Short and long-term effects of nanobiomaterials in fish cell lines. Applicability of RTgill-W1.

Nanobiomaterials (NBMs) are nanostructured materials for biomedical applications that can reach aquatic organisms. The short and long-term effects of these emerging contaminants are unknown in fish. The RTgill-W1 cell line has been proposed as a model to predict the acute toxicity of chemicals to fish (OECD Test Guideline nº 249). We assessed the applicability of this cell line to study the short and long-term toxicity of 15 NBMs based on hydroxyapatites (HA), lipid (LSNP/LNP), gold, iron oxide, carbon, poly l-Lactide acid (PLLA) fibers with Ag and poly (lactide-co-glycolide) acid. Two more rainbow trout cell lines (RTL-W1, from liver, and RTS-11, from spleen) were exposed, to identify possible sensitivity differences among cells. Exposures to a range of concentrations (0.78-100 µg/mL) lasted for 24 h. Additionally, the RTgill-W1 was used to perform long-term (28 d exposure) and recovery (14 d exposure/14 d recovery) assays. Cells were exposed to the 24 h-IC20 and/or to 100 µg/mL. A triple cytotoxicity assay was conducted. After 24 h, only PLLA Fibers-Ag showed cytotoxicity (IC50 < 100 µg/mL). However, the NBMs in general provoked concentration-dependent effects after long-term exposures, except the LSNPs. A recovery of viability was only observed for AuNPs, AuNRods, Fe3O4PEG-PLGA, MgHA-Collag_Scaffolds, Ti-HA and TiHA-Alg NPs.These results evidenced the need to test the long-term toxicity of NBMs and showed differences in cytotoxicity probably associated to different mechanisms of toxic action. The RTgill-W1 was useful to screen short and long-term toxicities of NBMs and appears as a promiseful model to assess possible toxicity of NBMs in fish.

Fish cell lines as screening tools to predict acute toxicity to fish of biocidal active substances and their relevant environmental metabolites.

Biocidal substances and their environmental relevant metabolites are highly toxic for fish. However, an important scarcity of toxicity data for metabolites is recognised. This article provides new data about the toxicity to fish of these compounds and evaluates the potential use of fish cell lines as screening tools to assess the acute toxicity of these compounds in fish. To this aim, acute toxicity of 7 substances was tested in Oncorhynchus mykiss (OECD TG203) and cytotoxicity of 16 substances was assessed in fish cell lines from two species; Poeciliopsis lucida (PLHC-1) and O. mykiss (RTH-149, RTG-2 and RTgill-W1) performing three cytotoxicity tests: Alamar-Blue, 5-carboxyfluorescein diacetate, acetoxymethyl ester and Neutral Red Uptake. Additionally, in vitro and in vivo data from the LIFE-COMBASE database were included in a dataset finally comprising 33 biocides and 14 metabolites. Hazard data were categorized into 4 toxicity groups, according to the intervals established in Regulation (EC) 1272/2008. Finally, the Spearman correlation test was performed and coincidences between in vitro-in vivo data established. In vitro and in vivo results revealed a high positive correlation, with a complete coincidence for 56.5% of the substances, a 2% of false positives (non-toxic in vivo) and a 13% of false negatives (toxic in vivo) for the 4 toxicity categories. However, when results were grouped in toxic or non-toxic coincidence was obtained for 85% of the substances. In conclusion, although fish denote a greater sensitivity, the use of at least two fish cell lines and three cytotoxicity endpoints appear to be valid approaches for fish acute toxicity screening of biocides and their metabolites.

Accuracy of tuberculosis diagnostic tests in small ruminants: A systematic review and meta-analysis.

