O R I G I N A L A R T I C L E

Accelerated testing of biological stain growth on external
concrete walls. Part 1: Development of the growth tests

Gilles Escadeillas Æ Alexandra Bertron Æ
Philippe Blanc Æ Arnaud Dubosc

Received: 27 March 2006 / Accepted: 16 October 2006 / Published online: 13 December 2006

RILEM 2006

Abstract

The development of biological stains has

a great influence on the durability of building
materials. For common concrete structures, the
main and most rapid disorder linked with this
development is aesthetic. In recent years, architects
have been increasingly using formwork surfaces for
external walls, so the search for aesthetic quality and
durability has become as important as the search for
mechanical quality and durability. Hence, there is a
demand from industry for the qualification of
concrete wall surface behaviour toward biological
growths. This paper aims to itemize the various
biological stains affecting concrete and to put
forward two accelerated tests for the growth of
algae, the organisms responsible for the first visible
stains. These tests enable a wall surface to be
qualified with respect to biological stains.

Keywords

Concrete

Algae
Biological stain

Laboratory tests

1 Introduction

The development of biological stains affects the
durability of many constructions [

1

]. The most

harmful degradation is due to heterotrophic
micro-organisms such as bacteria or fungi, which
massively excrete organic acids [

2

–

5

]. The degra-

dation may also be linked with direct or indirect
physical actions [

6

,

7

].

The greatest loss of durability for external wall

surfaces, in particular for concrete walls, concerns
aesthetic degradation. As architects are now
increasingly favouring formwork concrete for
wall surface frontage finishing, it is necessary for
concrete industry professionals and clients to
think seriously about the impact of concrete
formulation and casting on micro-organism pro-
liferation. Unfortunately, to our knowledge, no
accelerated test exists at the moment to qualify
the aesthetic evolution of external wall surfaces
exposed to biological stains, except for a few tests
developed for other purposes [

8

].

In response to this problem, laboratory accel-

erated tests of concrete wall surface biological
ageing have been developed. The first part of this
publication aims to specify the nature of biolog-
ical stains through a literature review and to
describe the tests developed. The second part will
concern the quantification of organic growths on
external wall surfaces. The results of a study
performed on mortars will also be presented.

G. Escadeillas

A. Bertron (

&)
A. Dubosc

Laboratoire Mate´riaux et Durabilite´ des
Constructions - INSA UPS Toulouse, 135 avenue de
Rangueil, Toulouse Cedex 4 31077, France
e-mail: [e-mail]

P. Blanc
Laboratoire Biotechnologie Bioproce´de´s - INSA,
Toulouse, France

Materials and Structures (2007) 40:1061–1071
DOI 10.1617/s11527-006-9205-x

2 Literature review

Biological stains result from the growth of
particular micro-organisms that may be found
on any building situated in polluted or non-
polluted areas anywhere in the world. Traces
have various colours (black, green, red, etc.) and
may spread over the whole frontage [

9

]. It

generally takes a year for stains to appear on
walls but, with favourable growth conditions,
development may be extremely rapid [

10

].

2.1 Colonisation of an external wall surface

Through ageing, external concrete wall surfaces
may be colonised successively by:

– bacteria: pioneer micro-organisms colonizing

every support [

11

]. However, they form invisible

biofilms and are not responsible for observed
discolouration (but they may degrade concrete);

– algae: micro-organisms that do not need an

organic supply for their synthesis (autotrophic
organisms) and may therefore develop on
exclusively mineral supports, such as concrete,
providing that the moisture level is sufficient.
Differently coloured (black, green or red)
stains, depending on the species, form along
water runoff, [

12

];

– fungi: micro-organisms that need a constant

organic supply, generally provided by the
support (heterotrophic organisms). Theoreti-
cally, they cannot be pioneer organisms. Fun-
gus is encountered either on organic supports
(e.g. paint), which are degraded by the meta-
bolic organic acids of the fungus, or on living or
dead organisms [

13

]. Very high moisture levels

are also required;

– lichens and mosses: lichens result from the

symbiosis between a fungus and an alga, the
fungus providing the moisture to the alga,
which gives back organic materials [

14

].

Mosses, which are classified among the lower
plants, generally develop in pads and lie on an
algal biological layer several years old [

10

].

