Heterologously expressed bacterial and human multidrug resistance proteins confer cadmium resistance to Escherichia coli

The human MDR1 gene is induced by cadmium exposure although no resistance to this metal is observed in human cells overexpressing hMDR1. To access the role of MDR proteins in cadmium resistance, human MDR1, Lactococcus lactis lmrA, and Oenococcus oeni omrA were expressed in an Escherichia coli tolC mutant strain which proved to be hypersensitive to cadmium. Both the human and bacterial MDR genes conferred cadmium resistance to E. coli up to 0.4 mM concentration. Protection was abolished by 100 mu M verapamil. Quantification of intracellular cadmium concentration by atomic absorption spectrometry showed a reduced cadmium accumulation in cells expressing the MDR genes. Inside-out membrane vesicles of L. lactis overexpressing lmrA displayed an ATP-dependent Cd-109(2+) uptake that was stimulated by glutathione. An evolutionary model is discussed in which MDR proteins have evolved independently from an ancestor protein displaying both organic xenobiotic- and divalent metal-extrusion abilities

Genotoxic and stress inductive potential of cadmium in Xenopus laevis larvae

The present investigation evaluates the toxic potential of Cd in larvae of the frog Xenopus laevis after 12 days of exposure to environmentally relevant contamination levels, close to those measured in the river Lot (France). Several genotoxic and detoxification mechanisms were analyzed in the larvae: clastogenic and/or aneugenic effects in the circulating blood by micronucleus (MN) induction, metallothionein (MT) production in whole larvae, gene analyses and Cd content in the liver and also in the whole larvae. The results show: (i) micronucleus induction at environmental levels of Cd contamination (2, 10, 30 μg L−1); (ii) an increased and concentration-dependent quantity of MT in the whole organism after contamination with 10 and 30 μg Cd L−1 (a three- and six-fold increase, respectively) although no significant difference was observed after contamination with 2 μg Cd L−1; (iii) Cd uptake by the whole organism and by the liver as a response to Cd exposure conditions; (4) up-regulation of the genes involved in detoxification processes and response to oxidative stress, while genes involved in DNA repair and apoptosis were repressed. The results confirm the relevance of the amphibian model and highlight the complementarity between a marker of genotoxicity, MT production, bioaccumulation and genetic analysis in the evaluation of the ecotoxicological impact

Elektromagnetsko polje na frekvenciji mobilnih telefona (900 MHz) izaziva stres i modifikacije DNA u gujavici Eisenia fetida

Eisenia fetida earthworms were exposed to electromagnetic field (EMF) at a mobile phone frequency (900 MHz) and at field levels ranging from 10 to 120 V m-1 for a period of two hours (corresponding to specific absorption rates ranging from 0.13 to 9.33 mW kg-1). Potential effects of longer exposure (four hours), field modulation, and a recovery period of 24 h after two hours of exposure were addressed at the field level of 23 V m-1. All exposure treatments induced significant DNA modifications as assessed by a quantitative random amplified polymorphic DNA-PCR. Even after 24 h of recovery following a two hour-exposure, the number of probe hybridisation sites displayed a significant two-fold decrease as compared to untreated control earthworms, implying a loss of hybridisation sites and a persistent genotoxic effect of EMF. Expression of genes involved in the response to general stress (HSP70 encoding the 70 kDa heat shock protein, and MEKK1 involved in signal transduction), oxidative stress (CAT, encoding catalase), and chemical and immune defence (LYS, encoding lysenin, and MYD, encoding a myeloid differentiation factor) were up-regulated after exposure to 10 and modulated 23 V m-1 field levels. Western blots showing an increased quantity of HSP70 and MTCO1 proteins confirmed this stress response. HSP70 and LYS genes were up-regulated after 24 h of recovery following a two hour-exposure, meaning that the effect of EMF exposure lasted for hours.U ovom istraživanju gujavice vrste Eisenia fetida bile su izložene elektromagnetskom polju (EMP) na frekvenciji mobilnih telefona (900 MHz) te poljima jačine 10 do 120 V m-1 u dvosatnom razdoblju (što odgovara specifičnim ratama apsorpcije od 0,13 do 9,33 mW kg-1). Utjecaj dužeg izlaganja (4 sata), modulacije polja te vrijeme oporavka od 24 sata nakon dva sata izlaganja proučavan je pri jačini polja od 23 V m-1. Metoda kvantitativne nasumično umnožene polimorfne DNA (engl. quantitative random amplified polymorphic DNA – qRAPD) otkrila je značajne modifikacije DNA na svim proučavanim tretmanima. Čak i nakon 24-satnog oporavka broj hibridizacijskih mjesta bio je dvostruko manji u odnosu na broj zabilježen u kontrolnim gujavicama, što upozorava na gubitak hibridizacijskih mjesta i na dugoročan utjecaj EMP-a. Ekspresija gena uključenih u odgovor na stres (HSP70: kodira za 70kDa heat shock protein i MEKK1: uključen u provođenje signala), oksidacijski stres (CAT: kodira za katalazu) te kemijsku i imunosnu obranu (LYS: kodira za lysenin i MYD: kodira za faktor mijeloidne diferencijacije) bila je povišena nakon izlaganja polju jačine 10 V m-1 te moduliranome polju jačine 23 V m-1. Western blot analiza potvrdila je odgovor na stres detekcijom povišene količine HSP70 i MTCO1 proteina. HSP70 i LYS geni imali su povišenu ekspresiju i nakon razdoblja oporavka, što upućuje na dugotrajan utjecaj EMP-a

