Sorbitol, the primary photosynthate and translocated carbohydrate in apple (Malus x domestica Borkh.), is converted to fructose by SORBITOL DEHYDROGENASE (SDH; EC 1.1.1.14) which is active in apple fruit throughout fruit development. Apple fruit set and early development is very sensitive to carbohydrate availability, but details on carbohydrate metabolism during this phase are limited. The first objective of this work was to determine if SORBITOL DEHYDROGENASE, the primary enzyme responsible for metabolism of the major phloem-transported carbohydrate sorbitol, is present and active during apple fruit set and early development. The second objective of this work was to determine if SDH genes are differentially expressed and how their patterns of expression may relate to SDH activity in apple seed and cortex during early fruit development. Nine different genes encoding SDH were determined from analysis of a cDNA library and genomic-clones. Northern, Western and ELISA analyses showed that SDH transcripts and SDH protein were present in the fruit during the first 5 weeks after bloom and comprised 7 to 8 % of the total extractable protein. Whole fruit SDH activity was highest at 2 to 3 weeks after bloom in each of three cultivars, Lodi, Redchief Delicious and Fuji. Seed SDH activity was found to be much higher than cortex SDH activity per mg and g FW, and seed SDH activity contributed significantly to whole fruit SDH activity during the first five weeks of development after bloom. Five of the nine SDH genes present in apple genome were expressed in apple fruit (SDH1, SDH2, SDH3, SDH6, SDH9). Expression of SDH6 and SDH9 was seed-specific and expression of SDH2 was cortex-specific. Using 2D SDS-PAGE and Western analyses, SDH isomers with pI values 4.2, 4.8, 5.5 and 6.3 were found in seeds, and SDH isomers with pI values 5.5, 6.3, 7.3 and 8.3 were found in cortex. The present work is the first to show that SDH is differentially expressed and highly active in seed and cortex during early development. Thus, SDH during apple fruit set and early development may play a primary role in defining fruit sink activity.

The effects of AVG, an inhibitor of ethylene synthesis, in combination with MCP or heat treatment (HT) on quality traits of several apple cultivars after harvest (AH) or cold storage (ACS), and the involvement of ethylene in the regulation of SDH activity during the last weeks of fruit development were studied. AVG was applied to Royal Gala, Lodi, Senshu, Redchief Delicious and Red Fuji trees 4 weeks before normal harvest (H1). Control and AVG-treated (AVG) fruit were harvested at H1 and treated with MCP or air-heated. Fruit were ripened at room temperature (RT) AH or ACS. Some AVG fruit were harvested at H1 and 1 to 2 weeks after H1 (H2), or at H2 only. Ethylene production (EP), respiration rate (RR), firmness, starch index (SI), titratable acidity (TA), volatile production (VP) and AAT activity, among others, were measured AH and ACS. Peel and cortex of Gala were alcohol-fed and ester production quantified. EP and SDH presence and activity were measured at various harvest dates on control and AVG Lodi, Red Delicious and Fuji apples. AVG plus MCP was more effective in reducing HEP, RR, firmness and TA loss than either treatment alone; it did not provide further control on SI and did not repress Gala red skin color development more than AVG alone, though it consistently repressed VP. AVG plus HT was generally more effective than single treatments in reducing HEP, RR and firmness loss during storage. It was not different than the single treatments on TA, SI, and VP. The effect of AVG plus HT on fruit quality ACS was cultivar-dependent. AVG plus HT was not enough to maintain the quality of the earlyharvest cultivars, and it did not improve fruit quality of late-harvest cultivars. The effects of AVG plus MCP but not of AVG plus HT were evident at H2. Precursor availability was the major factor limiting VP, suggested by the low VP when RR was low, the increase in ester production in alcohol-fed samples, and the lack of correlation between AAT and ester production. SDH activity or expression was not affected by a reduction in ethylene production.

