Production of methylmercury by sulphate-reducing bacteria in sediments from the orbetello lagoon in presence of high macroalgal loads

Milva Pepi, Claudio Leonzio, Silvano E. Focardi, Monia Renzi



Methylmercury is a potent neurotoxin affecting shallow-water ecosystems. Mercury polluted sediment samples were collected at six different sites in the Orbetello Lagoon (central Italy) characterized by high levels of silt, iron, manganese hydroxides, and organic matter originated the latter originated from the decomposition of macroalgae. Porous water pointed out the presence of sulphates, methylmercury, and sulphides. Slurries arranged in anaerobic conditions from sediment aliquots from the six sites, with the addition of ionic mercury, highlighted the production of methylmercury. Sulphate reducing bacteria (SRB) were quantified in lagoon sediments; furthermore, sediments cultured under anaerobic conditions showed SRBs active in mercury methylation. Anaerobic cultures of SRB, amended with ionic mercury, produced methylmercury during the growth of bacterial cells. The percentage of aerobic mercury resistant bacteria was pointed out at each sampling site, evidencing the presence of bioavailable mercury. Several aerobic mercury resistant bacteria were isolated and their level of resistance to inorganic and organic forms of mercury was evaluated. These isolates may be potentially used for eventual bioremediation processes. Mercury methylation by SRB in the Orbetello Lagoon sediments was described for the first time, focusing the attention on the need for possible bioremediation processes by using autochthonous mercury resistant bacteria. Moreover, the influence of algal biomass on mercury methylation was highlighted for the first time in this lagoon ecosystem. The importance of removing algal biomass, as it represents a source of organic matter favouring the process of mercury methylation, was strongly pointed out in this study.


Mercury-methylation; sulphate-reducing bacteria; mercury-resistant bacteria; macrophytes; Orbetello Lagoon

Full Text:



Aires T, Muyzer G, Serrão EA and Engelen AH (2019) Seaweed Loads Cause Stronger Bacterial Community Shifts in Coastal Lagoon Sediments Than Nutrient Loads. Front. Microbiol. 9:3283. doi: 10.3389/fmicb.2018.03283

Aldrige K.T. and Ganf G.G. 2003. Modification of sediment redox potential by three contrasting macrophytes: implications for phosphorus adsorption/desorption. Marine and Freshwater Research 54: 87-94.

Amsterdam, D. 1996. Susceptibility testing of antimicrobials in liquid media, p.52-111. In Loman, V., ed, Antibiotics in laboratory medicine, 4th ed. Williams and Wilkins, Baltimore, MD.

APAT-IRSA-CNR. 2003. Metodi analitici per le acque. APAT manuale linee guida. pp. 1153.

ARPAT – Agenzia Regionale per la Protezione Ambientale della Toscana, 2007a.Relazione finale.

ARPAT – Agenzia Regionale per la Protezione Ambientale della Toscana, 2007b. Programma di monitoraggio dell’ambiente marino costiero della Toscana Attività luglio 2006-gennaio 2007.

ASTM International Standards Worldwide. Method D854-06 Standard Test Methods for Specific Gravity of Soil Solids by Water Pycnometer. Disponibile al sito internet:

Baldi, F., M. Pepi and M. Filippelli. 1993. Methylmercury resistance in Desulfovibrio desulfuricans strains in relation to methylmercury degradation. Appl. Environ. Microbiol. 59: 2479-2485.

Barghigiani, C., D. Pellegrini, D. Gioffrè, S. De Ranieri and R. Bargagli. 1986. Preliminary results on the mercury content of Cytharus linguatula (L) in the northern Tyrrhenian Sea. Mar. Pollut. Bullet. 17: 424-427.

Barghigiani, C. and T. Ristori. 1995. The distribution of mercury in a Mediterranean area. In: C.J. Watras and J.W. Huckabee (Eds), Mercury Pollution: Integration and Synhtesis. Lewis Publisher, Boca Raton. pp 41.

Barkay, T. and A.J. Poulain. 2007. Mercury (micro)biogeochemistry in polar environments. FEMS Microbiol. Ecol. 59: 232-241.

Berner, R.A., Scott, M.R. and Thomlinson, C. 1970. Carbonate alkalinity in the pore water of anoxic marine sediments. Limnol Oceanogr 15: 544–549.

