Mohapatra S.,Indian Institute of Horticultural Research
Journal of environmental science and health. Part. B, Pesticides, food contaminants, and agricultural wastes | Year: 2011
Flubendiamide is a new insecticide that has been found to give excellent control of lepidopterous pests of tomato. This study has been undertaken to develop an improved method for analysis of flubendiamide and its metabolite des-iodo flubendiamide and determine residue retention in tomato and soil. The analytical method developed involved extraction of flubendiamide and its metabolite des-iodo flubendiamide with acetonitrile, liquid-liquid partitioning into hexane-ethyl acetate mixture (6:4, v v 1) and cleanup with activated neutral alumina. Finally the residues were dissolved in gradient high pressure liquid chromatography (HPLC) grade acetonitrile for analysis by HPLC. The mobile phase, acetonitrile-water at 60:40 (v v 1) proportion and the wavelength of 235 nm gave maximum peak resolution. Using the above method and HPLC parameters described, nearly 100 % recovery of both insecticides were obtained. There was no matrix interference and the limit of quantification (LOQ) of the method was 0.01 mg kg 1. Initial residue deposits of flubendiamide on field-treated tomato from treatments @ 48 and 96 g active ingredient hectare 1 were 0.83 and 1.68 mg kg 1, respectively. The residues of flubendiamide dissipated at the half-life of 3.9 and 4.4 days from treatments @ 48 and 96 g a.i. ha 1, respectively and persisted for 15 days from both the treatments. Des-iodo flubendiamide was not detected in tomato fruits at any time during the study period. Residues of flubendiamide and des-iodo flubendiamide in soil from treatment @ 48 and 96 g a.i. ha 1 were below detectable level (BDL, < 0.01 mg kg 1) after 20 days. Flubendiamide completely dissipated from tomato within 20 days when the 480 SC formulation was applied at doses recommended for protection against lepidopterous pests.
Thomas P.,Indian Institute of Horticultural Research
Plant Cell Reports | Year: 2011
The study was undertaken with a view to unravel the source of bacterial colony growth observed in a section of micropropagated triploid watermelon cultures that were supposedly cleansed of the associated endophytic bacteria through antibiotic treatment, and thereafter maintained under stringent sterility checks to prevent lateral intrusion of contaminants. Five different bacteria were retrieved from colony growth-displaying watermelon cultures that were previously treated with gentamycin and five isolates from cefazolin-treated stocks with the organisms showing tolerance to the respective antibiotic. These watermelon cultures were in degeneration phase (over 6 months after the previous sub-culturing), while the actively maintained counterpart stocks appeared healthy with no colony growth on different bacteriological media during tissue-screenings. The latter cultures, however, revealed abundant motile, tetrazolium-stained bacterial cells in microscopy, suggesting tissue colonization by non-culturable endophytes. PCR screening on healthy cultures endorsed tissue colonization by different bacterial phylogenic groups. A few organisms could be activated to cultivation from healthy watermelon stocks through host tissue extract supplementation, which also enhanced the growth of all the organisms. The study indicated that a fraction of antibiotic-tolerant bacteria survived intra-tissue in non-culturable form during the preceding cleansing activity, multiplied to substantial numbers thereafter, and turned cultivable in degenerating cultures contributed by tissue breakdown products. This study brings out the existence of a deep endophyte association in tissue cultures which is not easily dissociable. It also signifies the utility of in vitro system for investigations into plant-endophyte association and to bring normally non-culturable novel organisms to cultivation facilitating their future exploitation. © 2011 Springer-Verlag.
Nidiry E.S.J.,Indian Institute of Horticultural Research
Magnetic Resonance in Chemistry | Year: 2012
Octadecyl p-coumarates undergo E-Z isomerization in daylight. Although 1H NMR, 13C NMR and 1H-1H COSY gave indications about this isomerization, the overlapping of some signals in the 1H NMR of aromatic region prevented the delineation of signals of the individual isomers. However, heteronuclear spin quantum coupling correlation (HSQC) with the unique feature of two sets of nearby δC- δH correlations gave conclusive evidence for this isomerization and helped in the delineation of 1H NMR and 13C NMR signals of E-octadecyl p-coumarate and Z-octadecyl p-coumarate. Copyright © 2012 John Wiley & Sons, Ltd.