So it clearly appears that microscopic algae are

the pioneer organisms responsible for the first
visible stains on the surfaces of external concrete
walls.

2.2 Algal species found on external wall

surfaces

In England, Whitely [

15

] noted unsightly reddish

stains composed of microscopic algae (chlorophy-
ceae Trentepohlia) and linked with high humidity.
In Asia, Wee et al. [

16

] also identified chlorophy-

ceae Trentepohlia on concrete showing reddish
stains and cyanophyceae in black streaks. In
France, Robic [

17

] showed that several concrete

wall stains in the city of Brest were caused by
microscopic algae: cyanopheae forming black
covering growth, and chlorophyceae causing
green and red stains.

The same species are encountered on every

continent and on other kinds of mineral
supports such as stone [

7

] or brick [

18

] but

always in conditions of high humidity. The
genera encountered are mainly Phormidium,
Gloeocapsa, Chroococcus, Lyngbya and An-
abaena for the cyanophyceae, and Chlorho-
rmidium, Trentepohlia and Chlorella for the
chlorophyceae.

There are two similarities between these spe-

cies. Firstly, they contain chlorophyll, allowing
internal photosynthesis to take place, which
provides energy supplies and ensures carbonic
nutrition (using daylight and atmospheric carbon
in particular). Secondly, their humidity require-
ments are high.

Cyanophyceae, also called blue-green algae

because of the colour of their cells, can be
unicellular, surrounded by a sheath or filamen-
tous. They are responsible for black stains and
have features that account for their ability to
colonize

concrete

wall

surfaces.

They

are

equipped with accessory blue and red pigments
allowing them to exploit very low light intensities
and protecting them against intense solar radia-
tion. Their very hygroscopic mucilage membrane
retains rainwater or seepage thanks to the min-
erals it contains. This membrane is a protection
against desiccation during drought.

Chlorophyceae, green coloured, are devoid of

accessory pigments, coloration or protective
mucilage and can only live in environments with
constant humidity and adequate—but not too
intense—light. In these favourable conditions, a
wall may turn green in a few days.

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Materials and Structures (2007) 40:1061–1071

2.3 Factors influencing algal growth

on concrete

These factors are classified in three categories:

– biological factors: linked with the accessibility

of the walls to the various species [

19

] and with

the competition between these species. Con-
cerning the accessibility, algae are carried by
the wind and by run-off waters. The competi-
tion between species is linked with environ-
mental conditions (some chlorophyceae stains
gradually give way to cyanophyceae in winter
with the coming of drier periods [

15

]);

– environmental factors: essentially humidity,

temperature, luminosity and nutritive supplies.

Humidity on an external wall surface must reach
a certain threshold for algae to develop [

15

,

18

].

The different moisture sources are relative
humidity, which favours chlorophyceae growth
[

16

], water flow, often linked with design faults

and which favours cyanophyceae propagation on
the support [

10

], and rain and wind, which lead to

stain development on the sides the building
facing prevailing winds [

20

]. The support also has

an influence through its porosity (water reten-
tion in the pores after bad weather or capillary
ascent on parts of walls close to the soil).
Temperature influences growth speed (the opti-
mal growth temperature is about 23C [

21

]) but it

is not as important a factor as humidity (cover
may be greater in winter). Light is necessary for
the photosynthesis reaction (a total light lack acts
as an inhibitor). Moreover, the species present
may vary according to the wall surface exposure
(Trentepohlia chlorophyceae prefer sunlit wall
surfaces [

22

]). However, overlong or too strong

sunshine tends to be an inhibitor (too high a light
intensity and rapid drying of the wall).
The nutritive supplies necessary to algae are
nitrogen, phosphorus, sulphur, etc. These ele-
ments, provided in their mineral form, may
come from the support (calcicole algae), from
run-off waters or from ashes due to pollution [

7

].

– substrate factors: those which enable the micro-

organisms to establish themselves and to survive
on whatever substrate are porosity, roughness
and mineral composition of the support [

18

].

Roughness influences the adhesion of cells

brought by wind or conveyed by a water flow on a
surface wall (algae tend to develop better on
rough supports [

16

]).

