Tese (doutorado)—Universidade de Brasília, Instituto de Ciências Biológicas, Departamento de Fitopatologia, Programa de Pós-Graduação em Fitopatologia, 2016. / Submitted by Albânia Cézar de Melo (albania@bce.unb.br) on 2017-01-05T14:35:38Z No. of bitstreams: 1 2016_MaurícioRossato.pdf: 3410176 bytes, checksum: deb7c72c5b63bb15ba465dfeee816c69 (MD5) / Approved for entry into archive by Raquel Viana(raquelviana@bce.unb.br) on 2017-02-08T21:24:38Z (GMT) No. of bitstreams: 1 2016_MaurícioRossato.pdf: 3410176 bytes, checksum: deb7c72c5b63bb15ba465dfeee816c69 (MD5) / Made available in DSpace on 2017-02-08T21:24:38Z (GMT). No. of bitstreams: 1 2016_MaurícioRossato.pdf: 3410176 bytes, checksum: deb7c72c5b63bb15ba465dfeee816c69 (MD5) / A murcha bacteriana, atualmente reconhecida como sendo causada por um complexo de espécies do gênero Ralstonia (Ralstonia solanacearum, R. pseudosolanacearum e R. syzygii) apresenta uma ampla diversidade, podendo ser dividida em raças, biovares, filotipos e sequevares. O objetivo deste trabalho foi: (i) ampliar os conhecimentos sobre o importante complexo de espécies de Ralstonia por meio da caracterização biológica, bioquímica e molecular do isolado CNPH-RS 488, que se mostrou capaz de suplantar a resistência da linhagem ‘Hawaii 7996’; (ii) gerar um novo protocolo de identificação de biovares por meio de técnicas moleculares, visando aumentar a eficiência do processo e reduzir os custos associados com o teste de biovar; (iii) identificar novas fontes de resistência em tomateiro à murcha bacteriana em germoplasma selvagem de Solanum, considerando o melhoramento antecipatório contra possíveis novos variantes do patógeno, como o isolado CNPH-RS 488; (iv) gerar informações sobre o patossistema Coffea – Ralstonia. No presente estudo, o isolado CNPH-RS 488 foi classificado quanto ao biovar e filotipo, e foram caracterizados o seu perfil de virulência, a capacidade de suplantar de resistência da linhagem ‘Hawaii 7996’, a produção de EPS e de biofilme, bem como a sensibilidade a antibióticos e sua gama de hospedeiras. Os sistemas de marcadores RAPD/SCAR em associação com a estratégia de bulk segregant analysis foram inicialmente empregados visando o desenvolvimento de primers específicos para identificação de biovares. Para a busca por novas fontes de resistência avaliou-se uma coleção de 75 genótipos de Solanum peruvianum em comparação com as linhagens controle ‘Hawaii 7996’ (resistente) e ‘L390’ (suscetível). Os genótipos foram inoculados com os isolados CNPH-RS 488 (biovar 2A) e CNPH-RS 489 (biovar 1). O status do cafeeiro como hospedeira do complexo de espécies de Ralstonia foi demonstrado em bioensaios com nove isolados em três cultivares de Coffea arabica, comparando com a resposta do tomateiro suscetível ‘San Vito’. A confirmação da capacidade de suplantação da resistência do ‘Hawaii 7996’ pelo CNPH-RS 488 não pôde ser explicada pelos testes de produção de exopolissacarídeo e biofilme. Foi também constatado que o isolado apresenta uma maior virulência sobre espécies mais resistentes como jiló e berinjela. Onze primers RAPD foram selecionados com potencial valor diagnóstico para identificação de biovares. Amplicons obtidos com quatro primers que geraram marcadores estáveis foram clonados e a informação de sequência foi utilizada para o desenho dos primers do tipo SCAR. Os primers J1-B1-B e SCAR-i18-B2A apresentaram especificidade parcial para isolados de biovar 1 e 2A, respectivamente. Para a biovar 3, foi empregada uma estratégia de subtração in silico que permitiu o desenho de um par de primers específico para a região do gene polS (codificador da enzima sorbitol desidrogenase). Sete genótipos dos 75 de S. peruvianum foram considerados como promissoras fontes para o melhoramento do tomateiro visando à resistência à murcha bacteriana. O cafeeiro se mostrou suscetível exclusivamente a isolados de R. pseudosolanacearum, indicando os potenciais riscos que a murcha bacteriana pode apresentar para a cafeicultura brasileira. O presente trabalho destaca que a existência de isolados capazes de suplantar a resistência do tomateiro, como o CNPH-RS 488, e isso implica em potenciais riscos da disseminação dos mesmos pelo país, podendo inviabilizar