Phytoplasmas are bacterial intracellular plant pathogens that cause devasting yield losses in diverse crops worldwide. Phytoplasmas were detected in clover and Catharanthus roseus plants, pear, apple and apricot trees. SecA and 16S rRNA genes, spacer region and 23S rRNA gene of five phytoplasma isolates were sequenced.

O filo Myxozoa compreende organismos parasitários que infectam vertebrados, principalmente peixes, em diversas regiões do Brasil e do mundo. Com mais de 2180 espécies já descritas os mixosporídeos estão entre os patógenos mais importantes que infectam peixes tanto de ambiente natural como de sistemas de criação. Neste estudo, foram realizadas coletas no ano de 2012, onde foram capturados 13 espécimes de piraputanga (Brycon hilarii) e 14 espécimes de piauçu (Leporinus macrocephalus) oriundos de pisciculturas do estado de São Paulo. Foram encontrados mixosporídeos infectando o rim de B. hilarii e o filamento branquial de L. macrocephalus. Para a identificação das espécies de parasitos, foi utilizada a microscopia de luz para a análise morfológica e, também, histopatológica para a observação das alterações decorrentes do parasitismo. A análise ultraestrutural foi realizada para verificar a interação parasita-hospedeiro do mixosporídeo encontrado na piraputanga. A análise filogenética do gene 18S rDNA avaliou a relação entre as espécies identificadas com outras espécies de mixosporídeos já descritas. Uma nova espécie de Myxobolus foi descrita neste estudo infectando o rim de B. hilarii e a caracterização molecular e filogenética de Henneguya leporinicola, encontrado infectando o filamento branquial de L. macrocephalus, foi realizada. No estudo filogenético, utilizando o método de Máxima Verossimilhança para ambas as espécies, verificou-se que as espécies de mixosporídeos se agruparam primeiro de acordo com a ordem do hospedeiro Characiforme e pelo tropismo tecidual. A análise molecular demonstrou diferenças genéticas significativas em comparação com outras espécies já descritas na América do Sul e em outros países. / The Myxozoa phylum comprises parasitic organisms that infect vertebrates, mainly fish, in many areas from Brazil and from the world. With more than 2180 species already described, myxosporeans are among the most important fish pathogens which infect both natural environment and breeding systems. In this study, captures were made in 2012, where 13 specimens of piraputanga (Brycon hilarii) and 14 specimens of piauçu (Leporinus macrocephalus) proceeding from fish farms in the state of Sao Paulo were capured. Myxosporeans were found infecting the kidney of B. hilarii and the gill filament of L. Macrocephalus. For the identification of parasite species, it was used light microscopy for morphological and histopathological analyzes to observe the damage caused by parasitism. The ultrastructural analysis was performed to verify the host-parasite myxosporean interaction found on piraputanga. The phylogenetic analysis of 18S rDNA evaluated the relation between the species identified with other myxosporean species already described. A new Myxobolus species was described in this study infecting the kidney of B. hilarii and molecular characterization and phylogenetic of Henneguya leporinicola, infecting the gill filament of L. macrocephalus, was performed. In phylogenetic study, using the method of Maximum Likelihood for both species, it was observed that the species of myxosporean grouped according to the order of the host Characiform and the tissue tropism. Molecular analysis showed significant genetic differences with other described species in South America and other countries.