Characterization of the arcD Arginine:Ornithine Exchanger of Pseudomonas aeruginosa. Localization in the Cytoplasmic Membrane and a Topological Model

The arcDABC operon of Pseudomonas aeruginosa encodes the enzymes of the arginine deiminase pathway and is induced by oxygen limitation. The arcD gene specifies a 53-kDa protein with arginine: ornithine exchange activity. The ArcD protein of P. aeruginosa, like the LysI lysine transporter of Corynebacterium glutamicum, has 13 hydrophobic regions which could span the cytoplasmic membrane. Fusion of a Caa (colicin A) epitope to the N-terminal part of ArcD permitted the localization, by immunoblotting, of the hybrid protein in the inner membrane of P. aeruginosa. Fusion of PhoA (alkaline phosphatase) to the very C terminus of ArcD produced another hybrid protein, which exhibited PhoA activity. Both ArcD hybrid proteins retained arginine transport activity and served to support a topological model which proposes that the N terminus is oriented toward the cytoplasm and the C terminus faces the periplasm. Further ArcD-PhoA fusions were consistent with this model. When the Caa epitope was fused to a C-terminal ArcD fragment consisting of only 5 hydrophobic domains, the resulting hybrid protein could be recovered intact from the inner membrane, suggesting that the C-terminal part of ArcD contains sufficient information for insertion into the membrane. This study illustrates the utility of the Caa epitope to tag membrane proteins

The Chemokine CCL2 Protects Against Methylmercury Neurotoxicity

Industrial pollution due to heavy metals such as mercury is a major concern for the environment and public health. Mercury, in particular methylmercury (MeHg), primarily affects brain development and neuronal activity, resulting in neurotoxic effects. Because chemokines can modulate brain functions and are involved in neuroinflammatory and neurodegenerative diseases, we tested the possibility that the neurotoxic effect of MeHg may interfere with the chemokine CCL2. We have used an original protocol in young mice using a MeHg-contaminated fish-based diet for 3 months relevant to human MeHg contamination. We observed that MeHg induced in the mice cortex a decrease in CCL2 concentrations, neuronal cell death, and microglial activation. Knock-out (KO) CCL2 mice fed with a vegetal control food already presented a decrease in cortical neuronal cell density in comparison with wild-type animals under similar diet conditions, suggesting that the presence of CCL2 is required for normal neuronal survival. Moreover, KO CCL2 mice showed a pronounced neuronal cell death in response to MeHg. Using in vitro experiments on pure rat cortical neurons in culture, we observed by blockade of the CCL2/CCR2 neurotransmission an increased neuronal cell death in response to MeHg neurotoxicity. Furthermore, we showed that sod genes are upregulated in brain of wild-type mice fed with MeHg in contrast to KO CCL2 mice and that CCL2 can blunt in vitro the decrease in glutathione levels induced by MeHg. These original findings demonstrate that CCL2 may act as a neuroprotective alarm system in brain deficits due to MeHg intoxicatio

