Juglans Hindsii Research Articles

Phytophthora and Phytopythium species associated with walnut dieback in plantations of northern Patagonia, Argentina

AbstractThe walnut industry in Argentina has grown in recent years, especially in northern Patagonia. However, this expansion has also brought new challenges, including the emergence of diseases threatening the health and productivity of walnut plantations. One of the most important diseases affecting walnut crops worldwide is ‘ink disease’ or ‘walnut dieback'. The aim of this study was to characterize ink disease, identify the aetiological agents, and estimate its incidence in walnut orchards of northern Patagonia, Argentina. Symptomatic trees were observed in the seven surveyed walnut orchards, showing a range of symptoms, including chlorosis and wilting, stem cankers and desiccation. In vivo assay and molecular analyses revealed that not only Phytophthora (P.) but also Phytopythium (Ph.) species are the causal agents of the disease, with P. drechsleri, Ph. mercuriale and Ph. vexans being the most virulent over the commonly used rootstock Juglans hindsii. Notably, Ph. vexans was the most frequently isolated pathogen, suggesting its prevalence and potential impact on the regional walnut industry. Overall, this study provides critical insights into the prevalence and impact of Phytophthora and Phytopythium species on walnut health in northern Patagonia. The presence of multiple pathogenic oomycete species raises concerns about the health of walnut orchards, as well as the potential risk of pathogen transmission to other fruit crops in the region. The findings will help growers to develop better cultural practices and implement measures to guarantee the sustainability of the walnut industry in Patagonia.

Reference genome of California walnut, Juglans californica, and resemblance with other genomes in the order Fagales.

Juglans californica, California walnut, is a vulnerable small tree that is locally abundant but restricted to woodland and chaparral habitats of Southern California threatened by urbanization and land use change. This species is the dominant species in a unique woodland ecosystem in California. It is one of two endemic California walnut species (family Juglandaceae). The other species, Northern California black walnut (J. hindsii), has been suggested controversially to be a variety of J. californica. Here, we report a new, chromosome-level assembly of J. californica as part of the California Conservation Genomics Project (CCGP). Consistent with the CCGP common methodology across ~150 genomes, we used Pacific Biosciences HiFi long reads and Omni-C chromatin-proximity sequencing technology to produce a de novo assembled genome. The assembly comprises 137 scaffolds spanning 551,065,703 base pairs, has a contig N50 of 30Mb, a scaffold N50 of 37Mb, and BUSCO complete score of 98.9%. Additionally, the mitochondrial genome has 701,569 base pairs. In addition, we compare this genome with other existing high quality Juglans and Quercus genomes, which are in the same order (Fagales) and show relatively high synteny within the Juglans genomes. Future work will utilize the J. californica genome to determine its relationship with the Northern California walnut and assess the extent to which these two endemic trees might be at risk from fragmentation and/or climate warming.

Phenotypic Differences and Physiological Responses of Salt Resistance of Walnut with Four Rootstock Types.

Walnut is one of the world’s four largest nuts. Currently, the bottleneck in walnut breeding is the production of resistant variants. Soil salinization is a global problem, and the use of salt-tolerant rootstocks is a basic strategy to overcome the challenge of sustained walnut production. Providing a scientific basis for the selection of walnut salt-tolerant rootstocks is possible by studying the physiological and biochemical response characteristics and salt tolerance variations of different walnut genotypes under salt stress. In the present study, seedlings of four genotypes of walnut rootstocks, including J1 (Juglans hindsii), J2 (J. mandshurica), J3 (J. regia × J. mandshurica), and J4 (J. regia × J. hindsii), were employed as test materials to conduct a 28-day pot experiment under NaCl stress with five NaCl concentrations (0, 50, 100, 200, and 300 mmol/L). Under different NaCl treatment concentrations, seedling morphology, growth indices, chlorophyll content, photosynthetic parameters, relative electrical conductivity (REC), malondialdehyde (MDA), proline (Pro), soluble sugar (SS), and the activity of superoxide dismutase (SOD) and peroxidase (POD) in the leaves were examined. Salt stress altered the morphological characteristics and growth indices of seedlings from four genotypes to varying degrees. In addition, according to the analysis of physiological and biochemical data, salt stress had a considerable impact on both the physiological and biochemical processes of seedlings. Salt stress decreased the chlorophyll content and photosynthetic parameters of four genotypes, the REC, MDA content, Pro content, and SS content of each genotype increased by different degrees, and the enzymatic activities showed different trends. The salt tolerance of rootstocks was evaluated thoroughly using principal component analysis and membership function analysis based on the 16 parameters. The results of a comprehensive evaluation of salt tolerance showed that the order of salt tolerance of the four genotypes was J4 > J1 > J3 > J2, which corresponded to the order of the morphological symptoms of salt injury. In conclusion, J4 has strong salt tolerance and is an important germplasm resource for walnut salt-tolerant rootstock breeding.

