Search results for: endophytic fungi

Authors: Reine Suci Wulandari, Rosa Suryantini

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Albizia (Paraserianthes falcataria) is a woody plant species that has a high economic value and multifunctional. Albizia is important timber, medicinal plants and can also be used as a plant to rehabilitate critical lands. The demand value of Albizia is increased so that the large quantities and high quality of seeds are required. In vitro propagation techniques are seed propagation that can produce more seeds and quality in a short time. In vitro cultures require growth regulators that can be obtained from biological agents such as endophytic fungi. Endophytic fungi are micro fungi that colonize live plant tissue without producing symptoms or other negative effects on host plants and increase plant growth. The purposes of this research were to isolate and identify endophytic fungi isolated from the root of Albizia and to study the effect of endophytic fungus on the growth of Albizia in vitro. The methods were root isolation, endophytic fungal identification, and inoculation of endophytic fungi to Albizia plants in vitro. Endophytic fungus isolates were grown on PDA media before being inoculated with Albizia sprouts. Incubation is done for 4 (four) weeks. The observed growth parameters were live explant percentage, percentage of explant shoot, and percentage of explant rooted. The results of the research showed that 6 (six) endophytic fungal isolates obtained from the root of Albizia, namely Aspergillus sp., Verticillium sp, Penicillium sp., Trichoderma sp., Fusarium sp., and Acremonium sp. Statistical analysis found that Trichoderma sp. and Fusarium sp. affect in vitro growth of Albizia. Endophytic fungi from the results of this research were potential as plant growth promoting. It can be applied to increase productivity either through increased plant growth and increased endurance of Albizia seedlings to pests and diseases.

Keywords: Albizia, endophytic fungi, propagation, in vitro

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Authors: Akwu A. Nneka, Naidoo, Yougasphree

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A significantly high number of microbes exist in higher plants; these microbes include bacteria, fungi, and actinomycetes. There are reports on the benefits of endophytic fungi and their products of metabolism to the host plant and man, consequently, it is expedient to explore the changes that could arise as a result of manipulating their growth media. Grewia lasiocarpa E. Mey. ex Harv. (Malvaceae) is an indigenous Southern African plant, that belongs to a genus with known medicinal properties. Three media were used to culture the endophytic fungi viz., Potato Dextrose Agar (PDA), Malt Extract Agar (MEA), and Bacteriological Agar (BA) were used singly, and supplemented with three dilutions of the leaves and stem bark extracts. The manipulated growth media composition had a significant effect on the diversity of the isolated fungal populations. Several endophytic fungi were isolated; their distribution and diversity revealed a significant relatedness with the manipulated media. The media supplemented with the plant extracts was observed to give a significant increase in the growth rate and yield of the endophytes. To the best of our knowledge, this is the first study describing the endophytic fungi present in the leaves and stem bark of G. lasiocarpa E. Mey. ex Harv.

Keywords: Grewia lasiocarpa, plant-based extracts, endophytic fungi, Malvaceae

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Authors: Gauravi Agarkar, Mahendra Rai

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Endophytic fungi from medicinal plants are important source of numerous pharmacologically important compounds. In the present investigation, the endophytic fungi were isolated from three medicinal plants; Andrographis paniculata, Rauwolfia serpentina and Tridax procumbens. Endophytic Colletotrichum sp. were identified on the basis of cultural and morphological characteristics as well as internal transcribed spacer (ITS) sequence analysis. Antibacterial and antifungal activity of the ethyl acetate and methanol extract of endophytic Colletotrichum sp. was evaluated against seven different human pathogenic bacteria and six Candida sp. The extracts were effective and showed significant activity against all the test pathogens. In case of yeast Candida, the combined effect of extracts and standard antibiotic was enhanced greatly showing synergistic activity. Further, the extracts were assayed for Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal/Fungicidal Concentration (MBC/MFC) where, MIC values were in the range of 100-250 μg/ml. These results suggest that the endophytic Colletotrichum sp. isolated from the medicinal plants are capable of producing promising antimicrobial metabolites.

Keywords: antimicrobial, colletotrichum, endophytic fungi, medicinal plants

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Authors: Daomao Yang, Qizhi Wang

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Monoterpenoids are mainly found in plant essential oils and they are ideal substrates for biotransformation into oxygen-containing derivatives with important commercial value due to their low price and simple structure. In this paper, eleven strains of endophytic fungi from Praxelis clematidea were used as test strains to conduct the whole cell biotransformation of the monoterpenoids: (+)-limonene, (-)-limonene and myrcene. The fungi were inoculated in 50 ml Sabouraud medium and incubated at 30 ℃ with the agitation of 150 r/min for 6 d, and then 0.5% (v/v) substrates were added into the medium and biotransformed for further 3 d. Afterwards the cultures were filtered, and extracted using equal volume of ethyl acetate. The metabolites were analyzed by GC-MS technique with NIST database. The Total Ion Chromatogram of the extractions from the eleven strains showed that the main product of (+)- and (-)-limonene biotransformation was limonene-1,2-diol, while it is limonene and linalool oxide for biotransformation of myrcene. This work will help screen the microorganisms to biotransform the monoterpenes.

