Virulence phenotypes of Puccinia graminis on barley, wheat and oat in Canada from 2020 to 2022

Abstract

Stem rust of barley and wheat is caused by the obligate pathogen Puccinia graminis f. sp. tritici (Pgt) and stem rust of oat is caused by P. graminis f. sp. avenae (Pga). Stem rust can be a very destructive disease of cereal crops and, until the development of resistant host varieties, was responsible for millions of dollars of losses to producers. Genetic host resistance is the most common and effective method to control stem rust on cereals. Knowledge of the virulence genes and races present in the pathogen population is important to identify effective host resistance genes for use by cereal plant breeders. The objectives of this study were to determine the incidence and severity of P. graminis in wheat, barley and oat in Manitoba, Canada, and assess the virulence frequencies and races of isolates collected from Manitoba, Saskatchewan and Ontario, Canada during 2020 to 2022. Stem rust incidence and severity was very light in Manitoba in 2020, 2021 and 2022. Virulence was not observed on resistance genes Sr30, Sr24 and Sr31 from Canadian collections of Pgt isolates made during 2020 to 2022. Race QFCSC was the most common race of Pgt in Canada, and races MCCFC and TPMKC were also common in Manitoba in 2022. Virulence to resistance genes Pg6, Pg10 and Pg16 was not observed among Pga isolates. The most common race of Pga from Manitoba and Saskatchewan was TGN, followed by SGB and TJS. The most frequent Pga races were TDJ, TDD and TGN in Ontario.

Résumé

La rouille des tiges de l’orge et du blé est causée par le pathogène obligatoire Puccinia graminis f. sp. tritici (Pgt) et la rouille des tiges de l’avoine est causée par P. graminis f. sp. avenae (Pga). La rouille des tiges peut être une maladie très destructrice des cultures céréalières et, jusqu’au développement de variétés hôtes résistantes, elle était responsable de millions de dollars de pertes pour les producteurs. La résistance génétique de l’hôte est la méthode la plus courante et la plus efficace pour lutter contre la rouille de la tige sur les céréales. La connaissance des gènes de virulence et des races présentes dans la population du pathogène est importante pour identifier des gènes de résistance à l’hôte efficaces à utiliser par les sélectionneurs de plantes céréalières. Les objectifs de cette étude étaient de déterminer l’incidence et la gravité de P. graminis dans le blé, l’orge et l’avoine au Manitoba, Canada, et d’évaluer les fréquences de virulence et les races des isolats collectés au Manitoba, en Saskatchewan et en Ontario, Canada, entre 2020 et 2022. L’incidence et la gravité de la rouille des tiges ont été très faibles au Manitoba en 2020, 2021 et 2022. Aucune virulence n’a été observée sur les gènes de résistance Sr30, Sr24 et Sr31 dans les collections canadiennes d’isolats de Pgt réalisées entre 2020 et 2022. La race QFCSC était la race de Pgt la plus commune au Canada, et les races MCCFC et TPMKC étaient également communes au Manitoba en 2022. La virulence aux gènes de résistance Pg6, Pg10 et Pg16 n’a pas été observée parmi les isolats de Pga. La race de Pga la plus courante au Manitoba et en Saskatchewan était TGN, suivie de SGB et TJS. Les races de Pga les plus fréquentes étaient TDJ, TDD et TGN en Ontario.

Keywords:

• Barley
• oat
• races
• stem rust
• virulence
• wheat

Mots-clés:

• Avoine
• blé
• orge
• races
• rouille de la tige
• virulence

Introduction

Stem rust is caused by a highly specialized, obligate parasitic fungus which is separated into forma speciales based on the cereal host being attacked. Stem rust on barley and wheat is caused by Puccinia graminis Pers.:Pers. f. sp. tritici Eriks & E. Henn. (Pgt), and the causal agent on oat is P. graminis Pers.:Pers. f. sp. avenae Eriks. & E. Henn (Pga). This has historically been one of the most destructive diseases of wheat in Canada with major epidemics occurring in 1916 (estimated 43% yield loss) and 1923 (Johnson 1961), followed by major epidemics in 1953, 1954, and 1955 with estimated yield losses of 45 million, 150 million and 9 million bushels, respectively (Peturson 1958). The most effective tool that has been developed to mitigate the effects of stem rust on its cereal hosts in Canada has been the use of resistant crop host varieties, which has led to no significant yield losses attributable to stem rust since 1955 (Fetch et al. 2021b).

