Please read the abstract in the thesis / Thesis (PhD)--University of Pretoria, 2013. / gm2013 / Biochemistry / unrestricted

Submitted by Nathália Faria da Silva (nathaliafsilva.ufv@gmail.com) on 2017-07-05T20:01:47Z No. of bitstreams: 1 texto completo.pdf: 613081 bytes, checksum: 51a4d7528d9d2e7c73d3049bdf77a998 (MD5) / Made available in DSpace on 2017-07-05T20:01:47Z (GMT). No. of bitstreams: 1 texto completo.pdf: 613081 bytes, checksum: 51a4d7528d9d2e7c73d3049bdf77a998 (MD5) Previous issue date: 2001-01-25 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / Isolinhas de feijoeiro-comum obtidas por cruzamentos em que o cultivar Rudá foi o genitor recorrente foram inoculadas com diferentes patótipos de ferrugem, antracnose e mancha-angular, analisadas com marcadores moleculares de DNA e avaliadas por meio de características quantitativas. As inoculações feitas em isolinhas RC n F 2:3 indicaram que as proporções entre as famílias heterozigotas e homozigotas para a resistência foram 2:1 em todos os cruzamentos. Para resistência à ferrugem e antracnose simultaneamente, foram selecionadas 13 isolinhas do retrocruzamento Rudá x Ouro Negro. Para resistência à antracnose, foram selecionadas, ainda, outras 16 isolinhas, sendo 10 isolinhas provenientes do retrocruzamento Rudá x TO e seis do retrocruzamento Rudá x AB 136. Cinco isolinhas homozigotas do retrocruzamento Rudá x AND 277 foram selecionadas para resistência à mancha-angular. De todas as isolinhas homozigotas, foram selecionadas aquelas geneticamente mais próximas do genitor recorrente, pela utilização de marcadores RAPD (Random Amplified Polymorphic DNA). Essas isolinhas foram utilizadas para avaliar a reação de resistência a ferrugem, antracnose e mancha-angular causadas pelos patótipos Uromyces appendiculatus, Colletotrichum lindemuthianum e Phaeoisariopsis griseola , respectivamente. A s isolinhas ON-25-99 e ON-48-99 se comportavam como resistentes a todos os patótipos de ferrugem inoculados. As isolinhas TO-41-5-6-24 e AB-74-1-13 foram resistentes a todos os patótipos de antracnose testados. A isolinha AN-7-2-9-4-6 foi resistente aos seis isolados de mancha-angular testados. As isolinhas mais próximas geneticamente do recorrente foram caracterizadas com marcadores moleculares fortemente ligados aos genes de resistência dos genitores doadores. Treze marcadores foram testados, e oito deles (OPAZ20 940 , OPB03 1800 , OPH13 490 , ScarBA08 560 , ScarF10 1050 , OPX11 630 , OPY20 830 e Satt174) podem ser potencialmente utilizados para dar continuidade ao processo de piramidização dos genes de resistência no cultivar Rudá. Os outros cinco marcadores sofreram recombinação no processo de retrocruzamento. As isolinhas foram, ainda, testadas para características quantitativas, em que foram estimados diferentes parâmetros genéticos, correlações fenotípicas, genotípicas e ambientais e efeitos diretos e indiretos de cada característica sobre a produção, mediante o uso da análise de trilha conduzido em dois ambientes diferentes. Os caracteres “número de vagens por planta” e “número de sementes por vagem” apresentaram maiores correlações genotípicas com a produção por planta. A média das características e a precisão experimental foram semelhantes nos dois ensaios. Os resultados encontrados neste trabalho são importantes para dar continuidade ao processo de piramidização de genes visando à resistência a ferrugem, antracnose e mancha- angular no cultivar Rudá atualmente sendo conduzido pelo BIOAGRO/UFV. / Common bean isolines obtained from crosses in which cultivar Rudá was the recurrent parent were inoculated with different rust, anthracnose, and angular leaf spot pathotypes, and also characterized with DNA molecular markers and evaluated for quantitative traits. The inoculations done in BC n F 2:3 isolines showed that the proportion between heterozygous and homozygous resistant plants was 2:1 for all crosses. Thirteen homozygous isolines resistant to rust and anthracnose simultaneously were selected from the cross Rudá x Ouro Negro. Sixteen isolines resistant to anthracnose were selected, 10 from the cross Rudá x TO, and six isolines from the cross Rudá x AB 136. Six homozygous isolines resistant to angular leaf spot were selected from the cross Rudá x AND 277. The isolines genetically closer to the recurrent parent were identified with the use of the random amplified DNA (RAPD) technique. These lines were tested against different pathotypes of the causing agents of rust, anthracnose and angular leaf spot, Uromyces appendiculatus, Colletotrichum lindemuthianum and Phaeoisariopsis griseola , respectively. Isolines ON-25-99 and ON-48-99 were resistant to all rust pathotypes tested. Isolines TO-41-5-6-24 and AB-74-1-13 were resistant to all anthracnose pathotypes tested. Isoline AN-7-2-9-4-6 was resistant to six angular spot pathotypes tested. The isolines which were genetically closer to the recurrent parent were characterized with molecular markers previously shown to be tightly linked to disease resistance genes in the donor parents. Thirteen markers were tested and eight of them (OPAZ20 940 , OPB03 