Summary: Arbuscular mycorrhizal fungi (AMF) and dark septate endophytes (DSEF) have such an influence on plant development and on the edaphological characteristics and nutrients available in the soil that changes in these communities can drive modifications in the biotic and abiotic variables of an ecosystem, demonstrating the synergetic between species. Given its relevance, the objective of this work was to determine the status of colonization in the roots of three representative herbaceous species of the understory of a community of Abies religiosa (Kunth Schltdl. et Cham.) and Pinus harwegii (Lindl), and to contrast the coexistence of colonizers in three conservation-disturbance scenarios that exemplify the vulnerability situations of a forest ecosystem. A tripartite AMF-HESO-plant interaction is reported by the occurrence of coenocytic hyphae, vesicles, septate hyphae, moniliform cells, and sclerotia. The values of arbuscular mycorrhization exceed 50% and are higher than those found for endophytes (15.7-64.5%). The colonization of both types of fungi, such as AMF sporulation (687 spores 50 g-1 soil in the reforested area), seems to be related to the microclimatic conditions resulting from soil use. The edaphological variables that allow inferring the colonization behavior correspond mainly to available phosphorus, calcium, pH, organic matter, and cation exchange capacity. It is concluded that soil properties, its use, and the type of plants are determinant in the establishment of fungal communities. However, more studies on HESO-plant interaction are needed.

Resumen: Los hongos micorrícicos arbusculares (HMA) y los endófitos septados oscuros (DSEF) tienen tal influencia en el desarrollo de las plantas y en las características edafológicas y nutrientes disponibles en el suelo, que cambios en estas comunidades pueden impulsar modificaciones en las variables bióticas y abióticas de un ecosistema, demostrando la sinergia entre especies. Dada su relevancia, el objetivo de este trabajo fue determinar el estado de colonización en las raíces de tres especies herbáceas representativas del sotobosque de una comunidad de Abies religiosa (Kunth Schltdl. et Cham.) y Pinus harwegii (Lindl), y contrastar la coexistencia de colonizadores en tres escenarios de conservación-perturbación que ejemplifican las situaciones de vulnerabilidad de un ecosistema forestal. Se constata una interacción tripartita AMF-HESO-planta por la aparición de hifas coenocíticas, vesículas, hifas septadas, células moniliformes y esclerocios. Los valores de micorrización arbuscular superan el 50% y son superiores a los encontrados para los endófitos (15.7-64.5%). La colonización de ambos tipos de hongos, como la esporulación de AMF (687 esporas 50 g-1 de suelo en la zona reforestada), parece estar relacionada con las condiciones microclimáticas derivadas del uso del suelo. Las variables edafológicas que permiten inferir el comportamiento de la colonización corresponden principalmente al fósforo disponible, calcio, pH, materia orgánica y capacidad de intercambio catiónico. Se concluye que las propiedades del suelo, su uso y el tipo de plantas son determinantes en el establecimiento de las comunidades fúngicas. Sin embargo, son necesarios más estudios sobre la interacción HESO-planta.

Abstract Introduction. Endophytic fungi (EF) in grapevines (Vitis vinifera) have great potential for agricultural applications and may represent possible biological control agents for vineyards. Objective. To isolate and identify endophytic fungi (EF) from grapevine plants with antibiosis capacity against Botrytis cinerea. Materials and methods. Young and mature branches from forty healthy grapevine plants were collected from seven production lots in the Cañete province, Lima, Peru, during March to May 2022. EF isolation was performed on 702 sections of disinfected tissues using potato dextrose agar (PDA) culture medium, and identification was carried out at the genus level. Secondary metabolites were extracted from twenty-eight EF strains to determine the mycelial inhibition of the pathogen. An analysis of variance (ANOVA) was conducted on the evaluated parameters using the Minitab statistical program. Results. A total of 129 EF strains were isolated from leaves and stems of young and mature grapevine branches, encompassing seven genera (Trichoderma, Aspergillus, Alternaria, Cladosporium, Fusarium, Nigrospora, and Stemphylium). The highest frequency was found in leaves and mature branches. Trichoderma and Alternaria were the most frecuent genera in grapevine plants, with 22.48 % and 21.71%, respectively. Metabolites obtained from the EFVH-01 strain of the genus Cladosporium, and EF-99 and EF-70 strains of the genus Alternaria, had the greatest effect on mycelial inhibition of the pathogen. Conclusions. Strains of endophytic fungi from the genera Cladosporium and Alternaria showed antibiosis capacity against Botrytis cinerea at the laboratory scale, which could potential biocontrol agents of pathogens in grapevine cultivation.

Resumen Introducción. Los hongos endófitos (HE) de la vid (Vitis vinifera) tienen un gran potencial de aplicación en la agricultura y pueden representar posibles agentes de control biológico de los viñedos. Objetivo. Aislar e identificar los hongos endófitos (HE) de plantas de vid con capacidad de antibiosis hacia Botrytis cinerea. Materiales y métodos. Se colectaron ramas jóvenes y maduras de cuarenta plantas sanas de vid de siete lotes de producción en la provincia de Cañete, Lima, Perú, durante los meses de marzo a mayo 2022. El aislamiento de HE se realizó de 702 secciones de tejidos desinfectados en medio de cultivo papa dextrosa agar (PDA) y la identificación fue a nivel de género. Se extrajeron metabolitos secundarios de veintiocho cepas de HE para determinar la inhibición micelial del patógeno. Se llevó a cabo un análisis de varianza (ANDEVA) de los parámetros evaluados con el programa estadístico Minitab. Resultados. Se aislaron un total de 129 cepas de HE de hojas y tallos de ramas jóvenes y maduras de vid, que correspondieron a siete géneros (Trichoderma, Aspergillus, Alternaria, Cladosporium, Fusarium, Nigrospora y Stemphylium). La mayor frecuencia se presentó en las hojas y rama madura. Trichoderma y Alternaria fueron los géneros más frecuentes en las plantas de vid con 22,48 % y 21,71 %, respectivamente. Los metabolitos obtenidos de las cepas HEVH-01 del género Cladosporium, HEV-99 y HEV-70 del género Alternaria, tuvieron mayor efecto en la inhibición micelial del patógeno. Conclusiones. Cepas de hongos endofitos del género Cladosporium y Alternaria mostraron capacidad de antibiosis hacia Botrytis cinerea a escala de laboratorio, los cuales podrían ser potenciales biocontroladores de patógenos en el cultivo de vid.

