Plastid genomes of photosynthetic flowering plants are usually highly conserved in both structure and gene content. However, the plastomes of parasitic and mycoheterotrophic plants may be released from selective constraint due to the reduction or loss of photosynthetic ability. Here we present the greatly reduced and highly divergent, yet functional, plastome of the nonphotosynthetic holoparasite Hydnora visseri (Hydnoraceae, Piperales). The plastome is 27 kb in length, with 24 genes encoding ribosomal proteins, ribosomal RNAs, tRNAs, and a few nonbioenergetic genes, but no genes related to photosynthesis. The inverted repeat and the small single copy region are only approximately 1.5 kb, and intergenic regions have been drastically reduced. Despite extreme reduction, gene order and orientation are highly similar to the plastome of Piper cenocladum, a related photosynthetic plant in Piperales. Gene sequences in Hydnora are highly divergent and several complementary approaches using the highest possible sensitivity were required for identification and annotation of this plastome. Active transcription is detected for all of the protein-coding genes in the plastid genome, and one of two introns is appropriately spliced out of rps12 transcripts. The whole-genome shotgun read depth is 1,400× coverage for the plastome, whereas the mitochondrial genome is covered at 40× and the nuclear genome at 2×. Despite the extreme reduction of the genome and high sequence divergence, the presence of syntenic, long transcriptionally active open-reading frames with distant similarity to other plastid genomes and a high plastome stoichiometry relative to the mitochondrial and nuclear genomes suggests that the plastome remains functional in H. visseri. A four-stage model of gene reduction, including the potential for complete plastome loss, is proposed to account for the range of plastid genomes in nonphotosynthetic plants.

Extreme haustorial parasites have long captured the interest of naturalists and scientists with their greatly reduced and highly specialized morphology. Along with the reduction or loss of photosynthesis, the plastid genome often decays as photosynthetic genes are released from selective constraint. This makes it challenging to use traditional plastid genes for parasitic plant phylogenetics, and has driven the search for alternative phylogenetic and molecular evolutionary markers. Thus, evolutionary studies, such as molecular clock-based age estimates, are not yet available for all parasitic lineages. In the present study, we extracted 14 nuclear single copy genes (nSCG) from Illumina transcriptome data from one of the “strangest plants in the world”, Hydnora visseri (Hydnoraceae). A ∼15,000 character molecular dataset, based on all three genomic compartments, shows the utility of nSCG for reconstructing phylogenetic relationships in parasitic lineages. A relaxed molecular clock approach with the same multi-locus dataset, revealed an ancient age of ∼91 MYA for Hydnoraceae. We then estimated the stem ages of all independently originated parasitic angiosperm lineages using a published dataset, which also revealed a Cretaceous origin for Balanophoraceae, Cynomoriaceae and Apodanthaceae. With the exception of Santalales, older parasite lineages tend to be more specialized with respect to trophic level and have lower species diversity. We thus propose the “temporal specialization hypothesis” (TSH) implementing multiple independent specialization processes over time during parasitic angiosperm evolution.

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Hydtiora africana is a root holoparasite of southern Africa that emerges only to flovi'er. The trimerous flowers of H. africana have androecial and gynoecial chambers and attract fioral visitors with putrid odors emitted from prominent osmophores. We observed floral phenology and insect visitation for H. africana at two sites in southern Namibia and evaluated the insect imprisonment mechanism with beetle addition and pollen viability assays. Flowers are putatively protogynous for 3 d. We observed 18 Horal visitors, including 10 coleopteran species imprisoned by the smooth inner surface of the androecial chamber. The hide beetle Dermestes nuutilatiis (Tenebrionidae) accounted for 76.9% of tbe imprisoned insects, with a density of 2.2 ± 0.6 per flower. The D. maculatiis addition experiment {n = 9) clearly demonstrated imprisonment during the carpellate stage. Changes in the inner surfaces of the androecial chamber, stippling, and texturing allowed D. maculatus to escape after pollen release. More than 55.5% of tbe beetles escaped, dusted with pollen, within 3 d after pollen release. Pollen was still viable 3 d after pollen release. The beetle addition and pollen assays demonstrate the efficiency of the H. africana imprisonment mechanism.