Endophytic fungi diversity of aquatic/riparian plants and their antifungal activity <Emphasis Type="Italic">in vitro</Emphasis>

Two hundred and fourteen endophytic fungi were isolated from 500 segments of aquatic/riparian plants Ottelia acuminata, Myriophyllum verticillatum, Equisetum arvense, Cardamine multijuga, and Impatiens chinensis. They were identified to 31 taxa in which Cladosporium, Fusarium, and Geotrichum were the dominant genera. Among all isolates, 169 (79%) were anamorphic fungi, 1 (0.5%) was an teleomorphic ascomycete and 44 (21%) were sterile mycelia. There were significant differences in the colonization frequency of endophytes between the five plant species (X∼2=51.128, P<0.001, Chi-square test). The riparian plants harboured more endophytes than the submerged plants. The antifungal activity of these isolates against Fusarium solani and Phytophthora nicotianae in vitro were tested and 28 (13.1%) isolates showed antifungal activities with more than 30% growth inhibition rate against the two pathogens.     

Isolation and identification of endophytic bacteria from root tissues of Salvia miltiorrhiza Bge. and determination of their bioactivities

Endophytic bacteria are microorganisms that live in host plants, but do not cause diseases to the hosts. This study examined the occurrence, distribution, growth-promoting and antifungal activities of endophytes in the root of Salvia miltiorrhiza Bge. Six endophytic bacterial strains, which belong to genera of Pseudomonas, Rhizobium, Bacillus and Novosphingobium, were isolated from the root of healthy S. miltiorrhiza. Cell suspension (approx. 10⁹ cell?·?ml^?1) of two isolates and cell-free fermentation filtrate of four isolates substantially promoted the growth of hypocotyl and radicle of muskmelon seeds. The cell-free fermentation filtrate of six isolates had no inhibiting effect on tested pathogenic fungi, namely Fusarium solani, F. oxysporum f. sp. vasinfectum and F. oxysporum. Six compounds were isolated from one of the six endophytic bacteria, namely, Bacillus aryabhattai, and two of these compounds displayed certain antifungal activity against three tested S. miltiorrhiza pathogens. Our work indicates that endophytic bacteria occur in the root of S. miltiorrhiza, and that associated bacterial isolates have growth-promoting effect on muskmelon seeds and are expected to be a potential source for bioactive metabolites.     

Culturable Endophytes Associated with Soybean Seeds and Their Potential for Suppressing Seed-Borne Pathogens

Seed-borne pathogens in crops reduce the seed germination rate and hamper seedling growth, leading to significant yield loss. Due to the growing concerns about environmental damage and the development of resistance to agrochemicals among pathogen populations, there is a strong demand for eco-friendly alternatives to synthetic chemicals in agriculture. It has been well established during the last few decades that plant seeds harbor diverse microbes, some of which are vertically transmitted and important for plant health and productivity. In this study, we isolated culturable endophytic bacteria and fungi from soybean seeds and evaluated their antagonistic activities against common bacterial and fungal seed-borne pathogens of soybean. A total of 87 bacterial isolates and 66 fungal isolates were obtained. Sequencing of 16S rDNA and internal transcribed spacer amplicon showed that these isolates correspond to 30 and 15 different species of bacteria and fungi, respectively. Our antibacterial and antifungal activity assay showed that four fungal species and nine bacterial species have the potential to suppress the growth of at least one seed-borne pathogen tested in the study. Among them, Pseudomonas koreensis appears to have strong antagonistic activities across all the pathogens. Our collection of soybean seed endophytes would be a valuable resource not only for studying biology and ecology of seed endophytes but also for practical deployment of seed endophytes toward crop protection.     

Endophytic bacteria enhancing growth and disease resistance of potato (Solanum tuberosum L.)