Diss. (sammanfattning) Alnarp : Sveriges lantbruksuniv. / Härtill 7 uppsatser.

Memoria para optar al Título Profesional de Ingeniero Agrónomo / Se evaluó el control de Eriosoma lanigerum Hausmann infestando manzanos (Malus domestica Borkh) var. Granny Smith plantados en maceta y mantenidos en sombreadero, con aplicaciones repetidas (1 a 3) de un detergente agrícola (TS2035) en concentraciones bajas (0,5 y 1,0 % v/v), en período estival. El diseño experimental fue completamente al azar, con tres repeticiones (=1 maceta/repetición) y 8 tratamientos, incluyendo Lorsban 75 WG (estándar recomendado contra E. lanigerum; se aplicó sólo una vez) y agua pura (testigo). Se hizo una evaluación pre-, y una evaluación post-aspersiones (0,7 y 14 días post aplicación) de los individuos vivos y muertos, estimando el porcentaje de mortalidad bajo lupa estereoscópica (20x). También se evaluó el porcentaje de hojas con síntomas de fitotoxicidad. Los resultados de mortalidad se corrigieron (respecto de la inicial), se transformaron a grados Bliss y sometieron a ANDEVA factorial, seguidos del test de Tukey (p<0,05). Se encontró que la mortalidad inicial fue baja y relativamente homogénea, sin signos de fitotoxicidad, antes de las aplicaciones. Post aplicaciones, tanto el número de aplicaciones como la concentración del detergente, fueron significativos en la mortalidad final, mientras que sólo la concentración fue significativa en la ocurrencia de fitotoxicidad en el follaje, sin interacción entre los factores para ninguna de las variables medidas.

Submitted by Mendes Eduardo (dasilva@ufba.br) on 2013-08-16T20:48:15Z No. of bitstreams: 1 output.pdf: 2244230 bytes, checksum: 22d5a6ac671939479c5bc9d185de2593 (MD5) / Approved for entry into archive by Alda Lima da Silva(sivalda@ufba.br) on 2013-08-22T23:16:20Z (GMT) No. of bitstreams: 1 output.pdf: 2244230 bytes, checksum: 22d5a6ac671939479c5bc9d185de2593 (MD5) / Made available in DSpace on 2013-08-22T23:16:20Z (GMT). No. of bitstreams: 1 output.pdf: 2244230 bytes, checksum: 22d5a6ac671939479c5bc9d185de2593 (MD5) / Capes, Bagisa / Estima-se que um terço dos recursos alimentares do ser humano depende diretamente da polinização por insetos, assim o declínio de polinizadores é uma séria ameaça aos sistemas agrícolas (KEVAN, 2004). KLEIN et al (2007) analisando dados de 200 países, selecionados por uma lista da FAO com culturas que representavam 99% da produção global de alimentos, concluíram que frutas, vegetais e produção de sementes de 87 das culturas mais importantes em termos globais dependem da polinização por animais, enquanto 28 não dependem. Esses autores observaram ainda que a maioria dos cultivos poderia sofrer perdas na produção devido a limitação de polinizadores, e dentre estes 13 dependem essencialmente dos polinizadores, 30 têm alta dependência, 27 apresentam dependência moderada e 21 têm pouca dependência. A perda na produção de frutos e de sementes tem sido claramente relacionada à escassez dos serviços de polinização, principalmente em áreas de agricultura intensiva que são geralmente isoladas de áreas naturais (RICHARDS, 2001). Por isso, o desenvolvimento de programas de manejo sustentáveis dos polinizadores e de conservação dos seus habitat naturais, é de fundamental importância para garantir esses serviços (AGUILAR, 2006). Muitos produtores obtêm os serviços de polinização em seus plantios por intermédio do aluguel de colônias de Apis mellifera (KREMEN & OSFELD, 2005), pois além dessas abelhas serem facilmente transportadas entre as plantações, elas são consideradas polinizadores eficientes de diversas culturas, dentre essas a macieira (KHAN & KHAN, 2004). A macieira (Malus domestica Borkh) é uma espécie que apresenta muitos cultivares com alto grau de incompatibilidade, sendo necessário dois ou mais cultivares que permita uma polinização cruzada eficiente (SOSTER & LATORRE, 2007). O que torna os serviços de polinização ainda mais necessários para esta cultura. Portanto o declínio da diversidade e abundância de insetos polinizadores nessa cultura é um importante fator que afeta a sua produtividade (CUTHBERTSON & BROWN, 2006). A maçã pertence à família Rosaceae e subfamília Pomoideae (SOUZA & LORENZI, 2005). A origem da maçã contemporânea é incerta, contudo evidências indicam que foi originada de terras altas entre o Mar Negro, Turquia e Índia, onde se expandiu para o ocidente estabelecendo variedades que resultaram dos vários cruzamentos feitos entre diversas espécies (HOLA, 1981 apud FREITAS, 1995). Atualmente é uma das frutas que engloba maior quantidade de variedades conhecidas, em que 3 a 4 mil variedades são cultivadas em maior ou menor escala, em diferentes partes do mundo (SILVA et al, 2002). No Brasil, o cultivo da maçã é uma atividade relativamente recente, até o inicio da década de 1970, o País importava maçãs que abasteciam o mercado nacional, porém, nos últimos anos obteve um significativo crescimento com a implantação de pomares comerciais na região do sul do país (PAGANINI et al 2004), tornando-se um importante componente da renda agrícola para o País (BRAGA et al, 2001).Atualmente, a cultura da maçã é uma atividade econômica relevante, com repercussão no cenário internacional, contribuindo com cerca de 1,5% da produção mundial, ou seja, aproximadamente 887 mil toneladas (PEREZ, 2006; www.abpm.org.br). O Brasil é praticamente auto-suficiente na produção de maçã e ainda exporta cerca de 10% da produção para mercados externos exigentes, como a Europa e os Estados Unidos (PAGANINI et al 2004). Segundo BRAGA et al (2001), de acordo com informações do Ministério de Desenvolvimento, Industria e Comércio Exterior, as exportações tem se mantido crescente nos últimos anos, atingindo em 2000, 64,5 mil toneladas, com ingresso de aproximadamente 30,8 milhões de dólares no país. Em 2004 registrou-se o recorde nas exportações, na ordem de 153.043 toneladas (FIORAVANÇO, 2009) e com ingresso de 72 milhões de dólares, representando um aumento expressivo em poucos anos e podendo ser um exemplo real da possibilidade de substituição de importações e da ampliação do mercado interno e da conquista de mercado externo por produto de qualidade e competitividade (PEREZ, 2006). Na região Nordeste, a Bahia foi o primeiro a implantar a cultura da macieira (Figura 02), pela Empresa Bagisa S/A Agropecuária e Comércio no município de Ibicoara, Chapada Diamantina (Figura 03), em 2005, tendo como interesse inicial a produção no mercado regional (www.seagri.ba.gov.br). Tendo em vista a importância da polinização nas culturas agrícolas, o presente estudo se propõe a analisar a eficiência do manejo de colmeias empregadas por produtores de maçã (Malus domestica) na região da Chapada Diamantina, Bahia, mais especificamente será avaliado o efeito da distância das colmeias instaladas na área de estudo sobre a taxa de polinização e de visitação das abelhas. Pretendemos contribuir também com informações a cerca da biologia floral, dinâmica de produção de néctar, sistemas de polinização e visitantes florais dessa cultura em áreas tropicais, tendo em vista que a maioria dos estudos foi realizada em áreas temperadas e sabemos que a biologia e fenofases das variedades desta cultura podem ser influenciadas pelo clima, e ou condições físicas como umidade e temperatura. A partir dessas informações, pretendemos ainda, avaliar o déficit dos “serviços” de polinização para essa cultura na região e o papel dos recursos florais da variedade estudada na atratividade dos polinizadores. / Salvador, Bahia