1800 , OPH13 490 , ScarBA08 560 , ScarF10 1050 , OPX11 630 , OPY20 830 and Satt174) proved to be potentially useful in a breeding program to pyramid the resistance genes in the same cultivar. The other five markers separated from the resistance loci due to recombinations during the backcrossing process. The isolines were also tested for quantitative traits and different genetic parameters, phenotypic, genotypic and environmental correlations, and the direct effect of each characteristic on productivity by using path analysis in two different environments. The characters ‘number of pods per plant’ and ‘number of seeds per pod’ presented the highest correlations with productivity per plant. The mean values for the traits and the experimental precision were similar for both assays. Our results will be important to continue the pyramidation process of resistance genes for rust, anthracnose, and angular leaf spot in the cultivar Rudá presently being conducted at BIOAGRO/UFV.

Lumpy skin disease (LSD) is an economically important and debilitating disease of cattle caused by the lumpy skin disease virus (LSDV), a poxvirus in the genus Capripoxvirus. The disease is of economic importance to farmers in endemic regions and is a major constraint to international trade in livestock and their products. It is characterised by fever, enlargement of superficial lymph nodes, loss of weight, inappetence, salivation, lachrymation and formation of eruptive circumscribed skin lesions. The quality of meat and milk are reduced; there is infertility due to reduced sperm quality, abortions and reduced calving rates. The hides are permanently scarred, thereby reducing their quality and trade may be affected following movement restrictions from affected areas. v Lumpy skin disease has the potential to become an emerging disease because of global climate change and changes in patterns of trade in animals and animal products. The disease has become endemic in Africa except in countries like Libya, Algeria, Tunisia and Morocco, where the disease has never been reported. It has also spread to the Middle East where outbreaks were first reported in Israel (1989), Kuwait (1991), Saudi Arabia (1990) , Lebanon (1993), The United Arab Emirates (2000) and Oman (2010). In endemic areas, LSD outbreaks are common in summer. The persistence of LSDV between inter-epidemic periods has not been determined and there is no carrier state reported in either cattle or wild animals. Transmission of the disease has been associated with a high incidence of biting insects such as in wet conditions. The spread of LSD from Egypt to Israel e.g. was associated with movement of the stable fly, Stomoxys calcitrans. The virus has been recovered from S. calcitrans and Biomya fasciata, caught while feeding on infected animals and transmission by insects is suspected to be mechanical, which has been demonstrated in Aedes aegypti mosquitoes. During the 1957 outbreak of LSD in Kenya, affected animals were observed to have high tick infestations, especially of Amblyomma species. In a pilot trial in 2008 at the University of Pretoria (UP), Department of Veterinary Tropical Diseases (DVTD), Amblyomma hebraeum, Rhipicephalus appendiculatus and R. (B) decoloratus ticks were implicated in the transmission of LSDV. The overall objective of this study was to investigate the vector competence of three common sub-Saharan tick species (R. (B) decoloratus, R. appendiculatus and A. hebraeum) and their potential roles in the epidemiology of LSD. This was achieved by testing for persistence of LSDV in ticks and its subsequent transmission to recipient animals following interrupted feeding, transstadial and transovarial development of the ticks. The over-wintering of LSDV was also investigated during transstadial passage in A. hebraeum and transovarial passage in R. (B) decoloratus. During the study, seven cattle were artificially infected with LSDV to serve as source (donors) of infection to ticks. To test for mechanical / intrastadial transmission and persistence in ticks, adult ticks (A. hebraeum and R. appendiculatus) were partially fed on donor animals and then transferred to recipient animals or collected for testing. To test for transstadial transmission/passage, nymphal stages of A. hebraeum and R. appendiculatus were fed on donor animals until they engorged and dropped. Engorged nymphs were incubated to moult to adults. The emergent adults were placed on recipient animals and also tested for the virus. To test for transovarial transmission and passage R. (B) decoloratus (one- host tick) larvae were fed on donor animals until engorged adults. For R. appendiculatus and A. hebraeum (three-host ticks), adults were fed to repletion on the donor animals. Engorged females were collected and incubated to lay eggs and the eggs were allowed to hatch. The emergent larvae were placed to feed on recipient animals to test for transovarial transmission, while larvae were tested for the presence of the virus. Over-wintering of LSDV in ticks was tested by transstadial passage in A. hebraeum and transovarial passage in