Mercury concentration in larvae of Eustrongylides sp (Nematoda: Dioctophymatoidea) from fish of the Brazilian Amazon / Concentración de mercurio en larvas de Eustrongylides sp (Nematoda: Dioctophymatoidea) en peces de la Amazonía brasileña

Abstract Introduction: Chemical pollution represents a great concern to aquatic organisms, especially fish. Metals enter the aquatic environment from a variety of sources, including natural biogeochemical cycles and anthropogenic sources such as industrial and residential effluents, mining and atmospheric sources. Objective: To describe the Eustrongylides sp. larvae and the interaction with their fish hosts as indicators of mercury (Hg) contamination in the Brazilian Amazon, and the distribution of Hg in the internal organs of fish species Hoplias malabaricus and Pygocentrus nattereri collected in oxbow lakes on the Tapajós River, in the municipality of Santarém, in the state of Pará. Methods: Total Hg was analyzed using the Direct Hg Analyzer - DMA-80. Concentrations of Hg in Eustrongylides sp. were compared with those found in the tissues/organs of the hosts H. malabaricus and P. nattereri. Hg concentrations in the host/parasite system were statistically compared using Principal Component Analysis. The bioconcentration factor (BCF) was calculated to assess the bioaccumulation capacity of metals in Eustrongylides sp. larvae, comparing the concentration of Hg in the parasite with that accumulated in the musculature of infected hosts. Results: Hg concentrations in all tissues/organs analyzed were higher in the parasitic species Eustrongylides sp. larvae when compared with those found in tissues/organs of H. malabaricus and P. nattereri. There was an inversely proportional relationship, showing that when Eustrongylides sp. larvae are present, the concentration in the parasite is higher than in the musculature of host fish H. malabaricus and P. nattereri. The BCF of Hg was found by comparing Eustrongylides sp. larvae/H. malabaricus muscle and was observed during a flood (BCF Hg = 15 364). Conclusions: The results confirm the greater bioaccumulative capacity of Eustrongylides sp. compared to its host. The data indicated the viability of using Eustrongylides sp. larvae in biomonitoring programs. It is worth mentioning that fish samples for Hg analysis must be free of parasites since their presence can alter the results.

Resumen Introducción: La contaminación química del hábitat acuático representa un gran peligro para organismos acuáticos, especialmente para peces. Los metales ingresan al ambiente acuático desde una variedad de fuentes, incluidos los ciclos biogeoquímicos naturales y fuentes antropogénicas, como efluentes industriales y residenciales, minería y fuentes atmosféricas. Objetivo: Describir las especies de Eustrongylides sp. y la interacción con sus peces hospederos como indicadores de contaminación por mercurio en la Amazonía brasileña, y la distribución en los órganos internos de las especies de peces Hoplias malabaricus y Pygocentrus nattereri recolectadas en cochas del Río Tapajós, en el municipio de Santarém, del estado de Pará. Métodos: El Hg total se analizó utilizando el Direct Hg Analyzer - DMA-80. Las concentraciones de Eustrongylides sp. se compararon con las encontrados en los tejidos/órganos de los hospederos H. malabaricus y P. nattereri. Las concentraciones en el sistema hospedero/parásito se compararon estadísticamente utilizando el análisis de componentes principales. Se calculó el factor de bioconcentración (BCF) para evaluar la capacidad de bioacumulación de metales en larvas de Eustrongylides sp., comparando la concentración en el parásito con la acumulada en la musculatura de los hospederos infectados. Resultados: Las concentraciones de Hg en todos los tejidos/órganos analizados fueron mayores en las larvas de la especie parasitaria Eustrongylides sp. en comparación con las encontradas en los tejidos/órganos de H. malabaricus y P. nattereri. Hubo una relación inversamente proporcional, mostrando que cuando las larvas de Eustrongylides sp. están presentes, la concentración en el parásito es mayor que en la musculatura de los peces hospederos H. malabaricus y P. nattereri. El BCF de Hg se encontró comparando Eustrongylides sp. larvas/ músculo H. malabaricus y se observó durante una inundación (BCF Hg = 15 364). Conclusiones: Los resultados confirman la mayor capacidad bioacumulativa de Eustrongylides sp. en comparación con su hospedero. Los datos indicaron la viabilidad de utilizar larvas de Eustrongylides sp. en programas de biomonitoreo. Cabe mencionar que las muestras de pescado para análisis de Hg deben estar libres de parásitos ya que su presencia puede alterar los resultados.

Evaluation of the anti-snakebite, antimicrobial and antioxidant potential of Philodendron megalophyllum Schott (Araceae), traditionally used in accidents caused by snakes in the western region of Pará, Brazil.