Cell growth and proliferation are processes in the cell that must be tightly regulated. Transcription of ribosomal RNA and ribosomal biogenesis are directly linked to cell growth and proliferation, since the ribosomal RNA encodes for the majority of transcription in a cell and ribosomal biogenesis influences directly the number of proteins that are synthesized. In the work presented in this thesis, we have investigated the ribosomal RNA genes, namely the ribosomal DNA genes and the 5S rRNA genes, and their transcriptional regulation. One protein complex that is involved in RNA polymerase I and III transcription is the chromatin remodelling complex B‑WICH (WSTF, SNF2h, NM1). RNA polymerase I transcribes the rDNA gene, while RNA polymerase III transcribes the 5S rRNA gene, among others. In Study I we determined the mechanism by which B‑WICH is involved in regulating RNA polymerase I transcription. B‑WICH is associated with the rDNA gene and was able to create a more open chromatin structure, thereby facilitating the binding of HATs and the subsequent histone acetylation. This resulted in a more active transcription of the ribosomal DNA gene. In Study II we wanted to specify the role of NM1 in RNA polymerase I transcription. We found that NM1 is not capable of remodelling chromatin in the same way as B‑WICH, but we demonstrated also that NM1 is needed for active RNA polymerase I transcription and is able to attract the HAT PCAF. In Study III we investigated the intergenic part of the ribosomal DNA gene. We detected non-coding RNAs transcribed from the intergenic region that are transcribed by different RNA polymerases and that are regulated differently in different stress situations. Furthermore, these ncRNAs are distributed at different locations in the cell, suggesting that they have different functions. In Study IV we showed the involvement of B‑WICH in RNA Pol III transcription and, as we previously had shown in Study I, that B‑WICH is able to create a more open chromatin structure, in this case by acting as a licensing factor for c-Myc and the Myc/Max/Mxd network. Taken together, we have revealed the mechanism by which the B‑WICH complex is able to regulate RNA Pol I and Pol III transcription and we have determined the role of NM1 in the B‑WICH complex. We conclude that B‑WICH is an important factor in the regulation of cell growth and proliferation. Furthermore, we found that the intergenic spacer of the rDNA gene is actively transcribed, producing ncRNAs. Different cellular locations suggest that the ncRNAs have different functions. / <p>At the time of the doctoral defence the following papers were unpublished and had a status as follows: Paper 2: Manuscript; Paper 3: Manuscript</p>

Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2016-06-29T12:07:11Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação Livia Araujo.pdf: 1983609 bytes, checksum: cbba526737c10f6743b3fd015372721a (MD5) / Made available in DSpace on 2016-06-29T12:07:11Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação Livia Araujo.pdf: 1983609 bytes, checksum: cbba526737c10f6743b3fd015372721a (MD5) Previous issue date: 2014-02-20 / CAPEs / Zymomonas mobilis vem despertando um grande interesse no meio científico, industrial e biotecnológico devido ao seu alto potencial fermentativo. Do ponto de vista taxonômico, Z. mobilis é a única espécie do gênero Zymomonas, e é subdivida em três subespécies: Z. mobilis subsp. mobilis, Z. mobilis subsp. pomaceae e Z. mobilis subsp. francensis. A diferenciação destas subespécies é baseada em testes fisiológicos. Estes testes consomem tempo e são frequentemente duvidosos. Por isso, técnicas moleculares são propostas como uma alternativa rápida e confiável para diferenciação destas bactérias. O presente estudo teve como objetivo realizar a caracterização molecular das 32 linhagens de Zymomonas mobilis depositadas na Coleção de Microrganismos UFPEDA, através da análise das sequências do gene 16S rDNA e ARDRA. As linhagens foram cultivadas em meio SSDL por 24 horas à 30º, seguida de centrifugação e extração de DNA cromossômico. As reações de PCR foram realizadas com iniciadores e condições específicas para a amplificação do gene 16S rDNA. Os produtos do gene 16S rDNA amplificados foram purificados, sequenciados e clivados com as enzimas de restrição Hae III, NdeII e StuI. Os dados obtidos pelo sequenciamento do gene 16S rDNA foram analisados, comparados e alinhados, pelo programas BLASTn e MultiAlin, com sequências de linhagens de Z. mobilis previamente depositadas no banco de dados GenBank. Um dendograma foi construido através do programa ClustalW pelo método de neighbor-joining Os perfis de restrição teórico das enzimas de restrição Hae III, NdeII e StuI foram gerados a partir do WebCutter 2.0. Dendogramas foram construídos a partir da matriz de similaridade genética de Jaccard, calculada pela análise dos perfis de restrição teóricos de cada enzima. A análise das sequências obtidas no presente estudo revelou o elevado grau de conservação no gene 16SrDNA, confirmando a relação de proximidade das linhagens de Zymomonas mobilis depositadas na Coleção de Micro-organismos UFPEDA e a aproximidade com a Z. mobilis subsp. mobilis LMG445, sugerindo que as linhagens desta coleção pertencem a esta subespécie.Além