Priming plants by non-pathogenic bacteria allows the host to save energy and to reduce time needed for development of defense reaction during a pathogen attack. However, information on the role of endophytes in plant defense is limited. Here, the ability of endophytic bacteria to promote growth and resistance of potato plants towards infection by the necrotroph Pectobacterium atrosepticum was studied. A Pseudomonas sp. strain was selected due to antagonism towards bacterial pathogens and a Methylobacterium sp. strain because of efficient plant colonization. The aim of this study was to find if there is any correlation between plant growth promotion and induction of resistance by endophytes of potato, as well as to study the putative mechanisms of endophytes interacting with the plant during resistance induction. Both tested strains promoted growth of potato shoots but only the Pseudomonas sp. increased potato resistance towards the soft rot disease. Induction of disease resistance by the Methylobacterium sp. was inversely proportional to the size of bacterial population used for inoculation. The plant antioxidant system was moderately activated during the induction of resistance by the biocontrol strains. qPCR data on expression of marker genes of induced systemic resistance and acquired systemic resistance in endophyte-infected Arabidopsis plants showed activation of both salicylic acid and jasmonate/ethylene-dependent pathways after challenge inoculation with the pathogen. We suggest that some endophytes have the potential to activate both basal and inducible plant defense systems, whereas the growth promotion by biocontrol strains may not correlate with induction of disease resistance.     

Biocontrol potential of endophytes harbored in Radula marginata (liverwort) from the New Zealand ecosystem

Radula marginata and Cannabis sativa L. are two phylogenetically unrelated plant species containing structurally similar secondary metabolites like cannabinoids. The major objective of our work was the isolation, identification, biocontrol efficacies, biofilm forming potential and anti-biofilm ability of endophytic microbial community of the liverwort R. marginata, as compared to bacterial endophytic isolates harbored in C. sativa plants. A total of 15 endophytic fungal and 4 endophytic bacterial isolates were identified, including the presence of a bacterial endosymbiont within an endophytic fungal isolate. The endosymbiont was visible only when the fungus containing it was challenged with two phytopathogens Botrytis cinerea and Trichothecium roseum, highlighting a tripartite microbe–microbe interaction and biocontrol potency of endophytes under biotic stress. We also observed sixteen types of endophytic fungal-pathogen and twelve types of endophytic bacterial-pathogen interactions coupled to varying degree of growth inhibitions of either the pathogen or endophyte or both. This showed the magnitude of biocontrol efficacies of endophytes in aiding plant fitness benefits under different media (environmental) conditions. Additionally, it was ecologically noteworthy to find the presence of similar endophytic bacterial genera in both Radula and Cannabis plants, which exhibited similar functional traits like biofilm formation and general anti-biofilm activities. Thus far, our work underlines the biocontrol potency and defensive functional traits (in terms of antagonism and biofilm formation) of endophytes harbored in liverwort R. marginata as compared to the endophytic community of phylogenetically unrelated but phytochemically similar plant C. sativa.     

Response of Endophytic Bacterial Communities in Potato Plants to Infection with Erwinia carotovora subsp. atroseptica

The term endophyte refers to interior colonization of plants by microorganisms that do not have pathogenic effects on their hosts, and various endophytes have been found to play important roles in plant vitality. In this study, cultivation-independent terminal restriction fragment length polymorphism analysis of 16S ribosomal DNA directly amplified from plant tissue DNA was used in combination with molecular characterization of isolates to examine the influence of plant stress, achieved by infection with the blackleg pathogen Erwinia carotovora subsp. atroseptica, on the endophytic population in two different potato varieties. Community analysis clearly demonstrated increased bacterial diversity in infected plants compared to that in control plants. The results also indicated that the pathogen stress had a greater impact on the bacteria population than the plant genotype had. Partial sequencing of the 16S rRNA genes of isolated endophytes revealed a broad phylogenetic spectrum of bacteria, including members of the α, β, and γ subgroups of the Proteobacteria, high- and low-G+C-content gram-positive organisms, and microbes belonging to the Flexibacter-Cytophaga-Bacteroides group. Screening of the isolates for antagonistic activity against E. carotovora subsp. atroseptica revealed that 38% of the endophytes protected tissue culture plants from blackleg disease.     

Community structure,spatial distribution,diversity and functional characterization of culturable endophytic fungi associated with Glycyrrhiza glabra L.