The development of host varieties with effective resistance requires knowledge of the virulence variation in natural pathogen populations. Studies on the variation in virulence in Canadian populations of Pgt and Pga started in 1919 (Johnson and Green 1957) and have been conducted annually ever since. This manuscript reports on the virulence variation in these Canadian populations for 2020, 2021 and 2022.

Materials and methods

A description of the methods used to collect and process samples and determine physiologic races (virulence phenotypes) of stem rust was described by Fetch et al. (2021a); a brief description follows. Stems of barley (Hordeum vulgare L.), wild barley (H. jubatum L.), oat (Avena sativa L.), wild oat (A. fatua L.) and wheat (Triticum aestivum L.) infected with stem rust were collected from different fields from mid-July to late September each year, from Manitoba, Ontario, and Saskatchewan, Canada in 2020, 2021 and 2022. Infected stem samples were collected, depending on availability, from fields at least 20 km apart, placed in paper envelopes labelled with a unique identifier and air dried at room temperature overnight. Dried samples were stored in a refrigerator until processed. For trap plot nurseries, susceptible spring wheat (‘Little Club’, ‘Morocco’), barley (‘Wolfe’) and oat (‘AC Morgan’, ‘Makuru’) lines were planted in the spring in each province and, if infection occurred, infected stems were collected for race analysis.

Urediniospores were transferred from infected stems to seedlings of ‘McNair’ wheat (Pgt) or ‘Pc94’ oat (Pga) using a sterile spatula. Inoculated seedlings were incubated overnight in a dew box in which they were misted using an ultrasonic humidifier. In the morning the seedlings were illuminated using T5 lights for at least 2 hours to finish the infection process. The dew box was opened slightly to avoid overheating and provide a slow drying period. The seedlings were then transferred to a greenhouse at 20°C with a 16 hour photoperiod and a plastic lamp cover was placed on the pots to prevent cross-contamination. A rubber hose with light positive air pressure was inserted through a hole in the lamp cover to aid in preventing cross-contamination. Approximately 14 days after inoculation, two isolates (A and B), each from a single large random pustule, were made from each collection using a mini-cyclone spore collector. Isolates of Pgt and Pga were increased on ‘McNair’ and ‘Pc94’, respectively, to obtain enough urediniospores for inoculation of the differential set. Race or virulence phenotype characterization was conducted for both isolates.

Physiological race or virulence phenotype identification was conducted using 20 single gene differential lines for Pgt isolates (Table 1), and 15 lines for Pga isolates (Table 2). Five seeds per line were planted in separate RL98 containers containing moist Sunshine Mix #5 (SunGro Horticulture Canada, Seba Beach, Alberta), pressed to a uniform 2 cm depth to provide even emergence, gently watered, and kept under a ‘plastic tent’ to prevent contamination prior to inoculation. Seedlings were grown in a ‘clean’ greenhouse at 20°C with a 16 hour photoperiod. Each set of single gene lines were inoculated with a single isolate at approximately 7–8 days after seeding, when the first leaf had fully extended. About 3 mg of urediniospores of each isolate was suspended in 0.7 mL Soltrol oil (Soltrol 170 Isoparaffin; Chevron Phillips, The Woodlands, Texas, USA.) and sprayed onto the differential lines using house air pressured to 20 kPa. The Soltrol oil carrier was volatilized off the inoculated plants by letting them sit for at least 20 minutes before incubating them in the dew box. Inoculated plants were incubated in a dew box as described above, and then incubated in a greenhouse to allow expression of the disease. Infection type (IT) was determined 14 days post-inoculation using a 0–4 scale (Stakman et al. 1962). The physiologic races were given a letter code following the system of Fetch and Jin (2007) and Jin et al. (2008) using the IT on each of the differential lines. A high or a susceptible response were those with ITs of 3± or 4, and IT responses of 0 to 22+ were considered low or resistant responses. The Pga races were given a three letter code, with the first letter referring to the reactions on Pg1, Pg2, Pg3 and Pg4, the second letter referring to Pg6, Pg8, Pg9 and Pg10 and the third letter referring to Pg12, Pg13, Pg15 and Pg16. The Pgt races were given a five letter code, with the first letter referring to the reactions on Sr5, Sr21, Sr9e and Sr7b, the second letter referring to Sr11, Sr6, Sr8a and Sr9g, the third letter referring to Sr36, Sr9b, Sr30 and Sr17, the fourth letter referring to Sr9a, Sr9d, Sr10 and SrTmp and the fifth letter referring to Sr24, Sr31, Sr38 and SrMcN.