Boszke, L., Kowalski, A., Glosińska, G., Szarek, R. and Siepak, J. 2003. Environmental factors affecting speciation of mercury in the bottom sediments; an overview. Polish J. Environ. Studies 12: 5-13.

Boyd, E.S., Yu, R.-Q., Barkay, T., Hamilton, T.L., Baxter, B.K., Naftz, D.L. and Marvin-DiPasquale, M. 2017. Effect of salinity on mercury methylating benthic microbes and their activities in Great Salt Lake, Utah. Science of the Total Environment581–582: 495-506.

Bravo, A.G, Cosio, C., Amouroux, D., Zopfi, J., Chevalley, P.-A., Spangenberg, J.E., Ungureanu, V.-G. and Dominik, J. 2014. Extremely elevated methyl mercury levels in water, sediment and organisms in a Romanian reservoir affected by release of mercury from a chlor-alkali plant. Water Research49: 391-405.

Bravo, A.G., Bouchet, S., Tolu, J., Björn, E., Mateos-Rivera, A. and Bertilsson, S. 2016. Molecular composition of organic matter controls methylmercury formation in boreal lakes. Nature Communications 8:14255|DOI: 10.1038/ncomms14255|

Canario, J. and Vale, C. 2012. Mercury in wetlands: a contribution to the definition of a global mercury policy. SETAC North America 3rd Annual Meeting, November 11-15, 2012.

Caricchia, A.M., G. Minervini, P. Soldati, S. Chiavarini, C. Ubaldi, R. Morabitto. 1997. GC-ECD Determination of methylmercury in sediment samples using a SPB-608 capillary column after alkaline digestion. Microchem. Jour. 55: 44–55.

Caumette, P. 1986. Phototrophic sulphur bacteria and sulfate-reducing bacteria causing red water in a shallow brackish coastal lagoon (Prévost Lagoon, France).

Choi, S.C., J.T. Chase and R. Bartha. 1994. Metabolic pathways leading to mercury methylation in Desulfovibrio desulfuricans LS. Appl. Environ. Microbiol. 60: 4072-4077.

Coelho-Souza, S.A., Guimares, J.R., Mauro, J.B., Miranda, M.R. and Azevedo, S.M. 2006. Mercury methylation and bacterial activity associated to tropical phytoplankton. Science of Total Environment 364: 188-199.

Compeau, G.C. and R. Bartha. 1985. Sulfate-reducing bacteria: principle methylators of mercury in anoxic estuarine sediment. Appl. Environ. Microbiol. 50: 498-502.

Correia, R.R.S. and Guimarães, J.R.D. 2017. Mercury methylation and sulfate reduction rates in mangrove sediments, Rio de Janeiro, Brazil: The role of different microorganism consortia. Chemosphere 167: 438-443.

Dung T.T.T., Cappuyns V., Swennen R., Phung N.K. 2013. From geochemical background determination to pollution assessment of heavy metals in sediments and soils. Reviews in Environmental Science and Biotechnology: 12: 335-353.

Ekstrom, E.B., F.M. Morel and J.M. Benoit. 2003. Mercury methylation independent of the acethylcoenzyme A pathway in sulphate-reducing bacteria. Appl. Environ. Microbiol. 69: 5414-5422.

Ekstrom, E.B. and F.M. Morel. 2004. Mercury methylation by sulphate-reducing bacteria independent of vitamin B12. Mater Geoenviron. 51: 968-970.

Esiobu, N., Amund, O.O., Fakile, O.O. and Popoola, O.H. 1991. Occurrence and distribution of sulphate-reducing bacteria in a polluted lagoon. Biomedical Letters 46: 129-132.

Faganeli, J., Hines, M.E., Covelli, S., Emili, A. and Giani, M. 2012. Mercury in lagoons: An overview of the importance of the link between geochemistry and biology. Estuarine, Coastal and Shelf Science. 113: 126-132.

Fantozzi, L., Ferrara, R. and Dini, F. 2009. Dissolved gaseous mercury production in the dark: evidence for the fundamental role of bacteria in different types of Mediterranean water bodies. Science of Total Environment 407: 917-925.

