A chromosome-level, haplotype-phased Vanilla planifolia genome highlights the challenge of partial endoreplication for accurate whole-genome assembly.

Vanilla planifolia, the species cultivated to produce one of the world's most popular flavors, is highly prone to partial genome endoreplication, which leads to highly unbalanced DNA content in cells. We report here the first molecular evidence of partial endoreplication at the chromosome scale by the assembly and annotation of an accurate haplotype-phased genome of V. planifolia. Cytogenetic data demonstrated that the diploid genome size is 4.09 Gb, with 16 chromosome pairs, although aneuploid cells are frequently observed. Using PacBio HiFi and optical mapping, we assembled and phased a diploid genome of 3.4 Gb with a scaffold N50 of 1.2 Mb and 59 128 predicted protein-coding genes. The atypical k-mer frequencies and the uneven sequencing depth observed agreed with our expectation of unbalanced genome representation. Sixty-seven percent of the genes were scattered over only 30% of the genome, putatively linking gene-rich regions and the endoreplication phenomenon. By contrast, low-coverage regions (non-endoreplicated) were rich in repeated elements but also contained 33% of the annotated genes. Furthermore, this assembly showed distinct haplotype-specific sequencing depth variation patterns, suggesting complex molecular regulation of endoreplication along the chromosomes. This high-quality, anchored assembly represents 83% of the estimated V. planifolia genome. It provides a significant step toward the elucidation of this complex genome. To support post-genomics efforts, we developed the Vanilla Genome Hub, a user-friendly integrated web portal that enables centralized access to high-throughput genomic and other omics data and interoperable use of bioinformatics tools.

Phenolic Profiling for Traceability of Vanilla ×tahitensis.

Vanilla is a flavoring recovered from the cured beans of the orchid genus Vanilla. Vanilla ×tahitensis is traditionally cultivated on the islands of French Polynesia, where vanilla vines were first introduced during the nineteenth century and, since the 1960s, have been introduced to other Pacific countries such as Papua New Guinea (PNG), cultivated and sold as "Tahitian vanilla," although both sensory properties and aspect are different. From an economic point of view, it is important to ensure V. ×tahitensis traceability and to guarantee that the marketed product is part of the future protected designation of the origin "Tahitian vanilla" (PDO), currently in progress in French Polynesia. The application of metabolomics, allowing the detection and simultaneous analysis of hundreds or thousands of metabolites from different matrices, has recently gained high interest in food traceability. Here, metabolomics analysis of phenolic compounds profiles was successfully applied for the first time to V. ×tahitensis to deepen our knowledge of vanilla metabolome, focusing on phenolics compounds, for traceability purposes. Phenolics were screened through a quadrupole-time-of-flight mass spectrometer coupled to a UHPLC liquid chromatography system, and 260 different compounds were clearly evidenced and subjected to different statistical analysis in order to enable the discrimination of the samples based on their origin. Eighty-eight and twenty three compounds, with a prevalence of flavonoids, resulted to be highly discriminant through ANOVA and Orthogonal Projections to Latent Structures Discriminant Analysis (OPLS-DA) respectively. Volcano plot analysis and pairwise comparisons were carried out to determine those compounds, mainly responsible for the differences among samples as a consequence of either origin or cultivar. The samples from PNG were clearly different from the Tahitian samples that were further divided in two different groups based on the different phenolic patterns. Among the 260 compounds, metabolomics analysis enabled the detection of previously unreported phenolics in vanilla (such as flavonoids, lignans, stilbenes and other polyphenols).

DNA Remodeling by Strict Partial Endoreplication in Orchids, an Original Process in the Plant Kingdom.

