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The Iron Quadrangle (IQ) is one of the main iron ore producing regions of the world. The exploitation of its reserves jeopardizes the high biological endemism associated with this region. This work aimed to understand the diversity and bacterial potential associated with IQ caves. Floor and ceiling samples of seven ferruginous caves and one quartzite cave were collected, and their microbial relative abundance and diversity were established by 16S rRNA gene amplicon sequencing data. The results showed that ferruginous caves present higher microbial abundance and greater microbial diversity compared to the quartzite cave. Many species belonging to genera found in these caves, such as Pseudonocardia and Streptacidiphilus, are known to produce biomolecules of biotechnological interest as macrolides and polyketides. Moreover, comparative analysis of microbial diversity and metabolic potential in a biofilm in pendant microfeature revealed that the microbiota associated with this structure is more similar to the floor rather than ceiling samples, with the presence of genera that may participate in the genesis of these cavities, for instance, Ferrovum, Geobacter, and Sideroxydans. These results provide the first glimpse of the microbial life in these environments and emphasize the need of conservation programs for these areas, which are under intense anthropogenic exploration.

Digital, Biocontrol and plant growth-promoting bacteria (PGPB) are important agricultural bioinoculants. This study aimed to prospect new potential bioinoculants for a more sustainable agriculture from ferruginous caves of the Brazilian Iron Quadrangle. Culturable bacteria, from seven caves and one canga soil sample, were evaluated for biocontroller activity of the phytopathogens Xanthomonas citri subsp. Citri—Xcc306 (citrus canker), Fusarium oxysporum—Fo (fusariosis), and Colletotrichum lindemuthianum—Cl89 (bean anthracnose). The ability of the superior candidates to solubilize inorganic phosphate, fix nitrogen, and produce hydrolytic enzymes and siderophores was then analyzed. Out of 563 isolates, 47 inhibited the growth of Xcc306 in vitro, of which 9 reduced citrus canker up to 68% when co-inoculated with the pathogen on host plants. Twenty of the 47 inhibited Fo growth directly by 51–73%, and 15 indirectly by 75–81%. These 15 inhibited Cl89 growth in vitro (up to 93% directly and 100% indirectly), fixed nitrogen, produced proteases and siderophores, showed motility ability, produced biofilm, and all but one solubilized inorganic phosphate. Therefore, 15 (2.66%) bacterial isolates, from the genera Serratia, Nissabacter, and Dickeya, act simultaneously as biocontrollers and PGPBs, and could be important candidates for future investigations in planta as an alternative to minimize the use of pesticides and chemical fertilizers through sustainable agricultural management practices., Ciencias Exactas y Naturales

Serratia liquefaciens strain FG3 (SlFG3), isolated from the flower of Stachytarpheta glabra in the Brazilian ferruginous fields, has distinctive genomic, adaptive, and biotechnological potential. Herein, using a combination of genomics and molecular approaches, we unlocked the evolution of the adaptive traits acquired by S1FG3, which exhibits the second largest chromosome containing the largest conjugative plasmids described for Serratia. Comparative analysis revealed the presence of 18 genomic islands and 311 unique protein families involved in distinct adaptive features. S1FG3 has a diversified repertoire of genes associated with Nonribosomal peptides (NRPs/PKS), a complete and functional cluster related to cellulose synthesis, and an extensive and functional repertoire of oxidative metabolism genes. In addition, S1FG3 possesses a complete pathway related to protocatecuate and chloroaromatic degradation, and a complete repertoire of genes related to DNA repair and protection that includes mechanisms related to UV light tolerance, redox process resistance, and a laterally acquired capacity to protect DNA using phosphorothioation. These findings summarize that SlFG3 is well-adapted to different biotic and abiotic stress situations imposed by extreme conditions associated with ferruginous fields, unlocking the impact of the lateral gene transfer to adjust the genome for extreme environments, and providing insight into the evolution of prokaryotes. Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior -Brazil (CAPES)CAPES [001, CFP 51/2013, 3385/2013]; National Council of Technological and Scientific Development (CNPq)National Council for Scientific and Technological Development (CNPq) [481226/2013-3]; Foundation of Protection to Research of the State of Minas Gerais - FAPEMIG [APQ-02387-14, APQ-02357-17]; CNPqNational Council for Scientific and Technological Development (CNPq); Fundect-MS [TO 141/2016, TO 007/2015, 007/2015 SIAFEM 025139]; UFOP grants; Laboratory of Biochemistry and Molecular Biology of the Faculty of Agrarian and Veterinary Sciences UNESP, Jaboticabal campus Thanks to all members of the Laboratory of Biochemistry and Molecular Biology (LBBM, Federal University of Ouro Preto, UFOP) and to the Laboratory of Biochemistry and Molecular Biology of the Faculty of Agrarian and Veterinary Sciences UNESP, Jaboticabal campus, for their support. This study was financed in part by the Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior -Brazil (CAPES) - Finance Code 001 - (the BIGA Project, CFP 51/2013, process 3385/2013), National Council of Technological and Scientific Development (CNPq Process 481226/2013-3), and Foundation of Protection to Research of the State of Minas Gerais - FAPEMIG (process APQ-02387-14 and process APQ-02357-17). LMM, JCS, AMV, NFA and JAF have a research fellowship from CNPq. NFA has grants from Fundect-MS (TO 141/2016 and TO 007/2015) and Fundect-MS (007/2015 SIAFEM 025139). LAGM and LMM have UFOP grants. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Made available in DSpace on 2021-06-25T11:01:46Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-07-01 Xanthomonas arboricola pv. juglandis (Xaj) is the most significant aboveground walnut bacterial pathogen. Disease management uses copper-based pesticides which induce pathogen resistance. We examined the genetic repertoire associated with adaptation and virulence evolution in Xaj. Comparative genomics of 32 Xaj strains reveal the possible acquisition and propagation of virulence factors via insertion sequences (IS). Fine-scale annotation revealed a Tn3 transposon (TnXaj417) encoding copper resistance genes acquired by horizontal gene transfer and associated with adaptation and tolerance to metal-based pesticides commonly used to manage pathogens in orchard ecosystems. Phylogenomic analysis reveals IS involvement in acquisition and diversification of type III effector proteins ranging from two to eight in non-pathogenic strains, 16 to 20 in pathogenic strains, besides six other putative effectors with a reduced identity degree found mostly among pathogenic strains. Yersiniabactin, xopK, xopAI, and antibiotic resistance genes are also located near ISs or inside genomic islands and structures resembling composite transposons. Center of Research in Biological Science Federal University of Ouro Preto Department of Plant Sciences University of California Faculty of Agricultural and Veterinary Sciences of Jaboticabal (FCAV) Universidade Estadual Paulista (UNESP) Department of Technology Cell Cycle Laboratory Butantan Institute Department of Biochemistry Chemistry Institute University of Sao Paulo School of Computing Federal University of Mato Grosso do Sul Department of Biological Science Institute of Exact and Biological Science Federal University of Ouro Preto Faculty of Agricultural and Veterinary Sciences of Jaboticabal (FCAV) Universidade Estadual Paulista (UNESP) Department of Technology

