Bioremediation of weathered-buildingstone surfacesAlison Webster and Eric May

School of Biological Sciences, University of Portsmouth, Portsmouth, Hampshire, PO1 2DY, UK

Atmospheric pollution and weathering of stone surfaces

in urban historic buildings frequently results in disfig-

urement or damage by salt crust formation (often

gypsum), presenting opportunities for bioremediation

using microorganisms. Conventional techniques for the

removal of these salt crusts from stone have several

disadvantages: they can cause colour changes;

adversely affect the movement of salts within the

stone structure; or remove excessive amounts of the

original surface. Although microorganisms are com-

monly associated with detrimental effects to the

integrity of stone structures, there is growing evidence

that they can be used to treat this type of stone

deterioration in objects of historical and cultural

significance. In particular, the ability and potential of

different microorganisms to either remove sulfate crusts

or form sacrificial layers of calcite that consolidate

mineral surfaces have been demonstrated. Current

research suggests that bioremediation has the potential

to offer an additional technology to conservators work-

ing to restore stone surfaces in heritage buildings.

Introduction

The term bioremediation covers a range of processes thatuse microorganisms to return contaminated environmentsto their original condition. The use of microorganisms ortheir enzymes to deal with contaminated soils, oil spills orchemical waste are well-developed biotechnologies [1,2]but the application of bioremediation to ameliorate theeffects of stone deterioration is less well known. Deterio-ration of building stone begins from the moment it isquarried due to natural weathering processes [3]. Otherfactors, sometimes acting synergistically, includingcrystallization of soluble salts, pollution and biologicalcolonisation can accelerate natural deterioration [4–8].Whatever the cause of stone deterioration, many buildingsrequire remedial measures to stabilize the surface layerand prevent further loss from external sources. Thisreview offers an insight into how biotechnology researchhas addressed the needs of conservators working withbuilding stone and considers how far bacterially mediatedbioremediation can be used in practice.

Stone and soluble salts

Internal pressures created by crystallization, hydrationand thermal expansion of salts are a significant cause of

Corresponding author: May, E. (eric.may@port.ac.uk).Available online 2 May 2006

www.sciencedirect.com 0167-7799/$ - see front matter Q 2006 Elsevier Ltd. All rights reserved

damage to stone [9–11]. Stone is hydrophilic, and will takeup water from the ground and adjoining stone. Solublesalts from the soil, the atmosphere and applications on thesurface can dissolve in this aqueous environment andmove through the pores. Highly soluble salts are usuallydeposited on the surface, and can be brushed off; however,less soluble salts might expand below surface level,ultimately causing the loss of the outer layers [12,13].Accumulation of sulfates, many derived from the oxidationof sulfur dioxide, is of particular concern. Those tech-niques that attempt to remove salts from heritage stoneartefacts by washing with water are either not practicableor accelerate degradation [14]. Nitrates, which canoriginate from the numerous oxides of nitrogen presentin the atmosphere (N2O, NO, N2O3, NO2, N2O5), can alsoaccumulate but because they have a high solubility theymigrate from the surface or are washed away by rain [15].Thus, unlike sulfates, nitrate accumulation is not asurface phenomenon and has not been the subject ofextensive study.

Black sulfate crusts

Many of the additional factors that accelerate deterio-ration of stone are still being definitively characterised,and much work has been carried out to highlight the roleof biological organisms in the blackening of stone [16,17].It is, however, the effects of anthropogenic sources ofpollution on building stone that are arguably the singlemost important factor in the production of black discolor-ation [18]. The burning of fossil fuels has led to an increasein the concentrations of acid gases in the atmosphere and,of these, perhaps the most important is sulfur dioxide.When dissolved in water [19,20], sulphur dioxide formssulphurous acid, which is oxidised to sulfuric acid; this, inturn, reacts with calcium carbonate to form calciumsulfate, the mineralized form of which is known as gypsum[21]. The formation of gypsum leads to the creation ofcavities below the surface because of the migration ofcalcium ions to the surface [22]. Thus, when the solublegypsum is washed away it takes with it some of the stoneitself, initially causing loss of surface detail but eventuallyleading to a loss of structural integrity.