Resumen Los hongos endófitos son microorganismos simbiontes de plantas, diversos y abundantes en regiones tropicales, con gran potencial como agentes biocontroladores de enfermedades fúngicas agrícolas. El objetivo del presente trabajo fue evaluar la actividad antifúngica in vitro de los hongos endófitos asociados con cultivares criollos de Cucumis sativus y Lagenaria siceraria de Yucatán. Las cepas endófitas fueron aisladas e identificadas de diferentes órganos de las plantas, para posteriormente ser sometidas a enfrentamientos duales y pruebas de antibiosis. Se identificaron 20 especies de hongos endófitos con efectos antifúngicos contra Corynespora cassiicola, Colletotrichum truncatum y Fusarium equiseti, obteniendo rangos de inhibición de 17.72-100%, 16.51-91.55% y 4.64-91.89%, respectivamente. Aspergillus flavus, Lasiodiplodia pseudotheobromae y Trichoderma longibrachiatum exhibieron los mayores porcentajes de inhibición contra los tres fitopatógenos. Así mismo, estas tres especies de endófitos demostraron un efecto antibiótico en sus extractos, al inhibir principalmente el crecimiento de F. equiseti hasta en un 46.46%. La cepa de L. pseudotheobromae tiene características que le confieren un potencial como agente antifúngico, por lo que se deberán seguir explorando sus aplicaciones y condiciones de crecimiento. Este trabajo representa un primer esfuerzo por conocer la diversidad de hogos endófitos en los cultivos criollos de la región.

Abstract Endophytic fungi are plant symbiont microorganisms, diverse and abundant in tropical regions, with great potential as biocontrol agents of agricultural fungal diseases. The objective of the present work was to evaluate in vitro antifunfal activity of endophytic fungi associated with creole cultivars of Cucumis sativus and Lagenaria siceraria from Yucatán. The endophytic strains were isolated and identified from different organs, to be subjected to dual confrontations and antibiotic tests. Twenty species with antifungal effects against Corynespora cassiicola, Colletotrichum truncatum and Fusarium equiseti were identified, obtaining inhibition ranges of 17.72-100%, 16.51-91.55% and 4.64-91.89%, respectively. Aspergillus flavus, Lasiodiplodia pseudotheobromae and Trichoderma longibrachiatum exhibited the highest percentages of inhibition against the three phytopathogens. Likewise, these three species of endophytes demonstrated an antibiotic effect in their extracts, mainly inhibiting the growth of F. equiseti by up to 46.46%. The L. pseudotheobromae strain has characteristics that give it potential as an antifungal agent, so its applications and growth conditions should continue to be explored. This work represents a first effort to understand the diversity of endophytic fungi in native crops in the region.

RESUMEN El cultivo de quinua es afectado por diversas enfermedades, entre las cuales, el mildiu (Peronospora variabilis Gaum) es la más importante por su prevalencia, severidad e impacto económico. Los hongos endófitos antagonistas representan una opción sustentable para el manejo sanitario de los cultivos. El objetivo del trabajo fue determinar el comportamiento endofítico de cepas locales de Trichoderma asperelloides en plántulas de quinua. Se utilizó la técnica de cultivo para el re-aislamiento del hongo de secciones de raíz, tallo y hoja a los 15 y 30 días después de la aplicación del hongo (dda), y se realizó la observación microscópica de cortes histológicos para caracterizar la colonización del hongo a los 30 dda. El diseño definió los efectos fijos: genotipo de quinua (AMMA-18, RQ-252-18 y RQ-SAC-18), cepa de T. asperelloides: (T1, T15, T16); órgano: raíz, tallo, hoja, y momento de evaluación (15 y 30 dda). La mayor ocurrencia de re-aislamientos fue a los 15 dda en RQ-252-18 desde las tres secciones, seguido por AMMA-2018 y RQ-SAC-18. Se registró diferencias significativas en los niveles de interacción RQ-252-18/T16 siendo superiores los re-aislamientos desde raíz y hoja. A los 30 dda, no se observaron diferencias entre genotipos, y re-asliamientos eran significativamente mayores desde la raíz. El re-aislamiento fue dependiente del tiempo desde la aplicación y la combinación genotipo/cepa. Se observó colonización intercelular de hifas del hongo en el tejido epidérmico y del córtex de tallo y raíz. El trabajo amplía los registros de T. asperelloides como endófito de quinua para futuros estudios de interacción múltiple con P. variabilis u otros factores estrés biótico y/o abiótico.

ABSTRACT The quinoa crop is affected by various diseases, with mildew (Peronospora variabilis Gaum) being the most important due to prevalence, severity and economic impact. Antagonistic endophytic fungi represent a sustainable option for the sanitary management of crops. The objective of the work was to determine the endophytic behavior of local strains of Trichoderma asperelloides in quinoa seedlings. The culture technique was used to re-isolate the fungus from root, stem and leaf sections at 15 and 30 daa and microscopic observation of histological sections to characterize the colonization of the fungus at 30 daa. The design defined the fixed effects: quinoa genotype (AMMA-18, RQ-252-18 and RQ-SAC-18), T. asperelloides strain: (T1, T15, T16); organ: root, stem, leaf, and evaluation time 15 and 30 daa. The highest occurrence of re-isolations was at 15 daa in RQ-252-18 from the three sections, followed by AMMA-2018 and RQ-SAC-18. Significant differences were recorded in the RQ-252-18/T16 interaction levels, with re-isolations from root and leaf being higher. At 30 daa, there were no differences between genotypes, but between type of organ, always superior in root. Re-isolation was dependent on the time since application and the genotype/strain combination. Intercellular colonization of fungal hyphae was observed in the epidermal tissue and stem and root cortex. The work expands the records of T. asperelloides as a quinoa endophyte for future multiple interaction studies with P. variabilis or other biotic and/or abiotic stress factors.

