SSR assessment of Phytophthora infestans populations on tomato and potato in British gardens demonstrates high diversity but no evidence for host specialization

Phytophthora infestans populations can differ in composition as a result of host specialization on tomato and potato hosts. In Great Britain many amateur gardeners grow outdoor tomatoes but there is little or no commercial tomato production outdoors. This study analysed isolates of P. infestans from British gardens with 12 multiplexed simple sequence repeat markers that are used to monitor the disease on commercial potato crops. Samples of P. infestans from tomato hosts were collected in 3 years and from potato in 1 year from across Great Britain. Seven previously unreported clonal lineages were detected in garden populations and higher frequencies of unique clonal lineages (28–40%) were present compared with populations from British commercial potato crops reported elsewhere. Garden populations had a lower proportion (11–48% less) of the most common lineages (13_A2 and 6_A1) that together made up at least 86% of the commercial potato populations during the sampling period. Host species accounted for only 2·0% of molecular variance detected between garden potato‐ and tomato‐hosted samples. No significant difference in clonal lineage composition was found between host species in Great Britain and this could be due to the whole P. infestans population overwintering on potato. British garden populations on both hosts were much more diverse than those on commercial potato crops; this finding may be influenced by less frequent fungicide use by gardeners and a higher diversity of unsprayed susceptible potato cultivars, enabling metalaxyl‐sensitive and less aggressive genotypes to survive in gardens.     

Distribution of major clonal lineages EU_13_A2, EU_2_A1, and EU_23_A1 of Phytophthora infestans associated with potato late blight across crop seasons and regions in Algeria

Potato is one of the most important agricultural crops in Algeria and worldwide. Each year, potato late blight, caused by Phytophthora infestans, is responsible for significant damage that leads to large production losses, and is thus a direct threat to food security in Algeria. In this study, 131 isolates of P. infestans and 92 DNA fingerprints captured on FTA cards were sampled from commercial and seed production fields in three major potato production regions (western, eastern, and central) during the main-season and late-season in Algeria over six cropping seasons (2010–2016). Genotypes of P. infestans and population genetic diversity were analysed using a 17-plex simple-sequence repeat (SSR) marker assay, and the mating type of all isolates was characterized. Both mating types (A1 and A2) were found, and often occurred in the same field. Differences in mating type proportion were observed between regions and between sampling periods. Analysis with SSR markers showed the prevalence of the EU_13_A2 lineage (70%) over EU_2_A1 (16%), EU_23_A1 (10%), and 4% of unknown multilocus lineage (MLL). The EU_13_A2 showed differentiation within the group. EU_23_A1 was found mainly in late-season crops. However, the cropping region did not influence the distribution of lineages due to the dispersal of the pathogen in Algeria by seeds. Genetic structure did not reveal a clear variation in distribution of the three lineages throughout the sampling regions. These data provide important new information on the composition and change over time of P. infestans populations in Algeria and open the way for a better understanding of the local epidemiology of this important pathogen.     

Genotypic and phenotypic characterization of Phytophthora infestans in South Korea during 2009–2016 reveals clonal reproduction and absence of EU_13_A2 genotype

In order to better understand the Phytophthora infestans population structure in South Korea, 172 isolates were collected between 2009 and 2016 from four major potato cultivation areas. Fungicide (metalaxyl and dimethomorph) response, mating type, and microsatellite (SSR) genetic fingerprints were analysed to characterize these isolates. Ten isolates collected in Gyeongnam Province, which specializes in protected winter cultivation in polytunnels, were A2 mating type. All other isolates were A1 mating type. Overall, 42% of the isolates were resistant to metalaxyl, and 43% were sensitive. All isolates were sensitive to dimethomorph. From the SSR fingerprints, 45 distinct genotypes were identified, which could be clustered into four clonal lineages: KR_1_A1, KR_2_A2, SIB-1, and US-11. KR_1_A1 was the predominant P. infestans genotype in South Korea. KR_2_A2 was only found in Gyeongnam Province; all isolates were A2 mating type and resistant to metalaxyl. SIB-1 was dominant until 2013 but its frequency has gradually decreased in more recent years. US-11 was first found in 2014, after which its frequency has increased to become codominant with KR_1_A1. The calculated standardized index of association (I_A) suggests that the South Korean P. infestans population is undergoing clonal reproduction. When compared with populations of P. infestans from the Netherlands, it has less genetic diversity and the dominant Netherlands P. infestans genotype, EU_13_A2 (Blue_13), was not found in South Korea. Such monitoring of the pathogen population contributes to a more efficient integrated pest management-based control strategy for potato late blight control in South Korea.     

