Browsing by Author "Borrego, M.J."
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- Accuracy of prenatal culture in predicting intrapartum group B streptococcus colonization statusPublication . Florindo, C.; Damião, V.; Lima, J.; Nogueira, I.; Rocha, I.; Caetano, P.; Ribeiro, L.; Viegas, S.; Gomes, João Paulo; Borrego, M.J.Objective: To evaluate the positive predictive value (PPV) of group B Streptococcus (GBS) cultures at 35–37 weeks of gestation relative to GBS colonization status at delivery. Methods: Rectovaginal swabs from 221 women at labor in four Lisbon hospitals were collected for GBS screening according to the CDC guidelines. Results: The PPV was 24.4%. IAP was administered to 100% of prenatally GBS positive women. There was no case of early onset GBS disease (EOD). Conclusions: Poor accuracy of prenatal cultures in identifying true candidates for IAP highlights the need for Portuguese clinical and laboratory guidelines to prevent EOD and antibiotic overtreatment of pregnant women.
- Adaptive evolution of the Chlamydia trachomatis dominant antigen reveals distinct evolutionary scenarios for B- and T-cell epitopes: worldwide surveyPublication . Nunes, A.; Nogueira, P.J.; Borrego, M.J.; Gomes, João PauloBackground: Chlamydia trachomatis is one of the most disseminated human pathogens, for which no vaccine is available yet. Understanding the impact of the host pressure on pathogen antigens is crucial, but so far it was only assessed for highly-restricted geographic areas. We aimed to evaluate the evolutionary picture of the chlamydial key antigen (MOMP), which is one of the leading multi-subunit vaccine candidates, in a worldwide basis. Methodology/Principal Findings: Using genetics, molecular evolution methods and mathematical modelling, we analyzed all MOMP sequences reported worldwide, composed by 5026 strains from 33 geographic regions of five continents. Overall, 35.9% of variants were detected. The evolutionary pattern of MOMP amino acid gains/losses was found to differ from the remaining chromosome, reflecting the demanding constraints of this porin, adhesin and dominant antigen. Amino acid changes were 4.3-fold more frequent in host-interacting domains (P,10212), specifically within B-cell epitopes (P,1025), where 25% of them are at fixation (P,1025). According to the typical pathogen-host arms race, this rampant B-cell antigenic variation likely represents neutralization escape mutants, as some mutations were previously shown to abrogate neutralization of chlamydial infectivity in vitro. In contrast, T-cell clusters of diverse HLA specificities are under purifying selection, suggesting a strategy that may lead to immune subversion. Moreover, several silent mutations are at fixation, generating preferential codons that may influence expression, and may also reflect recombination-derived ‘hitchhikingeffect’ from favourable nonsilent changes. Interestingly, the most prevalent C. trachomatis genotypes, E and F, showed a mutation rate 22.3-fold lower than that of the remainder (P,10220), suggesting more fitted antigenic profiles. Conclusions/Significance: Globally, the adaptive evolution of the C. trachomatis dominant antigen is likely driven by its complex pathogenesis-related function and reflects distinct evolutionary antigenic scenarios that may benefit the pathogen, and thus should be taking into account in the development of a MOMP-based vaccine.
