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Dissertations |
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1
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Ricardo Gomes Tomáz
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Large-scale identification and ecological characterization of plant-nematodes in corn fields in the State of Goiás.
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Advisor : THAIS RIBEIRO SANTIAGO
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COMMITTEE MEMBERS :
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THAIS RIBEIRO SANTIAGO
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CLEBER FURLANETTO
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JUVENIL ENRIQUE CARES
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Andressa Cristina Zamboni Machado
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Data: Feb 27, 2023
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Show Abstract
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The maize (Zea mays L.) is the cereal of great importance in human and animal nutrition worldwide, however productivity in crop fields could be higher if it were not for the losses caused by phytonematodes. The control of plant parasitic nematodes is complex and the situation is even worse when little is known about the main morphological groups of phytonematodes present in a given location and the environmental modulators that positively influence the presence and abundance of these groups. With the aim of elucidating the main morphological groups of phytonematodes in the State of Goiás, as well as evaluating their incidence, abundance, density and quantifying the spatial heterogeneity these organisms at the level of region, municipality, and fields by mixed generalized linear models, a total of 354 commercial corn fields were collected in 26 municipalities in the State with different soil types, cultivation sequences, planting systems and altitude ranges. In addition, the influence of soil physicochemical components and the effect of different cultural practices on the abundance and damage threshold of the main morphological groups of phytonematodes present in the sampled areas were evaluated using logistic models. A total of 21 morphological groups of phytonematodes were detected, being the genera Aphelenchoides, Aphelenchus, Helicotylenchus, and Rotylenchulus and the Criconemaidae family present in all regions and with high abundance. The genus Helicotylenchus and Pratylenchus were the predominant groups in terms of fields, municipalities and geographic regions in the State of Goiás. It was possible to observe a greater heterogeneity in the incidence and population density of nematodes at field level, indicating that the variability of these organisms may be directly influenced by the physical-chemical characteristics of the soil and specific cultural practices in each area. The soil types, cultivation sequence, planting type and altitude had little influence on the population of the main morphological groups of phytonematodes. The pH, sand and clay variables expressed a greater effect on the incidence and ability of nematodes to reach the damage threshold in the crop field. This study contributed to the understanding of phytonematodes present in corn fields, the behavior and development of these organisms under the effect of different environmental modulating variables in the state of Goiás.
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2
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MARCOS SILVA DE QUEIROZ FERREIRA
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Diversity of Sida micrantha mosaic virus isolates and characterization of new begomoviruses in Malvaceae in Brazil
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Advisor : RITA DE CASSIA PEREIRA CARVALHO
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COMMITTEE MEMBERS :
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CLEBER FURLANETTO
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HELSON MARIO MARTINS DO VALE
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MIRTES FREITAS LIMA
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RITA DE CASSIA PEREIRA CARVALHO
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Data: Feb 28, 2023
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Show Abstract
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Begomovirus (family Geminiviridae) corresponds to the largest genus of plant viruses, encompassing viral species that have one or two single-stranded circular DNA molecules, separately encapsidated in 18-30 nanometer particles. Isolates with a bipartite genome (with DNA–A and DNA–B components) predominate in tomato in the New World. Begomoviruses are transmitted with great efficiency by the Bemisia tabaci Middle East Asian Minor 1 (MEAM-1 = biotype B) vector, which is widely distributed, with a polyphagous feeding habit and high adaptability to different environmental conditions. The introduction of B. tabaci MEAM 1 in the early 1990s was the triggering factor for begomovirus epidemics in tomato in Brazil, with a subsequent increase in the number of new viral species. Furthermore, distinct patterns of diversity and dynamics of viral subpopulations were observed in different environments. In this scenario, weeds play an important role as they function as natural reservoirs of begomovirus. New viral species have been reported in weeds, including areas in and around commercial tomato fields. Some weed begomoviruses have been identified and biologically characterized, however it is believed that these studies have not been extensive enough to estimate the actual diversity of new species in weeds. In the country, four begomoviruses were reported concomitantly infecting tomato and weeds. Globally, more than 40 begomoviruses have been described in Malvaceae species, and some of these have been detected originally infecting Solanaceae species. The small and bipartite genome, together with the transmission efficiency and polyphagy of the vector, provide favorable conditions for the occurrence of mixed infections and recombination and pseudo-recombination events between begomoviruses, thus contributing to the frequent alteration of the genetic structure of viral populations in our conditions. Different strategies have been employed to analyze the evolutionary processes capable of shaping the genetic-molecular structure of begomoviruses. The main strategy is to obtain the complete viral genome (DNA–A and DNA–B) for further analysis using different programs and evolutionary models. The metagenomics combined with High Throughput Sequencing (HTS) has allowed to determine a great viral diversity present in Brazil. A high frequency of mixed infections has been detected with many of them involving potential new species capable of inducing severe symptoms in plants. In the present work, four contigs obtained from HTS of Malvaceae leaf samples were selected for study and characterization, in according to the methodology carried out by the LVV-Fito team and summarized below. Initially foliar samples of weeds showing typical symptoms of begomovirus (generalized chlorosis, mosaics, and golden spots) were collected in tomato production areas and/or areas close to tomato cultivation, in the five regions of the country. Samplings were carried out from 2001 to 2020, with a total of 78 leaf samples from plants of the Malvaceae family analyzed (selected using the year/place of collection as criteria). Total DNA was extracted via CTAB and organic solvents and stored at -20 °C. The initial confirmation of the presence of begomovirus infection in the samples was made through PCR (polymerase chain reaction) assays using degenerate primers ‘PAR1c496’ and ‘PAL1v1978’. Viral circular DNAs were enriched in positive samples via rolling circle amplification (RCA). High-throughput sequencing (HTS) was performed on an Illumina HiSeq2500 platform. 5 The viral contigs were annotated and the reads were mapped back to the annotated genome using the ‘Map to reference’ tool available in the Geneious 11.1.5 program. About 7,391,728 million readings were obtained from the pool of 78 samples. After assembly, using the CLC Genomics Workbench 11 program, 10,679 contigs were obtained. Four contigs were selected. Three of these contigs corresponded to complete DNA–A and exhibited identity levels below 91%, consistent with the current taxonomic criteria for defining new species within the genus Begomovirus. The DNA component– Potential new species #1 showed 79% identity with Sidastrum golden leaf spot virus (HM357458), new species #2 showed 81% identity with Oxalis yellow vein virus (KM887907), while new #3 showed 78% identity with Sida yellow mosaic Alagoas virus (JX871383), confirming the occurrence of three new species of begomovirus in weeds. Sequences of DNA – B components were obtained and analyzes performed, including confirmation of cognates species. The fourth contig showed 98,24% identity with Sida micranta mosaic virus (SiMMV) (KC706535). After using specific primers, already available, a result of 41 positive samples for SiMMV was obtained. The characterization of these three species, as well as the occurrence and distribution of Sida micranta mosaic virus in five Brazilian regions will be presented in this dissertation.