Thomas P.,Indian Institute of Horticultural Research |
Sekhar A.C.,Indian Institute of Horticultural Research
AoB PLANTS | Year: 2014
It is generally believed that endophytic microorganisms are intercellular inhabitants present in either cultivable or non-cultivable form primarily as root colonizers. The objective of this study was to determine whether the actively mobile micro-particles observed in the intracellular matrix of fresh tissue sections of banana included endophytic bacteria. Tissue sections (50-100 mm) from apical leaf sheaths of surface-disinfected suckers (cv. Grand Naine) displayed 'Brownian motion'-reminiscent abundant motile micro-particles under bright-field and phase-contrast (×1000), which appeared similar in size and motility to the pure cultures of endophytes previously isolated from banana. Observations on callus, embryonic cells and protoplasts with intact cell wall/plasma membrane confirmed their cytoplasmic nature. The motility of these entities reduced or ceased upon tissue fixation or staining with safranin/crystal violet (0.5 % w/v), but continued uninterrupted following treatment with actin-disrupting drugs, ruling out the possibility of micro-organelles like peroxisomes. Staining with 2,3,5-triphenyl tetrazolium chloride (TTC) confirmed them to be live bacteria with similar observations after dilute safranin (0.005 %) treatment. Tissue staining with SYTO-9 coupled with epi-fluorescence or confocal laser scanning microscopy showed bacterial colonization along the peri-space between cell wall and plasma membrane initially. SYTO-9 counterstaining on TTC-or safranin-treated tissue and those subjected to enzymatic permeabilization revealed the cytoplasmic bacteria. These included organisms moving freely in the cytoplasm and those adhering to the nuclear envelope or vacuoles and the intravacuolar colonizers. The observations appeared ubiquitous to different genomes and genotypes of banana. Plating the tissue homogenate on nutrient media seldom yielded colony growth. This study, supported largely by live cell video-imaging, demonstrates enormous intracellular colonization in bananas by normally non-cultivable endophytic bacteria in two niches, namely cytoplasmic and periplasmic, designated as 'Cytobacts' and 'Peribacts', respectively. The integral intracellular association with their clonal perpetuation suggests a mutualistic relationship between endophytes and the host. © 2014 The Authors.
Swarupa V.,Indian Institute of Horticultural Research |
Ravishankar K.V.,Indian Institute of Horticultural Research |
Rekha A.,Indian Institute of Horticultural Research
Planta | Year: 2014
Soil-borne fungal pathogen, Fusarium oxysporum causes major economic losses by inducing necrosis and wilting symptoms in many crop plants. Management of fusarium wilt is achieved mainly by the use of chemical fungicides which affect the soil health and their efficiency is often limited by pathogenic variability. Hence understanding the nature of interaction between pathogen and host may help to select and improve better cultivars. Current research evidences highlight the role of oxidative burst and antioxidant enzymes indicating that ROS act as an important signaling molecule in banana defense response against Fusarium oxysporum f.sp. cubense. The role of jasmonic acid signaling in plant defense against necrotrophic pathogens is well recognized. But recent studies show that the role of salicylic acid is complex and ambiguous against necrotrophic pathogens like Fusarium oxysporum, leading to many intriguing questions about its relationship between other signaling compounds. In case of banana, a major challenge is to identify specific receptors for effector proteins like SIX proteins and also the components of various signal transduction pathways. Significant progress has been made to uncover the role of defense genes but is limited to only model plants such as Arabidopsis and tomato. Keeping this in view, we review the host response, pathogen diversity, current understanding of biochemical and molecular changes that occur during host and pathogen interaction. Developing resistant cultivars through mutation, breeding, transgenic and cisgenic approaches have been discussed. This would help us to understand host defenses against Fusarium oxysporum and to formulate strategies to develop tolerant cultivars. © 2014 Springer-Verlag Berlin Heidelberg.
