Multi-omics revealed molecular mechanism of biphenyl phytoalexin formation in response to yeast extract-induced oxidative stress in Sorbus aucuparia suspension cells.

KEY MESSAGE: Yeast extract-induced oxidative stress in Sorbus aucuparia suspension cells leads to the biosynthesis of various hormones, which activates specific signaling pathways that augments biphenyl phytoalexin production. Pathogen incursions pose a significant threat to crop yield and can have a pronounced effect on agricultural productivity and food security. Biphenyl phytoalexins are a specialized group of secondary metabolites that are mainly biosynthesized by Pyrinae plants as a defense mechanism against various pathogens. Despite previous research demonstrating that biphenyl phytoalexin production increased dramatically in Sorbus aucuparia suspension cells (SASCs) treated with yeast extract (YE), the underlying mechanisms remain poorly understood. To address this gap, we conducted an in-depth, multi-omics analysis of transcriptome, proteome, and metabolite (including biphenyl phytoalexins and phytohormones) dynamics in SASCs exposed to YE. Our results indicated that exposure to YE-induced oxidative stress in SASCs, leading to the biosynthesis of a range of hormones, including jasmonic acid (JA), jasmonic acid isoleucine (JA-ILE), gibberellin A4 (GA4), indole-3-carboxylic acid (ICA), and indole-3-acetic acid (IAA). These hormones activated specific signaling pathways that promoted phenylpropanoid biosynthesis and augmented biphenyl phytoalexin production. Moreover, reactive oxygen species (ROS) generated during this process also acted as signaling molecules, amplifying the phenylpropanoid biosynthesis cascade through activation of the mitogen-activated protein kinase (MAPK) pathway. Key genes involved in these signaling pathways included SaBIS1, SaBIS2, SaBIS3, SaPAL, SaB4H, SaOMT, SaUGT1, SaLOX2, SaPR1, SaCHIB1, SaCHIB2 and SaCHIB3. Collectively, this study provided intensive insights into biphenyl phytoalexin accumulation in YE-treated SASCs, which would inform the development of more efficient disease-resistance strategies in economically significant cultivars.

Genome-Wide Identification and Expression Profiling of Heat Shock Protein 20 Gene Family in Sorbus pohuashanensis (Hance) Hedl under Abiotic Stress.

Small heat shock proteins (HSP20s) are a significant factor in plant growth and development in response to abiotic stress. In this study, we investigated the role of HSP20s' response to the heat stress of Sorbus pohuashanensis introduced into low-altitude areas. The HSP20 gene family was identified based on the genome-wide data of S. pohuashanensis, and the expression patterns of tissue specificity and the response to abiotic stresses were evaluated. Finally, we identified 38 HSP20 genes that were distributed on 16 chromosomes. Phylogenetic analysis of HSP20s showed that the closest genetic relationship to S. pohuashanensis (SpHSP20s) is Malus domestica, followed by Populus trichocarpa and Arabidopsis thaliana. According to phylogenetic analysis and subcellular localization prediction, the 38 SpHSP20s belonged to 10 subfamilies. Analysis of the gene structure and conserved motifs indicated that HSP20 gene family members are relatively conserved. Synteny analysis showed that the expansion of the SpHSP20 gene family was mainly caused by segmental duplication. In addition, many cis-acting elements connected with growth and development, hormones, and stress responsiveness were found in the SpHSP20 promoter region. Analysis of expression patterns showed that these genes were closely related to high temperature, drought, salt, growth, and developmental processes. These results provide information and a theoretical basis for the exploration of HSP20 gene family resources, as well as the domestication and genetic improvement of S. pohuashanensis.

Cryptic population structure at the northern range margin of the service tree Sorbus domestica.

