Origin and distribution of cr2, a gene for resistance to white pine blister rust in natural populations of Western white pine

ABSTRACT The distribution and frequency of the Cr2 gene for resistance to white pine blister rust (Cronartium ribicola) in western white pine (Pinus monticola) was surveyed in natural populations of the host by inoculation of open-pollinated seedlings from 687 individual seed parents from throughout most of the species' range. Because Cr2 is dominant and results in a conspicuous hypersensitive reaction (HR) in pine needles, the phenotype can readily be detected in offspring of susceptible seed parents fertilized by unknown Cr2 donors in the ambient pollen cloud. Gametic frequencies of Cr2 were thus determined as the proportion of total challenged seedlings that were pollen receptors exhibiting the Cr2 phenotype. Zygotic frequencies, the proportion of seed parents with progeny that segregated in Mendelian ratios for the Cr2 phenotype to the total number of parents, were a complementary, though less precise, measure. Cr2 frequency was rare overall, ranging from 0.004 to 0.008 in the Sierra Nevada to about 0.001 in the central Cascade Range; it was undetectable further north in the Cascades, as well as in the Rocky Mountains and Coast Mountains of the United States and Canada. The diminishing frequency of Cr2 from the southern and central Sierra Nevada northward mirrors that of Cr1 in sugar pine (P. lambertiana) and points to this region as the origin of both genes. We rationalize that this coincidence may have resulted from protection that these genes may have conferred on both species to an endemic pine stem rust congeneric with C. ribicola (C. occidentale) in recent geologic epochs.     

Genetic specificity in the white pine-blister rust pathosystem

ABSTRACT Four of eight white pine species native to western North America surveyed for resistance to white pine blister rust by artificial inoculation showed classical hypersensitive reactions (HR) at frequencies ranging from very low to moderate. Mendelian segregation, indicating a single dominant allele for resistance (Cr3), was observed in southwestern white pine (Pinus strobiformis), as it was previously in sugar pine (P. lambertiana, Cr1) and western white pine (P. monticola, Cr2). HR was present at a relatively high frequency (19%) in one of five bulk seed lot sources of limber pine (P. flexilis), and was also presumed to be conditioned by a single gene locus, by analogy with the other three species. HR was not found in whitebark pine (P. albcaulis), Mexican white pine (P. ayacahuite), foxtail pine (P. balfouriana), or Great Basin bristlecone pine (P. longaeva), but population and sample sizes in these species may have been below the level of detection of alleles in low frequency. When challenged by (haploid) inocula from specific locations known to harbor virulence to Cr1 or Cr2, genotypes carrying these alleles and Cr3 reacted differentially, such that inoculum virulent to Cr1 was avirulent to Cr2, and inoculum virulent to Cr2 was avirulent to Cr1. Neither of these two inocula was capable of neutralizing Cr3. Although blister rust traditionally is considered an exotic disease in North America, these results, typical of classic gene-for-gene interactions, suggest that genetic memory of similar encounters in past epochs has been retained in this pathosystem.     

Virulence gene distribution and dynamics of the white pine blister rust pathogen in Western north america

ABSTRACT We assayed the distribution and frequency of two genes of the blisterpathogen with specific virulence to major resistance genes in sugar pine and western white pine in inoculum from extensive parts of the hosts' ranges. The genes, vcr1 and vcr2, differentially neutralize the cognate resistance alleles Cr1 and Cr2 of the two respective hosts and are clearly marked by their interaction phenotypes. Basidiospores from each inoculum source were cast over Cr1 and Cr2 host genotypes simultaneously, and interaction phenotypes scored when developed. vcr1 was confined to sites with high concentrations of Cr1 (mostly plantations) where frequencies tended toward fixation. vcr2 showed a similar tendency, except high frequencies were occasionally observed from natural and planted stands of western white pine with very low frequencies of Cr2. Otherwise, no pattern was evident for either allele: frequencies were very erratic from site to site within short distances (<1 to 7 km) of each other and oscillated with high amplitudes at the same sites measured in consecutive years. Intense selection for virulence by Cr alleles occurs locally, but spread of vcr alleles over the landscape is mitigated by remarkably low gene flow. Absence of heterozygotes among single telia inoculum on Cr2 genotypes indicated cytoplasmic inheritance of vcr2, similar to vcr1(previously reported).     