ETHNOPHARMACOLOGICAL RELEVANCE: In the region of Western Pará, Amazonia, Brazil, Philodendron megalophyllum is widely used for the treatment of envenomations caused by bites from venomous snakes. The traditional use of plants is usually done through oral administration of an infusion (decoction) soon after the bite occurs. The efficiency of aqueous extracts of P. megalophyllum was demonstrated for blocking the activity of the venom of Bothrops sp., but only for a pre-incubation protocol (venom:extract), which fails to simulate the real form of use of this species. In this context, the objective of this research was to evaluate the anti-snakebite potential of the aqueous extract of P. megalophyllum to inhibit for the biological activity induced by Bothrops atrox venom (BaV) using traditional treatment methods. MATERIAL AND METHODS: Initially, an aqueous extract using the stem of P. megalophyllum (AEPm) was prepared following the standard procedure used by the residents of the rural area along the Tapajós River (Eixo Forte region) in Santarém, PA, Brazil. The phytochemical profile of AEPm was conducted using thin layer chromatography (TLC) and phenolic compounds were quantified through colorimetric trials. The cytotoxicity of AEPm was evaluated using the MRC-5 human fibroblast line, and the antioxidant potential was measured using DPPH methods and cell culture. AEPm antimicrobial action was evaluated by the 96-well plate microdilution and the minimum inhibitory concentration (MIC) methods using 18 types of microorganisms including bacteria that are present in the oral cavity of snakes. AEPm blocking potential was tested against BaV activity in vitro (fibrinolytic) and in vivo (defibrinating and hemorrhagic). In order to test for an interaction between BaV and AEPm SDS-PAGE electrophoresis was conducted. RESULTS: The presence of coumarins, fatty acids, and hydrolysable tannins were detected in the AEPm. The colorimetric trials showed that AEPm had a high concentration of condensed tannins (20.1 ± 1.2%). The potential of AEPm for blocking of hemorrhagic and fibrinolytic activity of BaV showed a maximum reduction of 86.1% and 96.5%, respectively, for the pre-incubation protocol (1:10, venom:extract). However, when the extract was administered orally there was no significant blocking of these activities. The interaction of BaV and AEPm showed a modification of the profile of proteic bands when compared to the pattern of bands obtained from the BaV alone. The AEPm was not considered toxic, demonstrated antioxidant activity, and was capable of reducing the growth of 10 of the 18 studied microorganisms. CONCLUSION: Although the stem of P. megalophyllum is indicated by traditional medicine techniques as effective against snakebites, the extract, when tested orally was not able to significantly inhibit (p ˃ 0.05) hemorrhage and defibrinating activity induced by the B. atrox venom. On the other hand, the extract yielded a promising result with respect to antioxidant and antimicrobial potential, and after further studies it could be used as a complementary treatment for localized action and secondary infections that frequently occur with snakebites from the genus of Bothrops sp.

A comparison of the ability of Bellucia dichotoma Cogn. (Melastomataceae) extract to inhibit the local effects of Bothrops atrox venom when pre-incubated and when used according to traditional methods.

Bellucia dichotoma Cogn. (Melastomataceae) is one of various plant species used in folk medicine in the west of the state of Pará, Brazil, to treat snake bites. Many studies have been carried out to evaluate the effectiveness of anti-snake bite plants, but few of these use the same preparation methods and doses as those traditionally used by the local populations. This study therefore compared inhibition of the main local effects of B. atrox venom (BaV) by aqueous extract of B. dichotoma (AEBd) administered according to traditional methods and pre-incubated with BaV). The concentrations of phenolic compounds (tannins and flavonoids) in AEBd were determined by colorimetric assays. The effectiveness of AEBd in inhibiting the hemorrhagic and edematogenic activities of BaV was evaluated in mice in four different experimental in vivo protocols: (1) pre-incubation (venom:extract, w/w); (2) pre-treatment (p.o.); (3) post-treatment (p.o.); and (4) AEBd (p.o.) in combination with Bothrops antivenom (BA) (i.v.). To assess in vitro inhibition of BaV phospholipase A2 activity, the pre-incubation method or incorporation of AEBd or BA in agarose gels were used. The effect of AEBd on BaV was determined by SDS-PAGE, zymography and Western blot. Colorimetric assays revealed higher concentrations of (condensed and hydrolyzable) tannins than flavonoids in AEBd. Hemorrhagic activity was completely inhibited using the pre-incubation protocol. However, with pre-treatment there was no significant inhibition for the concentrations tested, and with the post-treatment only the 725 mg/kg dose of AEBd was able to inhibit 40.5% (p = 0.001) of the hemorrhagic activity of BaV. Phospholipase A2 activity was only inhibited when AEBd was pre-incubated with BaV. BaV-induced edema was completely inhibited with pre-incubation (p < 0.05) and significantly reduced (p < 0.05) with pre- and post-treatment (p.o.) for the concentrations tested. The reduction in local edema was even greater when AEBd was administered in combination with BA. The SDS-PAGE profiles showed that several of the BaV protein (SDS-PAGE) and enzyme (zymography) bands were not detected when the venom was pre-incubated, and Western blot revealed that this was not caused by the AEBd enzymes observed in the zymogram. The "pseudo inhibition" observed after pre-incubation in this study may be due to the presence of tannins in the extract, which could act as chelating agents, removing metalloproteins and Ca²âº ions and thus inhibiting hemorrhagin and PLA2 activity. However, when administered according to traditional methods, B. dichotoma extract was effective in blocking BaV-induced edematogenic activity and had an additional effect on inhibition of this activity by BA.