Percorrer por autor "Carneiro, Sofia"
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- Genome-scale analysis of Mycobacterium avium complex isolates from Portugal reveals extensive genetic diversityPublication . Carneiro, Sofia; Pinto, Miguel; Rodrigues, Joana; Gomes, João Paulo; Macedo, RitaOpportunist infections caused by nontuberculous mycobacteria (NTM) have emerged as a significant public health problem. Among these, species of the Mycobacterium avium complex (MAC) are the main responsible for the increase in the number of human disease cases. In order to address the current needs in the detection and surveillance of MAC disease cases, we evaluated different species classification methodologies (BLASTn-based marker-gene approach, Kraken v2, rMLST and MLST databases) and their congruence with a core-SNP phylogenetic approach, based on whole genome sequencing (WGS) data. For this purpose, we used a collection of 142 MAC isolates from Portuguese patients diagnosed between 2014 and 2022. The marker-gene approach (based on the rpoB, hsp65 and groEL genes), showed the best results, allowing the identification of the 142 MAC isolates to the species/subspecies level (M. avium subsp. hominissuis, M. intracellulare, M. intracellulare subsp. chimaera, M. intracellulare subsp. yongonense, M. marseillence and M. colombiense). Additionally, we performed drug susceptibility testing that confirmed clarithromycin efficacy as a first-line treatment for MAC disease, as 93 % of the Portuguese isolates were susceptible. Using a core-SNP approach we also performed an in-depth phylogenetic analysis within each identified species group, and despite the high genetic diversity within the MAC species, we were able to clearly distinguish all the species/subspecies and identify genetic clusters with epidemiological potential. We highlight not only the need for the standardization of an appropriate genotyping approach for species identification and management of MAC disease, but also a more robust large-scale WGS data analysis, in a One Health perspective, in order to identify potential routes of transmission.
- Genome-Scale Characterization of Mycobacterium abscessus Complex Isolates from PortugalPublication . Carneiro, Sofia; Pinto, Miguel; Silva, Sónia; Santos, Andrea; Rodrigues, Irene; Santos, Daniela; Duarte, Sílvia; Vieira, Luís; Gomes, João Paulo; Macedo, RitaThe Mycobacterium abscessus complex (MABC) is an emerging, difficult to treat, multidrug-resistant nontuberculous mycobacteria responsible for a wide spectrum of infections and associated with an increasing number of cases worldwide. Dominant circulating clones (DCCs) of MABC have been genetically identified as groups of strains associated with higher prevalence, higher levels of antimicrobial resistance, and worse clinical outcomes. To date, little is known about the genomic characteristics of MABC species circulating in Portugal. Here, we examined the genetic diversity and antimicrobial resistance profiles of 30 MABC strains isolated between 2014 and 2022 in Portugal. The genetic diversity of circulating MABC strains was assessed through a gene-by-gene approach (wgMLST), allowing their subspecies differentiation and the classification of isolates into DCCs. Antimicrobial resistance profiles were defined using phenotypic, molecular, and genomic approaches. The majority of isolates were resistant to at least two antimicrobials, although a poor correlation between phenotype and genotype data was observed. Portuguese genomes were highly diverse, and data suggest the existence of MABC lineages with potential international circulation or cross-border transmission. This study highlights the genetic diversity and antimicrobial resistance profile of circulating MABC isolates in Portugal while representing the first step towards the implementation of a genomic-based surveillance system for MABC at the Portuguese NIH.
- Microevolution of a Mycobacteroides abscessus subsp. bolletii strain in a clinical persistent infectionPublication . Santos, Andrea; Pinto, Miguel; Carneiro, Sofia; Silva, Sónia; Rodrigues, Irene; Munhá, João; Gomes, João Paulo; Macedo, RitaMycobacteroides abscessus complex (MAB), a fast-growing nontuberculous mycobacterium, is emerging as a significant infectious disease threat, due to both intrinsic and acquired resistance mechanisms to antibiotics and disinfectants and the need for extensive and multidrug regimens for treatment. Despite the prolonged regimens, outcomes are poor and persistence cases have been reported. Here, we describe clinical, microbiologic and genomic features of a M. abscessus subsp. bolletii (M. bolletii) strain consecutively isolated from a patient within an eight-year infection period. From April 2014 to September 2021, the National Reference Laboratory for Mycobacteria received eight strains isolated from a male patient. Species identification, molecular resistance profile and phenotypic drug susceptibility were determined. Five of these isolates were recovered for further in-depth genomic analysis. Genomic analysis confirmed the multidrug resistant pattern of the strain and also other genetic changes associated with adaptation to environment and defence mechanisms. We highlight the identification of new mutations in locus MAB_1881c and in locus MAB_4099c (mps1 gene), already described as associated with macrolides resistance and morphotype switching, respectively. Additionally, we also observed the emergence and fixation of a mutation in locus MAB_0364c that appeared at a frequency of 36% for the 2014 isolate, 57% for the 2015 isolate and 100% for the 2017 and 2021 isolates, clearly illustrating a fixation process underlying a microevolution of the MAB strain within the patient. Altogether these results suggest that the observed genetic alterations are a reflection of the bacterial population's continuous adaptation and survival to the host environment during infection, contributing to persistence and treatment failure.