On the surface of the stone, particulate matter from theatmosphere can combine with gypsum to leave unsightlyblack crusts [23,24]. Carbonaceous particles were thoughtto be the most significant element in black crusts butrecent work has shown that they also contain a complexmixture of aliphatic and aromatic carboxylic acids and

Review TRENDS in Biotechnology Vol.24 No.6 June 2006

. doi:10.1016/j.tibtech.2006.04.005

Review TRENDS in Biotechnology Vol.24 No.6 June 2006256

polycyclic aromatic hydrocarbons [25]. Sabbioni et al. [26]concluded that the oxalate found in these crusts probablyoriginated from a combination of microbial metabolismand protective treatments; formate and acetate anionsfound in the black crust were believed to be fromatmospheric pollutants. Furthermore, Schiavon et al.noted that the types of pollutants found in urban buildingstones are changing to reflect the atmospheric pollutantsof the time [27].

Figure 1. Examples of biorestoration. Left: Marble balcony support arm from a

building in Athens undergoing restoration showing black sulfated crusts. Right:

detail of a balcony scroll before (top) and after (bottom) application. Removal of

black crust is apparent after a single application of sulfate-reducing bacteria,

suspended in gel, after 48 hours. Further removal can be achieved with repeat

applications and can be tailored to the needs of the conservator.

Bioremediation of black sulfated crusts

Attempts to restore stone surfaces by removal of blackcrusts and salts through mechanical stone cleaning hasbegun to be called into question by some conservators asthe effects of past cleaning regimes become more evident[28]. It is clear that such cleaning can result in severaltypes of damage, some of which is immediately apparent(e.g. loss of surface), whereas in other cases the damagemight not become apparent for several years, for example,where a single dominant species of microorganismreplaces complex established microbiota removed in thecleaning process [29,30]. Indeed, some conservatorsbelieve certain surface deterioration should not beremoved. For example, patinas (surface layers developedover extended periods by biological and material-relatedfactors) removed from the Parthenon were associated withthe best-preserved surfaces and it was therefore rec-ommended that these should remain intact [31].

Thus, the removal of black crusts is problematic forconservators because conventional cleaning techniquescan potentially remove a portion of the underlying stone[32]. Skoulikidis and Beloyannis reported that gypsumcould be converted back to calcite using carbonate anionsin aqueous solution [33]. They found that the gypsumlayer was consolidated by the new calcite, which showedsimilar behaviour to the underlying marble. Although noother authors reported success using purely chemicalreactions, the use of microorganisms, a relatively recenttechnique, has been successful in removing sulfate fromblack gypsum crusts. Sulfate-reducing bacteria are able todissociate gypsum into Ca2C and SO4

2C ions, and the SO42C

ions are then reduced by the bacteria, whereas the Ca2C

ions react with carbon dioxide to form new calcite [32]:

6CaSO4 C4H2OC6CO2/6CaCO3 C4H2SC2SC11O2:

In 1970, Moncreiff and Hempel [34] described the use ofa ‘biological pack’ and the role of microorganisms in apoultice. The first successful application of the anaerobicsulfate reducer Desulfovibrio desulfuricans was reportedby Atlas et al. [35] and Gauri and Chowdhury [36].Heselmeyer et al. applied Desulfovibrio vulgaris togypsum crusts, which brought about the conversion tocalcite [14]. D. desulfuricans was again used by Gauri etal. to remove sulfates from the black crust on marble [37],and Kouzeli, also working with marble, reported goodresults compared with chemically based pastes [38].Saiz-Jimenez presented evidence that demonstratedmicroorganisms removed some of the most abundantcomponents of black crusts, such as gypsum and polycyclicaromatic hydrocarbons [39]; however, he did not consider

www.sciencedirect.com

that microorganisms would be of use in the bioremediationof buildings because of the practical difficulties imposed byfactors such as the size of the buildings and thetime required.

Ranalli et al. [40,15,41], unlike other researchers, havenot reported the deposition of calcite but, using D.desulfuricans and D. vulgaris, they demonstrated thesuccessful removal of black crusts from marble. A singleapplication of sulfate-reducing bacteria to an urbanmarble structure illustrates this ability (Figure 1).

