Fonsecaea and Cladophialophora are genera of black yeast-like fungi harboring agents of a mutilating implantation disease in humans, along with strictly environmental species. The current hypothesis suggests that those species reside in somewhat adverse microhabitats, and pathogenic siblings share virulence factors enabling survival in mammal tissue after coincidental inoculation driven by pathogenic adaptation. A comparative genomic analysis of environmental and pathogenic siblings of Fonsecaea and Cladophialophora was undertaken, including de novo assembly of F. erecta from plant material. The genome size of Fonsecaea species varied between 33.39 and 35.23 Mb, and the core genomes of those species comprises almost 70% of the genes. Expansions of protein domains such as glyoxalases and peptidases suggested ability for pathogenicity in clinical agents, while the use of nitrogen and degradation of phenolic compounds was enriched in environmental species. The similarity of carbohydrate-active vs. protein-degrading enzymes associated with the occurrence of virulence factors suggested a general tolerance to extreme conditions, which might explain the opportunistic tendency of Fonsecaea sibling species. Virulence was tested in the Galleria mellonella model and immunological assays were performed in order to support this hypothesis. Larvae infected by environmental F. erecta had a lower survival. Fungal macrophage murine co-culture showed that F. erecta induced high levels of TNF-α contributing to macrophage activation that could increase the ability to control intracellular fungal growth although hyphal death were not observed, suggesting a higher level of extremotolerance of environmental species.

Four strains of rhizobia isolated from nodules of Vigna unguiculata (UFLA03-321, UFLA03-320 and UFLA03-290) and Macroptilium atropurpureum (UFLA04-0212) in Brazilian soils were previously reported as a new group within the genus Bradyrhizobium. To determine their taxonomic position, these strains were characterized in this study using a polyphasic approach. The analysis of the 16S rRNA gene grouped the four strains with Bradyrhizobium pachyrhizi PAC48. However, the concatenated sequence analysis of the two (recA and glnII) or three (atpD, gyrB and recA) housekeeping genes indicated that these strains represent a novel species of Bradyrhizobium, which is very closely related to B. pachyrhizi PAC48 and B. elkanii USDA 76. Genomic relatedness analyses between the UFLA03-321 strain and B. elkanii USDA 76 and B. pachyrhizi PAC48 revealed an average nucleotide identity below 96% and values of estimated DNA-DNA hybridization below 70%, confirming that they represent genomically distinct species. Analysis of MALDI-TOF MS (Matrix-assisted laser desorption ionization-time-of-flight mass spectrometry) profiles and phenotypic characteristics also allowed differentiation of the novel species from its two neighboring species. In phylogenetic analysis of nodC and nifH genes, UFLA03-321 exhibited maximum similarity with B. tropiciagri CNPSo 1112. The data suggest that these four UFLA strains represent a novel species, for which the name Bradyrhizobium brasilense sp. nov. is proposed, with UFLA03-321 (=LMG 29353 =CBAS645) as type strain. G + C content in the DNA of UFLA03-321 is 63.9 mol %.

Cowpea (Vigna unguiculata L.) is a legume species that considerably benefits from inoculation with nitrogen fixing bacteria of the genus Bradyrhizobium. One of the strains recommended for inoculation in cowpea in Brazil is UFLA03-84 (Bradyrhizobium sp.). The aim of our study was to define the taxonomic position of the UFLA03-84 strain and of two other strains of Bradyrhizobium (UFLA03-144 and INPA237B), all belonging to the same phylogenetic group and isolated from soils of the Brazilian Amazon. Multilocus sequence analysis (MLSA) of the housekeeping genes atpD, gyrB, recA, and rpoB grouped (with similarity higher than 99%) the three strains with Bradyrhizobium viridifuturi SEMIA 690 T. The analyses of average nucleotide identity and digital DNA-DNA hybridization supported classification of the group as Bradyrhizobium viridifuturi. The three strains exhibited similar behavior in relation to the most of the phenotypic characteristics evaluated. However, some characteristics exhibited variation, indicating phenotypic diversity within the species. Phylogenetic analysis of the nodC and nifH genes showed that the three strains are members of the same symbiovar (tropici) that contains type strains of Bradyrhizobium species coming from tropical soils (SEMIA 690 T B. viridifuturi, CNPSo 1112 T B. tropiciagri, CNPSo 2833 T B. embrapense, and B. brasilense UFLA03-321 T).

The species belonging to the genus Fonsecaea are the main causative agents of chromoblastomycosis. The invasive potential of Fonsecaea differs significantly among its various sibling species. Moreover, the lack of clarity on the virulence and availability of precise markers to distinguish and detect Fonsecaea species is attributed to the different ways of dissemination and pathogenicity. Therefore, the present study aimed to propose new molecular tools to differentiate between sibling species causing chromoblastomycosis. We used an infection model of chromoblastomycosis in BALB/c to study speciesspecific molecular markers for the in vivo detection of Fonsecaea species in biological samples. Specific primers based on the CBF5 gene were developed for Fonsecaea pedrosoi, Fonsecaea monophora, Fonsecaea nubica, and Fonsecaea pugnacius. In addition, a padlock probe was designed for F. pugnacius based on ITS sequences. We also assessed the specificity of Fonsecaea species using in silico, in vitro, and in vivo assays. The results showed that markers and probes could effectively discriminate the species in both clinical and environmental samples, enabling Handling Editor: Abdullah Mohammed Said Al-Hatmi.Mycopathologia (2019) 184:493-504 https://doi.org/10.1007/s11046-019-00359-2( 0123456789().,-volV) ( 01234567 89().,-volV) bioprospecting of agents of chromoblastomycosis, thereby elucidating the infection route of the disease.

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