Vol.3, No.6, 478-483 (2011) Natural Science
http://dx.doi.org/10.4236/ns.2011.36066
Indices to monitor biological soil crust growth rate—lab
and field experiments
Avraham Dody1*, Roni Hakmon1, Boaz Asaf1, Eli Zaady2
1Nuclear Research Center-Negev, Beer Sheva, Israel; *corresponding author:
2Department of Natural Resources and Agronomy, Gilat Research Center, Israel.
Received 28 March 2011; revised 23 April 2011; accepted 8 May 2011.
ABSTRACT
The aim of this work was to identify test meth-
ods for accelerating growth of biological soil
crust (BSC). The BSC in the Yamin Plateau in
the north-east of the Negev Desert is composed
of cyanobacteria such as microcoleus spp.
nostoc spp. and others. Cyanobacteria are well
adapted to dry environments, owing to their
ability to survive desiccation, high temperatures
and solar radiation. Since the BSC is a live
component in the ecosystem, it can repair itself
in the event of failures such as environmental
disturbances by living things. In the lab, we
used five different treatments and mediums:
natural BSC, pure sand as reference, pure sand
with spores and propagules, pure sand with
whey, and pure sand with spores and propa-
gules and whey. The spores were collected from
specified collecting areas in the field. Each Petri
dish was irrigated daily with 10 mL of double-
distilled water. The testing period ran for 4.5
months with 10 samples taken from each
treatment at 1.5 month intervals. The analyses
criteria were: NDVI for chlorophyll content by
remote sensing techniques, polysaccharide
content, infiltration rate through the crust, pro-
tein and organic content. The results showed
that NDVI, polysaccharides and infiltration rates
are good indicators for showing growth accel-
eration of the crust; while protein and organic
content were found to be less indicative. The
treatments using whey for preliminary crust
failed in the lab since cracks were observed, but
succeeded in the field experiments. In the field,
we measured only the chlorophyll content with
a time interval of 20 months. The methodology
of how to accelerate the growth of BSC was
found to be effective.
Keywords: Microcoleus Spp; Spores; NDVI;
Infiltration Rate, Polysaccharides Content
1. INTRODUCTION
The main motivation for this study was to identify
how to reduce the migration of contaminants into the
biosphere from waste disposal sites (WDS). According
to International Atomic Energy Agency recommenda-
tions, radioactive (and/or other hazardous materials)
WDS should be under institutional control for a period
of several hundred years after site closure [1]. After that
time, the site should be given back to the public for dif-
fering land uses. We are suggesting the use of a biologi-
cal soil crust (BSC) as a top layer over WDS as soon as
possible after site closure for the following reasons: 1)
BSC has been shown to reduce water infiltration and,
therefore, may reduce the leaching of the contaminants
toward groundwater; 2) The BSC, when grown over
WDS, may reduce the erosion of the topmost layer, and
may also reduce the migration of contaminants into the
biosphere; 3) Since the BSC is composed of live organ-
isms that renew themselves, its longevity and durability
is much longer and greater than any common geo-textile
layer; 4) Since the BSC is composed of live organisms in
the natural, dry ecosystem (with the ability to survive
desiccation, high temperatures and solar radiation) it has
the capability of repairing itself in the case of damage or
disturbance; 5) The BSC can survive climate changes.
It must be stated very clearly that the idea of using the
BSC is not to replace any other engineered barriers, but
as an additional layer that should be applied to the top of
the WDS.
In semiarid and arid regions of the world, when land-
scapes are undisturbed, the soil surface is covered with
BSC. The BSC community varies significantly by pre-
cipitation regime [2-4]. The BSC cover is characterized
by a tightly structured surface and typically varies from
2 mm thick, relatively homogeneous cyanobacterial
crusts, to complex crust composition communities of
mosses, lichens, soil algae, fungi and cyanobacteria of
about 15 mm thickness [5,6]. The primary colonizers of
the crust community, which are composed of cyanobac-

A. Dody et al. / Natural Science 3 (2011) 476-483 479
teria and soil algae, secrete polysaccharide mucilaginous
sheaths on the soil surface that bind together the soil
particles [7-9]. Consequently, they play an important
role in soil surface stabilization for preventing both wind
and water erosion [10,11]. In the Negev desert (Israel),
the combination of relatively high contents of silt and
clay particles and the cyanobacterial exudates change the
soil/water regime by affecting runoff, rain interception,
water-holding capacity and soil moisture content [12-14].