Severe outbreaks of late blight on potato and tomato in South India caused by recent changes in the Phytophthora infestans population

Late blight, caused by Phytophthora infestans, has emerged as the most destructive disease of potato and tomato in South India since 2008. One hundred and fifty‐seven isolates of Phytophthora infestans, 63 from potato and 94 from tomato, were collected from major potato and tomato production areas of South India between 2010 and 2012. Their phenotypic and genotypic characteristics were determined and compared with reference isolates. Isolates were characterized based on mating type, in vitro metalaxyl sensitivity, mitochondrial DNA haplotype, RG57 DNA fingerprinting patterns, SSR markers and aggressiveness on potato and tomato, in order to monitor population changes in P. infestans. All isolates were A2 mating type, metalaxyl resistant, mtDNA haplotype Ia and had RG57 and SSR fingerprints almost identical to the 13_A2 clonal lineage reported in Europe. Variation at the D13 and SSR4 loci allowed discrimination of minor variants, designated as 13_A2_3, 13_A2_3b, 13_A2_3c and 13_A2_1. A comparison of the lesion diameters caused by 157 isolates on detached leaflets of three potato and tomato cultivars showed all isolates to be equally aggressive, confirming that the same clonal population is infecting both hosts. This study demonstrates that the 13_A2 lineage was responsible for severe late blight outbreaks on potato and tomato in South India and has replaced the prior population represented by the US‐1 and other genotypes. Revised management strategies will be required to combat this destructive 13_A2 clonal lineage and monitoring of the population across other potato‐ and tomato‐growing regions of India is warranted.     

Reduced efficacy of fluazinam against <Emphasis FontCategory="SansSerif" Type="Italic">Phytophthora infestans</Emphasis> in the Netherlands

Phytophthora infestans is the causal organism of potato late blight, the most important disease in potato, the second most important arable crop in Europe. The P. infestans population in Europe is well known for its sudden changes in composition. Currently it is composed of a wide variety of genotypes, some of which are dominant clonal lines while others are rare or even unique to a year or location. Fungicides play a crucial role in the integrated control of late blight. Since its introduction in the Netherlands in 1992, fluazinam has been used in late blight control strategies in ware and starch potatoes. It has a broad spectrum of activity and is effective against a range of diseases including potato late blight. Fluazinam interrupts the pathogen cell’s energy production process by an uncoupling effect on oxidative phosphorylation. It is considered to have a low resistance risk. Until recently, reduced efficacy against fluazinam was not detected in P. infestans surveys in Europe. In this paper we present the finding of a new clonal lineage (EU_33_A2) of P. infestans in the Netherlands and the reduced efficacy of fluazinam to control one of the EU_33_A2 isolates in field experiments carried out in 2011 and 2015 under high disease pressure. The potential effects of this finding on practical late blight control strategies are discussed.     

Population structure of Phytophthora infestans collected on potato and tomato in Italy

Late blight caused by the oomycete Phytophthora infestans is a disease of potato and tomato of worldwide relevance and is widespread throughout Europe and the Mediterranean region. While pathogen populations in northern Europe have been sampled and characterized for many years, the genetic structure of populations from southern Europe, including Italy, has been less studied. Between 2018 and 2019, we collected 91 samples of P. infestans from potato and tomato crops in Italy, Algeria, and Tunisia on FTA cards and genotyped them using 12-plex microsatellites. These samples were compared to genotypes of P. infestans previously collected within the framework of the EuroBlight network and from published sources. Four clonal lineages were identified: 13_A2 (Blue 13), 2_A1, 23_A1, and 36_A2. Two other isolates collected could not be matched to any currently known clonal lineage. The 13_A2 and 36_A2 lineages were found exclusively in southern Italy and Algeria, while 2_A1 was only found in Algeria. This is the first report of the 36_A2 lineage in Italy. Two isolates from Solanum nigrum were 13_A2, suggesting this weed host could be a reservoir of inoculum. The 23_A1 lineage was found widely on infected tomato crops in Italy and is the same as the lineage US-23 that is widespread in North America. Differences in genotypes across the country suggests that there may be different sources of introduction into Italy, possibly via infected seed tubers from other countries in Europe, tubers for consumption from North Africa, or tomatoes.     

