Endophytic fungal community of Rosa damascena Mill. as a promising source of indigenous biostimulants: Elucidating its spatial distribution, chemical diversity, and ecological functions.

The role of endophytes in maintaining healthy plant ecosystems and holding promise for agriculture and food security is deeply appreciated. In the current study, we determine the community structure, spatial distribution, chemical diversity, and ecological functions of fungal endophytes of Rosa damascena growing in the North-Western Himalayas. Culture-dependent methods revealed that R. damascena supported a rich endophyte diversity comprising 32 genera and 68 OTUs. The diversity was governed by climate, altitude, and tissue type. Species of Aspergillus, Cladosporium, Penicillium, and Diaporthe were the core endophytes of the host plant consisting of 48.8% of the endophytes collectively. The predominant pathogen of the host was Alternaria spp., especially A. alternata. GC-MS analyses affirmed the production of diverse arrays of volatile organic compounds (VOC) by individual endophytes. Among the primary rose oil components, Diaporthe melonis RDE257, and Periconia verrucosa RDE85 produced phenyl ethyl alcohol (PEA) and benzyl alcohol (BA). The endophytes displayed varied levels of plant growth-promoting, colonization, and anti-pathogenic traits. Between the selected endophytes, P. verrucosa and D. melonis significantly potentiated plant growth and the flavonoids and chlorophyll content in the host. The potential of these two endophytes and their metabolites PEA and BA was confirmed on Nicotiana tabacum. The treatments of the metabolites and individual endophytes enhanced the growth parameters in the model plant significantly. The results imply that P. verrucosa and D. melonis are potential plant growth enhancers and their activity may be partially due to the production of PEA and BA. Thus, R. damascena harbors diverse endophytes with potential applications in disease suppression and host growth promotion. Further investigations at the molecular level are warranted to develop green endophytic agents for sustainable cultivation of R. damascena and biocontrol of leaf spot disease.

A Secondary Metabolite of Cercospora sp., Associated with Rosa damascena Mill., Inhibits Proliferation, Biofilm Production, Ergosterol Synthesis and Other Virulence Factors in Candida albicans.

Here we describe the antimicrobial potential of secondary metabolites, fulvic acid (F.A.) and anhydrofulvic acid (AFA), produced by RDE147, an endophyte of Rosa damascena Mill. The endophyte was identified as Cercospora piaropi by ITS and ß-tubulin-based phylogenetic analyses, while chemoprofiling of the endophyte by column chromatography and spectroscopy yielded two pure compounds, F.A. and AFA. The compounds demonstrated different antimicrobial profiles, with AFA suppressing the growth of C. albicans at 7.3 µg ml-1 IC50. Further studies revealed that AFA strongly restricted the biofilm production and hyphae formation in C. albicans by down-regulating several biofilm and morphogenesis-related genes. The time-kill assays confirmed the fungicidal activity of AFA against C. albicans, killing 83.6% of the pathogen cells in 24 h at the MIC concentration, and the post-antibiotic effect (PAE) experiments established the suppression of C. albicans growth for extended time periods. The compound acted synergistically with amphotericin B and nystatin and reduced ergosterol biosynthesis by the pathogen, confirmed by ergosterol estimation and comparative expression profiling of selected genes and molecular docking of AFA with C. albicans squalene epoxidase. AFA also suppressed the expression of several other virulence genes of the fungal pathogen. The study determines the anti-C. albicans potential of AFA and its impact on the biology of the pathogen. It also indicates that Cercospora species may yield potential bioactive molecules, especially fulvic acid derivatives. However, it is imperative to conduct in vivo studies to explore this molecule's therapeutic potential further.

Ultrasound-assisted facile synthesis of Boron-Heck-coupled sclareol analogues as potential antibacterial agents against Staphylococcus aureus.

AIM: To evaluate the antimicrobial capability of sclareol and its derivatives against Staphylococcus aureus and its Methicillin-resistant strain (MRSA). METHODS AND RESULTS: A new series of Boron-Heck-coupled sclareol analogues were prepared by structural modifications at the C-15 terminal double bond of sclareol using ultrasonication. The structural modifications were designed to keep the stereochemistry of all the five chiral centres of sclareol intact. A two-step reaction scheme consisting of Boron-Heck coupling of sclareol followed by Wittig reaction was carried out to produce novel sclareol congeners for antimicrobial evaluation. Three compounds SAJ-1, SAJ-2 and SB-11 exhibited strong antibacterial activity against S. aureus and Methicillin-resistant strain (MRSA) with MIC values between 3 and 11 µM. Among all the screened compounds, SAJ-1 and SAJ-2 showed the best antibiofilm profiles against both strains. Moreover, SAJ-1 and SAJ-2 acted synergistically with streptomycin against S. aureus while creating varying outcomes in combination with ciprofloxacin, penicillin and ampicillin. SAJ-1 also acted synergistically with ampicillin against S. aureus, while SB-11 showed synergism with ciprofloxacin against both pathogens. Moreover, SAJ-1 and SAJ-2 also inhibited staphyloxanthin production in S. aureus and MRSA and induced postantibiotic effects against both pathogens. CONCLUSIONS: It can be inferred that SAJ-1, SAJ-2 and SB-11 may act as potential chemical entities for the development of antibacterial substances. The study revealed that SAJ-1 and SAJ-2 are the most suitable sclareol analogues for further studies towards the development of antibacterial substances. SIGNIFICANCE AND IMPACT OF THE STUDY: SAJ-1, SAJ-2 and SB-11 show promising antibacterial properties against Staphylococcus aureus. Efforts should be made and more research should be done utilizing in vivo models to determine their efficacy as antibiotics.

