Electromagnetic Field as Agent Moving Bioactive Cations. A New Antimicrobial System in Architecture Technology

Featured Application

Paint with Zn and Cu (patent Galvi PL229012) can be used inside damp lodgings (butchers, winery, bathrooms, cellars, storehouses, museums etc.) as well as elevations of buildings. Using paint with Zn, Bi, Cu (three electrode systems) for covering the lower side of the stretchers is suggested. This system is the source of bioactive “ions cocktail” and it works only at a very fine level on the lower surface of the stretcher. According to a pessimistic prediction, building material in architecture could be largely destroyed at the end of XXI century. Our last investigations can help in this matter. Two-phase media with SNA (synthetic low-nutrient agar) medium + sterile grandiorite, melabasanite, sandstone, marl and sodium glass (as control) were used. This enables the investigation of the processes of colonization, course, and development of selected microorganisms in the emerging rock microhabitats (patent pending). For all analyzed rocks, their color may also be important. A dark or even black rock (i.e., melabasanite) exposed to solar radiation heats up much more than a light or even white one. In this particular case, granodiorite is light gray, sandstone is beige, and marl is pale beige.

Abstract

There is a new described antifungal system (GALVI) involving the moving of bioactive ions of Zn, Cu and Bi for the protection of cultural heritage objects such as buildings, sculptures and stretchers. There were two kinds of galvanic cells that were used: the first composed of a two-electrodes system, Zn, Cu, and second one composed of a three-electrodes system, Zn, Bi and Cu. Moreover, two-phase media are proposed with various kinds of rocks used in architectonical objects. Microorganisms inhabit the boundaries of two liquid and solid phases. This enables the investigation of the process of rock colonization. Possible applications of the suggested GALVI system are mentioned.

1. Introduction

Cultural heritage contains not only buildings but also mines, caves, graves, old canals, bridges, ships, boats, historical glass, ceramics, sculptures, and artistic painting. Historical architectural objects are mostly made of stone and wood and are rarely made out of glass, metals and ceramics. We will try to describe methods of building protection against fungi. Various microorganisms are involved in the biodeterioration of the architecture objects. These are algae, lichens, fungi, and bacteria. Damp buildings are mostly colonized by hydrophilic fungi [1].

In architectural objects, various kinds of rocks such as granite, basalt, sandstones, and calcareous stones were used. Among the various rock types, calcareous stones are the most widely used, while other kinds are less common in architecture technology [2]. Embry & Klovan [3] and more recently Flügel [4] presented a classification of carbonate rocks (limestones, dolomites, marls, etc.). Siegesmund and Török [2] put these rocks in chemically and biologically precipitated rocks. Gadd [5] pointed out that microcolonial fungi can significantly change a rock’s surficial appearance, such as discoloration, staining, biodeterioration and the formation of new biogenic minerals. Calcareous stones are often investigated by mycologists [6,7,8,9,10,11,12,13,14,15]. Some publications are devoted to other stones: basalt [16], granite [17,18], sandstone [19,20] and minerals such as löllingite FeAs₂ and arsenopyrite FeAsS [21,22]. Brandl [23] cited 82 articles that were devoted to bioleaching of metals from mineral resources (albite, amphibolite, andesite, apophylite, argentite, armtone, augite, basalt, bauxite, biotite, calamine, calcium, carbonaceous ore, chalcocite, chrysocolla, chrysolite, copper molybdenum ore, cobalt, cuprite, diorite, dunite, dysprosium, ferromanganese sea nodules, galena, garnierite, genthite, geothite, glauconite, gold bearing ore, gypsum, grandiorite, granite, halloysite, harmtone, hematites, hemetitic lateritic ore, heulandite, illite, kaolonite, labradorite, lateritic nickel ore, leucite, limescale, limonite, lydite, malachite, manganese, manganese ore, microcline, montmorillonmite, muscovite, natrolite, nepheline, neodymium, olivine, orthoclase, pearceite, pegmatite, peridoptite, phlogopite, phosphates of aluminium, polybasite, piritic ore, proutite, pyromorphite, quartz sand, rhodochrosite, rhyolite, samarium, sandstone, saponite, scheelite, serpentine, silicate nickel ore, spodumene, stilbite, vermiculite, wollastonite, yttrium, zinc oxide, zinc). Among stone colonizing fungi we can find mostly anamorphic states of ascomycete and basidiomycete fungi, and Zygomycetes rarely occurs. Coincidentally, teleomorphs of Basidiomycetes occur [24]. There are different names for rock-inhabiting fungi (lithobionts), such as microcolonial fungi = MCF [5,11], rock-eating fungi = REF [25], rock-inhabiting fungi = RIF [14], and rock-building fungi = RBF [26]. Most of them belong to black yeasts and their relatives [27,28]. Black yeasts are mostly extremotolerant anamorphs that lost their teleomorphic stadia. These RIF microorganisms are present both in hot deserts and cold Arctic and Antarctic places. Stone deterioration in the presence of microorganisms is approximately ten thousand times faster than without their presence [29]. Cyanobacteria creating biofilms are often reported from diverse architectural buildings (