Population structure of Phytophthora infestans in China – geographic clusters and presence of the EU genotype Blue_13

The population structure of Phytophthora infestans in China was studied and three mitochondrial haplotypes (Ia, IIa, IIb) were observed. Genetic analysis with 10 highly informative SSR markers identified 68 different genotypes, including three dominant clonal lineages. In the Chinese P. infestans population, the genotypes were strongly clustered according to their geographic origin. One of dominant clonal lineages was genetically similar to Blue_13, a dominant clonal lineage found in Europe since 2004. This is the first report of Blue_13 outside Europe. Only one mating type (A1) was found in the northern and southeastern provinces, but in southern and northwestern China both mating types were observed. The mating type ratio and SSR allele frequencies indicate that in China the sexual cycle of P. infestans is rare. These results emphasize that the migration of asexual propagules and the generation of subclonal variation are the dominant driving factors of the population structure of P. infestans in China. They may also have implications for the role of monitoring P. infestans populations in potato late blight management strategies in China.     

Genotypic and phenotypic characterization of Phytophthora infestans populations on Java,Indonesia

Phytophthora infestans is endemic to Indonesia and can infect potato crops at any stage in the growing season. Little is known about P. infestans populations in Indonesia. The objectives of this study were first to identify the genotypes causing late blight in the main potato-growing regions on Java in Indonesia, and secondly to examine genotypic diversity in the P. infestans populations in those regions. Samples were collected on FTA cards (n = 140) or in tubers (n = 6) from 15 locations in nine regencies over four years (2016–2019). Microsatellite analysis revealed that late blight outbreaks in these regencies were caused by EU_2_A1 and other genotypes that are unique to Indonesia. Eighty percent of the samples that amplified with CAPS markers were the A1 mating type. Cultures of six isolates were determined to be the A1 mating type based on the pairing test, and of these, two isolates were intermediate and four were sensitive to metalaxyl-M (mefenoxam). The mode of reproduction of the P. infestans population on Java, Indonesia, was found to be clonal. However, as the sample size in this study was small, more isolates need to be tested to confirm this. Microsatellite analysis revealed that 90% of Indonesian samples had trisomic loci. A high number of multilocus genotypes (MLGs) were found in all nine regencies (131 MLGs out of 146 samples). Results indicate that there is ongoing polyploidization in these populations due to a high mutation rate and no selection pressure from the susceptible potato hosts that are being grown in Indonesia.     

Population structure of Phytophthora infestans in Turkey reveals expansion and spread of dominant clonal lineages and virulence

Late blight, caused by the oomycete Phytophthora infestans, is considered the most important and destructive disease of potato in Turkey. In this study, characterization of 367 isolates of P. infestans obtained from the potato-growing areas of the country was carried out to evaluate the pathogen population structure over the 2017–2019 production seasons. The isolates were characterized by numerous features including mating type, in vitro mefenoxam sensitivity, simple-sequence repeat (SSR) markers, and virulence against a set of potato differential lines. Most isolates were A2 mating type (353 isolates). Also, 68% of isolates were resistant to mefenoxam; the remainder were intermediate in their sensitivity and there were no sensitive isolates. SSR-based genotypic analysis of P. infestans populations showed a low genetic diversity. The 13_A2 clonal lineage predominated with a frequency of 92.1%, followed by 34_A1 (3.3%) and 37_A2 (2.7%). Genotypes 34_A1 and 37_A2 were detected only in 2019. This is the first report of 34_A1 and 37_A2 clonal lineages causing late blight disease of potato in Turkey. The most abundant virulence type was one overcoming resistance genes R1, R2, R3, R4, R6, R7, R10, and R11. These results emphasized that the migration of individuals and the asexual generation of subclonal differences were the main factors driving the population structure of P. infestans in Turkey.     