Tuberculosis (TB) in small ruminants is a neglected disease despite its major impact on goat and sheep production and the global public health. The awareness of the role of small ruminants in the epidemiology of animal TB has increased in the last two decades, however, there is a lack of standardization of procedures and robust quantitative estimates on the accuracy of diagnostic TB tests in the scientific literature. To address this knowledge gap, all the available information regarding the use of ante-mortem diagnostic techniques in small ruminants was collected and summarized through a systematic review process. Furthermore, a random-effects meta-analysis was conducted to separately estimate the sensitivity (Se) and specificity (Sp) of cell-based tests among the retrieved studies in goats. Studies included in the meta-analysis were also evaluated using the Quality Assessment of Diagnostic Accuracy Studies included in systematic reviews adapted for animal diagnostic tests (VETQUADAS). Median pooled Se estimates of the single intradermal tuberculin (SIT) test (ranged from 0.51 to 0.59), the comparative intradermal tuberculin (CIT) test (ranged from 0.30 to 0.50) and the interferon-gamma (IFN-γ) release assay (IGRA) (ranged from 0.66 to 0.72) were lower than that reported previously in cattle, regardless the interpretation criteria and the reporting of MAP infection or vaccination. However, the specificity was adequate for all the tests (ranged from 0.95 to 0.99), except for the SIT test in MAP vaccinated herds (ranged from 0.78 to 0.90). This study provides an overview of the accuracy of diagnostic tests for TB in goats, however, the considerable between-study heterogeneity found hampered the conclusive interpretation of the pooled Se and Sp estimates. Therefore, further studies in small ruminants are necessary to optimize the diagnostic Se, which could help to design effective control strategies, accelerate the eradication of TB in these species and harmonize test procedures.

Cytotoxicity against fish and mammalian cell lines and endocrine activity of the mycotoxins beauvericin, deoxynivalenol and ochratoxin-A.

The toxicity of mycotoxins is well recognized in mammals, but their effects on fish have received less attention. Moreover, in the last years several studies have reported that some mycotoxins may act as endocrine disruptors. The aim of this study was to determine the cytotoxic effects and endocrine activities of three mycotoxins: beauvericin, deoxynivalenol and ochratoxin-A. Cytotoxicity in two fish hepatoma and one mammalian hepatoma cell lines was determined by the AlamarBlue, Neutral Red Uptake and CFDA-AM assays. For the assessment of androgenic, estrogenic and thyroidal agonistic/antagonistic effects three cell lines stably expressing luciferase as reporter gene under the control of hormone receptors were used. Results showed that both fish and mammalian cell lines were very sensitive to the mycotoxins tested. OTA was the least toxic mycotoxin and DON and BEA showed similar acute toxicity. None of the three mycotoxins tested presented agonistic effects at the receptors studied, but all of them showed strong antagonistic effect at the thyroid receptor. BEA showed a weak antagonistic effect at the androgen receptor and OTA produced a biphasic dose-response curve at the estrogen receptor. The data obtained in this work are of high interest for aquaculture industries and for regulators.

Evaluation of the McLintock syringe as a cause of non-specific reactions in the intradermal tuberculin test used for the diagnosis of bovine tuberculosis.

The objective of this study was to elucidate whether the use of the McLintock syringe, used to inject tuberculin in cattle in several countries and based on an intradermal inoculation by needle, may, in itself, cause skin reactions that can be interpreted as positive reactions regardless of the real tuberculosis (TB) infection status of the animals. Forty-four cattle from an officially TB-free (OTF) herd were selected for the experiment. Each animal received four inoculations [one with sterile phosphate buffer saline (PBS) with 10% of glycerol and three with bovine purified protein derivative (PPD), as performed during the single intradermal tuberculin (SIT) test], two on each side of the neck (n = 176 inoculations). Three different McLintock syringes (n = 132 inoculations, PBS and bovine PPD) and one Dermojet syringe (n = 44 inoculations, PBS) were used to carry out the inoculations. No positive reactions (increase in skin-fold thickness > 3 mm) in response to the bovine PPD or PBS inoculations were observed regardless of the syringe used. No significant differences (p > 0.05) in the skin fold thickness increase (in mm) were observed between inoculation sites. Significant differences (p < 0.05) in the skin fold thickness were observed when PPD was injected in comparison to the PBS but no differences between McLintock and Dermojet were detected when PBS was injected. The McLintock syringe did not cause reactions per se that could be misunderstood as positive in TB-free cattle demonstrating that it is not a significant factor associated with the previously reported imperfect specificity of the SIT test.

Evaluation of the use of a needle-free injection syringe as a cause of non-specific reactions in the intradermal tuberculin test used for the diagnosis of bovine tuberculosis.

