Endophytic Species Research Articles

FruiTemp: Design, Implementation and Analysis for an Open-Source Temperature Logger Applied to Fruit Fly Host Experimentation

FruiTemp is an open-source prototype developed to study the response of endophytic insect species such as fruit flies (Diptera: Tephrtidae) to variable temperature conditions including the controlled laboratory and fluctuating field settings. The system is a three-channel temperature sensor that consists of two precision thermistors that measure the temperature in the core and the surface of a fruit on a tree and a Harsh Environment thermistor that measures the air temperature surrounding the host at a rate of one measurement per 15 min. The sample rate can be adjusted according to the researcher’s needs. The system was successfully tested in field and laboratory experimental conditions using apples as the fruit model. The measurements on apples on trees lasted five consecutive days and produced a range of reliable data. After assessing statistical agreement and precision, the results revealed a differential bias of 0.331 °C and a proportional bias of a magnitude of 0.982. This work promotes open-source implementations allowing inexpensive solutions aiding experimentation procedures by significantly lowering operating costs.

Diversity and community structure of endophytic Bacillus with antagonistic and antioxidant activity in the fruits of Xisha Wild Noni (Morinda citrifolia L.)

Noni (Morinda citrifolia L.) is a tropical crop with strong antibacterial, antioxidant and other abilities, and its fruit has a strong potential for adjuvant treatment of diseases. This study aimed to explore the dynamic change of endophytic bacteria in Noni fruit at different stages and the correlation between the antagonistic and antioxidant activity of the Bacillus which was screened and the change of the host's growth stage. In this study, though the high-throughput sequencing technology (HTS), 106 endophytic bacteria species were found in A, B, C, D, E and F stages of Noni fruit, among which the dominant group were Pantoea (0.3%–20.9%), and Candidatus_Uzinura (2.3%–35.2%) etc. The endophytic bacteria were isolated by culture-dependent method. Through their antagonistic experiments on Staphylococcus aureus and Escherichia coli, the results of 16S polyphasic taxonomic identification showed that the 34 antagonistic strains belonged to Bacillus. Five species of these Bacillus were identified by gyrA polyphase taxonomy, including Bacillus subtilis (76% of all Bacillus), Bacillus licheniformis (9%), Bacillus amyloliquefaciens (6%), Bacillus velezensis (6%) and Bacillus mojavensi (3%), and the RAPD showed these Bacillus are no signs of stable passage. In C, D, E and F stages, the average total antioxidant activity of Bacillus endophytic antagonists against Noni was 7.812 U/mL, 8.144 U/mL, 7.817 U/mL and 7.144 U/mL, which was much higher than that of Noni fruit, and antioxidant activity of Noni juice and Bacillus bacterial liquid vary with host's growth period showed the same trend, both rose slowly at first, and reached the highest in period E, then declined slightly in period F, it showed that the antagonistic Bacillus of Noni had synergistic function with Noni fruit. This study clarified the relationship of function between Noni fruit and endophytic bacteria, and laid a foundation for future study on the dynamic change of endophytic flora succession and efficacy.

Bioremoval capacity of phenol by some selected endophytic fungi isolated from Hibiscus sabdariffa and batch biodegradation of phenol in paper and pulp effluents.

Background and Objectives:The use of endophytic fungi for management of phenol residue in paper and pulp industries has been shown as cost-effective and eco-friendly approach. In this study, isolation of endophytic fungi from roots, stems, and leaves of Hibiscus sabdariffa was conducted. Additionally, the isolated fungi were examined for their ability to degrade phenol and its derivatives in paper and pulp industrial samples, using different growth conditions.Materials and Methods:Out of 35 isolated endophyitc fungi, 31 were examined for their phenol biodegradation capacity using Czapek Dox broth medium containing Catechol and Resorcinol as a sole carbon source at final concentrations of 0.4, 0.6 and 0.8%.Results:A total of 35 fungal species belonging to 18 fungal genera were isolated and identified from different parts of H. sabdariffa plants. All strains have the capability for degrading phenol and their derivatives with variable extents. The optimum condition of degrading phenol in paper and pulp effluent samples by Fusarium poae11r7 were at pH 3–5, temperature at 28–35°C, good agitation speed at no agitation and 100 rpm.Conclusion:All endophytic fungal species can utilize phenol and its derivatives as a carbon source and be the potential to degrade phenol in industrial contaminants.