Insights into the Endophytic Bacterial Microbiome of Crocus sativus: Functional Characterization Leads to Potential Agents that Enhance the Plant Growth, Productivity, and Key Metabolite Content.

The study was undertaken to unravel the culturable endophytic bacterial microbiome of Crocus sativus L. (saffron crocus) and consequently obtain potential leads to develop plant growth-promoting and biocontrol agents for increased productivity and sustainable cultivation. The endophytes formed 47 different operational taxonomic units (OTUs), spanning over 28 genera. The host was preferentially colonized by the genus Bacillus, followed by Burkholderia and Pantoea, respectively. Several endophytes possessed potential plant growth-promoting properties and inhibitory activities against the specific fungal pathogens of saffron. The endophytes, except for Microbacterium oxydans, did not cause any disease symptoms in the pot experiments. The selected cultures, Burkholderia gladioli, Streptomyces achromogenes, and three species of Bacillus, enhanced the host plant growth significantly. Based on the pot experiment results, two isolates, Bacillus mojavensis CS4EB32 and Burkholderia gladioli E39CS3, were selected for the field experiments. We obtained an increase of 67.5%, 69.8%, and 68.3% in the production of flowers with the individual and collective treatments, respectively. The treatments also enhanced the biomass of the plant and the length and weight of stigmas significantly. The endophyte treatments induced the expression of the pathway genes, resulting in a marked increase in the concentration of apocarotenoids. The study indicates that the dominant endophytes support plant growth and development in nature and present an opportunity for developing microbial formulations for the sustainability of saffron cultivation.

Burkholderia gladioli E39CS3, an endophyte of Crocus sativus Linn., induces host resistance against corm-rot caused by Fusarium oxysporum.

AIM: To investigate the role of the leading saffron endophyte Burkholderia gladioli strain E39CS3 (BG-E39) in the inhibition of corm-rot and induced systemic resistance (ISR) in the host against the saffron specific pathogen, Fusarium oxysporum. METHODS AND RESULTS: We studied the interaction between BG-E39 and the corm-rot pathogen F. oxysporum in vitro and in vivo. BG-E39 strongly inhibited both the F. oxysporum strains and other saffron-specific and non-specific pathogens used in this study. Confrontation and microscopic analyses revealed that the endophyte possessed fungicidal activity against the pathogens and effectively induced cell death in the mycelia. The endophyte produced chitinases as well as ß-1,3-glucanase that may be involved in the pathogen cell wall degradation. BG-E39 did not cause corm-rot in Crocus sativus and the closely related plant, Gladiolus, thus establishing that it is non-pathogenic to these plants. The endophyte reduced corm-rot through antibiosis and enhanced the endogenous jasmonic acid (JA) levels and expression of JA-regulated and other plant defence genes. CONCLUSIONS: The bacterial endophyte BG-E39 provides resistance to the host plant against F. oxysporum corm-rot in nature. SIGNIFICANCE AND IMPACT OF THE STUDY: The current study discovers the role of the saffron endophyte BG-E39 in providing resistance to the host against corm-rot. Therefore, this endophyte is a potential candidate for developing a microbial formulation for the biocontrol of the most common disease of C. sativus.

Discovery of a Secalonic Acid Derivative from Aspergillus aculeatus, an Endophyte of Rosa damascena Mill., Triggers Apoptosis in MDA-MB-231 Triple Negative Breast Cancer Cells.

A new secalonic acid derivative, F-7 (1), was isolated from the endophytic Aspergillus aculeatus MBT 102, associated with Rosa damascena. The planar structure of 1 was established on the basis of 1D and 2D NMR and ESI-TOF-MS spectra. The relative configuration of 1 was determined applying a combined quantum mechanical/NMR approach and, afterward, the comparison of calculated and experimental electronic circular dichroism spectra determined the assignment of its absolute configuration. The compound possesses strong cytotoxic activity against triple negative breast cancer (TNBC) cells. It was found to induce apoptosis, as evidenced by scanning electron microscopy and phase contrast microscopy. Furthermore, flow cytometry analyses demonstrated that 1 induced mitochondrial damage and reactive oxygen species mediated apoptosis, arresting the G1 phase of the cells in a dose-dependent manner. Also, the compound causes significant microtubule disruption in TNBC cells. Subsequently, 1 restricted the cell migration leading to the concomitant increase in expression of cleaved caspase and PARP.