Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Staphylococcus aureus resistente à meticilina (MRSA) é um importante patógeno pulmonar em pacientes com fibrose cística (FC). Caracteriza-se pela resistência a todos os β-lactâmicos, devido a presença do elemento genético móvel SCCmec o qual abriga o gene mecA. Além disso, é reconhecido por vários fatores de virulência o qual destacamos a toxina Panton-Valentine Leukocidin (PVL), uma citolisina formadora de poros na célula hospedeira, e por apresentar diversos clones epidêmicos envolvidos em surtos hospitalares. O objetivo desse estudo foi caracterizar a epidemiologia de MRSA, isolados de pacientes com FC referente a dois centros de referência no Rio de Janeiro a partir da aplicação de técnicas fenotípicas e genotípicas. Um total de 57 amostras de MRSA foi submetido ao teste de difusão em ágar para 11 antimicrobianos a fim de avaliar perfil de resistência, com aplicação da técnica da PCR foi tipificado o SCCmec e investigado a presença do gene LukS-PV responsável pela codificação da toxina PVL com intuito de estabelecer uma melhor caracterização epidemiológica dos clones identificados pela técnica do MLST (Multilocus Sequence Typing). Os antimicrobianos não β-lactâmicos apresentaram um percentual de resistência abaixo de 50%, em que destacamos a eritromicina com o maior percentual 45,6% e quanto ao perfil de resistência 24,6% foram multirresistentes. Com exceção do SCCmec II, os outros tipos foram encontrados (I, III, IV e V) com os respectivos percentuais de 22,8% (n=13), 7,1% (n=4), 61,4% (n=35) e 3,5% (n=2) e apenas 5,3% (n=3) das amostras não foram caracterizadas, não há dados da prevalência do SCCmec IV. Vinte (35,1%) amostras apresentaram produtos de amplificação compatível com a presença do gene lukS, aproximadamente metade dessas amostras (55%) estava correlacionada ao SCCmec IV. Com a análise do MLST, obtivemos os STs 1 (n=1, 1,7%), 5 (n=28, 49,1%), 30 (n=11, 19,3%), 72 (n=1, 1,7%), 398 (n=1, 1,7%), 1635 (n=7, 12,3%), 1661 (n=2, 3,5%), 239 (n=5, 8,8%), e ainda identificamos um novo ST (2732) presente em 1 amostra. A partir de uma análise associativa entre o MLST e o SCCmec foi possível observar a presença de linhagens características de clones epidêmicos, como o UK-EMRSA-3 (ST5, SCCmec I), USA 800/pediátrico (ST5, SCCmec IV), Oceania Southwest Pacific Clone - OSPC (ST30, SCCmec IV) e Brazilian Epidemic Clone - BEC (ST239, SCCmec III). Em conclusão este estudo é o primeiro a caracterizar linhagens epidêmicas de MRSA nos centros de atendimento a pacientes com FC no Rio de Janeiro, sendo necessário um monitoramento constante a fim de evitar a disseminação desses clones. / Methicillin-resistant Staphylococcus aureus (MRSA) is a major pulmonary pathogen in patients with cystic fibrosis (CF). It is characterized by resistance to all β-lactam antibiotics due to the presence of the mobile genetic element SCCmec which harbors the mecA gen. Furthermore, MRSA is recognized by several virulence factors, such as the toxin Panton-Valentine Leukocidin (PVL), pore-forming cytolysin in the host cell, and produces various epidemic clones involved in hospital outbreaks. The aim of this study was to characterize the epidemiology of MRSA, using phenotypic and genotypic methods of isolates from CF patients from two reference centers in Rio de Janeiro. A total of 57 MRSA isolates were tested by the Agar diffusion test for 11 antibiotics. SCCmec and the presence of the Luks-PV gene, responsible for encoding the PVL