disso, conclui-se que a análise do perfil restrição teórico do gene 16S rDNA possibilita a diferenciação de Z. mobilis,a nível de subespécie, mas não é eficaz para analisar a variabilidade genética entre as linhagens de Z. mobilis UFPEDA. Baseados nestes resultados, outros marcadores filogenéticos devem ser empregados para analisar a variabilidade genética destas linhagens, possibilitando um melhor conhecimento da diversidade desta bactéria. / Zymomonas mobilis has attracted great interest in the scientific, industrial and biotechnological medium due to its high fermentation potential. The taxonomic viewpoint, Z. mobilis is the only species of the genus Zymomonas , and is subdivided into three subspecies : Z. mobilis subsp. mobilis, Z. mobilis subsp. pomaceae and Z. mobilis subsp. francensis. The differentiation of these subspecies is based on physiological tests. These tests are time consuming and often unreliable. Therefore, molecular techniques are proposed as a fast and reliable alternative to differentiation of these bacteria. This study aims to perform molecular characterization of 32 strains of Zymomonas mobilis deposited in the Collection of Microorganisms UFPEDA by sequence analysis of 16S rDNA gene and the theoretical restriction profile of this gene. The strains were grown in SSDL for 24 hours at 30 ° , followed by centrifugation and extraction of chromosomal DNA . PCR reactions were performed with primers and specific conditions for amplification of the 16S rDNA gene. The amplified products of 16S rDNA were purified, sequenced, and cleaved with restriction enzymes Hae III, NdeII and StuI . The data obtained by 16S rDNA gene were analyzed, compared and aligned by BLASTn and MultiAlin programs with sequences of strains of Z. mobilis previously deposited in the GenBank databas . A dendogram was constructed using the program ClustalW method by neighbor-joining. Profiles theoretical restriction of restriction enzyme Hae III, NdeII and StuI were generated from WebCutter 2.0. Dendrograms were constructed from the genetic Jaccard similarity matrix, calculated by analyzing the theoretical restriction profiles of each enzyme. The analysis of the sequences obtained in this study revealed the high degree of conservation in 16SrDNA gene, confirming the close relationship of strains of Zymomonas mobilis deposited in the Collection of Micro-organisms UFPEDA and closeness with Z. mobilis subsp. mobilis LMG445, suggesting that the strains in this collection belong to this subespécie. In addition, it is concluded that the theoretical restriction profile analysis of the 16S rDNA gene allows differentiation of Z. mobilis , the level of subspecies , but it is not effective to analyze the genetic variability between strains of Z. mobilis UFPEDA . Based on these results , other phylogenetic markers should be employed to analyze the genetic variability of these strains , allowing a better understanding of the diversity of this bacteria.

O filo Myxozoa compreende organismos parasitários que infectam vertebrados, principalmente peixes, em diversas regiões do Brasil e do mundo. Com mais de 2180 espécies já descritas os mixosporídeos estão entre os patógenos mais importantes que infectam peixes tanto de ambiente natural como de sistemas de criação. Neste estudo, foram realizadas coletas no ano de 2012, onde foram capturados 13 espécimes de piraputanga (Brycon hilarii) e 14 espécimes de piauçu (Leporinus macrocephalus) oriundos de pisciculturas do estado de São Paulo. Foram encontrados mixosporídeos infectando o rim de B. hilarii e o filamento branquial de L. macrocephalus. Para a identificação das espécies de parasitos, foi utilizada a microscopia de luz para a análise morfológica e, também, histopatológica para a observação das alterações decorrentes do parasitismo. A análise ultraestrutural foi realizada para verificar a interação parasita-hospedeiro do mixosporídeo encontrado na piraputanga. A análise filogenética do gene 18S rDNA avaliou a relação entre as espécies identificadas com outras espécies de mixosporídeos já descritas. Uma nova espécie de Myxobolus foi descrita neste estudo infectando o rim de B. hilarii e a caracterização molecular e filogenética de Henneguya leporinicola, encontrado infectando o filamento branquial de L. macrocephalus, foi realizada. No estudo filogenético, utilizando o método de Máxima Verossimilhança para ambas as espécies, verificou-se que as espécies de mixosporídeos se agruparam primeiro de acordo com a ordem do hospedeiro Characiforme e pelo tropismo tecidual. A análise molecular demonstrou diferenças genéticas significativas em comparação com outras espécies já descritas na América do Sul e em outros países. / The Myxozoa phylum comprises parasitic organisms that infect vertebrates, mainly fish, in many areas from Brazil and from the world. With more than 2180 species already described, myxosporeans are among the most important fish pathogens which infect both natural environment and breeding systems. In this study, captures were made in 2012, where 13 specimens of piraputanga (Brycon hilarii) and 14 specimens of piauçu (Leporinus macrocephalus) proceeding from fish farms in the state of Sao Paulo were capured. Myxosporeans were found infecting the kidney of B. hilarii and the gill filament