Climate change has aroused interest in planting warm- and drought-adapted trees in managed forests and urban areas. An option is to focus on tree species that occur naturally, but have centers of distribution in warmer and drier areas. However, in order to protect the species pool of genetic diversity, efforts of planting and promotion should be informed by knowledge on the local genetic diversity. Here, we studied the macro- and micro-scale population genetic structure of the rare European fruit tree Sorbus domestica at its northern range margin, in western Switzerland. New microsatellite data were combined with published data from across the European distribution of the species. Analyses revealed the presence of mainly one of two species-wide ancestral clusters, i.e., the western European cluster, with evidence that it consists of two cryptic sub-clusters. Average pairwise F ST of 0.118 was low across the range, and only allelic richness was reduced in the northern margin compared to more southern and southeastern areas of Europe. Based on our finding of considerable genetic diversity of the species in western and northern Switzerland, we suggest that a national propagation program should focus on collecting seeds from natural, high-density tree stands and propagate locally. More generally, our study shows that rare tree species in marginal areas of their distributions do not necessarily have low genetic diversity or heightened levels of inbreeding, and in those cases probably need no assisted migration in efforts to propagate them.

Metabolic and Transcriptional Stress Memory in Sorbus pohuashanensis Suspension Cells Induced by Yeast Extract.

Plant stress memory can provide the benefits of enhanced protection against additional stress exposure. Here, we aimed to explore the responses of recurrent and non-recurrent yeast extract (YE) stresses in Sorbus pohuashanensis suspension cells (SPSCs) at metabolomics and transcriptional levels. Biochemical analyses showed that the cell wall integrity and antioxidation capacity of SPSCs in the pretreated group were evidently improved. Metabolic analysis showed that there were 39 significantly altered metabolites in the pretreated group compared to the non-pretreated group. Based on the transcriptome analysis, 219 differentially expressed genes were obtained, which were highly enriched in plant-pathogen interaction, circadian rhythm-plant, oxidative phosphorylation, and phenylpropanoid biosynthesis. Furthermore, the correlation analysis of the transcriptome and metabolome data revealed that phenylpropanoid biosynthesis involved in the production of biphenyl phytoalexins may play a critical role in the memory response of SPSC to YE, and the key memory genes were also identified, including PAL1, BIS1, and BIS3. Collectively, the above results demonstrated that the memory responses of SPSC to YE were significant in almost all levels, which would be helpful for better understanding the adaptation mechanisms of medicinal plants in response to biotic stress, and laid a biotechnological foundation to accumulate favorable antimicrobial drug candidates from plant suspension cells.

Molecular Structure and Phylogenetic Analyses of the Plastomes of Eight Sorbus Sensu Stricto Species.

Sorbus L. is a genus of controversy on its taxonomic concept due to great variations in morphological characteristics. Sorbus sensu stricto species, being monophyletic, are characterized as pinnate leaves. However, phylogenetic relationships of these species are somewhat fluid based on morphological characteristics and genetic markers. In this study, the chloroplast (cp) genomes of eight Sorbus s. str. species were characterized and compared with those of twelve released species in this genus. Our results indicated that the plastomes of the twenty Sorbus species had a conserved quadripartite structure, and 129 annotated genes had the same order and showed a good collinearity. Additionally, numerous SSRs were observed in the cp genome of each Sorbus species; most of the sequence variations appeared in non-coding regions, and four intergenic regions were identified as mutation spots. By contrast, protein-coding genes showed low variations under purifying selection. The phylogenetic and molecular-dating analysis showed that Sorbus was resolved into two major clades, in which S. americana forms one clade originating at 51.78 Ma, and the rest of Sorbus formed another clade, splitting at 36.77 Ma into two sister groups with high support values. These results provide a basis for further studying the phylogenetic relationship and biogeography of Sorbus species.

Complete chloroplast genomes of Sorbus sensu stricto (Rosaceae): comparative analyses and phylogenetic relationships.