Faust, S.D. and M.A. Osman. 1981. Mercury, arsenic, lead, cadmium, selenium, and chromium in aquatic environments, p. 200-205. In: Chemistry of natural water. Ann. Arbor. Science Publishers. Inc., Ann Arbor, Mich.

Fergusson, J.E. 1990. The heavy elements – chemistry, environmental impact and health effects. Oxford, New York, Beijing, Frankfurt, San Paulo, Tokyo, Toronto. Pergamon Press.

Figueiredo, N.L., Canário, J., O’Driscoll, N.J., Duarte, A. and Carvalho, C. 2016. Aerobic mercury-resistant bacteria alter mercury speciation and retention in the Tagus Estuary (Portugal). Ecotoxicology and Environmental Safety 124: 60-67.

Fleming, E.J., E.E. Mack, P.G. Green and D.C. Nelson. 2006. Mercury methylation from unexpected sources: molybdate-inhibited freshwater sediments and an iron-reducing bacterium. Appl. Environ. Microbiol. 72: 457-464.

Folk, R.L. 1954. The distinction between grain size and mineral composition in sedimentary-rock nomenclature. J. Geol. 62: 344-359.

Folk, R.L. and Ward, W.C. 1957. Brazos river bar: A study in the significant of grain size parameters. J. Sedimentol. Petrol. Tusla 27: 3-26.

Förstner, U. and Wittmann, G.T.W. 1983. Metal Pollution in the Aquatic Environment. Springer-Verlag, Heidelberg, 486 p.

Gagnon, C., Pelletier, E., Mucci, A. and Fitzgerald, W.F. 1996. Diagenetic behaviour of methylmercury in organic-rich coastal sediments. Limnol. Oceanogr. 41: 428.

Gardiner, E.E. 1972. Differences between ducks, pheasants, and chicken tissue Hg retention, depletion, and tolerance to increasing levels of dietary mercury. Can. J. Anim. Sci. 52: 419-427.

Gaudette, H.E. and Flight, W.R. 1974. An inexpensive titration method for the determination of organic carbon in recent sediments. J. Sediment. Petrol. 44: 249-253.

Gilmour, C.C., Henry, E.A. and Mitchell, R. 1992. Sulfate stimulation of mercury methylation in freshwater sediments. Environ. Sci. Technol. 26: 2281-2287.

Gilmour, C.C., Elias, D.A., Kucken, A.M., Brown, S.D., Palumbo, A.V., Schadt, C.W. and Wall, J.D. 2011. Sulfate-reducing bacterium Desulfovibrio desulfuricans ND132 as a model for understanding bacterial mercurymethylation. Applied and Environmental Microbiology 77: 3938-3951.

Giovani A, Mari E, Specchiulli A, Cilenti L, Scirocco T, Breber P, Renzi M, Focardi SE, Lenzi M. 2010. Factors affecting changes in phanerogam distriburion patterns of Orbetello lagoon, Italy. Transit Waters Bull 4(1): 35-52.

Giusti, E., Marsili-Libelli, S., Renzi, M., Focardi, S.2010. Assessment of spatial distribution of submerged vegetation in the Orbetello lagoon by means of a mathematical model. Ecological Modelling 221: 1484-1493.

Grassi, S. and Netti, R. 2000. Seawater intrusion and mercury pollution of some coastal aquifers in the province of Grosseto (Southern Tuscany e Italy). Journal of Hydrology 237: 198-211.

Guentzel, J.L., Powell, R.T., Landing, W.M. and Mason, R.P. 1996. Mercury associated with colloidal material in estuarine and open-ocean environment. Mar. Chem. 55: 177.

Guerranti, C., Cannas, S., Scopetani, C., Fastelli, P., Cincinelli, A., Renzi, M., 2017. Plastic litter in aquatic environments of Maremma Regional Park (Tyrrhenian Sea, Italy): Contribution by the Ombrone river and levels in marine sediments. Marine Pollution Bulletin, 117: 366-370.

Hamasaki, T., Nagase, H., Yoshioka, Y. and Sato, T. 1995. Formation, distribution, and ecotoxicology of methylmetals of tin, mercury, and arsenic in the environment. Critical Review in Environ. Sci. Technol. 25: 45.

