Thesis (PhD(Agric))--University of Stellenbosch, 2000. / ENGLISH ABSTRACT: The purpose of this study was to characterize the race and vegetative compatibility of Fusarium oxysporum f. sp. melonis (FOM) isolates collected in the major melon producing areas, to report on their geographical distribution, and their possible relatedness to isolates from other countries. Seventy two FOM isolates obtained from 30 fields in 17 melon producing regions were race-typed using the differential cultivars Topmark (susceptible to all races), Doublon (Fomi), CM 17187 (Fom2) and Perlita (Fom3) and grouped by means of vegetative compatibility. All isolates belonged to vegetative compatibility group 0134, indicating a high degree of genetic homogeneity among the South African FOM population. Fifty four isolates were identified as race 0, eight as race 1, and 10 as race 2. Race 0 occurred in 15 of the regions whereas race 1 was sporadically recovered. Race 2, on the other hand, was obtained only from four fields located in one geographical region. Perlita plants (carrying the gene Fom3) inoculated with local isolates ofrace 0 and race 2 and reference isolates of race 0 became stunted, their leaves turned yellow, and became thickened and brittle. These results suggested that Fom3 in Perlita confers a tolerant reaction compared to the resistant reaction of gene FornI in Doublon. The disease reaction of cultivar Perlita to FOM was therefore reinvestigated. Twenty isolates, including the four FOM races (0, 1, 2, and 1,2) obtained from different countries, were used. The differential cultivars were included to verify virulence of the isolates. Perlita plants inoculated with three isolates of race 2 remained asymptomatic. The remaining race 2 and 0 isolates, induced severe stunting of Perlita plants, but mean percentage stunting values did not differ significantly (P = 0.05) and ranged between 25.1 and 50.0. Leaves of stunted plants were chlorotic, thickened and brittle. Disease reaction of Perlita was verified at a lower inoculum concentration with two race 2 (pipette method) and two race 0 isolates (root dip method). Results proved that Fom3 does not confer similar resistance towards race 0 and some race 2 isolates as FornI in Doublon. Cultivars possessing Fom3, should therefore be considered tolerant to FOM races 0 and 2. The ability of a nit mutant isolate, generated from FOM race 0 which belongs to VCG 0134, to change its virulence during infection of melon plants, was investigated under quarantine. Seedlings of melon cultivars Imperial 45 and Early Sweet (no resistance genes), Amber (Fom2) and Fiata (FomI, Fom2) were consecutively grown in two cement troughs in a gauzehouse. Each planting was terminated when plants had advanced Fusarium wilt or after the fruit were harvested. In the first planting, Imperial 45 seedlings were transplanted and artificially inoculated with the nil mutant isolate. In the consecutive plantings, seeds were sown in the infested soil to enable natural infection. For each crop, representative plants showing Fusarium wilt were selected for isolation. All F. oxysporum isolates recovered were single-spored and their nit mutant and VCG status verified. Virulence of the labelled isolates was determined using differential cultivars. In trough A, all plants of the susceptible cultivars Imperial 45 and Early Sweet crops showed Fusarium wilt. The labelled isolates recovered from the selected plants were all designated race O. In the first crop (planting No.5) of the resistant cultivar Amber, 6.7% of the plants developed Fusarium wilt. In the second Amber crop the disease incidence increased to 56.6%, and to 81.8% in the final crop. Contrary to the susceptible cultivars, only race 2 isolates were obtained from the symptomatic Amber plants. Similar data were found with the susceptible cultivar Imperial 45 and the resistant cultivar Amber in trough B. Planting of Fiata caused a dramatic reduction in Fusarium wilt incidence in trough B. However, 1.2% of plants were affected by Fusarium wilt in the first Fiata crop (planting No.6), whereas 4% of the plants were symptomatic in the final planting. From these symptomatic Fiata plants only race 1,2 isolates were obtained. These findings, and the fact that the symptomatic plants represented a substantial proportion of the first Amber (approximately 7-15%) and Fiata (approximately 2%) crops, provedthat changes in the race structure of this fungal pathogen occurred rapidly when confronted with a resistant cultivar. The potential of RAPD analysis to differentiate between the isolates displaying virulence changes was evaluated. Four F. oxysporum f. sp. niveum isolates were included as an outgroup. A histopathological study was conducted to verify whether these isolates retain their ability to behave as true vascular pathogens. The three primers used clearly distinguished the 12 FOM isolates from the four F. oxysporum f. sp. niveum isolates. However, the primers showed a highly conserved and characteristic banding pattern for the FOM isolates which represented three physiological races (race 0, race 2, race 1,2), indicating that RAPD analysis cannot detect race-specific groupings in FOM. Disease reactions on the three differential cultivars confirmed the virulence of FOM isolates. The histopathological data furthermore proved that the two FOM races (race 2, race 1,2), which derived from the race 0 parent isolate, retained their ability to behave as true vascular pathogens. / AFRIKAANSE OPSOMMING: DIE KARAKTERISERING EN PATOGENESITEIT VAN SUID-AFRIKAANSE ISOLATE