- A Bioinformática na Identificação dos Processos Moleculares de Interacção entre o Agente Patogénico e o HomemPublication . Borges, V.; Nunes, A.; Ferreira, R.; Borrego, M.J.; Gomes, João PauloQualquer processo infeccioso caracteriza-se por um “braço de ferro” contínuo entre o agente patogénico e o Homem. Se por um lado, o Homem tenta debelar a infecção através da complexa rede celular que caracteriza a resposta imunitária, por outro, o agente patogénico tenta escapar a essa resposta através da acumulação de mutações no seu genoma. De um modo geral, as mutações num gene podem ser sinónimas (quando não originam uma alteração de aminoácido) ou não-sinónimas (quando a proteína correspondente é alterada), sendo estas últimas, quando vantajosas, as principais responsáveis pela adaptação do agente infeccioso ao hospedeiro. A fixação de alterações não-sinónimas vantajosas resulta de um processo evolutivo denominado de “selecção positiva”. A bioinformática surge nos últimos anos como uma ferramenta acessível e indispensável para a análise dos dados genómicos que diariamente são gerados de forma exponencial. Neste âmbito, a bioinformática tem sido essencial, por exemplo, para a identificação dos processos moleculares de interacção entre o agente patogénico e o Homem, que decorrem durante o processo infeccioso. Na presente comunicação, a utilidade desta valência computacional é ilustrada tomando como modelo o processo infeccioso despoletado pela bactéria Chlamydia trachomatis. De facto, esta bactéria intracelular é caracterizada por um perfil bem definido de infecções no Homem, cujos diversos genótipos afectam distintamente o tecido ocular, os órgãos genitais e os nódulos linfáticos inguinais (via macrófagos), orgãos estes que exercem uma pressão selectiva distinta (resposta imunitária, pH, flora comensal, etc) sobre a bactéria. Através da utilização de várias plataformas de bioinformática para a análise específica das mutações que distinguem os vários genótipos de Chlamydia trachomatis, é possível identificar genes, que por força de uma selecção positiva, estão hipoteticamente envolvidos na adaptação/invasão aos vários tipos de células humanas infectadas, nomeadamente, células epiteliais das mucosas ocular e genital, e macrófagos. Noutras vertentes, este tipo de análise de detecção de selecção positiva teve já aplicações no desenvolvimento de uma vacina para o VIH bem como no esclarecimento da evolução da virulência do vírus Influenza, e pode ser aplicado a todo o sistema biológico.
- Chlamydia trachomatis infection in patients selected for HPV detectionPublication . Santo, I.; Azevedo, J.; Borrego, M.J.; Gomes, João Paulo; Verdasca, N.; Pista, A.Background: The significance of the association between the human papillomavirus (HPV) and other sexually transmitted infections in the development of cervical, penile or anal neoplasias has been investigated, and the more consistent data have pointed to an association with Chlamydia trachomatis. In Portugal, the lack of information on STI precludes any knowledge on this subject. Objective: To determine CT infection in a group of individuals selected for HPV detection in the major Portuguese STD clinic. Methods: This opportunistic screening comprehended 177 outpatients (148 women, 29 men; age: 16-61 years) suspected of HPV infection (warts, abnormal histology) between 2008 and 2010. Demographic and sexual behaviour data and a full medical history were obtained at enrolment. Genital samples (cervical, vulvar, vaginal, penile or anal) were collected from all the subjects. HPV DNA was detected by CLART HPV2 assay, which allowed the detection of 35 genotypes. CT DNA was detected by Cobas 4800. Results and Discussion: Overall, 84.5% of the individuals had at least one of the infections. Evidencing an excellent correlation with clinical signs, HPV infection was detected in 68.2% of the women and in 75.9% of the men, where CT positivity was 10.1% and 13.8%, respectively. Coinfection was observed in 8.9% of the women and in 13.8% of the men. No correlation with HPV or CT genotypes could be established. HPV infection was more frequent in CT negative (87.1%) than in CT positive women (13.8%), and the same was observed for men (81.8% versus 18.2%). Full results will be presented and discussed. Conclusions: No correlation between HPV-CT coinfection, and clinical signs was observed. However, further long-term studies are needed to elucidate the effects of HPV-CT coinfection in the clinical history of the infected patient, which would greatly contribute towards a better management of patients.
- Complete Genome Sequence of Chlamydia trachomatis Ocular Serovar C Strain TW-3Publication . Borges, V.; Pinheiro, M.; Vieira, Luís; Sampaio, Daniel A.; Nunes, A.; Borrego, M.J.; Gomes, João PauloChlamydia trachomatis is the etiological agent of trachoma, the leading infectious cause of blindness worldwide. We report here the first complete and annotated genome of a C. trachomatis trachoma-causing serovar C strain (strain TW-3). The chromosome and plasmid are 1,043,554 bp and 7,501 bp in length, respectively.