Consolidation of stone surfaces

The desire to protect and consolidate stone surfaces has along history, with evidence for Roman use of pastes appliedto recently sculpted stone [42]. Traditional methods such asshelter coatings are made from slaked lime (calciumhydroxide), which combines with carbon dioxide in theatmosphere and hardens to form calcium carbonate in aprocess known as carbonation. The formation of calciumcarbonate crystals in the form of calcite is not an immediateprocess and can take up to 80 days when applied to wallpaintings [43]. External shelter coatings, such as limewash,last between 10 and 15 years but because they are subject tothe same deterioration processes as the underlying stone,they are eventually washed away and only providerelatively short-term protection [44]. Another method thatworks with the underlying stone is the generation of acalcium oxalate patina. Surface patinas can be formed,naturally, during time by oxidation of calcite with oxalicacid, and some authors consider a biological origin the mostprobable cause, for example, those observed at TarragonaCathedral, Spain [45]; other patinas might be anthropo-genic in origin, the result of protective treatments applied inthe past [46]. Cezar reported on work that has been carriedout in the laboratory to generate calcium oxalate layers,chemically, in a matter of hours rather than years [44].

Biomineralization

An alternative to the chemical generation of calciumcarbonate is the exploitation of a common phenomenon inliving organisms – biomineralization. This activity is

Figure 2. Crystal production under laboratory conditions. Left: crystal formation by

two different bacterial cultures growing on B4 solid medium [48]. Top: unidentified

bacteria isolated from a karstic stream. Bottom: Sphingomonas paucimobilis.

Crystal formation occurs within 10 hours of inoculation (magnification!400). Right

panel is a scanning electron micrograph showing a layer of calcified Pseudomonas

putida cells on Portland limestone, following the contours of the underlying

substrate. Magnification!5000, BarZ1 mm.

Review TRENDS in Biotechnology Vol.24 No.6 June 2006 257

widespread across many phyla, and molecular studies ofbone and shell suggest that there might be a commongenetic ancestry [47]. Biomineralization by bacteria(Figure 2) has provoked controversy, with some authorsbelieving that crystal production is a purely chemicalby-product, whereas others assert that microorganismsare actively involved in the process. Ehrlich [49] definedmicrobial mineral formation as either ‘active’, involvingenzymes or metabolic products, or ‘passive’, where evendead cells can produce minerals. Rivadeneyra et al.propose that the formation of calcium carbonate crystalsin the presence of Nesterenkonia halobia involves bothbiological and inorganic processes [50]. Given thatorganisms are in contact with the available precursorsrequired for crystal formation, it is perhaps unsurprisingthat new mineral material is produced on stone, and thishas, indeed, been observed in cyanobacteria followinginvasion of stone and lichens [51,52]. Urzi et al. isolatedmicroorganisms from stone surfaces and found that themajority precipitate CaCO3 in the form of calcite [53];furthermore, most of the common bacteria associated withbuilding stones can also induce precipitation in thelaboratory. Such examples demonstrate the complexinter-relationship between biological organisms andminerals, leading to elements of both destruction andconsolidation of the substratum.

Biomineralization of stone

Although biomineralization has been observed for manyyears, the potential for its use in stone consolidation hasonly been explored relatively recently. In 1990, Adolpheand others applied for a European patent for thegeneration of calcite through the action of bacteria. Orialet al. [54,55] also examined the formation of sacrificiallayers by bacteria and considered that it offered apromising avenue for treatment of historic buildings.Le Metayer-Levrel et al. used several different strains ofbiocalcifying bacteria to promote successful bacterialcarbonatogenesis on the surface of limestone buildings,statuary and monuments [56]. A variety of uses have beenfound for biocalcifying bacteria, as seen in the work ofBang et al. who used Bacillus pasteurii immobilised in

www.sciencedirect.com

polyurethane to fill cracks in concrete [57] and Ramachan-dran et al. who found that an increase in compressivestrength and stiffness can be achieved by combiningbacteria with sand in cracks [58]. Rodriguez-Navarroet al. also proposed the use of Myxococcus xanthus in stoneconservation and observed newly formed carbonate, whichcreated a cement that adhered strongly to the substratum:the bacteria induced carbonate cementation to a depth ofO500 mm and no plugging or blocking of pores wasobserved [59]. This newly formed carbonate was moreresistant to mechanical stress in the form of sonication,possibly because of the incorporation of organic moleculesproduced by bacterial metabolism into the crystals. Thisagrees with the work of Morse, who discovered thatbacterially induced calcite was less soluble that inorgani-cally produced calcite [60]. Castanier et al. reported thegeneration of bioconsolidating cement a few micrometresthick, with significant carbonate production occurringwithin 5–10 days [61]. Limited EPS (extracellularpolymeric substances) formation was observed whenusing buffered media M-3 but small quantities wereformed when growing the bacteria using unbuffered M-3,indicating the importance of the choice of pH andmedia components.