of L. Macrocephalus. For the identification of parasite species, it was used light microscopy for morphological and histopathological analyzes to observe the damage caused by parasitism. The ultrastructural analysis was performed to verify the host-parasite myxosporean interaction found on piraputanga. The phylogenetic analysis of 18S rDNA evaluated the relation between the species identified with other myxosporean species already described. A new Myxobolus species was described in this study infecting the kidney of B. hilarii and molecular characterization and phylogenetic of Henneguya leporinicola, infecting the gill filament of L. macrocephalus, was performed. In phylogenetic study, using the method of Maximum Likelihood for both species, it was observed that the species of myxosporean grouped according to the order of the host Characiform and the tissue tropism. Molecular analysis showed significant genetic differences with other described species in South America and other countries.

Water quality in the Vaalharts region in the Northern Cape Province, South Africa, decreased over the past few years and there was a need for the microbiological and physico-chemical assessment. This problem was identified through discussions with Vaalharts Water (Vaalharts Water User Association) in 2010 when the issue of the impact of deteriorating water quality on drinking water production was raised. It was thus important to investigate concerns of the water users association pertaining to water quality issues. The aim of this study was to assess the physico-chemical and microbiological quality of household water in the Vaalharts irrigation scheme. The main residential areas were Hartswater, Pampierstad, Jan Kempdorp and Warrenton. Faecal coliforms were detected in the raw water of all the drinking water distribution systems during 2011 and 2012. No faecal coliforms were detected in the household water during 2011. This was a very positive result, because not only did the household water comply with the SANS 241 (2011) standard (0 CFU/100ml), but the purification processes were successful by removing all the E. coli’s from the raw water. However, during March 2012 faecal coliforms were detected in the household water of Jan Kempdorp (191CFU/100ml). This could be due to point pollution and possible breakage of faecal coliforms in the distribution system. Low amounts of total coliforms were detected in the raw water of some of the drinking water distribution systems. This could be due to high amounts of other colonies (pink and purple) growing on the m-Endo agar which suppress the growth of the metallic green sheen (total coliform) colonies. The total coliform numbers complied with the SANS 241 (2011) standard of ≤10 CFU/100ml at most of the distribution systems, except for Hartswater during July 2011 (14CFU/100ml) and Warrenton during March 2012 (256 CFU/100ml). Heterotrophic plate count bacteria were very high in the household water of some of the distribution systems during 2011 and 2012 which exceeded the SANS 241 (2011) standard of ≤1000 CFU/ml. A large number of pigmented (yellow, orange, pink) and non-pigmented (white) colonies were isolated on R2A agar. This can be an indication of some failure in treatment processes. Other microbiological parameters that were tested such as faceal streptococci, Clostridia, Pseudomonas aeruginosa and fungi did not indicate any danger, but there were high levels of total anaerobic bacteria in the raw water during 2011 and 2012. A high level of anaerobic bacteria was detected in the household water of Hartswater during July 2011. Clostridia were also present in the household water of some of the distribution systems during 2011 and 2012. Sequencing results of the mdh, lacZ and uidA genes indicated that one of the isolates was identified as Enterobacter cloacae and the other isolates were E. coli. Four of the isolates were identified as Escherichia coli O104:H4. This is a pathogenic strain and raised concern. The physicochemical parameters that were measured complied with the SANS 241 (2011) standards during 2011 and 2012, but some of the parameters increased gradually from 2011 to 2012. Statistical analysis indicated that physico-chemical parameters had an influence on microbiological parameters and that deteriorating raw water may have an impact on drinking water quality. Another concern currently is that there is no SANS 241 (2011) for faecal streptococci, Clostridia, Pseudomonas aeruginosa, fungi and anaerobic bacteria. These are all opportunistic pathogenic bacteria and consuming water with high levels of these bacteria may cause health problems. This study indicated good progress in the treatment processes of the distribution systems over the two years. This may be due to the feedback given to Vaalharts Water during this study regarding the water quality of the residential areas. The physico-chemical and microbiological results of the present study indicated possible biofilm formation in the distribution systems. This may have impacts on the drinking water quality of the distribution systems. It was also evident that deteriorating raw water sources may have an impact on drinking water production. / Thesis (MSc (Environmental Sciences))--North-West University, Potchefstroom Campus, 2013.

