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Disease severity and drought due to climate change present significant challenges to orchard productivity. This study examines the effects of spring inoculation with Pseudomonas syringae pv. syringae ( Pss ) on sweet cherry plants, cvs. Bing and Santina with varying defense responses, assessing plant growth, physiological variables (water potential, gas exchange, and plant hydraulic conductance), and the levels of abscisic acid (ABA) and salicylic acid (SA) under two summer irrigation levels. Pss inoculation elicited a more pronounced response in 'Santina' compared to 'Bing' at 14 days post-inoculation (dpi), and those plants inoculated with Pss exhibited a slower leaf growth and reduced transpiration compared to control plants during 60 dpi. During differential irrigations, leaf area was reduced 14% and 44% in Pss inoculated plants of 'Bing' and 'Santina' respectively, under well-watered (WW) conditions, without changes in plant water status or gas exchange. Conversely, water-deficit (WD) conditions led to gas exchange limitations and a 43% decrease in plant biomass compared to that under WW conditions, with no differences between inoculation treatments. ABA levels were lower under WW than under WD at 90 dpi, while SA levels were significantly higher in Pss -inoculated plants under WW conditions. These findings underscore the influence on plant growth during summer in sweet cherry cultivars that showed a differential response to Pss inoculations and how the relationship between ABA and SA changes in plant drought level responses.

One of the causal agents of bacterial canker is Pseudomonas amygdali pv. morsprunorum -Pam (formerly Pseudomonas syringae pv. morsprunorum ). Recently detected in Chile, Pam is known to cause lesions in the aerial parts of the plant, followed by more severe symptoms such as cankers and gummosis in the later stages of the disease. This study presents the design of PCR and LAMP detection methods for the specific and sensitive identification of Pseudomonas amygdali pv. morsprunorum (Pam) from cherry trees. Twelve Pseudomonas isolates were collected, sequenced, and later characterized by Multi-locus Sequence Analysis (MLSA) and Average Nucleotide Identity by blast (ANIb). Three of them (11116B2, S1 Pam, and S2 Pam) were identified as Pseudomonas amygdali pv. morsprunorum and were used to find specific genes through RAST server, by comparing their genome with that of other Pseudomonas , including isolates from other Pam strains. The effector gene HopAU1 was selected for the design of primers to be used for both techniques, evaluating sensitivity and specificity, and the ability to detect Pam directly from plant tissues. While the PCR detection limit was 100 pg of purified bacterial DNA per reaction, the LAMP assays were able to detect up to 1 fg of purified DNA per reaction. Similar results were observed using plant tissues, LAMP being more sensitive than PCR, including when using DNA extracted from infected plant tissues. Both detection methods were tested in the presence of 30 other bacterial genera, with LAMP being more sensitive than PCR.

Chile is an important producers of sweet cherry ( Prunus avium L.), with a total of 356,385 t exported in the 2021 to 2022 season. The production area includes most of the country's regions. Bacterial samples were isolated in 2017 and 2018 from 18 commercial sweet cherry orchards with canker disease. From one of this samples collected in the spring of 2018, was isolated the strain A2M176 from buds of trees that presented canker and gomosis in Malloa locality (34°23' 46'' S 71°01' 39'' W). The strain produced fluorescent pigment on King's B agar medium. Is oxidase and arginine dihydrolase negative, potato soft rot positive and showed a slight degree of tobacco hypersensitivity. It was able to growth up to 0.8 mM (200 ppm) of CuSO4·5H2O. The strain A2M176 was deposited in the Colección Chilena de Recursos Genéticos Microbianos (CChRGM) under the no. RGM 3342. The DNA of this strain was extracted from a pure culture using silica spin columns (Epoch Life Science Inc., Sugar Land, USA). The complete DNA was sequenced using HiSeq with 150 bp paired-end at GENEWIZ (New York, USA). Raw data was checked using FASTQC and trimmed with BBDuk. The genome was assembled using Unicycler v0.4.9 with defaulf settings and annotated with Prokaryotic Genome Annotation Pipeline (PGAP) v4.3. The reads and genomes were uploaded to GenBank under the BioProyect no. PRJNA750090, BioSample no. SAMN26870984 and assembly no. GCA_022936465.1. The sequenced genome was compared through Average Nucleotide Identity algorithm (ANI) using FastANI v1.33 to compare with closest complete genome available on NCBI. The strain A2M176 was identified as P. viridiflava with ANI value of 98.06% with the strain p22.E7 (GCF_900585495). Maximum likelihood phylogenetic estimation clustered strain A2M176 with other P. viridiflava strains with 95% bootstrap. The pathogenicity of the strain was tested inoculating immature cherry fruits with a needle with a bacterial suspension (1x10 8 CFU/ml). The inoculated fruits were placed at room temperature in a humid chamber for 10 d. Soft rot lesions were observed, which appeared at 6 days post-inoculation (DPI). The control fruits treated with sterile water did not show symptoms. Further analyses in the genome of strain A2M176 led to identify genes related to pathogenicity, such as the effector gene avrE and the regulator gen HrpL, suggesting the pathogenic capacity of the strain. Also, there were identify genes of two known Pseudomonas copper resistance mechanisms, the cus and cop operon. These genes were found part of the copABCDns cluster similar to what was observed in Pseudomonas from Mango. Presence of P. viridiflava strains causing fruit rot in P. avium is not surprising, since P. viridiflava has a wide host range and causes a variety of symptoms in different plant parts, including stems, leaves, and blossoms. P. viridiflava represents one of the multiple phylogroups found within the P. syringae species complex.  To our knowledge, this is the first report of a strain of P. viridiflava copper resistant causing infection on sweet cherries in Chile.