RESUMEN El programa de mejoramiento de alfalfa requiere alternativas más amigables con el ambiente para incrementar la sanidad del cultivo. Las rlzobacterlas promotoras del crecimiento vegetal (RPCV) y endófltos de semillas constituyen una opción para inhibir hongos patógenos por la producción de compuestos antifúngicos o enzimas hidrolíticas. En Argentina, la alfalfa es afectada principalmente por Colletotr¡chum trifolii (Ct) y Phytophthora megasperma (Pm). Los objetivos fueron seleccionar bacterias con capacidad antifúngica contra Ct y Pm, e identificar una compatible con el rizobio INTA5 con baja emisión de N2O para constituir un consorcio bacteriano sustentable. Diez cepas mostraron capacidad antifúngica con medias de porcentaje de inhibición (PI) entre 41,74-91,28 para Ct y 56,23-77,59 relativos al control para Pm. B2, B4, SC6 y MN1 fueron seleccionadas por su mayor capacidad de biocontrol y se estudió la compatibilidad con INTA5. Sólo B4 resultó compatible, por ello fue evaluada contra Ct sobre plántulas de alfalfa en experimentos bajo condiciones controladas. Esta cepa incrementó el número de plántulas sobrevivientes cuando las mismas fueron inoculadas con Ct. La disponibilidad de un consorcio bacteriano que estimule tanto el biocontrol como la fijación biológica de nitrógeno representa la estrategia preferida por el programa de mejoramiento de alfalfa.

ABSTRACT Alfalfa breeding programs require environmentally friendly alternatives to improve plant growth and health. Plant Growth Promoting Rhizobacteria (PGPR) and endophytic bacteria offer a biological solution as they can inhibit phytopathogenic fungi by producing antifungal compounds or enzymes. In Argentina, alfalfa is primarily affected by Colletotrchum trifolii (Ct) and Phytophthora megasperma (Pm). The objectives of this study were to select bacterial isolates with antifungal inhibition against Ct and Pm and identify a biocontroller isolate compatible with the rhizobia INTA5 has and with low-N2O emissions, to constitute an eco-friendly bacterial consortium. Ten isolates demonstrated antifungal inhibition, with mean Inhibition Percentage (IP) values between 41.74-91.28 for Ct and 56.23-77.59 for Pm compared to the control. Among these isolates, B2, B4, SC6, and MN1 showed the highest inhibition performance and were selected to evaluate their compatibility with INTA5. Only B4 exhibited compatibility, which was further evaluated against Ct in an experiment with alfalfa seedlings under controlled conditions. The alfalfa seedlings treated with B4 increased the plant survival number against Ct. The availability of a bacterial consortium that promotes biocontrol and biological nitrogen fixation represents the preferred strategy for alfalfa breeding programs.

Resumo Fungos endofíticos colonizam as regiões inter e/ou intracelulares de tecidos vegetais saudáveis e possuem uma relação de simbiose com seus hospedeiros. Esses microrganismos produzem antibióticos, enzimas e outros compostos bioativos que os permitem sobreviver em habitats competitivos com outros microrganismos. Além disso, os metabólitos secundários conferem proteção à planta hospedeira contra outros patógenos bacterianos e fúngicos e/ou podem promover o crescimento vegetal. Os fungos endofíticos são considerados uma fonte promissora de produtos naturais bioativos, que podem ser otimizados por meio de mudanças nas condições de cultivo. A exploração de novas moléculas bioativas produzidas por esses microrganismos tem chamado a atenção dos pesquisadores. A diversidade química e funcional dos produtos naturais de fungos endofíticos exibe um amplo espectro de aplicações na medicina, agricultura, indústria e meio ambiente. Os fungos endofíticos também podem aumentar os efeitos fotoprotetores e a eficiência fotoquímica nas plantas hospedeiras. As abordagens ômicas modernas têm facilitado as pesquisas sobre as interações simbióticas entre plantas e fungos endofíticos. Portanto, a pesquisa sobre fungos endofíticos pode ajudar na descoberta de novas biomoléculas para diversas aplicações biotecnológicas e a desenvolver uma agricultura sustentável.

Abstract Endophytic fungi colonize the inter- and/or intracellular regions of healthy plant tissues and have a close symbiotic relationship with their hosts. These microorganisms produce antibiotics, enzymes, and other bioactive compounds that enable them to survive in competitive habitats with other microorganisms. In addition, secondary metabolites confer protection to their host plant against other bacterial and fungal pathogens and/or can promote plant growth. Endophytic fungi are viewed as a promising source of bioactive natural products, which can be optimized through changes in growing conditions. The exploration of novel bioactive molecules produced by these microorganisms has been attracting attention from researchers. The chemical and functional diversity of natural products from endophytic fungi exhibits a broad spectrum of applications in medicine, agriculture, industry and the environment. Fungal endophytes can also enhance the photoprotective effects and photochemical efficiency in the host plants. Modern omic approaches have facilitated research investigating symbiotic plant-endophytic fungi interactions. Therefore, research on endophytic fungi can help discovery novel biomolecules for various biotechnological applications and develop a sustainable agriculture.