DNA remodeling during endoreplication appears to be a strong developmental characteristic in orchids. In this study, we analyzed DNA content and nuclei in 41 species of orchids to further map the genome evolution in this plant family. We demonstrate that the DNA remodeling observed in 36 out of 41 orchids studied corresponds to strict partial endoreplication. Such process is developmentally regulated in each wild species studied. Cytometry data analyses allowed us to propose a model where nuclear states 2C, 4E, 8E, etc. form a series comprising a fixed proportion, the euploid genome 2C, plus 2-32 additional copies of a complementary part of the genome. The fixed proportion ranged from 89% of the genome in Vanilla mexicana down to 19% in V. pompona, the lowest value for all 148 orchids reported. Insterspecific hybridization did not suppress this phenomenon. Interestingly, this process was not observed in mass-produced epiphytes. Nucleolar volumes grow with the number of endocopies present, coherent with high transcription activity in endoreplicated nuclei. Our analyses suggest species-specific chromatin rearrangement. Towards understanding endoreplication, V. planifolia constitutes a tractable system for isolating the genomic sequences that confer an advantage via endoreplication from those that apparently suffice at diploid level.

Diversity and dynamics of plant genome size: an example of polysomaty from a cytogenetic study of Tahitian vanilla (Vanilla xtahitensis, Orchidaceae).

PREMISE OF THE STUDY: Abnormal mitotic behavior with somatic aneuploidy and partial endoreplication were previously reported for the first time in the plant kingdom in Vanilla planifolia. Because vanilla plants are vegetatively propagated, such abnormalities have been transmitted. This study aimed to determine whether mitotic abnormalities also occur in Vanilla hybrid or are suppressed by sexual reproduction. METHODS: Twenty-eight accessions of Vanilla ×tahitensis, one V. planifolia, and hybrid V. planifolia × V. ×tahitensis were analyzed by chromosome counts, cytometry, and fluorescent in situ hybridization of 18S-5.8S-26S rDNA. KEY RESULTS: In a single root meristem of V. ×tahitensis, chromosome number varied from 22 to 31 in diploids (mean 2C = 5.23 pg), 31 to 41 in triploids (2C = 7.82 pg) and 43 to 60 in tetraploids (2C = 10.27 pg). Morphological diversity is apparently related to ploidy changes. Aneuploidy and partial (asymmetrical) endoreduplication were observed in root meristems of both V. ×tahitensis and the hybrid V. planifolia × V. ×tahitensis, but pollen grains had the euploid chromosome number (n = 15 in diploids). CONCLUSIONS: Genome irregularities may be transmitted not only during vegetative propagation but also by sexual reproduction in Vanilla. However, there must be a complex regulation of genome size and organization between the aneuploidy in somatic tissues and subsequently euploid gametic tissue. This is a novel example of polysomaty with developmentally regulated partial endoreplication.

Neotropical roots of a Polynesian spice: the hybrid origin of Tahitian vanilla, Vanilla tahitensis (Orchidaceae).

Absent in the wild, Tahitian vanilla (V. tahitensis) is a gourmet spice restricted in distribution to cultivated and feral stands in French Polynesia and Papua New Guinea. Its origins have been elusive. Our objective was to test the purported hybrid derivation and parentage of V. tahitensis from aromatic, neotropical progenitors. Nucleotide sequences from V. tahitensis and neotropical Vanilla were assayed for phylogenetic relatedness in two independently inherited genomic regions, the nuclear ITS region, and the trnH-psbA noncoding region of chloroplast DNA. As predicted to occur for early generation hybrids, placement of V. tahitensis was nonconcordant. All V. tahitensis clustered with V. planifolia from analysis of cpDNA sequences, suggesting V. planifolia as the maternal genome contributor. Phylogenetic reconstruction of ITS sequences showed that most V. tahitensis nested incongruently with V. odorata, but others remained sister to V. planifolia. Recovery of ITS clones in V. tahitensis related to both V. planifolia and V. odorata also supports its biphyletic origin from these two taxa. We interpret the high percentage (95%) of additive polymorphic sites in V. tahitensis relative to its parents as indication of a recent, and probably human-mediated, evolutionary origin.