A total of 266 endophytic fungal isolates were recovered from 1019 tissue segments of Glycyrrhiza glabra collected from four different locations in the North-Western Himalayas. The endophytes grouped into 21 genera and 38 different taxa. The host had strong affinity for the genus Phoma, followed by Fusarium. The species richness was highest at the sub-tropical location, followed by the sub-temperate location and the temperate locations, respectively. The tissue specificity of endophytes was also evident. Some endophytes showed potential antimicrobial activity against phyto-pathogens indicating that they may be helpful to the host in evading pathogens. All the endophytic taxa produced the plant growth promoting hormone, indole acetic acid (IAA), though in varying concentrations. None of these endophytes caused any symptoms of disease in co-cultivation with the tissue cultured plants. Further, all the endophytes had a positive influence on the phenolic and flavonoid content of the host. Three endophytes, Stagonosporopsis cucurbitacearum, Bionectria sp. and Aspergillus terreus also increased the host root (rhizome) and shoot growth visibly. Such endophytes are potential candidates for developing endophyte-based technologies for sustainable cultivation and enhanced productivity of G. glabra. This is the first report of community structure and biological properties of fungal endophytes associated with G. glabra.     

Endophytic fungi from the root tubers of medicinal plant Stephania dielsiana and their antimicrobial activity

The adverse effects of chemical synthetic fungicides on agricultural fields and the environment are driving a need to search for safer and less environmentally harmful plant protectants to move toward more sustainable development of agriculture. The endophytic fungal community associated with the medicinal plant Stephania dielsiana, and its potential for providing antimicrobial secondary metabolites were investigated. A total of 26 isolates of endophytic fungi were obtained, and 21 isolates were identified and classified into eight different genera, including Briansuttonomyces, Glomerella, Pleosporales, Diaporthe, Phoma, Penicillium, Periconia and Colletotrichum, and the most frequent endophytic species obtained were Diaporthe phaseolorum, Penicillium sp., Periconia igniari and Colletotrichum sp. The ethyl acetate (EtOAc) extract of the endophytic fungus Diaporthe phaseolorum Stdif6 displayed the most significant antifungal activity against all tested phytopathogens, with EC₅₀ values ranging from 0.0138 to 0.3103 mg/mL. While the EtOAc extract of the endophytic fungus Penicillium sp. Stdif9 exhibited greater potential for antibacterial activity, with the minimum inhibitory concentration (MIC) values against seven bacteria ranging from 1.25 to 6 mg/mL. The remarkable antimicrobial activity of fungal endophytes suggests that fungal endophytes harbored inside the root tubers of S. dielsiana hold great promise as biocontrol agents against a broad spectrum of economically significant pathogens.     

Isolation and characterization of plant growth promoting endophytic diazotrophic bacteria from Korean rice cultivars

We have isolated 576 endophytic bacteria from the leaves, stems, and roots of 10 rice cultivars and identified 12 of them as diazotrophic bacteria using a specific primer set of nif gene. Through 16S rDNA sequence analysis, nifH genes were confirmed in the two species of Penibacillus, three species of Microbacterium, three Bacillus species, and four species of Klebsiella. Rice seeds treated with these plant growth-promoting bacteria (PGPB) showed improved plant growth, increased height and dry weight and antagonistic effects against fungal pathogens. In addition, auxin and siderophore producing ability, and phosphate solubilizing activity were studied for the possible mechanisms of plant growth promotion. Among 12 isolates tested, 10 strains have shown higher auxin producing activity, 6 isolates were confirmed as strains with high siderophore producing activity while 4 isolates turned out to have high phosphate-solubilizing activity. These results strongly suggest that the endophytic diazotrophic bacteria characterized in this study could be successfully used to promote plant growth and inducing fungal resistance in plants.     

Fungal endophytes of turmeric (<Emphasis Type="Italic">Curcuma longa</Emphasis> L.) and their biocontrol potential against pathogens <Emphasis Type="Italic">Pythium aphanidermatum</Emphasis> and <Emphasis Type="Italic">Rhizoctonia solani</Emphasis>

