Non-nodulated bacterial leaf symbiosis promotes the evolutionary success of its host plants in the coffee family (Rubiaceae).

Every plant species on Earth interacts in some way or another with microorganisms and it is well known that certain forms of symbiosis between different organisms can drive evolution. Within some clades of Rubiaceae (coffee family), a specific plant-bacteria interaction exists in which non-pathological endophytes are present in the leaves of their hosts. It is hypothesized that the bacterial endophytes, either alone or by interacting with the host, provide chemical protection against herbivory or pathogens by producing toxic or otherwise advantageous secondary metabolites. If the bacteria indeed have a direct beneficial influence on their hosts, it is reasonable to assume that the endophytes may increase the fitness of their hosts and therefore it is probable that their presence also has an influence on the long-term evolution of the particular plant lineages. In this study, the possible origin in time of non-nodulated bacterial leaf symbiosis in the Vanguerieae tribe of Rubiaceae is elucidated and dissimilarities in evolutionary dynamics between species with endophytes versus species without are investigated. Bacterial leaf symbiosis is shown to have most probably originated in the Late Miocene, a period when the savannah habitat is believed to have expanded on the African continent and herbivore pressure increased. The presence of bacterial leaf endophytes appears to be restricted to Old World lineages so far. Plant lineages with leaf endophytes show a significantly higher speciation rate than plant lineages without endophytes, while there is only a small difference in extinction rate. The transition rate shows that evolving towards having endophytes is twice as fast as evolving towards not having endophytes, suggesting that leaf symbiosis must be beneficial for the host plants. We conclude that the presence of bacterial leaf endophytes may also be an important driver for speciation of host plants.

α-Glucosidase Inhibitors from a Xylaria feejeensis Associated with Hintonia latiflora.

Two new compounds, pestalotin 4'-O-methyl-ß-mannopyranoside (1) and 3S,4R-(+)-4-hydroxymellein (2), were isolated from an organic extract of a Xylaria feejeensis, which was isolated as an endophytic fungus from Hintonia latiflora. In addition, the known compounds 3S,4S-(+)-4-hydroxymellein (3), 3S-(+)-8-methoxymellein (4), and the quinone derivatives 2-hydroxy-5-methoxy-3-methylcyclohexa-2,5-diene-1,4-dione (5), 4S,5S,6S-4-hydroxy-3-methoxy-5-methyl-5,6-epoxycyclohex-2-en-1-one (6), and 4R,5R-dihydroxy-3-methoxy-5-methylcyclohexen-2-en-1-one (7) were obtained. The structures of 1 and 2 were elucidated using a set of spectroscopic and spectrometric techniques. The absolute configuration of the stereogenic centers of 1 and 2 was determined using ECD spectroscopy combined with time-dependent density functional theory calculations. In the case of 1, comparison of the experimental and theoretical (3)J6-7 coupling constants provided further evidence for the stereochemical assignments. Compounds 2 and 3 inhibited Saccharomyces cerevisiae α-glucosidase (αGHY), with IC50 values of 441 ± 23 and 549 ± 2.5 µM, respectively. Their activity was comparable to that of acarbose (IC50 = 545 ± 19 µM), used as positive control. Molecular docking predicted that both compounds bind to αGHY in a site different from the catalytic domain, which could imply an allosteric type of inhibition.

Biologically active eremophilane-type sesquiterpenes from Camarops sp., an endophytic fungus isolated from Alibertia macrophylla.

Two new eremophilane-type sesquiterpenes, xylarenones F (3) and G (4), have been isolated from solid substrate cultures of a Camarops sp. endophytic fungus isolated from Alibertia macrophylla, together with the known compounds xylarenones C (1) and D (2). The structures and relative configurations of 1-4 were elucidated by extensive NMR and HRESIMS spectroscopic analysis. Due to their effects on the respiratory burst of neutrophils, which included inhibition of the reactive oxygen species production, these sesquiterpenes exhibited potential anti-inflammatory and antioxidant properties.

