Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgico / O presente trabalho foi conduzido no perÃodo de abril de 2006 a marÃo de 2008, nos departamentos de Zootecnia e de Biologia, da Universidade Federal do CearÃ. O objetivo desta pesquisa foi investigar, atravÃs de dados moleculares, as relaÃÃes filogenÃticas de algumas abelhas indÃgenas sem ferrÃo do tÃxon Melipona Illiger, 1806, nativas do Brasil. Procurou-se fornecer subsÃdios para facilitar uma futura revisÃo taxonÃmica sobre essas abelhas, e desse modo gerar informaÃÃes para o desenvolvimento de um criatÃrio racional, adequado Ãs diferentes espÃcies deste tÃxon, dessa forma contribuindo para o melhor aproveitamento comercial e conservaÃÃo dessas abelhas. As amostras de abelhas foram coletadas em vÃrios estados das regiÃes Norte, Nordeste e Sudeste do Brasil. Por meio da extraÃÃo, amplificaÃÃo e seqÃenciamento parcial da regiÃo ITS1 do DNA ribossÃmico nuclear dessas amostras, somadas Ãs seqÃÃncias parciais da regiÃo ITS1 de outras abelhas do mesmo tÃxon retiradas do GenBank, pÃde-se verificar os seguintes aspectos: alinhamento mÃltiplo, composiÃÃo nucleotÃdica, matriz de distÃncia genÃtica e reconstruÃÃo filogenÃtica entre as mesmas. Os resultados mostraram que o alinhamento mÃltiplo produziu uma interseÃÃo central com o comprimento de apenas 141 pb e a mÃdia da distÃncia genÃtica entre todas as seqÃÃncias estudadas do tÃxon Melipona foi de 7,6%. As Ãrvores construÃdas usando algoritmos baseados nos mÃtodos de agrupamento do vizinho mais prÃximo (NJ), mÃxima parcimÃnia (MP) e mÃxima verossimilhanÃa (MV) para as seqÃÃncias parciais da regiÃo pesquisada mostraram essencialmente a mesma topologia, sendo esta bem definida em trÃs grandes clados: Clado 1- contendo as sequÃncias de M. subnitida, M. quadrifasciata e M. mandacaia (todas pertencendo ao subgÃnero Melipona); Clado 2 â abrangendo as seqÃÃncias de M. quinquefasciata e M. fasciculata (ambas pertencendo ao subgÃnero Melikerria); Clado 3 â tendo como representantes no presente trabalho, as seqÃÃncias de M. mondury, M. flavolineata e M. scutellaris (todas pertencentes ao subgÃnero Michmelia). Todas as trÃs Ãrvores filogenÃticas foram capazes de recuperar a monofilia tanto do gÃnero Melipona como tambÃm a dos trÃs clados, que apareceram como grupos monofilÃticos / The present work was carried out from April 2006 to March 2008, in the departments of Biology and Animal Science in the Universidade Federal do CearÃ. The aim of this research was to investigate, through molecular data, the phylogenetic relationships among some Brazilian stingless bee species belonging to the taxon Melipona Illiger, 1806. It was attempted to obtain information that could facilitate a taxonomic revision of this bee group in the future and to generate information useful to the development of rational rearing adequate to the distinct species of this taxon, contributing to a better commercial exploitation and conservation of these bee species. Bee samples were collected in various states of the North, Northeast and Southeast regions of Brazil. Through the extraction, amplification and partial DNA sequencing of the ITS1 region of nuclear ribosomal DNA of those samples and the partial sequences of the ITS1 region of other bees of the same taxon searched in the GenBank, it was possible to observe the following parameters: multiple alignment, nucleotide composition, matrixes of genetic distances and phylogenetic reconstruction. Results showed that multiple alignment resulted produced a central intersection of only 141 bp long and the average of the genetic distance among all the studied sequences of the taxon Melipona was of 7,6%. The phylogenetic trees built using algorithms based on the methods of the Neighbor-Joining (NJ), Maximum Parsimony (MP) and Maximum Likelihood (ML) for the partial sequences showed essentially the same topology, which was clearly distinct in three great clados: Clado 1 - contained the sequences of M. subnitida, M. quadrifasciata and M. mandacaia (all belonging to the subgenus Melipona); Clado 2 - included the sequences of M. quinquefasciata and M. fasciculata (both belonging to the subgenus Melikerria); and Clado 3 â represented in this work by the sequences of M. mondury, M. flavolineata and M. scutellaris (all belonging to the subgenus Michmelia). The three phylogenetic trees were capable to recover the monophyly of the genus Melipona as well as of the three clados, that appeared as monophyletic groups