The application of microorganisms and a suitablegrowing media directly to stone surfaces has severalpotential problems, including the formation of EPS,blocking of pores and promotion of microbial growth onexcess media. To avoid some of these problems, Tiano et al.examined carbonatogenesis using organic matrix macro-molecules extracted from seashells, a procedure thatunfortunately proved complex and produced a low yieldof usable product [62]. Further work compared the effectsof Mytilus californianus with Ca(OH)2 and CaCl2 in termsof porosity, capillary water absorption and superficialcohesion on Pietra di Lecce and Pietra d’Angera (bioclasticlimestone): M. californianus gave the best results [63]. In1999, Tiano et al. examined methods of evaluatingbiologically mediated precipitation of calcite using poredimension, stone strength and colour [64]. Using Pietra diLecce with Micrococcus sp. and Bacillus subtilis, calcitewas identified using X-ray diffraction (XRD) and Fouriertransform–infrared spectroscopy (FT-IR) He found therehad been a decrease in stone porosity but considered thathalf of this was probably due to physical obstruction ofpores rather than newly precipitated calcite. A side effectof the treatment was the formation of stained patches dueto the growth of airborne contaminants exploiting thepresence of organic nutrients in the media used to growthe biocalcifying bacteria. Concerns regarding contami-nation of stone during induced biomineralization led toattempts to define the genetic mechanism as a possiblealternative to the direct application of live cells [65].

It is now understood that application of a physicalbarrier on the stone surface will hinder the movement ofsalts, which can then build up, leading initially tounsightly discolouration and ultimately to physicaldamage [66]. Even coatings that permit evaporation cancause problems of salt accumulation and crystallizationand, therefore, a protective coating must be sympatheticto the nature of the stone itself. The production of

Review TRENDS in Biotechnology Vol.24 No.6 June 2006258

a calcium carbonate layer generated by bacteria (Figure 2)might offer a solution to this dilemma because the layerwould not block the natural pore structure, thus permit-ting free passage of soluble salts through the stone. Thelife of sacrificial layers generated by microorganismsis unclear but because of accelerated aging tests, LeMetayer-Levrel suggests that protection acquired bybiomineralization tends to increase with age by offeringlonger-term resistance to the weathering caused bygrowth of acid-producing microbial populations [55].

Current work and future prospects

Bioremediation is less harsh than the use of environmen-tally toxic chemicals or aggressive mechanical procedures,which are considered to be destructive methods. Crustremoval by microorganisms takes place in a natural waybecause these microorganisms have an active role in theenvironment, where they contribute to the closure ofbiogeochemical cycles or stabilization of dynamic equili-bria. However, research issues that have and will continueto be addressed are the effectiveness of these procedures,in addition to appropriate risk analyses, to provideconservators with confidence in the technology. Thus,technology transfer from the scientific community canonly be achieved by direct engagement of heritage end-users across national boundaries.

Recent research has explored the factors that influencebiological precipitation of calcium carbonate. Studies inSpain, France, USA, Italy and the UK have shown thatconsolidation of stones by biocalcifying bacteria can becontrolled by application of safe bacteria that pose littlethreat to the heritage object [50,56,57,65,67]. Acceptanceand satisfactory use of these technologies in conservationpractice requires adequate knowledge of the risk factorsto the heritage object in addition to conservators handlingthe active components. There have been concerns aboutthe long-term effects of the applied bacteria and theirnutrient media. Such issues are best addressed insystematic studies involving collaboration among scien-tists and end-users active in conservation practice.Practical application of bioremedial technologies hasrecently progressed, through the work of two projectsfunded by the European Community, which also usefullyillustrate the differences of approach that have beenadopted to evaluate the potential of bioremediation forconservation work: BIOREINFORCE (http://www.ub.es/rpat/bioreinforce/bioreinforce.htm); and BIOBRUSH(http://www.biobrush.org). The BIOREINFORCE partner-ship has successfully demonstrated that dead cells fromactive biocalcifying strains showed a much higher and/orfaster production of CaCO3 crystals than less activestrains. By elucidating the genetic expression of crystalformation in bacteria, the project aimed to produce bio-derived, low-cost, renewable macromolecules that willinduce calcification on stones without using livingbacterial cells [68]. The novel approach of the BIOBRUSHproject was to use live but low-hazard bacteria to linkthe mineralization processes (that remove salt crusts)to biomineralization (that can consolidate the stone).Bacteria were applied directly to stone surfaces usingtechniques that minimized the risk to the environment