Submitted by Reginaldo Soares de Freitas (reginaldo.freitas@ufv.br) on 2016-04-28T14:37:35Z No. of bitstreams: 1 texto completo.pdf: 399247 bytes, checksum: 2bb5a04ca9827bc635208a00b51d1fe0 (MD5) / Made available in DSpace on 2016-04-28T14:37:35Z (GMT). No. of bitstreams: 1 texto completo.pdf: 399247 bytes, checksum: 2bb5a04ca9827bc635208a00b51d1fe0 (MD5) Previous issue date: 2014-12-16 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / O objetivo desse trabalho foi avaliar o efeito da aplicação do ácido naftaleno acético (ANA) e do inibidor de transporte de auxina ácido 2,3,5-triiodobenzóico (TIBA) na diferenciação e funcionalidade dos vasos do xilema, expressão de genes que codificam para proteínas de transporte de Ca no tonaplasto, atividade da pectinametilesterase (PME) e a ocorrência de “bitter pit” (BP), bem como sua influência sobre os atributos físico-químicos em maçãs ‘Braeburn’. Macieiras ‘Braeburn’ foram pulverizadas com duas concentrações de ANA (10 e 20 mg L-1) e TIBA (10 e 20 mg L-1) e como tratamento controle, água destilada. As aplicações foram realizadas semanalmente, durante o período de 50 a 106 dias após a plena floração (DAPF). Os frutos foram colhidos semanalmente, durante o período de 60 a 106 DAPF, e a cada 15 dias dos 107 aos 164 DAPF (até a colheita comercial), e avaliados quanto a funcionalidade do xilema e composição mineral. Os frutos colhidos aos 164 DAPF foram avaliados quanto à permeabilidade de membrana, nível de expressão de genes que codificam para bombas eletrogênicas e transportadores de Ca e atividade da PME. Seis repetições de 100 frutos foram armazenadas em atmosfera do ar a 1 °C e UR 90-95%, por dois meses, e avaliados quanto a coloração da epiderme, respiração, produção de etileno, pH, firmeza de polpa, sólidos solúveis, acidez titulável, Ca solúvel total e incidência e severidade de BP. A aplicação de ANA 20 mg L-1 proporcionou maior funcionalidade do xilema nos sistemas cortical primário e secundário das regiões proximal e distal dos frutos, quando comparado ao das plantas tratadas com TIBA 20 mg L-1. As concentrações de Ca total, aos 60, 89 e 164 DAPF, e de Ca solúvel total aos 164 DAPF, na polpa da região distal dos frutos, foram maiores no tratamento com ANA (10 e 20 mg L-1), comparadas com os demais tratamentos. Frutos tratados com ANA (10 e 20 mg L-1) tiveram redução de ~50% na atividade da PME, na polpa da região distal dos frutos, em relação ao tratamento com TIBA (10 e 20 mg L-1). Os tratamentos com TIBA (10 e 20 mg L-1) aumentaram a expressão de CAX2 e H+-PPase em comparação aos tratamentos com ANA (10 e 20 mg L-1). Após dois meses de armazenamento, os frutos tratados com ANA (10 e 20 mg L-1) apresentaram menores taxas respiratórias e de produção de etileno e exibiram menor coloração vermelha da epiderme no lado mais exposto a radiação solar na planta, em relação aos frutos tratados com TIBA. Plantas tratadas com ANA 20 mg L-1 apresentaram frutos com maiores valores de acidez titulável e sólidos solúveis do que nos demais tratamentos. Os frutos submetidos a aplicação de auxina, mantiveram níveis mais elevados de Ca solúvel total após o armazenamento, o que contribui para a manutenção da integridade da membrana plásmatica e redução na incidência e severidade de BP. De acordo com os resultados obtidos as auxinas contribuem para a redução da ocorrência de BP e para a manutenção da qualidade dos frutos em maçãs 'Braeburn'. / The aim of this study was to evaluate the effect of naphataleneacetic acid (NAA) and auxin inhibitor transport 2,3,5-triidobenzoic acid (TIBA) on xylem vessel differentiation and function , expression of genes encoding calcium (Ca) transport proteins into the vacuole, pectin methylesterase activity (PME) and bitter pit (BP) incidence, as well as, the influence on physical-chemical attributes in Braeburn apples.