R. (B) decoloratus under fluctuating reduced temperatures, simulating wintery climatic conditions. Engorged A. hebraeum nymphs and R. (B) decoloratus females were infected by intracoelomic injection. The presence of the virus in LSDV- infected animals was tested by real-time PCR, virus isolation (VI), and the serum neutralisation test (SNT). Tick saliva was tested by real-time PCR and VI while ticks were tested by immunohistochemistry, transmission electron microscopy, VI and real-time PCR. Mechanical/intrastadial and transstadial transmission is reported in A. hebraeum and R. appendiculatus. Transovarial transmission was reported in A. hebraeum, R. appendiculatus and R. (B) decoloratus. The virus was demonstrated in saliva and tick organs of A. hebraeum and R. appendiculatus adults following both mechanical/intrastadial and transstadial persistence. Transovarial passage of LSDV was demonstrated in R. (B) decoloratus, R. appendiculatus and A. hebraeum larvae. The virus also persisted through cold temperature exposure during transstadial passage in A. hebraeum and transovarial passage in R. appendiculatus. This study confirms the vector competency of A. hebraeum, R. appendiculatus and R. (B) decoloratus ticks for LSDV. It also shows the potential for LSDV to over-winter in ticks and demonstrates that LSDV may persist in ticks during inter-epidemic periods. / Thesis (PhD)--University of Pretoria, 2013. / gm2014 / Veterinary Tropical Diseases / unrestricted

Thesis (MTech (Horticulture))--Cape Peninsula University of Technology, 2018. / Ticks and tick-borne diseases are important limiting factors to the attainment of sustainable animal and human health, affecting livelihood of resource poor farming communities in developing countries. Rhipicephalus appendiculatus and Amblyomma variegatum (Ixodidae) are among the most troubling tick species in Africa. While ticks can be controlled by applying chemical acaricides, these chemicals are quite expensive, especially, for small-scale famers in developing countries. Hence, the quest for alternative tick control over the years have revealed that entomopathogenic fungi (EPF) and plant extracts have huge prospects as sustainable alternatives for tick control. Beauveria bassiana (Hypocreales) is a fungal entomopathogen with the ability to colonize plants endophytically and induce secondary metabolite production in plants, and it has been found to be a potential biological control agent against a wide range of arthropods. Several plant species including plant species belonging to the family Alliaceae possess anti-tick activities (repellent and toxic); therefore, integrating the two strategies by inoculating Tulbaghia violacea with B. bassiana could enhance secondary metabolite contents in extracts obtained from the plant and increase medicinal materials. The primary purpose of the present study was to evaluate the effect of indigenous endophytic entomopathogens inoculation of T. violacea on the plant growth, tissue nutrient contents and secondary metabolites, and anti-tick activities of extracts of T. violacea. The specific objectives were: (i) to carry out a study on the effects of inoculating T. violacea with fungal strains of B. bassiana and Clonostachys rosea on plant growth and tissue nutrient contents in extracts of T. violacea with the view of selecting one of the two fungi for further investigation (ii) to compare secondary metabolite profiles of extracts obtained from plants exposed to fungus (EPF) inoculum and control treatment during cultivation, and (iii) to determine whether exposure of T. violacea to an endophytic fungus (B. bassiana) during cultivation affects the repellent and acaricidal activities of extracts of T. violacea on A. variegatum and R. appendiculatus. The study had two experiments, presented in chapters two and three. In the first experiment, eight weeks old potted seedlings of T. violacea were inoculated separately with B. bassiana (strain SM3) and C. rosea (strain SM8) conidia suspended at concentrations of 1 x 106 conidia mL-1. Plant growth parameters, such as number of leaves, plant height (aerial part), fresh weights of aerial parts, and tissue nutrient contents were assessed. Results indicated that B. bassiana induced higher growth of plants than C. rosea. Inoculation with B. bassiana did not significantly (P ˃ 0.05) influence most of the growth parameters, number of leaves, fresht weight of roots and of fresh weight aerial parts (leaves) of T. violacea assessed in the current study. However, mean plant heights and root lengths were significantly (P < 0.05) higher in favour of B. bassiana treated plants compared to those in C. rosea. Fe contents in the roots (1416.3 ± 305.10 mg/kg) were found to be positively influenced (P < 0.05) by the fungal inoculation. There was a significant difference in roots (P < 0.05) on the uptake of Mn in C. rosea treated plants (243 ± 19 mg/kg) compared to the control group (169 ± 16.37 mg/kg) and B. bassiana treated roots (161.3 ± 14.44 mg/kg). Macro nutrients up take did not differ significantly among treatments on both leaves and roots (P > 0.05).

Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Agrárias, Programa de Pós-Graduação em Recursos Genéticos Vegetais, Florianópolis, 2011 / Made available in DSpace on 2012-10-25T18:06:05Z (GMT). No. of bitstreams: 0 / O uso de indutores de resistência na agricultura apresenta-se como uma das alternativas eficientes para o controle de doenças. O presente trabalho teve por objetivo estudar a resistência induzida por sacarina ou ulvana à ferrugem (Uromyces appendiculatus) e à mancha angular (Pseudocercospora griseola) do feijão (Phaseolus vulgaris L.). Realizaram-se três experimentos em diferentes épocas sob condições de infecção natural com duas cultivares SCS 202 Guará (suscetível à ferrugem) e FT 911249 (resistente à ferrugem). Iniciaram-se as pulverizações das plantas com o primeiro trifólio expandido, em intervalos de quinze dias com água, sacarina ou ulvana. Avaliou-se a severidade final e Área Abaixo da Curva de Progresso da Doença (AACPD) da ferrugem e mancha angular e os componentes de rendimento. Para a avaliação dos efeitos na severidade e nos mecanismos de defesa envolvidos na resistência raça-específica ou induzida da ferrugem, em casa de vegetação, plantas suscetíveis e resistentes com o primeiro trifólio expandido, foram tratadas 6 dias antes da inoculação (DAI) com água e sacarina e 6 e 3 DAI com ulvana e inoculadas com U. appendiculatus. Às 24 e 48 horas após a inoculação (HAI) foram retiradas amostras para avaliação da atividade de peroxidases, germinação uredósporos, formação de apressórios e células com acúmulo de peróxido de hidrogênio. Aos 15 dias após a inoculação quantificou-se a porcentagem de área foliar afetada, o número e diâmetro de pústulas e o número de uredósporos por pústula. Pulverizações de sacarina reduziram a severidade final e a AACPD da ferrugem. Para mancha angular reduziu a AACPD no segundo experimento, e a severidade final e AACPD no terceiro experimento. A sacarina aumentou o peso de 100 sementes no terceiro experimento. A ulvana não afetou a ferrugem, mas reduziu a AACPD da mancha angular na cultivar FT 911249 no terceiro experimento. Os componentes de produção nos três experimentos não foram afetados pela ulvana. No experimento em casa de vegetação, a sacarina e ulvana reduziram a severidade da ferrugem. A ulvana elevou a atividade de POX às 24 HAI. Plantas resistentes tiveram maior atividade de POX às 48 HAI. Os tratamentos não afetaram a germinação de uredósporos, formação de apressórios, ou número de células com reação de hipersensibilidade às 24 e 48 HAI respectivamente. Em conclusão sacarina atuou como um indutor de resistência contra a ferrugem e a mancha angular e aumentou o peso de 100 sementes. Sob condição natural a ulvana não afeta a ferrugem. Em casa de vegetação a ulvana induziu resistência reduzindo a severidade da ferrugem e elevando a atividade de peroxidases.