Pseudomonas sp. strain RGM 3321 is a phyllosphere endophyte from Fragaria chiloensis subsp. chiloensis f. patagonica that harbors genes associated with plant growth promotion pathways, as well as genes typically found in plant pathogens.

Color acquisition is one of the most distinctive features of fruit development and ripening processes. The color red is closely related to the accumulation of polyphenolic compounds, mainly anthocyanins, during sweet cherry fruit maturity. In non-climacteric fruit species like sweet cherry, the maturity process is mainly controlled by the phytohormone abscisic acid (ABA), though other hormones may also play a role. However, the coordinated stage-specific production of polyphenolic compounds and their relation with hormone content variations have not been studied in depth in sweet cherry fruits. To further understand the accumulation dynamics of these compounds (hormones and metabolites) during fruit development, two sweet cherry cultivars ("Lapins" and "Glenred") with contrasting maturity timing phenotypes were analyzed using targeted metabolic analysis. The ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) approach revealed that phenolic acids, flavonols, and flavan-3-ols accumulated mainly until the straw-yellow stage in the early-maturing cultivar, while accumulation was mainly at the green stage in the mid-maturing cultivar, suggesting a cultivar-dependent stage-specific production of secondary metabolites. In the mid-maturing cultivar, anthocyanins were detected only from the red stage onward, whereas detection began at the pink stage in the early-maturing cultivar. ABA negatively correlated ( p -value < 0.05) with the flavonols and flavan-3-ols in both cultivars. ABA and anthocyanin content increased at the same time in the early-season cultivar. In contrast, anthocyanins accumulated and the pink color initiation started several days after the ABA increase in the mid-maturing cultivar. Differential accumulation patterns of GA 4 , a ripening antagonizing hormone, between the cultivars could explain this difference. These results showed that both red-colored cultivars presented different accumulation dynamics of phenolic compounds and plant hormones during fruit development, suggesting underlying differences in the sweet cherry fruit color evolution.

Gibberellin (GA) negatively affects color evolution and other ripening-related processes in non-climacteric fruits. The bioactive GA, gibberellic acid (GA 3 ), is commonly applied at the light green-to-straw yellow transition to increase firmness and delay ripening in sweet cherry (Prunus avium L.), though causing different effects depending on the variety. Recently, we reported that GA 3 delayed the IAD parameter (a ripening index) in a mid-season variety, whereas GA 3 did not delay IAD but reduced it at ripeness in an early-season variety. To further explore this contrasting behavior between varieties, we analyzed the transcriptomic responses to GA 3 applied on two sweet cherry varieties with different maturity time phenotypes. At harvest, GA 3 produced fruits with less color in both varieties. Similar to our previous report, GA 3 delayed fruit color initiation and IAD only in the mid-season variety and reduced IAD at harvest only in the early-season variety. RNA-seq analysis of control- and GA 3 -treated fruits revealed that ripening-related categories were overrepresented in the early-season variety, including 'photosynthesis' and 'auxin response'. GA 3 also changed the expression of carotenoid and abscisic acid (ABA) biosynthetic genes in this variety. In contrast, overrepresented categories in the mid-season variety were mainly related to metabolic processes. In this variety, some PP2Cs putative genes were positively regulated by GA 3 , which are negative regulators of ABA responses, and MYB44-like genes (putative repressors of PP2Cs expression) were downregulated. These results show that GA 3 differentially modulates the transcriptome at the onset of ripening in a variety-dependent manner and suggest that GA 3 impairs ripening through the modification of ripening associated gene expression only in the early-season variety; whereas in the mid-season variety, control of the ripening timing may occur through the PP2C gene expression regulation. This work contributes to the understanding of the role of GA in non-climacteric fruit ripening.

