Next Article in Journal
The Characterization Analysis of the Oil-Immersed Transformers Obtained by Area Elimination Method Design
Next Article in Special Issue
The Role of Fungi in Biodeterioration of Cultural Heritage: New Insights for Their Control
Previous Article in Journal
Concept for Generating Energy Demand in Electric Vehicles with a Model Based Approach
Previous Article in Special Issue
Fungi Affecting Wall Paintings of Historical Value: A Worldwide Meta-Analysis of Their Detected Diversity
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Applied Sciences
Volume 12
Issue 8
10.3390/app12083969
applsci-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Review

Black Fungi on Stone-Built Heritage: Current Knowledge and Future Outlook

by
Filomena De Leo
*,
Alessia Marchetta
and
Clara Urzì
Department of Chemical, Biological, Pharmaceutical and Environmental Sciences, University of Messina, Viale F. Stagno d’Alcontres, 31, 98166 Messina, Italy
*
Author to whom correspondence should be addressed.
Appl. Sci. 2022, 12(8), 3969; https://doi.org/10.3390/app12083969
Submission received: 1 March 2022 / Revised: 6 April 2022 / Accepted: 11 April 2022 / Published: 14 April 2022
(This article belongs to the Special Issue The Role of Fungi in Biodeterioration of Cultural Heritage: New Insights for Their Control)
Browse Figures
Review Reports Versions Notes

Abstract

:

Featured Application

This is an updated review on black fungi as main biodeteriogens of cultural heritage stone artifacts. Colonization pattern, taxonomy, and methods to eradicate their settlement are discussed here.

Abstract

Black fungi are considered as one of the main group of microorganisms responsible for the biodeterioration of stone cultural heritage artifacts. In this paper, we provide a critical analysis and review of more than 30 years of studies on black fungi isolated from stone-built heritage from 1990 to date. More than 109 papers concerning the fungal biodeterioration activity of stone were analysed. The main findings were a check list of the black fungal taxa involved in the biodeterioration of stone-built heritage, with a particular reference to meristematic black fungi, the main biodeterioration pattern attributed to them, and the methods of study including the new molecular advances. A particular focus was to discuss the current approaches to control black fungi from stone-built heritage and future perspectives. Black fungi are notoriously hard to remove or mitigate, so new methods of study and of control are needed, but it is also important to combine classical methods with new approaches to improve current knowledge to implement future conservation strategies.

1. Background

Stonework, such as artistic sculptures, historical buildings, monuments, archaeological sites, caves, etc., are ubiquitous across the globe, being an expression of culture, religion, aesthetics, and building techniques of populations, typical of certain historical construction periods. Due to their unicity and intrinsic value, ensuring the integrity of stone-built heritage for posterity is a critical issue. The study of biodeterioration of cultural heritage is a hot topic of broad interest to the researcher’s community and the implementation of safeguard measures is one of the main goals. All materials are subjected to a natural weathering, and “biodeterioration” of stones should be considered as an integral part of bio-geo-morphogenesis [1,2,3,4]. The term “biodeterioration” defines any irreversible transformation of inorganic or organic material with economic, commercial, historic, and artistic loss caused by macro- and micro-organisms [5].
“Biodeterioration” is a very complex matter and conservators should also take into account whether the observed biologically driven phenomena can even be considered positive for the artifact.
In fact, in some cases, the presence of subaerial biofilm (SABs) may have a protective effect on the surface [6]; on the other hand, SABs developed at the interface between rock surface and air is considered the main cause of biodeterioration of stone monuments [7,8].
Microbial biodeterioration of stones is often associated with the presence of a complex community formed by chemoorganotrophic microorganisms (bacteria and microfungi) and autotrophic microorganisms (such as algae and cyanobacteria and to lesser extent autotrophic bacteria) usually embedded in an extracellular matrix EPS (in which are present DNA, enzymes, pigments, lipids, proteins, etc.). Microbial cells in the EPS show a typical biofilm lifestyle that confers resistance to hostile environments and reinforces the attachment of microorganisms on the surface [6,9].
The prevalence of one or more group of microorganisms depends on numerous factors which include the intrinsic characteristics of the material (such as lithotype, porosity, roughness, and state of preservation) that affect its “bioreceptivity” sensu Guillitte [10]. The species composition can vary greatly depending on climatic and microclimatic conditions such as temperature, solar irradiation, shining, nutrient and water availability, and, last but not least, the characteristics of species involved [6]. However, microbial colonization is a very dynamic process in time and space, that is the result of the interactions between microbial species and substrates. It varies continuously during the year following the seasons, and it is also under the influence of the dispersion ability of propagules in the air [11,12,13].
In recent years, much knowledge has been gained about rock-inhabiting black fungi, and important issues concerning their taxonomy, physiology, phylogeny, and weathering processes [14] have been clarified. However, the majority of studies concerned black fungi from natural environments [4,7,15].
In the field of cultural heritage, most reviews had as a topic the biodeterioration of stone caused by fungi in general [6,16,17]; some have focused on the microbial and fungal deterioration of various type of substrata (both organic and inorganic such as textile, parchment, wood, paper, metals, and stone) used for artworks [18]; few concerned exclusively black fungi as a cause of biodeterioration of stone monuments [19,20].
This paper aims to give an overview on the present knowledge of rock-inhabiting black fungi in the field of stone cultural heritage with reference to their taxonomy, biodeterioration pattern, methods of study, and control, with a look to a future perspectives.
A bibliographic search was carried out using such databases as Scopus (https://www.scopus.com accessed on 17 March 2022), Science Direct (https://www.sciencedirect.com accessed on 17 March 2022), Web of Science (http://www.webofknowledge.com accessed on 17 March 2022), and Google Scholar (https://scholar.google.com accessed on 17 March 2022), that were consulted by using keywords such as ‘black fungi’, ‘meristematic fungi’, ‘stone monuments’, ‘stone artworks’, ‘stone biodeterioration’, ‘biodeteriogenic fungi’, ‘fungal treatment’, and ‘fungal control’.
The search produced about 500 papers of which 109 were included in this paper. The updates of fungal nomenclature were searched in the databases Index Fungorum (http://www.indexfungorum.org accessed on 22 March 2022), National Center for Biotechnology Information (NCBI) (https://www.ncbi.nlm.nih.gov accessed on 24 February 2022) and in the recent literature [14,21].
Nucleotide sequences were retrieved from GenBank database that is accessible from NCBI platforms (http://www.ncbi.nlm.nih.gov accessed on 14 February 2022). Molecular Evolution Genetic Analyses (Mega 11 Software) free downloadable via the URL. http://www.megastoftware.net accessed on 14 February 2022 was employed for alignments and phylogenetic tree constructions.

