Aida Duarte | Universidade de Lisboa (original) (raw)

Papers by Aida Duarte

Research paper thumbnail of Spread of In58 containing blavim-2 among Pseudomonas aeruginosa

Clinical Microbiology and Infection, 2006

Research paper thumbnail of Persistência de Klebsiella pneumoniae em doentes de unidades pediátricas do Hospital de Santa Maria, em Lisboa

Portuguese Journal of Pediatrics, 2011

A Klebsiella pneumoniae é responsável por infecções nosocomiais e adquiridas na comunidade. A pro... more A Klebsiella pneumoniae é responsável por infecções nosocomiais e adquiridas na comunidade. A produção de β-lactamases de espectro alargado (ESBLs) e os factores de virulência assumem particular importância nestas infecções. Métodos: Este estudo incluiu 47 isolados de K. pneumoniae, obtidos de dez pacientes de unidades pediátricas de um hospital central, entre 2001 e 2009. Os antibiogramas foram determinados pelo método da difusão em disco. A caracterização genotípica foi efectuada por M13 fingerprinting e, nos perfis representativos, por Multilocus Sequence Typing (MLST). Os genes de resistência bla CTX-M-1 , bla CTX-M-2 e bla SHV , e os genes de virulência k 2 A, rmpA, magA, fimH, mrkD, khe e iucC, foram identificados por reacção em cadeia da polimerase e sequenciados. Resultados: Encontraram-se três antibiotipos predominantes, R1, R2 e R3, de acordo com a susceptibilidade dos isolados à cefotaxima, ceftazidima, ciprofloxacina e gentamicina. Todos os isolados eram sensíveis ao imipenemo. O M13 fingerprinting distinguiu dois perfis genómicos principais, M1 e M2, detectados em vários momentos entre 2001 e 2009. Entre os isolados com perfil M1, 38,5% tinham o antibiotipo R1 e 53,8% tinham o antibiotipo R2; no perfil M2, 12,5% e 56,25% dos isolados apresentavam os antibiotipos R1 e R3, respectivamente. Verificou-se uma associação entre o perfil M1 e o Sequence Type (ST) 15, assim como entre o perfil M2 e os STs 133 e 276. 89,4% dos isolados possuíam genes codificantes para ESBLs; a sequenciação identificou os genes das enzimas CTX-M-15, SHV-1,-2a e-11. A incidência dos genes fimH, mrkD e khe foi elevada; k 2 A, rmpA, magA e iucC não foram detectados. Discussão e conclusão: A presença dos mesmos perfis de M13 fingerprinting durante alguns anos sugere uma endemia nas unidades pediátricas do hospital. Além disso, a elevada incidência dos genes das adesinas fimbriais pode justificar a persistência do mesmo clone no mesmo doente durante longos períodos de tempo.

Research paper thumbnail of Starch-based Pickering emulsions for topical drug delivery: A QbD approach

Colloids and Surfaces B: Biointerfaces, Nov 1, 2015

Pickering emulsions are stabilized by solid particles instead of surfactants and have been widely... more Pickering emulsions are stabilized by solid particles instead of surfactants and have been widely investigated in pharmaceutical and cosmetic fields since they present less adverse effects than the classical emulsions. A quality by design (QbD) approach was applied to the production of w/o emulsions stabilized by starch. A screening design was conducted to identify the critical variables of the formula and the process affecting the critical quality properties of the emulsion (droplet size distribution). The optimization was made by establishing the design space, adjusting the concentration of starch and the quantity of the internal aqueous phase. The emulsion production process was, in turn, adjusted by varying the time and speed of stirring, to ensure quality and minimum variability. The stability was also investigated, demonstrating that an increase in starch concentration improves the stability of the emulsion. Rheological and mechanical studies indicated that the viscosity of the emulsions was enhanced by the addition of starch and, to a higher extent, by the presence of different lipids. The developed formulations was considered non-irritant, by an in vitro assay using human cells from skin (Df and HaCat) with the cell viability higher than 90% and, with self-preserving properties. Finally, the QbD approach successfully built quality in Pickering emulsions, allowing the development of hydrophilic drug-loaded emulsions stabilized by starch with desired organoleptic and structural characteristics. The results obtained suggest that these systems are a promising vehicle to be used in products for topical administration.

