Delimitation of species and the search for a proper threshold for defining phylogenetic species in fungi are under discussion. In this study, morphological and molecular data are correlated to delimit species of Tulasnella, the most important mycobionts of Orchidaceae, which suffer from poor taxonomy. Resupinate basidiomata of Tulasnella species were collected in Ecuador and Germany, and 11 specimens (seven from Ecuador, four from Germany) were assigned to traditional species concepts by use of morphological keys. The specimens were compared by micro-anatomical examination with 75 specimens of Tulasnella borrowed from fungaria to obtain better insights on variation of characters. Sequences of the ITS region (127) were obtained after cloning from the fresh basidiomata and from pure cultures. Proportional variability of ITS sequences was analyzed within and among the cultures and the specimens designated to different morphospecies. Results suggested an intragenomic variation of less than 2%, an intraspecific variation of up to 4% and an interspecific divergence of more than 9% in Tulasnella. Cryptic species in Tulasnella, mostly from Ecuador, were revealed by phylogenetic analyses with 4% intraspecific divergence as a minimum threshold for delimiting species. Conventional diagnostic morphological characters appeared insufficient for species characterization. Arguments are presented for molecular delimitation of the established species Tulasnella albida, T. asymmetrica, T. eichleriana, T. cf. pinicola, T. tomaculum and T. violea.

• Premise of the study: The presence of compatible fungi is necessary for epiphytic orchid recruitment. Thus, identifying associated mycorrhizal fungi at the population level is essential for orchid conservation. Recruitment patterns may also be conditioned by factors such as seed dispersal range and specific environmental characteristics.• Methods: In a forest plot, all trees with a diameter at breast height >1 cm and all individuals of the epiphytic orchid Epidendrum rhopalostele were identified and mapped. Additionally, one flowering individual of E. rhopalostele per each host tree was randomly selected for root sampling and DNA extraction.• Key results: A total of 239 E. rhopalostele individuals were located in 25 of the 714 potential host trees. Light microscopy of sampled roots showed mycorrhizal fungi in 22 of the 25 sampled orchids. Phylogenetic analysis of ITS1‐5.8S‐ITS2 sequences yielded two Tulasnella clades. In four cases, plants were found to be associated with both clades. The difference between univariate and bivariate K functions was consistent with the random labeling null model at all spatial scales, indicating that trees hosting clades A and B of Tulasnella are not spatially segregated. The analysis of the inhomogenous K function showed that host trees are not clustered, suggesting no limitations to population‐scale dispersal. χ² analysis of contingency tables showed that E. rhopalostele is more frequent on dead trees than expected.• Conclusions: Epidendrum rhopalostele establishes mycorrhizal associations with at least two different Tulasnella species. The analysis of the distribution patterns of this orchid suggests a microsite preference for dead trees and no seed dispersal limitation.

Bacterial resistance to antibiotics is a serious public health problem that needs new antibacterial compounds for control. Fungi, including resupinated fungi, are a potential source to discover new bioactive compounds efficient again to bacteria resistant to antibiotics. The inhibitory capacity against the bacterial species was statistically evaluated. All the species (basidiomata and strains) were molecularly characterized with the ITS1-5.8S-ITS2 barcoding marker. The strains Ceraceomyces sp., Fuscoporia sp., Gloeocystidiellum sp., Oliveonia sp., Phanerochaete sp., and Xenasmatella sp. correspond to resupinate Basidiomycetes, and only the strain Hypocrea sp. is an Ascomycete, suggesting contamination to the basidiome of Tulasnella sp. According to the antagonistic test, only the Gloeocystidiellum sp. strain had antibacterial activity against the bacterial species Escherichia coli of clinical interest. Statistically, Gloeocystidiellum sp. was significantly (<0.001) active against two E. coli pathotypes (O157:H7 and ATCC 25922). Contrarily, the antibacterial activity of fungi against other pathotypes of E. coli and other strains such as Serratia sp. was not significant. The antibacterial activity between 48 and 72 h increased according to the measurement of the inhibition halos. Because of this antibacterial activity, Gloeocystidiellum sp. was taxonomically studied in deep combined morphological and molecular characterization (ITS1-5.8S-ITS2; partial LSU D1/D2 of nrDNA). A new species Gloeocystidiellum lojanense, a resupinate and corticioid fungus from a tropical montane rainforest of southern Ecuador, with antibacterial potential against E. coli, is proposed to the science.

Organic and conventional agriculture are vital for the development of human society; however, the use of contaminated water and the inappropriate use of organic chemical fertilizers can lead to an increase in the microbial load (potentially pathogenic) of the normal microbiota of the agricultural soil. In this context, the aim of our study was to isolate Gram-negative bacteria from the superficial soil layer and irrigation water of agricultural areas (11 organic farms and nine conventional farms) and consider their potential ecological and health risk importance. Through culture isolation using three bacterial media (TSA) trypticase soy agar (general nutritive media); MacConkey Gram-negative bacteria and (EMB) eosin methylene blue agar (selective for Enterobacteriaceae) and classical biochemical tests, we recorded a total of 12 bacterial species, most belonging to the Enterobacteriaceae family, such as Enterobacter, Escherichia, Klebsiella, Salmonella and Shigella, which can be pathogenic for humans and animals. In contrast, bacteria such as Pantoea agglomerans, Pseudomonas aeruginosa, P. fluorescens and Burkholderia mallei could facultatively work as diazotrophic or plant growth-promoting rhizobacteria. Soil bacteria richness detected with the media applied was significantly higher than water bacteria, but we found no significant differences between organic and conventional agriculture. We conclude that the isolated bacteria in water and soil mostly belongs to enteropathogenic bacteria which could be pathogenic to animals and humans. While other bacteria like Pseudomonas aeruginosa could be viewed as useful by improving nutrient availability in agricultural soil.

Coffee (Coffea arabica), produced and marketed in Ecuador and worldwide, can be organoleptically improved by means of microorganisms such as well-characterized yeasts. This study aimed to isolate and characterize yeasts from three postharvest fermentation processes (i.e., Natural aerobic at room temperature; Carbonic maceration with a CO₂ atmosphere at room temperature; and Carbonic refrigerated maceration with a CO₂ atmosphere to 10 °C) of coffee fruits in Ecuador. Phenotypic and molecular analyses were conducted on 329 yeast isolates obtained from coffee farms in Loja, Olmedo, and Gonzanamá. Three universal media were used for yeast isolation diversity, and phenotypic characterization included morphology, sugar fermentation, salt tolerance, and ethanol resistance. Molecular characterization involved DNA analysis. The isolated diversity was classified into 12 morphotypes, nine distinct biochemical groups and nine genetic species. Only six species (i.e., Kurtzmaniella quercitrusa, Hanseniaspora opuntiae, Pichia. kluyveri, Torulaspora delbrueckii, T. quercuum, and Wickerhamomyces anomalus) identified phylogenetically corresponded to the designated morphotypes. But surprisingly, nine genetic species matched with the nine biochemical groups determined phenotypically analyzed using principal component analysis (PCA). Most of this diversity was found in the coffee plantation located in Gonzanamá, in contrast to Olmedo and Loja, without statistical significance (p value: 0.08295). On the other hand, the richness is not similar statistically (p value: 0.02991) between postharvest fermentation treatments. The findings suggest that the application of biochemical tests is useful for species determination, although morphological data may be ambiguous. Notably, Pichia kluyveri, detected in this study, holds potential for biotechnological evaluation in coffee fermentation processes.