Resumo Os fungos endofíticos são um grupo ubiquituo que colonizam todas as espécies de plantas na terra. Os estudos que comparam a localização dos fungos endofíticos dentro das folhas de Manihot esculenta Crantz (mandioca) e o tempo de amostragem são muito escassos. Neste estudo, folhas maduras de M. esculenta foram coletadas do Panamá com a finalidade de comparar a diversidade cultivável de endófitos e determinar sua distribuição dentro das folhas. Um total de 170 endófitos foram isolados de 97 espécies que representam 13 gêneros e 8 morfoespécies determinadas como micélios esterilizados contendo 63 isolados. Os fungos Cladosporium, Nigrospora, Periconia e mycelia sterilia 1 e 3 foram os isolados mais predominantes. Também detectamos que os endófitos variaram ao longo do tempo de amostragem, mas não entre os locais dentro das folhas. A composição de endófitos na amostragem e localização de endófitos dentro da folha foi semelhante, exceto para Periconia e mycelia sterilia 3 e 7. Os dados gerados neste estudo contribuem para o conhecimento da biodiversidade de fungos endofíticos no Panamá e estabelecem as bases para pesquisas sobre o entendimento da função de endófitos em culturas de M. esculenta.

Abstract Endophytic fungi are a ubiquituos group that colonize all plant species on earth. Studies comparing the location of endophytic fungi within the leaves and the sampling time in Manihot esculenta Crantz (cassava) are limited. In this study, mature leaves of M. esculenta from Panama were collected in order to compare the cultivable diversity of endophytic fungi and to determine their distribution within the leaves. A total of one hundred sixty endophytes belonging to 97 species representing 13 genera and 8 morphospecies determined as mycelia sterilia that containing 63 isolates were isolated. Cladosporium, Nigrospora, Periconia, and mycelia sterilia 1 and 3 were the most predominant isolated endophytes. We detected that endophytes varied across the sampling time, but not amongst locations within leaves. The endophytes composition across sampling and the location of endophytes within leaf was similar, except for Periconia and mycelia sterilia 3 and 7. The data generated in this study contribute to the knowledge on the biodiversity of endophytic fungi in Panama, and establish the bases for future research focused on understanding the function of endophytes in M. esculenta crops.

Abstract Endophytic microorganisms are those that inhabit the interior of plant tissues and organs without causing damage to the plant, triggering a mutualistic interaction. These endophytes are known to produce compounds with various applications in the areas of biotechnology, pharmaceutical industry and agriculture; therefore, molecular methods are used to investigate the phylogeny of these organisms. The Brazilian Cerrado is the second largest biome in Brazil and is considered a hotspot, with a high diversity of endemic fauna and flora. The objective of this work was to analyze, through in silico analysis, the phylogeny of cultivable endophytic fungi isolated from plants found in the Brazilian Cerrado biome. A search was carried out for articles that worked with endophytes in the period between 2012 and 2022. The dendrogram was generated using the Neighbor-joining method based on the ITS1-5.8S-ITS2 conserved region obtained by GenBank codes of isolated endophytes. The genera Xylaria, Diaporthe, and Colletotrichum were isolated in more than three plants in the seven articles found. Most of the articles found related to the biodiversity of endophytic fungi from the Brazilian Cerrado aim at the bioprospecting of bioactive compounds, through culture-dependent methods; as such, a part of endophytic diversity may be lost due to the inability of certain endophytes to grow in artificial media. In silico analysis can assist in the investigation of phylogenetic relationships between endophytic fungi and has the potential to guide future work aimed at prospecting for bioactive compounds, phylogenetic identification, or biodiversity of this group of endophytes. interaction biotechnology agriculture therefore organisms hotspot flora analyze 201 2022 Neighborjoining Neighbor joining ITS15.8SITS2 ITS158SITS2 ITSSITS ITS1 5.8S ITS2 5 8S ITS S ITS1-5.8S-ITS Xylaria Diaporthe culturedependent culture dependent such media identification 20 202 ITS15 8SITS2 SITS ITS15.8SITS ITS158SITS 58S 2 8SITS

Resumo Microrganismos endofíticos são aqueles que vivem no interior de tecidos e órgãos vegetais sem causar dano à planta, desencadeando uma interação mutualística. Esses endófitos são conhecidos por produzir compostos com diversas aplicações nas áreas de biotecnologia, indústria farmacêutica e agricultura; logo, métodos moleculares são utilizados para investigar a filogenia destes organismos. O Cerrado brasileiro é o segundo maior bioma do Brasil e é considerado um hotspot, com uma alta diversidade de fauna e flora endêmica. O objetivo deste trabalho foi analisar, por meio da análise in silico, a filogenia de fungos endofíticos cultiváveis isolados de plantas encontradas no bioma Cerrado brasileiro. Foi realizada uma busca por artigos que trabalharam com endófitos no período entre 2012 e 2022. O dendrograma foi gerado usando o método Neighbor-joining baseado na região conservada ITS1-5.8S-ITS2 obtida pelos códigos GenBank de endófitos isolados. Os gêneros Xylaria, Diaporthe e Colletotrichum foram isolados em mais de três plantas nos sete artigos encontrados. A maior parte dos artigos encontrados relacionados à biodiversidade de fungos endofíticos do Cerrado brasileiro visam à bioprospecção de compostos bioativos, através de métodos dependentes de cultura; como tal, uma parte da diversidade endofítica pode ser perdida devido à incapacidade de certos endófitos crescerem em meios artificiais. A análise in silico pode auxiliar na investigação das relações filogenéticas entre fungos endofíticos e tem potencial para orientar trabalhos futuros voltados à prospecção de compostos bioativos, identificação filogenética ou biodiversidade deste grupo de endófitos. planta mutualística biotecnologia agricultura logo organismos hotspot endêmica analisar 201 2022 Neighborjoining Neighbor joining ITS15.8SITS2 ITS158SITS2 ITSSITS ITS1 5.8S ITS2 5 8S ITS S ITS1-5.8S-ITS Xylaria bioativos cultura tal artificiais 20 202 ITS15 8SITS2 SITS ITS15.8SITS ITS158SITS 58S 2 8SITS