Research paper thumbnail of Bioactive Diterpenoids, a New Jatrophane and Two <i>e</i><i>nt</i>-Abietanes, and Other Constituents from <i>Euphorbia </i><i>p</i><i>ubescens</i>

Journal of Natural Products, Apr 23, 2004

A new jatrophane diterpene, pubescenol (1), known ent-abietane lactones, helioscopinolide A (2) a... more A new jatrophane diterpene, pubescenol (1), known ent-abietane lactones, helioscopinolide A (2) and B (3), and taraxerone, 24-methylenecycloartanol, and vanillin have been isolated from Euphorbia pubescens. Diterpenes 1-3 and previously described pubescene D (4) were shown to be moderate inhibitors of the growth of MCF-7, NCI-H460, and SF-268 human tumor cell lines, whereas compounds 2 and 3 also exhibited significant antibacterial activity against Staphylococcus aureus.

Research paper thumbnail of Characterization of Portuguese <i>Thymbra capitata</i> , <i>Thymus caespititius</i> and <i>Myrtus communis</i> essential oils in topical formulations

Flavour and Fragrance Journal, Jun 5, 2017

As the personal care industry evolves, formulators are seeking innovative solutions for their for... more As the personal care industry evolves, formulators are seeking innovative solutions for their formulations' needs, and for ingredients that can offer multiple functions within formulations. Considering that essential oils (EOs) may present a wide spectrum of biological activities, the composition, antimicrobial and antioxidant activity of Portuguese Thymbra capitata, Thymus caespititius and Myrtus communis EOs were assessed in order to evaluate them as preservatives and antioxidants in topical emulsions. The in vivo safety application of some emulsions was also tested. T. capitata EO was mainly constituted by carvacrol (73%), whereas α-terpineol (27%), p-cymene (14%) and carvacrol (10%) dominated Th. caespititius EO, and 1,8-cineole (37%) was dominant in M. communis EO. The minimum inhibitory concentration of T. capitata, Th. caespititius and M. communis EOs was 0.4 μg/mL against C. albicans and ranged between 0.4 and 30.7 μg/mL against A. brasiliensis. Gram-positive bacteria were more susceptible to each EO than Gram-negative bacteria. T. capitata EO showed significantly higher antioxidant activity than Th. caespititius and M. communis EOs. EOs incorporated in emulsions showed preservative activity against all microorganisms tested and T. capitata EO emulsions showed powerful reactive oxygen species (ROS) scavenging effects. A safety evaluation study was performed with 0.10% and 0.01% T. capitata EO emulsions according to the Human Repeat Insult Patch Test (HRIPT). All emulsions were considered safe for topical application. T. capitata, Th. caespititius and M. communis significantly improved the microbiological quality of the prepared emulsions and may constitute a powerful alternative to the current preservatives and antioxidants for use in topical formulations.

Research paper thumbnail of <i>In vitro</i> Antimicrobial Activity of Royleanone Derivatives Against Gram-Positive Bacterial Pathogens

Phytotherapy Research, Mar 11, 2013

Infections caused by multiresistant bacterial pathogens are a significant problem worldwide, turn... more Infections caused by multiresistant bacterial pathogens are a significant problem worldwide, turning the search for natural compounds to act as alternatives to antibiotics of major importance. The aim of the present study was to investigate the in vitro antimicrobial activity of 7α-acetoxy-6β-hydroxyroyleanone (1), isolated from Plectranthus grandidentatus (Lamiaceae), and 11 additional royleanone abietane derivatives of 1 (2-12) against important Gram-positive human bacterial pathogens. Results showed that the aromatic and alkylic esters 2, 3 and 5 are more active than 1 against Enterococcus and Staphylococcus (minimum inhibitory concentration (MIC) values ranging from 0.98 to 62.50 µg/mL). Moreover, 7α-acetoxy-6β-hydroxy-12-O-(4-chloro)benzoylroyleanone (2) gave rise to a new antibacterial-prototype (MIC values of 3.91-15.63 µg/mL against Staphylococcus and of 0.98-3.91 µg/mL against Enterococcus). The results showed that the compounds under analysis also present antimicrobial activity against resistant bacteria. The hydrophobic extra-interactions with bacterial targets seem to play an important role on the activity of royleanones derivatives.

