Control of pathogens of Triticum aestivum using endophytic fungal isolates. A. ABAYA AND T. HSIANG. Environmental Sciences, Bovey 3224, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada
More than 100 endophytic fungi were collected from root and leaf tissues of field grown winter wheat ‘AC Morley’ and ‘25R34’ and spring wheat ‘Sumai’ and ‘Scotia’ which are all considered to be moderately resistant to fusarium head blight. The plants were collected from Ariss, Ontario in June 2016. These fungal strains were screened using dual culture plates for antagonism against three wheat pathogens Fusarium graminearum, Waitea circinata and Microdochium majus. Out of 101 strains, 38 were found to be antagonistic showing distinct inhibition zones with at least one of the three wheat pathogens. These antagonistic strains were then tested for pathogenicity on wheat leaves in lab tests, and 16 showed no pathogenicity. Among these non-pathogenic strains, three were selected and identified by internal transcribed spacer-polymerase chain reaction (ITS-PCR) as Valsa friesii, Simplicillium lamellicola and Cladorrhinum flexuosum. These three were then tested for their ability to inhibit disease in growth room tests along with a positive control, Clonostachys rosea ACM941. When agar plug inocula were applied to 21-day-old wheat leaves (four-leaf stage) at 3 days prior to inoculation with agar plugs of Fusarium graminearum, S. lamellicola and ACM941 significantly reduced disease severity. The three wheat endophytes and ACM941 were effective against W. circinata for disease reduction, but only ACM941 was effective against M. majus. Simplicillium lamellicola is a promising biocontrol agent against F. graminearum and W. circinata.
Preliminary sensitivity assessment of Botrytis cinerea isolates to four fungicides from wild blueberry fields. J. A. ABBEY, D. PERCIVAL, S. K. ASIEDU, B. PRITHIVIRAJ AND A. SCHILDER. Department of Plant, Food, and Environmental Sciences, Dalhousie University, Faculty of Agriculture, 50 Pictou Road, P.O. Box 550, Truro, NS B2N 5E3, Canada; and (A.S.) Department of Plant, Soil and Microbial Sciences, Michigan State University, Center for Integrated Plant Systems 578 Wilson, Room 105 CIPS East Lansing, MI 48824, USA
Botrytis cinerea is a high risk pathogen capable of developing resistance to various groups of fungicides. Fifteen single-spore isolates of B. cinerea were collected from commercial wild blueberry fields in Nova Scotia. Eight baseline isolates were also collected to evaluate resistance development. The isolates were evaluated for their sensitivity to cyprodinil, fludioxonil, boscalid and penthiopyrad, using mycelium growth assay. The EC50 values for the 15 isolates ranged from 0.04–10.03, 0.0047–0.0073, 0.47–9.25 and 0.15–1.88 for cyprodinil, fludioxonil, boscalid and penthiopyrad, respectively. Results from this study revealed the potential existence of cyprodinil and boscalid-resistant strains at frequencies of 100 and 73.3%, respectively. Compared with the baseline isolates, reduced sensitivity to penthiopyrad was found in two isolates, whereas 11 isolates exhibited reduced sensitivity to cyprodinil and boscalid, respectively. No isolate with reduced sensitivity to fludioxonil was detected. Significant cross-resistance existed between the SDHI fungicides boscalid and penthiopyrad (r = 0.671, P = 0.006). A negative linear correlation was observed between fludioxonil and boscalid (r = −0.583, P = 0.023). Though some isolates had reduced sensitivity to more than one fungicide, no cross-resistance was detected in the remaining fungicide pairs. This study reveals a possible shift of B. cinerea isolates towards resistance development to cyprodinil and boscalid. It also suggests a prompt occurrence of B. cinerea populations resistant to penthiopyrad unless suitable resistant management strategies are employed to curb future resistance challenges.
Distribution of viable resting spores of Plasmodiophora brassicae in infested fields. F. AL-DAOUD, B. D. GOSSEN AND M. R. MCDONALD. Department of Plant Agriculture, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada; and (B.D.G.) Saskatoon Research and Development Centre, Agriculture and Agri-Food Canada, 107 Science Place, Saskatoon, SK S7N 0X2, Canada
Resting spores of Plasmodiophora brassicae Woronin, the causal agent of clubroot of Brassicaceae crops, can survive in soil for many years. The objectives of this study were to assess the vertical distribution of viable spores in newly infested and heavily infested fields in Alberta, Canada, and the horizontal distribution of viable spores in a hot-spot in one infested field. Soil cores (1–3 cores per site, 2–3 sites per field) were collected at 7.5 cm increments from 0–45 cm depth from three fields. Also, samples (five per site) to 15 cm depth were collected from 15 sites within a 0.5 ha section of a hot-spot in one field. Spores were quantified using qPCR and propidium monoazide-assisted qPCR (PMA-PCR). PMA inhibits amplification of DNA of non-viable cells in qPCR. High variability in the vertical and horizontal distribution of spores was observed in each field. In general, spores were detected at greater depths in heavily infested fields (103–105 spores g−1 soil at 30–45 cm deep) as compared with the newly infested field (105 spores g−1 soil, only in 0–7.5 cm sample). Most of the spores found in the heavily infested field were viable. The horizontal distribution of viable spores was highest in the middle of the hot-spot (105–106 spores g−1 soil) and declined rapidly towards the outer sampling sites (103–104 spores g−1 soil). This supports previous reports on the high site to site variability of spore concentration within a field, and that resting spores move downward in the soil profile over time.
Survey of clubroot (Plasmodiophora brassicae) pathotypes in canola and Brassica vegetable fields in Ontario in 2017. F. AL-DAOUD, M. MORAN, T. J. CRANMER, M. J. CELETTI, B. D. GOSSEN, A. TENUTA AND M. R. MCDONALD. Department of Plant Agriculture, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada; (M.M.) Ontario Ministry of Agriculture, Food, and Rural Affairs, 63 Lorne Avenue East, Stratford, ON N5A 6S4, Canada; (T.J.C., M.J.C., A.T.) Ontario Ministry of Agriculture, Food, and Rural Affairs, Guelph, ON N1G 4Y2; and (B.D.G.) Saskatoon Research and Development Centre, Agriculture and Agri-Food Canada, 107 Science Place, Saskatoon, SK S7N 0X2, Canada
Plasmodiophora brassicae Woronin is the causal agent of clubroot, a major disease affecting several economically important members of the family Brassicaceae. Clubroot is endemic on Brassica vegetables in many regions of Ontario, and in 2016 it was reported for the first time on canola in the province. The pathotype of P. brassicae was assessed from six canola fields and seven vegetable fields in Ontario in 2017. The inoculum from clubbed roots was increased on a susceptible host (Shanghai pak choy cv. ‘Mei Qing Choi’, Brassica rapa var. chinensis) grown under controlled conditions (25/20ºC day/night, 60% relative humidity, 18 h photoperiod). The resulting clubs were harvested at 6 weeks post inoculation (wpi) and used to inoculate cultivars that comprise Williams’ differential set. Four replicates were used with five to six plants per experimental unit. Plants were rated for clubroot symptoms using a 0–3 scale at 5 wpi, and a disease severity index (DSI) was calculated. A host was resistant if DSI + 95% confidence interval < 50%; otherwise it was susceptible. Samples from four canola fields were pathotype 2, one was pathotype 5 and one was pathotype 8. Samples from five vegetable fields were pathotype 6, one was pathotype 5, and one is being determined. Therefore, the most common pathotype in canola fields is pathotype 2 and in vegetable fields it is pathotype 6. This is similar to survey results from 1969 where pathotype 6 was found on cabbage and cauliflower and pathotype 2 was identified from a field of clubroot-infected rutabagas.
The cross-kingdom biosynthesis and signalling of cytokinins during the formation of tumours in the Ustilago maydis–Zea mays pathosystem. I. O. ALIMI, R. J. N. EMERY AND B. J. SAVILLE. Environmental and Life Sciences Graduate Program, Trent University, 1600 West Bank Drive, Peterborough, ON K9J 0G2, Canada; (R.J.N.E.) Biology Department, Trent University, 1600 West Bank Drive, Peterborough, ON K9J 0G2, Canada; and (B.J.S.) Forensic Science Program, Trent University, 1600 West Bank Drive, Peterborough, ON K9J 0G2, Canada
Cytokinins are plant hormones that stimulate cell growth and cell division. During the Ustilago maydis infection of Zea mays, the levels of cytokinins rise dramatically and are thought to be integral to tumour growth. Since both organisms can produce cytokinins, we determined the timing of cytokinin production and the expression of biosynthetic and response regulator genes in both organisms during pathogenesis in order to determine the biosynthetic origins of cytokinin and the biological responses of each organism to these cytokinins. Primers were designed for species and gene specific amplification allowing us to use reverse transcription PCR to detect and estimate levels of transcripts at different stages of infection and in the organisms grown independently. We also identified and determined the levels of cytokinin present at the different stages of infection. The results indicate that both organisms contribute to biosynthesis of cytokinins and that both respond to cytokinin production. The alteration in timing of cytokinin production, and the expression of response regulators during infection with a U. maydis strain that does not produce cytokinins, suggests a model that involves early synthesis of cytokinins by the fungus. This production may stimulate increased pathosystem production of cytokinins and enable response to the cytokinins at specific stages in pathogenesis. We hypothesize that response to cytokinins is a component of the signalling that stimulates changes in cell growth and cell division in both the host and the pathogen. Progress on experimental results that support this model of interaction were presented.
Virulence of Puccinia striiformis f. sp. tritici in Western Canada. E. AMUNDSEN, K. GHANBARNIA AND R. ABOUKHADDOUR. Lethbridge Research and Development Centre, Agriculture and Agri-Food Canada, 5403–1 Avenue South, Lethbridge, AB T1J 4B1, Canada
The virulence of 35 different isolates of Puccinia striiformis f. sp. tritici was tested on 18 near-isogenic wheat lines in the Avocet background. These isolates are part of a collection obtained mainly from western Canada during 2015, 2016 and 2017. The seedlings were inoculated with a spore/talc mixture (ratio 1:20) and infection types (ITs), on the second leaf, were recorded 18–21 days after inoculation based on a scale of 0–9. In total, 21 different virulence patterns were observed. Near-isogenic wheat lines with resistance genes Yr1, Yr 5, Yr15 and Yr76 remain effective against all tested isolates, and the line harbouring YrSp was defeated twice. Lines possessing Yr6, Yr7, Yr8 and Yr9 genes, were defeated by most tested isolates maybe due to worldwide use of these genes in commercial cultivars. This study represents results on a small subset of collected isolates, but it shows wide spectrum virulence in tested isolates and high variability in the yellow rust pathogen populations in Canada.
Introducing an odd pathogen of coriander: Heterosphaeria. C. L. ARMSTRONG-CHO AND S. BANNIZA. Crop Development Centre, University of Saskatchewan, 51 Campus Drive, Saskatoon, SK S7N 5A8, Canada
Renewed research on blossom blight of coriander in Saskatchewan has led to identification of a pathogen implicated in the majority of disease outbreaks observed from 2015–2017. This fungal organism is very slow-growing, difficult to recover using conventional plating, and easily overlooked due to its lack of distinct structures when grown on various agar-based media. A tentative identification was reached using sequence information of the ITS and TUB regions in consultation with taxonomists, placing this fungus in the Leotiomycetes, close to Heterosphaeria patella (Tode) Grev. Macroconidia have been successfully induced to form on wheat straw, and efforts to induce teleomorph formation are underway in order to better assess the taxonomic position of this organism. The sensitivity of this pathogen to strobilurin fungicides was investigated as well as its ability to infect other Apiaceae crop flowers. Primers for molecular detection of the pathogen in plant and seed samples will be used to assess seed to seedling transmission. Germplasm collections of coriander were screened in 2017 for their reaction to Heterosphaeria infection in the hopes of finding a source of disease resistance.
Genotypic and phenotypic diversity of Phytophthora sojae isolates in Canadian soybean fields. G. ARSENAULT-LABRECQUE, C. DUSSAULT-BENOIT, H. SONAH, F. BELZILE AND R. R. BELANGER. Département de phytologie, Université Laval, 2425 rue de l’Agriculture, Québec, QC G1V 0A6, Canada
Phytophthora sojae, causing phytophthora root rot (PRR), has been present in Canada since 1950. The expansion of Canadian soybean in recent years gave this pathogen a new niche to establish its devastating presence. The best method to control it is through the use of soybean varieties carrying resistance genes (Rps) that provide immunity against P. sojae isolates carrying the corresponding avirulence genes (Avr). Breeders are thus confronted with the need to introgress Rps genes based on the P. sojae pathotypes found in the environment, while this information is constantly incomplete because of the rapid evolution of the pathogen versus the unwieldiness of phenotyping methods. This project aimed to determine the presence and distribution of virulence profile (pathotypes) of P. sojae in Canada, based on the seven most important Rps/Avr genes relationships. For this purpose, a collection of 31 P. sojae isolates representing the most common pathotypes found in Canadian fields were targeted for whole-genome sequencing. Different gene mutations directly linked to those seven Avr genes from P. sojae, mostly gene suppression and SNPs, were discovered. These findings corroborate some previous reports, and highlight new findings, while demonstrating the reliability of genomic markers to predict phenotypes of P. sojae isolates.
Interaction between soybean cyst nematode (Heterodera glycines) and phytophthora root rot (Phytophthora sojae) on soybean. C. AUDETTE, R. R. BÉLANGER AND B. MIMEE. (C.A., B.M.) Saint-Jean-sur-Richelieu Research and Development Centre, Agriculture and Agri-Food Canada, 430 boulevard Gouin, Saint-Jean-sur-Richelieu, QC J3B 3E6, Canada; and (C.A., R.R.B.) Département de phytologie, Faculté des Sciences de l’Agriculture et de l’Alimentation, Université Laval, Québec, QC G1V 0A6, Canada
In Canada, soybean acreage has been increasing steadily in recent years. However, this broad expansion has contributed to the establishment and spread of several diseases and pests. The soybean cyst nematode (SCN), Heterodera glycines, is the leading cause of economic losses in the USA. It has been present in Ontario since 1987 and was reported in Quebec in 2013. Currently, phytophthora root rot (PRR) caused by Phytophthora sojae is still the most problematic disease of soybean in Canada. The best tool to control these pathogens is the use of resistant cultivars. It is known that SCN may facilitate the development of certain diseases (e.g. Fusarium virguliforme) or repress some host resistance genes (e.g. Cadophora gregata). Thus, the objective of this project was to determine if cultivars with resistance to PRR and SCN will remain effective if both pathogens are present. This study revealed that the presence of P. sojae negatively affected the number of cysts produced by SCN on susceptible host (more than 50% reduction). This suggests that P. sojae may activate host defence mechanisms that could be effective against SCN or directly decrease SCN virulence. On the other hand, plant resistance was not affected by the presence of either pathogens and remained effective against both pathogens. This confirms that the use of resistant cultivars is a valid option for controlling and preventing SCN and PRR even when both organisms co-occur in a field.
Field testing of alfalfa populations carrying root-rot resistance and freezing tolerance. P. AUDY, S. ROCHER AND A. CLAESSENS. Quebec Research and Development Centre, Agriculture and Agri-Food Canada, 2560 Hochelaga Boulevard, Québec, QC G1V 2J3, Canada
Two important traits affecting productivity in alfalfa under cold climate conditions are root-rot disease resistance and freezing tolerance. In the last few years, Castonguay’s group in Quebec has improved several alfalfa backgrounds for their tolerance to freezing (TF populations) using recurrent selections under controlled conditions (Castonguay et al., 2009). In the present study, we have used two alfalfa cultivars, ‘Apica’ and ‘Caribou’ that has previously gone through three cycles of recurrent selection for freezing tolerance (TF3-derived populations). We proceeded with three additional cycles of selection for resistance to phytophthora root rot (PRR) caused by Phytophthora medicaginis. PRR is one of the major causes of decline of established alfalfa in North America. At each cycle of selection, 1500 genotypes of these two PRR-sensitive backgrounds were screened using a blend of four PRR isolates coming from diverse locations in Canada. PRR-resistant cultivar Amerigraze and PRR-sensitive cultivar ‘Saranac’ seedlings were used as controls. The 100 most PRR-tolerant genotypes of each background were selected and intercrossed to generate seeds for the next cycle of selection (e.g. APRR1 for Apica TF3 after one cycle of selection, CPRR1 for Caribou TF3 after one cycle of selection, etc.). We assessed these alfalfa populations (for Apica: ATF3, APRR1, APRR2, APPR3; for Caribou: CTF3, CPRR1, CPRR2, CPRR3; and controls: Amerigraze, Saranac) for their yield performance under clay-rich field conditions for 2 years (third year now) in plots near Quebec City. A 3-summer cut schedule was used. Phytophthora medicaginis is a naturally occurring pathogen in the chosen field and therefore, the test plot was not further inoculated. Aphanomyces euteiches was also present in the soil but to a lesser extent. Alfalfa populations with improved PRR-resistance were significantly more productive than the original TF3-Apica and TF3-Caribou populations (improved tolerance to freezing). The notable yield increase was mostly due to the fact that many more alfalfa plants survived in the PRR-improved populations whereas no significant yield difference between the surviving plants was found for all treatments.
