A genome variation map provides insights into the genetics of walnut adaptation and agronomic traits

Background Common walnut (Juglans regia L.) is one of the top four most consumed nuts in the world due to its health benefits and pleasant taste. Despite its economic importance, the evolutionary history and genetic control of its adaptation and agronomic traits remain largely unexplored. Results We report a comprehensive walnut genomic variation map based on whole-genome resequencing of 815 walnut accessions. Evolutionary analyses suggest that Chinese J. regia diverged from J. sigillata with extensive hybridizations after the split of the two species. In contrast to annual crops, the genetic diversity and heterozygous deleterious mutations of Chinese common walnut trees have continued to increase during the improvement process. Selective sweep analyses identify 902 genes uniquely selected in the improved common walnut compared to its progenitor population. Five major-effect loci are identified to be involved in walnut adaptations to temperature, precipitation, and altitude. Genome-wide association studies reveal 27 genomic loci responsible for 18 important agronomic traits, among which JrFAD2 and JrANR are the potentially major-effect causative genes controlling linoleic acid content and color of the endopleura of the nut, respectively. Conclusions The largest genomic resource for walnuts to date has been generated and explored in this study, unveiling their evolutionary history and cracking the genetic code for agronomic traits and environmental adaptation of this economically crucial crop tree.

Genome-Wide Identification and Transcriptional Expression Profiles of Transcription Factor WRKY in Common Walnut (Juglans regia L.)

The transcription factor WRKY is widely distributed in the plant kingdom, playing a significant role in plant growth, development and response to stresses. Walnut is an economically important temperate tree species valued for both its edible nuts and high-quality wood, and its response to various stresses is an important factor that determines the quality of its fruit. However, in walnut trees themselves, information about the WRKY gene family remains scarce. In this paper, we perform a comprehensive study of the WRKY gene family in walnut. In total, we identified 103 WRKY genes in the common walnut that are clustered into 4 groups and distributed on 14 chromosomes. The conserved domains all contained a WRKY domain, and motif 2 was observed in most WRKYs, suggesting a high degree of conservation and similar functions within each subfamily. However, gene structure was significantly differentiated between different subfamilies. Synteny analysis indicates that there were 56 gene pairs in J. regia and A. thaliana, 76 in J. regia and J. mandshurica, 75 in J. regia and J. microcarpa, 76 in J. regia and P. trichocarpa, and 33 in J. regia and Q. robur, indicating that the WRKY gene family may come from a common ancestor. GO and KEGG enrichment analysis showed that the WRKY gene family was involved in resistance traits and the plant-pathogen interaction pathway. In anthracnose-resistant F26 fruits (AR) and anthracnose-susceptible F423 fruits (AS), transcriptome and qPCR analysis results showed that JrWRKY83, JrWRKY73 and JrWRKY74 were expressed significantly more highly in resistant cultivars, indicating that these three genes may be important contributors to stress resistance in walnut trees. Furthermore, we investigate how these three genes potentially target miRNAs and interact with proteins. JrWRKY73 was target by the miR156 family, including 12 miRNAs; this miRNA family targets WRKY genes to enhance plant defense. JrWRKY73 also interacted with the resistance gene AtMPK6, showing that it may play a crucial role in walnut defense.

Identification and Quantification of Naphthoquinones and Other Phenolic Compounds in Leaves, Petioles, Bark, Roots, and Buds of Juglans regia L., Using HPLC-MS/MS

The present study was designed to identify and quantify the major phenolic compounds in different Juglans regia L. (common walnut) tissues (leaves, petioles, bark, roots, buds), to define the compositions and contents of phenolic compounds between these tissues. A total of 91 individual phenolic compounds were identified and quantified, which comprised 8 hydroxycinnamic acids, 28 hydroxybenzoic acids, 11 flavanols, 20 flavonols, 22 napthoquinones, and 2 coumarins. Naphthoquinones were the major phenolic group in leaves, petioles, bark, and buds, as >60% of those identified, while hydroxybenzoic acids were the major phenolic group in side roots, as ~50% of those identified. The highest content of phenolic compounds was in the J. regia main root, followed by side roots and buds, leaves, and 1-year-old bark; the lowest content was in petioles and 2-year-old bark. Leaves, roots, and buds of J. regia represent a valuable source of these agro-residues.