Research paper thumbnail of Neoclerodane and Labdane Diterpenoids from <i>Plectranthus </i><i>o</i><i>rnatus</i>

Journal of Natural Products, Aug 22, 2002

Three new diterpenoids, a neoclerodane and two labdane derivatives, have been isolated from an ac... more Three new diterpenoids, a neoclerodane and two labdane derivatives, have been isolated from an acetone extract of Plectranthus ornatus. The structures of these compounds (plectrornatins A-C, 1-3, respectively) were established mainly by spectroscopic means, particularly by 1D and 2D NMR studies and, in the case of the neoclerodane 1, by an X-ray diffraction analysis. Compounds 1 and 3 showed moderate antimicrobial activity against five Candida species and selected Gram negative and Gram positive bacteria strains.

Research paper thumbnail of Antibiotic activity of five probiotics strains against five multirresistant clinical isolates

Research paper thumbnail of Liposomal Delivery of Newly Identified Prophage Lysins in a Pseudomonas aeruginosa Model

International Journal of Molecular Sciences

Pseudomonas aeruginosa is a Gram-negative opportunistic bacterium that presents resistance to sev... more Pseudomonas aeruginosa is a Gram-negative opportunistic bacterium that presents resistance to several antibiotics, thus, representing a major threat to human and animal health. Phage-derived products, namely lysins, or peptidoglycan-hydrolyzing enzymes, can be an effective weapon against antibiotic-resistant bacteria. Whereas in Gram-positive bacteria, lysis from without is facilitated by the exposed peptidoglycan layer, this is not possible in the outer membrane-protected peptidoglycan of Gram-negative bacteria. Here, we suggest the encapsulation of lysins in liposomes as a delivery system against Gram-negative bacteria, using the model of P. aeruginosa. Bioinformatic analysis allowed for the identification of 38 distinct complete prophages within 66 P. aeruginosa genomes (16 of which newly sequenced) and led to the identification of 19 lysins of diverse sequence and function, 5 of which proceeded to wet lab analysis. The four purifiable lysins showed hydrolytic activity against Gr...

Research paper thumbnail of Genomic epidemiological analysis of Klebsiella pneumoniae from Portuguese hospitals reveals insights into circulating antimicrobial resistance

Scientific Reports

Klebsiella pneumoniae (Kp) bacteria are an increasing threat to public health and represent one o... more Klebsiella pneumoniae (Kp) bacteria are an increasing threat to public health and represent one of the most concerning pathogens involved in life-threatening infections and antimicrobial resistance (AMR). To understand the epidemiology of AMR of Kp in Portugal, we analysed whole genome sequencing, susceptibility testing and other meta data on 509 isolates collected nationwide from 16 hospitals and environmental settings between years 1980 and 2019. Predominant sequence types (STs) included ST15 (n = 161, 32%), ST147 (n = 36, 7%), ST14 (n = 26, 5%) or ST13 (n = 26, 5%), while 31% of isolates belonged to STs with fewer than 10 isolates. AMR testing revealed widespread resistance to aminoglycosides, fluoroquinolones, cephalosporins and carbapenems. The most common carbapenemase gene was blaKPC-3. Whilst the distribution of AMR linked plasmids appears uncorrelated with ST, their frequency has changed over time. Before year 2010, the dominant plasmid group was associated with the extende...