‘Braeburn’ trees were sprayed with two concentrations of NAA (10 and 20 mg L -1) and TIBA (10 and 20 mg L-1). The control treatment was sprayed with distilled water. The applications were carried out weekly from 50-106 days after full bloom (DAFB). The fruit were harvested weekly from 60-106 DAFB and each 15 days from 107-164 DAFB (comercial harvest) and were evaluated for xylem functionality and mineral composition. Fruits harvested at 164 DAFB were evaluated for membrane permeability, expression of genes encoding Ca pumps and carriers and PME activity. Six replications of 100 fruits were stored in a cold room at 1°C and RH 90-95% for two months. The fruit were evaluated for skin color, respiration, ethylene production, pH, firmness, total soluble solid, titratable acidity, total soluble Ca, BP incidence and severity. The use of 20 mg L-1 NAA increased the functionality of the xylem of proximal and distal regions as compared to 20 mg L -1 TIBA-treated fruit. Ca content at 60, 89 and 164 DAFB and the total soluble Ca at 164 DAFB in the distal flesh of the fruit, were higher in fruit treated with NAA (10 and 20 mg L-1) than in TIBA-treated fruits and water (control). NAA-treated fruit had 50% lower PME activity in the distal flesh of the fruits, compared to TIBA treatment (10 and 20 mg L-1). The TIBA-treated fruits (10 and 20 mg L-1) showed higher CAX2 and H+- PPase expression than the NAA treatment (10 and 20 mg L -1). After two months of storage, the NAA-treated fruit had lower respiratorion and ethylene production and showed lower red coloration of the sun exposed skin compared to TIBA-treated fruit. Plants treated with ANA 20 mg L-1 showed fruit with higher total soluble solids and titratable acidity. Fruit subjected to application of auxin maintained higher levels of soluble Ca after storage, which contributed to the main to membrane integrity, and reduced BP incidence and severity. According our results auxin reduces BP incidence and maintains 'Braeburn' fruit quality after harvest.

DIPLOMA THESIS Purification of phlorizin from Malus domestica Borkh.by solid-phase extraction and semi-preparative high-performance liquid chromatography Eva Prachařová Charles University, Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Botany The aim of this diploma thesis was to find the best conditions for purification of a flavonoid extract from leaves of Malus domestica Borkh., and obtaining the purest fraction of phlorizin. Phlorizin may be used in the treatment of diabetes mellitus type 2 in the future, it has the ability to reduce glycemia by reducing the absorption of glucose in the small intestine and by increasing urinary glucose excretion. The first step was to find an SPE cartridge with a suitable sorbent and a suitable eluent for solid phase extraction. The DPA-6S cartridge and 100% methanol as an eluentwerefound to be the most suitable for SPE. The next step was to find the best possible conditions for semi-preparative HPLC using an ACE 5 C18 column (5 μm, C18, 150 x 10 mm i.d., 150 mm length). The mobile phase consisted of 1% (v/v) acetic acid in water (solvent A) and ethanol 100% (v/v) (solvent B), and a linear gradient elution was used (10-100% B), 0-60 min, the flow: 1mL/min. This method resulted in the 91.05% purity of phlorizin. Keywords: phlorizin, SPE,...