Submitted by Cleber Casali (clebercasali@ufv.br) on 2017-06-01T18:39:15Z No. of bitstreams: 1 texto completo.pdf: 2869939 bytes, checksum: 08096f9e604c1fe651e880958476ddb4 (MD5) / Made available in DSpace on 2017-06-01T18:39:15Z (GMT). No. of bitstreams: 1 texto completo.pdf: 2869939 bytes, checksum: 08096f9e604c1fe651e880958476ddb4 (MD5) Previous issue date: 2004-08-20 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / O principal objetivo deste trabalho foi caracterizar, fenotipicamente, linhagens de feijão do cruzamento Ouro Negro x Pérola, quanto à reação a Colletotrichum lindemuthianum (raças 73, 81 e 89) e Uromyces appendiculatus (mistura das raças 15, 35, 45, 49, 50 e 59) e avaliar, em campo, o desempenho das linhagens que mostraram-se resistentes. Paralelamente, objetivou-se também avaliar a eficiência dos marcadores moleculares SCARF10 1050 (relacionado à resistência à antracnose e à ferrugem) e OPX11 550 (relacionado à resistência à ferrugem), identificados e mapeados em outras populações, com vistas à seleção assistida na população Ouro Negro x Pérola. Inicialmente, 400 linhagens (progênies F 7:8 ) do cruzamento Ouro Negro x Pérola foram inoculadas com a raça 89 de C. lindemuthianum. Dentre as linhagens resistentes, 68 com grãos de boa aceitação comercial foram inoculadas com as raças 73 e 81 de C. lindemuthianum e com a referida mistura de raças de U. appendiculatus. No experimento de campo, foram avaliadas 75 linhagens, que, sob inoculação artificial, mostraram-se resistentes à raça 89 de C. lindemuthianum. As testemunhas, constituídas pelos genitores (Pérola e Ouro Negro), o cultivar Talismã e as linhagens Vi 4899, VC4, e Rudá-Piramidado foram, também, avaliadas em campo. utilizou-se um látice quadrado triplo, com parcelas constituídas de duas linhas com 2m de comprimento cada uma. Na avaliação da eficiência dos marcadores moleculares, foram caracterizadas 150 linhagens do cruzamento Ouro Negro x Pérola, fenotipicamente, por meio de inoculação artificial de C. lindemuthianum (raça 89). Paralelamente, uma planta de cada linhagem foi genotipada, utilizando-se os marcadores SCARF10 1050 e OPX11 550 . A partir da inoculação artificial, foram identificadas 42 linhagens resistentes às raças de C. lindemuthianum e de U. appendiculatus, utilizadas no estudo. Considerando os resultados da caracterização fenotípica e da avaliação de campo, foram identificadas 10 linhagens que apresentaram grãos de bom aspecto comercial, iguais ou superiores àqueles do cultivar Pérola quanto à produtividade de grãos. Estas linhgens mostraram-se resistentes à antracnose e ferrugem, além de, moderadamente resistentes à mancha- angular. Das 150 linhagens avaliadas por meio de inoculação artificial, 61 mostraram-se resistentes às raças 73, 81 e 89 de C. lindemuthianum, sendo que, 42 destas foram resistentes a uma mistura de raças de U. appendiculatus. Vinte e cinco linhagens apresentaram as bandas correspondentes aos marcadores SCARF10 1050 e OPX11 550 . A eficiência de seleção para resistência à antracnose, com base na genotipagem de 150 linhagens utilizando o marcador SCARF10 1050 foi de 78%. Considerando que este marcador encontra-se a uma distância de 19,6 cM do gene de resistência, esperava-se uma eficiência de 72%, portanto, próxima àquela obtida no presente estudo. Para o marcador OPX11 550 , verificou-se uma eficiência de 85%. Esta maior eficiência era esperada, em razão de este marcador localizar-se mais próximo (11,6 cM) dos genes ou do bloco gênico, os quais conferem resistência à antracnose e ferrugem, presentes no cultivar Ouro Negro. Além disso, foi observado que o marcador OPX11 550 também pode ser utilizado no monitoramento do gene de resistência à antracnose. Embora estivessem a uma distância relativamente grande dos genes de resistência, esses marcadores proporcionaram boa eficiência de seleção. / The main objective of this study was to phenotypically characterize the common bean lines from the crossing Ouro Negro x Pérola for the reaction to Colletotrichum lindemuthianum (races 73, 81 and 89) and Uromyces appendiculatus (a mixture of races 15, 35, 45, 49, 50 and 59), as well as to evaluate the performance of the lines showing to be resistant ones. Another target was to evaluate the efficiency of the molecular markers SCARF10 1050 (related to the resistance against anthracnose and rust) and OPX11 550 (associated to the resistance against rust), which were identified and mapped in other populations and targeted to the assisted selection in the population Ouro Negro x Pérola. Initially, all 400 lines (progenies F 7:8 ) from the Ouro Negro x Pérola crossing were inoculated with the race 89 of C. lindemuthianum. Sixty eight lines of the resistant ones, from which the common beans are commercially well