Keywords: endophytic fungi, (+)–limonene, (-)–limonene, myrcene

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Authors: Mohd Sidek Ahmad, Zainon Mohd Noor, Zaidah Zainal Ariffin

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Fibrin degradation is an important part in prevention or treatment of intravascular thrombosis and cardiovascular diseases. Plasmin like fibrinolytic enzymes has given new hope to patient with cardiovascular diseases by treating fibrin aggregation related diseases with traditional plasminogen activator which have many side effects. Various researches involving wide range of sources for production of fibrinolytic proteases, from bacteria, fungi, insects and fermented foods. But few have looked into endophytic fungi as a potential source. Sixteen (16) endophytic fungi were isolated from Hibiscus sp. leaves from six different locations in Shah Alam, Selangor. Only two endophytic fungi, FH3 and S13 showed positive fibrinolytic protease activities. FH3 produced 5.78cm and S13 produced 4.48cm on Skim Milk Agar after 4 days of incubation at 27°C. Fibrinolytic activity was observed; 3.87cm and 1.82cm diameter clear zone on fibrin plate of FH3 and S13 respectively. 18srRNA was done for identification of the isolated fungi with positive fibrinolytic protease. S13 had the highest similarity (100%) to that of Penicillium citrinum strain TG2 and FH3 had the highest similarity (99%) to that of Fusarium sp. FW2PhC1, Fusarium sp. 13002, Fusarium sp. 08006, Fusarium equiseti strain Salicorn 8 and Fungal sp. FCASAn-2. Media composition variation showed the effects of carbon nitrogen on protein concentration, where the decrement of 50% of media composition caused drastic decrease in protease of FH3 from 1.081 to 0.056 and also S13 from 2.946 to 0.198.

Keywords: isolation, identification, fibrinolytic protease, endophytic fungi, Hibiscus leaves

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Authors: Lyudmila V. Ignatova, Yelena V. Brazhnikova, Togzhan D. Mukasheva, Ramza Zh. Berzhanova, Anel A. Omirbekova

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Endophytic microorganisms are presented in plants of different families growing in the foothills and piedmont plains of Trans-Ili Alatau. It was found that the maximum number of endophytic micromycetes is typical to the Fabaceae family. The number of microscopic fungi in the roots reached (145.9±5.9)×103 CFU/g of plant tissue; yeasts - (79.8±3.5)×102 CFU/g of plant tissue. Basically, endophytic microscopic fungi are typical for underground parts of plants. In contrast, yeasts more infected aboveground parts of plants. Small amount of micromycetes is typical to inflorescence and fruits. Antagonistic activity of selected micromycetes against Fusarium graminearum, Cladosporium sp., Phytophtora infestans and Botrytis cinerea phytopathogens was detected. Strains with a broad, narrow and limited range of action were identified. For further investigations Rh2 and T7 strains were selected, they are characterized by a broad spectrum of fungicidal activity and they formed the large inhibition zones against phytopathogens. Active antagonists are attributed to the Rhodotorula mucilaginosa and Beauveria bassiana species.

Keywords: endophytic micromycetes, fungicidal activity, prevalence, plants

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Authors: Walaa Al-Maghraby

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Endophytes are organisms that colonize internal plant tissues without causing apparent harm to their host. Almost all groups of microorganisms have been found in endophytic association with plants may be fungi. They stimulate the production of secondary metabolites with a diverse range of biological activities. Leptadenia pyrotechnica is a more or less leafless, erect shrub with straight stems which is highly distributed in Saudi Arabia. Four endophytes fungi were isolated from Leptadenia pyrotechnica and identified using 18S ribosomal RNA sequences, which revealed four fungi genuses, namely Aspergillus terreus; Aspergillus welwitschiae; Aspergillus fumigatus and Aspergillus flavus. In this present study, four endophytic fungi from Leptadenia pyrotechnica were used for obtaining crude aqueous and ethyl acetate extracts for antimicrobial screening against 6 human pathogens, the antibacterial tests presented satisfactory results, where the pathogenic bacteria were inhibited by the four extracts tested, except for Escherichia coli that was inhibited by all extracts except ethyl acetate extract of Aspergillus terreus. Analysis of variance showed that the extract produced by endophyte Leptadenia pyrotechnica was the most effective against all bacteria, either gram-negative or positive. However, the extract was not efficient against pathogenic fungi. Therefore, this study indicates that endophytes from medicinal plant Leptadenia pyrotechnica could be potential sources of antibacterial substances.

Keywords: antimicrobial activity, Aspergillus sp, endophytes, Leptadenia pyrotechnica

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Authors: Gauravi Agarkar, Mahendra Rai

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Fungi are closely associated with the plants in various types of interactions such as mycorrhizal, parasitic or endophytic. Some of these interactions are beneficial and a few are harmful to the host plants. It has been suggested that these plant-associated fungi are able to change their lifestyle abd this means endophyte may become parasite or vice versa. This phenomenon may have profound effect on plant-fungal interactions and various ecological niches. Therefore, it is necessary to identify the factors that trigger the change in fungal lifestyle and understand whether these different lifestyles are interconnected at some points either by physiological, biochemical or molecular routes. This review summarizes the factors affecting plant fungal interactions and discusses the possible mechanisms for lifestyles switching of fungi based on available experimental evidences. Research should be boosted in this direction to fetch more advantages in future and to avoid the severe consequences in agriculture and other related fields.

Keywords: endophytic, lifestyle switching, mycorrhizal, parasitic, plant-fungal interactions

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Authors: Kanika Chowdhary, Satyawati Sharma

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Endophytic microbes are hosted inside plants in a symbiotic and hugely benefitting relationship. Exploring agriculturally beneficial endophytes is quite a prospective field of research. In the present work fungal endophytes associated with aromatic plant Ocimum basicilium L. were investigated for biocontrol potential. The anti-plant pathogenic activity of fungal endophytes was tested against causal agent of stem rot Sclerotinia sclerotiorum. 75 endophytic fungi were recovered through culture-dependent approach. Fungal identification was performed both microscopically and by rDNA ITS sequencing. Curvuaria lunata (Sb-6) and Colletotrichum lindemuthianum (Sb-8) inhibited 86% and 72% mycelia growth of S. sclerotinia on Sabouraud dextrose agar medium at 7.4 pH. Small-scale fermentation was carried out on sterilised oatmeal grain medium. In another set of experiment, fungi were grown in oatmeal grain medium amended with certain biostimulants such as aqueous seaweed extract (10% v/w); methanolic seaweed extract (5% v/w); cow urine (20% v/w); biochar (10% w/w) in triplicate along with control of each to ascertain the degree of metabolic difference and anti-plant pathogenic activity induced. Phytochemically extracts of both the fungal isolates showed the presence of flavanoids, phenols, tannins, alkaloids and terpenoids. Ethylacetate extract of C. lunata and C. lindemuthianum suppressed S. sclerotinia conidial germination at IC50 values of 0.514± 0.02 and 0.913± 0.04 mg/ml. Therefore, fungal endophytes of O. basicilium are highly promising bio-resource agent, which can be developed further for sustainable agriculture.