English

The diversity and genetic differentiation of populations of Fusarium species associated with sorghum fields, both endophytes obtained from sorghum performing and non performing plants and isolates obtained from two sampling periods were investigated. Fusarium specific Internal Transcribed Spacer 2 (FITS2) primers set were used to assess genetic variability of 32 isolates from susceptible Fusarium spp. endophytes from Sorghum tissues. Fusarium thapsinum (Gibberella thapsina) with 68.75% of the isolates constituted the majority of Fusarium spp. isolated in performing plants. Gibberella thapsina species identified are described as non-pathogenic and associated to performing plant of sorghum. Previously, some species of Fusarium thapsinum have been recognized as pathogenic and responsible for yield losses in several cereal crops including Sorghum bicolor produced in Burkina Faso. The other Fusarium spp. identified in this study including Fusarium subglutinans, Fusarium chlamydosporum, Gibberella intermedia, Fusarium dlaminii, Fusarium oxysporum, Fusarium proliferatum, and Fusarium spp. An additional unknown fungi species were also identified. A diverse population of 10 sequence types was found, although 8 sequence types represented nearly two-thirds of the isolates studied. The sequence types were placed in different phylogenetic clades within Fusarium spp., and endophytic isolates were not monophyletic. Phylogenetic analysis from Neighbor-Joining/UnWeighted Neighbor-Joining showed a high genetic relationship among these 32 isolates of Fusarium spp. and high variation in FITS sequence of them. The use of specific phylomarker of the genus Fusarium allowed to identify the endophytic species of this genus and to establish the phylogenetic relationships between the endophytic species of Fusarium. The phylogenetic analysis revealed three groups of the fungi. However, no relationship between these groups and the geographical origins of these fungi has been established.   Key words: Fusarium thapsinum, endophyte, FITS2 marker, sorghum.

Carbon utilization and growth-inhibition of citrus-colonizing Phyllosticta species

The genus Phyllosticta includes both endophytic and phytopathogenic species that occur on a broad range of plant hosts, including Citrus. Some pathogenic species cause severe disease, such as Phyllosticta citricarpa, the causal agent of Citrus Black Spot (CBS). In contrast, other species, such as Phyllosticta capitalensis, have an endophytic lifestyle in numerous plant hosts. Carbon utilization capabilities are hypothesized to influence both host range and lifestyle, and are in part determined by the set of Carbohydrate Active Enzyme (CAZyme) encoding genes of a species. In this study, carbon utilization capabilities of five Phyllosticta species were determined, as well as the CAZyme repertoire (CAZome) encoded in their genomes. Little variation was found among species in terms of carbon utilization capabilities and CAZome. However, one of the tested carbon sources, sugar beet pulp (SBP), inhibited growth of the plant pathogens, also when combined with another carbon source, while endophytic species remained unaffected.

From simple and specific zymographic detections to the annotation of a fungus Daldinia caldariorum D263 that encodes a wide range of highly bioactive cellulolytic enzymes

BackgroundLignocellulolytic enzymes are essential for agricultural waste disposal and production of renewable bioenergy. Many commercialized cellulase mixtures have been developed, mostly from saprophytic or endophytic fungal species. The cost of complete cellulose digestion is considerable because a wide range of cellulolytic enzymes is needed. However, most fungi can only produce limited range of highly bioactive cellulolytic enzymes. We aimed to investigate a simple yet specific method for discovering unique enzymes so that fungal species producing a diverse group of cellulolytic enzymes can be identified.ResultsThe culture medium of an endophytic fungus, Daldinia caldariorum D263, contained a complete set of cellulolytic enzymes capable of effectively digesting cellulose residues into glucose. By taking advantage of the unique product inhibition property of β-glucosidases, we have established an improved zymography method that can easily distinguish β-glucosidase and exoglucanase activity. Our zymography method revealed that D263 can secrete a wide range of highly bioactive cellulases. Analyzing the assembled genome of D263, we found over 100 potential genes for cellulolytic enzymes that are distinct from those of the commercially used fungal species Trichoderma reesei and Aspergillus niger. We further identified several of these cellulolytic enzymes by mass spectrometry.ConclusionsThe genome of Daldinia caldariorum D263 has been sequenced and annotated taking advantage of a simple yet specific zymography method followed by mass spectrometry analysis, and it appears to encode and secrete a wide range of bioactive cellulolytic enzymes. The genome and cellulolytic enzyme secretion of this unique endophytic fungus should be of value for identifying active cellulolytic enzymes that can facilitate conversion of agricultural wastes to fermentable sugars for the industrial production of biofuels.