Phenotypic and genotypic changes in French populations of Phytophthora infestans: are invasive clones the most aggressive?

The main objective of this investigation was to test the ‘always more aggressive’ hypothesis, often advocated to explain lineage replacements in clonal populations of the potato late blight oomycete Phytophthora infestans. To this end, genotypic and pathogenicity data on 1274 French isolates of P. infestans, collected over the period 2001–2008, were analysed. Overall, the populations sampled showed limited genetic diversity, with four multilocus lineages (1_A1, 2_A1, 8_A1 and 13_A2) accounting for over 80% of the isolates collected. As in other West European countries, drastic changes in these dominant clonal lineages were observed over the course of the 8 years, particularly in the appearance and propagation of the clone 13_A2. However, invasiveness of clones was not associated with higher aggressiveness; on the contrary, dominant clones had generally low or moderate aggressiveness relative to others present at the same time within the same populations. This finding challenges the link between invasive behaviour and increased aggressiveness often assumed for this biotrophic pathogen, and could reflect the existence of a trade‐off between intra‐season and inter‐season transmission. This would be consistent with the concept of inclusive fitness, which involves the abilities to both reproduce and survive.     

Large subclonal variation in Phytophthora infestans populations associated with Ecuadorian potato landraces

The population of Phytophthora infestans on potato landraces in three provinces (Carchi, Chimborazo and Loja) of Ecuador was analysed. All isolates (n = 66) were of the A1 mating type. Simple sequence repeats (SSR) were used to assess the genetic diversity of the isolates. The P. infestans isolates from the potato landraces grouped in a single clade together with reference isolates belonging to the clonal lineage EC‐1. In the 66 SSR profiles obtained, 31 multilocus genotypes were identified. The 66 isolates constituted 49 different races according to the Solanum demissum differential set ( R1 to R11). The P. infestans population was complex and virulent on 4 to 11 R genes. Analysis showed that the subclonal variation in the Ecuadorian EC‐1 clone is increasing over time and is much larger than clonal variation in lineages in the Netherlands and Nicaragua, suggesting high mutation rates and little or no selection in Ecuador.     

Population genetics of Phytophthora infestans in Denmark reveals dominantly clonal populations and specific alleles linked to metalaxyl‐M resistance

Control of the potato late blight pathogen Phytophthora infestans relies heavily on chemicals. The fungicide metalaxyl‐M (Mefenoxam) has played an important role in controlling the disease, but insensitivity to the fungicide in certain isolates is now of major concern. A genetic basis for resistance to metalaxyl suggests the possibility for linking resistance phenotypes to specific population genetic markers, but in order to do this, the population genetic structure and mode of reproduction in a population must first be well described. The dynamics of metalaxyl‐M resistance in the Danish population of P. infestans was characterized over the course of the 2013 growing season, as was the population genetic structure, using simple sequence repeat (SSR) genotypes and single nucleotide polymorphism (SNP)‐based mitochondrial haplotyping of over 80 isolates. Both mating types A1 and A2 were present in most fields, but tests for recombination showed that clonal reproduction dominates in Danish populations. Genotype was not linked to haplotype and no differentiation was observed at the haplotype level, but rather between fields. Resistance phenotypes were linked to specific SSR alleles, demonstrating the potential for a more precise SNP‐based marker system for predicting resistance to metalaxyl‐M.     

Population structure and host range of the potato late blight pathogen Phytophthora infestans in Peru spanning two decades

The genetic diversity of the late blight pathogen Phytophthora infestans infecting cultivated potato and alternative hosts growing in the vicinity of fields in the main potato-growing areas of the Peruvian Andes was characterized using collections from 1997–2013 as reference. The Peruvian P. infestans population, including previously collected and current isolates, consists of four clonal lineages (EC-1, US-1, PE-3 and PE-7) that belong to the A1 mating type and have been present in the country for decades. The first report of US-1 was in isolates collected between 1982 and 1986; meanwhile, EC-1 and PE-3 appeared for the first time in isolates from 1992 and PE-7 was found in 1997. The pathogen has a very broad host range among the solanaceous plants infecting cultivated potato, tomato, pear melon and several wild species. The solanaceous species growing in the vicinity of the potato fields sampled were identified and surveyed for late blight-like symptoms. Phytophthora infestans was isolated from nine wild species, including three new host species: Solanum zahlbruckneri, Solanum grandidentatum and Iochroma grandiflorum. There was no clear host specialization, but geographical substructuring was found as well as changes in the pathogen populations at the regional level. The clonal lineage EC-1, which is mostly resistant to metalaxyl, has complex virulence and contains a high level of subclonal variation, continues to dominate the population. Some multilocus genotypes of the EC-1 lineage were sampled in high frequencies and were found among the previously collected and new samples.     