Keywords: endophytic fungi, ocimum basicilium, sclerotinia sclerotiorum, biostimulants

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Authors: Ahlem Nefzi, Rania Aydi Ben Abdallah, Hayfa Jabnoun-Khiareddine, Ammar Nawaim, Rabiaa Haouala, Mejda Daami-Remadi

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Seven endophytic fungi were isolated from the wild Solanaceous species Lycium arabicum growing in the Tunisian Centre-East and were assessed for their ability to suppress Fusarium Crown and Root Rot disease caused by Fusarium oxysporum f. sp. radicis lycopersici (FORL) and to enhance plant growth. Fungal isolates were shown able to colonize tomato cv. Rio Grande roots, crowns, and stems. A significant promotion in all studied growth parameters (root length, shoot height, and roots and shoots fresh weight) was recorded in tomato plants treated with fungal conidial suspensions or their cell-free culture filtrates compared to FORL-inoculated or pathogen-free controls. I15 and I18 isolates were shown to be the most effective leading to 85.7-87.5 and 93.6-98.4% decrease in leaf and root damage index and the vascular discoloration extent, respectively, over FORL-inoculated and untreated control. These two bioactive and growth-promoting isolates (I15 and I18) were morphologically characterized and identified using rDNA sequencing gene as being Alternaria alternata (MF693801) and Fusarium fujikuroi (MF693802). These fungi significantly suppressed FORL mycelial growth and showed chitinolytic, proteolytic and amylase activities whereas only F. fujikuroi displayed a lipolytic activity. This study clearly demonstrated the potential use of fungi naturally associated with L. arabicum as biocontrol and bio-fertilizing agents.

Keywords: biocontrol, endophytic fungi, Fusarium oxysporum f. sp. radicis-lycopersici, tomato promotion, Lycium arabicum

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Authors: Gloria M. Ntuba-Jua, Afui M. Mih, Eneke E. T. Bechem

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Prunus africana (Hook. F.) Kalkman, commonly known as Pygeum or African cherry belongs to the Rosaceae family. It is a medium to large, evergreen tree with a spreading crown of 10 to 20 m. It is used by the traditional medical practitioners for the treatment of over 45ailments in Cameroon and sub-Sahara Africa. In modern medicine, it is used in the treatment of benign prostrate hyperplasia (BPH), prostate gland hypertrophy (enlarged prostate glands). This is possible because of its ability to produce some secondary metabolites which are believed to have bioactivity against these ailments. The ready international market for the sale of Prunus bark, uncontrolled exploitation, illegal harvesting using inappropriate techniques and poor timing of harvesting have contributed enormously to making the plant endangered. It is known to harbor a large number of endophytic fungi with the potential to produce similar secondary metabolites as the parent plant. Alternative sourcing of medicinal principles through endophytic fungi requires succinct knowledge of the endophytic fungi. This will serve as a conservation measure for Prunus africana by reducing dependence on Prunus bark for such metabolites. This work thus sought to compare the production of some major secondary metabolites produced by P. africana and some of its associated endophytic fungi. The leaves and stem bark of the plant from different provenances were soaked in methanol for 72 hrs to yield the methanolic crude extract. The phytochemical screening of the methanolic crude extracts using different standard procedures revealed the presence of tannins, flavonoids, terpenoids, saponins, phenolics and steroids. Pure cultures of some predominantly isolated endophyte species from the difference Prunus provenances such as Curvularia sp, and Morphospecies P001 were also grown in Potato Dextrose Broth (PDB) for 21 days and later extracted with Methylene dichloride (MDC) solvent after 24hrs to produce crude culture extracts. Qualitative assessment of crude culture extracts showed the presence of tannins, terpenoids, phenolics and steroids particularly β-Sitosterol, (a major bioactive metabolite) as did the plant tissues. Qualitative analysis by thin layer chromatography (TLC) was done to confirm and compare the production of β-Sitosterol (as marker compounds) in the crude extracts of the plant and endophyte. Samples were loaded on TLC silica gel aluminium barked plate (Kieselgel 60 F254, 0.2 mm, Merck) using acetone/hexane, (3.0:7.0) solvent system. They were visualized under an ultra violet lamp (UV254 and UV360). TLC revealed that leaves had a higher concentration of β-sitosterol in terms of band intensity than stem barks from the different provenances. The intensity of β-sitosterol bands in the culture extracts of endophytes was comparable to the plant extracts except for Curvularia sp (very minute) whose band was very faint. The ability of these fungi to make β-sitosterol was confirmed by TLC analysis with the compound having chromatographic properties (retention factor) similar to those of β-sitosterol standard. The ability of these major endophytes to produce secondary metabolites similar to the host has therefore been demonstrated. There is, therefore, the potential of developing the in vitro production system of Prunus secondary metabolites thereby enhancing its conservation.

Keywords: Caneroon, endophytic fungi, Prunus africana, secondary metabolite

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Authors: Rasha Saad, Jean Frederic Weber, Fatimah Bebe, Sadia Sultan

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The goal of study was to screen the effects of candesartan and four endophytic fungi for their potential in microbial biotransformation. In this experiment, four types of unidentified fungi with the codes of TH2L1, TH2R10, TH1P35 and TH1S46 were used in screening process by MECFUS (Microtiter plate, Elicitors, Combination, Freeze-drying, UHPLC, Statistical analysis) protocol. The experiment was carried out by using 96-well microtiter plate (MTP) with different media and elicitors. Various media with two concentrations of Potato Dextrose Broth (PDB) and elicitors used were to induce the production of secondary metabolites from the fungi as well as the biotransformation of the drug compound. After incubation, cultures were extracted by freeze drying method and finally analyzed by ultra-High performance Liquid Chromatography (uHPLC). The extracts analyzed by uHPLC followed by LC/Ms, demonstrated the presence of biotransformation products from the drug compound and elicitation of the secondary metabolism from the fungi by the occurrence of the additional peaks. From the four fungi, TH1S46 showed highly potential produced secondary metabolites as well as the biotransformation of candesartan. For other fungi, they responded when candesartan was introduced. Moreover, the additional peaks produced in uHPLC need to be further investigation by using LC-MS or NMR.

