Past endosymbiotic events allowed photosynthetic organisms to flourish and evolve in terrestrial areas. The precursor of chloroplasts was an ancient photosynthetic cyanobacterium. Presently, cyanobacteria are still capable of establishing successful symbioses in a wide range of hosts. One particular host plant among the gymnosperms is cycads (Order Cycadales) in which a special type of root system, referred to as coralloid roots, develops to house symbiotic cyanobacteria. A number of studies have explained coralloid root formation and cyanobiont invasion but the questions on mechanisms of this host-microbe association remains vague. Most researches focus on diversity of symbionts in coralloid roots but equally important is to explore the underlying mechanisms of cycads-Nostoc symbiosis as well. Besides providing an overview of relevant areas presently known about this association and citing putative genes involved in cycad-cyanobacteria symbioses, this paper aims to identify the limitations that hamper attempts to get to the root of the matter and suggests future research directions that may prove useful.

The culture-based approach was used to characterize the fungal endophytes associated with the coralloid roots of the endemic Cycas debaoensis and Cycas fairylakea from various population sites in China. We aim to determine if the assemblages of fungal endophytes inside these endemic plant hosts are distinct and could be explored for bioprospecting. The isolation method yielded a total of 284 culturable fungal strains. Identification based on the analysis of the internal transcribed spacer (ITS) rDNA showed that they belonged to two phyla, five classes, eight orders and 22 families. At least 33 known genera and 62 different species were confirmed based on >97% ITS sequence similarity. The most frequent and observed core taxa in the two host species regardless of their population origin were Talaromyces, Penicillium, Fusarium, Pochonia and Gliocladiopsis. Seventy percent was a rare component of the fungal communities with only one or two recorded isolates. Contrary to common notions, diversity and fungal richness were significantly higher in C. debaoensis and C. fairylakea collected from a botanical garden, while the lowest was observed in C. debaoensis from a natural habitat; this provides evidence that garden management, and to a minor extent, ex-situ conservation practice, could influence fungal endophyte communities. We further selected nineteen fungal isolates and screened for their antagonistic activities via a co-cultivation approach against the phytopathogens, Diaporthe sp. and Colletotrichum sp. Among these, five isolates with high ITS similarity matches with Hypoxylon vinosupulvinatum (GD019, 99.61%), Penicillium sp. (BD022, 100%), Penicillifer diparietisporus (GD008, 99.46%), Clonostachys rogersoniana (BF024, 99.46%) and C. rosea (BF011, 99.1%), which showed exceptional antagonistic activities against the phytopathogenic fungi with a significant inhibition rate of 70–80%. Taken together, our data presented the first and most comprehensive molecular work on culturable fungal endophytes associated with the coralloid roots of cycads. Our study also demonstrated that about 5% of fungal endophytes were not detected by the high-throughput sequencing approach, implying the equal importance of a culture-dependent approach to study fungal communities of cycads. We further highlighted the potential role of endemic and rare plants to discover and isolate unique plant-associated fungal taxa with excellent biocontrol properties.

Bougainvillea (Nyctaginaceae) is a popular ornamental plant group primarily grown for its striking colorful bracts. However, despite its established horticultural value, limited genomic resources and molecular studies have been reported for this genus. Thus, to address this existing gap, complete chloroplast genomes of four species (Bougainvillea glabra, Bougainvillea peruviana, Bougainvillea pachyphylla, Bougainvillea praecox) and one Bougainvillea cultivar were sequenced and characterized. The Bougainvillea cp genomes range from 153,966 bp to 154,541 bp in length, comprising a large single-copy region (85,159 bp–85,708 bp) and a small single-copy region (18,014 bp–18,078 bp) separated by a pair of inverted repeats (25,377–25,427 bp). All sequenced plastomes have 131 annotated genes, including 86 protein-coding, eight rRNA, and 37 tRNA genes. These five newly sequenced Bougainvillea cp genomes were compared to the Bougainvillea spectabilis cp genome deposited in GeBank. The results showed that all cp genomes have highly similar structures, contents, and organization. They all exhibit quadripartite structures and all have the same numbers of genes and introns. Codon usage, RNA editing sites, and repeat analyses also revealed highly similar results for the six cp genomes. The amino acid leucine has the highest proportion and almost all favored synonymous codons have either an A or U ending. Likewise, out of the 42 predicted RNA sites, most conversions were from serine (S) to leucine (L). The majority of the simple sequence repeats detected were A/T mononucleotides, making the cp genomes A/T-rich. The contractions and expansions of the IR boundaries were very minimal as well, hence contributing very little to the differences in genome size. In addition, sequence variation analyses showed that Bougainvillea cp genomes share nearly identical genomic profiles though several potential barcodes, such as ycf1, ndhF, and rpoA were identified. Higher variation was observed in both B. peruviana and B. pachyphylla cp sequences based on SNPs and indels analysis. Phylogenetic reconstructions further showed that these two species appear to be the basal taxa of Bougainvillea. The rarely cultivated and wild species of Bougainvillea (B. pachyphylla, B. peruviana, B. praecox) diverged earlier than the commonly cultivated species and cultivar (B. spectabilis, B. glabra, B. cv.). Overall, the results of this study provide additional genetic resources that can aid in further phylogenetic and evolutionary studies in Bougainvillea. Moreover, genetic information from this study is potentially useful in identifying Bougainvillea species and cultivars, which is essential for both taxonomic and plant breeding studies.