Table 1. Frequency (%) of virulence of Puccinia graminis f. sp. tritici isolates to single gene stem rust differential lines. Isolates were collected from trap plots (wheat, barley, oat), cultivated and wild barley, cultivated and wild oat, and wheat in Manitoba, Ontario and Saskatchewan, Canada in 2020 to 2022.

	2020		2021			2022
Resistance Gene	Manitoba	Ontario	Manitoba	Saskatchewan	Ontario	Manitoba	Ontario
Sr5	100	100	100	100	100	100	100
Sr21	100	100	100	0	100	56	100
Sr9e	0	0	0	0	0	84	0
Sr7b	0	0	0	100	100	24	0
Sr11	0	0	0	0	100	55	0
Sr6	0	0	0	0	100	13	0
Sr8a	100	100	100	0	100	56	100
Sr9g	100	100	100	100	100	100	100
Sr36	0	0	0	0	0	39	0
Sr9b	0	0	0	0	100	11	0
Sr30	0	0	0	0	0	0	0
Sr17	100	100	100	100	100	89	100
Sr9a	100	100	100	0	100	28	100
Sr9d	100	100	100	0	100	56	100
Sr10	100	100	100	100	100	88	100
SrTmp	0	0	0	100	0	71	0
Sr24	0	0	0	0	0	0	0
Sr31	0	0	0	0	0	0	0
Sr38	0	0	0	0	0	2.7	0
SrMcN	100	100	100	100	100	100	100
Total isolates assessed	8	4	17	1	1	72	4

Table 2. Frequency (%) of virulence of Puccinia graminis f. sp, avenae isolates to single gene stem rust differential lines and the Pg-a gene complex. Isolates were collected from trap plots, cultivated and wild oat in Manitoba, Saskatchewan and Ontario, Canada during 2020 to 2022.

	2020				2021				2022
Resistance Gene	Manitoba and Saskatchewan^a		Ontario		Manitoba and Saskatchewan^b		Ontario		Manitoba and Saskatchewan^c		Ontario
	No.	%	No.	%	No.	%	No.	%	No.	%	No.	%
Pg1	424	100	13	100	161	100	2	100	280	100	9	100
Pg2	424	100	13	100	161	100	2	100	273	98	9	100
Pg3	424	100	13	100	161	100	2	100	280	100	9	100
Pg4	318	75	13	100	131	81	2	100	246	88	9	100
Pg6	0	0	0	0	0	0	0	0	0	0	0	0
Pg8	424	100	5	33	161	100	1	50	280	100	2	20
Pg9	49	12	9	60	23	14	2	100	78	28	9	100
Pg10	0	0	0	0	0	0	0	0	0	0	0	0
Pg12	225	53	4	27	104	64	0	0	173	62	2	20
Pg13	51	12	8	53	33	21	1	50	88	32	1	10
Pg15	299	71	13	87	126	78	1	50	175	63	9	90
Pg16	0	0	0	0	0	0	0	0	0	0	0	0
Pg-a	206	49	2	13	79	49	0	0	163	58	2	20
alpha	147	35	2	13	64	40	0	0	140	50	2	20
omega	165	39	2	13	64	40	0	0	150	54	2	20
Total isolates assessed	415		13		161		2		280		9

^aTen isolates were assessed from Saskatchewan in 2020.

^bFour isolates were assessed from Saskatchewan in 2021.

^cFive isolates were assessed from Saskatchewan in 2022.

