Rubiaceae Plant Research Articles

A near-complete genome assembly of Cinchona calisaya

Rubiaceae plants are globally widespread and possess significant economic and medicinal value. Such as the globally important crop of coffee, and Cinchona calisaya Wedd., which is rich in alkaloids, is an important medicinal resource for treating malaria. In recent years, several genome resources of Rubiaceae have been reported. However, the comprehensive phylogenetic relationships of Rubiaceae are still unknown. In this study, we present a nearly complete diploid genome assembly of C. calisaya, characterized by a genome size of 869.93 Mb, and contig N50 length of 44.34 Mb. Notably, 99.75% of the sequences have been successfully anchored to 17 chromosomes, with only 12 gaps remaining. BUSCO assessment indicates that 97.40% of complete core genes are present in the assembly. We identified a total of 42,741 protein-coding genes, among which 38,022 (89.00%) have received functional annotation. The high continuity and integrity of the C. calisaya genome provide a robust foundation for functional genomics research, varietal improvement, and the conservation of genomic resources in medicinal plants.

Toxicological Insights into Myrmecodia platytyrea Tuber Aqueous Extract: An Oral Acute and Subacute Toxicity Studies in Mice

Despite advancements in medical science, the global burden of inflammation-related diseases remains high, necessitating the exploration of novel and effective remedies that can offer relief without significant side effects. The indigenous peoples of Papua and Borneo use the decoction of Myrmecodia platytyrea tubers, the ant plant, for its medical benefits, particularly in cancer therapy, demonstrating the deep traditional knowledge of natural treatments. The Rubiaceae plant has been used in traditional medicine to alleviate inflammation and other conditions. Thus, this study aims to investigate the toxicological profile of the aqueous extract of M. platytyrea tubers (MPAE). The study conducted adhered to recognised protocols for assessing oral acute and subacute toxicity, with necessary modifications to meet the specific experimental needs. In the acute toxicity study, male and female mice received a single oral dose of MPAE at 2000 mg/kg and monitored for 14 days. In the subacute toxicity study, male and female mice received 500 mg/kg/day MPAE orally for 28 days. Physical and behavioural changes were observed. Haematological parameters and biochemical parameters were analysed, including histology of the liver and kidneys. Acute and subacute toxicity studies found no clinical signs of toxicity, mortality, or weight changes. The subacute toxicity study found no significant changes in haematological parameters or biochemical parameters for both sexes. Acute and subacute toxicity tests revealed no treatment-related necropsy, gross, or histopathological abnormalities. In conclusion, MPAE is safe to consume and supports further development of M. platytyrea for treating inflammation-related diseases, particularly cancer.

Non-food renewable and bioactive products for pesticides: Synthesis, anti-oomycete and anti-fungal activities of 9R-acyloxyquinine derivatives bearing an 1,3,4-oxadiazole moiety

Quinine is the main alkaloids in the bark of Cinchona and its homologous plants of Rubiaceae. To discover biorational natural product-based pesticides, a series of 9R-acyloxyquinine derivatives containing 1,3,4-oxadiazole moieties were prepared by ingeniously introducing an active ester group and a 1,3,4-oxadiazole five membered heterocyclic ring at its C-9 position, and their structures were well characterized by 1H NMR, 13C NMR, HRMS, and m.p. The stereochemical configuration of target compound 6 t was unambiguously subjected to single-crystal X-ray diffraction. Furthermore, bioactivities of these compounds as anti-oomycete and anti-fungal agents against two serious agricultural diseases, Phytophthora capsici and Fusarium graminearum we assessed. Among all tested compounds, 1) Compounds 6c and 6d displayed promising anti-oomycete against P. capsici, with EC50 values of 38.42 and 35.49 mg/L, respectively. 2) Compound 6i exhibited promising anti-fungal against F. graminearum, with an EC50 value of 29.92 mg/L. Moreover, some interesting results of structure-activity relationships were also observed. The results of this study pave the way for further design, structural modification, and development of quinine derivatives as plant anti-oomycetes and anti-fungal agents in crop protection.