Bougainvillea Comm. ex Juss. is one of the renowned genera in the Nyctaginaceae, but despite its recognized horticultural value, the taxonomy and phylogeny of the genus is not well-studied. Phylogenetic reconstructions based on plastid genomes showed that B. pachyphylla and B. peruviana are basal taxa, while B. spinosa is sister to two distinct clades: the predominantly cultivated Bougainvillea clade (B. spectabilis, B. glabra, B. arborea, B. cultivar, B. praecox) and the clade containing wild species of Bougainvillea (B. berberidifolia, B. campanulata, B. infesta, B. modesta, B. luteoalba, B. stipitata, and B. stipitata var. grisebachiana). Early divergence of B. peruviana, B. pachyphylla and B. spinosa is highly supported, thus the previously proposed division of Bougainvillea into two subgenera (Bougainvillea and Tricycla) was not reflected in this study. Morphological analysis also revealed that leaf arrangement, size, and indumentum together with the perianth tube and anthocarp shape and indumentum are important characteristics in differentiating the species of Bougainvillea. In the present study, 11 species and one variety are recognized in Bougainvillea. Six names are newly reduced to synonymy, and lectotypes are designated for 27 names. In addition, a revised identification key and illustrations of the distinguishing parts are also provided in the paper.

Endogenously formed coralloid roots in living cycads are known to be infected and maintained by a mutual partnership with photosynthetic cyanobacteria. In this study, two strains of filamentous, nitrogen-fixing endosymbiotic cyanobacteria from the green zone of the coralloid roots of Cycas fairylakea were isolated and grown in culture. The phylogenetic placement based on the 16S rRNA gene region and the secondary structures of the ITS between the 16S and 23S rRNA gene, in addition to traditional morphological characterization, were used to analyse the two strains. Results showed that the 16S rRNA gene sequences of the two strains, CF01 and CF06, displayed ≥97% similarity to the genus Desmonostoc. The phylogeny inferred by maximum likelihood, bayesian inference and neighbour-joining placed the two strains on a separate node away from other species of Desmonostoc. Hormogonial filaments were evident in the two symbiotic species. In addition, a few unique morphological structures in the CF01 and CF06 strains were observed. The D1-D1ʹ, Box-B and V3 helices generated from the ITS of 16S–23S sequences did not fit those of any described species of Desmonostoc, and this led to proposing the two strains as novel species. In compliance with the International Code of Nomenclature for algae, fungi, and plants, the two species are named Desmonostoc lechangense sp. nov. and Desmonostoc aggregatum sp. nov.

Cycads have developed a complex root system categorized either as normal or coralloid roots. Past literatures revealed that a great diversity of key microbes is associated with these roots. This recent study aims to comprehensively determine the diversity and community structure of bacteria and fungi associated with the roots of two Cycas spp. endemic to China, Cycas debaoensis Zhong & Chen and Cycas fairylakea D.Y. Wang using high-throughput amplicon sequencing of the full-length 16S rRNA (V1-V9 hypervariable) and short fragment ITS region. The total DNA from 12 root samples were extracted, amplified, sequenced, and analyzed. Resulting sequences were clustered into 61 bacteria and 2128 fungal OTUs. Analysis of community structure revealed that the coralloid roots were dominated mostly by the nitrogen-fixer Nostocaceae but also contain other non-diazotrophic bacteria. The sequencing of entire 16S rRNA gene identified four different strains of cyanobacteria under the heterocystous genera Nostoc and Desmonostoc. Meanwhile, the top bacterial families in normal roots were Xanthobacteraceae, Burkholderiaceae, and Bacillaceae. Moreover, a diverse fungal community was also found in the roots of cycads and the predominating families were Ophiocordycipitaceae, Nectriaceae, Bionectriaceae, and Trichocomaceae. Our results demonstrated that bacterial diversity in normal roots of C. fairylakea is higher in richness and abundance than C. debaoensis. On the other hand, a slight difference, albeit insignificant, was noted for the diversity of fungi among root types and host species as the number of shared taxa is relatively high (67%). Our results suggested that diverse microbes are present in roots of cycads which potentially interact together to support cycads survival. Our study provided additional knowledge on the microbial diversity and composition in cycads and thus expanding our current knowledge on cycad-microbe association. Our study also considered the possible impact of ex situ conservation on cyanobiont community of cycads.