Malathion alters the transcription of target genes of the tumour suppressor tp53 and cancerous processes in Colossoma macropomum: Mechanisms of DNA damage response, oxidative stress and apoptosis.

Different classes of pesticides such as fungicides, herbicides, and insecticides, can induce differential expression of genes that are involved in tumorigenesis events in fish, including the expression of tumor suppressor tp53. The degree and duration of the stressful condition is decisive in defining which tp53-dependent pathway will be activated. Herein we evaluate the target genes expression that participates in the regulation pathway of the tumor suppressor tp53 and in the cancerous processes in tambaqui after exposure to malathion. Our hypothesis is that malathion promotes a gene response that is differentially regulated over time, with positive regulation of tp53 target genes related to the apoptotic pathway and a negative regulation of genes that promote antioxidant responses. The fish were exposed to a sublethal concentration of the insecticide for 6 and 48 h. Liver samples were used to analyze the expression of 11 genes using real-time PCR. Overall, the malathion promoted over time increases in tp53 expression and differential expression of tp53 related genes. The exposure resulted in the activation of damage response related genes, caused a positive expression of atm/atr genes. The pro-apoptotic gene bax was up-regulated and the anti-apoptotic bcl2 was down-regulated. Increased expression of mdm2 and sesn1 in the first hours of exposure and no effect on the antioxidant genes sod2 and gpx1 were also observed. We also witnessed an increase in the expression of the hif-1α gene, with no effect on ras proto-oncogene. The extension of this stressful condition accentuated tp53 transcription, and minimized the levels of mdm2, sens1 and bax; however, it down regulated the levels of bcl2 and the bcl2/bax ratio, which indicates the maintenance of the apoptotic response to the detriment of an antioxidant response.

Hypoxia tolerance in two amazon cichlids: mitochondrial respiration and cellular metabolism adjustments are result of species environmental preferences and distribution.

The amazon fishes' responses to hypoxia seem to be related to the Amazon basin diversity of aquatic environments, which present drastic daily and seasonal variations in the dissolved oxygen concentration. Among these fishes' adaptation to hypoxia, behavioral, metabolic, physiological, and biochemical responses are well known for some species. In this work, we aimed to identify how two different aquatic environments, normoxic forest streams and hypoxic lakes, dictate the responses to hypoxia for two cichlid species, Mesonauta festivus and Aequidens pallidus. In our results, we found that A. pallidus is less tolerant to hypoxia, which seems to be related to this animal's natural normoxic environment. Even though this species modulated the mitochondrial respiration in order to improve the oxygen use, it also showed a lower decrease in metabolic rate when exposed to hypoxia and no activation of the anaerobic metabolism. Instead, M. festivus showed a higher decrease in metabolic rate and an activation of the anaerobic metabolism. Our data reveal that the natural dissolved oxygen influences the hypoxia tolerance and the species' tolerance is related to its ability to perform metabolic depression. The interest results are the absence of mitochondrial respiration influences in these processes. The results observed with A. pallidus bring to light also the importance of preserving the forests, in which streams hold very specialized species acclimated to normoxia and lower temperature. The importance of hypoxia tolerance is, thus, important to keep fish assemblage and is thought to be a strong driver of fish biodiversity.

Biomarker responses and PAH ratios in fish inhabiting an estuarine urban waterway.

Many cities worldwide are established adjacent to estuaries and their catchments resulting in estuarine contamination due to intense anthropogenic activities. The aim of this study was to evaluate if fish living in an estuarine urban waterway were affected by contamination, via the measurement of a suite of biomarkers of fish health. Black bream (Acanthopagrus butcheri) were sampled in a small urban embayment and a suite of biomarkers of fish health measured. These were condition factor (CF), liver somatic index (LSI), gonadosomatic index (GSI), hepatic EROD activity, polycyclic aromatic hydrocarbon (PAH) biliary metabolites, serum sorbitol dehydrogenase (s-SDH) and branchial enzymes cytochrome C oxidase (CCO), and lactate dehydrogenase (LDH) activities. The biomarkers of exposure EROD activity, and pyrene- and B(a)P-type biliary metabolites confirmed current or recent exposure of the fish and that fish were metabolizing contaminants. Relative to a reference site, LSI was higher in fish collected in the urban inlet as was the metabolic enzyme LDH activity. CF, GSI, s-SDH, CCO, and naphthalene-type metabolites were at similar levels in the urban inlet relative to the reference site. PAH biliary metabolite ratios of high-molecular-weight to low-molecular-weight suggest that fish from the urban inlet were exposed to pyrogenic PAHs, likely from legacy contamination and road runoff entering the embayment. Similarly, the sediment PAH ratios and the freshness indices suggested legacy contamination of a pyrogenic source, likely originating from the adjacent historic gasworks site and a degree of contamination of petrogenic nature entering the inlet via storm water discharge. Biomarkers of exposure and effect confirmed that black bream collected in the Claisebrook Cove inlet, Western Australia, are currently exposed to contamination and are experiencing metabolic perturbations not observed in fish collected at a nearby reference site.

Large-scale SNP discovery and construction of a high-density genetic map of Colossoma macropomum through genotyping-by-sequencing.