Research paper thumbnail of A phylogenomic approach for the analysis of colistin resistance-associated genes in Klebsiella pneumoniae, its mutational diversity and implications for phenotypic resistance

International Journal of Antimicrobial Agents

The emergence of carbapenemase-producing Klebsiella pneumoniae strains has triggered the use of o... more The emergence of carbapenemase-producing Klebsiella pneumoniae strains has triggered the use of old antibiotics such as colistin which is consequently driving the emergence of colistin resistance in already multidrug resistant strains that can underlie life-threatening infections. This study analyses the mutational diversity of 22 genes associated with colistin resistance in 140 K. pneumoniae clinical isolates integrated in a high-resolution phylogenetic scenario. Colistin susceptibility was accessed by broth microdilution categorizing 98 isolates as susceptible and 16 as resistant, 10 of which were carbapenemase producers. Across the 22 genes examined, 171 non-synonymous mutations and 9 mutations associated with promoter regions were found. Eighty-five isolates had a truncation and/or deletion in at least one of the 22 genes. However, only seven mutations, the complete deletion of mgrB or insertion sequence mediated disruption were exclusively observed in resistant isolates, 4 of these (mgrBIle13fs, pmrBGly207Asp, phoQHis339Asp and ramAIle28Met) comprising novel mutations potentially involved in colistin resistance. The ISEcp1-blaCTX-M-15::mgrB disruption is herein noteworthy as a mechanism underlying co-resistance to third-generation cephalosporins and colistin. Moreover, the high-resolution phylogenetic context shows that most of the mutational diversity spans across multiple phylogenetic clades and that most of the mutations previously associated with colistin resistance are clade associated and present in susceptible isolates, showing no correlation with colistin resistance. In conclusion, the present study contributes with relevant data towards the genetic background of genes involved with colistin resistance deeply rooted across monophyletic groups providing a better understanding of the genes and mutations involved in colistin resistance.

Research paper thumbnail of Article Exploring Dangerous Connections between Klebsiella pneumoniae

Healthcare-associated infections (HAI) are a huge public health concern, particularly when the et... more Healthcare-associated infections (HAI) are a huge public health concern, particularly when the etiological agents are multidrug resistant. The ability of bacteria to develop biofilm is a helpful skill, both to persist within hospital units and to increase antibiotic resistance. Although the links between antibiotic resistance, biofilms assembly and HAI are consensual, little is known about biofilms. Here, electron microscopy was adopted as a tool to investigate biofilm structures associated with increased antibiotic resistance. The K. pneumoniae strains investigated are able to assemble biofilms, albeit with different kinetics. The biofilm structure and the relative area fractions of bacteria and extracellular matrix depend on the particular strain, as well as the minimal inhibitory concentration (MIC) for the antibiotics. Increased values were found for bacteria organized in biofilms when compared to the respective planktonic forms, except for isolates Kp45 and Kp2948, the MIC values for which remained unchanged for fosfomycin. Altogether, these results showed that the emergence of antimicrobial resistance among bacteria responsible for HAI is a multifactorial phenomenon dependent on antibiotics and on bacteria/biofilm features.

Research paper thumbnail of Antimicrobial activity of the diterpene 7 a-acetoxy-6b-hidroxyroyleanone and its 6b-carbamoyl derivative

Research paper thumbnail of Antimicrobial Activity of (11R*, 13 E)-11-Acetoxyhalima-5-13-dien-15-oic Acid and its Methyl Ester Derivative

Research paper thumbnail of Plasmid-mediated CMY-2 AmpC beta lactamase and CTX-M-14 extended-spectrum beta-lactamase in Proteus mirabilis clinical isolate

Clinical Microbiology and Infection, 2008

Research paper thumbnail of Identification of an OXA-58 carbapenemase in an unusual Portuguese Acinetobacter baumannii clinical isolate

Research paper thumbnail of Antimicrobial activity of 7 alpha-acetoxy-6 beta-hydroxyroyleanone 12-O-benzoyl esters

Research paper thumbnail of Replicon typing of plasmids from Klebsiella pneumoniae and Escherichia coli clinical isolates producing CTX-M-15 extended-spectrum b-lactamase

Clinical Microbiology and Infection, 2009

Research paper thumbnail of IMP-5, metalo-beta-lactamase, em estirpes hospitalares de Pseudomonas aeruginosa

Research paper thumbnail of Class 1 and class 3 integrons among Klebsiella oxytoca in a paediatric hospital

