As a highly economic berry fruit crop, blueberry is enjoyed by most people and has various potential health benefits, many of which are attributed to the relatively high concentrations of flavonoids. To obtain more accurate and comprehensive transcripts, the full-length transcriptome of half-highbush blueberry (Vaccinium corymbosum/angustifolium cultivar Northland) obtained using single molecule real-time and next-generation sequencing technologies was reported for the first time. Overall, 147,569 consensus transcripts (average length, 2738 bp; N50, 3176 bp) were obtained. After quality control steps, 63,425 high-quality isoforms were obtained and 5030 novel genes, 3002 long non-coding RNAs, 3946 transcription factor genes (TFs), 30,540 alternative splicing events, and 2285 fusion gene pairs were identified. To better explore the molecular mechanism of flavonoid biosynthesis in mature blueberry fruit, an integrative analysis of the metabolome and transcriptome was performed on the exocarp, sarcocarp, and seed. A relatively complete biosynthesis pathway map of phenylpropanoids, flavonoids, and proanthocyanins in blueberry was constructed. The results of the joint analysis showed that the 228 functional genes and 42 TFs regulated 78 differentially expressed metabolites within the biosynthesis pathway of phenylpropanoids/flavonoids. O2PLS analysis results showed that the key metabolites differentially accumulated in blueberry fruit tissues were albireodelphin, delphinidin 3,5-diglucoside, delphinidin 3-O-rutinoside, and delphinidin 3-O-sophoroside, and 10 structural genes (4 Vc4CLs, 3 VcBZ1s, 1 VcUGT75C1, 1 VcAT, and 1 VcUGAT), 4 transporter genes (1 VcGSTF and 3 VcMATEs), and 10 TFs (1 VcMYB, 2 VcbHLHs, 4 VcWD40s, and 3 VcNACs) exhibited strong correlations with 4 delphinidin glycosides. These findings provide insights into the molecular mechanisms of flavonoid biosynthesis and accumulation in blueberry fruit.

Blueberry belongs to the Vaccinium genus and is a highly popular fruit crop with significant economic importance. It was not until the early twentieth century that they began to be domesticated through extensive interspecific hybridization. Here, we collected 220 Vaccinium accessions from various geographical locations, including 154 from the United States, 14 from China, eight from Australia, and 29 from Europe and other countries, comprising 164 Vaccinium corymbosum, 15 Vaccinium ashei, 10 lowbush blueberries, seven half-high blueberries, and others. We present the whole-genome variation map of 220 accessions and reconstructed the hundred-year molecular history of interspecific hybridization of blueberry. We focused on the two major blueberry subgroups, the northern highbush blueberry (NHB) and southern highbush blueberry (SHB) and identified candidate genes that contribute to their distinct traits in climate adaptability and fruit quality. Our analysis unveiled the role of gene introgression from Vaccinium darrowii and V. ashei into SHB in driving the differentiation between SHB and NHB, potentially facilitating SHB’s adaptation to subtropical environments. Assisted by genome-wide association studies, our analysis suggested VcTBL44 as a pivotal gene regulator governing fruit firmness in SHB. Additionally, we conducted whole-genome bisulfite sequencing on nine NHB and 12 SHB cultivars, and characterized regions that are differentially methylated between the two subgroups. In particular, we discovered that the β-alanine metabolic pathway genes were enriched for DNA methylation changes. Our study provides high-quality genetic and epigenetic variation maps for blueberry, which offer valuable insights and resources for future blueberry breeding.

Sugars will eventually be exported transporters (SWEETs) are essential transmembrane proteins involved in plant growth, stress responses, and plant–pathogen interactions. Despite their importance, systematic studies on SWEETs in blueberries (Vaccinium corymbosum L.) are limited. Blueberries are recognized for their rapid growth and the significant impact of sugar content on fruit flavor, yet the role of the SWEET gene family in sugar accumulation during fruit development remains unclear. In this study, 23 SWEET genes were identified in blueberry, and their phylogenetic relationships, duplication events, gene structures, cis-regulatory elements, and expression profiles were systematically analyzed. The VcSWEET gene family was classified into four clades. Structural and motif analysis revealed conserved exon–intron organization within each clade. RT-qPCR analysis showed widespread expression of VcSWEETs across various tissues and developmental stages, correlating with promoter cis-elements. VcSWEET6a, in particular, was specifically expressed in fruit and showed reduced expression during fruit maturation. Subcellular localization indicated that VcSWEET6a is located in the endoplasmic reticulum. Functional assays in yeast confirmed its role in glucose and fructose uptake, with transport activity inhibited at higher sugar concentrations. Overexpression of VcSWEET6a in blueberries resulted in reduced sugar accumulation. These findings offer valuable insights into the role of VcSWEETs in blueberry sugar metabolism.