Evidence of Cytoplasmic Inheritance of Virulence in Cronartium ribicola to Major Gene Resistance in Sugar Pine

ABSTRACT Tests for Mendelian segregation of virulence and avirulence in Cronartium ribicola, causal agent of white pine blister rust, to a major gene (R) for resistance in sugar pine were made using haploid basidiospore progenies from single diploid telia as inoculum on resistant genotypes. The telia were sampled from a small deme in the Siskyou Mountains of northern California, where a few mature sugar pines known to be Rr genotypes had become infected after withstanding the chronic blister rust epidemic for several decades and where intermediate frequencies of virulence in the ambient basidiospore population were subsequently measured. Infection type on inoculated seedlings with R was qualitative: all progenies of 81 single telia tested over 3 different years were either virulent (compatible) or avirulent (inducing hypersensitive necrosis), never a mixture of both reactions. The complete absence of heterozygotes in the telia population is strong evidence that virulence is not controlled by a nuclear gene. The data are consistent with earlier tests showing that basidiospore inoculum derived from aeciospores isolated from infected Rr trees produced mostly (>90%) virulent reactions on R- seedlings. The evidence indicates that transmission of virulence is uniparental via the cytoplasm of aeciospores. Exchange of spermatia between haploid thalli does not appear to be involved.     

松材线虫对黑松一些生理生化指标的影响（英文）

 松材线虫病是松树的一种毁灭性病害。本文对接种松材线虫后3~4年生黑松茎部的一些生理生化指标的变化进行了研究。结果显示,接种线虫后,松苗的可溶性糖含量降低,在病情发展中期和后期比早期降低更显著。松苗的可溶性蛋白质和抗坏血酸含量逐渐降低,游离氨基酸含量也降低。松树茎部的可溶性蛋白质含量可用于病害的早期诊断。此外,对这4个生理生化指标在病害致病机理中的作用进行了讨论。     

Enhancement of sphaeropsis canker of aleppo pine by the Israeli pine bast scale

An interaction between the fungal pathogenSphaeropsis sapinea (Fr.) Dyko & Sutton, causal organism of Sphaeropsis canker, and the Israeli pine bast scaleMatsucoccus josephi (Homoptera: Margarodidae), causal agent of pine decline, was studied, since both were prevalent on Aleppo pine (Pinus halepensis) stands.Sphaeropsis sapinea was isolated from larvae (stage II) and molts of the Israeli pine bast scale collected in four Aleppo pine stands. When Aleppo pine saplings were infested with pine bast scale and subsequently inoculated withS. sapinea most plants died, whereas mortality of seedlings inoculated withS. sapinea alone without previous scale infestation was negligible. Scale infestation of saplings without subsequent fungal inoculation caused the death of one third of the plants. Wounding of pine shoot apices by removal of needle fascicles enhanced infection byS. sapinea. The results support the assumption thatM. josephi can play an important role in transmitting the fungal inoculum to the host tree as well as enabling its penetration to the shoots through wounds in the bark. http://www.phytoparasitica.org posting Dec. 20, 2004.     

黑松13个抗病家系对松材线虫的抗性反应及组织病理学观察

 为评估自日本引进的抗性黑松对松材线虫(Pine Wood Nematode,PWN)的抗病性,选用3个不同来源的PWN强毒和中强毒虫株采用皮接法接种,对1.5年生的黑松13个优良家系进行抗病性测定。结果表明,引进的黑松大部分家系对3个PWN虫株都表现出一定的抗病性,但针对不同虫株,抗病各家系间差异明显,其中有7个家系(3、4、5、6、9、11和12号)表现良好。同时,为探讨引进黑松家系感染PWN后的组织病理反应,对12号抗性家系进行组织病理学研究。观察表明,松材线虫在抗性黑松和普通黑松中的纵向扩散速度差别不明显,但其在抗性黑松内的横向扩散速度比在普通黑松内要慢,且抗性黑松体内木射线细胞和围绕树脂道泌脂细胞的薄壁细胞,出现病变要比普通黑松晚;在接种后同一时间取茎段的同一部位发现,抗性松苗组织解剖结构及细胞的病变和破坏程度要比普通松苗轻。     

松材线虫侵染对马尾松、黑松水分及其相关代谢的影响

 以盆栽马尾松(Pinus massoniana)幼苗和离体水培黑松(P.thunbergii)、马尾松为材料,研究了松材线虫(Bursaphe-lenchus xylophilus)侵染对寄主水分、游离脯氨酸及叶绿素含量的影响以及由此引起的幼苗萎蔫与干旱萎蔫间的差异。松苗和离体枝接种松材线虫后,茎部相对含水量逐渐下降,早期下降缓慢,中后期加快。叶片变化滞后于茎部,相对含水量的明显下降发生在叶片褐变萎蔫症状出现之后(病害发展后期)。游离脯氨酸含量、叶绿素含量变化完全随叶片相对含水量的变化而变化。水分状况是影响受侵植株叶片脯氨酸和叶绿素含量的决定因子。自然干旱枯萎和由松材线虫侵染引起枯萎的松幼苗,两者从枯萎方式到茎、叶相对含水量的变化均存在显著差异。     