Wild populations of Malus sieversii [Ldb.] M. Roem are valued genetic and watershed resources in Inner Eurasia. These populations are located in a region that has experienced rapid and on-going climatic change over the past several decades. We assess relationships between climate variables and wild apple radial growth with dendroclimatological techniques to understand the potential of a changing climate to influence apple radial growth. Ring-width chronologies spanning 48 to 129 years were developed from 12 plots in the Trans-Ili Alatau and Jungar Alatau ranges of Tian Shan Mountains, southeastern Kazakhstan. Cluster analysis of the plot-level chronologies suggests different temporal patterns of growth variability over the last century in the two mountain ranges studied. Changes in the periodicity of annual ring-width variability occurred ca. 1970 at both mountain ranges, with decadal-scale variability supplanted by quasi-biennial variation. Seascorr correlation analysis of primary and secondary weather variables identified negative growth associations with spring precipitation and positive associations with cooler fall-winter temperatures, but the relative importance of these relationships varied spatially and temporally, with a shift in the relative importance of spring precipitation ca. 1970 at Trans-Ili Alatau. Altered apple tree radial growth patterns correspond to altered climatology in the Lake Balkhash Basin driven by unprecedented intensified Arctic Oscillations after the late 1970s.

Pollination-dependent fruit trees grown in home gardens play an important role in the agricultural based economy of Central Asian countries, yet little is known about the status of pollinator communities, the cultivated plant composition or the factors that influence management practices in Kyrgyz home garden agroecosystems. As agricultural systems are human created and managed, a logical approach to their study blends anthropological and ecological methods, an ethnoecological approach. Over three years, I investigated how species richness and abundance of Hymenoptera, cultivated plants, and home garden management were related using quantitative and qualitative methods in the Issyk-kul Man and Biosphere reserve. Structured surveys were undertaken with heads of households using a random sample stratified by village. Gardens were then mapped with participation of household members to inventory edible species in gardens, most of which are pollinator-dependent, and to compare home garden diversity as reported by respondents during interviews. Apple diversity was studied to the variety level to understand respondents’ classification system in the context of in situ agrobiodiversity conservation. Household members identified 52 edible plant species when mapping the garden, compared with 32 reported when interviewed. The proportion of plant species received from others through exchange and the number of plots cultivated significantly explained the variation in edible plant diversity among gardens. Insects were sampled in gardens and orchards to determine potential pollinator community composition and the effect of different management practices on Hymenoptera richness and abundance. I collected 756 Hymenoptera individuals (56 bee; 12 wasp species); 12 species were new records for Kyrgyzstan or within Kyrgyzstan. Economic pressures to intensify cultivation could impact management practices that currently promote diversity. A home garden development initiative was undertaken to study management practice improvement. Participants in the initiative had higher adoption rates than controls of management practices that improve long-term yield, ecological sustainability and stability of home gardens. Home gardens, as currently managed, support abundant and diverse pollinator communities and have high cultivated plant diversity with few differences in community composition between garden management types.

