Identification of mating types and metalaxyl resistance in North American populations ofPhytophthora infestans

The A² mating type ofPhytophthora infestans was first reported in the United States in 1990. Concurrently,P. infestans strains resistant to metalaxyl ere found in the Pacific Northwest. Collaborative surveys were undertaken during 1991–1993 to investigate the frequency of occurrence of A² mating types and metalaxyl resistant strains in populations ofP. infestans isolated from outbreaks of late blight in potato and tomato crops in North America.In vitro testing indicated that isolates from the northeastern U.S. and Atlantic Canada were primarily (52/55) metalaxyl sensitive and all were A¹ mating types. Among 85 isolates from late blight epidemics in Florida and Texas, greater than 61% were both metalaxyl resistant and A² mating type. Metalaxyl resistance and A² mating types were identified also in a few tomato isolates from North Carolina. Although the majority of 134 isolates from the Pacific Northwest (British Columbia and Washington) were metalaxyl resistant, only 2 isolates from Washington were A² mating types. Among 111 isolates from 2 sites in central Mexico, 63% and 77% were both metalaxyl resistant and A² mating types. The data indicate also a higher frequency of metalaxyl resistance in A² isolates, than in A¹ isolates, among isolates from Florida and Texas. Highest metalaxyl resistance levels were found, however, in A¹ isolates from California, where no A² isolates were recovered.     

Assessment of mating types and resistance to metalaxyl of Canadian populations ofPhytophthora infestans in 1997

The distribution of A1 and A2 mating types, and of metalaxyl-sensitive (MS), metalaxyl-intermediate (MI), and metalaxyl-resistant (MR) strains ofPhytophthora infestans, the causal pathogen of late blight, were determined in Canada in 1997. Potato plants or tubers and tomato plants or fruits, suspected to have late blight infection, were received from seven Canadian provinces in 1997, and were assessed for the presence ofP. infestans. Among 220 samples suspected to have late blight,P. infestans was recovered from 132 samples (60%) received from British Columbia (BC), Manitoba, New Brunswick, Prince Edward Island, and Quebec. From the 132 samples, 364 isolates were obtained and tested for their mating type and response to metalaxyl. Populations ofP. infestans within Canadian provinces were homogeneous for mating types, except within BC, where populations comprised both A1 and A2 mating types both within fields and sometimes within samples. The A2 mating type was found in all provinces where late blight was present. The A1 mating type was found only in Manitoba (1 isolate) and in BC (38 isolates). Unlike the old A1 isolates, those found in BC in 1997 were highly resistant to metalaxyl. These A1 isolates are even more resistant than A2 isolates. Based on assays with 10 ug.ml^?1 of metalaxyl, 14% of all tested isolates were MS, 69.8% were MI and 16.2% were MR. In addition, all provinces contained MS, MI, and MR isolates. No specific mating type or metalaxyl sensitivity level were specific to tomato or potato.     

Survey ofPhytophthora infestans populations in Serbia: Mating type, metalaxyl resistance and virulence properties

Summary A total of 37 single-lesion isolates ofPhytophthora infestans were collected during 2000 from different potato growing regions in the central part of Serbia. They were paired in Al and A2 matings to determine their mating types. Of these 37 isolates, 28 were found to belong to the A2 mating type, eight to the Al type and one was found to be self-fertile (A1/A2). Using the in vitro mycelial radial growth method no metalaxyl resistantP. infestans isolate could be found. The rate of occurrence of intermediate isolates was not high, on the average 2.70%. All other isolates were sensitive to metalaxyl. There was no correlation between mating type and the intermediate metalaxyl resistance found.     

Factors affecting the survival of metalaxyl-resistant strains ofPhytophthora infestans (Mont.) de Bary in Ireland

Summary Five metalaxyl-resistant and five sensitive isolates ofPhytophthora infestans were compared for sporangium production per unit leaf area, sporangium germination and virulence to foliage and to tubers. Differences between the two groups of isolates were significant for sporangium production per unit leaf area only. The metalaxyl-resistant strains produced fewer sporangia per unit leaf area than the metalaxyl-sensitive strains. The implications under field conditions are discussed.     