The porosity of the support influences the quan-

tity of water absorbed and retained after bad weather
or any humidification (algal growth on construction
stones is proportional to their porosity [

7

]).

The surface mineral composition plays a role

since it may either be used as nutriments by algae
(silicon for diatoms for instance [

23

]) or, on the

contrary, prevent their development (the pH of
concrete before its carbonation [

12

,

13

] inhibits

algal growth, as do some products with biocidal
effect or with catalytic action like TiO

2

).

3 Development of laboratory accelerated tests

Laboratory tests aiming to study algal growth on
wall surfaces have to be as realistic as possible.
But, at the same time, these tests are required to
be accelerated, reproducible, low cost and easy to
implement in construction material laboratories.
They also have to discriminate among the support
parameters for biological growth.

For these reasons, two tests were developed [

24

]:

– a static test simulating growth conditions at the

base of a construction (humidity was provided
by water capillary ascent, the species accessi-
bility was not a limiting factor);

– a dynamic test simulating run-off on some parts of

constructions (this test corresponds to external
wall surfaces exposed to ‘‘bad weather’’, or leaky
parts of a building or design defects). The species
accessibility may be a limiting factor here (algae
have to hang from the support before developing).

The common part of the preparation stage of

the two tests is presented in section 3.1 and then
the specific aspects proper to each test are given
in the following sections.

3.1 Common preparation for growth tests on

cementitious support

3.1.1 Biological factors

a. Choice of algal species

Algae were chosen

from species belonging to both cyanophyceae and
chlorophyceae. The selection criteria were:

Materials and Structures (2007) 40:1061–1071

1063

– representativeness: the chosen species were

regularly quoted in the literature;

– growth rapidity: the selected species were able

to develop rapidly in favourable conditions;

– ease of liquid culture: the selected algal species

were easy to homogenise in liquid culture,
which allowed the quantity inoculated into
supports to be controlled.

Eight species were pre-selected. The selection

test consisted of a humidification simulation—in
the presence of algae—by capillary ascent of a
previously carbonated cementitious support. This
test corresponded to a very favourable real case
of colonisation.

The species studied and the results are pre-

sented in Table

1

.

Considering the results, the Chroococcidiopsis,

Chlorella and Chlorhormidium genera were rep-
resentative of the two species and were therefore
chosen.

b. Preparation of algae before inoculation

The

chosen species were ordered from a culture
collection and transported in the form of liquid
cultures in which their growth was slowed down
(preservation liquid cultures). On reception at the
laboratory, the cultures were first kept in a purely
mineral medium (BG11 medium see composition
in Table

2

) at a temperature between 15 and

20C, under low illumination (level less than 1000
lux) with an 8-h day and 16-h night photoperiod.

Still in the BG11 medium, the algae were then

placed in conditions allowing them to start active
growth again (inoculation liquid cultures) so that
they could be inoculated in an exponential growth
phase [

25

].

The preparation method was the following

(one culture per species):

– removal of 5 ml of the preservation liquid

culture;

– inoculation of the 5 ml into flasks containing

100 ml of BG11;

– culture at a temperature of 23C, under an

intensity of illumination of 1600 lux, a photo-
period of 16 h days and 8 h nights, and with
constant stirring (better illumination condi-
tions);

– growth quantification every week with an

appropriate technique.

The total period before inoculation (beginning

of the exponential growth phase) was generally
35 days.

c. Algae quantification before inoculation

The

methods for quantifying algal growth in liquid
culture are the measurement of chlorophyll a (or
chl a) and of dry biomass together with
nephelometry. Their principles, advantages and
drawbacks are described below:

– the chlorophyll assay consists in measuring of

the characteristic absorption peak height by
spectrophotometry of chl a at 665 nm after it
has been dissolved in ethanol. The main
advantage of this technique is that it also

Table 1 Description of pre-selected species and results of selection tests

Class

Genus

Type

Stain colour

Liquid culture

Growth

Cyanophyceae

Chroococcidiopsis

Unicellular

Black

Easy

Very strong

Lyngbya

Filamentous

Black

Difficult

Low

Nostoc

Filamentous

Black

Difficult

Nil

Chlorophyceae

Chlorella

Unicellular

Green

Easy

Very strong

Chlorhormidium

Filamentous

Green

Easy

Very strong

Stichococcus

Filamentous

Green

Easy

Very strong

Haematococcus

Unicellular

Red

Easy

Low

Trentepohlia

Filamentous

Red

Difficult

Nil

Table 2 Composition of the algal preservation and growth
medium (g/l)