BOMFIM, Isac Gabriel Abrahão. Relações filogenéticas de abelhas indígenas sem ferrão do táxon Melipona Illiger, 1806 (Apidae: Meliponina) baseadas em seqüências parciais da região its1 do DNA ribossômico nuclear. 2008. 97 f. Dissertação (Mestrado em zootecnia)- Universidade Federal do Ceará, Fortaleza-CE, 2008. / Submitted by Elineudson Ribeiro (elineudsonr@gmail.com) on 2016-05-17T17:48:21Z No. of bitstreams: 1 2008_dis_igabomfim.pdf: 1036482 bytes, checksum: eed11b61d455adc337c8c2475055b894 (MD5) / Approved for entry into archive by José Jairo Viana de Sousa (jairo@ufc.br) on 2016-05-27T19:59:42Z (GMT) No. of bitstreams: 1 2008_dis_igabomfim.pdf: 1036482 bytes, checksum: eed11b61d455adc337c8c2475055b894 (MD5) / Made available in DSpace on 2016-05-27T19:59:42Z (GMT). No. of bitstreams: 1 2008_dis_igabomfim.pdf: 1036482 bytes, checksum: eed11b61d455adc337c8c2475055b894 (MD5) Previous issue date: 2008 / The present work was carried out from April 2006 to March 2008, in the departments of Biology and Animal Science in the Universidade Federal do Ceará. The aim of this research was to investigate, through molecular data, the phylogenetic relationships among some Brazilian stingless bee species belonging to the taxon Melipona Illiger, 1806. It was attempted to obtain information that could facilitate a taxonomic revision of this bee group in the future and to generate information useful to the development of rational rearing adequate to the distinct species of this taxon, contributing to a better commercial exploitation and conservation of these bee species. Bee samples were collected in various states of the North, Northeast and Southeast regions of Brazil. Through the extraction, amplification and partial DNA sequencing of the ITS1 region of nuclear ribosomal DNA of those samples and the partial sequences of the ITS1 region of other bees of the same taxon searched in the GenBank, it was possible to observe the following parameters: multiple alignment, nucleotide composition, matrixes of genetic distances and phylogenetic reconstruction. Results showed that multiple alignment resulted produced a central intersection of only 141 bp long and the average of the genetic distance among all the studied sequences of the taxon Melipona was of 7,6%. The phylogenetic trees built using algorithms based on the methods of the Neighbor-Joining (NJ), Maximum Parsimony (MP) and Maximum Likelihood (ML) for the partial sequences showed essentially the same topology, which was clearly distinct in three great clados: Clado 1 - contained the sequences of M. subnitida, M. quadrifasciata and M. mandacaia (all belonging to the subgenus Melipona); Clado 2 - included the sequences of M. quinquefasciata and M. fasciculata (both belonging to the subgenus Melikerria); and Clado 3 – represented in this work by the sequences of M. mondury, M. flavolineata and M. scutellaris (all belonging to the subgenus Michmelia). The three phylogenetic trees were capable to recover the monophyly of the genus Melipona as well as of the three clados, that appeared as monophyletic groups. / O presente trabalho foi conduzido no período de abril de 2006 a março de 2008, nos departamentos de Zootecnia e de Biologia, da Universidade Federal do Ceará. O objetivo desta pesquisa foi investigar, através de dados moleculares, as relações filogenéticas de algumas abelhas indígenas sem ferrão do táxon Melipona Illiger, 1806, nativas do Brasil. Procurou-se fornecer subsídios para facilitar uma futura revisão taxonômica sobre essas abelhas, e desse modo gerar