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3
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EDUARDO SOARES DA SILVA LIMA
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" High Throughput Sequencing- based identification and characterization of Geminiviridae members in weeds associated with the tomato crop in Brazil ".
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Advisor : RITA DE CASSIA PEREIRA CARVALHO
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COMMITTEE MEMBERS :
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RITA DE CASSIA PEREIRA CARVALHO
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ROBERT NEIL GERARD MILLER
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HELSON MARIO MARTINS DO VALE
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MIRTES FREITAS LIMA
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Data: Apr 18, 2023
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Show Abstract
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The production of tomato (Solanum lycopersicum L.) has great economic and social importance for Brazil. Production of this vegetable is affected by many diseases, including those induced by species of Begomovirus (family Geminiviridae). Chapter 1 presents a review of these pathogens and the diseases they induce. In addition to tomato, begomoviruses have a wide range of hosts across different botanical families. Many weeds are natural hosts, playing an important role as reservoirs for such pathogens. Begomoviruses are transmitted by species of the Bemisia tabaci complex, which are characterized by having a wide geographic distribution and being extremely polyphagous. These characteristics of the vectors increase the potential for mixed viral infections and for events of recombination and pseudo-recombination. Such genetic events contribute to increased genetic variability of the viral populations, including the emergence of novel species. In addition, this large diversity can generate viral variants capable of overcoming resistance factors present in commercial varieties. In this context, monitoring viral diversity in tomato fields and in associated weeds is extremely important for effective management of this group of pathogens. Different molecular strategies have been employed for identification and characterization of begomoviruses, including High Throughput Sequencing (HTS) platforms. Herein, a broad metagenomics analysis of single-stranded, circular DNA viral species of the Geminiviridae family (especially from the genus Begomovirus) was conducted in weeds frequently occurring in tomato fields. Plants from the Malvaceae, Euphorbiaceae, Amaranthaceae, Solanaceae, Asteraceae, Rubiaceae, Fabaceae, Lamiaceae, Cucurbitaceae, Convolvulaceae and Brassicaceae families were collected in field surveys carried out in the five Brazilian regions. For this, 91 foliar samples of weeds exhibiting begomovirus-like symptoms (mosaic, mottled, chlorotic sectors, and dwarfism) were selected according to the year and collection site. Samples were collected in production areas and areas in the vicinity of tomato fields between 2003 and 2022. The samples were subjected to total DNA extraction, which was used as a template for performing Rolling Circle Amplification (RCA). The selected samples were submitted to PCR assays with specific primers for genomic regions of begomovirus species. After confirming results, the RCA pool was assembled and sent for sequencing on an Illumina Nova Seq 6000 platform – why? To enable species level identification?. In addition, the DNA of a subgroup of host plants was used for Barcoding using primers for two genes from the chloroplast genome (Rubisco and/or Maturase K) – why? To enable species level identification?. In chapter 2, there are results and analyses of viruses and subviral agents recovered via HTS. After the assembly and recovery of 100 contigs, 20 corresponded to 15 new species that will subsequently be characterized biologically and molecularly. For an isolate representing a potential new species (denominated CE–076), the complete genome was retrieved via HTS and Sanger sequencing using sequences derived from amplicons generated with five pairs of internal primers. Identity was 85.87% with tomato bright yellow mottle virus (ToBYMV). The molecular characterization of this new virus detected in the host species Bolusafra bituminosa is described in chapter 3. The information generated in this dissertation can contribute to the establishment of management systems and generate information of interest on viral diversity for tomato genetic improvement programs. Furthermore, the present work confirms the epidemiological importance of these invasive plants as reservoirs of viral species described infecting tomato as well as a potential role in the genetic evolution of populations of this group of pathogens in Brazil.
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4
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JULIANA GABRIELLE ISIDORIO DA SILVA
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Bioprospecção de viromas de plantas ornamentais das famílias Bromeliaceae e Orchidaceae no Planalto Central Brasileiro
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Advisor : RITA DE CASSIA PEREIRA CARVALHO
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COMMITTEE MEMBERS :
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RITA DE CASSIA PEREIRA CARVALHO
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MAURICIO ROSSATO
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MARILIA SANTOS SILVA PATRIOTA
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MIRTES FREITAS LIMA
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Data: Jun 1, 2023
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Show Abstract
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Viruses are widespread, yet little is understood about these biological entities. The evaluation is less than 1% of the total eukaryotic virosphere known. Common knowledge is constrained to viruses that cause diseases and economic losses. Other ecological relationships and rising factors of viruses are unexplored. The segment of plant viruses is not distinct. The understanding of diversity, evolutionary forces, and their impact is unknown. However, with the High throughput sequencing (HTS) advent, the study of the virome got tangible and fast-growing. Likewise, the bioinformatic tools improved the comprehension of viral diversity and evolution patterns. The SARS-CoV 2 pandemics, and the possible rising of new viral diseases in humans and in other economically important eukaryotes have demonstrated that it is necessary to explore the virosphere. The Orchidaceae family is the second-largest family of flowering plants. They grow in tropical areas as Bromeliaceae family specimens. The viral diversity of both families is not entirely investigated, except for the economic interest plants. In this work, the virome bioprospection research was conducted with specimens of Bromeliaceae and Orchidaceae family found in the Brazilian central plateau ornamental market. DNA and RNA HTS from these plants were performed using the Illumina NovaSeq 6000 and HiSeq 2000 platforms, respectively. Bioinformatics programs, CLC Genomic Workbench v. 20.0 and Geneious R11.1, were used to study the results found in the HTS. Two new DNA and seven RNA viruses were described infecting orchids and bromeliads. In the first chapter, a bibliographic review of the ornamental plant market and of the incidence of viruses on them was developed with the aim of exploring the status quo of research in the area. In the second chapter, the exploration of the RNA HTS results of 54,088,166 reads and 16,560 contigs was carried out with the goal of characterizing the possible RNA viruses present. For this, the contigs obtained were BLASTn confronted against a viral sequence 5 database. 