VAN FUSARIUMOXYSPORUMF. SP.MELONIS Die doel van die studie was om Fusarium oxysporum f. sp. melonis (FOM) isolate wat in die hoof spanspekproduserende gebiede versamel is, volgens ras en vegetatiewe verenigbaarheid te karakteriseer, en hul geografiese verspreiding en verwantskap met isolate van ander lande aan te dui. Twee en sewentig FOM isolate afkomstig vanaf 30 landerye wat 17 spanspekproduserende areas verteenwoordig, is gebruik. Die differensiële kultivars Topmark (vatbaar vir alle rasse), Doublon (Forni), CM 17187 (Fom2) en Perlita (Fond) is gebruik om die rasbepalings te doen asook om die vegetatiewe verenigbare groepe (VVG) te bepaal. Al die isolate is as VVG 0134 geklassifiseer, wat 'n hoë mate van genetiese homogenesiteit binne die Suid-Afrikaanse populasie aandui. Vier en vyftig isolate is as ras 0, agt as ras 1 en 10 as ras 2 geklassifiseer. Ras 0 is vanaf 15 gebiede afkomstig, terwyl ras 1 sporadies voorgekom het. Ras 2 is vanuit vier landerye binne dieselfde geografiese gebied verkry. Plante van die kultivar Perlita wat met plaaslike isolate van ras 0 en 2, asook verwysings-isolate van ras 0 geïnokuleer is, het verdwerg voorgekom. Die blare van die plante het vergeel, verdik en bros voorgekom. Hierdie siekte reaksie het aangedui dat Fond in Perlita toleransie bewerkstellig in teenstelling met die weerstandbiedende reaksie van geen Fomi in Doublon. Die siekte reaksie van Perlita teenoor FOM is dus verder ondesoek. Hiervoor is 20 isolate wat al vier FOM rasse insluit (0, 1, 2, en 1,2), en van verskillende wêrelddele afkomstig is, gebruik. Die virulensie van die isolate is met die differensiële kultivars bevestig. Drie van die ras 2 isolate het geen siektesimptome op Perlita veroorsaak nie. Die ander ras 2 isolate, en al die ras 0 isolate, het egter die Perlita plante aansienlik verdwerg en die blare vergeel en verdik. Laasgenoemde groep isolate het 'n gemiddelde verdwergingsindeks van tussen 25.1% en 50.0% veroorsaak. Die siekte reaksie by Perlita is verder bevestig deur plante teen 'n laer inokulumdigtheid van twee ras 2 (pipet metode), en twee ras 0 (wortel-doop metode) isolate, te inokuleer. Uit die resultate was dit duidelik dat die weerstand wat Fom3 teenoor ras 0 en sommige ras 2 isolate verskaf, van FornI verskil. Kultivars wat oor die weerstandsgeen Fom3 beskik moet dus as tolerant beskou word. 'n Ondersoek is geloods na die vermoë van 'n nil mutant isolaat, genereer vanaf die wilde ras 0 isolaat van FOM (VVG 0134), om onder kwarantyn sy virulensie gedurende infeksie van spanspekplante te verander. Saailinge van die spanspekkultivars Early Sweet (geen weerstandsgene), Amber (Fom2) en Fiata (FornI, Fom2) is opeenvolgens in twee sement trêe in 'n gaashuis verbou. Die afsonderlike aanplantings is beëindig sodra gevorderde Fusarium-verwelksimptome verkry is, of nadat vrugte ge-oes is. Vir die eerste aanplanting is oorgeplante Imperial 45 saailinge kunsmatig met die nil mutant isolaat geïnokuleer. Tydens die opeenvolgende aanplantings is saad direk in die besmette grond gesaai ten einde natuurlike infeksie te verkry. Met elke aanplanting is isolasies gedoen vanaf verteenwoordigende plante wat Fusarium-verwelksimptome getoon het. Alle F. oxysporum isolate wat verkry is, is ge-enkelspoor en hul nit mutant status en VVG is bevestig. Virulensie van die gemerkte isolate is bepaal deur inokulasie van die differensiële kultivars. Alle plante van die vatbare Imperial 45 en Early Sweet kultivars wat in trog A geplant is, het Fusarium-verwelksimptome getoon. Die gemerkte isolate wat vanaf die verteenwoordigende plante verkry is, is almal as ras 0 geklassifiseer. Tydens die eerste aanplanting van die weerstandbiedende kultivar, Amber (aanplanting No.5), het 6.7% van die plante Fusarium-verwe1ksimptome ontwikkel. Tydens die tweede en derde aanplanting van Amber het die frekwensie van siektevoorkoms verhoog na 56.6% en 81.8 %, onderskeidelik. In teenstelling met die vatbare kultivars, is slegs ras 2 vanuit die Amber plante met siektesimptome verkry. Soortgelyke resultate is met Imperial 45 en Amber in trog B verkry. Aanplanting van kultivar Fiata het egter 'n dramatiese verlaging in die voorkoms van Fusarium-verwelk bewerkstellig. Tydens die eerste Fiata aanplanting (aanplanting No.6) het 1.2% plante Fusarium-verwelksimptome ontwikkel, en 4% tydens die laaste aanplanting. Vanaf hierdie plante is slegs ras 1,2 isolate verkry. Hierdie bevindings, en die feit dat 'n aansienlike hoeveelheid van die Amber (ongeveer 7-15%) en Fiata plante (ongeveer 2%) siektesimptome getoon het, bewys dat FOM vinnig van virulensie verander wanneer die patogeen 'n weerstanbiedende kultivar infekteer. Die vermoë van RAPD analise om tussen isolate wat in virulensie verander het, te onderskei, is ondersoek. Vier isolate van F. oxysporum f. sp. niveum is as 'n buite-groep ingesluit. Om te bevestig dat die isolate wat van ras verander het wel egte vaskulêre patogene is, is 'n histopatologiese ondersoek gedoen. Die drie inleiers wat gebruik is, het die 12 FOM isolate duidelik van die vier F. oxysporum f. sp. niveum isolate onderskei. Die 12 FOM isolate wat drie fiosologiese rasse (ras 0, ras 2, ras 1,2) verteenwoordig het, is egter saam gegroepeer, wat aandui dat hierdie metode nie tussen rasse van FOM kan onderskei nie. Inokulasiestudies met die differensiële kultivars het die virulensie van die isolate bevestig. Die histopatologiese ondersoek het verder bewys dat beide FOM rasse (ras 2, ras 1,2) wat vanaf die wilde tipe ras ° isolaat ontstaan het, hul vermoë behou het om as egte vaskulêre patogene op te tree.