Keywords: biotransformation, candesartan, endophytes, secondary metabolites

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Authors: Dawang D. N., Dasat G. S., Nden D.

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Endophyte means “in the plant” are referred to all microorganisms that live in the internal tissues of stems, petioles, roots and leaves of plants causing no apparent symptoms of disease. Secondary metabolites from fungal endophytes have an enormous potential applications as antioxidant, antimicrobial, anticancer and antidiabeties. Thus, this study aimed to determine the antimicrobial activity of these metabolites against some clinical isolates. The fungi were subjected to fermentation medium and the metabolites were extracted using ethyl acetate. The fungal extracts showed both antibacterial and antifungal activities with maximum zone of inhibition diameter of 10.5mm against Aspergillus fumigatus. Staphylococcus aureus was inhibited by all the five crude extracts with inhibition zone diameter of 4mm. Endophytic fungal crude extract2 (EDF2) exhibited antimicrobial effect against all the test organisms used, EDF4 was active against all test organisms except on Penicillium chrysogenum and Klebsiella pneumoniae. Antibacterial standard of ciprofloxacin which is 15mm is comparable to the effect of endophytic extract of EDF1 and EDF2. Klebsiella pneumoniae was resistant to EDF4 and EDF5. EDF3 showed a wide range of antimicrobial activity against all the test organisms used. The highest inhibition zone diameter of 10.50mm recorded against Aspergillus fumigatus is comparable to antifungal standard of fluconazole (15.5mm). The result of this study suggests that endophytic fungi associated with the roots of Irish potato could be a promising source of novel bioactive compounds of pharmaceutical and industrial importance.

Keywords: endophyte, fungal extract, antimicrobial, potato

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Authors: Bandana Saikia, Ashok Bhattacharyya

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Endophytic microbes and their metabolites positively impact overall plant health, which may have a potential implication in agriculture. In the present study, 177 bacterial endophytes and 57 fungal endophytes were isolated, with the highest recovery rate from tomato roots. A maximum of 112 endophytes were isolated during monsoon, followed by 64 isolates and 58 isolates isolated during pre-monsoon and post-monsoon periods, respectively, indicating the rich diversity in bacterial and fungal endophytes of tomato crops from different locations of Assam, India. Further, the endophytes were evaluated for their antagonistic potential against Fusarium oxysporum f. sp. lycopersici. Fungal endophytic isolate AAUTLF (Endophytic Fungi of Tomato Leaf from Assam Agricultural University, Assam, India area) and bacterial endophyte D1B (Endophytic bacteria of tomato from Dhemiji, India district) showed the highest antifungal activity against the pathogen both in vitro and in vivo. Based on 5.8 rDNA sequence analysis of fungal and 16S rDNA sequence of bacteria endophytes, the most effective fungal and bacterial isolates against FOL were identified as Trichoderma asperellum AAUTLF and Stenotrophomonas maltophilia D1B, respectively. The isolates showed an antagonistic effect against Fusarium oxysporum f.sp. lycopersici in-vitro and reduced the disease index of Fusarium wilt in tomatoes by 64.4% under pot conditions. Trichoderma asperellum AAUTLF produced an antifungal compound viz., 6-pentyl-2H-pyran-2-one, which also possesses growth-promoting characteristics. The bacteria Stenotrophomonas maltophilia D1B produced antifungal compounds, including benzothiazole, oleic acid, phenylacetic acid, and 3-(Hydroxy-phenyl-methyl)-2,3-dimethyl-octan-4-one. This would be of high importance for the source of antagonistic strains and biocontrol of tomato Fusarium wilt, as well as other plant fungal diseases.

Keywords: root endophytes, Stemotrophomonas, Trichoderma, benzothiazole, 6-pentyl-2H-pyran-2-one

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Authors: Deepanwita Deka, Dhruva Kumar Jha

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Endophytes are the microbes that colonize living, internal tissues of plants without causing any immediate, overt negative effects. Endophytes are rich source of therapeutic substances like antimicrobial, anticancerous, herbicidal, insecticidal, immunomodulatory compounds. Brucea mollis, commonly known as Quinine in Assam, belonging to the family Simaroubaceae, is a shrub or small tree, recorded as endangered species in North East India by CAMP survey in 2003. It is traditionally being used as antimalarial and antimicrobial agent and has antiplasmodial, cytotoxic, anticancer, diuretic, cardiovascular effect etc. Being endangered and medicinal; this plant may host certain noble endophytes which need to be studied in depth. The aim of the present study was isolation and identification of potent endophytic fungi from Brucea mollis, an endangered medicinal plant, to protect it from extinction due to over use for medicinal purposes. Aseptically collected leaves, barks and roots samples of healthy plants were washed and cut into a total of 648 segments of about 2 cm long and 0.5 cm broad with sterile knife, comprising 216 segments each from leaves, barks and roots. These segments were surface sterilized using ethanol, mercuric chloride (HgCl2) and aqueous solution of sodium hypochlorite (NaClO). Different media viz., Czapeck-Dox-Agar (CDA, Himedia), Potato-Dextrose-Agar (PDA, Himedia), Malt Extract Agar (MEA, Himedia), Sabourad Dextrose Agar (SDA, Himedia), V8 juice agar, nutrient agar and water agar media and media amended with plant extracts were used separately for the isolation of the endophytic fungi. A total of 11 fungal species were recovered from leaf, bark and root tissues of B. mollis. The isolates were screened for antimicrobial, antioxidant and enzymatic activities using certain protocols. Cochliobolus geniculatus was identified as the most dominant species. The mycelia sterilia (creamy white) showing highest inhibitory activity against Candida albicans (MTCC 183) was induced to sporulate using modified PDA media. The isolate was identified as Geosmithia pallida. The internal transcribed spacer of rDNA was sequenced for confirmation of the taxonomic identity of the sterile mycelia (creamy white). The internal transcribed spacer r-DNA sequence was submitted to the NCBI (KU693285) for the first time from India. G. pallida and Penicillium showed highest antioxidant activity among all the isolates. The antioxidant activity of G. pallida and Penicillium didn’t show statistically significant difference (P˃0.05). G. pallida, Cochliobolus geniculatus and P. purpurogenum respectively showed highest cellulase, amylase and protease activities. Thus, endopytic fungal isolates may be used as potential natural resource of pharmaceutical importance. The endophytic fungi, Geosmithia pallida, may be used for synthesis of pharmaceutically important natural products and consequently can replace plants hitherto used for the same purpose. This study suggests that endophytes should be investigated more aggressively to better understand the endophyte biology of B. mollis.

