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.     

Genetic diversity and host differentiation among isolates of Phytophthora infestans from cultivated potato and wild solanaceous hosts in Peru

To test the hypothesis that isolates of Phytophthora infestans attacking wild Solanaceae exhibit specialization for particular host species, 115 isolates of P. infestans were collected from cultivated potatoes, nontuber-bearing Solanum spp. of the Basarthrum section and wild tomatoes from five departments in the northern and central highlands of Peru, and characterized using several neutral markers. All isolates belonged to one of four clonal lineages described previously in Peru: EC-1, US-1, PE-3 and PE-7. There was a strong association of three lineages with host species: PE-3 was only isolated from cultivated potato, while PE-7 and US-1 were only isolated from nontuber-bearing Solanum spp. ( Basarthrum section and wild tomatoes). EC-1 was isolated from all host groups sampled. A subset ( n  = 74) of the isolates was evaluated for metalaxyl resistance. High levels of resistance were found almost exclusively in EC-1 and PE-3, while US-1 and PE-7 isolates were generally sensitive. In a detached-leaf assay for lesion diameter using five EC-1 isolates from S. caripense and seven EC-1 isolates from cultivated potato, there was a significant interaction between isolate origin and inoculated host, caused by higher aggressiveness of EC-1 from cultivated potato on its host of origin. In a comparison of EC-1 (seven isolates from cultivated potato) and US-1 (three isolates from S. caripense ), each pathogen lineage was more aggressive on its original host species, causing a highly significant interaction between isolate origin and inoculated host. Wild tomatoes and nontuber-bearing Solanum spp. harbour several pathogen lineages in Peru and could serve as reservoirs of inoculum that might contribute to epidemics on potato or tomato. Potential risks associated with the use of wild Solanum hosts as sources of resistance to P. infestans are discussed .     

Host Specialization not Detected Among Isolates of the EC-1 Lineage of <Emphasis Type="Italic">Phytophthora infestans</Emphasis> Attacking Wild and Cultivated Potatoes in Peru

To determine whether populations of Phytophthora infestans attacking wild and cultivated potatoes in the highlands of Peru are specialized on their hosts of origin, we characterized isolates using several neutral markers, metalaxyl resistance and for aggressiveness in a detached leaf assay. One hundred and fifty-three isolates were collected from the northern and central highlands of Peru from different potato cultivars (both modern and native cultivars) and from different species of wild, tuber-bearing potatoes. All the isolates analyzed belonged to one of four clonal lineages that had been described previously in Peru: EC-1, US-1, PE-3 and PE-7. The EC-1 lineage (n = 133) was dominant and present in similar frequencies on wild and cultivated potatoes. PE-3 (n = 14) was found primarily on cultivated potatoes, with only one isolate coming from a wild host. US-1 (n = 2) and PE-7 (n = 4) were rare; all but one (PE-7) occurred on wild potatoes. Isolates from the EC-1 lineage from modern cultivars were compared in three separate detached leaf inoculation assays with EC-1 isolates from the wild potato species S. sogarandinum, S. bill-hookerii or S. huancabambense, respectively. No significant interactions between isolate type (from wild or cultivated potato) and host type (wild or cultivated) were measured for any assay. It appears that the pathogen genotypes in the EC-1 lineage indiscriminately attack both wild and cultivated tuber-bearing solanaceous hosts in Peru, and breeders should be able to select for resistance using the common EC-1 lineage.     

Potential of sexual reproduction among host-adapted populations of Phytophthora infestans sensu lato in Ecuador