A comparative study on antimicrobial effects of Rubiaceae plants

Traditional Chinese Medicine (TCM) has accumulated valuable experience from ancient China while mixed with impurities. Plants from the Rubiaceae family are widely distributed in China, with a long history of use and a significant amount of related literature on Traditional Chinese Medicine. This study used ethanol extracts from three Rubiaceae plants, namely Galium spurium L., Rubia cordifolia L., and Hedyotis corymbosa L., to conduct antimicrobial experiments against Escherichia coli and Bacillus subtilis. Among them, G. spurium and R. cordifolia were confirmed to have antimicrobial capabilities, while H. corymbosa did not show antimicrobial ability. In the E. coli experiment, the diameter of the inhibition zone of G. spurium extract was approximately 1.48 cm, and that of R. cordifolia extract was about 1.03 cm. For B. subtilis, the inhibition zone diameter of G. spurium extract was approximately 1.49 cm, and that of R. cordifolia extract was about 1.48 cm. This experiment, conducted under modern research methods, confirmed the antimicrobial abilities of these three plants.

Volatile organic composition of five Rubiaceae species: Insights into their phytochemical diversity

Natural volatile organic compounds (VOCs) obtained from five Rubiaceae plants native to south China, namely Damnacanthus indicus, Hamelia patens, Exallage chrysotricha, Psychotria serpens, and Spermacoce pusilla, were isolated and characterized in this study. The analysis revealed that each plant contained 71, 57, 80, 73, and 70 volatile organic chemical compounds, respectively. The main VOCs shared by five plants was palmitate (10.20%–36.26%). Methyl salicylate, caryophyllene oxide, tetradecanoic acid, 6,10,14-trimethyl-2-pentadecanone, pentadecanoic acid, pentacosane, pentadecanal, nonanal, hexadecanoic acid methyl ester, phytol, linalool, and (E)-β-ionone were shared by at least 4 Rubiaceae plants in this study. Besides, each of the five plants has its main components as follows: 12.81% ethyl linoleate and 6.13% (E)-13-octadecenoic acid for D. indicus; 35.17% methyl salicylate for H. patens; 9.50% (Z, Z)-9,12-octadecadienoic acid and 7.56% endo-borneol for E. chrysotricha; 13.93% methyl salicylate for P. serpens; 6.19% humulene and 6.08% humulene oxide II for S. pusilla. This research provides insights into the phytochemical diversity and classification of Rubiaceae plants, laying a solid foundation for future natural product development in the Rubiaceae family.

Biocontrol fungi induced stem-base rot disease resistance of Morinda officinalis How revealed by transcriptome analysis.

Morinda officinalis How (MO) is a Rubiaceae plant, and its medicinal part is dried root, which is one of the "Four Southern Medicines" in China. At present, the plant MO breed seedlings mainly by cutting methods. Long-term asexual propagation makes pathogenic fungi accumulate in MO, leading to stem-base rot, which is caused by Fusarium oxysporum (Fon). In this study, we used Trichoderma harzianum and Pestalotiopsis sp. as biocontrol fungi to investigate their antagonistic ability to Fon through in vitro antagonism and pot experiments, and combined with transcriptome sequencing to explore the mechanism of biocontrol. The results showed that both Trichoderma harzianum and Pestalotiopsis sp. could inhibit the growth of Fon. In addition, Trichoderma harzianum and Pestalotiopsis sp. could also enhance the basic immunity to Fon by increasing the activities of defensive enzymes such as POD and SOD, chlorophyll content, soluble sugar content, and oligosaccharide content of MO. The mechanism of biological control of stem-base rot of MO was discussed by transcriptome technology. MO was treated with two treatments, root irrigation with biocontrol fungi or inoculation with Fon after root irrigation with biocontrol fungi. Transcriptome sequencing revealed that nearly 11,188 differentially expressed genes (DEGs) were involved in the process of inducing MO systemic resistance to Fon by biocontrol fungi. Meanwhile, Gene Ontology (GO) classification and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, as well as transcription factor (TFs) prediction showed that there were significant differences in the expression levels of MO roots under different treatments. Also, the genes of the "MAPK signaling pathway" and "plant hormone signaling pathway" were analyzed, in which the ERFs gene of the ethylene signal transduction pathway participated in the metabolism of glycosyl compounds. It is speculated that the ethylene signal may participate in the immune response of the sugar signal to the infection of Fon. After qRT-PCR verification of 10 DEGs related to the ethylene signal transduction pathway, the expression trend is consistent with the results of transcriptome sequencing, which proves the reliability of transcriptome sequencing. In conclusion, this study preliminarily identified the molecular mechanism of the biological control of MO stem-base rot and provided a scientific basis for further research on the prevention and control mechanism of MO stem-base rot.