Keywords: Antimicrobial activity, antioxidant activity, Brucea mollis, endophytic fungi, enzyme activity, Geosmithia pallida

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Authors: Maryam Beiranvand, Sajad Yaghoubi

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Background: Some microbes can colonize plants’ inner tissues without causing obvious damage and can even produce useful bioactive substances. In the present study, the diversity of the endophytic bacteria associated with medicinal plants from Iran was investigated by culturing techniques, molecular gene identification, as well as measuring them for antibacterial activity. Results: In the spring season from 2013 to 2014, 35 herb pharmacology samples were collected, sterilized, meshed, and then cultured on selective media culture. A total of 199 endophytic bacteria were successfully isolated from 35 tissue cultures of medical plants, and sixty-seven out of 199 bacterial isolates were subjected to identification by the 16S rRNA gene sequence analysis method. Based on the sequence similarity gene and phylogenetic analyses, these isolates were grouped into five classes, fourteen orders, seventeen families, twenty-one genera, and forty strains. The most abundant group of endophytic bacteria was actinobacterial, consisting of thirty-two (47%) out of 67 bacterial isolates. Ten (22.3%) out of 67 bacterial isolates remained unidentified and classified at the genus level. The signature of the 16S rRNA gene formed a distinct line in a phylogenetic tree showing that they might be new species of bacteria. One (5.2%) out of 67 bacterial isolates was still not well categorized. Forty-two out of 67 strains were candidates for antimicrobial activity tests. Nineteen (45%) out of 42 strains showed antimicrobial activity multidrug resistance (MDR); thirteen (68%) out of 19 strains were allocated to classes actinobacteria. Four (21%) out of 19 strains belonged to the Bacillaceae family, one (5.2%) out of 19 strains was the Paenibacillaceae family, and one (5.2%) out of 19 strains belonged to the Pseudomonadaceae family. The other twenty-three strains did not show inhibitory activities. Conclusions: Our research showed a high-level phylogenetic diversity and the intoxicating antibiotic activity of endophytic bacteria in the herb pharmacology of Iran.

Keywords: Antibacterial activity, endophytic bacteria, multidrug-resistant bacteria, whole genom sequencing

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Authors: Maryam Beiranvand, Sajad Yaghoubi

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Background: Some microbes can colonize plants’ inner tissues without causing obvious damage and can even produce useful bioactive substances. In the present study, the diversity of the endophytic bacteria associated with medicinal plants from Iran was investigated by culturing techniques, molecular gene identification, as well as measuring them for antibacterial activity. Results: In the spring season from 2013 to 2014, 35 herb pharmacology samples were collected, sterilized, meshed, and then cultured on selective media culture. A total of 199 endophytic bacteria were successfully isolated from 35 tissue cultures of medical plants, and sixty-seven out of 199 bacterial isolates were subjected to identification by the 16S rRNA gene sequence analysis method. Based on the sequence similarity gene and phylogenetic analyses, these isolates were grouped into five classes, fourteen orders, seventeen families, twenty-one genera, and forty strains. The most abundant group of endophytic bacteria was actinobacterial, consisting of thirty-two (47%) out of 67 bacterial isolates. Ten (22.3%) out of 67 bacterial isolates remained unidentified and classified at the genus level. The signature of the 16S rRNA gene formed a distinct line in a phylogenetic tree showing that they might be new species of bacteria. One (5.2%) out of 67 bacterial isolates was still not well categorized. Forty-two out of 67 strains were candidates for antimicrobial activity tests. Nineteen (45%) out of 42 strains showed antimicrobial activity multidrug-resistance (MDR); thirteen (68%) out of 19 strains were allocated to classes actinobacteria. Four (21%) out of 19 strains belonged to the Bacillaceae family, one (5.2%) out of 19 strains was the Paenibacillaceae family, and one (5.2%) out of 19 strains belonged to the Pseudomonadaceae family. The other twenty-three strains did not show inhibitory activities. Conclusions: Our research showed a high-level phylogenetic diversity and the intoxicating antibiotic activity of endophytic bacteria in the herb pharmacology of Iran.