Research paper thumbnail of Spread of In58 containing blavim-2 among Pseudomonas aeruginosa

Clinical Microbiology and Infection, 2006

Research paper thumbnail of Persistência de Klebsiella pneumoniae em doentes de unidades pediátricas do Hospital de Santa Maria, em Lisboa

Portuguese Journal of Pediatrics, 2011

A Klebsiella pneumoniae é responsável por infecções nosocomiais e adquiridas na comunidade. A pro... more A Klebsiella pneumoniae é responsável por infecções nosocomiais e adquiridas na comunidade. A produção de β-lactamases de espectro alargado (ESBLs) e os factores de virulência assumem particular importância nestas infecções. Métodos: Este estudo incluiu 47 isolados de K. pneumoniae, obtidos de dez pacientes de unidades pediátricas de um hospital central, entre 2001 e 2009. Os antibiogramas foram determinados pelo método da difusão em disco. A caracterização genotípica foi efectuada por M13 fingerprinting e, nos perfis representativos, por Multilocus Sequence Typing (MLST). Os genes de resistência bla CTX-M-1 , bla CTX-M-2 e bla SHV , e os genes de virulência k 2 A, rmpA, magA, fimH, mrkD, khe e iucC, foram identificados por reacção em cadeia da polimerase e sequenciados. Resultados: Encontraram-se três antibiotipos predominantes, R1, R2 e R3, de acordo com a susceptibilidade dos isolados à cefotaxima, ceftazidima, ciprofloxacina e gentamicina. Todos os isolados eram sensíveis ao imipenemo. O M13 fingerprinting distinguiu dois perfis genómicos principais, M1 e M2, detectados em vários momentos entre 2001 e 2009. Entre os isolados com perfil M1, 38,5% tinham o antibiotipo R1 e 53,8% tinham o antibiotipo R2; no perfil M2, 12,5% e 56,25% dos isolados apresentavam os antibiotipos R1 e R3, respectivamente. Verificou-se uma associação entre o perfil M1 e o Sequence Type (ST) 15, assim como entre o perfil M2 e os STs 133 e 276. 89,4% dos isolados possuíam genes codificantes para ESBLs; a sequenciação identificou os genes das enzimas CTX-M-15, SHV-1,-2a e-11. A incidência dos genes fimH, mrkD e khe foi elevada; k 2 A, rmpA, magA e iucC não foram detectados. Discussão e conclusão: A presença dos mesmos perfis de M13 fingerprinting durante alguns anos sugere uma endemia nas unidades pediátricas do hospital. Além disso, a elevada incidência dos genes das adesinas fimbriais pode justificar a persistência do mesmo clone no mesmo doente durante longos períodos de tempo.

Research paper thumbnail of Starch-based Pickering emulsions for topical drug delivery: A QbD approach

Colloids and Surfaces B: Biointerfaces, Nov 1, 2015

Pickering emulsions are stabilized by solid particles instead of surfactants and have been widely... more Pickering emulsions are stabilized by solid particles instead of surfactants and have been widely investigated in pharmaceutical and cosmetic fields since they present less adverse effects than the classical emulsions. A quality by design (QbD) approach was applied to the production of w/o emulsions stabilized by starch. A screening design was conducted to identify the critical variables of the formula and the process affecting the critical quality properties of the emulsion (droplet size distribution). The optimization was made by establishing the design space, adjusting the concentration of starch and the quantity of the internal aqueous phase. The emulsion production process was, in turn, adjusted by varying the time and speed of stirring, to ensure quality and minimum variability. The stability was also investigated, demonstrating that an increase in starch concentration improves the stability of the emulsion. Rheological and mechanical studies indicated that the viscosity of the emulsions was enhanced by the addition of starch and, to a higher extent, by the presence of different lipids. The developed formulations was considered non-irritant, by an in vitro assay using human cells from skin (Df and HaCat) with the cell viability higher than 90% and, with self-preserving properties. Finally, the QbD approach successfully built quality in Pickering emulsions, allowing the development of hydrophilic drug-loaded emulsions stabilized by starch with desired organoleptic and structural characteristics. The results obtained suggest that these systems are a promising vehicle to be used in products for topical administration.

