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.     

Comparison of different culture media on the mycelial growth, sporangia and oospore production ofPhytophthora infestans

The purpose of this study was to evaluate various culture media, with or without β-sitosterol, on mycelial growth, sporangia and oospore production by isolates ofPhytophthora infestans. Nine media were compared to rye agar (RA) which is frequently used to cultureP. infestans. Poor fungal growth across most isolates was observed on soybean agar, carrot agar and carrot agar with sterols relative to RA. Only clarified V8 juice containing β-sitosterol (CLV8S) resulted in significantly more sporangia production compared to RA. All other media performed similar to the RA. No significant differences were found in the sizes (length and width) of sporangia produced among the isolates tested on various media. Oospore production was greatest whenP. infestans was cultured on carrot agar with β-sitosterol (CAS) or CLV8S when compared to RA. Responses of isolate matings varied somewhat with an A2 isolate from Nova Scotia consistently producing fewer oospores in matings with A1 isolates compared to the other A1/A2 mating combinations. Rye agar and oat meal agar with or without β-sitosterol as well as clarified V8 juice agar with β-sitosterol are useful forin vitro growth of mycelium.     

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.     

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.     

Characterization and Distribution of Three New Clonal Lineages of Phytophthora infestans Causing Late Blight in Wisconsin from 2009 to 2012

Phytophthora infestans causes late blight of potato and tomato, a disease that has been estimated to cost U.S. potato growers $287.8 million annually. We collected isolates of P. infestans from Wisconsin from 2009 to 2012 and determined distribution of clonal lineages and mating types and sensitivity to the systemic fungicide mefenoxam. We also sought to evaluate the current utility of an analysis of the Glucose-6-phosphate isomerase (Gpi) allozyme locus for predicting mefenoxam sensitivity with the aim of delivering timely information to growers. Overall, 143 isolates were collected from 52 locations in 20 Wisconsin counties from 2009 to 2012. Three clonal lineages, US-22, US-23, and US-24, were identified and were novel to Wisconsin and the U.S. US-22 is of the A2 mating type and sensitive to mefenoxam, with Gpi 100/122. US-23 and US-24 are of the A1 mating type and primarily intermediately sensitive to mefenoxam, with Gpi 100/100 and 100/100/111, respectively. Because of this close correlation and the unique Gpi patterns for each lineage present, we were able to predict mefenoxam sensitivity directly from samples using the allozyme assay and quickly deliver management information to growers. Both mating types were present in Wisconsin in 2009 and 2010 but were spatially separated and no evidence of sexual recombination or soil persistence was detected. The presence of new clonal lineages of P. infestans in Wisconsin indicates a need for continued close monitoring of late blight to facilitate generation of timely information for enhanced short-term and long-term late blight management.     

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.     

Characterization ofPhytophthora infestans from Maine during 1999 and 2000

A total of 77 isolates ofPhytophthora infestans was collected in 1999 and 93 isolates were collected in 2000 from growers’ fields, home gardens, and research plots in Maine. Genotype diversity based onglucose-6-phosphate isomerase (Gpi) allozyme analysis was greater in the collections from 2000, with six genotypes identified vs two genotypes identified in 1999. The US-8 (Gpi 100/111/122) genotype constituted 88% of the isolates collected in 1999 and 75% of those collected in 2000. The US-14 (Gpi 100/122) genotype constituted 12% and 7% of the isolates collected in 1999 and 2000, respectively. Three newGpi genotypes constituted 7% of the isolates collected from potato in northern Maine in 2000. A new genotype (A1 mating type,Gpi 100/100) that was aggressive on both tomato and potato was identified from home gardens in western Maine and constituted 11% of the isolates collected in 2000. A1, A2, and A1A2 mating types as well as self-fertile isolates were collected in both years. Self-fertility in artificial culture appeared to be relatively common in field isolatesof P. infestans. Potential origins of the new genotypes are discussed.     

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.     

