A2 Mating Type Isolates Research Articles

Phytophthora × cambivora as a Major Factor Inciting the Decline of European Beech in a Stand within the Southernmost Limit of Its Natural Range in Europe.

The objective of this study was to investigate the role of the oomycete Phytophthora × cambivora in the decline affecting European beech (Fagus sylvatica) in the Nebrodi Regional Park (Sicily, southern Italy). In a survey of a beech forest stand in the heart of the park, Phytophthora × cambivora was the sole Phytophthora species recovered from the rhizosphere soil and fine roots of trees. Both A1 and A2 mating type isolates were found. Direct isolation from the stem bark of trees showing severe decline symptoms and bleeding stem cankers yielded exclusively P. gonapodyides, usually considered as an opportunistic pathogen. The mean inoculum density of P. × cambivora in the rhizosphere soil, as determined using the soil dilution plating method and expressed in terms of colony forming units (cfus) per gm of soil, the isolation frequency using leaf baiting, and the percentage of infected fibrous roots from 20 randomly selected beech trees with severe decline symptoms (50 to 100 foliage transparency classes) were 31.7 cfus, 80%, and 48.6%, respectively. These were significantly higher than the corresponding mean values of 20 asymptomatic or slightly declining trees, suggesting P. × cambivora is a major factor responsible for the decline in the surveyed stand.

Mating Type and Aggressiveness of Phytophthora infestans (Mont.) de Bary in Potato-Growing Areas of Punjab, Pakistan, 2017\u20132018 and Identification of Genotype 13_A2 in 2019\u20132020

Samples of blighted potato leaves were collected from fields in six potato-growing districts of the Punjab Province of Pakistan in 2017–2018. A total of 149 isolates of Phytophthora infestans were obtained from six potato cultivars (Asterix, Barsenna, Burana, Caroda, Mazika, Sante). Of these isolates, 73% were A1 mating type, 23% were A2 mating type and 4% were self-fertile. Both mating types of P. infestans occurred in all six districts sampled, but in every case, the A1 mating type predominated. The foliar aggressiveness of 104 of these isolates (weakly pathogenic isolates were excluded) was tested on detached leaflets of potato cv. Caroda, and a composite aggressiveness index (CAI) calculated from lesion area (measured after 10 days), latent period and infection frequency was used to compare isolates. There was no difference in CAI between isolates obtained from different districts or cultivars. The A2 mating type isolates had significantly greater CAIs than A1 isolates but this does not imply a genetic linkage nor a general association between mating type and aggressiveness. It may be that the A2 isolates belonged to an aggressive clonal lineage such as 13_A2 that has been reported in other countries in the region. While it was not possible to test the isolates collected in 2017–2018, genotyping of samples collected in 2019–2020 showed the widespread occurrence of the EU_13_A2 clonal lineage in the same districts of Pakistan and supported this hypothesis. This is the first report of EU_13_A2 from Pakistan. The implications for potato late blight management in the Punjab are discussed.

A Microsatellite Analysis Used to Identify Global Pathways of Movement of Phytophthora cinnamomi and the Likely Sources of Wildland Infestations in California and Mexico.

The genetic structure of a sample of isolates of the oomycete plant pathogen Phytophthora cinnamomi from natural and agricultural outbreaks and the long-distance movement of individual genotypes were studied using four microsatellite markers to genotype 159 isolates of Californian, Mexican, and worldwide origins. Allelic profiles identified 75 multilocus genotypes. A STRUCTURE analysis placed them in three groups characterized by different geographic and host ranges, different genic and genotypic diversity, and different reproductive modes. When relationships among genotypes were visualized on a minimum spanning network (MSN), genotypes belonging to the same STRUCTURE group were contiguous, with rare exceptions. A putatively ancestral group 1 had high genic diversity, included all A1 mating type isolates and all Papuan isolates in the sample, was rarely isolated from natural settings in California and Mexico, and was positioned at the center of the MSN. Putatively younger groups 2 and 3 had lower genic diversity, were both neighbors to group 1 but formed two distinct peripherical sectors of the MSN, and were equally present in agricultural commodities and natural settings in Mexico and California. A few genotypes, especially in groups 2 and 3, were isolated multiple times in different locations and settings. The presence of identical genotypes from the same hosts in different continents indicated that long-distance human-mediated movement of P. cinnamomi had occurred. The presence of identical genotypes at high frequencies in neighboring wildlands and agricultural settings suggest that specific commodities may have been the source of recent wild infestations caused by novel invasive genotypes.