Thomas P.,Indian Institute of Horticultural Research |
Reddy K.M.,Indian Institute of Horticultural Research
AoB PLANTS | Year: 2013
This study was aimed at generating microscopic evidence of intra-tissue colonization in banana in support of the previous findings on widespread association of endophytic bacteria with the shoot tips of fieldgrown plants and micropropagated cultures, and to understand the extent of tissue colonization. Leaf-sheath tissue sections (50-100 mm) from aseptically gathered shoot tips of cv. Grand Naine were treated with Live/ Dead bacterial viability kit components SYTO 9 (S9) and propidium iodide (PI) followed by epifluorescence or confocal laser scanning microscopy (CLSM). The S9, which targets live bacteria, showed abundant green-fluorescing particles along the host cell periphery in CLSM, apparently in between the plasma membrane and the cell wall. These included non-motile and occasional actively motile single bacterial cells seen in different x-y planes and z-stacks over several cell layers, with the fluorescence signal similar to that of pure cultures of banana endophytes. Propidium iodide, which stains dead bacteria, did not detect any, but post-ethanol treatment, both PI and 4?,6-diamidino-2- phenylindole detected abundant bacteria. Propidium iodide showed clear nuclear staining, as did S9 to some extent, and the fluorophores appeared to detect bacteria at the exclusion of DNA-containing plant organelles as gathered from bright-field and phase-contrast microscopy. The S9-PI staining did not work satisfactorily with formalin- or paraformaldehyde-fixed tissue. The extensive bacterial colonization in fresh tissue was further confirmed with the suckers of different cultivars, and was supported by transmission electron microscopy. This study thus provides clear microscopic evidence of the extensive endophytic bacterial inhabitation in the confined cell wall-plasma membrane peri-space in shoot tissue of banana with the organisms sharing an integral association with the host. The abundant tissue colonization suggests a possible involvement of endophytes in the biology of the host besides recognizing cell wall-plasma membrane peri-space as a major niche for plant-associated bacteria. © The Authors 2013.
Mohapatra S.,Indian Institute of Horticultural Research
Journal of Environmental Science and Health - Part B Pesticides, Food Contaminants, and Agricultural Wastes | Year: 2014
The persistence and dissipation kinetics of trifloxystrobin and tebuconazole on onion were studied after application of their combination formulation at a standard and double dose of 75 + 150 and 150 + 300 g a.i. ha-1. The fungicides were extracted with acetone, cleaned-up using activated charcoal (trifloxystrobin) and neutral alumina (tebuconazole). Analysis was carried out by gas chromatograph (GC) and confirmed by gas chromatograph mass spectrometry (GC-MS). The recovery was above 80% and limit of quantification (LOQ) 0.05 mg kg-1 for both fungicides. Initial residue deposits of trifloxystrobin were 0.68 and 1.01 mg kg-1 and tebuconazole 0.673 and 1.95 mg kg-1 from standard and double dose treatments, respectively. Dissipation of the fungicides followed first-order kinetics and the half life of degradation was 6-6.6 days. Matured onion bulb (and field soil) harvested after 30 days was free from fungicide residues. These findings suggest recommended safe pre-harvest interval (PHI) of 14 and 25 days for spring onion consumption after treatment of Nativo 75 WG at the standard and double doses, respectively. Matured onion bulbs at harvest were free from fungicide residues. © 2014 Copyright © Taylor & Francis Group, LLC.