Rejuvenation remodels transcriptional network to improve rhizogenesis in mature Juglans tree.

Adventitious rooting of walnut species (Juglans L.) is known to be rather difficult, especially for mature trees. The adventitious root formation (ARF) capacities of mature trees can be significantly improved by rejuvenation. However, the underlying gene regulatory networks (GRNs) of rejuvenation remain largely unknown. To characterize such regulatory networks, we carried out the transcriptomic study using RNA samples of the cambia and peripheral tissues on the bottom of rejuvenated and mature walnut (Juglans hindsii × J. regia) cuttings during the ARF. The RNA sequencing data suggested that zeatin biosynthesis, energy metabolism and substance metabolism were activated by rejuvenation, whereas photosynthesis, fatty acid biosynthesis and the synthesis pathways for secondary metabolites were inhibited. The inter- and intra-module GRNs were constructed using differentially expressed genes. We identified 35 hub genes involved in five modules associated with ARF. Among these hub genes, particularly, beta-glucosidase-like (BGLs) family members involved in auxin metabolism were overexpressed at the early stage of the ARF. Furthermore, BGL12 from the cuttings of Juglans was overexpressed in Populus alba × P. glandulosa. Accelerated ARF and increased number of ARs were observed in the transgenic poplars. These results provide a high-resolution atlas of gene activity during ARF and help to uncover the regulatory modules associated with the ARF promoted by rejuvenation.

Walnut twig beetle landing rates differ between host and nonhost hardwood trees under the influence of aggregation pheromone in a northern California riparian forest

Abstract Host selection behaviour of the walnut twig beetle (WTB) among hardwood trees was investigated in a riparian forest in northern California by monitoring the landing rate of the beetle with sticky traps on branches baited with 3‐methyl‐2‐buten‐1‐ol, the male‐produced aggregation pheromone. The assay was conducted over 7 days (22 May to 29 May 2017) and compared landing rates on branches of six nonhost species paired with northern California black walnut, Juglans hindsii (the host). A total of 2242/1192 WTB were collected on branches of host/nonhost pairs, and more WTB landed on J. hindsii than on nonhosts in 42 of 58 instances. Female landing rate generally exceeded male landing rate, which underscores the influence of the male‐produced synthetic pheromone in this system. Landing rates of WTB males, females, and the combined sexes on boxelder, Acer negundo, and valley oak, Quercus lobata, did not differ significantly from the landing rates on J. hindsii, suggesting that these two nonhost riparian hardwoods do not repel WTB (in the context of the aggregation pheromone). Significantly fewer WTB landed on Oregon ash, Fraxinus latifolia, river red gum, Eucalyptus camaldulensis, Fremont cottonwood, Populus fremontii, and red willow, Salix laevigata, than on J. hindsii, which suggests that these four nonhosts may repel one or both sexes of WTB in the context of the aggregation pheromone. Future analysis of the volatiles from these four hardwood species may lead to the discovery of semiochemical repellents for WTB.

A study of landing behaviour by the walnut twig beetle,Pityophthorus juglandis, among host and nonhost hardwood trees in a northern California riparian forest

