Genetic and biochemical characterization of BIM-1, a novel acquired subgroup B1 MBL found in a Pseudomonas sp. strain from the Brazilian Amazon region.

OBJECTIVES: To characterize a novel acquired MBL, BIM-1, in a Pseudomonas #2 (subgroup P. guariconensis) strain isolated from the Aurá river located in the Brazilian Amazon hydrographic basin. METHODS: WGS using an Illumina® MiSeq System was used to characterize the genome of Pseudomonas sp. IEC33019 strain. Southern blotting/hybridization assays were performed to confirm the location of the MBL-encoding gene, blaBIM-1 (Belém Imipenemase). Antimicrobial susceptibility testing, cloning, and biochemical and phenotypic characterization were performed to determine BIM-1 kinetics. RESULTS: The IEC33019 strain showed high resistance rates to ß-lactams, ciprofloxacin and aminoglycosides, being susceptible only to polymyxins and susceptible, increased exposure to aztreonam. WGS analysis revealed a novel acquired MBL-encoding gene, blaBIM-1, found as a gene cassette inserted into a class 1 integron (In1326) that also carried qnrVC1 and aadA11e. In1326 was located in a complex transposon, Tn7122, carried by a 52.7 kb conjugative plasmid (pIEC33019) with a toxin/antitoxin system (vapB/vapC). BIM-1 belongs to the molecular subgroup B1 and shares 70.2% and 64.9% similarity with SIM-1 and IMP-1, respectively. Kinetics analysis of BIM-1 showed hydrolytic activity against all ß-lactams tested. CONCLUSIONS: BIM-1 is a novel acquired MBL encoded by a gene carried by mobile genetic elements, which can be transferred to other Gram-negative bacilli (GNB). Because the IEC33019 strain was recovered from a river impacted by a populous metropolitan region with poor basic sanitation and served by limited potable freshwater, it would be important to establish the role of the BIM-1-producing GNB as nosocomial pathogens and/or as colonizers of the riverside population in this geographical region.

Biodiversity and interannual variation of cyanobacteria density in an estuary of the brazilian Amazon.

The influence of environmental variables on planktonic biodiversity is widely known. However, the absence of information about the cyanobacterial community in tropical estuarine regions motivated this work, whose objective was to investigate the spatio-temporal variation of cyanobacterial density related to physicochemical factors in a Brazilian Amazonian estuary. For the qualitative and quantitative study of cyanobacteria and physicochemical variables, samples were collected in April/July/2009 and April/August/2010. We identified 31 species of the orders Chroococcales, Oscillatoriales and Nostocales. Species of the genera Aphanocapsa, Dolichospermum, Komvophoron, Microcystis, Pseudanabaena and Merismopedia were frequent and abundant throughout the study period. Some of the found genera have already been described as potential toxin producers. The dynamics of cyanobacteria were temporal, the highest densities occurred in 2010 (average= 1080.86 ± 702.86 cells.mL-1) mainly influenced by the high values of pH, temperature, electrical conductivity, total dissolved solids, ammonium nitrogen which led cyanobacteria to present different responses in terms of richness, density and diversity between the years.

Metabolic and oxidative impairments in human salivary gland cells line exposed to MeHg.

BACKGROUND/AIM: The ingestion of contaminated seafood by MeHg is considered the main route of human exposure, turning the salivary gland one important target organ. The salivary glands play critical roles in maintaining oral health homeostasis, producing saliva that maintains the oral microbiota, initiation of the digestion of macromolecules, and being essential in maintaining the integrity of the adjacent soft tissues and teeth. Thus, this study aimed to investigate the effects of MeHg exposure on human salivary gland cells line. METHODS: Cells were exposed to 1-6 µM of MeHg for 24 h, and analysis of toxicity was performed. Based on these results, the LC50 was calculated and two concentrations were chosen (0.25 and 2.5 µM MeHg) to evaluate intracellular mercury (Hg) accumulation (THg), metabolic viability and oxidative stress parameters (GSH:GSSG ratio, lipid peroxidation, protein oxidation and DNA damage). RESULTS: The results demonstrated accumulation of THg as we increased the MeHg concentrations in the exposure and, the higher the dose, the lower is the cell metabolic response. In addition, the 2.5 µM MeHg concentration also triggered oxidative stress in human salivary gland cells by depleting the antioxidant competence of GSH:GSSG ratio and increasing lipid peroxidation and proteins carbonyl levels, but no damages to DNA integrity. CONCLUSION: In conclusion, although these two elected doses did not show lethal effects, the highest dose triggered oxidative stress and new questionings about long-term exposure models are raised to investigate furthers cellular damages to human salivary gland cells caused by MeHg exposure to extrapolate in a translational perspective.