BACKGROUND: Sorbus sensu stricto (Sorbus s.s.) is a genus with important economical values because of its beautiful leaves, and flowers and especially the colorful fruits. It belongs to the tribe Maleae of the family Rosaceae, and comprises about 90 species mainly distributed in China. There is on-going dispute about its infrageneric classification and species delimitation as the species are morphologically similar. With the aim of shedding light on the circumscription of taxa within the genus, phylogenetic analyses were performed using 29 Sorbus s.s. chloroplast (cp) genomes (16 newly sequenced) representing two subgenera and eight sections. RESULTS: The 16 cp genomes newly sequenced range between 159,646 bp and 160,178 bp in length. All the samples examined and 22 taxa re-annotated in Sorbus sensu lato (Sorbus s.l.) contain 113 unique genes with 19 of these duplicated in the inverted repeat (IR). Six hypervariable regions including trnR-atpA, petN-psbM, rpl32-trnL, trnH-psbA, trnT-trnL and ndhC-trnV were screened and 44-53 SSRs and 14-31 dispersed repeats were identified as potential molecular markers. Phylogenetic analyses under ML/BI indicated that Sorbus s.l. is polyphyletic, but Sorbus s.s. and the other five segregate genera, Aria, Chamaemespilus, Cormus, Micromeles and Torminalis are monophyletic. Two major clades and four sub-clades resolved with full-support within Sorbus s.s. are not consistent with the existing infrageneric classification. Two subgenera, subg. Sorbus and subg. Albocarmesinae are supported as monophyletic when S. tianschanica is transferred to subg. Albocarmesinae from subg. Sorbus and S. hupehensis var. paucijuga transferred to subg. Sorbus from subg. Albocarmesinae, respectively. The current classification at sectional level is not supported by analysis of cp genome phylogeny. CONCLUSION: Phylogenomic analyses of the cp genomes are useful for inferring phylogenetic relationships in Sorbus s.s. Though genome structure is highly conserved in the genus, hypervariable regions and repeat sequences used are the most promising molecule makers for population genetics, species delimitation and phylogenetic studies.

High sorbic acid resistance of Penicillium roqueforti is mediated by the SORBUS gene cluster.

Penicillium roqueforti is a major food-spoilage fungus known for its high resistance to the food preservative sorbic acid. Here, we demonstrate that the minimum inhibitory concentration of undissociated sorbic acid (MICu) ranges between 4.2 and 21.2 mM when 34 P. roqueforti strains were grown on malt extract broth. A genome-wide association study revealed that the six most resistant strains contained the 180 kbp gene cluster SORBUS, which was absent in the other 28 strains. In addition, a SNP analysis revealed five genes outside the SORBUS cluster that may be linked to sorbic acid resistance. A partial SORBUS knock-out (>100 of 180 kbp) in a resistant strain reduced sorbic acid resistance to similar levels as observed in the sensitive strains. Whole genome transcriptome analysis revealed a small set of genes present in both resistant and sensitive P. roqueforti strains that were differentially expressed in the presence of the weak acid. These genes could explain why P. roqueforti is more resistant to sorbic acid when compared to other fungi, even in the absence of the SORBUS cluster. Together, the MICu of 21.2 mM makes P. roqueforti among the most sorbic acid-resistant fungi, if not the most resistant fungus, which is mediated by the SORBUS gene cluster.

Genome sequence and transcriptome of Sorbus pohuashanensis provide insights into population evolution and leaf sunburn response.

Sorbus pohuashanensis (Hance) Hedl. is a potential horticulture and medicinal plant, but its genomic and genetic backgrounds remain unknown. Here, we sequence and assemble the S. pohuashanensis reference genome using PacBio long reads. Based on the new reference genome, we resequence a core collection of 22 Sorbus spp. samples, which are divided into 2 groups (G1 and G2) based on phylogenetic and PCA analyses. These phylogenetic clusters are highly consistent with their classification based on leaf shape. Natural hybridization between the G1 and G2 groups is evidenced by a sample (R21) with a highly heterozygous genotype. Nucleotide diversity (π) analysis shows that G1 has a higher diversity than G2 and that G2 originated from G1. During the evolution process, the gene families involved in photosynthesis pathways expanded and the gene families involved in energy consumption contracted. RNA-seq data suggests that flavonoid biosynthesis and heat-shock protein (HSP)-heat-shock factor (HSF) pathways play important roles in protection against sunburn. This study provides new insights into the evolution of Sorbus spp. genomes. In addition, the genomic resources, and the identified genetic variations, especially those related to stress resistance, will help future efforts to produce and breed Sorbus spp.