2. Black Fungi and Stone Monuments: An Intimate Connection

Beginning in the 1990s, black fungi were described as one of the most likely groups of microorganisms responsible for the biodeterioration of the stone monuments [22,23,24,25] and it was confirmed in the following decades [6,18,26,27].
The term “black fungi” refers to a very huge group of dematiaceous fungi, unrelated phylogenetically, which have in common the presence of melanin in the cell wall that confers an olive brown appearance to the colony [28]. Another common characteristic is the ability to withstand hostile environments such as scarcity of nutrients, high solar irradiation, scarcity of water, high osmolarity, and low pH [15,19,29].
As reported by Gueidan et al. [30] the ancestors of black fungi were well adapted to live in oligotrophic environments such as rock surfaces or sub-surfaces, and currently they can also grow in anthropogenic habitats such as glass, silicon, organic surfaces, metals [31], or consolidants applied on the stone [9].
Their resilience is related to the extremotolerant or even polyextremotolerant characteristics of the species. The stress-tolerance is due to different factors such as: pigmentation, and in particular melanins production; mycosporine-like substances; morphological and metabolic versatility; meristematic development; and oligotrophy [32,33,34]. All these characteristics make them very suitable for colonizing outdoor rocks and built stones due to the fact that those surfaces can be exposed to extreme environments [17,18,19].
This group of fungi includes (a) fast growing hyphomycetes of epiphytic origin, recognizable under microscope by the presence of typical conidiophores and spores; (b) pleomorphic hyphomycetes that include the “black yeasts”, showing a yeast-like form, and the so-called “black meristematic fungi” with a Torula-like growth pattern (Figure 1).
Hyphomycetes and black yeasts are ubiquitous and widespread all over the world in very different habitats (e.g., soil, fresh water, sea, plants, animals, and humans) [28,35], while the meristematic black fungi, mostly isolated from stone or natural rocks, can be considered the true stone-inhabiting fungi [19,20,36,37].
In the literature, many of black fungi are reported as RIF (rock inhabiting fungi) to emphasize that the “rock” is their preferred or exclusive habitat. However, this terminology does not include their main features such as melanin production, pleomorphism, or meristematic development; for this reason, we do not use it in this context.
In the frame of cultural heritage the acronym MCF (MicroColonial Fungi) as first employed by Staley [38] is widely used for their description. It refers to the typical black cauliform-like colonies visible on the rocks and stones.
Humidity may affect the settlement of MCF on the stone artifacts as unique inhabitants or as associated with other stone colonizers. In fact, in lower or sheltered parts near the ground, where there is a sufficient availability of water, MCF are strictly associated with phototrophic microorganisms with whom, however, they do not establish a symbiotic relationship (Figure 2); in harsh, dry micro-environmental conditions, MCF become the unique colonizers [39,40,41].
Black fungi are currently classified in the Phylum of Ascomycota in the Class of Dothideomycetes and Eurotiomycetes, mainly in the order of Capnodiales, Dothideales, Chaetothyriales, Pleosporales and Cladosporiales, and Mycocaliciales [14,20,21,42].
In Table 1 are listed the genera of black fungi identified through molecular analyses that from 1997 up to date have been related to the biodeterioration of stone monuments.
In manuscripts published prior to 1999, black meristematic fungal species that were identified without molecular analyses, such as Hormonema dematioides, Lichenothelia sp. and Hortaea werneckii, Trimmatostroma sp., are listed among the most abundant fungal species present in arid and semiarid environments in association with biodeterioration of stone monuments [3].
The molecular analyses introduced at the end of the 20th century considerably increased the knowledge about the taxonomy of the black fungi isolated from stone monuments and allowed the description of twenty-six new species and three new genera.
The new species and genera described are listed below:
Sarcinomyces petricola Wollenzien and de Hoog [73]; S. sideticae Sert and Sterflinger [70]; Coniosporium apollinis Sterflinger, C. perforans Sterflinger [43]; C. uncinatum De Leo, Urzì and de Hoog [44]; C. sumbulii Sert and Sterflinger [47]; Phaeococcomyces chersonesos Bogomolova and Minter [74]; Pseudotaeniolina globosa De Leo, Urzì and de Hoog [54]; Capnobotryella antaliensis Sert and Sterflinger [50]; C. erdogani Sert and Sterflinger; C. kiziroglui Sert and Sterflinger [51]; Ochroconis lascauxensis Nováková and Martin-Sanchez; O. anomala Nováková and Martin-Sanchez [69]; Knufia marmoricola Onofri and Zucconi, K. vaticanii Zucconi and Onofri; K. karalitana Isola and Onofri; K. mediterranea Selbmann and Zucconi [37]; K. calcarecola Su, Sun and Xiang [72]; Exophiala bonarie Isola and Zucconi; Vermiconia calcicola de Hoog and Onofri [37]; Devriesia simplex Selbmann and Zucconi; D. modesta Isola and Zucconi [55]; and D. sardiniae Isola and de Hoog [37].
Three new genera and 4 species were also introduced as new: Saxophila tyrrenica Selbmann and de Hoog, Lithophila guttulata Selbmann and Isola [37], L. catenulata Su, Sun and Xiang [72], and Aeminium ludgeri Trovão, Tiago and Portugal [57].
Over the years, some of the above mentioned genera and species were reclassified: in particular, Sarcinomyces petricola and Phaecoccomyces chersonesos resulted identical, and they were reclassified as Knufia petricola [75,76]; Coniosporium perforans is now a synonym with Knufia perforans [76]; Devriesia species and Vermiconia species were included, respectively, in the new genera of Neodevriesia [77] and Vermiconidia [21]. Hao et al. [78] proposed a revision of the genus Ochroconis that was established as synonymous with the sister genus of Scolecobasidium. However, this taxonomic accommodation has been refused by Samerpitak et al. [79,80] on the basis of phylogenetic analyses and because the old generic name Scolecobasidium is considered of doubtful identity for the ambiguity of type specimens; therefore, the genus Ochroconis that is also characterized by oligotrophism and mesophilia was maintained.
However, many questions regarding the taxonomy and phylogeny of black fungi are still unresolved and further studies are required, especially to clarify the taxonomical position and phylogeny of many species of incertae sedis and of strains that are preserved in the mycological collections and are not yet identified (Figure 3, Table 2).

3. Mechanisms Involved in the Stone Biodeterioration

Being well adapted to the stone habitat and being oligotrophic, this group of fungi can often act as pioneer colonizer of the stone. In fact, for their growth it is sufficient to have just a little input of nutrient coming from the surrounding environment (e.g., animal and plant particles, air pollutants, guano droppings, etc.) [81,82]. Marble exposed to different environments and laboratory experiments demonstrated that black fungi such as Aureobasidium pullulans can be the first colonizer of freshly exposed marble surfaces in outdoor conditions [68,83].
The presence of a source of organic matter, such as the proximity of plants and trees, can considerably increase the chances of colonization by these fungi and the consequent rate of biodeterioration (Figure 4).
The mechanism of biodeterioration of stone monuments caused by black fungi is not fully understood [16,84].
The pattern of colonization of black fungi, and in particular of MCF, demonstrate once more that these microorganisms are well adapted to the stone habitat. In fact, they not only grow on the surface of stone, causing an aesthetic alteration due to the presence of melanin in the mycelium and conidia, described as discoloration, black staining, black spots, and black/greyish patinas [9,20,85]; but they can also act as true endolithic microorganisms by penetrating into the rocks/stones via intercrystalline spaces or through an active mechanism in which both mechanical and chemical aspects are hypothesized.
Microscopic observations show that where they settle is shaped accordingly to the morphology of these fungi (Figure 5). This fact can be explained by a local release of organic acids, followed by a precipitation of mineral phases; this buffer effect may be the reason why these fungi, in contrast to other ubiquitous hyphomycetes, such as Aspergillus niger and Penicillium spp., as reported by Salvadori and Municchia [16], do not show a marked organic acid production in laboratory conditions. However, Favero Longo et al. [86] demonstrated that some species of MCF (e.g., Knufia petricola) penetrate actively into freshly exposed stone probes through the production of iron-chelating molecules (siderophores such as compounds).
Another mechanism is due to the ability of these fungi to penetrate the stone using already existing fractures and cracks. The mechanical forces due to the expansion of hyphae may increase the fractures and cause the loss of materials [18,26,39]. To explain the mechanical penetration of the hyphae into the stone, a past hypothesis gave a crucial role to melanin present in the cell wall of hyphae and in the meristematic cells, similar to the process of penetration of phytopathogenic fungi in the host cells [16,23,25,39].
In fact, the melanin present in the appressoria of the phytopathogenic fungi confers turgor and rigidity to the cells, favouring their penetration of the host cells [87].
However, on the basis of the results obtained in a recent publication by Tonon et al. [88], this hypothesis should be rejected. In fact, non-melanized mutants of the black fungus Knufia petricola do not lose their ability to penetrate carbonate pellets regardless of porosity; on the contrary, thin, non-melanized, exploring hyphae showed an even higher penetration pattern into the stone, probably due to their nutrient-seeking role and thinness.

4. Multistep Analyses to Study Black Fungi from Stone Monuments

Contrary to studies carried out on natural rocks, in the frame of cultural heritage it is mandatory to carry out all the necessary multidisciplinary studies (chemical, geological, physical, biological analysis, etc.), by using very low destructive or non-destructive sampling methods [18,46,67,89]. This fact may limit the extent of the studies, but a careful planning of a sampling campaign leads to the right protocol of intervention and of assessing the risk of further biodeterioration processes.
A general useful multistep approach should include microscopy, cultural analyses, molecular analyses, and the laboratory evaluation of selected methods of control (chemical or physical) on isolated strains. The assessment of environmental conditions (temperature, humidity, shining, presence of surrounding vegetation, or other organic sources, atmospheric pollutants, etc.) should be also evaluated. In fact, these data not only allow us a better understanding of the physiology and ecology of fungi, but can help to control their growth indirectly, especially in indoor and/or in confined environments. At the end, a monitoring campaign over time should establish the level of the risk of the item and the frequency of intervention.
Further, a common language for the description of the alterations is also indispensable for sharing the results with the scientific community; to this purpose, there is a glossary [90,91] to obtain an objective and standardized description.

4.1. Methods of Isolation and Characterization

The sampling is critical. As reported in the previous paragraph, due to the value of the artifacts, non-invasive sampling methods have been developed over the years and are now widely used. Examples are a needle to take samples from black spots or cavities, scalpel or lancet to scrape fungi from the surface, and adhesive tape sample, useful both for microscopy and cultural and molecular analysis as it provides a mirror image of the stone colonization [22,92,93].
Microscopic examination of the samples is the most common practice to obtain evidence of black fungi directly from stone samples. In fact, due to their size, morphology, and pigmentation, black fungi can be directly visualized under optical microscope without a specific preparation. Even with a good light microscope (LM) or, better, with a scanning electron microscope (SEM), detailed information is obtained. Microscopy is useful for the direct visualization of the fungi in the stone sample, for determining what types of relationship they establish with stone material and with other types of microorganisms, and also for addressing the next step of analyses. Unfortunately, fluorescence microscopy (FM) cannot help in detecting and studying black fungi due to the presence of melanin that masks the fluorochrome fluorescence.
Still, cultural analyses remain the best way to study this group of fungi.
Black fungi, and in particular MCF, possess a poor ability to compete with fast growing fungi, being characterized by a slow growth rate, and very often require more than 1 month of incubation before visible colonies are seen. These reasons explain why they are difficult to isolate and maintain in culture [94]. Nevertheless, selective cultural media that inhibit the growth of bacteria and of fast-growing fungi are successfully employed both for qualitative and quantitative cultural analyses [22,95].
These culture techniques have the advantage of allowing the whole characterization of the isolates by microscopical, biochemical, physiological, and molecular analysis; these latter are indispensable for the identification of MCF that do not have recognizable morphological traits. Multilocus sequencing typing (mlst) is routinely carried out to resolve their taxonomic and phylogenetic position [55,56] by Blast search homology (Basic Local Alignment Search Tools) available online. However, many nucleotide sequences in the Genkank nucleotide database are not updated in the “definition”. Therefore, although this approach is within the reach of all laboratories, it requires a deep knowledge of the literature and a curated nucleotide database.
A deeper genetic characterization of the isolates can be obtained by whole genome analyses but, to date, only four genomes belonging to three species of black fungi (Coniosporium apollinis CBS 100218, Knufia petricola MA5789, K. petricola MA5790, and Aeminium ludgeri DSM 106916) isolated from stone monuments are available in the NCBI database (https://www.ncbi.nlm.nih.gov/genome/ accessed on 15 February 2022).
Cultural techniques also allow us to investigate the biodeteriorative abilities of the isolated species by setting up laboratory experiments that simulate their settlement, colonization, and biodeterioration pattern [96,97].