Resumo Este trabalho apresenta o isolamento de fungos endofíticos de folhas de Campomanesia adamantium(Cambess.) O. Berg (Myrtaceae), espécie nativa encontrada no Brasil, com distribuição abundante no Cerrado brasileiro, popularmente conhecida como “guabiroba-do-campo. Têm sido utilizadas popularmente por suas atividades anti-reumáticas anti-inflamatórias, antidiarreicas, redutoras de colesterol no sangue, antisséptica urinária e depurativas. Fungos endofíticos são microrganismos que vivem no interior de plantas superiores, ao menos por um período de seu ciclo de vida, ocupando os espaços intercelulares dos tecidos vegetais, como folhas e caules. Esses fungos, além de serem inofensivos à planta hospedeira, produzem benefícios a partir de metabólitos secundários que promovem proteção, regulam o crescimento, combatem bactérias, vírus e fungos e promovem resistência a estresse abiótico, além de efeitos inseticidas. Os fungos endófitos associados às folhas de C. adamantiumforam isolados utilizando-se a técnica de isolamento em meio de cultura. Logo após o crescimento, os fungos foram divididos em grupos baseados em morfotipos. O DNA genômico fúngico foi extraído A reação em cadeia da polimerase (PCR) foi conduzida para amplificação da região ITS-1-5.8S-ITS2 de rRNA e as sequências de nucleotídeos obtidas foram comparados com aqueles disponíveis na base de dados GenBank para a identificação molecular dos isolados. A construção da filogenia da árvore foi realizada utilizando o software MEGA 11. Os resultados apresentaram representantes do filo Ascomycota e em nível de gênero foi possível identificar 18 fungos dos gêneros Colletotrichum, Diaporthe, Epicoccum e Neofusicoccum. adamantiumCambess. adamantiumCambess adamantium Cambess. Cambess adamantium(Cambess. Myrtaceae, Myrtaceae , (Myrtaceae) Brasil brasileiro guabirobadocampo. guabirobadocampo guabiroba campo. campo “guabiroba-do-campo antireumáticas anti reumáticas antiinflamatórias, antiinflamatórias inflamatórias, inflamatórias anti-inflamatórias antidiarreicas sangue depurativas superiores vida vegetais caules hospedeira proteção crescimento bactérias abiótico inseticidas C utilizandose se cultura morfotipos PCR (PCR ITS15.8SITS2 ITS158SITS2 ITSSITS ITS 1 5.8S ITS2 5 8S S ITS-1-5.8S-ITS 11 Colletotrichum Diaporthe Neofusicoccum adamantium(Cambess (Myrtaceae ITS15 8SITS2 SITS ITS15.8SITS ITS158SITS 58S ITS1 8SITS

Abstract This work presents the isolation of endophytic fungi from the leaves of Campomanesia adamantium (Cambess.) O. Berg (Myrtaceae), a native species found in Brazil and popularly known as “guabiroba-do-campo”, with abundant distribution in the Brazilian Cerrado. It has been popularly used for its anti-rheumatic, anti-inflammatory, antidiarrheal, blood cholesterol-reducing, urinary antiseptic, and depurative properties. Theese fungi are microorganisms that live inside higher plants, at least for a period of their life cycle, occupying the intercellular spaces of plant tissues such as leaves and stems. These fungi are harmless to the host plant, and their secondary metabolites promote protection, regulate growth, combat bacteria, viruses, and fungi, and promote resistance to abiotic stress, as well as insecticidal effects. Endophytic fungi associated with the leaves of C. adamantium were isolated using the culture medium isolation technique. After growth, the fungi were divided into groups based on morphotypes. Fungal genomic DNA was extracted, and a polymerase chain reaction (PCR) was conducted to amplify the ITS1-5.8S-ITS2 regions of rRNA, and the nucleotide sequences obtained were compared with those available in the GenBank database for molecular identification of the isolates. The phylogenetic tree was constructed using MEGA 11 software. The results showed representatives of the Ascomycota phylum, and it was possible to identify at the genus level 18 fungi of the genera Colletotrichum, Diaporthe, Epicoccum, and Neofusicoccum. Cambess. Cambess (Cambess. O Myrtaceae, Myrtaceae , (Myrtaceae) guabirobadocampo, guabirobadocampo guabiroba do campo “guabiroba-do-campo” Cerrado antirheumatic, antirheumatic anti rheumatic, rheumatic anti-rheumatic antiinflammatory, antiinflammatory inflammatory, inflammatory anti-inflammatory antidiarrheal cholesterolreducing, cholesterolreducing cholesterol reducing, reducing cholesterol-reducing antiseptic properties plants cycle stems protection growth bacteria viruses stress effects C technique morphotypes extracted PCR (PCR ITS15.8SITS2 ITS158SITS2 ITSSITS ITS1 5.8S ITS2 5 8S ITS S ITS1-5.8S-ITS rRNA isolates 1 software phylum Colletotrichum Diaporthe Epicoccum Neofusicoccum (Cambess (Myrtaceae “guabiroba-do-campo ITS15 8SITS2 SITS ITS15.8SITS ITS158SITS 58S 8SITS

ABSTRACT Introduction: Bioproduct’s development to beneficiate economic importance crops such rice, contributes directly to our food sovereignty. The objective of the work was: To obtain and characterize bacterial strains associated with two rice cultivars widely distributed in Cuba, in terms of their potential as Plant Growth Promoting Bacteria. Methods: Bacteria associated to the rhizosphere and seeds of INCA LP-5 and INCA LP-7 rice cultivars were isolated and identified by partial 16S rRNA sequencing. The strains were characterized in terms of potentialities as biofertilizers, phytostimulants, biocontrol, and attributes of infection and colonization. The endophytic capacity of a Rhizobium strain was studied in rice seedlings, from inoculation tests and confocal fluorescence microscopy. Inoculation experiments were carried out under controlled, semi-controlled and field conditions. Results: We isolated 43 strains; 24 from the rhizosphere and 19 from rice seeds and eight associated bacterial genera were identified: Rhizobium, Pantoea, Pseudomonas, Acinetobacter, Mitsuaria, Enterobacter, Bacillus and Paenibacillus. Some strains solubilized calcium and potassium phosphate, grew in nitrogen-free media, and produced siderophores, indole compounds, hydrolytic enzymes, and formed biofilms. The endophytic capacity of a Rhizobium strain was verified by systemic colonization of rice seedlings. The inoculation of some strains increased the content of nutrients, some biomolecules and rice plant growth. Furthermore, the crop yield enhanced between 30% - 80% with reduction of 40% - 60% of nitrogen fertilization. Conclusions, the research constitutes a contribution to the knowledge of Rhizobium-rice interaction and in the identification, characterization and selection of bacterial strains with potential for the development of bioproducts that positively impact the environment, the economy and the society.

