Browsing by Author "Andrade, Joana"
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- Avaliação da resposta de bactérias heterotróficas presentes no meio aquático à presença de microcistinasPublication . Andrade, Joana; Valério, ElisabeteO objetivo deste estudo foi avaliar a razão pela qual bactérias heterotróficas aquáticas não são muito afectadas pela presença de microcistinas.
- Evaluation of the physiologic response of heterotrophic bacteria present in aquatic environments to the presence of microcystinsPublication . Andrade, Joana; Valério, Elisabete; Reis, Ana Maria GonçalvesMicrocystins (MC) are the hepatotoxins produced by cyanobacteria, which are photosynthetic organisms that usually colonize aquatic environments. Climate change has had a significant impact on the increased frequency of cyanobacterial blooms, which are often associated with the production of cyanotoxins. Studies show that the growth of aquatic heterotrophic bacteria that co-occur with cyanobacteria may not be affected by the presence of MC, or, on the other hand they may present a reduction in growth, never being totally inhibited by their presence (Miguéns, 2013; Pinto, 2016). The aim of this study was to examine the effects of three microcystin variants (MCLR, MCRR, MCYR) on heterotrophic aquatic bacteria living in the same ecosystem as cyanobacteria. In particular, (1) to observe the impact of microcystins on the growth of heterotrophic bacteria, (2) the impact on the enzymes of the antioxidant system of these bacteria and (3) to screen the presence of mlr genes. For this purpose, we performed the morphological and molecular characterization of 22 bacteria isolated from two reservoirs. It was intended to analyze the growth of bacterial isolates exposed to different concentrations of extract of each variant of MCs, and also to two concentrations of pure microcystin-LR. The search for the presence of MCs degradation genes (mlrA-D), was done using PCR. In order to analyze the effect of microcystins on the antioxidant system of the isolates, catalase (CAT) and superoxide dismutase (SOD1 and SOD2) activities were determined spectrophotometrically, at 240 nm and 550 nm, respectively, in the cells exposed to the different variants of microcystin. None of the isolates grown in liquid medium was completely inhibited by the exposure to microcystins, however, it was found that MCs can reduce the growth of most bacteria tested. While some bacteria grew without any induced effect, others reacted differently according to the variant and the concentration (in the same isolate). In some cases a slight growth stimulation was observed. In most cases the growth stimulation appears to be related with other molecules from the MC extracts. The results of the determination of CAT and SOD activities revealed that only three isolates have catalase, namely M17F, M17K and 594196, and only three isolates have superoxide dismutase, M17C, M17D and 594196. In this study, a heterotrophic bacteria, M17C, mlr+, was also isolated, mlr genes were amplified and sequenced. A new primer pair for the mlrA was also proposed in this study. In summary, the results from this study, showed that there is not a pattern characteristic of the species or genera analyzed, thus indicating that the response of the heterotrophic bacteria isolates to MCs is mainly related to strain characteristics. They corroborate previous studies, which indicate that the cluster of mlr genes is not the only cause to allow bacteria to grow in the presence of MC without being affected by them, even with no alteration in the main enzymes from the antioxidant system (CAT and SOD).
- Increased antibacterial properties of indoline-derived phenolic Mannich basesPublication . Rimpiläinen, Tatu; Nunes, Alexandra; Calado, Rita; Fernandes, Ana S.; Andrade, Joana; Ntungwe, Epole; Spengler, Gabriella; Szemerédi, Nikoletta; Rodrigues, João; Gomes, João Paulo; Rijo, Patricia; Candeias, Nuno R.The search for antibacterial agents for the combat of nosocomial infections is a timely problem, as antibiotic-resistant bacteria continue to thrive. The effect of indoline substituents on the antibacterial properties of aminoalkylphenols was studied, leading to the development of a library of compounds with minimum inhibitory concentrations (MICs) as low as 1.18 μM. Two novel aminoalkylphenols were identified as particularly promising, after MIC and minimum bactericidal concentrations (MBC) determination against a panel of reference strain Gram-positive bacteria, and further confirmed against 40 clinical isolates (Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, and Listeria monocytogenes). The same two aminoalkylphenols displayed low toxicity against two in vivo models (Artemia salina brine shrimp and Saccharomyces cerevisiae). The in vitro cytotoxicity evaluation (on human keratinocytes and human embryonic lung fibroblast cell lines) of the same compounds was also carried out. They demonstrated a particularly toxic effect on the fibroblast cell lines, with IC50 in the 1.7-5.1 μM range, thus narrowing their clinical use. The desired increase in the antibacterial properties of the aminoalkylphenols, particularly indoline-derived phenolic Mannich bases, was reached by introducing an additional nitro group in the indolinyl substituent or by the replacement of a methyl by a bioisosteric trifluoromethyl substituent in the benzyl group introduced through use of boronic acids in the Petasis borono-Mannich reaction. Notably, the introduction of an additional nitro moiety did not confer added toxicity to the aminoalkylphenols.