Table 1. The most frequent strains of Cyanobacteria identified in diverse architectural buildings.

Name of Species	Citation	Kind of Substratum	Climate
Chroococcus sp. Chlorococcus sp.	[30,31]	historic church in Porto Alegre, RS, Brazil Europe, UK. Historical building Windermere	Tropical Moderate
Fischerella sp., Gloeocapsa,	[32]	cyanobacteria from external stone walls, Mexico	Equatorial and subequatorial zone
Leptolyngbya gracillima	[33]	cooling tower of the power plant at Bełchatów, Poland	Moderata
Scytonema myochrous	[33]	cooling tower of the power plant at Bełchatów, Poland;	Moderate
Scytonema sp.	[34]	limestone walls at other low-pollution sites, the Great Jaguar Pyramid at Tikal, Guatemala	Equatorial and subequatorial zone
Gloeothece rupestris	[33]	wall of the cooling tower of the power plant et Bełchatow	Moderate
Nostoc sp., Gloeothece rupestris (Lyngbye)	[33]	wall of the cooling tower of the power plant at Bełchatów	Moderate
Trentepohlia sp.	[32]	stone altar in the architectural zone of Becan, Campeche state, Mexico	Tropical
Gloecapsa sp. Gloeothece, Aphanocapsa, and Chroococcus, and filamentous species like Scytonema and Tolypothrix.	[35]	on urban building surfaces	Tropics

).

Fungi are extremely corrosive endoliths [27,29,36,37]. They can produce acids (Figure 1) that can dissolve minerals. Fungi can cause the biodeterioration of stone elevations, surfaces, various kinds of monuments, buildings, sculptures, and other objects made of stone as well as wooden elements, artistic paintings, and even concrete walls [38]. There are a huge number of publications concerning the fungal deterioration of rock in architecture objects. Nielsen [39] noted 746 such publications in his own reference list. There are only some important articles [7,13,19,20,22,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55]. Recently Adams et al. [1] noted 111 species of fungi in damp buildings.

Table 2. The most frequent strains of fungi identified in diverse architectural buildings, with published references, focusing on the diverse climate of Earth. The most common fungal species are highlighted in italics.

Name of Fungi	Citations	Kind of Substratum	Climate
Alternaria	very common	gypsum, tombstone, damp buildings	widely distributed
Aspergillus	very common	walls	widely distributed
Aureobasidium	common, [15,56]	marble, damp buildings	widely distributed
Baudoinia	[50,57]	buildings, cars	temperate
Beauveria	[58]	biofilm in Roman catacombs	mediterranean
Botryomyces	[8]	stone monuments	mediterranean
Chaetomium	[1,59]	damp buildings	temperate
Cladosporium	very common	damp buildings, tombstone	widely distributed
Constantinomyces	[15]	tombstone	temperate
Epicoccum	[1]	damp buildings	widely distributed
Exophiala	[15,60]	gold mine, tombstones	temperate
Geomyces	[21]	gold mine	temperate
Hortea	[29]	marble, monuments Delos island	mediterranean
Knufia	[15,56]	marble, monuments, tombstone	temperate, mediterranean
Lecanicillium	[58]	biofilm in Roman catacombs	mediterranean
Neocatenulostroma	[15]	tombstone	temperate
Neodevresia	[15]	tombstone	temperate
Penicillium	very common	damp buildings	widely distributed
Phoma	very common	damp buildings	widely distributed
Rhizopus stolonifer	Ref. [1], Chlebicki personal inf.	damp wall after flood, packing houses	widely distributed
Rhodotorula	[15,39]	paints, tombstone	widely distributed
Sarcinomyces	[8]	marble	mediterrenean
Stachybotrys	very common	gypsum	temperate
Torrubiella	[58]	biofilm in Roman catacombs	mediterranean
Trichoderma	very common	damp buildings, tombstone	widely distributed
Trimmatostroma	[29]	rock	temperate
Zasmidium	[61,62]	vine cellar wall	temperate

presents only the most common (according to Nielsen, [39] and phenomenal fungi noted in buildings, sculptures, tombs, and mines.