accepted, were inoculated with the races 73 and 81 from C. lindemuthianum and with the mixture of races from U. appendiculatus. In the field experiment, the evaluations were performed for 75 lines, which under artificial inoculation showed to be resistant to the race 89 of C. lindemuthianum. The controls constituted by genitors (Pérola and Ouro Negro), the Talisman cv. and the lines Vi 4899, VC4, and Rudá-Piramidado were also inoculated. The triple square lattice design was used with the plots consisting into two rows with 2m each one. When evaluating the efficiency of the molecular markers, 150 lines from the crossing Ouro Negro x Pérola were phenotypically characterized, by artificial inoculation of C. lindemuthianum (race 89). At the same time, one plant of each line was genotyped, by using the markers SCARF10 1050 and OPX11 550 . From the artificial inoculation, 42 lines which were resistant to either races of C. lindemuthianum and U. appendiculatus were identified. Taking into account the results of both the phenotypic characterization and field evaluation, ten 10 lines were identified which were provided with commercially good beans, that were equal or superior to the Pérola cv. for bean productivity. These lines showed to be resistant to either anthracnose and rust, and moderately resistant to the angular leaf spot. From those 150 lines evaluated by artificial inoculation, 61 were resistant to the races 73, 81 and 89 of C. lindemuthianum, and 42 lines from these ones showed to be resistant to the mixture of the U. appendiculatus races. Twenty-five lines exhibited the bands corresponding to the markers SCARF10 1050 and OPX 11550 . The selection efficiency for the resistance to anthracnose, based upon genotyping of 150 lines, using the marker SCARF10 1050 was 78%. By considering this marker is at 19.6 cM distance from the resistance gene, an efficiency of 72% was expected, therefore the obtained value was close to the expected one. An efficiency of 85% was found for the marker OPX11 550 . This higher efficiency was expected because this marker is located near (11.6 cM) the genes or the gene block that confers resistance to both anthracnose and rust in Ouro Negro cv. Moreover, it was shown what this marker can be made useful, in the targeted of the resistance gene to anthracnose. These markers provided good selection efficiency, although they were at a relatively long distance from the resistance genes.

Text in English / Corridor disease is an economically important tick-borne disease of cattle in southern Africa. The disease is caused by Theileria parva and transmitted by the vectors, Rhipicephalus appendiculatus and R. zambeziensis. There is currently no vaccine to protect cattle against T. parva that is permitted in South Africa. To develop recombinant anti-tick vaccines against Corridor disease, comprehensive databases of genes expressed in the tick’s salivary glands are required. Therefore, in Chapters 2 and 3, mRNA from the salivary glands of R. appendiculatus and R. zambeziensis was sequenced and assembled using next generation sequencing technologies. Respectively, 12 761 and 13 584 non-redundant protein sequences were predicted from the sialotranscriptomes of R. appendiculatus and R. zambeziensis and uploaded to public sequence domains. This greatly expanded the number of sequences available for the two vectors, which will be invaluable resources for the selection of vaccine candidates in future. Further, in Chapter 3, differential gene expression analysis in R. zambeziensis revealed dynamic expression of secretory protein transcripts during feeding, suggestive of stringent transcriptional regulation of these proteins. Knowledge of these intricate expression profiles will further assist vaccine development in future. In Chapter 4, comparative sialotranscriptomic analyses were performed between R. appendiculatus and R. zambeziensis. The ticks have previously shown varying vector competence for T. parva and this chapter presents the search for correlates of this variance. Phylogenetic analyses were performed using these and other publically available tick transcriptomes, which indicated that R. appendiculatus and R. zambeziensis are closely related but distinct species. However, significant expression differences were observed between the two ticks, specifically of genes involved in tick immunity or pathogen transmission, signifying potential bioinformatic signatures of vector competence. Furthermore, nearly four thousand putative long non-coding RNAs (lncRNAs) were predicted in each of the two ticks. A large number of these showed differential expression and suggested a potential transcriptional regulatory function of lncRNA in tick blood feeding. LncRNAs are completely unexplored in ticks. Finally, in Chapter 5, concluding remarks are given on the potential impact the R. appendiculatus and R. zambeziensis sialotranscriptomes may have on future vaccine developments and some future research endeavours are discussed. / Life and Consumer Sciences / Ph. D. (Life Sciences)