Colossoma macropomum, or tambaqui, is the largest native Characiform species found in the Amazon and Orinoco river basins, yet few resources for genetic studies and the genetic improvement of tambaqui exist. In this study, we identified a large number of single-nucleotide polymorphisms (SNPs) for tambaqui and constructed a high-resolution genetic linkage map from a full-sib family of 124 individuals and their parents using the genotyping by sequencing method. In all, 68,584 SNPs were initially identified using minimum minor allele frequency (MAF) of 5%. Filtering parameters were used to select high-quality markers for linkage analysis. We selected 7,734 SNPs for linkage mapping, resulting in 27 linkage groups with a minimum logarithm of odds (LOD) of 8 and maximum recombination fraction of 0.35. The final genetic map contains 7,192 successfully mapped markers that span a total of 2,811 cM, with an average marker interval of 0.39 cM. Comparative genomic analysis between tambaqui and zebrafish revealed variable levels of genomic conservation across the 27 linkage groups which allowed for functional SNP annotations. The large-scale SNP discovery obtained here, allowed us to build a high-density linkage map in tambaqui, which will be useful to enhance genetic studies that can be applied in breeding programs.

Influence of the natural Rio Negro water on the toxicological effects of a crude oil and its chemical dispersion to the Amazonian fish Colossoma macropomum.

The increment in crude oil exploitation over the last decades has considerably increased the risk of polycyclic aromatic hydrocarbon (PAH) contamination to Amazonian aquatic environments, especially for the black water environments such as the Rio Negro. The present work was designed to evaluate the acute toxicity of the Urucu crude oil (CO), the chemically dispersed Urucu crude oil (CO + D), and the dispersant alone (D) to the Amazonian fish Colossoma macropomum. Acute toxicity tests were performed, using a more realistic approach, where fish were acclimated to both groundwater (GW), used as internal control, and natural Rio Negro water (RNW) and exposed to CO, CO + D and D. Then, biomarkers such as ethoxyresorufin-O-deethylase (EROD), superoxide dismutase (SOD), lipid peroxidation (LPO), serum sorbitol dehydrogenase (s-SDH) in liver, DNA damage in blood cells, and the presence of the benzo[a]pyrene-type, pyrene-type, and naphthalene-type metabolites in fish bile were assessed. Fish exposed to CO and CO + D, at both water types tested, presented increased biomarker responses and higher PAH-type metabolites in the bile. However, fish exposed to these treatments after the acclimation to RNW increased the levels of LPO, s-SDH (hepatotoxicity), DNA damage in blood cells (genotoxicity), and benzo[a]pyrene-type metabolites when compared to fish in GW. Our data suggests that some physicochemical properties of Rio Negro water (i.e., presence of natural organic matter (NOM)) might cause mild chemical stress responses in fish, which can make it more susceptible to oxidative stress following exposure to crude oil, particularly to those chemically dispersed.

Rapid regulation of blood parameters under acute hypoxia in the Amazonian fish Prochilodus nigricans.

Prochilodus nigricans, locally known as curimatã, is an Amazonian commercial fish that endures adverse environmental conditions, in particular low dissolved oxygen, during its migration. Poorer environmental conditions are expected in the near future. Prochilodus nigricans overcomes current seasonal and diurnal changes in dissolved oxygen by adjusting erythrocytic levels of ATP and GTP, modulators of Hb-O2 affinity. Will this fish species be endangered under more extreme environmental conditions as hypoxia and acidification tend to occur in a shorter period of time? As P. nigricans does not exhibit any apparent morphological alterations to exploit the air-water interface, it must rely on fast adjustments of blood properties. To investigate this aspect, basic hematology indices, pHe, pHi, plasma lactate, erythrocytic levels of ATP and GTP and functional properties of the hemolysate of P. nigricans were analyzed over a period of 6h in hypoxia and subsequent recovery in normoxia. The levels of erythrocytic GTP were four times higher than ATP and were reduced to » of the original level after 3h under hypoxia. Erythrocytic levels of ATP were unaffected over the experimental period. All other analyzed blood parameters exhibited a time-course change in animals under hypoxia and returned to normoxic levels. Considering the hemolysate functional properties and the ability to regulate the above mentioned blood characteristics, P. nigricans is able to endure short-term changes in dissolved oxygen.

Copper sensitivity of wild ornamental fish of the Amazon.

The copper sensitivity of 10 wild ornamental fish from Rio Negro, an ion-poor river in the Amazon, was analyzed in the absence of dissolved organic carbon (DOC). No mortality was observed at concentrations up to 700mug Cul(-1) for two species exposed to CuCl(2).2H(2)O dissolved in standard EPA-USA water. Copper sensitivity of fish was also tested in INPA groundwater, which is similar to that of the Rio Negro, but lacking typical DOC of Rio Negro water. In these experiments, all four characid fish species were more sensitive to copper than the catfish and the cichlids. LC(50) values ranged from 12.8mug Cul(-1) for the characid Hemigrammus rhodostomus to 74.1mug Cul(-1) for the callichtyid Dianema urostriatum. Copper sensitivity of Hyphessobrycon socolofi was 52-fold lower in EPA (1405.5mug Cul(-1)) than in INPA water (26.8mugCul(-1)). The differences in values of LC(50) may be related to species-specific physiological abilities to ion-regulate under the original conditions of the habitat of the analyzed species.