Water quality in the Vaalharts region in the Northern Cape Province, South Africa, decreased over the past few years and there was a need for the microbiological and physico-chemical assessment. This problem was identified through discussions with Vaalharts Water (Vaalharts Water User Association) in 2010 when the issue of the impact of deteriorating water quality on drinking water production was raised. It was thus important to investigate concerns of the water users association pertaining to water quality issues. The aim of this study was to assess the physico-chemical and microbiological quality of household water in the Vaalharts irrigation scheme. The main residential areas were Hartswater, Pampierstad, Jan Kempdorp and Warrenton. Faecal coliforms were detected in the raw water of all the drinking water distribution systems during 2011 and 2012. No faecal coliforms were detected in the household water during 2011. This was a very positive result, because not only did the household water comply with the SANS 241 (2011) standard (0 CFU/100ml), but the purification processes were successful by removing all the E. coli’s from the raw water. However, during March 2012 faecal coliforms were detected in the household water of Jan Kempdorp (191CFU/100ml). This could be due to point pollution and possible breakage of faecal coliforms in the distribution system. Low amounts of total coliforms were detected in the raw water of some of the drinking water distribution systems. This could be due to high amounts of other colonies (pink and purple) growing on the m-Endo agar which suppress the growth of the metallic green sheen (total coliform) colonies. The total coliform numbers complied with the SANS 241 (2011) standard of ≤10 CFU/100ml at most of the distribution systems, except for Hartswater during July 2011 (14CFU/100ml) and Warrenton during March 2012 (256 CFU/100ml). Heterotrophic plate count bacteria were very high in the household water of some of the distribution systems during 2011 and 2012 which exceeded the SANS 241 (2011) standard of ≤1000 CFU/ml. A large number of pigmented (yellow, orange, pink) and non-pigmented (white) colonies were isolated on R2A agar. This can be an indication of some failure in treatment processes. Other microbiological parameters that were tested such as faceal streptococci, Clostridia, Pseudomonas aeruginosa and fungi did not indicate any danger, but there were high levels of total anaerobic bacteria in the raw water during 2011 and 2012. A high level of anaerobic bacteria was detected in the household water of Hartswater during July 2011. Clostridia were also present in the household water of some of the distribution systems during 2011 and 2012. Sequencing results of the mdh, lacZ and uidA genes indicated that one of the isolates was identified as Enterobacter cloacae and the other isolates were E. coli. Four of the isolates were identified as Escherichia coli O104:H4. This is a pathogenic strain and raised concern. The physicochemical parameters that were measured complied with the SANS 241 (2011) standards during 2011 and 2012, but some of the parameters increased gradually from 2011 to 2012. Statistical analysis indicated that physico-chemical parameters had an influence on microbiological parameters and that deteriorating raw water may have an impact on drinking water quality. Another concern currently is that there is no SANS 241 (2011) for faecal streptococci, Clostridia, Pseudomonas aeruginosa, fungi and anaerobic bacteria. These are all opportunistic pathogenic bacteria and consuming water with high levels of these bacteria may cause health problems. This study indicated good progress in the treatment processes of the distribution systems over the two years. This may be due to the feedback given to Vaalharts Water during this study regarding the water quality of the residential areas. The physico-chemical and microbiological results of the present study indicated possible biofilm formation in the distribution systems. This may have impacts on the drinking water quality of the distribution systems. It was also evident that deteriorating raw water sources may have an impact on drinking water production. / Thesis (MSc (Environmental Sciences))--North-West University, Potchefstroom Campus, 2013.