Mycoleptones A-C and polyketides from the endophyte Mycoleptodiscus indicus.

Three new azaphilones with an unusual methylene bridge, named mycoleptones A, B, and C (2, 4, and 5), were isolated from cultures of Mycoleptodiscus indicus, a fungus associated with the South American medicinal plant Borreria verticillata. Additionally, four known polyketides, austdiol (1), eugenitin (3), 6-methoxyeugenin (6), and 9-hydroxyeugenin (7), were also isolated. The structural characterization of compounds was carried out by nuclear magnetic resonance spectroscopy, high-resolution mass spectrometry, electronic circular dichroism spectroscopy, time-dependent density functional theory calculations, and X-ray crystallography. Compounds 1-9 were weakly active when tested in antileishmanial and cytotoxicity assays.

Genome sequence of Fusarium oxysporum strain ByF01, the causal agent of root rot of Knoxia roxburghii in China.

OBJECTIVES: Knoxia roxburghii is a member of the madder (Rubiaceae) family. This plant is cultivated in different areas of China and recognized for its medicinal properties, which leads to its use in traditional Chinese medicine. The incidence of root rot was 10-15%. In June 2023, the causal agent of root rot on K. roxburghii was identified as Fusarium oxysporum. To the best of our knowledge, this is the first report of the complete genome of F. oxysporum strain ByF01 that is the causal agent of root rot of K. roxburghii in China. The results will provide effective resources for pathogenesis on K. roxburghii and the prevention and control of root rot on this host in the future. DATA DESCRIPTION: To understand the molecular mechanisms used by F. oxysporum to cause root rot on K. roxburghii, strain ByF01 was isolated from diseased roots and identified by morphological and molecular methods. The complete genome of strain ByF01 was then sequenced using a combination of the PacBio Sequel IIe and Illumina sequencing platforms. We obtained 54,431,725 bp of nucleotides, 47.46% GC content, and 16,705 coding sequences.

Phylogenetic lineages in Vanguerieae (Rubiaceae) associated with Burkholderia bacteria in sub-Saharan Africa.

PREMISE OF THE STUDY: It is well known that mutualistic bacteria can provide substantial benefits to their host plants. However, 'how,' 'why,' and the possible applications of such an interaction are only second to the questions 'who is involved?', and 'where does it occur?'. In the coffee family (Rubiaceae), certain species closely interact with endophytic leaf bacteria that are freely distributed among the mesophyll cells. This type of interaction was recently discovered in South Africa. Our aim is to document the bacterial diversity associated with Rubiaceae ('who') and to establish the geographic range of the interaction ('where'). METHODS: Representatives of the Vanguerieae tribe in Rubiaceae were investigated for the presence of endophytes with special emphasis on the distributional range of the plant-bacteria association by collecting specimens from different African regions. KEY RESULTS: The interaction is found in five genera and is restricted to three major host lineages. The endophytic bacteria belong to the genus Burkholderia and are part of the plant-associated beneficial and environmental group. Some endophytes are similar to B. caledonica, B. graminis, B. phenoliruptrix or B. phytofirmans, while others are classified in OTUs that show no similarity with any previously described Burkholderia species of bacteria. CONCLUSIONS: The association is not obligate from the bacterial point of view and is considered a loose and recent interaction, which is demonstrated by the fact that there is no evidence for coevolution. The geographical distribution of the association is restricted by the distributional range of the host plants covering the whole of sub-Saharan Africa.

New Meliolaceae from the Brazilian Atlantic forest 2: species on host families Annonaceae, Cecropiaceae, Meliaceae, Piperaceae, Rubiaceae, Rutaceae and Tiliaceae.

