PATCLIN - Artigos publicados em revistas não indexadas
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- Molecular epidemiology of HIV type 1 infection in Portugal: high prevalence of non-B subtypesPublication . Esteves, A; Parreira, R; Venenno, T; Franco, M; Piedade, J; Sousa, JG; Canas-Ferreira, WIn this study, we have investigated the diversity of current HIV-1 strains circulating in the metropolitan area of Lisbon, Portugal. A total of 217 HIV-1-positive blood samples, collected between October 1998 and December 2000, was genetically characterized in the gp120 C2V3C3 region (n = 205) or part of the gp41 N-terminal segment (n = 12) by heteroduplex mobility assay (HMA) and/or DNA sequencing. The HMA subtyping efficiency (number of samples unambiguously subtyped by HMA divided by the total number of samples subtyped) was 65.9% (143 of 217), with indeterminate migration patterns of subtype A and G strains contributing significantly to this value. On the overall, subtype B was the most prevalent (50.2%), followed by subtypes G (21.7%), A (17.5%), and F (5.5%), whereas subtypes C, D, H, and J accounted altogether for 5.1% of the infections. Non-B subtypes were responsible for 77.4 and 33.1% of the infections among African immigrants and Portuguese subjects, respectively. Angolan individuals (n = 25) were the only ones infected with all the HIV-1 subtypes documented, probably reflecting a high degree of viral genetic diversification in their country of origin. Phylogenetic analysis showed a predominance of IbNG-like viruses among subtype A sequences and two new major subclusters within subtype G (G(P) and G(P)'). The majority of the Portuguese G sequences described formed a well-defined subcluster (G(P)), supported by bootstrap values >90%, phylogenetically distant from clade G sequences in databases. gag (p24/p7) sequence analysis of these variants confirmed the maintenance of the subtype G subclusters. The multiple subclustering observed for the major clades A, B, D, and G, as well as the variety of subtypes found, indicate a high diversity of HIV-1 variants circulating in Portugal and suggest a need for continuous epidemiologic surveillance.
- Spreading of HIV-1 subtype G and envB/gagG recombinant strains among injecting drug users in Lisbon, Portugal.Publication . Esteves, A; Parreira, R; Piedade, J; Venenno, T; Franco, M; Sousa, JG; Patrício, L; Brum, P; Costa, A; Canas-Ferreira, WWe have evaluated the genetic diversity of HIV-1 strains infecting injecting drug users (IDUs) in Lisbon, Portugal. Heteroduplex mobility assay and/or phylogenetic analysis revealed that env (C2V3C3 or gp41) subtype B is present in 63.7% of the 135 viral samples studied, followed by subtypes G (23.7%), A (6.7%), F (5.2%), and D (0.7%). Similar analysis of gag (p24/p7) performed on 91 of the specimens demonstrated that 49.5% of the infections were caused by subtype G viruses; other gag subtypes identified were B (39.5%), F (3.3%), A and D (1.1.% each), and the recombinant circulating form CRF02_AG (5.5%). Discordant env/gag sub-types were detected in 34.1% of the strains and may reflect the presence of dual infections and/or recombinant viruses. The presumptive B/G recombinant form was highly predominant (21 of 31). The genetic pattern of HIV-1 subtype B and G strains is suggestive of multiple introductions and recombination episodes and of a longstanding presence of both subtypes in the country. C2V3C3 amino acid sequences from IDU-derived subtype G viruses presented highly significant signatures, which distinguish the variants from this transmission group. The unusually high prevalence of subtype G sequences (34.1%), independent of the geographic origin of the infected individuals, makes this IDU HIV-1 epidemic unique.
- Identificação molecular pelo método de Spoligotyping de estirpes do complexo Mycobacterium tuberculosis isoladas no Hospital Fernando FonsecaPublication . David, S; Portugal, C; Antunes, A; Cardoso, A; Calado, A; Barros, V; Sancho, LSpoligotyping was used in the genotyping of 219 isolates of the Mycobacterium tuberculosis complex, from patients of the Hospital Fernando Fonseca. This technique, based on PCR methodology, analyses a region of the chromosome specific of the Mycobacterium tuberculosis complex, the DR locus (Direct Repeat). With the aid of an international database, we showed that the predominant Spoligotypes belonged to the LAM family (Latino-American Mediterranean), 29.2 %. The LAM 9 family, with 12.3 %, left us attentive to the possible import of the disease through populations from South America, were it has been frequently identified. The genotypic families T1 and Haarlem, with 6.4 % and 8.7 % respectively, represented a frequency typical to Europe. The Beijing family, with 1.4 %, may represent an emerging problem in our country due to recent immigration of Asian and Eastern European populations. Isolates with a Spoligotype of the M. bovis type were found at a high percentage, 3.7 %. In Europe, this infection is extremely rare suggesting the result may not be due to M. bovis infection but to M. bovis BCG (due to vaccination or eventual recombinant BCG based therapies), or M. africanum (due to the proximity of the two species). A high percentage of the Spoligotypes were not identified by the database, 21.4 %. This is the first study of this type amongst us and may be the starting point for the creation of a data base with important consequences on the national program against tuberculosis.
- Novos dados sobre os Spoligotypes de estirpes do complexo Mycobacterium tuberculosis isoladas no Hospital Fernando Fonseca (Amadora-Sintra, Portugal)Publication . David, S; Barros, V; Portugal, C; Antunes, A; Cardoso, A; Calado, A; Sancho, L; Sousa, JGO presente estudo populacional, que decorreu entre 1999 e 2003, foi baseado na utilização do Spoligotyping na genotipagem de 452 isolados do complexo Mycobacterium tuberculosis de doentes com tuberculose internados no Hospital Fernando Fonseca. Spoligotypes foram identificados como shared types (ST) recorrendo a uma base de dados internacional. Onze ST raros, não identificados na base de dados, acomodaram 8,4% dos isolados. Aliás, particular a Portugal poderá ser a predominância de ST identificados na base de dados mas não previamente classificados como famílias genotípicas, tais como o ST244, ST150 e ST389, representando 13,3 % do total. A identificação de isolados clínicos de M. africanum de genótipo Afri1 e de M. tuberculosis de genótipo CAS1 poderá confirmar a importação de isolados de origem africana e asiática. M. tuberculosis da família Beijing foi pela primeira vez por nós assinalado a partir de 1999. Desde então, o número de isolados provenientes do hospital passou de um para cinco, anualmente, representando actualmente 2,2%, o que a coloca em décimo lugar em prevalência. M. tuberculosis Beijing poderá corresponder a um problema emergente em Portugal devido à recente imigração proveniente da Europa Oriental e da Ásia. Outros genótipos, ST150 e ST389, mostraram um incremento, cujo significado não é claro. No entanto, as frequências relativas das famílias predominantes LAM, T1 e Haarlem mantiveram-se relativamente estáveis. O presente estudo confirma a variabilidade genética em Portugal dos isolados do complexo M. tuberculosis. Estes estudos poderão contribuir para a definição de prioridades nos programas nacionais de luta contra a tuberculose.
- Direct application of the INNO-LiPA Rif.TB line-probe assay for rapid identification of Mycobacterium tuberculosis complex strains and detection of rifampin resistance in 360 Mycobacterium tuberculosis complex strains and detection of rifampin resistance in 360 smear-positive respiratory specimens from an area of high incidence of multidrug-resistant tuberculosisPublication . Viveiros, M; Leandro, C; Rodrigues, L; Almeida, J; Bettencourt, R; Couto, I; Carrilho, L; Diogo, J; Fonseca, A; Lito, L; Lopes, J; Pacheco, T; Pessanha, M; Quirim, J; Sancho, L; Salfinger, M; Amaral, LThe INNO-LiPA Rif.TB assay for the identification of Mycobacterium tuberculosis complex strains and the detection of rifampin (RIF) resistance has been evaluated with 360 smear-positive respiratory specimens from an area of high incidence of multidrug-resistant tuberculosis (MDR-TB). The sensitivity when compared to conventional identification/culture methods was 82.2%, and the specificity was 66.7%; the sensitivity and specificity were 100.0% and 96.9%, respectively, for the detection of RIF resistance. This assay has the potential to provide rapid information that is essential for the effective management of MDR-TB.
- Spoligtyping e polimorfismo do gene pncA: um cenário em duas etapas para o diagnóstico de mycobacterium bovis em PortugalPublication . David, S; Portugal, C; Antunes, A; Calado, A; Cardoso, A; Barros, V; Sancho, L; Sousa, JGThe differential diagnosis of Mycobacterium bovis is important in the control of transmission to the human population and also for treatment since M. bovis is naturally resistant to pyrazinamide. Eleven clinical isolates from the Fernando Fonseca Hospital with Spoligotypes indicative of M. bovis, through the absence of spacers 39-43 but that also counted with the absence of spacer 38, were analyzed. For the identification of these strains, the phenotypic analysis of pyrazinamide resistance and study of the polymorphisms of the pncA and gyrB genes were carried out. The study of the pncA polymorphism revealed that the strains analyzed did not contain the M. bovis specific mutation. In relation the gyrB polymorphisms, using the GenoTypeO MTBC kit, the strains were identified as belonging to the group M. tuberculosis, M. africanum subtipo II e M. canetti. The present investigation enabled us to define new genotypes on which future bacteriological studies should be based. Amongst these the study of the pncA polymorphism was considered important due to the immediate practical implications for the clinician. Evaluating transmission and defining groups of risk is an objective for which support from the veterinary services is considered relevant.
- Contribution of spoligotyping to the characterization of the population structure of Mycobacterium tuberculosis isolates in PortugalPublication . David, S; Ribeiro, D; Antunes, A; Portugal, C; Sancho, L; Sousa, JGTuberculosis is a major health problem in Portugal. To begin characterizing the population structure of Mycobacterium tuberculosis, spoligotyping was used for the systematic typing, through consecutive sampling, of patient isolates from the Amadora-Sintra area of Greater Lisbon. Distribution amongst major spoligotype families, including the Latin American Mediterranean (LAM), T, Haarlem and Beijing, was compared to that of the international spoligotype database SpolDB4 and to the European countries of traditional Portuguese immigration represented in SpolDB4. Spoligotypes from 665 isolates were analyzed and 97 shared international types (SITs) identified. In SpolDB4 Portugal is represented by part of the spoligotypes from this study explaining the reduced number of unidentified patterns. The importance of the LAM family, and especially of LAM1 and LAM9 sub-families that alone represented 38% of all the isolates in this study as compared to 8% relative to the European sub group, led us to believe that at least in this respect the population structure was closer to that of Africa and South America than to Europe. Spoligotypes characteristic of Portugal or Portuguese related settings were identified. These included SIT244 a T1 sub-family predominant in Portugal and Bangladesh, SIT64 a LAM 6 sub-family common to Portugal and Brazil, and SIT1106 a LAM 9 sub-family. These studies were the first in Portugal stressing the importance of monitoring the population structure of M. tuberculosis isolates, an important step towards gaining an understanding of tuberculosis and the dynamics of this disease.
- Evaluation of liquid biphasic Granada medium and instant liquid biphasic Granada medium for group B streptococcus detection.Publication . Martinho, F; Prieto, E; Pinto, C; Castro, R; Morais, A; Salgado, L; Exposto, FINTRODUCTION: Group B streptococci (GBS) are transmitted from the mother to the newborn. Prevention of neonatal infection is achieved by intrapartum prophylaxis given to mothers colonized with GBS at 35 to 37 weeks of pregnancy. MATERIALS AND METHODS: Liquid biphasic Granada medium (LB) and instant liquid biphasic Granada medium (ILB) were evaluated for GBS detection. Vaginal swabs obtained from 300 women were inoculated onto LB or ILB, or onto Todd-Hewitt broth and analyzed with the ATB system (comparison method). RESULTS: Prevalence of GBS was 20% (61/300). LB and Todd-Hewitt with ATB detected GBS in 20% of women, and ILB in 19% of women. No growth was observed at four hours in any of the media studied. At 10 h and 14 h, identification of GBS was possible in 43/300 (14%) and 53/300 (18%) of ILB cultures, respectively, and in 32/300 (11%) and 46/300 (15%) of LB cultures. CONCLUSION: All the media used are suitable for GBS detection. The majority of GBS were identified in ILB and LB cultures at 10 h and 14 h.
- Prevalence of hepatitis E virus antibody in an non endemic population: prospective studyPublication . Alberto, S; Pires, S; Félix, J; Figueiredo, A; Silva, L; Franco, M; Sousa, JG; Deus, JR
- High prevalence of ST121 in community-associated methicillin-susceptible Staphylococcus aureus lineages responsible for skin and soft tissue infections in Portuguese childrenPublication . Conceição, T; Aires-de-Sousa, M; Pona, N; Brito, MJ; Barradas, C; Coelho, R; Sardinha, T; Sancho, L; Sousa, JG; Machado, MC; Lencastre, HIn order to evaluate the incidence of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) in Portugal, we analyzed a collection of 38 S. aureus isolates recovered from 30 children attending the pediatric emergency department of a central hospital in Lisbon due to skin and soft tissue infections. Molecular characterization identified seven clonal lineages among the 35 methicillin-susceptible S. aureus (MSSA) isolates, of which the major lineage PFGE A/t159/ST121 included 63% of the isolates. The three MRSA isolates belonged to the Pediatric clone PFGE D/t535/ST5-IV (n = 2) and to the European CA-MRSA clone PFGE G/t044/ST80-IVc (n = 1). All isolates harbored several virulence factors, namely, leukocidins. Panton-Valentine leukocidin (PVL) was produced by isolates from five MSSA lineages and by the ST80 MRSA. Of interest, this is the first reported isolation of CA-MRSA ST80 in Portugal.
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