ORIGINAL ARTICLE Arsenic-resistant bacteria isolated from contaminated sediments of the Orbetello Lagoon, Italy, and their characterization M. Pepi 1 , M. Volterrani 1 , M. Renzi 1 , M. Marvasi 2 , S. Gasperini 3 , E. Franchi 1 and S.E. Focardi 1 1 Department of Environmental Sciences, University of Siena, Siena, Italy 2 Department of Genetics and Animal Biology, University of Florence, Florence, Italy 3 Services Centre, Faculty of Sciences, University of Siena, Siena, Italy Introduction Arsenic is a metalloid widely distributed in soils and nat- ural waters, released both from natural and anthropogenic sources, from the weathering of rocks or by mining industries and agricultural practices. It is found in the oxidation states +5 (arsenate), +3 (arsenite), 0 (elemental arsenic) and )3 (arsine). It has been classified as a human carcinogen by the International Agency for Research on Cancer (IARC 1987) and the US Environmental Protec- tion Agency (EPA 1988). The most common oxidation states of arsenic in the environment are the pentavalent As(V) and trivalent As(III) forms (Cullen and Reimer 1989). Of these two, As(III) is more toxic and can inhibit various dehydrogenases (Ehrlich 1996); arsenite (AsO 2- or AsO 3 2- ) is able to bind sulfhydryl groups of proteins and dithiols such as glutaredoxin. Arsenate (AsO 4 3- ) acts as a structural analogue of phosphate and inhibits oxidative phosphorylation by producing unstable arsenylated deriv- atives (Da Costa 1972). Its biogeochemical cycle strongly depends on microbial transformations, which affects the mobility and the distribution of arsenic species in the environment (Mukhopadhyay et al. 2002). Although arsenic is generally toxic to life, it has been demonstrated that micro-organisms can use arsenic compounds as elec- tron donors, electron acceptors or possess arsenic detoxi- fication mechanisms (Ji and Silver 1992; Ahmann et al. 1994; Cervantes et al. 1994; Newmann et al. 1997; Stolz Keywords Aeromonas, arsenic contamination, arsenic resistance, Bacillus, bioremediation, Orbetello Lagoon, Pseudomonas, 16S rRNA gene. Correspondence Milva Pepi, Department of Environmental Sciences, University of Siena, Via P.A. Mattioli, 4; 53100 Siena, Italy. E-mail: pepim@unisi.it 2006 ⁄ 1610: received 18 November 2006, revised 6 March 2007 and accepted 8 May 2007 doi:10.1111/j.1365-2672.2007.03471.x Abstract Aims: The aim of this study was to isolate arsenic-resistant bacteria from con- taminated sediment of the Orbetello Lagoon, Italy, to characterize isolates for As(III), As(V), heavy metals resistance, and from the phylogenetic point of view. Methods and Results: Enrichment cultures were carried out in the presence of 6Æ75 mmol l )1 of As(III), allowing isolation of ten bacterial strains. Four iso- lates, ORAs1, ORAs2, ORAs5 and ORAs6, showed minimum inhibitory con- centration values equal or superior to 16Æ68 mmol l )1 and 133Æ47 mmol l )1 in the presence of As(III) and As(V), respectively. Isolate ORAs2 showed values of 1Æ8 mmol l )1 in the presence of Cd(II) and 7Æ7 mmol l )1 of Zn(II), and isolate ORAs1 pointed out a value of 8Æ0 mmol l )1 in the presence of Cu(II). Analysis of 16S rRNA gene sequences revealed that they can be grouped in the three genera Aeromonas, Bacillus and Pseudomonas. Phylogenetic analysis of the four more arsenic-resistant strains was also performed. Conclusion: Isolates are highly resistant to both As(III) and As(V) and they could represent good candidates for bioremediation processes of native pol- luted sediments. Significance and Impact of the Study: This study provides original results on levels of resistance to arsenic and to assigning genera of bacterial strains isola- ted from arsenic-polluted sediments. Journal of Applied Microbiology ISSN 1364-5072 ª 2007 The Authors Journal compilation ª 2007 The Society for Applied Microbiology, Journal of Applied Microbiology 103 (2007) 2299–2308 2299