informações para o desenvolvimento de um criatório racional, adequado às diferentes espécies deste táxon, dessa forma contribuindo para o melhor aproveitamento comercial e conservação dessas abelhas. As amostras de abelhas foram coletadas em vários estados das regiões Norte, Nordeste e Sudeste do Brasil. Por meio da extração, amplificação e seqüenciamento parcial da região ITS1 do DNA ribossômico nuclear dessas amostras, somadas às seqüências parciais da região ITS1 de outras abelhas do mesmo táxon retiradas do GenBank, pôde-se verificar os seguintes aspectos: alinhamento múltiplo, composição nucleotídica, matriz de distância genética e reconstrução filogenética entre as mesmas. Os resultados mostraram que o alinhamento múltiplo produziu uma interseção central com o comprimento de apenas 141 pb e a média da distância genética entre todas as seqüências estudadas do táxon Melipona foi de 7,6%. As árvores construídas usando algoritmos baseados nos métodos de agrupamento do vizinho mais próximo (NJ), máxima parcimônia (MP) e máxima verossimilhança (MV) para as seqüências parciais da região pesquisada mostraram essencialmente a mesma topologia, sendo esta bem definida em três grandes clados: Clado 1- contendo as sequências de M. subnitida, M. quadrifasciata e M. mandacaia (todas pertencendo ao subgênero Melipona); Clado 2 – abrangendo as seqüências de M. quinquefasciata e M. fasciculata (ambas pertencendo ao subgênero Melikerria); Clado 3 – tendo como representantes no presente trabalho, as seqüências de M. mondury, M. flavolineata e M. scutellaris (todas pertencentes ao subgênero Michmelia). Todas as três árvores filogenéticas foram capazes de recuperar a monofilia tanto do gênero Melipona como também a dos três clados, que apareceram como grupos monofiléticos.

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2016-08-09T23:35:51Z No. of bitstreams: 1 VirginiaPenellopeMacedoESilva_TESE.pdf: 2028754 bytes, checksum: 8e6e495b2056248d59d55bf24abbf089 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2016-08-15T22:34:21Z (GMT) No. of bitstreams: 1 VirginiaPenellopeMacedoESilva_TESE.pdf: 2028754 bytes, checksum: 8e6e495b2056248d59d55bf24abbf089 (MD5) / Made available in DSpace on 2016-08-15T22:34:21Z (GMT). No. of bitstreams: 1 VirginiaPenellopeMacedoESilva_TESE.pdf: 2028754 bytes, checksum: 8e6e495b2056248d59d55bf24abbf089 (MD5) Previous issue date: 2015-04-23 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior (CAPES) / Nas Am?ricas, a infec??o por Leishmania tem como principal agente etiol?gico Leishmania infantum. Nos ?ltimos 30 anos o padr?o de distribui??o das leishmanioses tem mudado substancialmente e a doen?a tem apresentado um perfil emergente na periferia dos grandes centros urbanos. A infec??o por Leishmania pode evoluir com um amplo espectro cl?nico desde o acometimento da pele, mucosas e v?sceras. Dos indiv?duos infectados por L. infantum apenas 10% desenvolvem a doen?a, sabe-se que 90% da infec??o humana ? assintom?tica e diversos fatores est?o envolvidos no curso da infec??o. Os principais fatores envolvidos no desenvolvimento da doen?a s?o a resposta imune do hospedeiro, a esp?cie e o conte?do g?nico do parasita. O sequenciamento dos isolados de Leishmania poderia aumentar a compreens?o acerca da sintomatologia dos indiv?duos. Dessa forma, o objetivo desse estudo foi avaliar a diversidade gen?tica de cepas de Leishmania circulantes entre humanos, sintom?ticos e assintom?ticos e c?es de ?reas end?micas do Rio Grande do Norte. A variabilidade gen?tica de um grupo de amostras de humanos e caninos com a doen?a visceral, doen?a cut?nea e infec??o assintom?tica foi avaliado com os marcadores moleculares hsp70 e ITS1. O genoma completo de 20 isolados de Leishmania oriundos de humanos, sintom?ticos e assintom?ticos, e c?es foram sequenciados para avaliar a diversidade dessas amostras. Os fragmentos amplificados de hsp70 e ITS1 das amostras e foram analisados e montadas utilizando o pacote Phred/Phrap. Os dendogramas foram constru?dos aplicando o