To determine the potential of sexual reproduction among host-adapted populations of Phytophthora infestans sensu lato in Ecuador, 13 A1 isolates belonging to clonal lineages US-1, EC-1 and EC-3, and 11 A2 isolates belonging to the clonal lineage EC-2, were paired on agar plates to induce crossing. In the first experiment, six A1 isolates (three US-1, two EC-1 and one EC-3) were each crossed with three A2 isolates (total = 18 crosses). Matings involving isolates of the EC-1 lineage produced more oospores of healthy appearance than did matings with isolates of US-1 or EC-3. In the second experiment, the oospores of 35 crosses (21 EC-1 × EC-2; 10 US-1 × EC-2; four EC-3 × EC-2) were dispersed on water agar to assess oospore germination. Overall, germination percentages were low. Only one cross produced enough progeny for evaluation. Twenty-three single-oospore offspring were isolated and evaluated for mating type; electrophoretic patterns of glucose-6-phosphate isomerase ( Gpi ) and peptidase ( Pep ) alloenzyme loci; mitochondrial DNA haplotype; and genomic DNA fingerprint. Multilocus genotype data indicated that all 23 isolates resulted from meiotic recombination. Four progeny with homothallic phenotype appeared to be unstable heterokaryons. Markers at several loci segregated according to simple Mendelian expectations for a diploid organism, but the ratios of three RFLP loci and the Pep locus were not consistent with Mendelian expectations. All progeny were nonpathogenic on hosts of the parental genotypes. Reduced mating success and reduced pathogenic fitness of progeny appear to be postmating mechanisms of reproductive isolation in populations of P. infestans sensu lato in Ecuador.     

Loss of Production of the Elicitor Protein INF1 in the Clonal Lineage US-1 of Phytophthora infestans

ABSTRACT The extracellular protein INF1 of Phytophthora infestans is a member of the elicitin family of protein elicitors known to induce a hypersensitive response on some solanaceous and cruciferous plants. The presence of INF1 elicitin in culture filtrates of 102 P. infestans isolates from 15 countries was examined. All tested isolates produced INF1 except five isolates collected in 1976 and 1977 from infected potatoes in East Germany (the former German Democratic Republic). Based on hybridization to the multi-locus DNA fingerprint probe RG57, all the INF1-nonproducing isolates were shown to belong to the clonal lineage US-1 that dominated world populations until the 1980s. Phylogenetic analysis of a set of European US-1 isolates using amplified fragment length polymorphism fingerprint data indicated that loss of INF1 production evolved independently in separate lineages within US-1. DNA and RNA blot hybridizations showed that INF1-nonproducing isolates still retain a copy of the inf1 gene, whereas little inf1 mRNA could be detected. Hypothetical interpretations of the evolution in a restricted geographic area of P. infestans lineages deficient in the production of a specific elicitor protein are discussed.     

Simulation of Potato Late Blight in the Andes. I: Modification and Parameterization of the LATEBLIGHT Model

ABSTRACT LATEBLIGHT, a mathematical model that simulates the effects of weather, host growth and resistance, and fungicide use on asexual development and growth of Phytophthora infestans on potato foliage, was modified so that it can be used in the Andes and, eventually, worldwide. The modifications included (i) the incorporation of improved equations for the effect of temperature on lesion growth rate (LGR) and sporulation rate (SR); (ii) the incorporation of temperature-dependent latent period (LP); and (iii) the use of experimentally measured parameters of LGR, SR, and LP for specific potato cultivars and pathogen lineages. The model was parameterized for three Peruvian potato cultivars (Tomasa, Yungay, and Amarilis) infected with isolates of a new clonal lineage of P. infestans that is currently predominant in Ecuador and Peru (EC-1).     

Host Specificity of Phytophthora infestans on Tomato and Potato in Ecuador

ABSTRACT Sixty Ecuadorian isolates of Phytophthora infestans from potato and 60 isolates from tomato were compared for dilocus allozyme genotype, mitochondrial DNA haplotype, mating type, and specific virulence on 11 potato R-gene differential plants and four tomato cultivars, two of which contained different Ph genes. Restriction fragment length polymorphism (RFLP) fingerprints of subsamples of isolates from each host were compared by using RG57 as the probe. All potato isolates had the allozyme genotype, haplotype, and mating type of the clonal lineage EC-1, which had been previously described in Ecuador. With the same markers, only one isolate from tomato was classified as EC-1; all others belonged to the globally distributed US-1 clonal lineage. RFLP fingerprints of isolate subsets corroborated this clonal lineage classification. Specific virulence on potato differentials was broadest among potato isolates, while specific virulence on tomato cultivars was broadest among tomato isolates. Some tomato isolates infected all tomato differentials but no potato differentials, indicating that specific virulence for the two hosts is probably controlled by different avirulence genes in P. infestans. In two separate experiments, the diameters of lesions caused by nine isolates from potato and 10 from tomato were compared on three tomato and three potato cultivars. All isolates produced larger lesions on the host from which they were isolated. No isolates were found that were highly aggressive on both tomato and potato. We conclude that there are two different populations of P. infestans in Ecuador and that they are separated by host.     