15 hitherto uncharacterized viral RNA sequences were identified. A total of seven new viral species were characterized. One sequence belonging to the genus Alphaendornavirus (family Endornaviridae), two belonging to the genus Totivirus (family Totiviridae), three belonging to the genus Polerovirus (family Solemoviridae), two RNA 1 and two RNA 2 belonging to the genus Dichorhavirus (family Rhabdoviridae) and one RNA 1 and an RNA 2 belonging to the genus Nepovirus (family Secoviridae). Three RNA 1 sequences related to the genus Sadwavirus, family Secoviridae, were identified. However, no RNA 2 was found. According to the genus classification criteria adopted by the International Committee on Taxonomy of Viruses (ICTV), the information found on these previous sequences is insufficient to establish new species. The seven new species were named Alphaendornavirus monocotyledonae (OQ866133); Dichorhavirus monocotyledonae (OQ866129, OQ866130, OQ866131, OQ866132); Nepovirus monocotyledonae (OQ866134, OQ866135); Polerovirus monocotyledonae 1 (OQ866138); Polerovirus monocotyledonae 2 (OQ866139, OQ866140); Totivirus monocotyledonae 1 (OQ866136); Totivirus monocotyledonae 2 (OQ866137). The third chapter is a preface containing information about the collection sites, sample cataloging and methodologies used in subsequent chapters. The fourth and fifth chapters contains the result of the bioinformatic treatment of the 11,060,510 reads and 163,808 contigs generated by HTS. The fourth chapter is a disease note of a mixed infection of two known begomoviruses, tomato severe rugose virus (ToSRV) and tomato chlorotic mottle virus (ToCMoV), in orchids of the genus Dendrobium. The fifth chapter is a characterization of two high divergently new begomovirus-like pathogens infecting orchids of genus Spathoglottis. The two new viral sequences were named Spathoglottis-associated mottle virus 1 (OQ791967) and Spathoglottisassociated mottle virus 2 (OQ791968) and should be considered new species. Primers were designed for both sequences and confirmed the infection in orchids sample of the genus Spathoglottis.
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5
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Isabella Cristina Santos do Egito
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" Colorimetric LAMP for the detection of maize red stunt using a unique genomic region".
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Advisor : MAURICIO ROSSATO
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COMMITTEE MEMBERS :
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Adriane Wendland Ferreira
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ANGELA MEHTA DOS REIS
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MARISA ALVARES DA SILVA VELLOSO FERREIRA
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MAURICIO ROSSATO
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Data: Jun 27, 2023
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Show Abstract
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Brazil stands out for being the third largest corn producer in the world, in addition to having selfsufficiency in national supply. Even with the high production, the country has, throughout its extension, climatic conditions that favor the attack of several pathogens on maize. The red stunt of maize, caused by phytoplasma (Maize bushy stunt phytoplasma), is one of the most harmful diseases to the crop. Given the importance of the pathogen, there is a demand for detection methods that are fast and accurate. Loop-mediated isothermal amplification (LAMP) is one of these methods, being fast, sensitive, with high specificity and can be used in field analysis. The objective of the present work was the development of a LAMP protocol, through comparative genomics, for Maize bushy stunt phytoplasma (MBSP) in maize. To design the sets of primers, the entire MBSP genome sequence and other sequences of other pathogens were used in the RUCS software. It was possible to identify 3706 unique core sequences, of which the 60 most suitable were used for the design (NEB LAMP primer design) and synthesis of three sets of primers that presented the desired criteria. A collection of 52 corn samples with and without symptoms were collected, three of which had the 16S rRNA region amplified, sequenced and confirmed for the presence of MBSP through phylogenetic analysis. For the LAMP assay test, the most promising primer set 0_731_10_ID1, together with the Warmstart colorimetric LAMP 2X master mix (NEB) Kit and the MBSP positive samples were used to evaluate the best reaction conditions. The proportion of internal and external primers 1:4 and temperature of 65 °C were considered the most suitable for the reaction and were used for the other tests. The sample collection was tested with Nested-PCR with primers P1/Tint and R16mF2/R16mR2 and compared with the LAMP assay of the present work. Considering the presence and absence of symptoms, there was confirmation that symptomatic women were positive for LAMP in a greater proportion than for Nested-PCR. Assay sensitivity was evaluated using purified, quantified, and serially diluted PCR product with F3/B3 external primers in 10 concentrations. The proposed LAMP assay proved to be sensitive, detecting up to 0.1 fg µL-1 of DNA. To assess the potential of the assay with plant samples, without prior DNA extraction, discs of leaves from the samples were macerated and tested directly in the LAMP reaction, resulting in positive for symptomatic and negative for asymptomatic. This result not only confirms that the reaction resisted possible inhibitors in plant tissue, but also did not cross-react with the leaf microbiota of the samples.
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6
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DWILLIAN FIRMIANO CUNHA
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"Interactions of carrot and tomato with Meloidogyne species: Parasitism and genetic resistance".