4.2. Culture Independent Analyses

It is well known that only a very small percentage (<0.5%) of environmental microorganisms can be cultured. Therefore, culture independent molecular approaches have been developed to overcome this limitation; the metagenomic approaches allow the study of the microbiota (microbial community) and its microbiome (gene pool) by total acid nucleic extraction from the samples, both DNA and/or RNA, depending on the purpose.
Recently Sterflinger and Pinar [98] reviewed the main molecular-based techniques that are currently available and, although some of these have not yet been used in the field of cultural heritage, they could be adapted for future studies.
In the literature, the majority of papers are focused on Bacteria and Archaea, while there are very few papers about microfungi from stone monuments [99,100].
Large-scale genomic analyses, such as high-throughput-sequencing analyses, are becoming increasingly popular to study the microbiota (both for biodiversity and functional genes analyses) in different areas of research, and cultural heritage is not an exception [89,99]. These techniques were developed in the late 1990s and early 2000s, and today they are more accessible to many laboratories due to the lower costs and the facilities offered by many companies that can carry out all steps of analysis, from nucleic acid extraction to bioinformatic data analysis.
The culture-independent approach certainly contributes to the deep knowledge on the microorganisms associated with the biodeterioration processes. However, there is a very high risk to obtain a plethora of data that are not easily interpretable, because they cannot be directly connected with the biodeterioration phenomenon observed. In fact, it is obvious, but not trivial, that the discovery of a microbial agent on a monument could be not related to any biodeteriogenic activity. Therefore, the current need is to associate these techniques to the culture-based ones for a more complete characterization of the state of deterioration of the artifact and for implementation of the more suitable prevention strategies.

5. How to Control Black Fungi

Despite the wide literature regarding the control of biodeteriogens on inorganic surfaces as reported in recent books and reviews [6,9,101,102,103], very little is said regarding the effectiveness of treatments against black fungi.
Black fungi, especially meristematic ones, are very difficult to eradicate and tend to be one of the first colonizers after cleaning procedures [6,12,83]. In the Lascaux cave, a black yeast Ochroconis lascauxensis caused an important and extensive black discoloration on the cave’s walls whose origin and evolution were probably linked to the intensive biocide treatments [104].
In order to achieve protection of an artifact, both indirect and direct methods should be implemented. The first ones aim to control, or more realistically to mitigate, the fungal growth by modification of the chemical-physical parameters such as humidity, source of nutrients, and temperature, that are crucial key factors for fungal growth. It is obvious that this is rarely fully achievable, and only in particular circumstances, such as in indoor environments (churches, museums, etc.) or for movable artifacts and objects that can be moved if necessary, while in outdoor conditions this is quite impossible.
Direct treatments aiming to kill/reduce black fungi on the stone should be different on the basis of their colonization pattern (diffuse patina, spot-like colonization, or intercrystalline growth) and on the characteristics of the environment; for example, in an indoor environment, the air is often heavily contaminated by fungal spores and thus they need to be eliminated at the same time as those settled on the surfaces; otherwise, their presence in the air is a continued source of reinfection.
Among the potential methods commonly used to control biodeterioration, physical methods such as mechanical removal and UV and heat shock treatments [101,105], are not very effective against black fungi [102,106].
Regarding chemical methods, in laboratory conditions, classical biocides (e.g., Preventol RI 50, Biotin R, RocimaTM 103) are still the most effective [102,107] and in the field they produce efficient results during cleaning procedures. Plant based extracts show a scarce effectiveness against fungi, and this difficult group of microorganisms is not even taken into account to assess their activity [108]. Nanoparticles are commonly used as biocides due to their activity against algae, cyanobacteria, and most bacteria, but they are not really satisfactory against black fungi.
Protective coatings with antifouling properties may have various effects. In fact, TiO2 based coatings, pure or doped with Ag, show a good effect but are limited to a short/medium term after application [108]. However, in both laboratory and field conditions, after treatments with titania-based coatings, black fungi are the first to recolonize the stone surface in dry environments, while algae first appears in damping walls [63]. Very recently, in laboratory conditions, cholinium@Il based coatings have shown that the use of Il’s with a 12 C chains and DBS as anion in combination with nanosilica coatings (e.g., Nano Estel) could be effective against the colonization of black fungi for a period of time over 30 months [109].
One possible explanation of this scarce effectiveness of most treatments against black fungi is that they possess a genetic resistance to environmental stresses, as reported in the previous paragraphs. Therefore, the different mechanisms concurring to the stress protection response may interfere to the biocidal treatments.
Understanding the cause of their resilience could improve the strategies for their control.

6. Concluding Remarks

The study of biodeterioration of stone monuments is quite complex and cannot be improvised. For a correct understanding of biodeterioration phenomena and the implementation of measures aimed at the elimination and/or mitigation of biodeteriogenic microorganisms, it is important to consider the monument and its surrounding as a whole.
When working for the protection of cultural heritage artifacts, scientists should not follow the same protocols for all the situations. In general, it is necessary to:
(1)
Listen the conservators;
(2)
Evaluate the environmental climatic conditions and specific conditions, such as the type of material and the overall status of conservation of monument; the description of the type of alteration visible under naked eye should be also included;
(3)
Interact with the other experts involved;
(4)
Answer the questions posed by the conservators.
The analysis must be planned according to their questions.
It is also important to relate the presence of fungal species with the observed biodeterioration phenomenon; then, for treatments, it is possible to use well known protocols or propose new products/treatments that, however, need to be tested in laboratory with fungal isolates and in situ (on probes, not on the item!!!) before applying it to the CH item. Finally, consider evaluating the use of coatings that match Green conservation criteria and are effective to prevent or slow down new colonization.
As there is not only one method that is valid in all circumstances, we have to work out, case by case, the best solution and monitor the result over the time to avoid unexpected and/or undesirable effects as much as possible.
Black fungi, especially meristematic ones, are very dangerous for stone artifacts for several reasons:
(a)
They are responsible of discolouring of the stone surface. The extended colonization of surfaces changes the global vision of the artifact, especially if different material and colour of stones were used by the artist;
(b)
Moreover, black fungi show an inter-crystalline pattern of growth. This pattern causes crystals to detach (so called sugaring) with loss of precious material, especially because it involves the first surface layer (very important for bas-reliefs and sculptures);
(c)
They could determine the biopitting. Fungi excavate cavities on the stone where they can better settle, giving the surface a pockmarked aspect. The convergence of several biopitting can often lead to larger cavities;
(d)
Hyphae penetrate deep into the surface, even more than a few mm;
(e)
Chemical and physical treatments used for other microorganisms are often non efficient in eradication;
(f)
Black fungi are often the first colonizers after the treatments.
For all these reasons, new methods of study and of control are needed that also aim to search for more eco-friendly molecules and/or approaches.
Despite the increase in interest in black fungi as a cause of biodeterioration of stone monuments and artifacts, many aspects need a more in-depth analysis. For example, not much is known about the molecular mechanisms involved in stress tolerance, in colonization, and biodeterioration of stone. Very important results were achieved by laboratory experiments that, however, concern only some species, and are still too few to generalize the results obtained. Only four genomes of three species were sequenced, and they are not sufficient for a comparative analyses aiming to a better understanding of the above mentioned processes and mechanisms.
Furthermore, the creation of a curated database including the nucleotide sequences used for identification of black fungi from monuments could be very helpful, as the public databases are not curated and are outdated; they could even be misleading for people who are not interested in deepening the knowledge around the taxonomy and phylogeny of this group of fungi.
Finally, it is also important to combine the classical methods with the new ones to improve current knowledge useful for implementation of future conservation strategies.

Author Contributions

Conceptualization, F.D.L. and C.U.; methodology, F.D.L., A.M. and C.U.; resources, F.D.L. and C.U.; data curation F.D.L.; writing—original draft preparation, F.D.L., A.M. and C.U. writing—review and editing, F.D.L., A.M. and C.U. All authors have read and agreed to the published version of the manuscript.

Funding

No funding was used for this manuscript.

