Antimicrobial and antioxidant properties of a bacterial. We isolated three endophytic bacteria from the leaves of g. We previously found that the overexpression of viba a gene for transcription factor in e. The extension of nitrogenfixing symbioses to important crop plants such as the cereals has been a longstanding goal in the field of biological nitrogen fixation. Endophytes as sources of bioactive products sciencedirect. The endophyte neotyphodium coenophialum inhabits tall fescue, and produces the toxic alkaloid ergovaline, causing late abortion in broodmares, and impaired growth of youngstock lewis, et al 1997. Disease free parts of the plants were cut with the help of a sterile scalpel and placed in ziplock plastic bags to store the material at 4 0c until isolation. Given that the same sterilization procedure, growth media and incubation conditions were applied to all samples collected for this study, our comparisons of communities across varieties, times of the season and tissue type should be unaffected by any bias related to the specific fungal isolation methods. Nov 17, 2009 introduction to endophytes and their application to develop commercial products 1. Biodegradation of trichloroethylene by an endophyte of hybrid poplar jun won kang, zareen khan, and sharon l. Isolation and characterization of diazotrophic endophyte, asaia bogorensis from mangifera indica patil. Tested by the oregon department of agriculture for endophyte infection, kentucky 32 comes with an official green tag. The role of the oregon state university endophyte service laboratory in diagnosing clinical cases of endophyte toxicoses a.
Montana state university the promise of endophyte biology 2. Except for already established seedendophytes truyens et. Impact of endophytic microorganisms on plants, environment. Omonrice 12 2004 95 the two hosts are given in table 1. Every offshoot was taken 5 pieces, peeled and sectioned in order to result in 10 pieces. Fungal endophytes of wild barley and their effects on. It has been anticipated that more than eighty percent of land plant species form symbiotic association with fungi. The bacterial cells were used to prepare the lysate and ethyl acetate extract was prepared from 7 ml cellfree broth by mixing it with equal volume of. After a correct growth medium is selected, the isolation and characterization of bioactive substances from culture filtrates is done using bioassay guided fractionation and spectroscopic methods.
The uninoculated controls received the same treatment and were exposed to 424 hours to the appropriate media in the absence of endophytes. To isolate and characterize indigenous bacterial endophytes from cultivars of. Pdf isolation and analysis of endophytic microorganisms. Screening of endophytic bacteria isolated from leaves of sambung. Endophytes help plant to cope up with infections developed by pathogen and various environmental stress 15. The strain identity was confirmed by 16s rrnabased phylogeny and scanning electron microscopy sem. This endophyte associated resistance was the result of antibiosis effects or starvation. Analysis of plant growth promoting potential of endophytes isolated. Duringer department of biomedical sciences, college of veterinary medicine, and department of environmental and molecular toxicology, college of agricultural sciences, oregon state university, corvallis. Isolation and identification of bacterial endophytes from. A new isolate, e6, of this organism, with unusual biochemical and biological properties, has been obtained from the branches of a mature guazuma ulmifolia sterculiaceae. An affordable alternative to novelendophyte fescue.
To enrich for potential tce degraders, 1 milliliter of this sample was added into 25 ml of synthetic medium containing 5. Biodegradation of trichloroethylene by an endophyte of. Isolation and characterization of bacterial endophytes of. Transmission of class 1 endophytes is primarily vertical, with maternal plants passing fungi on to offspring via seed infections saikkonen et al. The natural ability of endophytes to degrade the xenobiotics has. The conventional isolation method involves surfacesterilisation of plant samples e. Endophytes were isolated from the stems, leaves, fruits of vitis vinifera, vitis quinquangularis and cayratia trifolia. Muscodor albus e6, an endophyte of guazuma ulmifolia making volatile antibiotics. The most frequently found genera of bacterial endophytes are pseudomonas, micrococcus, bacillus, stenotrophomonas, burkholderia, pantoea, microbacterium, 1622. The biology of this fungus is considered with special reference to host specificity and its occurrence as a free living soil micro.
Isolation, characterization and identification of endophytic. Isolation and identification of endophytic fungi from cocoa. Some of them are mutualistic symbionts with beneficial effects on their host, such as improved growth or resistance against disease or environmental stress, and are being used as microbial inoculants. Endophytes are ubiquitous and have been found in all species of plants studied to date. This was the first study to evaluate their cytotoxic and antitumour activities against brine shrimp and five types of tumour cells, respectively. Plant sample collection and endophyte isolation wheat samples were collected from farmland near the west campus of yangtze university in jingzhou, hubei province, china n30 21032. The mycorrhizal status of a range of isolates is assessed and the distinctive features of the true endophyte are described.