The interaction between the plant host, walnut (Juglans regia, Jr), and a deadly pathogen (Xanthomonas arboricola pv. juglandis 417, Xaj) can lead to walnut bacterial blight (WB), which depletes walnut productivity by degrading the nut quality. Here, we dissect this pathosystem using tandem mass tag quantitative proteomics. Walnut hull tissues inoculated with Xaj were compared to mock-inoculated tissues, and 3972 proteins were identified, of which 3296 are from Jr and 676 from Xaj. Proteins with differential abundance include oxidoreductases, proteases, and enzymes involved in energy metabolism and amino acid interconversion pathways. Defense responses and plant hormone biosynthesis were also increased. Xaj proteins detected in infected tissues demonstrate its ability to adapt to the host microenvironment, limiting iron availability, coping with copper toxicity, and maintaining energy and intermediary metabolism. Secreted proteases and extracellular secretion apparatus such as type IV pilus for twitching motility and type III secretion effectors indicate putative factors recognized by the host. Taken together, these results suggest intense degradation processes, oxidative stress, and general arrest of the biosynthetic metabolism in infected nuts. Our results provide insights into molecular mechanisms and highlight potential molecular tools for early detection and disease control strategies.

Following photosynthesis, sucrose is translocated to sink organs, where it provides the primary source of carbon and energy to sustain plant growth and development. Sugar transporters from the SWEET (sugar will eventually be exported transporter) family are rate-limiting factors that mediate sucrose transport across concentration gradients, sustain yields, and participate in reproductive development, plant senescence, stress responses, as well as support plant&ndash, pathogen interaction, the focus of this study. We identified 25 SWEET genes in the walnut genome and distinguished each by its individual gene structure and pattern of expression in different walnut tissues. Their chromosomal locations, cis-acting motifs within their 5&prime, regulatory elements, and phylogenetic relationship patterns provided the first comprehensive analysis of the SWEET gene family of sugar transporters in walnut. This family is divided into four clades, the analysis of which suggests duplication and expansion of the SWEET gene family in Juglans regia. In addition, tissue-specific gene expression signatures suggest diverse possible functions for JrSWEET genes. Although these are commonly used by pathogens to harness sugar products from their plant hosts, little was known about their role during Xanthomonas arboricola pv. juglandis (Xaj) infection. We monitored the expression profiles of the JrSWEET genes in different tissues of &ldquo, Chandler&rdquo, walnuts when challenged with pathogen Xaj417 and concluded that SWEET-mediated sugar translocation from the host is not a trigger for walnut blight disease development. This may be directly related to the absence of type III secretion system-dependent transcription activator-like effectors (TALEs) in Xaj417, which suggests different strategies are employed by this pathogen to promote susceptibility to this major aboveground disease of walnuts.

Made available in DSpace on 2021-06-25T10:44:05Z (GMT). No. of bitstreams: 0 Previous issue date: 2020-11-01 Here we present and analyze the complete genome of Alcaligenes faecalis strain Mc250 (Mc250), a bacterium isolated from the roots of Mimosa calodendron, an endemic plant growing in ferruginous rupestrian grasslands in Minas Gerais State, Brazil. The genome has 4,159,911 bp and 3,719 predicted protein-coding genes, in a single chromosome. Comparison of the Mc250 genome with 36 other Alcaligenes faecalis genomes revealed that there is considerable gene content variation among these strains, with the core genome representing only 39% of the protein-coding gene repertoire of Mc250. Mc250 encodes a complete denitrification pathway, a network of pathways associated with phenolic compounds degradation, and genes associated with HCN and siderophores synthesis; we also found a repertoire of genes associated with metal internalization and metabolism, sulfate/sulfonate and cysteine metabolism, oxidative stress and DNA repair. These findings reveal the genomic basis for the adaptation of this bacterium to the harsh environmental conditions from where it was isolated. Gene clusters associated with ectoine, terpene, resorcinol, and emulsan biosynthesis that can confer some competitive advantage were also found. Experimental results showed that Mc250 was able to reduce (∼60%) the virulence phenotype of the plant pathogen Xanthomonas citri subsp. citri when co-inoculated in Citrus sinensis, and was able to eradicate 98% of juveniles and stabilize the hatching rate of eggs to 4% in two species of agricultural nematodes. These results reveal biotechnological potential for the Mc250 strain and warrant its further investigation as a biocontrol and plant growth-promoting bacterium. Núcleo de Pesquisas em Ciências Biológicas (NUPEB) Universidade Federal de Ouro Preto Departamento de Ciências Biológicas (DECBI) Instituto de Ciências Exatas e Biológicas (ICEB) Universidade Federal de Ouro Preto (UFOP) Instituto Pristino Departamento de Bioquimica (DBQ) Instituto de Quimica (IQ) Universidade de São Paulo (USP) Instituto de Biotecnologia Aplicada a Agropecuária (BIOAGRO) Universidade Federal de Vicosa (UFV) Departamento de Tecnologia Faculdade de Ciências Agrárias e Veterinárias de Jaboticabal (FCAV) Universidade Estadual Paulista (UNESP) Faculdade de Computação (FACOM) Universidade Federal de Mato Grosso do Sul Departamento de Tecnologia Faculdade de Ciências Agrárias e Veterinárias de Jaboticabal (FCAV) Universidade Estadual Paulista (UNESP)

The ferruginous rupestrian grasslands (FRG) in the Iron Quadrangle (IQ) are ecosystems characterized by rocky soils with reduced availability of water and nutrients, but high levels of metals. In order to comprehend the interference of microorganisms on the adaptive process of endemic plant Mimosa calodendrum (Fabaceae), bacteria associated with its roots and rhizosphere were isolated. Fourteen isolates were obtained and subsequently grown in the presence of different concentrations of arsenic (As) species. The isolate Mc250, an Alcaligenes faecalis strain, resisted to 10 mM of As (III) and 800 mM of As (V). In the presence of this strain, atomic spectrometer detected a reduction of 55% for As (III) and 72% for As (V) respectively in 10 mM and 500 mM solution. Scanning electron microscopy of this isolate demonstrated morphological modification and EDX spectroscopy revealed the presence of both As species adsorbed on the membrane, justifying the removal observed in the in vitro assays. To validate this potential removal of As in vivo, tomato plants were used as grown model in the presence and absence of A. faecalis in soil previously contaminated with 5 mM of As (III). After 14 days, plants from contaminated soil had their growth improved when compared to untreated control plants. All these results suggest for the first time that plant-associated bacteria from FRG-IQ present potential for soil rhizoremediation and may benefit the adaptive processes of plants in extreme environments including application in recovering degraded areas.