m?todo neighbor joining com 500 bootstraps, seguido das infer?ncias sobre as rela??es entre as variantes de Leishmania. As sequ?ncias dos 20 isolados brasileiros foram mapeadas com o genoma de refer?ncia Leishmania infantum JPCM5, usando o programa Bowtie2, com identifica??o de 36 contigs. As informa??es dos SNPs v?lidos foram utilizadas na PCA. Os SNPs foram visualizados pelo Geneious 7.1 e IGV. As anota??es do genoma foram ent?o transferidas para seus respectivos cromossomos e visualizadas utilizando o Geneious. As sequ?ncias consenso de todos os cromossomos (com m?nimo de 75% das reads com a mesma base) foram alinhadas usando Mauve. As ?rvores filogen?ticas foram calculadas de acordo com c?lculos de m?xima verossimilhan?a e modelos JTT. Como resultados obtivemos que hsp70 e ITS1 n?o foram capazes de definir variabilidade gen?tica entre os isolados de humanos e c?es; nem para os isolados de cultura e de sangue perif?rico, oriundos de um mesmo paciente.O sequenciamento gen?mico dos 20 isolados brasileiros revelou uma forte rela??o entre as cepas de Leishmania circulantes em no Rio Grande do Norte. Os isolados da Grande Natal de humanos e caninos permaneceram agrupados em todas as an?lises, sugerindo que existe proximidade genot?pica e geogr?fica entre os isolados. As amostras isoladas na d?cada de 1990 apresentaram uma maior diversidade genot?pica quando comparadas as amostras recentemente isoladas. De forma geral, n?o encontramos correla??o entre as formas cl?nicas sintom?ticas e assintom?ticas e o conte?do g?nico dos isolados brasileiros de Leishmania. / Leishmania infantum is the main etiologic agent of visceral leishmaniasis in the New World. The pattern of distribution of leishmaniasis has changed substantially and has presented an emerging profile within the periphery of the Large Urban Centers. Leishmania infection can compromise skin, mucosa and viscera. Only 10% of the individuals infected develop the disease and 90% of human infection is asymptomatic. The main factors involved in the development of the disease are the host immune response, the vector?s species and the parasite?s genetic content. The sequencing of Leishmania isolated seeks to increase the understanding of the symptoms of individuals. The aim of this study was to evaluate the genetic diversity of circulating Leishmania strains among humans, and symptomatic and asymptomatic, and dogs from endemic areas of Rio Grande do Norte State and analyze sandflies from endemic areas for cutaneous and visceral disease. The genetic variability was evaluated by the use of markers hsp70 , ITS1 and a whole genome sequencing was also carried out. The amplified hsp70 and ITS1 of samples were analyzed and assembled using a Phred / Phrap package. The dendograms were constructed using the same methodology, but adding 500 bootstraps, followed by inferences on the relationships between Leishmania variants. The sequences of the 20 Brazilian isolates were mapped to the reference genome L. infantum JPCM5, using the Bowtie2 program and the identification of 36 contigs. The information of the valid SNPs were used in the PCA. SNPs were visualized by Geneious 7.1 and IGV. The genome annotations were transferred to their respective chromosomes and displayed on Geneious. The matching sequences of all chromosomes were aligned using Mauve. The phylogenetic trees were calculated according to maximum likelihood and JTT models. Sandflies were analyzed by PCR for the identification of Leishmania infection, a blood meal source and GAPDH sand fly. As a result, hsp70 and ITS1 were not capable of identifying genetic variability among human isolates from symptomatic and asymptomatic, and dogs. The complete sequencing of the 20 Brazilian isolates revealed a strong similarity between the circulating Leishmania strains in Rio Grande do Norte. The isolates collected in the city of Natal from humans and canines remained grouped in all analyzes, suggesting that there is genotypic and geographic proximity among the isolates. The isolated samples in the 1990s had a higher genotypic diversity when compared to freshly isolated samples. All isolates presented 36 chromosomes with variable ploidy among them, no correlation was found between the number of amastina genes copies, gp63, A2 and SSG with such clinic forms. In general, we did not find correlation between symptomatic and asymptomatic clinical forms and the gene content of the Brazilian isolates of Leishmania. 