Informed Consent Statement

Not applicable.

Data Availability Statement

Not applicable.

Acknowledgments

Authors would like to thank Sherron Collins for her revision of the English text.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Krumbein, W.E. Microbial interactions with mineral materials. In Biodeterioration 7, 1st ed.; Hougthon, D.R., Smith, R.N., Eggins, H.O., Eds.; Springer: Dordrecht, The Netherlands, 1988; pp. 78–100. [Google Scholar] [CrossRef]
  2. Saiz-Jimenez, C. Biogeochemistry of weathering processes in monuments. Geomicrobiol. J. 1999, 16, 27–37. [Google Scholar] [CrossRef]
  3. Sterflinger, K. Fungi as geologic agents. Geomicrobiol. J. 2000, 17, 97–124. [Google Scholar] [CrossRef]
  4. Gadd, G.M. Geomicrobiology of the built environment. Nat. Microbiol. 2017, 2, 16275. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  5. Urzì, C. Biodeterioramento dei manufatti artistici. In Microbiologia Ambientale ed Elementi di Ecologia Microbica; Barbieri, P., Bestetti, G., Galli, E., Zannoni, D., Eds.; Casa Editrice Ambrosiana: Milano, Italy, 2008; pp. 327–346. ISBN 978-88-08-18434-4. [Google Scholar]
  6. Pinna, D. Microbial growth and its effect on inorganic heritage material. In Microorganisms in the Deterioration and Preservation of Cultural Heritage, 1st ed.; Joseph, E., Ed.; Springer: Cham, Switzerland, 2021; pp. 3–35. [Google Scholar] [CrossRef]
  7. Gorbushina, A.A. Life on the rocks. Environ. Microbiol. 2007, 9, 1613–1631. [Google Scholar] [CrossRef] [PubMed]
  8. Villa, F.; Stewart, P.S.; Klapper, I.; Jacob, J.M.; Cappitelli, F. Subaerial biofilms on outdoor stone monuments: Changing the perspective toward an ecological framework. BioScience 2016, 66, 285–294. [Google Scholar] [CrossRef] [Green Version]
  9. Pinna, D. Coping with Biological Growth on Stone Heritage Objects: Methods, Products, Applications, and Perspectives, 1st ed.; Apple Academic Press: Oakville, ON, Canada, 2017; ISBN 9781771885324. [Google Scholar]
  10. Guillitte, O. Bioreceptivity: A new concept for building ecology studies. Sci. Total Environ. 1995, 167, 215–222. [Google Scholar] [CrossRef]
  11. Saiz-Jimenez, C. Deposition of anthropogenic compounds on monuments and their effect on airborne microorganisms. Aerobiologia 1995, 11, 161–175. [Google Scholar] [CrossRef]
  12. Urzì, C.; De Leo, F.; Salamone, P.; Criseo, G. Airborne fungal spores colonising marbles exposed in the terrace of Messina Museum, Sicily. Aerobiologia 2001, 17, 11–17. [Google Scholar] [CrossRef]
  13. Polo, A.; Gulotta, D.; Santo, N.; Di Benedetto, C.; Fascio, U.; Toniolo, L.; Villa, F.; Cappitelli, F. Importance of subaerial biofilms and airborne microflora in the deterioration of stonework: A molecular study. Biofouling 2012, 28, 1093–1106. [Google Scholar] [CrossRef] [Green Version]
  14. Liu, B.; Fu, R.; Wu, B.; Liu, X.; Xiang, M. Rock-inhabiting fungi: Terminology, diversity, evolution and adaptation mechanisms. Mycology 2022, 13, 1–31. [Google Scholar] [CrossRef]
  15. Selbmann, L.; Egidi, E.; Isola, D.; Onofri, S.; Zucconi, Z.; de Hoog, G.S.; Chinaglia, S.; Testa, L.; Tosi, S.; Balestrazzi, A.; et al. Biodiversity, evolution and adaptation of fungi in extreme environments. Plant Biosyst. 2013, 147, 237–246. [Google Scholar] [CrossRef] [Green Version]
  16. Salvadori, O.; Municchia, A.C. The role of fungi and lichens in the biodeterioration of stone monuments. Open Conf. Proc. J. 2016, 7, 39–54. [Google Scholar] [CrossRef]
  17. Liu, X.; Koestler, R.J.; Warscheid, T.; Katayama, Y.; Gu, J.-D. Microbial deterioration and sustainable conservation of stone monuments and buildings. Nat. Sustain. 2020, 3, 991–1004. [Google Scholar] [CrossRef]
  18. Sterflinger, K. Fungi: Their role in deterioration of cultural heritage. Fungal Biol. Rev. 2010, 24, 47–55. [Google Scholar] [CrossRef]
  19. Urzì, C.; De Leo, F.; de Hoog, S.; Sterflinger, K. Recent advances in the molecular biology and ecophysiology of meristematic stone-inhabiting fungi. In Of Microbes and Art: The Role of Microbial Communities in the Degradation and Protection of Cultural Heritage; Ciferri, O., Tiano, P., Mastromei, G., Eds.; Springer: Boston, MA, USA, 2000; pp. 3–19. [Google Scholar] [CrossRef]
  20. Onofri, S.; Zucconi, L.; Isola, D.; Selbmann, L. Rock-inhabiting fungi and their role in deterioration of stone monuments in the Mediterranean area. Plant Biosyst. 2014, 148, 384–391. [Google Scholar] [CrossRef]
  21. Crous, P.W.; Schumacher, R.K.; Akulov, A.; Thangavel, R.; Hernández-Restrepo, M.; Carnegie, A.J.; Cheewangkoon, R.; Wingfield, M.J.; Summerell, B.A.; Quaedvlieg, W.; et al. New and interesting fungi. Fungal Syst. Evol. 2019, 3, 57–134. [Google Scholar] [CrossRef]
  22. Wollenzien, U.; de Hoog, G.S.; Krumbein, W.E.; Urzì, C. On the isolation of microcolonial fungi occurring on and in marble and other calcareous rocks. Sci. Total Environ. 1995, 167, 287–294. [Google Scholar] [CrossRef]
  23. Diakumaku, E.; Gorbushina, A.A.; Krumbein, W.E.; Panina, L.; Soukharjevski, S. Black fungi in marble and limestones–an aesthetical, chemical and physical problem for the conservation of monuments. Sci. Total Environ. 1995, 167, 295–304. [Google Scholar] [CrossRef]
  24. Urzì, C.; Wollenzien, U.; Criseo, G.; Krumbein, W.E. Biodiversity of the rock inhabiting microflora with special reference to black fungi and black yeasts. In Microbial Diversity and Ecosystem Function; Allsopp, D., Colwell, R.R., Hawksworth, D.L., Eds.; CAB International: Wallingford, UK, 1995; pp. 289–302. [Google Scholar]
  25. Sterflinger, K.; Krumbein, W.E. Dematiaceous fungi as a major agent for biopitting on mediterranean marbles and limestones. Geomicrobiol. J. 1997, 14, 219–230. [Google Scholar] [CrossRef]
  26. De Leo, F.; Urzì, C. Microfungi from deteriorated materials of cultural heritage. In Fungi from Different Substrates; Misra, J.K., Tewari, J.P., Deshmukh, S.K., Vágvölgyi, C., Eds.; CRC Press: Boca Raton, FL, USA, 2015; pp. 144–158. [Google Scholar] [CrossRef]
  27. Isola, D.; Zucconi, L.; Cecchini, A.; Caneva, G. Dark-pigmented biodeteriogenic fungi in etruscan hypogeal tombs: New data on their culture-dependent diversity, favouring conditions, and resistance to biocidal treatments. Fungal Biol. 2021, 125, 609–620. [Google Scholar] [CrossRef]
  28. de Hoog, G.S.; Guarro, J.; Gené, S.A.; Al-Hatmi, A.M.S.; Figueras, M.J.; Vitale, R.G. Atlas of Clinical Fungi, 4th ed.; Westerdijk Institute/Universitat Rovira Virgili: Utrecht, The Netherlands, 2019. [Google Scholar]
  29. Palmer, F.E.; Emery, D.R.; Stemmler, J.; Staley, J.T. Survival and growth of microcolonial rock fungi as affected by temperature and humidity. New Phytol. 1987, 107, 155–162. [Google Scholar] [CrossRef]
  30. Gueidan, C.; Ruibal, C.; de Hoog, S.; Schneider, H. Rock-inhabiting fungi originated during periods of dry climate in the late Devonian and middle Triassic. Fungal Biol. 2011, 115, 987–996. [Google Scholar] [CrossRef] [PubMed]
  31. Gostinčar, C.; Grube, M.; Gunde-Cimerman, N. Evolution of fungal pathogens in domestic environments? Fungal Biol. 2011, 115, 1008–1018. [Google Scholar] [CrossRef] [PubMed]
  32. Gorbushina, A.A.; Whitehead, K.; Dornieden, T.; Niesse, A.; Schulte, A.; Hedges, J.I. Black fungal colonies as units of survival: Hyphal mycosporines synthesized by rock-dwelling microcolonial fungi. Can. J. Bot. 2003, 81, 131–138. [Google Scholar] [CrossRef]