Sensitivity to Protectant Fungicides and Pathogenic Fitness of Clonal Lineages of Phytophthora infestans in the United States

ABSTRACT Since 1991, dramatic changes have occurred in the genetic composition of populations of Phytophthora infestans in the United States. Clonal lineages recently introduced into the United States (US-7 and US-8) are more common now than the previously dominant lineage (US-1). To help determine why these changes occurred, four clonal lineages of P. in-festans common during the early 1990s in the United States and Canada were evaluated for sensitivity to the protectant fungicides mancozeb and chlorothalonil using amended agar assays for isolates collected from 1990 to 1994. No isolate or lineage was resistant to either mancozeb or chlorothalonil. There were significant differences among isolates for degree of sensitivity to one fungicide individually, but there were no significant (P = 0.05) differences among the US-1, US-6, US-7, and US-8 clonal lineages for degree of sensitivity to both fungicides. Therefore, resistance to protectant fungicides cannot explain the rapid increase in frequency of the US-7 and US-8 clonal lineages. Three components of pathogenic fitness (latent period, lesion area, and sporulation after 96 h) were tested for the three clonal lineages that were detected most commonly during 1994 (US-1, US-7, and US-8). All but one of the isolates in this analysis were collected during 1994 and evaluated within 10 months of collection by inoculating detached leaflets of the susceptible potato cultivar Norchip. There were significant differences between the US-1 and US-8 clonal lineages for lesion area and sporulation, and between US-1 and US-7 for latent period. The US-6 clonal lineage was excluded from the pathogenic fitness experiments, because no isolates of this lineage were collected during 1994. Compared with US-7 and US-8, US-1 had the longest latent period and the smallest lesions with the least sporulation. Incorporation of the differences between US-1 and US-8 in computer simulation experiments revealed that significantly more protectant fungicide (e.g., 25%) would be required to suppress epidemics caused by the US-8 clonal lineage compared with US-1. These differences in pathogenic fitness components probably contribute to the general predominance of the "new" clonal lineages (especially US-8) relative to the "old" US-1 lineage.     

Competitive Fitness of Phytophthora infestans Isolates Under Semiarid Field Conditions

ABSTRACT Spread of US-1 and US-8 isolates of Phytophthora infestans were observed in field plots of potato (cv. Russet Burbank) grown in Pullman, WA, in 1996 and 1997. Infected greenhouse-grown potato plants with similar lesion numbers for both strains were transplanted to field plots with four replications. Spread of the pathogen was favored by sprinkler irrigation during evening hours. Diseased leaves and stems were sampled over time to determine the spread of US-1 and US-8 isolates. In 1996, late blight developed in two of the four replications (105 and 87 total isolates recovered). From those two replications, two US-1 isolates were recovered, both from the same replication. Nine isolates from one replication and six isolates from another displayed a phenotype different from the initial isolates, as determined by compatibility type, allozyme genotype, and restriction fragment length polymorphism genotype. These putative recombinant isolates may have arisen from sexual recombination between the US-1 and US-8 isolates. The remaining isolates were of the US-8 strain. In 1997, late blight developed in all four replications (123, 122, 81, and 34 total isolates recovered). One US-1 isolate was recovered (out of 123) from one replication and three (out of 122) from another, and the remaining isolates were of the US-8 strain. Isolates with phenotypes differing from the initial isolates were not recovered in 1997. In both years, oospores were not observed in the plant tissue examined. The low number of putative recombinant isolates in 1996 and their absence in 1997 suggests that sexual reproduction between US-8 and US-1 isolates in a field setting is a rare event. The predominance of US-8 isolates recovered is a measure of the increased fitness and aggressiveness of the US-8 isolates relative to the US-1 isolate used in this study. This further substantiates the increased aggressiveness of the US-8 genotype observed on excised tissues and potted plants in previous laboratory and greenhouse studies.     