2. Biofilms

Natural biofilms are composed of many various species, crustose epilitic lichens, algae, cyanobacteria. Fungi are also present in biofilms as propagules [21], but they are not forms of biofilms. However, Aureobasidium pullulans sensu lato forms biofilm with extracellular polymers (EPS) composed of pullulan [63]. Biofilms are composed from bacterial cells embedded in an organic polymer matrix [64], known as extracellular polymers EPS [65]. The chemical composition of EPS is extremely diverse [65]. Moreover, natural biofilms can be composed of some layers with algae, lichens, bacteria and fungi. The bacteriological investigation of rock biofilm in Gertruda adit [66,67] showed that the Gram and DAPI staining methods showed all the bacterial shapes known in literature, whereas 16S rRNA gene amplification from total DNA showed more than one hundred OTUs, dominated by α-Proteobacteria [68]. Only seven species of fungi were isolated from this biofilm [10]. Among Antarctic sponge-associated bacteria, the Roseovarius genus plays a pivotal role in biofilm production [69], enabling adhesion and ecological competition with other microorganisms [70]. Artificial biofilms can be composed of some species of bacteria, algae and black yeasts [71]. The vertical distribution of these microorganisms was different: algae and black yeasts were more abundant in the outer layer, while in the inner layer only few algae were observed. However, it appeared that the inner layer harbored a higher amount of microorganisms than the outer one. Miao et al. [72] observed more complex bacterial networks on artificial substrates than on natural substrates. However, the keystone species on natural substrates were more abundant. Among Antarctic sponge-associated bacteria, the Roseovarius genus plays a pivotal role in biofilm production, enabling adhesion and ecological competition with other microorganisms.

3. GALVI Technology

The rapid development of drug resistance in pathogenic fungi forced us to develop a new antifungal technology [73]. Snethlage [74] described antimicrobial drugs that we can use at the present time in architectural technology, as well as the stone conservation of historic and artistic objects. Among these drugs, conservative materials such as water repellents, biocides, and consolidants are mentioned. Then, chemicals such as water glass, ethyl silicates, fluorosilicates, epoxy resins, acrylic resins, and silicon organic compounds are presented. Moreover, Castanier et al. [44] described a biomineralization process with a suitable bacterial suspension culture that creates a coating scale, the “biocalcin”, whereas Koo et al. [75] noted a wide variety of biofilm-targeting strategies, among them a surface charge, roughness and topography, hydrophobicity, as well as using inorganic metallic nanoparticles of Ag and Cu and organic nanoparticles (inhibitors such as liposomes and aptamers), and biofilm removal by using mechanical, energy- and light disruption (ultrasound, acoustic, electric waves, laser shockwaves). This can protect stone objects from deterioration. The next methods, new antifungal discovery—system RNA interference or the exogenous application of synthetic RNA, are in preparation [73]. Recently, De Leo et al. [71] suggested the application of surface active ionic liquids (SAILs), based on cholinium cations and dodecylbenzenesulfonate, as anions in the protection of cultural heritage.

The toxicity of metal ions (mostly silver and copper) is well documented, and results have been presented in many papers [46,76,77,78]. Zinc has been shown to inhibit the growth and respiration of fungi [79,80], as well as the germination of fungal spores [81]. The action of silver is proportional to the bioactive released silver ion (Ag⁺). Silver interacts with bacterial or fungal cell membranes [46]. We proposed using a new antimicrobial technology—an electromagnetic field created by galvanic cells that influences the moving of metal ions [82,83]. This method was named GALVI system. We used two kinds of galvanic cells: the first composed of a two-electrodes system, Zn, Cu [82], and the second one composed of a three-electrodes system, Zn, Bi and Cu [83] (Figure 2). The three-electrodes system has a stronger antibacterial activity than the first one (Figure 2). Unfortunately, this system is much more expensive because the use of expensive rheological additives securing the fall of bismuth is necessary.

