Descobrim els clarinetistes de Barcelona : de la segona meitat del segle XIX fins la Guerra Civil

La finalitat d’aquest Projecte Fi de Grau és descobrir els clarinetistes que hi havia a Barcelona des de la segona meitat del segle XIX fins a la Guerra Civil, tant a nivell d’intèrprets com a nivell de professors. A més, ens hem proposat conèixer en quines institucions educatives aquests professors impartien les seves classes, com l’Escola Municipal de Música de Barcelona o el Conservatori del Liceu. Això ens ha portat, també, a identificar en quines agrupacions musicals desenvolupaven la seva carrera —com l’Orquestra Pau Casals, la Banda Municipal de Barcelona, l’Orquestra del Liceu o diferents orquestres de ball en les quals oferien concerts populars al llarg del territori. La metodologia utilitzada ha estat l'extracció i l’anàlisi de ressenyes hemerogràfiques i la recerca específica sobre la carrera artística del clarinetista que possiblement va realitzar a nivell estatal els primers enregistraments per a aquest instrument: en Josep Nori.

TRIAGEM METABÓLICA POR PKS E NRPS EM ACTINOBACTÉRIAS ENDOFÍTICAS DE Citrus reticulata

Polyketides and non-ribosomal peptides are natural products widely found in bacteria, fungi and plants. The biological activities associated with these metabolites have attracted special attention in biopharmaceutical studies. Polyketide synthases act similarly to fatty acids synthetases and the whole multi-enzymatic set coordinating precursor and extending unit selection and reduction levels during chain growth. Acting in a similarly orchestrated model, non-ribosomal peptide synthetases biosynthesize NRPs. PKSs-I and NRPSs enzymatic modules and domains are collinearly organized with the parent gene sequence. This arrangement allows the use of degenerated PCR primers to amplify targeted regions in the genes corresponding to specific enzymatic domains such as ketosynthases and acyltransferases in PKSs and adenilation domains in NRPSs. Careful analysis of these short regions allows the classifying of a set of organisms according to their potential to biosynthesize PKs and NRPs. In this work, the biosynthetic potential of a set of 13 endophytic actinobacteria from Citrus reticulata for producing PKs and NRP metabolites was evaluated. The biosynthetic profile was compared to antimicrobial activity. Based on the inhibition promoted, 4 strains were considered for cluster analysis. A PKS/NRPS phylogeny was generated in order to classify some of the representative sequences throughout comparison with homologous genes. Using this approach, a molecular fingerprint was generated to help guide future studies on the most promising strains.

Isolado do vírus do mosaico do pepino obtido de bananeira no Estado de São Paulo pertence ao subgrupo Ia

Em 1996, foi feita a caracterização parcial de um isolado do vírus do mosaico do pepino (Cucumis mosaic virus, CMV) obtido de bananeira (Musa sp.) proveniente do município de Miracatu, SP. Com o objetivo de se determinar o subgrupo do isolado de CMV, recorreu-se às técnicas de ELISA, RT-PCR, RFLP e seqüenciamento de fragmentos de RNA genômico. Amostras de folhas infetadas, desidratadas com cloreto de cálcio e armazenadas à -20 °C desde 1994 na viroteca do Laboratório de Fitovirologia e Fisiopatologia, foram inoculadas em plantas de Nicotiana glutinosa. Dez dias após a inoculação, folhas apresentando mosaico foram utilizadas para DAS-ELISA e extração de RNAs totais. Em ELISA, houve reação apenas contra o anti-soro específico para CMV subgrupo I. Através de RT-PCR com primers desenhados para anelar em regiões conservadas da porção terminal 3' do gene da capa protéica, foi amplificado um fragmento de DNA com 486 pares de bases. O produto obtido via RT-PCR foi submetido à digestão com as enzimas EcoRI, HindIII, BamHI e MspI, obtendo-se um padrão de restrição esperado para o subgrupo I. Estes resultados foram confirmados através do seqüenciamento do produto de PCR, o qual apresentou homologia de 96% a 98% com os isolados do CMV pertencentes ao subgrupo I. Pelos sintomas observados na hospedeira diferencial Vigna unguiculata, o isolado foi confirmado como sendo do subgrupo Ia.