Endophytic fungi have been isolated from the healthy turmeric (Curcuma longa L.) rhizomes from South India. Thirty-one endophytes were identified based on morphological and ITS–rDNA sequence analysis. The isolated endophytes were screened for antagonistic activity against Pythium aphanidermatum (Edson) Fitzp., and Rhizoctonia solani Kuhn., causing rhizome rot and leaf blight diseases in turmeric respectively. Results revealed that only six endophytes showed >?70% suppression of test pathogens in antagonistic dual culture assays. The endophyte T. harzianum TharDOB-31 showed significant in vitro mycelial growth inhibition of P. aphanidermatum (76.0%) and R. solani (76.9%) when tested by dual culture method. The SEM studies of interaction zone showed morphological abnormalities like parasitism, shriveling, breakage and lysis of hyphae of the pathogens by endophyte TharDOB-31. Selected endophytic isolates recorded multiple plant growth promoting traits in in vitro studies. The rhizome bacterization followed by soil application of endophyte TharDOB-31 showed lowest Percent Disease Incidence of rhizome rot and leaf blight, 13.8 and 11.6% respectively. The treatment of TharDOB-31 exhibited significant increase in plant height (85 cm) and fresh rhizome yield/plant (425 g) in comparison with untreated control under greenhouse condition. The confocal microscopy validates the colonization of the TharDOB-31 in turmeric rhizomes. The secondary metabolites in ethyl acetate extract of TharDOB-31 were found to contain higher number of antifungal compounds by high resolution liquid chromatograph mass spectrometer analysis. Thereby, endophyte T. harzianum isolate can be exploited as a potential biocontrol agent for suppressing rhizome rot and leaf blight diseases in turmeric.     

Thaxtomin A-deficient endophytic Streptomyces sp. enhances plant disease resistance to pathogenic Streptomyces scabies

Each plant species in nature harbors endophytes, a community of microbes living within host plants without causing any disease symptom. However, the exploitation of endophyte-based phytoprotectants is hampered by the paucity of mechanistic understandings of endophyte-plant interaction. We here reported two endophytic Streptomyces isolates IFB-A02 and IFB-A03 recovered from a stress-tolerant dicotyledonous plant Artemisia annua L. After the determination of their non-pathogenicity at the genomic level and from the toxin (thaxtomin A, TXT) level, the endophytism of both isolates was supported by their successful colonization in planta. Of the two endophytes, IFB-A03 was further studied for the mechanism of endophyte-conferred phytoprotection owing to its plant growth promotion in model eudicot Arabidopsis thaliana. Using the endophyte-Arabidopsis co-cultivation system into which pathogenic Streptomyces scabies was introduced, we demonstrated that IFB-A03 pre-inoculation could activate the salicylic acid (SA)-mediated plant defense responses upon pathogen challenge. Moreover, IFB-A03 was shown to partially rescue the defense deficiency in eds5 (enhanced disease susceptibility 5) Arabidopsis mutants, putatively acting at the upstream of SA accumulation in the defense signaling pathway associated with the systemic acquired resistance (SAR). These data suggest that endophytic Streptomyces sp. IFB-A03 could be a promising candidate for biocontrol agents against S. scabies—a causative pathogen of common scab diseases prevailing in agronomic systems.     

In Vitro and In Vivo Plant Growth Promoting Activities and DNA Fingerprinting of Antagonistic Endophytic Actinomycetes Associates with Medicinal Plants

Endophytic actinomycetes have shown unique plant growth promoting as well as antagonistic activity against fungal phytopathogens. In the present study forty-two endophytic actinomycetes recovered from medicinal plants were evaluated for their antagonistic potential and plant growth-promoting abilities. Twenty-two isolates which showed the inhibitory activity against at least one pathogen were subsequently tested for their plant-growth promoting activities and were compared genotypically using DNA based fingerprinting, including enterobacterial repetitive intergenic consensus (ERIC) and BOX repetitive elements. Genetic relatedness based on both ERIC and BOX-PCR generates specific patterns corresponding to particular genotypes. Exponentially grown antagonistic isolates were used to evaluate phosphate solubilization, siderophores, HCN, ammonia, chitinase, indole-3-acetic acid production, as well as antifungal activities. Out of 22 isolates, the amount of indole-3-acetic acid (IAA) ranging between 10–32 μg/ml was produced by 20 isolates and all isolates were positive for ammonia production ranging between 5.2 to 54 mg/ml. Among 22 isolates tested, the amount of hydroxamate-type siderophores were produced by 16 isolates ranging between 5.2 to 36.4 μg/ml, while catechols-type siderophores produced by 5 isolates ranging from 3.2 to 5.4 μg/ml. Fourteen isolates showed the solubilisation of inorganic phosphorous ranging from 3.2 to 32.6 mg/100ml. Chitinase and HCN production was shown by 19 and 15 different isolates, respectively. In addition, genes of indole acetic acid (iaaM) and chitinase (chiC) were successively amplified from 20 and 19 isolates respectively. The two potential strains Streptomyces sp. (BPSAC34) and Leifsonia xyli (BPSAC24) were tested in vivo and improved a range of growth parameters in chilli (Capsicum annuum L.) under greenhouse conditions. This study is the first published report that actinomycetes can be isolated as endophytes from within these plants and were shown to have antagonistic and plant growth promoting abilities. These results clearly suggest the possibility of using endophytic actinomycetes as bioinoculant for plant growth promotion, nutrient mobilization or as biocontrol agent against fungal phytopathogens for sustainable agriculture.     