Phenotypes ofPhytophthora infestans in Nepal: mating types and metalaxyl sensitivity

Summary Potato late blight has appeared in epidemic proportions in Nepal since the mid 1990s and fungicides have been reported to be decreasingly effective in managing the disease.Phytophthora infestans isolates were collected from potato crops during 1999–2000 and analysis of 371 isolates for mating types and 270 isolates for metalaxyl sensitivity revealed the presence of both new and old populations. This is the first report on the presence of metalaxyl-resistant isolates in Nepal. The frequencies of A1 and A2 were 83 and 17%, respectively. Metalaxyl-resistant, intermediate and sensitive isolates were recorded as 10%, 12% and 78% respectively. Metalaxyl resistance was distributed in both mating types. Sites with a history of metalaxyl use had a significantly higher number of resistant and intermediate isolates ({ie337-1}, P<0.01) than sites where metalaxyl had not been used. This study confirms the changes in population structure ofP. infestans in Nepal.     

Changes in metalaxyl resistance among glucose phosphate isomerase genotypes ofPhytophthora infestans in Canada during 1997 and 1998

A total of 930 isolates ofPhytophthora infestans collected during 1997 and 1998 from commercial potato and tomato fields, home gardens, and potato storages in Canada was analyzed for glucose phosphate-isomerase (Gpi) genotype, mating type, and resistance to metalaxyl. Both mating types and diversity in resistance to metalaxyl were detected within and amongGpi-genotypes. Pathogen populations were dominated by the 100:111:122Gpi-genotype (characteristic of US-8) in all the provinces where late blight was detected, except in 1997 in British Columbia (BC) where the 100:100:111Gpi-genotype (characteristic of US-11) predominated.Gpi-genotypes 100:111, 111:122, and 100:122 (characteristic of US-7, US-10 and US-14/US-17, respectively) were less common. Nearly all of the isolates with the 100:100:111Gpi-genotype were the A1 mating type, except for a few collected in BC in 1997. A majority of isolates with the 100:111:122Gpi-genotype were A2 except for a few found in Québec in 1998. Based on colony growth on media amended with 1,10, or 100 μg/ml metalaxyl, the 100:100:111Gpi-genotype isolates had higher levels of resistance to the fungicide in 1998 compared to 1997. In 1998, level of metalaxyl resistance was higher within the 100:100:111Gpi-genotype whereas it was lower within the 100:111: 122Gpi-genotype, when compared to 1997. BC was the only Canadian province where an increase in metalaxyl resistance within the 100:111:122Gpi-genotype was recorded between 1997 and 1998. Isolates from leaves had higher metalaxyl resistance levels than stem or tuber isolates, regardless of genotype.     

Changes in the distribution of phenylamide-resistant strains ofPhytophthora infestans (Mont.) de Bary in Ireland 1985–89

Summary Phenylamide-based fungicides were reintroduced to the Irish market in 1985. Mixtures of phenylamides and mancozeb were recommended for no more than 3 sprays in potato late blight control programmes with the last systemic spray to be applied not later than 31 July. Samples of potato foliage infected withP. infestans were collected from the main potato growing areas in Ireland over a five-year period starting in 1985. Using the floating disc method, 11% of the crops sampled in 1985 were found to have phenylamide-resistant strains ofP. infestans present. The number of crops with resistant strains increased rapidly up to 1988 when 83% of the crops sampled had resistance. Factors affecting the distribution of resistance and the efficacy of a phenylamide-based spray programme are discussed.     

Relationships between pathotypes and RAPDs, Gpi-allozyme patterns, mating types, and resistance to metalaxyl ofPhytophthora infestans in Canada in 1997

Relationships between pathotypes and other markers such as mating types, metalaxyl resistance, Gpi-allozyme and random amplified polymorphic DNA (RAPD) patterns, were investigated among 33 isolates ofPhytophthora infestans obtained from both potato and tomato across Canada in 1997. Pathogenicity testings, based on virulence against potato differentials carrying single-resistance genes and the size of lesions on leaves, revealed higher pathogenic diversity in 1997 than in the few past years, with more pathotypes and more race-complex isolates. A higher pathogenic diversity was observed within A2 (US-8) than within A1 (US-11) isolates. There was no correlation between the level of resistance to metalaxyl of an isolate and its pathogenicity. Nevertheless, when grouped into metalaxyl-sensitive (MS), -intermediate (MI), and -resistant (MR) classes, both average pathotype complexity and average lesion size were lowest for MS group. RAPD data revealed a good distinction between A1 (US-11) and A2 (US-8) mating type groups, and between isolates from British Columbia and those from other provinces. The detection in the last few years of both A1 and A2 mating types in Canada and the occurrence in 1997 of some isolates sharing different characteristics of both of these two groups suggest that sexual reproduction may have occurred locally.     