White pine blister rust in north america: past and prognosis

ABSTRACT After a full century in North America, the blister rust epidemic has yet to stabilize, continuing to spread into warmer and drier areas previously considered climatically inhospitable. The disease apparently has no environmental limits wherever white pines and Ribes spp. cohabit and will eventually become pandemic. Although much timber value has been lost, more severe long-term damage is disruption caused to ecosystems by altered patterns of natural succession. During the last half of the century just past, development of genetic resistance superceded other direct control measures-mainly Ribes spp. eradication and antibiotics-which proved ineffective and/or unfeasible in large areas of the white pine range, especially in the West. Several mechanisms of complete (major gene) and partial resistance are common to at least several white pine species. Although North American populations of rust have low genetic variability overall, rust genotypes with specific virulence to major resistance genes exist in some local demes at high frequencies. The challenge will be to package and deploy resistance genes in ways that will dampen sudden increases in rust races of wide virulence. New introductions of blister rust from its gene center in Asia remain the gravest threat to genetic improvement programs.     

Gene cloning of a thaumatin-like (PR-5) protein of western white pine (Pinus monticola D. Don) and expression studies of members of the PR-5 group

To understand the molecular defence response of western white pine to the blister rust pathogen Cronartium ribicola, we have endeavoured to isolate and characterise pathogenesis-related (PR) proteins from western white pine. A full-length cDNA (Pin mTLP) was isolated from a cDNA library constructed from inoculated foliage. BLASTX search results indicated that the sequence shared a high degree of identity with the thaumatin-like proteins (TLPs) of the PR-5 family, and in particular with those of the ‘small-TLP’ subgroup mainly identified in monocots. The mature protein predicted from the cDNA sequence has a molecular mass of 16 kDa and pI of 4.1. Southern blot analysis showed that Pin mTLP is a single-copy gene member of a multi-gene family.Two-dimensional Western blot analysis of total western white pine protein extracts probed with a Douglas-fir anti-TLP antibody detected a major and a minor protein spot of 23 and 17 kDa, respectively. Both 23 and 17-kDa proteins were identified as TLPs by Nanospray MS/MS analysis. The 23-kDa protein was shown to accumulate in canker margins in the bark of infected seedlings indicating that it is locally induced in response to fungal invasion. Both local wounding and methyl jasmonate treatments induced expression of the protein, whereas salicylic acid treatment did not. These results suggest that the 23-kDa TLP plays a role in the molecular defence response of western white pine to wounding and pathogen attack.     

Survival time analysis of <Emphasis Type="Italic">Pinus pinaster</Emphasis> inoculated with <Emphasis Type="Italic">Armillaria ostoyae</Emphasis>: genetic variation and relevance of seed and root traits

Results of a greenhouse Armillaria ostoyae inoculation experiment, designed for screening resistant Pinus pinaster genotypes and for exploring the role of different phenotypic traits in seedling susceptibility, are reported. The experiment included 39 open-pollinated pine families that comprised a random subset of the breeding population of P. pinaster in Galicia (NW Spain). We employed a non-parametric survival-time analysis to analyze patterns of survival times during 14 months after inoculation with a local A. ostoyae strain. Results indicate (i) a significant correlation between seed weight and tree susceptibility, with seedlings originating from large seeds being more susceptible, (ii) a positive family mean correlation between secondary root weight and size and median life expectancy, and (iii) genetic variation of tree tolerance to A. ostoyae, with some families surviving significantly longer than others. Less susceptible families could be used in breeding programmes or directly in forest plantations to reduce the losses caused by A. ostoyae. Large within-family variation in tolerance to the disease was also observed, suggesting that non additive genetic variance was also important. Although being infected, 32 out of the 1200 inoculated trees survived the fungus infection. These tolerant genotypes comprise an attractive collection to further investigate genetic, phenotypic and environmental factors affecting pine susceptibility to Armillaria root rot.     