Background: The nematicidal effects of fluopyram were evaluated for the suppression of Meloidogyne ethiopica and migratory nematodes, Xiphinema americanum s. l., Hemicycliophora spp. and Pratylenchus spp., in two commercial tomato greenhouses in Chile. The effects of fluopyram on plant-parasitic nematodes, plant vigor and fruit yield were determined., Results: Fluopyram demonstrated good potential for the management of M. ethiopica and migratory nematodes, especially during the early stages of evaluation (30-60 days after planting). There were also improvements in vigor of treated plants. A general trend in improved fruit quality was also observed, however, no significant differences in total yield were found between treatments., Conclusions: Our study is one of the first evaluations of fluopyram under field conditions against M. ethiopica. The findings suggest that this new nematicide has good potential for the management of M. ethiopica and some migratory nematodes in tomatoes cropped in greenhouses in the Metropolitan Region of Chile. © 2021 Society of Chemical Industry., (© 2021 Society of Chemical Industry.)

Hexokinases (HXKs) and fructokinases (FRKs) are the only two families of enzymes in plants that have been identified as able to phosphorylate Glucose (Glc) and Fructose (Fru). Glc can only be phosphorylated in plants by HXKs, while Fru can be phosphorylated by either HXKs or FRKs. The various subcellular localizations of HXKs in plants indicate that they are involved in diverse functions, including anther dehiscence and pollen germination, stomatal closure in response to sugar levels, stomatal aperture and reducing transpiration. Its association with modulating programmed cell death, and responses to oxidative stress and pathogen infection (abiotic and biotic stresses) also have been reported. To extend our understanding about the function of HXK-like genes in the response of Prunus rootstocks to abiotic stress, we performed a detailed bioinformatic and functional analysis of hexokinase 3-like genes (HXK3s) from two Prunus rootstock genotypes, 'M.2624' (Prunus cerasifera Ehrh × P. munsoniana W.Wight & Hedrick) and 'M.F12/1' (P. avium L.), which are tolerant and sensitive to hypoxia stress, respectively. A previous large-scale transcriptome sequencing of roots of these rootstocks, showed that this HXK3-like gene that was highly induced in the tolerant genotype under hypoxia conditions. In silico analysis of gene promoters from M.2624 and M.F12/1 genotypes revealed regulatory elements that could explain differential transcriptional profiles of HXK3 genes. Subcellular localization was determinates by both bioinformatic prediction and expression of their protein fused to the green fluorescent protein (GFP) in protoplasts and transgenic plants of Arabidopsis. Both approaches showed that they are expressed in plastids. Metabolomics analysis of Arabidopsis plants ectopically expressing Prunus HXK3 genes revealed that content of several metabolites including phosphorylated sugars (G6P), starch and some metabolites associated with the TCA cycle were affected. These transgenic Arabidopsis plants showed improved tolerance to salt and drought stress under growth chamber conditions. Our results suggest that Prunus HXK3 is a potential candidate for enhancing tolerance to salt and drought stresses in stone fruit trees and other plants.

Several phytohormones modulate ripening in non-climacteric fruits, which is triggered by abscisic acid (ABA). Gibberellins (GAs) are present during the onset of ripening in sweet cherry fruits, and exogenous gibberellic acid (GA 3 ) application delays ripening, though this effect is variety-dependent. Although an ABA accumulation delay has been reported following GA 3 treatment, the mechanism by which GA modulates this process has not been investigated at the molecular level in sweet cherry. Therefore, the aim of this work is to analyze the effect of GA 3 on the fruit ripening process and the transcript levels of ABA pathway orthologs in two varieties having different maturity time phenotypes. The early-season variety had a rapid transition from yellow to pink fruit color, whereas pink color initiation took longer in the mid-season variety. GA 3 increased the proportion of lighter colored fruits at ripeness in both varieties, but it produced a delay in IAD-a ripening index-only in the mid-season variety. This delay was accompanied by an increased transcript abundance of PavPP2Cs , which are putative negative regulators of the ABA pathway. On the other hand, the early-season variety had increased expression of PavCYP707A2 -a putative ABA catabolic gene-, and reduced transcript levels of PavPP2Cs and SnRK2s after the GA 3 treatment. Together these results show that GA modulates fruit ripening, exerting its action in part by interacting with the ABA pathway in sweet cherry.