Keywords: medical plant, endophytic bacteria, antimicrobial activity, whole genome sequencing analysis

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Authors: Iyabode A. Kehinde, Joshua O. Oyekanmi, Jumoke T. Abimbola, Olajumoke E. Ayanda

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African yam bean (AYB), (Sphenostylis stenocarpa) is a leguminous crop that provides nutritionally rich seeds, tubers and leaves for human consumption. AYB potentials as an important food security crop is yet to be realized and thus classified as underutilized crop. Underutilization of the crop has been partly associated with scarce information on the incidence and characterization of fungal endophytes infecting vascular parts of AYB. Accurate and robust detection of these endophytic fungi is essential for diagnosis, modeling, surveillance and protection of germplasm (seed) health. This work aimed at isolating and identifying fungal endophytes associated with leaves, stems and roots of AYB in Ogun State, Nigeria. This study investigated both cultural and molecular properties of endophytic fungi in AYB for its characterization and diversity. Fungal endophytes were isolated and culturally identified. DNA extraction, PCR amplification using ITS primers and analyses of nucleotide sequences of ribosomal DNA fragments were conducted on selected isolates. BLAST analysis was conducted on consensus nucleotide sequences of 28 out of 30 isolates and results showed similar homology with genera of Rhizopus, Cunninghamella, Fusarium, Aspergillus, Penicillium, Alternaria, Diaporthe, Nigrospora, Purpureocillium, Corynespora, Magnaporthe, Macrophomina, Curvularia, Acrocalymma, Talaromyces and Simplicillium. Slight similarity was found with endophytes associated with soybean. Phylogenetic analysis by maximum likelihood method showed high diversity among the general. These organisms have high economic importance in crop improvement. For an instance, Purpureocillium lilacinum showed high potential in control of root rot caused by nematodes in tomatoes. Though some can be pathogens, but many of the fungal endophytes have beneficial attributes to plant in host health, uptake of nutrients, disease suppression, and host immunity.

Keywords: molecular characterization, African Yam Bean, fungal endophyte, plant parts

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Authors: Nefzi Ahlem, Aydi Ben Abdallah Rania, Jabnoun-Khiareddine Hayfa, Ammar Nawaim, Mejda Daami-Remadi

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Fusarium Crown and Root Rot (FCRR) caused by Fusarium oxysporum f. sp. radicis-lycopersici (FORL) is a serious tomato (Solanum lycopersicum L.) disease in Tunisia. Its management is very difficult due to the long survival of its resting structures and to the luck of genetic resistance. In this work, we explored the wild Solanaceae species Withania somnifera, growing in the Tunisian Centre-East, as a potential source of biocontrol agents effective in FCRR suppression and tomato growth promotion. Seven fungal isolates were shown able to colonize tomato roots, crowns, and stems. Used as conidial suspensions or cell-free culture filtrates, all tested fungal treatments significantly enhanced tomato growth parameters by 21.5-90.3% over FORL-free control and by 27.6-93.5% over pathogen-inoculated control. All treatments significantly decreased the leaf and root damage index by 28.5-92.8 and the vascular browning extent 9.7-86.4% over FORL-inoculated and untreated control. The highest disease suppression ability (decrease by 86.4-92.8% in FCRR severity) over pathogen-inoculated control and by 81.3-88.8 over hymexazol-treated control) was expressed by I6 based treatments. This endophytic fungus was morphologically characterized and identified using rDNA sequencing gene as Fusarium sp. I6 (MG835371). This fungus was shown able to reduce FORL radial growth by 58.5–83.2% using its conidial suspension or cell-free culture filtrate. Fusarium sp. I6 showed chitinolytic, proteolytic and amylase activities. The current study clearly demonstrated that Fusarium sp. (I6) is a promising biocontrol candidate for suppressing FCRR severity and promoting tomato growth. Further investigations are required for elucidating its mechanism of action involved in disease suppression and plant growth promotion.

Keywords: antifungal activity, associated fungi, Fusarium oxysporum f. sp. radicis-lycopersici, Withania somnifera, tomato growth

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Authors: Itji Diana Daud, Nuniek Widiayani

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The fungus Beauveria bassiana can be endophytic in maize. The fungus was recovered in culture from stems, leaves and roots after a month planting. This phenomenon was shown until the third generation of the corn. The result from laboratory shows that B. bassiana appear in F1, F2 and F3 in order 70, 80 and 90% in the roots, 80% in the stems in all generation, 90, 80 and 70% in leaves. In CFU’s ml-1 of B. bassiana in corn seed, show F1 was 8.9 x 106, F2 was 8.1 x 106 and F3 was 7.8 x 106. The research showed that B. Bassiana as endophyte still remain to the third generation. Innovation to the corn seed which is endophyte seed is essential to protect from the attack of corn borer and to avoid the usage of insecticide.

Keywords: endophytic, recovered, third generation, Beauveria bassiana

Procedia PDF Downloads 248

Authors: Sadhana Shukla, Pushplata Singh, Nidhi Didwania

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Arbuscular mycorrhizal (AM) fungi are a highly advantageous and versatile group of fungi that significantly contribute to the formation of soil organic matter by creating a demand for plant carbon (C) and distributing it through below-ground hyphal biomass, regardless of their substantial contribution in enhancing net primary productivity and accumulating additional photosynthetic fixed C in the soil. The genetic role of AM fungi in carbon cycling is largely unexplored. In our study, we propose that AM fungi significantly interact with the soil, particularly: the provision of photosynthates by plants. We have studied the expression of AM fungi genes involved in CO₂ sequestration during host-plant interaction was investigated by qPCR studies. We selected Rhizophagus proliferus (AM fungi) and Oryza sativa (Rice) (inoculated with or without 200ppg AMF inoculums per plant) and investigated the effect of AM fungi on soil organic carbon (SOC) and rice growth under field conditions. Results thus provided faster SOC turnover, 35% increased nutrient uptake in plants and pronounced hyphal biomass of AM fungi which enhanced soil carbon storage by 15% in comparison to uninoculated plants. This study will offer a foundation for delving into various carbon-soil studies while also advancing our comprehension of the relationship between AM fungi and the sustainability of agricultural ecosystems.

Keywords: arbuscular mycorrhizal (AM) fungi, carbon sequestration, gene expression, soil health, plant development.

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Authors: Njabe Christelle

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Fungal products are essential building blocks for change towards a more sustainable future for our planet. In nature, fungi are special in breaking down plant material by means of a rich spectrum of plant cell wall degrading enzymes. Enzymes serve as catalysts in organic synthesis. Imagine the immense benefits that the known 250000 plant genes might provide in the future through scientific investigation. Plants are the primary basis for human sustenance, used directly for food, clothing, and shelter or indirectly in processed form and through animal feeding. Fungi are the only organisms known to extensively degrade lignin, a major component of wood. Although humans cannot digest cellulose and lignin, many fungi, through their assimilation of these substances, produce food in the form of edible mushrooms.