- Identification of type III secretion substrates of Chlamydia trachomatis using Yersinia enterocolitica as a heterologous systemPublication . da Cunha, M.; Milho, C.; Almeida, F.; Pais, S.V.; Borges, V.; Borrego, M.J.; Gomes, João Paulo; Mota, L.J.Background: Chlamydia trachomatis is an obligate intracellular human pathogen causing ocular and urogenital infections that are a significant clinical and public health concern. This bacterium uses a type III secretion (T3S) system to manipulate host cells, through the delivery of effector proteins into their cytosol, membranes, and nucleus. In this work, we aimed to find previously unidentified C. trachomatis T3S substrates. Results: We first analyzed the genome of C. trachomatis L2/434 strain for genes encoding mostly uncharacterized proteins that did not appear to possess a signal of the general secretory pathway and which had not been previously experimentally shown to be T3S substrates. We selected several genes with these characteristics and analyzed T3S of the encoding proteins using Yersinia enterocolitica as a heterologous system. We identified 23 C. trachomatis proteins whose first 20 amino acids were sufficient to drive T3S of the mature form of β-lactamase TEM-1 by Y. enterocolitica. We found that 10 of these 23 proteins were also type III secreted in their full-length versions by Y. enterocolitica, providing additional support that they are T3S substrates. Seven of these 10 likely T3S substrates of C. trachomatis were delivered by Y. enterocolitica into host cells, further suggesting that they could be effectors. Finally, real-time quantitative PCR analysis of expression of genes encoding the 10 likely T3S substrates of C. trachomatis showed that 9 of them were clearly expressed during infection of host cells. Conclusions: Using Y. enterocolitica as a heterologous system, we identified 10 likely T3S substrates of C. trachomatis (CT053, CT105, CT142, CT143, CT144, CT161, CT338, CT429, CT656, and CT849) and could detect translocation into host cells of CT053, CT105, CT142, CT143, CT161, CT338, and CT429. Therefore, we revealed several C. trachomatis proteins that could be effectors subverting host cell processes.
- Identification of type III secretion substrates of Chlamydia trachomatis using Yersinia enterocolitica as a heterologous systemPublication . da Cunha, M.; Milho, C.; Almeida, F.; Pais, S.V.; Borges, V.; Maurício, R.; Borrego, M.J.; Gomes, João Paulo; Mota, L.J.Background: Chlamydia trachomatis is an obligate intracellular human pathogen causing ocular and urogenital infections that are a significant clinical and public health concern. This bacterium uses a type III secretion (T3S) system to manipulate host cells, through the delivery of effector proteins into their cytosol, membranes, and nucleus. In this work, we aimed to find previously unidentified C. trachomatis T3S substrates. Results: We first analyzed the genome of C. trachomatis L2/434 strain for genes encoding mostly uncharacterized proteins that did not appear to possess a signal of the general secretory pathway and which had not been previously experimentally shown to be T3S substrates. We selected several genes with these characteristics and analyzed T3S of the encoding proteins using Yersinia enterocolitica as a heterologous system. We identified 23 C. trachomatis proteins whose first 20 amino acids were sufficient to drive T3S of the mature form of β-lactamase TEM-1 by Y. enterocolitica. We found that 10 of these 23 proteins were also type III secreted in their full-length versions by Y. enterocolitica, providing additional support that they are T3S substrates. Seven of these 10 likely T3S substrates of C. trachomatis were delivered by Y. enterocolitica into host cells, further suggesting that they could be effectors. Finally, real-time quantitative PCR analysis of expression of genes encoding the 10 likely T3S substrates of C. trachomatis showed that 9 of them were clearly expressed during infection of host cells. Conclusions: Using Y. enterocolitica as a heterologous system, we identified 10 likely T3S substrates of C. trachomatis (CT053, CT105, CT142, CT143, CT144, CT161, CT338, CT429, CT656, and CT849) and could detect translocation into host cells of CT053, CT105, CT142, CT143, CT161, CT338, and CT429. Therefore, we revealed several C. trachomatis proteins that could be effectors subverting host cell processes.