A educação ambiental (EA) surgiu num contexto de mudanças sociais evidenciadas por volta de 1960. A princípio, a EA favorecia apenas o aprendizado de conceitos ecológicos. Mais tarde, em decorrência de uma redução da disponibilidade de recursos naturais, o foco da EA passou a se estabelecer no contexto cidadão e democrático. O objetivo da EA passou a ser o de transformar o indivíduo, estabelecendo um criticismo junto aos problemas socioambientais permitindo assim a tomada de decisões na promoção do desenvolvimento sustentável. Desde então, a EA tem sido vista como um caminho para mudanças atitudinais. Neste contexto, a interdisciplinaridade como uma estratégia de diálogo entre saberes passou a ser essencial nos trabalhos visando a EA. O propósito desse trabalho é apresentar contextualizações e reflexões sobre o surgimento e a importância da EA, atualmente caracterizada como um importante fator de transformações sociais, exigindo que profissionais da educação trabalhem esta temática numa perspectiva interdisciplinar. Environmental education (EE) emerged in a context of social changes evidenced around 1960. At first, EE favored only the learning of ecological concepts. Later, as a result of a reduction in the availability of natural resources, the focus of EE started to be established in the citizen and democratic context. EE's goal has been to transform the individual, establishing a criticism with the socio-environmental problems, thus allowing decision-making in the promotion of sustainable development. Since then, EE has been seen as a path to attitudinal change. In this context, interdisciplinarity as a strategy of dialogue between knowledge has become essential in the works aimed at EE. The purpose of this work is to present contextualization and reflections about the emergence and importance of EE, currently characterized as an important factor of social transformations, requiring education professionals to work this subject in an interdisciplinary perspective. La educación ambiental (EA) surgió en un contexto de cambios sociales evidenciados alrededor de 1960. Al principio, la EA sólo favoreció el aprendizaje de conceptos ecológicos. Más tarde, como consecuencia de una reducción de la disponibilidad de recursos naturales, el foco de EA pasó a establecerse en el contexto ciudadano y democrático. El objetivo de EA pasó a ser el de transformar al individuo, estableciendo un criticismo junto a los problemas socioambientales permitiendo así la toma de decisiones en la promoción del desarrollo sostenible. Desde entonces, EA ha sido vista como un camino para cambios actitudes. En este contexto, la interdisciplinariedad como una estrategia de diálogo entre saberes pasó a ser esencial en los trabajos hacia la EA. El propósito de este trabajo es presentar contextualizaciones y reflexiones sobre el surgimiento y la importancia de la EA, actualmente caracterizada como un importante factor de transformaciones sociales, exigiendo que profesionales de la educación trabajen esta temática desde una perspectiva interdisciplinaria.

A mutant library of the bacterium Xanthomonas citri subsp. citri strain 306 pathotype A (Xac), the causative agent of most aggressive Asiatic type A citrus canker, was screened regarding altered canker symptoms after inoculations into Citrus sinensis and Citrus limonia host leaves. Twenty-six mutants have shown phenotypic virulence changes and have respectively knocked out gene identified by sequencing. In vivo growth curves were obtained for nine mutants to quantify how the mutations could affect pathogen’s adaptability to growth inside and attack host plant infected tissue. Among identified genes in mutated strains, we could find those that until now had not been reported as being involved in Xac adaptation and/or virulence, such as predicted to encode for xylose repressor-like protein (XACΔxylR), Fe-S oxidoredutase (XACΔaslB), helicase IV (XACΔhelD), ubiquinol cytochrome c oxidoreductase iron-sulfur subunit (XACΔpetA), chromosome partitioning protein (XACΔparB) and cell division protein FtsB (XACΔftsB), in addition to genes predicted to encode for hypothetical proteins. The new genes found in this study as being relevant to adaptation and virulence, improve the understanding of Xac fitness during citrus plant attack and canker symptoms development.

Made available in DSpace on 2020-12-12T01:05:16Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-12-01 Fundação Grupo Boticário de Proteção à Natureza Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) The taxonomy of muriquis, the largest extant primates in the New World, is controversial. While some specialists argue for a monotypic genus (Brachyteles arachnoides), others favor a two-species classification, splitting northern muriquis (Brachyteles hypoxanthus) from southern muriquis (B. arachnoides). This uncertainty affects how we study the differences between these highly endangered and charismatic primates, as well as the design of more effective conservation programs. To address this issue, between 2003 and 2017 we collected over 230 muriqui fecal samples across the genus’ distribution in the Brazilian Atlantic Forest, extracted DNA from these samples, and sequenced 423 base pairs of the mitochondrial DNA (mtDNA) control region. Phylogenetic and species delimitation analyses of our sequence dataset robustly support two reciprocally monophyletic groups corresponding to northern and southern muriquis separated by an average 12.7% genetic distance. The phylogeographic break between these lineages seems to be associated with the Paraíba do Sul River and coincides with the transition between the north and south Atlantic Forest biogeographic zones. Published divergence estimates from whole mitochondrial genomes and nuclear loci date the split between northern and southern muriquis to the Early Pleistocene (ca. 2.0 mya), and our new mtDNA dataset places the coalescence time for each of these two clades near the last interglacial (ca. 120–80 kya). Our results, together with both phenotypic and ecological differences, support recognizing northern and southern muriquis as sister species that should be managed as distinct evolutionarily significant units. Given that only a few thousand muriquis remain in nature, it is imperative that conservation strategies are tailored to protect both species from extinction. Department of Anthropology New York University New York Consortium in Evolutionary Primatology (NYCEP) Escola de Ciências Pontifícia Universidade Católica do Rio Grande do Sul Centro Nacional de Pesquisa e Conservação de Primatas Brasileiros Instituto Chico Mendes de Conservação da Biodiversidade Laboratório de Ecologia Aplicada e Conservação (LECON) Departamento de Ciências Ambientais Universidade Federal de São Paulo Instituto Pró-Muriqui Department of Anthropology University of Wisconsin-Madison Diretoria de Biodiversidade Sociedade Ecoatlântica Muriqui Instituto de Biodiversidade Programa de Pós-graduação em Animais Selvagens Universidade Estadual Paulista (UNESP) Divisão de Meio Ambiente Departamento de Recursos Ambientais Instituto de Tecnologia para o Desenvolvimento (Lactec) Programa de Pós-Graduação em Desenvolvimento de Tecnologia Instituto de Tecnologia para o Desenvolvimento e Instituto de Engenharia do Paraná Programa de Pós-graduação em Ecologia Aplicada Universidade Federal de Lavras Centro de Primatologia do Rio de Janeiro Instituto Estadual do Ambiente Unidade Acadêmica Especial de Ciências Biológicas Universidade Federal de Goiás Departamento de Engenharia Florestal Universidade Federal de Viçosa Parque Zoológico Municipal Quinzinho de Barros Departamento de Ciências Biológicas Universidade Federal do Espírito Santo Departamento de Pesquisa e Conservação da Fauna Prefeitura Municipal de Curitiba Department of Anthropology Primate Molecular Ecology and Evolution Laboratory The University of Texas at Austin Programa de Pós-graduação em Animais Selvagens Universidade Estadual Paulista (UNESP) Fundação Grupo Boticário de Proteção à Natureza: 0017_2016 Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul: 16/0491 CNPq: 310472/2017-2 CNPq: 402975/2012-0 CAPES: BEX 1872-07-9