Dominance of a single clonal lineage in the Phytophthora infestans population from northern Shaanxi,China revealed by genetic and phenotypic diversity analysis

Late blight caused by Phytophthora infestans is the most serious disease of potato worldwide. To understand the P. infestans population structure in northern Shaanxi, an emerging potato production region in China, 125 single‐lesion isolates were randomly collected from farmers' fields in 2009 and characterized phenotypically and genotypically. A mating type assay showed that 94 isolates were A1 mating type. Virulence determination of selected isolates on a set of differential potato lines containing R1 to R11, respectively, showed the presence of two pathotypes, of which the pathotype lacking avirulence genes Avr3, Avr4 and Avr10 was dominant. Isolates lacking all avirulence factors Avr1 to Avr11 were detected but at lower frequency (13·6%). Analysis for mtDNA haplotype showed all 61 examined isolates were IIa. A total of seven multilocus genotypes were distinguished among 125 isolates, as determined with seven polymorphic microsatellite markers. The genotype SG‐1 was dominant in the population with a frequency of 75·2% and was present throughout the region. Analysis of the phenotypic and genotypic structures of P. infestans populations indicated strict clonal reproduction of the pathogen and suggested that sexual reproduction probably does not occur. Potential implications for disease management are discussed.     

Pineapple heart rot isolates from Ecuador reveal a new genotype of Phytophthora nicotianae

Pineapple heart rot disease, caused by Phytophthora nicotianae (syn. P. parasitica), is responsible for significant annual reductions in crop yield due to plant mortality. In Ecuador, new infections arise during the rainy season and increase production costs due to the need for biocontrol and fungicide applications. Studies of P. nicotianae population structure suggest that certain genetic groups are associated with host genera; however, it is not clear how many host‐specific lineages of the pathogen exist or how they are related. The objectives of this study were to determine the level of genetic variation in the P. nicotianae population causing heart rot disease of pineapple in Ecuador and compare the genotypes found on pineapple to those previously reported from citrus, tobacco and ornamentals. Thirty P. nicotianae isolates collected from infected pineapple leaves from four farms were genotyped using nine simple sequence repeat loci. In addition, the DNA sequences of mitochondrial loci cox2 + spacer and trnG‐rns were analysed. Together, these loci supported a single clonal lineage with two multilocus genotypes differing in a single allele and low mitochondrial diversity. This lineage was distinct but closely related to isolates collected from vegetables and ornamentals in Italy. The results support the hypothesis of host specialization of P. nicotianae in intensive cropping systems and contribute to the understanding of population structure of this important pathogen.     

Population genetic analysis of Phytophthora infestans in northwestern China

Late blight caused by Phytophthora infestans is the most devastating disease of potato worldwide. To understand the P. infestans population structure and dynamics in northwestern China, 959 single‐lesion isolates were purified in three consecutive years (2009–2011) and were characterized for mating type, pathotype, mtDNA haplotype and molecular variation at eight SSR loci. The results showed that the distribution of mating types changed significantly over years, with self‐fertile isolates dominant in 2010 and 2011. SSR genotyping distinguished 959 isolates into 151 genotypes, and association analysis indicated that P. infestans populations in 2010 and 2011 were strictly asexual while in 2009 they showed signs of sexual reproduction. Population analysis showed that the majority of genetic variation was within P. infestans populations. Isolates sharing identical SSR genotypes were detected in distant regions, indicating that migration of P. infestans could have occurred between regions. Pathogenicity assays on a set of potato differential lines containing R1 to R11 resistance genes detected four pathotypes from 74 selected isolates, with the pathotype virulent against all 11 R genes being dominant. Three mtDNA haplotypes (Ia, IIa, IIb) were detected with Ia being dominant among 507 isolates examined. Phylogenetic analysis indicated that P. infestans populations in northwestern China are distant from European lineages including 13‐A2 (blue‐13) at the time of this survey. The results have implications for the trade of healthy seed tubers as a means of managing late blight.     