Medium

NaNO

3

K

2

HPO

4

FeSO

4

CaCl

2

BG11

1.500

0.040

0.075

0.036

1064

Materials and Structures (2007) 40:1061–1071

provides the quantity of pheopigments (or pheo
a), which are the degraded form of chl a
(Pheopigments are found in dead cells). The
drawback is that it is heavy and not very rapid,
results being obtained after two days;

– the measurement of dry biomass amounts to

determining the dry mass of algae contained in
a given algal volume after filtration and drying
at 60C. The advantage is that the quantity of
algae is measured directly, rather than the
quantity of one of the cellular components as in
the previous method. The drawback is that
separate quantification of live and dead algae is
impossible;

– the nephelometry measurement relies on the

evaluation of the blur induced by an algal
development in the liquid culture. This mea-
surement is made by spectrophotometry, by
estimating the OD

750

or optical density of the

suspension at a wavelength of 750 nm. The
main advantage of this test is that it is easy and
quick. The drawback lies in the fact that the
value obtained has to be linked with a direct
parameter, such as the dry mass. For this, an
experimental calibration curve has to be estab-
lished showing the relation between the optical
density and the quantity of algae.

The different quantification methods were

tested for the three selected algae. The results are
presented in Figs.

1

and

2

.

The graphs show that:

– the increase of the various parameters versus

time was almost linear in the period considered;

– the quantity of chl a remained low for Chroo-

coccidiopsis compared to the other algae
(Fig.

1

a). This difference is linked with the fact

that chl a cell quantity depends on the alga, and it
is stronger for chlorophyceae than for cyano-
phyceae. Hence, it is not possible to compare the
results of the two species or to quantify a mixture
of several algae by measuring chl a content.

However, pheo a measurement by the same
method (Fig.

2

b) allowed variations in the

amount of dead algae to be followed and thus
illustrated the algae state of vitality (which is
impossible with other techniques).

– The curves obtained through nephelometry

(DO

750

) (Fig.

2

a) and dry mass measurements

were quite similar for all the species.

These results also brought out correlations

between the methods. Regression lines could be
plotted between dry mass and optical density at
750 nm (OD

750

) for each culture, as has already

been done by Piquemal [

26

]. However, these

correlations seemed to have a validity limit of
between 1 and 1.25 for the OD

750

value. Above

this limit, an inflexion of the nephelometric curve
was observed although dry biomass continued to
increase.

Table

3

gives the equations of these lines in the

determined validity domain.

For the chl a measurement, similar correlations

with the nephelometry measurement were ob-
served for Chlorhormidium and Chlorella chloro-
phyceae. The results were more random for
Chroococcidiopsis cyanophyceae. This may be
linked with the low content of chl a in the cells of
this algal class.

a

Chl a = f(t)

0

5

10

15

20

25

30

35

0

20

40

60

t in days

Ch

l

a

in

µ

g/ml

Chroococcidiopsis

Chlorhormidium

Chlorella

80

0

20

40

60

t in days

80

la

i

n

0

5

10

15

20

25

30

35

Chroococcidiopsis

Chlorhormidium

Chlorella

b

Pheo a = f(t)

Chroococcidiopsis

Chlorhormidium

Chlorella

Pheo a in

µ

g/ml

Fig. 1 Variation of chl a and pheo a according to time

Materials and Structures (2007) 40:1061–1071

1065

Considering the results, the nephelometry

method (OD

750

measurement) was preferred for

monitoring the variation of algal density in inoc-
ulation liquid cultures over time. The measure-
ment method of dry biomass was used when OD

750

values were greater than 1. Measurements of chl a
and pheo a quantities were taken periodically,
above all at the moment of inoculation, to check
the state of vitality in the inoculation culture.