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Advisor : LEONARDO SILVA BOITEUX
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COMMITTEE MEMBERS :
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LEONARDO SILVA BOITEUX
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JUVENIL ENRIQUE CARES
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Mara Rúbia da Rocha
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REGINA MARIA DECHECHI GOMES CARNEIRO
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Data: Aug 18, 2023
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Show Abstract
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Root-knot nematodes (Meloidogyne spp.) represent one of the most important groups of root pathogens due to the damage induced in a wide range of hosts. Parasitism by Meloidogyne spp. is a major obstacle in the commercial production of carrots (Daucus carota) in subtropical regions, causing yield and quality losses. In Brazil, the most important species in carrot crop are M. javanica and M. incognita. The cultivar ‘Brasília’ is the main source of resistance in carrot against these two species. However, molecular marker systems linked to resistance factors for Brazilian carrot cultivars are still not available, which would facilitate the selection process. Tomato (Solanum lycopersicum) is also severely affected by Meloidogyne spp. with special emphasis on M. enterolobii due to its recent geographic expansion and the ability to “break-down” the resistance conferred by the Mi-1.2 gene. However, alternative sources of resistance in tomato to M. enterolobii are not yet available. Furthermore, new methodologies for nematological evaluation are necessary for the selection of resistance plants in both vegetable crops. In this context, the present work aimed to study different aspects of carrot and tomato interactions with Meloidogyne species. In carrots, a study was carried out with an F2 population of 108 plants of the cultivar ‘Brasília #83147’ (which segregates for resistance against M. javanica). Plants were inoculated with 8000 eggs + J2 of M. javanica 30 days after sowing. Total DNA was extracted from foliar samples and evaluated (via PCR) with codominant SCAR/STS markers (SQ1850R / SQ1700S and SQ6650R / SQ6590S) previously identified in close linkage with a resistance locus (Mj-1) to M. javanica in ‘Brasilia’. As for the nematological analyses, the root systems of individual carrot plants were evaluated at 120 days after inoculation (DAI) for the reproduction factor (FR = pf/pi) and the individual quantification of galls and egg mass. Roots of individual contrasting plants (resistant and susceptible) were vernalized and subjected to selfing (S1 generation). Inoculation bioassay in conjunction with marker analyses confirmed that resistance is not fixed in this population, with many susceptible individuals being detected. However, it was possible to identify individuals with extreme resistance (immunity-like response) and with the presence of codominant SCAR/STS markers. These individuals can now be used as parents to facilitate genetic studies as well as the development of new Brazilian carrot cultivars. In addition, the scale proposed in this work was presented as an extremely useful tool, being easy and quick to use for germplasm assessment. In tomato, the reaction of 24 germplasm accessions was verified under different inoculum levels of M. enterolobii. The potential residual effect of the Mi-1.2 gene was also studied in comparative assays using isogenic lines (isolines) contrasting for this resistance factor: ‘Rio Grande’ (mi-1.2/mi-1.2) versus ‘Nemadoro’ (Mi-1.2/ Mi-1.2). The allelic status of each individual plant was confirmed via the use of molecular markers linked to the Mi1.2 locus. This molecular marker was also employed to evaluate the use of the breeding use of this gene in the Brazilian varietal panorama. Accessions were inoculated with four levels of M. enterolobii inoculum (1000, 2000, 4000, and 8000 eggs + J2s) 15 days after transplanting. For the nematological analyses, the tomato roots (at 45 days after inoculation) were evaluated according to the gall index (GI), egg mass index (EMI), number of eggs per gram of root (NOGR), and the reproduction factor (RF). All evaluated tomato accessions showed susceptible responses, demanding the search of novel sources of resistance against this emerging pathogen. In turn, the Mi-1.2 gene did not confer a significant residual effect against M. enterolobii. Although extremely effective against at least 13 Meloidogyne species, this gene was not able to interfere in the infection process of this nematode. In the M. enterolobii inoculation process, the most appropriate inoculum levels ranged from 1000 to 2000 eggs + J2 to obtain the maximum manifestation of nematological traits. Given the importance of tomato and carrot in Brazil, the results of this study contribute substantially to breeding programs aimed at developing cultivars resistant to different Meloidogyne species.
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7
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Cleberly Evangelista dos Santos
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The pathosystem Fusarium oxysporum f. sp. lycopersici-tomato in Brazil: Variability of effector genes, molecular markers and stability of the I-7 resistance gene.
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Advisor : LEONARDO SILVA BOITEUX
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COMMITTEE MEMBERS :
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LEONARDO SILVA BOITEUX
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JOSE RICARDO PEIXOTO
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CLEIA SANTOS CABRAL
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MILTON LUIZ DA PAZ LIMA
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Data: Aug 31, 2023
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Show Abstract
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Vascular wilt, caused by different races of the fungus Fusarium oxysporum f. sp. lycopersici (FOL), is one of the main diseases affecting the tomato crop (Solanum lycopersicum L.) in the world. This group of pathogens invades the root system in susceptible cultivars and colonizes the xylem vessels, preventing the ascending transport of sap and causing the wilt symptoms. Three physiological races of FOL have already been described and a hypothetical race 4 is in the process of being characterized in California. These races are defined based upon their ability to infect a set of differential tomato accessions with factors containing four dominant resistance genes (I, I-2, I-3, and I-7). The planting of cultivars with genes for resistance to physiological races is the main control measure adopted in production areas, since the soil infested with the pathogen is impossible to eradicate. The determination of the physiological races of FOL is also possible by means of molecular markers developed by Hirano & Aire (2006) that also allow the identification of F. oxysporum f. sp. radicislycopersici, quickly and reliably. However, the initial characterization of the physiological races of FOL in Brazil was based exclusively upon pathogenicity assays. Characterization studies conducted subsequently showed that FOL race 2 isolates present in Brazil have a molecular pattern distinct from FOL race 2 isolates occurring in other regions of the world. A group of small proteins rich in the amino acid cysteine, called SIX (= secreted into the xylem), are effector molecules of FOL that play a crucial role in host colonization and also in the expression of symptoms. An alternative molecular marker system was developed based on the use of primers derived from the Six series of genes. However, this system of molecular markers has not yet been evaluated with Brazilian isolates. A representative set of Brazilian FOL isolates was evaluated with these primers and a new marker (= Six 4) proved to be useful to discriminate between race 1 and race 2 FOL isolates from Brazil (Chapter 2). Another objective of the present work was to evaluate the stability and dosage effect of the I-7 gene in homozygosity and heterozygosity against Brazilian isolates of FOL race 3 (Chapter 3). The results indicated that a single dose of the I-7 gene is sufficient to prevent the expression of symptoms in all evaluated isolates. (Chapter 3). A novel set of codominant, CAPS-like functional molecular markers evaluated for use in monitoring the incorporation of the I-7 gene in tomato accessions (Chapter 4). For this, a segregating F2 population for the I-7 gene was obtained, which were inoculated with a suspension of FOL race 3 spores and evaluated using a rating scale. DNA from susceptible and resistant plants was extracted and used as a template for analysis of single nucleotide polymorphism on chromosome 8 and development enabled the development of a functional marker increased efficiency and offered greater security in the process of selection of disease resistant plants within breeding programs. In summary, this work presents new and relevant information about the FOL-tomato pathosystem that can be applied in genetic management strategies for this group of pathogens.
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8
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Giovana Curcio Guimarães
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"Diversity of ssDNA and alphasatellite viruses in solanaceae and molecular characterization of three new species of Begomovirus in tomato".