AbstractThe host selection behaviour of the walnut twig beetle,Pityophthorus juglandis, was assessed by monitoring the landing rates of the beetles with sticky sheet traps on the host and nonhost hardwood branches.Sticky sheet traps were deployed for 8 weeks from 6 June to 2 August. 2017 in the Putah Creek Riparian Reserve, Davis,CA. Branches from host northern California black walnut,Juglans hindsii, were paired with branches from six nonhost hardwood species.The landing rate ofP.juglandis(412 beetles trapped/8 weeks; 389 on host branches, 23 on nonhost branches) was significantly greater on the host branches for all nonhost hardwoods exceptPopulus fremontii. Proportional comparisons of beetle presence also revealed a significant preference for the host branches compared with all but two nonhost species,Acer negundoandP.fremontii.CapturingP.juglandiswithout the use of an aggregation pheromone was a rare event, underscoring the difficulty of studying the initial phases of host selection behaviour in bark beetles. Unbaited funnel traps adjacent to selected host trees in the experiment only captured five individuals over a 19‐week period. None were captured in traps adjacent to nonhost trees.This study provided evidence thatP.juglandisdiscriminates between host and nonhost branches while in‐flight. This directed flight behaviour is likely informed by the recognition of both host and nonhost volatile cues.This study established an ecological context for the development of a semiochemical‐based repellent system for protecting walnut trees from future attacks from this invasive bark beetle.

Effects of temperature and other environmental factors on the post‐diapause development of walnut husk fly Rhagoletis completa (Diptera: Tephritidae)

AbstractTo more effectively manage walnut husk fly Rhagoletis completa (Diptera: Tephritidae), in California walnut orchards, it is important to understand the factors that affect the timing of adult emergence. In the present study, we examine the effects of incubation temperature, pre‐chill and chill durations, latitude, cultivar and size on the post‐diapause development of R. completa puparia. The lower developmental threshold, upper developmental threshold and optimal temperature for puparial development are estimated to be 4, 34 and 26.6 °C, respectively. The thermal requirement for adult emergence after 120 days of chilling is estimated to be 2024 degree days. Percentage adult emergence declines at both higher and lower incubation temperatures. Chill duration at 5 °C for diapausing puparia has a nonlinear negative effect on the thermal requirement but no effect on percentage emergence. Insufficient chilling leads to poor synchronization of adult emergence. Greater pre‐chill duration at room temperature increases the thermal requirement and slightly decreased percentage emergence. Latitude has a negative effect on the thermal requirement. Puparia from northern California black walnut (Juglans hindsii) have a slightly greater thermal requirement than puparia from cultivated walnut (Julgans regia). There is no significant difference in puparial fresh weight or mean thermal requirement between males and females, although the positive correlation between thermal requirement and puparial fresh weight is stronger for females than males. The effects of temperature and other environmental factors on the post‐diapause development of R. completa are discussed in relation to observations from other Rhagoletis species.

Clarifying the Conservation Status of Northern California Black Walnut (Juglans hindsii) Using Microsatellite Markers

The conservation status of the northern California black walnut (Juglans hindsii (Jeps.) Jeps. ex R. E. Sm.) has been a source of considerable confusion and controversy. Although not currently legally protected by either Federal or State Endangered Species Acts, this species is given conservation status by the California Native Plant Society and the California Department of Fish and Wildlife, and some California counties require mitigation for removal of individuals of this species, especially older trees. Despite the current widespread distribution in northern California and southern Oregon of trees that match J. hindsii morphologically, there are only three or four sites where the species is known to have occurred prior to extensive settlement of California by Europeans in the mid-19th century. This has led to the suspicion that trees found in other places may not be genetically pure J. hindsii but may instead be descendants of lineages that experienced past gene flow from one or more other species. In addition, despite its more distant relationship, the cultivated walnut (J. regia L.) readily hybridizes (as the male parent) with J. hindsii, producing morphologically identifiable ‘Paradox' hybrids, which occur spontaneously and are widely planted as rootstocks and street trees. Finally, recent collections of J. hindsii from southern California have raised questions about the respective geographic distributions of J. hindsii and southern California black walnut (J. californica S. Watson). We analyzed genotypes at 10 microsatellite loci for 158 mostly wild J. hindsii trees, as well as some orchard waifs, from 10 counties in northern and southern California and one county in southern Oregon, including representatives of putative original native populations. We also sampled several Paradox hybrids, 10-20 standards for each of the five North American black walnut species, and six standards for J. regia. Bayesian cluster analyses with the program STRUCTURE revealed that at least 71.5% of the putatively wild J. hindsii represent genetically pure members of that species, while the remaining trees show evidence of past hybridizations with one or more of the other North American black walnut species. We found no evidence of introgression of J. regia into J. hindsii. The results suggest that individual J. hindsii trees should not have conservation status.

Genomic Variation Among and Within Six Juglans Species.