In vitro antagonism of biocontrol agents against fungal diseases affecting hemp and marijuana. C. BALTHAZAR, M. FILION AND D. L. JOLY. Université de Moncton, 18 avenue Antonine-Maillet, Moncton, NB E1A 3E9, Canada
Cannabis sativa L. is a herbaceous plant grown worldwide for its narcotic and medicinal uses (marijuana) or as a source of food and fibre (hemp). Following the legislative changes to legalize recreational marijuana in Canada this year, a renewed interest for this crop now raises concerns about potential recrudescence of its associated diseases and pests. Of particular interest are pathogenic fungi, due to the magnitude of their impact on C. sativa. Our goal is to develop and characterize wide-spectrum biological agents that would help control six fungi that have been isolated from hemp or marijuana: Botrytis cinerea Pers. causing grey mould, Sclerotinia sclerotiorum (Lib.) de Bary causing hemp canker, Fusarium sp. causing wilt, root rot and damping-off, Alternaria alternata (Fr.) Keissl. causing brown blight, Nigrospora sp. potentially causing leaf spot and Phoma glomerata (Corda) Wollenw. & Hochapfel causing brown leaf spot and stem canker. Our biological agents are soil-borne beneficial bacteria screened from 184 strains of Pseudomonas spp. and Bacillus spp. Confrontational assays were set up in vitro and revealed great biocontrol potential for at least two strains of Pseudomonas spp. and three of Bacillus spp. Whole genome sequencing and transcriptomic studies are currently underway, along with a validation of the reduction of symptoms in planta. Biological control could be a viable alternative to the use of chemical fungicides, the use of which is controversial in Cannabis production. Moreover, a better understanding of the molecular interactions between the host plant, the pathogenic fungi and the beneficial bacterium could offer new insights to control emerging threats against this crop.
Towards the identification of critical time-points of Colletotrichum lentis infection on lentil. P. BAWA, J. HALLIDAY, C. CHO, V. BHADAURIA, K. BETT, A. VANDENBERG AND S. BANNIZA. Department of Plant Sciences/Crop Development Centre, 51 Campus Drive, University of Saskatchewan, Saskatoon, SK S7N 5A8, Canada; and (V.B.) Swift Current Research and Development Centre, Agriculture and Agri-Food Canada, P.O. Box 1030, Swift Current, SK S9H 3X2, Canada
Anthracnose is one of the major foliar diseases of lentil in western Canada and remains an impediment to the production of high quality seeds and yields of lentil. Two pathogenic races (races 0 and 1) have been differentiated in western Canadian populations of the causal pathogen of anthracnose, Colletotrichum lentis. Previous microscopic studies revealed that race 1 isolates had lower conidial germination, fewer appressoria, and a slower and less destructive necrotrophic phase compared to isolates of the more virulent race 0 after inoculation on lentil cultivar ‘CDC Robin’ with partial resistance to the less virulent race 1. Consistent with this, we found that fungal biomass of a race 0 isolate in ‘CDC Robin’ assessed at different time points through quantitative real-time PCR (qPCR) was higher than that of the race 1 isolate. Accessions of Lens ervoides, a wild relative of lentil, have been identified with high levels of resistance to both pathogenic races of C. lentis. An intraspecific L. ervoides population (LR-66) was previously developed from accessions L01-827A × IG 72815 to study the genetic control of resistance to C. lentis. The growth and development of a virulent race 0 isolate in the most resistant and the most susceptible recombinant inbred lines through qPCR has been assessed to identify critical time-points for in-depth gene expression studies.
Genetic analysis of ergot resistance in a Canada Western Red Spring wheat population. S. BERRAIES, H. L. CAMPBELL, R. E. KNOX, R. D. CUTHBERT, Y. RUAN, V. BHADAURIA, B. MEYER, S. KUMAR AND R. M. DEPAUW. Swift Current Research and Development Centre, Agriculture and Agri-Food Canada, 1 Airport Road, Swift Current, SK S9H 3X2, Canada; (S.K.) Department of Genetics and Plant Breeding, Chaudhary Charan Singh University, Meerut, Uttar Pradesh, India; and (R.M.D.) Advancing Wheat Technology, 870 Field Drive, Swift Current, SK S9H 4N5, Canada
The fungal pathogen Claviceps purpurea (Fr.) Tul. infects wheat ovaries during flowering causing ergot disease. The disease is significant because toxic alkaloids present in ergot sclerotia are harmful to the circulatory system and neurotransmission of both humans and animals when infested grain is consumed. Few sources of resistance to C. purpurea have been reported in wheat and no specific resistance genes have been identified. We grew in the field near Swift Current, Canada over multiple years a doubled haploid hard red spring wheat population of 774 lines from a cross of ‘Carberry’ by ‘AC Cadillac’ and evaluated natural ergot infection. Genotyping was done with the Infinium II iSelect 90K wheat assay and 6806 single nucleotide polymorphisms were mapped. Composite interval mapping detected seven quantitative trait loci (QTL). ‘Carberry’ contributed resistance alleles for QTL on chromosomes 2B, 5A and 6A while QTL on chromosomes 2A, 3D2, 6B and 7B were contributed by ‘AC Cadillac’. The expression of the 2B, 5A and 6B QTL was more stable as they were detected in more than one environment. Among the seven QTL, the 7B QTL explained the least phenotypic variation of 1.3%, while the QTL on chromosome 6B accounted for the most phenotypic variation at 4.4%. This information will be valuable in marker-assisted breeding for ergot resistance in hexaploid wheat.
Genomic profiling of sudden oak death populations. G. J. BILODEAU, R. HEINZELMANN, N. FEAU, A. DALE AND R. C. HAMELIN. Canadian Food Inspection Agency, 3851 Fallowfield Road, Ottawa, ON K2H 8P9, Canada; (R.H., N.F., A.D.) University of British Columbia (UBC), 2424 Main Mall, Vancouver, BC V6T 1Z3, Canada; and (R.C.H.) UBC and Universite Laval, 1030, avenue de la Médecine, Quebec, QC G1V 0A6, Canada
Phytophthora ramorum is the causal agent of sudden oak death (SOD) and sudden larch death in the Western USA and the UK, respectively, as well as ramorum blight on several woody ornamental plants. Prior to 2009, three genetically divergent clonal lineages of this pathogen were known (EU1, NA1 and NA2), each named according to the continent where it was first detected. In 2009, a fourth lineage (EU2) was discovered in the UK. Several different markers have been developed for genotyping P. ramorum including ASO-PCR (Allele-specific oligonucleotide-PCR), microsatellites and TaqMan assays; however, SNP markers can be used to gain a better understanding of inter- and intra-lineage genetic diversity and population structure. By targeting multiple SNPs and SNPs unique to geographic populations, we can also gain a better understanding of migration patterns. One of the objectives of the BioSAFE (BioSurveillance of Alien Forest Enemies) project is to study the genomic epidemiology of SOD populations. We aim to sequence the genomes of approximately 500 P. ramorum individuals covering the entire Canadian outbreak, as well as some of the US and European outbreaks, across several years and nurseries in order to track the Canadian SOD outbreak, uncover migration patterns, and identify sources and pathways.
First report of radish (daikon) showing Verticillium longisporum infection in Manitoba, Canada. V. BISHT, M. PRADHAN, C. CAVERS, D. FERNANDO, Z. ZOU AND T. BARASUBIYE. Manitoba Agriculture, 65-3rd Avenue NE, Carman, MB R0G 0J0, Canada; (M.P.) Crop Diagnostic Centre, Manitoba Agriculture, 545 University Crescent, Winnipeg, MB R3T 5S6, Canada; (C.C.) Agriculture and Agri-Food Canada (AAFC), 370 River Road, Portage La Prairie, MB R1N 3V6, Canada; (D.F., Z.Z.) Plant Sciences, University of Manitoba, Room 205 FAFS, 66 Dafoe Road, Winnipeg, MB R3T 2N2, Canada; and (T.B.) Science and Technology Branch, Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada
Verticillium wilt (renamed verticillium stripe) disease was identified for the first time in 2014 from Manitoba, Canada on canola (Brassica napus). Since the first discovery, survey of crops and soils has confirmed the presence of the pathogen in several provinces in Canada. In 2015, in a Portage La Prairie field, there was a high incidence of verticillium stripe disease in a blackleg susceptible canola variety, but not in a blackleg resistant variety, suggesting interaction between the two pathogens. In 2016, in this infested field, various crucifer crops (broccoli, canola, cauliflower, daikon radish, mustard and rutabaga) were planted in small plots to test the host reaction to Verticillium. Daikon radish, Raphanus raphanistrum subsp. sativus, showed external stem discolouration and internal vascular tissue blackening in stem and root. The incidence appeared to be high, over 50%. In 2017, daikon radish was planted again on the farm, in an adjoining field. Survey of the field showed > 50% plants with internal vascular blackening. Isolation of the pathogen indicated Verticillium species. Based on DNA sequence of ITS barcode and partial sequence of Actin gene the strain isolated was identified as Verticillium longisporum Hybrid (A1 × D1). This appears to be the first report of V. longisporum infecting radish in Manitoba, Canada.
A stable quantitative trait locus conditioning leaf rust (Puccinia triticina) resistance on chromosome 2D of the wheat cultivar ‘Stettler’. F. E. BOKORE, R. D. CUTHBERT, R. E. KNOX, B. D. MCCALLUM, C. HIEBERT, A. N’DIAYE, R. DEPAUW, C. MCCARTNEY, C. J. POZNIAK, Y. RUAN, C. MUNRO, H. L. CAMPBELL AND B. MEYER. Swift Current Research and Development Centre, Agriculture and Agri-Food Canada (AAFC), 1 Airport Road, P.O. Box 1030, Swift Current, SK S9H 3X2, Canada; (B.D.M., C.H., C.M.) Morden Research and Development Centre, AAFC, 101 Route 100, Unit 100, Morden, MB R6M 1Y5, Canada; (A.N., C.J.P.) Department of Plant Sciences and Crop Development Centre, University of Saskatchewan, 51 Campus Drive, Saskatoon, SK S7N 5A8, Canada; (R.D.) Advancing Wheat Technology, 870 Field Drive, Swift Current, SK S9H 4N5, Canada; and (C.M.) Plant and Food Research, Canterbury Agriculture and Science Centre, Gerald Street, Lincoln 7608, New Zealand
‘Stettler’ is moderately susceptible to prevalent races of leaf rust (Puccinia triticina Erikss.) but expresses uncharacterized resistance. The objective of this study was to investigate the genetic basis of resistance to leaf rust in ‘Stettler’. A doubled haploid population of 218 lines was developed from a cross of ‘Stettler’ with the heritage cultivar ‘Red Fife’. The population was evaluated for leaf rust reaction in four field nursery environments in Canada near Swift Current, SK in 2014 and 2015, Morden MB in 2015 and New Zealand near Lincoln in 2014. Genotyping was performed using the 90K Infinium iSelect assay and linkage maps were constructed by JoinMap using 1548 non-redundant markers. Quantitative trait locus (QTL) analysis was performed in MapQTL6. ‘Stettler’ generally exhibited lower leaf rust infection than ‘Red Fife’ and most DH lines. A stable QTL conditioning leaf rust resistance was identified from ‘Stettler’ on chromosome 2DS. QTL confined to single environments were also contributed by ‘Stettler’ on chromosome 6B and ‘Red Fife’ on chromosomes 7A and 7B. Although SNP markers associated with the QTL on 2DS are in the proximity of SSR markers that co-segregate with Lr22, based on pedigree and level of gene expression, the ‘Stettler’ locus is more likely Lr2a than Lr22, but could be another unique gene. With the genetic markers reported here, the potential exists for the stacking of the 2D minor resistance gene with other genes.
Evaluation of antagonistic activity of Bacillus pumilus and Bacillus subtilis against Botrytis cinerea. M. BOUCHARD-ROCHETTE, T. T. A. NGUYEN, R. NAASZ, H. ANTOUN AND R. J. TWEDDELL. Département de phytologie, Université Laval, Québec, QC G1V 0A6, Canada; (T.T.A.N., H.A.) Département des sols et de génie agroalimentaire, Université Laval, Québec, QC G1V 0A6, Canada; et (R.N.) Premier Tech, 1 avenue Premier, Rivière-du-Loup, QC G5R 6C1, Canada
Grey mould, caused by the fungus Botrytis cinerea Pers., is one of the most damaging diseases affecting horticultural crops. Bacillus pumilus Meyer and Gottheil strain PTB 180 and Bacillus subtilis (Ehrenberg) Cohn strain PTB 185 have shown antifungal activity against several soil-borne fungal pathogens. This study aims to investigate in vitro the antagonistic activity of PTB 180, PTB 185 and a mix (1:1) of both strains against B. cinerea. The antagonistic activity of PTB 180 and PTB 185 was evaluated on agar (using a double layer technique) and on tomato (Solanum lycopersicum L.)/cucumber (Cucumis sativus L.) leaf discs placed in Petri plates. Both strains were shown on agar to inhibit B. cinerea mycelial growth by more than 90% and spore germination by more than 50%. When applied (1 × 107 colony forming units mL−1; 1 mL) prior to B. cinerea inoculation (1 × 106 propagules mL−1; 100 µL), PTB 180, PTB 185 and a mix (1:1) of both strains inhibited significantly mycelial growth of the fungus on tomato/cucumber leaf discs as compared with the control. In conclusion, PTB 180 and PTB 185 showed strong antagonistic activity against B. cinerea, suggesting that they could eventually find application as biocontrol agents against grey mould. Greenhouse tests will be conducted to evaluate their efficiency in controlling the disease on tomato/cucumber plants.
Survival of Bacillus pumilus and Bacillus subtilis on the phyllosphere of greenhouse tomato and cucumber. M. BOUCHARD-ROCHETTE, T. T. A. NGUYEN, R. NAASZ, H. ANTOUN AND R. J. TWEDDELL. Département de phytologie, Université Laval, Québec, QC G1V 0A6, Canada; (T.T.A.N., H.A.) Département des sols et de génie agroalimentaire, Université Laval, Québec, QC G1V 0A6, Canada; et (R.N.) Premier Tech, 1 avenue Premier, Rivière-du-Loup, QC G5R 6C1, Canada
Bacillus pumilus Meyer and Gottheil strain PTB 180 and Bacillus subtilis (Ehrenberg) Cohn strain PTB 185 showed in vitro antagonistic activity against Botrytis cinerea Pers., suggesting that these bacteria could show antagonistic activity against the pathogen in planta. As part of ongoing research on the biocontrol capability of strains PTB 180 and PTB 185 against grey mould (B. cinerea), the objective of the study was to evaluate their survival, when applied alone and in combination, on tomato (Solanum lycopersicum L.) and cucumber (Cucumis sativus L.) plants. Tomato and cucumber plants, aged 4 and 2 weeks respectively, were sprayed until runoff with a suspension (1 × 107 colony forming units (CFU) mL−1) of either strain PTB 180, strain PTB 185 or a mix (1:1) of both strains and were then grown in a greenhouse for 21 days. Three leaves of each tomato/cucumber plant were collected after 0 (1 h after the plants were sprayed), 7, 14 and 21 days. Leaves were shredded, placed in sterile water, and mixed thoroughly. The suspensions were submitted to serial dilutions, heated (55ºC, 15 min) and plated on nutrient agar. After an incubation period of 24 h at 37ºC, the number of CFU was determined. Populations of both strains of Bacillus estimated at about 1 × 107 CFU g−1 of fresh leaf (0 day) remained as high as 5 × 106 CFU g−1 of fresh leaf 21 days following their application on tomato and cucumber plants. In addition, when applied as a mix (1:1), the ratio of each bacterial population remained unchanged.