Race Characterization of Phytophthora Root Rot on Capsicum in Taiwan as a Basis for Anticipatory Resistance Breeding.

Peppers (Capsicum sp.) are an increasingly important crop because of their use as a vegetable, spice, and food colorant. The oomycete Phytophthora capsici is one of the most devastating pathogens to pepper production worldwide, causing more than $100 million in losses annually. Developing cultivars resistant to P. capsici is challenging because of the many physiological races that exist and new races that are continuously evolving. This problem is confounded by the lack of a universal system of race characterization. As a basis to develop a global anticipatory breeding program, New Mexico recombinant inbred lines (NMRILs) functioned as a host differential for Phytophthora root rot to characterize the race structure of P. capsici populations in Taiwan. Using the NMRILs, 24 new races were identified, illustrating the utility and usefulness of the NMRILs for anticipatory breeding. Virulence of P. capsici was observed to be geographically specific and in two virulence clusters. Interestingly, all but two isolates collected in 2016 were the A2 mating type, which is a shift from the predominantly A1 mating type isolates collected prior to 2008. The NMRILs host differential provides an approach for scientists to work together on a global scale when breeding for resistance as well as on a local level for regional gene deployment. Additionally, we propose that the current race numbering system, which has no biological meaning, be supplemented with the virulence phenotype, based on the susceptible NMRILs to a given isolate. This work provides insights into the population dynamics of P. capsici and interactions within the highly complex Capsicum-Phytophthora pathosystem, and offers a basis for similar research in other crops.

Evaluation\xa0of PCR markers for Phytophthora infestans mating type determination

Late blight is a devastating potato disease caused by Phytophthora infestans. This organism can reproduce asexually and sexually between the strains of two mating types named A1 and A2. Mating type is an important strain characteristic affecting the pathogen’s population structure. We validate different PCR markers for P. infestans mating type determination by comparison of the results obtained with the markers (W16, S1, PHYB) with the pairing test results for 26 isolates collected worldwide and for a group of 146 Polish isolates. This study identifies an interesting feature of the isolates of genotype US-1. For all these A1 mating type isolates, the product specific for A2 isolates is amplified using the marker W16. Analysis of sequences of W16 PCR products indicates high similarity of the US-1 isolates with modern A2 mating type isolates. When US-1 isolates are excluded from analysis, 95 and 96% of isolates are correctly assigned by markers W16 and S1, respectively, when compared with the pairing test results. Marker PHYB produces 14% of discrepant results with the pairing test. Our results show that molecular markers can be useful tools for P. infestans mating type determination, but their application should be preceded by validation in each local population since their efficiency may vary depending on a pathogen’s genotype.

Characterization of Phytophthora infestans populations in northwestern Algeria during 2008–2014

A total of 161 Phytophthora infestans isolates, collected from infected potato and tomato plants during 2008–2014, were characterized based on mating type, metalaxyl sensitivity and polymorphism at 12 simple sequence repeat (SSR) loci, in order to investigate the population of P. infestans in the north-west of Algeria, an emerging potato production region. The majority of isolates were of A2 mating type (112 isolates). A high percentage (89 %) of resistance to metalaxyl among isolates was detected. The metalaxyl resistant phenotype was present in both mating types with a higher percentage in A2 mating type isolates. SSR-based genotypic analysis of P. infestans population showed a low diversity. Genotype 13_A2 was the predominant in the population with a frequency of 67 % followed by 2_A1 (21 %) and 23_A1 (5 %). Genotype 23_A1 was detected only in tomato and potato isolates collected in 2013 and 2014.

High genotypic diversity found among population of Phytophthora infestans collected in Estonia

Potato late blight, caused by the oomycete pathogen Phytophthora infestans, is one of the most important diseases of potato worldwide. This is the first study characterising Estonian P. infestans population using the SSR marker genotyping method. 70 P. infestans isolates collected during the growing season in 2004 from eight potato fields in three different regions of Estonia were characterised with nine polymorphic SSR markers. A1 and A2 mating type isolates were detected from every studied field indicating the high potential for sexual reproduction, which raises the genotypic diversity in P. infestans population. Results revealed highly diverse P. infestans population in Estonia resembling the Northern European populations. Most of the multilocus genotypes were detected only once among the collected isolates. Subpopulations collected from different geographical regions of Estonia showed no differentiation from each other but instead formed one highly diverse group.

