Departamento de Doenças Infecciosas
Permanent URI for this community
Browse
Browsing Departamento de Doenças Infecciosas by advisor "Caeiro, Maria Filomena"
Now showing 1 - 2 of 2
Results Per Page
Sort Options
- Activities of an aqueous plant extract against herpes simplex virus type2 (HSV 2): role in virus replication and progeny yieldPublication . Mota Mendes, Ana Rita; Caeiro, Maria Filomena; Lopo, SílviaHerpes simplex virus type 2 (HSV-2) is widely distributed through the human population, infecting more than 500 million people globally. Although causing generally mild infections this virus may cause severe symptoms occasionally, mainly in immunocompromised patients. Presently, there are a number of systemic antiviral agents against herpesvirus, the most commonly used being acyclovir (ACV) and its prodrugs. However, long term treatments with these drugs may result in the development of resistance, especially in immunocompromised patients, which leads to a continuous search for new and better therapeutic alternatives. According to the World Health Organization (WHO) plants would be the best sources for obtaining a wide variety of drugs. In fact, in the last decades many pharmacological and chemical studies have focused on medicinal plants and the discovery of new natural therapeutic compounds. In this study the anti-herpetic action of an aqueous extract was evaluated. The product was obtained by decoction of stem and leaves from Solidago virgaurea, a perennial herb member of the Asteraceae family. The study of the aqueous extract activity included a preliminary evaluation of its cytotoxicity in Vero cells – by the MTT assay - in the same conditions that are applied for viral production. Extract direct effect on viral particles – virucidal effect – was also assessed, having proved null. Anti-herpetic activity was investigated through two kinds of experiments: treatment of infected cells during virus production that revealed a mean yield reduction of 94 % in treated relatively to non-treated cells and an IC50 of 35.1 μg/mL; and treatment of infected cells during virus titration, which revealed slighter inhibition but significant size differences between virus plaques formed in treated and control conditions (smaller in treatment conditions). To further evaluate the mechanisms that mediate the aqueous extract inhibitory effect, infected cells – treated and non-treated - and virus particles – produced in treated and non-treated cells - were visualized through Transmission Electron Microscopy (TEM), revealing less damage due to infection in treated cells and a reduced amount of viral particles in HSV-2 suspensions produced in treated cells, relatively to controls. A kinetic of the first hours of the infection was performed with and without treatment, to assess possible differences in DNA production. Extracted samples were subjected to qPCR and results showed that the amount of viral DNA raises significantly slower in treated versus non-treated infected cells, throughout the infection. This is consistent with the effective reduction of the extract when added at later infection times - 4-6 h p.i. - when DNA replication is already in an advanced stage. Our results suggest that the aqueous extract inhibits HSV-2 replication, when present at the beginning of the infection, possibly by interfering with the viral DNA synthesis.
- Dynamics of β-lactamases in Gram-negative bacteriaPublication . Manageiro, Vera; Caniça, Manuela; Caeiro, Maria Filomenaβ-Lactamase production is the most important resistance mechanism among Gram-negative bacteria. The overall aim of this PhD thesis was to contribute to the knowledge of molecular epidemiology of β-lactamases and to the understanding of their diversity in a structural-functional level. To accomplish this aim, several studies with different approaches were performed. The emergence of β-lactamase-producing isolates, as well as the appearance of new epidemic clones, is of great concern. The studies presented in the first chapter of results, have clearly shown that specific extended-spectrum β-lactamase (ESBL)-, plasmid-mediated AmpC β-lactamase (PMAβ)- and carbapenem-hydrolyzing class D β-lactamase (CHDL)-producing clones are able to persist in clinical settings for long periods, resulting in a complex β-lactamase endemic situation. A high diversity of β-lactamases was encountered, specifically: CTX-M family which is the most prevalent ESBL, and PMAβ (e.g., DHA-1, CMY-2, CMY-39, MIR-1, MIR-3, FOX-5 and the novel CMY-46 and CMY-50), both in Enterobacteriaceae, as well as CHDLs OXA-23 and OXA-24/40 in Acinetobacter baumannii. The results obtained in this thesis also highlight different strategies for bacterial spread of resistance that can occur through either clonal spread or horizontal gene transfer of mobile genetic elements. In the second chapter of results, structure/function correlation of five novel clinical important β-lactamases, namely three inhibitor-resistant SHV (SHV-72, SHV-84 and SHV-107), one ESBL (SHV-55) and one parental SHV (SHV-99), are presented. One of the key findings we can infer from results is that the conserved motif Lys234-Thr/Ser235-Gly236, present in class A β-lactamases, is a hot-spot for β-lactamase inhibition, meaning that new compounds can be designed to address this structural feature. In summary, the work performed in this thesis allows the elucidation on the dynamics of β-lactamases in Gram-negative bacteria, in Portugal. Molecular characterization together with biochemical data is essential for understanding the emergence of new resistance mechanisms and their spread.