Temperature Effects on Developmental Stages of Isolates from Three Clonal Lineages of Phytophthora infestans

ABSTRACT Sporangia germination of Phytophthora infestans isolates belonging to three clonal lineages (US-1, -7, and -8) was assessed at temperatures ranging from 10 to 25 degrees C. At 10 degrees C there were no significant differences in germination percents among US-1, -7, and -8. At 18 or 20 degrees C US-7 and -8 had significantly lower germination percents than US-1. At 21, 24, or 25 degrees C all clonal lineages had low germination percents. Sporangia of the US-7 and -8 lineages germinated more quickly at 15 degrees C (P = 0.001) during the first 2 h than did the US-1 lineage. The incubation period (IP), lesion area (LA), and sporulation per unit of lesion area (SPU) of the isolates were assessed on inoculated detached leaflets of susceptible potato cv. Norchip kept at 10, 15, 20, or 25 degrees C. In general, IP declined exponentially and LA increased exponentially with increasing temperatures. SPU had a quadratic shape, with the maximum at 15 degrees C. Averaged over all temperatures, the US-7 lineage had the shortest IP (59.3 h compared to 66.4 h for US-1 [P = 0.012] and 71.7 h for US-8 [P = 0.026]). Again, averaged over all temperatures, the US-8 lineage had a larger LA (P = 0.030) than US-1. There was no significant difference between US-7 and -1 for LA. There were no significant differences among lineages in terms of SPU. These results indicate that clonal lineages differ from each other in epidemiological attributes, but the differences can be complex.     

Population Genetic Structure of Phytophthora infestans in Ecuador

ABSTRACT The population genetic structure of Phytophthora infestans in Ecuador was assessed from 101 isolates collected from 1990 to 1992 and 111 isolates collected in 1993. All isolates were analyzed for mating type and allozyme genotype. Both samples were dominated (>95%) by a clonal lineage (EC-1) defined from neutral markers: 90/100 genotype for glucose-6-phosphate isomerase, 96/100 genotype for peptidase, A1 mating type, and a previously unreported nuclear DNA fingerprint. The remaining isolates belonged to the US-1 clonal lineage, which has a worldwide distribution. Isolates in the 1993 sample were analyzed for virulence and metalaxyl sensitivity. All representatives of EC-1 had complex patho-types, with three pathotypes representing >60% of the collection. There was variation for metalaxyl sensitivity. There was no evidence for geographical substructuring on the basis of neutral markers, but there was evidence for limited substructuring based on metalaxyl sensitivity and specific virulence. We hypothesize that EC-1 has been recently introduced to Ecuador.     

Identification of an A2 population of Phythophthora andina attacking tree tomato in Peru indicates a risk of sexual reproduction in this pathosystem

Tree tomato, Solanum betaceum, is an Andean fruit crop previously shown to be attacked by Phytophthora andina in Ecuador and Colombia. Blight‐like symptoms were discovered on tree tomato plants in the central highlands of Peru in 2003 and shown to be caused by P. andina. Isolates of P. andina, collected from three different plantations in Peru over a 6‐year time span (2003–2008), were compared genetically with P. andina isolates from Colombia and Ecuador to test whether the pathogen population is geographically structured in the Andes. Restriction fragment length polymorphism (RFLP), mitochondrial DNA and simple sequence repeat (SSR) genetic markers, and mating type behaviour indicated that the Peruvian P. andina population from tree tomato is genetically distinct from populations infecting tree tomato in Colombia (CO‐1) and Ecuador (EC‐3, Ia, A1), but is more similar to the population infecting solanaceous hosts of the Anarrhichomenum complex (EC‐2, Ic, A2). Such geographic substructuring within this pathogen species could result from spatial isolation. Most strikingly, in contrast to the Ecuadorian and Colombian P. andina isolates from tree tomato, the Peruvian isolates have the A2 mating type. The presence of both mating types in the Andean population of P. andina attacking tree tomato indicates a risk of sexual reproduction and the presence of long‐lasting oospores in this pathosystem.     