Research paper thumbnail of Bioactive Diterpenoids, a New Jatrophane and Two <i>e</i><i>nt</i>-Abietanes, and Other Constituents from <i>Euphorbia </i><i>p</i><i>ubescens</i>

Journal of Natural Products, Apr 23, 2004

A new jatrophane diterpene, pubescenol (1), known ent-abietane lactones, helioscopinolide A (2) a... more A new jatrophane diterpene, pubescenol (1), known ent-abietane lactones, helioscopinolide A (2) and B (3), and taraxerone, 24-methylenecycloartanol, and vanillin have been isolated from Euphorbia pubescens. Diterpenes 1-3 and previously described pubescene D (4) were shown to be moderate inhibitors of the growth of MCF-7, NCI-H460, and SF-268 human tumor cell lines, whereas compounds 2 and 3 also exhibited significant antibacterial activity against Staphylococcus aureus.

Research paper thumbnail of Characterization of Portuguese <i>Thymbra capitata</i> , <i>Thymus caespititius</i> and <i>Myrtus communis</i> essential oils in topical formulations

Flavour and Fragrance Journal, Jun 5, 2017

As the personal care industry evolves, formulators are seeking innovative solutions for their for... more As the personal care industry evolves, formulators are seeking innovative solutions for their formulations' needs, and for ingredients that can offer multiple functions within formulations. Considering that essential oils (EOs) may present a wide spectrum of biological activities, the composition, antimicrobial and antioxidant activity of Portuguese Thymbra capitata, Thymus caespititius and Myrtus communis EOs were assessed in order to evaluate them as preservatives and antioxidants in topical emulsions. The in vivo safety application of some emulsions was also tested. T. capitata EO was mainly constituted by carvacrol (73%), whereas α-terpineol (27%), p-cymene (14%) and carvacrol (10%) dominated Th. caespititius EO, and 1,8-cineole (37%) was dominant in M. communis EO. The minimum inhibitory concentration of T. capitata, Th. caespititius and M. communis EOs was 0.4 μg/mL against C. albicans and ranged between 0.4 and 30.7 μg/mL against A. brasiliensis. Gram-positive bacteria were more susceptible to each EO than Gram-negative bacteria. T. capitata EO showed significantly higher antioxidant activity than Th. caespititius and M. communis EOs. EOs incorporated in emulsions showed preservative activity against all microorganisms tested and T. capitata EO emulsions showed powerful reactive oxygen species (ROS) scavenging effects. A safety evaluation study was performed with 0.10% and 0.01% T. capitata EO emulsions according to the Human Repeat Insult Patch Test (HRIPT). All emulsions were considered safe for topical application. T. capitata, Th. caespititius and M. communis significantly improved the microbiological quality of the prepared emulsions and may constitute a powerful alternative to the current preservatives and antioxidants for use in topical formulations.

Research paper thumbnail of <i>In vitro</i> Antimicrobial Activity of Royleanone Derivatives Against Gram-Positive Bacterial Pathogens

Phytotherapy Research, Mar 11, 2013

Infections caused by multiresistant bacterial pathogens are a significant problem worldwide, turn... more Infections caused by multiresistant bacterial pathogens are a significant problem worldwide, turning the search for natural compounds to act as alternatives to antibiotics of major importance. The aim of the present study was to investigate the in vitro antimicrobial activity of 7α-acetoxy-6β-hydroxyroyleanone (1), isolated from Plectranthus grandidentatus (Lamiaceae), and 11 additional royleanone abietane derivatives of 1 (2-12) against important Gram-positive human bacterial pathogens. Results showed that the aromatic and alkylic esters 2, 3 and 5 are more active than 1 against Enterococcus and Staphylococcus (minimum inhibitory concentration (MIC) values ranging from 0.98 to 62.50 µg/mL). Moreover, 7α-acetoxy-6β-hydroxy-12-O-(4-chloro)benzoylroyleanone (2) gave rise to a new antibacterial-prototype (MIC values of 3.91-15.63 µg/mL against Staphylococcus and of 0.98-3.91 µg/mL against Enterococcus). The results showed that the compounds under analysis also present antimicrobial activity against resistant bacteria. The hydrophobic extra-interactions with bacterial targets seem to play an important role on the activity of royleanones derivatives.