Keywords: plants, fungi, sustainable use, planet earth

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Authors: Ashraf Y. Z. Khalifa, Mohammed A. Almalki

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Medicago sativa (Alfalfa) is an important forage crop legume worldwide including Saudia Arabia due to its high nutritive value. Soil bacteria exist in root or root-nodules of Medicago sativa in either symbiotic relationships or in associations. The aim of the present study was to isolate and characterize endophytic bacteria that live in association with non-nodulated roots of Medicago sativa growing in Al-Ahsaa region, Saudia Arabia. Several bacterial strains were isolated from sterilized roots of Medicago sativa. Strains were characterized using 16S rRNA gene sequences, phylogenetic relationships analysis, morphological and biochemical characteristics. The strains utilized 50% (10 out of 20) of the different chemical substrates contained in the API20E strip. In general, many strains had the ability to ferment/oxidise all the carbohydrate tested except for rhamnose and the polyol carbohydrate, inositol. Comparative sequence analysis of the 16S rDNA gene indicated that the strains were closely related to the genus Bacillus. Furthermore, the growth parameters of Vigna sinensis were enhanced upon single-inoculation of the isolated strains, compared to the uninoculated control plants. The results highlighted that the root-nodules of Medicago sativa harbor non-nodulating bacterial strains that could have significant agricultural applications.

Keywords: Medicago sativa, endophytic bacteria, Pisum sativum, Vigna sinensis

Procedia PDF Downloads 346

Authors: Dahliz Abderrahmène, Lakhdari Wassim, Bouchikh Yamina, Hammi Hamida, Soud Adila, M’lik Randa, Benglia Sara

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Devastating insects constitute one of strains for cultivate tomato. Among this vandal insects, the tomato leafminer (T. absoluta), which has been introduced in Algeria constitute a challenge for both agricultures and scientists. Firstly, this insect is introduced without their natural enemies which may reduce their damage. Secondly, this species has developed insecticide resistance to many active matters. To contribute to establish a control strategy for T. absoluta we have mad an inventory for their enthomopathogenic fungi. Two fungi were identified among others taken from adults and pupae. These fungi are Aspergillus flavus and Metarhizium sp. A study was conducted in laboratory to recognize the efficiency of these antagonists. These species had unregistered a mortality mounts of 42% and 56% respectively.

Keywords: Tuta absoluta, enthomopathogenic fungi, Aspergillus flavus, Metarhizium sp, control strategy

Procedia PDF Downloads 416

Authors: H. Necef, A. Benayad

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Fungi are becoming more and more important in our life. Therefore, as a start for the symposium on filamentous fungi in biotechnology a short survey of the role of fungi in biotechnology. Salin soils occupy about 7% of land area; they are characterized by unsuitable physical conditions for the growth of living organisms. However, researches showed that some microorganisms especially fungi are able to grow and adapt to such extreme conditions; it is due to their ability to develop different physiological mechanisms in their adaptation. This is the first study on the physiological and biological characteristics of El-Beida marsh. Nine soil samples were taken at different points in two steps, the first was in winter (low temperature), and the second was in summer (high temperature). The physicochemical analyses of the soil were conducted, then the isolation process was applied using two methods, direct method and dilution method (10-1, 10-2, 10-3, 10-4). Different species of fungi were identified belong to 21 genera in addition to 3 yeast species, Aspergillus showed the highest proportion by 43%, then Penicillium by 20% then Alternaria by 7%, in addition to various genera in different proportions. As for the sampling periods, it was observed that the spread of fungi in winter was higher than in summer with the proportion 75.47% and 24.53% respectively. Some halotolerant fungi have a biotechnological importance especially if the salinity of the medium is necessary for the fermentation, and if the halotolerance genes of the fungus will define, this will open the research to study and improve this property for the industrial important micro-organisms.

Keywords: salinity, identification, aspergillus oryzae, halotolerance, fungi

Procedia PDF Downloads 370

Authors: Muhammad Naveed, Sohail Yousaf, Zahir Ahmad Zahir, Birgit Mitter, Angela Sessitsch

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Most bacterial endophytes originate from the soil and enter plants via the roots followed by further spread through the inner tissues. The mechanisms allowing bacteria to colonize plants endophytically are still poorly understood for most bacterial and plant species. Specific bacterial functions are required for plant colonization, but also the plant itself is a determining factor as bacterial ability to establish endophytic populations is very often dependent on the plant genotype (cultivar) and inoculums density. The effect of inoculum density (107, 108, 109 CFU mL-1) of Burkholderia phytofirmans strain PsJN was evaluated on growth and endophytic colonization of different maize and potato cultivars under axenic and natural soil conditions. PsJN inoculation significantly increased maize seedling growth and tuber yield of potato at all inoculum density compared to uninoculated control. Under axenic condition, PsJN inoculation (108 CFU mL-1) significantly improved the germination, root/shoot length and biomass up to 62, 115, 98 and 135% of maize seedling compared to uninoculated control. In case of potato, PsJN inoculation (109 CFU mL-1) showed maximum response and significantly increased root/shoot biomass and tuber yield under natural soil condition. We confirmed that PsJN is able to colonize the rhizosphere, roots and shoots of maize and potato cultivars. The endophytic colonization increased linearly with increasing inoculum density (within a range of 8 x 104 – 3 x 107 CFU mL-1) and were highest for maize (Morignon) and potato (Romina) as compared to other cultivars. Efficient colonization of cv. Morignon and Romina by strain PsJN indicates the specific cultivar colonizing capacity of the bacteria. The findings of the study indicate the non-significant relationship between colonization and plant growth promotion in maize under axenic conditions. However, the inoculum level (109 CFU mL-1) that promoted colonization of rhizosphere and plant interior (endophytic) also best promoted growth and tuber yield of potato under natural soil conditions.