Continuing the study of black mildews in fragments of the Atlantic forest, three new species and five new records are described herein. Irenopsis luheae-grandiflorae, Meliola vicosensis and Meliola xylopia-sericiae are new species. Cecropia hololeuca, Piper gaudichaudianum and Trichilia lepidota are new hosts for Asteridiella leucosykeae, Asteridiella glabroides and Meliola trichiliae respectively. Asteridiella obesa and Meliola psychotriae var. chiococcae are reported for the first time from Brazil. The new species are described and illustrated based on light and scanning electron microscopy and tables with main characteristics of morphologically similar specimens with species collected in Viçosa are provided. Other species belonging to Meliolaceae collected on hosts belonging to the Annonaceae, Meliaceae and Tiliaceae in Brazil also were studied.

Identification of the bacterial endosymbionts in leaf nodules of Pavetta (Rubiaceae).

Three genera in the Rubiaceae (Pavetta, Psychotria and Sericanthe) harbour bacterial endosymbionts within leaf nodules or galls. The present paper identifies the bacterial endophytes in three leaf-nodulating Pavetta species. In order to reveal their identity and assess their phylogenetic position, 16S rRNA, recA and gyrB genes were sequenced from an extensive sampling of Burkholderia strains. This multigene approach results in a robust phylogeny, which places the bacterial endosymbionts of Pavetta at two distinct positions within the genus Burkholderia (class Betaproteobacteria), suggesting that leaf-nodulating endosymbionts within Pavetta have different origins. The endophytes of nodulated Psychotria species were recognized as the closest relatives to the Pavetta endosymbionts. Our results suggest that the endosymbionts of Pavetta represent novel species, which can be classified as 'Candidatus Burkholderia hispidae', 'Candidatus Burkholderia rigidae' and 'Candidatus Burkholderia schumannianae'.

Meroterpenes from endophytic fungus A1 of mangrove plant Scyphiphora hydrophyllacea.

Four new meroterpenes, guignardones F-I (1-4), together with two known compounds guignardones A (5) and B (6) were isolated from the endophytic fungus A1 of the mangrove plant Scyphiphora hydrophyllacea. Their structures and relative configurations were elucidated by spectroscopic data and single-crystal X-ray crystallography. A possible biogenetic pathway of compounds 1-6 was also proposed. All compounds were evaluated for inhibitory activity against methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus aureus.

A fatty acid glycoside from a marine-derived fungus isolated from mangrove plant Scyphiphora hydrophyllacea.

To study the antimicrobial components from the endophytic fungus A1 of mangrove plant Scyphiphora hydrophyllacea Gaertn. F., a new fatty acid glucoside was isolated by column chromatography from the broth of A1, and its structure was identified as R-3-hydroxyundecanoic acid methylester-3-O-α-l-rhamnopyranoside (1) by spectroscopic methods including 1D and 2D NMR (HMQC, (1)H-(1)H COSY and HMBC) and chemical methods. Antimicrobial assay showed compound 1 possessed modest inhibitory effect on Saphylococcus aureus and methicillin-resistant S. aureus (MRSA) using the filter paper disc agar diffusion method.

Bioproduction of Cinchona alkaloids by the endophytic fungus Diaporthe sp. associated with Cinchona ledgeriana.

We report that an endophytic filamentous fungus species of the genus Diaporthe isolated from Cinchona ledgeriana (Rubiaceae) produces Cinchona alkaloids (quinine, quinidine, cinchonidine, and cinchonine) upon cultivation in a synthetic liquid medium. This study provides evidence that Cinchona alkaloids are produced not only in Cinchona plant cells, but also in the endophytic microbe cells, and will help to elucidate the relationship between endophytic microbes and their host plants.

Two new meroterpenes from endophytic fungus A1 of Scyphiphora hydrophyllacea.

Two new meroterpenes, guignardone D (1) and guignardone E (2), were isolated from endophytic fungus A1 of Scyphiphora hydrophyllacea Gaertn. F. Their structures were established based on spectroscopic methods including 1D and 2D NMR (HMQC, (1)H-(1)H COSY, HMBC, and ROESY).

(R)-(+)-Lasiodiplodin isolated from the endophytic fungus Sordaria tamaensis exhibits potent antimycobacterial and anti-inflammatory activities in vitro and in vivo: a dual approach for the treatment of severe pulmonary tuberculosis.