Research paper thumbnail of Neoclerodane and Labdane Diterpenoids from <i>Plectranthus </i><i>o</i><i>rnatus</i>

Journal of Natural Products, Aug 22, 2002

Three new diterpenoids, a neoclerodane and two labdane derivatives, have been isolated from an ac... more Three new diterpenoids, a neoclerodane and two labdane derivatives, have been isolated from an acetone extract of Plectranthus ornatus. The structures of these compounds (plectrornatins A-C, 1-3, respectively) were established mainly by spectroscopic means, particularly by 1D and 2D NMR studies and, in the case of the neoclerodane 1, by an X-ray diffraction analysis. Compounds 1 and 3 showed moderate antimicrobial activity against five Candida species and selected Gram negative and Gram positive bacteria strains.

Research paper thumbnail of Antibiotic activity of five probiotics strains against five multirresistant clinical isolates

Research paper thumbnail of Liposomal Delivery of Newly Identified Prophage Lysins in a Pseudomonas aeruginosa Model

International Journal of Molecular Sciences

Pseudomonas aeruginosa is a Gram-negative opportunistic bacterium that presents resistance to sev... more Pseudomonas aeruginosa is a Gram-negative opportunistic bacterium that presents resistance to several antibiotics, thus, representing a major threat to human and animal health. Phage-derived products, namely lysins, or peptidoglycan-hydrolyzing enzymes, can be an effective weapon against antibiotic-resistant bacteria. Whereas in Gram-positive bacteria, lysis from without is facilitated by the exposed peptidoglycan layer, this is not possible in the outer membrane-protected peptidoglycan of Gram-negative bacteria. Here, we suggest the encapsulation of lysins in liposomes as a delivery system against Gram-negative bacteria, using the model of P. aeruginosa. Bioinformatic analysis allowed for the identification of 38 distinct complete prophages within 66 P. aeruginosa genomes (16 of which newly sequenced) and led to the identification of 19 lysins of diverse sequence and function, 5 of which proceeded to wet lab analysis. The four purifiable lysins showed hydrolytic activity against Gr...

Research paper thumbnail of Genomic epidemiological analysis of Klebsiella pneumoniae from Portuguese hospitals reveals insights into circulating antimicrobial resistance

Scientific Reports

Klebsiella pneumoniae (Kp) bacteria are an increasing threat to public health and represent one o... more Klebsiella pneumoniae (Kp) bacteria are an increasing threat to public health and represent one of the most concerning pathogens involved in life-threatening infections and antimicrobial resistance (AMR). To understand the epidemiology of AMR of Kp in Portugal, we analysed whole genome sequencing, susceptibility testing and other meta data on 509 isolates collected nationwide from 16 hospitals and environmental settings between years 1980 and 2019. Predominant sequence types (STs) included ST15 (n = 161, 32%), ST147 (n = 36, 7%), ST14 (n = 26, 5%) or ST13 (n = 26, 5%), while 31% of isolates belonged to STs with fewer than 10 isolates. AMR testing revealed widespread resistance to aminoglycosides, fluoroquinolones, cephalosporins and carbapenems. The most common carbapenemase gene was blaKPC-3. Whilst the distribution of AMR linked plasmids appears uncorrelated with ST, their frequency has changed over time. Before year 2010, the dominant plasmid group was associated with the extende...