- Rapid drug resistance prediction in positive clinical samples using an extensive targeted next-generation sequencing panelPublication . Rosendal, Ebba; Isidro, Joana; Carneiro, Sofia; Gomes, João Paulo; Macedo, RitaTuberculosis (TB) remains a global health challenge, exacerbated by the emergence of drug-resistant strains. Most methods for drug susceptibility testing (DST) are culture-dependent and time consuming, possibly delaying optimal TB-treatment. This study aimed to develop an extensive targeted next-generation sequencing (tNGS) approach for rapid genotypic DST directly from clinical samples. We designed a tNGS panel comprising 30 amplicons targeting 19 genomic regions associated with resistance to 20 antibiotics. This method was applied to 71 smear-positive (0-3+) pulmonary TB clinical samples collected at the Portuguese National Reference Laboratory. DNA was extracted and amplified using multiplex PCRs, followed by sequencing on Oxford Nanopore Technologies MinION platform. Sequencing data were using TB-Profiler and the tNGS results compared to phenotypic DST and whole genome sequencing (WGS) data from corresponding isolates. The tNGS demonstrated high concordance with both phenotypic and WGS-based DST across different sample types and smear positivity levels. For first-line drugs, tNGS showed 88% categorical agreement (CA) with pDST, increasing to 97% when excluding undetermined results. Compared to WGS across all analysed antibiotics, tNGS achieved 92% CA, increasing to >99% when excluding undetermined results. Validation of the tNGS panel showed 90% (1,895/2,076) of amplicons reaching >10x coverage at all analysed positions and 43 (61%) samples with all complete amplicons above this threshold. Non-specific amplification of contaminant bacterial DNA was minimal, with most mapped off-target reads being of human origin. This method enables comprehensive resistance prediction directly from clinical samples and signifies an important development in TB diagnostics and resistance monitoring.
- Seroprevalence of meningococcal serogroup C bactericidal antibodies in the Portuguese population, a decade after vaccine introduction in the National Immunisation ProgrammePublication . Gonçalves, Paulo; Sáez-López, Emma; Carneiro, Sofia; Simões, Maria JoãoBackground: The incidence of invasive meningococcal disease due to serogroup C (MenC) decreased in Portugal since the introduction of the conjugate vaccine (MCC) in the free market in 2001 and in the National Immunisation Plan in 2006. Considering the potential waning of the antibody response reported in the literature, the different vaccination schemes that were used in our country over the past decade, and that Neisseria meningitidis serogroup C continues to circulate, the Portuguese population may currently be at increased risk of infection. In the absence of national data, we evaluated the seroprotection level of the Portuguese population against MenC, in order to identify the protected fraction of the population and ponder on the necessity of a booster dose of the MCC vaccine. Methods: We measured serum bactericidal antibody levels against MenC in a representative sample of the population (n = 1500) aged 2-64 years who participated in the 2015/2016 National Serological Survey. Results: A total of 31.1% (466/1500, 95%CI: 29-33%) of the individuals studied were protected against MenC. The geometric mean titre was 6.5. The proportion of seroprotected was particularly low in children aged 2-4 years (<16%) who received a single dose of the vaccine at 12 months of age (vaccination strategy since 2012). The proportion of seroprotected was higher (44.7% to 53.5%) in adolescent and young adults (15-24 years of age), resulting from vaccination during the catch-up campaign at 5-15 years of age. The highest protection rates were observed when the vaccine was administered during adolescence. Conclusion: The small fraction of population seroprotected, combined with the already known waning effect of the antibody response over time, may indicate that the Portuguese population will become progressively more exposed to the risk of infection. Taking in consideration our results, we recommend to change the current vaccination strategy and introduce a booster dose of the MCC vaccine during adolescence.