www.sciencedirect.com

and conservators applying the treatment. Work at theUniversities of Milan and Molise, respectively, havedemonstrated that multiple short-term applications ofaerotolerant sulfate-reducing bacteria, within an appro-priate delivery system, can be successful in removingblack crusts from marble, both in the laboratory andin situ on buildings [69]. Results suggest that thebioremediation is at least as good as existing methods.Work with biocalcifying bacteria that were isolated fromthe environment has shown that deposition of a calcitelayer can be achieved without significant reduction inporosity or growth of contaminating microorganisms [67].This approach uses environmental isolates and controlledrelease of nutrients to minimize an adverse stimulation ofmicrobial growth.

Concluding remarks

The controlled use of microorganisms as agents ofbioremediation offers new approaches for conservators tohelp preserve, protect and restore building stone. Suchtechniques are intended to supplement rather thanreplace existing conservation technologies, which canoften be ineffective or toxic to end-users or the environ-ment. In the past 10 years, research has explored theconstraints of applying bacteria to stone to remove saltcrusts and consolidate the damaged pore structure.Suitable organisms are now known, and the environmen-tal factors have been identified; however, the risks posedby aesthetic and mineral changes are still beingaddressed. The challenge for the immediate future is totranslate a wide range of promising results into thepractical technology that has been achieved in other fieldsof biotechnology. Although the technology is still in itsinfancy and, therefore, not readily available, the results sofar indicate that it promises to offer a viable alternative tothose working to preserve our cultural heritage.

References

1 Atlas, R.M. (1995) Bioremediation. Chem. Eng. News 73, 32–422 Jordening, H-J. and Winter, J. (2005) Environmental Biotechnology:

Concepts and Applications, Wiley-VCH, Weinheim3 Gauri, K.L. and Bandyopadhyay, J.K. (1999) Carbonate Stone:

Chemical Behaviour, Durability, and Conservation, John Wiley &Sons, New York

4 Leysen, L. et al. (1987) A study of the weathering of an historicbuilding. Anal. Chim. Acta 195, 247–255

5 Doornkamp, J.C. and Ibrahim, H.A.M. (1990) Salt weathering. Prog.Phys. Geogr. 14, 335–348

6 Warscheid, T. and Braams, J. (2000) Biodeterioration of stone: areview. International Biodeterioration & Biodegradation 46, 343–368

7 Gaylarde, C. et al. (2003) Microbial impact on building materials: anoverview. Materials and Structures 36, 342–352

8 McNamara, C.J. and Mitchell, R. (2005) Microbial deterioration ofhistoric stone. Frontiers in Ecology and the Environment 3, 445–451

9 Rodriguez-Navarro, C. et al. (2000) How does sodium sulfate crystal-lize? Implications for the decay and testing of building materials.Cement Concrete Res. 30, 1527–1534

10 Rodriguez-Navarro, C. et al. (2002) Effects of ferrocyanide ions onNaCl crystallization in porous stone. J. Cryst. Growth 243, 503–516

11 Papida, S. et al. (2000) Enhancement of physical weathering ofbuilding stones by microbial populations. Int. Biodeterior. Biodegrada-tion 46, 305–317

12 Lewin, S. (1989) The susceptibility of calcareous stones to salt decay.In The Conservation of Monuments in the Mediterranean Basin(Zezza, F., ed.), pp. 59–64, Grafo, Brescia

Review TRENDS in Biotechnology Vol.24 No.6 June 2006 259

13 Lopez-Acevedo, V. et al. (1997) Salt crystallization in porousconstruction materials. II. Mass transport and crystallizationprocesses. J. Cryst. Growth 182, 103–110