The objective of the study was to elucidate whether the use of the needle-free Dermojet syringe, which is based on a high pressure inoculation and is used to inject tuberculin in cattle in several countries, may, in itself, cause skin reactions that can be interpreted as positive reactions to the intradermal tests that are not, in fact, related to the real infection status of the animals. Forty-four cattle from an officially tuberculosis-free (OTF) herd were selected, and four single intradermal tuberculin (SIT) tests were performed on each animal, two on each side of the neck. Three different Dermojet (D1, D2 and D3) and one McLintock (M4) syringes were used to carry out sterile phosphate buffer saline (PBS) with 10% of glycerol and bovine PPD injections. No positive reactions to the SIT test were observed when using the D1-D3 syringes in the case of either bovine PPD or PBS. With regard to M4 (PBS), all the tests were negative when using a standard interpretation but three were positive in the case of the severe interpretation. Significant differences (p < 0.05) in the skin fold thickness measured were found only between certain Dermojet and McLintock syringes at certain inoculation sites. The results showed that the needle-free Dermojet syringe used for PPD intradermal testing in cattle did not cause significant reactions that could be misunderstood as positives.

Development of a new tool for the long term in vitro ecotoxicity testing of nanomaterials using a rainbow-trout cell line (RTL-W1).

The current wide use of manufactured nanomaterials (MNs) is leading to the release of nanoparticles (NPs) to water bodies. Aquatic organisms, including fish, are exposed to low concentrations of NPs for long periods of time being necessary to develop laboratory toxicity tests reflecting realistic conditions. Additionally, today there is a demand of in vitro assays respecting the 3Rs principle. Thus, the main aim of this work was to stablish an in vitro tool for the assessment of long-term NPs ecotoxicity. Considering the key role of liver in detoxification, a rainbow trout liver cell line, RTL-W1, was used. CuO NPs were chosen to validate this tool taking into account their important production level. Cells were exposed for 21 days to 25 or 100 µg CuO NPs/ml. Every seven days cells were split and one fourth of them transferred to a new plate with appropriate concentrations of NPs in culture medium. Lower concentrations of CuO NPs did not cause any deleterious effect, whereas higher concentrations led to significant mortality after 14 days and to the intracellular accumulation of Cu particles. Identical results were observed in cells exposed to CuSO4 at the same Cu concentrations. Therefore, the observed toxic effects might be mainly due to Cu2+ ions.

Fish cell lines as a tool for the ecotoxicity assessment and ranking of engineered nanomaterials.

Risk assessment of engineered nanomaterials (ENMs) is being hindered by the sheer production volume of these materials. In this regard, the grouping and ranking of ENMs appears as a promising strategy. Here we sought to evaluate the usefulness of in vitro systems based on fish cell lines for ranking a set of ENMs on the basis of their cytotoxicity. We used the topminnow (Poeciliopsis lucida) liver cell line (PLHC-1) and the rainbow trout (Oncorhynchus mykiss) fibroblast-like gonadal cell line (RTG-2). ENMs were obtained from the EU Joint Research Centre repository. The size frequency distribution of ENM suspensions in cell culture media was characterized. Cytotoxicity was evaluated after 24 h of exposure. PLHC-1 cells exhibited higher sensitivity to the ENMs than RTG-2 cells. ZnO-NM was found to exert toxicity mainly by altering lysosome function and metabolic activity, while multi-walled carbon nanotubes (MWCNTs) caused plasma membrane disruption at high concentrations. The hazard ranking for toxicity (ZnO-NM > MWCNT ≥ CeO2-NM = SiO2-NM) was inversely related to the ranking in size detected in culture medium. Our findings reveal the suitability of fish cell lines for establishing hazard rankings of ENMs in the framework of integrated approaches to testing and assessment.

Vaccination as a control strategy against Salmonella infection in pigs: A systematic review and meta-analysis of the literature.