Fusarium oxysporum Schl. var. vanillae (Tucker) Gondon is known to cause root rot in Vanilla planifolia Andrews in most regions where it is grown, including the major plantations in Xishuangbanna, Yunnan Province of China. This is of serious economic concern to the Province since the vanilla flavouring extractable from the beans of the plant is a valuable food product and an important export commodity. There are no fungicides registered for the control of Fusarium root rot and the only available chemical control methods are ineffective and cause serious contamination of the soil. Breeding for resistance is difficult when no dominant gene is known or where little information is available on fungal pathogenicity. Biocontrol is the main alternative for disease control in this crop, an attractive approach because of increasing concerns for environmental protection. The investigation considers two biocontrol strategies: first the introduction of virulent, antagonistic, non-pathogenic strains, closely-related to the pathogen, to overcome pathogenic populations in infected soils; second the use of essential oils with antimicrobial properties when applied to infected soils. Pathogenicity tests have been done on 81 out of 87 F. oxysporum isolates collected in Yunnan Province. Among these, 32 isolates were non-pathogenic and 49 were pathogenic. The pathogenicity results showed the complexity of F. oxysporum in Yunnan. Seventeen isolates were recovered from the Daluo plantation, of which 14 were pathogenic isolates and 3 non-pathogenic isolates; 26 from the Menglun plantation, in which 12 were pathogenic and 14 were non-pathogenic; 18 isolates from the Manjingdai plantation, in which 12 isolates were pathogenic, whilst the other 6 were non-pathogenic and 20 were obtained from the plantation in Hekou i County, of which 11 were pathogenic isolates and 9 were non-pathogenic. Genetic diversity within this population of F. oxysporum has been investigated with respect to vegetative compatibility and to determine the relationship between VCGs and virulence. The VCG results showed that the 87 strains of Fusarium oxysporum f.sp vanillae isolated from Yunnan Province were complex. They could be distributed into 12 different VCGs and that a direct relationship between VCGs group and virulence could not be drawn. Two non-pathogenic strains, ML-5-2 and HK-5b-4-1, have been screened from 87 strains as candidate biocontrol agents by pathogenicity and VCG, which are self-incompatible and closely related to the pathogens. These two strains were effective in vanilla root rot control in controlled environments, but their effects in field experiments were less conclusive. Seven essential oils, which have long been regarded as having inhibitory effects on pathogens in nature, have also been investigated as biocontrol agents. Three oils, cinnamon oil, thyme oil and clove oil, were effective in inhibiting the growth of pathogen in vitro. These oils may develop into useful components of different management strategies with non-pathogenic strains. For the future, consideration will need to be given to the mechanism(s) of the interaction of the antagonistic components with the soil microbe population and host plant and also to appropriate formulation, to take account of soil type, crop status, cultural practices, environmental and economic factors. Biocontrol methods have considerable potential but must be acceptable to farmers as part of an overall crop management programme.

Dioxygenase-cytochrome P450 (DOX-CYP) fusion enzymes are present in diverse human and plant pathogenic fungi. They oxygenate fatty acids to lipid mediators which have regula­tory functions in fungal development and toxin production. These enzymes catalyze the for­mation of fatty acid hy­droperoxides which are subsequently converted by the P450 activities or reduced to the corresponding alcohols. The N-terminal DOX domains show catalytic and structural homology to mammalian cyclooxygenases, which belong to the most thoroughly studied human enzymes. 7,8-Linoleate diol synthase (LDS) of the plant pathogenic fungus Gaeumannomyces graminis was the first characterized member of the DOX-CYP fusion enzyme family. It catalyzes the conversion of linoleic acid to 8R-hydroperoxylinoleic acid (HPODE) and subse­quently to 7S,8S-dihy­droxylinoleic acid by its DOX and P450 domains, respectively. By now, several enzymes with homology to 7,8-LDS have been identified in im­portant fungi, e.g., psi fac­tor-producing oxygenase (ppo)A, ppoB, and ppoC, of Aspergillus nidulans and A. fumigatus. By cloning and recombinant expression, ppoA of A. fumigatus was identi­fied as 5,8-LDS. Partial expression of the 8R-DOX domains of 5,8-LDS of A. fumigatus and 7,8-LDS of G. graminis yielded active protein which demonstrates that the DOX activities of LDS are independent of their P450 domains. The latter domains were shown to contain a conserved motif with catalytically important amide residues. As judged by site-directed mutagene­sis studies, 5,8- and 7,8-LDS seem to facilitate heterolytic cleavage of the oxygen-oxygen bond of 8R-HPODE by aid of a glutamine and an asparagine residue, respectively. Cloning and expression of putative DOX-CYP fusion proteins of A. terreus and Fusarium oxysporum led to the discovery of novel enzyme activities, e.g., linoleate 9S-DOX and two allene oxide synthases (AOS), specific for 9R- and 9S-HPODE, respectively. The fungal AOS are present in the P450 domains of two DOX-CYP fusion enzymes and show higher se­quence homology to LDS than to plant AOS and constitute therefore a novel class of AOS. In summary, this thesis describes the discovery of novel fatty acid oxy­genases of human and plant pathogenic fungi and the characterization of their reaction mechanisms.