Han, S., Obraztsova, A., Pretto, P., Choe, K.-Y., Gieskes, J., Deheyn, D.D. and Tebo, B.M. 2007. Biogeochemical factors affecting mercury methylation in sediments of the Venice lagoon, Italy. Environmental Toxicology and Chemistry 26: 655-663.

Han, S., Narasingarao, P., Obraztsova, A., Gieskes, J., Hartmann, A.C., Tebo, B.M., Allen, E.E. and Deheyn, D.D. 2010. Mercury speciation in marine sediments under sulfate-limited conditions.Environmental Science and Technology 44: 3752-3757.

Heinz, G. 1974. Effects of low dietary levels of methylmercury on mallard reproduction. Bull. Environ. Contam. Toxicol. 13: 554-559.

Hosokawa, J. 1995. Remediation work for Hg contaminated bay-experiences of Minamata Bay Project, Japan. Water Sci. Technol. 28: 338-348.

ICMGP: 8th International Conference on Mercury as a Global Pollutant (2006). Madison Conference Declaration on Mercury Pollution. J.P. Hurley, D.P. Krabbenhoft co-chairs. Madison, WI, USA. 8 pp.

Kannan, K. and Falandysz, J. 1998. Speciation and concentrations of mercury in certain coastal marine sediments. Water, Air & Soil Pollut. 103: 129.

Kim, M., Han, S., Gieskes, J. and Deheyn, D.D. 2011. Importance of organic matter lability for monomethylmercury production in sulfate-rich marine sediments. Science of the Total Environment 409: 778-784.

King, J.K., J.E. Kostka, M.E Frisher and F.M. Saunders. 2000. Sulfate-reducing bacteria methylate mercury at variable rates in pure culture and in marine sediments. Appl. Environ. Microbiol. 66: 2430-2437.

Köpke, B., Wilms, R., Engelen, B., Cypionka, H. and Sass, H. 2005. Microbial diversity in coastal subsurface sediments: a cultivation approach using various electron acceptors and substrate gradients. Appl. Environ. Microbiol. 71: 7819-7830.

Krumbein, W.C. 1934. Size frequency distributions of sediments, J. Sedim. Petrol. 4: 65-77.

Janssen, S.E., Schaefer, J.K., Barkay, T. and Reinfelder, J.R. 2016. Fractionation of mercury stable isotopes during microbial methylmercury production by iron- and sulfate-reducing bacteria. Environmental Science and Technology 50: 8077-8083.

Jay, J.A., Morel, F.M.M. and Hemond, H.F. 2000. Mercury speciation in the presence of polysulfides. Environ. Sci. Technol. 34: 2196.

ICRAM (triennio 2001–2003), Sedimenti-scheda 3- Analisi delle caratteristiche granulometriche, in Metodologie analitiche di riferimento, Ministero dell’Ambiente e dellaTutela del Territorio, Servizio Difesa Mare, Ed. ICRAM.

ICRAM Istituto Centrale per le Ricerche Applicate al Mare. 2001–2003. Sedimenti-scheda 4- Analisi di carbonio totale e organico (metodo analizzatore elementare), in Metodologie analitiche di riferimento, Ministero dell’Ambiente e Tutela del Territorio, Servizio Difesa Mare, Ed. ICRAM.

Llobet-Brossa, E., Rabus, R., Böttcher, M.E., Könneke,M., Finke, N., Schramm, A., Meyer, R.L., Grötzschel, S., Rosselló-Mora, R., Amann, R. 2002. Community structure and activity of sulfate-reducing bacteria in an intertidal surface sediment: a multi-method approach. Aquat. Microb. Ecol. 29: 211–226.

Meson, R., N. Bloom, S. Cappellino, G. Gill, J. Benoit and C. Dobbs. 1998. Investigation of pore water sampling methods for mercury and methylmercury. Environ. Sci. Technol. 32: 4031-4040.

Matida, Y., H. Kumada, S. Kimura, Y. Saiga, T. Nose, M. Yokote and H. Kawatsu. 1971. Toxicity of mercury compounds to aquatic organisms and accumulation of the compounds by the organisms. Bull. Freshwater Fish. Res. Lab. (Tokyo) 21: 197-227.