Age-related susceptibility of grapevine leaves and berries to infection by Elsinoe ampelina. O. CARISSE, A. LEVASSEUR AND C. PROVOST. Saint-Jean-sur-Richelieu Research Centre, Agriculture and Agri-Food Canada, 430 Gouin Boulevard., Saint-Jean-sur-Richelieu, QC J3B 3E6, Canada; and (C.P.) Centre de recherche agroalimentaire de Mirabel, 9850 rue Belle-Rivière, Mirabel, QC J7N 2X8, Canada
Anthracnose, caused by Elsinoe ampelina Shear, is an important disease of grapevine. In recent years, there have been regular outbreaks in Eastern Canada on grape cultivars that are gaining in popularity. Young leaves and berries are reported to be highly susceptible to E. ampelina, but the time of onset and young leaves’ susceptibility among cultivar variation in ontogenic resistance have remained undefined. Age-related susceptibility was studied under greenhouse conditions by inoculating 1- to 19-day-old grape leaves of the cultivars ‘Vidal’, ‘Marquette’ and ‘Vandal-Cliche’. Similarly, flowers/berries were inoculated under field conditions on 10 occasions from flower formation until berries at approximately 8°Brix. For all cultivars, there was a significant effect of leaf and flower/berry age on anthracnose severity. Susceptibility was highest on 1-day-old leaves and diminished as the leaves aged to reach 20%, 10% and 5% of the maximum susceptibility on 4-, 6- and 8-day-old leaves. The influence of leaf age on anthracnose severity was described with an exponential decay model (R2 = 0.98). Susceptibility was highest at the early stage of flower formation and diminished to reach 50%, 40–20% and 5% at the stages of flowers separating (stage 17), fruit set (stage 27) and 4–6 mm berries (stage 29), respectively. These results suggest that the risks of anthracnose are high from bud-break to fruit set, and on newly emerged leaves either early in the season or following pruning. More knowledge on anthracnose epidemiology is needed, but these results could be used to improve timing of fungicide applications and of pruning activities.
First report of verticillium wilt of faba bean (Vicia faba) caused by Verticillium dahliae in western Canada. K. F. CHANG, S. F. HWANG, H. FU, H. U. AHMED, Q. X. ZHOU, H. T. YU, A. J. HO, G. D. TURNBULL AND S. E. STRELKOV. Alberta Agriculture and Forestry, Crop Diversification Centre North, 17507 Fort Road, Edmonton, AB T5Y 6H3, Canada; (H.T.Y.) Institute of Food Crops, Yunnan Academy of Agricultural Science, Kunming, Yunnan, 650205 China; and (A.J.H., S.E.S.) Department of Agricultural, Food and Nutritional Science, 410 Agriculture/Forestry Centre, University of Alberta, Edmonton, AB T6G 2P5, Canada
Faba bean (Vicia faba L.) with tannin-free seed has potential for use as food, feed and aquaculture in the Canadian prairies. However, the crop is susceptible to vascular wilts caused by soil-borne pathogens. In a disease survey of faba bean conducted in 2017, plants with wilt symptoms were observed near Namao, Alberta. Symptomatic plants were collected, and their infected stem tissues were surface-sterilized and plated onto potato dextrose agar (PDA) medium. Five isolates which developed colony morphology similar to that of Verticillium species were purified by hyphal tip culture and single-spore isolation. On PDA, the colonies were dark with compact mycelium embedded with numerous microsclerotia and conidial masses 14 days after plating. The isolates formed conidiophores with 4–5 verticillate phialides. The conidia were hyaline, elliptical, aseptate, with a mean length of 3.34 µm, and range of 2.56–5.14 µm. The mean width was 1.94 µm with a range of 1.31–2.56 µm. The isolates produced irregular to elliptical microsclerotia of various sizes. The DNA sequence of seven isolates, obtained with a primer set (ITS5/4), revealed 99% similarity with Verticillium dahliae Kleb. sequences available in GenBank, confirming the identity of the isolates. Inoculation of one of the isolates onto the faba bean cultivars ‘Earlibird’ and ‘Snowbird’ resulted in typical symptoms and signs of verticillium wilt. The pathogen was re-isolated from all parts of the infected plants. This is the first report of natural infection of faba bean by V. dahliae in western Canada.
Resistance characterization of potato fusarium dry rot. D. CHEN, H. H. TAI, K. GARDNER, B. BIZIMUNGU, S. SOUCY AND R. PETERS. Fredericton Research and Development Centre, Agriculture and Agri-Food Canada (AAFC), 850 Lincoln Road, Fredericton, NB E3B 4Z7, Canada; and (R.P.) Charlottetown Research and Development Centre, AAFC, 440 University Avenue, Charlottetown, PE C1A 4N6, Canada
Potato fusarium dry rot (FDR) is one of the most important storage diseases in potatoes. Annual losses to FDR were estimated at 6–25%, and occasionally up to 60%. Multiple Fusarium spp. can cause FDR with different aggressiveness. Fungicide application is commonly used to control Fusarium diseases, however fungicide resistance has been developed in some Fusarium spp. Use of host resistance is the most effective and environmentally sound approach to manage diseases. However, resistance to FDR in potatoes is not well understood. In the present study, six advanced potato breeding lines and standard variety Jemseg or Russet Burbank were inoculated with three Fusarium spp. (F. sambucinum, F. oxysporum and F. coeruleum) to screen lines with resistance to the FDR, using two inoculated methods. The inoculated tubers were incubated at 13ºC and 95% humidity in the first 48 h after inoculation, and then were kept at 13ºC and 65% humidity for 6 weeks at dark before disease assessment. Two lines, F14034 and F14028 were resistant or highly tolerant to the infection of F. sambucinum, the most aggressive species, compared with the susceptible variety Jemseg. The F. sambucinum was used to investigate the gene differential expression using transcriptome profiling approach to identify host and pathogen genes involved in initial infection responses. The Fusarium-infected tuber samples from the six lines and Jemseg were taken at 0, 4, 10, 24 and 48 h after inoculation with a 7 mm diameter punch. The RNA samples were extracted from the tuber samples for transcriptome profiling.
Towards improved recovery efficiency and identification accuracy of cereal rust fungal pathogens in environmental samples. W. CHEN, S. HAMBLETON, H.-Y. ZHANG, K. CHUENG, M.-L. DE GRAAF, Q. EGGERTSON AND G. BAKKEREN. (W.C., S.H., H.-Y.Z., K.C.) Ottawa Research and Development Centre, Agriculture and Agri-Food Canada (AAFC), 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada; (H.-Y.Z.) Inner Mongolia Agricultural University, 306 Zhaowuda Road, Hohhot, Inner Mongolia, 010000, China; and (M.-L.deG., G.B.) Summerland Research and Development Centre, AAFC, 4200 Highway 97, Summerland, BC V0H 1Z0, Canada
The rust fungus family Pucciniaceae includes devastating pathogens to cereal crops which produce airborne spores disseminated by wind and storms, generating a source of inocula to nearby and distant host plants. The abundance and dispersal pathways of the inocula can be monitored by analysing DNA from spore collectors using metabarcoding approaches, providing early warning of potential disease epidemics. From 2007–2011 (n = 344) and 2015–2016 (n = 117), air/rain-borne spore samples were collected by a network of equipment in agricultural fields across Canada. ITS DNA barcodes, obtained using universal primers ITS5/ITS4 to amplify both internal transcribed spacers (ITS1 and ITS2) and 454-pyrosequencing technology in 2007–2011, recovered only 17 Pucciniaceae Operational Taxonomic Units (OTUs) (0.2% richness, 0.38% total abundance). Using rust-enhanced primers targeting ITS2 (Rust2inv/ITS4var) and the Illumina MiSeq platform in 2015–2016, 1594 Pucciniaceae OTUs (10.5% richness, 7.65% total abundance) were recovered and they showed spatial-temporal distribution patterns in Canada’s air. The identity of these metabarcodes were evaluated utilizing signature oligonucleotides designed from a curated in-house reference database using the automated oligonucleotide design pipeline (AODP). Our results show that the universal primer set, commonly used for fungal DNA barcoding studies, did not recover the near-complete diversity of rusts from environmental DNAs, partially due to low primer efficiency and the challenge in sequencing the ITS1 region for this group. Considering the severe economic loss related to cereal rust epidemics, improved recovery efficiency and identification accuracy of the causal agents are essential knowledge for establishing a reliable pest management protocol.
Adoption of major-gene resistance groups for managing blackleg (Leptosphaeria maculans) of canola in Canada. J. E. J. CORNELSEN, C. JURKE AND C. REMPEL. Canola Council of Canada, 400–167 Lombard Avenue, Winnipeg, MB R3B 0T6, Canada
The fungal pathogen Leptosphaeria maculans (Desmaz.) Ces. & de Not. causes blackleg, one of the most economically important diseases of canola (Brassica napus L.) in Canada. Management for the disease has focused on pathogen race monitoring, extension of crop rotation, and use of resistant (R) cultivars. R cultivars have been deployed since the 1990s, which helped to minimize the disease impact by using major-gene and race-nonspecific resistance. Recently, an increase in blackleg incidence and severity has been observed on the prairies, even on R-rated cultivars. The recommendation for canola producers finding increased blackleg was to select a different cultivar, which could be a gamble; it may or may not be effective since the new cultivar may carry the same set of resistance gene(s). The Canadian canola industry has decided to adopt a new resistance labelling scheme to identify the major resistance genes deployed in canola cultivars. If substantial increases in blackleg are seen with one cultivar, producers will be recommended to switch to a cultivar with an alternative major-gene resistance group to target the dominant L. maculans races within their fields. A new diagnostic test has also been developed for producers to identify L. maculans races and select cultivars carrying effective resistance genes. The adoption of major-gene resistance groups and the L. maculans race diagnostics test will provide producers with new tools to help manage and mitigate blackleg on their farms.
Évaluation de la détection moléculaire et de l’observation visuelle pour établir le seuil de nuisibilité de la tache argentée et de la dartrose sur les semences de pommes de terre. J. D’ASTOUS-PAGÉ, S. MORISETTE, A. GAGNON ET R. HOGUE. Institut de recherche et de développement en agroenvironnement, 2700 rue Einstein, Québec, QC G1P 3W8, Canada; (S.M.) Groupe Pousse-Vert, 301-49 rue de l’Église, Saint-Arsène, QC G0L 2K0, Canada; et (A.G.) Progest2001, 6833 route Marie-Victorin, Sainte-Croix, QC G0S 2H0, Canada
La tache argentée (Helminthosporium solani Durieu & Mont.; Hs) et la dartrose (Colletotrichum coccodes (Wallr.) Hughes; Cc) sont deux maladies reconnues pour causer des anomalies de coloration de l’épiderme des tubercules et réduire la valeur de la récolte. En se basant sur l’observation des taches et la présence de microsclérotes, il est difficile de faire la distinction entre les deux maladies et d’en prédire le développement à l’entreposage. Les symptômes sont parfois visibles à la récolte mais c’est à l’entreposage qu’ils se développent. Comme les traitements au champ et en entrepôt sont limités, la prévention des sources d’inoculum demeure la méthode la plus économiquement efficace à ce jour. Nous avons développé une approche moléculaire de détection de Hs ou de Cc sur les pelures des tubercules prélevés avant la mise en entrepôt de la récolte. Cette détection sensible et économiquement efficiente a permis d’élaborer les bases d’un modèle prévisionnel des dommages de Hs et Cc sur les tubercules en entrepôt. Nos résultats suggèrent que les seuils de nuisibilité de Hs et Cc devraient être adaptés selon le niveau de sensibilité du cultivar. Il a aussi été observé que Hs et le Cc sont détectés par qPCR sur les pelures de tubercules d’apparence saine et sans symptôme, même après plusieurs mois d’entreposage. La compétition entre les deux pathogènes sur le même tubercule est peu connu et pourrait masquer des symptômes. Ainsi, nous proposons une nouvelle approche d’observation et de détection combinée de Hs et Cc.
Identification of QTLs associated with horizontal resistance against Phytophthora sojae in early maturing soybeans. M. DE RONNE, A. LEBRETON, C. LABBÉ, J. LAUR, A. RASOOLIZADEH, C. DUSSAULT-BENOIT, D. GOVARE-MONROE, F. BELZILE, L. O’DONOGHUE AND R. R. BÉLANGER. Envirotron, Université Laval, 2480 Boulevard Hochelaga, Québec, QC G1V 0A, Canada; (F.B.) Pavillon Charles-Eugène-Marchand, 1030 Rue de la Médecine, Québec, QC G1V 0A6, Canada; and (L.O.) CEROM, 740 Chemin Trudeau, Saint-Mathieu-de-Beloeil, QC J3G 0E2, Canada
Although the deployment of resistance genes that confer complete immunity against Phytophthora sojae is currently the most attractive means to reduce soybean losses, the constant evolution of new avirulence genes lead to a breakdown in resistance. A complementary approach is to rely on horizontal resistance, or partial resistance, that is not dependent on a gene-for-gene interaction. The objective of this study is to identify QTLs associated with horizontal resistance against P. sojae, in a soybean recombinant inbred line (RIL) population that has been obtained from one early maturing line adapted to Canadian conditions, and one line showing high levels of horizontal resistance both in field observations and greenhouse testing. For phenotyping purposes, we relied on a new hydroponic bioassay that reproduces the natural infection process of the pathosystem soybean–P. sojae and allows a reproducible evaluation of horizontal resistance among the RILs. In parallel, genotyping-by-sequencing (GBS) was performed on all RILs and the resulting reads (~ 1M/line) were used for SNP calling and construction of a genetic map. The combination of these two innovative approaches led to the identification of new QTLs involved in horizontal resistance of soybean, which will help breeders to develop new varieties adapted for Canadian conditions with a higher durable resistance against P. sojae.
Bacillus pumilus and Bacillus subtilis for the biocontrol of soil-borne diseases of greenhouse cucumber. E. DEMEULE, T. T. A. NGUYEN, R. NAASZ, H. ANTOUN AND R. J. TWEDDELL. Département de phytologie, Université Laval, Québec, QC G1V 0A6, Canada; (T.T.A.N., H.A.) Département des sols et de génie agroalimentaire, Université Laval, Québec, QC G1V 0A6, Canada; et (R.N.) Premier Tech, 1 avenue Premier, Rivière-du-Loup, QC G5R 6C1, Canada
Soil-borne diseases cause important economic losses in greenhouse cucumber (Cucumis sativus L.) production. Chemical fungicides are widely used to control these diseases. However, concerns about their negative effects (development of resistance, residues in food, occupational exposure, environmental impacts) have motivated the development of new alternatives such as biocontrol agents. As part of ongoing research on the biocontrol capability of Bacillus pumilus Meyer and Gottheil strain PTB180 and Bacillus subtilis (Ehrenberg) Cohn strain PTB185, we investigated their antagonistic activity against Rhizoctonia solani Kühn, Pythium ultimum Trow, Sclerotinia sclerotiorum (Lib.) de Bary, Phytophthora capsici Leonian and Fusarium oxysporum Schltdl. on agar using a double layer technique. Greenhouse experiments were subsequently conducted to determine the efficacy of the strains to control collar and root rot on cucumber plants inoculated with R. solani. Results showed that both PTB180 and PTB185 strongly inhibited mycelial growth of R. solani, P. ultimum, S. sclerotiorum, P. capsici and F. oxysporum on agar. PTB180, PTB185 and a mix (1:1) of both strains reduced significantly the severity of collar and root rot caused by R. solani on cucumber plants grown in greenhouse. This study suggests that strains PTB180 and PTB185 could eventually represent a sustainable alternative to the use of chemical fungicides for the control of soil-borne pathogens affecting greenhouse cucumber production.
Date palm–Fusarium oxysporum albedinis interaction: enhancement of defence responses with different non-pathogenic microorganisms. M. El HASSNI, A. EL HADRAMI, F. DAAYF, A. DIHAZI AND K. NAAMANI. Laboratoire de Biotechnologie et Développement Durable des Ressources Naturelles, Faculté Polydisciplinaire, Mghila BP. 592, Université Sultan Moulay Slimane Beni Mellal 23000, Maroc; (A.E.) OMEX Agriculture Inc. 290 Agri Park Road, Oak Bluff, MB R4G 0A5, Canada; (F.D.) Department of Plant Science, 222 Agriculture Building, 66 Dafoe Road, University of Manitoba, Winnipeg, MB R3T 2N2, Canada; and (A.D., K.N.) Laboratorie de Protection et Valorisation des Ressources Végétales, Faculté des Sciences Semlalia, Université Cadi Ayyad Avenue My Abdellah BP 2390, Marrakech 40000, Maroc
The Bayoud, caused by Fusarium oxysporum f. sp. albedinis (Foa), is the most devastating disease of date palm (Phoenix dactylifera L.) in Morocco and south-west of Algeria. Since no effective treatments are currently available, biological control and the enhancement of plant defence responses provide an alternative for disease management strategy. The first part of this work consisted of eliciting defence reactions in roots with a hypo-aggressive isolate of Fusarium oxysporum (AHD). Pre-treatment of the seedlings with AHD enhanced faster enzymatic activities (PPO and POX) to reach levels as high as those obtained in response to the inoculation with the aggressive isolate ZAG in both the susceptible (JHL) and resistant (BSTN) cultivar. Inoculation with AHD also induced a rapid increase in caffeoylshikimic acids and an accumulation of non-constitutive hydroxycinnamic acid derivatives and chiefly a sinapic derivative known as I2. In the second part, four bacteria Bacillus pumilus W1, Bacillus cereus X16, Rahnella aquatilis W2 and not yet identified S1 were selected among 21 microorganisms exhibiting a high inhibition toward mycelia growth of Foa (70–77%) and its sporulation (80–95%). These antagonists in addition to Bacillus subtilis B1 and Pseudomonas sp. P1 have presented the potential in the induction and synthesis of non-constitutive hydroxycinnamic derivatives in the roots of the inoculated seedlings. The level of their accumulation varied based on the antagonist and the time of incubation. Results are discussed in the perspective of implementing a control strategy that relies on the use of biocontrol agents to trigger defence responses against bayoud.