Genomic analysis in Juglans (walnuts) is expected to transform the breeding and agricultural production of both nuts and lumber. To that end, we report here the determination of reference sequences for six additional relatives of Juglans regia: Juglans sigillata (also from section Dioscaryon), Juglans nigra, Juglans microcarpa, Juglans hindsii (from section Rhysocaryon), Juglans cathayensis (from section Cardiocaryon), and the closely related Pterocarya stenoptera. While these are ‘draft’ genomes, ranging in size between 640Mbp and 990Mbp, their contiguities and accuracies can support powerful annotations of genomic variation that are often the foundation of new avenues of research and breeding. We annotated nucleotide divergence and synteny by creating complete pairwise alignments of each reference genome to the remaining six. In addition, we have re-sequenced a sample of accessions from four Juglans species (including regia). The variation discovered in these surveys comprises a critical resource for experimentation and breeding, as well as a solid complementary annotation. To demonstrate the potential of these resources the structural and sequence variation in and around the polyphenol oxidase loci, PPO1 and PPO2 were investigated. As reported for other seed crops variation in this gene is implicated in the domestication of walnuts. The apparently Juglandaceae specific PPO1 duplicate shows accelerated divergence and an excess of amino acid replacement on the lineage leading to accessions of the domesticated nut crop species, Juglans regia and sigillata.

Effects of Fire-retardant Treatment and Burl Wood Structure on Three-dimensional Changes of Sandwich Panels Made from Walnut Decorative Veneer

The effects of a fire-retardant treatment and burl wood structure on the three-dimensional changes of aircraft sandwich panels were evaluated. Unvarnished and varnished panels with an outer decorative layer made from walnut burl (Juglans hindsii L.) were studied. Half of the samples from each type of panel received a fire-retardant treatment (phosphate-based) on all three layers of the decorative plywood. The other half had the two inner layers treated and the outer layer was left untreated. Three different wood areas formed by rotary peeling and by the grain orientation from the burl structure were identified and their veneer surfaces were separately studied. Samples pre-conditioned at 20 °C and 40% relative humidity (RH) underwent adsorption (25 °C, 90% RH) and then desorption (25 °C, 40% RH) treatments. Changes in the moisture content (MC), swelling, shrinkage, roughness, and waviness were measured after each moisture exposure condition. The results showed that the fire-retardant treatment significantly increased the MC, swelling, shrinkage, roughness, and waviness of the unvarnished and varnished panels. This treatment also affected the roughness and waviness of the burl wood structure for the unvarnished panels. The effect of this anatomical feature was not noticeable in the varnished panels.

A SURVEY OF CHERRY LEAF ROLL VIRUS IN INTENSIVELY MANAGED GRAFTED ENGLISH (PERSIAN) WALNUT TREES IN ITALY

Blackline disease, caused by Cherry leaf roll virus (CLRV), is considered a serious threat limiting English walnut (Juglans regia) production in Italy and the EU if walnut species other than J. regia e.g. ‘Paradox’ hybrid (J. regia × J. hindsii), French hybrid (J. regia × J. major or J. regia × J. nigra) or northern California black walnut (J. hindsii) are used as the rootstock. The virus transmissibility by pollen as well as latent infections can result in the spread of CLRVcontaminated propagative material, which is a major means of the virus dispersal by human activities. In 2014 and 2015 to ascertain the presence and the distribution of blackline symptoms in commercial orchards and to provide a description of the symptomatology, visual inspections and double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) analyses were carried out on 1,684 walnut trees in four different intensively managed grafted English walnut orchards in northeast Italy (Veneto Region). Trees with clear blackline symptoms at the scion-rootstock junction, often associated with general decline of the plant, were found only in one commercial orchard in northeast Italy on trees older than ten years of cvs. ‘Tulare’ and ‘Chandler’, grafted onto ‘Paradox’ rootstock. To our knowledge this is the first report of CLRV (blackline) decline and death in a commercial walnut orchard in Italy.

