RESUMEN
Western redcedar (Thuja plicata Donn ex D. Don) is one of the most important commercial tree species in British Columbia, generates more than $1 billion in economic activity annually and about 8-10 million trees are planted in reforestation efforts (Gregory et al. 2018). It has been selected as the provincial tree of British Columbia (BC) because of its tremendous economic, ecoogical and cultural value. However, foliar diseases such as leaf blights have serious impact on redcedar growth and may cause significant loss of tree volume (Russell, 2007). Our 2014 - 2015 surveys of western redcedar forests in coastal areas of BC indicated high incidence of a distinctive type of blight. We observed the incidence of this disease on more than 80% of western redcedar (approximately 493) trees from late May to early December. Early symptoms appeared as circular to oval, brownish to black spots (2-3 mm), 1-5 spots per branch tip, scattered at the tip margins. Sequentially, the spots enlarged and developed into necrotic lesions on both young and old leaves. More than 50 symptomatic leaves from 10 different trees were collected and rinsed in distilled water then surface-sterilized with three times washing in Tween 20 (%5 solution) for 2 minutes (each time) and %70 ethanol for 30 second (3 times repeat). Tissues from under lesions were placed on MEA (Malt Extract Agar; Phyto Tech® labs-Product ID: M498) and PDA (Potato Dextrose Agar; Phyto Tech® Labs-Product ID: P772). The plates were incubated at 21°C in the dark. They developed distinct dull white to brown, cottony colonies with each black acervuli approximately 450-500µm. The isolates produced fusiform conidia with four cells. They didn't have any distinct color. The conidiophore size was approximately 23-24 x 2-3 µm with mostly hyaline to light brown color, branched and conidiogenous was hyaline and not branched and simple. The spore size was approximately 15-20µm by 7-10µm with three transverse septa and endogenous papillae with hyaline apical appendages. Next, we collected spores and replated them on fresh MEA media culture and placed back in the incubator to produce pure cultures. We studied conidia from leaves of trees mentioned above using light and electron microscopy using Hitachi S-3500N Scanning Electron Microscope (Noshad et al. 2023). After morphological study, further identification to the species level conducted using Zambounis and Wenneker's approach (Zambounis 2019; Wenneker,2017). Genomic DNA from two single-spore isolates were isolated and sequenced. Sequences of ITS (Internal Transcribed Spacer) region amplified using primers ITS1/ITS4 and sequenced. Final sequences were deposited in Genbank and published (accession numbers OP086244 and OP086251). Blast analysis of these sequences showed 99% and 99% resemblances with T. angustata sequence (Sutton 1980). To verify its pathogenicity, we performed a comprehensive pathogenicity test to fulfill Koch's postulates. We collected their distinctive spores in an aseptic environment and standardized them (5000/ml) using a haemocytometer. Then we inoculated 100 western redcedar seedlings (three years old) by injecting standardized spore suspension solution (inoculum) using ultra-fine 0.3ml, 31G, 8mm syringes (approximately 0.1ml per inoculation site). Ten positive control seedlings were inoculated with distilled water and ten negative control seedlings were not inoculated at all. All inoculated (experimental) seedlings demonstrated same symptoms (black spots and characteristic spores) after eight weeks. None of the control seedlings showed any similar symptoms. In the next stage, we isolated and cultured spores from inoculated seedlings and studied them. The identity of reisolates confirmed using DNA sequencing. We used these spores for our next set of disease screening which was successful again. We identified Truncatella angustata (Pers.) Hughes as the causal agent for shoot-tip blight (STB) on western redcedar by examining morphological and molecular characteristics of the pathogen. This is the first report of T. angustata as a primary pathogen on western redcedar in British Columbia, Canada.
