Assessment of chemical and genetic variability in Tanacetum gracile accessions collected from cold desert of Western Himalaya.

Genetic diversity is essential for survival and adaptation of high altitude plants such as those of Tanacetum genus, which are constantly exposed to environmental stress. We collected flowering shoots of ten accessions of Tanacetum gracile Hook.f. & Thomson (Asteraceae) (Tg 1-Tg 10), from different regions of cold desert of Western Himalaya. Chemical profile of the constituents, as inferred from GC-MS, exhibited considerable variability. Percentage yield of essential oil ranged from 0.2 to 0.75% (dry-weight basis) amongst different accessions. Tg 1 and Tg 6 were found to produce high yields of camphor (46%) and lavandulol (41%), respectively. Alpha-phellendrene, alpha-bisabool, p-cymene and chamazulene were the main oil components in other accessions. Genetic variability among the accessions was studied using RAPD markers as well as by sequencing and analyzing nuclear 18S rDNA, and plastid rbcL and matK loci. The polymorphic information content (PIC) of RAPD markers ranged from 0.18 to 0.5 and the analysis clustered the accessions into two major clades. The present study emphasized the importance of survey, collection, and conservation of naturally existing chemotypes of medicinal and aromatic plants, considering their potential use in aroma and pharmaceutical industry.

Mortierella alpina CS10E4, an oleaginous fungal endophyte of Crocus sativus L. enhances apocarotenoid biosynthesis and stress tolerance in the host plant.

Crocus sativus is the only plant species which produces apocarotenoids like crocin, picrocrocin and safranal in significant amounts. These compounds impart organoleptic properties to saffron (dried stigmas of Crocus flower) making it world's costliest spice. Crocus apocarotenoids have tremendous medicinal properties as well. Effect of endophytes on Crocus apocarotenoid production and the molecular mechanism involved has not been reported so far. Here we studied the effect of an oleaginous fungal endophyte, Mortierella alpina CS10E4 on Crocus growth, apocarotenoid metabolism and tolerance to corm rot disease. The results demonstrated that there was a significant improvement in many morphological and physiological traits in endophyte treated Crocus plants including total biomass and size of corms, stigma biomass, number of apical sprouting buds, and number of adventitious roots. The endophyte also shifted metabolic flux towards enhanced production of apocarotenoids by modulating the expression of key pathway genes. Further, M. alpina CS10E4 enhanced tolerance to corm rot disease by releasing arachidonic acid which acts as conserved defense signal and induces jasmonic acid production in endophyte treated Crocus corms. This is first report on effect of a fungal endophyte on Crocus apocarotenoid metabolism and stress tolerance.

An endophytic Fusarium sp. isolated from Monarda citriodora produces the industrially important plant-like volatile organic compound hexanal.

An endophytic fungus, MC_25L, has been isolated from the leaves of MonardacitriodoraCerv. ex Lag., a medicinal and aromatic herb from the northwestern Himalayas. It produces a fruity fragrance while growing on potato dextrose agar, suggesting that it is producing volatile organic compounds (VOCs). The endophyte inhibited the growth of plant pathogens such asSclerotiniasp. and Aspergillusflavus by virtue of VOCs. Identification of MC_25L based on morphological and microscopic features, as well as ITS-based rDNA sequence analysis, revealed that it is a Fusariumsp. GC-MS analysis revealed that this endophyte produces a unique array of VOCs, in particular hexanal, p-fluoroanisole, pentafluoropropionic acid 2-ethylhexyl, (5E)-5-ethyl-2-methyl-5-hepten-3-one, 2-butyl-2-hexanol, (7E)-2-methyl-7-hexadecene and acoradiene. Three major compounds were hexanal, (5E)-5-ethyl-2-methyl-5-hepten-3-one and acoradiene, and they account for around 84.57 % of the total VOCs. Moreover, of interest was the presence of hexanal, which has applications in the food and cosmetic industries, as well as in mycofumigation. This is the first report of a fungal endophyte producing the industrially important plant-like VOC hexanal. Hexanal is also active biologically. Thus this study indicates that Fusariumsp. (MC_25L) is a potential candidate for the up-scaling of hexanal.

An endophyte of Picrorhiza kurroa Royle ex. Benth, producing menthol, phenylethyl alcohol and 3-hydroxypropionic acid, and other volatile organic compounds.

An endophytic fungus, PR4 was found in nature associated with the rhizome of Picrorhiza kurroa, a high altitude medicinal plant of Kashmir Himalayas. The fungus was found to inhibit the growth of several phyto-pathogens by virtue of its volatile organic compounds (VOCs). Molecular phylogeny, based on its ITS1-5.8S-ITS2 ribosomal gene sequence, revealed the identity of the fungus as Phomopsis/Diaporthe sp. This endophyte was found to produce a unique array of VOCs, particularly, menthol, phenylethyl alcohol, (+)-isomenthol, ß-phellandrene, ß-bisabolene, limonene, 3-pentanone and 1-pentanol. The purification of compounds from the culture broth of PR4 led to the isolation of 3-hydroxypropionic acid (3-HPA) as a major metabolite. This is the first report of a fungal culture producing a combination of biologically and industrially important metabolites­menthol, phenylethyl alcohol, and 3-HPA. The investigation into the monoterpene biosynthetic pathway of PR4 led to the partial characterization of isopiperitenone reductase (ipr) gene, which seems to be significantly distinct from the plant homologue. The biosynthesis of plant-like-metabolites, such as menthol, is of significant academic and industrial significance. This study indicates that PR4 is a potential candidate for upscaling of menthol, phenylethyl alcohol, and 3-HPA, as well as for understanding the menthol/monoterpene biosynthetic pathway in fungi.