Submitted by Gabriela Lopes (gmachadolopesufpel@gmail.com) on 2017-06-20T16:50:30Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertação Dediel.pdf: 766607 bytes, checksum: 1d7fec62b5b0587e0accd22056511b85 (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2017-06-21T18:58:18Z (GMT) No. of bitstreams: 2 Dissertação Dediel.pdf: 766607 bytes, checksum: 1d7fec62b5b0587e0accd22056511b85 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-06-21T18:58:18Z (GMT). No. of bitstreams: 2 Dissertação Dediel.pdf: 766607 bytes, checksum: 1d7fec62b5b0587e0accd22056511b85 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2012-03-30 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / A busca por alternativas no controle de doenças em substituição ao uso intensivo de agrotóxicos tem sido objeto da pesquisa agrícola. Rizobactérias são conhecidas pelo biocontrole de doenças e promoção de crescimento em diversos cultivos. Os objetivos deste trabalho foram avaliar a eficácia de seis rizobactérias, pré- selecionadas, no controle de Ralstonia solanacearum e Fusarium oxysporum f. sp. lycopersici (FOL), em casa de vegetação e relacionar este comportamento a produção de compostos “in vitro”, bem como avaliar a proteção de plantas de tomate contra doenças da parte aérea que ocorreram naturalmente em condições de campo. Foi avaliada a capacidade destas rizobactérias em produzir quitinases, amilases, lipases, compostos antibióticos e de solubilizar fosfato. A microbiolização das sementes com a rizobactéria DFs1420 (Bacillus sp.) reduziu a severidade da murcha bacteriana em 66% aos 14 dias após a inoculação, no primeiro ensaio e valores de AACPD em 70%, no segundo ensaio. Este controle pode ser associado à produção de compostos responsáveis pela antibiose observada “in vitro”. Streptomyces (DFs1296 e DFs1315) e Bacillus sp. (DFs1414) reduziram significativamente a murcha de fusário. O controle observado pode ser atribuído à atividade quitinolítica e/ou antibiótica por compostos voláteis. Foram instalados três ensaios de campo. A severidade das doenças foliares foi monitorada ao longo do tempo. As rizobactérias foram capazes de proteger as plantas contra Septoria lycopersici. DFs1414 e DFs1421 (Pseudomonas sp.) foram as mais estáveis, proporcionando proteção em dois ensaios consecutivos. As rizobactérias DFs1296 e DFs1420 foram capaz de controlar a requeima (Phytophthora infestans) em dois ensaios. A bactéria DFs1296 também apresentou capacidade de proteção contra estas e outras doenças quando pulverizada semanalmente na parte aérea. De modo geral, o controle alcançado por estas bactérias, microbiolizadas às sementes ou em aplicação foliar, não diferiu dos tratamentos utilizando produtos químicos recomendados para a cultura em aplicações semanais. / The search on alternative control of diseases to replace the use of pesticides has been the subject of agricultural research. Rhizobacteria are known by controlling diseases and promoting growth in several crops. The objectives of this work were to evaluate the efficacy of six rhizobacteria pre-selected for the control of Ralstonia solanacearum and Fusarium oxysporum f. sp. lycopersici (FOL) in greenhouse, and to relate this behavior with the production of compounds "in vitro", as well as evaluate the protection of tomato plants against foliar diseases under field conditions. It was evaluated the ability of these rhizobacteria to produce chitinases, amylases, lipases, antibiotic compounds and to solubilize phosphate. The seed microbiolization with rhizobacterium DFs1420 (Bacillus sp.) reduced the severity of the bacterial wilt 66% at 14 days after inoculation and AUDPC 70%, respectively, the first and second assays. This control may be associated with the production of compounds responsible for the antibiosis observed "in vitro". Streptomyces (DFs1296 and DFs1315) and Bacillus sp. (DFs1414) significantly reduced fusarium wilt. The control observed can be attributed to the chitinolytic activity and/or antibiosis in the presence of volatile compounds. Three field trials were carried out in field. The foliar diseases severity was monitored over time. The rhizobacteria were capable of protecting plants against Septoria lycopersici. DFs1414 and DFs1421 (Pseudomonas sp.) were the most stable, providing protection in two consecutive trials. The rhizobacteria DFs1296 and DFs1420 were able to control late blight (Phytophthora infestans) in two trials. DFs1296 also had the ability to protect against these and other diseases when sprayed weekly in the plant canopy.In general, the control achieved by these rhizobacteria, by seed microbiolization or by foliar application, did not differ from treatments using recommended chemical in weekly applications.

Submitted by (lucia.rodrigues@ufrpe.br) on 2017-03-10T15:46:17Z No. of bitstreams: 1 Frederick Mendes Aguiar.pdf: 412210 bytes, checksum: 4f5f91c02dea9d844bac53a51c2cafa0 (MD5) / Made available in DSpace on 2017-03-10T15:46:17Z (GMT). No. of bitstreams: 1 Frederick Mendes Aguiar.pdf: 412210 bytes, checksum: 4f5f91c02dea9d844bac53a51c2cafa0 (MD5) Previous issue date: 2011-02-24 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Okra is a vegetable crop with high nutritional value, being a rich source of vitamins and mineral salts. Fusarium wilt (caused by Fusarium oxysporum f. sp. vasinfectum – FOV) is one of the major field diseases of okra in tropical areas. In the present work, 53 okra accessions and the commercial cultivar Santa Cruz 47 were evaluated aiming to identify sources of resistance to Brazilian FOV isolates and to study the resistance stability of the selected accessions to different pathogen isolates. An initial screening was carried out with only one FOV isolate (Fus-194). In the second assay, thirty-three accessions identified in the first screening were re-evaluated in two assays (in two different seasons) using two FOV isolates (Fus-194 and Fus-201). The resistance evaluation was carried out with 21 day-old plantlets, using the root-dipping inoculation procedure, utilizing a spore suspension of 1x106 conidia/mL. The evaluation was done using a disease severity grade system with grades ranging from 0 to 4. These grades were used to generate a disease index that was employed for clustering the accessions according to their reaction to FOV. In the evaluation carried out in August (average temperature of 19,8°C) only 12 out the 32 accessions (i.e. 37%) were rated as having high to intermediate resistant response to Fus-194 isolate. Only 28% of the accessions were classified within the high to intermediate resistance cluster when using the Fus-201 isolate. In the assay carried out in October (higher temperatures) 72% of the accessions were classified as resistant and intermediate resistant to Fus-194 isolate, whereas 32% were resistant to isolate Fus-201. Our results indicated that the Fus-201 isolate was more aggressive than Fus-194. Comparative analysis of the assays indicated that the overall aggressiveness of the isolates was higher in August than in October assay. The accessions BR-2399 and BR-1449 as well as the commercial cultivar Santa Cruz 47 were the most promising accessions displaying higher levels of stable resistance against the two Brazilian FOV isolates.Okra is a vegetable crop with high nutritional value, being a rich