Genome mining of metabolic gene clusters in the Rubiaceae family

The Rubiaceae plant family, comprising 3 subfamilies and over 13,000 species, is known for producing significant bioactive compounds such as caffeine and monoterpene indole alkaloids. Despite an increase in available genomes from the Rubiaceae family over the past decade, a systematic analysis of the metabolic gene clusters (MGCs) encoded by these genomes has been lacking. In this study, we aim to identify and analyze metabolic gene clusters within complete Rubiaceae genomes through a comparative analysis of eight species. Applying two bioinformatics pipelines, we identified 2372 candidate MGCs, organized into 549 gene cluster families (GCFs). To enhance the reliability of these findings, we developed coexpression networks and conducted orthology analyses. Using genomic data from Solanum lycopersicum (Solanaceae) for comparative purposes, we provided a detailed view of predicted metabolic enzymes, pathways, and coexpression networks. We bring some examples of MGCs and GCFs involved in biological pathways of terpenes, saccharides and alkaloids. Such insights lay the groundwork for discovering new compounds and associated MGCs within the Rubiaceae family, with potential implications in developing more robust crop species and expanding the understanding of plant metabolism. This large-scale exploration also provides a new perspective on the evolution and structure-function relationship of these clusters, offering opportunities for the highly efficient utilization of these unique metabolites. The outcome of this study contributes to a broader comprehension of the biosynthetic pathways, elucidating multiple aspects of specialized metabolism and offering innovative avenues for biotechnological applications.

Plant-based red colouration of shell beads 15,000 years ago in Kebara Cave, Mount Carmel (Israel).

Decorating the living space, objects, body and clothes with colour is a widespread human practice. While the habitual use of red mineral pigments (such as iron-oxide, e.g., ochre) by anatomically modern humans started in Africa about 140,000 years ago, the earliest documentation of the use of organic plant or animal-based red pigments is known from only 6,000 years ago. Here, we report the oldest reliable evidence of organic red pigment use 15,000 years ago by the first sedentary hunter-gatherers in the Levant. SEM-EDS and Raman Spectroscopy analyses of 10 red-stained shell beads enabled us to detect and describe the use of a colourant made of Rubiaceae plants roots (Rubia spp., Asperula spp., Gallium spp.) to colour personal adornments from the Early Natufian of Kebara cave, Mount Carmel, Israel. This adds a previously unknown behavioural aspect of Natufian societies, namely a well-established tradition of non-dietary plant processing at the beginning of the sedentary lifestyle. Through a combined multidisciplinary approach, our study broadens the perspectives on the ornamental practices and the chaînes opératoires of pigmenting materials during a crucial period in human history.

Differential Volatile Organic Compound Expression in the Interaction of Daldinia eschscholtzii and Mycena citricolor.

Fungi exhibit a wide range of ecological guilds, but those that live within the inner tissues of plants (also known as endophytes) are particularly relevant due to the benefits they sometimes provide to their hosts, such as herbivory deterrence, disease protection, and growth promotion. Recently, endophytes have gained interest as potential biocontrol agents against crop pathogens, for example, coffee plants (Coffea arabica). Published results from research performed in our laboratory showed that endophytic fungi isolated from wild Rubiaceae plants were effective in reducing the effects of the American leaf spot of coffee (Mycena citricolor). One of these isolates (GU11N) from the plant Randia grandifolia was identified as Daldinia eschscholtzii (Xylariales). Its antagonism mechanisms, effects, and chemistry against M. citricolor were investigated by analyzing its volatile profile alone and in the presence of the pathogen in contactless and dual culture assays. The experimental design involved direct sampling of agar plugs in vials for headspace (HS) and headspace solid-phase microextraction (HS-SPME) gas chromatography-mass spectrometry (GC-MS) analysis. Additionally, we used ultrahigh-performance liquid chromatography coupled to high-resolution mass spectrometry (UHPLC-HRMS/MS) to identify nonvolatile compounds from organic extracts of the mycelia involved in the interaction. Results showed that more volatile compounds were identified using HS-SPME (39 components) than those by the HS technique (13 components), sharing only 12 compounds. Statistical tests suggest that D. eschscholtzii inhibited the growth of M. citricolor through the release of VOCs containing a combination of 1,8-dimethoxynapththalene and terpene compounds affecting M. citricolor pseudopilei. The damaging effects of 1,8-dimethoxynaphthalene were corroborated in an in vitro test against M. citricolor pseudopilei; scanning electron microscopy (SEM) photographs confirmed structural damage. After analyzing the UHPLC-HRMS/MS data, a predominance of fatty acid derivatives was found among the putatively identified compounds. However, a considerable proportion of features (37.3%) remained unannotated. In conclusion, our study suggests that D. eschscholtzii has potential as a biocontrol agent against M. citricolor and that 1,8-dimethoxynaphthalene contributes to the observed damage to the pathogen's reproductive structures.