Plant growth promotion by bacterial consortia. L. EMAD, P. BEAUREGARD AND C. BEAULIEU. Centre SÈVE, Département de biologie, Faculté des sciences, Université de Sherbrooke, Sherbrooke, QC J1K 2R1, Canada
Plant growth promoting rhizobacteria (PGPR) can be used as active ingredients of biofertilizers as they promote plant growth by different mechanisms such as auxin production. However, individual PGPR strains sometimes show inconsistent results under field conditions while bacterial consortia of PGPR have been shown to provide more consistency. The aim of this project was to establish PGPR consortia from actinobacteria and Bacillus isolates. Members of these collections were screened for auxin production. The capacity of the highest auxin-producing strains to promote plant growth was tested on Lemna minor, which was used as a model plant as it is characterized by a rapid growth in addition to its importance as animal fodder. Respectively, 73% and 11% of the selected auxin-producing actinobacteria and Bacillus isolates promoted the growth of L. minor. The compatibility between the selected strains was determined by using a double agar overlay technique. It was not possible to form bacterial consortia containing more than three strains due to antagonism between strains. A total of 16 consortia were tested and nine promoted L. minor growth. No synergy between consortia members was observed when these consortia were applied to the plant growth medium since the ability of a combination of compatible isolates to promote L. minor growth was found to be equal or lower than the ability of the single strains composing the consortia. Preliminary data indicated that the selected consortia could also promote the growth of lettuce seedlings indicating that L. minor is an interesting model to screen PGPR.
Ethylene-mediated resistance to fusarium head blight of wheat. N. A. FOROUD, R. K. GOYAL, D. RYABOVA, A. ERANTHODI, D. GONZÁLEZ-PEÑA FUNDORA, Y. PAN, R. PORDEL, I. KOVALCHUK AND S. CHATTERTON. Lethbridge Research and Development Centre, Agriculture and Agri-Food Canada, 5403-1 Avenue South, Lethbridge, AB T1J 4B1, Canada; (Y.P.) Information and Communications Technologies, National Research Council of Canada, Ottawa, ON K1A 0R6, Canada; and (I.K.) Department of Biological Sciences, University of Lethbridge, 4401 University Drive West, Lethbridge, AB T1K 3M4, Canada
Ethylene is a gaseous plant hormone involved in both plant defence and development. Often ethylene-mediated plant defence responses to necrotrophic fungi involve synergistic interactions with the jasmonate signalling pathway. On the other hand, ethylene is also an inducer of senescence and cell death, which could be beneficial for some invading necrotrophic pathogens. Fusarium graminearum is a hemibiotrophic pathogen, with both biotrophic and necrotrophic phases, that can infect the wheat inflorescence and cause a disease known as fusarium head blight (FHB). Interestingly, the role of ethylene signalling in the host response to FHB is unclear: some studies indicate that ethylene mediates resistance, while others have shown that it is associated with susceptibility. Preliminary results in our group suggested that there may be a genotype-dependent role for ethylene, explaining some of these discrepancies. To test this hypothesis, FHB experiments were carried out in six wheat genotypes with different levels of resistance or susceptibility. Detached wheat heads from each genotype were treated with ethylene inhibitors or enhancers and then inoculated with F. graminearum to assess both resistance to initial infection and disease spread. The results suggest that ethylene signalling promotes resistance, regardless of genotype: ethylene inhibition broke down resistance in three resistant wheat genotypes, whereas enhancer treatments resulted in reduced susceptibility in three susceptible genotypes. Additional work is underway to determine whether time-dependent expression of ethylene signalling or cross-talk with other hormone pathways can affect ethylene-mediated resistance to FHB.
Preliminary results of the 2017 Manitoba soybean cyst nematode survey. N. GHAVAMI, M. TENUTA AND D. LANGE. Department of Soil Science, University of Manitoba, 13 Freedman Crescent, Winnipeg, MB R3T 2N2, Canada; and (D.L.) Manitoba Department of Agriculture, Altona, MB R0G 0B0, Canada
Soybean cyst nematode (SCN), Heterodera glycines, is one of the most devastating disease/pest organisms of soybean worldwide. The nematode is expected to soon be present in Manitoba as the pest is in every major soybean growing region of the world, including Ontario and Quebec in Canada, and North Dakota and Minnesota that border Manitoba. Early detection of SCN before it establishes in Manitoba is critical to limit yield losses. Therefore, the objective of this on-going study is to survey soybean fields in Manitoba for the presence of SCN. The project continues from surveys conducted from 2012 to 2015 that did not find the nematode. In the current study, 30 commercial soybean fields in Manitoba near the USA border with history of soybean and edible bean cultivation were sampled. Each field was sectioned into areas prone for the establishment of SCN, including headlands, entrances, in-field drainage courses, depressions and remaining field. A total of 90 composite soil samples were obtained for about three samples for analysis per field. A modified Fenwick elutriator (soil washing unit) based on the USDA soil cyst extractor was used to recover nematode cysts. Cysts were extracted from debris obtained from the elutriator by using ethanol flotation. Overall, 17 of the composite samples from 12 fields had nematode cysts. One to a few cysts were recovered from each of these 17 composite samples. In total, 42 cysts were recovered and most of the cysts were round and not lemon-shaped as expected of SCN, the cysts also were intact for morphological and molecular examination. Cyst identification based on morphology and genetic structure is on-going and will be presented. We hope to address the question, ‘Is Manitoba still free of the soybean cyst nematode?’
Proteome analysis of Streptomyces scabies grown in the presence of potato tuber. L. GIROUX, I. ISAYENKA, N. BEAUDOIN AND C. BEAULIEU. Centre SÈVE, Département de biologie, Faculté des sciences, Université de Sherbrooke, Sherbrooke, QC J1K 2R1, Canada
Streptomyces scabies is the major causal agent of potato common scab. The disease is characterized by suberized lesions on potato tubers that decrease their market value. A proteomics study aiming to identify S. scabies proteins produced during potato infection was carried out. Potato microtubers from the resistant cultivar ‘Russet Burbank’ and the sensitive cultivar ‘Yukon Gold’ were produced on Murashige–Skoog medium with 8% sucrose. Streptomyces scabies was then grown in a liquid growth medium in the absence or presence of 6-week-old potato microtubers of each cultivar. After 5 days of growth, S. scabies proteins were extracted and analysed by LC-MS/MS. The similarity index of S. scabies proteome profiles between a culture grown in the absence or presence of microtubers was 48% and 54% for ‘Russet Burbank’ and ‘Yukon Gold’, respectively. The proteomes of S. scabies grown in the presence of ‘Yukon Gold’ or ‘Russet Burbank’ exhibited a higher similarity index (70%). Proteins linked to the production of the toxins concanamycins and the siderophore pyochelin were detected only in the presence of tubers. These proteins were found to be more abundant when the bacterium was grown with the resistant cultivar than with the sensitive one. The production of concanamycins and the ability to scavenge iron thus appear to participate in the infection process. Proteins involved in the SOS response and DNA repair were also found to be more abundant in the presence of microtubers than when the bacterium was grown in pure culture, suggesting a bacterial stress adaptation during host infection.
Effects of over-expressing the Mgv1 mitogen-activated protein kinase in Fusarium graminearum. D. GONZÁLEZ-PEÑA FUNDORA, A. ERANTHODI, R. K. GOYAL, R. G. SUBRAMANIAM, C. RAMPITSCH, N. THAKOR AND N. A. FOROUD. (D.G.-P.F., A.E, R.K.G., N.A.F.) Lethbridge Research and Development Centre, Agriculture and Agri-Food Canada (AAFC), Lethbridge, 5403 1st Avenue South, Lethbridge AB T1J 4B1, Canada; (D.G.F., N.T.) University of Lethbridge, Department of Chemistry and Biochemistry, 4401 University Drive West, Lethbridge, AB T1K 6T5, Canada; (A.E.) University of Lethbridge, Department of Biological Sciences, 4401 University Drive West, Lethbridge, AB T1K 6T5, Canada; (R.G.S.) Ottawa Research and Development Centre, AAFC, 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada; and (C.R.) Morden Research and Development Centre, AAFC, 101 Route 100, Morden, MB R6M 1Y5, Canada
Mitogen-activated protein kinase (MAPK) cascades are involved in signal transduction processes in the cell, through sequential phosphorylation. MAPK kinase kinases phosphorylate MAPK kinases, which in turn phosphorylate MAPKs. In Fusarium graminearum, one of the causal agents of fusarium head blight disease of cereals, three MAPK pathways have been identified. Among these, the Mgv1 (MAP kinase for growth and virulence 1) pathway is part of the Bck1-Mkk2-Mgv1 kinase cascade, and previous reports have shown that deletion of MGV1 affects female fertility, mycelial growth, mycotoxin accumulation, and virulence (possibly due to reduced fitness). Despite the significance of research focused on these pathways, little is known about the downstream components of the Mgv1 cascade. With the aim of identifying Mgv1 targets, F. graminearum strains over-expressing MGV1 under a constitutive promoter were generated by Agrobacterium-mediated transformation of the wild-type (WT) strain, GZ3639. The colony pattern and mycelial growth of the transformants on potato dextrose agar were similar to the WT, even in the presence of chemicals affecting cell wall integrity, such as congo red and calcofluor white. Quantitative RT-PCR analysis showed a higher relative gene expression for MGV1 in the transformants compared to WT, but not for RLM1, a known downstream target of Mgv1. The down-regulation of TRI5, a key gene involved in trichothecene biosynthesis, may explain the low levels of the mycotoxin deoxynivalenol detected in the transformant compared with the WT. The active form of Mgv1 will be used in future proteomics studies to identify and further characterize the downstream signalling pathway.
The elusive Puccinia tritici-duri – pathology, taxonomy and relationship to Puccinia recondita. S. HAMBLETON AND M. LIU. Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada
Common usage of the name Puccinia triticina Erikss. specifically for the fungus causing leaf rust of wheat is relatively recent in its taxonomic history. Described in 1899, subsequent names were as a forma specialis, variety or subspecies of P. dispersa Erikss. & Henning, P. rubigo-vera (DC.) G. Winter, P. perplexans Plowr. or P. persistens Plowr., until it was included as one of 52 synonyms in the P. recondita Roberge ex Desm. complex in 1971. By the 1980s in North America, the concept of a species restricted to wheat as its primary host was clearly articulated and accepted, with alternate hosts in the Ranunculaceae and clear morphological differences of the spores and sori. Another leaf rust species on wheat, Puccinia tritici-duri, was described by Viennot-Bourgin in 1941. It infects durum wheat but is thought to be restricted to Morocco, Spain and Portugal and to alternate hosts in the Boraginaceae. Tracking down information about the species was not straightforward. The name was not listed online in the fungal nomenclature databases, the original description was published in a difficult-to-locate journal, and references to this species were not always by name but rather implied. A review of published pathogenicity studies and phylogenetic analyses of brown leaf rust fungi suggests that Puccinia tritici-duri is closely related to Puccinia recondita sensu stricto and as such is one member of a lineage restricted to alternate hosts in the Boraginaceae. Analyses of DNA data for authentic specimens are needed to test this hypothesis and anchor the name taxonomically.
Evans blue staining is a rapid and accurate method for evaluating inactivation of Plasmodiophora brassicae resting spores by chemical disinfectants. M. W. HARDING, T. B. HILL, G. C. DANIELS, S. E. STRELKOV, S. F. HWANG AND J. FENG. Alberta Agriculture and Forestry, Crop Diversification Centre South, 301 Horticultural Station Road East, Brooks, AB T1R 1E6, Canada; (S.E.) University of Alberta, Agriculture Food and Nutritional Sciences, 410 Agriculture-Forestry Centre, Edmonton, AB T6G 2P5, Canada; and (S.F.H., J.F.) Alberta Agriculture and Forestry, Crop Diversification Centre North, 17 507 Fort Road NW, Edmonton, AB T5Y 6H3, Canada
Clubroot is an important disease on canola caused by the protist Plasmodiophora brassicae Woronin. Over the past 15 years, it has spread rapidly across much of central and northern Alberta and was more recently confirmed in Saskatchewan and Manitoba. It is widely accepted that the rapid spread in Alberta has been due in large part to movement of infested soil on farm or construction equipment. Equipment sanitization is the recommended practice for preventing unintentional spread of infested soil to new fields. The thick-walled resting spores produced by P. brassicae are very capable of surviving harsh physical and chemical treatments, therefore it is important to know which disinfectants, if any, can quickly and effectively inactivate resting spores. Evans blue, a vital stain that can discriminate viable from non-viable resting spores, was used to evaluate 10 chemical disinfectants for their efficacies versus resting spores. Repeated experiments, comparing bioassay results with spore staining, indicated that Evans blue staining was a reliable and rapid method to evaluate efficacies of most disinfectants tested. Only two of the disinfectants tested, sodium hypochlorite and ethanol, were capable of achieving greater than 95% inactivation of resting spores.
Population dynamics of fungal pathogens on wheat heads in Alberta in 2015 and 2016. M. W. HARDING, G. C. DANIELS, T. GRÄFENHAN, T. K. TURKINGTON AND J. FENG. Alberta Agriculture and Forestry, Crop Diversification Centre South, 301 Horticultural Station Road East, Brooks, AB T1R 1E6, Canada; (T.G.) Canadian Grain Commission, Grain Research Laboratory, 196 Innovation Drive, Richardson Centre, Winnipeg, MB R3T 6C5, Canada; (T.K.T.) Lacombe Research and Development Centre, Agriculture and Agri-Food Canada, 6000 C and E Trail, Lacombe, AB T4L 1W1, Canada; and (J.F.) Alberta Agriculture and Forestry, Crop Diversification Centre North, 17 507 Fort Road NW, Edmonton, AB T5Y 6H3, Canada
Fusarium head blight, caused by Gibberella zea (Schwein.) Petch (syn. Fusarium graminearum), is a serious disease in cereals that reduces yield, grade and market acceptance. A comprehensive survey for the prevalence, incidence and distribution of wheat head pathogens was conducted in Alberta in 2015 and 2016. Over 800 wheat fields were sampled each year and grain analysed for the presence of head blight pathogens such as F. graminearum, and other fungi commonly associated with cereal spikes. Dry conditions were reported in 2015 for much of Alberta during the heading and flowering stages of cereal crops, whereas 2016 was relatively wet during these stages for much of the province. The environmental conditions, along with other factors, caused shifts in fungal populations on wheat heads. For example, the incidences of Fusarium spp. and Phaeosphaeria nodorum (Mull.) Hedjar., were significantly higher in 2016 when compared with those of 2015. Interestingly, the incidences of Alternaria spp. were significantly lower in 2016 compared with 2015 and those of Pyrenophora spp. were relatively unaffected.
Integrated genomics, plant pathology and breeding research for improvement of fusarium head blight, leaf rust and stripe rust resistance in Canadian cereals. M. A. HENRIQUEZ, G. HUMPHREYS, B. D. MCCALLUM, H. S. RANDHAWA, T. FETCH, J. M. FETCH, X. WANG, M. F. BELMONTE, C. A. MCCARTNEY AND M. KANG-CHOI. Morden Research and Development Centre, Agriculture and Agri-Food Canada (AAFC), Unit 101 Route 100, Morden, MB R6M 1Y5, Canada; (G.H., M.K.-C.) Ottawa Research and Development Centre, AAFC, 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada; (H.S.R.) Lethbridge Research and Development Centre, AAFC, 5403–1st Avenue South, P.O. Box 3000, Lethbridge, AB T1J 4B1, Canada; (J.M.F.) Brandon Research and Development Centre, AAFC, 2701 Grand Valley Road, Brandon, MB R7A 5Y3, Canada; and (M.F.B.) Department of Biological Sciences, University of Manitoba, 50 Sifton Road, Winnipeg, MB R3T 2N2, Canada
Development of cereal crop varieties with resistance to fusarium head blight (FHB) (Fusarium spp.) and rust (Puccinia spp.) is a main breeding goal worldwide. This project’s objectives are: (1) development and genotyping of a winter wheat germplasm collection, (2) development of a doubled haploid mapping population from the winter wheat cross: AC Morley × Emerson, (3) phenotyping of the DH population for its response to FHB, (4) genotype the AC Morley × Emerson doubled haploid population to provide SNPs to generate a molecular map of the population, (5) QTL mapping of FHB resistance, (6) identify and characterize Fusarium spp. from FHB-infected winter wheat crops, (7) transcriptome analyses of winter wheat–F. graminearum interaction, (8) validation of differentially expressed genes, (9) development/validation of SNP markers diagnostic for FHB resistance of winter wheat, (10) development of FHB, leaf rust and stripe rust resistant germplasm, and linked molecular markers, that can be used by wheat breeding programmes, and (11) analysis of Fusarium spp. complex in oat and screening of oat germplasm for resistance to major Fusarium spp. found in western Canada. This knowledge will be transferred to breeding programmes for marker-assisted selection and will be an important resource for breeding new elite disease-resistant wheat and oat cultivars for Canada.