RT-PCR and Dot Blot hybridization methods for a universal detection of tospoviruses

Transcriptase reverse - polymerase chain reaction (RT-PCR) and dot blot hybridization with digoxigenin-labeled probes were applied for the universal detection of Tospovirus species. The virus species tested were Tomato spotted wilt virus, Tomato chlorotic spot virus, Groundnut ringspot virus, Chrysanthemum stem necrosis virus, Impatiens necrotic spot virus, Zucchini lethal chlorosis virus, Iris yellow spot virus. Primers for PCR amplification were designed to match conserved regions of the tospovirus genome. RT-PCR using distinct primer combinations was unable to simultaneously amplify all tospovirus species and consistently failed to detect ZLCV and IYSV in total RNA extracts. However, all tospovirus species were detected by RT-PCR when viral RNA was used as template. RNA-specific PCR products were used as probes for dot hybridization. This assay with a M probe (directed to the G1/G2 gene) detected at low stringency conditions all Tospovirus species, except IYSV. At low stringency conditions, the L non-radioactive probe detected the seven Tospovirus species in a single assay. This method for broad spectrum detection can be potentially employed in quarantine services for indexing in vitro germplasm.

Garlic viral complex: identification of Potyviruses and Carlavirus in Central Brazil

Garlic viruses often occur in complex infections in nature. In this study, a garlic virus complex, collected in fields in Brazil, was purified. RT-PCR was performed using specific primers designed from the consensus regions of the coat protein genes of Onion yellow dwarf virus, a garlic strain (OYDV-G) and Leek yellow stripe virus (LYSV). cDNA of Garlic common latent virus (GCLV) was synthesized using oligo-dT and random primers. By these procedures individual garlic virus genomes were isolated and sequenced. The nucleotide sequence analysis associated with serological data reveals the presence of two Potyvirus OYDV-G and LYSV, and GCLV, a Carlavirus, simultaneously infecting garlic plants. Deduced amino acid sequences of the Brazilian isolates were compared with related viruses reported in different geographical regions of the world. The analysis showed closed relations considering the Brazilian isolates of OYDV-G and GCLV, and large divergence considering LYSV isolate. The detection of these virus species was confirmed by specific reactions observed when coat protein genes of the Brazilian isolates were used as probes in dot-blot and Southern blot hybridization assays. In field natural viral re-infection of virus-free garlic was evaluated.

Sequence analysis of the capsid protein gene of an isolate of Apple stem grooving virus, and its survey in Southern Brazil

Apple stem grooving virus (ASGV) is one of the most important viruses infecting fruit trees. This study aimed at the molecular characterization of ASGV infecting apple (Malus domestica) plants in Santa Catarina (SC). RNA extracted from plants infected with isolate UV01 was used as a template for RT-PCR using specific primers. An amplified DNA fragment of 755 bp was sequenced. The coat protein gene of ASGV isolate UV01 contains 714 nucleotides, coding for a protein of 237 amino acids with a predicted Mr of approximately 27 kDa. The nucleotide and the deduced amino acid sequences of the coat protein gene showed identities of 90.9% and 97.9%, respectively, with a Japanese isolate of ASGV. Very high amino acid homologies (98.7%) were also found with Citrus tatter leaf capillovirus (CTLV), a very close relative of ASGV. These results indicate low coat protein gene variability among Capillovirus isolates from distinct regions. In a restricted survey, mother stocks in orchards and plants introduced into the country for large scale fruit production were indexed and shown to be infected by ASGV (20%), usually in a complex with other (latent) apple viruses (80%).

Identificação de xanthomonas axonopodis pv. phaseoli e x. axonopodis pv. phaseoli var. fuscans através da técnica de pcr

Este trabalho teve por objetivo verificar a viabilidade de utilização da técnica de PCR, usando primers considerados específicos, para identificação de Xanthomonas axonopodis pv. phaseoli (Xap) e x. axonopodis pv. phaseoli var. fuscans (Xapf), visando criar bases para sua utilização em diagnose rotineira e em programas de certificação de sementes. Foram incluídos no estudo 42 isolados da bactéria provenientes de locais distintos, além de outros gêneros, espécies e patovares, para ser verificada a especificidade dos primers. Os resultados obtidos permitem concluir que os primers utilizados são adequados para identificação de Xap e Xapf, embora dois isolados patogênicos não tenham sido amplificados. Foi observada também a amplificação de uma banda fraca para X. axonopodis pv. vitians, com o mesmo número de pares de bases, o que sugere existir homologia entre estes patovares, na região amplificada.

Identification of rapd marker linked to blast resistance gene in a somaclone of rice cultivar Araguaia

The gene Pi-ar confers resistance to Pyricularia grisea race IB-45 in a somaclone derived from immature panicles of the susceptible rice (Oryza sativa) cultivar Araguaia. RAPD technique was used to identify molecular markers linked to this gene utilizing bulked segregant analysis. Initially, the two parental DNAs from the resistant donor SC09 and 'Araguaia' were analyzed using random primers. Of the 240 primers tested, 203 produced amplification products. The two parental DNAs along with the resistant and susceptible bulks of F2 population were screened using 48 primers that differentiated resistant and susceptible parents. Even though eight primers differentiated the resistant bulk from the susceptible bulk, as well as somaclone SC09 and 'Araguaia', only one primer, OPC02 ('GTGAGGCGTC'), was found to be tightly linked (1.7cM) to the resistance gene of somaclone SC09.