Diversity of Culturable Plant Growth-Promoting Bacterial Endophytes Associated with Sugarcane Roots and Their Effect of Growth by Co-Inoculation of Diazotrophs and Actinomycetes

Application of environmentally friendly agents to reduce the use of chemicals and to enhance growth of plants is an ultimate goal of sustainable agriculture. The use of plant growth-promoting endophytes has become of great interest as a way to enhance plant growth and additionally protect plants from phytopathogens. In this study, 135 isolates of endophytic bacteria including actinomycetes were isolated from roots of commercial sugarcane plants cultivated in Thailand and were characterized for plant growth-promoting (PGP) traits. Based on morphological and 16S rRNA sequence analysis, the endophytes were distributed into 14 genera of which the most dominant species belong to Bacillus, Enterobacter, Microbispora, and Streptomyces. Two strains of endophytic diazotrophs, Bacillus sp. EN-24 and Enterobacter sp. EN-21; and two strains of actinomycetes, Microbispora sp. GKU 823 and Streptomyces sp. GKU 895, were selected based on their PGP traits including 1-aminocyclopropane-1-decarboxylate deaminase, indole-3-acetic acid, nitrogen fixation, phosphate solubilization, and siderophore production for evaluation of sugarcane growth enhancement by individual and co-inoculation. Sixty days after co-inoculation by endophytic diazotrophs and actinomycetes, the growth parameters of sugarcane plants were significantly greater than that of individual and un-inoculated plants. The results indicated that these endophytes have high potential as PGP agents that could be applied to promote sugarcane growth and could be developed as active added value biofertilizers in the future.     

Novel bacterial endophytes from Hevea brasiliensis as biocontrol agent against Phytophthora leaf fall disease

Bacterial endophytes offer control against many diseases of crop plants as potential biocontrol agents. Antagonistic bacterial endophytes acting against Phytophthora meadii have been screened from leaf, petiole and root tissues of Hevea brasiliensis. Six bacterial endophytes could exhibit more than 50 % inhibition of P. meadii, among which EIL-2, from disease-free zones showed a maximum of 62.5 % inhibition. The isolate EIL-2 was characterized as Alcaligenes sp. and the other isolates were identified as Pseudomonas aeruginosa. 16S rDNA sequence analysis showed that there existed genetic variation among the five isolates of P. aeruginosa from different tissues of the plant indicating the tissue type adaptation of the isolates. Dual culture technique with endophyte EIL-2 completely arrested the growth of P. meadii when inoculated prior to pathogen. The bioassay with EIL-2 in H. brasiliensis clones, RRII 105 showed 43 % reduction of lesion size on infected leaves whereas in RRIM 600 it was only 30 %.     

Methylobacterium-Induced Endophyte Community Changes Correspond with Protection of Plants against Pathogen Attack

Plant inoculation with endophytic bacteria that normally live inside the plant without harming the host is a highly promising approach for biological disease control. The mechanism of resistance induction by beneficial bacteria is poorly understood, because pathways are only partly known and systemic responses are typically not seen. The innate endophytic community structures change in response to external factors such as inoculation, and bacterial endophytes can exhibit direct or indirect antagonism towards pathogens. Earlier we showed that resistance induction by an endophytic Methylobacterium sp. in potato towards Pectobacterium atrosepticum was dependent on the density of the inoculum, whereas the bacterium itself had no antagonistic activity. To elucidate the role of innate endophyte communities in plant responses, we studied community changes in both in vitro and greenhouse experiments using various combinations of plants, endophyte inoculants, and pathogens. Induction of resistance was studied in several potato (Solanum tuberosum L.) cultivars by Methylobacterium sp. IMBG290 against the pathogens P. atrosepticum, Phytophthora infestans and Pseudomonas syringae pv. tomato DC3000, and in pine (Pinus sylvestris L.) by M. extorquens DSM13060 against Gremmeniella abietina. The capacities of the inoculated endophytic Methylobacterium spp. strains to induce resistance were dependent on the plant cultivar, pathogen, and on the density of Methylobacterium spp. inoculum. Composition of the endophyte community changed in response to inoculation in shoot tissues and correlated with resistance or susceptibility to the disease. Our results demonstrate that endophytic Methylobacterium spp. strains have varying effects on plant disease resistance, which can be modulated through the endophyte community of the host.     

