Acute and subchronic exposure of diplopods to substrate containing sewage mud: Tissular responses of the midgut

Abstract

Several discussions regarding the agricultural use of sewage mud have occurred; however, its use has been questioned due to the risks of soil contamination mainly by metals. Diplopods are saprophagous invertebrates, agile in colonizing several layers of soil. Due to the habit of this group, several researchers have proposed their use in ecotoxicological analyses. This study aimed to expose these invertebrates to substrate containing sewage mud of a STS (Sewage Treatment Station) from São Paulo State, Brazil and analyzed its toxic potential by morphological analysis of the midgut of the diplopod Rhinocricus padbergi, region of the digestive tube where digestion and nutrient absorption occur. The animals were exposed in mud concentrations at 1%, 10% and 50% mixed with soil from the collection site for periods of 7 (acute exposure), 15 (intermediary exposure) and 90 (subchronic exposure) days in each treatment. The animals from the control group did not present alterations at all the exposition times. As an acute response (7 days), it was observed in the animals exposed to mud at 1% and 10% an increase in the epithelial renovation and in the liberation of secretory vesicles of glycoprotein content. In the animals exposed to mud at 50%, the main alteration observed was the increase in the number of cytoplasmatic granules in the fat body cells; the epithelium seemed to be in reorganization process in all the observed individuals, suggesting an intense epithelial renovation. In the intermediary period of exposure (15 days), in the animals exposed to mud at 1% it was observed an increase in the number of haemocytes among the cells of the fat body, distributed both separately and grouped; the animals exposed to mud at 10% presented the same evidence of epithelial renovation observed in the animals exposed for 7 days at a concentration of 50% of mud; it was also observed an increase in the number of haemocytes, which reacted strongly to the PAS technique. Cytoplasmatic granules in the hepatic cells, fat body cells and haemocytes were also observed in great number, being that great part presented calcium in their constitution. In the animals exposed to mud at 50%, the alteration observed was the presence of numerous secretory vesicles of the apocrine type through all the epithelia of vacuolated aspect; hepatic cells with cytoplasmatic granules were also observed. As subchronic response (90 days), it was observed that all the fat body cells were completely taken by cytoplasmatic granules, into which it was, detected the presence of calcium; such granules were also observed in the apical region of the principal epithelial cells. These results suggest the existence of toxic agents in the analyzed mud sample, whose different concentrations trigger responses that occur in different manners, intensities and velocities that might prevent the entrance and action of toxic agents in the organism of the studied invertebrate.

Introduction

Morphological alterations may be used in the investigation of the toxicity of specific chemical compounds and in the monitoring of the chronic and acute effects in impacted environments. Such morphological alterations may provide qualitative evidences of a functional adaptation to an external environment (Meyers and Hendricks, 1985). The qualitative assessment of these changes before the death of the organism provides early indications of toxicity. In this context, histology has become widely used in studies with invertebrates aiming to identify different damages caused by harmful substances to the organisms (Triebskorn et al., 1999).

Paoletti et al. (1991) emphasize, in general, soil invertebrates efficiently indicate certain alterations in the terrestrial ecosystems since they are sensitive to environmental changes. Thus, physiological changes of organs and tissues may occur being promptly detected in the analysis of the cell (Köhler and Triebskorn, 1998). Depending on the intensity of the stressing factor and of the type of the affected organ, the cellular alterations may be diverse.

Diplopods are saprophagous invertebrates, agile in colonizing several layers of soil living in humid places that offer them nutrients for their development (Hopkin and Read, 1992). Colonization of diplopods in the soil provides a greater aeration and favorable conditions to the decomposition work of fungi and bacteria promoting a mineralization and dynamics of nutrients. Besides this, the diplopods excretion products (ammonia and uric acid), when degraded, enrich the soil with nitrates (Boccardo and Penteado, 1997).

Due to the habits of the diplopods, they have been considered as good environmental indicators used as test organisms in some soil analyses (Hopkin et al., 1985, Triebskorn et al., 1991, Köhler and Triebskorn, 1998).

The growing population of urban centers is an important producer of several residues among which we can highlight the sewage mud that presents potential for agricultural use and recovery of degraded soils since it contains a considerable percentage of organic matter and essential elements for plants, and can replace, even partially, the mineral fertilizers. However, sewage mud may also contain different concentrations of elements, potentially toxic, such as organic and inorganic contaminants, besides pathogens, which are retained in the nutrients of the sewage. Thus, in order to make viable the application of sewage mud in agriculture and in the recovery of degraded soils, it is necessary that the sewage mud passes by a characterization regarding its agronomic potential, the inorganic and organic substances potentially toxic, the presence of pathogenic agents, besides studies about its stability in the environment (MMA, 2006).

In this sense, considering the habits of the diplopods and the reduced number of studies using them as tools for the assessment of potential environmental contaminants, this study used such invertebrates as soil bioindicators by the analysis of the toxic effects of sewage mud of a STS (Sewage Treatment Station) from São Paulo State, Brazil in the midgut of the diplopod Rhinocricus padbergi, region of the digestive tube where the digestion occurs and nutrients are absorbed. This species was chosen due to its abundance and easy collection in the region where the study was developed, and it has already been studied under different viewpoints (Camargo-Mathias et al., 1998, Camargo-Mathias et al., 2004, Fantazzini et al., 1998, Fantazzini et al., 2002, Camargo-Mathias and Fontanetti, 2000, Fontanetti and Camargo-Mathias, 2004a, Fontanetti and Camargo-Mathias, 2004b, Fontanetti et al., 2004, Fontanetti et al., 2006).