Misaghi ij, donndelinger cr 1990 endophytic bacteria in symptomfree. Some of the more interesting compounds produced by endophytic microbes, with which strobles lab have dealt, are. Isolation and identification of endophytic bacteria from. Pdf isolation and identification of fungal endophytes. Natural products from endophytic microorganisms pubmed.
Endophytefree fescue once the effects of fescue on grazing animals were understood, scientists and plant breeders set to work removing the endophyte from the fescue. One of the approaches that has been used to try to achieve this goal involves the isolation and characterization of stable endophytic bacteria from a variety of wild and cultivated rice species that either have a natural ability to. Isolation and identification of a novel endophyte from a. There are several basic steps in the procedure to reestablish with endophyte free tall fescue. Vegab and soroush parsaa aentomology, international center for tropical agriculture ciat, cali, colombia. Exploration of endophytic microorganisms from selected. Isolation of endophytic fungi from healthy branches cocoa branches chosen in this study were infected free, symptomless, healthy, and wooden tissue. Endophyte free there are many varieties of forage and turf type tall fescue that contain the endophyte fungus, which produces toxic alkaloids that are detrimental to livestock. The role of the oregon state university endophyte service. Isolation of bacillus pumilus from in vitro grapes as a. Cohen sd 1999 technique for large scale isolation of discula umbrinella and other foliar endophytic fungi from quercus species. University of groningen bacterial endophytes of rice hardoim. Bacterial endophytes were sampled from across two wheat types winter, spring.
All samples were surface sterilization by 7% of sodium hypochlorite for 57. Muscodor albus e6, an endophyte of guazuma ulmifolia making. Dormant cuttings of several varieties of young poplars were collected from tcecontaminated sites. An affordable alternative to novel endophyte fescue. A read is counted each time someone views a publication summary such as the title, abstract, and list of authors, clicks on a figure, or views or downloads the fulltext. Isolation of endophytes a surface sterilization of the plant matertial the method most frequently utilised to detect and quantify endophytes involves isolation from surfacesterilized. Endophyte bacillus subtilis evade plant defense by. Isolation of microbial endophytes from some ethnomedicinal.
Bacterial endophyte colonization and distribution within plants mdpi. Isolation of endophytic endosporeforming bacteria from. With high yields and extended longevity, cajun ii produces an abundance of nontoxic forage for many years, providing a fresh option to. Fungal diversity 53 genetic variation of alternaria alternata, an endophytic fungus isolated from pinus tabulaeformis as determined by random amplified microsatellites rams liangdong guo1, li xu1, weihong zheng1 and kevin d. Reestablishment with endophyte free tall fescue if it is decided that reestablishment with endophyte free tall fescue is the best option, 25 percent or less of the total acreage should be replanted each year. There are several basic steps in the procedure to reestablish with endophytefree tall fescue. Isolation procedures for endophytic microorganisms springerlink. Fermentation conditions of an antagonistic endophyte to.
Isolation and identification of fungal endophytes from. Endophyte bacillus subtilis evade plant defense by producing lantibiotic subtilomycin to mask selfproduced flagellin. Pdf isolation and identification of endophytic bacteria from rare. Muscodor albus is an endophytic fungus, represented by a number of isolates from tropical tree and vine species in several of the worlds rainforests, that produces volatile organic compounds vocs with antibiotic activity.