RESUMEN
Western redcedar (WRC; Thuja plicata) is a conifer of the Pacific Northwest of North America prized for its durable and rot-resistant wood. WRC has naturally low outcrossing rates and readily self-fertilizes in nature. Challenges faced in WRC breeding and propagation involve selecting trees for accelerated growth while also ensuring enhanced heartwood rot resistance and resistance to ungulate browsing, as well as mitigating potential effects of inbreeding depression. Terpenes, a large and diverse class of specialized metabolites, confer both rot and browse resistance in the wood and foliage of WRC, respectively. Using a Bayesian modelling approach, we isolated single nucleotide polymorphism (SNP) markers estimated to be associated with three different foliar terpene traits and four different heartwood terpene traits, as well as two growth traits. We found that all traits were complex, being associated with between 1700 and 3600 SNPs linked with putatively causal loci, with significant polygenic components. Growth traits tended to have a larger polygenic component while terpene traits had larger major gene components; SNPs with small or polygenic effect were spread across the genome, while larger-effect SNPs tended to be localized to specific linkage groups. To determine whether there was inbreeding depression for terpene chemistry or growth traits, we used mixed linear models for a genomic selection training population to estimate the effect of the inbreeding coefficient F on foliar terpenes, heartwood terpenes and several growth and dendrochronological traits. We did not find significant inbreeding depression for any assessed trait. We further assessed inbreeding depression across four generations of complete selfing and found that not only was inbreeding depression not significant but that selection for height growth was the only significant predictor for growth during selfing, suggesting that inbreeding depression due to selfing during operational breeding can be mitigated by increased selection intensity.
RESUMEN
Western redcedar (WRC) is an ecologically and economically important forest tree species characterized by low genetic diversity with high self-compatibility and high heartwood durability. Using sequence capture genotyping of target genic and non-genic regions, we genotyped 44 parent trees and 1520 offspring trees representing 26 polycross (PX) families collected from three progeny test sites using 45,378 SNPs. Trees were phenotyped for eight traits related to growth, heartwood and foliar chemistry associated with wood durability and deer browse resistance. We used the genomic realized relationship matrix for paternity assignment, maternal pedigree correction, and to estimate genetic parameters. We compared genomics-based (GBLUP) and two pedigree-based (ABLUP: polycross and reconstructed full-sib [FS] pedigrees) models. Models were extended to estimate dominance genetic effects. Pedigree reconstruction revealed significant unequal male contribution and separated the 26 PX families into 438 FS families. Traditional maternal PX pedigree analysis resulted in up to 51% overestimation in genetic gain and 44% in diversity. Genomic analysis resulted in up to 22% improvement in offspring breeding value (BV) theoretical accuracy, 35% increase in expected genetic gain for forward selection, and doubled selection intensity for backward selection. Overall, all traits showed low to moderate heritability (0.09-0.28), moderate genotype by environment interaction (type-B genetic correlation: 0.51-0.80), low to high expected genetic gain (6.01%-55%), and no significant negative genetic correlation reflecting no large trade-offs for multi-trait selection. Only three traits showed a significant dominance effect. GBLUP resulted in smaller but more accurate heritability estimates for five traits, but larger estimates for the wood traits. Comparison between all, genic-coding, genic-non-coding and intergenic SNPs showed little difference in genetic estimates. In summary, we show that GBLUP overcomes the PX limitations, successfully captures expected historical and hidden relatedness as well as linkage disequilibrium (LD), and results in increased breeding efficiency in WRC.
RESUMEN
We assembled the 9.8-Gbp genome of western redcedar (WRC; Thuja plicata), an ecologically and economically important conifer species of the Cupressaceae. The genome assembly, derived from a uniquely inbred tree produced through five generations of self-fertilization (selfing), was determined to be 86% complete by BUSCO analysis, one of the most complete genome assemblies for a conifer. Population genomic analysis revealed WRC to be one of the most genetically depauperate wild plant species, with an effective population size of approximately 300 and no significant genetic differentiation across its geographic range. Nucleotide diversity, π, is low for a continuous tree species, with many loci showing zero diversity, and the ratio of π at zero- to fourfold degenerate sites is relatively high (approximately 0.33), suggestive of weak purifying selection. Using an array of genetic lines derived from up to five generations of selfing, we explored the relationship between genetic diversity and mating system. Although overall heterozygosity was found to decline faster than expected during selfing, heterozygosity persisted at many loci, and nearly 100 loci were found to deviate from expectations of genetic drift, suggestive of associative overdominance. Nonreference alleles at such loci often harbor deleterious mutations and are rare in natural populations, implying that balanced polymorphisms are maintained by linkage to dominant beneficial alleles. This may account for how WRC remains responsive to natural and artificial selection, despite low genetic diversity.