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.

Characterisation of Phytophthora colocasiae isolates associated with leaf blight of taro in India

Leaf blight and corm rot caused by Phytophthora colocasiae are the most devastating diseases of taro. Fourteen P. colocasiae isolates collected from different states of India were characterised for mating type, metalaxyl fungicide sensitivity and aggressiveness on taro. These organisms were identified as P. colocasiae based on morphological characteristics, internal transcribed spacer (ITS) sequence homology and pathogenicity to taro plants. Ribosomal DNA (rDNA) ITS regions of P. colocasiae isolates were examined, and compared with additional Phytophthora species. All isolates of P. colocasiae fell within a single cluster in phylogenetic trees, regardless of their geographic origins. These isolates were more closely related to Phytophthora capsici. All isolates except the isolate collected from Sikkim state of India (98–111) were sensitive to metalaxyl at 100 ppm and it was also scored for causing higher average foliar disease and tuber rot ratings. Isolates tested exhibited four growth patterns in cultures: cottony, rosaceous, petaloid and stellate. P.colocasiae isolates, including an IISR isolate (PC-73), with cottony growth pattern did not grow at 36°C. The mean oospore diameter of A1 mating type isolates was greater than that of A2 mating type isolates. Six of 14 isolates tested produced chlamydospores in V8-CaCO3 liquid medium.

Root and Basal Stem Rot of Rose Caused by Phytophthora citrophthora in Italy.

Approximately 800 ha of cut flower roses are cultivated for commercial production in Italy. During autumn of 2004 in an experimental greenhouse in western Sicily (southern Italy), 60% of 2-year-old plants of rose cv. Red France on Rosa indica cv. Major rootstock grown in soil showed leaf chlorosis and wilt. A dark brown lesion lined by a water-soaked area was noticeable at the stem base near the soil surface. Root rot was found consistently associated with aboveground symptoms and plants collapsed within 4 months after the appearance of the first symptoms. The same symptoms were observed sporadically on rose plants of the same cultivar during the last 6 years in commercial nurseries in western Sicily. In all cases, a Phytophthora species has been consistently isolated from rotted roots and stems on Phytophthora-selective media. Pure cultures were obtained by single-hypha transfers. The species was identified as Phytophthora citrophthora on the basis of morphological characters and electrophoretic analysis of mycelial proteins on polyacrylamide gel (1). On potato dextrose agar, isolates produced petaloid colonies with optimum growth temperature at 25°C. On V8 agar, mono- and occasionally bipapillate, ovoid to limoniform sporangia, measuring 44 to 55 × 27 to 28 μm, with a mean length/breadth ratio of 1.4:1 were produced. All isolates were heterothallic but did not produce gametangia in dual cultures with P. nicotianae isolates of A1 and A2 mating type. Electrophoretic patterns of total mycelial proteins and four isozyme (acid and alkaline phosphatases, esterase, and malato dehydrogenase) of the isolates from rose were identical to those of reference isolates of P. citrophthora, but clearly distinct from isolates of other heterothallic species with papillate sporangia, including P. capsici, P. nicotianae, P. palmivora, and P. tropicalis. All isolates from rose showed the same electrophoretic profiles. Blast search of rDNA-ITS sequence from PCR-amplified ITS4/ITS6 primers (2) of a representative isolate from rose (IMI 392044) showed 98% homology with a reference isolate of P. citrophthora (GenBank No. EU0000631), thus confirming the identification. Pathogenicity of isolate IMI 392044 was tested on 10 12-month-old plants of rose cv. Red France grafted on R. indica cv. Major transplanted in pots containing a mixture of sphagnum peat moss and sandy loam soil (1:1 vol/vol) infested with 80 g of inoculum per liter of mixture. Inoculum was produced by growing the isolate on wheat kernels. Plants transplanted in pots containing noninfested soil served as controls. Plants were kept in a greenhouse at 22 ± 3°C and watered to soil saturation once a week. Inoculated plants developed symptoms of leaf chlorosis and root and crown rot within 15 to 30 days and wilted within 40 to 80 days after inoculation. Control plants remained healthy. P. citrophthora was consistently reisolated from inoculated plants. Root and basal stem rot of rose may be caused by several Phytophthora spp. and has been reported in various countries of Asia, Europe, and North America (3,4). However, to our knowledge, this is the first report in Italy. The occurrence of this disease may be attributed to excessive irrigation practices.