Nature-based tourism elicits a phenotypic shift in the coping abilities of fish

Nature-based tourism is gaining extensive popularity, increasing the intensity and frequency of human-wildlife contacts. As a consequence, behavioral and physiological alterations were observed in most exposed animals. However, while the majority of these studies investigated the effects of punctual exposure to tourists, the consequences of constant exposition to humans in the wild remains overlooked. This is an important gap considering the exponential interest for recreational outdoor activities. To infer long-term effects of intensive tourism, we capitalized on Odontostilbe pequira, a short-lived sedentary Tetra fish who spends its life close to humans, on which it feeds on dead skin. Hence, those fish are constantly exposed to tourists throughout their lifecycle. Here we provide an integrated picture of the whole phenomenon by investigating, for the first time, the expression of genes involved in stress response and neurogenesis, as well as behavioral and hormonal responses of animals consistently exposed to tourists. Gene expression of the mineralocorticoid (and cortisol) receptor (mr) and the neurogenic differentiation factor (NeuroD) were significantly higher in fish sampled in the touristic zone compared to those sampled in the control zone. Additionally, after a simulated stress in artificial and controlled conditions, those fish previously exposed to visitors produced more cortisol and presented increased behavioral signs of stress compared to their non-exposed conspecifics. Overall, nature-based tourism appeared to shift selection pressures, favoring a sensitive phenotype that does not thrive under natural conditions. The ecological implications of this change in coping style remain, nevertheless, an open question

Navike u konzumaciji ribe u trudnica iz Itaitube, grada na slijevu rijeke Tapajós u Brazilu – rizici od kontaminacije živom procijenjeni mjerenjem ukupne žive u piscivornim ribljim vrstama koje se često konzumiraju i u kosi trudnica

The Tapajós River basin in the Amazon region, Brazil is one of the most active gold mining areas in the world. In this study, we evaluated fish consumption habits and mercury exposure in 110 pregnant women in the city of Itaituba by measuring their total hair mercury concentrations. In addition, we investigated seasonal differences in mercury concentrations in two highly consumed piscivorous fish species, tucunaré (Cichla spp.) and pescada (Plagioscion squamosissimus). Total fish mercury concentrations (THg) during the dry season were 0.62±0.07 mg/kg for Cichla spp. and 0.73±0.08 mg/kg for P. squamosissimus. During the rainy season they were 0.39±0.04 and 0.84±0.08 mg/kg, respectively. Of our participants 44 % declared that they ate Cichla spp. and 67 % P. squamosissimus. Mean mercury concentration in their hair was 1.6±0.2 mg/kg and was above the US Environmental Protection Agency (US EPA) reference dose of 1 mg/kg in 48 % of them. Mean fish THg concentrations were also above the joint Food and Drug Administration and US EPA safety limit of 0.5 mg/kg for P. squamosissimus during both seasons and for Cichla spp. during the dry season only. These results show that pregnant women should avoid consumption of these piscivorous fish species during pregnancy and call for a regular programme to monitor Hg levels in that area.Slijev rijeke Tapajós u amazonskom području u Brazilu jedno je od najaktivnijih područja iskopavanja zlata na svijetu. U ovoj smo studiji procijenili navike konzumiranja ribe i razinu izloženosti metil-živi u 110 stanovnica Itaitube u peripartumu mjerenjem ukupne koncentracije žive u kosi (THg). Osim toga, istražili smo sezonske razlike u koncentracijama žive u dvjema najčešćim vrstama konzumnih piscivornih riba: tucunaré (Cichla spp.) i pescada (Plagioscion squamosissimus). U tih je ribljih vrsta THg tijekom sušne sezone iznosio 0,62±0,07 mg/kg za Cichla spp. te 0,73±0,08 mg/kg za P. squamosissimus. Tijekom kišne sezone koncentracija THg bila je 0,39±0,04 mg/kg za Cichla spp. I 0,84±0,08 mg/kg za P. squamosissimus. Sudionice su izjavile da su konzumirale Cichla spp. (44 %) i P. squamosissimus (67 %). Srednja koncentracija THg izmjerena u kosi bila je 1,6±0,2 mg/kg±std. Prosječna koncentracija THg u kosi bila je viša od referentne doze Američke agencije za zaštitu okoliša (krat. US EPA) koja iznosi 1 mg/kg, a čak u 48 % sudionica izmjerena je viša koncentracija. Također, srednja vrijednost koncentracija THg u ribi bila je iznad sigurnosne granice (0,5 mg/kg) za P. squamosissimus i tijekom sušne i kišne sezone, a za Cichla spp. samo tijekom sušne sezone. Rezultati istraživanja pokazuju da bi trudnice trebale izbjegavati konzumaciju ovih piscivornih ribljih vrsta. Stoga je razvidna potreba za redovitim programom praćenja razine žive na tom području

Feeding mice with diets containing mercury-contaminated fish flesh from French Guiana: a model for the mercurial intoxication of the Wayana Amerindians

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