Fusarium oxysporum f. sp. cubense Consists of a Small Number of Divergent and Globally Distributed Clonal Lineages

ABSTRACT A worldwide collection of Fusarium oxysporum f. sp. cubense was analyzed using anonymous, single-copy, restriction fragment length polymorphism (RFLP) loci. Several lines of evidence indicated that this pathogen has a clonal population structure. Of the 165 isolates examined, only 72 RFLP haplotypes were identified, and nearly half the isolates were represented by the five most common haplotypes. Individuals with identical haplotypes were geographically dispersed, and clone-corrected tests of gametic disequilibrium indicated significant nonrandom association among pairs of alleles for 34 of 36 loci tested. Parsimony analysis divided haplotypes into two major branches (bootstrap value = 99%) that together contained eight clades supported by significant bootstrap values. With the exception of two isolates, all isolates within a vegetative compatibility group were in the same clade and clonal lineage. Clonal lineages were defined by isolates having coefficients of similarity between 0.94 and 1.00. Ten clonal lineages were identified, and the two largest lineages had pantropical distribution. Minor lineages were found only in limited geographical regions. Isolates composing one lineage (FOC VII) may represent either an ancient genetic exchange between individuals in the two largest lineages or an ancestral group. The two largest lineages (FOC I and FOC II) and a lineage from East Africa (FOC V) are genetically distinct; each may have acquired the ability to be pathogenic on banana independently.     

Genetic Relationships Between Cercospora kikuchii Populations from South America and Japan

ABSTRACT A collection 160 isolates of Cercospora kikuchii was made from South America and 245 from Japan. DNA fingerprint patterns were analyzed based on amplified fragment length polymorphism among the sample isolates, dividing the isolates into seven lineages (I to VII). Partial nucleotide sequence analyses of the beta-tubulin gene supported this division into seven lineages. Lineages I and III commonly existed in South America and Japan. In all, 136 of the 160 isolates from South America and 223 of the 245 isolates from Japan belonged to lineage I, indicating that lineage I was the major lineage in each area; 5 isolates from South America and 8 isolates from Japan belonged to lineage III. Lineages II (12 isolates) and IV (2 isolates) were specific to Japan and lineages V (3 isolates), VI (1 isolate), and VII (15 isolates) specifically existed in South America. These results suggest that the population genetic structure of C. kikuchii was different between South America and Japan, but the dominance of lineage I was common between the two areas.     

Characterization of Pyricularia grisea in the United States Using Independent Genetic and Molecular Markers

ABSTRACT A total of 540 isolates of Pyricularia grisea from rice in the United States were examined for vegetative compatibility, MGR586 DNA fingerprint diversity, and mating type based on hybridization with the mat1-1 and mat1-2 sexual mating type alleles. The collections contained both archived and contemporary field isolates representative of the known MGR586 lineages and races that occur throughout the United States. Complementary nitrate nonutilizing (nit) or sulfate nonutilizing (sul) mutants were used to assess vegetative compatibility in P. grisea. There was a complete correspondence between vegetative compatibility groups (VCGs), MGR586 lineage, and mating type among 527 contemporary isolates (collected between 1991 and 1997) from Arkansas, Louisiana, Missouri, Mississippi, and Texas; all isolates in MGR586 lineages A, B, C, and D belonged to VCGs US-01, US-02, US-03, and US-04, respectively. In addition, all isolates tested in VCGs US-01 and US-04 had the mat1-1 mating type allele whereas those in VCGs US-02 and US-03 had the mat1-2 allele. The strict association of independent markers during this sample period was consistent with a strictly asexual mode of reproduction. However, examination of archived isolates collected in the 1970s and 1980s and contemporary isolates revealed an incongruent relationship between the independent markers. MGR586 C and E isolates were vegetatively compatible which indicated that multiple robust MGR586 delineated lineages could be nested within certain VCGs. Although isolates in lineages C and E were vegetatively compatible, they were of opposite mating type. Several hypotheses, including recombination, could account for the incongruence between the various markers. Among the eight MGR586 lineages (A through H) that occur in the United States, all isolates in lineages A, D, E, G, and H had the mat1-1 allele, whereas isolates in lineages B, C, and F had the mat1-2 allele. Nit mutants can be recovered relatively easy from P. grisea and should allow large numbers of individuals within a population to be assessed for vegetative compatibility. VCGs may prove to be an effective multilocus marker in P. grisea. Thus, VCGs should be a useful means for characterizing genetic structure in populations of the rice blast fungus worldwide, provide a useful genetic framework to assist in interpreting molecular population data, and may provide insight into potential sexual or asexual recombination events.     