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Advisor : RITA DE CASSIA PEREIRA CARVALHO
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COMMITTEE MEMBERS :
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DANIEL MENDES PEREIRA ARDISSON DE ARAUJO
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MIRTES FREITAS LIMA
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MÁRCIO MARTINELLO SANCHES
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RITA DE CASSIA PEREIRA CARVALHO
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Data: Dec 19, 2023
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Show Abstract
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In Brazil, the tomato (Solanum lycopersicum L.) is one of the most important vegetable crops both economically and socially. The production costs and yield levels of this crop suffer recurrent fluctuations due to several factors, among which diseases such as begomoviruses (caused by a complex of Begomovirus species) stand out. These pathogens have one or two single-stranded circular DNA molecules, encapsidated in 18- 30 nanometer particles. Begomoviruses with a bipartite genome (presenting DNA–A and DNA–B components) predominate in tomato in the New World, being transmitted by different cryptic species of the Bemisia tabaci complex. These vectors have polyphagous feeding habits, being widely distributed, with an extensive circle of hosts, including several weeds. The introduction of B. tabaci Middle East Asia Minor 1 – MEAM-1 (= biotype B) took place in the early 1990s, triggering epidemics of begomoviruses in tomato, with an exponential increase in the number of Begomovirus species. The presence of weeds, mainly from the Solanaceae family, plays a very important role as reservoirs of tomato-infecting begomoviruses. New viral species have been reported in weeds, and such interactions increase the occurrence of natural mechanisms that generate genetic variability, which can lead to the surfacing of new viral variants. The advent of High Throughput Sequencing (HTS) technology has led to the discovery of new viral species associated with tomato cultivation. In this context, one of the objectives of this work is to study viral diversity in tomato and weeds from the Solanaceae family. One hundred and thirty-five (135) tomato and weed leaf samples exhibiting typical begomovirus-like symptoms (apical chlorosis, golden mosaic, and chlorotic spots) were collected in the North (15), Northeast (43), South (11), Center West (45), Southeast Brazil (18) and Uruguay (3). Collections were carried out from 2003 to 2022, and leaf samples were analyzed from S. lycopersicum (92), S. aethiopicum (4), S. paniculatum (3), S. sessiliflorum (1), S. melongena (4), S. mammosum (1), Capsicum species (12), Nicandra physalodes (8), Physalis species (6), S. betaceum (2), S. viarum (1), and S. americanum (1). These samples were selected using year/place of collection as criteria. The extracted total DNA was used as template for rolling circle amplification (RCA). Initially, the RCA of each sample was used to confirm the presence of begomovirus via PCR assays using degenerate primers for DNA–A (PAR1c496 and PAL1v1978) and for DNA–B (PBL and CRC). After this step, the RCA products of the 135 samples were used to form a pool and sequenced on an Ilumina NovaSeq 6000 platform. A total of 19,735,294 million reads was obtained. These reads were used to assemble contigs in CLC Genomics Workbench 11. 41,659 contigs were obtained, 173 of which showed (after analysis with RefSeqViral) identities with begomoviruses (170), topilevirus (1), gemycircularvirus (1) and one alphasatellite. Sixteen out of the 170 begomovirus-like contigs showed nucleotide identity below 91%, being consistent with the current taxonomic criteria for defining new species within the genus Begomovirus. Contigs C195, C18367 and C2046 showed identities of 99.1%, 99.67% and 99.92% with tomato apical leaf curl virus (ToALCV; MH539677), plant associated genomovirus 12 (MT214094), and Alphasatellitidae species (MT214093), respectively. With the exception of ToALCV, the other viruses will be fully characterized in the future. Three contigs were considered as potential novel species within the Begomovirus genus. These contigs (C16, C21 and C230) showed 80%, 94% and 80% identity with melochia mosaic virus (KT201151), tomato bright yellow mottle virus (KC791691) and ToBYMV (KC791691), respectively. The alignment of these three contigs showed a typical genomic organization of bipartite begomoviruses. The common regions of C16 and C230 were determined and analyzes were carried out with their iterons and other motifs verifying them as cognate bipartite genomes. On the other hand, contig C21 displayed a typical genomic organization of a monopartite begomoviruses, also representing a potential new viral species. In conclusion, with the employment of HTS it was possible to observe the presence of high levels of genetic variability and diversity of begomoviruses in members of the Solanaceae family, indicating their relevant role as viral reservoirs. Research actions are being conducted aimed at producing infectious clones to uncover the range of hosts as well as sources of resistance against these new viruses.
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9
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JOYCE PEREIRA DE SOUZA PAES
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“Monitoring of nematodes in commercial cultivation of irrigated Nanica banana and evaluation of biological and chemical control agents for root-knot nematodes in Cv banana trees. Nanica and BRS Princesa”.
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Advisor : JUVENIL ENRIQUE CARES
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COMMITTEE MEMBERS :
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JUVENIL ENRIQUE CARES
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CLEBER FURLANETTO
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LARISSA DE BRITO CAIXETA VASCONCELOS
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JADIR BORGES PINHEIRO
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Data: Dec 20, 2023
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Show Abstract
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Banana crops are often affected by the attack of plant-parasitic nematodes. Therefore, growers have been seeking control options that minimize losses and are not harmful to the environment. This study aimed to evaluate the effectiveness of control measures adopted in a 200-hectare banana plantation in the western region of Bahia. For this purpose, the population fluctuation of both plant-parasitic and free-living nematodes was monitored through soil and root samples collected at 54 georeferenced points on the farm every 3-4 months for two years. Nematodes were extracted from both roots and soil. Plant-parasitic nematodes were quantified and identified at the species level, while freeliving nematodes were identified at the trophic group level. Maps of distribution and population level were then generated, and the population fluctuation of nematodes was analyzed in relation to the application of biological control agents (Bacillus subtilis, B. licheniformis, B. amyloliquefaciens, and Trichoderma asperellum). After six collection cycles, it was possible to observe a general trend of population reduction among plantparasitic nematodes and a progressive increase in the presence of free-living nematodes in the banana plantation. However, this reduction was more pronounced for the species Helicotylenchus multicinctus than for Radopholus similis, Meloidogyne incognita, and M. javanica, indicating a higher vulnerability of nematodes that spend longer periods in the soil. Root-knot nematode populations were less affected over time, and in some areas of the farm, their presence increased. When evaluating the isolated effect of antagonists in a controlled environment, a loss of efficiency was noted, suggesting that the test conditions in pots favor nematode multiplication at the expense of antagonists. Antagonists require the interaction of multiple factors to achieve the desired control effect. Biological control agents contribute to reducing the impact of nematode parasitism in banana cultivation, but greater efforts are needed to develop effective strategies for controlling root-knot nematodes.
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Thesis |
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1
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Leticia Dias de Freitas
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Advisor : ROBERT NEIL GERARD MILLER
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COMMITTEE MEMBERS :
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ROBERT NEIL GERARD MILLER
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JUVENIL ENRIQUE CARES
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LEONARDO SILVA BOITEUX
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MARIA EUGENIA LISEI DE SA
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JANSEN RODRIGO PEREIRA SANTOS
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Data: Jan 27, 2023
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Show Abstract
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2
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Débora Gonçalves Pereira
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"Phytophthora capsici: Diversity and resistance in Solanum (Lycopersicon)".
