Biomonitoring of heavy metal pollution in Malta

Abstract

Heavy metals have been classified amongst the most potentially deleterious contaminants to biota and human consumers of sea food (Martin and Richardson, 1991 ). The need to gain further information on the heavy metal pollution status of coastal waters has thus been recognised on both a national and regional scale. In this regard, Malta is directing its efforts towards monitoring heavy metal pollution within the framework of a biomonitoring programme for the Mediterranean Region. This investigation aimed to assess the feasibility of detecting heavy metal pollution using a biomonitoring approach. This entails measuring the biochemical response of an indicator species to contaminant exposure and thus provides an early warning signal of environmental metal contamination. The biological response or stress index which is being proposed is the induced synthesis of the metal-binding protein metallothionein in the presence of heavy metal contamination. The suitability for heavy metal biomonitoring of two locally abundant mollusc species, the limpet Patella rustica and the neogastropod snail Hexaplex truncu/us was assessed. Both species were used in previous studies for monitoring of metals and tributyltin respectively. They were sampled from six locations along the north east coast of Malta. These included Qawra Point (reference area); Wied Ghammieq, Xghajra and Marsascala (constituting an environmental pollution gradient); Bahar ic-Caghaq (Maghtab Landfill area) and Manoel Island, two suspected 'hot spots' of metal pollution. The molluscs were found to accumulate significantly high levels of cadmium, copper and zinc in Xghajra, Marsascala and Bahar ic-Caghaq when compared to the reference site. The high copper and zinc levels measured in the limpet in the present study were comparable to values in similar biota from foreign industrialised polluted areas. Hexaplex trunculus accumulated inherently high levels of copper and cadmium, the latter metal probably playing a physiological role in shell formation. On the basis of chemical data (i.e. total metal content in tissues) and biological data (the role of metallothioneins in metal sequestration), Patella rustica was selected as the more suitable sentinel species for monitoring purposes and a valuable substitute to Mytilus galloprovincialis, the official MEDPOL indicator species. Metallothioneins (MTs) constituted suitable biomarkers of metal pollution in P.rustica since measurements of protein levels, performed using gel-filtration chromatography, were generally correlated with heavy metal concentrations in the cytosol and limpet soft parts and thus provided a clear indication of the pollution status of the sites investigated. The correlation was weaker for zinc since this metal was bound mainly to high molecular weight proteins. Moreover, zinc was preferentially distributed in the non-cytosolic compartment of the cell. The kinetics of cadmium accumulation and sequestration were studied in Patella rustica via in vitro exposure of the limpet to 500µgCd/L for three weeks. The induction of metallothioneins provided a quantifiable biological response to sub-lethal levels of cadmium. Also, it was observed that the accumulated cadmium was transferred preferentially to the particulate fraction of the cell (presumably in lysosomes or insoluble granules). The application of a relatively simple spectrophotometric assay (described by Viarengo et al., 1997) for the semi-quantitation of metallothioneins was evaluated. This technique can be considered as a compromise between sensitivity and cost and sees a practical application for routine monitoring purposes. It yielded the same results in terms of the identification of 'hot spots' of metal contamination as those obtained from gelfiltration chromatography. Significant differences in protein levels amongst the sites investigated were observed in both species. Limpets and snails collected from Xghajra, Marsascala, Bahar ic-Caghaq and Manoel Island exhibited significantly higher MT levels than those in the reference site. Data of MT concentrations were correlated with total tissue metal levels and with cytosolic cadmium and zinc in P.rustica. MT levels in Xghajra limpets were tentatively linked not only to metal contamination but also to pollution by organic compounds suggesting that MTs could be used as an index of general environmental stress. Seasonal fluctuations in protein levels in the limpet were obtained with the highest levels measured in spring and the lowest in autumn. These variations were related to the reproductive and growth cycles of the limpet. Spring was suggested as the season which lends itself best to biomonitoring since the variations in protein levels due to the reproductive cycle can be excluded. The largest quantitative differences in MT levels between the sampling points and reference site were most evident in this season. In the conclusion, a set of recommendations and guidelines are proposed to assist in the planning and application of a local biomonitoring programme using Patella rustica as the sentinel species.