Consumption or handling of improperly processed or cooked pork is considered one of the top sources for foodborne salmonellosis, a common cause of intestinal disease worldwide. Asymptomatic carrier pigs may contaminate pork at slaughtering; therefore, pre-harvest reduction of Salmonella load can contribute to reduce public health risk. Multiple studies have evaluated the impact of vaccination on controlling Salmonella in swine farms, but results are highly variable due to the heterogeneity in vaccines and vaccination protocols. Here, we report the results of an inclusive systematic review and a meta-analysis of the peer-reviewed scientific literature to provide updated knowledge on the potential effectiveness of Salmonella vaccination. A total of 126 articles describing the use of Salmonella vaccines in swine were identified, of which 44 fulfilled the inclusion criteria. Most of the studies (36/44) used live vaccines, and S. Typhimurium and S. Choleraesuis were the predominant serotypes evaluated. Vaccine efficacy was most often measured through bacteriological isolation, and pooled estimates of vaccine efficacy were obtained as the difference in the percentage of positive animals when available. Attenuated and inactivated vaccines had similar efficacy [Risk Difference=-26.8% (-33.8, -19.71) and -29.5% (-44.4, -14.5), respectively]. No serotype effect was observed on the efficacy recorded for attenuated vaccines; however, a higher efficacy of inactivated vaccines against S. Choleraesuis was observed, though in a reduced sample. Results from the meta-analysis here demonstrate the impact that vaccination may have on the control of Salmonella in swine farms and could help in the design of programs to minimize the risk of transmission of certain serotypes through the food chain.

Effects of a silver nanomaterial on cellular organelles and time course of oxidative stress in a fish cell line (PLHC-1).

Among the nanomaterials currently in commercial products, those based on silver are the most used, and so there is a high probability that silver nanoparticles (AgNPs) will be released into aquatic environments where they could adversely affect aquatic organisms, including fish. Taking this into account, the aim of the present work was to characterize in depth the mechanisms underlying the toxic action of AgNPs using fish cell lines, determining specifically the contribution of alterations in cellular structures and oxidative stress time course to the cytotoxicity of AgNPs. Since liver plays a key role in detoxification, the hepatoma cell line PLHC-1 was used. Exposure to AgNPs (NM-300K, obtained from the Joint Research Centre Repository) caused alterations at the lysosomal and mitochondrial levels at lower concentrations than those that disrupted plasma membrane (evaluated by means of neutral red, alamarBlue, and 5-carboxyfluorescein diacetate, acetoxymethyl ester assays respectively). AgNO3, used as a control Ag+ ion source, produced similar cytotoxic effects but at lower concentrations than AgNPs. Both silver forms caused oxidative disruption but the initial response was delayed in AgNPs until 6h of exposure. Transmission electron microscopy analysis also evidenced the disruption of mitochondrial structures in cells exposed to cytotoxic concentrations of both forms of silver. At non-cytotoxic concentrations, AgNPs were detected inside the nucleoli and mitochondria, thereby pointing to long-term effects. The present work evidences the mutual interaction between the induction of oxidative stress and the alterations of cellular structures, particularly mitochondria, as cytotoxicity mechanisms not exclusively associated to NPs.

Molecular characterization and antimicrobial resistance of STEC strains isolated from healthy cattle in 2011 and 2013 in Spain.

Prevention of Shiga toxin-producing Escherichia coli (STEC) foodborne outbreaks is hampered by its complex epidemiology. We assessed the distribution of virulence genes (VGs), main serogroups/serotypes for public health [haemolytic uraemic syndrome (HUS)-related], antimicrobial resistance (AMR) profiles and pulsed-field gel electrophoresis (PFGE) patterns in a collection of STEC isolates obtained from cattle hide (n = 149) and faecal (n = 406) samples collected during a national survey conducted in Spain in 2011 and 2013. Isolates were cultured using McConkey and CT-SMAC agar after enrichment, and confirmed as STEC by PCR. STEC prevalence in hides (15·4%) was higher than in faeces (10·7%) and O157:H7 was more frequent in the former (2·7% vs. 0·99%). Non-O157 HUS-related serogroups were present albeit at low frequencies. The non-O157 isolates were more heterogeneous than O157:H7 in their VG patterns, with 25/64 presenting VGs from both STEC and enterotoxigenic pathotypes (hybrid isolates). Of the STEC isolates, 62·5% were resistant at least to one antimicrobial, and no differences in AMR between O157:H7 and non-O157 were detected. All isolates had different profiles by PFGE and did not form a cluster. Overall, our results demonstrated that STEC in the cattle reservoir is still a matter of concern for human health due to the presence of HUS-related serogroups, the occurrence of certain VGs, AMR and the additional risks that hybrid isolates may pose, and thus warrants further investigation.

Major allergen from Amaranthus palmeri pollen is a profilin: Isolation, partial characterisation and IgE recognition.