toxin, were evoluted by PCR, in order to establish a better epidemiological clone characterization by MLST (Multilocus Sequence Typing) technique. Non-β-lactam antimicrobials showed less than 50% of resistance, which included erythromycin with the highest percentage was 45.6%, beside, multirresistant profile was observed in 24.6% of isolates. We found SCCmec types I, III, IV and V with the corresponding percentage of 22.8% (n = 13), 7.1% (n = 4), 61.4% (n = 35) and 3.5% (n = 2) respectively and just 5.3% (n = 3) isolates were not typified. SCCmec II was not detected among our isolates. Twenty (35.1%) isolates showed amplification products consistent with the presence of the lukS gen, approximately half of these samples (55%) were correlated with SCCmec IV. Using MLST analysis, we obtained STs 1 (n = 1, 1.7%), 5 (n = 28, 49.1%), 30 (n = 11, 19.3%), 72 (n = 1, 1.7%), 398 (n = 1, 1.7%), 1635 (n = 7, 12.3%), 1661 (n = 2, 3.5%), 239 (n = 5, 8, 8%), and further identified a new ST (2732) present in one isolate. Associating MLST and SCCmec, it was possible to observe the presence of epidemic clones, such as, UK-EMRSA-3 (ST5, SCCmec I), USA800/pediatric (ST5, SCCmec IV), Oceania Southwest Pacific Clone - OSPC (ST30, SCCmec IV) and Brazilian Epidemic Clone - BEC (ST239, SCCmec III). In conclusion this study is the first one to characterize epidemic strains of MRSA in care centers of CF patients in Rio de Janeiro, that require constant monitoring in order to prevent the spread of these clones.

Quarenta e nove amostras EPEC típica (tEPEC) e atípica (aEPEC) pertencentes a diferentes sorotipos, isoladas de humanos e animais (cães, gatos, bovinos, ovinos, coelhos e sagüis) foram investigadas quanto ao perfil de virulência pela PCR e similaridade clonal por Multilocus Sequence Typing (MLST) e Pulsed-Field Gel Electrophoresis (PFGE). O objetivo deste estudo foi verificar se animais atuam como reservatório e fonte infecção de aEPEC para humanos. Os marcadores de virulência analisados revelaram que cepas aEPEC isoladas de animais possuem potencial para causar diarréia em humanos. As técnicas MLST e PFGE revelaram que amostras isoladas de animais e humanos compartilham relações clonais próximas ou idênticas. Estes resultados indicam que os animais estudados atuam como reservatório de aEPEC e representam fonte de infecção para humanos. Pelo fato de humanos, também atuarem como reservatório de aEPEC, ciclos de infecção cruzada animal-humano não podem ser descartados, pois a dinâmica de transmissão entre reservatórios de aEPEC não é muito bem compreendida. / Forty-nine typical and atypical EPEC strains belonging to different serotypes, isolated from humans, pets (cats and dogs), farm (bovines, sheep and rabbits) and wild animals (monkeys) were investigated for virulence markers and clonal similarity by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). The virulence markers analyzed revealed that atypical EPEC strains isolated from animals have the potential to cause diarrhea in humans. Close clonal relationship between human and animal isolates was found with MLST and PFGE. These results indicate that these animals act as atypical EPEC reservoirs and may represent sources of infection for humans. Since humans also act as a reservoir of atypical EPEC strains, the cycle of mutual infection of atypical EPEC between animals and humans, mainly pets and their owners, cannot be ruled out, since the transmission dynamics between the reservoirs are not yet clearly understood.

