Introduction
•Biodeterioration, bioalternation, biocorrosion, biodecay
•Biodeteriogen
•Building stone
•Cyanobacterial and algal growths
•The habitat concept
•Floristic studies
•Taxonomy
•Ecology and autecology
•Assemblages
•Biodeterioration of stone associated with cyanobacteria and algae
•Remarks on control and removal of algae
•
1

Biodeterioration
•(Bio)alternation, (bio)corrosion, (bio)deterioration, (bio)decay
•
•An exchange of material and energy
•
•Two heterogenous open dynamic system: The solid substrate and its atmospheric (indoor and
autdoor), aquatic or chthonic environment
•
•The natural limit of turnover activities is given by the penetration depths and speeds of physical
gradients, gases, solutions, and organisms (including their extracellular products) into mineral
material.
2

Biodeteriogen
•We could not debar any organism from being an actual or potential biodeteriogen because of no
special criteria for biodeteriogen.
•
•Therefore the range of potential biodeteriogens is huge.
•
•The deteriogenic effects are mainly a minor incidental part of their activities.
•
•The biodeteriogenic organism usually inhabit secondarily an artificial man-made surface and
participate on cycling of elements (Winkler, 1994).
3

Building Stone
•Stone is the oldest entity of Earth
•
•Characterised by a wide range of mineral compositions, textures, and rock structures
•
•Creates ecosystem which includes also environmental factors (light, nutrition, climate, pollution)
•
•Significant deterioration of monuments has been begun since years 1870-1880, this phenomenon has
not been possible to stop since 1950
4

Cyanobacterial and algal growths
•Primarily concerned with the cyanobacteria and algae growing on the exposed surfaces of buildings
constructed of man-made materials
•
•Special consideration is given to those obscuring coloured stains on building stone
•
•This account includes the tasks caused by them and the method used for their investigation,
control and elimination
5

The Habitat Concept
•Most of the cyanobacteria and algae are ubiquitous
•
•Cyanobacterial and algal taxa growing on man-made surfaces
•
•Exactly, it is difficult to define the basis of habitat
•
•Growing in non-aquatic environments have been termed terrestrial, aerial or subaerial forms
•
•Man-made subaerial environment form stressing factors - temperature, dessication, high or low pH,
toxicants (pollutants), high or very low irradiance
6

Floristic Studies
•More than 200 algal genera had been recorded world wide in floristic studies
•
•The investigation of colonisations of stone by algae
•
•Most species lists include a number of unidentified organisms
•
•Difficult to make exact comparisons of the floristic data
•
•Differences in the homogeneity of the surface sampled, the intensity of sampling
•
•The experiences of collector
•
•The method used for the identification of samples, inconsistencies in the taxonomy of some groups,
and in the competence and special interests of the phycologist identifying the algae
7

Taxonomy
•subaerial cyanobacteria and algae are of a simple organization, exhibit a simple unicellular or
filamentous habit
•
•Cyanobacteria and green algae dominate on subaerial habitat
•
•No systematic effort has been made to distinguish which genera are truly subaerial
•
•Many taxonomic problems
8

Ecology and Autecology
•Main question are the adaptations to the subaerial environment
•
•The capacity to tolerance desiccation is a feature of many cyanobacteria and algae of both aquatic
and terrestrial origin
•
•The most fundamental feature of all truly aubaerial algae is the ability to resist extremes of
temperature, high light intensity and considerable water loss
•
•Many opportunists in subaerial habitats, colonising and growing very fast over wetter or aquatic
periods
9

Assemblages
•In subaerial habitats some cyanobacteria and algae were typically much more abundant than others
•
•The assemblages were named by using the names of the most abundant forms: the blue-green
assemblage, the Trentepohlia assemblage, Desmococcus assemblage, Prasiola assemblage, Stichococcus
assemblage
10

Biodeterioration of stone associated
with cyanobacteria and algae
•Excessive biofilm accumulation in porous substrata changes or reduces their properties or
effectiveness
•
•The pedogenic significance of cyanobacteria and algae is very important by biogeophysical
weathering of stone surfaces
•
•Mechanical damage caused by active penetration of euendoliths and chemical damage caused by
substances and products of metabolism and of extracellular matrix
•
•Aesthetic biodeterioration or soiling
•
11