Marine endophytic fungi associated with Halopteris scoparia (Linnaeus) Sauvageau as producers of bioactive secondary metabolites with potential dermocosmetic application.

Marine fungi and, particularly, endophytic species have been recognised as one of the most prolific sources of structurally new and diverse bioactive secondary metabolites with multiple biotechnological applications. Despite the increasing number of bioprospecting studies, very few have already evaluated the cosmeceutical potential of marine fungal compounds. Thus, this study focused on a frequent seaweed in the Portuguese coast, Halopteris scoparia, to identify the endophytic marine fungi associated with this host, and assess their ability to biosynthesise secondary metabolites with antioxidative, enzymatic inhibitory (hyaluronidase, collagenase, elastase and tyrosinase), anti-inflammatory, photoprotective, and antimicrobial (Cutibacterium acnes, Staphylococcus epidermidis and Malassezia furfur) activities. The results revealed eight fungal taxa included in the Ascomycota, and in the most representative taxonomic classes in marine ecosystems (Eurotiomycetes, Sordariomycetes and Dothideomycetes). These fungi were reported for the first time in Portugal and in association with H. scoparia, as far as it is known. The screening analyses showed that most of these endophytic fungi were producers of compounds with relevant biological activities, though those biosynthesised by Penicillium sect. Exilicaulis and Aspergillus chevalieri proved to be the most promising ones for being further exploited by dermocosmetic industry. The chemical analysis of the crude extract from an isolate of A. chevalieri revealed the presence of two bioactive compounds, echinulin and neoechinulin A, which might explain the high antioxidant and UV photoprotective capacities exhibited by the extract. These noteworthy results emphasised the importance of screening the secondary metabolites produced by these marine endophytic fungal strains for other potential bioactivities, and the relevance of investing more efforts in understanding the ecology of halo/osmotolerant fungi.

Diversity and functional characterization of endophytic Methylobacterium isolated from banana cultivars of South India and its impact on early growth of tissue culture banana plantlets.

This study aimed at determining the distribution, colonization and growth promoting nature of Methylobacterium spp. in tissue culture banana plantlets. Leaf samples from different field grown banana cultivars were used for Methylobacterium spp., isolation. Metabolic profile and functional characterization for plant growth-promoting traits of the isolates were assessed. The isolates were confirmed using 16S rRNA gene sequencing analysis, which resulted in six distinct species of Methylobacterium namely M. radiotolerans, M. salsuginis, M. thiocyanatum, M. rhodesianum, M. rhodinum and M. populi. Methylobacterium spp. inoculation experiment was conducted under hydroponic system in tissue culture banana plantlets (germ free) with eight selected isolates. A significant increase in growth parameters of Methylobacterium treated plantlets compared to uninoculated control was observed. Methylobacterium salsuginis TNMB03-gfp29 was developed and colonization micrograph was obtained using confocal laser scanning microscopy (CLSM) and scanning electron microscopy in different parts of banana plantlets (root, stem and leaves). Field grown banana plants found to harbour diverse endophytic Methylobacterium population. Our finding suggests that endophytic Methylobacterium species may provide significant plant growth promoting compounds/nutrients to the banana plants. The experimental results demonstrated the efficacy of Methylobacterium spp. as a potential bioinoculant and can be exploited as a phyllosphere and rhizosphere based bioinoculant for the initial establishment and growth of tissue culture banana plantlets. This study extended our knowledge on the distribution of Methylobacterium spp. in banana plants and endophytic colonization nature of this particular genus in plants. In addition, efficient isolate (M. salsuginis TNMB03) identified in this study may be promoted as bio-inoculants for banana plants after field evaluation.