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Advisor : LEONARDO SILVA BOITEUX
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COMMITTEE MEMBERS :
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LEONARDO SILVA BOITEUX
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RITA DE CASSIA PEREIRA CARVALHO
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CLEIA SANTOS CABRAL
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JADIR BORGES PINHEIRO
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KAMILA CAMARA CORREIA
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Data: Feb 28, 2023
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Show Abstract
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Phytophthora capsici can induce severe losses in several crops, including tomato (Solanum lycopersicum). Chapter I: reviews this pathogen and how difficult is the management of this oomycete, mainly due to the lack of resistant cultivars and the wide variation in the virulence profile of the pathogen’s isolates. However, few studies have studied sources of resistance as well as interaction patterns between Solanum accessions (Lycopersicon) and neotropical isolates of this pathogen. In chapter II: the genetic structure of 48 isolates classified as P. capsici based on morphological aspects was investigated. Subsequently, ten isolates from different hosts were evaluated for their ability to cause disease in sweet pepper fruits and seedlings of the ‘Santa Clara’ and ‘Tico’ sweet peppers. Isolates from different Brazilian states and hosts were genotyped for cox2 (mitochondrial gene). All 48 isolates evaluated with primer-specific produced amplicons of the expected size, confirming the identity of the isolates as P. capsici. The two compatibility groups (A1 and A2) were observed among the isolates, even among collections from the same location. The isolates ‘PCa-31’, ‘PCp-183’, ‘PCp-167’, ‘PCa-29’, ‘PCa-31’, ‘PCp-183’, ‘PCp-167’, and ‘PCa-29’ caused diseases in bell-pepper fruits, being possible to visualize mycelium on the infected tissue, four days after inoculation. Some isolates that caused disease in bell-pepper fruits did not cause disease in seedlings of ‘Tico’ and ‘Santa Clara’, demonstrating that there is an apparent tissue-specific interaction for the occurrence of disease. The analysis of the cox2 sequences obtained from this collection of isolates indicated that this locus is efficient as a precise “barcoding” at the species level in P. capsici. The phylogenetic relationships observed among isolates in the phylogenetic trees for cox2 did not show clusters correlated to compatibility group, collection site or original host plant. In chapter III: studies were conducted to identify the presence of potential pathotypes as well as the definition of an adequate panel of accessions of differential hosts in this pathosystem. Seventeen virulent isolates (from different host plants and geographic regions) were used to evaluate accessions of Solanum (Lycopersicon) that showed superior levels of resistance to one or more isolates in previous bioassays. Eight potential pathotypes were identified according to their interaction patterns with nine accessions of Solanum (Lycopersicon). The cultivar S. lycopersicum ‘Santa Clara’ exhibited a “universal” susceptible reaction (100% mortality for all isolates), while ‘Hawaii 7996’ followed by S. habrochaites ‘PI 127826’ and ‘PI 127827’ were the main sources broad-spectrum resistance, exhibiting superior performance against a wide range of isolates. Inheritance studies and mapping of these pathotype-specific resistance factors for each accession will facilitate their incorporation into commercial cultivars. These eight informative accessions of Solanum (Lycopersicon) are suggested as differential host accessions for this pathosystem and may provide a more accurate picture of the pathotypes that occur in the field through simple bioassays based on the capacity of the isolates to “breakdown” this unique set of specific genes of this germplasm. In chapter IV: three controlled bioassays were conducted to evaluate the reaction of 28 accessions of Solanum (Lycopersicon) against a collection of seven P. capsici isolates. Capsicum annuum ‘Tico’ was used as a susceptible control. Inoculations were performed by depositing a suspension (2 x 104 zoospores per mL) around the crown area of the seedlings. The mortality rate was evaluated 14 days after inoculation. All isolates (in all bioassays) induced severe symptoms in ‘Tico’ (100% mortality). The accession S. lycopersicum ‘Hawaii 7996’ (resistant to Ralstonia species) showed superior levels of isolate-specific resistance to four out of six isolates, while S. habrochaites ‘WIR 7924’ exhibited resistance to five out of seven isolates. Two of the 18 accessions of S. habrochaites (‘PI 127826’ and ‘PI 127827) displayed immunity-like resistance against two P. capsici isolates. The resistance responses of these accessions were not coincident, indicating the potential presence of pathotypes. Unstable responses of some accessions were observed in the trials, indicating complex inheritance or incomplete penetrance of the resistance. The development of cultivars with a broad-spectrum of resistance to multiple P. capsici isolates is essential for the sustainable management of this highly variable pathogenic oomycete. Therefore, pyramiding resistance factors from ‘Hawaii 7996’ and S. habrochaites accessions into a single genome would be a promising breeding strategy aimed at developing tomato cultivars with stable, broad-spectrum resistance to a wide range of P. capsici isolates. Extensive variation in the virulence profile has been observed for many pathogen isolates, inducing sharp contrasting reactions among host accessions. However, no extensive work has investigated the interaction patterns between Solanum (Lycopersicon) and pathogen isolates. In Chapter V: Inheritance studies were conducted to determine the genetic basis of S. habrochaites ‘PI 127827’ resistance. Crosses were made with the susceptible accession ‘Ponderosa’. The F1 and F2 populations were obtained and inoculated with a zoospore suspension of isolate ‘PCp-182’. The frequency distribution of resistant and susceptible plants in the F2 generation (evaluated by the Chi-square test) indicated a good fit for an epistatic model (15:1) involving two genetic factors in duplicate. In this way, the information generated in the present work provides crucial elements for the establishment of a scientific and technological base for the development of tomato cultivars with stable and durable resistance against a pathogen with a wide range of host plants.
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3
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Rildo Alexandre Fernandes da Silva
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Diversity and identification of phytopathogenic fungi in unconventional fruits in Brazil.