Submitted by Marco Antônio de Ramos Chagas (mchagas@ufv.br) on 2017-06-02T17:41:45Z No. of bitstreams: 1 texto completo.pdf: 461198 bytes, checksum: 1983c9ff254d751fb9f67e35fbabb1fc (MD5) / Made available in DSpace on 2017-06-02T17:41:45Z (GMT). No. of bitstreams: 1 texto completo.pdf: 461198 bytes, checksum: 1983c9ff254d751fb9f67e35fbabb1fc (MD5) Previous issue date: 2004-08-03 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / Visando piramidar genes de resistência à ferrugem (Uromyces appendiculatus), antracnose (Colletotrichum lindemuthianum) e mancha-angular (Phaeoisariopsis griseola) em cultivar de feijão do tipo carioca, foram intercruzadas quatro isolinhas de feijoeiro-comum. As isolinhas foram obtidas por meio de retrocruzamento nos quais foi utilizado o cv. Rudá como genitor recorrente. As isolinhas continham os seguintes genes de resistência: linha ON-48-99 - genes Co-10 e Ur-ON provenientes do cultivar Ouro Negro, linha AB-74-1-18 - gene Co-6 proveniente do cultivar AB-136, linha TO- 41-5-6-24 - gene Co-4 proveniente do cultivar TO e linha AND-7-2-9-7-10 - gene Phg- 1 proveniente do cultivar AND 277. Após seleção para homozigose e avaliação das características altura de plantas, número de dias para floração, número de dias para maturação, produtividade de grãos, peso de 100 sementes, número de sementes por vagens, número de vagens por plantas, e da reação de resistência aos patótipos de Uromyces appendiculatus, Colletotrichum lindemuthianum e Phaeoisariopsis griseola, as isolinhas foram intercruzadas duas a duas. Em seguida, as plantas F 1 foram intercruzadas para obter os híbridos duplos. Os marcadores moleculares SCAR-F10 1150a , SCAR-BA8 560a , OPX11 550a , OPY20 830a , OPB03 1800r , OPAZ20 940a e OPH13 490a associados aos respectivos genes de resistência Ur-ON, Co-10, Co-6, Co-4 e Phg-1 proveniente foram utilizados para identificar as plantas contendo todos os genes de interesse as quais foram autofecundadas para obtenção de sementes F 2 . As sementes F 2 foram semeada em casa de vegetação, e as plantas F 2 foram genotipadas utilizando-se de marcadores moleculares ligados aos genes de resistência. A resistência das plantas foi também confirmada por inoculação dos patógenos. Na geração F 3 foi feita nova avaliação com os marcadores moleculares, selecionando-se apenas as plantas que apresentavam todas as marcas. Este procedimento foi também utilizado na geração F 4 . No final deste processo foi obtida uma população constituída de 40 famílias F 4 denominadas genericamente por ́Rudá R`. Paralelamente, foi feito o cruzamento de ́Rudá R` com o cv. Pérola, obtendo-se 30 famílias F 4 . Sementes das famílias F 4:5 foram multiplicadas para realização de experimento a campo. As famílias F 4:6 foram testadas quanto ao seu desempenho agronômico em ensaio em condição de campo e foram selecionadas 4 e 3 linhagens F 4:7 de cada população, respectivamente. Estas famílias foram avaliadas quanto à resistência a diferentes patótipos de U. appendiculatus, C. lindemuthianum e P. griseola. As inoculações feitas em famílias F 4:7 mostraram que as famílias R-127-10-14, R-97-13-5, R-97-13-6, R-127-4-13, P-33-5-1, P-49-8-2 e P-49-2- 2 se comportaram como resistentes a todos os patótipos de U. appendiculatus e C. lindemuthianum, e a cinco dos sete patótipos de P. griseola inoculados. As melhores familias serão avaliadas em uma rede de ensaio de Valor de Cultivo e Uso (VCU) para uma possível recomendação de um novo cultivar de feijão tipo carioca. A seleção de linhagens de feijão com grãos do tipo carioca, resistentes à ferrugem, antracnose e mancha-angular confirmam o grande potencial e a importância do uso da seleção assistida por marcadores moleculares em programas de piramidação de genes de resistência. / The objective of this work was to pyramid resistance genes to rust (Uromyces appendiculatus), anthracnose (Colletotrichum lindemuthianum) and angular leaf spot (Phaeoisariopsis griseola) in the "carioca-type" common bean cultivar Rudá. Four isolines were obtained in four backcross programs and intercrossed. The isolines contained the following resistance genes: isoline ON-48-99 - genes Co-10 and Ur-ON from cultivar Ouro Negro (ON), isoline AB-74-1-18 - gene Co-6 from cultivar AB 136, isoline TO-41-5-6-24 - gene Co-4 from cultivar TO and isoline AND-7-2-9-7-10 - gene Phg-1 from cultivar AND 277. After selection for homozygosis and evaluation of different quantitative, morphologic, and molecular characteristics, and for resistance to