible). Standard soybean cultivars, Sloan and Conrad, were included with each test in order that comparisons could be made with all of the accessions evaluated with the different P.sojae races.

The inoculum consisted of two-week-old P.sojae cultures grown on dilute lima bean agar (extract from 50g ground lima beans in 1 L) in glass petri plates. The P.sojae colonized agar was removed intact from the petri plate and placed 5 cm below the seed (12 soybean seeds per pot) in course vermiculite in 1.2 L polystyrene containers with bottom drainage. The pots were watered to run-through twice daily. Seedling roots were inoculated with P.sojae as they grew through the agar layer.

Three weeks after planting, the root mass was removed from the pot, the vermiculite shaken off, and the plants scored on the following scale: 1 = no root rot; 2 = up to 10% root mass rotted; 3 = up to 25% of root mass rotted; 4 = 50% of root mass rotted; 5 = all roots rotted, up to 20% seedlings killed; 6 = up to 50% seedlings killed; 7 = up to 75% of plants killed; 8 = up to 90% seedlings killed; 9 = all plants killed.

In order to compare the results from multiple tests as well as isolates with different levels of aggressiveness, the partial resistance rating of three replications were adjusted for the results of the standard check cultivars, Sloan and Conrad, as follows: the means of the partial resistance rating for the standard cultivars were all adjusted by adding or substracting 0.5 to 1.5 to equal the following scores: Conrad (3.5) susceptible cultivar Sloan (6.0).

The scores of the controls were analyzed with the PROC GLM procedure at SAS (SAS Institute, Cary, N.C.) for the effects of test and test by cultivar interaction. The corrected means for all of the accession data was then pooled and analyzed with PROC GLM using SAS (SAS Institute, Cary, N.C.).

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