Induction of resistance against pine wilt disease caused by Bursaphelenchus xylophilus using selected pine endophytic bacteria

Pine wilt disease (PWD) is the most destructive disease threatening pine worldwide. The disease is mainly caused by the pine wood nematode, Bursaphelenchus xylophilus, which is vectored by pine sawyer longhorn-beetles, Monochamus spp. This study aimed to select resistance-inducing pine endophytic bacteria for management of PWD. To set up a defence-related genes expression pattern for screening, four chemical inducers (salicylic acid, γ-aminobutyric acid (GABA), β-aminobutyric acid and α-aminobutyric acid) were tested in vitro on pine calli and in vivo on pine seedlings. Treatment with GABA had the greatest reduction in PWD severity on pine seedlings. The pattern of defence-related gene expression in calli treated with GABA was used to select potential resistance-inducing bacterial strains. In addition, 92 bacterial strains were isolated from pine tree needles and stems and were tested for expression of defence-related genes in pine calli in vitro. Among the tested strains, 13 showed a similar pattern to GABA treatment in at least four tested defence-related genes and were selected for the seedling assay. From the seedling assay, three bacterial strains (16YSM-E48, 16YSM-P180 and 16YSM-P39) showed significant reduction in PWD severity compared to the untreated control. Moreover, among the selected strains, cell-free culture supernatant of strain 16YSM-P180 significantly reduced PWD severity in inoculated pine seedlings. The selected strains were identified based on the 16S rRNA sequence as Pseudomonas putida 16YSM-E48, Curtobacterium pusillum 16YSM-P180 and Stenotrophomonas rhizophila 16YSM-P39. These selected strains are suggested as potential alternatives for management of PWD by induction of systemic resistance.     

Genotype-isolate interaction for resistance to black stem in sunflower (Helianthus annuus)

Two experiments were undertaken to determine the partial resistance of sunflower genotypes to seven isolates of Phoma macdonaldii . In the first experiment, 28 genotypes, including recombinant inbred lines and their parents, M6 mutant lines developed by gamma irradiation, and some genotypes from different geographical origins, were used. The experiment consisted of a split-plot design with three replications, each with 12 seedlings per genotype per isolate, in controlled conditions. Seven days after inoculation, plantlets were scored on a 1–9 scale for percentage necrotic area. Highly significant differences were observed among genotypes, isolates and their interactions. The presence of a differential interaction between genotypes and P. macdonaldii isolates was confirmed in a second experiment using 12 genotypes representing large variability for partial resistance to P. macdonaldii isolates, as identified in the first experiment. Inbred lines B454/03, ENSAT-B5 and LC1064C were the most susceptible sunflower genotypes, whereas two American lines SDR19 and SDR18 presented high partial resistance to all P. macdonaldii isolates studied. The least and most aggressive isolates were MA6 and MP3, respectively. Isolates interacted differentially with sunflower genotypes. This study identified two genotypes (AS613 and PAC2) presenting specific resistance to isolate MP8. The results also showed a wide range of isolate-nonspecific partial resistances among the lines tested. The information presented here could assist sunflower breeders to choose parents of crosses for breeding of durable resistance to phoma black stem disease.     

Genetic and molecular analyses in crosses of race 2 and race 7 of albugo Candida

ABSTRACT The inheritance of avirulence and polymorphic molecular markers in Albugo candida, the cause of white rust of crucifers, was studied in crosses of race 2 (Ac2), using isolates MiAc2-B1 or MiAc2-B5 (metalaxyl-insensitive and virulent to Brassica juncea cv. Burgonde) with race 7 (Ac7), using isolate MsAc7-A1 (metalaxyl-sensitive and virulent to B. rapa cv. Torch). Hybrids were obtained via co-inoculation onto a common susceptible host. Putative F(1) progeny were selfed to produce F(2) progeny. The parents and F(1) progeny were examined for virulence on the differential cultivars B. juncea cv. Burgonde and B. rapa cv. Torch. Segregation of avirulence or virulence of F(2) populations was analyzed on cv. Torch. Putative F(1) hybrids were confirmed by random amplified polymorphic DNA markers specific for each parent. Avirulence or virulence of F (2) progeny to B. rapa cv. Torch suggested 3:1 in each of three populations, supporting the hypothesis of a single dominant avirulence gene. Amplified fragment length polymorphism markers also segregated in regular Mendelian fashion among F(2) progeny derived from two F(1) hybrids (Cr2-5 and Cr2-7) of Cross-2. This first putative avirulence gene in A. candida was designated AvrAc1. These results suggest that a single dominant gene controls avirulence in race Ac2 to B. rapa cv. Torch and provides further evidence for the gene-for-gene relationship in the Albugo-Brassica pathosystem.     

