S. Senthilkumar

Bio: S. Senthilkumar is an academic researcher. The author has contributed to research in topics: Fluorescence microscope & Aerides. The author has an hindex of 2, co-authored 2 publications receiving 22 citations.

Visualization of orchid mycorrhizal fungal structures with fluorescence dye using epifluorescence microscopy.

Abstract: The mycorrhizal association is a wellknown mutual relationship between two organisms, a fungus and the root of a vascular plant. A number of mycorrhizal types have been recognized based on the fungal group involved and by the morphological features, resulting in the complex interaction between the symbionts. Various microscopic methods have been used to characterize morphological, anatomical and molecular features of orchid mycorrhizae. Fluorescence microscopy allows optical sectioning of images obtained from different reconstructions which are useful for studying complex fungal structures within the host. Recently, Laser Scanning Confocal Microscopy has stimulated research in the development of new fluorochromes and dyes used routinely for light microscopy. Many dyes have been used routinely for light microscopy. We report here the use of acridine orange for fluorescence microscopy, which is an effective method for a rapid screening of cellular details to study the complex fungal structures of orchid mycorrhizae. Spathoglottis plicata Blume, an ornamental orchid commonly cultivated in several parts of the world, was selected for this study. Its underground bulbs produce several roots that have mycorrhizal association. The plants were maintained in pots with natural inoculum of the mycorrhizal fungus Eupholorhiza repens. The mycorrhizal roots were collected, washed thoroughly in water, cut into small pieces, fixed in formalin–acetic acid–alcohol, dehydrated, embedded in paraffin and sectioned by microtome. Slides with sections were heated gently on a water bath and stained with 0.05% acridine orange (Sigma Chemical Co) for 10 min. Slides were then rinsed with distilled water and allowed to dry. Stained tissues were viewed and photographed directly using Nikon microscope with a filter B-2A excitation and emission at 450 nm In addition, fresh hand sections were also used. Infection and peloton formation were noticed in the cortical cells and it was more in the outer cortical cells. Free hand section also showed a peloton of the mycorrhizal fungus appeared as spherical balls (Figures 1 b, c and 2 c). Individual hyphae forming the peloton could be easily differentiated (Figure 1) from completely filled up cells where recognition of individual hyphae was gradually lost, indicating its slow lysis and digestion (Figures 1 b and 2 c) which supports our earlier findings. Entry of the fungus into the cortex is always through the passage cells of the exodermis and no instance of the presence of fungal mycelium in the thick-walled cells of the exodermis was noticed (Figure 2 a, c). In fact, the exodermal passage cells do have a control on the entry of fungus into the cortex, since they are the only living cells of the exodermis whose cell walls do not have lignin and suberin to the extent that the other exodermal cells have. Like the rhizodermal and hypodermal cells, even in passage cells of the exodermis the fungal hypae do not settle down to form coils, indicating that these cells are meant only as a channel for hyphal entry into the cortex and do not have the appropriate physiological conditions necessary for coil formation and hyphal degradation, although they are more active metabolically. Entry into passage cells is through the outer tangential walls of these cells. From these cells, the hypha spreads into the cortex intracellularly, branches repeatedly and some of these branches get into the cortical parenchyma cells, forming pelotons. More often, the SCIENTIFIC CORRESP

Recent developments in the study of orchid mycorrhiza

Abstract: Orchids are mycoheterotrophic during their seedling stage and in many species the dependency on fungi as a carbohydrate source is prolonged into adulthood. The mycobionts in orchid mycorrhiza belong in at least 5 major taxonomic groups of basidiomycetes. Traditional records have mainly focused on saprotrophic mycobionts but the participation of both ectomycorrhizal and parasitic fungi in orchid mycorrhiza has been corroborated. There is an increasing evidence of specific relationships between orchids and fungi, though usually not on a species-to-species level. Physiological compatibility demonstrated under artificial conditions, as in vitro, may be much broader, however. Recent development of field sowing techniques has improved the possibilities of evaluating orchid-fungal relations in an ecological context. Although the general nutrient flow in orchid mycorrhiza is well known, some questions remain regarding breakdown processes of fungi within orchid tissues, especially the ptyophagic syndrome that has recently been illustrated at the ultrastructural level for the first time.

Comparative anatomy of the absorption roots of terrestrial and epiphytic orchids

Abstract: The present study compared roots of terrestrial and epiphytic Orchidaceae, analyzing the anatomical characteristics from an ecological point of view. The material was collected at three different sites in Minas Gerais / Brazil and was fixed in FAA. Transverse sections were obtained by freehand sections or from material previously embedded in Paraplast® or Historesin®. The prominent characteristics of the epiphytic group were: significant smaller perimeter, epidermis with 3 or more cell layers, U-thickened exodermal cell walls, O-thickened endodermal cell walls, and a low ratio between the caliber and the number of protoxylem arches. The terrestrial group presented simple or multiseriate epidermis, and exodermis and endodermis with typical Casparian strips. The anatomical characteristics should have evolved with several adaptations to distinct environments during evolutionary process.