34,28% of the sandflies collected in the upper west region were L. longipalpis and the main sources of blood meal were humans, dogs and chickens.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The Pampa is the newest Brazilian biomes. It is one of the lest studied and the one of the most economically exploited. Among these forms of exploitation, there is the cultivation of great dominated grassy fields with exotic forest species. Researches involving animal and plant species have revealed great biodiversity, however, they are still rare for microorganisms. The aim of the study was to collect, to isolate and to analyze morphologically and molecularly three wood-decaying fungi growing on Pinus sp. in Pampa biome: Pseudomerulius curtisii (Berk.) Redhead & Ginns, Gelatoporia subvermispora (Pilat) Niemelä and Sarcoporia polyspora P. Karsten. It was conducted the morphological characterization and the sequencing of ITS1-5.8S-ITS2 of species allowing their identification and phylogenetic comparisons with the literature. P. curtisii calls attention to the rarity and beauty of the basidiome and morphological similarities with specimens of the Northern Hemisphere and Oceania, but with a genetic distance, and may show the geographic isolation and the new taxon to be described. G. subvermispora, widely studied by the high biotechnology and industrial potential. It has high similarity with morphogenetic U.S. specimens, but with genetic divergence of Eurasian specimens. S. polyspora shows high morphological similarity with the specimens of the northern hemisphere, but there is a lack of sequences available for phylogenetic comparison. These species were registered for the first time in South America. P. curtisii, G. subvermispora and S. polyspora, fungi Basidiomycota show high potential for nutrient cycling and soil formation by the capacity or lignin and cellulose degradation. / O Pampa é o mais novo dos biomas brasileiros, sendo o menos estudado e um dos mais explorados comercialmente. Dentre estas formas de exploração, destaca-se o plantio de grandes áreas com espécies florestais exóticas. Os estudos envolvendo espécies animais e vegetais têm revelado grande biodiversidade, mas ainda são escassos para microrganismos. O objetivo deste trabalho foi coletar, isolar e analisar morfológica e molecularmente três fungos decompositores de madeira de Pinus sp. no bioma Pampa: Pseudomerulius curtisii (Berk.) Redhead & Ginns, Gelatoporia subvermispora (Pilát) Niemelä e Sarcoporia polyspora P. Karsten. Efetuou-se a caracterização morfológica e o sequenciamento da região ITS1-5.8S-ITS2 das espécies, permitindo suas identificações e comparações filogenéticas com a literatura. P. curtisii chama atenção pela raridade e beleza dos basidiomas e similaridade morfológica com os espécimes do Hemisfério Norte e Oceania, mas com uma distância genética, podendo mostrar o isolamento geográfico e novo taxon a ser descrito. G. subvermispora, amplamente estudado pelo elevado potencial biotecnológico e industrial, apresenta alta similaridade morfogenética com espécimes dos EUA, porém com divergência genética dos espécimes da Eurásia. S. polyspora apresenta alta similaridade morfológica com os espécimes do hemisfério Norte, mas há falta de sequências disponíveis para comparação filogenética. Essas espécies foram registradas pela primeira vez na América do Sul. P. curtisii, G. subvermispora and S. polyspora (Basidiomycota) apresentam grande potencial para estudos de ciclagem de nutrientes e formação do solo pela capacidade de degradação de lignina e/ou celulose.