  33. Gostinčar, C.; Muggia, L.; Grube, M. Polyextremotolerant black fungi: Oligotrophism, adaptive potential and a link to lichen symbioses. Front. Microbiol. 2012, 3, 390. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  34. Zakharova, K.; Tesei, D.; Marzban, G.; Dijksterhuis, J.; Wyatt, T.; Sterflinger, K. Microcolonial fungi on rocks: A life in constant drought? Mycopathologia 2013, 175, 537–547. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  35. Marchetta, A.; van den Ende, G.B.; Al-Hatmi, A.M.S.; Hagen, F.; Zalar, P.; Sudhadham, M.; Gunde-Cimerman, N.; Urzì, C.; de Hoog, S.; De Leo, F. Global molecular diversity of the halotolerant fungus Hortaea werneckii. Life 2018, 8, 31. [Google Scholar] [CrossRef] [Green Version]
  36. Sterflinger, K.; Piñar, G. Microbial deterioration of cultural heritage and works of art—tilting at windmills? Appl. Microbiol. Biot. 2013, 97, 9637–9646. [Google Scholar] [CrossRef] [Green Version]
  37. Isola, D.; Zucconi, L.; Onofri, S.; Caneva, G.; de Hoog, G.S.; Selbmann, L. Extremotolerant rock inhabiting black fungi from Italian monumental sites. Fungal Divers. 2016, 76, 75–96. [Google Scholar] [CrossRef]
  38. Staley, J.T.; Palmer, F.; Adams, J.B. Microcolonial fungi: Common inhabitants on desert rocks? Science 1982, 215, 1093–1095. [Google Scholar] [CrossRef] [Green Version]
  39. De Leo, F.; Antonelli, F.; Pietrini, A.M.; Ricci, S.; Urzì, C. Study of the euendolithic activity of black meristematic fungi isolated from a marble statue in the Quirinale Palace’s Gardens in Rome, Italy. Facies 2019, 65, 18. [Google Scholar] [CrossRef]
  40. Santo, A.P.; Cuzman, O.A.; Petrocchi, D.; Pinna, D.; Salvatici, T.; Perito, B. Black on white: Microbial growth darkens the external marble of Florence cathedral. Appl. Sci. 2021, 11, 6163. [Google Scholar] [CrossRef]
  41. Marvasi, M.; Donnarumma, F.; Frandi, A.; Mastromei, G.; Sterflinger, K.; Tiano, P.; Perito, B. Black microcolonial fungi as deteriogens of two famous marble statues in Florence, Italy. Int. Biodeterior. Biodegrad. 2012, 68, 36–44. [Google Scholar] [CrossRef]
  42. Abdollahzadeh, J.; Groenewald, J.Z.; Coetzee, M.P.A.; Wingfield, M.J.; Crous, P.W. Evolution of lifestyles in Capnodiales. Stud. Mycol. 2020, 95, 381–414. [Google Scholar] [CrossRef]
  43. Sterflinger, K.; de Baere, R.; de Hoog, G.S.; de Wachter, R.; Krumbein, W.E.; Haase, G. Coniosporium perforans and C. apollinis, two new rock-inhabiting fungi isolated from marble in the Sanctuary of Delos (Cyclades, Greece). Antonie Van Leeuwenhoek J. Microb. 1997, 72, 349–363. [Google Scholar] [CrossRef]
  44. De Leo, F.; Urzì, C.; de Hoog, G.S. Two Coniosporium species from rock surfaces. Stud. Mycol. 1999, 43, 70–79. [Google Scholar]
  45. Sterflinger, K.; Prillinger, H. Molecular taxonomy and biodiversity of rock fungal communities in an urban environment (Vienna, Austria). Antonie Van Leeuwenhoek J. Microb. 2001, 80, 275–286. [Google Scholar] [CrossRef]
  46. Ricca, M.; Urzì, C.E.; Rovella, N.; Sardella, A.; Bonazza, A.; Ruffolo, S.A.; De Leo, F.; Randazzo, L.; Arcudi, A.; La Russa, M.F. Multidisciplinary approach to characterize archaeological materials and status of conservation of the roman Thermae of Reggio Calabria site (Calabria, south Italy). Appl. Sci. 2020, 10, 5106. [Google Scholar] [CrossRef]
  47. Sert, H.B.; Sterflinger, K. A new Coniosporium species from historical marble monuments. Mycol. Prog. 2010, 9, 353–359. [Google Scholar] [CrossRef]
  48. Sert, H.B.; Sümbül, H.; Sterflinger, K. Microcolonial fungi from antique marbles in Perge/Side/Termessos (Antalya/Turkey). Antonie Van Leeuwenhoek J. Microb. 2007, 91, 217–227. [Google Scholar] [CrossRef]
  49. Sazanova, K.V.; Zelenskaya, M.S.; Vlasov, A.D.; Bobir, S.Y.; Yakkonen, K.L.; Vlasov, D.Y. Microorganisms in superficial deposits on the stone monuments in Saint Petersburg. Microorganisms 2022, 10, 316. [Google Scholar] [CrossRef]
  50. Sert, H.B.; Sümbül, H.; Sterflinger, K. A new species of Capnobotryella from monument surfaces. Mycol. Res. 2007, 111, 1235–1241. [Google Scholar] [CrossRef] [PubMed]
  51. Sert, H.B.; Sümbül, H.; Sterflinger, K. Two new species of Capnobotryella from historical monuments. Mycol. Prog. 2011, 10, 333–339. [Google Scholar] [CrossRef]
  52. Sert, H.B.; Wuczkowski, M.; Sterflinger, K. Capnobotryella isiloglui, a new rock-inhabiting fungus from Austria. Turk. J. Bot. 2012, 3, 401–407. [Google Scholar] [CrossRef]
  53. Owczarek-Kościelniak, M.; Krzewicka, B.; Piątek, J.; Kołodziejczyk, Ł.M.; Kapusta, P. Is there a link between the biological colonization of the gravestone and its deterioration? Int. Biodeterior. Biodegrad. 2020, 148, 104879. [Google Scholar] [CrossRef]
  54. De Leo, F.; Urzì, C.; de Hoog, G.S. A new meristematic fungus, Pseudotaeniolina globosa. Antonie Van Leeuwenhoek J. Microb. 2003, 83, 351–360. [Google Scholar] [CrossRef]
  55. Egidi, E.; de Hoog, G.S.; Isola, D.; Onofri, S.; Quaedvlieg, W.; de Vries, M.; Verkley, G.J.M.; Stielow, J.B.; Zucconi, L.; Selbmann, L. Phylogeny and taxonomy of meristematic rock-inhabiting black fungi in the Dothideomycetes based on multi-locus phylogenies. Fungal Diver. 2014, 65, 127–165. [Google Scholar] [CrossRef]
  56. Rizk, S.M.; Magdy, M.; De Leo, F.; Werner, O.; Rashed, M.A.-S.; Ros, R.M.; Urzì, C. A new extremotolerant ecotype of the fungus Pseudotaeniolina globosa isolated from Djoser Pyramid, Memphis Necropolis, Egypt. J. Fungi 2021, 7, 104. [Google Scholar] [CrossRef]
  57. Trovão, J.; Tiago, I.; Soares, F.; Paiva, D.S.; Mesquita, N.; Coelho, C.; Catarino, L.; Gil, F.; Portugal, A. Description of Aeminiaceae fam. nov., Aeminium gen. nov. and Aeminium ludgeri sp. nov. (Capnodiales), isolated from a biodeteriorated art-piece in the old cathedral of Coimbra, Portugal. MycoKeys 2019, 45, 57–73. [Google Scholar] [CrossRef]
  58. Nuhoglu, Y.; Oguz, E.; Uslu, H.; Ozbek, A.; Ipekoglu, B.; Ocak, I.; Hasenekoglu, I. The accelerating effects of the microorganisms on biodeterioration of stone monuments under air pollution and continental-cold climatic conditions in Erzurum, Turkey. Sci. Total Environ. 2006, 364, 272–283. [Google Scholar] [CrossRef] [Green Version]
  59. Cappitelli, F.; Principi, P.; Pedrazzani, R.; Toniolo, L.; Sorlini, C. Bacterial and fungal deterioration of the Milan cathedral marble treated with protective synthetic resins. Sci. Total Environ. 2007, 385, 172–181. [Google Scholar] [CrossRef] [PubMed]
  60. Cappitelli, F.; Nosanchuk, J.; Casadevall, A.; Toniolo, L.; Brusetti, L.; Florio, S.; Principi, P.; Borin, S.; Sorlini, C. Synthetic consolidants attacked by melanin-producing fungi: Case study of the biodeterioration of Milan (Italy) cathedral marble treated with acrylics. Appl. Environ. Microbiol. 2007, 73, 271–277. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  61. Suihko, M.L.; Alakomi, H.L.; Gorbushina, A.; Fortune, I.; Marquardt, J.; Saarela, M. Characterization of aerobic bacterial and fungal microbiota on surfaces of historic Scottish monuments. Syst. Appl. Microbiol. 2007, 30, 494–508. [Google Scholar] [CrossRef] [PubMed]
  62. Ortega-Morales, B.O.; Narváez-Zapata, J.; Reyes-Estebanez, M.; Quintana, P.; De la Rosa-García del, C.S.; Bullen, H.; Gómez-Cornelio, S.; Chan-Bacab, M.J. Bioweathering potential of cultivable fungi associated with semi-arid surface microhabitats of mayan buildings. Front. Microbiol. 2016, 7, 201. [Google Scholar] [CrossRef]
  63. Ruffolo, S.A.; De Leo, F.; Ricca, M.; Arcudi, A.; Silvestri, C.; Bruno, L.; Urzì, C.; La Russa, M.F. Medium-term in situ experiment by using organic biocides and titanium dioxide for the mitigation of microbial colonization on stone surfaces. Int. Biodeterior. Biodegrad. 2017, 123, 17–26. [Google Scholar] [CrossRef]