Metagenomics of African Empogona and Tricalysia (Rubiaceae) reveals the presence of leaf endophytes.

Leaf symbiosis is a phenomenon in which host plants of Rubiaceae interact with bacterial endophytes within their leaves. To date, it has been found in around 650 species belonging to eight genera in four tribes; however, the true extent in Rubiaceae remains unknown. Our aim is to investigate the possible occurrence of leaf endophytes in the African plant genera Empogona and Tricalysia and, if present, to establish their identity. Total DNA was extracted from the leaves of four species of the Coffeeae tribe (Empogona congesta, Tricalysia hensii, T. lasiodelphys, and T. semidecidua) and sequenced. Bacterial reads were filtered out and assembled. Phylogenetic analysis of the endophytes was used to reveal their identity and their relationship with known symbionts. All four species have non-nodulated leaf endophytes, which are identified as Caballeronia. The endophytes are distinct from each other but related to other nodulated and non-nodulated endophytes. An apparent phylogenetic or geographic pattern appears to be absent in endophytes or host plants. Caballeronia endophytes are present in the leaves of Empogona and Tricalysia, two genera not previously implicated in leaf symbiosis. This interaction is likely to be more widespread, and future discoveries are inevitable.

A metabolomic platform to identify and quantify polyphenols in coffee and related species using liquid chromatography mass spectrometry.

Products of plant secondary metabolism, such as phenolic compounds, flavonoids, alkaloids, and hormones, play an important role in plant growth, development, stress resistance. The plant family Rubiaceae is extremely diverse and abundant in Central America and contains several economically important genera, e.g. Coffea and other medicinal plants. These are known for the production of bioactive polyphenols (e.g. caffeine and quinine), which have had major impacts on human society. The overall goal of this study was to develop a high-throughput workflow to identify and quantify plant polyphenols. First, a method was optimized to extract over 40 families of phytochemicals. Then, a high-throughput metabolomic platform has been developed to identify and quantify 184 polyphenols in 15 min. The current metabolomics study of secondary metabolites was conducted on leaves from one commercial coffee variety and two wild species that also belong to the Rubiaceae family. Global profiling was performed using liquid chromatography high-resolution time-of-flight mass spectrometry. Features whose abundance was significantly different between coffee species were discriminated using statistical analysis and annotated using spectral databases. The identified features were validated by commercially available standards using our newly developed liquid chromatography tandem mass spectrometry method. Caffeine, trigonelline and theobromine were highly abundant in coffee leaves, as expected. Interestingly, wild Rubiaceae leaves had a higher diversity of phytochemicals in comparison to commercial coffee: defense-related molecules, such as phenylpropanoids (e.g., cinnamic acid), the terpenoid gibberellic acid, and the monolignol sinapaldehyde were found more abundantly in wild Rubiaceae leaves.

In vitro study of antioxidant and antibacterial properties of Phyllanthaceae and Rubiaceae plant families from Simeuleu Island, Aceh, Indonesia