source of vitamins and mineral salts. Fusarium wilt (caused by Fusarium oxysporum f. sp. vasinfectum – FOV) is one of the major field diseases of okra in tropical areas. In the present work, 53 okra accessions and the commercial cultivar Santa Cruz 47 were evaluated aiming to identify sources of resistance to Brazilian FOV isolates and to study the resistance stability of the selected accessions to different pathogen isolates. An initial screening was carried out with only one FOV isolate (Fus-194). In the second assay, thirty-three accessions identified in the first screening were re-evaluated in two assays (in two different seasons) using two FOV isolates (Fus-194 and Fus-201). The resistance evaluation was carried out with 21 day-old plantlets, using the root-dipping inoculation procedure, utilizing a spore suspension of 1x106 conidia/mL. The evaluation was done using a disease severity grade system with grades ranging from 0 to 4. These grades were used to generate a disease index that was employed for clustering the accessions according to their reaction to FOV. In the evaluation carried out in August (average temperature of 19,8°C) only 12 out the 32 accessions (i.e. 37%) were rated as having high to intermediate resistant response to Fus-194 isolate. Only 28% of the accessions were classified within the high to intermediate resistance cluster when using the Fus-201 isolate. In the assay carried out in October (higher temperatures) 72% of the accessions were classified as resistant and intermediate resistant to Fus-194 isolate, whereas 32% were resistant to isolate Fus-201. Our results indicated that the Fus-201 isolate was more aggressive than Fus-194. Comparative analysis of the assays indicated that the overall aggressiveness of the isolates was higher in August than in October assay. The accessions BR-2399 and BR-1449 as well as the commercial cultivar Santa Cruz 47 were the most promising accessions displaying higher levels of stable resistance against the two Brazilian FOV isolates. / O quiabo é uma hortaliça de alto valor alimentício e uma importante fonte de vitaminas e sais minerais. Dentre os problemas fitossanitários que ocorrem no quiabeiro, a murcha-de-fusário, causada pelo fungo Fusarium oxysporum f. sp. vasinfectum (FOV) é uma das mais importantes por proporcionar grandes perdas na produção. Este trabalho objetivou avaliar, em condições de casa de vegetação, a cultivar comercial Santa Cruz 47 e uma coleção de 53 acessos de quiabeiro, visando identificar fontes de resistência a FOV e analisar a estabilidade da resistência de acessos promissores a diferentes isolados do patógeno. Inicialmente foi conduzida uma triagem utilizando um único isolado de FOV (Fus-194). Em uma segunda etapa, trinta e dois acessos obtidos na primeira etapa, foram reavaliados para resistência ao FOV utilizando dois isolados (Fus-194 e Fus-201), em duas épocas do ano. A inoculação foi realizada em mudas com 21 dias após semeadura, pela imersão das raízes em suspensão de conídios (1x106 conídios/mL) do patógeno. A avaliação da severidade da doença foi realizada usando uma escala de notas, variando de 0 a 4. As notas foram transformadas em índice de doença (ID) e agrupadas em classes de acordo com a reação da doença. Na segunda etapa do trabalho realizado no mês de agosto de 2010 (temperatura média de 19,8°C), dos 32 acessos avaliados, 12 (37%) foram considerados altamente resistentes a medianamente resistentes ao Fus-194. Para o isolado Fus-201 apenas 28% foram classificados dentro dessas duas classes. No ensaio realizado no mês de outubro (temperatura média de 25°C) referente à segunda etapa desse trabalho, 72% dos acessos foram considerados medianamente a altamente resistentes ao Fus-194. Neste ensaio, 32% dos acessos foram resistentes ao isolado Fus-201. Os resultados demonstraram uma maior agressividade do isolado Fus-201 em relação ao isolado Fus-194. Os níveis de agressividade dos isolados testados foram maiores no ensaio realizado no mês de agosto do que em outubro. Os acessos BR-2399, BR-1449 e a cultivar Santa Cruz 47 apresentaram resistência estável aos dois isolados do patógeno tanto na etapa preliminar, quanto nas duas épocas de avaliação da segunda etapa do trabalho.

Submitted by (lucia.rodrigues@ufrpe.br) on 2017-03-14T13:17:49Z No. of bitstreams: 1 Josiene Silva Veloso.pdf: 976122 bytes, checksum: 7669da38d458979fe12860f67c3c1a9c (MD5) / Made available in DSpace on 2017-03-14T13:17:49Z (GMT). No. of bitstreams: 1 Josiene Silva Veloso.pdf: 976122 bytes, checksum: 7669da38d458979fe12860f67c3c1a9c (MD5) Previous issue date: 2013-02-18 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / The cowpea (Vigna unguiculata) is grown predominantly in the North and Northeast of brazil, with low yields due to a variety of factors, including the fusarium wilt, caused by Fusarium oxysporum f. sp. tracheiphilum. The present study aimed to characterize isolates of F. oxysporum associated with wilt in cowpea through observations of morphological characters, pathogenicity test and analysis of sequences of the translation elongation factor 1-α (tef1) gene. The colony color of the isolates varied from white to violet with dense aerial mycelium. The microconidia were oval to ellipsoid, slightly curved and unicelular, arranged in false heads formed on short monophyalides. The macroconidiaformed on sporodochia were falcate, slightly curved with three to five septa. Of the 27 isolates of F. oxysporum analyzed 20 were pathogenic to cowpea, causing disease with severity ranging from 1 to 99%. The neighbor-joining analysis based on tef1 grouped the isolates into six haplotypes that were not correlated with pathogenicity.. It is suggested from the results of the present study that f. sp. tracheiphilum represents at least three phylogenetic lineages behaving as polyphyletic and paraphyletic, indicating they have independent evolutionary origins. / O Feijão-caupi é uma leguminosa cultivada predominantemente nas regiões Norte e Nordeste do brazil , apresentando baixo rendimento devido a uma diversidade de fatores, dentre eles a murcha-de-fusário, causada pelo fungo Fusarium oxysporum f. sp. tracheiphilum. O presente estudo teve como objetivo caracterizar isolados de F. oxysporum associados à murcha em feijão-caupi por meio de observações de caracteres morfológicos, teste de patogenicidade e análise de sequencias do gene fator de elongação 1-α (tef1). Os isolados apresentaram coloração variando do branco ao violeta com micélio aéreo bastante denso. Os microconídios apresentaram formato oval a elipsóide, ligeiramente curvado e sem septo, dispostos em falsas cabeças produzidos em monofiálides curtas. Os macroconídios formados em esporodóquios apresentaram formato falcado, ligeiramente curvado com três a cinco septos. Dos 27 isolados de F. oxysporum analisados 20 se mostraram patogênicos ao feijão-caupi, causando doença com severidade variando de 1 a 99%. A análise de neighbor-joining baseada na região tef1 agrupou os isolados em seis haplótipos independentemente de serem ou não patogênicos. Pode-se sugerir a partir dos resultados obtidos no presente estudo que a f. sp. tracheiphilum representa pelo menos três linhagens filogenéticas comportando-se como polifilética e parafilética, indicando assim terem origens evolucionárias independentes.