as cultivares de tomateiro atualmente disponíveis no mercado e que usam a linhagem ‘Hawaii 7996’ como parental. Estudos adicionais sobre esse variante do patógeno serão necessários bem como o desenvolvimento de programas de melhoramento com novas fontes de resistência efetivas contra isolados com um perfil de virulência similar ao apresentado pelo CNPH-RS 488. / The bacterial wilt is currently recognized as being caused by a complex of three species: Ralstonia solanacearum, R. pseudosolanacearum, and R. syzygii. The causal agents of the bacterial wilt display a wide diversity, being classified in races, biovars, phylotypes, and sequevars. The objective of the present thesis was: (i) add new information about this important bacterial complex via biological, biochemical and molecular characterization of the isolate R. solanacearum CNPH-RS 488, which was found to be able to break down the resistance of the tomato (Solanum lycopersicum) inbred line ‘Hawaii 7996’; (ii) develop a new protocol for molecular characterization of biovars, focusing on making a faster and cheaper biovar identification test; (iii) search for new sources of resistance to bacterial wilt disease within a germplasm collection of wild Solanum (section Lycopersicon) species, using the preemptive breeding and developing new resistant cultivar to new and atypical strains like the CNPH-RS 488; (iv) study and characterize a new pathosystem: Coffea – Ralstonia. Studies with the isolate R. solanacearum CNPH-RS 488 involved different approaches: (1) determination of its biovar and phylotype; (2) determination of its virulence profile, including the capability of breaking down the resistance of ‘Hawaii 7996’; (3) production of exopolysaccharides (EPS) and biofilm; (4) antibiotic sensitivity, and (5) host range. The RAPD/SCAR marker systems in association with bulk segregant analysis was the strategy initially employed aiming to develop biovar-specific PCR-based markers. The search for new sources of resistance to bacterial wilt was carried using 75 Solanum peruvianum accessions and two standard lines ‘Hawaii 7996’ (resistant) and ‘L390’ (susceptible). Bioassays were carried out with two isolates, CNPH-RS 488 (biovar 2A) and CNPH-RS 489 (biovar 1). For the study of coffee as a host of the Ralstonia species complex, plants of three C. arabica cultivars were inoculated with a collection of nine isolates and their reactions were compared with that of the susceptible standard (tomato ‘San Vito’). The ‘Hawaii 7996’-resistant breaking ability of the isolate CNPH-RS 488 could not be explained by EPS and biofilm production. In addition, it was found that this isolate displayed a wider virulence profile being also able to infect accessions of scarlet eggplant and eggplant (which are naturally more tolerant to bacterial wilt). Eleven RAPD primers with potential diagnostic value for biovar were selected. Four stable amplicons (generated by four RAPD primers) were gel-purified and cloned. The sequence information was used for the design of SCAR-like primers. The primers J1-B1-B and SCAR-i18-B2A displayed partial specificity to isolates biovar 1 and 2A, respectively. For the biovar 3, it was employed the in silico subtraction technique, which allowed the design of a polS (sorbitol dehydrogenase) gene-specific primer. Seven out of the 75 S. peruvianum accessions were considered as promising sources for future use in tomato breeding programs to bacterial wilt resistance. Coffee accessions were susceptible exclusively to R. pseudosolanacearum isolates, indicating the potential risk of the bacterial wilt for the Brazilian coffee production. The present work emphasizes the existence of isolates (such as the CNPH-RS 488), which are able to break down the major sources of resistance available in tomato breeding programs. The present work also points out the potential risks of the spread of isolates with similar virulence profile across the country, which may preclude the employment of ‘Hawaii 7996’ as parental material. More studies on these variants will be necessary aiming to help the tomato breeding programs in the search for new sources of resistance to isolates with virulence profiles similar to CNPH-RS 488.

Submitted by Marco Antônio de Ramos Chagas (mchagas@ufv.br) on 2015-10-19T13:25:55Z No. of bitstreams: 1 texto completo.pdf: 1740275 bytes, checksum: dbf657dfff35fdf68a02c45937d7f4ba (MD5) / Made available in DSpace on 2015-10-19T13:25:55Z (GMT). No. of bitstreams: 1 texto completo.pdf: 1740275 bytes, checksum: dbf657dfff35fdf68a02c45937d7f4ba (MD5) Previous issue date: 2014-08-22 / Fundação de Amparo à Pesquisa do Estado de Minas Gerais / A murcha bacteriana, causada por Ralstonia solanacearum, causa perdas diretas e indiretas na produção de várias culturas. Apesar de o uso de variedades resistentes ser a melhor opção para o manejo da doença, nãoraro encontram-se relatos de suplantação da resistência. O primeiro relato de murcha bacteriana no Brasil é antigo, porém pouco se conhece sobre os biovares, filotipos, sequevares e variabilidade genética do patógeno. Por essa razão realizou-se o presente estudo. Isolados brasileiros de R.solanacearum foram caracterizados como biovar 1, 2 e 3, filotipo II, que está disperso por todo o país e filotipo I, que encontra-se predominantemente na região Norte. Sete sequevares foram identificados: 1, 4, 18, 27, 28, 41 e 50. Além disso, nós classificamos quatro novos sequevares no filotipo IIB como sequevar 53, 54, 55 e 56. Inicialmente, utilizou-se rep-PCR para estimar a variabilidade. Obtiveram- se 282 haplótipos. Uma possível explicação para o alto número de haplótipos é a ocorrência de recombinação. Isolados das regiões Sul, Sudeste e Central formam um grande grupo genético, enquanto aqueles das regiões Norte e Nordeste formam outro grupo. Tal fato evidencia fluxo gênico entre essas regiões, provavelmente mediado pelo transporte de tubérculos e mudas contaminadas. Detectou-se diferenciação genética entre isolados de tomate e batata coletados na região Sul-Sudeste-Central. Os mecanismos evolutivos foram parametrizados a partir de análises de genealogia de genes por processo coalescente utilizando sequências parciais de sete genes. Detectou- se evidência de subdivisão dos filotipos I, IIA e IIB, mas não subdivisão por hospedeiro. O fluxo gênico é mais intenso da região Sul para a região Norte. O filotipo II é uma linhagem ancestral que teve origem no Brasil e o filotipo I foi recentemente introduzido, possivelmente por ação antropogênica. O filotipo IIA deu origem ao filotipo IIB e essa diferenciação ocorreu possivelmente devido a fatores ecológicos que precisam ser eBacterial wilt, caused by the Ralstonia solanacearum, cause direct and indirect losses in several crops. Planting of resistant varieties is the best option for disease management, but reports of resistance breakdown are commonly found in the literature. Although many years have passed after the first report of this pathogen in Brazil, information on biovars, phylotypes, sequevars and genetic variability of the pathogen is scarce. In the present study isolates were characterized as biovar 1, 2 and 3. Phylotype II is spread throughout Brazil and phylotype I is predominantly found in the North. Seven sequevars were identified: 1, 4, 18, 27, 28, 41 and 50. Moreover, we classified four new sequevars in the phylotype IIB as sequevar 53, 54, 55 and 56. Initially, we used rep-PCR to estimate the variability of the pathogen and 282 haplotypes were obtained. The high number of haplotypes could be due to the occurrence of recombination. Isolates of the South/Southeast/Central regions formed a genetically related cluster. Isolates from the North/Northeast regions formed another group. Gene flow occurs through the transportation of contaminated tubers and seedlings. Genetic differentiation was detected among isolates from tomato and potato collected in the South/Central/Southeast regions. In addition, gene genealogies based on the coalescent process were used to infer about evolutionary mechanisms. We detected evidence of subdivision of phylotypes I, IIA and IIB, but no subdivision by hosts. The gene flow is predominantly from the southern to the northern regions. We confirmed that phylotype II is an ancestral lineage that originated in Brazil and probably phylotype I was recently introduced by anthropogenic actions. Phylotype IIA originated phylotype IIB and this difference was probably due to ecological factors that need to be studied in more detail. Mutation, migration, recombination and selection occur in the population of R. solanacearum in Brazil, however mutation is more important than recombination. We conclude that the use of resistant varieties is a major challenge in all regions and hosts. studados com mais detalhes futuramente. Detectaram-se evidências de mutação, migração, recombinação e seleção na população de R.solanacearum no Brasil. Entretanto, mutação é mais importante que recombinação. Conclui-se que a utilização de variedades resistentes será uma grande desafio em todas regiões e hospedeiros. / Bacterial wilt, caused by the Ralstonia solanacearum, cause direct and indirect losses in several crops. Planting of resistant varieties is the best option for disease management, but reports of resistance breakdown are commonly found in the literature. Although many years have passed after the first report of this pathogen in Brazil, information on biovars, phylotypes, sequevars and genetic variability of the pathogen is scarce. In the present study isolates were characterized as biovar 1, 2 and 3. Phylotype II is spread throughout Brazil and phylotype I is predominantly found in the North. Seven sequevars were identified: 1, 4, 18, 27, 28, 41 and 50. Moreover, we classified four new sequevars in the phylotype IIB as sequevar 53, 54, 55 and 56. Initially, we used rep-PCR to estimate the variability of the pathogen and 282 haplotypes were obtained. The high number of haplotypes could be due to the occurrence of recombination. Isolates of the South/Southeast/Central regions formed a genetically related cluster. Isolates from the North/Northeast regions formed another group. Gene flow occurs through the transportation of contaminated tubers and seedlings. Genetic differentiation was detected among isolates from tomato and potato collected in the South/Central/Southeast regions. In addition, gene genealogies based on the coalescent process were used to infer about evolutionary mechanisms. We detected evidence of subdivision of phylotypes I, IIA and IIB, but no subdivision by hosts. The gene flow is predominantly from the southern to the northern regions. We confirmed that phylotype II is an ancestral lineage that originated in Brazil and probably phylotype I was recently introduced by anthropogenic actions. Phylotype IIA originated phylotype IIB and this difference was probably due to ecological factors that need to be studied in more detail. Mutation, migration, recombination and selection occur in the population of R. solanacearum in Brazil, however mutation is more important than recombination. We conclude that the use of resistant varieties is a major challenge in all regions and hosts.

Ph.D. (Biochemistry) / Please refer to full text to view abstract

La bactérie phytopathogène du sol, Ralstonia solanacearum cause la maladie du flétrissement bactérien sur un grand nombre de plante hôtes. Un des déterminants clefs de son pouvoir pathogène est le système de sécrétion de type III. Celui-ci permet à la bactérie d'injecter des protéines directement dans les cellules de l'hôte. Dans ma thèse je me suis attaché à décrire et analyser finement la contribution au pouvoir pathogène de la bactérie de certains de ces substrats de l'appareil de sécrétion de type III (RipAX2, RipH2, RipH3 et RipG). Des expériences de double-hybride nous ont permis d'identifier des protéines des plantes hôtes pouvant être ciblées par ces effecteurs de type III. Dans une autre partie de mon travail j'ai contribué à l'étude de RipAX2 qui est induit spécifiquement la résistance dans des lignées d'aubergines porteur d'un locus de résistance. J'ai également travaillé sur l'identification des cellules de plantes soumises à l'injection de type III dans une interaction compatible. Pour cela j'ai utilisé la plante hôte Medicago truncatula, en exprimant dans certaines lignées cellulaires un effecteurs (RipG7) pour lequel nous avions démontré que son expression constitutive das M. truncatula pouvait restaurer l'infection de ces plantes par un mutant bactérien dans ce même effecteur. Enfin, j'ai aussi contribué à l'analyse structure-fonction fine de l'effecteur RipG7 dans sa fonction de contribution au pouvoir pathogène de R. solanacearum sur la plante hôte M. truncatula. Ce travail nous a permis d'identifier les acides aminés de RipG7 qui sont sous sélection positive, et parmi ceux-là, ceux qui contribuent directement à la fonction de RipG7 sur M. truncatula. / The soil-borne pathogen Ralstonia solanacearum causes bacterial wilt in a broad range of plants. The type III secretion system (T3SS) and its associated type III effectors (T3Es) are the main virulence determinants of R. solanacearum. In my PhD study, to understand the mode of action of several "core" effectors (RipH2, RipH3, RipG7, RipG6) from R. solanacearum in host cells. We performed yeast two-hybrid screening of plant cDNA library to identify their protein targets. Besides, we also collaborated on the identification and characterization of a specific type III effector RipAX2 which is an avirulence factor that triggers a hypersensitive response in specific Eggplant lines. To understand which plant root cells are actually subjected to type III injection during the compatible interaction, I have generated transiently transformed Medicago truncatula lines (hairy root), expressing a host specificity and core T3E RipG7 in different root cell layers. When the transformed plant expressing RipG7 under 35S promoter, the plant can be infected and colonized by the ripG7 single mutant strain. The study could be refined by using specific root cell layer promoter to identify the root cell layers that are key players in this interaction. We also worked on the characterization of the structure-function of RipG7 from R. solanacearum. Our work revealed the genetic and functional variation of RipG7. Furthermore, positive selection study and mutagenesis analysis enabled us to identify essential functional residues which likely to have been differentially selected during the host-pathogen co-evolution. The potential plant targets of RipG7 were also studies further in our study by differential yeast two-hybrid.

Les bactéries phytopathogènes produisent des effecteurs de type III qui sont des facteurs de virulence injectés dans la cellule hôte afin de moduler les défenses et ainsi favoriser l'infection. L'effecteur PopP2 de Ralstonia solanacearum possède une activité acétyl-transférase qui est perçue par la paire de protéines de résistance RRS1-R et RPS4 dans le noyau des cellules végétales. Cette étape de perception conduit à l'activation des réponses de défenses. GTE9 et GTE11, deux protéines à bromodomaine d'Arabidopsis thaliana, s'apparentent à des lecteurs épigénétiques ciblés par PopP2. Nos données démontrent que ces protéines GTEs (i) interagissent avec PopP2 dans le noyau des cellules végétales et (ii) sont acétylées en présence de l'effecteur. De plus, GTE9 et GTE11 se lient in vivo et in vitro à des histones H4, suggérant que PopP2 interfère avec des processus de remodelage de la chromatine. / Microbial pathogens infect host cells by delivering virulence factors (effectors) that interfere with defenses, thereby favouring infection. The Ralstonia solanacearum PopP2 effector displays an acetyltransferase activity perceived by the Arabidopsis immune receptor pair, RRS1-R with RPS4, in the plant nucleus. This recognition step leads to the activation of immunity. Two Arabidopsis bromodomain-containing proteins , called General Transcription factor with Extra-terminal domain 9 and 11 (GTE9 and GTE11), were identified as PopP2-interacting partners. GTE9 and GTE11 are considered as epigenetic readers. Our data demonstrate that these GTEs proteins (i) interact with PopP2 in the plant nucleus and (ii) are acetylated in presence of the effector. Moreover, bromodomains of GTE9 and GTE11 bind in vivo and in vitro to Histone H4. Together, these data suggest that PopP2, through the targeting of GTE9 and GTE11, likely interfere with chromatin remodeling processes to affect ETS and/or ETI-related processes.

Please read the abstract in the section 00front of this document / Dissertation (MSc)--University of Pretoria, 2008. / Plant Science / unrestricted

Submitted by MARCOS LEANDRO TEIXEIRA DE OLIVEIRA (marcosteixeira@ufv.br) on 2018-11-01T13:24:40Z No. of bitstreams: 1 texto completo.pdf: 1410315 bytes, checksum: 018838e11a8a7e3cca3d440cf4f025db (MD5) / Made available in DSpace on 2018-11-01T13:24:40Z (GMT). No. of bitstreams: 1 texto completo.pdf: 1410315 bytes, checksum: 018838e11a8a7e3cca3d440cf4f025db (MD5) Previous issue date: 2018-09-27 / Ralstonia spp. são conhecidas por causar murcha bacteriana em várias plantas de interesse econômico. O patógeno possui alta variabilidade genética, ampla variedade de hospedeiros e pode sobreviver no solo mesmo na ausência de hospedeiros. A compreensão das potenciais fontes de inóculo, que contribuem para a variabilidade genética no centro de origem do patógeno é interessante para o manejo da doença. O papel dos rios, plantas daninhas e da população nativa de Ralstonia spp. em áreas de vegetação natural no desenvolvimento de epidemias de murcha bacteriana é pouco compreendido. A variabilidade genética entre cepas de Ralstonia spp. em uma região onde a doença é endêmica pode elucidar a contribuição dos meios de dispersão e fatores associados à sobrevivência. No presente estudo, a detecção de Ralstonia spp. em rios de diferentes biomas do Brasil revelou o potencial destes recursos naturais para dispersar o patógeno. As plantas invasoras mostraram ser importantes reservatórios de ambas as espécies de Ralstonia que ocorrem no Brasil e colaboram para sua sobrevivência. Métodos de detecção não foram sensíveis para confirmar a presença de Ralstonia spp. em amostras de solo de áreas sem ocorrência de murcha bacteriana. Quando se analisaram 204 isolados de R. solanacearum e 60 isolados de R. pseudosolanacearum obtidos do município de Coimbra, Minas Gerais, constatou-se haver baixa variabilidade genotípica e clonalidade. Nenhuma estruturação foi observada para as regiões do município, mas a composição genotípica variou entre os anos amostrados. Para o controle alternativo da murcha bacteriana, cinco fagos pertencentes à família Siphoviridae, ordem Caudovirales, foram isolados em amostras de solo. A análise molecular e a gama de hospedeiros com diferentes isolados de Ralstonia spp., representando o Brasil, revelaram diferenças entre os vírus. Adicionalmente, houve diferenças quanto à gama de hospedeiros quando os cinco fagos foram expostos a 24 isolados de Ralstonia spp. Os fagos não foram capazes de prevenir a infecção e controlar o número de células de Ralstonia spp. no solo. Outros métodos de aplicação são necessários para avaliar a eficiência dos fagos no controle da murcha bacteriana. / Ralstonia spp. are known to cause bacterial wilt in several plants of economic interest. The pathogen has high genetic variability, wide host range and can survive in the soil even in the absence of hosts. Understanding potential inoculum sources that contribute to genetic variability in the center of origin is interesting to the management of the disease. The importance of rivers, weeds and native population of Ralstonia spp. in areas of natural vegetation in the development of epidemics of bacterial wilt is poorly understood. Genetic variability among strains of Ralstonia spp. in a local region where the disease is endemic can elucidate the contribution of the means of dispersal and factors of survival. In the present study, the detection of Ralstonia spp. was attempted in water of rivers of different biomes of Brazil and revealed the potential of these natural resources to disperse the pathogen. Weeds were important reservoirs of both species of Ralstonia that occur in Brazil, and collaborate to their survival. Methods of detection were not sensitive to confirm the presence of Ralstonia spp. in soil samples from areas without the occurrence of bacterial wilt. The genetic variability of 204 strains of R. solanacearum and 60 strains of R. pseudosolanacearum from the municipality of Coimbra, Minas Gerais, was low and there was evidence of clonality in the population. The population was not genetically structured according to the geographic region in the municipality, however the genotypic composition varied in time. To assess an alternative measure to control bacterial wilt, five phages were isolated. All phages belong to the Siphoviridae family, Caudovirales order. Molecular analysis and host range with different R. solanacearum strains revealed differences among the viruses. There were differences in the host range when the five phages were exposed to 24 Ralstonia spp. strains. The phages were not able to prevent tomato infection and control the number of cells of Ralstonia spp. in the soil. Other methods of application are necessary to evaluate the efficiency of the phages to control of bacterial wilt.

Over the past decade, the Eastern Shore of Virginia (ESV) has been implicated in at least four outbreaks of Salmonellosis associated with tomato all originating from the same strain, Salmonella enterica serovar Newport.  In addition to S. Newport contamination, the devastating plant disease, bacterial wilt, caused by the phytopathogen Ralstonia solanacearum threatens the sustainability of ESV tomato production.  Bacterial wilt is present in most ESV tomato fields and causes devastating yield losses each year.  Due to the ESV\'s endemic population of R. solanacearum and S. Newport, the relationship between the two pathogens is of interest and has never been investigated.  Two separate studies were conducted to assess the relationship between these two bacteria.  One study consisted of a series of greenhouse trials that involved root-dip inoculations of tomato plants with one of four treatments: 1) S. Newport, 2) R. solanacearum, 3) a co-inoculation of S. Newport + R. solanacearum, and 4) a control group with no inoculation. Leaf, stem, and fruit samples were collected from the plants and S. enterica presence from the internal tissues was observed.  S. enterica was recovered from a low percentage of fruit and leaf samples.  There were significantly more stem samples from plants co-inoculated with S. Newport + R. solanacearum positive for S. enterica (17.46%) than from other treatments.  Another study examined the relationship between the two bacteria via vacuum infiltration inoculations of tomato fruit collected from commercial production fields on the ESV with S. Newport.  Tomato fruit were collected from plants expressing symptoms of bacterial wilt (symptomatic) and plants not expressing bacterial wilt symptoms (asymptomatic).  After fruit infiltration with S. Newport, recovery concentration of S. enterica from internal tissues was measured.  S. enterica populations were greater in fruit originating from asymptomatic (5.15 log CFU/g) versus symptomatic (4.91 log CFU/g) plants across five studies.  Fruit collected from asymptomatic plants had a significantly higher internal pH (4.60) than fruit collected from symptomatic plants (4.37).  These results suggest that R. solanacearum can influence S. enterica survival and transportation throughout the internal tissues of tomato plants as well as the influence internal tomato fruit pH, which could potentially impact S. Newport survival in the fruit. / Master of Science in Life Sciences

In 2007 and 2008 more than 100 million dollars of fresh market tomatoes were grown in Virginia, with the majority of production occurring on the Eastern Shore of Virginia (ESV), according to the National Agricultural Statistics Service. Bacterial wilt of tomato, caused by Ralstonia solanacearum (Smith) and Yabucchi et al., is the most devastating disease of tomato on the ESV. Four 'observational trials' were conducted on the ESV over three growing seasons to determine the temporal and spatial distribution of this disease in commercial tomato fields. Plants were assessed at approximately one-week intervals throughout the growing seasons and the incidence of bacterial wilt for each individual plant was recorded. A steady increase in both disease incidence and clustered distribution of the disease within rows was observed as the growing season progressed. Positive correlations between disease incidence and percentage of rows exhibiting a significant clustered distribution occurred in all trials, which indicated an increase in clustered distribution as disease incidence increased. Research trials were conducted over three years, beginning in the summer of 2007, to investigate the effects of tomato bacterial wilt resistant cultivars on the ESV. In 2008 and 2009, the selective, systemic compound which induces host plant resistance, acibenzolar-S-methyl (ASM) was incorporated into resistant cultivar trials. Results from the 2007 trial revealed significant resistance in some of the breeding lines, CRA 66 and PI 126408. The 2008 and 2009 trials revealed that ASM was not effective at reducing levels of bacterial wilt. Grafted transplants in the spring trials of 2008 and 2009 had varied results in resistance and yield. Results revealed the tomato cultivar BHN 669 was an excellent resistant cultivar with promising yield potential and fruit quality. / Master of Science

Ralstonia solanacearum est une bactérie vasculaire d'origine tellurique dévastatrice chez les Solanacées maraîchères. L'une des difficultés pour la sélection variétale réside dans le contournement de la résistance, en raison de la grande variabilité et plasticité génotypique du pathogène. Trente accessions de tomate, aubergine et piment (Core-TEP) testées vis-à-vis de 12 souches représentatives des phylotypes I, II et III de R. solanacearum (Core-Rs2) ont été classées de très sensibles à très résistantes. Des groupes de souches définis en fonction de leur virulence et agressivité ont été nommés pathoprofils, d'une part, lorsque établis sur les 3 espèces et pathotypes, d'autre part, lorsque établis sur chacune des trois espèces. Aucune résistance universelle n'a pu être mise en évidence chez aucune des trois espèces. Le déterminisme génétique de la résistance à R. solanacearum chez l'aubergine a été étudié sur une population de lignées recombinantes F6 ainsi que sur les générations issues du croisement intraspécifique S. melongena MM738 (S) x AG91-25 (R), testées vis-à-vis de quatre souches du phylotype I. Une carte génétique composée de 119 marqueurs AFLP, SSR et SRAP, positionnés sur 18 groupes de liaisons, pour une longueur totale de 884 cM, a été construite. Les analyses génétiques montrent que la résistance aux souches CMR134, PSS366, GMI1000 est contrôlée par un même gène majeur expliquant jusqu’à 87% de la variance phénotypique. Celui-ci n'est pas détecté vis-à-vis de la souche virulente PSS4 qui contourne cette résistance. Dans ce cas, seul un QTL à effet intermédiaire expliquant jusqu’à 38 % de la variance phénotypique est trouvé sur un autre groupe de liaison. / The plant pathogenic soil-borne bacteria Ralstonia solanacearum causes heavy damage to solanaceous crops. One of the dilemmas for varietal breeding of solanaceous crops is the circumvention of the resistance to R. solanacearum due to the genotypic variability and plasticity of this pathogen. Thirty accessions of Tomato, Eggplant, and Pepper (Core-TEP), screened with a collection of 12 representatives strains of R. solanacearum phylotype I, II and III (Core-Rs2) within the species complex ranked from highly resistant to highly susceptible. The strains were grouped according to there profile of virulence on all three solanaceous, and by considering each species individually. We refer to these groups respectively, as pathoprofiles and pathotypes. No universal resistance was found for the three host species. Genetic analysis of resistance to R. solanacearum in eggplant was conducted against four strains of phylotype I. We used a population of recombinant lines F6 as well as the generations derived from the intraspecific cross between S. melongena MM738 (S) and AG91-25 (R). A genetic map was built with 119 markers a combination of AFLPs, SSRs and SRAPs positioned on 18 linkage groups, for a total length of 884 cM. The genetic analysis revealed that resistance to strains CMR134, PSS366, and GMI1000 was controlled by one single major gene, which explained up to 87% of the phenotypic variation. When the resistance against the highly virulent PSS4 strain is tested, this gene is not detected. In this case, a significant QTL with intermediate effect that explains up to 38% of phenotypic variation was found on another linkage group for this strain.