Paula Sampaio | Universidade do Minho (original) (raw)

Papers by Paula Sampaio

Curr Clin Micro Rpt, 2014

Candida species are an increasing cause of invasive candidiasis (IC), resulting in significant mo... more Candida species are an increasing cause of invasive candidiasis (IC), resulting in significant mortality and morbidity. Although Candida albicans remains the predominant etiologic agent of invasive fungal infections in hospital settings, the incidence of new infections from non-albicans Candida species is steadily increasing, and important geographical differences in species distribution and patterns of in vitro antifungal susceptibilities have been reported. In this article, we review the recent epidemiology of Candida invasive infections, and discuss trends in incidence, mortality, species distribution, and antifungal resistance, emphasizing the particular situation of C. glabrata. We also address the techniques available for diagnosis of IC, the classic culture techniques and the non-culture based methods, including the recent recommendation of the European Fungal Infection Study Group (EFISG) of the European Society of Clinical Microbiology and Infectious Diseases (ESCMID) about different techniques and procedures to detect IC, considering specificities with C. glabrata detection.

PLoS ONE, 2010

The incidence of Candida albicans infections and the relapse episodes after antifungal treatment ... more The incidence of Candida albicans infections and the relapse episodes after antifungal treatment have increased in recent decades. Recurrences are mainly due to the persistence of the original infecting strain that may present genetic and genomic rearrangements during interaction with the host, reflecting strain adaptation. In this study, four isolates recovered from a patient during recurrent candidemia episodes were genotyped by microsatellite length polymorphism (MLP) and by multilocus sequence typing (MLST) and found to be genetic variants of the same strain. Using experimental mouse infections, a progressive reduction in the virulence of the four isolates was observed, with the first two isolates more virulent than the third and fourth. Additionally, in the mouse model, the first isolate resisted host control more efficiently, resulting in higher kidney fungal burdens and necrosis as compared to the third isolate. The resolution of inflammation was delayed in mice challenged with the first isolate and the message for IFN-c and TNF-a in the spleen was lower within the first few hours post-infection. Original and recurrent isolates also displayed different phenotypes regarding activity of secreted enzymes and response to stress agents. Overall, the comparative analysis indicated that the virulence decrease of these isolates was related to a lower ability to resist to the host anticandida effector mechanisms. We showed for the first time that C. albicans genetic variants of the same strain, sequentially isolated from an immunocompromised patient, underwent adaptations in the human host that resulted in virulence attenuation when tested in mice.

Brazilian Journal of Microbiology, 2014

Microbial culture collections underpin biotechnology applications and are important resources for... more Microbial culture collections underpin biotechnology applications and are important resources for clinical microbiology by supplying reference strains and/or performing microbial identifications as a service. Proteomic profiles by MALDI-TOF MS have been used for Candida spp. identification in clinical laboratories and demonstrated to be a fast and reliable technique for the routine identification of pathogenic yeasts. The main aim of this study was to apply MALDI-TOF MS combined with classical phenotypic and molecular approaches to identify Candida clinical isolates preserved from 1 up to 52 years in a Brazilian culture collection and assess its value for the identification of yeasts preserved in this type of collections. Forty Candida spp. clinical isolates were identified by morphological and biochemical analyses. Identifications were also performed by the new proteomic approach based on MALDI-TOF MS. Results demonstrated 15% discordance when compared with morphological and biochemical analyses. Discordant isolates were analysed by ITS sequencing, which confirmed the MALDI-TOF MS identifications and these strains were renamed in the culture collection catalogue. In conclusion, proteomic profiles by MALDI-TOF MS represents a rapid and reliable method for identifying clinical Candida species preserved in culture collections and may present clear benefits when compared with the performance of existing daily routine methods applied at health centres and hospitals.

PLoS ONE, 2014

Candida albicans cell wall is important for growth and interaction with the environment. RLM1 is ... more Candida albicans cell wall is important for growth and interaction with the environment. RLM1 is one of the putative transcription factors involved in the cell wall integrity pathway, which plays an important role in the maintenance of the cell wall integrity. In this work we investigated the involvement of RLM1 in the cell wall biogenesis and in virulence. Newly constructed C. albicans D/Drlm1 mutants showed typical cell wall weakening phenotypes, such as hypersensitivity to Congo Red, Calcofluor White, and caspofungin (phenotype reverted in the presence of sorbitol), confirming the involvement of RLM1 in the cell wall integrity. Additionally, the cell wall of C. albicans D/Drlm1 showed a significant increase in chitin (213%) and reduction in mannans (60%), in comparison with the wild-type, results that are consistent with cell wall remodelling. Microarray analysis in the absence of any stress showed that deletion of RLM1 in C. albicans significantly down-regulated genes involved in carbohydrate catabolism such as DAK2, GLK4, NHT1 and TPS1, up-regulated genes involved in the utilization of alternative carbon sources, like AGP2, SOU1, SAP6, CIT1 or GAL4, and genes involved in cell adhesion like ECE1, ALS1, ALS3, HWP1 or RBT1. In agreement with the microarray results adhesion assays showed an increased amount of adhering cells and total biomass in the mutant strain, in comparison with the wild-type. C. albicans mutant D/Drlm1 strain was also found to be less virulent than the wild-type and complemented strains in the murine model of disseminated candidiasis. Overall, we showed that in the absence of RLM1 the modifications in the cell wall composition alter yeast interaction with the environment, with consequences in adhesion ability and virulence. The gene expression findings suggest that this gene participates in the cell wall biogenesis, with the mutant rearranging its metabolic pathways to allow the use of alternative carbon sources.