Mazrui, N.M., Jonsson, S., Thota, S., Zhao, J. and Mason, R.P. 2016. Enhanced availability of mercury bound to dissolved organic matter for methylation in marine sediments. Geochimica et Cosmochimica Acta194: 153-162.

Miniero, R., Beccaloni, E., Carere, M., Ubaldi, A., Mancini, L., Marchegiani, S., Cicero, M.R., Scenati, R., Lucchetti, D., Ziemacki, G.and De Felip, E.2013.Mercury (Hg) and methyl mercury (MeHg) concentrations in fish from the coastal lagoon of Orbetello, central Italy. Marine Pollution Bulletin 76: 365-369.

Moreau, J.W., Gionfriddo, C.M., Krabbenhoft, D.P., Ogorek, J.M., DeWild, J.F., Aiken, G.R. and Roden, E.E. 2015. The effect of natural organic matter on mercurymethylation by Desulfobulbus propionicus 1pr3. Frontiers in Microbiology 6: Article number 01389.

Nelson, J.D., W.R. Blair, F.E. Brinckman, R.R. Colwell, W.P. Iverson. 1973. Biodegradation of phenylmercury acetate by mercury-resistant bacteria. Appl. Environ. Microbiol. 26: 321-326.

Olsen, M., Schaanning, M.T., Veiteberg Braaten, H.F., Eek, E., Moy, F.E. and Lydersen, E. 2018. The influence of permanently submerged macrophytes on sediment mercury distribution, mobility and methylation potential in a brackish Norwegian fjord. Science of the Total Environment 610-611: 1364-1374.

Pepi, M., Gaggi, C., Bernardini, E., Focardi, S., Lobianco, A., Ruta, M., Nicolardi, V., Volterrani, M., Gasperini, S., Trinchera, G., Renzi, P., Gabellini, M. and Focardi S.E. 2011. Mercury-resistant bacterial strains Pseudomonas and Psychrobacter spp. isolated from sediments of Orbetello Lagoon (Italy) and their possible use in bioremediation processes. Int. Biodeter. Biodegr. 65: 85-91.

Pepi, M., Focardi, S., Tarabelli, A., Volterrani, M. and Focardi, S.E. 2013. Bacterial strains resistant to inorganic and organic forms of mercury isolated from polluted sediments of the Orbetello Lagoon, Italy, and their possible use in bioremediation processes. E3S Web of Conferences 1, 31002.

Perra, G., Renzi, M., Guerranti, C., Focardi, S.E. 2010. Polycyclic aromatic hydrocarbons pollution in sediments: distribution and sources in a lagoon system (Orbetello, Central Italy). Transitional Waters Bulletin 38(1): 45-58.

Petranich, E., Covelli, S., Acquavita, A., Faganeli, J., Horvat, M., Contin, M. 2018. Evaluation of mercury biogeochemical cycling at the sediment–water interface in anthropogenically modified lagoon environments. Journal of Environmental Sciences (China) 68: 5-23.

Porrello S, Lenzi M, Ferrari G, Persia E, Tomassetti P, 2005. Loading of nutrient from a land based fish farm (Orbetello, Italy) at different time. Aquaculture International 13(1-2): 97-108.

Renzi, M., 2007. Distribuzione dei banchi di fanerogame nella laguna di Orbetello inrelazione alle caratteristiche chimico-fisiche dei sedimenti, PhD thesis, Italianlanguage, pp. 200.

Renzi, M., Guerranti, C. 2018. Seasonal fluctuations of trace elements from different habitats of Orbetello Lagoon (Thyrrenian Sea, Italy). Archives of Environmental Contamination and Toxicology, 74(1):92-113.

Renzi M, Lenzi M, Franchi E, Tozzi A, Porrello S, Focardi S, Focardi SE. 2007. Mathematical modelling of sediment chemico-physical parameters in a coastal lagoon to estimate high density seagrass meadow (Ruppia cirrhosa) distribution Int J Environ Health 1(3): 360-374.

Renzi M, Perra G, Guerranti C, Franchi E, Focardi S. 2009. Abatement efficiency of municipal wastewater treatment plants using different technologies (Orbetello Lagoon, Italy) Int J Environ Health 3(1): 58-70.