BACKGROUND: Pollens represent a rich source of proteins that are also potential elicitors of IgE-mediated pollen allergy. Sensitisation to panallergens could play an important role in diagnosis and specific immunotherapy, because these molecules are present in different plant pollens and plant foods and have marked structural similarity in different species. Profilins are one of the most common panallergens to be studied because they are responsible for a large number of sensitisations and are clearly related to cross-reactivity and co-sensitisation. This study aimed to isolate and characterise a new allergen of Amaranthus palmeri pollen and to determine its allergenicity. METHODS: A. palmeri pollen profilin was purified using poly-l-proline-Sepharose affinity chromatography followed by anion exchanger chromatography. Identification of purified protein was carried out by mass spectrometry. Specific IgE was estimated in sera of patients with positive skin prick test to A. palmeri pollen extract, by enzyme-linked immunosorbent assay (ELISA). PRINCIPAL FINDINGS: Purified protein appeared as a single band at 14 kDa in SDS-PAGE gel. Mass spectrometric analysis of the gel band identified two highly conserved peptides corresponding to allergenic profilins from pollen of other plants. Sera from about 60% of allergic patients have IgE that recognises the purified A. palmeri protein. CONCLUSION: A 14 kDa protein of A. palmeri pollen was purified and identified as allergenic profilin, which was recognised by sera from pollen allergic patients.

Cytotoxicity of the mycotoxins deoxynivalenol and ochratoxin A on Caco-2 cell line in presence of resveratrol.

Exposure to mycotoxins through dietary food intake involves a highly complex scenario where co-contamination of different mycotoxins has been frequently demonstrated. On the other hand, the effect of the interaction of mycotoxins with other generally considered beneficial food components, as the antioxidants, has been scarcely studied. The main goal of the present work was to assess the cytotoxic effects on Caco-2 cells of the mycotoxins deoxynivalenol (DON) and ochratoxin A (OTA), alone or combined, and to explore potential protective effects of resveratrol (RES), an antioxidant frequently found in wine. In parallel, reactive oxygen species (ROS) production has also been studied as a first approach to understand the underlying mechanism of cytotoxicity. Results indicate a higher toxic effect of the mycotoxins when they are co-exposed. This increase in cytotoxicity was not accompanied by an increase in ROS production. The co-exposure of OTA or DON with RES did not result in a decrease in cytotoxicity; on the contrary, it resulted in increased cytotoxicity not associated with an increase in ROS production.

In vitro assessment of thyroidal and estrogenic activities in poultry and broiler manure.

Among the many chemicals found in avian manure, endocrine disruptors (EDs), of natural or anthropogenic origin, are of special environmental concern. Nowadays, an increasing amount of estrogens is being released into the environment via the use of manure to fertilize agricultural land. While most research in this field has focused on estrogenic phenomena, little is known about alterations related to other endocrine systems, such as the thyroidal one. Here we simultaneously assessed the potential estrogenic and thyroidal activity of poultry and broiler litter manure using in vitro approaches based on estrogen receptor (Er) and thyroid receptor (Tr) transactivation assays. In addition, leaching experiments were performed to assess whether the EDs present in the manure pass through a soil column and potentially reach the groundwater. Manure from four broiler and four poultry farms was collected in two sampling campaigns carried out in two seasons (fall and spring). Extracts from broiler and poultry manure exhibited strong thyroidal activity. Only poultry manure showed estrogenic activity, which is consistent with the low levels of estrogens expected in hatchlings. Leakage experiments were performed in columns with two kinds of arable soils: sandy and loamy. No estrogenicity or thyroidal activity was detectable in soils treated with the manure or in the corresponding leachates. These results indicate that substances with estrogenic or thyroidal activity were degraded in the soil under our experimental conditions. However, the long-term effects associated with the constant and intensive application of manure to agricultural land in some regions require further research.

Risk assessment of coccidostatics during feed cross-contamination: animal and human health aspects.