Characterisation of Phytophthora infestans Isolates Collected from Potato and Tomato Crops in Tunisia During 2006–2008

Severe late blight epidemics in Tunisia in recent years prompted population studies on the pathogen responsible for this disease, Phythophthora infestans. Characterisation of 165 Tunisian P. infestans isolates collected from 2006 to 2008 was performed for the mating type and mt haplotype, while subsets were analysed for metalaxyl sensitivity (n?=?65), virulence on differential set of 11 R genes of Solanum demissum (n?=?31), aggressiveness on cv. Bintje (n?=?36) and measurement of the radial growth on agar medium at three temperatures (n?=?38). Most isolates from potato and all isolates from tomato had the A1 mating type. The A2 mating type was detected in the north-east and northern areas, but not in the north-west. All the A2 mating type isolates were metalaxyl resistant and seem to be part of a new generation of the P. infestans isolates which are more aggressive, with more complex races, and tolerant to higher temperatures. The increased severity of epidemics during 2006 to 2008 can be attributed to favourable weather conditions during growing seasons, adaptation of new genotypes, widespread phenylamide resistance in potato production regions and most probably incorrect spray programmes. In contrast to the presence of complex pathotypes in two major potato crop regions (north-east and northern areas), the P. infestans population detected in the other regions and in tomato crops was still relatively simple. Compared with the situation in Europe and the American continent, or even compared with neighbouring countries such as Algeria, the genetic changes in Tunisia are still comforting and require strict management decision on late blight control to avoid the spread of new P. infestans populations from Europe or neighbouring countries.     

Incidence of the A2 mating type ofPhytophthora infestans on potato crops in Northern Ireland

Summary Of 250 isolates ofPhytophthora infestans obtained from Northern Ireland potato crops between 1981 and 1993, only six proved to be of the A2 mating type, the remainder being A1. The first A2 isolate was obtained from a tuber in 1987. The frequency of A2 isolates from 1987 onwards was 3%, a lower incidence than has been reported for England and Wales and the Republic of Ireland. There was no association between phenylamide resistance and mating type.     

Identification of late blight, Colorado potato beetle, and blackleg resistance in three Mexican and two South American wild 2x (1EBN)Solanum species

Wild potatoes are important sources of genes for resistance to disease and insect pests. A collection of wild Mexican and South AmericanSolarium species from the US potato Genebank was evaluated under laboratory and/or field conditions for their reaction to late blight (Phytophthora infestans), Colorado potato beetle (CPB,Leptinotarsa decemlineata Say), and blackleg (Erwinia carotovora subsp.atroseptica (van Hall) Dye) in order to identify individual genotypes with multiple resistance genes. Late blight inoculations using aggressive isolates (US-8/A2 and US-11/A1 mating types) of P.infestans revealed a wide range of variation for resistance between and within the accessions of the wild species tested. For late blight, susceptible as well as moderately to highly resistant genotypes were observed in all the species tested. However, at least one accession from the three Mexican and one South American wild diploid species tested showed a relatively uniform high level of resistance toP. infestans. These includedS. bulbocastanum, S. pinnatisectum, S. cardiophyllum, andS. circaeifolium. Two accessions from South American speciesS. commersonii were highly susceptible to late blight. For the Colorado potato beetle test, only one species,S. pinnatisectum appeared uniformly resistant to CPB under field conditions. Results of screening for blackleg resistance showed that there were major differences between genotypes in the wild species. Accessions ofS. circaeifolium PI 498119 andS. bulbocastanum PI 243504 were identified as having significantly higher blackleg resistance than cultivated potato and the other wild species tested. However, genotypes from these two accessions were more susceptible to late blight and CPB. Characterization of theP. infestans isolate P1801C.16 used for late blight evaluation and multi-locus isolate tests using US-8/A2 and US-11/A1 races revealed that the resistance inS. pinnatisectum genotypes tested corresponded to a race-non-specific genetic system, which was different from any existing R genes.Solanum pin-natisectum genotypes with both high levels of late blight and CPB resistance as well as blackleg resistance genotypes identified in the present study represent a diverse gene pool that may be useful for development of new potato cultivars with multiple disease and insect resistance. The potential utilization of these valuable sources for improvement of cultivated potato is discussed.     