Luther Burbank’s Contributions to Walnuts

Luther Burbank began making controlled crosses between walnut species in the late 19th century after hearing about a “supposed natural European hybrid walnut.” He crossed Juglans hindsii (northern California black walnut) × J. regia (Persian walnut) and produced progeny that he named ‘Paradox’ because of its extremely fast growth and other “anomalies.” He also crossed two American species, J. hindsii × J. nigra (eastern black walnut), producing ‘Royal’ walnut progeny that were fast-growing and prolific nut producers. A third interspecific hybrid was a cross between J. ailantifolia (Japanese walnut) × J. regia that resulted in extremely vigorous progeny but was not named. He observed segregation in the F2 populations and described giants and dwarfs as reversions to ancestral forms. Luther Burbank also made selections for walnut scion cultivars and was especially interested in thin-shelled nuts. He collected seeds from a J. regia growing in San Francisco because it produced regularly and had very high-quality nuts with relatively thin but poorly sealed shells. He selected one of its seedlings as ‘Santa Rosa Soft-Shell’ and described it as bearing large crops of nuts that were nearly white with thin shells and delicious white meat. Burbank’s contributions to the walnut industry endure to this day, especially through the widespread use of seedling and clonal ‘Paradox’ walnut rootstocks.

THE CALIFORNIA WALNUT IMPROVEMENT PROGRAM: SCION BREEDING AND ROOTSTOCK DEVELOPMENT

The goals of the University of California-Davis Walnut Improvement Program are to develop improved scion and rootstock cultivars for the California walnut industry and to utilize emerging genomic tools to increase breeding efficiency. Scion breeding objectives include new cultivars with early harvest dates, light kernel color, and resistance to blight and the development of scions with resistance to Cherry leaf roll virus. Rootstock development is directed at genetic resistance to soil-borne pathogens including Phytophthora, nematodes, and crown gall. Advances in clonal propagation methods, particularly micropropagation, have allowed replicated testing of genotypes of interest and have facilitated an expanding commercial production of improved clonal rootstocks. RX1, a Juglans microcarpa × Juglans regia hybrid with resistance to Phytophthora, and VX211, a very vigorous Juglans hindsii × J. regia hybrid exhibiting nematode tolerance, have been released for commercial use. Phenotyping, DNA mapping, and bioinformatic analyses are being combined to generate markers for traits of interest, including lateral bearing, kernel color, and harvest date, and will be used to improve selection efficiency. The Walnut Improvement Program is an integrated effort that depends on research cooperation with pathologists, molecular biologists, and UC Extension farm advisors, and relies on the assistance of commercial nurseries and walnut growers for planting and evaluation of statewide trials.

Incidence of Agrobacterium tumefaciens Biovar 1 in and on 'Paradox' (Juglans hindsii × Juglans regia) Walnut Seed Collected from Commercial Nurseries.

The walnut rootstock 'Paradox' (Juglans hindsii × J. regia) is susceptible to Agrobacterium tumefaciens, which often results in a high incidence of crown gall in nursery or walnut production orchards. Though A. tumefaciens is susceptible to the commonly used preplant soil fumigants, crown gall incidence can rise above acceptable levels. We examined the ability of Paradox seed to acquire A. tumefaciens as a function of harvest method used prior to planting. Over a 2-year period at two participating commercial nurseries, Paradox seed were collected directly from the mother tree without contacting the soil or gathered after sitting on the orchard floor for up to 28 days. A. tumefaciens was never detected in or on the 2,650 seeds collected directly from the mother tree. Both virulent and avirulent A. tumefaciens strains were detected in and on the husk of nuts incubated on the orchard floor at a frequency directly proportional to the time spent on the orchard floor. Regardless of A. tumefaciens contamination in or on the husk, A. tumefaciens was never detected in the seed interior. Avoiding soilborne populations of A. tumefaciens at the time of seed collection will play an important role in managing crown gall.