Fine-level genetic structure of white pine blister rust populations

ABSTRACT The fine-level genetic structure of the white pine blister rust agent, Cronartium ribicola, was investigated by sampling multiple monokaryotic spermogonia directly on cankers in four eastern Canadian white pine (Pinus strobus) plantations and assessing genetic variability, using random amplified polymorphic DNA (RAPD) markers. Ninety-eight percent of the cankers surveyed contained a single DNA haplotype, suggesting spermogonia within cankers are the result of clonal reproduction. A single canker contained two haplotypes that were divided between the upper and lower parts of the canker, suggesting it represented two confluent cankers. In contrast, genotypic diversity was high among cankers. Thirty-seven haplotypes were found among forty-three cankers sampled, and an analysis of molecular variance indicated that 93% (P < 0.001) of the total genetic diversity was attributable to sampling of different cankers, strongly suggesting that multiple infections do not take place in the white pine blister rust pathosystem, i.e., a canker is the result of infection by a single genotype. This result is in contrast with the high level of genetic diversity previously reported among dikaryotic aecidia within cankers and is consistent with the hypothesis that variability in the aecidial stage is the result of outcrossing between resident spermogonia and alien spermatia. The genetic structure of the spermogonial stage, which is the vegetative extension of infection by basidiospores and, therefore, the indirect result of meiosis, was consistent with random mating; the observed genotypic diversity was not significantly different (P > 0.05) from the genotypic diversity expected under the assumption of panmixis. The results indicate that monokaryotic cankers can be genotyped by sampling a single unopened spermogonia per canker.     

Variation in Virulence Within Meloidogyne chitwoodi, M. fallax, and M. hapla on Solanum spp

ABSTRACT The virulence of Meloidogyne hapla, M. chitwoodi, and M. fallax was studied on genotypes of Solanum spp. in a greenhouse. Juveniles of 11 M. hapla race A isolates, 3 M. hapla race B isolates, and 5 mono-female lines of a M. hapla race A isolate were inoculated on S. chacoense, S. hougasii, and S. sparsipilum. Juveniles of eight M. chitwoodi isolates, five M. fallax isolates, and six mono-female lines of a M. chitwoodi isolate were inoculated on S. bulbocastanum, S. chacoense, S. hougasii, S. stoloniferum, and S. tuberosum. Virulence was expressed as nematode reproduction 8 weeks after inoculation. Nematode reproduction was estimated by the number of egg masses and, in one experiment, by the number of hatched second-stage juveniles per inoculated juvenile. Considerable variation in virulence and resistance was observed among M. hapla isolates and plant genotypes, respectively. The M. hapla isolate-plant species interaction was highly significant. The response to M. chitwoodi ranged from susceptible (S. tuberosum and S. chacoense) to highly resistant (S. bulbocastanum and S. hougasii). S. tuberosum was susceptible to M. fallax, whereas all four wild species were resistant. In contrast to M. hapla, no significant isolate-plant genotype interaction was obtained for M. chitwoodi or M. fallax, indicating no or little intraspecific variation in virulence. M. chitwoodi juveniles in species mixtures with M. fallax isolates appeared to be able to break the resistance of S. bulbocastanum and S. hougasii. Significant differences among mono-female lines of M. hapla and M. chitwoodi were observed, indicating heterogeneity of pathogenicity within meiotic parthenogenic Meloidogyne populations.     

番茄细菌性溃疡病苗期接种新方法的研究

 在剪叶、浸根和针刺等细菌病害传统接种方法基础上设计了打顶接种新方法;以番茄细菌性溃疡病菌中国菌株和美国菌株借助打顶法接种佳粉10、合作908和华南红宝石3个国内主栽番茄品种2~3片真叶期幼苗,在昼夜最低、最高温度15~18℃、32~35℃和相对湿度为30%~60%的温室条件下,打顶法接种番茄幼苗后溃疡病发病率为87.5%~100.0%,病情指数随接种菌悬液浓度提高而增大,达到23.96~82.29,3个供试品种之间表现稳定一致;剪叶、浸根和针刺接种方法的发病率普遍在40%以下,病情指数在20以下,3个供试品种之间未表现明显差异;进一步研究显示,使用选择性培养基mSCM能够从打顶法接种后的发病植株上获得培养性状与原接种体一致的分离物,专化性免疫凝聚试剂盒检测到特定的测试线,PCR扩增到614bp的特异性条带,证实该分离物为番茄细菌性溃疡病菌,发病植株为接种体侵染所致。该研究结果表明,打顶接种方法比剪叶法、针刺法和浸根法更适合用于番茄溃疡病菌的致病性测定和评价不同番茄品种苗期的抗病性,具有应用价值。     