RESUMEN Introducción: El desarrollo de bioproductos para beneficiar cultivos de importancia económica como el arroz tributa directamente a nuestra soberanía alimentaria. El objetivo del trabajo fue obtener y caracterizar cepas bacterianas asociadas a 2 cultivares de arroz de amplia distribución en Cuba, en cuanto a sus potencialidades como bacterias promotoras del crecimiento vegetal. Métodos: Se realizó el aislamiento de bacterias asociadas a la rizosfera y semillas de los cultivares de arroz INCA LP-5 e INCA LP-7 y se identificaron por secuenciación parcial del ARNr 16S. Las cepas se caracterizaron en cuanto a potencialidades como biofertilizantes, fitoestimulantes, de biocontrol y atributos de infección y colonización. Se estudió la capacidad endofítica de una cepa de Rhizobium en plántulas de arroz, a partir de ensayos de inoculación y microscopía confocal de fluorescencia. Se desarrollaron experimentos de inoculación en condiciones controladas, semicontroladas y de campo. Resultados: Se aislaron 43 cepas; 24 de la rizosfera y 19 del interior de semillas de arroz y se identificaron 8 géneros bacterianos asociados: Rhizobium, Pantoea, Pseudomonas, Acinetobacter, Mitsuaria, Enterobacter, Bacillus y Paenibacillus. Algunas cepas solubilizaron fosfato de calcio y potasio, crecieron en medios libres de nitrógeno y produjeron sideróforos, compuestos indólicos, enzimas hidrolíticas y formaron biopelículas. Se comprobó la capacidad endofítica de una cepa de Rhizobium y su colonización sistémica en plántulas de arroz. La inoculación de algunas cepas incrementó el contenido de nutrientes, algunas biomoléculas y el crecimiento de plantas de arroz. Además, el rendimiento del cultivo se favoreció entre 30 % y 80 %, con la reducción del 40 % a 60 % de la fertilización nitrogenada. Conclusiones, la investigación constituye un aporte al conocimiento de la interacción Rhizobium-arroz y a la identificación, caracterización y selección de cepas bacterianas con potencialidades para el desarrollo de bioproductos que impacten positivamente en el medio ambiente, la economía y la sociedad.

Resumen Los microorganismos están naturalmente asociados con las plantas. El presente experimento se llevó a cabo para aislar bacterias endófitas de las células internas de raíces, tallos, hojas y Tejido capitulear de Senecio glaucus recolectadas en 2 hábitats diversos (costeros y desérticos) de Egipto. Del aislamiento se obtuvieron un total de 10 bacterias endófitas; la mayor diversidad de endófitos bacterianos se observó en raíces y hojas de muestras del desierto. Los aislamientos se reconocieron con base en la morfología, la bioquímica y los genes de la secuencia del ARNr 16S. Todos estos aislados indican la capacidad de producir enzimas como amilasa, celulasa, lipasa, catalasa y proteasa en sus descripciones bioquímicas; los análisis también mostraron una indicación significativa de su potencial para producir hormonas de crecimiento vegetal; como su capacidad para disolver el fosfato. En el mundo y en Egipto, somos los primeros en reportar endófitos bacterianos aislados de Senecio glaucus. Este estudio podría ayudar a determinar el papel de las bacterias endófitas en hábitats severos, así como sus posibles aplicaciones en medicina, biorremediación, agricultura e industria.

Abstract Microorganisms are naturally associated with plants in several ways. The study was conducted to isolate bacteria endophytes from the internal cells of roots, stems, leaves, and capitula of Senecio glaucus collected from 2 diverse (coastal and desert) habitats in Egypt. A total of 10 endophytic bacteria were obtained from the isolation; the highest diversity of bacterial endophytes was observed in desert samples roots and leaves. The isolates were recognized based on morphology, biochemical and 16S rRNA sequence genes. All isolates indicated the ability for enzyme production as amylase, cellulase, lipase, catalase, and protease in their biochemical descriptions; analyses also gave a significant indication of their potential to produce plant growth hormones, as their ability to dissolve Phosphate. In the world and Egypt, we are the first to report bacterial endophytes isolated from Senecio glaucus. This study could aid in determining the role of endophytic bacteria in severe habitats, as well as their potential applications in medicine, bioremediation, agriculture, and industry.