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Advisor : DANILO BATISTA PINHO
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COMMITTEE MEMBERS :
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MARIA ALVES FERREIRA
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Flávia Rodrigues Barbosa
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DANILO BATISTA PINHO
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ROBERT NEIL GERARD MILLER
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WILLIE ANDERSON DOS SANTOS VIEIRA
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Data: Jul 28, 2023
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Show Abstract
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Unconventional fruits in Brazilian agriculture, such as Syzygium jambos (Myrtaceae), are widely studied for medicinal use, but the diseases that deteriorate these fruits are neglected. Fungal genera from the families Nectriaceae, Bionectriaceae, and Botryosphaeriaceae cause significant diseases in agriculturally and commercially important plants, but they are rarely reported in native species and/or causing fruit rot. The identification of these fungi is challenging, especially the Calonectria-like group (Nectriaceae) that includes phylogenetically and morphologically related fungi to the genus Calonectria. Since 1995, the submission of sequences from different genomic regions has been consolidated in GenBank for the recognition of phylogenetic species. This rule has facilitated species identification, but the lack of uniformity and the increasing volume of data make it difficult or even impossible to study host range and geographical distribution comprehensively. Therefore, this study aimed to conduct a meta-analysis of the information available in GenBank for Calonectria-like fungal complex and to morpho-molecularly identify the isolates from the Calonectria-like, Clonostachys, Cylindrocladiella, Gliocephalotrichum, Gliocladiopsis and Neofusicoccum obtained from fruits collected in Distrito Federal, Goiás, Minas Gerais, Paraná, Pernambuco, Santa Catarina and São Paulo. The meta-analysis revealed discrepancies among the genomic regions deposited for each isolate, with the sequences from the elongation factor region (n=5,125) and beta-tubulin region (n=5,287) being more representative compared to calmodulin, histone, RNA Polymerase 2, internal transcribed spacer, and the large ribosomal subunit (28S rDNA). The exchange of samples between research groups led to the "Matryoshka Effect," evidenced by the over-representation of isolates. The analysis reveals that 78.3% of the analyzed sequences have information about the collection location, while only 12% have information about the substrate/host. While most genera have a broad geographical distribution, the genera Curvicladiella and Xenocylindrocladium are restricted to South America and Asia, respectively, with the majority of species found in Brazil (90%) and China (100%), respectively. The 86 morphomolecularly identified isolates belong to the following genera: Calonectria (n=15), Clonostachys (n=8), Cylindrocladiella (n=28), Gliocladiopsis (n=18), Gliocephalotrichum (n=2), and Neofusicoccum (n=15). Among these, seven known species have been documented, while one, two, and three new species will be proposed for Clonostachys, Calonectria and Cylindrocladiella genera, respectively. The species Cylindrocladiella vitis, Gliocladiopsis hennebertii, Neofusicoccum occulatum, and N. umdonicola are reported for the first time in Brazil while new host combinations were recorded for C. pseudocholeuca, C. rogersoniana, Cy. infestans, Cy. lageniformis, Cy. peruviana, Gliocephalotrichum simplex, Gliocladiopsis tenuis, N. batangarum, and N. parvum on fruits of the Dypsis madagascariensis, Eugenia aggregata, Eugenia involucrata, Euterpe edulis, Spondias mombin, Syzygium jambos and Terminalia catappa. This study demonstrates the need for comprehensive surveys on the diversity of fungal species in native plants and highlights the risk of transmission to economically important plants.
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4
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Tiago Silva Jorge
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Groundnut ringspot orthotospovirus (GRSV) in lettuce and new hosts: Identification of susceptibility genes via transcriptomic analysis, search for new sources of natural resistance and mobilization of the Sw-5b resistance gene from tomato to lettuce via transgenics.
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Advisor : LEONARDO SILVA BOITEUX
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COMMITTEE MEMBERS :
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LEONARDO SILVA BOITEUX
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RITA DE CASSIA PEREIRA CARVALHO
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ROBERT NEIL GERARD MILLER
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Rosana Rodrigues
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SIMONE DA GRAÇA RIBEIRO
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Data: Aug 30, 2023
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Show Abstract
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Lettuce (Lactuca sativa L.) is one of the main leafy vegetables in Brazil and worldwide. Diseases caused by a complex of Orthotospovirus species (including groundnut ringspot orthotospovirus – GRSV and tomato spotted wilt orthotospovirus – TSWV) are among the most important of this vegetable. GRSV is the predominant viral species in Brazil and, up to now, lettuce cultivars with adequate levels of resistance are not available. This thesis was organized into five chapters. Chapters I & II present a review of this viral complex and the advances obtained through classical and biotechnological resistance breeding to orthotospoviruses in lettuce, Capsicum, and tomato crops. In Chapter III, experiments were conducted to identify new sources of genetic resistance to orthotospoviruses in germplasm of cultivated and wild lettuce species. Sixty-five (65) Lactuca accessions were initially evaluated under field (natural) inoculum conditions. Eleven (11) accessions (displaying low incidence/severity of symptoms) were selected for greenhouse trials via mechanical inoculation. Five different orthotospovirus isolates (three GRSV isolates and two TSWV isolates) were used. Three evaluated accessions (‘Bedford’, ‘Belíssimo’, and ‘UC12100’) showed low values of viral incidence (= high tolerance levels). No accessions showed immunity-like responses. This is the first study identifying sources of genetic tolerance to GRSV. In chapter IV, a transcriptomic analysis was conducted to identify genes differentially expressed across inoculated versus non-inoculated lettuce plants during the interaction between the susceptible cultivar ‘Salinas’ and a GRSV isolate. The total RNA of the plants was collected in five periods after inoculation (1 hour, 6 hours, 24 hours, 48 hours, and 7 days). The dynamics of viral replication over different times was determined via real-time PCR. The best time for transcriptomic analysis was identified at 48 hours after inoculation, when viral multiplication in the host tissues was initiated. The samples were then submitted to the RNA-seq. A total of 273 genes was found to be differentially expressed in the contrasts between inoculated versus non-inoculated plants, including different categories of gene functions. In chapter V, cisgenic lettuce events containing the tomato Sw-5b resistance gene were challenged under controlled inoculation conditions with a GRSV isolate. The evaluation of the effectiveness of the events was carried out by confirming the presence of the virus by visual and serological evaluation of diseased plants, with subsequent nucleic acid extraction and PCR with virus-specific primers. The genomic insertion of the Sw-5b gene and the presence of its transcripts were also evaluated in lettuce leaf tissues. Lettuce transformed with the Sw-5b gene showed high susceptibility to GRSV, with only a few asymptomatic plants. The Sw-5b gene product and its transcripts were detected in both symptomatic and asymptomatic plants, indicating that the transfer of this gene alone was not effective to provide resistance to GRSV in lettuce. In chapter VI, tests were conducted aiming at the characterization of new natural or experimental host species of GRSV among crops and weeds. Three crops have been reported as new natural hosts, including Cichorium endive, Cichorium intybus, and eggplant (Solanum melongena). Thirty accessions from the germplasm bank of jurubebas were challenged with a GRSV isolate to identify new potential experimental hosts. All accessions were susceptible. This result was confirmed by dot-ELISA test. Accessions of S. macrocarpon, S. acanthodes, S. viarum, S. subinerme, S. scuticum, S. stramoniifolium, and S. sisymbriifolium were reported as new experimental hosts of GRSV. This thesis provides new information that will help to establish more effective breeding and management strategies of diseases induced by orthotospoviruses in lettuce and other host crops.