Bacterial endophyte colonization and distribution within plants. B international journal of environmental sciences volume 3 no. Endophytic fungi have been classified into two broad groups discriminated based on phylogeny and life history traits as clavicipitaceous c which infect some grasses and the nonclavicipitaceous endophytes ncendophytes, which can be recovered from asymptomatic tissues of nonvascular plants, ferns and allies, conifers, and angiosperms. Aug 19, 20 endophyte free fescue once the effects of fescue on grazing animals were understood, scientists and plant breeders set to work removing the endophyte from the fescue. Endophyte infected grasses are resistant to sod webworms, chinchbugs, billbugs and numerous other surfacefeeding insects. Isolation and identification of a novel endophyte from a plant amaranthus spinosus shilpa sharma and shikha roy department of botany, university of rajasthan, jaipur, india corresponding author a b s t r a c t keywords number of fungal endophytes than either roots or leaves. In other experiments, endophytefree plants of pi 269406 and pi 4404 h. Endophytic bacteria are relevant symbionts that contribute to plant growth and development. The revelation that general recommendation of 7080% ethanol may not be the most effective bactericidal concentration for all bacteria, elucidation of the possibility of covert bacterial survival in vitro plant cultures and isolation of a potential plant growth promoting endophyte in grape. Endophytes are microorganisms bacteria or fungi or actinomycetes that dwell within robust plant tissues by having a symbiotic association. This study reports on the isolation and identification of methylobacterium radiotolerans mamp 4754 from the seeds of the medicinal plant, combretum erythrophyllum, for the purposes of investigating antimicrobial and antioxidant activities from this endophyte. Conventionally, endophytes were isolated from surface sterilized plant tissue and cultivated in. Pdf isolation and identification of endophytes from.
Endophyteinfected grasses are resistant to sod webworms, chinchbugs, billbugs and numerous other surfacefeeding insects. Isolation, characterization, and insecticidal activity of an. Suryanarayanan vivekananda institute of tropical mycology vinstrom, ramakrishna mission vidyapith, 45a, oliver road, chennai 600 004, india article info article history. Endophytic fungi from the chinese medicinal plant actinidia macrosperma were isolated and identified for the first time. Ne novel endophyte fescue in the late 1990s, scientists identified strains of the tall fescue endophyte that do not produce the alkaloids that. The modified method revealed better result for dna isolation, as clear from strands of dna at the end of isolation protocol. Then 32 endophyteinfected genotypes and 32 endophytefree genotypes of lolium perenne were planted in a farm in isfahan.
Five different taxa were represented by 11 isolates, and six isolates were. Received 28 may 20 revision received 24 august 20 accepted 26 september 20 available. It has been suggested that bacterial endophytes are one step removed from specialized rhizosphere living bacteria, as they are able to interact in a very efficient way with their plant hosts ali et al. Endophytic definition of endophytic by medical dictionary. The effect of endophytic fungi in drought resistance of.
Doty id school of environmental and forest sciences, college of the environment, university of washington. Endophytes isolated either on cmm agar or in m102 produced ergot alkaloids whentransferred to m104t. Endophyte of hybrid poplar biodegradation of trichloroethylene by an. The tissue preparation that yielded the least endophyte was nonmacerated sheath tissue placed in m102. Kit invitrogen, carlsbad, ca, usa following the manufacturers protocol. Jadhav modern college of arts, science and commerce, ganeshkhind, pune, maharashtra, india. Pdf endophytic bacteria are ubiquitous in most plant species, residing latently or. All samples were processed within 48 h of collection.
Any dead plant tissue and most of the roots were removed from the plant. Plant growthpromoting bacterial endophytes sciencedirect. Some commonly found endophytes are those belonging to the genera enterobacter sp. Endophyte reside in the living plant tissue are mostly unstudied and potential sources of new natural products for therapeutic preparations. It also describes the threats that some endophyte plant associations have in animal production. Isolation procedures for endophytic microorganisms. Samples were collected in an aseptic bag and were taken back to the laboratory immediately. Isolation and characterisation of bacterial root endophytes. Muscodor albus e6, an endophyte of guazuma ulmifolia. For the isolation of endophytic bacterial isolates, the rhizomes of c. Endophytes are known to supply nutrients to plant by fixing atmospheric nitrogen and solubilizing ion marx 2004.
Fresh and healthy rhizomes were washed thoroughly under running tap water and surface sterilized 70 % c 2 h 5 oh, 3 min, 0. Reestablishment with endophytefree tall fescue if it is decided that reestablishment with endophytefree tall fescue is the best option, 25 percent or less of the total acreage should be replanted each year. Endophytic bacteria are those bacteria that reside inside plant tissues and showed no visible external signs of invasion or negative effects on plant growth schulz and boyle, 2006. Procedure for isolating the endophyte tall and screening. A number of methods are used to achieve this, including heat, humidity, and various chemicals. Isolation and identification of endophytic fungi from. Previous research at the ohio state university showed that excellent control bluegras of s billbug can be obtained in kentucky bluegrass lawns overseeded with endophyte infected perennial rye. A fivefold serial dilution was performed after which 100l was inoculated on yeast manitol agar yem, nitrogen free medium, and nutrient agar. Isolation of endophytic endosporeforming bacteria from theobroma cacao as potential biological control agents of cacao diseases rachel l. Cajun ii endophyte free forage tall fescue is an affordable alternative to both toxic ky31 and novel endophyte tall fescues.