Resumo As bactérias endofíticas desempenham papel fundamental na manutenção da saúde e do crescimento das plantas. Poucos estudos até o momento, no entanto, exploraram as propriedades antagônicas e promotoras de crescimento de plantas (PGP) de endófitos de Prunus cerasifera. Para esse fim, isolamos bactérias endofíticas de amostras de tecido de P. cerasifera e usamos um ensaio de placa de cultura dupla para rastrear esses micróbios quanto à atividade antagonista contra Verticillium dahliae, Botryosphaeria dothidea, Fusarium oxysporum, F. graminearum e F. moniliforme. Das 36 cepas de bactérias isoladas, quatro (cepas P1, P10, P16 e P20) exibiram efeitos antagônicos contra todos os cinco patógenos modelo, e a cepa P10 exibiu o antagonista mais forte para cinco patógenos. Essa cepa P10 foi então caracterizada em profundidade por meio de avaliações fenotípicas, análises fisiológicas e sequenciamento de rDNA 16s, revelando ser uma cepa de Bacillus subtilis. A aplicação de uma suspensão de células P10 (1 × 108 UFC / mL) aumentou significativamente a germinação das sementes e o crescimento das mudas de tomate em casa de vegetação. Essa cepa P10 suprimiu ainda mais a murcha de Verticillium do tomate com incidência de doença muito menor e pontuações de índice de doença sendo observadas após o tratamento com P10 em relação a plantas não tratadas em experimentos baseados em vasos. As plantas de tomate que foram tratadas com a cepa P10 também aumentaram as enzimas relacionadas à defesa, peroxidase, superóxido dismutase e atividade da catalase após o desafio de V. dahliae em relação às plantas que não foram tratadas com essa bactéria endofítica. Os resultados revelaram que a cepa bacteriana P10 tem valor potencial como agente de biocontrole para uso na prevenção da murcha de Verticillium em tomate.

Abstract Endophytic bacteria serve key roles in the maintenance of plant health and growth. Few studies to date, however, have explored the antagonistic and plant growth-promoting (PGP) properties of Prunus cerasifera endophytes. To that end, we isolated endophytic bacteria from P. cerasifera tissue samples and used a dual culture plate assay to screen these microbes for antagonistic activity against Verticillium dahliae, Botryosphaeria dothidea, Fusarium oxysporum, F. graminearum, and F. moniliforme. Of the 36 strains of isolated bacteria, four (strains P1, P10, P16, and P20) exhibited antagonistic effects against all five model pathogens, and the P10 strain exhibited the strongest antagonistic to five pathogens. This P10 strain was then characterized in-depth via phenotypic assessments, physiological analyses, and 16s rDNA sequencing, revealing it to be a strain of Bacillus subtilis. Application of a P10 cell suspension (1×108 CFU/mL) significantly enhanced the seed germination and seedling growth of tomato in a greenhouse setting. This P10 strain further significantly suppressed tomato Verticillium wilt with much lower disease incidence and disease index scores being observed following P10 treatment relative to untreated plants in pot-based experiments. Tomato plants that had been treated with strain P10 also enhanced defense-related enzymes, peroxidase, superoxide dismutase, and catalase activity upon V. dahliae challenge relative to plants that had not been treated with this endophytic bacterium. The results revealed that the P10 bacterial strain has potential value as a biocontrol agent for use in the prevention of tomato Verticillium wilt.

Abstract Extraradical spores of arbuscular mycorrhizal (AM) fungi are often parasitized by a wide range of soil microorganisms. However, reports on the parasitization of intraradical structures of AM fungi are very rare. While examining AM colonization in roots of Cuminum cyminum and Sambucus nigra growing in the medicinal plants garden of the Centre of Medicinal Plants Research in Homeopathy of the Nilgiris, we found that AM fungal hyphal coils, arbusculate coils, vesicles, and intraradical spores are colonized by dark septate endophytic (DSE) fungi which were co-occurring with AM fungi. The AM fungal hyphae were significantly wider than the DSE fungi. Colonization of different AM fungal structures significantly varied between the two plant species. Moreover, the percentage of colonized AM fungal structures was linearly correlated to the abundance of these structures. The colonization of the intraradical structures of AM fungi is illustrated and the significance of this phenomenon needs further elucidation. (AM microorganisms However rare Nilgiris coils vesicles (DSE cooccurring co occurring species Moreover elucidation

Resumen Las esporas extrarradicales de hongos micorrícicos arbusculares (AM) a menudo son parasitados por una amplia gama de microorganismos del suelo. Sin embargo, los informes sobre la parasitación de estructuras intrarradicales de hongos AM son muy raros. Al examinar la colonización de AM en raíces de Cuminum cyminum y Sambucus nigra que crecen en el jardín de plantas medicinales del Centro de Investigación de Plantas Medicinales en Homeopatía de Nilgiris, encontramos bobinas de hifas de hongos AM, bobinas arbusculadas, vesículas y esporas intrarradicales colonizadas por hongos endófitos septados oscuros (DSE) que coexistían con hongos AM. Las hifas de hongos AM eran significativamente más anchas que los hongos DSE. La colonización de diferentes estructuras fúngicas AM varió significativamente entre las dos especies de plantas. Además, el porcentaje de estructuras fúngicas AM colonizadas se correlacionó linealmente con la abundancia de estas estructuras. Se ilustra la colonización de las estructuras intrarradicales de hongos AM y la importancia de este fenómeno necesita mayor aclaración. (AM suelo embargo raros Nilgiris arbusculadas DSE (DSE Además aclaración

Resumen Molina, L.; M. Rajchenberg, M. C. Aime & M. B. Pildain. 2023. Evaluación de la diversidad de endófitos fúngicos: un estudio comparativo de tres flujos de trabajo automatizados de metabarcoding. Darwiniana, nueva serie 11(2): 402-419. La utilización de la secuenciación de alto rendimiento se ha vuelto frecuente en el estudio de comunidades endófitas. Estas tecnologías han permitido la descripción acumulativa de diversidad fúngica a lo largo de la última década. No obstante, también han implicado nuevos desafíos para los investigadores de las áreas involucradas en términos de la necesidad de contar con herramientas de programación y habilidades desarrolladoras. Hoy en día no existe un consenso sobre las herramientas bioinformáticas más adecuadas para procesar los datos crudos de secuencias que estas tecnologías arrojan. El objetivo de este trabajo fue comparar el rendimiento de tres flujos de trabajo realizados en dos plataformas gratuitas diseñadas para ser amigables con usuarios que no son programadores y desarrolladas específicamente para estudios de hongos: AMPtk y PIPITS. Evaluamos los ensambles de hongos que habitan en la albura de dos especies de Nothofagus de los bosques patagónicos y comparamos el conjunto de datos de metabarcoding del espaciador transcrito interno (ITS) con un conjunto de datos de secuencias existente, obtenido de la prospección de cultivos de los mismos árboles y sitios de estudio. La plataforma AMPtk se desempeñó mejor con respecto a la descripción de la comunidad, en términos de precisión del agrupamiento de taxones, principalmente debido al algoritmo DADA2. El flujo de trabajo PIPITS evidenció una mayor sensibilidad en la detección de taxones conocidos presentes, por lo que es potencialmente útil para futuros estudios que persigan la detección de taxones específicos. Debido a la falta de información que exhiben las bases de datos de referencia sobre el ecosistema en estudio, ambas plataformas tuvieron un desempeño deficiente en cuanto a la asignación de taxonomías. Es imperativo seguir estudiando estos ecosistemas y mejorar las bases de datos para aumentar el potencial explicativo de las nuevas tecnologías.