Remarks on control and removal
of algae
•Indirect methods
•Reduction of humidity, firing, cover, isolation...
•Direct methods
•Eliminate crusts, patinas; UV-rays, X-rays, biocide chemicals...
12

The Aims
•Floristic screening and taxonomic study of microalgae causing the biological attack on building
stone;
•
•Which factor of urban environment (substrata, pollution, humidity, etc.) have an influence on the
composition of the subaerial taxa;
•
•To set a growth inhibition test on subaerial microalgae;
•
•To test the microalgae causing actively biodeterioration which potentially damage the building
stone.
13

Individual studies
forming the results
•Gravestones in Cemetery
•Tombstone in a historic cemetery
•Subterranean Jewish cemetery
•The walls of building in Bratislava (SK)
•The walls of building in Murcia (ESP)
•The walls of building in Brno (CZ)
•Growth inhibition test on subaerial microalgae
14

Materials and methods
•Research area
•The Bratislava City
•The Murcia Region
•The Brno Region
•Sampling
•Laboratory cultivation
•Analysis for determination
•Ultrastructural analysis (SEM)
15

Research area
•Bratislava - capital city is located in West Slovakia (48°10´N, 17°10´E)
•Murcia - This region is located in South-east Spain (the Murcia City - 36°56´N, 1°07´W)
•Brno - city in south Moravia eastern part of Czech Republic
16

Sampling
•Different substrata - horizontal distribution
•One substrata - vertical distribution
•North and south exposition on one building
•Spatial distribution in a city
•
•
17

Sampling Schema
DomSvMaritna PresbyteriumSever Mramor Hrubozrnny
Monument
Sampling Place
Detail of Substrate
1st Cultivation
Culture
Mikroskop
17a

In Laboratory
LabakEspana ExperimentEspana Flowbox DSCN0531 Dscn0535
Flowbox
Isolation
Experiment
Cultivation
Consultation
17b

Cintorin Kozia Brana Litavsky Vapenec Cintorin Kozia Brana 2 Travertin Pieskovec s Vapnitym tmelom
Vápnitý Pieskovec
Figs 3-8. Kozia brána Cemetery: Figs. 3-4. Landform of the Kozia brána Cemetery; Fig. 3. The
southern part of the cemetery; Fig. 4. Near the entrance; Fig. 5. Litava limestone; Fig. 6.
Travertine; Fig. 7. Sandstone with calcareous cement; Fig. 8. Calcarenite.
3
4
5
6
7
8
18
Stone Substrata

Ryolit 2 Tehla Granodiorit Cierny Dolerit 1 Cierny Dolerit 2 Neogenny Pieskovec 2
9
10
11
12
13
14
Figs 9-14. Kozia brána Cemetery: Fig. 9. Black dolerite – smooth surface; Fig. 10. Black dolerite –
ragged surface; Fig. 11. Neogene calcarenite; Fig. 12. Brick; Fig. 13. Ryolite; Fig. 14.
Granodiorite.
19
Stone Substrata

Hluznaty Vapenec - Tata Madarsko Mramor Hrubozrnny
15
16
17
18
19
Figs 15-19. Figs 15-16. Kozia brána Cemetery: Fig. 15. Pink nodular limestone from Tata (Hungary);
Fig. 16. Marble. Figs. 17-19. Chatam Sófer Mausoleum: Fig. 17. Concrete shall of the Mausoleum
before total reconstruction (arrow); Fig. 18. Gravestones on the soil; Fig. 19. Central part,
Chatam Sófer’s sarcophagus.
20
Underground

DomSvMaritna PresbyteriumSever DSCN0893 DSCN0924
20
21
22
23
Figs 20-23. Figs 20-21. St. Martin’s Cathedral in Bratislava, Slovakia: Fig. 20. Northern part of
Cathedral; Fig. 21. Socle part of Presbyterium. Figs. 22-23. Islamic Castle Monteagudo in Region of
Murcia, Spain: Fig. 22. The NW view of the Castle; Fig. 23. The central walls of the Castle.
21
Historical Monuments