This study applies molecular and morphological approaches addressing the identification of morphologically similar larval stages (cercariae) of Plagiorchis spp. (Digenea: Plagiorchiidae) parasitising lymnaeid snail populations in the freshwater ecosystems of central Europe. Five morphologically homogeneous and genetically distinct lineages of Plagiorchis spp. were identified via matching molecular data for the mitochondrial cox1 gene with detailed morphometric data. Phylogenetic and comparative sequence analyses using partial 28S rDNA and ITS1-5.8S-ITS2 sequences allowed molecular identification of three species (P. elegans, P. maculosus and P. koreanus) via matching sequences from larval and adult digenean stages. A key for the identification of the cercariae of Plagiorchis spp. parasitising lymnaeid populations in central Europe is provided.

No description available.

Le genre Pneumocystis désigne un groupe de champignons opportunistes présentant une étroite spécificité d'hôte. Il détermine lors d'immunodépression sévère une infection pulmonaire grave, la pneumonie à Pneumocystis (PPC). La transmission de Pneumocystis par voie aérienne d'un hôte développant une PPC à un hôte susceptible a été démontrée à l'aide des modèles murins. Les travaux menés chez la souris ont montré également que des sujets immunocompétents colonisés par Pneumocystis murina peuvent transmettre le champignon à des souris immunodéprimées qui développeront une PPC ultérieurement. Les individus colonisés par Pneumocystis sp., ainsi que ceux développant une PPC, participeraient au réservoir du champignon. La survenue de cas groupés de PPC en milieu hospitalier est en faveur de la transmission interindividuelle de Pneumocystis jirovecii (P.jirovecii) chez l’homme. La détection de l'ADN de P.jirovecii dans l'air exhalé par les patients développant une PPC suggère que cette transmission se fait par voie aérienne. La caractérisation des populations infectées par P.jirovecii et la caractérisation génotypique du champignon au sein de son réservoir humain constituent la base de ce travail de recherche. Nous avons montré que la prévalence de la colonisation par P.jirovecii est faible chez les patients atteints de mucoviscidose et suivis dans notre CHU. La participation de ces patients au réservoir de P.jirovecii à Brest serait donc marginale. Cette faible prévalence pourrait être le reflet d'une faible circulation du champignon dans les communautés humaines dans notre région. Nous avons évalué le dosage du ß-1,3-D glucane sérique pour dépister les populations infectées. Ce dosage couplé à la détection de P.jirovecii dans les prélèvements respiratoires par la microscopie et la PCR, permet de différencier les patients développant une PPC et les patients présentant une colonisation pulmonaire par P.jirovecii. De plus, les premières données sur le ß-1,3-D glucane au cours de la primo-infection chez le nourrisson ont été obtenues.En termes de caractérisation de P.jirovecii dans notre région, l'analyse du locus dihydropteroate synthase (DHPS) a montré que: i) le lieu habituel de résidence plutôt que le lieu de diagnostic de l’infection à P.jirovecii serait un facteur prédictif d’infection par un mutant, ii) P.jirovecii pourrait circuler en France d’une région à une autre via des voyageurs infectés, iii) la prévalence de mutants potentiellement résistants chez les patients vivant effectivement à Brest était de 0%. L'analyse des séquences des "internal transcribed spacers" (ITS) 1 et 2 de P. jirovecii conforte l'hypothèse que les patients développant une PPC et les patients colonisés sont infectés par des populations fongiques présentant des caractéristiques identiques. Tous les patients, quelle que soit la présentation clinique de leur infection, constitueraient un réservoir unique et commun de P.jirovecii. Les travaux de génotypage ont constitué l'étape préalable nécessaire à l'analyse de cas groupés d'infections à P.jirovecii survenus chez des patients transplantés rénaux au CHU de Brest. Nous avons apporté des données originales sur le rôle des patients colonisés en tant que source potentielle de P. jirovecii dans un contexte d'acquisition et de transmission nosocomiales du champignon. Par ailleurs, la concordance partielle ou complète des génotypes ITS et DHPS dans les couples "prélèvements d'air–LBA" réalisés chez des patients développant une PPC est compatible avec l’exhalation du champignon et sa diffusion aérienne dans l’environnement hospitalier. Ces données apportent des arguments pour l'application de mesures de prévention des infections