Coccidiosis, an intestinal plasmodium infection, is a major infectious disease in poultry and rabbits. Eleven different coccidiostats are licensed in the EU for the prevention of coccidiosis in these animal species. According to their chemical nature and main biological activity, these compounds can be grouped as ionophoric (monensin, lasalocid sodium, salinomycin, narasin, maduramicin and semduramicin) or non-ionophoric (robenidine, decoquinate, nicarbazin, diclazuril, and halofuginone) substances. Coccidiostats are used as feed additives, mixed upon request into the compounded feed. During the technical process of commercial feed production, cross-contamination of feed batches can result in the exposure of non-target animals and induce adverse health effects in these animals due to a specific sensitivity of mammalian species as compared to poultry. Residue formation in edible tissues of non-target species may result in unexpected human exposure through the consumption of animal products. This review presents recent risk assessments performed by the Scientific Panel on Contaminants in the Food Chain (CONTAM) of the European Food Safety Authority (EFSA). The health risk to non-target species that would result from the consumption of cross-contaminated feed with coccidostats at levels of 2, 5 or 10% was found to be negligible for most animal species with the exception of salinomycin and monensin in horses because of the particular sensitivity for which toxicity may occur when cross-contamination exceeds 2% and 5% respectively. Kinetic data and tissue analyses showed that residues of coccidiostats may occur in the liver and eggs in some cases. However, the level of residues of each coccidiostat in edible animal tissues remained sufficiently low that the aggregate exposure of consumers would not exceed the established acceptable daily intake (ADI) of each coccidiostat. It could be concluded that technical cross-contamination of animal feeds would not be expected to adversely affect the health of consumers.

In vitro dose-response effects of poly(amidoamine) dendrimers [amino-terminated and surface-modified with N-(2-hydroxydodecyl) groups] and quantitative determination by a liquid chromatography-hybrid quadrupole/time-of-flight mass spectrometry based method.

This article presents a dose-response study of the effects of two types of third-generation (G3) and fourth-generation poly(amidoamine) (PAMAM) dendrimers on two cell lines (RTG-2 and H4IIE) by in vitro cytotoxicity assays with 3-(4,5-dimethylthizol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), neutral red uptake (NRU), and lactate dehydrogenase (LDH) assays. We particularly investigated the potential cytotoxic effect of positive surface charge, which a cationic amino-terminated PAMAM dendrimer can display, on the marked ability of PAMAM dendrimers to cross the cell membrane compared with PAMAM dendrimers functionalized with chains of N-(2-hydroxydodecyl). Quantification of dose-response effects was performed by use of mass spectrometry analysis. The analytical method using liquid chromatography-hybrid quadrupole/time-of-flight mass spectrometry that we developed allowed characterization of defective dendrimers instead of "ideal structures." Identification was based on accurate mass measurement, assignment of elemental composition, and the fully resolved (13)C/(12)C isotopic clusters of the multiply charged ions of PAMAM dendrimers. Validation of the liquid chromatography-mass spectrometry method made possible reliable and reproducible quantification of the extracellular and intracellular concentration of dendrimers at a micromolar level (limits of detection from 0.14 to 1.34 µM and from 0.43 to 1.82 µM in standard and culture medium, respectively). A higher cytotoxicity was found with the H4IIE cell line for surface-modified PAMAM dendrimers. The LDH assay was significantly more sensitive than the MTT and NRU assays, with half-maximal inhibitory concentrations (IC(50)) of 12.96 and 38.31 µg mL(-1) for surface-modified G3 and G4 dendrimers, respectively. No cytotoxic effects, in terms of IC(50), of amino-terminated PAMAM dendrimers were observed on both H4IIE and RTG-2 cells when the concentration was below 500 µg mL(-1) for G3 and G4 dendrimers.

Differences in the induction of cyp1A and related genes in cultured rainbow trout Oncorhynchus mykiss. Additional considerations for the use of EROD activity as a biomarker.

Two rainbow trout Oncorhynchus mykiss fish farms were repeatedly sampled in order to observe the variability of ethoxyresorufin-O-deethylase (EROD) activity and of related genes in the liver. Fish coming from fish farm A exhibited EROD levels that could be considered as basal according to the scientific literature, however, EROD activity in fish coming from fish farm B was significantly increased. This was accompanied by augmented aryl hydrocarbon receptor (ahr) and cytochrome P4501A (cyp1A) messenger RNA expression and reduced oestrogen receptor (er) and vitellogenin (vtg) transcription. Only sediment extracts from the entry channel of fish farm B induced EROD activity in O. mykiss cultured cells, however, this induction could not be explained by the levels of polyaromatic hydrocarbons (PAH) and polychlorinated biphenyls (PCB) measured in the sediments. The results of this study point out that O. mykiss cultured in fish farms could be used as sentinels for indication of pollution. In this particular work, however, no conclusive evidence has been found for a relationship between the presence of PAHs and PCBs and the observed EROD induction.