Research paper thumbnail of A phylogenomic approach for the analysis of colistin resistance-associated genes in Klebsiella pneumoniae, its mutational diversity and implications for phenotypic resistance

International Journal of Antimicrobial Agents

The emergence of carbapenemase-producing Klebsiella pneumoniae strains has triggered the use of o... more The emergence of carbapenemase-producing Klebsiella pneumoniae strains has triggered the use of old antibiotics such as colistin which is consequently driving the emergence of colistin resistance in already multidrug resistant strains that can underlie life-threatening infections. This study analyses the mutational diversity of 22 genes associated with colistin resistance in 140 K. pneumoniae clinical isolates integrated in a high-resolution phylogenetic scenario. Colistin susceptibility was accessed by broth microdilution categorizing 98 isolates as susceptible and 16 as resistant, 10 of which were carbapenemase producers. Across the 22 genes examined, 171 non-synonymous mutations and 9 mutations associated with promoter regions were found. Eighty-five isolates had a truncation and/or deletion in at least one of the 22 genes. However, only seven mutations, the complete deletion of mgrB or insertion sequence mediated disruption were exclusively observed in resistant isolates, 4 of these (mgrBIle13fs, pmrBGly207Asp, phoQHis339Asp and ramAIle28Met) comprising novel mutations potentially involved in colistin resistance. The ISEcp1-blaCTX-M-15::mgrB disruption is herein noteworthy as a mechanism underlying co-resistance to third-generation cephalosporins and colistin. Moreover, the high-resolution phylogenetic context shows that most of the mutational diversity spans across multiple phylogenetic clades and that most of the mutations previously associated with colistin resistance are clade associated and present in susceptible isolates, showing no correlation with colistin resistance. In conclusion, the present study contributes with relevant data towards the genetic background of genes involved with colistin resistance deeply rooted across monophyletic groups providing a better understanding of the genes and mutations involved in colistin resistance.

Research paper thumbnail of Article Exploring Dangerous Connections between Klebsiella pneumoniae

Healthcare-associated infections (HAI) are a huge public health concern, particularly when the et... more Healthcare-associated infections (HAI) are a huge public health concern, particularly when the etiological agents are multidrug resistant. The ability of bacteria to develop biofilm is a helpful skill, both to persist within hospital units and to increase antibiotic resistance. Although the links between antibiotic resistance, biofilms assembly and HAI are consensual, little is known about biofilms. Here, electron microscopy was adopted as a tool to investigate biofilm structures associated with increased antibiotic resistance. The K. pneumoniae strains investigated are able to assemble biofilms, albeit with different kinetics. The biofilm structure and the relative area fractions of bacteria and extracellular matrix depend on the particular strain, as well as the minimal inhibitory concentration (MIC) for the antibiotics. Increased values were found for bacteria organized in biofilms when compared to the respective planktonic forms, except for isolates Kp45 and Kp2948, the MIC values for which remained unchanged for fosfomycin. Altogether, these results showed that the emergence of antimicrobial resistance among bacteria responsible for HAI is a multifactorial phenomenon dependent on antibiotics and on bacteria/biofilm features.

Research paper thumbnail of Antimicrobial activity of the diterpene 7 a-acetoxy-6b-hidroxyroyleanone and its 6b-carbamoyl derivative

Research paper thumbnail of Antimicrobial Activity of (11R*, 13 E)-11-Acetoxyhalima-5-13-dien-15-oic Acid and its Methyl Ester Derivative

Research paper thumbnail of Plasmid-mediated CMY-2 AmpC beta lactamase and CTX-M-14 extended-spectrum beta-lactamase in Proteus mirabilis clinical isolate

Clinical Microbiology and Infection, 2008

Research paper thumbnail of Identification of an OXA-58 carbapenemase in an unusual Portuguese Acinetobacter baumannii clinical isolate

Research paper thumbnail of Antimicrobial activity of 7 alpha-acetoxy-6 beta-hydroxyroyleanone 12-O-benzoyl esters

Research paper thumbnail of Replicon typing of plasmids from Klebsiella pneumoniae and Escherichia coli clinical isolates producing CTX-M-15 extended-spectrum b-lactamase

Clinical Microbiology and Infection, 2009

Research paper thumbnail of IMP-5, metalo-beta-lactamase, em estirpes hospitalares de Pseudomonas aeruginosa

Research paper thumbnail of Class 1 and class 3 integrons among Klebsiella oxytoca in a paediatric hospital