Selection of antagonistic bacteria from pea root and rhizosphere to manage aphanomyces root rot. Z. HOSSAIN, L. D. BAINARD AND Y. GAN. Swift Current Research and Development Centre, Agriculture and Agri-Food Canada, 1 Airport Road East, Swift Current, SK S9H 3X2, Canada
Aphanomyces root rot caused by Aphanomyces euteiches Drechs is a serious disease of pulse crops worldwide and is becoming a major constraint to pea (Pisum sativum L.) and lentil (Lens culinaris Medik) production in Canada. Seed treatments are not effective, resistance is limited and use of crop rotation to minimize the disease pressure has a limited scope. The current project was designed to identify endemic bacteria that were antagonistic toward A. euteiches. Rhizosphere and root samples were collected from diseased and healthy pea plants from nine locations across southern Saskatchewan. Approximately 6000 rhizosphere and endophytic bacterial colonies were isolated on various media (Luria Bertani, potato dextrose, Pseudomonas and tryptic soy agar). Bioassays were conducted with 410 selected bacteria in vitro to evaluate their antagonistic potential toward A. euteiches. Thirty-five isolates were identified based on their ability to completely inhibit growth of A. euteiches. These isolates were then tested in a replicated greenhouse study and nine were selected for field evaluations. Each of these selected antagonistic bacteria suppressed the pathogen, such that treated plants were as healthy as the control plants. Also, a gene expression study was conducted to investigate the disease mechanism and formulate a disease management strategy. Initial results showed stronger expression of several pathogenesis-related genes in pathogen-inoculated plant leaves compared with the healthy control. A similar study on lentil is in progress. Management of Aphanomyces root rot using antagonistic bacteria may provide a disease control strategy ensuring a sustainable pulse production system on the Canadian prairie.
First report of clubroot (Plasmodiophora brassicae) on canola in the Peace Region, Alberta. S. F. HWANG, H. U. AHMED, Q. X. ZHOU, V. P. MANOLII, G. D. TURNBULL, R. FREDUA-AGYEMAN, S. KAUS AND S. E. STRELKOV. Crop Diversification Centre North, Alberta Agriculture and Forestry, 17507 Fort Road NW, Edmonton, AB T5Y 6H3, Canada; (V.P.M., S.E.S.) Department of Agricultural, Food and Nutritional Sciences, 410 Agriculture/Forestry Centre, University of Alberta, Edmonton, AB T6G 2P5, Canada; and (S.K.) Big Lakes County, 5305–56 Street, High Prairie, AB T0G 1E0, Canada
Clubroot of canola (Brassica napus L.), caused by Plasmodiophora brassicae Woronin, has spread to more than 2700 fields in Alberta since it was first identified in 2003. The disease, however, had not been reported in the Peace Region of north-west Alberta until 2017, when 46 cases of clubroot were identified in the municipal district of Big Lakes. Surveys of the infested crops found a clubroot incidence of < 10% in 40 fields and > 10% in the other six. Individual plants were rated for clubroot severity on a 0–3 scale. In most (32) fields, clubroot symptoms were mild, with all affected plants rated as a 1, although severely infected plants (with a rating of 3) also were found in 10 fields. Rotation histories were obtained for 20 of the affected fields, which indicated that canola had been grown either back-to-back or in alternating years in at least 63% of the cases. A PCR-based quantitative analysis with primers developed for pathotype 5X, which is able to overcome clubroot resistance in canola, found it (or similar strains) was present in 88% of the fields tested, but at a very low frequency (0.5%) relative to the old pathotype 3H. Phenotypic evaluation for pathotype classification on the Canadian Clubroot Differential (CCD) Set revealed that of 17 P. brassicae populations tested from Big Lakes, 11 were pathotype 3H, two each were pathotype 8N and pathotype 5I, and two appeared to have novel virulence patterns which require further investigation. The identification of pathotype 5X at a very low frequency suggests the potential for shifts in the virulence of the P. brassicae populations in Big Lakes, particularly if clubroot resistant canola is grown in short rotations in the region.
Evaluating the potential of arbuscular mycorrhizae to manage carrot leaf blight. U. ILYAS, M. N. RAIZADA, L. DU TOIT AND M. R. MCDONALD. Department of Plant Agriculture, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada; and (L.T.) Department of Plant Pathology, University of Washington State, Washington, WA 98273–4768, USA
The symbiosis of plants and arbuscular mycorrhizae (AM) is known to increase nutrient uptake and improve resistance to biotic and abiotic stresses. Commercial AM products are available and carrot growers are using these products. However, the ability of AM to suppress disease or increase yield in carrots in Ontario is currently unknown. The present study evaluated the efficacy of AM inoculant Glomus intraradices, on carrot seeds, to manage carrot leaf blight. Leaf blight is caused by one or both fungi Alternaria dauci Kühn and Cercospora carotae Pass. Field trials were conducted on high organic matter soils at two sites, one with low phosphorus (P) content (46 ppm) and one with high P content (68 ppm) near Bradford, ON in 2017. Mono-ammonium phosphate was applied at 100 kg actual P/ha to high P soils only. Fungicide was applied biweekly in August and September. Disease severity was assessed throughout the growing season and canopy health and yield was determined at harvest. There were no differences in disease progress among the treatments during the growing season. At harvest, the treatment with AM applied to low P soils had less disease (25%), more healthy leaves per plant (22%), and higher canopy fresh (46%) and dry weight (55%) compared with the no AM check. However, there were no differences in yield or the percentage of AM colonization in roots of AM treated and untreated carrots. Results suggest that AM association might be beneficial in low P soils.
Identification of non-host crops for the management of stem and bulb nematode (Ditylenchus dipsaci) in garlic (Allium sativum). L. IVES, M. J. CELETTI, K. JORDAN AND M. R. MCDONALD. Department of Plant Agriculture, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada; and (M.J.C.) Ontario Ministry of Agriculture, Food and Rural Affairs, University of Guelph, Guelph, ON N1G 2W1, Canada
Stem and bulb nematode (SBN), Ditylenchus dipsaci (Kuhn) Filip’ev, is problematic for garlic (Allium sativum L.) growers in Ontario, where this nematode reduces yield and infests seed cloves for successive planting. There are no garlic cultivars with resistance to SBN, and no nematicides are registered for use on garlic seed cloves. Rotating garlic with non-host crops could reduce the soil population between cropping seasons. However, the susceptibility of rotation crops to the SBN strain affecting Ontario garlic is unknown. In growth room trials, six potential rotation crops: carrot, corn, potato, onion, soybean and wheat were evaluated for their response to infection by SBN. The experiment contained a positive (susceptible garlic cv. ‘Music’) and negative (inoculated soil without vegetation) control. At planting, each pot was inoculated using 100 nematodes of mixed developmental stages. Plants were harvested 6 weeks post-inoculation, and nematodes were recovered from soil and plant tissue using the Baermann funnel method. Nematode reproduction factor (RF) was calculated to identify non-host crops. Soil RF values suggest that potato (RF = 7.3), carrot (RF = 6.1), onion (RF = 4.3) and corn (RF = 3.5) were hosts. However, based on plant tissue extraction, onion appeared to be a very susceptible host (RF = 23.8) compared with corn (RF = 1.1), garlic (RF = 1.0) and potato (RF = 0.6). Soybean and wheat were non-hosts, since no nematodes were recovered from plant tissue, and fewer nematodes were recovered from soil than were used for inoculation. Wheat and soybean may be good choices of crops to rotate with garlic to reduce SBN population levels in the soil.
Chocolate spot disease risk periods in faba beans in Alberta and Saskatchewan. S. KAUR, R. BOWNESS, S. BANNIZA AND S. CHATTERTON. Lethbridge Research and Development Centre, Agriculture and Agri-Food Canada (AAFC), 5403–1 Avenue South, P.O. Box 3000, AB T1J 4P4, Canada; (R.B.) Lacombe Research and Development Centre, AAFC, 6000 C and E Trail, Lacombe, AB T4L 1W1, Canada; and (S.B.) Crop Development Centre, University of Saskatchewan, 51 Campus Drive, Saskatoon, SK S7N 5A8, Canada
Chocolate spot (CS), caused by Botrytis fabae Sard., is one of the most important diseases of faba bean (Vicia faba L.) affecting its productivity in Alberta and Saskatchewan. Cool, wet and humid weather conditions favour sporulation and secondary infection, but information of CS disease risk periods and inoculum release under prairie conditions is lacking. Therefore, this study was conducted to determine the duration of inoculum discharge and infectious periods under field conditions by using 3-week-old faba bean plants (cv. ‘Malik’ (tannin) and ‘Snowdrop’ (zero tannin)) as spore traps. A total of 16 trap periods from mid-June to mid-August were assessed in 2017. Five plants of each cultivar were placed within the canopy for a period of 4 days at each trap period at locations in Lethbridge and Lacombe, AB and Saskatoon, Melfort and Scott, SK. After the exposure period, plants were incubated in the greenhouse and CS severity was rated after 14 days. Data analysis showed significant effects of location and trap period on CS severity. However, the effect of cultivar on CS severity was not always significant. Infectious periods occurred earlier than expected (mid-June) at the Lethbridge sites, but was later (late July–August) at all other locations in AB and SK. High temperatures were negatively correlated with disease severity across all locations, whereas the significance of correlations between severity and precipitation or humidity were not consistent across locations. Experiments will be conducted for two more cropping seasons to assess the disease risk period, and model infectious periods to weather parameters.
Contrasting aboveground and belowground responses to PAMP-triggered immunity. A. LACAZE, A. CULL AND D. L. JOLY. Université de Moncton, 18 avenue Antonine-Maillet, Moncton, NB E1A 3E9, Canada
Deciphering the molecular responses to pathogen-associated molecular patterns (PAMPs) is an active research area in the field of plant–microbe interactions. Much of the progress that has been made in puzzling out PAMP-triggered immunity (PTI) pathways and the microbial factors involved in the elicitation or suppression of PTI has relied solely on aerial parts to generate insights, despite the organ-specificity of plant defences. In this study, we focus on one oomycete PAMP, Pep-13/Pep-25, and contrast it to the well-characterized bacterial PAMP flg22 in aerial and below-ground organs of Solanum tuberosum L. and Arabidopsis thaliana (L.) Heynh. By systematic analyses of defence responses such as ROS burst, transcript changes or accumulated hormones, we found that flg22 triggers fast and strong responses in the different organs tested, while Pep-13/Pep-25 treatments elicit organ-specific responses. Using measurement of PAMP-induced ROS generation, we screened collections of A. thaliana T-DNA insertional mutants for those suppressed for Pep-25 responsiveness, including the regulatory leucine-rich repeat-receptor-like kinases BAK1 and SOBIR1. This study will improve our global understanding of PAMP-triggered immunity and allow the discovery of novel components of the plant immune system.
Gene flow as a character in polyphasic classification of phytopathogenic fungi: a case study in wheat leaf rust species complex. M. LIU, S. HAMBLETON, Y. ANIKSTER AND J. KOLMER. Biodiversity and Bioresources, Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada; (Y.A.) Institute for Cereal Crops Improvement, George S. Wise Faculty of Life Science, Tel Aviv University, Tel Aviv 69978, Israel; and (J.K.) U. S. Department of Agriculture, Cereal Disease Laboratory, 1551 Lindig Street, St. Paul, MN 5508, USA
The combination of phylogenetics and molecular technologies has dramatically increased the resolution of fungal classification over the past 30 years. Numerous species defined by morphology turn out to be complexes. Along with the achievements of more refined species recognition, a commonly encountered dilemma and debate is whether paraphyletic species should be recognized. Topology based species recognition considers that species must be supported by evidence of monophyly. A study of four host-associated forms of P. triticina Erikss. by Liu et al. 2013 showed that tree topologies varied when different loci and analytical methods were used. The combined ITS and EF1-α analyses grouped all four forms as one clade whereas phylogenetic analysis of 239 SNPs from 15 loci resolved two strongly supported monophyletic groups and coalescence-based analysis (BEAST) resolved three monophyletic groups. In this case, one, two or three species could be recognized based on various tree topologies. The form on Aegilops speltoides Tausch was previously recognized as a forma specialis based on infection experiments by Anikster et al. 2005, which showed that A. speltoides was resistant to P. triticina isolated from wheat. Gene flow analysis (IMa2) indicated that there was no detectable gene flow between the Aegilops form and any other forms while constant gene flow was detected among other forms. We consider the lack of gene flow as strong evidence of genetic separation and propose to recognize the Aegilops form as a separate species: Puccinia speltoides sp. nov., while other forms belong to one species.
Integrated management of grey mould on greenhouse tomato: efficacy of biological control. O. CARISSE, T. ARSENEAULT AND G. MARCHAND. Saint-Jean-sur-Richelieu Research and Development Centre, Agriculture and Agri-Food Canada (AAFC), 430 Gouin Boulevard, Saint-Jean-sur-Richelieu, QC J3B 3E6, Canada; and (G.M.) Harrow Research and Development Centre, AAFC, 2585 County Road 20, Harrow, ON N0R 1G0, Canada
Table tomato (Lycopersicon esculentum Mill.) is the leading greenhouse vegetable crop in Canada. Grey mould (Botrytis cinerea) is endemic and can result in high yield losses due to stem wound infections, which may cause the death of an entire plant. In greenhouse tomato production, de-leafing of plants is necessary to manage vegetative/reproductive balance. However, this practice can result in B. cinerea spores being shaken from lesions, becoming airborne, and settling onto the fresh stem wounds. Considering the high level of fungicide resistance in B. cinerea populations, biological control should play a key role in integrated grey mould management. The objective of this study was to evaluate the efficacy of Botector, a bio-fungicide containing two strains of Aureobasidium pullulans (de Bary) Arnaud, a yeast-like fungus. Results from this study from competition experiments confirmed the ability of Botector to establish itself in stem lesions in the presence or absence of B. cinerea. Efficacy of Botector was significantly higher when it was applied prior to B. cinerea inoculation, and efficacy increased with increasing time between sprays of Botector and B. cinerea with 87.6%, 92.4% and 99.0% reduction of symptoms for intervals of 1, 6 and 24 h, respectively. However, when Botector was applied after inoculation with B. cinerea, efficacy was reduced and decreased with increasing time between sprays of Botector and B. cinerea, with efficacies of 75.3%, 69.5% and 29.8% for intervals of 1, 6 and 24 h, respectively. Results from these experiments suggest that Botector should be used as a preventative, pre-infection treatment.
Genetic mapping of leaf rust resistance genes LrCen and LrMar. B. McCALLUM, M. Z. CHE, M. BOYCE, C. W. HIEBERT, A. L. BRULE-BABEL, C. A. McCARTNEY AND Z. J. ZHANG. Morden Research and Development Centre, Agriculture and Agri-Food Canada, 101 Route 100, Unit 100, Morden, MB R6M 1Y5, Canada; (M.Z.C., Z.J.Z.) Department of Plant Pathology, China Agricultural University, Beijing, 100193, People’s Republic of China; (M.B.) Global Edible Oil Solutions – Specialties, Cargill Ltd. Canada, 701 Central Avenue, Aberdeen, SK S0K 0A0, Canada; and (A.L.B.-B.) Department of Plant Science, 222 Agriculture Building, 66 Dafoe Road, University of Manitoba, Winnipeg, MB R3T 2N2, Canada
A group of Puccinia triticina Eriks. isolates with similar virulence phenotypes, represented by TDBG, predominated in the annual Canadian virulence surveys from 2004 to the present. These had an unusual mesothetic infection type on most plants within the Thatcher-Lr1 line, RL6003, which was due to a previously undetected second resistance gene in this line, temporarily named LrCen. LrCen was mapped to chromosome arm 7AL, 1.1 cM from the closest marker, cfa2240. RL6071, a rust susceptible Marquis derivative, was also found to carry a phenotypically similar resistance gene effective only against this same group of virulence phenotypes. However, this gene from RL6071 was mapped on 7BL at 1.8 cM from marker barc182 in a doubled haploid population from the cross RL6071/KU168-2, and was temporarily named LrMar. These genes are widely distributed in Canadian wheat, are effective against the same small number of virulence phenotypes, and map to similar locations on homologous chromosomes.