Fungal Endophyte Diversity and Bioactivity in the Indian Medicinal Plant Ocimum sanctum Linn

Endophytic mycopopulation isolated from India’s Queen of herbs Tulsi (Ocimum sanctum) were explored and investigated for their diversity and antiphytopathogenic activity against widespread plant pathogens Botrytis cinerea, Sclerotinia sclerotiorum, Rhizoctonia solani and Fusarium oxysporum. 90 fungal isolates, representing 17 genera were recovered from 313 disease-free and surface sterilised plant segments (leaf and stem tissues) from three different geographic locations (Delhi, Hyderabad and Mukteshwar) during distinct sampling times in consequent years 2010 and 2011 in India. Fungal endophytes were subjected to molecular identification based on rDNA ITS sequence analysis. Plant pathogens such as F. verticillioides, B. maydis, C. coarctatum, R. bataticola, Hypoxylon sp., Diaporthe phaseolorum, Alternaria tenuissima and A. alternata have occurred as endophyte only during second sampling (second sampling in 2011) in the present study. Bi-plot generated by principal component analysis suggested tissue specificity of certain fungal endophytes. Dendrogram revealed species abundance as a function of mean temperature of the location at the time of sampling. Shannon diversity in the first collection is highest in Hyderabad leaf tissues (H'' = 1.907) whereas in second collection it was highest from leaf tissues of Delhi (H'' = 1.846). Mukteshwar (altitude: 7500 feet) reported least isolation rate in second collection. Nearly 23% of the total fungal isolates were considered as potent biocontrol agent. Hexane extract of M. phaseolina recovered from Hyderabad in first collection demonstrated highest activity against S. sclerotiorum with IC₅₀ value of 0.38 mg/ml. Additionally, its components 2H-pyran-2-one, 5,6-dihydro-6-pentyl and palmitic acid, methyl ester as reported by GC-MS Chromatogram upon evaluation for their antiphytopathogenic activity exhibited IC₅₀ value of 1.002 and 0.662 against respectively S. sclerotiorum indicating their significant role in antiphytopathogenic activity of hexane extract. The production of 2H-pyran-2-one, 5,6-dihydro-6-pentyl from M. phaseolina, an endophytic fungus is being reported for the first time.     

Plant growth promoting potential of bacterial endophytes in novel association with <Emphasis Type="Italic">Olea ferruginea</Emphasis> and <Emphasis Type="Italic">Withania coagulans</Emphasis>

Microbially unexplored medicinal plants can have a genetically diverse microbial population with multi-functional plant growth promoting traits. In this aspect, 75 endophytic bacterial isolates with plant growth promoting traits were isolated from Withania coagulans Dunal and Olea ferruginea Royal. Many of these bacteria were able to solubilize phosphate, produce indole-3-acetic acid, ammonia as well as hydrogen cyanide, synthesize extracellular enzymes and show antagonistic activities against plant pathogenic fungi under in vitro conditions. These isolates were also characterized by morphological and biochemical analysis. Furthermore, four representative isolates with pronounced plant growth promoting activities were identified as Enterobacter cloacae, Enterobacter dissolvens, Enterobacter hormaechei and Cronobacter sakazakii by 16S rDNA sequencing analysis. This work for the first time, reported the isolation of endophytic bacteria, the novel association form selected plants, Withania coagulans and Olea ferruginea. The explored endophytes might have great potential in the field of biocontrol and plant growth promoting for sustainable agricultural practices.     