Testing for resistance to metalaxyl inPhytophthora infestans isolates from northwestern Washington

Outbreaks of late blight occurred in commercial potato fields in northwestern Washington in 1989 and reached epidemic proportions in 1990. Since most blighted fields reportedly had received 2–4 late-season applications of metalaxyl, the possibility existed that pathogenic strains ofPhytophthora infestans had developed resistance to this systemic fungicide. In testing this hypothesis,P. infestans was isolated consistently from diseased leaves, stems, and tubers of potato when small tissue pieces were surface-disinfected in 0.5% NaOCl for 3 min. Virulence of isolates was assessed on detached leaflets of eight differential (R-gene) potato genotypes. Although simple races (single and double) were recovered, there was also a high frequency of complex race combinations. When tested in metalaxyl-amended media, 81% of the 73 isolates recovered from diseased tissues were highly resistant to 10 μg metalaxyl/ml and 19% yielded intermediate resistance. Sensitive isolates were not recovered. Fifty-five percent of 40 isolates sporulated on potato tuber disks in the presence of ≥10 ug metalaxyl/ml. On leaf tissues, 85% of 20 isolates sporulated in the presence of 10 μg metalaxyl/ml, and 50% sporulated in the presence of 100 μg/ml. This is the first report ofP. infestans resistance to metalaxyl in the U.S.     

Transmission ofPhytophthora infestans in cut potato seed

One or two cycles of late blight transmission may occur in cut potato seed between cutting and emergence. If seed are not planted immediately, mycelium from diseased pieces can infect across adjoining cut surfaces within 8 hr. Sporangia production begins on freshly cut diseased tubers in 1–2 days, on mycelium-infected seed pieces within 4–5 days, and on spore-infected seed pieces in about 1 wk. These spores are further dispersed during handling and planting, and may infect contaminated seed in the ground, regardless of suberization. Sporangia can be transferred to healthy tubers on cutting knives, although this appears infrequent. The risk and degree of secondary spread increase with the length of time cut seed are held before planting. Such secondarily infected seed are more likely to survive and produce infected stems than the original infected tubers. Treatment of seed with appropriate fungicides immediately after cutting substantially reduces or prevents transmission of late blight to healthy seed. Certain seed treatments withoutPhytophthora-specific components may increase the risk of tuber blight.     

Indications of soil borne inoculum ofPhytophthora infestans

Summary In this paper we present observations made during three years in a potato field in Sweden that indicated the presence of soil borne inoculum ofPhytophthora infestans. In 1994, a fungicide trial against late blight was left in the field with various degrees of late blight infection. In 1995, cereals was grown in the field. In 1996, certified potato seed tubers were planted and blight appeared early in the season in a pattern corresponding to the heavily infected areas in 1994. No volunteer potato plants were found in 1995 or 1996. Both mating types A1 and A2 were isolated in 1996 and 1997 and oospores were found in leaves, stems as well as in stolons.     

Variation in tuber pathogenicity ofPhytophthora infestans in the Netherlands

Summary Variation in aggressiveness to tubers among isolates ofPhytophthora infestans sampled from three potato growing regions in the Netherlands was compared. Variation in the ability to infect tubers of cv. Bintje was found between isolates of each of the three regional populations. The most aggressive isolate of the old population matched the average level of the new population in its ability to infect tubers. As a consequence, the commonly used reference isolate VK 6C can no longer be considered to be representative of the present population ofP. infestans. Therefore it is recommended that testing tuber resistance for the official list of potato cultivars with this isolate should be discontinued. Tuber infection and subsequent spread of the fungus in the tuber tissues were not found to be correlated. The components of tuber pathogenicity studied were not correlated to pathogenicity factors in the foliage, as measured under growth chamber conditions.     

Race diversity and virulence complexity ofPhytophthora infestans in Nepal

Summary A total of 251Phytophthora infestans isolates collected during 1999–2000 potato growing seasons from nine potato growing areas of Nepal were tested for virulence using a set of eleven potato differentials. The virulence test revealed the presence of 30 races with the most frequent being 4.11, 4 and 11. Twenty-nine percent of isolates belonged to simple races having none to one virulence factor, whereas 71% had complex virulence factors governed by 2 to 7 genes. Gleason and Shannon indices of 5.24 and 2.59, respectively, showed a moderate level of diversity in NepaleseP. infestans populations. Shannon indices comparison among regional populations revealed significant differences (t-test, P<0.001) in the diversity. The population from hills were the most diverse followed byterai and the high hill populations. The virulence complexities were minimal in Nepalese populations. The implications of these findings in association with the potato cultivation system in Nepal are discussed.     