Opportunities for technology, new and old, to reduce, improve or replace pesticides. M. R. MCDONALD AND B. D. GOSSEN. Department of Plant Agriculture, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada; and (B.D.G.) Saskatoon Research and Development Centre, Agriculture and Agri-Food Canada, 107 Science Place, Saskatoon, SK S7N 0X2, Canada
A consistent shift to larger farms, bigger equipment and reduced crop diversity has been occurring in Canadian agriculture from many years. As a result, the natural biological control provided by (i) crop rotation and (ii) biological diversity from wetlands, headlands, fencerows and windbreaks has been replaced by increased reliance on genetic resistance and pesticides. This has often resulted in rapid erosion of cultivar resistance and loss of sensitivity to single-site pesticides. The increasing expense of finding new genetic resistance and pesticides indicates that this approach is not sustainable in the long term. Fortunately, several new technologies may help to change this balance. Gene editing (e.g. using CRISPR/Cas 9) will provide a source of novel resistance, and marker-assisted selection will facilitate selection for complex traits. RNA interference (gene silencing) may provide effective reductions in some pests and pathogens. A shift to autonomous field equipment has the potential to reverse the trend for larger farm equipment. Smaller autonomous equipment could seed and harvest small areas, making intercropping, deployment of multi-lines, and variable plant spacing easier and cost-effective. Remote sensing of the host, microenvironment and individual pests, combined with autonomous, targeted application of pesticides, could improve the efficacy of both biocontrol agents and synthetic pesticides in these small crop areas. Robots may also remove or treat individual weeds, insects or infected plants. These changes could shift the balance back towards natural biological control and biological diversity, and so reduce the need for large-scale pesticide application.
Physiological races of Puccinia coronata var. avenae f. sp. avenae in Canada during 2010 to 2017. J. MENZIES, A. XUE AND C. AZAR. Morden Research and Development Centre, Agriculture and Agri-Food Canada (AAFC), Unit 101 Route 100, Morden, MB R6M 1Y5, Canada; (A.X.) Ottawa Research and Development Centre, AAFC, 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada; and (C.A.) Crop Production Research Farm, La Coop fédérée, Saint-Hyacinthe, QC J2T 5J4, Canada
Crown rust of oat, caused by Puccinia coronata Corda var. avenae f. sp. avenae (Urban & Marková) (Pca), is a problematic disease of oat. Breeding for resistant varieties has been an important method of controlling this disease. Since Pca is highly genetically variable, a thorough knowledge of the races in the pathogen population is necessary to identify effective genes for resistance for breeding and gene deployment. This study’s objective was to determine the frequency of virulence genes present in Pca isolates collected from oat plants in Canada during 2010 to 2017. Single pustule isolates from infected leaves were inoculated onto a standard set of 16 differential isolines (Pc28, Pc39, Pc40, Pc45, Pc46, Pc48, Pc50, Pc51, Pc52, Pc54, Pc56, Pc58, Pc59, Pc62, Pc64, Pc68) and eight additional isolines (Pc91, Pc94, Pc96, Pc97, Pc98, Pc101, Pc103-1, Pc104) at the seedling stage. Virulence frequencies varied between eastern Canada (Ontario and Quebec) and western Canada (Manitoba and Saskatchewan). Virulence to Pc38 and Pc68 was observed at frequencies of > 50% in Canada. Pc94 was the most effective resistance gene in western Canada, with virulence frequencies to Pc94 of < 2%. Virulence to Pc98 and Pc101 was not observed in eastern Canada. The most dramatic increase in virulence was to Pc91 in western Canada with frequencies that increased from 0% in 2010 and 2011 to 80% in 2016.
Soybean cyst nematode in Canada: genomics applied to biovigilance. B. MIMEE. Saint-Jean-sur-Richelieu Research and Development Centre, Agriculture and Agri-Food Canada, 430 Gouin Boulevard, Saint-Jean-sur-Richelieu, QC J3B 3E6, Canada
Soybean production has had a spectacular growth over the last 5 years in Canada. One of the main soybean pests, Heterodera glycines, responsible for over a billion-dollar yield loss in the USA annually, is already well established in south-western Ontario. Is this microscopic worm a real threat to soybean production in northern latitudes? Climate warming will promote the expansion of soybean to northern areas but will also increase the number of generations the nematode can produce in a single year, thus speeding up its establishment and spread in new areas. Even if the nematode was found in all regions of Quebec, population densities are still very low and it is anticipated that yield-limiting populations will not be reached for several years. However, the adaptive potential of this alien invasive species to new environments was found to be very high. Winter, drought or high precipitations will not stop its spread. Worst, it appears that strong selective pressure resulting from the overuse of resistant cultivars in the Midwest is causing a large-scale homogenization with virulent populations. Thus, these cultivars will become increasingly ineffective more rapidly in new areas than before. It is therefore critical to include new promising parental lines with new resistance genes in breeding programmes. Even more important is the comprehension of the mechanisms used by the nematode to overcome plant resistance. To this end, recent advances in SCN genomics suggest that gene duplication and transposition could be responsible for the rapid adaptation and evolution of the pest.
Assessing incidence of eastern filbert blight in Ontario hazelnuts. A. MUNAWAR, C. BAKKER, M. FILOTAS AND K. S. JORDAN. Department of Plant Agriculture, Simcoe Research Station, University of Guelph, 1283 Blueline Road, Simcoe, ON N3Y 4N5, Canada; and (M.F.) Ontario Ministry of Agriculture, Food and Rural Affairs, 1283 Blueline Road, Simcoe, ON N3Y 4N5, Canada
Eastern filbert blight (EFB), caused by the fungus Anisogramma anomala (Peck) Müller, is the main disease affecting hazelnut trees in eastern North America. This pathogen is an obligate, biotrophic parasite and is known to attack only species of the genus Corylus L. To support increased production of this crop in Ontario, there is a need to better understand the overall incidence and severity of this disease in commercial orchards in the province, and how that varies among cultivars, ages of orchards and management practices. It is also important to determine the incidence of latent infection in asymptomatic trees as symptom development usually takes place 1–1.5 years after the initial infection. Six hazelnut orchards in Ontario were scouted for visible symptoms of EFB infection from February–March 2018. The incidence of EFB varied from orchard to orchard and ranged from less than 1% to 12%. The young trees in orchard 1–3 had severe cankers although the number of symptomatic trees was small. The cultivars ‘Yamhill’ and ‘Jefferson’ were also found infected although these cultivars are reported to be resistant in Oregon. The latent infection was assessed in the twigs of 25 asymptomatic trees from orchards 1–4 using polymerase chain reaction method. Orchard 1 had the highest percentage of latent infection (64%) followed by orchard 3 (44%) and orchard 4 (20%). Orchard 2 had the lowest infection (4%). The trees that tested positive for latent infection have been marked and will be monitored for symptom development in autumn 2018.
Assessment of transmission pathways for the presence of the cucumber downy mildew pathogen, Pseudoperonospora cubensis. A. MUNAWAR, C. BAKKER, C. MCCREARY AND K. S. JORDAN. Department of Plant Agriculture, Simcoe Research Station, University of Guelph, 1283 Blueline Road, Simcoe, ON N3Y 4N5, Canada; and (C.M.) Ontario Ministry of Agriculture, Food and Rural Affairs, Harrow Research Station, 2585 County Road 20, Harrow, ON N0R 1G0, Canada
Pseudoperonospora cubensis (Berkeley & Curtis) Rostovtsev, is responsible for one of the most important foliar diseases of cucumber, namely downy mildew (DM). Symptoms include yellow angular lesions on the leaf surface and production of sporangia on the underside of the leaf. Control of DM depends on an intensive fungicide programme which is not always feasible due to high cost of fungicides and evolution of resistant pathogen populations. The current project is focused on investigating cucumber seeds and fruit and alternative DM hosts as inoculum sources in a Canadian environment. In year 1–2 of the study, 295 fruit collected from infected cucumber fields were tested microscopically and none were found infected. Approximately 3000 cucumber seeds were grown for 4 weeks in a greenhouse under ideal conditions for DM symptom development. Leaves of 108 plants developed yellow lesions similar to DM but did not show sporulation. However, a polymerase chain reaction (PCR) showed all tested symptomatic leaf samples were positive for ribosomal internal transcribed spacer DNA of Pseudoperonospora. PCR analysis on individual seeds, harvested from fruit of severely infected cucumber plants showed 96% of them positive for Pseudoperonospora. Four cucurbits namely, golden-creeper, bitter-melon, wild and bur cucumber were tested as alternative hosts using a detached leaf inoculation method. All wild cucurbits developed DM symptoms 3–5 days after inoculation and showed sporangial production. The last year of this study will generate data to confirm cucumber fruit and seeds, and alternative hosts as possible inoculum sources for DM and provide recommendations for disease management.
Incidence and management of hop downy mildew in Ontario in 2016 and 2017. A. MUNAWAR, M. FILOTAS, C. BAKKER, M. R. MCDONALD AND K. S. JORDAN. Department of Plant Agriculture, Simcoe Research Station, University of Guelph, 1283 Blueline Road, Simcoe, ON N3Y 4N5, Canada; (M.R.M.) Department of Plant Agriculture, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada; and (M.F.) Ontario Ministry of Agriculture, Food and Rural Affairs, 1283 Blueline Road, Simcoe, ON N3Y 4N5, Canada
Hop downy mildew (HDM), caused by Pseudoperonospora humuli, is one of the most devastating diseases of hops. The pathogen can overwinter in dormant buds or rhizomes causing a persistent systemic infection and subsequent infection of bines and cones. Due to the systemic nature of the infection, applying fungicides with systemic activity in the plant is an important management tool. The only systemic fungicide registered for hops in Canada is metalaxyl but resistance of P. humuli against this fungicide is unknown in Ontario. This project is designed to determine the incidence of HDM in Ontario commercial hop yards and to evaluate the resistance of the pathogen to metalaxyl. In 2016 and 2017, the incidence of HDM varied among cultivars and was greatly affected by weather conditions. Eighty-four rhizomes were screened for systemic presence of P. humuli through polymerase chain reaction (PCR) using primers designed for ribosomal internal transcribed spacer DNA of the fungus. Approximately 30% of the 84 rhizomes showed systemic infection in PCR and percentage of infection varied from yard to yard. In preliminary work on resistant populations of P. humuli, eight downy mildew infected spikes from a conventional hop yard and one spike from an organic hop yard were tested for metalaxyl sensitivity. All spikes from the conventional yard were found resistant to metalaxyl at 50 and 100 ug mL−1, whereas the spike from the organic yard was sensitive to the fungicide at both tested concentrations. Future work will focus on testing P. humuli populations from more hop yards.
Practical solutions for managing clubroot (Plasmodiophora brassicae) on canola in western Canada. D. ORCHARD, B. D. GOSSEN, M. R. MACDONALD AND S. E. STRELKOV. Canola Council of Canada, 400–167 Lombard Avenue, Winnipeg, MB R3B 0T6, Canada; (B.D.G.) Saskatoon Research and Development Centre, Agriculture and Agri-Food Canada, 107 Science Place, Saskatoon, SK S7N 0X2, Canada; (M.R.M.) Department of Plant Agriculture, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada; and (S.E.S.) Department of Agricultural, Food and Nutritional Science, 410 Agriculture/Forestry Centre, University of Alberta, Edmonton, AB T6G 2P5, Canada
Since it was first identified on canola (Brassica napus L.) in Alberta in 2003, the spread of clubroot caused by Plasmodiophora brassicae Woronin has been relentless, with about 3000 infested fields confirmed as of 2017. Genetic resistance has been the main approach used for managing clubroot since the commercial release of resistant cultivars in 2009, but numerous pathotypes able to overcome the existing resistance have recently been identified at sites across the region. In the long term, management of clubroot will require adoption of management practices beyond simple major-gene resistance. Research indicates that strategies such as raising soil pH with lime, mapping infestations and recognizing hot-spots, rotating to non-host crops, seeding a densely rooted grass crop, knocking soil off of equipment, and other tactics can also be employed to reduce the spread of clubroot and the concentration of resting spores in soil. Deploying the results of this research in a practical combination with genetic resistance on fields and farms should result in more effective clubroot management and slower spread of the pathogen compared with reliance on major gene resistance alone. Potentially effective strategies include: (i) grassing and/or liming field entrances, hot-spots and low or wet areas, (ii) reducing tillage, (iii) minimizing field traffic, (iv) creating new exits far removed from entrances and (v) dedicating a sanitation zone to reduce the spread of resting spores. These approaches are being recommended in current extension efforts, and several options are also being assessed in replicated research studies.
Transcriptomic analysis of Brassica napus–Leptosphaeria maculans pathosystem by dual RNA-seq using a single-R-gene cultivar and a single-Avr-gene isolate. K. R. E. PADMATHILAKE, H. SONAH, Z. ZOU, J. R. TUCKER, A. CARTER, R. R. BELANGER AND W. G. D. FERNANDO. Department of Plant Science, 222 Agriculture Building, 66 Dafoe Road, University of Manitoba, Winnipeg, MB R3T 2N2, Canada; (H.S., R.R.B.) Department of Plant Science, Laval University, Québec, QC G1V 0A6, Canada; and (A.C.) Brandon Research and Development Centre, Agriculture and Agri-Food Canada, 2701 Grand Valley Road, Brandon, MB R7A 5Y3, Canada
Blackleg disease caused by Leptosphaeria maculans remains a significant threat to canola (Brassica napus) cultivation. Understanding plant resistant mechanisms is crucial to counter this disease effectively. Qualitative resistance of the plant, which is controlled by a single resistant (R) gene activates when the infecting pathogen contains the corresponding avirulence (Avr) gene. The R-Avr incompatible interaction leads to a hypersensitive reaction as the plant recognizes the pathogen at early onset. This study focused on identifying genes associated with the B. napus–L. maculans pathosystem using dual RNA-sequencing. B. napus genotype, ‘01–23-2-1‘ that carries only Rlm7 was inoculated with a L. maculans isolate UMAvr7 that carries only AvrLm7, and with the AvrLm7 knockout mutant umavr7 of the same isolate in order to study incompatible and compatible interactions, respectively. This study will investigate differential gene expression during biotrophic over necrotrophic stage, differentially expressed effector genes in the pathogen, receptor-genes and genes associated with signal transduction of the host. Plants were inoculated followed by sample collection at time of inoculation, 1 d, 3 d, 7 d and 11 d after inoculation. Total RNA was extracted and sequenced on an Illumina 4000 HiSeq. The results were presented at the conference.
Sensitivity of Monilinia vaccinii-corymbosi to propiconazole from wild blueberry fields. D. C. PERCIVAL, L. GUO, S. JOSE, A. SCHILDER, B. PRITHVIRAJ AND A. R. OLSON. Department of Plant, Food & Environmental Sciences, Dalhousie University Agriculture Campus, Truro, NS B2N 5E3, Canada; and (A.S.) Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI 48823, USA
Monilinia blight, caused by Monilinia vaccinii-corymbosi (Reade) (M.vc) Honey, is one of the most devastating fungal diseases of wild blueberry. Chemical control of Monilinia blight is necessary to achieve high yield and quality berries in the absence of adequate host plant resistance. Propiconazole, the most effective fungicide against Monilinia blight has been used throughout the wild blueberry industry for the past 20 years. However, long-term use may lead to the development of resistance among fungal populations. Given the extensive usage of propiconazole, the occurrence of fungicide resistance in M.vc population was examined in 2011, 2012 and 2013 from five commercial fields in Nova Scotia. The effective concentration (EC50) of fungicide were determined for 102 single spore isolates (isolated from mummy berries and Monilinia blighted shoots) and compared with the baseline EC50 value of isolates from unmanaged and conventionally managed fields in Maine, USA. All isolates, regardless of collection date, were sensitive (ranged from 0.007 to 0.036 μg mL−1) with an average of 0.015 μg mL−1. The value was not significantly different from the mean propiconazole EC50 value of baseline isolate (0.016 μg mL−1), but was lower than the mean propiconazole EC50 value of isolates (0.021 μg mL−1) from conventionally managed fields in Maine. Although variation in sensitivity was observed within and among isolates from different locations, it can be concluded that M.vc isolates from Nova Scotia have not developed reduced sensitivity to propiconazole. Furthermore, the results from this study can serve as a benchmark for assessing any future decline in sensitivity to propiconazole.
Genome analysis of Pseudomonas sp. strain S1Bt23, a potent fungal antagonist, reveals the presence of phenazine and pyrrolnitrin gene clusters. S. F. PINTO, H. BALASUNDARAM, R. XU AND J. T. TAMBONG. Ottawa Research and Development Research Centre, Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada
Species of the genus Pseudomonas have been proven to be environmentally significant as e.g. phytopathogens, biological control agents and xenobiotic degraders. Certain species such as Pseudomonas chlororaphis are able to synthesize metabolites implicated in bioprotection of crops against fungal pathogens. We demonstrated that strain S1Bt23 inhibited, in vitro, in dual cultures, the growth of Rhizoctonia solani, Fusarium graminearum, Alternaria solani, Pythium ultimum and Pythium arrhenomanes. The objectives of this study were (1) to determine the taxonomic position of the strain using a polyphasic approach and (2) to analyse the genome of strain S1Bt23 for antifungal gene clusters. 16S rRNA sequence analyses suggested that strain S1Bt23 was affiliated with the genus Pseudomonas. Multi-locus sequence analyses (MLSA) using recA, gyrB, rpoB and rpoD clustered strain S1Bt23 uniquely within the subspecies of P. chlororaphis. The genome of S1Bt23 was sequenced and genome-based DNA-DNA hybridization (dDDH) and MuMmer-based average nucleotide identity (ANIm) performed with four putative P. chlororaphis subspecies. Based on dDDH, ANIm and proteome data, we conclude that strain S1Bt23 is an authentic novel subspecies within the species P. chlororaphis. In-depth genome analysis of strain S1Bt23 identified the presence of phenazine and pyrrolnitrin gene clusters known to produce antifungal metabolites. Work is now focused on determining whether these gene clusters are involved in the potent antifungal properties of strain S1Bt23.