Evaluating Paradox Walnut Rootstocks for Resistance to Armillaria Root Disease

The most common rootstock for Juglans regia (Persian or “English” walnut) in California is Paradox, typically a hybrid of J. hindsii (Northern California black walnut) × J. regia. Unfortunately, Paradox is very susceptible to Armillaria root disease. The relative resistance to Armillaria mellea of six clonally propagated Paradox rootstocks (AX1, Px1, RR4 11A, RX1, Vlach, VX211) was evaluated and compared with that of clonally propagated J. hindsii rootstock selection W17, J. regia scion cultivar Chandler, and Pterocarya stenoptera (Chinese wingnut). In a growth-chamber assay, plants were micropropagated and rooted in vitro before inoculating the culture medium with A. mellea. At two months post-inoculation, the most resistant and susceptible Paradox rootstocks were AX1 and VX211, respectively, with 9% vs. 70% mortality, and this finding was consistent across three isolates of A. mellea and three replicate experiments. This broad range of resistance within Paradox is consistent with past field trials that tested other genotypes. Our finding of similarly high susceptibility of ‘Chandler’ and W17 (61% vs. 69% mortality) is in contrast to two field trials, in which other J. regia genotypes were more susceptible than those of J. hindsii. A third trial, however, identified some J. regia genotypes as more resistant than those of J. hindsii. Therefore, it is possible that W17, which was not previously tested, is an Armillaria-susceptible genotype of J. hindsii. Based on our findings of repeatable mortality levels across three isolates of A. mellea and three replicate experiments, the growth-chamber assay has promise, albeit with confirmed resistant and susceptible controls, for identifying putative resistant rootstocks (e.g., AX1) in preparation for a field trial with controlled inoculations.

Role of Systemic Agrobacterium tumefaciens Populations in Crown Gall Incidence on the Walnut Hybrid Rootstock 'Paradox'.

Greater than 75% of English walnut production in the United States occurs on the walnut rootstock Juglans hindsii × J. regia 'Paradox', which is highly susceptible to infection by Agrobacterium tumefaciens. When seed were germinated and grown in the presence of A. tumefaciens, in the absence of wounding, 94% of the seedlings exhibited tumors while 89% contained systemic A. tumefaciens populations. When seedlings were wound inoculated, A. tumefaciens established endophytic populations in stem tissue and often migrated from the site of infection. Distribution of A. tumefaciens in the stem was random and may exhibit seasonal variation. A. tumefaciens populations in root tissue were more readily detected than in stem tissue and may serve as a reservoir for subsequent infection of the aerial portions of the tree. Importantly, 7% of inoculated, asymptomatic seedlings contained endophytic populations of A. tumefaciens. In all, 17% of seedlings inoculated as seeds developed galls at secondary stem-wound sites. These results provide an ecological and epidemiological foundation upon which to modify existing tree-handling practices in both nursery and orchard production environments to manage crown gall incidence.

An analysis of the larval instars of the walnut twig beetle, Pityophthorus juglandis Blackman (Coleoptera: Scolytidae), in northern California black walnut, Juglans hindsii, and a new host record for Hylocurus hirtellus

By measuring and analyzing larval head capsule widths, we determined that a northern California population of the walnut twig beetle, Pityophthorus juglandis Blackman (Coleoptera: Scolytidae), has three larval instars. We also developed rules to classify P. juglandis larval instars. Overlap in the ranges of widths among consecutive instars was addressed by fitting a distribution model to the data and determining limits for head capsule width classes and probabilities of instar misclassification. Growth ratios for laboratory and field samples of P. juglandis were consistent with the Brooks-Dyar rule and with results in the literature for other bark and ambrosia beetle species. As a consequence of branch dissection to retrieve larvae of P. juglandis, we established that northern California black walnut, Juglans hindsii (Jeps.) Jeps. ex R.E. Sm., is a host for the xylophagous “California hardwood bark beetle,” Hylocurus hirtellus (LeConte) (Coleoptera: Scolytidae), and that H. hirtellus is associated with P. juglandis beneath the bark of small diameter branches. It appears that in northern California, univoltine H. hirtellus is active in flight from early March through early May, with the majority of the adults dispersing in April. The females are the colonizing sex.

Resistance to Phytophthora and Graft Compatibility with Persian Walnut among Selections of Chinese Wingnut