Substantial heritable variation for susceptibility to Dothistroma septosporum within populations of native British Scots pine (Pinus sylvestris)

The threat from pests and pathogens to native and commercially planted forest trees is unprecedented and expected to increase under climate change. The degree to which forests respond to threats from pathogens depends on their adaptive capacity, which is determined largely by genetically controlled variation in susceptibility of the individual trees within them and the heritability and evolvability of this trait. The most significant current threat to the economically and ecologically important species Scots pine (Pinus sylvestris) is dothistroma needle blight (DNB), caused by the foliar pathogen Dothistroma septosporum. A progeny‐population trial of 4‐year‐old Scots pine trees, comprising six populations from native Caledonian pinewoods each with three to five families in seven blocks, was artificially inoculated using a single isolate of D. septosporum. Susceptibility to D. septosporum, assessed as the percentage of non‐green needles, was measured regularly over a period of 61 days following inoculation, during which plants were maintained in conditions ideal for DNB development (warm; high humidity; high leaf wetness). There were significant differences in susceptibility to D. septosporum among families indicating that variation in this trait is heritable, with high estimates of narrow‐sense heritability (0.38–0.75) and evolvability (genetic coefficient of variation, 23.47). It is concluded that native Scots pine populations contain sufficient genetic diversity to evolve lower susceptibility to D. septosporum through natural selection in response to increased prevalence of this pathogen.     

Genetic analysis of partial resistance to bacterial leaf streak (Xanthomonas campestris pv. cerealis) in wheat

Genetic variability of partial resistance to bacterial leaf streak was investigated in hexaploid winter wheat ( Triticum aestivum. ), using 16 parental genotypes and 48 pure lines (F10) derived from a composite cross programme. Two experiments were undertaken in a controlled growth chamber. Seeds of all genotypes were grown under controlled conditions using a randomized block design with three replications. Each replication consisted of a row of 20 seedlings of each parent and pure line. An Iranian strain of bacterial leaf streak was used for the inoculation of 12-day-old seedlings. In a third experiment, eight genotypes from parents and F10 pure lines representing a large variability for partial resistance were inoculated with four other Iranian strains of bacterial leaf streak. A large genetic variability was observed amongst the 64 genotypes for partial resistance to the disease. Partial resistance heritability estimates were rather high (70%), indicating that the resistance factors may be transmitted by crossing. Amongst all genotypes investigated, 'DC²-30-N2' and 'IBPT-66' displayed the highest partial resistance to the disease. Significant correlations between strains in the third experiment show that a genotype resistant or susceptible to one strain will have similar reactions with other strains. No significant genetic gain was observed for partial resistance in the best pure line of the 48 lines studied, when compared with the best parental line. Increasing the number of pure lines is likely to result in the identification of genotypes that might prove to be more resistant.     

Identification and Characterization of Random Amplified Polymorphic DNA Markers Linked to a Major Gene (Cr2) for Resistance to Cronartium ribicola in Pinus monticola

ABSTRACT DNA markers tightly linked to resistance (R) genes provide a very powerful tool for both marker-assisted selection in plant breeding and positional cloning of R genes. In the present study, a linkage of random amplified polymorphic DNA (RAPD) markers to the single dominant gene (Cr2) for resistance to white pine blister rust fungus (Cronartium ribicola) was investigated in western white pine (Pinus monticola). A mapping population of 128 individual megagametophytes was generated from seeds of a heterozygous resistant tree (Cr2/cr2), and the corresponding seedlings of each megagametophyte were subjected to the test of phenotype segregation by inoculation with C. ribicola. Bulked segregant analysis and haploid segregation analysis identified eight robust RAPD markers linked to Cr2. This constitutes the first Cr2 genetic linkage map spanning 84.7 cM with four markers only 3.2 cM from Cr2. One sequence (U256-1385) of these linked markers was significantly similar to the Ty3/gypsy-like long terminal direct repeats retrotransposons. Another marker, U570-843, had no significant similarity to any entry in either GenBank or the loblolly genomics data bank. As presumed that the average physical distance per centimorgan is about 10 Mb in P. monticola, it is probably unrealistic to use these DNA markers for positional cloning of the Cr2 gene.