Genetic analyses of Rhizoctonia solani isolates from Phaseolus vulgaris grown in the Atlantic Rainforest Region of São Paulo, Brazil

Rhizoctonia solani isolates obtained from common beans (Phaseolus vulgaris) grown in the mountainous Atlantic Rainforest (Mata Atlântica) region of São Paulo, Brazil, were analyzed to determine their genetic diversity using internal transcribed spacer (ITS), microsatellite and telomere sequence-based PCR primers. Restriction digestion of the ITS1/5.8S/ITS2 ribosomal regions yielded unique banding patterns specific for AG4 and its subgroups. The ITS restriction digestion (ITS/RFLP), telomere and microsatellite primers identified five to 11 genotypes within the isolates of R. solani. While all isolates were pathogenic on beans, there was no correlation found between genotypic differences and pathogenicity. The different PCR primers revealed a number of isolates that were genetically similar. Some of these genetic groups were supported by more than one of the primers utilized in this study, thus confirming their relationship.

Detection and coat protein gene characterization of an isolate of Grapevine virus B from corky bark-affected grapevines in Southern Brazil

An isolate of Grapevine virus B (GVB), obtained by indexing Vitis labrusca and V. vinifera grapevines on the indicator LN33, was transmitted mechanically to several Nicotiana species. The virus was partially purified from N. cavicola and the coat protein estimated at 23 kDa by SDS-PAGE. In negatively stained leaf extracts of experimentally inoculated N. cavicola and N. occidentalis, flexuous particles with cross banding were observed, predominantly measuring 750-770 x 12 nm, with a modal length of 760 nm. Decoration indicated a clear, positive reaction against AS-GVB. In DAS-ELISA, GVB was detected in N. cavicola and grapevine extracts, and Western blots showed homologous and cross reaction of GVB and GVA antisera with GVB coat protein. Using specific primers for GVB, a fragment of 594 bp, comprising the coat protein gene coding for 197 amino acids, was amplified by RT-PCR with viral RNA extracted from GVB-infected N. occidentalis. The nucleotide and the deduced amino acid sequences of the coat protein gene showed high identities with Italian and Japanese isolates of GVB.

Análise de restrição de DNA ribossomal amplificado (ARDRA) pode diferenciar Fusarium solani f. sp. phaseoli de F. solani f. sp. glycines

Métodos moleculares têm sido utilizados para caracterizar a diversidade entre isolados de Fusarium spp. patogênicos e não patogênicos a uma cultura e, para determinar relações genéticas entre formae speciales. Testes de patogenicidade realizados em soja (Glycine max) e feijoeiro (Phaseolus vulgaris) com 17 isolados de Fusarium solani não demonstraram especificidade de hospedeiros. Utilizou-se a técnica ARDRA (Amplified Ribosomal DNA Restriction Analysis) para analisar a região ITS1 - 5,8S rDNA - ITS2, amplificada com os primers ITS5 e ITS4. Os produtos amplificados foram digeridos com as enzimas de restrição Hae III e Msp I. Os padrões de bandas gerados pela digestão com a enzima Hae III permitiram diferenciar três grupos entre os isolados de F. solani, sendo um grupo específico para isolados de F. solani f. sp. phaseoli com 100% de similaridade entre os 11 isolados. Entre os isolados de F. solani f. sp glycines foram observados dois padrões distintos de restrição. A técnica de ARDRA utilizando a enzima Hae III apresenta, portanto, potencial para utilização como um marcador para diferenciação entre as formae specialesphaseoli e glycines, dentro do complexo F. solani.

An isolate of Apple stem grooving virus associated with Cleopatra mandarin fruit intumescence

A citrus tatter leaf isolate (CTLV-Cl) of Apple stem grooving virus (ASGV) has been found to be associated with a fruit rind intumescence in Cleopatra mandarin (Citrus reshni) in Limeira (SP). The CTLV-Cl was mechanically transmitted to the main experimental herbaceous hosts of CTLV. Chenopodium quinoa and C. amaranticolor reacted with local lesions and systemic symptoms while other test plants reacted somewhat differently than what is reported for CTLV. A pair of primers designed for specific detection of ASGV and CTLV amplified the expected 801 bp fragment from the CTLV-Cl-infected plants. Typical capillovirus-like particles were observed by the electron microscope in experimentally infected C. quinoa and C. amaranticolor leaves.