Philosophiae Doctor - PhD / Apple scab, powdery mildew and woolly apple aphid are a major concern for apple breeders and producers. Control of these diseases is a significant economic and marketing priority for the South African apple industry. Application of chemicals and orchard management practices are the main methods for controlling these diseases. These diseases require an average of 15 chemical sprays per season, which leads to increased production costs for the farmer. The increased cost of chemical based control programs and demand from consumers for ‘organic apples’ grown with very little to no chemical sprays makes it important to breed for commercial apple cultivars with endogenous disease resistance genes (R-genes). The use of genetic tools (apple genetic linkage maps and the apple genome sequence) to track and introgress endogenous R-genes in breeding and to confer durable disease resistance in commercial apple cultivars will lead to a more cost effective means of disease control for apple producers. Historically, most breeding programmes rely on recurrent conventional breeding systems. This involves the crossing of apple selections showing resistance to a given disease with a susceptible elite variety. This is followed by phenotyping the progeny to identify trees exhibiting segregating field resistance. Several crosses and backcrossing are required to produce resistant varieties and to fix the resistance trait using this breeding strategy. This breeding technique is time consuming, especially in perennial tree species such as apples, which have a long juvenile period. Molecular markers have enabled the building of genetic maps, which has allowed for tracking of the inheritance of genes contributing towards the observed resistances. This has given breeders the opportunity to start the implementation of marker-assisted-breeding (MAB) and marker-assistedselection (MAS). MAB and MAS greatly reduce the time required to select for favourable genotypes, given that MAB facilitates efficient selection for inherited traits at the seedling stage. With the publication of the apple genome sequence, the identification of the genes involved in disease resistances has been made possible and this will allow researchers to venture into cisgenics for apples, which will further reduce the time required for the introgression of desirable genes into commercial cultivars. The main thrust of this research was to generate dense genetic linkage maps for three mapping populations segregating for apple scab, woolly apple aphid and powdery mildew resistance. The three mapping populations are ‘Mildew Resistant’ x ‘Golden Delicious’, ‘Russian Seedling’ x ‘Golden Delicious’ and Malus platycarpa x ‘Mildew Resistant’ and are Malus full-sib outbreed mapping populations. The generation of the genetic maps was for use in the subsequent identification candidate disease resistance QTLs/genes that can be implemented in apple cisgenics. Integrated genetic maps using SSRs, DArTs and SNP marker data were generated for all the three crosses. The integrated map of ‘Mildew Resistance’ x ‘Golden Delicious’ consists of 1, 563 markers with a total map length of 1, 298.8 cM. The ‘Russian Seedling’ x ‘Golden Delicious’ genetic map is composed of 979 markers with a total map length of 1, 729.9 cM. The Malus platycarpa x ‘Mildew Resistant’ integrated map has 616 markers and a total map length of 1,324.3 cM. Due to the fragmentation of some of the linkage groups in the ‘Russian Seedling’ x ‘Golden Delicious’ and in the Malus platycarpa x ‘Mildew Resistant’ genetic maps, a phylogenetic analysis was performed to evaluate the genetic distances between the parents of the crosses in order to understand the cause of the fragmentation of these two integrated genetic maps. QTLs were detected through the statistical correlation of the phenotypic and map data using restricted Multiple QTL Mapping (rMQM) from MapQTL® 6.0. The genome-wide LOD score minimum QTL detection thresholds were determined using 10 000 permutations for each population. The minimum QTL detection threshold for accepting a putative QTL was then determined to be 4.5 for ‘Mildew Resistant’ x ‘Golden Delicious’ and 4.6 for both the ‘Malus platycarpa’ x ‘Mildew Resistant’ and ‘Russian Seedling’ x ‘Golden Delicious’ mapping populations. A total of 17 putative QTLs were detected for the ‘Mildew Resistant’ x ‘Golden Delicious’ population, 10 putative QTLs for the Malus platycarpa x ‘Mildew Resistant’ population and nine putative QTLs for the ‘Russian Seedling’ x ‘Golden Delicious’ population were detected for the three diseases under study. The two putative QTLs for apple scab resistance detected on LG 02 of the ‘Russian Seedling’ x ‘Golden Delicious’ map coincided with the loci previously identified as encoding two apple scab resistance genes Vh2 and Vh4 on ‘Russian apple’. The QTL for apple scab resistance identified on the proximal QTL of LG 02 co-localized with SNP marker R_8936738_Lg2 on the loci where Vh4 was previously