rust, anthracnose and angular leaf spot, the isolines were intercrossed. The F 1 plants were intercrossed to obtain the double hybrid. The molecular markers SCAR-F10 1150a , SCAR-BA8 560a , OPY20 830a , OPAZ20 940a , OPH13 490a , OPX11 550a and OPB03 1800r associated to the resistance genes were used to identify the plants containing all the genes of interest which were selfed to obtain the F 2 seeds. The F 2 seeds were sown, and the corresponding plants were selected with molecular markers linked to the resistance genes and resistance was confirmed by inoculation of the pathogens. The selection based on molecular markers was repeated in the F 3 and F 4 generations, only plants containing all the markers were selected. At the end of this process a population of 40 families was obtained and designated ́Rudá R`. In a parallel procedure, ́Rudá R` were crossed with cv. Pérola. Thirty F 4 families ( ́Rudá R` x Pérola) were obtained. Seeds of the F 4:5 families were multiplied and used for agronomic evaluation in preliminary field tests. Four and three lines were selected from populations ́Rudá R` and ́Rudá R` x Pérola, respectively. These lines were tested against different pathotypes of U. appendiculatus, C. lindemuthianum and P. griseola. The inoculations done in F 4:7 lines showed that the lines R-127-10-14, R-97-13-5, R-97-13-6, R-127-4-13, P-33-5-1, P-49- 8-2 and P-49-2-2 were resistant to all pathotypes of U. appendiculatus and C. lindemuthianum, and to five of the seven pathotypes of P. griseola tested. The lines are xstill being analyzed for quantitative characteristics in field trials. The best lines will be tested in an official trial to be released as new varieties of "carioca-type" bean. The selection of bean lines with "carioca-type" grains, resistant to rust, anthracnose and angular leaf spot confirm the power and importance of the use of marker assisted selection in breeding programs aiming to pyramid disease resistance genes.

Attività antropiche e pressioni di mercato stanno rapidamente riducendo la biodiversità. Per questa ragione, conservare il patrimonio ecosistemico, tassonomico e genetico risulta fondamentale al fine di garantire potenziale adattativo alle specie, e, in ultima analisi, un futuro sostenibile per il pianeta. Al fine di minimizzare la perdita di biodiversità, numerosi metodi sono stati proposti per priorizzare ecosistemi, specie e popolazioni. Il presente lavoro di tesi fornisce in primo luogo una revisione di tali approcci, proponendo un albero decisionale volto a favorirne un corretto utilizzo. Secondariamente, la variabilità genomica neutrale del bufalo d’acqua (Bubalus bubalis L.) è investigata per mezzo di un pannello di marcatori SNP a media densità, rivelando due centri di domesticazione (India Nord-occidentale, Cina-Indocina) e possibili rotte di migrazione per gli ecotipi ‘river’ e ‘swamp’. L’adattamento locale ad East Coast Fever, patologia endemica delle popolazioni bovine in Africa Sub-sahariana, è stato inoltre studiato in bovini autoctoni Ugandesi (Bos taurus L.) combinando tecniche di modellizzazione delle nicchie ecologiche e di genomica del paesaggio. L’approccio ha portato ad indentificare PRKG1 e SLA2 come possibili geni di adattamento. I risultati sono discussi alla luce delle possibili implicazioni nella conservazione del bufalo e nella gestione delle risorse genetiche animali Ugandesi. / Biodiversity is quickly disappearing due to human impact on the biosphere, and to market pressure. Consequently, the protection of both wild and domestic species needs to become a priority in order to preserve their evolutionary potential and, ultimately, guarantee a sustainable future for coming human generations. To date, tens of methods have been proposed to prioritize biodiversity for conservation purposes. Here, an ontology for priority setting in conservation biology is provided with the aim of supporting the selection of the most opportune methodologies given specific conservation goals. Further, two case studies are presented characterizing neutral and adaptive genomic diversity in water buffalo (Bubalus bubalis L.) and indigenous Ugandan cattle (Bos taurus L.), respectively. In particular, two independent domestication centres (North-western India and Indochina) and separate migration routes are suggested for the ‘river’ and ‘swamp’ water buffalo types. In the case of indigenous Ugandan cattle, the integration of species distribution modelling and landscape genomics techniques allowed the identification of PRKG1 and SLA2 as candidate genes for local adaptation to East Coast Fever, a vector-borne disease affecting bovine populations of Sub-Saharan Africa. Results are discussed for their implications in water buffalo conservation and Ugandan cattle adaptive management.