Genetic Structure of the Population of Phytophthora infestans Attacking Solanum ochranthum in the Highlands of Ecuador

Thirty-nine isolates of Phytophthora infestans were collected from the wild host Solanum ochranthum in the highland tropics of Ecuador and characterized with a set of phenotypic and molecular markers (mating type, metalaxyl sensitivity, the allozyme loci Gpi, and Pep, mitochondrial DNA haplotype, RFLP, and SSR), as well as for pathogenicity on various hosts. Three groups of isolates (A, B, and C) were identified based on their multilocus genotypes and variable abilities to cause disease on different hosts. Group A had a combination of alleles for the Gpi (86/100), Pep (96/100) and mtDNA (Ia) loci, as well as an RFLP fingerprint, that have not been reported for P. infestans in Ecuador, or elsewhere. Group B shares many marker characteristics with the US-1 lineage described in Ecuador on tomato, pear melon (S. muricatum), and S. caripense, but has SSR alleles not present in typical US-1 isolates. Group C for all markers tested is identical to the EC-1 lineage described on cultivated and wild potatoes in Ecuador. All isolates from S. ochranthum were able to re-infect their host of origin in the detached leaf assay; however, we did not draw clear conclusions as to the relative aggressiveness of the three groups on this host. Isolates of group A were the most specialized and were generally non-pathogenic or weakly pathogenic on all hosts other than S. ochranthum. Groups B and C infected tuber-bearing hosts, including the cultivated potato but were generally non-pathogenic on other non-tuber bearing hosts. Solanum ochranthum was infected by isolates coming from tuber-bearing Solanum hosts (i.e., the EC-1 lineage of P. infestans) and some US-1 isolates from non-tuber bearing hosts. Thus, in nature this species might be a potential reservoir of inoculum of different pathogen populations able to infect the cultivated hosts potato, tomato and pear melon (S.␣muricatum). Unlike potato and tomato in Ecuador, each of which is primarily attacked by a highly specialized pathogen population, S. ochranthum appears to harbour at least three pathogen groups of␣different genetic make-up. The unresolved issue of potential host specificity in isolates found on S.␣ochranthum could complicate efforts to use this species in tomato improvement.     

Aggressiveness and host specificity of Brazilian isolates of Phytophthora infestans

The population of Phytophthora infestans in Brazil consists of two clonal lineages, US-1 associated with tomatoes and BR-1 associated with potatoes. To assess whether host specificity in these lineages resulted from differences in aggressiveness to potato and tomato, six aggressiveness-related epidemiological components – infection frequency (IF), incubation period (IP), latent period (LP), lesion area (LA), lesion expansion rate (LER) and sporulation at several lesion ages (SSLA) – were measured on detached leaflets of late blight-susceptible potato and tomato plants. Infection frequency of US-1 was similar on potato and tomato leaflets, but IF of BR-1 was somewhat reduced on tomato. Incubation period was longer on both hosts with US-1, although this apparent lineage affect was not significant. Overall there was no host effect on IP. On potato, BR-1 had a shorter LP (110·3 h) and a larger LA (6·5 cm²) than US-1 (LP = 162·0 h; LA = 2·8 cm²). The highest LER resulted when isolates of BR-1 (0·121 cm² h⁻¹) and US-1 (0·053 cm² h⁻¹) were inoculated on potato and tomato leaflets, respectively. The highest values of the area under the sporulation capacity curve (AUSC) were obtained for isolates of US-1 inoculated on tomato leaflets (6146) and for isolates of BR-1 on potato leaflets (3775). In general, higher values of LA, LER, SSLA and AUSC, and shorter values of LP were measured when isolates of a clonal lineage were inoculated on their original host than with the opposite combinations. There is evidence that there are quantitative differences in aggressiveness components between isolates of US-1 and BR-1 clonal lineages that probably contribute to host specificity of P. infestans populations in Brazil.     