Renzi M, Specchiulli A, Baroni D, Scirocco T, Cilenti L, Focardi S, Breber P, Focardi SE. 2012. Trace elements in sediments and bioaccumulation in fish (Anguilla anguilla) in a Mediterranean lagoon (SE Italy) Int J Environ An Ch 92(6): 676-697.

Renzi, M., Guerranti, C., Giovani, A., Perra, G., Focardi, S.E. 2013. Perfluorinated compounds: Levels, trophic web enrichments and human dietary intakes in transitional water ecosystems. Marine Pollution Bulletin 76: 146-157.

Renzi M, Mariottini M, Specchiulli A, Perra G, Guerranti C, Giovani A, Focardi SE. 2013. Occurrence of POPs in sediments and tissues of European eels (Anguilla anguilla L.) from two Italian lagoons: Varano and Orbetello. Transit Waters Bull 6(3): 1-18

Romano, E., Bergamin, L., Croudace, I.W., Ausili, A., Maggi, C.and Gabellini, M. 2015. Establishing geochemical background levels of selected trace elements in areas having geochemical anomalies: The case study of the Orbetello lagoon (Tuscany, Italy). Environmental Pollution 202: 96-103.

Schartup, A.T., Mason, R.P., Balcom, P.H., Hollweg, T.A. and Chen, C.Y. 2013. Methylmercury Production in Estuarine Sediments: Role of Organic Matter. Environ. Sci. Technol.47: 695–700.

Sand-Jensen K, Prahl C, Stokholm H, 1982.Oxygen release from roots of submerged aquaticmacrophytes. Oikos 38: 349-354.

Siciliano, S.D., N.J. O’Driscoll, R. Tordon, J. Hill, S. Beauchamp and D.R. Lean. 2005. Abiotic production of methylmercury by solar radiation. Environ. Sci. Technol. 39: 1071-1077.

Simpson, S.L. 2001. A rapid screening method for acid-volatile sulfide in sediments. Environ. Toxicol. Chem. 20: 2657-2661.

Specchiulli A, Focardi S, Renzi M, Scirocco T, Cilenti L, Breber P, Bastianoni S. 2008. Environmental heterogeneity patterns and assessment of trophic levels in two Mediterranean lagoons: Orbetello and Varano, Italy. Sci Total Environ 402: 285-298.

Tessier, A. and Campbell, P. 1988. Partitioning of trace metals in sediments. In: Kramer J.R., Allen H. (eds) Metal speciation: theory, analysis and application. Lewis Publishers, Chelsea, MI, USA, pp 183–199.

UNEP/FAO/WHO, 1987. Assessment of the state of pollution of the Mediterranean Sea by mercury and mercury compounds. MAP Tech. Rep. Series N° 18.

US EPA, U.S. Method 3051, Microwave assisted acid digestion of sediments, sludges, soil, and oils, Environmental Protection Agency, Washington, DC, 1994,

US EPA, U.S. Method 245.5, Mercury in sediments by manual cold vapour atomic absorption (CVAA), Environmental Protection Agency, Washington, DC, 2001,

U.S. Environmental Protection Agency (U.S. EPA) Environmental Protection Agency, 1997.

Udden, J.A. 1914. Mechanical composition of clastic sediments, Bull. Geol. Soc.Am. 25: 655-744.

Viaroli, P., Bartoli, M., Bondavalli, C., Christian, R., Giordani, G., Naldi, M, 1996. Macrophyte communities and their impact on benthic fluxes of oxygen, sulphide and nutrients in shallow eutrophic environment. Hydrobiologia 329: 93-103.

Weber, J.H. 1993. Review of possible paths for abiotic methylation of mercury (II) in the aquatic environment. Chem. 26: 2063-2077.

Wentworth, C.K. 1922. A scale of grade and class terms for clastic sediments, J. Geol. 30: 377-392.

Zhang, T., Kucharzyk, K.H., Kim, B., Deshusses, M.A. and Hsu-Kim, H. 2014. Net methylation of mercury in estuarine sediment microcosms amended with dissolved, nanoparticulate, and microparticulate mercuric sulfides. Environmental Science and Technology. 48: 9133-9141.

Partnerzy platformy czasopism