Made available in DSpace on 2019-10-06T16:50:25Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-09-09 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Background: Xanthomonas citri subsp. citri pathotypes cause bacterial citrus canker, being responsible for severe agricultural losses worldwide. The A pathotype has a broad host spectrum, while A∗and Aw are more restricted both in hosts and in geography. Two previous phylogenomic studies led to contrasting well-supported clades for sequenced genomes of these pathotypes. No extensive biogeographical or divergence dating analytic approaches have been so far applied to available genomes. Results: Based on a larger sampling of genomes than in previous studies (including six new genomes sequenced by our group, adding to a total of 95 genomes), phylogenomic analyses resulted in different resolutions, though overall indicating that A + AW is the most likely true clade. Our results suggest the high degree of recombination at some branches and the fast diversification of lineages are probable causes for this phylogenetic blurring effect. One of the genomes analyzed, X. campestris pv. durantae, was shown to be an A∗strain; this strain has been reported to infect a plant of the family Verbenaceae, though there are no reports of any X. citri subsp. citri pathotypes infecting any plant outside the Citrus genus. Host reconstruction indicated the pathotype ancestor likely had plant hosts in the family Fabaceae, implying an ancient jump to the current Rutaceae hosts. Extensive dating analyses indicated that the origin of X. citri subsp. citri occurred more recently than the main phylogenetic splits of Citrus plants, suggesting dispersion rather than host-directed vicariance as the main driver of geographic expansion. An analysis of 120 pathogenic-related genes revealed pathotype-associated patterns of presence/absence. Conclusions: Our results provide novel insights into the evolutionary history of X. citri subsp. citri as well as a sound phylogenetic foundation for future evolutionary and genomic studies of its pathotypes. Departamento de Bioquímica Instituto de Química Universidade de Saõ Paulo Laboratório Especial de Ciclo Celular Instituto Butantan Departamento de Fitopatologia e Nematologia Escola Superior de Agricultura luiz de Queiroz Universidade de Saõ Paulo Escola de Artes Ciências e Humanidades Universidade de Saõ Paulo Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista (UNESP) Departamento de Pesquisa e Desenvolvimento Fundo de Defesa da Citricultura (Fundecitrus) Citrus Research and Education Center Department of Microbiology and Cell Science University of Florida Faculdade de Computacaõ Universidade Federal de Mato Grosso Do sul Núcleo de Pesquisas em Ciências Biológicas Universidade Federal de Ouro Preto Biocomplexity Institute of Virginia Tech Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista (UNESP)

Made available in DSpace on 2020-12-12T01:43:17Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-01-01 Fundo de Defesa da Citricultura Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG) Background. In Citrus cultures, three species of Xanthomonas are known to cause distinct diseases. X. citri subsp. citri patothype A, X. fuscans subsp. aurantifolii pathotypes B and C, and X. alfalfae subsp. citrumelonis, are the causative agents of cancrosis A, B, C, and citrus bacterial spots, respectively. Although these species exhibit different levels of virulence and aggressiveness, only limited alternatives are currently available for proper and early detection of these diseases in the fields. The present study aimed to develop a new molecular diagnostic method based on genomic sequences derived from the four species of Xanthomonas. Results. Using comparative genomics approaches, primers were synthesized for the identification of the four causative agents of citrus diseases. These primers were validated for their specificity to their target DNA by both conventional and multiplex PCR. Upon evaluation, their sensitivity was found to be 0.02 ng/µl in vitro and 1.5 × 104 CFU ml−1 in infected leaves. Additionally, none of the primers were able to generate amplicons in 19 other genomes of Xanthomonas not associated with Citrus and one species of Xylella, the causal agent of citrus variegated chlorosis (CVC). This denotes strong specificity of the primers for the different species of Xanthomonas investigated in this study. Conclusions. We demonstrated that these markers can be used as potential candidates for performing in vivo molecular diagnosis exclusively for citrus-associated Xanthomonas. The bioinformatics pipeline developed in this study to design specific genomic regions is capable of generating specific primers. It is freely available and can be utilized for any other model organism. Núcleo de Pesquisas em Ciências Biológicas Universidade Federal de Ouro Preto Departamento de Tecnologia Faculdade de Ciências Agrárias e Veterinárias de Jaboticabal Universidade Estadual Paulista-Unesp Departamento de Fitopatologia e Nematologia Escola Superior de Agricultura ‘‘Luiz de Queiroz’’ Departamento de Bioquímica Instituto de Química Universidade de São Paulo Instituto de Computação Universidade Estadual de Campinas Faculdade de Computação Universidade Federal de Mato Grosso do Sul Departamento de Ciências Biológicas Universidade Federal de Ouro Preto Departamento de Tecnologia Faculdade de Ciências Agrárias e Veterinárias de Jaboticabal Universidade Estadual Paulista-Unesp Fundo de Defesa da Citricultura: 007/2015 SIAFEM 025139 CAPES: 3385/2013 CNPq: 481226/2013-3 FAPEMIG: APQ-02387-14 CAPES: CFP 51/2013

Type IV pili (Tfp) are widely distributed adhesins of bacterial surfaces. In plant pathogenic bacteria, Tfp are involved in host colonization and pathogenesis. Xanthomonas citri subsp. citri (Xcc) is the phytopathogen responsible for citrus canker disease. In this work, three Tfp structural genes, fimA, fimA1, and pilA from Xcc were studied. A pilA mutant strain from Xcc (XccΔpilA) was constructed and differences in physiological features, such as motilities, adhesion, and biofilm formation, were observed. A structural study of the purified Tfp fractions from Xcc wild-type and Xcc∆pilA showed that pilins are glycosylated in both strains and that FimA and FimA1 are the main structural components of the pili. Furthermore, smaller lesion symptoms and reduced bacterial growth were produced by Xcc∆pilA in orange plants compared to the wild-type strain. These results indicate that the minor pilin-like gene, pilA, is involved in Tfp performance during the infection process. Fil: Petrocelli, Silvana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina Fil: Arana, Maite Rocío. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina Fil: Cabrini, Marcela N.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Centro de Investigaciones en Hidratos de Carbono. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Centro de Investigaciones en Hidratos de Carbono; Argentina Fil: Casabuono, Adriana Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Centro de Investigaciones en Hidratos de Carbono. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Centro de Investigaciones en Hidratos de Carbono; Argentina Fil: Moyano, Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina Fil: Beltramino, Matias. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina Fil: Moreira, Leandro M.. Universidade Federal de Ouro Preto; Brasil Fil: Couto, Alicia Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Centro de Investigaciones en Hidratos de Carbono. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Centro de Investigaciones en Hidratos de Carbono; Argentina Fil: Orellano, Elena Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina

Aedes aegypti females ingest sugar or blood to obtain the nutrients needed to maintain cellular homeostasis. During human blood ingestion, female mosquitoes may transmit different viruses such as dengue, yellow fever and, more recently, zika and chikungunya. Here, we report changes in protein expression in the heads of adult female Ae. aegypti mosquitoes in response to the ingestion of blood or sugar. Proteins extracted from the heads of Ae. aegypti fed exclusively on blood (BF) or sugar (SF) were trypsin hydrolyzed (off-gel) and analyzed by the reverse-phase nano-liquid chromatography coupled with hybrid mass spectrometry. A total of 1139 proteins were identified in female heads, representing 7.4% of the predicted proteins in Ae. aegypti genome (total = 15 419 active genes). Gene ontology annotation and categories showed that, in this insect, the head was rich in proteins involved in the metabolic process, proton transport, organelle, macromolecular complex, structural molecule activity, antioxidant activity, and catalytic activity. Our report is the first indicating that many of the annotated genes are translated into functional proteins in heads of adult female Ae. aegypti. Interestingly, we identified 8.7 times more exclusively expressed proteins involved in signal transduction, replication-transcription-translation (5.5 x), and transport (2.9 x) activity in BF than in SF groups. This paper discusses the protein profile of Ae. aegypti female heads and its implications for blood ingestion and carbohydrate intake.