Introduction: As a high-value small fruit crop worldwide, blueberry is prized for its health-promoting properties. Glutathione S-transferases (GSTs) are a large and complex family of enzymes that play vital roles in flavonoid metabolism, plant growth and development, and responses to abiotic stress. However, relatively little is known about the GST gene family in blueberry. Methods: In this study, genome-wide identification and bioinformatics analysis of the blueberry GST gene family were conducted. The candidate gene VcGSTU53 was further analyzed for its response to abiotic stress using transcriptome profiling, subcellular localization, and heterologous expression assays. Results: A total of 190 VcGST genes were identified and classified into 14 subclasses: tau (U, 75), phi (F, 28), EF1G (17), lambda (L, 10), MAPEG (10), zeta (Z, 8), GHR (8), Metaxin (M, 7), GST2N (6), DHAR (5), hemerythrin (H, 4), mPGES2 (4), TCHQD (4) and theta (T, 4); however, no iota (I) class GST was found. The lengths of the VcGST genes ranged from 366 bp to 4386 bp, encoding polypeptides of 121 to 1461 amino acids, with predicted molecular weights ranging from 13.86 kDa to 163.02 kDa, and theoretical isoelectric points ranging from 4.77 to 9.56. Approximately 46.32% of VcGST proteins were predicted to localize in the cytoplasm, with the remainder predicted to localize in the endomembrane system, chloroplasts, nucleus, mitochondria, extracellular space, and plasma membrane. Based on a comparative transcriptome profiling of 30 AlCl ₃ -treated root samples from two cultivars, VcGSTU53 was screened as a key aluminum-stress-responsive gene. VcGSTU53 encodes a protein of 230 amino acids, is localized in the cytoplasm, and exhibits transferase activity. Heterologous expression of VcGSTU53 in Escherichia coli did not inhibit bacterial growth, but enhanced tolerance to aluminum stress. Notably, it also increased bacterial resistance to drought and cadmium stresses. Conclusion: These findings contribute to a deeper understanding of the functions of the GST family in plants under abiotic stress, which is of great significance for the development of new stress-resistant blueberry varieties.

Sulfur metabolism plays a major role in plant growth and development, environmental adaptation, and material synthesis, and the sulfate transporters are the beginning of sulfur metabolism. We identified 37 potential VcSULTR genes in the blueberry genome, encoding peptides with 534 to 766 amino acids. The genes were grouped into four subfamilies in an evolutionary analysis. The 37 putative VcSULTR proteins ranged in size from 60.03 to 83.87 kDa. These proteins were predicted to be hydrophobic and mostly localize to the plasma membrane. The VcSULTR genes were distributed on 30 chromosomes; VcSULTR3;5b and VcSULTR3;5c were the only tandemly repeated genes. The VcSULTR promoters contained cis-acting elements related to the fungal symbiosis and stress responses. The transcript levels of the VcSULTRs differed among blueberry organs and changed in response to ericoid mycorrhizal fungi and sulfate treatments. A subcellular localization analysis showed that VcSULTR2;1c localized to, and functioned in, the plasma membrane and chloroplast. The virus-induced gene knock-down of VcSULTR2;1c resulted in a significantly decreased endogenous sulfate content, and an up-regulation of genes encoding key enzymes in sulfur metabolism (VcATPS2 and VcSiR1). These findings enhance our understanding of mycorrhizal-fungi-mediated sulfate transport in blueberry, and lay the foundation for further research on blueberry–mycorrhizal symbiosis.

Introduction: Fruit softening is a major factor limiting the postharvest quality, transportability, and shelf life of blueberry, but the physiological and molecular basis of firmness variation remains incompletely understood. Methods: The soft-fruited cultivar ‘Duke’ and the firm-fruited genotype ‘2017D’ were compared using phenotypic, physiological, histological, biochemical, transcriptomic, and transient functional analyses. Results: ‘2017D’ maintained substantially higher firmness during fruit development, together with greater retention of several texture attributes, including chewiness, gumminess, resilience, and cohesiveness. Histological observations showed that ‘2017D’ retained a comparatively more continuous mesocarp organization at the blue stage. Cell wall analysis further revealed greater retention of structural polysaccharides and insoluble pectin in ‘2017D’, whereas ‘Duke’ accumulated more soluble pectin during the later developmental stages. Lower PG and β-galactosidase activities in ‘2017D’ were consistent with reduced cell wall polysaccharide degradation during ripening. RNA-seq analysis identified 6,168 differentially expressed genes between the two genotypes, with significant enrichment of pathways and biological processes associated with hormone signaling, carbohydrate metabolism, cell wall organization, and related ripening processes. Among the cell wall-related candidate genes, VcβGAL369 showed a pronounced expression difference between the two genotypes and a distinct developmental expression pattern and was therefore selected for further characterization. Transient silencing of VcβGAL369 increased fruit firmness and promoted cell wall component retention, whereas transient overexpression reduced fruit firmness and promoted cell wall disassembly. Discussion: These results indicate that the contrasting firmness of ‘2017D’ and ‘Duke’ is associated with differences in cell wall remodeling and ripening-related transcriptional processes and support a role for VcβGAL369 in promoting blueberry fruit softening.