Browsing by Author "Carneiro, Catarina"
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- Isolates from hospital environments are the most virulent of the Candida parapsilosis complexPublication . Sabino, Raquel; Sampaio, Paula; Carneiro, Catarina; Rosado, Laura; Pais, CéliaBackground: Candida parapsilosis is frequently isolated from hospital environments, like air and surfaces, and causes serious nosocomial infections. Molecular studies provided evidence of great genetic diversity within the C. parapsilosis species complex but, despite their growing importance as pathogens, little is known about their potential to cause disease, particularly their interactions with phagocytes. In this study, clinical and environmental C. parapsilosis isolates, and strains of the related species C. orthopsilosis and C. metapsilosis were assayed for their ability to induce macrophage cytotocixity and secretion of the pro-inflammatory cytokine TNF-a, to produce pseudo-hyphae and to secrete hydrolytic enzymes. Results: Environmental C. parapsilosis isolates caused a statistically significant (p = 0.0002) higher cell damage compared with the clinical strains, while C. orthopsilosis and C. metapsilosis were less cytotoxic. On the other hand, clinical isolates induced a higher TNF-a production compared with environmental strains (p < 0.0001). Whereas the amount of TNF-a produced in response to C. orthopsilosis strains was similar to the obtained with C. parapsilosis environmental isolates, it was lower for C. metapsilosis strains. No correlation between pseudo-hyphae formation or proteolytic enzymes secretion and macrophage death was detected (p > 0.05). However, a positive correlation between pseudo-hyphae formation and TNF-a secretion was observed (p = 0.0119). Conclusions: We show that environmental C. parapsilosis strains are more resistant to phagocytic host defences than bloodstream isolates, being potentially more deleterious in the course of infection than strains from a clinical source. Thus, active environmental surveillance and application of strict cleaning procedures should be implemented in order to prevent cross-infection and hospital outbreaks.
- Using whole genome sequencing to understand host-nontuberculous mycobacteria interactionPublication . Sousa, Sara; Borges, Victor; Faria, Sónia; Carneiro, Catarina; Vieira, Luís; Gomes, João Paulo; Jordão, LuísaBackground: Nontuberculous mycobacteria (NTM) are a large group of Mycobacterium species that don’t belong to the Mycobacterium tuberculosis (Mtb) complex. These bacteria are mostly environmental being regarded as etiological agents of opportunistic infection in humans mainly immunocompromised. Distinguishing NTM from Mtb is still a challenge and identification at the species level is essential for an accurate diagnostic and effective treatment. The growth rate of NTM is very important for the onset of treatment being these bacteria divided into rapidly growing mycobacteria (RGM) and slowly growing mycobacteria (SGM). Mycobacterium fortuitum, M. abscessus and the model organism M. smegmatis are RGM whereas M. avium is SGM that take more than 7 days to form CFU on growth media. The knowledge of NTM infections is still reduced being needed more studies to understand the host-pathogen interaction during the infectious process in order to establish more effective therapeutic schemes. Recently our group conducted a study using human alveolar macrophages as a model. The obtained data showed that both RGM (747/08) and SGM (60/08) were able to persist and even replicate within this macrophages whereas others do not (M.smegmatis and M.abscessus)1. Here we used NGS as a tool to identify bacterial factor responsible for the observed outcome. Materials and Methods: Three reference strains (M.fortuitum ATCC6841, M.avium ATCC25291, M.smegmatis (ATCC700084) and 3 strains from Ricardo Jorge mycobacterial collection isolated from patients (M.fortuitum 747/08, M. avium 60/08 and Mtb70/09) were used. For DNA extraction bacteria were grown in Middlebrook 7H9 supplemented with 10% OADC and 0.05% Tween80. Full genome sequence was performed using NGS platform MiSeq (Illumina Inc., San Diego,CA, USA) according to the manufacturer’s instructions. Data analysis: RAST (www.rast.nmpdr.org) and MAUVE platforms were used for annotation and multiple alignments, respectively.3 Results: Data analysis suggest a link between mycobacteria growth rate and genome size with RGM (6.7 million bp) having longer genomes than SGM (4.8 million bp) what might reflect bacteria adaptation to the host(s). Mtb with an exclusive host has a shorter genome (4.3 million bp) than NTM which exhibit a wider host tropisms and the ability to persist within the environment Therefore, we can find genes associated to the Lactate fermentation like MAV_2543 and Methanogenesis xfp that cannot be found in Mtb. The two NTM (747/08 and 60/08) strains sequenced and that show intramacrophage persistence displayed also a longer size, which is associated to the persistence related genes. In specific gene subsystems were observed intra-species differences between clinical and reference NTM. Fatty acid metabolism cluster and cell envelope related genes subsystems illustrate this result. Conclusion: The preliminary results suggest a link between genome size and intracellular persistence and support the fact that clinic NTM share virulence factors with Mtb. However, only a transcriptional analysis could ensure the evolvement of these genes in intracellular persistence.