Made available in DSpace on 2019-10-06T15:59:04Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-10-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG) Microorganisms associated with plants have a great biotechnological potential, but investigations of these microorganisms associated with native plants in peculiar environments has been incipient. The objective of this study was to analyze the plant growth-promoting bacteria potential of cultivable bacteria associated with rare plants from the ferruginous rocky fields of the Brazilian Iron Quadrangle. The roots and rhizospheres of nine endemic plants species and samples of a root found in a lateritiric duricrust (canga) cave were collected, the culturable bacteria isolated and prospected for distinct biotechnological and ecological potentials. Out of the 148 isolates obtained, 8 (5.4%) showed potential to promote plant growth, whereas 4 (2.7%) isolates acted as biocontrol agents against Xanthomonas citri pathotype A (Xac306), reducing the cancrotic lesions by more than 60% when co-inoculated with this phytopathogen in Citrus sinensis plants. Moreover, other 4 (2.7%) isolates were classified as potential bioremediation agents, being able to withstand high concentrations of arsenite (5 mM As3+) and arsenate (800 mM As5+), by removing up to 35% and 15% of this metalloid in solution, respectively. These same four isolates had a positive influence on the growth of both the roots and the aerial parts when inoculated with tomato seeds in the soil contaminated with arsenic. This is the first time that an investigation highlights the potentialities of bacteria associated with rare plants of ferruginous rocky fields as a reservoir of microbiota of biotechnological and ecological interest, highlighting the importance of conservation of this area that is undergoing intense anthropic activity. Graphical abstract: [Figure not available: see fulltext.]. Programa de Pós-graduação em Biotecnologia Núcleo de Pesquisas em Ciências Biológicas Universidade Federal de Ouro Preto Departamento de Ciências Biológicas (DECBI) Instituto de Ciências Exatas e Biológicas (ICEB) Universidade Federal de Ouro Preto (UFOP) Departamento de Tecnologia Universidade Estadual Paulista UNESP, Campus de Jaboticabal Instituto Prístino Departamento de Tecnologia Universidade Estadual Paulista UNESP, Campus de Jaboticabal CAPES: 3385/2013 CNPq: 481226/2013-3 FAPEMIG: APQ-02387-14 CAPES: CFP 51/2013

Mobile genetic elements (MGEs) are an important feature of prokaryote genomes but are seldom well annotated and, consequently, are often underestimated. MGEs include transposons (Tn), insertion sequences (ISs), prophages, genomic islands (GEIs), integrons, and integrative and conjugative elements (ICEs). They are intimately involved in genome evolution and promote phenomena such as genomic expansion and rearrangement, emergence of virulence and pathogenicity, and symbiosis. In spite of the annotation bottleneck, there are so far at least 75 different programs and databases dedicated to prokaryotic MGE analysis and annotation, and this number is rapidly growing. Here, we present a practical guide to explore, compare, and visualize prokaryote MGEs using a combination of available software and databases tailored to small scale genome analyses. This protocol can be coupled with expert MGE annotation and exploited for evolutionary and comparative genomic analyses.

A educação ambiental (EA), enquanto campo epistemológico, pode auxiliar no despertar da compreensão do papel do indivíduo no ambiente. Neste trabalho, apresenta-se o resultado da parceria entre o Mestrado Profissional em Ensino de Ciências da Universidade Federal de Ouro Preto e o Instituto Prístino, com o objetivo de desenvolver uma cartilha de EA, envolvendo o ecossistema de canga, para professores e alunos do ensino fundamental II. Em Minas Gerais, os ecossistemas ferruginosos – conhecidos como “cangas” – apresentam características abióticas que resultaram em um dos ambientes com maior endemismo vegetal do Brasil. Atuando como reserva hídrica que abastece mananciais da região metropolitana de Belo Horizonte, as cangas estão na lista dos ecossistemas mais ameaçados, em especial pela mineração. Este artigo fornece conceitos, peculiaridades, importâncias, ameaças e desafios, além de exercícios de fixação e discussão contextualizada, sendo, portanto, uma ferramenta diferenciada para o ensino de Ciências. Environmental Education (EE), while an epistemological field, may assist to awake the comprehension of individuals’ role within the environment. In this work, we present the result of a partnership between the Professional Masters Studies in Science Teaching from the Federal University of Ouro Preto and the Prístino Institute, aiming to develop an EE textbook involving the canga ecosystem for teachers and students of the basic education. In Minas Gerais, ferruginous ecosystems – known as “Cangas” – present abiotic characteristics that resulted in one of the environments with higher plant endemism in Brazil. Acting as a water reservoir that supplies water sources of the metropolitan region of Belo Horizonte, the Cangas are on the list of the most endangered ecosystems, especially by mining. This article provides concepts, peculiarities, importance, threats and challenges, in addition to fixation exercises and contextualized discussion being, therefore, a differentiated tool for Science teaching.

Soil is a habitat where plant roots and microorganisms interact. In the region of the Brazilian Iron Quadrangle (IQ), studies involving the interaction between microbiota and plants have been neglected. Even more neglected are the studies involving the holoparasite plant Langsdorffia hypogaea Mart. (Balanophoraceae). The geomorphological peculiarities of IQ soil, rich in iron ore, as well as the model of interaction between L. hypogaea, its hosts and the soil provide a unique niche that acts as selective pressure to the evolution of plant growth-promoting bacteria (PGPB). The aim of this study was to prospect the bacterial microbiota of holoparasitic plant L. hypogaea, its plant host and corresponding rhizosphere of IQ soil, and to analyze the potential of these isolates as PGPB. We obtained samples of 11 individuals of L. hypogaea containing fragments of host and rhizosphere remnants, resulting in 81 isolates associated with Firmicutes and Proteobacteria phyla. The ability to produce siderophores, hydrocyanic acid (HCN), indole-3-acetic acid (IAA), nitrogen (N2) fixation, hydrolytic enzymes secretion and inhibition of enteropathogens, and phytopathogens were evaluated. Of the total isolates, 62, 86, and 93% produced respectively siderophores, IAA, and were able to fix N2. In addition, 27 and 20% of isolates inhibited the growth of enteropathogens and phytopathogens respectively, and 58% were able to produce at least one hydrolytic activity investigated. The high number of isolates that produce siderophores and indole-3-acetic acid suggests that this microbiota may be important for adaptation of plants to IQ. The results demonstrate for the first time the biological importance of Brazilian IQ species as reservoirs of specific microbiotas that might be used as plant growth-promoting bacteria on agricultural land or antropized soils that needs to be reforested.

