Comparative analysis of miRNA expression profiles in flowering and non-flowering tissue of Crocus sativus L.

Crocus sativus is a valuable plant due to the presence of apocarotenoids in its stigma. Considerable work has been done in the past to understand the apocarotenoid biosynthetic pathway in saffron. However, the reports on understanding the regulation of flowering at the post-transcriptional level are meagre. The study aimed to discover the candidate miRNAs, target genes, transcription factors (TFs), and apocarotenoid biosynthetic pathway genes associated with the regulation and transition of flowering in C. sativus. In the present investigation, miRNA profiling was performed in flowering and non-flowering corms of saffron, along with expression analysis of apocarotenoid genes and transcription factors involved in the synthesis of secondary metabolites. Significant modulation in the expression of miR156, miR159, miR166, miR172, miR395, miR396, miR399, and miR408 gene families was observed. We obtained 36 known miRNAs (26 in flowering and 10 in non-flowering) and 64 novel miRNAs (40 in flowering and 24 in non-flowering) unique to specific tissues in our analysis. TFs, including CsMADS and CsMYb, showed significant modulation in expression in flowering tissue, followed by CsHB. Additionally, the miRNAs were predicted to be involved in carbohydrate metabolism, phytohormone signalling, regulation of flower development, and response to stress, cold, and defence. The comprehensive study has enhanced our understanding of the regulatory machinery comprising factors like phytohormones, abiotic stress, apocarotenoid genes, transcription factors, and miRNAs responsible for the synthesis of apocarotenoids and developmental processes during and after flowering.

Green synthesis and biological evaluation of glaucanic acid and dihydrocompactin acid by endophytic fungus Penicillium polonicum from Zingiber officinale.

Fungal endophytes are valued for biosynthesizing chemically diverse metabolic cascade with interesting biological activities. In the current investigation, two compounds were isolated from Penicillium polonicum, an endophyte of Zingiber officinale. The active moieties, glaucanic acid (1) and dihydrocompactin acid (2) were isolated from the ethyl acetate extract of P. polonicum and characterized by NMR and mass spectroscopy. Further, bioactive potential of the isolated compounds was evaluated by antimicrobial, antioxidant and cytotoxicity assays. Compounds 1 and 2 displayed antifungal activity against phytopathogen Colletotrichum gloeosporioides with more than 50% reduction in its growth. Both the compounds exhibited antioxidant activity against free radicals (DPPH and ABTS) and cytotoxicity activity against cancer cell lines respectively. The compounds, glaucanic acid and dihydrocompactin acid are being reported for the first time from an endophytic fungus. This is the first report on the biological activities of Dihydrocompactin acid produced by endophytic fungal strain.

Fungal endophyte bioinoculants as a green alternative towards sustainable agriculture.

Over the past half century, limited use of synthetic fertilizers, pesticides, and conservation of the environment and natural resources have become the interdependent goals of sustainable agriculture. These practices support agriculture sustainability with less environmental and climatic impacts. Therefore, there is an upsurge in the need to introduce compatible booster methods for maximizing net production. The best straightforward strategy is to explore and utilize plant-associated beneficial microorganisms and their products. Bioinoculants are bioformulations consisting of selected microbial strains on a suitable carrier used in the enhancement of crop production. Fungal endophytes used as bioinoculants confer various benefits to the host, such as protection against pathogens by eliciting immune response, mineralization of essential nutrients, and promoting plant growth. Besides, they also produce various bioactive metabolites, phytohormones, and volatile organic compounds. To design various bioformulations, transdisciplinary approaches like genomics, transcriptomics, metabolomics, proteomics, and microbiome modulation strategies like gene editing and metabolic reconstruction have been explored. These studies will refine the existing knowledge on the diversity, phylogeny and beneficial traits of the microbes. This will also help in synthesizing microbial consortia by evaluating the role of structural and functional elements of communities in a controlled manner. The present review summarizes the beneficial aspects associated with fungal endophytes for capitalizing agricultural outputs, enlists various multi-omics techniques for understanding and modulating the mechanism involved in endophytism and the generation of new bioformulations for providing novel solutions for the enhancement of crop production.

A Brief Review of Plant Cell Transfection, Gene Transcript Expression, and Genotypic Integration for Enhancing Compound Production.