3.1.2 Cement-based slab preparation

Characterisation tests were performed on mor-
tars. This choice was a simplification with respect
to the initial study, which concerned concrete
external walls, but in practice, the assimilation of
the first few millimetres of a concrete wall to a
mortar is not absurd as regards wall effect
phenomena. Moreover, the use of mortars
allowed small sized specimens to be made.

a. Dimension and design of the mould

The

choice of the slab dimensions (5 * 5 * 1 cm

3

)

was based on the following elements:

– the cast face in the bottom of the mould which

received the inoculation at the beginning of the
tests was small (5 * 5 cm

2

) so as to allow a large

number of slabs with different formulations to
be used in the same test without requiring a
very large installation. The high number of
slabs allowed representative measurements to
be taken for growth quantification;

– the very small thickness (1 cm) was chosen to

allow total and rapid humidification of the
mortar surface during the test of capillary
ascent and to accelerate the mortar carbon-
ation prior to the test.

The mould intended for slab fabrication was

composed of:

– a removable bottom made of the cast material

to be tested (steel, wood, PVC);

– a grid system composed of PVC bars 2 cm wide

and 1.2 cm high, which was fixed in grooves
0.2 cm deep provided for the purpose in the
bottom of the mould. This grid divided the
mould into 30 parts each 5 * 5 * 1 cm

3.

b. Fabrication

The specimens were processed as

follows:

– the mould was sprayed with form oil;
– the divisions were filled with mortar;
– each side of the mould was subjected to about

ten shocks (by gravity) to help the mortar fill
the divisions properly;

– the mould was put in a storage room (100%

relative humidity and temperature 21C)

a

Chroococcidiopsis
Chlorhormidium
Chlorella

Chroococcidiopsis
Chlorhormidium
Chlorella

OD

750

= f(t)

0

0,5

1

1,5

2

t in days

OD

750

Chroococcidiopsis
Chlorhormidium
Chlorella

Chroococcidiopsis
Chlorhormidium
Chlorella

b

Dry mass = f(t)

0

0,5

1

1,5

2

2,5

0

20

40

60

80

0

20

40

60

80

t in days

Dr

y

m

ass

in

m

g

/m

l

Chroococcidiopsis
Chlorhormidium
Chlorella

Fig. 2 Evolution of nephelometry and dry mass according to time

Table 3 Equations of the correlation lines between dry
mass and nephelometry

Species

Regression lines

Correlation
coefficient

Chroococcidiopsis Dry mass (g/l) = 1.042.

OD

750

+ 0.118

0.99

Chlorhormidium

Dry mass (g/l) = 1.131.

OD

750

–0.068

0.94

Chlorella

Dry mass (g/l) = 1.134.

OD

750

–0.053

0.99

1066

Materials and Structures (2007) 40:1061–1071

c. Early days conservation

Hardened mortars

were treated as follows in the early days:

– form removal at 1 day;
– storage in a room with regulated atmosphere

(20C and 50% R.H.) for 15 days (correspond-
ing to the usual ageing conditions for concrete
walls);

– accelerated carbonation in regulated chamber

(50% air and 50% CO

2

mixture, relative

humidity between 60 and 70%) until complete
carbonation was obtained (1–3 months depend-
ing on the mortar composition);

– 7-days leaching in demineralised water, chan-

ged daily (elimination of strongly basic com-
pounds and water saturation of the specimens).

d. Mortar characterisation

The initial charac-

terisation of mortars concerned the various
parameters which could directly or indirectly
influence

the

biological

growth

on

a

cementitious support or its visual perception:
chemical

composition,

mechanical

strength,

porosity, roughness, bug holes (surface bubbles),
colour and water absorption-desorption capacity.

3.1.3 Mortar moistening simulation test by water

capillary ascent

This method simulated algal growth at a wall base,
where water supplies essentially occur by capillary
ascent. It highlighted the influence of the material
parameters such as the mineral composition and the
pore-size distribution in the mortars. Here, algae
were tested individually (one algal species per box).

Figure

3

a and

b

show a schematic diagram and

photograph of the device.