The floral biology and reproductive phenology of Langsdorffia hypogaea were investigated. Flowering is annual and high synchronicity was observed along two years, with significant correlation with rainfall. The post-reproductive death of rhizomes suggests monocarpy. Flowers have anthers with extrorse dehiscence and a stigma exposed above the perigonium, and are arranged in dense inflorescences which produce extrafloral nectar. A total of 259 floral visitors were observed, mostly Hymenoptera/Formicidae (149 individuals, 17 ant species), and eight species of Araneae. Although ants were frequent floral visitors, a Coleoptera (Nitidulidae, Stelidota; 28% of total visits) species was the most abundant pollinator. In consequence of herbivory only 12.5% of all inflorescence clusters fruited, and we could not evaluate the percentage of pollinated plants due to insect damage. By being a rich resource in a season of scarcity, activity on inflorescences of L. hypogaea of both pollinators and herbivores was high.

Rede de Ensino Doctum, Caratinga, Minas Gerais, Brasil Conservation International. Demographic data are essential to assessments of the status of endangered species. However, establishing an integrated monitoring program to obtain useful data on contemporary and future population trends requires both the identification of priority areas and populations and realistic evaluations of the kinds of data that can be obtained under different monitoring regimes. We analyzed all known populations of a critically endangered primate, the muriqui (genus: Brachyteles) using population size, genetic uniqueness, geographic importance (including potential importance in corridor programs) and implementability scores to define monitoring priorities. Our analyses revealed nine priority populations for the northern muriqui (B. hypoxanthus) and nine for the southern muriqui (B. arachnoides). In addition, we employed knowledge of muriqui developmental and life history characteristics to define the minimum monitoring intensity needed to evaluate demographic trends along a continuum ranging from simple descriptive changes in population size to predictions of population changes derived from individual based life histories. Our study, stimulated by the Brazilian government's National Action Plan for the Conservation of Muriquis, is fundamental to meeting the conservation goals for this genus, and also provides a model for defining priorities and methods for the implementation of integrated demographic monitoring programs for other endangered and critically endangered species of primates. Univ Fed Espirito Santo, Dept Ciencias Biol, Vitoria, ES, Brazil Preserve Muriqui, Caratinga, MG, Brazil Muriqui Inst Biodiversidade, Caratinga, MG, Brazil Ctr Primatol Rio De Janeiro CPRJ INEA, Guapimirim, RJ, Brazil Univ Fed Rio de Janeiro, Programa Posgrad Ecol, Rio De Janeiro, Brazil Univ Fed Goias, Inst BioCiencias, Jatai, Go, Brazil Univ Fed Sao Paulo, Lab Primatol & Conservacao Especies, Dept Ciencias Ambientais, Campus Diadema, Diadema Sao Paulo, Brazil Programa Posgrad Anal Ambiental Integrada, Diadema Sao Paulo, Brazil Inst Promuriqui, Sao Miguel Arcanjo, SP, Brazil Ecoatlantica, Rio De Janeiro, RJ, Brazil Inst Pesquisas Mate Atlantica IPEMA, Vitoria, ES, Brazil Inst Chico Mendes Conservacao Biodiversidade ICMB, Ctr Nacl Pesquisa & Conservacao Primatas Brasilei, Joao Pessoa, Paraiba, Brazil Univ Fed Sao Paulo, Lab Primatol & Conservacao Especies, Dept Ciencias Ambientais, Campus Diadema, Diadema Sao Paulo, Brazil Web of Science

Motivation Information about metabolic pathways in a comparative context is one of the most powerful tool to help the understanding of genome-based differences in phenotypes among organisms. Although several platforms exist that provide a wealth of information on metabolic pathways of diverse organisms, the comparison among organisms using metabolic pathways is still a difficult task. Results We present TabPath (Tables for Metabolic Pathway), a web-based tool to facilitate comparison of metabolic pathways in genomes based on KEGG. From a selection of pathways and genomes of interest on the menu, TabPath generates user-friendly tables that facilitate analysis of variations in metabolism among the selected organisms. Availability and implementation TabPath is available at http://200.239.132.160:8686.

Fundacao de Amparo a Pesquisa do Estado de Sao Paulo Fundacao de Amparo a Pesquisa do Estado de Minas Gerais BIGA grant-CAPES Background: Xanthomonas citri subsp. citri (Xac) is the causal agent of citrus canker. A proteomic analysis under in planta infectious and non-infectious conditions was conducted in order to increase our knowledge about the adaptive process of Xac during infection. Results: For that, a 2D-based proteomic analysis of Xac at 1, 3 and 5 days after inoculation, in comparison to Xac growth in NB media was carried out and followed by MALDI-TOF-TOF identification of 124 unique differentially abundant proteins. Among them, 79 correspond to up-regulated proteins in at least one of the three stages of infection. Our results indicate an important role of proteins related to biofilm synthesis, lipopolysaccharides biosynthesis, and iron uptake and metabolism as possible modulators of plant innate immunity, and revealed an intricate network of proteins involved in reactive oxygen species adaptation during Plants'Oxidative Burst response. We also identified proteins previously unknown to be involved in Xac-Citrus interaction, including the hypothetical protein XAC3981. A mutant strain for this gene has proved to be non-pathogenic in respect to classical symptoms of citrus canker induced in compatible plants. Conclusions: This is the first time that a protein repertoire is shown to be active and working in an integrated manner during the infection process in a compatible host, pointing to an elaborate mechanism for adaptation of Xac once inside the plant. Univ Fed Ouro Preto, Inst Ciencias Exatas & Biol, Dept Ciencias Biol DECBI, Ouro Preto, MG, Brazil Univ Fed Ouro Preto, Nucleo Pesquisas Ciencias Biol NUPEB, Ouro Preto, MG, Brazil Univ Fed Rio de Janeiro, Inst Quim, Dept Bioquim DBq, Rio De Janeiro, RJ, Brazil Univ Estadual Paulista, UNESP, Dept Tecnol, Fac Ciencias Agr & Vet Jaboticabal, Jaboticabal, SP, Brazil Univ Estadual Campinas, UNICAMP, Inst Quim, Campinas, SP, Brazil Univ Sao Paulo, Inst Quim, Dept Bioquim, Sao Paulo, SP, Brazil Univ Fed Sao Paulo UNIFESP, Dept Ciencias Biol, Diadema, SP, Brazil Virginia Tech, Biocomplex Inst, Blacksburg, VA USA Univ Sao Paulo, Inst Quim, Dept Bioquim, Sao Paulo, SP, Brazil Univ Fed Sao Paulo UNIFESP, Dept Ciencias Biol, Diadema, SP, Brazil CAPES FAPESP: 04/02006-7 Fundacao de Amparo a Pesquisa do Estado de Minas Gerais: CBB-APQ-04425-10 BIGA grant-CAPES: 3385/2013 Web of Science