BSC dramatically influences the runoff yield, mainly in
arid and semi arid zones [15].
Field measurements, at several study sites in Israel,
showed that BSC decreased water infiltration [16,17], by
producing a tough cover-layer on the soil surface [4].
The aim of this work was to find the parameters indi-
cating the growth rate of BSC. In nature, the growth rate
of the BSC is very slow (years and more) [18]. For this
study, BSC spores and propagules were collected from
overland runoff in nearby, naturally undisturbed areas
and transferred to the laboratory experiments. The natu-
ral conditions of the field study area are: soil pH around
8, low rainfall amounts (80 mm/y) and high potential
evapotranspiration (2200 mm/y) are the main reasons
that the soil surface is dominated by cyanobacteria
(>80% of the soil cover).
2. METHOD AND MATERIALS
Runoff water containing spores and propagules of the
BSC were collected from a specific collecting system
built in the field at Yamin Plateau, north-east Negev De-
sert, Israel. The dominant cyanobacteria in our study
area are microcoleus spp. and Nostoc spp.
Pure sand (from 0.3 meter below the soil surface) was
collected from the natural, undisturbed nearby area and
used as substrate. Five treatments were considered: 1)
BSC as reference, 2) Pure sand, 3) Pure sand with spores,
4) Pure sand with whey (whey used as coagulating sub-
stance), 5) Pure sand with spores and whey. We worked
with five sets of Petri-dishes, each containing 30 dishes
and totaling 150 samples; each Petri-dish contained 80
grams of pure sand. Runoff water from nearby natural
plots containing BSC spores were used to irrigate treat-
ments 3 to 5 (only once). Then, for the next 4.5 months,
each Petri-dish was irrigated every day with 10 ml of
double-distilled water (DDW). Analysis samples were
taken at intervals of 1.5 months, 3 months and 4.5
months. Similar experiments were run in the field on
fifteen separate 2 × 1 m sand-dune plots where the crust
layer of the soil surface was removed.
Growth rates of the BSC were studied using five dif-
ferent variables: infiltration rate, polysaccharide content,
protein, organic content and Normalized Difference Ve-
getation Index (NDVI) and chlorophyll content. A col-
umn of 100 mm of DDW was placed on each Petri-dish
(with 5 holes in the bottom) and the infiltration rates
were measured. The polysaccharides were measured
with a spectrophotometer (UV-VIS mini-1240 spectro-
photometer, Shimadzu), using Anthron reagent and Sul-
furic acid [19]. Protein content was measured using the
Lowry method [20]. The chlorophyll content was ex-
tracted with acetone and quantified [21]. Organic content
was measured by washing the sample with 0.1 M of HCl
and 5 hours in a 550˚C furnace [22]. Vegetation index
(VI) was developed during recent decades [23], based on
different combinations of the ratio between the R-band
(which corresponds to the region of maximum chloro-
phyll absorption), and the NIR-band (which corresponds
to maximum reflectance of incident light by living ve-
getation). The most widely used index is the NDVI and
is defined as:
NDVI = [NIR – R]/[NIR + R] (1)
where R and NIR are the radiance and reflectance, or at
least “apparent reflectance” in the R and the NIR spec-
tral bands, respectively [24]. The NDVI values lie in the
range –1.0 to +1.0, with denser and/or healthier vegeta-
tion having higher positive values.