Submitted by Marco Antônio de Ramos Chagas (mchagas@ufv.br) on 2017-04-27T17:21:31Z No. of bitstreams: 1 texto completo.pdf: 301848 bytes, checksum: 965b4a5e98b835e25f72696745c97da7 (MD5) / Made available in DSpace on 2017-04-27T17:21:31Z (GMT). No. of bitstreams: 1 texto completo.pdf: 301848 bytes, checksum: 965b4a5e98b835e25f72696745c97da7 (MD5) Previous issue date: 2004-04-27 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / O presente trabalho objetivou investigar o potencial de uso do controle biológico no patossistema F. oxysporum f. sp. phaseoli e feijão a partir das seguintes etapas: 1-Selecionar um isolado de F. oxysporum não-patogênico (FO), um isolado de Trichoderma sp. (TR) e outro de Pseudomonas fluorescentes (PF) mais competentes e com alta competitividade rizosférica. 2-Estudar o efeito do tratamento de sementes de feijão com isolados selecionados de Pseudomonas fluorescentes, no desenvolvimento da murcha-vascular. 3-Estudar a relação entre a densidade de inóculo de F. oxysporum f. sp. phaseoli e de F. oxysporum não- patogênicos adicionada no solo para supressão da forma patogênica. 4-Estudar, em solo infestado com F. oxysporum f. sp. phaseoli, o efeito combinado de sementes de feijão tratadas com a bactéria e plantadas em solo infestado com F. oxysporum não-patogênico em casa de vegetação, no desenvolvimento da murcha-vascular. 5-Estudar, em solo infestado com F. oxysporum f. sp. phaseoli, o efeito combinado de sementes de feijão tratadas ou não-tratadas com a bactéria e plantadas em solo infestado com F. oxysporum não-patogênico ou Trichoderma sp. e o efeito de cada potencial antagonista sozinho, no campo, no desenvolvimento da murcha-vascular. Foram testados três isolados de FO, 11 de TR e 150 de PF, obtidos da rizosfera (80), rizoplano (30) e internamente as raízes (40). Nos testes de antibiose “in vitro” foram selecionados 22 isolados de PF e nos testes posteriores de competência e competitividade selecionou-se o isolado PF22 (Pseudomonas putida). Testando-se a competência e competitividade de FO e TR foram selecionados os isolados TR11 de Trichoderma (T harzianum) e FO3 de Fusarium não patogênico. Observou-se que não teve diferença entre as concentrações de FO para um mesmo isolado, no entanto, foi observada diferença entre os isolados e destes com o controle. O isolado FO3 teve uma maior área abaixo da curva de decréscimo da clorofila (AACDCL) para as clorofilas A, B e total (5,38, 1,71 e 7,10, respectivamente), menor severidade da doença (2,37) e maior produção de grãos (6,65g) em relação ao FOPm sozinho, com AACDCL de 2,41, 0,87 e 3,25, respectivamente, severidade 4,25 e produção de grãos de 2,23g. O isolado PF22 sozinho não reduziu a severidade da doença. Quando FO3 e PF22 foram usados em conjunto obteve-se uma maior AACDCL paras as clorofilas A, B e total (4,52, 1,05 e 5,57, respectivamente), maior produção de grãos (12,59g) e redução da severidade da doença (2,50) enquanto que FOPm sozinho teve AACDCL de 0,92, 0,23 e 1,16, respectivamente, severidade de 4,30 e produção de 5,71g de grãos. No campo, os isolados TR11 e FO3 sozinhos ou em conjunto com PF22, não diferiram entre si quanto a AACDCL, mas diferiram do FOPm sozinho. Quanto à produção, observou-se que a associação dos microrganismos FO3 + PF22 e TR11 + PF22 não diferiram entre si, com uma produção de 15 e 14 vagens e 14,7 e 13,1 g de grãos, respectivamente. A produção do tratamento apenas com FOPm foi de 8,3 vagens e 4,5 g de grãos. Esses valores mostram um aumento de aproximadamente 200% na produção de grãos quando utilizados os microrganismos em conjunto no controle da doença no campo. A ação conjunta dos microrganismos usados neste trabalho mostrou-se uma estratégia de controle biológico promissora para a redução no solo da densidade de inóculo de F. oxysporum f. sp. phaseoli e, com isso, propiciar uma redução da ocorrência e dos danos causados por essa doença. / This study aimed to investigate the potential of the biological control upon the pathosystem F. oxysporum f. sp. phaseoli and bean, by the following procedure: 1) Selecting an isolate of non-pathogenic F. oxysporum (FO), an isolate of Trichoderma sp. (TR) and one of fluorescent Pseudomonas (FP), which should be more competent and have a higher degree of rhizosphere competitiveness; 2) Studying the effect of the seed treatment with those selected fluorescent Pseudomonas isolates upon the development of the vascular wilt; 3) Studying the relationship between the inoculum densities of the non-pathogenic F. oxysporum and pathogenic F. oxysporum f. sp. phaseoli added to the soil in order to suppress the pathogenic form. 4) Studying the combined effect of bean seed treatment with F. oxysporum f. sp. phaseoli and sowing in a non-pathogenic F. oxysporum-infested soil upon the development of the vascular wilt, under greenhouse conditions. 5) Comparing the combined effect of bean seed treatment with F. oxysporum f. sp. phaseoli or no treatment followed by sowing in soil infested by either non-pathogenic F. oxysporum or Trichoderma sp., as well as the effect of each antagonistist alone upon the development of the vascular wilt under field conditions. The following isolates were tested: three from FO, 11 from TR and 150 from PF [which were obtained from the rhizosphere (80), rhizoplane (30) and internally from the roots (40)]. In the in vitro antibiosis test, twenty two PF isolates were selected, whereas in the following tests for competence and competitiveness the isolate PF22 (Pseudomonas putida) was selected. When testing FO and TR for competence and competitiveness, the isolates TR11 from Trichodema (T. harzianum) and FO3 from the non-pathogenic Fusarium were selected. No differences were observed among the FO concentrations