- In silico scrutiny of genes revealing phylogenetic congruence with clinical prevalence or tropism properties of Chlamydia trachomatis strainsPublication . Ferreira, R.; Antelo, M.; Nunes, A.; Borges, V.; Damião, V.; Borrego, M.J.; Gomes, João PauloMicrobes possess a multiplicity of virulence factors that confer them the ability to specifically infect distinct biological niches. Contrary to what is known for other bacteria, for the obligate intracellular human pathogen Chlamydia trachomatis, the knowledge of the molecular basis underlying serovars’ tissue specificity is scarce. We examined all ~900 genes to evaluate the association between individual phylogenies and cell-appetence or ecological success of C. trachomatis strains. Only ~1% of the genes presented a tree topology showing the segregation of all three disease groups (ocular, urogenital, and lymphatic) into three wellsupported clades. Approximately 28% of the genes, which include the majority of the genes encoding putative type III secretion system effectors and Inc proteins, present a phylogenetic tree where only lymphogranuloma venereum strains form a clade. Similarly, an exclusive phylogenetic segregation of the most prevalent genital serovars was observed for 61 proteins. Curiously, these serovars are phylogenetically cosegregated with the lymphogranuloma venereum serovars for ~20% of the genes. Some clade-specific pseudogenes were identified (novel findings include the conserved hypothetical protein CT037 and the predicted a-hemolysin CT473), suggesting their putative expendability for the infection of particular niches. Approximately 3.5% of the genes revealed a significant overrepresentation of nonsynonymous mutations, and the majority encode proteins that directly interact with the host. Overall, this in silico scrutiny of genes whose phylogeny is congruent with clinical prevalence or tissue specificity of C. trachomatis strains may constitute an important database of putative targets for future functional studies to evaluate their biological role in chlamydial infections.
- In Silico Scrutiny of Genes Revealing Phylogenetic Congruence with Clinical Prevalence or Tropism Properties of Chlamydia trachomatis StrainsPublication . Ferreira, R.; Antelo, M.; Nunes, A.; Borges, V.; Damião, V.; Borrego, M.J.; Gomes, João PauloMicrobes possess a multiplicity of virulence factors that confer them the ability to specifically infect distinct biological niches. Contrary to what is known for other bacteria, for the obligate intracellular human pathogen Chlamydia trachomatis, the knowledge of the molecular basis underlying serovars' tissue specificity is scarce. We examined all ~900 genes to evaluate the association between individual phylogenies and cell-appetence or ecological success of C. trachomatis strains. Only ~1% of the genes presented a tree topology showing the segregation of all three disease groups (ocular, urogenital, and lymphatic) into three well-supported clades. Approximately 28% of the genes, which include the majority of the genes encoding putative type III secretion system effectors and Inc proteins, present a phylogenetic tree where only lymphogranuloma venereum strains form a clade. Similarly, an exclusive phylogenetic segregation of the most prevalent genital serovars was observed for 61 proteins. Curiously, these serovars are phylogenetically cosegregated with the lymphogranuloma venereum serovars for ~20% of the genes. Some clade-specific pseudogenes were identified (novel findings include the conserved hypothetical protein CT037 and the predicted α-hemolysin CT473), suggesting their putative expendability for the infection of particular niches. Approximately 3.5% of the genes revealed a significant overrepresentation of nonsynonymous mutations, and the majority encode proteins that directly interact with the host. Overall, this in silico scrutiny of genes whose phylogeny is congruent with clinical prevalence or tissue specificity of C. trachomatis strains may constitute an important database of putative targets for future functional studies to evaluate their biological role in chlamydial infections.
- Molecular epidemiology of group B streptococcalPublication . Florindo, C.; Gomes, João Paulo; Rato, M.G.; Bernardino, L.; Spellerberg, B.; Santos-Sanches, I.; Borrego, M.J.Streptococcus agalactiae is a major pathogen of neonates and immunocompromised adults. Prior studies have demonstrated that, beyond the neonatal period, S. agalactiae rarely causes invasive infections in children. However, during 2004–2005, S. agalactiae was the causative agent of 60 meningitis episodes in children aged 3 months to 12 years from Angola. To identify and study the specific causative genetic lineages of S. agalactiae childhood meningitis, which lack characterization to date, we conducted an extensive molecular analysis of the recovered isolates (n521). This constitutes what we believe to be the first molecular study of the population structure of invasive S. agalactiae isolates from Africa. A low genetic diversity was observed among the isolates, where the majority belonged to clonal complex (CC) 17 presenting the capsular subtype III-2 (86% of cases) and marked by the intron group II GBSi1, which has previously been observed to be associated with neonatal hosts. The predominance of single-locus variants of sequence type (ST) 17 suggested the local diversification of this hypervirulent clone, which displayed novel alleles of the fbsB and sip virulence genes. The absence of the scpB–lmb region in two S. agalactiae isolates with the Ia/ST23 genotype is more typical of cattle than human isolates. Globally, these data provide novel information about the enhanced invasiveness of the CC17 genetic lineage in older children and suggest the local diversification of this clone, which may be related to the future emergence of a novel epidemic clone in Angola.