OBJECTIVES: This study aimed to evaluate endophytic fungi isolated from Tocoyena bullata and Humiria balsamifera plant species for their antimycobacterial and anti-inflammatory activities, focusing on severe pulmonary tuberculosis cases which are often associated with exacerbated inflammation. METHODS: Mycobacterium suspensions were incubated with the samples for 5 days. RAW 264.7 macrophages stimulated with LPS were also incubated with them for 24 h to assess the inhibition of inflammatory mediator production and cytotoxicity. C57BL/6 mice were infected with Mtb M299 and treated for 15 days with lasiodiplodin (Lasio). KEY FINDINGS: Endophytic fungus Sordaria tamaensis, obtained from T. bullata, was the most promising. Its ethanolic extract impaired mycobacterial growth with MIC50 (µg/ml): 1.5 ± 0.6 (BCG), 66.8 ± 0.1 (H37Rv) and 80.0 ± 0.1 (M299). (R)-(+)-Lasio showed MIC50 92.2 ± 1.8 µg/ml (M299). In addition, Lasio was able to inhibit NO, IL-1ß and TNF-α production and was not cytotoxic for macrophages. M. tuberculosis-infected C57BL/6 animals treated by Lasio reduced the number of acid-fast bacilli, lung pathology, leucocyte influx and proinflammatory cytokine production in the lungs. The class IIa fructose 1,6-bisphosphate aldolase was the predicted hypothetical target of Lasio. CONCLUSIONS: (R)-(+)-Lasio stood out as a promising anti-TB compound, exhibiting anti-inflammatory and antimycobacterial effects, as well as low cytotoxicity.

Xylarenones C-E from an endophytic fungus isolated from Alibertia macrophylla.

Xylarenones C-E (2-4), three new eremophilane sesquiterpenes, have been isolated from solid substrate cultures of a Camarops-like endophytic fungus isolated from Alibertia macrophylla. The structures were elucidated by analysis of spectroscopic data. Compounds were evaluated in subtilisin and pepsin protease assays, and compound 2 showed potent inhibitory activity against both proteases.

Mainland and island populations of Mussaenda kwangtungensis differ in their phyllosphere fungal community composition and network structure.

We compared community composition and co-occurrence patterns of phyllosphere fungi between island and mainland populations within a single plant species (Mussaenda kwangtungensis) using high-throughput sequencing technology. We then used 11 microsatellite loci for host genotyping. The island populations differed significantly from their mainland counterparts in phyllosphere fungal community structure. Topological features of co-occurrence network showed geographic patterns wherein fungal assemblages were less complex, but more modular in island regions than mainland ones. Moreover, fungal interactions and community composition were strongly influenced by the genetic differentiation of host plants. This study may advance our understanding of assembly principles and ecological interactions of phyllosphere fungal communities, as well as improve our ability to optimize fungal utilization for the benefit of people.

Bioactive metabolites produced by Penicillium sp. 1 and sp. 2, two endophytes associated with Alibertia macrophylla (Rubiaceae).

In the course of our continuous search for bioactive metabolites from endophytic fungi living in plants from the Brazilian flora, leaves of Alibertia macrophylla (Rubiaceae) were submitted to isolation of endophytes, and two species of Penicillium were isolated. The acetonitrile fraction obtained in corn from a culture of Penicillium sp. 1 afforded orcinol (1). On the other hand, Penicillium sp. 1 cultivated in potato-dextrose-broth furnished two different compounds, cyclo-(L-Pro-L-Val) (2) and uracil (3). The chromatographic fractionation of the acetonitrile fraction obtained from Penicillium sp. 2 led to three dihydroisocoumarins, 4-hydroxymellein (4), 8-methoxymellein (5) and 5-hydroxymellein (6). Compounds 5 and 6 were obtained from the Penicillium genus for the first time. Additionally, metabolites 1-6 were evaluated for their antifungal and acetylcholinesterase (AChE) inhibitory activities. The most active compounds 1 and 4 exhibited detection limits of 5.00 and 10.0 microg against Cladosporium cladosporioides and C. sphaerospermum, respectively. Compound 2 showed a detection limit of 10.0 microg, displaying potent AChE inhibitory activity.