Plants possess a plethora of important secondary metabolites, which are unique sources of natural pigments, pharmaceutical compounds, food additives, natural pesticides, and other industrial components. The commercial significance of such metabolites/compounds has directed the research toward their production and exploration of methods for enhancement of production. Biotechnological tools are critical in selecting, integrating, multiplying, improving, and analyzing medicinal plants for secondary metabolite production. Out of many techniques that are being explored to enhance secondary metabolite production, "plant cell transfection" is the latest tool to achieve maximum output from the plant source. It is based upon the introduction of foreign DNA into the plant cell relying on physical treatment such as electroporation, cell squeezing, sonoporation, optical transfection nanoparticles, magnetofection, and chemical treatment or biological treatment that depends upon carrier. One of the promising tools that have been exploited is CRISPR-Cas9. Overall, the abovementioned tools focus on the stable transfection of desired gene transcripts. Since the integration and continuous expression of transfected gene of particular trait represents stable transfection of host cell genome, resulting from transfer of required trait to daughter cells ultimately leading to enhanced production of secondary metabolites of interest. This chapter will review a set of biotechnological tools that are candidates for achieving the enhanced bioactive compound production indicated here to be used for drug discovery.

Microbial dysbiosis and epigenetics modulation in cancer development - A chemopreventive approach.

An overwhelming number of research articles have reported a strong relationship of the microbiome with cancer. Microbes have been observed more commonly in the body fluids like urine, stool, mucus of people with cancer compared to the healthy controls. The microbiota is responsible for both progression and suppression activities of various diseases. Thus, to maintain healthy human physiology, host and microbiota relationship should be in a balanced state. Any disturbance in this equilibrium, referred as microbiome dysbiosis becomes a prime cause for the human body to become more prone to immunodeficiency and cancer. It is well established that some of these microbes are the causative agents, whereas others may encourage the formation of tumours, but very little is known about how these microbial communications causing change at gene and epigenome level and trigger as well as encourage the tumour growth. Various studies have reported that microbes in the gut influence DNA methylation, DNA repair and DNA damage. The genes and pathways that are altered by gut microbes are also associated with cancer advancement, predominantly those implicated in cell growth and cell signalling pathways. This study exhaustively reviews the current research advancements in understanding of dysbiosis linked with colon, lung, ovarian, breast cancers and insights into the potential molecular targets of the microbiome promoting carcinogenesis, the epigenetic alterations of various potential targets by altered microbiota, as well as the role of various chemopreventive agents for timely prevention and customized treatment against various types of cancers.

Secretomic Insights into the Pathophysiology of Venturia inaequalis: The Causative Agent of Scab, a Devastating Apple Tree Disease.

Apple scab, caused by Venturia inaequalis, is one of the world's most commercially significant apple diseases. The fungi have a catastrophic impact on apples, causing considerable losses in fruit quality and productivity in many apple-growing locations despite numerous control agents. Fungi secrete various effectors and other virulence-associated proteins that suppress or alter the host's immune system, and several such proteins were discovered in this work. Using state-of-the-art bioinformatics techniques, we examined the V. inaequalis reference genome (EU-B04), resulting in the identification of 647 secreted proteins, of which 328 were classified as small secreted proteins (SSPs), with 76.52% of SSPs identified as anticipated effector proteins. The more prevalent CAZyme proteins were the enzymes engaged in plant cell wall disintegration (targeting pectin and xylanase), adhesion and penetration (Cutinases/acetyl xylan esterase), and reactive oxygen species formation (multicopper oxidases). Furthermore, members of the S9 prolyl oligopeptidase family were identified as the most abundant host defense peptidases. Several known effector proteins were discovered to be expressed during the V. inaequalis infection process on apple leaves. The present study provides valuable data that can be used to develop new strategies for controlling apple scab.

Endophytic Fungi-Mediated Biocatalysis and Biotransformations Paving the Way Toward Green Chemistry.

Catalysis is a process carried out in the presence of a heterogenous catalyst for accelerating the rate of a chemical reaction. It plays a pivotal role in transition from take, make, and dispose technology to sustainable technology via chemo- and biocatalytic processes. However, chemocatalyzed reactions are usually associated with copious amounts of perilous/hazardous environmental footprints. Therefore, whole-cell biotransformations or enzyme cocktails serve as cleaner biocatalytic alternatives in replacing the classical chemical procedures. These benchmark bioconversion reactions serve as important key technology in achieving the goals of green chemistry by eliminating waste generation at source. For this, nature has always been a driving force in fuelling natural product discovery and related applications. The fungal endophytic community, in particular, has undergone co-evolution with their host plant and has emerged as a powerful tool of genetic diversity. They can serve as a treasure trove of biocatalysts, catalyzing organic transformations of a wide range of substances into enantiopure compounds with biotechnological relevance. Additionally, the biocatalytic potential of endophytic fungi as whole-intact organisms/isolated enzyme systems has been greatly expanded beyond the existing boundaries with the advancement in high-throughput screening, molecular biology techniques, metabolic engineering, and protein engineering. Therefore, the present review illustrates the promising applications of endophytic fungi as biocatalysts for the synthesis of new structural analogs and pharmaceutical intermediates and refinement of existing proteins for novel chemistries.