강원지역 감자 역병균 Phytophthora infestans의 교배형 및 약제저항성 변화

Potato late blight caused by Phytophthora infestans was the most constrain disease at potato cultivation areas. The mating type distribution and fungicides response of P. infestans were investigated to elucidate the changes of pathogen from Gangwon province. On the fungal isolates in 2006, 58.7% were A1 mating type and 41.3% were A2 mating type. In 2007, A1 mating type isolates increased to 93.3% and A2 mating type isolates were collected from Jinbu areas as much as 6.7%. About 234 isolates analysed for metalaxyl response, the results was resistance 73.7%, intermediate 18.8% and sensitive 7.5% in 2006. And it was resistance 59.4%, intermediate 4.0% and sensitive 36.6% in 2007. It meant that mating type distribution and fungicide response were very different over the collection sites. Minimal inhibition concentration (MIC) of dimethomorph examined with 126 isolates in 2006 and 106 isolates in 2007. MIC over was 56.3% in 2006 and it was 3.8% in 2007. The average value of P. infestans was in 2006, but it decreased to in 2007. Fungicides response and pathogenesis of P. infestans should be monitored continuously to enhance the chemical efficacy at potato fields.

Formation and Pathogenicity Variation of Oospores of Phytophthora capsici Infecting Black Pepper

Phytophthora capsici Leonian is the causal agent of foot rot disease of black pepper (Piper nigrum L.). Foot rot disease is the most destructive disease which can cause significant economic losses of black pepper. Two mating types of P. capsici were found in black pepper plantations in Lampung. This research was aimed at examining the effects of temperature, light, and polycarbonate membrane on oospore formation. Also the effect of light on germination and determining both mating types of 30 randomly selected progenies. The results showed that oospores were produced through either hyphal contact or without any contact. Oospores were produced abundantly on V8 agar in the dark at 16-24 oC. Oospores germinated after 3-weeks incubation in the dark followed by 1-week incubation under TL fluorescent light. All progeny were heterothallic, and consisted of 16 isolates of A1 mating type and 14 isolates of A2 mating type. There was no correlation between mating type categories and their pathogenicities on black pepper leaves. Twenty-six progeny isolates may infect unwounded pepper leaves, whereas another four isolates may only infect wounded pepper leaves. Pathogenicities of all progenies were lower than those of their parents.

Characterization of Phytophthora capsici Isolates from Processing Pumpkin in Illinois.

This study was conducted to investigate pathogenic, morphologic, and genetic variations among Phytophthora capsici isolates from processing pumpkin (Cucurbita moschata) fields in Illinois. Random amplified polymorphic DNA (RAPD) markers were employed to assess genetic variation among 24 isolates of P. capsici from 10 individual fields at six locations. Unweighted mean pair group analysis clustered isolates into six groups. The genetic distances ranged from 0.03 to 0.45. Inoculation of pumpkin seedlings in the greenhouse revealed that the isolates belonged to six distinct genetic groups differing significantly (P = 0.05) in virulence. Isolates tested exhibited four growth patterns in culture: cottony, rosaceous, petaloid, and stellate. P. capsici isolates, including an ATCC isolate (ATCC-15427), with cottony growth pattern did not grow at 36°C. The mean oospore diameter of A1 mating type isolates was greater than that of A2 mating type isolates. Nine of 24 isolates tested produced chlamydospores in V8-CaCO3 liquid medium.

Changes in the Sensitivity to Metalaxyl, Dimethomorph and Ethaboxam of Phytophthora infestans in Korea