As mentioned above, we sprayed whey on two treat-
ments. Whey is a byproduct of the cheese- and ca-
sein-manufacturing industry [25]. There is continuing
interest in utilizing this byproduct as a fermentation sub-
strate for the production of value-added products. In
order to check its feasibility to create crust in a short
time, we also wanted to study if there was any interrela-
tionship between spores and whey, and how growth rate
may be affected.
Data were processed using analysis of variance with
the ANOVA statistical package. One-way ANOVA, with
Tukey Test’s [26] were used to test the differences be-
tween treatments (crust-control, sand, sand with spores
and sand with spores and whey). The dependent vari-
ables were: infiltration rate, polysaccharide content,
protein, organic content and NDVI for chlorophyll con-
tent. Differences were considered statistically significant
if p < 0.05.
3. RESULTS
The analyses for all treatments were made via the time
sequence sets presented in the following figures.
1) Polysaccharides
In all five treatments, the concentrations of the poly-
saccharides became higher with time (Figure 1).
Changes are almost linear in all treatments, but with
different rate. The highest value of polysaccharides was
found in the whey treatments, and the lowest value in the
sand treatment. The relative highest change was found in

A. Dody et al. / Natural Science 3 (2011) 476-483 480
Figure 1. Polysaccharide content with time. The bars indicate
the ±SE.
the spores’ treatment from 31 to 100 mg/g during 3.5
months. In the ANOVA test, significant differences were
found among the five treatments. On Feb. 06 p < 0.001.
On April 06 p < 0.001. On June 06 p < 0.001.
2) NDVI
The chlorophyll content was measured using remote
sensing techniques—NDVI (Figure 2). The relatively
higher change was found in the medium treated with
spores, even higher than the natural BSC. In the ANOVA
test, significant differences were found among the five
treatments. On Feb. 06 p < 0.005. On April 06 p < 0.005.
On June 06 p < 0.005.
3) Infiltration rates
Infiltration rates were reduced with time in all ex-
periments (Figure 3). The significant change from 9.5 to
4 ml/min occurred during month 3.5, in the medium
treated with spores. In the ANOVA test, significant dif-
ferences were found among the four treatments. On Feb.
06 p < 0.005. On April 06 p < 0.005. On June 06 p <
0.005.
4) Protein content
The changes of the protein content as a function of
time are presented in Figure 4. As seen in April, three
treatments had low concentrations when compared with
February. High concentrations were found in June. Two
weeks between 23.02.06 to 27.4.06 the Petri dishes were
not irrigated due to a technical problem. This might be
explaining the drop in NDVI in some of the treatments.
5) Chlorophyll in the field
We ran limited experiments in the field in order to
gain assumptions about the growth rate in the field
compared to the laboratory experiments. In specific 2 ×
1 m plots (Figure 5), we measured the chlorophyll con-
tent by direct extraction from the top most crust layer of
the soil surface (Figure 6). Twenty months after the be-
ginning of the experiment showed the importance of
sowing spores and propagules on the sandy dunes. As
expected, the crust has the highest chlorophyll content
with the spores’ treatment being significantly higher than
the sand treatments, which is statistically similar to the
Figure 2. Changes in NDVI value with time. The bars indicate
the ± SE.
Figure 3. The rate of infiltration in each of the treatments over
time. The bars indicate the ±SE.
Figure 4. Protein content with time. The bars indicate the ±SE.
whey treatment.
6) Regression line between dependent variables
NDVI was plotted against infiltration rate and poly-
saccharide content (Figures 7(a-b)) and polysaccharides
was plotted versus infiltration rate (Figure 7(c)).
4. DISCUSSIONS
In this study, we examined physical and bio-physio-
logical methods in order to find useful and reliable tech-
niques for monitoring crust development. The findings
showed that NDVI is the favored parameter for assessing

A. Dody et al. / Natural Science 3 (2011) 476-483 481
Figure 5. The experiment plots in the field.