3.1.4 Test process

The different environmental parameters to be
controlled during the tests were the available
moisture on the support surface, the intensity of
illumination and the nutritive supplies.

a. Box preparation

Boxes were prepared as

follows:

– 160 g of dry vermiculite was placed in each

transparent (polycarbonate) simulation box of

dimensions 23.5 * 17.5 * 9.5 cm

3

. 650 ml of BG11

medium diluted 20 times was added to guarantee
the algae mineral supplies, and the vermiculite
was homogenised and then compacted;

– the mortars, previously water saturated, were

put on the vermiculite, formwork face up,
3 days before the beginning of the tests (so
that capillary ascent moistening could start up).
For each type of inoculum, 3 prisms of each
formulation were inoculated (a box allowed 4
mortar formulations to be tested at the same
time).

b. Inoculation of the prisms

The process was the

following:

– the algae were cultivated in liquid medium for

35 days (beginning of the exponential growth
phase);

– two hours before the inoculation, the mortars

were removed from the vermiculite so that
their surface was dry at the inoculation time (so
that the liquid containing the algae was more
easily absorbed and the initial algal distribution
remained homogenous);

– the inoculation was performed on the form-

work surface of each prism by spreading the
chosen

quantities

(0.2–0.5 ml/25 cm

2

)

uni-

formly with the point of a pipette.

c. Growth parameters

The box storage conditions

were optimized to accelerate the algal potential
growth on the cementitious materials:

– the test enclosure (1.5 m long, 1 m wide and

1.5 m high) was placed in an air conditioned
room at 21C and it cut off from outside natural
or artificial lighting.

– lighting was provided by 4 fluorescent lamps

(OSRAM ‘‘daylight’’). These lamps ensured an
illumination of 1600 ± 200 lux on the cultures,
which corresponded to the optimal growth
condition for the selected species. The photo-
period (day-night daily cycle) was fixed to 16 h
day and 8 h night (lighting was turned on form
6 a.m. to 10 p.m. every day);

– the temperature in the boxes varied between 21

and 23C according to the lighting (Fig.

4

), and

was close to the growth optimum for the
selected species (23C);

Materials and Structures (2007) 40:1061–1071

1067

– the boxes were closed, which allowed a relative

humidity greater than 98% to be maintained on
the prisms;

– some 20 times diluted BG11 was used as the

moistening liquid for the vermiculite, so that
mineral supplies did not constitute a growth
limiting factor.

3.1.5 Evaluation of the accelerated test

In practice and in favourable growth conditions
(porous support), it takes 1 year to see the first
visible covering and 2–5 years for cover to
become really intense.

In the conditions of the test and for a porous,

bioreceptive support, a total covering appeared
between 15 and 30 days after inoculation with the
two chlorophyceae, whereas 20% of the surface
only was colonized after 60 days by the cyano-
phyceae.

This great acceleration was partly connected

with the support conditioning but, above all, with
the fact that the support was directly inoculated,
and was kept moist. So the accessibility was no
longer a limiting factor

3.2 Mortar moistening simulation tests

by water run-off

In this test, the mortar prisms were tilted at 45 in
a polycarbonate transparent chamber. The faces
to be studied were subjected to intermittently
applied, uniform run-off of a BG11 solution
inoculated with a mixture of the three algae.

Figure

5

a and

b

give a schematic diagram and a

photograph of the device.

3.2.1 Test process

a.

Chamber

preparation

The

experimental

device, designed to test 4 mortar compositions
at the same time, was stored in an air-conditioned
room at 21C. The device characteristics were the
following:

– polycarbonate transparent chamber 1 * 0.5 *

0.5 m

3

, divided into 4 alveoli, each one con-

taining a 45 tilted support for the test prisms.
This chamber, fitted with a lid, was stored in an
air-conditioned room (21C) where no outside
light could enter;

– the lighting inside the chamber was provided by

two OSRAM ‘‘daylight’’ fluorescent lamps. The
prisms were lit by one lamp (about 1600 lux on
the prisms whatever the position, upper or lower
ramp). The inoculated solution was lit by the
second lamp. The imposed photoperiod was 12 h
day and 12 h night (lighting was turned on from
8 a.m. to 8 p.m. every day).