Changes of control efficacy of chemical to potato late blight caused by Phytophthora infestans in potato fields from 2001 to 2004 were examined. Control efficacy of metalaxyl was suddenly decreased from 100% in 2002 to 50% in 2004 and that of dimethomorph also was similar to those of metalaxyl. However, the control efficacy of ethaboxam no great change. Both A1 and A2 mating type isolates were isolated from 2001 to 2004 in several areas in Korea. The majority of the P. infestans isolates were A1 mating type. Total 939 isolates of P. infestans obtained from several areas in Korea from 2001 to 2004 were examined for changes of sensitivity to metalaxyl. Frequencies of metalaxyl resistance isolates were gradually increased from 17% in 2001 to 84.2% in 2004, but isolation frequencies of metalaxyl sensitive and intermediate resistant isolate were decreased. Cause of decreasing control efficacy of metalaxyl was thought by increase of resistance isolates in A1 mating type population according to increasing metalaxyl use. Most isolates were grown at 0.5 <TEX>${\mu}g</TEX>/ml of dimethomorph and isolates grown at 1 <TEX>${\mu}g</TEX>/ml of dimethomorph were approximately 10.2-22.9%. However, no isolate was able to grow at 5.0 <TEX>${\mu}g</TEX>/ml. Based on these results, minimum inhibitory concentrations (MIC) of dimethomorph to P. infestans were determined to be 0.5-1.0 <TEX>${\mu}g</TEX>/ml. Our results indicated that the reason decreasing control efficacy of dimethomorph was not caused by occurrence of resistant isolates. About 5% and 12.1% isolates among the total isolates collected in 2003 and 2004 were grown on V-8 juice rye agar containing 1.0 <TEX>${\mu}g</TEX>/ml ethaboxam. The 2.1 and 25.4% isolates had MICs of 0.2-0.4 <TEX>${\mu}g</TEX>/ml, and MIC values of 87.9% and 74.3% isolates were less than 0.2 <TEX>${\mu}g</TEX>/ml concentrations of ethaboxam. Therefore, resistance development by P. infestans to ethaboxam is not likely to occur in the natural condition.

Phenotypic and genotypic diversity of Phytophthora infestans populations in Scotland (1995–97)

In a survey of Scottish potato late blight (Phytophthora infestans) populations from 1995 to 1997, nearly 500 isolates were collected from over 80 disease outbreaks in commercial potato crops and gardens/allotments. The isolates were characterized by mating type, resistance to the fungicide metalaxyl and almost 300 were examined by DNA‐based AFLP fingerprinting. These data were examined alongside cropping details to determine the population structure in the context of existing disease management strategies. A1 and A2 mating type isolates were present in both commercial potato crops and gardens or allotments although they coexisted more frequently in the latter sites. One‐fifth of the isolates collected were of the A2 mating type and the frequency was similar over the 3 years and amongst sites. In 1995 the proportions of isolates that were sensitive and resistant to metalaxyl were equal (∼40%) but, over the following 2 years, the frequency of resistant isolates decreased and that of intermediate isolates increased. The mating type response to metalaxyl differed markedly, with 52% of A1 and only 5% of A2 isolates being resistant. Considerable molecular diversity was observed, with over half of the isolates having unique AFLP patterns. Analysis of the molecular and phenotypic data revealed a broad clustering of the population into three groups. Many factors point to an A2 population restricted by its sensitivity to phenylamides. The majority of the A2 isolates were found in a single AFLP group, but the presence of mixed mating type samples, an increasing frequency of isolates of intermediate metalaxyl resistance and the extent of the AFLP diversity suggest occasional sexual recombination, and thus gene flow, between groups.

Phytophthora infestans: populations, pathogenicity and phenylamides.

Isolates of Phytophthora infestans (Mont) de Bary (the potato and tomato late blight pathogen) resistant to phenylamides appeared in Europe and North America in the late 1970s and early 1990s respectively. Concurrent, but coincidentally, with both these events there were radical structural shifts in the pathogen populations as immigrant genotypes from Mexico displaced the indigenous populations. Both A1 and A2 mating type isolates are now present in blighted crops, permitting alternative inoculum via germinating sexually produced oospores to influence dynamics of late blight populations. Studies of inheritance of ploidy, host-specific pathogenicity, mating type and resistance to antibiotics and phenylamide fungicides have provided insight into mechanisms of variation in this potent pathogen.

AFLP and RFLP (RG57) fingerprints can give conflicting evidence about the relatedness of isolates of Phytophthora infestans

Selected isolates of Phytophthora infestans from around England and Wales were fingerprinted using both RG57, a multi-locus RFLP probe, and Amplified Fragment Length Polymorphisms (AFLPs). The larger number of polymorphisms detectable with the AFLP method allowed resolution of several similar AFLP genotypes among isolates with identical RG57 fingerprints. However, some isolates with the same RG57 genotype had remarkably dissimilar AFLP genotypes, suggesting that there has been convergent evolution of some RG57 fingerprints. Also, some isolates with dissimilar RG57 fingerprints had similar or identical AFLP fingerprints. Both techniques distinguished isolates of mitochondrial DNA haplotype la from those of haplotype IIa. However, with AFLPs only, most of the isolates of A2 mating type were very similar and were distinguished from those of A1 mating type, suggesting that gene flow between A1 and A2 genotypes is limited and that sexual recombination is rare.