Gene flow among wild and cultivated common walnut (Juglans regia) trees in the Qinling Mountains revealed by microsatellite markers

Patterns of gene flow and gene introgression can be used to assess the risk of genetic pollution of wild forest trees from widespread cultivated trees. A comprehensive understanding of the genetic relationships and levels of gene flow among wild and cultivated common walnut (Juglans regia) has become an urgent issue. Using twelve microsatellite markers, we investigated the genetic diversity and gene flow between cultivated and wild trees of J. regia in the Qinling Mountains, China. A high level of genetic variation was detected in both cultivated and wild trees. The mean number of alleles per locus was 17.5. Observed heterozygosity (HO) and expected heterozygosity (HE) were 0.777 and 0.800, respectively. Pollination of mother trees was not by nearest neighbors, and a paternity of 60.7% of offspring evaluated could not be assigned to a local, sampled tree. Pollen flow from cultivated trees to wild trees was infrequent (5.4%), and selfing rates ranged from zero to 25.0%. Male parents were located from 0 to 1005 m from their female partners, with an average pollination distance of 285.1 m. These results are discussed in light of the cultivated species' diversity, outlining the frequent spontaneous genetic contributions from the wild to the cultivated compartment. In addition, the pollen flow parameters provide useful information about the dynamics of pollen movement within J. regia populations.

Preliminary results about the efficacy of abamectin trunk injection against the walnut husk fly (Rhagoletis completa)

The walnut husk fly (Rhagoletis completa) is one of the main pests affecting common walnut in both Europe and America. This work examines the effects of abamectin on the development of walnut husk fly larvae by injecting the product Vertimec 1.8 EC (Syngenta) into the trunks of walnut trees in Hungary. In the case of properly injected trees, the infection rate was negligible and the abamectin content in the husk samples ranged between 1.54 and 3.00 ng/g; controls show a very high (> 90%) infestation rate and an abamectin content under the detection limit. Abamectin residue measured in walnut kernel did not exceed the maximum residue limit; moreover, the active ingredient content was below the detection limit (0.0003 mg/kg) in all the collected kernel samples. Our results confirm that trunk injection is a viable method for walnut pest control.

Investigation of high-frequency vacuum drying on physical and mechanical properties of common oak (Quercus robur) and common walnut (Juglans regia) lumber

In conventional drying kilns, the drying of especially thick lumber is not ideal in terms of drying quality and drying time. Therefore, it is important to dry thick and dense lumber by high-frequency electromagnetically heated vacuum drying. In this study, common oak (Quercus robur) and common walnut (Juglans regia) selected from local sources used in rifle stock were used as experimental material. These materials were technologically examined by drying the timber for a short time using a fully automated, high frequency vacuum (HF + V) drying kiln. Physical and mechanical properties of these tree species were compared after air drying and HF + V drying processes. There were no significant changes in physical properties. However, the mechanical properties such as bending strength, modulus of elasticity, compression strength, and dynamic bending strength were decreased. According to these results, non-overloaded areas like rifle stock can use high frequency vacuum drying as the preferred drying method compared to conventional drying methods.

First record of the walnut leaf miner Caloptilia roscipennella (Lepidoptera: Gracillariidae) in Belarus, with a note on synonymy

The walnut leaf miner Caloptilia roscipennella (Hübner, 1796) is recorded for the first time in Belarus. The larvae of this species damage leaves of the common walnut (Juglans regia L.). In 2015, C. roscipennella was registered in a number of geographical locations in the Brest and Gomel provinces. In subsequent years, the species was found only in the same places. It is shown that the name C. roseipennella Anikin, Zolotukhin et Kirichenko, 2016, syn. nov., being an unjustified emendation, is a junior synonym of C. roscipennella (Hübner, 1796) in accordance with the International Code of Zoological Nomenclature, Articles 32.5.1 and 33.2.3.