Oxidative Damage in Human Periodontal Ligament Fibroblast (hPLF) after Methylmercury Exposure.

Human exposure to mercury (Hg) is primary associated with its organic form, methylmercury (MeHg), through the ingestion of contaminated seafood. However, Hg contamination is also positively correlated with the number of dental restorations, total surface of amalgam, and organic mercury concentration in the saliva. Among the cells existing in the oral cavity, human periodontal ligament fibroblast (hPLF) cells are important cells responsible for the production of matrix and extracellular collagen, besides sustentation, renewal, repair, and tissue regeneration. In this way, the present study is aimed at investigating the potential oxidative effects caused by MeHg on hPLF. Firstly, we analyzed the cytotoxic effects of MeHg (general metabolism status, cell viability, and mercury accumulation) followed by the parameters related to oxidative stress (total antioxidant capacity, GSH levels, and DNA damage). Our results demonstrated that MeHg toxicity increased in accordance with the rise of MeHg concentration in the exposure solutions (1-7 µM) causing 100% of cell death at 7 µM MeHg exposure. The general metabolism status was firstly affected by 2 µM MeHg exposure (43.8 ± 1.7%), while a significant decrease of cell viability has arisen significantly only at 3 µM MeHg exposure (68.7 ± 1.4%). The ratio among these two analyses (named fold change) demonstrated viable hPLF with compromised cellular machinery along with the range of MeHg exposure. Subsequently, two distinct MeHg concentrations (0.3 and 3 µM) were chosen based on LC50 value (4.2 µM). hPLF exposed to these two MeHg concentrations showed an intracellular Hg accumulation as a linear-type saturation curve indicating that metal accumulated diffusively in the cells, typical for metal organic forms such as methyl. The levels of total GSH decreased 50% at exposure to 3 µM MeHg when compared to control. Finally, no alteration in the DNA integrity was observed at 0.3 µM MeHg exposure, but 3 µM MeHg caused significant damage. In conclusion, it was observed that MeHg exposure affected the general metabolism status of hPLF with no necessary decrease on the cell death. Additionally, although the oxidative imbalance in the hPLF was confirmed only at 3 µM MeHg through the increase of total GSH level and DNA damage, the lower concentration of MeHg used (0.3 µM) requires attention since the intracellular mercury accumulation may be toxic at chronic exposures.

Assessment of DDT and mercury levels in fish and sediments in the Iriri River, Brazil: Distribution and ecological risk.

In order to assess the risk of exposure of human populations to dichlorodiphenyltrichloroethane (DDT) and mercury, muscles of five fish species were analysed, along with the surface sediment of 14 Iriri River sampling sites. The fish specimens were sacrificed by the spinal section, prior to sex identification, body weight determination and total length. Considering the fish specimens studied, 11% of them showed concentrations of mercury higher than the maximum established by the World Health Organization for safe human consumption. A positive correlation between fish body weight and mercury concentration was observed, besides a positive correlation between the fish size and Hg concentration. Significant differences (P < 0.05) were found between mean concentrations of DDT and metabolites among species of fish studied. In the Plagioscion squamossissimus species, the highest concentration of total DDT (151.4 ng/g) was found, while in Eugerres Brasilianus species, the lowest. However, the DDT levels in fish muscle of studied species are below the maximum set by FAO-Alimentarius CODEX. In the sediments, total DDT ranged from 11.58 ng/g to 48.4 ng/g, which is associated with the historical DDT use in the Amazon. According to sediment quality guidelines, these levels have a moderate toxic effect in almost all of the studied region.