Keywords: crop genotype, inoculum density, Burkholderia phytofirmans PsJN, colonization, growth, potato

Procedia PDF Downloads 447

Authors: Ifueko Oghogho Ukponmwan, Mike O. Orji

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This study investigated the antifungal activity of Bacilluss subtilis in the control of postharvest fungal rot of white yam (Dioscorea spp). Bacillus subtilis was isolated from the soil and fungi (Aspergillus spp, Mucor and yeasts) were isolated from rotten yam. The organisms were paired in yam nutrient agar (YNA) and yam Sabourraud dextrose agar media. In the yam dextrose agar media (YSDA) plates, the Bacillus grew rapidly and established itself and restricted the growth of the fungi organisms, but there was no zone of inhibition. This behaviour of Bacillus on the plates of YSDA was also observed in the yams where the fungi caused rot but the rot was suppressed by the presence of the Bacillus as compared to the degree of rot observed in the control that had only spoilage fungi. The control yam showed greater rot than other yams that contained a combination of Bacillus and fungi. The t-Test analysis showed that the difference in the rot between the treated samples and the control sample is significant and this implies that the presence of Bacillus significantly reduced the growth of fungi in the samples (yams). It was revealed from this study that Bacillus subtilis treatment can be successfully used to preserve white yams in storage. Its fast growth and early establishment in the sample accounts for its antifungal strength.

Keywords: Bacillus subtilis, rot, fungi, yam

Procedia PDF Downloads 151

Authors: Gabriel Roveda-Hoyos, Margarita Ramirez-Gomez, Adrian Perez, Diana Paola Serralde

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The objective of this study was to determine the variability of arbuscular mycorrhizal fungi (HFMA) communities associated with wild agraz plants (Vaccinium meridionale Swartz) in the Colombian Andes. This species is one of the most promising fruits within the genus Vaccinium because of the high content of anthocyanins and antioxidants in its fruits, and like other species of the Ericaceae family, it depends on the association with HFM for its development in the natural environment. In this study, the presence of mycorrhizae in wild communities of V. meridionale was evaluated, and their relationship with the edaphic and climatic conditions of the study area was analyzed. Sampling was conducted in the rural area of the municipalities of Raquira, and Chiquinquira, Chia, and Tabio in the departments of Cundinamarca and Boyaca, Colombia. Seven sites were selected, and in each site, 5 plants were randomly selected, root and soil samples were taken from each plant in the rhizosphere zone for the quantification of colonization and the presence of spores. The samples were collected on different soils, taxonomic orders Entisols, Inceptisols, and Alfisols, located at altitudes between 2,600 and 3,000 above sea level in the Eastern Cordillera of Colombia. The physicochemical characteristics of the soil were compared with the density of spores and the percentage of presence of mycorrhizae in the roots and variables with the morphometric and physiological characteristics of the plants. Four types of mutual associations were found: arbuscular mycorrhizae, ectendomycorrhiza, ericoid mycorrhizae, and endophytic septate fungi. The main results obtained show a predominance of spores of the genera Glomus and Acaulsopora, in most of the soils analyzed. The spore density of Glomeromycete fungi in the soil varied considerably between the different sites; it was higher ( > 50 spores/g of dry soil) in soil samples with lower bulk density and higher content of organic matter; in these soils a higher cation exchange capacity was found, as well as of nitrogen, calcium, magnesium, manganese and zinc concentration. It can be concluded that Vaccinium meridionale is able to establish in a natural way, association with HFMA.

Keywords: Ericaceae, Arbuscular mycorrhizae, Andes, soils, Glomus sp.

Procedia PDF Downloads 148

Authors: H. Heydari, B. Konuklugil, P. Proksch

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Marine-derived fungi can produce interesting bioactive secondary metabolites that can be considered the potential for drug development. Turkey is a country of a peninsula surrounded by the Black Sea at the north, the Aegean Sea at the west, and the Mediterranean Sea at the south. Despite the approximately 8400 km of coastline, studies on marine secondary metabolites and their biological activity are limited. In our ongoing search for new natural products with different bioactivities produced by the marine-derived fungi, we have investigated secondary metabolites of Turkish collection of the marine sea slug (Peltodoris atromaculata) associated fungi Hypocrea nigricans collected from Seferihisar in the Egean sea. According to the author’s best knowledge, no study was found on this fungal species in terms of secondary metabolites. Isolated from ethyl acetate extract of the culture of Hypocrea nigricans were (isodihydroauroglaucin,tetrahydroauroglaucin and dihydroauroglaucin. The structures of the compounds were established based on an NMR and MS analysis. Structural elucidation of another isolated secondary metabolite/s continues.

Keywords: Hypocrea nigricans, isolation, marine fungi, secondary metabolites

Procedia PDF Downloads 131

Authors: W. Lakhdari, B. Doumendji-Mitich, A. Dahliz, S. Doumendji, Y. Bouchikh, R. M'lik, H. Hammi, A. Soud

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This work was done in order to fight against Brachytrupes megacephalus, a major pest in the Algerian oasis and promote one aspect of biological control against it. He wears a hand on the isolation and identification of indigenous fungi on imagos of this insect harvested in the station of INRAA Touggourt and secondly, the study of the pathogenicity of these strains fungal on this orthoptère adults. The results obtained showed the presence of six different species of entomopathogenic fungi, it is: Aspergillus flavus, Fusarium sp, Beauveria bassiana, Penicillium sp, Metharizium anisopliae and Aspergillus Niger. The pathogenicity test using fungi Beauveria bassiana strains and Metharizium anisopliae. On adult of B. megacephalus highlights the effectiveness of these strains of predatory adults, with a mortality rate approaching 100% after 11 days.

Keywords: biological control, brachytrupes megacephalus, entomopathogenic fungi, Southeastern Algeria

Procedia PDF Downloads 386