Plants are still the preferred solution to solve several of the main problems the world is facing today, such as antibiotic resistance and free radicals. The plant chemical compounds have the potential as antibacterial and antioxidant agents. Therefore, four plants representing Phyllantaceae family ( Glochidion varians Miq., Glochidion zeylanicum (Gaertn.) A.Juss) and two plants from Rubiaceae family ( Uncaria lanosa var. glabrata, and Uncaria cordata (Lour.) Merr.)Â were collected from Simeulue island, Aceh. We obtained 32 extracts from several plants with various solvents, such as hexane, dichloromethane, ethyl acetate, and methanol. The methods applied to investigate antibacterial activities were Thin Layer Chromatography (TLC) and Minimum Inhibitory Concentration (MIC). Meanwhile, TLC and Microdilution Assay were utilized to analyze antioxidant capacity against 1,1-diphenyl-2-picrylhydrazyl (DPPH). The result of antibacterial activity shows 32 extracts displayed very strong to weak activity against Staphylococcus aureus and Escherichia coli . Among all extracts, methanol extracts of G. varians (stem and leaf) , methanol extracts of G. zeylanicum (stem and leaf) , methanol and ethyl acetate extracts of U. cordata (leaf), and ethyl acetate extract of G. varians (leaf) indicate potent antioxidant activity with the range of antioxidant activity index (AAI) values of 2.16 -11.73. Furthermore, all extracts exhibit moderate to weak antibacterial evaluation. The methanol extract of G. varians stem has the highest MIC value of 128 µg/ml against S. aureus. The methanol extracts of G. varians stem and leaf and methanol extract of G. zeylanicum leaf against E. coli have MIC value of 256 µg/ml, whereas other extracts have >256 µg/ml MIC value. To sum up, four plants from Phylantaceae and Rubiaceae families have the potential to be expanded as antioxidant agents rather than antibacterial agents.

Tolerance and Biological Removal of Fungicides by Trichoderma Species Isolated From the Endosphere of Wild Rubiaceae Plants

The transition from conventional to organic agriculture is often challenged by the adaptation of biological control agents to environments heavily exposed to agrochemical pollutants. We studied Trichoderma species isolated from living leaf tissues of wild Rubiacaeae (coffee family) plants to determine their fungicide tolerance and potential for bioremoval. First, we assessed the in vitro tolerance to fungicides of four Trichoderma isolates (Trichoderma rifaii T1, T. aff. crassum T2, T. aff. atroviride T3, and T. aff. strigosellum T4) by placing mycelial plugs onto solid media supplemented with seven different systemic and non-systemic fungicides. After a week, most of the fungicides did not significantly inhibit the growth of the isolates, except in the case of cyproconazole, where the only isolate able to grow was T1; however, the colony morphology was affected by the presence of fungicides. Second, biological removal potential was established for selected isolates. For this experiment, the isolates T1, T2, and T4 were independently inoculated into liquid media with the fungicides azoxystrobin, chlorothalonil, cyproconazole, and trifloxystrobin. After 14 days of incubation, a removal of up to 89% was achieved for chlorothalonil, 46.4% for cyproconazole, and 33.1% for trifloxystrobin using viable biomass. In the case of azoxystrobin, the highest removal (82.2%) occurred by adsorption to fungal biomass. Ecotoxicological tests in Daphnia magna revealed that T1 has the highest removal potential, achieving significant elimination of every fungicide, while simultaneously detoxifying the aqueous matrix (except in the case of cyproconazole). Isolate T4 also exhibited an intermediate efficiency, while isolate T2 was unable to detoxify the matrix in most cases. The removal and detoxification of cyproconazole failed with all the isolates. These findings suggest that endosphere of wild plants could be an attractive guild to find new Trichoderma species with promising bioremediation capabilities. In addition, the results demonstrate that attention should be placed when combining certain types of agrochemicals with antagonistic fungi in Integrated Pest and Disease Management strategies or when transitioning to organic agriculture.

Characterization and phylogenetic analysis of the complete chloroplast genome of Ophiorrhiza pumila (Rubiaceae)

Ophiorrhiza pumila (Rubiaceae) is an herbaceous plant that grows streamside in forest gullies or wetlands in the shade. Complete chloroplast genome of O. pumila was obtained and analyzed its phylogeny relationship within Rubiaceae plants. The results showed that the genome had a typical quadripartite structure of 154,385 bp, and contained a total of 112 unique genes, including 79 protein-coding genes, 29 tRNA genes, and 4 rRNA genes. Phylogenetic analysis suggested that O. pumila is sister to a highly supported clade composed of 10 species including Morinda officinalis, Gynochthodes cochinchinensis, Saprosma merrillii, Hedyotis ovata, Foonchewia guangdongensis, Dunnia sinensis, Paederia scandens, Leptodermis scabrida, Rubia cordifolia, and Galium mollugo. The complete chloroplast genome provides valuable information for the phylogenetic analysis of O. pumila.