Effects of mixing Neolamarckia cadamba leaves on fermentation quality, microbial community of high moisture alfalfa and stylo silage.

Neolamarckia cadamba is not only a fodder of high nutritional value, but also a source of natural antimicrobial agent. The silage quality of high moisture alfalfa and stylo with or without N. cadamba leaves (NCL) was investigated, and microbial community after ensiling was analysed. Results showed that the silage samples with NCL have lower pH (4.32 versus 4.88, 4.26 versus 4.71 in alfalfa and stylo silage, respectively), ammonia-N content (67.5 versus 146, 42.2 versus 95.1 g kg-1 total N) and higher lactic acid (13.3 versus 10.4, 17.3 versus 13.6 g kg-1 dry matter), true protein N (592 versus 287, 815 versus 589 g kg-1 total N). The addition of NCL also influenced the bacterial community distribution. The relative abundance of Clostridium and Enterobacter decreased, whereas Lactobacillus abundance increased when NCL was added. In conclusion, NCL could inhibit undesirable microorganisms in high moisture alfalfa and stylo silage. Mixing with NCL could be a feasible way to improve the quality of silage.

An endophytic Neurospora sp. from Nothapodytes foetida producing camptothecin.

The medicinal plant, Nothapodytes foetida contains a number of important alkaloids like camptothecin (an anticancer drug molecule) but its concentration is less to meet the existing demand of this important molecule, so in an effort for accessible availability of camptothecin. An endophyte (designated ZP5SE) was isolated from the seed of Nothapodytes foetida and was examined as potential source of anticancer drug lead compound i.e. camptothecin, when grown in Sabouraud liquid culture media under shake flask conditions. The presence of anticancer compound (camptothecin) in this fungus was confirmed by chromatographic and spectroscopic methods in comparison with authentic camptothecin. Isolated endophyte (Neurospora crassa) producing camptothecin may become an easily accessible source for the production of precursor anticancer drug molecule in future at large scale.

Leaf nodule symbiosis: function and transmission of obligate bacterial endophytes.

Various plant species establish intimate symbioses with bacteria within their aerial organs. The bacteria are contained within nodules or glands often present in distinctive patterns on the leaves, and have been used as taxonomic marker since the early 20th century. These structures are present in very diverse taxa, including dicots (Rubiaceae and Primulaceae) and monocots (Dioscorea). The symbionts colonize the plants throughout their life cycles and contribute bioactive secondary metabolites to the association. In this review, we present recent progress in the understanding of these plant-bacteria symbioses, including the modes of transmission, distribution and roles of the symbionts.

Intraspecific variation in Burkholderia caledonica: Europe vs. Africa and soil vs. endophytic isolates.

The best-known interaction between bacteria and plants is the Rhizobium-legume symbiosis, but other bacteria-plant interactions exist, such as between Burkholderia and Rubiaceae (coffee family). A number of bacterial endophytes in Rubiaceae are closely related to the soil bacterium Burkholderia caledonica. This intriguing observation is explored by investigating isolates from different geographic regions (Western Europe vs. sub-Saharan Africa) and from different niches (free-living bacteria in soil vs. endophytic bacteria in host plants). The multilocus sequence analysis shows five clades, of which clade 1 with two basal isolates deviates from the rest and is therefore not considered further. All other isolates belong to the species B. caledonica, but two genetically different groups are identified. Group A holds only European isolates and group B holds isolates from Africa, with the exception of one European isolate. Although the European and African isolates are considered one species, some degree of genetic differentiation is evident. Endophytic isolates of B. caledonica are found in certain members of African Rubiaceae, but only in group B. Within this group, the endophytes cannot be distinguished from the soil isolates, which indicates a possible exchange of bacteria between soil and host plant.