Colonization with non-mycorrhizal culturable endophytic fungi enhances orchid growth and indole acetic acid production.

BACKGROUND: Symbiotic associations of endophytic fungi have been proved by possessing an ability to produce hormones and metabolites for their host plant. Members of the Orchidaceae are obligate mycorrhizal species but a non-mycorrhizal association needs more investigation for their ability to promote plant growth and produce plant growth hormones. In the present study, endophytic fungi were isolated from the roots of Dendrobium longicornu Lindl., to investigate the root colonizing activity and role in plant growth and development. RESULTS: Among 23 fungal isolates were identified both by morphological and molecular technique as Penicillium sp., Fusarium sp., Coniochaeta sp., Alternaria sp., and Cladosporium sp. The dominate species were Coniochaeta sp. and Cladosporium sp. The dominant species as per the isolation was Coniochaeta sp. These fungal strains were screened for growth-promoting activity of Cymbidium aloifolium (plantlet) consider as cross genus interaction and Dendrobium longicornu (protocorms) as a host plant in in-vitro condition. Importantly, Cladosporium sp., and Coniochaeta sp. showed successful colonization and peloton formation with roots of C. aloifolium. Moreover, it also enhanced acclimatization of plantlets. Fungal elicitors from nine fungal isolates enhanced the growth of the in vitro grown protocorms of D. longicornu. Key bioactive compounds detected in the fungal colonized plant extract were 2H-pyran-2-one, Cyclopropanecarboxylic acid, Oleic Acid and d-Mannitol, which may have a potential role in plant-microbe interaction. All fungal endophytes were able to synthesize the indole acetic acid (IAA) in presence of tryptophan. Moreover, fungal extract DLCCR7 treated with DL-tryptophan yielded a greater IAA concentration of 43 µg per ml than the other extracts. The iaaM gene involved in IAA synthesis pathway was amplified using iaaM gene primers successfully from Alternaria sp., Cladosporium sp., and Coniochaeta sp. CONCLUSIONS: Hence, this study confirms the production of IAA by endophytes and demonstrated their host as well as cross-genus plant growth-promoting potential by producing metabolites required for the growth of the plant.

Structure-Activity Relationship of Carbon Nitride Dots in Inhibiting Tau Aggregation.

Due to the numerous failed clinical trials of anti-amyloid drugs, microtubule associated protein tau (MAPT) now stands out as one of the most promising targets for AD therapy. In this study, we report for the first time the structure-dependent MAPT aggregation inhibition of carbon nitride dots (CNDs). CNDs have exhibited great promise as a potential treatment of Alzheimer's disease (AD) by inhibiting the aggregation of MAPT. In order to elucidate its structure-activity relationship, CNDs were separated via column chromatography and five fractions with different structures were obtained that were characterized by multiple spectroscopy methods. The increase of surface hydrophilic functional groups is consistent with the increase of polarity from fraction 1 to 5. Particle sizes (1-2 nm) and zeta potentials (~-20 mV) are similar among five fractions. With the increase of polarity from fraction 1 to 5, their MAPT aggregation inhibition capacity was weakened. This suggests hydrophobic interactions between CNDs and MAPT, validated via molecular dynamics simulations. With a zebrafish blood-brain barrier (BBB) model, CNDs were observed to cross the BBB through passive diffusion. CNDs were also found to inhibit the generation of multiple reactive oxygen species, which is an important contributor to AD pathogenesis.

A prospectus of plant growth promoting endophytic bacterium from orchid (Vanda cristata).

BACKGROUND: A plant growth-promoting endophytic bacterium PVL1 isolated from the leaf of Vanda cristata has the ability to colonize with roots of plants and protect the plant. PVL1 was isolated using laboratory synthetic media. 16S rRNA gene sequencing method has been employed for identification before and after root colonization ability. RESULTS: Original isolated and remunerated strain from colonized roots were identified as Bacillus spp. as per EzBiocloud database. The presence of bacteria in the root section of the plantlet was confirmed through Epifluorescence microscopy of colonized roots. The in-vitro plantlet colonized by PVL1 as well as DLMB attained higher growth than the control. PVL1 capable of producing plant beneficial phytohormone under in vitro cultivation. HPLC and GC-MS analysis suggest that colonized plants contain Indole Acetic Acid (IAA). The methanol extract of Bacillus spp., contains 0.015 µg in 1 µl concentration of IAA. PVL1 has the ability to produce antimicrobial compounds such as ethyl iso-allocholate, which exhibits immune restoring property. One-way ANOVA shows that results were statistically significant at P ≤ 0.05 level. CONCLUSIONS: Hence, it has been concluded that Bacillus spp. PVL1 can promote plant growth through secretion of IAA during root colonization and ethyl iso-allocholate to protect plants from foreign infections. Thus, this study supports to support Koch's postulates of bacteria establishment.

Interfacial behavior of Proteinase K enzyme at air-saline subphase.

This study investigates the interfacial behavior of the proteinase K enzyme at air-water interface. Adsorption of enzyme on the surface was induced using saline subphase. The surface packing and stability of the enzyme was investigated using of surface pressure-area (π-A) and surface potential-area (ΔV-A) isotherms. Proteinase K enzyme forms film at air-aqueous interface and demonstrates good stability as shown through compression-decompression cycle experiments. To characterize the surface assembly morphology of the interfacial enzymes UV-vis and fluorescence spectroscopic techniques were used. The data revealed that the enzyme Langmuir monolayer has good homogeneity with no evidence of aggregates during compression. The secondary structure of the enzyme at interface was determined to be α-helix using p-polarized infrared-reflection absorption spectroscopy. This was confirmed through Circular dichroism spectra of the enzyme Langmuir-Blodgett (LB) film which showed that the major conformation present were α-helices.