Results and discussion

The incidence and severity of stem rust in Manitoba was very light in 2020 (Fetch, Zegeye, et al. 2021), 2021 and 2022, resulting in no economic losses to wheat, barley or oat. One of the factors that may have resulted in a low incidence and severity of stem rust in these years is very low precipitation. According to the ‘Drought Watch’ website of Agriculture and Agri-Food Canada (https://agriculture.canada.ca/en/agricultural-production/weather; accessed 14 September 2022), most areas of Manitoba were abnormally dry to suffering severe drought in June and July in 2020 and 2022, with conditions improving slightly in August of those years. Most areas of Manitoba were under conditions of moderate to exceptional drought throughout June, July and August 2021. Another factor that may have resulted in low incidence and severity of stem rust in Manitoba was low levels of inoculum being blown in from the USA. Stem rust on wheat and barley was of low severity in the USA in 2020, with some isolated severe hot spots in Louisiana and South Dakota, and stem rust on oat was only reported from Louisiana and Texas (Cereal Disease Laboratory 2020). Stem rust on wheat and barley was only reported on wheat in 2021 in Texas, USA, while oat stem rust was also not common, being reported from Texas, USA, and from a disease nursery in Minnesota, USA (Cereal Disease Laboratory 2021). Stem rust of wheat and barley was only reported on wheat in 2022 in the USA, with the only report being from a nursery in South Dakota, which had a 100% incidence and 60–70% severity on susceptible lines (Cereal Disease Laboratory 2022). Oat stem rust in the USA in 2022 was reported at low incidence and severity in Louisiana and Florida, as well as in a nursery in South Dakota, which had a 20% incidence and 30% severity on susceptible lines. Fungicide applications by producers over this time period may also have contributed to the low incidence and severity of stem rust.

Physiological specialization

Puccinia graminis f. sp. tritici

The virulence of the Pgt isolates collected from Manitoba, Ontario and Saskatchewan collected in 2020 to 2022 to single gene differential lines is presented in Table 1. All isolates collected in Manitoba had virulence to genes Sr5, Sr9g and SrMcN, whereas none of the isolates had virulence to genes Sr30, Sr24 and Sr31. An isolate of Pgt was collected from winter wheat in 2020, which was virulent to the same genes as all other isolates collected in 2020. One isolate was collected from cultivated wheat in a commercial field in Saskatchewan in 2021, which had a different virulence phenotype from the isolates collected in Manitoba in 2021. The Saskatchewan isolate was avirulent to Sr21, Sr8a, Sr9a and Sr9d, while being virulent to Sr7b and SrTmp. An isolate was also collected from a commercial wheat plant in Manitoba in 2022, which did not contain any unique virulence as compared to the other isolates collected.

The virulence of the isolates collected in Manitoba in 2022 was much more diverse than the isolates collected in Manitoba in 2020 and 2021 (Table 1). Virulence to Sr9e, Sr7b, Sr11, Sr6, Sr36, Sr9b, SrTmp and Sr38 was observed in 2022, which was not noted in 2020 and 2021. Virulence was noted to Sr21, Sr8a, Sr17, Sr9a, Sr9d and Sr10 in 2022, but not at 100% frequency as in 2020 and 2021. The situation in 2022 was similar to the virulence situations in the Manitoba Pgt populations in 2019, 2017 and 2013 (Fetch et al. 2021a, 2021b), with no previously unreported virulence occurring in 2022. The virulence situation in 2020 and 2021 was the same as the virulence situation in 2018 (Fetch et al. 2021b).

Nine isolates were assessed for virulence phenotype from Ontario in 2020 to 2022. All isolates were virulent to Sr5, Sr21, Sr8a, Sr9g, Sr17, Sr9a, Sr9d, Sr10 and SrMcN (Table 1). The isolate collected in 2021 also expressed virulence to Sr7b, Sr11, Sr6, Sr9b and Sr38. The virulence patterns found in 2020 and 2022 were the same as those reported in 2018 (Fetch et al. 2021b) and 2013 (Fetch et al. 2021a). Virulence to Sr7b, Sr11, Sr6, Sr9b and Sr38 has been previously reported in Ontario (Fetch et al. 2021b). Virulence to Sr30, Sr24 and Sr31 had not been reported in Ontario during 2013 to 2022 (Table 1; Fetch et al. 2021a, 2021b).

