Effects of heavy metals on ultrastructure and Hsp70 induction in Lemna minor L. exposed to water along the Sarno River, Italy A. Basile a,1 , S. Sorbo b , M. Cardi a,2 , M. Lentini a,2 , D. Castiglia a,c,2 , P. Cianciullo a,1 , B. Conte a,d,2 , S. Loppi e , S. Esposito a,n,2 a Dipartimento di Biologia, Università di Napoli “Federico II”, Italy b Ce.S.M.A, Microscopy Section, University of Naples Federico II, Via Foria 223, I-80139 Napoli, Italy c Institute of Biosciences and Bioresources – CNR – UOS Portici – Via Università 133, I-80055 Portici, Naples, Italy d Dipartimento di Scienze e Tecnologie, Università degli Studi del Sannio – Via Port'Arsa 11 – 82100 Benevento, Italy e Dipartimento di Scienze della Vita, Università di Siena, Via Mattioli 4, I- 53100 Siena, Italy article info Article history: Received 30 July 2014 Received in revised form 8 January 2015 Accepted 9 January 2015 Keywords: Lemna minor Freshwaters Heavy metals pollution HSPs TEM Sarno River abstract The effects of freshwater pollution in the highly contaminated river Sarno (Campania, Southern Italy) have been evaluated using bags containing the aquatic plant Lemna minor (Lemnacee, Arales), in order to determine morpho-physiological modifications as a response to pollutants. The exposition of Lemna bags for 7 days on three different sites along the river path showed al- terations in chloroplasts and vacuoles shape and organization. Moreover, some specimens were exposed in vitro at the same heavy metal (HM) concentrations measured in the polluted sites of the river, and compared with data from the bag experiment; to verify the dose and time dependent effects, samples were exposed to HM in vitro at concentrations ranging from 10 À6 to 10 À4 M up to 7 days. Transmission electron microscopy (TEM) observations on in vitro plants confirmed that ultra- structural alterations affected most of plastids and the shape of different subcellular structures, namely vacuoles; in in vitro stressed specimens, Heat Shock Proteins 70 (Hsp70) levels changed, in dependence of changing levels of HM measured in different sites along the river path. Thus L. minor exhibited a possible correlation between the levels of HM pollution and Hsp70 oc- currence; interestingly, the data presented showed that copper specifically increased Hsp70 levels at concentrations detected in polluted river waters, whereas cadmium and lead did not; on the other side, the latter represent highly toxic elements when specimens were exposed to higher levels in vitro. The effects of specific elements in vitro are compared to those observed in bags exposed along the river path; thus results are examined in order to propose L. minor as an organism able to be utilized to monitor heavy metals pollution; the possibility of using Hsp70 s as specific markers of HM pollution is discussed. & 2015 Elsevier Inc. All rights reserved. 1. Introduction Environmental pollution is a major issue for human health: namely, poisoning of freshwaters is of special concern, since rivers may transport contaminants very far away from the pollution source, thus hazardously modifying uncorrupted biosystems. Sarno River (Campania, Southern Italy) is about 25 km long, passes through a large plan comprising highly populated towns from Sarno, where the river springs, to Nocera, Pompei, Scafati and finally flows at Rovigliano in the gulf of Naples; the catchment area of the Sarno is 500 km 2 with a population of about 700,000 in- habitants living along the river basin (Albanese et al., 2013a). The Sarno is actually supplied by three springs, Foce, Palazzo and Santa Marina, originating three separate torrents joining in Sarno and forming a single main river stream; southwards, at San Marzano – Scafati, the river collects the waters from Alveo No- cerino, an artificial canal, collecting Cavaiola and Solofrana rivers, the latter being concerned by tanneries and therefore by chro- mium pollution (De Pippo et al., 2006; Albanese et al., 2013a). The Sarno basin could be divided in two environmental units: the first, corresponding to the hilly and mountain areas is char- acterized by a low anthropic pressure; the second, the econom- ically developed and densely populated areas of the river plain path, corresponds to the river valley where from moderate to very Contents lists available at ScienceDirect journal homepage: www.elsevier.com/locate/ecoenv Ecotoxicology and Environmental Safety http://dx.doi.org/10.1016/j.ecoenv.2015.01.009 0147-6513/& 2015 Elsevier Inc. All rights reserved. n Corresponding author. E-mail address: sergio.esposito@unina.it (S. Esposito). 1 Via Foria 223, I-80139 Napoli, Italy. 2 Complesso Universitario Monte Sant'Angelo, Via Cinthia 4, I-80126 Napoli, Italy. Ecotoxicology and Environmental Safety 114 (2015) 93–101