Samples were rinsed with water to remove soil and debris, swirled in a beaker containing. Isolation and purification of endophytic however, endophytes are a rich and reliable source of genetic diversity, novel nondescribed species and novel bioactive compounds like new drugs for effective treatment of diseases in humans, plants and animals9. Jun 11, 20 given that the same sterilization procedure, growth media and incubation conditions were applied to all samples collected for this study, our comparisons of communities across varieties, times of the season and tissue type should be unaffected by any bias related to the specific fungal isolation methods. Preparation of culture media yeast manitol agar sigma aldrich, steinheim, germany, nitrogen free medium, and nutrient agar sigma. This ultimately leads to increase in plant immune system as well as protects plant from infection by plant pathogens. They are ubiquitously associated with almost all plants studied till date. The healthy plant samples were collected and cut into small pieces approximately 0. A protocol for a newlydeveloped dilutiontoextinction method for isolation and cultivation of horizontallytransmitted endophytes is detailed in the experimental procedures, and protocols for. Previous research at the ohio state university showed that excellent control bluegras of s billbug can be obtained in kentucky bluegrass lawns overseeded with endophyteinfected perennial rye. Read department of botany, the university of sheffield received 3 october 1972 summary techniques are described for the isolation of fungi inhabiting ericaceous roots and for the culture of mycorrhizas under aseptic conditions. Thus, endophytes employ different mechanisms to gain entry into the plant tissues, particularly in roots. Isolation and characterization of diazotrophic endophyte. In order to study the role of endophytic fungi in drought resistance of lolium perenne, an experiment was conducted in a randomized complete block design with two replications. A novel method to scale up fungal endophyte isolations.
Isolation, characterization and identification of endophytic bacteria by 16s rrna partial sequencing technique from roots and leaves of prosopis cineraria plant rekha m. In total, 168 bacterial strains were isolated and assigned to 8 otus. Botanical samples of seven aquilaria species were collected, and endophytic bacteria were isolated from surfacesterilizedtissue samples. Diversity free fulltext isolation and identification of endophytic.
Bills gf 1996 isolation and analysis of endophytic fungal communities from woody. An endophyte is an endosymbiont, often a bacterium or fungus, that lives within a plant for at least part of its life cycle without causing apparent disease. Genetic variation of alternaria alternata, an endophytic. However, pathogens and saprophytes may also be endophytic at some point of. Introduction to endophytes and their application to develop. The offshoots were hand carried to laboratorium, washed by using drop water, and cutted into small pieces 4mm. Pdf isolation and identification of fungal endophytes researchgate. Misaghi ij, donndelinger cr 1990 endophytic bacteria in symptom free. Isolation of micropropagated strawberry endophytic bacteria and assessment of their potential for plant growth promotion. Endophytes are microorganisms living within the tissue of a plant as endosymbionts, without causing symptoms of disease. Criteria for isolation of endophytes are closely related to isolation of bioactive molecules, e.
No dna strands were visible after isolation steps, as indicated by established protocol. The causal agent of dutch elm disease ceratocystis ulmi 3. Bacterial endophyte antagonism toward a fungal pathogen in. Samples were rinsed with water to remove soil and debris, swirled in a beaker containing distilled water and two drops of tween 20100 ml, and rinsed again prior to cutting the tissue. Isolation and culturing of bacterial endophytic isolates the samples were aseptically ground in a motor and pestle in potassium dihydrogen phosphate buffer ph 6. Spatial and temporal variation in fungal endophyte. Identification and characterization of endophytic bacteria. Genomic dna extraction for molecular identification of.
Isolation and characterization of indigenous endophytic bacteria. Isolation and purification of endophytic however, endophytes are a rich and reliable source of genetic diversity, novel non. Endophyte isolation samples of roots, leaves, flowers and fruit from healthy plants were washed with water to remove soil contaminants then surface sterilized by soaking in 70% ethanol for 2 min, sodium hypochlorite solution 525% for 5 min, and finally 70% ethanol for a further 30 s before being air. Isolation and characterization of endophytes from different.
1476 40 724 1023 1387 59 689 658 42 1474 1164 1101 1037 545 258 3 1530 953 47 616 530 321 113 1369 497 610 180 312 324 1195 731