Development of multiplex protocols to detect grapevine wood diseases. C. PROVOST, K. OZAKI, C. GUERTIN AND E. DEZIEL. Centre de recherche agroalimentaire de Mirabel. 9850 rue Belle-rivière, Mirabel, QC J7N 2X8, Canada; and (K.O., C.G., E.D.) INRS – Institut Armand-Frappier, 531 boulevard des Prairies, Laval, QC H7V 1B7, Canada
Grapevine wood diseases are considered very damaging to the sustainability of viticultural heritage in all major wine regions of the world. Several fungi are responsible for these diseases and attack the perennial organs of the vine, which causes plant death in a short or medium term. Grapevine wood diseases can affect young plantations as well as ageing vineyards where the risk related to the presence of these diseases is growing across the province of Quebec. The main objective of this project is therefore to develop laboratory methods to rapidly detect 10 pathogenic fungi associated with five grapevine wood diseases that are, or are expected to become, prevalent in Canada. The diseases targeted in the project are Esca, Black foot, Black dead arm, Eutypa dieback and Dead-arm disease. To achieve this goal, we chose an approach that uses multiplex real-time PCR, in which TaqMan® hybridization probes can target and bind to different genomic markers of the pathogenic agents. Four multiplex systems were developed, one for each disease, and are ready to use with field samples. For example, the first duplex system, targeting the internal transcribed spacer 1 (ITS1) region of the studied pathogens, can detect DNA from Phaemoniella chlamydospora and Phaoacremonium aleophilum, which are two fungal species that cause Esca. The benefits of this project will enable the various stakeholders in the vineyard to have access to accurate, reliable and rapid diagnostic methods, which will have a positive impact on the overall health of Quebec’s vineyards.
Survival and detection of Colletotrichum species on celery and common weeds species. S. REYNOLDS, M. J. CELETTI, K. S. JORDAN AND M. R. MCDONALD. Department of Plant Agriculture, University of Guelph, 50 Stone Road East, Guelph, ON N1G 2W1, Canada; and (M.J.C.) Ontario Ministry of Agriculture, Food and Rural Affairs, University of Guelph, Guelph, ON N1G 2W1, Canada
Fungal pathogens Colletotrichum fioriniae and C. nymphaeae cause anthracnose on several agriculturally important crops but can also asymptomatically colonize non-host species. The objective of this study was to evaluate the pathogenicity of C. fioriniae and C. nymphaeae and determine their survival on common weed species of Ontario. Conidia of one isolate of C. fioriniae from celery and apples and one isolate of C. nymphaeae from strawberry were inoculated onto the foliage of celery (cv. ‘TZ 9779’) and five weed species: Amaranthus retroflexus, Chenopodium album, Chenopodium glaucum, Cyperus esculentus and Senecio vulgaris. Leaf tissue sections were excised 10 and 20 days post inoculation (dpi), and were either plated directly or surface-sterilized and plated on semi-selective media to determine per cent pathogen recovery. Quantitative PCR was also used to detect and quantify the pathogen from leaf samples, and conidia germination was characterized by light microscopy. C. fioriniae caused typical anthracnose symptoms on celery, whereas C. nymphaeae only caused minor lesions on young leaves. Weeds remained asymptomatic, however, plated leaf segments from all weeds and celery showed Colletotrichum growth. Growth was observed from surface-sterilized leaves, suggesting endophytic or latent infection. On all plant species tested, most conidia germinated, developed melanized appressoria and produced secondary conidia by 4 dpi. Quantitative PCR was able to detect C. fioriniae and C. nymphaeae, and therefore can be used as a tool for rapid detection of the pathogen on asymptomatic leaves. Both Colletotrichum species colonized and reproduced on weeds, which could be a potential inoculum source for surrounding hosts.
Compartmentalization barriers in trees: some reflections on possible routes exploited by pathogens to overcome these defensive tissues. D. RIOUX AND M. BLAIS. Natural Resources Canada, Canadian Forest Service, Laurentian Forestry Centre, 1055 du P.E.P.S., P.O. Box 10380, Stn. Sainte-Foy, QC G1V 4C7, Canada
Compartmentalization barriers (CBs), in the broad sense, occur as non-specific defensive responses in the bark and xylem of trees reacting to various injuries or attacks by pathogens. Many studies report that the entry of air which accompanies this damage would be the main trigger to the formation of CBs. Canker pathogens can directly penetrate necrophylactic periderms, the main CBs in the bark. However, it has also been proposed that they can colonize beyond these CBs indirectly, through disruptions in them or by bypassing these defensive tissues. As we showed in a recent study involving artificial inoculations with the butternut canker pathogen, and no matter the route taken, fungal colonization that occurs during dormancy seems to be crucial in disease development. In some cases, when the pathogen is already present in the xylem, or when it can reach it, propagules can be transported over long distances beyond the CBs through the long open vessels. Time-course microscope studies are needed to follow the path of these pathogens and promising insights will likely be gained using fluorescent species-specific markers, but this only when coupled with the use of reagents capable of quenching the autofluorescence of the numerous compounds present in trees.
Does inoculation with the biocontrol agent Pseudomonas fluorescens LBUM223 impact the rhizosphere and geocaulosphere microbiomes of potato? R. ROQUIGNY, A. NOVINSCAK, M. FILION AND D. L. JOLY. Université de Moncton, 18 avenue Antonine-Maillet, Moncton, NB E1A 3E9, Canada
The phenazine-1-carboxylic acid (PCA)-producing Pseudomonas fluorescens Migula strain LBUM223 shows biocontrol capacity by reducing symptoms of the causal agent of common scab of potato, Streptomyces scabies (Thaxter) Waksman & Henrici. The aim of this study is to better characterize the impact of inoculating a specific biocontrol agent under natural field conditions on the microbiomes of the rhizosphere and the geocaulosphere of potato plants using Next-Generation sequencing. Single or biweekly applications of LBUM223 were performed up to 11 weeks after planting (in addition to non-inoculated plants). Rhizosphere and geocaulosphere (when potato tubers were produced) soils were sampled every 2 weeks. Following soil DNA extractions, 16S rRNA gene amplification and sequencing were performed using the Illumina MiSeq technology. The QIIME pipeline was used for data analyses. Results were generated from 45 rhizosphere and 27 geocaulosphere samples, for which 63,502 and 44,469 different operational taxonomical units were observed. Diversity comparisons were performed between both datasets. To our knowledge, this is the first time that the geocaulosphere microbiome is characterized and compared to the rhizosphere. Eleven phyla accounted for 95% of the diversity, with Actinobacteria, Proteobacteria, Chloroflexi and Acidobacteria being the most important ones. Overall, the results obtained suggest that Pseudomonas fluorescens LBUM223 does not significantly interfere with the autochthonous rhizosphere or geocaulosphere microbiomes, providing first insights on its non-target safety in the field.
Évaluation des besoins en phytoprotection des cultures ornementales en serres. N. ROULLÉ, A. BÉLANGER, B. CHAMPAGNE, M. A. LAPLANTE ET M. É. TOUSIGNANT. Institut Québécois du développement en horticulture ornementale du Québec,3230 rue Sicotte, E-307, St-Hyacinthe, QC J2S 2M2, Canada
Au Québec, les cultures ornementales en serres représentent près de 136 millions de dollars de vente par an. Ce sont des productions qui incluent plus de 300 espèces de plantes et plus de 50 ennemis des cultures. Mandaté par le MAPAQ depuis 2016 pour réaliser la surveillance phytosanitaire dans les cultures ornementales du Québec, l’IQDHO a entrepris de réaliser une évaluation des besoins en phytoprotection du secteur des serres. L’objectif de cette évaluation était tout d’abord de mieux orienter les efforts de l’IQDHO en termes de choix de projets de recherche, de thèmes de formation et de sujets de publications techniques. L’objectif était également de mieux communiquer les besoins du secteur aux différents intervenants provinciaux et fédéraux. Tout au long de la démarche, les treize conseillers et conseillères des cultures ornementales en serres du Réseau d’avertissements phytosanitaires du Québec (RAP) ont été consultés. La démarche s’est déroulée en 4 étapes: 1-Établissement de la liste des 13 ennemis des cultures les plus problématiques; 2-Pour ces ennemis, évaluation des pertes économiques, de l’utilisation en pesticides et de la disponibilité en méthode de contrôle; 3-Évaluation des besoins en projets de recherche, en formations et en demandes d’homologation de pesticides; 4-Proposition de projets de recherche et de thèmes de formation. Cette consultation des conseillers et conseillères a permis d’établir un portrait des principales problématiques du secteur. Ce portrait sera un guide pour viser une réduction significative des impacts des ravageurs et des maladies, ainsi qu’une réduction de l’usage des pesticides.
Modelling and mapping the suitability of Canada’s climate for an insect-vectored, exotic blue-stain fungal pathogen, Endoconidiophora polonica. K. R. SAMBARAJU, R. SAINT-AMANT, C. CÔTÉ AND B. FILION. Natural Resources Canada, Canadian Forest Service, Laurentian Forestry Centre, 1055 du P.E.P.S., P.O. Box 10380, Stn. Sainte-Foy, QC G1V 4C7, Canada
Dramatic increases in the numbers of non-indigenous pest invasions have been observed globally over the past decades owing to increased global trade. At risk in forest ecosystems are ‘naïve’ trees that are susceptible to damage as they may lack defence mechanisms to counter harmful invasives. Introductions of non-native forest insects and pathogens can cause catastrophic tree declines, endangering ecological balance and causing serious economic losses. In case of an accidental invasion or when a risk of introduction exists, it is critical to understand the climatic suitability of susceptible forests in order to develop proactive plans for pest detection and spread assessment. In this work, we assessed the suitability of Canada’s climate for an exotic blue-stain fungal pathogen, Endoconidiophora polonica (Siemaszko) de Beer, Duong & Wingf., that is vectored by the European spruce bark beetle, Ips typographus L. We collected occurrence data of E. polonica from several sources including online databases, journal articles and culture collections. We then generated 22 bioclimatic variables that summarized the tolerances of the pathogen in its native habitat. Two different modelling approaches were used to develop climatic suitability indices that were used to create maps for the pathogen for Eurasia under the current climate. We compared the outputs from the two models with a random model to select the ‘best’ model for mapping Canada’s climatic suitability. Mapping suggests that current climatic conditions are conducive for the establishment of the pathogen in Canada, and forests further north will become favourable in the future.
Battling biology – overcoming challenges with spore germination. A. M. SETO, M. E. DONALDSON AND B. J. SAVILLE. Environmental and Life Sciences Graduate Program, Trent University, 1600 West Bank Drive, Peterborough, ON K9L 0G2, Canada; and (B.J.S.) Forensic Science Program, Trent University, 2140 East Bank Drive, DNA Building, Peterborough, ON K9L 0G2, Canada
Fungal diseases are a major threat to sustainable crop production. The success of these fungal pathogens stems from their production of spores, which allows them to disperse over long distances and survive for extended periods of time outside the host. The processes of spore dormancy, dispersal and germination are integral to the emergence of infectious fungal diseases. Despite their importance, studies on fungal spores have been limited. We use Ustilago maydis D.C. Corda as a model for studying plant–pathogen interactions. This fungus is dispersed as thick-walled dormant teliospores, which germinate, and complete meiosis to initiate new rounds of Zea mays infection. Teliospore germination is asynchronous, which presents a challenge when identifying changes in gene expression. To overcome this challenge, we used RNA-seq to analyse transcript level changes at 0, 9 and 18 h after teliospore germination was induced. These analyses identified 18 patterns of transcript level change, nine for genes up-regulated in the teliospore and nine for genes down-regulated in the teliospore. These patterns suggest transcriptional and post-transcriptional control of gene expression during teliospore development and germination. Reverse transcriptase quantitative PCR (RT-qPCR) confirmed some of the patterns of transcript level change. Gene ontology (GO) enrichment analysis identified biological processes that were enriched for in each pattern. A summary of the GO enrichment and RT-qPCR analysis will be presented, along with a model of functional changes during teliospore development and germination. This model can be used to inform future investigations of smut and rust teliospore germination.
Novel techniques for screening cercospora leaf spot resistant fenugreek (Trigonella foenum-graecum) genotypes. U. SUBEDI, S. N. ACHARYA, S. CHATTERTON, J. THOMAS, R. BARENDREGT AND D. FRIEBEL. (U.S., J.T.) Department of Biological Sciences, University of Lethbridge, 4401 University Drive West, Lethbridge, AB T1K 6T5, Canada; (U.S., S.N.A., S.C., D.F.) Lethbridge Research and Development Centre, Agriculture and Agri-Food Canada, 5403 1 Avenue South, Lethbridge, AB T1J 4P4, Canada; and (R.B.) Department of Geography, University of Lethbridge, 4401 University Drive West, Lethbridge, AB T1K 6T5, Canada
Cercospora leaf spot (CLS) caused by Cercospora traversiana is an important phyto-pathological problem of fenugreek (Trigonella foenum graecum L.), a multi-use legume crop. Field screenings for resistant plants, although accurate and effective, demand significant time and a sizable workforce to accomplish the goal. Also, weather conditions in the field may not always be favourable for uniform disease spread which eventually may lead to failure of the overall experiment. Whole plant assay (WPA) and detached leaf assay (DLA) with artificial inoculation not only help in scaling up the number of plants screened but also reduce the space, time and the amount of inoculum needed for the experiment. The results in our experiment indicated that both WPA and DLA methods could be used reliably to differentiate the resistant and susceptible genotypes of fenugreek. In addition, the correlation coefficient (r = 0.784, P < 0.01) derived from the mean disease score from each genotype, between WPA and DLA showed that they can be used interchangeably while screening fenugreek for CLS. The DLA was found to be temperature sensitive for development of CLS symptoms and wounded leaves developed symptoms faster than un-wounded leaves. These indoor methods can be used for development of CLS resistant fenugreek cultivar in areas where disease development is difficult under field condition.
CRISPR/Cas9 gene editing of the Dutch elm disease pathogen Ophiostoma novo-ulmi. P. TANGUAY. Natural Resources, Canadian Forest Service, Laurentian Forestry Centre, 1055 rue du PEPS, P.O. 10380 Ste-Foy stn., QC G1V 4C7, Canada
Strategies employed to establish a CRISPR/Cas9 system to efficiently disrupt target genes in the tree pathogen Ophiostoma novo-ulmi are being described. We achieved successful CRISPR/Cas9 knock-out of the ADE2 gene. O. novo-ulmi protoplasts were co-transformed by PEG/CaCl2 using synthetic sgRNAs and a plasmid expressing a O. novo-ulmi codon-optimized Cas9 nuclease. Furthermore, we are testing, in O. novo-ulmi, the ability of a linear plasmid with telomeric ends to behave as an artificial acentric minichromosome, rapidly lost under non-selective conditions. This would allow recycling of the selection marker, and prevent the potential negative effects of constitutive Cas9 expression. A CRISPR/Cas9 gene editing system is an important advance to investigate gene function in O. novo-ulmi, a species with extremely low rates of homologous recombination.
Evaluation of different oak leaf extracts for the control of bacterial leaf spot of lettuce. V. TREMBLAY, V. TOUSSAINT AND R. J. TWEDDELL. Département de phytologie, Université Laval, Québec, QC G1V 0A6, Canada; and (V.T.) Saint-Jean-sur-Richelieu Research and Development Centre, Agriculture and Agri-Food Canada, 430 boulevard Gouin, Saint-Jean-sur-Richelieu, QC J3B 3E6, Canada
Oak (Quercus spp.) leaf extracts were reported in several studies to have antimicrobial properties. Such properties could be exploited for the control of bacterial diseases affecting horticultural crops for which very few antibacterial chemicals are available. As part of ongoing research aimed to develop new strategies to control bacterial leaf spot (Xanthomonas campestris (Pammel) Dowson pv. vitians) of lettuce (Lactuca sativa L.), the objectives of the study were (1) to test different red oak (Quercus rubra L.) leaf extracts (aqueous, ethanol:water, acetone:water) for their antibacterial activity against X. campestris pv. vitians and (2) to evaluate their efficacy to control bacterial leaf spot of lettuce. Antibacterial activity of the extracts was determined using standard procedures. Efficacy of the extracts to control bacterial leaf spot was tested on lettuce plants grown in greenhouse. With a minimum bactericidal concentration of 12.5 mg mL−1, ethanol extracts (40:60 and 20:80, ethanol:water) showed the strongest antibacterial activity. Foliar application of aqueous, ethanol (80:20, 60:40 and 40:60), and acetone (50:50 and 20:80, acetone:water) extracts allowed a marked reduction in disease severity when the disease pressure was low to moderate. Moreover, addition of sodium bicarbonate (0.2 M) was shown to increase the efficacy of ethanol extracts (80:20 and 60:40) to control the disease. In conclusion, oak leaf extracts show potential for controlling bacterial leaf spot of lettuce and could eventually find application to control bacterial diseases affecting horticultural crops.