Physiological and transcriptomic analyses characterized high temperature stress response mechanisms in Sorbus pohuashanensis.

Sorbus pohuashanensis (Hance) Hedl. is a Chinese native alpine tree species, but the problem of introducing S. pohuashanensis to low altitude areas has not been solved. In this study, we aimed to explore the molecular regulatory network of S. pohuashanensis in response to high-temperature stress using RNA-Sequencing technology and physiological and biochemical determination. Based on transcriptomic data, we obtained 1221 genes (752 up-regulated and 469 down-regulated) that were differentially expressed during 8 h 43℃ treatment and candidate genes were related to calcium signaling pathway, plant hormone signal transduction, heat shock factors, chaperones, ubiquitin mediated proteolysis, cell wall modification, ROS scavenging enzymes, detoxification and energy metabolism. The analysis of high temperature response at the physiological level and biochemical level were performed. The chlorophyll fluorescence parameters of leaf cells decreased, the content of osmotic regulators increased, and the activity of ROS scavenging enzymes decreased. The molecular regulatory network of S. pohuashanensis in response to high-temperature stress was preliminarily revealed in this study, which provides fundamental information improving introducing methods and discovering heat-tolerant genes involved in high-temperature stress in this species and provides a reference for other plants of the genus Sorbus.

Molecular characterization and expression analysis of small heat shock protein 17.3 gene from Sorbus pohuashanensis (Hance) Hedl. in response to abiotic stress.

Sorbus pohuashanensis, a native tree species in China that is distributed at high altitudes. However, the problem of adaptability when introducing S. pohuashanensis to low altitude areas has not been solved. sHSPs can respond and play an essential role when exposing to abiotic stresses for plants. In this study, we aimed to investigate the expression patterns underlying the abiotic stress response of the small heat shock protein 17.3 gene from S. pohuashanensis (SpHSP17.3) at growing low altitude. 1 to 4 years old seedlings of S. pohuashanensis were used as materials for the gene cloning, the tissue-specific expression and the expression analysis underlying the response to abiotic stress using the transgentic methods and qPCR. We identified the open reading frame (ORF) sequence of SpHSP17.3 of 471 bp, which encodes a 17.3 kD protein of 156 amino acids that is located in cytoplasmic. We found that SpHSP17.3 had the highest expression in the stem, followed sequentially by fruit, root, and flower. The expression level of SpHSP17.3 in the leaves was significantly induced by the high temperature (42 °C), NaCl salt and drought stress of S. pohuashanensis. Notably, the same SpHSP17.3 expression trend was detected in the SpHSP17.3-overexpressing homozygous transgenic Arabidopsis underlying the high temperature, NaCl salt and drought stress, and the SpHSP17.3-overexpressing homozygous transgenic Arabidopsis also showed higher seed germination rates under the NaCl salt stress conditions. Our results suggested that SpHSP17.3 is involved in the response to high temperature, Nacl salt, and drought stress which would play a certain effect in the adaptability of introduction and domestication of S. pohuashanensis.

Large-scale genetic admixture suggests high dispersal in an insect pest, the apple fruit moth.