Sexual recombination in Phytophthora cinnamomi in vitro and aggressiveness of single‐oospore progeny to Eucalyptus

Fifty single oospore progeny were established from an in vitro mating of A1 and A2 mating type isolates of Phytophthora cinnamomi from South Africa. Forty‐nine progeny were identified as F1 hybrids using seven random amplified polymorphic DNA (RAPD) primers, and one was a selfed isolate of the A1 mating type parent. Among the hybrid progeny, 24 and 25 were A1 and A2 mating type, respectively. Aggressiveness of progeny and parental isolates was assessed on 1‐year‐old seedlings of Eucalyptus smithii. The mean aggressiveness of hybrid oosporic isolates, expressed as lesion length, was significantly (P = 0·0001) lower than that of the parental isolates. No significant difference in aggressiveness of A1 and A2 mating type F1 hybrid isolates was observed. This is the first report demonstrating sexual recombination in vitro in P. cinnamomi.

Environmental and Genetic Factors Influencing Self-Fertility in Phytophthora infestans

ABSTRACT Phytophthora infestans is generally regarded as heterothallic-requiring physical proximity of two individuals of different mating type (A1 and A2) for oosporogenesis. Recent reports of limited selfing in young cultures of this oomycete stimulated us to investigate factors contributing to the phenomenon. The ability to produce oospores rapidly (within 2 weeks) in pure, single individual cultures (self-fertility) was tested in 116 individual isolates. The 116 isolates were from geographically diverse locations (16 countries) and were genetically diverse. Mating type and growth medium were the most prominent factors in determining if an isolate would be self-fertile. The majority of A2 isolates (45 of 47 tested) produced oospores when grown on a 50:50 mixture of V8 and rye B medium. In contrast, the majority of A1 isolates (65 of 69 tested) did not produce oospores on this medium. None of the 116 isolates produced oospores when grown on rye B medium (with no V8 juice). Further tests on representative A1 and A2 isolates revealed that oatmeal agar, tomato juice agar, and V8-juice agar all induced the A2 mating type isolate to produce oospores but did not induce the A1 mating type isolate to produce oospores. Calcium carbonate and pH did not alter the self-fertile oospore production in either A1 or A2 mating type isolates. For in vivo tests, the application of fungicide to potato or tomato leaf tissue either before or after inoculation did not stimulate any individual isolate (one A2 and one A1 isolate) to produce oospores in infected tissue. However, in all of the controls for all experiments (in vivo and in vitro), many oospores were produced rapidly if both strains grew in physical proximity.

Characterization of Phytophthora infestans Populations in Western Washington.

The first detection in the United States of isolates of Phytophthora infestans having metalaxyl insensitivity and complex pathotypes occurred in western Washington during the early 1990s. To determine the genetic structure of the current population in western Washington, a total of 115 isolates of P. infestans were obtained during 1996 from infected tubers or foliage of potato, tomato, nightshade, and bittersweet throughout the region. An additional 45 isolates were collected from a single field. Based on mating type, metalaxyl-insensitivity, and molecular markers (allozymes of glucose-6-phosphate isomerase, peptidase, and RG57 DNA fingerprint), all of the isolates were A1 mating type and had the US-11 multilocus genotype. Analyses of an additional 120 isolates collected during 1997 from potato, tomato, and nightshade were performed. As in 1996, US-11 was the predominant genotype detected on all three hosts. However, three additional A2 mating type genotypes were also detected: US-7, US-8, and US-14. These three genotypes represent the first A2 mating type isolates detected in western Washington. Most of a subset of 60 isolates infected 4 to 7 of the 10 potato differentials tested. This included 90% of the isolates collected in 1996 (all US-11), plus 72% of the US-11 and 100% of the US-8 and US-14 isolates collected during 1997. Virulence phenotypes in this region are complex even without the selection pressure of R-genes in the local commercial cultivars. The apparent increase in genetic variation observed in populations of P. infestans in western Washington from 1996 to 1997 most likely occurred by migration rather than by sexual recombination.