Mother and two doughters P1010045 Detail Doughter with book Mother with Doughter Left
Figs 24-27. Tombstone (statue represent mother with two doughters) and tomb cover in the historic
cemetery of Pressburg Evangelicals Kozia brána in the centre of Bratislava. Fig. 24. Sampling
places of algae (1-14) in different microhabitat zones (A-E); Fig. 25. Tomb cover with growths of
liverworts and mosses (arrows); Figs 26-27. Detail views on weathered parts of statue (arrows).
24
25
26
27
Nakres Socha
22
Tombstone

Laboratory Cultivation
•Zehnder medium (Zehnder in Staub, 1961)
•BG11 medium (Rippka et al., 1979)
•BG110 (Rippka, 1988)
•BBM (Smith & Bold, 1966)
•either liquid or agarised at 20-22 °C
•The unialgal strains were used for detail taxonomic study to obtain the data of whole life cycle
23

Analysis for Determination
•Type of stone substrata
•10 scores for each stone sample
•Structure of algal assemblage
•The phenotype features
•Original drawings made by camera lucida from field and from cultured material
•Olympus BH2 Photomicrography system and Nikon DXM1200 (F) colour camera by LUCIA imaging system
(Y/C and composite) – image analysis software developed for image capture, archiving and analysis
(© 2004 Laboratory Imaging s.r.o.)
•
•
24

Ultrastructural Analysis (SEM)
•Pieces of stone substrata were taken from outer walls of monuments and buildings in urban sites.
•
•Samples were fixed, dehydrated, coated with gold as described previously (Garty and Delarea, 1987)
•
•Examined by a JEOL JXA 840A scanning electron microscope (SEM) operating at 200 kV
25

General Results and Discussion
•Cca 115 taxa of microalgae
•The competitive „first come, first serve“ relationships
•Cyanobacteria 35%
•Chlorophytes 42%
•Chrysophytes 17%
•Charophytes 8%
•The north-facing or humid side of buildings
•
•
26

Chroococcidiopsis umbratilis Leptolyngbya nostocorum1JSP4 CHS98-48 CHS98-35d 131 84
28
Figs 28-33. Cyanobacteria: Fig. 28. Chroococcidiopsis umbratilis, a vegetative cells, b
endosporangium; Fig. 29. Chroococcidiopsis sp.; Fig. 30. Leptolyngbya nostocorum, a false
branching; Fig. 31. L. fragilis; Fig. 32. Nostoc sphaericum; Fig. 33. N. microscopicum
31
29
30
32
33
a
b
a
20 µm
20 µm
20 µm
20 µm
40 µm
40 µm
27
Cyanobacteria

Porphyridium7(100x) Porphyridium9(100x) 144 111 113 164
36
34
35
37
38
39
Figs 34-39. Fig. 34. Cyanobacterium: Scytonema julianum, a apical globular cell ; Fig. 35-36.
Rhodophyta (red algae): Fig. 35. The colony of Porphyridium purpureum, a cell division; Fig. 36.
Solitary globular cells of P. purpureum; Fig. 37-39. Heterokontophyta: Fig. 37. One-celled
representatives of Xanthonema sp.; Fig. 38. Representatives of Xanthonema sp., a one-celled, b
two-celled, c four-celled; Fig. 39. Fragmentation of filament of Xanthonema sp., a H-particles, b
unicelled fragments of filament
c
a
b
a
b
a
a
20 µm
40 µm
40 µm
20 µm
10 µm
10 µm
28
Rhodophyta and Heterokontophyta

Figs 40-45. Figs 40-44. Chlorophyta: Fig. 40. Chlorosarcinopsis  cf. arenicola; Fig. 41.
Tetracystis sarcinalis, a globular zoosporangia with tetrades of zoospores, b zoospores, c
pyrenoid; Fig. 42. Oocystis assymmetrica, a vegetative cells, b berry-like colonies; Fig. 43.
Chlorosarcinopsis pseudominor; Fig. 44. Scenedesmus obliquus, unicelled stage; Fig. 45. Charophyta:
Fig. 45. Stichococcus bacillaris.
25 173 156 CHS98-18a CHS98-12
42
40
41
43
45
a
b
c
a
b
CHS98-9b
44
40 µm
20 µm
20 µm
40 µm
40 µm
20µm
29
Chlorophyta