The only race identified from Pgt isolates collected in Manitoba in 2020 and 2021 was race QFCSC (Table 3). The most common race identified in 2022 was MCCFC at 41%, followed by TPMKC (25%), QFCSC (20%), and RKQQC (8%). One isolate from Saskatchewan isolated in 2021 from cultivated wheat in a commercial field was identified as MCCFC. One isolate each was isolated from winter wheat plants (race QFCSC) and commercial spring wheat plants (race TPMKC) in Manitoba in 2020 and 2022, respectively. Race QFCSC has been the dominant race identified from Pgt isolates from Manitoba since 2009, except in 2015 when no race QFCSC isolates were identified (no isolates were assessed in 2014) (Fetch et al. 2015, 2018, 2021a, 2021b). Only race QFCSC was identified in 2018, which is similar to the results of 2020 and 2021 (Table 3). Races MCCFC, TPMKC, RKQQC and RKQQF, which were identified in 2022 (Table 3), were identified in the virulence assessments of Pgt during 2013 to 2019 (Fetch et al. 2021a, 2021b).

Table 3. Frequency (%) of races of Puccinia graminis f. sp. tritici obtained from trap plots (wheat, barley), cultivated and wild barley and wheat in Manitoba, Ontario and Saskatchewan in Canada in 2020 to 2022.

Race^a	Virulence spectrum (ineffective SR genes)	Total isolates assessed	Percent isolates assessed
2020
Manitoba^b
QFCSC	5, 21, 8a, 9 g, 17, 9a, 9d, 10, McN	8	100
Ontario
QFCSC	5, 21, 8a, 9 g, 17, 9a, 9d, 10, McN	4	100
2021
Manitoba and Saskatchewan^c
MCCFC	5, 7b, 9 g, 17, 10, Tmp, McN	1	6
QFCSC	5, 21, 8a, 9 g, 17, 9a, 9d, 10, McN	17	94
Ontario
RTHSC	5, 21, 7b, 11, 6, 8a, 9 g, 9b, 17, 9a, 9d, 10, McN	1	100
2022
Manitoba^b
MCCFC	5, 7b, 9 g, 17, 10 Tmp, McN	32	41
TPMKC	5, 21, 9e, 7b, 11, 8a, 9 g, 36, 17, 9d, 10, Tmp, McN	20	25
QFCSC	5, 21, 8a, 9 g, 17, 9a, 9d, 10, McN	16	20
RKQQC	5, 21, 7b, 6, 8a, 9 g, 36, 9b, 9a, 9d, McN	6	8
RKQQF	5, 21, 7b, 6, 8a, 9 g, 36, 9b, 9a, 9d, 38, McN	2	3
Ontario
QFCSC	5, 21, 8a, 9 g, 17, 9a, 9d, 10, McN	4	100

^aThe Pgt races were given a five letter code, with the first letter referring to the reactions on Sr5, Sr21, Sr9e and Sr7b, the second letter referring to Sr11, Sr6. Sr8a and Sr9g, the third letter referring to Sr36, Sr9b, Sr30 and Sr17, the fourth letter referring to Sr9a, Sr9d, Sr10 and SrTmp and the fifth letter referring to Sr24, Sr31, Sr38 and SrMcN (Fetch and Jin 2007).

^bNo isolates were collected from Saskatchewan from wheat, cultivated or wild barley in 2020 or 2022.

^cOne isolate was collected from wheat in Saskatchewan in 2021; the race was MCCFC.

Nine isolates of Pgt from Ontario were assessed for virulence during 2020 to 2022, with eight of the isolates identified as QFCSC (Table 3). An isolate from Ontario collected in 2021 was identified as RTHSC. Race QFCSC was the dominant race in 2019, and 2017, and the only race identified from Ontario in 2018 and 2013 (Fetch et al. 2021a, 2021b). Race RTHSC was not identified during 2013 to 2019. All isolates identified to race in the USA in 2021 and 2020 were QFCSC (Cereal Disease Laboratory 2021, 2022).

Puccinia graminis f. sp. avenae

All isolates of Pga from Manitoba from 2020 to 2022 were virulent to resistance genes Pg1, Pg3 and Pg8 (Table 2). Similar results were obtained with virulence to Pg2, except in 2022, when the virulence frequency was at 98%. No virulence was observed towards resistance genes Pg6, Pg10 or Pg16 over these 3 years. Virulence was noted at ≥75% for Pg4 during 2020 to 2022. The results obtained for 2020 to 2022 are similar to those obtained for 2013 to 2019 for most of the resistance genes tested (Fetch et al. 2021a, 2021b). The percent of isolates with virulence declined for Pg9, Pg12, Pg13, Pg15 and Pg-a, in 2020 to 2022 compared to 2013 to 2018, although virulence to these genes was still common.