O gênero Yersinia compreende 12 espécies. Y. enterocolitica, Y. pseudotuberculosis e Y. pestis são patógenos de vários animais, incluindo os humanos. Y. aldovae, Y. bercovieri, Y. frederiksenii, Y. intermedia, Y. kristensenii, Y. aleksiciae, Y. mollaretti e Y. rhodei são encontradas sobretudo no meio ambiente e alimentos e consideradas, usualmente, como bactérias oportunistas não-patogênicas e Y. ruckeri é um importante patógeno de peixes. Usualmente, as linhagens de Yersinia são classificadas em espécies de acordo com suas características bioquímicas. O Laboratório Nacional de Referência em Yersinia spp. outras que Y. pestis recebeu mais de 700 linhagens que foram identificadas bioquimicamente. Entretanto, sete linhagens de Yersinia não puderam ser identificadas pelos testes bioquímicos convencionais em nenhuma das espécies até o momento conhecidas e, por esse motivo, foram denominadas Yersinia atípicas. Os objetivos desse trabalho foram identificar as linhagens atípicas de Yersinia spp. em espécies por técnicas moleculares como Enterobacterial Repetitive Intergenic Consensus PCR (ERIC-PCR), Eletroforese em Campo Pulsado (PFGE), sequenciamento do gene 16S rRNA e Multilocus Sequencing Typing (MLST) e definir dentre as metodologias empregadas a que mais contribui para a identificação precisa dessas linhagens. Foi estudado um total de 59 linhagens de Yersinia spp., sendo 52 linhagens representantes das diferentes espécies do gênero e sete as linhagens bioquimicamente atípicas de Yersinia. As técnicas de ERIC-PCR, sequenciamento do gene 16S rRNA e o MLST foram eficientes na identificação molecular do gênero Yersinia, uma vez que conseguiram reunir todas as espécies em ramos espécie-específicos, com exceção de algumas linhagens de Y. frederiksenii e Y. kristensenii. A técnica de PFGE, pelo contrário, não agrupou as linhagens estudadas em clusters espécie-específicos. Os dados de ERIC-PCR, sequenciamento do gene 16S rRNA e MLST, sugerem que as linhagens atípicas FCF 229 e FCF 231 pertençam à espécie Y. ruckeri. Os dados de ERIC-PCR e MLST sugerem que a linhagem atípica FCF 487 pertença à espécie Y. enterocolitica. Ademais, os dados de ERIC-PCR, sequenciamento do gene 16S rRNA e MLST sugerem que as linhagens atípicas FCF 216, FCF 465, FCF 457 e FCF 494 pertençam a espécie Y. massiliensis. Os resultados obtidos nesse trabalho fornecem dados importantes para a caracterização molecular de linhagens bioquimicamente atípicas de Yersinia e contribuem para uma melhor descrição do gênero quanto a sua diversidade e reforçam o MLST como uma técnica confiável e reprodutível a ser usada na identificação de bactérias pertencentes a esse gênero, sendo dentre as metodologias utilizadas nesse estudo a mais indicada para tipagem molecular de yersiniae. / The genus Yersinia comprises 12 species. Y. enterocolitica, Y. pseudotuberculosis and Y. pestis are pathogens of various animals, including humans. Y. aldovae, Y. bercovieri, Y. frederiksenii, Y. intermedia, Y. kristensenii, Y. aleksiciae, Y. mollaretti and Y. rohdei have been mostly found in the environment and food sources and are commonly considered to be opportunistic nonpathogenic bacteria and Y. ruckeri is an important fish pathogen. Usually, Yersinia strains are classified into species according to their biochemical characteristics. The Brazilian Reference Center on Yersinia spp. other than Y. pestis received more than 700 strains that were biochemically identified. However, seven strains that were typed as Yersinia could not be biochemically identified in any one of the currently known Yersinia species and for this reason they were named as atypical strains. The aims of this work were to identify into species the atypical Yersinia strains using molecular techniques as Enterobacterial Repetitive Intergenic Consensus PCR (ERIC-PCR), Pulsed Field Gel Electrophoresis (PFGE), 16S rRNA gene sequencing and Multilocus Sequencing Typing (MLST) and to define which methodology better contribute to the identification of those strains. A total of 59 Yersinia spp. strains were studied, being 52 representative strains of the defined Yersinia species and seven atypical Yersinia strains. ERIC-PCR, 16S rRNA gene sequencing and MLST were efficient in molecular identifying the genera Yersinia once they grouped the strains into species-specific clusters, with exception of some Y. frederiksenii and Y. kristensenii strains. However, PFGE was not capable to cluster the defined Yersinia strains into species-specific clusters. The data obtained by ERIC-PCR, 16S rRNA gene sequencing and MLST suggest that the atypical strains FCF 229 and FCF 231 belong to Y. ruckeri species. The data obtained by ERIC-PCR and MLST suggest that FCF 487 belong to the Y. enterocolitica species. Additionally, ERIC-PCR, 16S rRNA gene sequencing and MLST suggest that the atypical strains FCF 216, FCF 465, FCF 457 and FCF 494 belong to the Y. massiliensis species. The results obtained provide important data for the molecular characterization of biochemically atypical strains and contribute for a better description of the genera regardless its diversity. Furthermore, the results reinforce MLST as a trustful and reproducible technique to be used in the identification of bacteria of this genus, being among the methodologies studied the most recommended one to molecular type yersiniae.