Changes in race structure of Canadian populations ofPhytophthora infestans based on specific virulence to selected potato clones

Summary A collection of 80 isolates ofPhytophthora infestans, representing multilocus genotypes of the pathogen found in Canada from 1994 to 1996, was screened on a series of nine differential potato host cultivars to determine the physiological race of the isolates. A total of 28 pathotypes were found among the 11 genotypes present in the collection. There was a significant increase in the complexity of pathotypes from 1994 to 1996 which reflected the displacement of the US-1 genotype (mean of 2.2 host differentials infected) by the US-8 genotype (mean of 8.1 host differentials infected). Eighteen of the US-8 isolates could overcome all nine of the resistance genes tested. Virulence genes which overcame host resistance conferred by R8 were the rarest in the populations studied. The predominance of complex populations ofP. infestans in Canada implies that the introduction of cultivars with specific R genes for resistance would be an ineffective control measure. AAFC Contribution No. 901     

Novel genotypes ofPhytophthora infestans in Canada during 1994 and 1995

Populations ofPhytophthora infestans, causing late blight of potato, changed significantly in Canada in the mid-1990s. The US-1 genotype (A1, metalaxylsensitive [MS]) was displaced by novel genotypes (mainly US-8) in much of the country in 1994 and 1995. Many of the new genotypes were insensitive to metalaxyl and were of the A2 mating type, although metalaxyl-insensitive (MI) Al isolates of the gll (or US-11) genotype were found in British Columbia. A total of 14 unique multilocus genotypes were described over the two years of the study based on mating type (Al or A2), metalaxyl sensitivity (MS or MI), allozyme banding patterns (Gpi andPep), and random amplified polymorphic DNA (RAPD) markers. Six of these genotypes could be distinguished by allozyme banding patterns at theGpi locus alone; RAPD analysis revealed additional variation within allozyme groupings and defined additional genotypes. Among 82 isolates collected from various Canadian provinces in 1994, US-1 and US-8 constituted 30% and 49% of isolates, respectively; other genotypes occurred in minor frequency. Among 50 isolates collected in 1995, US-1 and US-8 constituted 10% and 74%, respectively. Isolates of gll (US-11) were found in British Columbia in 1995. Diversity of genotypes was greater in 1994 (11 genotypes) than in 1995 (six genotypes). We hypothesize that sexual reproduction may have been responsible for generating some of the genetic variability inP. infestans populations, during a time period in which A2 isolates first migrated into many provinces in Canada.     

Metalaxyl-resistant strains ofPhytophthora infestans (Mont.) de bary in ireland

Summary Isolates ofPhytophthora infestans obtained from different sources were tested for pathogenicity on cv. Kerr's Pink after different fungicide treatments. All isolates were pathogenic on untreated Kerr's Pink. Disease development after inoculation with all isolates was effectively controlled by a mancozeb spray. One isolate was effectively controlled by metalaxyl. The remaining two isolates were resistant to all levels of metalaxyl (50–700μl/l). Similar results were obtained with cv. Home Guard. In vitro tests showed that metalaxyl-resistant strains grew faster on metalaxyl-containing medium than sensitive strains.

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Zusammenfassung Isolate vonPhytophthora infestans, von verschiedenen Herkünften, wurden auf Resistenz gegen Metalaxyl geprüft. Eingetopfte Pflanzen (Sorte Kerr's Pink) wurden mit den gewünschten Konzentrationen von Metalaxyl (50–700μl/l), Mancozeb (6000 μl/l) oder Wasser besprüht. 24 Stunden nach der Fungizidan wendung wurden alle Pflanzen mit einer Zoosporensuspension der Isolate 1, 2 und 3 vonPhytophthora infestans inokuliert. 7 und 10 Tage nach der Inokulation wurde der Befall der Pflanzen bestimmt. Isolat I wurde durch Metalaxyl wirkungsvoll bek?mpft. Sieben Tage nach der Inokulation mit den Isolaten 2 und 3 zeigten die mit Metalaxyl behandelten Pflanzen einen geringeren Befallals die unbehandelten Kontrollen (Tabelle 1). 10 Tage nach der Inokulation war jedoch kein signifikanter Unterschied zwischen den Kontrollen und den mit Metalaxyl behandelten Pflanzen (Tabelle 1). Die Behandlung mit Mancozeb hemmte effektiv den Befall nach Inokulation mit allen Isolaten. Gleiche Ergebnisse wurden mit der Sorte Home Guard erhalten (Tabelle 2). In vitro Prüfungen zeigten, dass Metalaxyl resistente St?mme schneller auf Metalaxyl haltigem Medium wuchsen als sensible (Tabelle 3). Von diesen Mersuchen ausgehend wird gefolgert, dass Metalaxyl-resistente St?mme vonPhytophthora infestans vorhanden waren und sie für den Misserfolg dieses Fungizids, die Krautf?ule in Irland 1980 zu bek?mpfen, verantwortlich gewesen sein k?nnen.