Race TGN was the most common race of Pga identified from Manitoba isolates during 2020 to 2022 (Table 4). The percentage of isolates identified to race TGN during the 3 years was 38.9%. Races SGB and TJS were the next most common races over the 3 years, being identified in each of the 3 years, with more isolates being identified as SGB (12.9%) than TJS (8.8%). There were 19 races identified from the Pga isolates in Manitoba in 2020, six of which were found at a frequency of >5% (Table 4). Thirteen races were identified in 2021 and 2022, with four and seven races having a frequency of >5% of all isolates, respectively. The Pga populations identified from Manitoba during 2020 to 2022 appear to be more diverse than previous populations, as the number of races identified each year from 2013 to 2019 ranged from five (2014) to eleven (2017) (Fetch et al. 2021a, 2021b). Races TGN and TJS were identified each year in Manitoba during 2013 to 2022 (Fetch et al. 2021a, 2021b, Table 4). Races TJJ and SGD were observed in nine (not in 2019) and eight (not in 2013 and 2014) of the 10 years, respectively. Races SGB and TGS were not identified in 2013 to 2015, but were identified every year from 2016 to 2022. All isolates identified to race in the USA were TGN in 2020 and 2022 (Cereal Rust Laboratory 2020, 2022), and 2021, races TJS and SGD were identified from isolates collected in Texas and race TGN was identified from Minnesota (Cereal Rust Laboratory 2021).

Table 4. Frequency (%) of races of Puccinia graminis f. sp. avenae obtained from trap plots, cultivated and wild oat in Manitoba, Ontario and Saskatchewan during 2020 to 2022.

	2020				2021				2022
Race	Manitoba and Saskatchewan^a		Ontario		Manitoba and Saskatchewan^b		Ontario		Manitoba and Saskatchewan^c		Ontario
	No.	%	No.	%	No.	%	No.	%	No.	%	No.	%
NGB^d									7	2.5
SGB	70	16.9			24	14.9			16	5.7
SGD	30	7.2			6	3.7			11	3.9
TDD			1	7.7							6	66.7
TDJ			7	53.8			1	50			1	11.1
TGB	29	7.0	1	7.7	4	2.5			29	10.4
TGD	35	8.4			18	11.2			1	0.4
TGL	16	3.9	1	7.7	2	1.2			45	16.1
TGN	183	44.1	3	23.1	77	47.8			73	26.1	2	22.2
TGS	3	0.7			6	3.7			20	7.1
TJB	4	1.0			3	1.9			6	2.1
TJD					1	0.6
TJG	2	0.5							2	0.7
TJJ	12	2.9			1	0.6	1	50	35	12.5
TJN	3	0.7			3	1.9			4	1.4
TJS	28	6.7			16	9.9			31	11.1
Total isolates assessed	415		13		161		2		280		9

^aTen isolates were assessed from Saskatchewan in 2020; four were race SGB, one was race TGL, and five were race TGN.

^bFour isolates were assessed from Saskatchewan in 2021; all were race SGB.

^cFive isolates were assessed from Saskatchewan in 2022; four were race TGN and one was race TJJ.

^dThe Pga races were given a three letter code, with the first letter referring to the reactions on Pga1, Pga2, Pga3 and Pga4, the second letter referring to Pga6, Pga8, Pga9 and Pga10 and the third letter referring to Pga12, Pga13, Pga15 and Pga16 (Fetch and Jin 2007).

The most common races identified from Ontario in 2020 to 2022 were TDJ (33%), TDD (26%), and TGN (14%), although only 27 isolates were identified to race over these 3 years (Table 4). Comparisons to race frequencies in 2013 to 2019 were not done because of the low number of isolates from Ontario identified to race during this period (Fetch et al. 2021a, 2021b).

Acknowledgments

The authors thank all of the cooperators who grew trap plot nurseries or submitted stem rust infected samples. This work was supported by Agriculture and Agri-Food Canada.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

The work was supported by Agriculture and Agri-Food Canada.