Résumé : Le Campylobacter est la première cause de gastro-entérites bactériennes dans les pays industrialisés. La grande majorité des cas sont des infections sporadiques dont la source est rarement identifiée. Le Campylobacter fait partie de la flore intestinale normale d’une large diversité d’animaux et peut également se retrouver dans l’eau. Le but de mon projet de recherche était d’étudier l’épidémiologie clinique et moléculaire des infections à Campylobacter en Estrie afin de déterminer les principales sources d’infections sporadiques et de comparer les génotypes des isolats de Campylobacter selon les différentes niches écologiques. Nous avons déterminé le profil de sensibilité aux antibiotiques d’isolats de différentes sources. Nous avons observé un haut taux de résistance à l’érythromycine et à la tétracycline et un faible taux de résistance à la ciprofloxacine chez les isolats de poulet, pouvant refléter l’utilisation de ces antibiotiques dans cet élevage. Le fait que le taux de résistance à l’érythromycine parmi les isolats humains soit significativement moins élevé que chez les isolats de poulet suggérait l’importance d’autres sources de Campylobacter chez l’humain. Afin de déterminer quelle méthode de typage moléculaire serait la mieux adaptée pour notre devis de recherche, nous avons comparé quatre méthodes (AFLP, MLST, typage du gène fla et EGCP). Seul le MLST a pu attribuer des isolats humains à des niches écologiques particulières comme le poulet, le lait cru et l’eau. Afin d’optimiser la technique de MLST, nous avons développé un système complémentaire basé sur le HRM, qui est beaucoup plus rapide et moins coûteux que le MLST. Nous avons démontré que le HRM a le potentiel de complémenter les méthodes d’analyses basées sur du séquençage pour l’étude des mutations ponctuelles et de faciliter une vaste gamme d’études basées sur des méthodes génotypiques, telle la détection de mutations ponctuelles qui confèrent de la résistance aux antibiotiques. Nous avons entrepris par la suite une étude cas-cas et un vaste projet d’isolement et de caractérisation moléculaire de souches de Campylobacter en Estrie, afin de véritablement cerner les mécanismes de transmission de la bactérie et de comparer les sources d’infections sporadiques chez les cas acquis en régions rurales vs urbaines. Nous avons confirmé que le poulet était responsable de la majorité des cas de campylobactérioses. Cependant, nos résultats suggèrent que la saisonnalité ainsi que le gradient urbain-rural de la campylobactériose sont dus à l’exposition aux souches bovines, particulièrement chez le groupe d’âge des 15-34 ans via l’exposition professionnelle. Par la détermination des sources d’infections, nous avons établi des pistes d’interventions utilisables par les autorités de santé publique, afin de diminuer l’incidence de la campylobactériose au Québec. / Abstract : Campylobacteriosis is the leading notifiable enteric disease in industrialised countries. It colonizes a wide range of animal which in turn spread the disease. The majority of campylobacteriosis cases are sporadic infections for which the source is rarely apparent. The main goal of my research project is to determine contamination sources of Campylobacter in the Eastern Townships, to identify the sources and routes of transmission and to establish the main sources of sporadic infections. We determined antimicrobial susceptibility profiles of Campylobacter isolates in order to predict which bacterial population will be resistant, caused by antimicrobial selective pressure administered to the host. High levels of resistance of chicken isolates to erythromycin and tetracycline, and low levels of resistance to ciprofloxacin reflect the use of the former antibiotics in animal husbandry. The fact that the erythromycin and tetracycline resistance levels were significantly lower among human isolates suggests that other transmission sources are important for human infection. In order to determine which molecular typing method will be the most relevant for our research design, we compared four typing methods (AFLP, MLST,/7a typing and PFGE). Only MLST has the potential to link isolates to a particular ecological niche, such as chicken, raw milk and water. In order to optimize MLST, we developed a complementary system based on HRM. We demonstrated that HRM has the potential to complement the analysis methods based on sequencing for SNP and facilitate a wide range of studies based on genotypic methods. We have subsequently undertaken a major project of isolation and molecular characterization of Campylobacter in the Eastern Townships, in order to truly understand the mechanisms of transmission of the bacteria and determine the source of sporadic cases. We confirmed that chicken was responsible for the majority of cases of campylobacteriosis. However, we have shown that the urban-rural gradient of campylobacteriosis in the Eastern Townships could be explained by exposure to bovine, especially for the 15-34 year old age group through occupational exposure. By the identification of infection sources, we proposed courses of action that could be used by public health authorities to reduce the incidence of campylobacteriosis in Quebec.