nosocomiales à P. jirovecii. Les précautions "gouttelettes" recommandées par la Société Française d'Hygiène Hospitalière devraient être appliquées a minima aux patients développant une PPC. Nous proposons leur extension aux patients colonisés par le champignon. / The genus Pneumocystis represents a group of opportunistic fungi that show strong host specificity. It is the cause of severe pneumonia (Pneumocystis Pneumonia [PCP]) in immunocompromised subjects. Pneumocystis transmission from a host with PCP to another susceptible host via the airborne route has been demonstrated in rodent models. Moreover, it has been established that Pneumocystis murina can be transmitted from immunocompetent mice, transiently colonized by the fungus, to immunocompromised susceptible mice that subsequently develop PCP. Colonized subjects and those developing PCP may be part of the fungus reservoir. Reports of PCP case cluster in hospital strongly suggest that Pneumocystis jirovecii (P.jirovecii) transmission in humans may also occur. P.jirovecii DNA detection in the air surrounding PCP patients is consistent with the transmission of P.jirovecii via the airborne route.Our goals were to characterize human populations infected with P.jirovecii and to characterize P.jirovecii within its human reservoir. We showed that P.jirovecii was rarely involved in pulmonary colonization in patients with cystic fibrosis monitored in the Brest Hospital. Thus this patient population was not part of the human reservoir of the fungus in our region (Brittany, Western France). This low prevalence of colonization may reflect a low level of P.jirovecii circulation within human communities in Brittany. In order to improve the identification of patients infected with P.jirovecii, we evaluated ß-1,3-D glucan detection in serum samples. We showed that serum ß-1,3-D glucan levels combined with P.jirovecii detection in pulmonary samples using microscopic examination and a PCR assay make it possible to distinguish between PCP and pulmonary colonization. Moreover the first data on ß-1,3-D glucan levels during primary infection were obtained.In order to characterize P.jirovecii in our region, we performed the typing of P.jirovecii isolates from infected patients monitored at Brest hospital, using the dihydropteroate synthase (DHPS) and the internal transcribed spacer (ITS) 1 and 2 locus analysis. DHPS typing showed that i) the usual city of patient residence rather that the city in which the diagnosis of P.jirovecii infection has been made is a predictor of mutants, ii) mutants can be imported from one region to another through infected visitors, iii) the prevalence of mutants potentially resistant to sulfonamides was 0% in patients who effectively lived in the Brest geographic area. Results of ITS analysis in PCP patients and colonized patients are consistent with the hypothesis that these 2 patient groups are infected with similar P.jirovecii populations. All infected patients, whatever their clinical presentation, may be part of a common and unique reservoir of the fungus. We investigated an outbreak of P.jirovecii infections in 18 renal transplant recipients using the same typing method combined with patient encounter analysis. The results provided evidence of the role of colonized patients as potential sources of P.jirovecii. The same typing method was applied to pairs of pulmonary samples and room air samples of PCP patients. Full or partial matches of P.jirovecii types in pulmonary and air sample pairs were observed. These results are consistent with P.jirovecii exhalation by PCP patients in their close environment. These data support arguments for applying droplet precautions, at least to PCP patients, to prevent P.jirovecii transmission, as recommended by the "Société française d'hygiène hospitalière". We suggest extending droplet precautions to colonized patients to achieve the prevention of P.jirovecii nosocomial infections.

Green algae are quite important primary producers in fresh waters. The genus Chlorella represents one of algae most frequently utilized in algal biotechnologies to produce biomass, using either autotrophic or heterotrophic cultivation systems. It is than exploited as a food supplement for humans or animals. However, particular species within the genus are morphologically indistinguishable and molecular markers should be used to characterize production strains. This work is aimed to molecularly characterize three production strains of Chlorella for patent protection purposes and to specify their phylogenetic and taxonomic position.