Abstract Molina, L.; M. Rajchenberg, M. C. Aime & M. B. Pildain. 2023. Assessing fungal endophyte diversity: a comparative study of three automated metabarcoding pipelines. Darwiniana, nueva serie 11(2): 402-419. High-throughput sequencing approaches have become frequent in the study of endophyte communities allowing the cumulative description of fungal diversity in the last decade. However, they brought new challenges to researchers in terms of programming and developing of informatics tools. Currently, there is no consensus concerning the appropriate bioinformatics to process such sequence data. The aim of this study was to compare the performance of three pipelines of two cost-free toolkits designed to be friendly to non-programmer users, and specifically developed for fungal data: AMPtk and PIPITS. The sapwood-inhabiting fungal assemblages of two Nothofagus species from the Patagonian Forests were assessed through metabarcoding of the internal transcribed spacer (ITS) and compared with an extant sequence dataset obtained from culture prospection in the same study sites and trees. The AMPtk toolkit has performed better concerning community description in terms of precision of taxa clustering, mainly due to the DADA2 algorithm; PIPITS evidenced a higher sensitivity in detecting taxa known to be present, hence it is potentially useful for future specific taxa detection surveys. Because of a current lack of information of the reference databases, both bioinformatic toolkits performed poorly as to taxonomy assignment. It is imperative to continue studying these ecosystems to, concomitantly, improve databases and the explanatory potential of the new technologies.

RESUMEN Nuestro objetivo fue evaluar la producción y partición de biomasa de plantas de Festuca arundinacea (festuca alta) infectadas con el hongo endófito Epichloë coenophiala e inoculadas con hongos micorrícicos arbusculares (HMA) en respuesta a la fertilización fosfatada (P). Para ello, se realizó un experimento en macetas en cámara de crecimiento, a través de un diseño completamente aleatorizado con arreglo factorial con 5 repeticiones. Se evaluó el crecimiento de F. arundinacea ante la ausencia o presencia de E. coenophiala (SE- y SE+, respectivamente), la inoculación con HMA (HMA+ y HMA-, con y sin inoculación, respectivamente) y la fertilización con fósforo (P15 y P0, con y sin fertilización, respectivamente). Luego de 160 días de crecimiento se registró la biomasa fresca total (BFT) y su partición en biomasa fresca aérea (BFA) y radical (BFR), y el peso seco aéreo (PSA). Además, se cuantificó la colonización micorrícica en raíces de festuca alta. Las plantas HMA+ produjeron menor BFT, BFA y BFR que las HMA- en las condiciones SE- y P0. No se detectaron diferencias en BFT y PSA entre plantas HMA+ y HMA- en P15. En P0, las plantas HMA- presentaron mayor PSA que las HMA+. Las plantas SE- presentaron mayor PSA que las SE+. En las plantas SE+ inoculadas con HMA para la condición P0, se registró abundante micelio extraradical, aunque no se evidenció colonización interradical ni formación de arbúsculos o vesículas. En P15, solamente en una muestra de SE- se observó micelio extraradical alrededor de las raíces (que presentaron menor longitud en relación a las raíces de las plantas sin fertilizar). Bajo las condiciones del presente experimento, no se registraron las respuestas esperadas en producción de biomasa en plantas infectadas con el hongo endófito y la inoculación con HMA ni sinergismo entre ambos simbiontes. Además, no se evidenció efecto modulador de la fertilización fosfatada en las respuestas esperadas.

SUMMARY Our objective was to evaluate the biomass production and partitioning of Festuca arundinacea (tall fescue) plants infected with endophyte Epichloë coenophiala and inoculated with arbuscular mycorrhizal fungi (HMA) in response to phosphate fertilization (P). For this, an experiment was carried out in pots in a growth chamber, through a completely randomized design with a factorial arrangement with 5 repetitions. The growth of F. arundinacea was evaluated in presence or not of E. coenophiala (SE- and SE+, respectively), inoculated with HMA (HMA+ and HMA-, with and without inoculation, respectively) and fertilized with phosphorus (P15 and P0, with and without fertilization, respectively). After 160 days of growth, the total fresh biomass (TFB) and its partitioning into aboveground (AFB) and belowground (BFB), and air dry weight (ADW) were quantified. Mycorrhizal colonization was quantified in roots. HMA+ plants showed lower BFT, BFA and BFR than HMA- plants in SE- and P0 conditions. No differences in BFT and PSA were detected between HMA+ and HMA- plants at P15. At P0, HMA- plants showed higher PFA than HMA+ plants. SE- plants showed higher PSA than SE+ plants. In SE+ plants inoculated with HMA for the P0 condition, abundant extraradical mycelium was recorded, although there was no evidence of interradical colonization or formation of arbuscules or vesicles. At P15, extraradical mycelium was observed around the roots (which were shorter than the roots of unfertilized plants) in only one sample of SE-. Under the conditions of the present experiment, expected responses in biomass production in plants infected with the endophyte fungus and inoculation with HMA and synergism between both symbionts were not recorded. Furthermore, no modulating effect of phosphate fertilization on the expected responses was found.