Geniposide Achieved an Anti-Inflammatory Effect Through the NF-κB/HIF-1α Signaling Pathway Mediated by VEGFA and HMOX1 Genes

Geniposide is a kind of iridoid glycoside compound mainly extracted from the dried and mature fruit of a Rubiaceae plant, Gardenia jasminoides J.Ellis, which has a certain anti-inflammatory effect, but mechanism of this effect remains unclear.In this study, we utilized the network pharmacology approach, molecular docking, and in vitro experiments on the pharmacodynamics and anti-inflammatory mechanism of Geniposide. We leveraged the public databases to uncover putative targets across Geniposide and inflammatory diseases and constructed the anti-inflammatory protein–protein interaction (PPI) network of Geniposide following the intersection targets between them, further molecular docking, and enrichment analysis were performed. Finally, in vitro experiments were conducted to validate the predicted target genes and pathways. A total of 59 Geniposide-related targets and 8954 inflammatory diseases related targets were screened out, and 50 intersection targets were eventually obtained. We took the key target of the top 10 for molecular docking, and vascular endothelial growth factor A (VEGFA), matrix metalloproteinase-2, recombination human heme oxygenase 1 (HMOX1), adenosine receptor A1, adenosine deaminasegenes were identified based on the free binding energy. Hypoxia-inducible factor 1 (HIF-1) was predicted as the critical path of Geniposide anti-inflammatory by the Kyoto Encyclopedia of Genes and Genomessignaling pathway enrichment analysis. The results showed that Geniposide could regulate the gene expression of VEGFA and HMOX1 and influence the protein expression levels of HIF-1α, VEGFA,HOMX1,and p-p65 in the NF-κB/HIF-1α signaling pathway. Our data revealed that the anti-inflammatory effect of Geniposide was at least through the NF-κB/HIF-1α signaling pathway mediated by VEGFA and HMOX1 genes.

Neotropical Rubiaceae: Synthesis of Chromosome Data from Costa Rican Taxa, with Insights on the Systematics of the Family

Recent molecular systematic studies have significantly improved our understanding of the large, complex, and cosmopolitan plant family Rubiaceae, comprising about 13,000 species. Besides the obvious importance of DNA phylogenetic data, cytological studies have long added important basic information on the circumscription of clades and relationships within the family. In light of recent changes affecting a large number of tribes and genera, the current knowledge on the systematics of Neotropical Rubiaceae is reviewed with a focus on Costa Rica, which harbors an exceptionally rich Rubiaceae flora including most of the genera and biogeographic elements present in the Neotropics. Based on this systematic framework, previously published chromosome counts on Costa Rican taxa are reviewed and 49 new chromosome counts are reported. In total, 110 accessions of 75 species or infraspecific taxa representing 36 genera of Costa Rican Rubiaceae are discussed and supplemented by new counts for extraterritorial taxa when appropriate. Altogether the present study includes the first chromosome counts reported for the tribes Cordiereae and Hillieae, as well as for 10 genera and 27 species, providing new aspects of Rubiaceae systematics.

Cytotoxicity Activities of Ethanol Extract of Hooks Uncaria tomentosa West Kalimantan

Uncaria tomentosa is a member of the plant family Rubiaceae. It has been used as medicinal plants in West Kalimantan. The cytotoxic of ethanol extract from the hooks of U. Tomentosa was determined. This study used Brine Shrimp Lethality Test (BSLT) method with solution concentration 1,000; 5,000 and 10,000 ppm. The extract has LC50 values of 21,754 ppm. It is indicated the extract not toxic. This extract is potent to be used as drugs.

Genetic and phenotypic characterization of Xanthomonas axonopodis pv. maculifoliigardeniae causing bacterial leaf spot of Ixora in Taiwan