Engineering Host Microbiome for Crop Improvement and Sustainable Agriculture.

Dynamic consortium of microbial communities (bacteria, fungi, protists, viruses, and nematodes) colonizing multiple tissue types and coevolving conclusively with the host plant is designated as a plant microbiome. The interplay between plant and its microbial mutualists supports several agronomic functions, establishing its crucial role in plant beneficial activities. Deeper functional and mechanistic understanding of plant-microbial ecosystems will render many "ecosystem services" by emulating symbiotic interactions between plants, soil, and microbes for enhanced productivity and sustainability. Therefore, microbiome engineering represents an emerging biotechnological tool to directly add, remove, or modify properties of microbial communities for higher specificity and efficacy. The main goal of microbiome engineering is enhancement of plant functions such as biotic/abiotic stresses, plant fitness and productivities, etc. Various ecological-, biochemical-, and molecular-based approaches have come up as a new paradigm for disentangling many microbiome-based agromanagement hurdles. Furthermore, multidisciplinary approaches provide a predictive framework in achieving a reliable and sustainably engineered plant-microbiome for stress physiology, nutrient recycling, and high-yielding disease-resistant genotypes.

Development of a system for efficient callus production, somatic embryogenesis and gene editing using CRISPR/Cas9 in Saffron (Crocus sativus L.).

BACKGROUND: Crocus sativus is a recalcitrant plant for genetic transformation and genetic improvement, largely due to difficulties in Agrobacterium mediated transformation and vegetative reproduction. Effective genome editing requires proficient callus production and an efficient method to deliver Cas9 and sgRNAs into the plant. Here, we demonstrate Agrobacterium-mediated transformation of saffron. Further, we developed a CRISPR-Cas9 based system in this plant, for efficient gene knockout or edits in future. RESULTS: Efficient callus production and regeneration confers important benefits in developing competent transformation system in plants. More than 70% multiplication rate of callus initiation was achieved from corm slices of saffron subjected to a two-step sterilization procedure and grown on complete MS medium supplemented with 2,4-D (0.5 mg/L), BAP (1 mg/L), IAA (1 mg/L), photoperiod of 16/8 h and 45% relative humidity at 20 ± 2 °C. In vitro cormlet generation was accomplished in 8 weeks by using mature somatic embryos on MS medium supplemented with TDZ (0.5 mg/L) + IAA (1 mg/L) + Activated charcoal (0.1 g/L) at 15 ± 2 °C. The attempt of using Agrobacterium-mediated transformation resulted in successful integration of the binary vector into the somatic embryos of saffron with a transformation efficiency of 4%. PCR and Southern blot analysis confirmed the integration of Cas9 into saffron. CONCLUSION: The protocol for callus production, somatic embryogenesis and regeneration was standardised. Successful demonstration of integrated Cas9 in this study constitutes first step in developing strategies for genetic manipulation of saffron, which has so far been considered recalcitrant. Furthering the development of this technology holds significant potential for advancing genetic research in saffron by integrating multigene targeting and/or use of recyclable cassettes.

Tissue-specific transcriptional regulation and metabolite accumulation in tomato (Solanum lycopersicum L.).

Tomato is an excellent model for studying fruit development, ripening, and other secondary metabolic pathways such as carotenoid biosynthetic pathway, flavonoid pathway, and many more. Tomato fruit development and ripening occurs under tight genetic control and involves the expression of thousands of genes affecting fruit quality and accumulation of pigments and metabolites. Here, we have described the development of a microarray platform that has allowed establishment of a framework for quantification of the expression of large number of genes and transcription factors possibly regulating various secondary metabolic pathways in tomato. To unravel the molecular mechanisms of fruit development and ripening, a tomato 60-mer oligonucleotide 44 K microarray along with the custom array for many genes and transcription factors was designed and validated in the fruit and leaf tissues. Comparative profiling of gene expression studies has allowed us to identify a large number of differentially expressed genes and transcription factors. Gene ontology revealed the involvement of these genes in various biological, cellular, and molecular processes like isoprenoid, terpenoid, pigment, ethylene biosynthesis, phytohormone signaling, and fruit ripening. Further, correlation, as well as differential expression studies, has revealed that several transcription factors like RIN, AGAMOUS, TAGL1, MYB, MADS-box etc. could be the possible regulators of various secondary metabolic pathways. The present study has identified various metabolites, their biosynthetic pathways and genes which may possibly be controlled by different transcription factors. The present findings have laid a base for understanding the transcriptional and metabolic shifts which occur in parallel during programmed fruit ripening and developmental processes in tomato.

Transcriptome analysis of Snow Mountain Garlic for unraveling the organosulfur metabolic pathway.

Snow Mountain Garlic grows in the high altitudes of the Himalayas under low temperature conditions. It contains various bioactive compounds whose metabolic pathways have not been worked out at genomic level. The present work is the first report on the transcriptome sequencing of this plant. >43 million paired-end reads (301 × 2) were generated using Illumina Miseq sequencing technology. Assembling of the sequencing data resulted in 326,785 transcripts. Differentially expressed genes between the clove and leaf tissues were identified and characterized. Besides, greater emphasis was laid on the genes, which were highly expressed in clove since the latter is assumed to contain high content of the bioactive compounds. Further analysis led to the identification of the genes plausibly involved in the organosulfur metabolism. We also identified several simple sequence repeats and single nucleotide polymorphism. These constitute valuable genetic resource for research and further genetic improvement of the plant.

Plant carotenoid cleavage oxygenases: structure-function relationships and role in development and metabolism.

A plant communicates within itself and with the outside world by deploying an array of agents that include several attractants by virtue of their color and smell. In this category, the contribution of 'carotenoids and apocarotenoids' is very significant. Apocarotenoids, the carotenoid-derived compounds, show wide representation among organisms. Their biosynthesis occurs by oxidative cleavage of carotenoids, a high-value reaction, mediated by carotenoid cleavage oxygenases or carotenoid cleavage dioxygenases (CCDs)-a family of non-heme iron enzymes. Structurally, this protein family displays wide diversity but is limited in its distribution among plants. Functionally, this protein family has been recognized to offer a role in phytohormones, volatiles and signal production. Further, their wide presence and clade-specific functional disparity demands a comprehensive account. This review focuses on the critical assessment of CCDs of higher plants, describing recent progress in their functional aspects and regulatory mechanisms, domain architecture, classification and localization. The work also highlights the relevant discussion for further exploration of this multi-prospective protein family for the betterment of its functional understanding and improvement of crops.

Carbohydrate Modifications of Neoandrographolide for Improved Reactive Oxygen Species-Mediated Apoptosis through Mitochondrial Pathway in Colon Cancer.

Modifications at the carbohydrate moiety of neoandrographolide, isolated from the medicinal plant Andrographis paniculata, result in more potent and less toxic derivatives, namely, 4',6'-benzylidene neoandrographolide (2b) and 4'6'-p-methoxybenzylidene neoandrographolide (2c). These showed improved cytotoxicity against SW-620, PC-3, and A549 cancer cell lines. Nuclear morphology studies were conducted on compound 2b by 4',6-diamidino-2-phenylidole staining and detection of intracellular reactive oxygen species (ROS) accumulation. It showed an increase in the generation of cellular and mitochondrial ROS level. The probable relation of B-cell lymphoma-2 (Bcl-2, an apoptosis inhibitor) to B-cell lymphoma-2-associated X protein (Bax, an apoptosis promoter) ratio with caspase-3 (apoptosis coordination enzyme) in the colon cancer cell line SW-620 was investigated, and it was discovered that upon 2b treatment, the expression of caspase-3 Bax increased remarkably. However, in 2b-treated cells, the expression of Bcl-2 was downregulated as compared to untreated cells.

Identification and in silico characterization of cis-acting elements of genes involved in carotenoid biosynthesis in tomato.

Carotenoids, the widespread and structurally diverse class of pigments, accumulate in the fruits of tomato plants in a tissue specific manner. The carotenoid biosynthetic pathway genes have been cloned and characterized in tomato and other plants, however, its regulation is still obscure. We collected and analyzed forty different accessions of tomato for the present study. HPLC analysis revealed differential accumulation of major carotenoids (lycopene and ß-carotene) in the ripe fruit tissue. In order to understand the underlying regulatory mechanisms in carotenoid biosynthesis and accumulation, we sequenced the cis-acting elements i.e. promoter, 5' and 3' untranslated regions of the carotenoid pathway genes, in all accessions, followed by their in silico validation. Major differences observed in the CAAT Box, Opaque-2 Box and L-box in the promoters of carotenoid isomerase and lycopene-beta cyclase genes, respectively, along with the variations in musashi binding element of 5' untranslated regions of the carotenoid isomerase gene, suggest their differential role in regulating the carotenogenesis process in tomato. The binding sites for various transcription factors namely RIN, AGAMOUS, CRY, RAP2.2 and PIF1 on the promoters of important carotenoid pathway genes were predicted in silico. We propose that expression of carotenoid genes and also the formation of protein product in ripe tomato fruits, is regulated efficiently by the binding of these transcription factors at selected sites in the promoter region. Finally, the differential expression of the above-mentioned genes in different developmental tissues supports the possible involvement of promoters and untranslated regions in carotenoid biosynthesis and accumulation process. The present study has generated significant information concerning regulatory players involved in the carotenoid biosynthesis in tomato.

Identification and expression profiling of miRNAs in two color variants of carrot (Daucus carota L.) using deep sequencing.

microRNAs represent small endogenous RNAs which are known to play a crucial role in various plant metabolic processes. Carrot being an important vegetable crop, represents one of the richest sources of carotenoids and anthocyanins. Most of the studies on microRNAs have been conducted in the aerial parts of the plants. However, carrot has the rare distinction of storing these compounds in roots. Therefore, carrot represents a good model system to unveil the regulatory roles of miRNAs in the underground edible part of the plant. For the first time, we report the genome wide identification and expression profiling of miRNAs in two contrasting color variants of carrot namely Orange Red and Purple Black using RNA-seq. Illumina sequencing resulted in the generation of 25.5M and 18.9M reads in Orange Red and Purple Black libraries, respectively. In total, 144 and 98 (read count >10), conserved microRNAs and 36 and 66 novel microRNAs were identified in Orange Red and Purple Black, respectively. Functional categorization and differential gene expression revealed the presence of several miRNA genes targeting various secondary metabolic pathways including carotenoid and anthocyanin biosynthetic pathways in the two libraries. 11 known and 2 novel microRNAs were further validated using Stem-Loop PCR and qRT-PCR. Also, target validation was performed for selected miRNA genes using RLM-RACE approach. The present work has laid a foundation towards understanding of various metabolic processes, particularly the color development in carrot. This information can be further employed in targeted gene expression for increasing the carotenoid and anthocyanin content in crop plants.

Origin, Behaviour, and Transmission of B Chromosome with Special Reference to Plantago lagopus.

B chromosomes have been reported in many eukaryotic organisms. These chromosomes occur in addition to the standard complement of a species. Bs do not pair with any of the A chromosomes and they have generally been considered to be non-essential and genetically inert. However, due to tremendous advancements in the technologies, the molecular composition of B chromosomes has been determined. The sequencing data has revealed that B chromosomes have originated from A chromosomes and they are rich in repetitive elements. In our laboratory, a novel B chromosome was discovered in Plantago lagopus. Using molecular cytogenetic techniques, the B chromosome was found to be composed of ribosomal DNA sequences. However, further characterization of the chromosome using next generation sequencing (NGS) etc. revealed that the B chromosome is a mosaic of sequences derived from A chromosomes, 5S ribosomal DNA (rDNA), 45S rDNA, and various types of repetitive elements. The transmission of B chromosome through the female sex track did not follow the Mendelian principles. The chromosome was found to have drive due to which it was perpetuating in populations. The present paper attempts to summarize the information on nature, transmission, and origin of B chromosomes, particularly the current status of our knowledge in P. lagopus.

Transcript profiling of carotenoid/apocarotenoid biosynthesis genes during corm development of saffron (Crocus sativus L.).

The dried stigmas of saffron constitute the world's costliest spice. Saffron has many therapeutic applications due to the presence of apocarotenoids. The latter are synthesized at different stages of development, and the biosynthetic pathway involves several genes encoding different enzymes. In order to understand the differential expression of various genes of the pathway, eight distinct developmental stages (S1-early to S8-late) were identified. The corms were assorted into three groups (I, II, and III) based on corm weight. Expression profiles of 12 carotenoid/apocarotenoid genes were studied. The expression of all genes was minimum/least in groups I and II corms during bud development. Lowest expression of carotenogenic genes (CsPSY, CsPDS, CsZDS, CsCRTISO, CsLYC-ß1, CsLYC-ε, CsBCH2, and CsNCED) was observed during early stages (S1-S3) of corm growth (dormant period). In group III corms, increased expression of apocarotenoid genes (CsZCO, CsCCD2, CsUGT, and CsALDH) was observed during S4 to S8 stages (reproductive period, floral differentiation). Besides, expression profiles of genes in apical and axillary buds were also examined. Of all the genes studied, apocarotenoid biosynthesis genes (CsBCH2, CsZCO, CsCCD2, CsALDH, and CsUGT) were found to be upregulated in apical bud than in the axillary bud. The results indicated that interaction of phytohormones and sugars, mother corm reserves and the influence of internal and external factors may be contributing to the growth of saffron corm/bud. The study has laid a foundation for further research on the molecular mechanisms underlying bud dormancy/growth in saffron.

Identification and characterization of some putative genes involved in arabinoxylan biosynthesis in Plantago ovata.

Plantago ovata is an important source of Psyllium (Isabgol), which swells upon contact with water forming mucilaginous mass, largely composed of arabinoxylans. In this study, we analyzed the expression pattern of arabinoxylan biosynthetic pathway genes at different stages of seed development in P. ovata. Besides, arabinoxylans were quantified at different stages of seed development in water extractable and water unextractable fractions. The expression analysis revealed 5-8 fold increase in the levels of expression of some genes involved in arabinoxylan biosynthetic pathway such as UDP-arabinopyranose mutase, UDP-xylosyltransferase 2 and xylan glucuronosyltransferase at 15 days after pollination stage in seed. The xylose and arabinose units were analyzed at different stages of seed development and also in water-soluble (cold water and hot water), alkali and ethanolic fractions. The concentration of xylose and arabinose units increased steadily after pollination. Overall, alkali extract had high concentration of xylose (0.70 ± 0.022 mg/g) and arabinose units (0.10 ± 0.01 mg/g) at 15 days after pollination stage.

Elucidation and functional characterization of CsPSY and CsUGT promoters in Crocus sativus L.

The dried stigmas of Crocus sativus constitute the saffron, which is considered to be the costliest spice of the world. Saffron is valuable for its constituents, which are mainly apocarotenoids. In order to enhance the production of apocarotenoids, it is imperative to understand the regulation of apocarotenoid biosynthetic pathway. In C. sativus, although the pathway has been elucidated, the information regarding the regulation of the pathwaygenes is scanty. During the present investigation, the characterization of promoters regulating the expression of two important genes i.e. CsPSY and CsUGT was performed. We successfully cloned the promoters of both the genes, which were functionally characterized in Crocus sativus and Nicotiana tabaccum. In silico analysis of the promoters demonstrated the presence of several important cis regulatory elements responding tolight, hormonesand interaction with transcription factors (TFs). Further analysis suggested the regulation of CsPSY promoter by Abscisic acid (ABA) and that of CsUGT by Gibberellic acid (GA). In addition, we also observed ABA and GA mediated modulation in the expression of significant TFs and CsPSY and CsUGT transcripts. Overall, the study addresses issues related to regulation of key genes of apocarotenoid pathway in C.sativus.

Detection of some new Trichosporon species from the dystrophied nails of three female members of a family from North Indian State of Jammu and Kashmir.

Dermatophytes are considered as the main pathogens responsible for onychomycosis, but recently successive isolations of yeast-like fungi from the infected nails has led to consider these also as primary agents of nail infections. Trichosporon species which are non-candidal, basidiomycetous, yeast-like, anamorphic fungi are commonly isolated from soil but they are also emerging as important etiological agents of onychomycosis. Three species of Trichosporon viz., T. asahii, T. asteroides and T. faecale were isolated from the infected nails of three female members of a family from district Doda of Jammu and Kashmir State. Among the isolated species of Trichosporon, T. asahii was recovered from the nail samples of all the three members, thus confirming its recognition as a main pathogenic species of onychomycosis. So far, there is no report of T. asteroides and T. faecale causing onychomycosis and hence they constitute new additions to the list of onychomycotic fungi. Some of the predisposing factors like low socio-economic condition, poor hygiene, frequent exposure of finger nails to water and dirt, climatic conditions and nail trauma were observed to be the main causes of nail infection in these patients. However, a link between the pathogenic genus and the genetic makeup of the patients is also probable.