– The run-off was set at 3 h a day for the bottom

ramp (between 8 and 11 a.m.) and 1 h a day for
the upper ramp (between 9 and 10 a.m.). It was
ensured by immersed pumps (300 l/h aquar-
ium-type pump) linked with spraying ramps

Fig. 3 Mortar moistening
system (a) diagram, and
(b) photo

lighting

Daily temperatures

20

21

22

23

0:00

4:00

8:00

12:00

16:00

20:00

0:00

Time

tem

p

era

tur

e

(°

C)

Fig. 4 Daily temperature cycle on the prisms in the
capillary ascent test

1068

Materials and Structures (2007) 40:1061–1071

allowing perfect run-off on the mortar prisms.
The liquid from the spraying ramps first fell on
PVC prisms situated above the mortars under
test and having the same size.

– 10 l of BG11 medium were placed in each of the

alveoli. An immersed, closed circuit pump ensured
constant movement and mixing of the medium.

b. Inoculation of the prisms

The prisms were

inoculated through the run-off (mixture of the 3
algae in the same liquid or each alga individually
tested). 25 ml of each previously prepared algal
culture was inoculated into the 10 l of BG11
medium situated in the bottom of each alveolus in
the chamber (i.e. about 75 mg of algae dry mass
per alveolus).

c. Growth parameters

In this test, the algal

potential growth was accelerated thanks to the
following parameters:

– the average temperature of the chamber , vary-

ing between 21 and 25C depending on the
lighting (see Fig.

6

);

– the illumination (controlled by a luxmeter)

whose intensity had been fixed at 1600±200 lux,
and the photoperiod (daily day-night cycle) of
12 h day and 12 h night (the lighting was on
from 8 a.m. to 12 p.m. every day);

– the run-off period of the liquid containing the

algae, which had been fixed at 1 h or at 3 h in
order to study the influence of the moistening
and colonisation time;

– sufficient nutritive elements, brought by the

BG11 solution run-off so that the algal growth
was not limited.

3.2.2 Evaluation of the test acceleration factor

In practice and in favourable growth conditions
(porous support, frequent water run-off), it takes
about 1 year to see the first visible covering and
2 years for it to become really intense.

In the conditions of the test and for a porous,

bioreceptive support, total coverage was observed
after 15 days in run-off conditions of 3 h/day and
35 days in 1 h/day run-off conditions. It took

Lamps

Pumps

PVC Prisms

Water +

algae

Watering
system

Mortar
prisms

Water +

algae

a

b

Fig. 5 (a) Run-off test
schematic diagram. (b)
Run-off test picture

Materials and Structures (2007) 40:1061–1071

1069

2 months in 3 h/day run-off conditions and
3 months in 1 h/day run-off conditions to reach
total coverage.

4 Conclusion

Biological stain development on external wall
surfaces, and particularly on concrete walls, can
quickly lead to considerable aesthetic deteriora-
tion and also, in the long term, to chemical and
physical degradation of the skin of the concrete. It
is therefore important to have an accelerated test
for concrete ageing by live species colonisation at
one’s disposal.

The literature shows that the first visible stains

correspond to considerable algal growth (algae
are autotrophic organisms) and that the growth
factors are temperature, support humidity and
mineral salt content. It also highlights the relevant
parameters for laboratory tests.

Two accelerated algal growth tests (results

obtained in less than 3 months) for mortar sup-
ports have been developed. The first recreates
stains observed at the base of the walls, which are
linked to capillary ascent, and the second simu-
lates stains observed at the top of walls or in some
particular areas (window sills, balcony under-
surfaces, bridge piers) and linked with repeated
run-off.

The important points in these two tests are the

following:

•

the species selection: three ubiquist species
were chosen for their ease of culture: Chroo-

coccidiopsis cyanophyceae and Chlorhormidi-
um and Chlorella chlorophyceae;

•

the preparation method, and choice of algae
(liquid cultures on BG11), mortars (acceler-
ated carbonation, saturation) and inoculation
techniques (deposit, run-off);

•

the choice of the test conditions (temperature
of 23C, illumination of 1600 lux, BG11
mineral supplies).

The quantification of stain growth on mortars

will be the subject of the second article in this
series: choice and development of mortar char-
acterisation methods (roughness measurement,
bug hole (surface bubble) estimation), choice and
development of algal quantification techniques on
mortars (image analysis, reflection factor mea-
surement).

Acknowledgements

The authors are grateful to ATILH,

and particularly Mr. Baron, for financial assistance. They
thank Mr. Haehnel, and Mr. Boulon (ITECH) for their
technical assistance.

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