The inhibition zones produced by the first system were compared with Cassini ovals [82]. However, the last investigation showed an additional agent that also inhibited the growth of fungi (Figure 3). Using a pH indicator (dibromothymolsulphonaphtalein), we can observe changes in the pH distribution and the influence of pH on fungal growth. H⁺ distributed in half of the Petri plate inhibited fungal growth similarly to ions of Zn²⁺.

Inhibition zones are larger when this system works in an environment with a higher salinity (Figure 4). Moreover, in such an environment, Cu electrodes work more effectively than those made of Zn (Figure 4). Babich & Stotzky [84] noted that the anionic composition of the environment influenced the toxicity of heavy metals. In particular, high-concentration NaCl can provoke the formation of complex Zn-Cl species (ZnCl⁺, ZnCl₂, ZnCl₃⁻, ZnCl₄²⁻) with increased toxicity.

Some metals produce ions that inhibit or kill bacteria and other microorganisms. This is known as the oligodynamic effect, discovered by von Nageli in 1893 [85]. The galvanic system (which is used by us) enables one to move metal ions over a much longer distance and creates a large, inhibited zone. Nine different metals for the experiments were used. Of them, Zn and Cu created the biggest inhibition zone (Figure 5). The classical galvanic system of Cu Zn is known very well, but the optimal effect of the electromagnetic field depends on the distance between electrodes. Thirty-two distances between these electrodes were investigated. Very common indoor fungi such as Aureboasidium pullulans, Alternaria atra, Cladosporium cladosporioides, and Rhodotorula mucilaginosa were used in the experiments as indicators of the size of the inhibition zone. It appears that a distance of 4.2 mm creates the biggest inhibition zone [82].

In the case of the three-electrode systems, their configurations (Figure 6) have fundamental implications for obtaining an optimal inhibition zone [83].

4. Two-Phase Media

The use of GALVI technology in natural building stones requires knowledge of the stone composition and its deterioration by microorganisms. Earlier microbiologists isolated microorganisms and cultivated them on artificial one-phase media afterwards [13,25,37,86]. Mirocha & DeVay [87] observed the autotrophical growth of Cephalosporium sp. and Fusarium sp. on a medium devoid of any known organic carbon source. Recently started experiments with media would allow us to observe the development of fungi in a two-phase environment. These two-phase media were used with SNA (synthetic low-nutrient agar) medium + sterile grandiorite¹, melabasanite², sandstone³, marl⁴, and sodium glass (as control). Microorganisms inhabit the boundaries of two liquid and solid phases. This enables the investigation of the processes of colonization, course, and development of selected microorganisms in the emerging rock microhabitats (Figure 7). Two-phase media with granodiorite and sandstone were much more colonized by fungi than media with melabasanite and marl. Both marl and melabasanite have a lower porosity and higher content of Ca; moreover, melabasanite is dark, has a low water absorption, high pH (45% SiO₂), and contains toxic Fe (see Figure 5). In addition, melabasanite can be a good habitat for special microorganisms that, in addition to Ca, still require Mg, Fe, and/or Al [88,89,90].

The content of elements in the mentioned rocks is very rich, and possible interactions between cations and anions during changing pH are difficult to interpret. For example, a ferric hydroxide can be created at pH > 4.5, whereas poor coagulation occurs in the pH range between 7 and 8.5 [91]. The influence of environs on cations’ toxicity was mentioned earlier by Babich & Stotzky [85]. Metal transformations were thoroughly described by Gadd [92].

1. Granodiorite is a light gray igneous acidic rock, the main components of which are quartz (SiO₂), potassium feldspar (KAlSi₃O₈), sodium plagioclase (Na[AlSi₃O₈]), and biotite (K(Mg,Fe₂)₃(OH,F)₂AlSi₃O₁₀). The average analysis of the chemical composition shows the dominance of silica (71–74%), followed by large amounts of aluminium (12–16%) and smaller but significant amounts of sodium (3–4%) and potassium (2–3%).