Fungal communities associated with species of Fraxinus tolerant to ash dieback,and their potential for biological control

Ash dieback, caused by the fungus Hymenoscyphus fraxineus, has threatened ash trees in Europe for more than two decades. However, little is known of how endophytic communities affect the pathogen, and no effective disease management tools are available. While European ash (Fraxinus excelsior) is severely affected by the disease, other more distantly related ash species do not seem to be affected. We hypothesise that fungal endophytic communities of tolerant ash species can protect the species against ash dieback, and that selected endophytes have potential as biocontrol agents. These hypotheses were tested by isolating members of the fungal communities of five tolerant ash species, and identifying them using ITS regions. Candidate endophytes were tested by an in vitro antagonistic assay with H.fraxineus. From a total of 196 isolates we identified 9 fungal orders, 15 families, and 40 species. Fungi in orders Pleosporales, such as Boeremia exigua and Diaporthe spp., and Hypocreales (e.g., Fusarium sp.), were recovered in most communities, suggesting they are common taxa. The in vitro antagonistic assay revealed five species with high antagonistic activity against H. fraxineus. These endophytes were identified based on ITS region as Sclerostagonospora sp., Setomelanomma holmii, Epicoccum nigrum, B. exigua and Fusarium sp. Three of these taxa have been described previously as antagonists of plant pathogenic microbes, and are of interest for future studies of their potential as biological control agents against ash dieback, especially for valuable ash trees in parks and urban areas.     

Biodiversity and antimicrobial potential of bacterial endophytes from halophyte Salicornia brachiata

Extreme natural habitats like halophytes, marsh land, and marine environment are suitable arena for chemical ecology between plants and microbes having environmental impact. Endophytes are an ecofriendly option for the promotion of plant growth and to serve as sustainable resource of novel bioactive natural products. The present study, focusing on biodiversity of bacterial endophytes from Salicornia brachiata, led to isolation of around 336 bacterial endophytes. Phylogenetic analysis of 63 endophytes revealed 13 genera with 27 different species, belonging to 3 major groups: Firmicutes, Proteobacteria, and Actinobacteria. 30% endophytic isolates belonging to various genera demonstrated broad-spectrum antibacterial and antifungal activities against a panel of human, plant, and aquatic infectious agents. An endophytic isolate Bacillus amyloliquefaciens 5NPA-1, exhibited strong in-vitro antibacterial activity against human pathogen Staphylococcus aureus and phytopathogen Xanthomonas campestris. Investigation through LC–MS/MS-based molecular networking and bioactivity-guided purification led to the identification of three bioactive compounds belonging to lipopeptide class based on ¹H-, ¹³C-NMR and MS analysis. To our knowledge, this is the first report studying bacterial endophytic biodiversity of Salicornia brachiata and the isolation of bioactive compounds from its endophyte. Overall, the present study provides insights into the diversity of endophytes associated with the plants from the extreme environment as a rich source of metabolites with remarkable agricultural applications and therapeutic properties.

     

Antagonistic bacteria of composted agro-industrial residues exhibit antibiosis against soil-borne fungal plant pathogens and protection of tomato plants from Fusarium oxysporum f.sp. radicis-lycopersici

Rhizospheric and root-associated/endophytic (RAE) bacteria were isolated from tomato plants grown in three suppressive compost-based plant growth media derived from the olive mill, winery and Agaricus bisporus production agro-industries. Forty-four (35 rhizospheric and 9 RAE) out of 329 bacterial strains showed in vitro antagonistic activity against at least one of the soil-borne fungal pathogens, Fusarium oxysporum f.sp. radicis-lycopersici (FORL), F. oxysporum f.sp. raphani, Phytophthora cinnamomi, P. nicotianae and Rhizoctonia solani. The high percentage of total isolates showing antagonistic properties (13%) and their common chitinase and β-glucanase activities indicate that the cell wall constituents of yeasts and macrofungi that proliferate in these compost media may have become a substrate that favours the establishment of antagonistic bacteria to soil-borne fungal pathogens. The selected bacterial strains were further evaluated for their suppressiveness to tomato crown and root rot disease caused by FORL. A total of six rhizospheric isolates, related to known members of the genera Bacillus, Lysinibacillus, Enterobacter and Serratia and one RAE associated with Alcaligenes faecalis subsp. were selected, showing statistically significant decrease of plant disease incidence. Inhibitory effects of extracellular products of the most effective rhizospheric biocontrol agent, Enterobacter sp. AR1.22, but not of the RAE Alcaligenes sp. AE1.16 were observed on the growth pattern of FORL. Furthermore, application of cell-free culture extracts, produced by Enterobacter sp. AR1.22, to tomato roots led to plant protection against FORL, indicating a mode of biological control action through antibiosis.