Molecular identification of a 16SrIII-B phytoplasma associated with cassava witches’ broom disease

Since its arrival in the British Isles in 1845 Phytophthora infestans has remained the most destructive pathogen of potato. In the ensuing period, the British and Irish P. infestans populations have undergone major displacements following the immigration of novel strains. Here we report the re-emergence of the Ib mitochondrial DNA haplotype in the British and Irish P. infestans populations associated with the 6_A1 genotype. Historically associated with the previously panglobally distributed clonal lineage US-1, the Ib haplotype has not been detected (with the exception of a single isolate in the mid 1990s) in the British or Irish P. infestans populations since the early 1980s. The 6_A1 isolates analysed possessed mtDNA Ib, but were otherwise quite unlike US-1, having the Pep allozyme genotype 96/96 and novel RG57 and SSR fingerprints. These genetic characteristics strongly suggest that the appearance of the 6_A1 genotype in these populations has resulted from migration (possibly after a recombination event elsewhere). This study highlights the advantages of utilising a range of different markers in pathogen monitoring.     

Diversity of populations of Phytophthora infestans in relation to patterns of potato crop management in Latvia and Lithuania

Potato crop losses can be substantial when conditions for late blight (Phytophthora infestans) development and spread are favourable. In this study, drivers of differences between the P. infestans population structures in Latvia and Lithuania, two neighbouring countries with similar potato-growing traditions, were investigated. Genotypes of P. infestans and population genetic diversity were analysed using a 12-plex simple sequence repeat (SSR) marker assay. High genetic diversity was demonstrated in both populations, with population diversity being higher in Latvia. It would appear that local populations established from soilborne oospores early in the season are well adapted to the conditions in the region. However, somewhat greater spread and survival of local clones was detected in Lithuania, suggesting that potato cropping there is more vulnerable to clonal invasion than in Latvia. For effective disease management, current strategies should be adjusted according to the specific pathogen populations in the region, considering the reproduction and survival of the pathogen. Potato growers should implement late blight preventive measures such as longer field rotation to prevent oospore infections, especially in Latvia, and should use more disease resistant cultivars and high-quality seed potatoes.     

What is the evidence for sexual reproduction of Phytophthora infestans in Europe?

The biology of late blight of potato and tomato, caused by Phytophthora infestans, changed when sexual reproduction by the pathogen became possible in many parts of the world, including Europe. In northern Europe, especially Scandinavia, there is increasing evidence that the pathogen is reproducing sexually on a regular basis, although in other regions further south or to the west it appears to reproduce primarily in a clonal manner. The presence of both mating types, the production of viable oospores, and observations of fields with soilborne sources of inoculum are consistent with sexual reproduction. Studies with different marker systems have revealed a population structure without any dominating clonal lineages in Scandinavia, and that is most easily explained by sexual reproduction. Phytophthora infestans recovered from the soil can also be linked to parental genotypes using likelihood‐based methods when codominant markers are used. A synthesis of all the available data points to a second centre of sexual reproduction in northern Europe.     

Tracking Phytophthora infestans with SSR markers within and between seasons – a field study in Sweden

The dynamics of a late blight epidemic and sexual reproduction in Phytophthora infestans were studied in an experimental field in mid‐Sweden. The field was inoculated with six isolates of P. infestans taken from another potato field where sexual reproduction of the pathogen was suspected. Three weeks after inoculation single‐lesion leaflets were sampled and the resulting isolates characterized using microsatellites (SSRs) and mating type as markers. Among the 151 isolates analysed, the inoculum genotypes constituted more than 80% of the genotypes found, with three other genotypes making up the remainder. The following year, P. infestans obtained from soil samples taken from this field were analysed, and six novel genotypes were identified. Genotypes from the previous summer’s population were not detected. Analysis of the genotypes recovered was consistent with them being recombinants, with the previous summer’s population acting as parents. These findings are consistent with the hypothesis that oospores produced during a summer epidemic in Sweden can overwinter and cause infection the next year.