Molecular Phylogeny of Endophytic Fungi from Rattan (Calamus castaneus Griff.) Spines and Their Antagonistic Activities against Plant Pathogenic Fungi.

Calamus castaneus is a common rattan palm species in the tropical forests of Peninsular Malaysia and is noticeable by the yellow-based spines that cover the stems. This study aimed to determine the prevalence of fungal endophytes within C. castaneus spines and whether they inhibit the growth of fungal pathogens. Twenty-one genera with 40 species of fungal endophytes were isolated and identified from rattan palm spines. Based on molecular identification, the most common isolates recovered from the spines were Colletotrichum (n = 19) and Diaporthe spp. (n = 18), followed by Phyllosticta spp., Xylaria sp., Trichoderma spp., Helminthosporium spp., Penicillium spp., Fusarium spp., Neopestalotiopsis spp., Arthrinium sp., Cyphellophora sp., Cladosporium spp., Curvularia sp., Bionectria sp., and Acremonium spp. Non-sporulating fungi were also identified, namely Nemania primolutea, Pidoplitchkoviella terricola, Muyocopron laterale, Acrocalymma fici, Acrocalymma medicaginis, and Endomelanconiopsis endophytica. The isolation of these endophytes showed that the spines harbor endophytic fungi. Most of the fungal endophytes inhibited the growth of several plant pathogenic fungi, with 68% of the interactions resulting in mutual inhibition, producing a clear inhibition zone of <2 mm. Our findings demonstrate the potential of the fungal endophytes from C. castaneus spines as biocontrol agents.

Diversity and biotechnological potential of endophytic Bacillus species originating from the stem apoplast fluid of sugarcane plants

The genus Bacillus has been isolated from a wide range of environments and comprises species with biotechnological applications, such as bioinseticides, biofungicides, and biofertilizers. This study investigated the endophytic occurrence and diversity of Bacilli in the apoplastic fluid of sugarcane cultivated in Brazil. Their biocontrol activity against fungi and their plant growth promotion (PGP) potential in sugarcane mini-set plants were also evaluated.Forty-two Bacillus colonies were isolated from the apoplastic fluid of one-year-old commercial sugarcane varieties and analyzed by sequencing the 16S rRNA gene. The ability to produce indols and siderophores and to solubilize phosphate and zinc were evaluated in vitro, along with their cellulose and protease activity and, antagonistic activity against sugarcane phytopatogenic fungi. The PGP potential was determined using strains which were inoculated in sugarcane mini-set plants grown in sterile substrate and later in unsterilized soil pots.The 16S rRNA analyses demonstrated high bacterial diversity with the majority closely related to B. megaterium followed by B. aryabhattai and B. cereus. B. pumilus, B. velezensis, B. safensis, B. flexus and Paenibacillus taichungensis species were also identified. Most strains were able to produce indol, solubilize zinc oxide and presented cellulase activity, while phosphate solubilization and siderophore production was observed in only few strains. The strains LMC21, LMC44 and LMC48 presented high biocontrol activity against the sugarcane fungi Colletotrichum falcatum. The spore formation was observed for strains grown for 8 days in the HCT medium but not in presence of sugarcane juice. Some strains also accumulated higher biomass in sugarcane (sterile conditions) and a mixture of three Bacillus strains performed better in the soil-pot inoculation experiment. These findings show for the first time that many Bacillus species are present in the apoplastic fluid of sugarcane grown in Brazil and indicate that some strains have PGP and biocontrol potential for application as biofertilizer in sugarcane plants.

Identification and Histological Observation of Endophytic Fungi Isolated from Medicinal Plant, Physalis angulata L.

Some endophytic fungi species live in medicinal plant tissue and does not make any damage, but live in symbiotic mutualism relationship with the host plant. This research was done to: 1) identify the endophytic fungi species isolated from P. angulata leaf, twig, and stem bark tissues, 2) determine the endophytic fungi colonization in the P. angulata plant tissue by histologic observation. The endophytic fungi was isolated from healthy P. angulata plant parts, then inoculated on Potato Dextrose Agar medium and incubated in 27°C for 7-14 days. Each endophytic fungi isolates were identified. The histologic observation was done by microscopic observation to determine the endophytic fungi position in the plant tissue. The conclusion are: 1) seven endophytic fungi species were found: Penicillium verrucosum, Colletotrichum alienum, Fusarium subglutinans, Aspergillus nidulans, Mycelia sterilia 1, Mycelia sterilia 2, and Rhizoctonia sp.; 2) the endophytic fungi micelium was found on the leaf epidermis cell wall, on the twig epidermis cell wall, and parenchyma cell wall, on the stem bark epidermis cell wall. The suggestion of the study: it is need to make the next research about secondary metabolites content produced by endophytic fungi species isolated from P. angulata and their antimicrobial activity.

Molecular phylogeny and morphology reveal three novel species of Curvularia (Pleosporales, Pleosporaceae) associated with cereal crops and weedy grass hosts

The genus Curvularia comprises phytopathogenic, saprobic, epiphytic and endophytic fungal species associated with cereal crops and their wild relatives. Recently, multi-locus phylogenetic studies have been widely implemented for accurate identification of Curvularia, at the species level. Although the genus is taxonomically diverse, the species associated with cereal crops and weeds are poorly known in Sri Lanka. In this study, symptomatic specimens of cereals and associated weedy grass hosts were collected from selected locations in Sri Lanka. The isolates obtained were initially identified based on microscopic characters. The nuclear ribosomal internal transcribed spacers 1 and 2 with 5.8S region (ITS), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and translation elongation factor 1-α (TEF1) loci were sequenced and used in multi-locus phylogenetic analyses. Three novel evolutionary lineages were identified, distinct from all the currently accepted species of Curvularia. To accommodate the novel phylogenetic lineages, three novel species of Curvularia are described, namely Curvularia eleusinicola, C. panici-maximi and C. simmonsii. Morphological descriptions and illustrations are provided for the newly described taxa. In addition, host records are updated for recently introduced C. plantarum and C. pseudointermedia. This study highlights the need for extensive collections and molecular identifications of tropical species of hyphomycetous fungi associated with cereals, fibre crops and weeds in order to support effective disease management and surveillance measures.

Effect of long-term fertilization on bacterial communities in wheat endosphere

Fertilization has been shown to exert a significant influence on soil microorganisms and directly and indirectly influences plant growth and survival in agroecosystems. However, it is unknown whether fertilization affects endophytic microbial communities, which are ubiquitous and intimately associated with plant growth and health. Herein, we investigated endophytic bacterial communities in wheat leaves and roots under different long-term fertilization regimes, including NPK chemical fertilizer and NPK chemical fertilizer combined with wheat straw, pig manure, or cow manure. Endophytic bacterial community composition considerably differed in leaves and roots. Although different fertilization treatments did not affect the endophytic bacterial species richness or phylogenetic diversity in either leaves or roots, the community composition was significantly altered, particularly in roots. The endophytic bacterial co-occurrence network in leaves was more complex and stable than that in roots. Furthermore, many of the keystone species that were identified by their topological positions in the co-occurrence networks of leaves and roots were involved in plant growth and fitness. The total relative abundance of keystone species was the highest in the NPK plus cow manure treatment in both leaves and roots. Overall, our results suggest that different fertilization regimes can strongly affect endophytic bacterial communities, and the combination of NPK fertilizer and cow manure promoted the relative abundance of the key endophytic bacterial microbiota in both leaves and roots, which might be beneficial for plants in agroecosystems.

Risk assessment of secondary metabolites produced by fungi in the genus Stemphylium.

The fungal genus Stemphylium (phylum Ascomycota, teleomorph Pleospora) includes plant pathogenic, endophytic, and saprophytic species with worldwide distributions. Stemphylium spp. produce prodigious numbers of airborne spores, so are a human health concern as allergens. Some species also produce secondary metabolites, such as glucosides, ferric chelates, aromatic polyketides, and others, that function as toxins that damage plants and other fungal species. Some of these compounds also exhibit a low level of mammalian toxicity. The high production of airborne spores by this genus can result in a high incidence of human exposure. Concern about toxin production appears to be the reason that Stemphylium vesicarium, which is a pathogen of several vegetable crops, was classified in Canada as a potential risk of harm to humans for many years. A detailed assessment of the risk of exposure was provided to the relevant regulatory body, the Public Health Agency of Canada, which then determined that Stemphylium spp. in nature or under laboratory conditions posed little to no risk to humans or animals, and the species was re-assigned as a basic (level 1) risk agent.

Bioprospecting of endophytic microorganisms for bioactive compounds of therapeutic importance.

Presently, several drug discovery investigations on therapeutic management of human health are aimed at bioprospecting for microorganisms, especially endophytic microbes of biotechnological importance. This review investigates the benefits of endophytes, especially in producing bioactive compounds useful in modern medicine by systematically reviewing published data from 12 databases. Only experimental studies investigating either or both bacterial and fungal endophytes and within the scope of this review were selected. The published data from the last 2 decades (2000-2019) revealed diverse endophytes associated with different plants produce a broad spectrum of bioactive compounds with therapeutic benefits. Notably, antibacterial, followed by anticancer and antifungal activities, were mostly reported. Only three studies investigated the anti-plasmodial activity. The variation observed in the synthesis of bioactive compounds amongst endophytes varied with host type, endophyte species, and cultivation medium. Fungal endophytes were more investigated than bacterial endophytes, with both endophytes having species diversity amongst literature. The endophytes were predominantly from medicinal plants and belonged to either Ascomycota (fungi) or Proteobacteria and Firmicutes (bacteria). This review presents excellent prospects of harnessing endophytes and their unique bioactive compounds in developing novel and effective compounds of medicinal importance.

Species and geographic specificity between endophytic fungi and host supported by parasitic Cynomorium songaricum and its host Nitraria tangutorum distributed in desert.

This study was aimed to investigate whether host plant species and lifestyles, and environmental conditions in the desert affect endophytic fungi composition. Endophytic fungal communities from parasitic plant Cynomorium songaricum and its host Nitraria tangutorum were investigated from three sites including Tonggu Naoer, Xilin Gaole, and Guazhou in Tengger and Badain Jaran Deserts in China using the next-generation sequencing of a ribosomal RNA gene region. Similarity and difference in endophytic fungal composition from different geographic locations were evaluated through multivariate statistical analysis. It showed that plant genetics was a deciding factor affecting endophytic fungal composition even when C. songaricum and N. tangutorum grow together tightly. Not only that, the fungal composition was also greatly affected by the local environment and rainfall. However, the distribution and richness of fungal species indicated that the geographical distance exerted little influence on characterizing the fungal composition. Overall, the findings suggested that plant species, parasitic or non-parasitic lifestyles of the plant, and local environment strongly affected the number and diversity of the endophytic fungal species, which may provide valuable insights into the microbe ecology, symbiosis specificity, and the tripartite relationship among parasitic plant, host, and endophytic fungi, especially under desert environment.

Endophytic Aspergillus species from corn kernels in Peninsular Malaysia

Endophytes are micro-organisms that infect and colonize internal tissues of host plants without causing obvious disease symptoms. Although most endophyte-plant relationships occur in the absence of the manifestation of disease, infections by some endophytic Aspergillus species may occur, leading to the production of mycotoxins in infected plant tissues by toxigenic species. In this study, endophytic Aspergillus species from kernels of corn plants in six states of Peninsular Malaysia were isolated and identified. A total of 178 isolates of endophytic Aspergillus belonging to two species, were recovered from surface disinfected corn kernels after 4-7 days of incubation on Potato Dextrose Agar (PDA), and identified using morphological characteristics on different growth media. Endophytic Aspergillus flavus was the most commonly isolated species (n=177), followed by Aspergillus tubingensis (n=1). Measures to control seed infection by endophytic Aspergillus species are required to improve corn seed health and preserve corn yield in Peninsular Malaysia.

Molecular profiling and anti-infective potency of endophytic actinomycetes inhabiting Madhuca insignis Radlk., from Western Ghats of India

BackgroundEndophytic actinomycetes are well known for their diverse bioactive entities and considered as an important source for drug development research. ResultsWe isolated and identified four potential endophytic Streptomyces species, i.e., Streptomyces misionensis MI22, Streptomyces roietensis MI24, Streptomyces glaucescens MI29, and Streptomyces sp. MI04 inhabiting Madhuca insignis by its characteristic morphological features and 16S rRNA gene sequence analysis. S. misionensis MI22 exhibits a broad spectrum of anti-microbial activity against methicillin-resistant Staphylococcus aureus (25.00 ± 1.00 mm) followed by Bacillus subtilis (23.66 ± 0.57 mm), Escherichia coli (22.00 ± 0.00 mm), and Candida albicans (18.00 ± 0.00 mm). Minimum inhibitory concentrations of the ethyl acetate fraction of S. misionensis MI22 against test pathogens were ranged from 25 to 100 μg/mL. Indeed, strain MI22 also exhibited significant anti-proliferative activity against HeLa cell line with IC50 value 98 μg/mL and showed no cytotoxicity effect to the normal human embryonic kidney cell line in the MTT assay. The anti-microbial metabolites from strain MI22 were detected at Rf 0.55 as depicted by the inhibition zone on the intensive band in TLC-bioautography assay. ConclusionThe study indicates that, anti-microbial metabolites of these endophytic Streptomyces species, especially S. misionensis MI22 as a prolific source to discover novel bioactive metabolites to combat multidrug-resistant pathogens.

Endophytic Diaporthe Associated With Citrus grandis cv. Tomentosa in China.

Diaporthe species are associated with Citrus as endophytes, pathogens, and saprobes worldwide. However, little is known about Diaporthe as endophytes in Citrus grandis in China. In this study, 24 endophytic Diaporthe isolates were obtained from cultivated C. grandis cv. “Tomentosa” in Huazhou, Guangdong Province in 2019. The nuclear ribosomal internal transcribed spacer (ITS), partial sequences of translation elongation factor 1-α (tef1), β-tubulin (tub2), and partial calmodulin (cal) gene regions were sequenced and employed to construct phylogenetic trees. Based on morphology and combined multigene phylogeny, eleven Diaporthe species were identified including two new species, Diaporthe endocitricola and D. guangdongensis. These are the first report of D. apiculata, D. aquatica, D. arecae, D. biconispora, D. limonicola, D. masirevicii, D. passifloricola, D. perseae, and D. sennae on C. grandis. This study provides the first intensive study of endophytic Diaporthe species on C. grandis cv. tomentosa in China. These results will improve the current knowledge of Diaporthe species associated with C. grandis. The results obtained in this study will also help to understand the potential pathogens and biocontrol agents and to develop a platform in disease management.

Fungal communities of grapevine decline in the main grape-growing regions of Iran

Grapevine decline is a worldwide problem of viticulture, leading to significant yield loss in the grape-growing regions of Iran. Several species of fungal endophytes are known as the causal agents of the disease. Vines with internal and external symptoms were sampled from Qazvin, Lorestan, North Khorasan and Razavi Khorasan provinces, showing different climate regimes. Regarding geographical regions with their variable climatic background, the cultivars, and vineyard age, the dominant fungi were different. In Bojnord, Phaeomoniella chlamydospora and Phaeoacremonium minimum, the known causal agents of esca disease, were the most isolated fungi obtained from 27 and 24 symptomatic plants, respectively. The establishment of Phaeomoniella and Phaeoacremonium species in Takestan and Tajareh might be prevented by different environmental and biological factors. Fomitiporia mediterranea isolates, the white rot agent, were isolated from Takestan with 25–30 years old vineyards, and the average summer temperature, precipitation, and humidity were about 1 mm, 25 °C, 38% respectively. In jovein with 15 years old vineyards, the most important esca related fungi, Phaeomoniella, Phaeoacremonium, and Fomitiporia species, were isolated in the summer, obtained from 31, 23 and 18 symptomatic plants, respectively. This study gives some insights into the possible relationship between grapevine decline related fungi and, especially, environmental factors, which leads to a more diffused picture of esca and the related fungi communities, making it challenging to clearly separate the effects of other factors.