2. Melabasanite (with a transition to ankaratrite) is a dark volcanic rock belonging to ultrabasic rocks (<45% SiO₂). The main minerals are pyroxenes (Ca silicates, Mg and Fe silicates), olivines (Mg,Fe)₂[SiO₄], amphiboles (hydrated Ca-silicates, Mg, Fe, and Na), and calcium plagioclases Ca[Al₂Si₂O₈]. In a variety closer to ankaratrite (a type of nephfinite), one of the main minerals is nepheline (sodium and potassium aluminosilicate (KNa₃(AlSiO₄)₄). The average chemical composition analysis shows, apart from silica (40–42%), large amounts of aluminium (13–14%), iron (approx. 12%), magnesium (approx. 12%), calcium (10–12%), and significant amounts of sodium (3–4%).

3. Sandstone is a coarse-grained beige clastic rock in which the spaces between quartz grains (SiO₂) are filled with silica-iron-clay binder (SiO₂/Fe₂O₃/hydrated aluminosilicates). Its technical parameters are variable, depending on the content of Fe₂O₃. Therefore, the main components of this sandstone are silica, aluminum, and iron.

4. Marl is a pale beige sedimentary rock with variable proportions of two main components: calcite (CaCO₃) or dolomite (CaMg(CO₃)₂) in the amount of 50–75% and clay minerals (hydrated aluminosilicates) in the amount of 25–50%. Marl may also contain 0–50% SiO₂. Its main components are therefore calcium, aluminium ± silica.

This was elaborated on the basis of the following books: [93,94,95].

Each of the analyzed types of rock, due to its chemical composition and color, is likely to be suitable for microorganisms requiring appropriate elements. Although marl and melabasanite abound in Ca, both may have the same calcium-demanding microorganism. For all the analyzed rocks, their color may also be important. A dark or even black rock, exposed to solar radiation, heats up much more than a light or even white one. For the samples analyzed, low-porous and low-absorbing melabasanite, which is rich in minerals Al, Fe, Mg, Ca and Na, will heat up more than slightly less porous and less absorbing red sandstone containing Si, Al, and Fe minerals. The lower temperature will reach a bright, low-porous, and very absorbing marl, built mostly of CaCo₃, while the relatively lowest one will reach also bright, but much less porous and less absorbing granodiorite, which in addition to SiO₂ contains minerals rich in Al, Na and K.

5. Possible Application of Galvanic Systems

Galvanic macrocells. Our first experiments were constructed with electrodes in the form of long bars of 2–3 mm-diameter and 5–10 cm length. This GALVI system can be used in wooden buildings against colonization by Serpula lacrymans, Gloeophyllum trabeum, and Schizophyllum commune.

Galvanic microcells. This next GALVI system was formed with electrodes that were in the form of metal grains of Zn and Cu deposited randomly on the surface. The zinc grains (anode) had diameters under 90 μm and copper grains (cathode) with a diameter under 63 μm [84]. Galvanic microcells were used in the paints production of GALVI technology in many variants:

1. Paint with Zn and Cu that can be used inside damp lodgings (butchers, winery, bathrooms, cellars, storehouses, museums etc.).

2. Paints with Zn and Cu that can be used outside in modern building elevations. This is a watertight system for use on facades that are at risk of fouling, with harmful microorganisms in objects with a higher exposure to biocorrosion. Grains of metals will be dipped in the matrix (Figure 8, Figure 9 and Figure 10), but their tops will be free and stand up. This is necessary for the function of the electromagnetic field when the paint surface will be in damp conditions.

3. We also suggest using paint with Zn, Bi, and Cu (three-electrode systems) on the lower side of the stretchers. This system is the source of bioactive “ions cocktail” [78], and it works only at a very fine level on the lower surface of the stretcher. Metal ions cannot come through the canvas and damage the upper part of the stretcher.

Metal grains should be added to the GALVI systems by the users directly before the application. Then, the paint must be mixed with a manual or mechanical stirrer to ensure an even distribution of electrodes in the coating. All GALVI systems do not work in very dry conditions, but in such conditions harmful biodeterioration fortunately does not occur. When a place with GALVI coating is damp, then the action of the electromagnetic field starts.

We are planning future experiments, forming a GALVI system with a transparent base composed of ethyl silicates, fluorosilicates, epoxy resins, or acrylic resins. It can be used in decorative stone walls or elevations, as well as in artistic concrete. It is possible that our methods could be used in antimicrobial technology in architecture and art. Microorganisms have never adapted to the high electromagnetic field’s strength, which contrasts with traditional biocides.