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5
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Karoliny de Almeida Souza Americo
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"Nematicidal action of Bacillus methylotrophicus on Meloidogyne enterelobii and physiological changes induced by the nematode in guava trees treated with the bionematicide".
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Advisor : JUVENIL ENRIQUE CARES
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COMMITTEE MEMBERS :
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JUVENIL ENRIQUE CARES
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MAURICIO ROSSATO
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SUELI CORRÊA MARQUES DE MELLO
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JADIR BORGES PINHEIRO
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JANSEN RODRIGO PEREIRA SANTOS
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Data: Dec 12, 2023
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Show Abstract
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Photosynthesis, stomatal conductance, and transpiration are plant physiological activities that influence the productive potential of crops. The interference of root-knot nematodes in plant physiology is still poorly studied, as well as the capacity of rhizobacteria to mitigate physiological damage caused by nematodes. The lack of alternative methods of control to the nematode Meloidogyne enterolobii hinders efficient and sustainable management friendly to the environment. The objectives of this study were to evaluate in vitro the effect of a bionematicide, with the active ingredient Bacillus methylotrophicus, on the eggs and second stage juveniles (J2s) of M. enterolobii; and to monitor changes in the physiology of guava plants inoculated with M. enterolobii and treated with the bionematicide over time. In vitro experiments were conducted on M. enterolobii eggs and J2s. The treatments included concentrations of 1%, 10%, 25%, 50%, and 70% of the bionematicide, with water as control treatment. The experimental design was completely randomized with six replicates and two repetitions over time. The treatments with eggs were evaluated at 2, 4, 6, 8, and 10 days after incubation by counting the number of damaged eggs. The treatments with juveniles were evaluated at intervals of 24 to 96 hours by counting the number of dead juveniles. The monitoring of guava tree physiological activity was assessed in greenhouse and field conditions. The treatments included: 1) non-inoculated and non-treated guava cv. Pedro Sato (control); 2) non-inoculated and non-treated guava cv. Paluma guava tree (control); 3) guava 'Pedro Sato' treated with B. met and noninoculated; 4) 'Paluma' treated with B. met and non-inoculated; 5) Paluma' inoculated and non-treated with B. met; 6) 'Pedro Sato' inoculated and non-treated with B. met; 7) 'Pedro Sato' inoculated and treated with B. met, and 8) 'Paluma' inoculated and treated with B. met. The treatments were arranged in a completely randomized design (DIC) and randomized block design (DBC) in the greenhouse and field, respectively. Four applications of the bionematicide were made, at doses of 3 mL/L of water per plant in the greenhouse and 1 mL/L of water per plant in the field experiment. Between the second and third application, the plants were inoculated with 5000 eggs and eventual juveniles of M. enterolobii in the greenhouse, and 6000 eggs and eventual juveniles of the nematodes in the field. In the greenhouse, seven evaluations of physiological parameters of the guava trees (stomatal conductance, water use efficiency, photosynthesis, and transpiration) and six evaluations of total chlorophyll content were performed throughout cycle of the guava plant development. At 132 days after inoculation (DAI), the roots were weighed, and the gall index, egg mass index, and reproduction factor were determined. In the field, ten evaluations of physiological parameters of the guava trees (stomatal conductance, water use efficiency, carboxylation efficiency, photosynthesis, and transpiration) were performed, and productivity was estimated through harvest and subsequent fruit weighing. In vitro, damage to the eggs increased for all treatments, with no difference between doses after six days of exposure. J2 mortality increased up to 48 hours in the lower doses (1%, 10%, and 25%), remaining close to 100% in the higher doses (50% and 70%), with no difference between doses after 48 hours of exposure. In the greenhouse, changes in physiological parameters were concentrated in the first 44 DAI. Meloidogyne enterolobii and B. met combined reduced stomatal conductance and transpiration at 26 and 44 DAI, respectively. Photosynthesis was lower at 26 DAI in treatments that received both nematode and bacterium in combination. Meloidogyne enterolobii and B. met, either isolated or combined, showed reduced water use efficiency at 26 and 44 DAI. Gall index, egg mass index, and reproduction factor were higher in treatments that received the nematode. In the field, there was no difference over time for physiological variables for all treatments, and there was no pattern in the changes caused by isolated or combined M. enterolobii and B. met. Meloidogyne enterolobii reduced stomatal conductance in eight evaluations, water use efficiency and photosynthesis in three evaluations, and transpiration in seven evaluations. The only increase conditioned by the nematode was observed in water use efficiency at 369 DAI. Bacillus methylotrophicus reduced photosynthesis at 421 DAI but increased it at 527 DAI; at 318 and 369 DAI, there was an increase in water use efficiency. Meloidogyne enterolobii and B. met combined reduced stomatal conductance in eight evaluations and transpiration in four evaluations. Reduction in water use efficiency and photosynthesis was observed at 421 DAI. Meloidogyne enterolobii and B. met combined increased water use efficiency at 369 DAI and carboxylation efficiency at 224 and 465 DAI. The application of B. met did not mitigate the negative effects of M. enterolobii on the physiological activities of the guava trees, and isolated or combined M. enterolobii and B. met did not alter guava tree productivity. Bacillus methylotrophicus caused damage to the eggs and mortality of M. enterolobii J2. Under controlled conditions, M. enterolobii and B. met combined reduced stomatal conductance, water use efficiency, photosynthesis, and transpiration of guava trees during the initial phase of fruit development. Isolated M. enterolobii and B. met reduced water use efficiency at 26 and 44 DAI. The nematological parameters were not influenced by the bacterium. In the field, the absence of alterations may be due to the influence of environmental factors, inoculum density, bacterium application method, and combinations of nematode and bacterium application over time. Reduction in physiological parameters by the nematode was dominant. The application of B. met did not mitigate the negative effects of the nematode on the physiological activity of the guava trees. Isolated or combined M. enterolobii and B. met did not alter guava productivity.
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