14 Heselmeyer, K. et al. (1991) Application of Desulfovibrio vulgaris forthe bioconservation of rock gypsum crusts into calcite. BIOforum 1, 89

15 Ranalli, G. et al. (2000) Bioremediation of cultural heritage: removal ofsulphates, nitrates, and organic substances. In Of Microbes and Art -The Role of Microbial Communities in the Degradation and Protectionof Cultural Heritage (Ciferri, O., Tiano, P. and Mastromei, G., eds), pp.231–245, Kluwer Academic-Plenum Publisher

16 Dornieden, T.H. et al. (2000) Patina: physical and chemical inter-actions of sub-aerial biofilms with objects of art. In Of Microbes andArt - The Role of Microbial Communities in the Degradation andProtection of Cultural Heritage (Ciferri, O., Tiano, P. andMastromei, G., eds), pp. 105–119, Kluwer Academic-PlenumPublisher

17 Gaylarde, P.M. et al. (2001) Biodeterioration of Mayan buildings atUxmal and Tulum, Mexico. Biofouling 17, 41–46

18 Ghedini, N. et al. (2000) Determination of elemental and organiccarbon on damaged stone monuments. Atmos. Environ. 34, 4383–4391

19 Hakkarinen, C. (1975) Fate of Atmospheric Sulphur and Nitrogenfrom Fossil Fuels. Am. Chem. Soc. Div. Fuel Chem. Prepr. 20, 109–115

20 Foyn, E. (1977) Problems of fossil fuel combustion and waste.Environmental Pollution Management 7, 48–49

21 Ashurst, J. and Dimes, F.G. (1990) Conservation of Building andDecorative Stone (Vols I and II), Butterworth

22 Gauri, K.L. et al. (1989) The sulphatation of marble and treatment ofgypsum crusts. Studies in Conservation 34, 201–206

23 Torok, A. (2003) Surface strength and mineralogy of weatheringcrusts on limestone buildings in Budapest Building and Environment38, 1185–1192

24 Primerano, P. et al. (2000) Possible alterations of monuments causedby particles emitted into the atmospheres carrying strong primaryacidity. Atmos. Environ. 34, 3889–3896

25 Gavino, M. et al. (2004) Composition of the black crusts from the SaintDenis Basilica, France, as revealed by gas chromatography–massspectrometry. Journal of Separation Science 27, 513–523

26 Sabbioni, C. et al. (2003) Organic anions in damage layers onmonuments and buildings. Atmos. Environ. 37, 1261–1269

27 Schiavon, N. et al. (2004) Soiling of limestone in an urbanenvironment characterized by heavy vehicular exhaust emissions.Environ. Geol. 46, 448–455

28 Webster, R.G.M. (1992) Stonecleaning and the nature, soiling anddecay mechanisms of stone. In Proceedings of the InternationalConference, Donhead

29 Maxwell, I. (1992) Stonecleaning – for better or worse? An overview. InStonecleaning and the Nature, Soiling and Decay Mechanisms ofStone (Webster, R.G.M., ed.), pp. 3–49, Donhead

30 MacDonald, J. et al. (1992) Chemical cleaning of sandstone:comparative laboratory studies. In Stonecleaning and the Nature,Soiling and Decay Mechanisms of Stone (Webster, R.G.M., ed.), pp.217–226, Donhead

31 Maravelaki-Kalaitzaki, P. (2005) Black crusts and patinas on Pentelicmarble from the Parthenon and Erechtheum (Acropolis, Athens):characterization and origin. Anal. Chim. Acta 532, 187–198

32 Gauri, K.L. and Bandyopadhyay, J.K. (1999) Carbonate Stone,Chemical Behaviour, Durability, and Conservation, John Wiley &Sons

33 Skoulikidis, T.N. and Beloyannis, N. (1984) Inversion of marblesulfation: reconversion of gypsum films into calcite on the surfaces ofmonuments and statues. Studies in Conservation 29, 197–204

34 Moncrieff, A. and Hempel, K. (1970) “Biological Pack”. Conservation ofStone and Wooden Objects 1, 103–114, ICC

35 Atlas, R.M. et al. (1988) Microbial calcification of gypsum-rock andsulfated marble. Studies in Conservation 33, 149–153