BMC Microbiology, 2011

Background Candida parapsilosis is frequently isolated from hospital environments, like air and s... more Background 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-α, 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...

Journal of Microbiological Methods, 2014

Herein we developed a method based on the quenching effect of propidium iodide over Sytox-Green f... more Herein we developed a method based on the quenching effect of propidium iodide over Sytox-Green fluorescence to assess yeast phagocytosis by flow cytometry. It allows accurate quantification of living from dead phagocytes; internalized from non-internalized cells, maintaining yeast fluorescence within phagocytes; and the different associations between phagocytes and fungal cells.

Immunogenetics of Fungal Diseases

The advances in medicine have achieved great benefits by improving or even eliminating various de... more The advances in medicine have achieved great benefits by improving or even eliminating various debilitating diseases or malignancies, expanding life expectancy. However, it has also originated the development of a compromised population susceptible to opportunistic diseases. A global major concern is the emergence and spread of life-threatening invasive infections in immunocompromised patients, in which the opportunistic fungal infections have greatly increased in the last decades. The high mortality rates associated with these infections, which remain as high as 40%, are due to the limited therapeutic options and the emergence of drug resistant fungi, but also due to the lack of efficient early diagnosis. Consequently, these facts led to the opinion that new approaches are needed to improve the outcome of these patients, such as immunopreventive strategies that could even be combined with standard antifungal treatment. In view of the proven effectiveness of various antibacterial and antiviral vaccines in preventing the respective diseases, several works have been developed to induce protective immunity against fungal infections as well. The better understanding of how the immune system works against fungal pathogens has made possible to explore immunomodulatory strategies that can protect both immunocompetent and immunocompromised hosts and generate memory. Recently, two fungal vaccines against Candida have advanced through clinical trials. However, there are still many challenges in the development of an efficient vaccine against invasive fungal infections. We will provide an update on the progress made in immunization against fungal infections, reviewing host-fungi interactions, antigens and adjuvants exploited in vaccine strategies, and discuss concerns that need to be overcome to further advance in the area of fungal vaccines.

Microorganisms

Four yeast isolates from the species—Apiotrichum brassicae, Candida tropicalis, Metschnikowia pul... more Four yeast isolates from the species—Apiotrichum brassicae, Candida tropicalis, Metschnikowia pulcherrima, and Pichia kudriavzevii—previously selected by their oleaginous character and growth flexibility in different carbon sources, were tested for their capacity to convert volatile fatty acids into lipids, in the form of single cell oils. Growth, lipid yields, volatile fatty acids consumption, and long-chain fatty acid profiles were evaluated in media supplemented with seven different volatile fatty acids (acetic, butyric, propionic, isobutyric, valeric, isovaleric, and caproic), and also in a dark fermentation effluent filtrate. Yeasts A. brassicae and P. kudriavzevii attained lipid productivities of more than 40% (w/w), mainly composed of oleic (>40%), palmitic (20%), and stearic (20%) acids, both in synthetic media and in the waste-derived effluent filtrate. These isolates may be potential candidates for single cell oil production in larger scale applications by using alterna...

Journal of Fungi

A new and easy polymerase chain reaction (PCR) multiplex strategy, for the identification of the ... more A new and easy polymerase chain reaction (PCR) multiplex strategy, for the identification of the most common fungal species involved in invasive fungal infections (IFI) was developed in this work. Two panels with species-specific markers were designed, the Candida Panel for the identification of Candida species, and the Filamentous Fungi Panel for the identification of Aspergillus species and Rhizopusarrhizus. The method allowed the correct identification of all targeted pathogens using extracted DNA or by colony PCR, showed no cross-reactivity with nontargeted species and allowed identification of different species in mixed infections. Sensitivity reached 10 to 1 pg of DNA and was suitable for clinical samples from sterile sites, with a sensitivity of 89% and specificity of 100%. Overall, the study showed that the new method is suitable for the identification of the ten most important fungal species involved in IFI, not only from positive blood cultures but also from clinical sampl...

Immunogenetics of Fungal Diseases, 2017

BMC Microbiology

Background Over the last years oleaginous yeasts have been studied for several energetic, oleoche... more Background Over the last years oleaginous yeasts have been studied for several energetic, oleochemical, medical and pharmaceutical purposes. However, only a small number of yeasts are known and have been deeply exploited. The search for new isolates with high oleaginous capacity becomes imperative, as well as the use of alternative and ecological carbon sources for yeast growth. Results In the present study a high-throughput screening comprising 366 distinct yeast isolates was performed by applying an optimised protocol based on two approaches: (I) yeast cultivation on solid medium using acetic acid as carbon source, (II) neutral lipid estimation by fluorimetry using the lipophilic dye Nile red. Conclusions Results showed that, with the proposed methodology, the oleaginous potential of yeasts with broad taxonomic diversity and variety of growth characteristics was discriminated. Furthermore, this work clearly demonstrated the association of the oleaginous yeast character to the stra...