Figure 6. The changes in chlorophyll a+b in the field from
January 2006 to August 2007.
biological soil-crust growth rate. It is a reliable method
for showing the increase of crust development in time,
good correlations were found between the variables; in-
filtration rates and polysaccharide content (as presented
in Figure 7). This result concurs with other studies which
found that NDVI is well correlated with various vegeta-
tion parameters such as green biomass [27] and photo-
synthetic activity [28]. It has also been found to be useful
for various image analyses like crop classification, green
coverage and change detection. The reflectance spectra of
BSC in semi-arid regions of Australia was studied on a
variety of soils [29]. Noticeable differences upon wetting
of the BSC were observed using her method. Conse-
quently, it is assumed that a vegetation index, such as
NDVI, can serve as a second indicator for recovery of
BSC. In [30,31] the authors tested whether the high
NDVI values may be caused by the photosynthetic activ-
ity of BSCs, which cover most of the soil surfaces in the
semi-arid regions of the Negev Desert. They found that
the spectral reflectance curves of lower plants can be
similar to those of higher ones and their derived NDVI
values can be as high as 0.30 units. A high correlation (r
= 0.79) was reported between NDVI values and chloro-
phyll content of a wet BSC [6,32].
A similar reliable method was found with infiltration
rate measurements. Decreases of the infiltration rate
(a)
(b)
(c)
Figure 7. (a) The correlation between the dependent variables:
NDVI vs. infiltration rate. (b) The correlation between the
dependent variables: NDVI vs. polysaccharides. (c) The corre-
lation between the dependent variables: polysaccharides vs.
infiltration rate.
were shown with crust development over time. Both of
the last two analyses used NDVI and infiltration rate
measurements, and were nondestructive techniques for
the soil surface in the Petri-dishes.
During the growth and the laboratory experiments we
used double distilled water (DDW) in order to eliminate
any side effect of the photoautotrophic organisms and
their exudates that may dissolve within the water and
affect the results. For this reason we did not use sterile
runoff water. It was reported in the literature that runoff
water contain nutrients and organic matter other than
propagules [33]. Polysaccharide content in the soil as an
indicator of the presence of cyanobacteria microcoleus

A. Dody et al. / Natural Science 3 (2011) 476-483 482
spp. was found to be a good parameter to present the
development of the crust (although only 1 gram of soil
surface was taken for each replicate). Because protein
and organic content are related to two other variables,
the standard deviations were relatively too high, in order
to identify clear trends in the development of the crusts.
The sandy soil used for the experiments was taken
from a depth of 30 cm, under the assumption that no
spores and propagules existed at that level. The fact that
there were changes in all the parameters in the pure sand
treatment used as reference proves that our assumption
was wrong.
The next stage of our study focuses on the develop-
ment of a bio-reactor to grow BSC spores and propa-
gules, as well as to figure out optimum conditions (bio-
logical and cost-beneficial). The spores and propagules
will be seeded on the target areas in order to shorten the
coverage time of the area with BSC (compared to natural
growth).
More field experiments are needed to study the appli-
cations under field conditions, but our preliminary re-
sults showed fast establishment of the spores on sand
dune soil at the north-eastern Negev Desert (Yamin pla-
teau). Again, for the point of view of WDS, it must be
stated very clearly that the idea of using the BSC is not
to replace any other engineered barriers, but to add an
additional top layer that increases the soil surface stabil-
ity by using natural components of the ecosystem in the
arid landscape.
5. CONCLUSIONS
 NDVI, polysaccharides and infiltration rates are good
indicators for showing growth acceleration of the
crust;
 Protein and organic content were found to be less
indicative.
 The treatments using whey for preliminary crust
failed in the lab since cracks were observed, but suc-
ceeded in the field experiments.
 In the field, we measured only the chlorophyll con-
tent with a time interval of 20 months.
 The methodology of how to accelerate the growth of
BSC was found to be effective.
6. ACKNOWLEDGEMENTS
The authors wish to thank Faina Tziperman from Gilat-ARO for her
assistance in the laboratory work and Catherine Kelly for reviewing the
English.
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