Prunus rootstock belonging to subgenera Amygdalus (peach), Prunus (plum) and Cerasus (cherry) are either from the same species as the scion or another one. The number of inter-species (including inter-subgenera) hybrids has increased as a result of broadening the genetic basis for stress (biotic and abiotic) resistance/tolerance. Identifying genes associated with important traits and responses requires expression analysis. Relative quantification is the simplest and most popular alternative, which requires reference genes (housekeeping) to normalize RT-qPCR data. However, there is a scarcity of validated housekeeping genes for hybrid Prunus rootstock species. This research aims to increase the number of housekeeping genes suitable for Prunus rootstock expression analysis. Twenty-one candidate housekeeping genes were pre-selected from previous RNAseq data that compared the response of root transcriptomes of two rootstocks subgenera to hypoxia treatment, 'Mariana 2624' (P. cerasifera Ehrh.× P. munsoniana W. Wight & Hedrick), and 'Mazzard F12/1' (P. avium L.). Representing groups of low, intermediate or high levels of expression, the genes were assayed by RT-qPCR at 72 hours of hypoxia treatment and analyzed with NormFinder software. A sub-set of seven housekeeping genes that presented the highest level of stability were selected, two with low levels of expression (Unknown 3, Unknown 7) and five with medium levels (GTB 1, TUA 3, ATPase P, PRT 6, RP II). The stability of these genes was evaluated under different stress conditions, cold and heat with the hybrid 'Mariana 2624' and N nutrition with the hybrids 'Colt' (P. avium × P. pseudocerasus Lindl.) and 'Garnem' [P. dulcis Mill.× (P. persica L.× P. davidiana Carr.)]. The algorithms of geNorm and BestKeeper software also were used to analyze the performance of these genes as housekeepers. Stability rankings varied according to treatments, genotypes and the software for evaluation, but the gene GBT 1 often had the highest ranking. However, most of the genes are suitable depending on the stressor and/or genotype to be evaluated. No optimal number of reference genes could be determined with geNorm software when all conditions and genotypes were considered. These results strongly suggest that relative RT-qPCR should be analyzed separately with their respective best housekeeper according to the treatment and/or genotypes in Prunus spp. rootstocks., Competing Interests: The authors have declared that no competing interests exist.

In sweet cherry trees, flowering is commercially important because the flowers, after fertilization, will generate the fruits. In P. avium, the flowering induction and flower organogensis are the first developmental steps towards flower formation and they occur within specialized organs known as floral buds during the summer, nine months before blooming. During this period the number of floral buds per tree and the bud fruitfulness (number of flowers per bud) are stablished affecting the potential yield of orchards and the plant architecture. The floral bud development is sensitive to any type of stress and the hotter and drier summers will interfere with this process and are calling for new adapted cultivars. A better understanding of the underlying molecular and hormonal mechanisms would be of help, but unlike the model plant Arabidopsis, very little is known about floral induction in sweet cherry. To explore the molecular mechanism of floral bud differentiation, high-throughput RNA sequencing was used to detect differences in the gene expression of P. avium floral buds at five differentiation stages. We found 2,982 differentially expressed genes during floral bud development. We identified genes associated with floral initiation or floral organ identity that appear to be useful biomarkers of floral development and several transcription factor families (ERF, MYB, bHLH, MADS-box and NAC gene family) with novel potential roles during floral transition in this species. We analyzed in deep the MADS-box gene family and we shed light about their key role during floral bud and organs development in P. avium. Furthermore, the hormonal-related signatures in the gene regulatory networks and the dynamic changes of absicic acid, zeatin and indolacetic acid contents in buds suggest an important role for these hormones during floral bud differentiation in sweet cherry. These data provide a rich source of novel informacion for functional and evolutionary studies about floral bud development in sweet cherry and new tools for biotechnology and breeding., Competing Interests: The authors have declared that no competing interests exist.