US-8 and US-11 genotypes ofPhytophthora infestans from potato and tomato respond differently to commercial fungicides

Isolates ofPhytophthora infestans collected in Canada in 1997 from both potatoes and tomatoes, were tested on potato leaf discs for their response to an equal active ingredient concentration (10 μg a.i./mL) of the following commercial fungicides: Acrobat MZ (Dimethomorph and Mancozeb), Ridomil Gold (metalaxyl-m), Dithane (Mancozeb), Curzate (Cymoxanil), Bravo (Chlorothalonil), and Tattoo C (Propamocarb and Chlorothalonil). Relative percent leaf infection values, estimated on fungicide-treated vs fungicide-free leaf discs, were compared among isolates from the US-8 and US-11 genotypes isolated from the two host plants. Based on an equal concentration of each fungicide’s active ingredients, variations in relative percent leaf infection were recorded between US-8 and US-11 genotypes, and between potato and tomato isolates within each genotype. Bravo and Tattoo C used with similar active ingredients concentrations were the most inhibitory to all groups of isolates. Dithane and Ridomil Gold provided uniform low inhibition againstP. infestans when tested on potato leaf discs. The different behavior ofP. infestans isolates from potato vs tomato suggests that management of late blight in these two important crops must take such differences into consideration. In particular, the nature and concentration of the fungicides to be applied must take into account any information available about genotypes present on each crop.     

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.     

Effects of hot and cold temperatures on the survival of oospores produced by United States strains ofPhytophthora infestans

Oospores produced from matings between US strains ofPhytophthora infestans were subjected to hot or cold temperatures and then evaluated for viability, based on percent germinationin vitro. Of 35 crosses, 3 produced oospores with sufficient germination for replicated assessments of temperature effects. Germination rates were as high as 26%, but the maximum germination percentage varied among crosses. Two additional crosses, one involving isolates from Mexico also had sufficient germination for assessment of temperature effects. Oospores did not germinate after 2 hr treatments at or above 46C, or after 12 hr treatments at 40C. Temperature treatment at -80C suppressed germination. Temperature had a similar effect on oospore germination from all crosses.     

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.     

Epidemiology and Integrated Control of Potato Late Blight in Europe

Phytophthora infestans, the causal agent of late blight, is a major threat to potato production in northwestern Europe. Before 1980, the worldwide population of P. infestans outside Mexico appeared to be asexual and to consist of a single clonal lineage of A1 mating type characterized by a single genotype. It is widely believed that new strains migrated into Europe in 1976 and that this led to subsequent population changes including the introduction of the A2 mating type. The population characteristics of recently collected isolates in NW Europe show a diverse population including both mating types, sexual reproduction and oospores, although differences are observed between regions. Although it is difficult to find direct evidence that new strains are more aggressive, there are several indications from experiments and field epidemics that the aggressiveness of P. infestans has increased in the past 20?years. The relative importance of the different primary inoculum sources and specific measures for reducing their role, such as covering dumps with plastic and preventing seed tubers from becoming infected, is described for the different regions. In NW Europe, varieties with greater resistance tend not to be grown on a large scale. From the grower??s perspective, the savings in fungicide input that can be achieved with these varieties are not compensated by the higher (perceived) risk of blight. Fungicides play a crucial role in the integrated control of late blight. The spray strategies in NW Europe and a table of the specific attributes of the most important fungicides in Europe are presented. The development and use of decision support systems (DSSs) in NW Europe are described. In The Netherlands, it is estimated that almost 40% of potato growers use recommendations based on commercially available DSS. In the Nordic countries, a new DSS concept with a fixed 7-day spray interval and a variable dose rate is being tested. In the UK, commercially available DSSs are used for c. 8% of the area. The validity of Smith Periods for the new population of P. infestans in the UK is currently being evaluated.     

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.     

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.     

Characterization of <Emphasis Type="Italic">Phytophthora infestans</Emphasis> Isolates from Jersey,Channel Islands

Potato production on the island of Jersey, in the English Channel, is dominated by Jersey Royal, a selection of the early cultivar International Kidney. Jersey Royal is very susceptible to Phytophthora infestans, the cause of potato late blight, and Jersey’s climate is frequently conducive to infection. During 2004–2006, isolates of P. infestans were obtained from Jersey Royal plant samples from 41 different sites (24 crops, one infected tuber, 16 volunteers) and from five outdoor tomato crops and characterised by mating type, mtDNA haplotype, Gpi and Pep allozyme genotype, metalaxyl resistance and RG57 fingerprint. A subset of 17 isolates was additionally characterised by simple sequence repeat (SSR). All but one isolate from potato belonged to a single A1 multilocus genotype or a variant. The five isolates from tomato represented three distinct genotypes, none of which was found on potato and included one which was A2 mating type. The populations of P. infestans on potato and tomato in Jersey appear distinct, with that on potato being highly clonal.