for the same isolate, but differences between the isolates as well as between these and the control were significant. The FO3 isolate occupied a higher area below the chlorophyll decreasing curve (AACDCL) for A, B and total chlorophylls (5.38, 1.71 and 7.10, respectively), as well as a lower disease severity (2.37) and higher bean yields (6.65g) in relation to FOPm alone, with the respective values of 2.41, 0,87 and 3,25 for AACDCL, 4.25 severity, and 2.23g for bean yield. The isolate PF22 did not reduce by itself the severity of the disease. When using FO3 and PF22 together, a higher AACDCL was obtained for the chlorophylls A, B and total (4.52, 1.05 and 5.57, respectively), as well as higher bean yields (12.59g) and reduced severity of the disease (2.50), whereas FOPm alone showed an AACDCL of 0.92, 0.23 and 1.16, respectively, 4.30 severity, and bean yield of 5.71g. Under field conditions, both the isolates TR11 and FO3 by themselves or combined with PF did not differ from each other for AACDCL, but from the FOPm alone. In relation to the yield, no differences were found for the combination of the microorganisms FO3 + PF22 and TR11 + PF22, since a total of 15 and 14 pods and 14.7 and 13.1 beans were yielded, respectively. In the treatment with FOPm alone, a total of 8.3 pods and 4.5 beans were yielded. These values show an increase of around 200% in bean yield, when the microorganisms were used in combination to control the disease in the field. In this study, the combined action of the microorganisms showed to be a promising biological control strategy for reducing the density of the F. oxysporum f. sp. phaseoli inoculum in the soil providing a reduction in both occurrence and damages caused by this disease. / Tese importada do Alexandria

Submitted by Reginaldo Soares de Freitas (reginaldo.freitas@ufv.br) on 2016-04-22T10:42:12Z No. of bitstreams: 1 texto completo.pdf: 2175949 bytes, checksum: cbe19c7e16e1d8e5f10097aa9663d4f9 (MD5) / Made available in DSpace on 2016-04-22T10:42:12Z (GMT). No. of bitstreams: 1 texto completo.pdf: 2175949 bytes, checksum: cbe19c7e16e1d8e5f10097aa9663d4f9 (MD5) Previous issue date: 2015-11-16 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / O melhoramento do feijoeiro para a obtenção de cultivares resistentes é a estratégia mais efetiva e econômica contra fitopatógenos, sobretudo habitantes de solo, onde o controle químico é inexistente e práticas culturais ineficientes. Os objetivos com este trabalho foram: i) estimar a capacidade geral e específica de combinação de genitores visando melhoramento do feijoeiro para resistência à murcha-de-fusarium (Fusarium oxysporum f. sp. phaseoli - Fop) com base na severidade da doença e redução do crescimento da planta, ii) estudar a herança da resistência do feijoeiro-comum à murcha-de-fusarium a partir de sete cruzamentos contrastantes e iii) transformação genética de Fop e uso desses transformantes para analisar o processo de colonização em tecidos do feijoeiro. Oito genótipos de feijoeiro grupo carioca foram combinados em esquema de dialelo parcial (3 x 5). Os genitores e 15 híbridos F1’s foram avaliados quanto a severidade da murcha-de- fusarium isolado FOP UFV 01, porcentagem de redução da altura e da massa fresca de parte aérea da planta e produtividade de grãos. A resistência do feijoeiro a Fop é controlada por poucos genes dominantes, enquanto a redução do crescimento da planta é governada por um conjunto diferente de genes, incluindo genes dominantes e recessivos. A redução do crescimento da planta, como resposta de suscetibilidade a Fop, aumentou a eficácia na seleção de genitores visando o melhoramento do feijoeiro para resistência à murcha-de-fusarium. A população segregante oriunda do cruzamento VC 25 / CVIII 8511 // VC 25 / Pérola, é promissora visando o melhoramento do feijoeiro para os caracteres produtividade de grãos, resistência à murcha-de-fusarium e redução do crescimento. Para o estudo de herança da resistência a Fop isolado FOP UFV 01, 210 plantas de cada população F2 resultante dos sete cruzamentos contrastantes do dialelo parcial foram avaliadas individualmente quanto à severidade da murcha-de-fusarium. Duas classes marcantes (resistente e suscetível) foram observadas nos parentais e gerações F1 e F2 dos cruzamentos, indicando herança oligogênica da resistência. A partir da segregação da geração F2, houve contraste entre as metodologias, porém pelos estimadores de máxima verossimilhança, metodologia mais adequada, conclui-se que a resistência é governada por um gene dominante de efeito maior com a presença de poligenes. Altas estimativas de herdabilidade indicam maiores chances de sucesso com a seleção. Por retrocruzamentos é possível à transferência do gene dominante de efeito maior das fontes de resistência deste trabalho. As fontes de resistência citadas neste trabalho direcionarão o melhoramento visando à desenvolvimento de cultivares de feijoeiro com resistência durável a Fop através da piramidação do gene dominante de efeito maior e dos poligenes. Para a análise do processo de colonização de Fop em tecidos da raiz e caule do feijoeiro, Fop raça 2 brasileira foi transformado para a expressão do gene da proteína verde fluorescente (egfp). O uso combinado de Driselase e Lyzing Enzyme foi eficiente para a protoplastização de micélio de Fop. O protocolo de transformação com base em PEG- CaCl2 foi eficiente para a obtenção de transformantes estáveis com colônias expressando o gene egfp. Aos 6 dias após a inoculação (DAI), o fungo penetrou na planta via pelos radiculares e aos 11 DAI cresceu em células parenquimáticas de maneira intercelular. Aos 19 DAI, atingiu o xilema bloqueando a passagem de água e sais minerais resultando em murcha e morte da planta aos 25 