The Xanthomonadaceae family consists of species of non-pathogenic and pathogenic γ-proteobacteria that infect different hosts, including humans and plants. In this study, we performed a comparative analysis using 69 fully sequenced genomes belonging to this family, with a focus on identifying proteins enriched in phytopathogens that could explain the lifestyle and the ability to infect plants. Using a computational approach, we identified seven phytopathogen-enriched protein families putatively secreted by type II secretory system: PheA (CM-sec), LipA/LesA, VirK, and four families involved in N-glycan degradation, NixE, NixF, NixL, and FucA1. In silico and phylogenetic analyses of these protein families revealed they all have orthologs in other phytopathogenic or symbiotic bacteria, and are involved in the modulation and evasion of the immune system. As a proof of concept, we performed a biochemical characterization of LipA from Xac306 and verified that the mutant strain lost most of its lipase and esterase activities and displayed reduced virulence in citrus. Since this study includes closely related organisms with distinct lifestyles and highlights proteins directly related to adaptation inside plant tissues, novel approaches might use these proteins as biotechnological targets for disease control, and contribute to our understanding of the coevolution of plant-associated bacteria.

Most of the computational biology analysis is made comparing genomic features. The nucleotide and amino acid sequence alignments are frequently used in gene function identification and genome comparison. Despite its widespread use, there are limitations in their analysis capabilities that need to be considered but are often overlooked or unknown by many researchers. This paper presents a gene based whole genome comparison toolkit which can be used not only as an alternative and more robust way to compare a set of whole genomes, but, also, to understand the tradeoff of the use of sequence local alignment in this kind of comparison. A study case was performed considering fifteen whole genomes of the Xanthomonas genus. The results were compared with the 16S rRNA-processing protein RimM phylogeny and some thresholds for the use of sequence alignments in this kind of analysis were discussed. Grande parte das analises realizadas na biologia computacional é feita comparando características genômicas. Os alinhamentos de nucleotídeos e de aminoácidos são frequentemente usados na identificação de funções gênicas e na comparação de genomas. Apesar de seu uso generalizado, há limitações em suas capacidades de analise que precisam ser consideradas, mas são frequentemente negligenciadas ou desconhecidas por muitos pesquisadores. Este artigo apresenta um conjunto de ferramentas de comparação de genomas completos baseado em genes que pode ser usado não somente como uma maneira alternativa e mais robusta de comparar um conjunto de genomas completos, mas também para entender as vantagens e desvantagens do uso do alinhamento local de sequencias neste tipo de comparação. Um estudo de caso foi realizado considerando quinze genomas completos do gênero Xanthomonas. Os resultados foram comparados com a filogenia produzida utilizando a proteína 16S rRNA-processing protein RimM e alguns limiares para o uso de alinhamentos de sequencias neste tipo de analise foram discutidos.

Made available in DSpace on 2018-11-26T16:27:54Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-02-23 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Citrus canker is a major disease affecting citrus production in Brazil. It's mainly caused by Xanthomonas citri subsp. citri strain 306 pathotype A (Xac). We analysed the differential expression of proteins secreted by wild type Xac and an asymptomatic mutant for hrpB4 (Delta hrpB4) grown in Nutrient Broth (NB) and a medium mimicking growth conditions in the plant (XAM1). This allowed the identification of 55 secreted proteins, of which 37 were secreted by both strains when cultured in XAMI. In this secreted protein repertoire, the following stand out: Virk, Polyphosphate-selective porin, Cellulase, Endoglucanase, Histone-like protein, Ribosomal proteins, five hypothetical proteins expressed only in the wild type strain, Lytic murein transglycosylase, Lipoprotein, Leucyl-tRNA synthetase, Co-chaperonin, Toluene tolerance, C-type cytochrome biogenesis membrane protein, Aminopeptidase and two hypothetical proteins expressed only in the Delta hrpB4 mutant. Furthermore, Peptidoglycan-associated outer membrane protein, Regulator of pathogenicity factor, Outer membrane proteins, Endopolygalacturonase, Chorismate mutase, Peptidyl-prolyl cis-trans isornerase land seven hypothetical proteins were detected in both strains, suggesting that there was no relationship with the secretion mediated by the type III secretory system, which is not functional in the mutant strain. Also worth mentioning is the Elongation factor Tu (EF-Tu), expressed only the wild type strain, and Type IV pilus assembly protein, Flagellin (FliC) and Flagellar hook-associated protein, identified in the wild-type strain secretome when grown only in NB. Noteworthy, that FliC, EF-Tu are classically characterized as PAMPs (Pathogen-associated molecular patterns), responsible for a PAMP-triggered immunity response. Therefore, our results highlight proteins potentially involved with the virulence. Overall, we conclude that the use of secretome data is a valuable approach that may bring more knowledge of the biology of this important plant pathogen, which ultimately can lead to the establishment of new strategies to combat citrus canker. Univ Estadual Paulista, Dept Tecnol, Jaboticabal, SP, Brazil Univ Fed Ouro Preto, Dept Ciencias Biol, Nucleo Pesquisas Ciencias Biol NUPEB, Ouro Preto, MG, Brazil Univ Fed Rio de Janeiro, Dept Bioquim, Rio De Janeiro, Rio De Janeiro, Brazil Univ Fed Vales Jequitinhonha & Mucuri, Dept Engn Florestal, Diamantina, MG, Brazil Univ Fed Sao Paulo, Dept Ciencias Biol, Diadema, SP, Brazil Univ Estadual Paulista, Dept Tecnol, Jaboticabal, SP, Brazil

Members of the genus Xanthomonas are among the most important phytopathogens. A key feature of Xanthomonas pathogenesis is the translocation of type III secretion system (T3SS) effector proteins (T3SEs) into the plant target cells via a T3SS. Several T3SEs and a murein lytic transglycosylase gene (mlt, required for citrus canker symptoms) are found associated with three transposition-related genes in Xanthomonas citri plasmid pXAC64. These are flanked by short inverted repeats (IRs). The region was identified as a transposon, TnXax1, with typical Tn3 family features, including a transposase and two recombination genes. Two 14-bp palindromic sequences within a 193-bp potential resolution site occur between the recombination genes. Additional derivatives carrying different T3SEs and other passenger genes occur in different Xanthomonas species. The T3SEs include transcription activator-like effectors (TALEs). Certain TALEs are flanked by the same IRs as found in TnXax1 to form mobile insertion cassettes (MICs), suggesting that they may be transmitted horizontally. A significant number of MICs carrying other passenger genes (including a number of TALE genes) were also identified, flanked by the same TnXax1 IRs and delimited by 5-bp target site duplications. We conclude that a large fraction of T3SEs, including individual TALEs and potential pathogenicity determinants, have spread by transposition and that TnXax1, which exhibits all of the essential characteristics of a functional transposon, may be involved in driving MIC transposition. We also propose that TALE genes may diversify by fork slippage during the replicative Tn3 family transposition. These mechanisms may play a crucial role in the emergence of Xanthomonas pathogenicity., IMPORTANCE Xanthomonas genomes carry many insertion sequences (IS) and transposons, which play an important role in their evolution and architecture. This study reveals a key relationship between transposons and pathogenicity determinants in Xanthomonas. We propose that several transposition events mediated by a Tn3-like element carrying different sets of passenger genes, such as different type III secretion system effectors (including transcription activation-like effectors [TALEs]), were determinant in the evolution and emergence of Xanthomonas pathogenicity. TALE genes are DNA-binding effectors that modulate plant transcription. We also present a model for generating TALE gene diversity based on fork slippage associated with the replicative transposition mechanism of Tn3-like transposons. This may provide a mechanism for niche adaptation, specialization, host-switching, and other lifestyle changes. These results will also certainly lead to novel insights into the evolution and emergence of the various diseases caused by different Xanthomonas species and pathovars.