Knowledge about population genetic structure and dispersal capabilities is important for the development of targeted management strategies for agricultural pest species. The apple fruit moth, Argyresthia conjugella (Lepidoptera, Yponomeutidae), is a pre-dispersal seed predator. Larvae feed on rowanberries (Sorbus aucuparia), and when rowanberry seed production is low (i.e., inter-masting), the moth switches from laying eggs in rowanberries to apples (Malus domestica), resulting in devastating losses in apple crops. Using genetic methods, we investigated if this small moth expresses any local genetic structure, or alternatively if gene flow may be high within the Scandinavian Peninsula (~850.000 km2, 55o - 69o N). Genetic diversity was found to be high (n = 669, mean He = 0.71). For three out of ten tetranucleotide STRs, we detected heterozygote deficiency caused by null alleles, but tests showed little impact on the overall results. Genetic differentiation between the 28 sampling locations was very low (average FST = 0.016, P < 0.000). Surprisingly, we found that all individuals could be assigned to one of two non-geographic genetic clusters, and that a third, geographic cluster was found to be associated with 30% of the sampling locations, with weak but significant signals of isolation-by-distance. Conclusively, our findings suggest wind-aided dispersal and spatial synchrony of both sexes of the apple fruit moth over large areas and across very different climatic zones. We speculate that the species may recently have had two separate genetic origins caused by a genetic bottleneck after inter-masting, followed by rapid dispersal and homogenization of the gene pool across the landscape. We suggest further investigations of spatial genetic similarities and differences of the apple fruit moth at larger geographical scales, through life-stages, across inter-masting, and during attacks by the parasitoid wasp (Microgaster politus).

Speciation by triparental hybridization in genus Sorbus (Rosaceae).

Hybridization associated with polyploidization and apomixis is a frequent mechanism of speciation. Sorbus is a genus with ongoing hybridization resulting in a polyploid complex with different parental lineage. Triparens is the smallest hybridogenous subgenus of Sorbus so far known to comprise only two taxa, S. intermedia and S. × liljeforsii that combine the genomes of three taxa (S. aria agg., S. aucuparia and S. torminalis). To elucidate the origins of S. dacica, S. paxiana and S. tauricola, three new trigenomic candidates formerly believed to be of biparental origin with either S. aria agg. × S. aucuparia or S. aria agg. × S. torminalis lineage we combined data from HPLC and chloroplast DNA analysing additional 33 related taxa as well. We concluded that the 'torminalis-type' flavonoid profile and the 'aucuparia-type' plastid indicate the participation of both S. torminalis and S. aucuparia resulting in the formation of S. dacica, S. paxiana and S. tauricola. Sorbus aria agg. as the third ancestor and as a necessary link to meet genes of S. torminalis and S. aucuparia in one genome is obvious from morphological features (densely tomentose undersides of leaves). The tetraploid cytotypes and obligate pseudogamy of S. dacica and S. paxiana were determined by flow cytometry and are published here for the first time. The most probable evolutionary scenario for Triparens species is: 1. a diploid sexual S. aucuparia as pollen acceptor hybridized with a tetraploid apomictic taxon from the S. aria agg. producing a triploid apomictic taxon with 'aucuparia-type' plastid inherited maternally; 2. during a second crossing event this subgenus Soraria hybrid as maternal progenitor hybridized with the sexual diploid S. torminalis (providing gene(s) of apigenin O-glucuronide synthesis) forming a tetraploid Triparens hybrid with 'aucuparia-type' plastid and 'torminalis-type' flavonoids.

De novo transcriptomic analysis and development of EST-SSRs for Sorbus pohuashanensis (Hance) Hedl.

Sorbus pohuashanensis is a native tree species of northern China that is used for a variety of ecological purposes. The species is often grown as an ornamental landscape tree because of its beautiful form, silver flowers in early summer, attractive pinnate leaves in summer, and red leaves and fruits in autumn. However, development and further utilization of the species are hindered by the lack of comprehensive genetic information, which impedes research into its genetics and molecular biology. Recent advances in de novo transcriptome sequencing (RNA-seq) technology have provided an effective means to obtain genomic information from non-model species. Here, we applied RNA-seq for sequencing S. pohuashanensis leaves and obtained a total of 137,506 clean reads. After assembly, 96,213 unigenes with an average length of 770 bp were obtained. We found that 64.5% of the unigenes could be annotated using bioinformatics tools to analyze gene function and alignment with the NCBI database. Overall, 59,089 unigenes were annotated using the Nr database(non-redundant protein database), 35,225 unigenes were annotated using the GO (Gene Ontology categories) database, and 33,168 unigenes were annotated using COG (Cluster of Orthologous Groups). Analysis of the unigenes using the KEGG (Kyoto Encyclopedia of Genes and Genomes) database indicated that 13,953 unigenes were involved in 322 metabolic pathways. Finally, simple sequence repeat (SSR) site detection identified 6,604 unigenes that included EST-SSRs and a total of 7,473 EST-SSRs in the unigene sequences. Fifteen polymorphic SSRs were screened and found to be of use for future genetic research. These unigene sequences will provide important genetic resources for genetic improvement and investigation of biochemical processes in S. pohuashanensis.