Comparative Cytotoxic Activity of Wild Harvested Stems and In Vitro-Raised Protocorms of Dendrobium chryseum Rolfe in Human Cervical Carcinoma and Glioblastoma Cell Lines.

From the medicinal orchid Dendrobium chryseum Rolfe, which is used in traditional and folk Chinese medicine, the protocorms were raised in Murashige and Skoog (MS) media in three strengths, full strength (FMS), half strength (1/2 MS), and quarter strength (1/4 MS), with or without the phytohormones 6-benzylaminopurine (BAP) and 1-naphthaleneacetic acid (NAA) and coconut water (CW). The comparative cytotoxic activities of the wild and in vitro-raised protocorms were evaluated in human cervical carcinoma (HeLa) and human glioblastoma (U251) cell lines by MTT assay. In in vivo and in vitro, the methanol extracts of D. chryseum showed significant cytotoxic activities. Significant growth inhibition (%) and potent IC50 values were demonstrated in HeLa cell lines (49.79% (210.5 µg/mL) for in vitro-raised Dendrobium chryseum (DCT) versus 46.97% (226.5 µg/mL) for wild Dendrobium chryseum (DCW)). Similarly, activities against U251 cell lines exhibited also significant inhibition (28.76% (612.54 µg/mL) for DCW and 17.15% (1059.92 µg/mL) for DCT). The cytotoxic activities of both, wild and tissue-cultured samples, were superior in HeLa cells. In U251 cells, the wild sample was more active than the tissue-cultured one with a moderate cytotoxic effect. Hence, protocorm culture may therefore be a promising future tool for producing pharmacologically bioactive compounds in medicinal orchids. Such sustainable technology approach will minimize the pressure on the natural population of threatened but commercially important medicinal orchids.

Isolation, characterization, and plant growth-promoting activities of endophytic fungi from a wild orchid Vanda cristata.

Endophytism is one of the widely explored phenomena related to orchids and fungi. Endophytic fungi assist plants by supplementing nutrient acquisition, and synthesis of plant growth regulators. Vanda cristata is an epiphytic orchid that has a great diversity of endophytic fungi. Endophytic fungi were isolated from roots, stems, and leaves of V.cristata and identified by both morphological and molecular study. Furthermore, the isolated endophytic fungi were subjected to auxin synthesis, phosphate solubilization, ammonia synthesis, and elicitor growth test for understanding their growth-promoting effect in a qualitative and quantitative manner. Altogether, 12 different endophytic fungi were isolated from roots, stems, and leaves of V. cristata of which most species belonged to Ascomycota. Unidentified II fungi were found to be most effective for auxin synthesis and phosphate solubilization while Agaricus bisporous and Mycolepto discus were most effective for ammonia synthesis. We have tested the plant growth-promoting activity of the twelve isolated endophytic fungi on Cymbidium aloifolium protocorms (12 weeks old). All the endophytic fungi showed growth-promoting activity. Plant growth of Cymbidium aloifolium was found higher on the MS medium supplemented with all fungal elicitors. Fungal elicitor CVS4, however, showed the highest plant growth-promoting activity toward C. aloifolium.

Piriformospora indica promotes the growth of the in-vitro-raised Cymbidium aloifolium plantlet and their acclimatization.

Cymbidium aloifolium is known for its ornamental and medicinal values. It has been listed as threatened orchid species. In this study, in vitro propagated C. aloifolium plantlets were interacted with the Piriformospora indica. The growth assay was performed for 45 days; the plant growth pattern such as number and length of roots and shoots were measured. Microscopic study of the root section stained by trypan blue was done to detect the peloton formation. The methanol extracts of the fungal colonized plant as well as uncolonized (control) plant were prepared and various metabolites were identified by gas chromatography mass spectroscopy. Acclimatization was done in a substrate composition of coco peat: gravel: charcoal in ratio 2:2:1. P. indica-colonized plantlet showed the highest growth with the formation of clamdospore in the root section. The growth regulator such as auxin, ascorbic acid, andrographolide, hexadecanoic acid, and DL-proline were identified. After three months of field transfer, plantlet colonized by P. indica survived and remained healthy as compared to uncolonized control plantlet.

Isolation and Characterization of Plant Growth-Promoting Endophytic Fungi from the Roots of Dendrobium moniliforme.

The present study aims to identify the diverse endophytic fungi residing in the roots of Dendrobium moniliforme and their role in plant growth and development. Nine endophytic fungi were isolated from the root sections and characterized by molecular technique. Quantification of the indole acetic acid (IAA) compound by these endophytes was done. Further, Chemical profiling of R11 and R13 fungi was done by Gas Chromatography-Mass Spectroscopy (GC-MS). Asymbiotic seed derived protocorms of Rhynchostylis retusa was used for the plant growth assay to investigate the growth promoting activities of the fungal elicitor prepared from the isolated fungi from D. moniliforme. Among the isolated fungi, the relative dominant fungus was Fusarium sp. The R13 and R6 fungi were identified only at the genus level which concludes the fungi are of new species or strain. The indole acetic acid production was relatively higher in R10. Bioactive compound diversity was observed in the organic extract of R11 and R6. The presence of phenolic compound and essential oil suggest their contribution for the antimicrobial and antioxidant properties to their host plant, D. moniliforme. The plant growth assay result concluded, the fungal elicitor prepared from R10, Colletotrichum alatae was the best among all other for the plant growth activities.