Effects of cerium oxide nanoparticles to fish and mammalian cell lines: An assessment of cytotoxicity and methodology.

Two cerium oxide nanoparticles (CeO(2) NPs) and one micro-sized CeO(2) particle were thoroughly characterized in their pristine form, in water and in cell culture medium. The particles were tested for cytotoxicity to the H4IIE rat hepatoma cell line or the RTG-2 rainbow trout gonadal cell line by means of four standard cytotoxicity assays. Nominal concentrations were verified by inductively coupled plasma mass spectrometry (ICP-MS) and methods were assessed for their suitability to detect reliably adverse effects due to particle exposure. All three particles showed aggregation in water and media. In the H4IIE cell line, the MTT cytotoxicity test revealed that negative effects could be observed for the CeO(2) NPs after 24h and for all particles after 72h of exposure, making the effects size, concentration and time dependent. No negative effect for the concentrations tested was detected for the remaining three assays and the RTG-2 cell line, making the MTT assay and the H4IIE cell line an appropriate system to assess adverse effects of CeO(2) NPs. A verification of the nominal concentration through ICP-MS revealed that there was a discrepancy between nominal and measured concentration depending on concentration and particle tested. Interferences of particles with assays were found to be present and need to be taken into consideration.

Factors associated with viral load suppression in HIV-infected pregnant women in Rio de Janeiro, Brazil.

Viral load (VL) near delivery is a determinant of mother-to-child transmission (MTCT) of HIV. To evaluate factors associated with an undetectable VL near delivery in HIV-infected pregnant women receiving highly active antiretroviral therapy (HAART) and non-HAART regimens, HIV-infected pregnant women with a detectable VL at entry and having used antiretrovirals for ≥4 weeks before delivery were selected. Multivariate analysis was employed using binary logistic unconditional models; the dependent variable was having a VL <400 copies/mL near delivery. VL suppression was achieved in 403/707 women (57%): 65.4% in the HAART group, but only 26% in the non-HAART group P = 0.001. Duration of HAART was correlated with VL suppression, with maximum benefit seen after ≥12 weeks of therapy (odds ratio [OR]: 2.51; 95% confidence interval [CI]: 1.72-3.65). CD4+ cell count near delivery (OR: 1.53; 95% CI: 1.06-2.20) and baseline VL (OR: 0.74; 95% CI: 0.58-0.94) were also independently associated with VL suppression. Overall MTCT rate was 1.6%. HAART for ≥12 weeks, baseline VL and CD4 cell count near delivery were independently associated with viral suppression near delivery.

Cardiorespiratory evaluation of juvenile rats experimentally envenomed with Tityus serrulatus venom

Accidental envenomation caused by Tityus serrulatus scorpions is very common in Brazil and may result in serious cardiorespiratory alterations that are frequently fatal to children. In the present study, the effects of T. serrulatus venom on the cardiorespiratory system of recently weaned male Wistar rats were evaluated. Fifteen animals were distributed into three groups (n = 5). The control group A received 400 miuL ultrapure water by subcutaneous injection, while the experimental groups B and C were injected with scorpion venom (100 and 450 miug, respectively, in 400 miuL water). Electrocardiogram (ECG) traces were obtained prior to the experiment, at five-minute intervals up to 30 minutes after treatment. At 40 minutes after envenomation, the animals had severe acute symptoms and were subsequently anesthetized for blood collection by means of intracardiac puncture. Biochemical profiles for the cardiac muscle were established by colorimetric analysis of creatine kinase (CK) and CK-MB isoenzyme. Semiquantitative analysis of troponin was performed using the immunochromatographic assay. Following euthanasia, the lungs and hearts were removed and subjected to histopathological examination. All experimental animals had ECG alterations compatible with electrolytic imbalance, myocarditis and alterations of the cardiac conduction system. Envenomed animals had accentuated bradycardia at 25 and 30 minutes after venom inoculation. All experimental animals had myocardial lesions, which were confirmed by increased serum levels of CK and CK-MB, although there were no alterations in the serum concentration of troponin. Pulmonary hemorrhage was detected in whole lungs and microscopically confirmed by the presence of congested capillaries and erythrocytes in the alveolar parenchyma. In conclusion, T. serrulatus venom caused great cardiorespiratory damage to weaned rats.