Orientador: Maria de Lourdes Ribeiro de Souza da Cunha / Resumo: Infecções causadas por Staphylococcus aureus representam um dos principais problemas de saúde pública, com elevadas taxas de morbidade e mortalidade principalmente entre indivíduos com deficiência de sistema imunológico como os diabéticos, em especial aqueles que fazem uso diário de insulina. Além da alta patogenicidade e facilidade de aquisição de resistência aos antimicrobianos, o patógeno tem grande habilidade de colonizar indivíduos de forma assintomática, favorecendo sua disseminação e tornando esses indivíduos fonte de risco para infecções. O estudo teve como objetivo analisar a prevalência e fatores de risco para o carreamento nasal e orofaríngeo, bem como caracterizar o perfil de resistência, virulência e tipagem molecular de S. aureus sensível à meticilina (MSSA) e S. aureus resistente à meticilina (MRSA) isolados de indivíduos diabéticos insulino-dependentes do município de Botucatu, SP. S. aureus foram obtidos da nasofaringe e orofaringe de 312 indivíduos diabéticos insulino-dependentes da comunidade e analisados quanto à presença do gene mecA, genes das enterotoxinas (sea, seb, e sec-1), esfoliatinas A e B (eta e etb), toxina 1 da síndrome do choque tóxico (tst), leucocidina Panton–Valentine (pvl), hemolisinas alfa (hla) e delta (hld) e biofilme (operon icaADBC) através da técnica da reação em cadeia da polimerase (PCR) e a tipagem do SCCmec através de PCRmultiplex. O perfil de sensibilidade à oxacilina, cefoxitina, linezolida, quinupristina/dalfopristina e sulfam... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Infections caused by Staphylococcus aureus is a major public health problem and infections with this microorganism are associated with high morbidity and mortality rates, especially among individuals with immune deficiency disorders such as diabetes and particularly those who take insulin daily. In addition to its high pathogenicity and ability to acquire antimicrobial resistance, asymptomatic infection with this pathogen is common, favoring its dissemination and rendering these individuals a source of infection. The objective of this study was to analyze the prevalence and risk factors for nasal and oropharyngeal carriage, as well as to characterize the resistance profile, virulence, clonal profile and sequence type of methicillin-susceptible (MSSA) and methicillin-resistant S. aureus (MRSA) isolated from insulin-dependent diabetic individuals in the city of Botucatu, SP, Brazil. Staphylococcus aureus was collected from the nasopharynx and oropharynx of 312 community insulin-dependent diabetic individuals. The isolates were analyzed for the presence of the mecA, enterotoxin (sea, seb and sec-1), exfoliatin A and B (eta and etb), toxic shock syndrome toxin 1 (tst), Panton-Valentine leukocidin (pvl), alpha and delta hemolysin (hla and hld), and biofilm (icaADBC operon) genes by the polymerase chain reaction (PCR). SCCmec typing was performed by multiplex PCR. The susceptibility profile against oxacillin, cefoxitin, linezolid, quinupristin/dalfopristin and sulfamethoxazole/trim... (Complete abstract click electronic access below) / Doutor

<p>Bacteria of the genus <i>Bartonella</i> inhabit the red blood cells of many mammals, including humans, and are transmitted by blood-sucking arthropod vectors. Different species of <i>Bartonella</i> are associated with different mammalian host species, to which they have adapted and normally do not cause any symptoms. Incidental infection of other hosts is however often followed by various disease symptoms, and several <i>Bartonella</i> species are considered as emerging human pathogens.</p><p>In this work, I have studied the genomic diversity within and between different <i>Bartonella</i> species, with focus on the feline-associated human pathogen <i>B. henselae</i> and its close relatives, the similarly feline-associated <i>B. koehlerae</i> and the trench-fever agent <i>B. quintana</i> which is restricted to humans.</p><p>In <i>B. henselae</i>, the overall variability in sequence and genome content was modest and well correlated, suggesting low levels of intra-species recombination in the core genome. The variably present genes were located in the prophage and the genomic islands, which are also absent from <i>B. quintana</i> and <i>B. koehlerae</i>, indicating multiple independent excision events. In contrast, diversity of genome structures was immense and probably associated with rearrangements between the repeated genomic islands located around the terminus of replication, possibly to avoid the host’s immune system. In both <i>B. henselae</i> and the mouse-associated species <i>B. grahamii</i> a large portion of the chromosome was manifold amplified in long-time cultures and packaged into phage particles, allowing for different recombination rates for different chromosomal regions.</p><p>In B<i>. quintana</i>, diversity was studied by sequencing non-coding spacers. The low variability might be due to the recent emergence of this species. Surprisingly, also this species displayed high variability in genome structures, despite its lack of repeated sequences.</p><p>The results indicate that genome rearrangements and gain or loss of mobile elements are major mechanisms of evolution in <i>Bartonella</i>.</p>