AbstractIxora spp. are Rubiaceae plants commonly planted as hedges or potted flower. Recently, incidents of bacterial leaf spot of Ixora were observed in central parts of Taiwan. Previous research on the disease has been scarce and focused mainly on its diagnosis. Therefore, many characteristics of the causal agent remain unclear. The present study aims to improve our understanding of this lesser‐characterized pathogen and provide information useful for its identification and management. Bacterial strains Ixo1, Ixo2 and Ixo3 were isolated from infected Ixora x westii. All three isolates were able to grow and induce leaf spot symptoms on Ixora. They also exhibited morphological and physiological characteristics typical of Xanthomonads. Biolog analysis indicated that Ixo1 to Ixo3 have metabolic fingerprints similar to X. axonopodis pv. poinsettiicola. Multilocus sequence analysis and inoculation assays identified Ixo1 to Ixo3 as X. axonopodis pv. maculifoliigardeniae, albeit their gene sequences were very similar to other species/pathovars belonging to the X. euvesicatoria species complex; members of this species complex have different plant hosts, yet share similar housekeeping gene sequences. A semi‐specific PCR assay evaluated in this work was able to differentiate Ixo1 to Ixo3 from bacteria not belonging to the X. euvesicatoria species complex, suggesting that the assay may be used in diagnosing bacterial leaf spot of ixoras. Finally, the sensitivity of the isolated pathogen to multiple commercial pesticides was tested, and the results showed that the bacterium is sensitive to streptomycin + tetracycline (10％ SP), thiophanate methyl + streptomycin (68.8％ WP) and oxolinic acid (20% WP), but more tolerant against copper‐based chemicals. Overall, the findings from this work may facilitate the identification and management of X. axonopodis pv. maculifoliigardeniae.

Conflicting phylogenetic signals in genomic data of the coffee family (Rubiaceae)

AbstractReconstructions of phylogenetic relationships in the flowering plant family Rubiaceae have up until now relied heavily on single‐ or multi‐gene data, primarily from the plastid compartment. With the availability of cost‐ and time‐efficient techniques for generating complete genome sequences, the opportunity arises to resolve some of the relationships that, up until now, have proven problematic. Here, we contribute new data from complete 58 plastid genome sequences, representing 55 of the currently 65 recognized tribes of the Rubiaceae. Also contributed are new data from the nuclear rDNA cistrons for corresponding taxa. Phylogenetic analyses are conducted on two plastid data sets, one including data from the protein coding genes only, and a second where protein coding data are combined with non‐coding regions, and on a nuclear rDNA data set. Our results clearly show that simply adopting a “more characters” approach does not resolve the relationships in the Rubiaceae. More importantly, we identify conflicting phylogenetic signals in the data. Analyses of the same plastid data, treated as nucleotides or as codon‐degenerated data, resolve and support conflicting topologies in the subfamily Cinchonoideae. As these analyses use the same data, we interpret the conflict to result from erroneous assumptions in the models used to reconstruct our phylogenies. Conflicting signals are also identified in the analyses of the plastid versus the nuclear rDNA data sets. These analyses use data from different genomic compartments, with different inheritance patterns, and we interpret the conflicts as representing “real” conflicts, reflecting biological processes of the past.

Is the bacterial leaf nodule symbiosis obligate for Psychotria umbellata? The development of a Burkholderia-free host plant.

Background & aimsThe bacterial leaf nodule symbiosis is an interaction where bacteria are housed in specialised structures in the leaves of their plant host. In the Rubiaceae plant family, host plants interact with Burkholderia bacteria. This interaction might play a role in the host plant defence system. It is unique due to its high specificity; the vertical transmission of the endophyte to the next generation of the host plant; and its supposedly obligatory character. Although previous attempts have been made to investigate this obligatory character by developing Burkholderia-free plants, none have succeeded and nodulating plants were still produced. In order to investigate the obligatory character of this endosymbiosis, our aims were to develop Burkholderia-free Psychotria umbellata plants and to investigate the effect of the absence of the endophytes on the host in a controlled environment.MethodsThe Burkholderia-free plants were obtained via embryo culture, a plant cultivation technique. In order to analyse the endophyte-free status, we screened the plants morphologically, microscopically and molecularly over a period of three years. To characterise the phenotype and growth of the in vitro aposymbiotic plants, we compared the growth of the Burkholderia-free plants to the nodulating plants under the same in vitro conditions.Key resultsAll the developed plants were Burkholderia-free and survived in a sterile in vitro environment. The growth analysis showed that plants without endophytes had a slower development.ConclusionsEmbryo culture is a cultivation technique with a high success rate for the development of Burkholderia-free plants of P. umbellata. The increased growth rate in vitro when the specific endophyte is present cannot be explained by possible benefits put forward in previous studies. This might indicate that the benefits of the endosymbiosis are not yet completely understood.