  64. Trovão, J.; Portugal, A.; Soares, F.; Paiva, D.S.; Mesquita, N.; Coelho, C.; Pinheiro, A.C.; Catarino, L.; Gil, F.; Tiago, I. Fungal diversity and distribution across distinct biodeterioration phenomena in limestone walls of the old cathedral of Coimbra, UNESCO World Heritage Site. Int. Biodeterior. Biodegrad. 2019, 142, 91–102. [Google Scholar] [CrossRef]
  65. Trovão, J.; Gil, F.; Catarino, L.; Soares, F.; Tiago, I.; Portugal, A. Analysis of fungal deterioration phenomena in the first Portuguese King tomb using a multi-analytical approach. Int. Biodeterior. Biodegrad. 2020, 149, 104933. [Google Scholar] [CrossRef]
  66. Mang, S.M.; Scrano, L.; Camele, I. Preliminary studies on fungal contamination of two rupestrian churches from Matera (Southern Italy). Sustainability 2020, 12, 6988. [Google Scholar] [CrossRef]
  67. Urzì, C.; De Leo, F.; Bruno, L.; Albertano, P. Microbial diversity in paleolithic caves: A study case on the phototrophic biofilms of the Cave of Bats (Zuheros, Spain). Microb. Ecol. 2010, 60, 116–129. [Google Scholar] [CrossRef]
  68. Urzì, C.; De Leo, F.; Lo Passo, C.; Criseo, G. Intra-specific diversity of Aureobasidium pullulans strains isolated from rocks and other habitats assessed by physiological methods and by random amplified polymorphic DNA (RAPD). J. Microbiol. Meth. 1999, 36, 95–105. [Google Scholar] [CrossRef]
  69. Martin-Sanchez, P.M.; Nováková, A.; Bastian, F.; Alabouvette, C.; Saiz-Jimenez, C. Two new species of the genus Ochroconis, O. lascauxensis and O. anomala isolated from black stains in Lascaux Cave, France. Fungal Biol. 2012, 116, 574–589. [Google Scholar] [CrossRef] [PubMed]
  70. Sert, H.B.; Sümbül, H.; Sterflinger, K. Sarcinomyces sideticae, a new black yeast from historical marble monuments in Side (Antalya, Turkey). Bot. J. Linn. Soc. 2007, 154, 373–380. [Google Scholar] [CrossRef] [Green Version]
  71. Isola, D.; Selbmann, L.; de Hoog, G.S.; Fenice, M.; Onofri, S.; Prenafeta-Boldú, F.X.; Zucconi, L. Isolation and screening of black fungi as degraders of volatile aromatic hydrocarbons. Mycopathologia 2013, 175, 369–379. [Google Scholar] [CrossRef] [PubMed]
  72. Sun, W.; Su, L.; Yang, S.; Sun, J.; Liu, B.; Fu, R.; Wu, B.; Liu, X.; Cai, L.; Guo, L.; et al. Unveiling the hidden diversity of rock-inhabiting fungi: Chaetothyriales from China. J. Fungi 2020, 6, 187. [Google Scholar] [CrossRef] [PubMed]
  73. Wollenzien, U.; de Hoog, G.S.; Krumbein, W.E.; Uijthof, J.M.J. Sarcinomyces petricola, a new microcolonial fungus from marble in the Mediterranean basin. Antonie Van Leeuwenhoek J. Microb. 1997, 71, 281–288. [Google Scholar] [CrossRef]
  74. Bogomolova, E.V.; Minter, D.W. A new microcolonial rock-inhabiting fungus from marble in Chersonesos (Crimea, Ukraine). Mycotaxon 2003, 86, 195–204. [Google Scholar]
  75. Tsuneda, A.; Hambleton, S.; Currah, R.S. The anamorph genus Knufia and its phylogenetically allied species in Coniosporium, Sarcinomyces, and Phaeococcomyces. Can. J. Bot. 2011, 89, 523–536. [Google Scholar] [CrossRef]
  76. Nai, C.; Wong, H.Y.; Pannenbecker, A.; Broughton, W.J.; Benoit, I.; de Vries, R.P.; Guedain, C.; Gorbushina, A.A. Nutritional physiology of a rock-inhabiting, model microcolonial fungus from an ancestral lineage of the Chaetothyriales (Ascomycetes). Fungal Genet. Biol. 2013, 56, 54–66. [Google Scholar] [CrossRef]
  77. Quaedvlieg, W.; Binder, M.; Groenewald, J.Z.; Summerell, B.A.; Carnegie, A.J.; Burgess, T.I.; Crous, P.W. Introducing the consolidated species concept to resolve species in the Teratosphaeriaceae. Persoonia 2014, 33, 1–40. [Google Scholar] [CrossRef] [Green Version]
  78. Hao, L.; Chen, C.; Zhang, R.; Zhu, M.; Sun, G.; Gleason, M.L. A new species of Scolecobasidium associated with the sooty blotch and flyspeck complex on banana from China. Mycol. Prog. 2013, 12, 489–495. [Google Scholar] [CrossRef]
  79. Samerpitak, K.; Van der Linde, E.; Choi, H.J.; Gerrits van den Ende, A.H.G.; Machouart, M.; Gueidan, C.; de Hoog, G.S. Taxonomy of Ochroconis, genus including opportunistic pathogens on humans and animals. Fungal Diver. 2014, 65, 89–126. [Google Scholar] [CrossRef]
  80. Samerpitak, K.; Duarte, A.P.M.; Attili-Angelis, D.; Pagnocca, F.C.; Heinrichs, G.; Rijs, A.J.M.M.; Alfjorden, A.; Gerrits van den Ende, A.H.G.; Menken, S.B.J.; de Hoog, G.S. A new species of the oligotrophic genus Ochroconis (Sympoventuriaceae). Mycol. Progress 2015, 14, 6. [Google Scholar] [CrossRef] [Green Version]
  81. De Leo, F.; Criseo, G.; Urzì, C. Impact of Surrounding Vegetation and Soil on the Colonization of Marble Statues by Dematiaceous Fungi. In Proceedings of the 8th International Congress on Deterioration and Conservation of Stone, Berlin, Germany, 30 September–4 October 1996; Reiderer, Ed.; Möller Druck und Verlag GMBH: Berlin, Germany, 1996; pp. 625–630. [Google Scholar]
  82. Scheerer, S.; Ortega-Morales, O.; Gaylarde, C. Microbial deterioration of stone monuments--an updated overview. Adv. Appl. Microbiol. 2009, 66, 97–139. [Google Scholar] [CrossRef] [PubMed]
  83. De Leo, F.; Urzì, C. Fungal colonization on treated and untreated stone surfaces. In Molecular Biology and Cultural Heritage, 1st ed.; Saiz-Jimenez, C., Ed.; Routledge: London, UK, 2003. [Google Scholar] [CrossRef]
  84. Chertov, O.; Gorbushina, A.A.; Deventer, D. A model for microcolonial fungi growth on rock surfaces. Ecol. Model. 2004, 177, 415–426. [Google Scholar] [CrossRef]
  85. Urzì, C.; Realini, M. Colour changes of Noto’s calcareous sandstone as related to its colonization by microorganisms. Int. Biodeterior. Biodegrad. 1998, 42, 45–54. [Google Scholar] [CrossRef]
  86. Favero-Longo, S.E.; Gazzano, C.; Girlanda, M.; Castelli, D.; Tretiach, M.; Baiocchi, C.; Piervittori, R. Physical and chemical deterioration of silicate and carbonate rocks by meristematic microcolonial fungi and endolithic lichens (Chaetothyriomycetidae). Geomicrobiol. J. 2011, 28, 732–744. [Google Scholar] [CrossRef]
  87. Jacobson, E.S. Pathogenic roles for fungal melanins. Clin. Microbiol. Rev. 2000, 13, 708–717. [Google Scholar] [CrossRef]
  88. Tonon, C.; Breitenbach, R.; Voigt, O.; Turci, F.; Gorbushina, A.A.; Favero-Longo, S.E. Hyphal morphology and substrate porosity -rather than melanization- drive penetration of black fungi into carbonate substrates. J. Cult. Herit. 2021, 48, 244–253. [Google Scholar] [CrossRef]
  89. Sterflinger, K.; Little, B.; Pinar, G.; Pinzari, F.; de los Rios, A.; Gu, J.-D. Future directions and challenges in biodeterioration research on historic materials and cultural properties. Int. Biodeterior. Biodegrad. 2018, 129, 10–12. [Google Scholar] [CrossRef]
  90. UNI. Beni Culturali—Materiali Lapidei Naturali Ed Artificiali—Descrizione della Forma di Alterazione—Termini e Definizioni. Available online: http://store.uni.com/catalogo/uni-11182-2006/ (accessed on 28 February 2022).
  91. ICOMOS-ISCS. Illustrated Glossary on Stone Deterioration Patterns. Available online: http://iscs.icomos.org/glossary.html (accessed on 28 February 2020).
  92. Urzì, C.; De Leo, F. Sampling with adhesive tape strips: An easy and rapid method to monitor microbial colonization on monument surfaces. J. Microbiol. Meth. 2001, 44, 1–11. [Google Scholar] [CrossRef]
  93. Ding, X.; Lan, W.; Gu, J.-D. A review on sampling techniques and analytical methods for microbiota of cultural properties and historical architecture. Appl. Sci. 2020, 10, 8099. [Google Scholar] [CrossRef]
  94. Quan, Y.; van den Ende, B.G.; Shi, D.; Prenafeta-Boldu, F.X.; Liu, Z.; Al-Hatmi, A.M.S.; Ahmed, S.A.; Verweij, P.E.; Kang, Y.; de Hoog, G.S. A comparison of isolation methods for black fungi degrading aromatic toxins. Mycopathologia 2019, 184, 653–660. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  95. Urzi, C.; Lisi, S.; Criseo, G.; Zagari, M. Comparazione di terreni per l’enumerazione e l’isolamento di funghi deteriogeni isolati da materiali naturali. Ann. Microb. Enzymol. 1992, 42, 185–193. [Google Scholar]