Evidence of parasexual exchange of DNA in the rice blast fungus challenges its exclusive clonality

ABSTRACT We applied DNA markers to determine whether parasexual recombination may contribute to the extreme genetic diversity and variability observed in Magnaporthe grisea, the causal agent of rice blast disease. Dispersed repetitive elements and mapped, low-copy restriction fragment length polymorphism (RFLP) probes were used to detect transfers of DNA between cultured isolates of M. grisea. Low-copy RFLP probes also were used to detect putative recombinants among isolates from well-characterized field populations of the pathogen. Microscopic examination of tufted mycelium between cocultured isolates revealed frequent hyphal fusions. Hyphal tips and conidia were recovered without selection from tufted zones in two separate vegetative pairings involving isolates with dissimilar haplotypes, based on the repetitive element MGR586. Haplotypic changes were observed at a higher frequency in tuft derivatives than in subcultures of each isolate alone. From 136 tuft derivatives analyzed, 5 putative recombinant haplotypes were identified. Introgression was demonstrated with two independent repetitive elements, fosbury and MGR586, as probes on DNA digested with several restriction enzymes. Introgressions were characterized by addition of 1 to 10 MGR586 bands, and 1 to 3 fosbury bands from one parent into the background of the other. Polymorphic single-copy probes were used to analyze putative recombinants. One probe detected an introgression event as predicted by analysis with MGR586. To assess the possible role of parasexual recombination in field populations of the pathogen, isolates in the Philippines previously grouped based on DNA fingerprinting were analyzed with low-copy RFLP markers. Polymorphism in single-copy loci typically was seen between, but not within, putative pathogen lineages. One lineage (designated lineage 4), however, was polymorphic for several probes. For some isolates, alleles at these loci comigrated with alleles characteristic of other lineages, suggesting the transfer of DNA fragments between lineages. One isolate was apparently a merodiploid, carrying an allele typical of lineage 4 plus another allele characteristic of a different lineage. In a survey of isolates from the Indian Himalayas, a merodiploid also was found with single- or low-copy probes. Examination of MGR586 profiles of the putative recombinant and its putative donor strains showed the expected introgression of MGR586 bands. The detection of parasexual DNA exchanges in wild-type strains under unselected conditions and the existence of merodiploids in nature suggest that parasexual recombination occurs in field populations of M. grisea. This raises questions concerning exclusive clonality in the blast fungus.     

Phytophthora infestans Populations from Tomato and Potato in North Carolina Differ in Genetic Diversity and Structure

ABSTRACT Phytophthora infestans causes a destructive disease on tomato and potato. In North Carolina (NC) potatoes are mostly grown in the east, whereas tomatoes are grown in the mountainous areas in the western part of the state. Five genotypes of P. infestans were identified from 93 and 157 isolates collected from tomato and potato over a 5 year period between 1993 and 1998. All isolates collected from potato in eastern NC were the US-8 genotype, whereas only a single isolate was the US-1 genotype. Tuber blight was found on immature daughter tubers in a single field in 1997, however infection on mature tubers was not observed. Within potato fields, a range of sensitivity to metalaxyl was observed among isolates but all were either intermediate or highly resistant to the fungicide. In contrast, isolates from tomatoes included previously reported US-7 and US-8 genotypes and two new genotypes called US-18 and US-19 (A2 mating type, allozyme genotype Gpi 100/100 and Pep 92/100). These genotypes had unique restriction fragment length polymorphism banding patterns, were sensitive to metalaxyl, and have not been reported elsewhere. All genotypes, with the exception of the US-1, were the Ia mitochondrial haplotype. Thus, isolates of P. infestans from tomato were more genetically diverse over time in NC than those from potato and include two new genotypes that are sensitive to metalaxyl.