Seedlings from seven open-pollinated selections of Chinese wingnut (Pterocarya stenoptera) (WN) representing collections of the USDA-ARS National Clonal Germplasm Repository at Davis, CA, and the University of California at Davis were evaluated as rootstocks for resistance to Phytophthora cinnamomi and P. citricola and graft compatibility with scions of five cultivars of Persian walnut (Juglans regia). Seedlings of Northern California black walnut (NCB) (J. hindsii) and Paradox hybrid (PH) (typically J. hindsii × J. regia) were used as standards. In greenhouse experiments, potted plants of the rootstocks were subjected to intermittent flooding in soil artificially infested with the pathogens. All WN seedlings were relatively resistant to the pathogens (means of 0% to 36% of root and crown length rotted) compared with NCB (44% to 100%) and PH seedlings (11% to 100%). Negligible disease occurred in flooded control soil without the pathogens. In 9-year graft compatibility trials in an orchard, NCB and PH rootstocks supported relatively good survival and growth of all tested scion cultivars (‘Chandler’, ‘Hartley’, ‘Serr’, ‘Tulare’, and ‘Vina’; final scion survival 80% to 100%, mean scion circumference increase 292 to 541 mm), whereas results with WN were mixed. Wingnut rootstocks from all sources were incompatible with ‘Chandler’ (final scion survival 20% to 60%, scion circumference increase 17 to 168 mm). Conversely, all WN rootstocks from all sources were compatible with ‘Tulare’ and ‘Vina’ (final scion survival 80% to 100%, scion circumference increase 274 to 556 mm). Use of the WN rootstocks produced variable results in ‘Hartley’ and ‘Serr’ (final scion survival 10% to 100%, mean scion circumference increase 69 to 542 mm). There was a tendency for more rootstock sprouts on WN selections than on NCB or PH. In a commercial walnut orchard infested with P. cinnamomi, ‘Hartley’ survived and grew markedly better on WN selections than on PH. High resistance to P. cinnamomi and P. citricola was common to all of the WN selections. The results indicate that WN selections may be useful rootstocks for cultivars Tulare and Vina in soils infested with P. cinnamomi or P. citricola and that WN selections may contribute valuable resistance to these pathogens in walnut rootstock breeding efforts.

DETECTION AND IMPLICATION OF EARLY AGROBACTERIUM TUMEFACIENS INFECTION OF PARADOX SEEDS AND SEEDLINGS

Paradox (Juglans hindsii × J. regia), the dominant rootstock used in California, USA walnut production, has many desirable horticultural characteristics, but is highly susceptible to crown gall. Crown gall, caused by the bacterium Agrobacterium tumefaciens, is not consistently controlled with soil fumigation. This observation, in conjunction with observations of galls occurring in above ground grafting wounds, led to the hypothesis that A. tumefaciens inoculum may be endophytic in walnut seedlings. Paradox seeds collected directly from commercial Paradox mother trees (i.e., Juglans hindsii) were free of A. tumefaciens. Sampling conducted at multiple times during seed production revealed that seed-borne A. tumefaciens populations increased as a function of contact time with the orchard floor. In greenhouse experiments, seeds inoculated with A. tumefaciens produce seedlings with galls at the crown and/or roots in the absence of artificial wounding. Endophytic populations of A. tumefaciens were also found in shoot and root tissue. These data suggest that A. tumefaciens inoculum may exist as a resident endophyte and that timely seed collection, limiting contact with the orchard floor, may be an important first step in preventing crown gall disease of walnut.

CROWN GALL INCIDENCE: SEEDLING PARADOX WALNUT ROOTSTOCK VERSUS OWN-ROOTED ENGLISH WALNUT TREES

Seedling 'Paradox' (Juglans hindsii × J. regia) has been the rootstock of choice for English walnut in California because of its vigor and greater tolerance of wet soil conditions. However, seedling 'Paradox' rootstock is highly susceptible to crown gall, a disease caused by the soil-borne bacterium Agrobacterium tumefaciens. In regions where tree death from walnut blackline disease (cherry leafroll virus) is prevalent, seedling English rootstock is used to avoid the hypersensitive response at the graft union associated with 'Paradox'. Own-rooted English walnut trees have replaced seedling English in the nursery trade and are now often used in counties where walnut blackline disease is severe. Early observations revealed no crown gall on own-rooted trees. In 2006-2007, we conducted a systematic statewide crown gall survey of own-rooted English walnut trees and 'Paradox' rooted walnut trees at five rootstock research sites and 14 commercial orchards planted in 2003 or earlier. Crown gall incidence was determined by visual inspection, and rated as present or absent at the crown level. Every own-rooted tree site was compared to 100 English trees on 'Paradox' rootstock growing within or near each site. Across the five research sites, seedling 'Paradox' had significantly more crown gall (20.5%) compared to the own-rooted trees (0.9%). The commercial orchards had a similar pattern with seedling 'Paradox' exhibiting 22.2% crown gall versus 0.3% in the own-rooted trees.