  96. Wiktor, V.; De Leo, F.; Urzì, C.; Guyonnet, R.; Grosseau, P.; Garcia-Diaz, E. Accelerated laboratory test to study fungal biodeterioration of cementitious matrix. Int. Biodeterior. Biodegrad. 2009, 63, 1061–1065. [Google Scholar] [CrossRef] [Green Version]
  97. Urzì, C.; De Leo, F. Evaluation of the efficiency of water-repellent and biocide compounds against microbial colonization of mortars. Int. Biodeterior. Biodegrad. 2007, 60, 25–34. [Google Scholar] [CrossRef]
  98. Sterflinger, K.; Piñar, G. Molecular-based techniques for the study of microbial communities in artworks. In Microorganisms in the Deterioration and Preservation of Cultural Heritage; Joseph, E., Ed.; Springer: Cham, Switzerland, 2022; pp. 59–77. [Google Scholar] [CrossRef]
  99. Marvasi, M.; Cavalieri, D.; Mastromei, G.; Casaccia, A.; Perito, B. Omics technologies for an in-depth investigation of biodeterioration of cultural heritage. Int. Biodeterior. Biodegrad. 2019, 144, 104736. [Google Scholar] [CrossRef]
  100. Li, Q.; Zhang, B.; He, Z.; Yang, X. Distribution and diversity of bacteria and fungi colonization in stone monuments analyzed by high-throughput sequencing. PLoS ONE 2016, 11, e0163287. [Google Scholar] [CrossRef] [Green Version]
  101. Cappitelli, F.; Cattò, C.; Villa, F. The control of cultural heritage microbial deterioration. Microorganisms 2020, 8, 1542. [Google Scholar] [CrossRef]
  102. Lo Schiavo, S.; De Leo, F.; Urzì, C. Present and future perspectives for biocides and antifouling products for stone-built cultural heritage: Ionic liquids as a challenging alternative. Appl. Sci. 2020, 10, 6568. [Google Scholar] [CrossRef]
  103. De Leo, F.; Jurado, V. Editorial for the special issue “Microbial Communities in Cultural Heritage and Their Control”. Appl. Sci. 2021, 11, 11411. [Google Scholar] [CrossRef]
  104. Martin-Sanchez, P.M.; Novákova, A.; Bastian, F.; Alabouvette, C.; Saiz-Jimenez, C. Use of biocides for the control of fungal outbreaks in subterranean environments: The case of the Lascaux Cave in France. Environ. Sci. Technol. 2012, 46, 3762–3770. [Google Scholar] [CrossRef] [PubMed]
  105. Cuzman, O.A.; Olmi, R.; Riminesi, C.; Tiano, P. Preliminary study on controlling black fungi dwelling on stone monuments by using a microwave heat system. Int. J. Cons. Sci. 2013, 4, 133–144. [Google Scholar]
  106. Gazzano, C.; Favero-Longo, S.E.; Iacomussi, P.; Piervittori, R. Biocidal effect of lichen secondary metabolites against rock-dwelling microcolonial fungi, cyanobacteria and green algae. Int. Biodeterior. Biodegrad. 2013, 84, 300–306. [Google Scholar] [CrossRef]
  107. Isola, D.; Bartoli, F.; Meloni, P.; Caneva, G.; Zucconi, L. Black fungi and stone heritage conservation: Ecological and metabolic assays for evaluating colonization potential and responses to traditional biocides. Appl. Sci. 2022, 12, 2038. [Google Scholar] [CrossRef]
  108. Chobba, M.B.; Weththimuni, M.L.; Messaoud, M.; Urzi, C.; Bouaziz, J.; De Leo, F.; Licchelli, M. Ag-TiO2/PDMS nanocomposite protective coatings: Synthesis, characterization, and use as a self-cleaning and antimicrobial agent. Prog. Org. Coat. 2021, 158, 106342. [Google Scholar] [CrossRef]
  109. De Leo, F.; Marchetta, A.; Capillo, G.; Germanà, A.; Primerano, P.; Schiavo, S.L.; Urzì, C. Surface active ionic liquids based coatings as subaerial anti-biofilms for stone built cultural heritage. Coatings 2021, 11, 26. [Google Scholar] [CrossRef]
Applsci 12 03969 g001 550
Figure 1. Main morphological characteristic traits of MicroColonial Fungi, MCF. Dark black colonies due to the melanin production as seen (a) for the unidentified strain MC 655 on different cultural media after 1 month of incubation. (b–d) characteristic meristematic pattern of growth described also as Torula-like hyphae observed under Light Microscope. Bar is 10 μm.
Figure 1. Main morphological characteristic traits of MicroColonial Fungi, MCF. Dark black colonies due to the melanin production as seen (a) for the unidentified strain MC 655 on different cultural media after 1 month of incubation. (b–d) characteristic meristematic pattern of growth described also as Torula-like hyphae observed under Light Microscope. Bar is 10 μm.
Applsci 12 03969 g001
Applsci 12 03969 g002 550
Figure 2. Close association between MCF and phototrophic microorganisms. (a) Association seen directly on a marble sample. Magnification 400X and (b) after growth in the isolation medium PDA: Chlorella-like alga and Coniosporium apollinis MC 728. Magnification 80X.
Figure 2. Close association between MCF and phototrophic microorganisms. (a) Association seen directly on a marble sample. Magnification 400X and (b) after growth in the isolation medium PDA: Chlorella-like alga and Coniosporium apollinis MC 728. Magnification 80X.
Applsci 12 03969 g002
Applsci 12 03969 g003 550
Figure 3. Phylogenetic tree (Neighbour-joining, Kimura two-parameters) showing the genetic divergence among ITS rDNA sequences of meristematic black fungi retrieved from GenBank database (https://www.ncbi.nlm.nih.gov/nucleotide accessed on 14 February 2022) and listed in Table 2.
Figure 3. Phylogenetic tree (Neighbour-joining, Kimura two-parameters) showing the genetic divergence among ITS rDNA sequences of meristematic black fungi retrieved from GenBank database (https://www.ncbi.nlm.nih.gov/nucleotide accessed on 14 February 2022) and listed in Table 2.
Applsci 12 03969 g003
Applsci 12 03969 g004 550
Figure 4. Extended black-greyish patina due to black fungi colonization on marble statue located in the inner yard of the Museum of Messina, Italy, under a tree of Pittosporum tobira. Fungal strains isolated from the statue were attributed to meristematic black fungi and genera of Cladosporium, Alternaria and Phoma [81].
Figure 4. Extended black-greyish patina due to black fungi colonization on marble statue located in the inner yard of the Museum of Messina, Italy, under a tree of Pittosporum tobira. Fungal strains isolated from the statue were attributed to meristematic black fungi and genera of Cladosporium, Alternaria and Phoma [81].
Applsci 12 03969 g004
Applsci 12 03969 g005 550
Figure 5. Microscopic observation of the endolithic behaviour of meristematic black fungi. (a) Settled on a thin section of Carrara marble sarcophagus flake showing the meristematic chains deepening inside the marble, magnification 400X; (b) SEM image showing MCF into the marble with dissolution pattern following the same shape of meristematic cells.
Figure 5. Microscopic observation of the endolithic behaviour of meristematic black fungi. (a) Settled on a thin section of Carrara marble sarcophagus flake showing the meristematic chains deepening inside the marble, magnification 400X; (b) SEM image showing MCF into the marble with dissolution pattern following the same shape of meristematic cells.
Applsci 12 03969 g005
Table
Table 1. Genera of black fungi isolated from stone monuments in the period from 1997–2022 in association with visible alterations.
Table 1. Genera of black fungi isolated from stone monuments in the period from 1997–2022 in association with visible alterations.
Class/OrderGenera *SubstrateEnvironmental and Climatic FeaturesAlterations Associated to Fungal ColonizationRefs
Dothideomycetes incertae sedisConiosporiumCalcarenite, granite, limestone, marbleMediterranean climate, urban environmentGrayish-black patina, pitting, black spots, greenish to dark green patina, crater shaped lesions, chipping, exfoliation, sugaring, crumbling, superficial deposit, and biofilm[37,39,43,44,45,46,47,48,49]