Toxoplasma gondii, the causal agent of toxoplasmosis, is an important water and food borne protozoan parasite. T. gondii was previously shown to have a distinct clonal population structure composed of Type I, II and III lineages in North America and Europe. But more recent studies demonstrated high diversity in South America. In the present project we have conducted an intensive study of the population diversity of T. gondii and surveyed the extent of genetic variation among natural T. gondii isolates on a global scale in order to better understand the population dynamics and pathogenesis of this parasite. To this end, 948 T. gondii isolates have been collected from a broad range of animal hosts and different sites worldwide. Our initial multilocus PCR-RFLP genotyping analysis revealed high diversity (~140 distinct genotypes) with abundant unique genotypes in South America and a strong clonal population structure in North America, Europe, Asia and Africa. It also showed that the Type II is the most common lineage worldwide, followed by the type III strain. The Type I strain, though widely distributed, has been infrequently isolated. Several new clonal genotypes have been identified from South America. The newly identified 140 RFLP genotypes have been further analyzed by multilocus microsatellites and intron sequencing methods. The composite data set identified 11 different haplotypes, providing a framework for future study of molecular epidemiology and population genetics of T. gondii . Multilocus DNA sequencing of markers from each of the 14 chromosomes covering the entire genome has also been completed to help reveal more information about genome evolution and the origin of T. gondii . Taken together, this comprehensive epidemiological and population genetic study has revealed significant details on the diversity and extent of sexual recombination, which provides the basis for future studies to understand transmission patterns, population dynamics and origin of this successful apicomplexan parasite Toxoplasma gondii.

Bacteria of the genus Bartonella inhabit the red blood cells of many mammals, including humans, and are transmitted by blood-sucking arthropod vectors. Different species of Bartonella are associated with different mammalian host species, to which they have adapted and normally do not cause any symptoms. Incidental infection of other hosts is however often followed by various disease symptoms, and several Bartonella species are considered as emerging human pathogens. In this work, I have studied the genomic diversity within and between different Bartonella species, with focus on the feline-associated human pathogen B. henselae and its close relatives, the similarly feline-associated B. koehlerae and the trench-fever agent B. quintana which is restricted to humans. In B. henselae, the overall variability in sequence and genome content was modest and well correlated, suggesting low levels of intra-species recombination in the core genome. The variably present genes were located in the prophage and the genomic islands, which are also absent from B. quintana and B. koehlerae, indicating multiple independent excision events. In contrast, diversity of genome structures was immense and probably associated with rearrangements between the repeated genomic islands located around the terminus of replication, possibly to avoid the host’s immune system. In both B. henselae and the mouse-associated species B. grahamii a large portion of the chromosome was manifold amplified in long-time cultures and packaged into phage particles, allowing for different recombination rates for different chromosomal regions. In B. quintana, diversity was studied by sequencing non-coding spacers. The low variability might be due to the recent emergence of this species. Surprisingly, also this species displayed high variability in genome structures, despite its lack of repeated sequences. The results indicate that genome rearrangements and gain or loss of mobile elements are major mechanisms of evolution in Bartonella.