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Résumé Des souches dePhytophthora infestans provenant de différentes sources ont été examinées pour leur résistance au métalaxyl. Des plantes en pots (var. Kerr's Pink) ont été pulvérisées à différentes concentrations de métalaxyl (50–700 μl/l), de mancozèbe (6000μl/l) ou d'eau. 24 heures après l'application de fongicide, toutes les plantes ont été inoculées avec une suspension de spores provenant de souches 1, 2 et 3 dePhytophthora infestans. Les plantes ont été examinées, en ce qui concerne l'incidence de la maladie, 7 et 10 jours après inoculation. La souche 1 était effectivement contr?lée par le métalaxyl. 7 jours après inoculation, les souches 2 et 3 présentaient sur les plantes traitées au métalaxyl un niveau de maladie plus bas que sur celles non traitées (tableau 1). Cependant, dix jours après inoculation, il n'y a avait aucune différence significative entre les plantes non tra?tées au métalaxyl lorsque l'inoculation était réalisée avec les souches 2 et 3 (tableau 1). Le développement de la maladie, après inoculation avec toutes les souches était effectivement contr?lé par le traitement au mancozèbe. Des résultats identiques ont été obtenus avec le cultivar Home Guard (tableau 2). In vitro, les tests montraient que les races résistantes au métalaxyl se développaient plus vite que les races sensibles, dans des milieux contenant du métalaxyl (tableau 3). Il ressort de ces expérimentations qu'il existe des races dePhytophthora infestans résistantes au métalaxyl et qu'elles peuvent être responsables des échecs enregistrés dans le contr?le du mildiou en Irlande en 1980.

     

Occurrence of the A2 mating type ofPhytophthora infestans in India

Summary One hundred and four isolates ofPhytophthora infestans were collected during 1990–93 from different potato growing regions of India. They were paired with mating type A1 ofP. capsici to determine their mating type. All isolates from the Indo-Gangetic plains and Assam were the A1 mating type, whereas a mixed population of A1 and A2 types occurred at Shillong, Darjeeling and Shimla hills. The frequency of A2 isolates was 5–30% in the Shimla hills and 60% and 58% respectively in the Shillong and Darjeeling hills.     

Evaluation of wildSolanum species for resistance to the US-8 genotype ofPhytophthora infestans utilizing a fine-screening technique

Greenhouse assays were carried out to characterizeSolanum accessions previously reported to be late blight resistant and to identify individuals within the accessions with high levels of resistance to late blight. Foliage of wildSolanum species accessions or hybrid cultivated potato x wild species (diploid or tetraploid) from Mexico, Russia, or South America was inoculated withPhytophthora infestans (US-8) and was scored for severity of foliar symptoms at 7 and 14 days after inoculation (DAI). Mexican accessions (S. brachycarpum,S. pinnatisectum,S. guerreroense, andS.fendleri) were significantly more resistant (1.1% infection at 14 DAI) than either Russian (59.1% infection at 14 DAI) or South American (53.4% infection at 14 DAI) accessions. Moreover, the genotypes within the Mexican accessions tended to be more uniformly resistant whereas genotypes within the Russian and South American accessions tended to segregate for resistance. The more resistant genotypes of Russian and South American accessions (176/546) were retested, and 56 genotypes were identified as having potential for use in potato breeding programs for resistance to late blight. Fifty percent of the selected genotypes were fromS. microdontum accessions PI498124, PI595510, and PI595511. The potential of these species for incorporation into breeding programs for late blight resistance is discussed.