As pesquisas no campo da genômica produzem uma grande quantidade de dados. Entretanto, o conhecimento genético acerca de certos fenótipos é limitado. Além disso, parte considerável dos genomas estudados possuem sequências codificantes (CDSs) com funções desconhecidas, representando um desafio adicional para a compreensão dos pesquisadores. Organismos provenientes de um mesmo ramo evolutivo compartilham muitas de suas CDSs, e certos fenótipos únicos a um grupo desses indivíduos podem ser resultado de um conjunto único de genes exclusivos. Neste trabalho é apresentado o arcabouço computacional GTACG, uma ferramenta com foco em uma usabilidade facilitada e destinada a pesquisas para identificação de características genéticas únicas em subgrupos de genomas de bactérias que possuem um determinado fenótipo em comum, encontrando dados que diferenciam eles dos outros organismos de forma simples. A análise do GTACG é baseada na formação de grupos de CDSs homólogas com base em alinhamentos locais. O front-end é simples de usar e a instalação de pacotes foi projetada para que usuários com pouco conhecimento em computação possam fazer análises complexas usando esta ferramenta. A validação dos resultados do GTACG se baseou em dois estudos de caso envolvendo um conjunto com 161 genomas da família Xanthomonadaceae e 45 genomas de Streptococcus pyogenes. No primeiro estudo de caso, buscava-se descobrir porque algumas Xanthomonadaceae se associam a plantas e outras não, e de fato foram encontradas 19 famílias de proteínas ortólogas exclusivas aos genomas associados a plantas (representando mais de 90% desses genomas), permitindo a identificação de proteínas potencialmente associadas com a adaptação e a virulência dessas bactérias nos tecidos das plantas. No segundo estudo, buscou-se encontrar marcadores filogenéticos para a proteína emm dos Streptococcus pyogenes, e foram encontrados 15 famílias de proteínas ortólogas que serviriam para este papel. Além disso, também foram encontrados algumas famílias combinadas que poderiam explicar parte das doenças causadas pelo Streptococcus pyogenes em seres humanos. Os resultados mostram o potencial de uso do GTACG para encontrar novos objetos de pesquisa para estudos moleculares de genômica comparativa de bactérias. Research in the field of genomics produces a large amount of data. However, genetic knowledge about certain phenotypes is limited. Besides, a considerable part of the studied genomes has coding sequences (CDSs) with unknown functions, representing an additional challenge for researchers. Organisms from the same evolutionary branch share many of their CDSs, and certain phenotypes specific to a group of these individuals may be the result of a unique set of unique genes. In this work the GTACG computational framework is presented, a user-friendly tool to help researches to identify unique genetic characteristics in subgroups of bacterial genomes that have a common phenotype, finding data that differentiate them from other organisms in a simple way. GTACG analysis is based on the formation of homologous CDS groups based on local alignments. The front end is simple to use, and the package installation is designed to allow users with little knowledge of computer science can do complex analysis using this tool. The validation of the GTACG results was based on two case studies involving a set of 161 genomes of the Xanthomonadaceae family and 45 Streptococcus pyogenes genomes. In the first case study, we attempted to find out why some Xanthomonadaceae are associated with plants and others not, and, in fact, 19 families of orthologous proteins unique to plant-associated genomes were found (representing over 90% of these genomes), allowing the identification of proteins potentially associated with the adaptation and virulence of these bacteria in plant tissues. In the second study, we attempted to find phylogenetic markers for the protein emm of Streptococcus pyogenes, and found 15 families of orthologous proteins that would play this role. In addition, some combined families were also found that could explain some of the diseases caused by Streptococcus pyogenes in humans. The results show the potential use of GTACG to find new research objects for molecular studies of bacterial comparative genomics.

Elementos móveis influenciam expressivamente a evolução do grupo Bacteria, uma vez que são integrantes importantes do processo de geração da variabilidade genética e promovem a diversidade da população frente à estocasticidade do ambiente. Investigamos a influência de elementos genéticos móveis em linhagens de X. oryzae sob três diferentes aspectos nesta tese. Primeiramente, investigamos a transferência horizontal de um operon adquirido exclusivamente por Xoc, porém ausente em Xoo e linhagens filogenéticamente próximas. A importância da produção do fitormônio GA4 por Xoc é indicada pela alta prevalência do operon em diferentes populações, e pela ampla distrubuição geográfica em um amplo perído de tempo. Também abordamos a influência de elementos genéticos móveis mais amplamente no genoma de X. oryzae. Descrevemos como elementos de transposição, representados por elementos do tipo IS, são capazes de interferir em um sistema central do metabolismo bacteriano, como o sistema de reparo de DNA. Por fim, descrevemos um novo elemento encontrado em X. oryzae e analisamos sua organização, distribuição, filogenia, o nível transcricional e o potencial funcional na interação com seu hospedeiro Nosso conjunto de resultados reitera a importância dos elementos de transposição no aumento de variabilidade genética de populações, e dessa forma, infere sobre seu papel relevante na diversificação em X. oryzae. Descrevemos a variabilidade, exploramos as consequências da amplificação de elementos de transposição nesta espécie, evidenciamos a complexidade desta variação em nível populacional. Mobile elements are genomic components that trigger genetic variability and promote the diversity of the population against the stochasticity of the environment and therefore, significantly influence the evolution of the Bacteria group. We investigate the influence of mobile genetic elements on X. oryzae lineages under three different aspects in this thesis. First, we investigated the horizontal transfer event of a operon, acquired exclusively by Xoc, but absent in Xoo and phylogenetically close lineages. The importance of the production of the phytohormone GA4 by Xoc is indicated by its high prevalence in different populations, in a wide geographic distribution and in a large time period.We also address the influence of mobile genetic elements more comprehensively in the X. oryzae genome. We describe how transposable elements, represented by IS elements, are capable of interfering in a central system of bacterial metabolism, such as the DNA repair system. Finally, we describe a new element found in X. oryzae and we analyze its organization, distribution, phylogeny and its transcriptional level. We also speculate its function in the X. oryzae-host interaction. Our results reiterate the importance of transposable elements enriching genetic variability of populations and infers on their relevant role in the diversification of X. oryzae lineage. We describe the variability, explore the consequences resulting from the amplification of transposition elements in the X. oryzae species and we show the complexity of this variation at the population level.