Breeding system and spatial isolation from congeners strongly constrain seed set in an insect-pollinated apomictic tree: Sorbus subcuneata (Rosaceae).

In plants, apomixis results in the production of clonal offspring via seed and can provide reproductive assurance for isolated individuals. However, many apomicts require pollination to develop functional endosperm for successful seed set (pseudogamy) and therefore risk pollination-limitation, particularly in self-incompatible species that require heterospecific pollen. We used microsatellite paternity analysis and hand pollinations to investigate pollen-limitation in Sorbus subcuneata, a threatened endemic tree that co-occurs with its congener, S. admonitor. We confirmed that S. subcuneata is an obligate pseudogamous apomict, but open-pollinated flowers rarely produced seed (flower-to-seed conversion < 1%) even though they rapidly accumulated pollen on their stigmas. Manual heterospecific pollination by S. admonitor resulted in a high flower-to-seed conversion rate (65%), however, we estimate that the ratio of self: heterospecific pollination in open-pollinated flowers was at least 22:1. Despite the efficacy of heterospecific pollination, the contribution of S. admonitor trees to paternity in seed from open-pollinated flowers of S. subcuneata decreased rapidly with the spatial separation between paternal and maternal trees. Conservation efforts aimed at maintaining species with this breeding system must therefore manage the congeners in tandem which will also maintain the potential for rare heterospecific fertilisation that typically cause rapid diversification in these lineages.

Origin and genetic differentiation of pink-flowered Sorbus hybrids in the Western Carpathians.

Background and Aims: Diversity of the genus Sorbus has been affected by interspecific hybridizations. Pink-flowered hybrid species have been insufficiently studied so far. They comprise bigenomic hybrid species derived from crosses S. aria s.l. × S. chamaemespilus and trigenomic ones, where S. aucuparia was involved as well. The main objective of the present study was to reconstruct their hybrid origins as well as to assess genetic distinction among several morphologically recognized hybrid species. Methods: Samples from putative maternal species and eight pink-flowered and two white-flowered hybrid species were collected in the Western Carpathians and the Sudetes. In total, 370 specimens were analysed. Six chloroplast microsatellites were used to infer parentage, whereas nuclear amplified fragment length polymorphism (AFLP) markers were employed for the identification of clones and patterns of genetic variation. Ploidy levels were estimated by flow cytometry on a subset of 140 individuals. Key Results: Genetic data supported their hybrid origins proposed based on flower and leaf morphology, and chloroplast DNA (cpDNA) revealed recurrent origins ( S. caeruleomontana , S. haljamovae ), even from bidirectional hybridization events ( S. zuzanae ). All bigenomic and trigenomic hybrid species (except triploid S. zuzanae ) were found to be tetraploid. In addition to polyploidy, low genetic variation and the presence of clones within and among populations were observed, suggesting predominantly apomictic reproduction of the hybrid species. Most of the described hybrid species appeared also genetically distinct. Conclusions: The data suggest that multiple hybridization events in the Western Carpathian Sorbus have led to the formation of separate, partially reproductively isolated genetic lineages, which may or may not be discriminated morphologically. Even bidirectional hybridization can produce individuals classified to the same taxon based on phenotype. For some hybrid taxa, hybridization pathways were proposed based on their genetic proximity to parental species and differences in genome sizes.

Molecular phylogenetics and historical biogeography of Sorbus sensu stricto (Rosaceae).

Explaining how plants from eastern Asia migrated to other Northern Hemisphere regions is still challenging. The genus Sorbus sensu stricto (including c. 88 species) is considered as a good example to illuminate this scenario, due to the wide distribution in the temperate zone and high diversity in the Himalayas and Hengduan Mountains. Based on four nuclear markers (LEAFY-2, GBSSI-1, SBEI and WD) and one chloroplast marker (rps16-trnK), we reconstructed the phylogenetic relationship of Sorbus using 54 taxa (60% of the genus representing all subgenera, sections, or series and geographical areas in the previous classifications), and estimated divergence time and historical biogeography of the genus. Phylogenetic analyses supported that the subgenera Sorbus and Albo-carmesinae, as defined by traditional morphological characters (such as fruit color), are suitable. However, five clades defined by phylogenetic analysis of the genus are not consistent with traditionally defined sections or series. Molecular dating and biogeographic reconstruction showed that the age of the most recent common ancestor was estimated at 41mya (95% HPD: 49-35mya) in eastern Asia. Four dispersal events are assumed to explain the wide distribution of Sorbus in the temperate zone and diversification in the edges of Qinghai-Tibet Plateau (QTP). Species dispersed from eastern Asia to the Tianshan Mountains, North America and Europe during the Oligocene and Miocene period. We found that polyploidization occurred multiple times in the subgenus Albo-carmesinae, in the Tianshan Mountains, Himalayas, and H-D Mountains. Finally, we suggest that apomixis, polyploidization, and hybridization may have combined with the multistage uplifting of Himalayas and H-D Mountains (and the subsequent increases in geologic, ecological and climatic heterogeneity) to drive the striking species diversity of Sorbus in this region.

When sexual meets apomict: genome size, ploidy level and reproductive mode variation of Sorbus aria s.l. and S. austriaca (Rosaceae) in Bosnia and Herzegovina.

BACKGROUND AND AIMS: Allopolyploidy and intraspecific heteroploid crosses are associated, in certain groups, with changes in the mating system. The genus Sorbus represents an appropriate model to study the relationships between ploidy and reproductive mode variations. Diploid S. aria and tetraploid apomictic S. austriaca were screened for ploidy and mating system variations within pure and sympatric populations in order to gain insights into their putative causalities. METHODS: Flow cytometry was used to assess genome size and ploidy level among 380 S. aria s.l. and S. austriaca individuals from Bosnia and Herzegovina, with 303 single-seed flow cytometric seed screenings being performed to identify their mating system. Pollen viability and seed set were also determined. KEY RESULTS: Flow cytometry confirmed the presence of di-, tri- and tetraploid cytotype mixtures in mixed-ploidy populations of S. aria and S. austriaca. No ploidy variation was detected in single-species populations. Diploid S. aria mother plants always produced sexually originated seeds, whereas tetraploid S. austriaca as well as triploid S. aria were obligate apomicts. Tetraploid S. aria preserved sexuality in a low portion of plants. A tendency towards a balanced 2m : 1p parental genome contribution to the endosperm was shared by diploids and tetraploids, regardless of their sexual or asexual origin. In contrast, most triploids apparently tolerated endosperm imbalance. CONCLUSIONS: Coexistence of apomictic tetraploids and sexual diploids drives the production of novel polyploid cytotypes with predominantly apomictic reproductive modes. The data suggest that processes governing cytotype diversity and mating system variation in Sorbus from Bosnia and Herzegovina are probably parallel to those in other diversity hotspots of this genus. The results represent a solid contribution to knowledge of the reproduction of Sorbus and will inform future investigations of the molecular and genetic mechanisms involved in triggering and regulating cytotype diversity and alteration of reproductive modes.

Biphenyl 4-Hydroxylases Involved in Aucuparin Biosynthesis in Rowan and Apple Are Cytochrome P450 736A Proteins.

Upon pathogen attack, fruit trees such as apple (Malus spp.) and pear (Pyrus spp.) accumulate biphenyl and dibenzofuran phytoalexins, with aucuparin as a major biphenyl compound. 4-Hydroxylation of the biphenyl scaffold, formed by biphenyl synthase (BIS), is catalyzed by a cytochrome P450 (CYP). The biphenyl 4-hydroxylase (B4H) coding sequence of rowan (Sorbus aucuparia) was isolated and functionally expressed in yeast (Saccharomyces cerevisiae). SaB4H was named CYP736A107. No catalytic function of CYP736 was known previously. SaB4H exhibited absolute specificity for 3-hydroxy-5-methoxybiphenyl. In rowan cell cultures treated with elicitor from the scab fungus, transient increases in the SaB4H, SaBIS, and phenylalanine ammonia lyase transcript levels preceded phytoalexin accumulation. Transient expression of a carboxyl-terminal reporter gene construct directed SaB4H to the endoplasmic reticulum. A construct lacking the amino-terminal leader and transmembrane domain caused cytoplasmic localization. Functional B4H coding sequences were also isolated from two apple (Malus × domestica) cultivars. The MdB4Hs were named CYP736A163. When stems of cv Golden Delicious were infected with the fire blight bacterium, highest MdB4H transcript levels were observed in the transition zone. In a phylogenetic tree, the three B4Hs were closest to coniferaldehyde 5-hydroxylases involved in lignin biosynthesis, suggesting a common ancestor. Coniferaldehyde and related compounds were not converted by SaB4H.

High molecular diversity in the true service tree (Sorbus domestica) despite rareness: data from Europe with special reference to the Austrian occurrence.

BACKGROUND AND AIMS: Sorbus domestica (Rosaceae) is one of the rarest deciduous tree species in Europe and is characterized by a scattered distribution. To date, no large-scale geographic studies on population genetics have been carried out. Therefore, the aims of this study were to infer levels of molecular diversity across the major part of the European distribution of S. domestica and to determine its population differentiation and structure. In addition, spatial genetic structure was examined together with the patterns of historic and recent gene flow between two adjacent populations. METHODS: Leaf or cambium samples were collected from 17 populations covering major parts of the European native range from north-west France to south-east Bulgaria. Seven nuclear microsatellites and one chloroplast minisatellite were examined and analysed using a variety of methods. KEY RESULTS: Allelic richness was unexpectedly high for both markers within populations (mean per locus: 3·868 for nSSR and 1·647 for chloroplast minisatellite). Moreover, there was no evidence of inbreeding (mean Fis = -0·047). The Italian Peninsula was characterized as a geographic region with comparatively high genetic diversity for both genomes. Overall population differentiation was moderate (FST = 0·138) and it was clear that populations formed three groups in Europe, namely France, Mediterranean/Balkan and Austria. Historic gene flow between two local Austrian populations was high and asymmetric, while recent gene flow seemed to be disrupted. CONCLUSIONS: It is concluded that molecular mechanisms such as self-incompatibility and high gene flow distances are responsible for the observed level of allelic richness as well as for population differentiation. However, human influence could have contributed to the present genetic pattern, especially in the Mediterranean region. Comparison of historic and recent gene flow may mirror the progress of habitat fragmentation in eastern Austria.

Phenolic and antioxidant profiles of rowan (Sorbus L.) fruits.

Fruits of different Sorbus L. species have been traditionally used in food production and in the treatment of various ailments. Besides common antioxidant active phytochemicals such as ascorbic acid, tocopherols and carotenoids, fruits are rich in phenolic compounds that are of particular interest. In this study, profiles of separate antioxidant active compounds (reducing and radical-scavenging activities) were investigated using online assays with post-column reactions. Significant variations in phenolic acids and flavonoids contents and antioxidant activity values were observed in the fruits of Sorbus L. species studied. Neochlorogenic and chlorogenic acids were determined as markers of antioxidant activity. Characteristic patterns of antioxidant profiles obtained using HPLC post-column FRAP and ABTS assays significantly depend on specific Sorbus L. species and are suitable for equivalency research of Sorbus L. fruits.