Mapeamento de genes de resistência do feijoeiro à ferrugem, antracnose e mancha-angular usando marcadores RAPD

A organização de diferentes genes de resistência da cultivar Ouro Negro de feijoeiro-comum (Phaseolus vulgaris) à ferrugem, antracnose e mancha-angular foi estudada com o auxílio de marcadores moleculares. Uma população de 154 linhas endogâmicas recombinantes (RIL's) obtidas do cruzamento entre as cultivares Ouro Negro e Rudá foram inoculadas com sete raças fisiológicas de Uromyces appendiculatus, três de Colletotrichum lindemuthianum, e quatro de Phaeoisariopsis griseola. Amostras de DNA de cada uma das RIL's foram amplificadas via PCR utilizando 70 diferentes primers. A análise da segregação da resistência à ferrugem, antracnose e mancha-angular na população de 154 RIL's revelou diferentes modos de herança para a resistência a cada uma das raças fisiológicas. A análise de ligação genética revelou que os diferentes genes de resistência à ferrugem e à antracnose estão no mesmo grupo de ligação. Os genes de resistência à mancha-angular também foram mapeados juntos, porém em outro grupo de ligação. Verificou-se neste trabalho que a utilidade dos marcadores RAPD, previamente identificados como ligados a genes de resistência do feijoeiro a doenças foi restrita. Apenas cinco dos 38 marcadores moleculares testados foram validados na população de RIL's como ligados aos genes de resistência à ferrugem e à antracnose. Três novos marcadores (OBA16(669) e OBA16(583) a 10,4 cM em acoplamento e OAD9(3210) a 13,9 cM em repulsão) ligados ao bloco gênico de resistência da cultivar Ouro Negro à mancha-angular foram identificados.

Genotypic diversity among brazilian isolates of Macrophomina phaseolina revealed by RAPD

Macrophomina phaseolina has been considered one of the most prevalent soybean (Glycine max) pathogens in Brazil. No genetic resistance has been determined in soybean and very little is known about the genetic diversity of this pathogen in tropical and sub-tropical regions. Fifty-five isolates from soybean roots were collected in different regions and analyzed through RAPD for genetic diversity. The UPGMA cluster analysis for 74 loci scored permitted identification of three divergent groups with an average similarity of 99%, 92% and 88%, respectively. The three groups corresponded to 5.45%, 59.95% and 34.6%, respectively of all isolates used. A single plant had three different haplotypes, while 10.9% of the analyzed plants had two different haplotypes. In another study the genetic similarity was evaluated among isolates from different hosts [soybean, sorghum (Sorghum bicolor), sunflower (Helianthus annuus), cowpea (Vigna unguiculata), corn (Zea mays) and wheat (Triticum aestivum)] as well as two soil samples from native areas. Results showed that more divergent isolates originated from areas with a single crop. Isolates from areas with crop rotation were less divergent, showing high similarity values and consequently formed the largest group. Amplification of the ITS region using primers ITS1 and ITS4 produced only one DNA fragment of 620 bp. None of the isolates were differentiated through PCR-RFLP. Our results demonstrated genetic variability among Brazilian isolates of M. phaseolina and showed that one single root can harbor more than one haplotype. Moreover, cultivation with crop rotation tends to induce less specialization of the pathogen isolates. Knowledge of this variation may be useful in screening soybean genotypes for resistance to charcoal rot.

Uso de marcadores RAPD na classificação de isolados de Phytophthora spp. causadores da podridão parda do cacaueiro no Brasil

Objetivou-se neste trabalho propor uma metodologia para a utilização de marcadores RAPD como uma ferramenta auxiliar na classificação de isolados de Phytophthora spp. causadores da podridão-parda do cacaueiro (Theobroma cacao) no Brasil. Existe uma necessidade constante de monitorar populações de Phytophthora spp. nas regiões cacaueiras do Brasil e a tarefa de classificação dos isolados é difícil e demorada. Com base em estudos de diversidade genética de isolados de Phytophthora capsici, P. palmivora e P. citrophthora por meio de marcadores RAPD, foram escolhidos três isolados de cada espécie como padrões e dois "primers" decâmeros mais informativos na diferenciação das espécies (OPA 13 e OPH 18). O DNA genômico dos isolados padrões e de três isolados não classificados foi extraído e amplificado, utilizando-se os dois "primers" decâmeros mais informativos. Os padrões de marcadores RAPD obtidos permitiram uma diferenciação visual clara dos isolados de cada espécie e mostraram-se úteis na classificação de isolados de Phytophthora spp. A metodologia proposta já está sendo utilizada no Centro de Pesquisas do Cacau, solucionando eventuais dúvidas resultantes da caracterização morfológica dos isolados.