Evaluation of different crop residues for the control of bacterial leaf spot of lettuce. V. TREMBLAY, A. BELLEY, V. TOUSSAINT AND R. J. TWEDDELL. Département de phytologie, Université Laval, QC G1V 0A6, Canada; and (V.T.) Saint-Jean-sur-Richelieu Research and Development Centre, Agriculture and Agri-Food Canada, 430 boulevard Gouin, Saint-Jean-sur-Richelieu, QC J3B 3E6, Canada
Over the past decades, numerous studies have reported that many plant extracts show toxicity against fungi and bacteria. The exploitation of the bactericidal properties of plant extracts represents an interesting approach that could find application for the control of bacterial diseases affecting lettuce (Lactuca sativa L.), namely bacterial leaf spot (BLS) (Xanthomonas campestris (Pammel) Dowson pv. vitians). The objectives of the study were (1) to test different extracts produced from crop residues for their antibacterial activity against X. campestris pv. vitians and (2) to test their efficacy to control BLS. Determination of antibacterial activity of the extracts was carried out using standard procedures. Efficacy of the extracts to control BLS was determined on lettuce plants grown in greenhouse. Aqueous extracts of radish (Raphanus sativus L.) leaf and cranberry (Vaccinium macrocarpon Ait.), and ethanol:water (80:20) extract of kale (Brassica oleracea var. sabellica L.) with respective minimum bactericidal concentrations (MBC) of 12.5, 50 and 50 mg mL−1 showed the strongest antibacterial activities. Aqueous extracts of broccoli (Brassica oleracea L. var. italica Plenck), canola (Brassica napus L.), kale, radish and rocket (Eruca sativa Mill.) showed MBC higher than 100 mg mL−1. When applied on lettuce plants inoculated with X. campestris pv. vitians, aqueous, ethanol:water (80:20, 60:40 and 20:80) and acetone:water (40:60 and 20:80) extracts of kale allowed a marked reduction of BLS severity that was observed only under low to moderate disease pressure.
Deoxynivalenol-3-Glucoside content of two-row barley infected with Fusarium graminearum measured by ultra-performance liquid chromatography – tandem mass spectrometry. J. R. TUCKER, A. BADEA, R. BLAGDEN, K. PLESKACH, S. A. TITTLEMIER AND W. G. D. FERNANDO. Brandon Research and Development Centre, Agriculture and Agri-Food Canada, 2701 Grand Valley Road, P.O. Box 1000A, R.R. 3, Brandon, MB R7A 5Y3, Canada; (R.B., K.P., S.A.T.) Grain Research Laboratory, Canadian Grain Commission, 303 Main Street, Winnipeg, MB R3C 3G8, Canada; and (J.R.T., W.G.D.F.) Department of Plant Science, 222 Agriculture Building, 66 Dafoe Road, University of Manitoba, Winnipeg, MB R3T 2N2, Canada
Fusarium graminearum Schwabe produces a range of secondary metabolites such as mycotoxins, which can be associated with pathogenicity. Deoxynivalenol (DON) is a common mycotoxin associated with infected barley grains. This mycotoxin is known to inhibit protein synthesis in eukaryotes through ribosomal interference, thereby limiting resistance response of the host. A common host defence mechanism is induction of detoxification genes that code for enzymes which facilitate conjugation of mycotoxins to alternate molecules. While these conjugated mycotoxins demonstrate reduced toxicity in planta, such forms represent a risk to industry as they can be transformed back to toxic state under processing. Strict guidelines for maximum limits have been set by malting and brewing and livestock feed industries, and grains are monitored. However, bound-mycotoxins are not detected by standard analytical procedures and are typically not an assayed target. A study was conducted at Brandon, MB in 2016 and 2017 to evaluate mycotoxin profile of 16 barley genotypes that differed in their resistance response to fusarium head blight and DON production. Plots were artificially inoculated with conidia suspension using a mixture of isolates and grown under irrigation to promote disease. Plots were split-harvested at soft-dough and ripe stage followed by desiccation in a high capacity drier. The soft dough samples were divided into rachis and seed components. A panel of 13 mycotoxins were assessed on all samples via UPLC-MS/MS, which included targets: DON, 3ADON, 15ADON, ZEA and DON-3-Glucoside. Results from mycotoxin analyses were discussed in reference to level barley host resistance.
Sudden death syndrome of soybean: Know your enemy. O. S. WALLY AND A. TENUTA. Harrow Research and Development Centre, Agriculture and Agri-Food Canada, 2585 County Road 20, Harrow, ON N0R 1G0, Canada; and (A.T.) Ontario Ministry of Agriculture, Food and Rural Affairs, 120 Main Street East, Ridgetown, ON N0P 2C0, Canada
The fungal pathogen Fusarium virguliforme causes soybean sudden death syndrome (SDS) in North America. The disease was first observed in Arkansas and has spread in a general northern direction through the majority of soybean growing areas in North America, being first identified in Ontario in 1998. Fusarium virguliforme colonizes the roots of soybeans during seed germination, as the soybeans mature the fungi produces toxins that translocate up the xylem causing the classic interveinal chlorosis and leaf cupping, eventually leading to premature defoliation and seed abortion. Disease is rated as a disease index (DX) which is a measure of severity (0–9, 0 = asymptomatic, 9 = defoliation) multiplied by the incidence (%) divided by 9, giving a DX from 0–100. Yield losses from SDS are directly correlated to DX, with an increase of 10 DX resulting in ~ 10% reduction in yield; losses due to SDS make it the second or third most damaging soybean pest in the northern USA and Ontario depending on the year. There are a number of factors that influence the disease which include: varietal resistance, soil composition, soil moisture and interactions with the soybean cyst nematode (SCN). Control measures for mitigating SDS damage include planting tolerant varieties as there currently are no completely resistant varieties; delaying planting date and the use of certain seed treatments have proven effective under moderate SDS pressure. Additional control may be obtained through the study of soils grown under continuous soybeans which have been discovered to be highly suppressive to SDS in south-western Ontario.
Impact of blackleg (Leptosphaeria maculans) on canola yield. Y. WANG, S.-F. HWANG AND S. E. STRELKOV. Department of Agricultural, Food and Nutritional Science, 410 Agriculture/Forestry Centre, University of Alberta, Edmonton, AB T6G 2P5, Canada; and (S.F.H.) Crop Diversification Centre North, Alberta Agriculture and Forestry, 17507 Fort Road NW, Edmonton, AB T5Y 6H3, Canada
Blackleg, caused by Leptosphaeria maculans (Desm.) Ces & de Not., is one of the most devastating diseases of canola (Brassica napus L.) worldwide. Yield losses as high as 30–50% have been reported in susceptible cultivars, and total crop failure can occur in severe epidemics. In Canada, blackleg is managed mainly by the cultivation of resistant or moderately resistant canola hybrids. Field experiments were conducted in 2017 to determine the relationship between blackleg severity and yield in two moderately resistant hybrids ‘73-15RR’ and ‘1950RR’. Blackleg disease severity gradients were generated by the application of different levels of L. maculans inoculum. The experiment was designed as a split plot and disease severity (on a 0–5 scale), pod number per plant and seed yield per plant were recorded at crop maturity. Regression analysis showed that pod number and seed yield had non-linear relationships with blackleg severity, and these relationships were best explained by second degree quadratic equations. Plants with a blackleg severity of 0 had a slightly lower seed yield than plants with a severity of 1 for both canola hybrids. Yield decreased by 15.6–100%, however, in plants with disease severities of 3–5 compared with plants with disease severities of 0–2. These results suggest that even in moderately resistant hybrid canola cultivars, seed yield per plant decreases as a result of L. maculans infection. Thus, alternative disease management strategies are required to mitigate yield losses due to blackleg. The experiment will be repeated in 2018 to confirm the results.
Identification of rutabaga accessions resistant to new and old pathotypes of Plasmodiophora brassicae from Alberta. Z. YU, R. FREDUA-AGYEMAN, S. F. HWANG AND S. E. STRELKOV. Department of Agricultural, Food and Nutritional Sciences, 410 Agriculture/Forestry Centre, University of Alberta, Edmonton, AB T6G 2P5, Canada; and (R.F.-A., S.F.H.) Crop Diversification Centre North, Alberta Agriculture and Forestry, 17507 Fort Road NW, Edmonton, AB T5Y 6H3, Canada
Clubroot, caused by Plasmodiophora brassicae Woronin, is a devastating yield-limiting disease of Brassica crops worldwide. The identification of new pathotypes of P. brassicae that can cause disease on clubroot-resistant canola (Brassica napus L.) cultivars in Canada has necessitated a search for new resistance sources to be used in breeding. Sixty rutabaga (B. napus spp. napobrassica) accessions were evaluated for resistance to 10 field- or single-spore isolates representing several of the ‘new’ and ‘old’ pathotypes. The new pathotypes included three field isolates classified as pathotype 5X and one field isolate each of pathotypes 5L and 5I. The old pathotypes included one single-spore isolate of each of pathotypes 2F, 3H 5I, 6M and 8N. One accession FGRA107 was resistant (index of disease ≤ 30%) to all 10 isolates while two accessions FGRA037 and FGRA044 were each resistant to nine and moderately resistant (30% < index of disease ≤ 50%) to one isolate. In addition, 58–68% of the rutabaga accessions were resistant to pathotypes 5L, 5I and one isolate of 5X. In contrast, only 5–10% of the rutabaga accessions were resistant to the old pathotypes. The observation that the rutabaga accessions showed significantly (P < 0.05) higher resistance to isolates representing the new vs. old pathotypes suggests that clubroot resistance to the new and old pathotypes may be under different genetic control.
Assessment of genetic variation in new strains of Plasmodiophora brassicae in Alberta by ITS sequence analysis. Q. ZHOU, S. F. HWANG, H. FU, R. FREDUA-AGYEMAN, V. P. MANOLII AND S. E. STRELKOV. Crop Diversification Centre North, Alberta Agriculture and Forestry, 17507 Fort Road NW, Edmonton, AB T5Y 6H3, Canada; and (V.P.M., S.E.S.) Department of Agricultural, Food and Nutritional Science, 410 Agriculture/Forestry Centre, University of Alberta, Edmonton, AB T6G 2P5, Canada
Clubroot of crucifers, caused by Plasmodiophora brassicae Woronin, is an important soil-borne disease of canola (Brassica napus L.) in Alberta, Canada. The disease is managed mainly by growing clubroot-resistant cultivars in the field. However, new strains of P. brassicae capable of overcoming this resistance were identified from more than 60 fields in Alberta between 2013 and 2016, suggesting shifts in the virulence of the pathogen populations. Evaluation of the virulence phenotypes of selected P. brassicae populations on the hosts of the Canadian Clubroot Differential Set revealed the occurrence of pathotypes A-P and X, but information on genetic variation is lacking. Such information is important for assessing diversity in pathogen populations and understanding the relationship between pathotypes. In the present study, the 18S and internal transcribed spacer (18S-ITS) regions of P. brassicae populations representing a total of 62 fields were amplified with a pathogen-specific forward primer RP2F and a universal reverse primer ITS4. The resultant amplicons were cloned and sequenced, and genetic variation was analysed by the neighbour-joining and bootstrap methods. The results of this analysis were presented.
Quantitative trait loci associated with pasmo resistance in flax. F. M. YOU, L. HE, K. Y. RASHID, Z. YAO, P. LI, J. XIAO, X. WANG AND S. CLOUTIER. (F.M.Y., L.H., S.C.) Ottawa Research and Development Centre, Agriculture and Agri-Food Canada (AAFC), 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada; (L.H., J.X., X.W.) State Key Lab of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, Nanjing, China; and (K.Y.R., Z.Y., P.L.) Morden Research and Development Centre, AAFC, 101 Route 100, Morden, MB R6M 1Y5, Canada
Pasmo is one of the most prevalent and widespread diseases limiting flax production. To improve genetic resistance to this disease, we phenotyped 370 accessions of the flax core collection in the field for pasmo resistance (PR) from 2012 to 2015 in Morden, MB, Canada. Genome-wide association study (GWAS) analyses using 258,873 SNPs were performed using 10 different statistical models including the traditional single-locus and the most recent multi-loci methods. A total of 355 unique quantitative trait nucleotides (QTNs) with significant effects, corresponding to 296 potential quantitative trait loci (QTL), were identified from the four individual years’ datasets. Different QTNs were obtained from the various methods and datasets, indicative of the complementation between analytical methods and/or genotype × environment interaction of the QTL effects. From these putative QTL, 47 were stable across all 4 years, had relatively large effects (> 5%), and explained 22–46% of the total variation for PR. Of these, 40 span resistance gene analogues. The number of positive-effect QTL (NPQTL) in accessions was significantly correlated to PR, indicative of their additive effects. NPQTL was also significantly associated with morphotype where most of the positive-effect QTL for PR were present in the fibre type accessions, thus making this germplasm an important source for PR improvement of linseed. The identified QTL can be used as molecular markers for germplasm evaluation, and parent and offspring selection in flax PR molecular breeding.
Plant pathogenic fungi in bentonite-amended sandy soil under corn monoculture in Northern China. H.-Y. ZHANG, J. H. LIU AND W. CHEN. (H.-Y.Z., W.C.) Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, ON K1A 0C6, Canada; and (H.-Y.Z., J.H.L.) Inner Mongolia Agricultural University, 306 Zhaowuda Road, Hohhot, Inner Mongolia, 010000, China
Sandy soils of aerolian origin are light-textured, alkaline, with poor water and nutrient retention and prone to compaction. The economic loss associated with sandification was estimated at 54 million RMB/year in China, with Horqin Sandy Land (a farming-pastoral ecotone in Northern China) being most severely affected. Over 70% arable lands in Horqin are under continuous corn production, often constrained by water deficits. Our long-term experiments on corn monocultural plots showed the addition of natural bentonite improved the aggregate stability, water content and enzymatic activities of sandy soil and maize production. The dynamics of the soil mycobiota, especially soil-borne fungal plant pathogens, in response to bentonite amendment was investigated using the fungal internal transcribed spacer 1 (ITS1) metabarcodes. The results showed that Ascomycota abundance was twice that of Basidiomycota in corn fields, while the most abundant fungal genera included Guehomyces, Alternaria, Mortierella, Cladosporium and Exophiala. The pathogenic mycobiota (annotated by FUNGuild) was dominated by Alternaria (5.8%), Cladosporium (4.24%) and Fusarium (3.57%) in abundance. Fungal plant pathogens in bentonite-amended soils had higher diversity and prevalence and differed significantly in composition from those in non-amended soils. Further analyses revealed the key soil properties and bacterial OTUs that may enhance the soil fungistatic potential. Notwithstanding bentonite-amendment improves the overall performance of sandy soils in corn production, however the increased diversity and abundance of fungal pathogens, possibly due to increased nutrient levels, requires pest management attention.
Identification and characterization of Verticillium isolates from Brassica crops in Manitoba, Canada. Z. ZOU, V. BISHT AND W. G. D. FERNANDO. Department of Plant Science, 222 Agriculture Building, 66 Dafoe Road, University of Manitoba, Winnipeg, MB R3T 2N2, Canada; and (V.B.) Primary Agriculture Branch, Manitoba Agriculture, P.O. Box 667, 65-3rd Avenue NE, Carman, MB R0G 0J0, Canada
Verticillium stem striping of canola (Brassica napus L.), caused by Verticillium longisporum, was first reported in Manitoba in 2014. In this study, Brassica crops including canola, mustard (B. juncea), and radish (Raphanus sativus) with wilt symptoms were collected and pathogen isolated. Isolates from canola and radish were characterized to V. longisporum, which produced longer conidia (7.92–12.00 µm) than conidia of V. dahliae (4.32–7.04 µm). Isolates derived from mustard were characterized to V. dahliae. Molecular diagnostics with primers 18S rDNA, 5.8S rDNA, mating type were used to confirm the identification of verticillium isolates. PCR-RFLP of mitochondrial small subunit rDNA and cytochrome b gene were also employed to distinguish V. longisporum isolates to V. dahliae. Subtypes of V. longisporum isolates indicated that isolates from canola and radish are in A1/D1 group. V. longisporum was inoculated on to canola cultivar ‘Westar’, which caused stem striping, stunting, shorter plant height (P < 0.05), significant yield loss including total seed weight and 1000 seed weight (P < 0.05), lower oil content (P < 0.05) and a higher glucosinolate content.
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