CHS98-103r CHS98_103v20x CHS98-103a CHS98-103f CHS98-103t CHS98-103q
48
46
47
49
50
51
Figs 46-51. Chlorophyta – Kentrosphaera gibberosa var. gibberosa:Fig. 46. Autospores with parietal
chloroplast; Fig. 47. Young vegetative cells with parietal and central chloroplast; Fig. 48. Adult
vegetative cells with central chloroplast, a cell wall protrusion; Fig. 49. Oval adult vegetative
cell, a cell wall protrusion; Fig. 50. Empty autosporagium, a autosporangium wall, b autospores;
Fig. 51. Old vegetative cell.
a
a
a
b
40 µm
40 µm
40 µm
40 µm
40 µm
40 µm
30
Chlorophyta

CHS98-20e
52
Figs 52-57. Charophyta: Fig. 52-54. Klebsormidium flaccidum, Fig. 52. Stright filamment with
parietal chloroplasts; Fig. 53. The protruding zoosporangial cells in asexual filament (arrow);
Fig. 54. Empty zoosporangial cells with fissured surface (arrows); Fig. 55-57. Klebsormidium
crenulatum, Fig. 55. Maturing filaments with short cells (arrow); Fig. 56. Begining of
pseudobiseriating in filament (arrow); Fig. 57. Pattern of fragmentation of filament.
Klebsormidium3SSP3 Klebsormidium2JSP Klebsormidium4JSP10 Klebsormidium4JSP8 Klebsormidium4JSP5
53
54
55
56
57
40 µm
40 µm
40 µm
40 µm
40 µm
40 µm
31
Charophyta

Figs 58-66. Life history of Kentrosphaera gibberosa var. gibberosa (orig. Uher B.). Figs 58-60.
Young vegetative cells; Figs 61-63. Adult vegetative cells; Fig. 64. Zoosporangium; Fig. 65.
Authosporangium; Fig. 66. Authospores.
58
59
60
61
62
63
64
65
66
32
Life history

Biofilm cyanobakterii na tehle Biofilm na mramore Biofilm tvoriaci rozsievky a zelena vlaknita
riasa Klebsormidium Endospory cyanobakterie Chroococcidiopsis Rozsievka Diadesmis - detail Povrch
cierneho doleritu Povrch granodioritu 2 MRAMOR___STRUKTURA_3
67
68
69
70
71
72
73
74
Figs 67-74. SEM-micrographs. Fig. 67. Surface of dolerite, Ca-felspars (arrows); Fig. 68. Surface
of granodiorite, Na,K-felspars; Fig. 69. Surface of marble; Fig. 70. Diatom Diadesmis sp.; Fig. 71.
Biofilm on marble surface, a bacterial layer, b cyanobacterial cells of Chroococcidiopsis  sp.;
Fig. 72. Cyanobacterial biofilm on brick, filaments of Leptolyngbya sp. (arrows); Fig. 73.
Endospores of Chroococcidiopsis sp., endolithic cyanobacterium (arrows); Fig. 74. Biofilm on the
brick, a filament of klebsormidium flaccidum, b cells of diatom Diadesmis sp.
a
b
a
b
33
SEM-micrographs

Original drawings
•Life cycle
•Vegetative stages
•Generative stages
•Young habit
•Adult habit
•Old habit
34

Coccal Cyanobacteria
01
35

Trichal Cyanobacteria
04
36

Heterokontophytes
10
37

Green Algae
17
38

Charophytes
18
39

The New Growth Inhibition Test
•To develop the techniques of new growth inhibition test
•
•The biotest used on subaerial phototrophic microorganisms (autochtonous, isolated from
deteriorated monuments)
•
•A type of growth inhibitor was used white powder called in Spain “el estuco”
40

Experiment Scheme
experimentPowderFinal
41

Experiment Realisation
42
Experimentgray

The View of The Realisation
EL ESTUCO RESULTS
43

Results
•This material Calcium Ruthenium Oxide Hydrate totally inhibited and eradicated growths of algae
•
•Active interests in developing miniaturized algal bioassays, algal biosensors and development of
cultivation tests
•
•Work on subaerial algae caused biodeterioration has highlighted their practical potencial
44

Acknowledgement
•Financial support for this study was provided by the Funding of Ministry of Education of Czech
Republic; project No. MSM0021622416
•
•Prof. Dr. František Hindák
•
•Prof. Dr. Marina Aboal Sanjurjo
•
•Dr. Ľubomír Kováčik

Thank you
for your attention!