Growth promotion-related miRNAs in Oncidium orchid roots colonized by the endophytic fungus Piriformospora indica.

Abstract: Piriformospora indica, an endophytic fungus of Sebacinales, colonizes the roots of a wide range of host plants and establishes various benefits for the plants. In this work, we describe miRNAs which are upregulated in Oncidium orchid roots after colonization by the fungus. Growth promotion and vigorous root development were observed in Oncidium hybrid orchid, while seedlings were colonized by P. indica. We performed a genome-wide expression profiling of small RNAs in Oncidium orchid roots either colonized or not-colonized by P. indica. After sequencing, 24,570,250 and 24744,141 clean reads were obtained from two libraries. 13,736 from 17,036,953 unique sequences showed homology to either 86 miRNA families described in 41 plant species, or to 46 potential novel miRNAs, or to 51 corresponding miRNA precursors. The predicted target genes of these miRNAs are mainly involved in auxin signal perception and transduction, transcription, development and plant defense. The expression analysis of miRNAs and target genes demonstrated the regulatory functions they may participate in. This study revealed that growth stimulation of the Oncidium orchid after colonization by P. indica includes an intricate network of miRNAs and their targets. The symbiotic function of P. indica on Oncidium orchid resembles previous findings on Chinese cabbage. This is the first study on growth regulation and development of Oncidium orchid by miRNAs induced by the symbiotic fungus P. indica.

Mycorrhizal association and morphology in orchids

Abstract: We investigated the mycorrhizal associations in 31 adult wild or cultivated green orchids (22 epiphytic, 8 terrestrial, and 1 species with both epiphytic and lithophytic life-forms) from different vegetation types of Western Ghats, southern India. All the orchids examined were mycorrhizal with the extent of colonization varying with species and life-forms. Mycorrhizal association has been reported for the first time in 25 orchids. The entry of mycorrhizal fungi into the roots was mostly through root hairs. In certain epiphytic species, the fungal entry was directly through the epidermis. The fungi formed highly coiled hyphal structures (pelotons) within the root cortex, and their size was related to the cell size. The fungal invasion of the cortical cells was through cell-to-cell penetration. The cortical cells contained intact and lysed pelotons, and their ratio varied with species and life-forms. No significant relationship existed between root hair characteristics and the extent of colonization. Chlamy...

Structural adaptations of two sympatric epiphytic orchids (Orchidaceae) to a cloudy forest environment in rocky outcrops of Southeast Brazil

Abstract: Adaptaciones estructurales de dos orquideas epifitas simpatricas (Orchidaceae) en un bosque nuboso de un campo rupestre del sureste de Brasil. The survival of plants in epiphytic environments depends on vegetative adaptations capable to defraud different stresses. Based on the structural diversity of the Orchidaceae, the current study has the objective of relating the anatomical structure of Dichaea cogniauxi- ana and Epidendrum secundum with the distinct environments where they live. It was expected that, despite structural similarities as strategies for resource acquisition, some peculiar variations related to the distinct light microenvironments (inside or in the edge of the nebular forest, near to "campo rupestre" area) might be found. Leaves and roots of both species were collected in a nebular forest located at a "campo rupestre" area at Serra da Piedade, Brazil), in January and February 2005. D. cogniauxiana is adhered to trunks, in sites with high atmo- spheric humidity and shaded, while E. secundum is located at the edge of the nebular forest, in more luminous sites. The leaves of E. secundum had thicker cuticle and higher number of stomata per area than those of D. cogniauxiana, characteristics coherent with their distinct pattern of exhibition to sun light. The suprastomatic chambers formed by the thicker cuticle may function as a barrier of resistance to water evaporation. The suc- culence of the leaves of E. secundum propitiates organic acids storage at night, and the storage of starch may be involved in PEP-carboxylase metabolism, both propitiating CAM mechanism. Roots with larger number of cell layers of the velamen, and specialized thick walled cortical cells (both in E. secundum) help water absorption and indicate better adaptation to an environment with intense solar radiation and a probable higher water deficit. The remarkable cell wall thickening of E. secundum exodermis can confer more efficient protection against the excess of transpiration at the border of the nebular forest. On the other hand, besides D. cogniauxiana be epiphyte, it is in a low position - in a shaded environment and with high relative humidity. Its thin thickened velamen permits the entrance of the low available light, and photosynthesis, producing oxygen and helping to avoid hypoxia condition. As features registered for D. cogniauxiana and E. secundum roots, we can depict the velamen, distinct exodermis and endodermis, and specialized thick walled cortical cells as characteristic of epiphytic plants. Rev. Biol. Trop. 61 (3): 1053-1065. Epub 2013 September 01.