identified. The distal QTL on LG 02 shown to encode the Vh2 R-gene was linked with the SNP marker R_32981524_Lg2. With ‘Russian apple’ being known to carry a natural pyramid of R-genes for apple scab on LG 02, therefore, the ‘Russian Seedling’ used in this study was screened by a set of 14 SSR markers to determine if it was related to ‘Russian apple. The 14 SSRs produced identical alleles to those amplified by ‘Russian apple’, which means “Russian Seedling’ and ‘Russian apple’ are closely related or identical. The LG 02 pseudo-chromosome sequence was extracted from the NCBI database housing the apple genome sequence and was then used to mine for the putative R-genes within the two QTL regions. The region corresponding to the Vh2 loci, which was roughly a 600 kb region, had two clusters of ABC (PDR) disease resistance related genes. These were predicted using a full Pfam domain search and were only detected on the negative strand. The 60 kb region corresponding to the Vh4 loci comprised a cluster of LRR domains that were also detected on the negative strand using a full Pfam domain search. This 60 kb region was further analysed using Phytozome and Genome Database for Rosaceae (GDR) leading to two candidate disease resistance genes being identified. Ten consensus gene sequences were present within the 60 kb region, with only two transcripts MDP0000657246 and MDP0000128458 identified as being disease resistance related genes. The MDP0000657246 was identified on the contig MDC000294 of the Malus x domestica reference genome as being a Leucine Rich Repeat protein kinase family, which is one of the most abundant disease resistance family mainly involved in the gene-for-gene resistance mechanism. The MDP0000128458 locus was identified on contig MDC015161 as being a Ser/Thr phosphatase 7. The Ser/Thr phosphatase genes have been associated with the regulation of MAP kinase cascades that have been shown to induce the hypersensitive response (HR) in tobacco. Therefore these two genes are likely to be the loci associated with the hypersensitive response associated with the infection of apples with race 4 of apple scab, carrying the Vh4 apple scab resistance gene. Recurrent putative QTLs were detected that still need to be validated in order to be used for MAB. The ‘Russian Seedling’ x ‘Golden Delicious’ cross produced a single powdery mildew resistance QTL located on LG08 and conferring a 1:1 resistance to susceptible phenotypic segregation ratio. These results indicate that the source of the resistance thus was a single dominant resistance gene. The ‘Mildew Resistant’ x ‘Golden Delicious’ mapping population also showed two stable QTLs one for powdery mildew on LG 03, which co-segregated with SNP GD_LG03snp00866 and in addition SNP R_13071892_Lg10 was also identified to be co-segregating with the QTL for apple scab resistance on LG10. However, none of these recurrent QTLs co-localized with known genes or QTLs. For the phylogenetic analysis, re-sequenced data using the Illumina® sequencing technologies and the apple SNP chip data for ‘Russian Seedling’, ‘Mildew Resistant’, Malus platycarpa, a Chinese accession of Malus sieversii and ‘Anna’ where used to infer relatedness of the five genotypes. The Chinese accession of Malus sieversii was included in the analysis since ‘Russian Seedling’ was thought to be relatively close genetically. Whilst ‘Anna’ is known to be a low chilling cultivar of Malus x domestica (Borkh) and therefore would add in the phylogenetic placement of ‘Mildew Resistant’ and Malus platycarpa. These were sequenced to coverage of approximately 60X for ‘Russian Seedling’ and 6X for the other four genotypes. The sequence data was aligned to the reference Malus x domestica cv Golden Delicious mitochondrial genome sequence. Phylogenetic analysis was then performed using both the data from the apple SNP-chip and the aligned mitochondrial genomes. The results from both sets of data supported the putative evolutionary distances between the five genotypes. ‘Russian Seedling’ and M. sieversii were closely related, while both were genetically divergent from the closely related ‘Anna’ and ‘Golden Delicious’ commercial cultivars. This analysis however indicated that ‘Mildew Resistant’ was relatively closely related to ‘Golden Delicious’ and hence the low number of markers showing segregation distortions for the ‘Mildew Resistant’ x ‘Golden Delicious’ population in the 17 LGs of the integrated map. However, the other two mapping population exhibited a high number of markers with segregation distortions. Markers which are closely associated with disease resistance to apple scab powdery mildew and woolly apple aphid resistance will play a major role in the identification of the genes responsible for the resistances being observed. The identification of the two candidate genes for the Vh4 gene associated with apple scab resistance will be the platform from which a cisgenic programme can be implemented in the South African apple breeding program.