Frontiers in Immunology

Candida albicans causes systemic life-threatening infections, particularly in immunocompromised i... more Candida albicans causes systemic life-threatening infections, particularly in immunocompromised individuals, such as patients in intensive care units, patients undergoing chemotherapy, and post-surgical and neutropenic patients. The proliferation of invading Candida cells is mainly limited by the action of the human innate immune system, in which phagocytic cells play a fundamental role. This function is, however, limited in neutropenic patients, who rely mainly on the protective immunity mediated by macrophages. Macrophages have been shown to release extracellular DNA fibers, known as macrophage extracellular traps (METs), which can entrap and kill various microbes by a process called ETosis. In this study, we observed that, upon contact with C. albicans, macrophages became active in phagocyting and engulfing yeast cells. ETosis was induced in 6% of macrophages within the first 30 min of contact, and this percentage increased with the multiplicity of infection until a plateau was reached. After 2.5 h incubation, the presence of extracellular macrophage DNA was observed in approximately half of the cells. This study suggests that the formation of METs occurs before pyroptosis (first 6-8 h) and macrophage cell death (up to 24 h), and thus, METs could be included in models describing C. albicans-macrophage interactions. We also observed that macrophage ETosis and phagocytosis can occur simultaneously and that, in the first hours of infection, both processes are similarly important in controlling the proliferation of yeast cells, this being critical in neutropenic patients. Finally, it can also be concluded that, since C. albicans can degrade DNA, the structural component of METs, yeast extracellular DNase activity can be considered as an important virulence factor.

FEMS Yeast Research

This is one of the first studies showing the capacity of natural yeast isolates to produce releva... more This is one of the first studies showing the capacity of natural yeast isolates to produce relevant amounts of succinic acid, giving clues for further strain improvement.

FEATURES Location/Qualifiers source 1..164 /organism="Candida albicans" /mol_type="genomic DNA" /... more FEATURES Location/Qualifiers source 1..164 /organism="Candida albicans" /mol_type="genomic DNA" /db_xref="taxon:5476" /chromosome="4" repeat_region 4..52 /note="microsatellite CAI-16" /rpt_type=tandem ORIGIN 1 actgcaacaa ctgcaacaac aacaacaaca acaacagcag cagcagcagc agaatgccca 61 acagcaacta gaagtacctg ttatcccaat acaaacacaa acatcacaac aaatggcttc 121 aactaccaat cacaaatcag ctaatctaat accagggttt ttac //

FEATURES Location/Qualifiers source 1..176 /organism="Candida albicans" /mol_type="genomic DNA" /... more FEATURES Location/Qualifiers source 1..176 /organism="Candida albicans" /mol_type="genomic DNA" /db_xref="taxon:5476" /chromosome="4" repeat_region 4..64 /note="microsatellite CAI-20" /rpt_type=tandem ORIGIN 1 actgcaacaa ctgcaacaac aacaacaaca acaacaacaa caacaacagc aacaacagca 61 gcagaatgcc caacagcaac tagaagtacc tgttatccca acacaaacac aaacatcaca 121 acaaatggct tcaactacca atcacaaatc agctaatcta ataccagggt ttttac //

FEATURES Location/Qualifiers source 1..197 /organism="Candida albicans" /mol_type="genomic DNA" /... more FEATURES Location/Qualifiers source 1..197 /organism="Candida albicans" /mol_type="genomic DNA" /db_xref="taxon:5476" /chromosome="4" repeat_region 4..85 /note="microsatellite CAI-27" /rpt_type=tandem ORIGIN 1 actgcaacaa ctgcaacaac aacaacaaca acaacaacaa caacaacaac aacaacaaca 61 acaacaacag caacaacagc agcagaatgc ccaacagcaa ctagaagtac ctgttatccc 121 aacacaaaca caaacatcac aacaaatggc ttcaactacc aatcacaaat cagctaatct 181 aataccaggg tttttac //

FEATURES Location/Qualifiers source 1..200 /organism="Candida albicans" /mol_type="genomic DNA" /... more FEATURES Location/Qualifiers source 1..200 /organism="Candida albicans" /mol_type="genomic DNA" /db_xref="taxon:5476" /chromosome="4" repeat_region 4..88 /note="microsatellite CAI-28" /rpt_type=tandem ORIGIN 1 actgcaacaa ctgcaacaac aacaacaaca acaacaacaa caacaacaac aacaacaaca 61 acaacaacaa cagcaacaac agcagcagaa tgcccaacag caactagaag tacctgttat 121 cccaatacaa acacaaacat cacaacaaat ggcttcaact accaatcaca aatcagctaa 181 tctaatacca gggtttttac //