dias. / Common bean improvement for development of resistant cultivars is the most effective and economic strategy against plant pathogens, especially soil inhabitants where there is not chemical control and cultural practices is an incomplete control measure. The objectives of this study were: i) to estimate the general and specific combining ability of parents for common bean breeding for resistance to Fusarium wilt (Fusarium oxysporum f. sp. phaseoli - Fop), based on disease severity and reduced plant growth, ii) to study the inheritance of resistance to Fusarium wilt in common bean in seven contrasting crosses and iii) Fop genetic transformation and colonization in bean tissue. Eight bean genotypes of carioca group were combined in a partial diallel scheme (3 x 5). The parents and their 15 F1’s hybrids were evaluated for severity of Fusarium wilt, strain FOP UFV 01, percentage of height reduction and fresh weight of plant shoots and grain yield. Resistance to Fop in common bean is controlled by a few dominant genes, while the reduction in plant growth by a different set of genes, including dominant and recessive genes. Measuring the reduction in plant growth, a response of Fop susceptibility, increased the efficiency in parents selecting for common bean breeding for Fusarium wilt resistance. The segregating population, from the cross VC 25 / CVIII 8511 // VC 25 / Pérola, is promising for common bean breeding for the traits grain yield, resistance to Fusarium wilt and stunted growth. To study the inheritance of resistance to Fop strain FOP UFV 01, 210 plants of each F2 population of the seven contrasting partial diallel crosses were individually evaluated for severity of Fusarium wilt. Two prevalent classes (resistant and susceptible) were observed in parental, F1 and F2 generations of the crosses, indicating oligogenic inheritance of resistance. From the segregation of the F2 generation, there was contrast results between the methodologies, but by maximum likelihood estimators, most appropriate methodology, it is concluded that resistance is governed by a single dominant major gene with the presence of polygenes. High heritability estimates indicate higher chances of success with selection. By backcrossing is possible to transfer the major dominant gene of resistance sources of this work. Resistance sources cited in this paper will direct improvement in order to development of bean cultivars with durable resistance to Fop by pyramiding of the major gene and the polygenes. For following the Fop colonization process in bean root and stem tissues, Fop 2 Brazilian race was transformed for the green fluorescent protein gene (egfp) expression. The combined use of Driselase and Lyzing Enzyme was efficient for Fop mycelium protoplastization. The PEG-CaCl2 transformation protocol was efficient to obtain stable transformants with colonies expression the egfp gene. At 6 days post-inoculation (DPI), the fungus penetrated the plant via root hair and grew 11 DPI in parenchyma cells in a intercellular way. At 19 DAI, reached the xylem, blocked the passage of water and minerals resulting in wilting and plant death at 25 days.

El cafeto (Coffea arábica, L.) es uno de los principales productos de la canasta agroexportadora peruana, hecho que ha contribuido con el desarrollo económico y social del Perú. Sin embargo este se ve afectado por enfermedades fúngicas como la pudrición vascular causado por Fusarium oxysporum, reduciendo el sistema radicular en más de 90% afectando la producción y calidad. Por ello, se busca aislar cepas nativas de Bacillus y Trichoderma de rizósfera de café con capacidad antagonista frente a Fusarium oxysporum que puedan utilizarse en programa de manejo integrado de plagas del cafeto, en el distrito de Monzón, región Huánuco. Se logró aislar 30 cepas de Bacillus ssp. y 20 cepas de Trichoderma spp las mismas que fueron evaluadas por su capacidad antagonista frente al patógeno F. oxysporum, siguiendo la metodología de enfrentamiento directo dual en el medio Agar Papa Dextrosa . Se realizó pruebas bioquímicas, caracterización macroscopica y microscópica a las cepas con actividad antagonista y se identificó por claves taxonómicas. Las 9 cepas de Trichoderma (M3PI.I, B1.T, MTS.I, MT5.A, MT12.A, PLSO1, TL7.R, MT24.I, MT14.e) y 6 cepas de Bacillus (MT1.I, MT1.II, MT2.I, MT2.II, MT7, MT3P3) con actividad antagonista; siendo la cepa M3PI.I la que alcanzó 80 % de actividad de inhibición del fitopatogeno. Las cepas con mayor actividad antagonista (MT3P3, B1.T, MTS.I) fueron identificadas molecularmente como Trichoderma sp, Trichoderma asperellum, Trichoderma sp respectivamente. Las cepas aisladas de Bacillus sp y Trichoderma sp podrían considerarse como potenciales biocontroladores de pudrición vascular en cafeto representando una nueva vía de trabajo en pro de una agricultura sostenible. / Tesis

The diploma thesis concerns observing the health conditions of pea plants. The experiments were established in 2016–2017 in cooperation with AGRITEC, research, breeding and services, s.r.o. company in Šumperk. There were 9 varieties and 3 lines selected for the experimentations. The observation for each experiment was divided into two timing terms. The resistance to pea powdery mildew caused by pathogen Eryshipe pisi, was observed in greenhouse conditions. The highest level of resistance occurred line 1109/13 and variety FRANKLIN. The lowest level of resistance had variety ARVIKA. However this variety also occured to have the highest level of resistance in laboratory tests to resistance to Fusarium solani and Fusarium oxysporum. The pathogens causing the complex of root rot and black stem deseases were observed in conditions of infectious field. Variety ATLAS was infected the least.