Dothideomycetes/Capnodiales incertae sedisCapnobotryellaLimestone, marble Mediterranean climate,
continental climate, and urban environment		Black spots, crater shaped lesions, chipping, exfoliation, sugaring, crumbling, pitting, superficial deposit, and biofilm formation[45,48,50,51,52]
ConstantinomycesSandstoneUrban environment, temperate climateDiscolorations, patina[53]
PseudotaeniolinaMarble, sandstone Mediterranean climate, arid and desert climateBiological green patina[54,55,56]
Dothideomycetes/CapnodialesAeminiumLimestoneTemperate climateBlack discoloration with salt efflorescence[57]
Dothideomycetes/CladosporialesCladosporiumCalcarenite, granite, limestone, marble, plaster, sandstone, tufaUbiquitous worldwide distribution in indoor environments and outdoorDark alterations, black spots, black patinas, detachment of marble grains, light grayish patina, crater shaped lesions, chipping, exfoliation, sugaring, crumbling, pitting, superficial deposit, biofilm, black crusts, green biofilm with salt efflorescence, stone erosion and disintegration, and discoloration[27,40,46,48,49,58,59,60,61,62,63,64,65,66,67]
VerrucocladosporiumLimestone, marble, sandstoneMediterranean climate, temperate climate, and urban environmentBlack patina, discoloration[37,53]
Dothideomycetes/DothidealesAureobasidiumGranite, limestone, marble,
plaster, sandstone		Urban environment, Mediterranean climate, temperate climate, indoor environment, and urban environmentBlack patina, black spots, detachments, superficial deposit, biofilm, discolorations with or without salt efflorescence, black crusts, and stone erosion and disintegration[37,40,45,49,53,63,64,65,68]
Dothideomycetes/MycosphaerellalesSalinomycesMarble, sandstoneMediterranean climateBlack patina[37]
NeocatenulostromaLimestone, sandstoneTemperate climate, urban environmentDiscolorations and/or patina, structural damage[53]
NeodevresiaLimestone, marble, plaster, tufaMediterranean climateBlack patina, discolorations, structural damage[37,53,55,63]
SaxophilaMarbleMediterranean climateBlack patina[37]
VermiconidiaLimestone, marble, travertineMediterranean climate, urban environmentBlack patina[37]
Dothideomycetes/NeophaeothecalesNeophaeothecaMarbleMediterranean climateBlack patina[37]
Dothideomycetes/PleosporalesAlternariaCalcarenite, granite, limestone, marble, plaster, tufaUbiquitous worldwide distribution in indoor environments and outdoorBlack spots, black patina, detachment of marble grains, greenish to dark green patina, biofilm, black crusts, green-black patina; and blackish patina[40,46,49,58,59,60,63,64,66,67]
EpicoccumGranite, limestone, marbleUrban environment, mediterranean climate, and temperate climateBlack spots, black patinas, detachment, superficial deposit, biofilm, blackish patina, green biofilm, and dark and green biofilm with salt efflorescence[40,45,49,60,64]
PhomaCalcarenite, granite, limestone, marble, plaster, tufaMediterranean climate, temperate climate, urban environment, continental-cold climate, and indoor and outdoor environmentsBlack spots, black patinas, detachment of marble grains; color changes, crater shaped lesions, chipping and exfoliation, sugaring, crumbling, pitting, superficial deposit, biofilm, and black crusts[40,46,48,49,58,63]
Dothideomycetes/VenturialesOchroconisCalcareniteSubterranean environmentBlack patina[69]
Eurotiomycetes incertae sedisSarcinomycesMarbleMediterranean climateBlack spots[70]
Eurotiomycetes/ChaetothyrialesCyphellophora sp.PlasterMediterranean climateBlack/grayish patina[63]
ExophialaCalcarenite, limestone, marble,
sandstone		Mediterranean climate, urban environment, temperate climate, and hypogean environmentDark alterations, black spots, black patinas, detachment of marble grains, discolorations, and visible structural damage[27,37,40,45,53,71]
LithophilaLimestone, marbleMediterranean climate, urban environment, and
dry continental climate		Black spots, black patinas, detachment of marble grains[37,40,72]
KnufiaLimestone, marble, sandstone
travertine		Mediterranean climate, urban environment, continental temperate climate, and dry continental climateBlack and grey spots, dark macropitting, biopitting, crater shaped lesions, chipping, exfoliation, sugaring, crumbling, discolorations, patina, and visible structural damage[37,41,43,45,48,53,72,73,74]
RhinocladiellaMarbleMediterranean climateBlack spots, crater shaped lesions, chipping and exfoliation, sugaring, crumbling, and pitting[48]
Eurotiomycetes/MycocalicialesMycocaliciumMarbleMediterranean climate, urban environmentBlack spots, crater shaped lesions, chipping and exfoliation, sugaring, crumbling, and pitting[45,48]
* According to the current taxonomic nomenclature.
Table
Table 2. ITS rDNA sequences of representative MCF isolated from stone monuments aligned in Figure 3.
Table 2. ITS rDNA sequences of representative MCF isolated from stone monuments aligned in Figure 3.
TaxonStrainITS rDNA
Capnobotryella antalyensisMA 4615AJ972858
Capnobotryella antalyensisMA 4624AJ972850
Capnobotryella antalyensisMA 4766AJ972851
Capnobotryella antalyensisMA 4775AJ972860
Capnobotryella isiloguiMA 3619AM746201
Capnobotryella erdoganiMA 4625AJ972857
Capnobotryella kirizogluiMA 4899AJ972859
Capnobotryella sp.MA 4701AJ972856
Capnobotryella sp.MA 4697AJ972855
Capnobotryella sp.MA 3615AM746203
Neodevriesia modestaCCFEE 5672KF309984
Neodevriesia simplexCCFEE 5681KF309985
Neodevriesia sardiniaeCCFEE 6202KP791765
Neodevriesia sardiniaeCCFEE 6210KP791766
Saxophila tyrrhenicaCCFEE 5935KP791764
Aeminium ludgeriE12MG938054
Aeminium ludgeriE16MG938061
Neocatenulostroma sp.CR1KY111907
Constantinomyces sp.CR21KY111911
Pseudaeniolina globosaDPS10MH396690
Pseudotaeniolina globosaCBS109889NR136960
Pseudotaeniolina globosaCCFEE5734KF309976
Vermiconidia calcicolaCBS 140080NR_145012
Vermiconidia calcicolaCCFEE 5780KP791761
Vermiconidia flagransCCFEE 5922KP791753
Coniosporium uncinatumCBS 100219AJ244270
Coniosporium apollinisCBS 100213AJ244271
Coniosporium apollinisCBS 352.97NR159787
Coniosporium apollinisCBS 100216AJ244272
Coniosporium apollinisQIIIaMH023395
Lithophila catenulataBJ10118JN650519
Lithophila guttulataM1MW361305
Lithophila guttulataCCFEE 5884KP791768
Lithophila guttulataCCFEE 5907KP791773
Knufia mediterraneaCCFEE 5738KP791791
Knufia mediterraneaCCFEE 6211KP791793
Knufia vaticaniiCCFEE 5939KP791780
Knufia calcarecolaSL11033JQ354925
Knufia calcarecolaCGMCC 3.17222KP174862
Knufia marmoricolaCCFEE 5895KP791775
Knufia marmoricolaCCFEE 5716KP791786
Knufia perforansCBS 885.95AJ244230
Knufia karalitanaCCFEE 5732KP791782
Knufia karalitanaCCFEE 5929KP791783
Knufia petricolaCCFEE 726.95KC978746
Knufia petricolaCBS 600.93KC978744
Knufia petricolaIMI38917AJ507323
Knufia petricolaD1JF749183
Knufia petricolaM4FJ556910
Knufia sp.QIIaMH023393
Knufia sp.QIIbMH023394
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

De Leo, F.; Marchetta, A.; Urzì, C. Black Fungi on Stone-Built Heritage: Current Knowledge and Future Outlook. Appl. Sci. 2022, 12, 3969. https://doi.org/10.3390/app12083969

AMA Style

De Leo F, Marchetta A, Urzì C. Black Fungi on Stone-Built Heritage: Current Knowledge and Future Outlook. Applied Sciences. 2022; 12(8):3969. https://doi.org/10.3390/app12083969

Chicago/Turabian Style

De Leo, Filomena, Alessia Marchetta, and Clara Urzì. 2022. "Black Fungi on Stone-Built Heritage: Current Knowledge and Future Outlook" Applied Sciences 12, no. 8: 3969. https://doi.org/10.3390/app12083969

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop