Composition of endophytic fungal community associated with leaves of maize cultivated in south Brazilian field

The objective of this study was to conduct a survey about fungi associated with leaves from two different maize plant lineages and to analyze their microbiota diversity. Isolated fungi were identified by morphological analysis and molecular taxonomy was performed using ITS1-5.8S-ITS2 rDNA. About 27 fungi morphotypes were obtained, 15 of them were from the first maize lineage. About 86.7% of the individuals belonged to the Dothideomycetes class (Phoma sorghina, Epicocum nigrum, Cladosporium sp., Bipolaris zeicola, and Alternaria alternata complex) and 13.3% to the Sordariomycetes class (Diaporthe/Phomopsis sp. and Nigrospora sp.). This ratio was opposite in the other maize lineage with 25.0% of Dothideomycetes (E. nigrum and Pleosporales) and 75.0% of Sordariomycetes (Gibberella fujikuroi complex, Fusarium graminearum complex, Diaporthe/Phomopsis sp., and Nigrospora sp.). By concerning the analyses of morphological characteristics and molecular phylogeny, this study intended to identify the groups of saprophytic, phytopathogenic, and mycotoxin fungi, which differently co-inhabit leaf tissue of maize plants in both tested lineages

Analysis of the genetic diversity of Candida isolates obtained from diabetic patients and kidney transplant recipients

Yeasts of the genu

Endophytes of Brazilian Medicinal Plants with Activity against Phytopathogens

Plant diseases caused by phytopathogens are responsible for significant crop losses worldwide. Resistance induction and biological control have been exploited in agriculture due to their enormous potential. In this study, we investigated the antimicrobial potential of endophytic fungi of leaves and petioles of medicinal plants Vochysia divergens and Stryphnodendron adstringens located in two regions of high diversity in Brazil, Pantanal, and Cerrado, respectively. We recovered 1,304 fungal isolates and based on the characteristics of the culture, were assigned to 159 phenotypes. One isolate was selected as representative of each phenotype and studied for antimicrobial activity against phytopathogens. Isolates with better biological activities were identified based on DNA sequences and phylogenetic analyzes. Among the 159 representative isolates, extracts from 12 endophytes that inhibited the mycelial growth (IG) of Colletotrichum abscissum (≥40%) were selected to expand the antimicrobial analysis. The minimum inhibitory concentrations (MIC) of the extracts were determined against citrus pathogens, C. abscissum, Phyllosticta citricarpa and Xanthomonas citri subsp. citri and the maize pathogen Fusarium graminearum. The highest activity against C. abscissum were from extracts of Pseudofusicoccum stromaticum CMRP4328 (IG: 83% and MIC: 40 μg/mL) and Diaporthe vochysiae CMRP4322 (IG: 75% and MIC: 1 μg/mL), both extracts also inhibited the development of post-bloom fruit drop symptoms in citrus flowers. The extracts were promising in inhibiting the mycelial growth of P. citricarpa and reducing the production of pycnidia in citrus leaves. Among the isolates that showed activity, the genus Diaporthe was the most common, including the new species D. cerradensis described in this study. In addition, high performance liquid chromatography, UV detection, and mass spectrometry and thin layer chromatography analyzes of extracts produced by endophytes that showed high activity, indicated D. vochysiae CMRP4322 and P. stromaticum CMRP4328 as promising strains that produce new bioactive natural products. We report here the capacity of endophytic fungi of medicinal plants to produce secondary metabolites with biological activities against phytopathogenic fungi and bacteria. The description of the new species D. cerradensis, reinforces the ability of medicinal plants found in Brazil to host a diverse group of fungi with biotechnological potential

Antibacterial Activity of Endophytic Actinomycetes Isolated from the Medicinal Plant \u3cem\u3eVochysia divergens\u3c/em\u3e (Pantanal, Brazil)

Endophytic actinomycetes from medicinal plants produce a wide diversity of secondary metabolites (SM). However, to date, the knowledge about endophytes from Brazil remains scarce. Thus, we analyzed the antimicrobial potential of 10 actinomycetes isolated from the medicinal plant Vochysia divergens located in the Pantanal sul-mato-grossense, an unexplored wetland in Brazil. Strains were classified as belonging to the Aeromicrobium, Actinomadura, Microbacterium, Microbispora, Micrococcus, Sphaerisporangium, Streptomyces, and Williamsia genera, through morphological and 16S rRNA phylogenetic analyzes. A susceptibility analysis demonstrated that the strains were largely resistant to the antibiotics oxacillin and nalidixic acid. Additionally, different culture media (SG and R5A), and temperatures (28 and 36°C) were evaluated to select the best culture conditions to produce the active SM. All conditions were analyzed for active metabolites, and the best antibacterial activity was observed from metabolites produced with SG medium at 36°C. The LGMB491 (close related to Aeromicrobium ponti) extract showed the highest activity against methicillin-resistant Staphylococcus aureus (MRSA), with a MIC of 0.04 mg/mL, and it was selected for SM identification. Strain LGMB491 produced 1-acetyl-β-carboline (1), indole-3-carbaldehyde (2), 3-(hydroxyacetyl)-indole (4), brevianamide F (5), and cyclo-(L-Pro-L-Phe) (6) as major compounds with antibacterial activity. In this study, we add to the knowledge about the endophytic community from the medicinal plant V. divergens and report the isolation of rare actinomycetes that produce highly active metabolites

Genome of Herbaspirillum seropedicae Strain SmR1, a Specialized Diazotrophic Endophyte of Tropical Grasses

The molecular mechanisms of plant recognition, colonization, and nutrient exchange between diazotrophic endophytes and plants are scarcely known. Herbaspirillum seropedicae is an endophytic bacterium capable of colonizing intercellular spaces of grasses such as rice and sugar cane. The genome of H. seropedicae strain SmR1 was sequenced and annotated by The Paraná State Genome Programme—GENOPAR. The genome is composed of a circular chromosome of 5,513,887 bp and contains a total of 4,804 genes. The genome sequence revealed that H. seropedicae is a highly versatile microorganism with capacity to metabolize a wide range of carbon and nitrogen sources and with possession of four distinct terminal oxidases. The genome contains a multitude of protein secretion systems, including type I, type II, type III, type V, and type VI secretion systems, and type IV pili, suggesting a high potential to interact with host plants. H. seropedicae is able to synthesize indole acetic acid as reflected by the four IAA biosynthetic pathways present. A gene coding for ACC deaminase, which may be involved in modulating the associated plant ethylene-signaling pathway, is also present. Genes for hemagglutinins/hemolysins/adhesins were found and may play a role in plant cell surface adhesion. These features may endow H. seropedicae with the ability to establish an endophytic life-style in a large number of plant species

Not avaliable (sem título inglês)

Guignardia citricarpa é um fungo ascomiceto frequentemente isolado como endófito de citros, sendo no entanto o agente causal da Mancha Preta dos Citros (MPC). Na literatura há referências sobre a existência de dois tipos de G. citricarpa, um deles patogênico, e outro não patogênico. Em 1994 obtivemos isolados de G. citricarpa provenientes de folhas de tangerineiras assintomáticas de pomares do Estado de São Paulo, nos quais a doença não era encontrada. A variabilidade morfológica e genética constatada entre eles foi explicada por duas hipóteses, a primeira de que eles pertenciam a uma população não patogênica, mas endofitica deste fungo, e a segunda de que estes isolados eram patógenos latentes. Como em 1996 a doença começou a ser relatada em diversas regiões do país, inclusive em pomares dos quais os isolados endofiticos foram obtidos, o presente trabalho foi realizado com a finalidade de, por meio de estudos genéticos, citológicos e morfológicos, tentar verificar similaridades e diferenças entre os isolados obtidos como endofiticos e isolados patogênicos. Desta forma, 103 isolados de Guignardia spp obtidos de diferentes hospedeiros, regiões e épocas, foram comparados por meio de marcadores de RAPO. Os isolados foram separados em pelo menos 3 grupos distintos geneticamente, sendo agrupados preferencialmente pela forma de isolamento, ou seja, de lesão de MPC ou como endofiticos de folhas aparentemente sadias. Das 343 bandas de RAPO obtidas, uma encontrada exclusivamente em isolados patogênicos, foi clonada e seqüenciada. A partir desta seqüência um par de "primers" foi sintetizado e utilizado com sucesso para discriminar os isolados patogênicos dos demais. Este par de "primers" após testes futuros poderá ser utilizado como diagnóstico molecular de isolados patogênicos de G. citricarpa em pomares de citros. Foi obtido também o cariótipo eletroforético de quatro isolados de G. citricarpa, revelando que este fungo apresenta genoma de no mínimo 21,5 Mb distribuídos em pelo menos 8 cromossomos, não sendo encontrada variabilidade entre os isolados ensaiados. Características morfológicas também foram analisadas. A taxa de crescimento não diferenciou os isolados analisados, mas foi possível separá-los quanto ao tipo de esporulação in vitro, o que concordou com o agrupamento sugerido por meio de marcadores de RAPD. Estudos de germinação in vitro e in vivo, revelaram que os conídios de G. citricarpa possuem germinação apical com formação de apressório, sendo sugerida a adesão como pré-requisito para a germinação in vitro. Neste processo de adesão deve participar uma glucana β 1,6 produzida por um dos isolados estudados. A produção deste e outros exopolissacarídios por dois outros isolados, parece ser dependente do pH e da fonte de carbono utilizada no meio de cultura. Pelos resultados obtidos, sugere-se que os isolados obtidos como endófitos, não são patógenos latentes, podendo inclusive pertencerem a outra espécie de Guignardia. Sendo assim, os isolados patogênicos poderão ser diferenciados dos endofíticos via PCR, utilizando-se o par de "primers" obtido neste trabalhoGuignardia citricarpa is an ascomycete fungus frequently isolated from citrus as endophyte, being however the causal agent of the Black Spot of citrus (BSC). There are in the literature references concerning the existence of two types of G. citricarpa, one pathogenic and one non pathogenic. ln 1994 we obtained isolates of G. citricarpa from leaves of asintomatic tangors in orchards of São Paulo state where no disease had been found. The morphologic and genetic variabilities found among them were explained by two hypothesis, the first suggesting that they belong to a non pathogenic or endophytic population, and the second as a latent pathogen. ln 1996 the disease started to be observed in several region of the country, including orchards from which the endophytic isolates were previously obtained. Thus, this present work was developed with the aim of, by means of genetic, cytological and morphologic studies, verify the similarities and differences between G. citricarpa obtained as endophyte and as pathogen. On this basis, 103 isolates of G. citricarpa obtained from different hosts, regions and seasons, were compared by RAPD markers. Isolates were separated in at least 3 distinct groups, being clustered preferentially by isolation protocol, that is, from BSC lesion (pathogen) or from apparently healthy leaves (endophyte). Of 343 RAPD bands observed, one was found exclusively in pathogenic isolates and therefore it was cloned and sequenced. Based on the revealed sequence, a pair of primers was synthesized and used with success to discriminate pathogenic isolates from the others. With additional tests, this pair of primers will be able to help the molecular diagnosis of pathogenic isolates of G. citricarpa in citrus orchards. We have also obtained the electrophoretic karyotype of 4 isolates of G. citricarpa, revealing that this fungus has a genome of at least 21,5 Mb, divided in at least 8 chromosomes. Morphologic characteristics were also analyzed in the two types of isolates. The growth rate did not discriminate the isolates, but it was possible to separate them in terms of sporulation in vitro, resulting in distinct groups already suggested by RAPD markers. Studies of germination in vitro and in vivo showed that G. citricarpa conidium germinates apically, with apressorium formation, and that adhesion seems to be a necessary step for germination in vitro. During adhesion, a β3 1-6 glucan produced by the isolates seems to play a role in the process. The production of this and other exopolysaccharides were anaiyzed by two isolates and seems to be pH and carbon source dependent. Based on the results obtained in this work, we suggest that the isolates referred as endophytes are not latent pathogens. Furthermore, they could even belong to another Guignardia species whereas, pathogenic isolates can be distinguished: from those endophytes by PCR, using a pair of primers developed in this wor