Upreti R.,Indian Institute of Horticultural Research |
Thomas P.,Indian Institute of Horticultural Research
Frontiers in Microbiology | Year: 2015
This study was undertaken to assess if the root-associated native bacterial endophytes in tomato have any bearing in governing the host resistance to the wilt pathogen Ralstonia solanacearum. Internal colonization of roots by bacterial endophytes was confirmed through confocal imaging after SYTO-9 staining. Endophytes were isolated from surface-sterilized roots of 4-week old seedlings of known wilt resistant (R) tomato cultivar Arka Abha and susceptible (S) cv. Arka Vikas on nutrient agar after plating the tissue homogenate. Arka Abha displayed more diversity with nine distinct organisms while Arka Vikas showed five species with two common organisms (Pseudomonas oleovorans and Agrobacterium tumefaciens). Screening for general indicators of biocontrol potential showed more isolates from Arka Abha positive for siderophore, HCN and antibiotic biosynthesis than from Arka Vikas. Direct challenge against the pathogen indicated strong antagonism by three Arka Abha isolates (P. oleovorans, Pantoea ananatis and Enterobacter cloacae) and moderate activity by three others, while just one isolate from Arka Vikas (P. oleovorans) showed strong antagonism. Validation for the presence of bacterial endophytes on three R cultivars (Arka Alok, Arka Ananya, Arka Samrat) showed 8-9 antagonistic bacteria in them in comparison with four species in the three S cultivars (Arka Ashish, Arka Meghali, Arka Saurabhav). Altogether 34 isolates belonging to five classes, 16 genera and 27 species with 23 of them exhibiting pathogen antagonism were isolated from the four R cultivars against 17 isolates under three classes, seven genera and 13 species from the four S cultivars with eight isolates displaying antagonistic effects. The prevalence of higher endophytic bacterial diversity and more antagonistic organisms associated with the seedling roots of resistant cultivars over susceptible genotypes suggest a possible role by the root associated endophytes in natural defense against the pathogen. © 2015 Upreti and Thomas.
Upreti K.K.,Indian Institute of Horticultural Research |
Murti G.S.R.,Indian Institute of Horticultural Research
Biologia Plantarum | Year: 2010
Effects of salinity (0, 50, 100 and 250 mM NaCl) on growth, root:shoot dry mass ratio, osmotic potential (ψπ), electrolyte leakage and contents of Na+ and K+, polyamines and abscisic acid (ABA) were studied in the grape rootstocks Dogridge, 1613, St. George and Salt Creek. In control rootstocks, the root length was highest in Dogridge and contents of K+ and ABA in Salt Creek. Salinity treatments increased root Na+ and decreased K+ content and St. George exhibited highest Na+ content and Na+:K+ ratio. The root:shoot dry mass ratio in all rootstocks increased upto 100 mM NaCl. With increasing NaCl concentration, putrescine, spermine and spermidine contents showed consistent increase and putrescine increase was highest in St. George and spermidine and spermine in the Dogridge and Salt Creek. Under salinity, the ABA content increased in all the rootstocks but more in Salt Creek and Dogridge than in St. George. © 2010 Springer Science+Business Media B.V.
Mohandas S.,Indian Institute of Horticultural Research
Scientia Horticulturae | Year: 2012
The rhizospheres of three year old-mango (Mangifera indica L.) rootstocks namely, Vellakulamban, Bappakai, Olour, Chandrakaran, Necker, Peach, Totapuri and Vellakulamban were studied for the spore load of arbuscular mycorrhizal (AM) fungi and root colonization at 15. cm, and 30. cm depths. Mycorrhizal spores were highest in Totapuri followed by Bappakai, Olour and Peach and Vellakulamban at 15. cm depth. Spores belonged to the genera Glomus and Acaulospora and few other genera, the predominant ones being Glomus fasciculatum and Glomus mosseae as identified by their morphology. The colonization of the root was higher in Vellakullamban and Totapuri rootstocks. Frequently occurring AM species were initially multiplied on finger millet (Eleucine coracona L.) in paper cups filled with soilrite and then in 12. in. pots that contained 1:1 sand soil mixture. The rootstock cultivars predominantly used for grafting mango scions in southern India were screened for their response to AM inoculation in pot culture. All the rootstock seedlings responded to mycorrhizal inoculations showed varied intensity of root colonization and improved plant height, growth and nutrient content compared to non-mycorrhizal in pot culture. Under field conditions, rootstock cv Totapuri inoculated with AM fungi and scions of mango hybrids Arka Aruna and Arka Puneeth grafted on them produced shoots earlier compared to non-mycorrhizal plants. Within two years of application of AM fungi yearly, clear difference in growth performance of mycorrhizal and nonmycorrhizal plants was observed. Plant growth studied in terms of number of branches, available soil P, leaf P, Zn and Cu improved significantly in AM colonized plants compared to uninoculated plants. This trend continued in the 8th year of sampling. The root acid and alkaline phosphatase activity was higher in six month old Arka Puneeth grafted on AM colonized Totapuri rootstock. Mycorrhizal inoculums can be easily multiplied on-farm on finger millet and applied yearly for desired results. © 2012 Elsevier B.V.