The study of fungal diversity may lead to many fundamental discoveries and conclusions. Molecular genetics, and particularly high throughput sequencing methods using short DNA fragments as barcodes, has recently experienced a boom. The most frequently used marker for fungal research is the partial region of nuclear ribosomal DNA called ITS (Internal Transcribed Spacer). It occurs in the form of tandem repetitions of up to 200 copies. This fact greatly simplifies its amplification from the environment but also introduces some negatives. One of them can be an existence of intragenomic and intraspecific variability which confounds diversity estimates by exaggerating the real number of species. Using alternative low-copy markers can easily prevent these problems. In this study EF-1α and RPB2 protein- coding genes were compared with traditionally used ITS1 and ITS2 markers. An artificial mock community was created by blending genomic DNA of different fungal lineages. The community was sequenced for all markers and the data were processed according to guidelines commonly used in environmental studies. The results show that ITS2 is unequivocally a more suitable marker for environmental studies than other compared markers. The average coefficient of overestimation was deemed to be approximately two for ITS1, ITS2,...

In der vorliegenden Arbeit wurde die Sensitivität des Nachweises von Leishmanien in Giemsa-gefärbten Bioptaten aus Hautulzerationen mittels direkter Mikroskopie mit der Sensitivität der ITS1-PCR verglichen. Bei der ITS1-PCR wurde eine Sensitivität von 87 % mit einem positiven predictive value von 100 %, sowie eine Spezifität von 100 % mit einem negativen predictive value von 85 % nachgewiesen. Weiterhin wurden vier verschiedene Nachweismethoden miteinander verglichen: die in vitro Kultivierung in NNN Medium, die direkte Mikroskopie von Giemsa gefärbten Hautbioptaten, die PCR Amplifizierung der ITS1 Region aus auf Filterpapier aufgetragenen Hautbioptaten (FP) sowie die ITS1-PCR von ungefärbten Hautbioptaten (US). Die PCR der US erwies sich als die sensitivste Methode. Die Verbreitung von Leishmanien Arten in Jericho wurde mittels molekularer Epidemiologie untersucht. Die räumliche (Spatial) Analyse zeigte drei statistisch relevante Cluster innerhalb der kutanen Leishmaniose (CL): ein Cluster mit L. major und zwei L. tropica Cluster. Bei der Raum-Zeit–Analyse wurden vier Cluster von Kutanen Leishmaniose, zwei L. major und drei L. tropica Cluster nachgewiesen. Insgesamt 106 Stämme, die aus verschiedenen endemischen Regionen in Zentralasien, im Nahen Osten und Afrika stammen, wurden mit 10 Mikrosatellitenmarkern untersucht. Die Auswertung erfolgte über zwei Analysemethoden: die Distanz-basierte und die Modell-basierte Methode. Anhand der L. major Genomsequenz wurden PCR-Primer zur Amplifizierung von Mikrosatellitenloci von L. major entwickelt, die auf den Chromosomen 1, 3, 5, 21 und 35 liegen. Sieben unterschiedliche L. major Populationen einschließlich zweier genetisch isolierter Populationen im Nahen Osten wurden mit diesen Markern nachgewiesen. / In this study we compared the sensitivity of the diagnosis of Giemsa-stained skin scrapings by standardized graded direct microscopy with that of ITS1-PCR. ITS1-PCR showed a sensitivity of 87% with positive predictive value of 100% and a specificity of 100% with negative predictive value of 85%. In-vitro cultivation using NNN medium and direct smear microscopy of Giemsa-stained slides, PCR amplifying region 1 of internal transcribed spacer (ITS1) using skin scrapings spotted on filter papers (FP) and unstained tissue smears (US) were compared. PCR using US was more sensitive than all other methods Molecular epidemiology was used to study the distribution of Leishmania species in Jericho. Spatial analysis showed three statistically significant clusters of CL, one cluster for L. major and two clusters for L. tropica. In the case of space-time, four clusters for CL, two for L. major and three for L. tropica were detected. A total of 106 strains isolated in different endemic regions of Central Asia, Middle East and Africa were analysed using 10 pairs of microsatellite markers under two cluster methods: distance and model-based. Markers were designed to amplify microsatellite loci identified in the genome sequence of L. major on chromosomes 1, 3, 5, 21 and 35. Seven discrete populations of L. major including two genetically isolated populations in the Middle East were revealed.