A general aim was to combine molecular typing methods with clinical background information to increase epidemiological knowledge about Chlamydia trachomatis infections. An outbreak of Lymfogranuloma venereum (LGV), caused by a more invasive variant of C. trachomatis, was reported from the Netherlands in 2003 among men who have sex with men (MSM). All Chlamydia positive specimens from a venereal disease clinic for MSM in Stockholm during one year were genotyped. No spread of LGV was found, apart from three symptomatic cases. The same ompA genotypes were found among MSM in Melbourne, but the genotype distribution was different compared to findings among the heterosexual population in Sweden. The standard method for genotyping of Chlamydia is ompA-sequencing, but it has low resolution because one genotype predominates. A multilocus sequence typing (MLST) system based on five targets was developed. In a sample of 47 specimens, 32 variants were found with MLST, but only 12 variants with ompA-sequencing. The polymorphisms in the hctB gene, one MLST target, are caused by an element of 108 bp that is present in two to four repetitions and in different variants. Although the DNA-binding function of Hc2 that is encoded by hctB has been studied, our findings of a considerable size variation show that new studies are needed. In 2006, specimens with a 377 bp deletion in the cryptic plasmid covering the target region for diagnostic test systems from Abbott and Roche were discovered in Sweden. Applying MLST to these specimens indicated that there was a single clone, denoted nvCT. The proportion of nvCT in all detected Chlamydia cases was higher (20% to 65%) in counties using Abbott/Roche compared to counties using the BectonDickinson test system (7% to 20%). The proportions of nvCT converge in counties with high or low levels when detection systems were adjusted to detect nvCT.

Geotrichum candidum est une levure hémiascomycète ubiquitaire longtemps considérée comme un champignon filamenteux. C'est l'une des levures les plus fréquemment trouvées dans les fromages dans les quelles elle contribue à l'affinage. Dans le cadre du projet ANR ALIA Food Microbiomes en partenariat avec des industriels fromagers et producteur de levain, nous avons caractérisé l'espèce G. candidum par une étude phylogénétique et placé de manière non ambigüe G. candidum parmi les levures hémiascomètes. Une analyse MLST a permis de séparer les souches étudiées en deux groupes. Le premier contient essentiellement des souches environnementales tandis que le second ne contient que des souches isolé du fromage. Cela suggère une certaine sélection ou spécialisation d'un groupe de souche dans la fabrication du fromage. Une méthode de typage inter LTR plus discriminante a permis de typer l'ensemble des souches et peut fournir aux industriels un outil robuste pour le suivi d'une souche en production. Le génome de G. candidum CLIB 918 = ATCC 204307 a été séquencé. Les premières analyses ont mis en évidence des discontinuités évolutives parmi les gènes qui le composent. Parmi les 6802 gènes identifiés, 315 gènes présentent des orthologues chez les champignons filamenteux et non chez les levures. Cela suggère que durant l'évolution, G. candidum a conservé un grand nombre de gènes qui a été perdu chez les autres levures ou en a reçu certain par transfert horizontal de gènes. L'existence de ce même type de gènes chez d'autre levure ayant une position basale dans l'arbre des hémiascomycètes, suggère que G. candidum et ces levures ont une position intermédiaire lors de la transition évolutive champignon vers levure. Il est à noter que certains d'entre eux sont impliqués dans le métabolisme et pourraient jouer un rôle dans l'adaptation de cette levure à la fabrication du fromage.