Maqui ( Aristotelia chilensis [Molina] Stunz) is a small dioecious tree, belonging to the Elaeocarpaceae family. Maqui fruit has high levels of antioxidant activity, which are due to elevated anthocyanin and polyphenol content. Here we describe a draft genome sequence data of maqui ( A. chilensis ). The genomic sequence datasets were obtained using Illumina NextSeq platform. Nucleotide sequences of raw reads and the assembled draft genome are available at NCBI's Sequence Read Archive as BioProject PRJNA544858. Also, a total of 210067 microsatellite or simple sequence repeat (SSR) markers were identified., (© 2019 The Authors.)

In this work, we partially sequenced genomes of two Atriplex species ( A. deserticola Phil. and A. atacamensis Phil.), using Illumina technology (Hiseq 2500 paired-end system) and de novo assembly strategy. Raw data of A. deserticola and A. atacamensis are available from NCBI-Bioproject, PRJNA495747 and PRJNA495763 accessions, respectively. A total of 127086 and 134984 microsatellite or simple sequence repeat (SSR) markers were identified within A. deserticola and A. atacamensis genomic DNA, respectively. In addition, predicted putative genes in A. deserticola and A. atacamensis sequences are also presented in this article.

Maqui (Aristotelia chilensis [Molina] Stunz) is a small dioecious tree native to South America with edible fruit characterized by very high antioxidant capacity and anthocyanin content. To preserve maqui as a genetic resource it is essential to study its genetic diversity. However, the complete genome is unknown and only a few gene sequences are available in databases. Simple sequence repeats (SSR) markers, which are neutral, co-dominant, reproducible and highly variable, are desirable to support genetic studies in maqui populations. By means of identification and characterization of microsatellite loci from a maqui genotype, using 454 sequencing technology, we develop a set of SSR for this species. Obtaining a total of 165,043 shotgun genome sequences, with an average read length of 387 bases, we covered 64 Mb of the maqui genome. Reads were assembled into 4,832 contigs, while 98,546 reads remained as singletons, generating a total of 103,378 consensus genomic sequences. A total of 24,494 SSR maqui markers were identified. Of them, 15,950 SSR maqui markers were classified as perfects. The most common SSR motifs were dinucleotide (31%), followed by tetranucleotide (26%) and trinucleotide motifs (24%). The motif AG/CT (28.4%) was the most abundant, while the motif AC (89 bp) was the largest. Eleven polymorphic SSRs were selected and used to analyze a population of 40 maqui genotypes. Polymorphism information content (PIC) ranged from 0.117 to 0.82, with an average of 0.58. Non-significant groups were observed in the maqui population, showing a panmictic genetic structure. In addition, we also predicted 11150 putative genes and 3 microRNAs (miRNAs) in maqui sequences. This results, including partial sequences of genes, some miRNAs and SSR markers from high throughput next generation sequencing (NGS) of maqui genomic DNA, constitute the first platform to undertake genetic and molecular studies of this important species.

Linkage maps are valuable tools in genetic and genomic studies. For sweet cherry, linkage maps have been constructed using mainly microsatellite markers (SSRs) and, recently, using single nucleotide polymorphism markers (SNPs) from a cherry 6K SNP array. Genotyping-by-sequencing (GBS), a new methodology based on high-throughput sequencing, holds great promise for identification of high number of SNPs and construction of high density linkage maps. In this study, GBS was used to identify SNPs from an intra-specific sweet cherry cross. A total of 8,476 high quality SNPs were selected for mapping. The physical position for each SNP was determined using the peach genome, Peach v1.0, as reference, and a homogeneous distribution of markers along the eight peach scaffolds was obtained. On average, 65.6% of the SNPs were present in genic regions and 49.8% were located in exonic regions. In addition to the SNPs, a group of SSRs was also used for construction of linkage maps. Parental and consensus high density maps were constructed by genotyping 166 siblings from a 'Rainier' x 'Rivedel' (Ra x Ri) cross. Using Ra x Ri population, 462, 489 and 985 markers were mapped into eight linkage groups in 'Rainier', 'Rivedel' and the Ra x Ri map, respectively, with 80% of mapped SNPs located in genic regions. Obtained maps spanned 549.5, 582.6 and 731.3 cM for 'Rainier', 'Rivedel' and consensus maps, respectively, with an average distance of 1.2 cM between adjacent markers for both 'Rainier' and 'Rivedel' maps and of 0.7 cM for Ra x Ri map. High synteny and co-linearity was observed between obtained maps and with Peach v1.0. These new high density linkage maps provide valuable information on the sweet cherry genome, and serve as the basis for identification of QTLs and genes relevant for the breeding of the species.