36 Gauri, K.L. and Chowdhury, A.N. (1988) Experimental studies onconversion of gypsum to calcite by microbes. In Proceedings of 6thInternational Congress on Deterioration and Conservation of Stone(Ciabach, J., ed.), pp. 545–550, Nicholas Copernicus University

37 Gauri, K.L. et al. (1992) Removal of sulfated-crust from marble usingsulfate reducing bacteria. In Stonecleaning and the Nature, Soilingand Decay Mechanisms of Stone (Webster, R.G.M., ed.), pp. 160–165,Donhead

www.sciencedirect.com

38 Kouzeli, K. (1992) Black crust removal methods in use: their effects onPentelic marble surfaces. In Proceedings of 7th International Congresson Deterioration and Conservation of Stone (Vol. 2), pp. 1147–1156

39 Saiz-Jimenez, C. (1997) Biodeterioration vs biodegradation: the role ofmicroorganisms in the removal of pollutants deposited on historicbuildings. Int. Biodeterior. Biodegradation 40, 225–232

40 Ranalli, G. et al. (1997) The use of microorganisms for the removal ofsulphates on artistic stoneworks. Int. Biodeterior. Biodegradation 40,255–261

41 Ranalli, G. et al. (2005) Biotechnology applied to cultural heritage:biorestoration of frescoes using viable bacterial cells and enzymes.J. Appl. Microbiol. 98, 73–83

42 Martin-Gil, J. et al. (1999) Ancient pastes for stone protection againstenvironmental agents. Studies in Conservation 44, 58–62

43 Brajer, I. and Kalsbeek, N. (1999) Limewater absorption and calcitecrystal formation on a limewater-impregnated secco wall painting.Studies in Conservation 44, 145–156

44 Cezar, T.M. (1998) Calcium oxalate: a surface treatment for limestone.Journal of Conservation and Museum Studies 4 (http://palimpsest.stanford.edu/jcms/issue4/cezar.html)

45 Garcia-Valles, M. et al. (1997) Coloured mineral coatings onmonument surfaces as a result of biomineralisation: the case ofTarragona Cathedral (Catalonia). Appl. Geochem. 12, 255–266

46 Alvarez de Buergo, M. and Fort Gonzalez, R. (2003) Protective patinason stony facades of historical buildings in the past. Construction and

Building Materials 17, 83–8947 Milet, C. et al. (2004) Conservation of signal molecules involved in

biomineralisation control in calcifying matrices of bone and shell.Comptes rendus Palevol 3, 493–501

48 Boquet, E. et al. (1973) Production of calcite (calcium carbonate)crystals by soil bacteria is a general phenomenon. Nature 246,527–529

49 Ehrlich, H.L. (1999) Microbes as geologic agents: their role in mineralformation. Geomicrobiol. J. 16, 135–153

50 Rivadeneyra, M.A. et al. (2000) Precipitation of carbonates byNesterenkonia halobia in liquid media. Chemosphere 41, 617–624

51 Duane, M.J. (2001) Biomineralisation and phytokarst development oncavernous quaternary carbonate terraces, Mohammedia, NorthwestMorocco. Carbonates and Evaporates 16, 107–116

52 Schiavon, N. (2002) Biodeterioration of calcareous and graniticbuilding stones in urban environments. In Natural Stone, WeatheringPhenomena, Conservation Strategies and Case Studies(Siegesmund, S., Vollbrecht, A. and Weiss, T., eds), pp. 195–205,GSL Special Publication

53 Urzi, C. et al. (1999) Biomineralization processes of the rock surfacesobserved in field and in laboratory. Geomicrobiol. J. 16, 39–54

54 Orial, G. et al. (1992) The biomineralisation: a new process to protectcalcareous stone: applied to historic monuments. In Proceedings of2nd International Conference on Biodeterioration of Cultural Heritageproperty (Arai, H., Kenjo, T. and Yamano, K., eds), pp. 98–116,International Communications Specialists

55 Orial, G. and Marie-Victoire, E. (1997) Fabrication de carbonate decalcium par voie biologique: la carbonatogenese. In Proceedings ofInternational Seminar on Deterioration of Concrete andNatural Stoneof Historical Monuments, pp. 59–76

56 Le Metayer-Levrel, G. et al. (1999) Applications of bacterialcarbonatogenesis to the protection and regeneration of limestones inbuildings and historic patrimony. Sediment. Geol. 126, 25–34

57 Bang, S.S. et al. (2001) Calcite precipitation induced by poly-urethane-immobilized Bacillus pasterii. Enzyme Microb. Technol.28, 404–409

58 Ramachandran, S.K. et al. (2001) Remediation of concrete usingmicro-organisms. ACI Materials Journal 98, 3–9

59 Rodriguez-Navarro, C. et al. (2003) Conservation of ornamental stoneby Myxococcus xanthus – induced carbonate biomineralization. Appl.Environ. Microbiol. 69, 2182–2193

60 Morse, J.W. (1983) Kinetics of calcium carbonate dissolution andprecipitation. In Carbonates, Mineralogy and Chemistry (Reeder, R.J.,ed.), pp. 227–264, Mineralogical Society of America, BookCrafters

61 Castanier, S. et al. (2000) Bacterial carbonatogenesis and applicationto preservation and restoration of historic property. In Of Microbesand Art – The Role of Microbial Communities in the Degradation and

Review TRENDS in Biotechnology Vol.24 No.6 June 2006260

Protection of Cultural Heritage (Ciferri, O., Tiano, P. andMastromei, G., eds), pp. 203–218, Kluwer Academic-PlenumPublisher

62 Tiano, P. et al. (1992) Stone reinforcement by induction of calcitecrystals using organic matrix molecules, feasibility study. InProceedings of 7th International Congress on Deterioration andConservation of Stone (Vol. 2), pp. 1317–1326

63 Tiano, P. (1995) Stone reinforcement by calcite crystals precipitationinduced by organic matrix macromolecules. Studies in Conservation40, 171–176

64 Tiano, P. et al. (1999) Bacterial bio-mediated calcite precipitation formonumental stones conservation: methods of evaluation. J. Microbiol.Methods 36, 139–145

65 Perito, B. et al. (2000) Bacterial genes involved in calcite crystalprecipitation. In Of Microbes and Art – The Role of Microbial

Endea

The quarterly magaziand philosophy

You can access EndeScienceDirect, whbook reviews, editand a collection

illustrated artihistory of s

Featur

Waxworks and the performance of anatomy inRepresenting revolution: icons o

Myths about moths: a study inThe origins of research into tIn search of the sea monste

Michael Faraday, med

Coming

The Livingstone story and the IndIntertwined legacies: Pierre CurieProvincial geology and the Indus

The history of computer climaThe history of NASA’s exobio

And much, mu

Endeavour is available on Science

www.sciencedirect.com

Communities in the Degradation and Protection of Cultural Heritage(Ciferri, O., Tiano, P. and Mastromei, G., eds), pp. 217–228, KluwerAcademic-Plenum Publisher

66 Doornkamp, J.C. and Ibrahim, H.A.M. (1990) Salt weathering. Prog.Phys. Geogr. 14, 335–348

67 Webster, A.M. et al. (2004) Bacteria and the bioremediation of stone:the potential for saving cultural heritage. In Proceedings of theEuropean Symposium on Environmental Biotechnology(Verstraete, W., ed.), pp. 793–796, Taylor and Francis Group

68 Barabesi, C. et al. (2003) Microbial calcium carbonate precipitation forreinforcement of monumental stones. In Molecular Biology andCultural Heritage (Saiz-Jimenez, C., ed.), pp. 209–212, Balkema

69 Cappitelli, F. et al. Improved methodology for bioremoval of blackcrusts on historical stone artworks by use of sulphate–reducingbacteria. Appl. Environ. Microbiol. (in press)

vour

ne for the historyof science

avour online onere you’ll findorial commentof beautifullycles on thecience.

ing

mid-eighteenth-century Italy by L. Dacomef industrialization by P. Faracontrasts by D.W. Rudge

he origins of life by I. Fryr by K. Hvidtfelt Nielsenia man by P. Fara

soon

ustrial Revolution by L. Dritsasand radium by D.H. Rouvraytrial Revolution by L. Veneerte modeling by M. Greenelogy program by S.J. Dick

ch more . . .

Direct www.sciencedirect.com