Ecological Indicators 10 (2010) 840–847

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Ecological Indicators
journal homepage: www.elsevier.com/locate/ecolind

Ecological quality scales based on phytoplankton for the implementation of Water

Framework Directive in the Eastern Mediterranean
Sofie Spatharis *, George Tsirtsis
Department of Marine Sciences, University of the Aegean, University Hill, 81100 Mytilene, Greece

A R T I C L E I N F O A B S T R A C T

Article history: Structural changes of phytoplankton communities, often expressed through ecological indices,
Received 2 October 2009 constitute one of the metrics for the implementation of the European Water Framework Directive
Received in revised form 28 December 2009 (WFD). In the current study a thorough analysis of the efficiency of 22 ecological indices was performed
Accepted 17 January 2010
and a small number was selected for the development of five-level water quality scales (High, Good,
Moderate, Poor, and Bad). The analysis was performed on simulated communities free of the noise of
Keywords: field communities due to uncontrolled factors or stochastic processes. Two criteria were set for the
Community structure
sensitivity of indices, namely their monotonicity and linearity across the studied eutrophication
Diversity
Ecological assessment
spectrum. The whole procedure was based on the development of a five-level quality assessment scheme
Indices based on phytoplankton abundance. Among the indices tested, the Menhinick diversity index and three
Log series distribution indices of evenness were the most efficient, showing consistency (monotonic behavior) and linearity and
Model communities were therefore used for the development of quality scales for the WFD. An Integrated Phytoplankton
Sensitivity analysis Index (IPI) based on three phytoplankton metrics, chlorophyll a, abundance, and diversity is also
proposed. The efficiency of these indices was evaluated for a number of sites in the Aegean, already
classified in the past by various methods based on nutrient concentrations or phytoplankton data. The
results indicate that the various phytoplankton metrics (chlorophyll a, abundance, and diversity)
assessed or proposed in the current study, carry their own information showing differences in the final
classification of areas. Therefore the establishment of synthetic indices as the IPI seems to be
advantageous for the integrated assessment of coastal water quality in the framework of European
policies as the WFD.
ß 2010 Elsevier Ltd. All rights reserved.

1. Introduction waters, the only planktic element referred to in the WFD is

Surface waters including streams, rivers, lakes, estuaries, and
coastal waters are under increasing ecological stress due to
anthropogenic activities worldwide (UNEP, 1999), leading to the
enforcement of major environmental policies, including the US
Federal Water Pollution Control Act (2008), the US Oceans Act
(2000), the European Water Framework Directive 2000/60 (EC,
2000), and the European Marine Strategy (EC, 2008). The
implementation of the Water Framework Directive (WFD) in
particular, both regionally and nationally, assumes the develop-
ment of a five-level water quality classification scheme (High,
Good, Moderate, Poor, and Bad) with the environmental objective
to achieve good surface water status for all European waters by
2015 (EC, 2000). The levels are defined using the Ecological Quality
Ratio (EQR) for a number of biological and chemical quality
elements. Among the key biological elements specified for coastal
* Corresponding author. Tel.: +30 22510 36835; fax: +30 22510 36809.
E-mail address: spathari@marine.aegean.gr (S. Spatharis).
phytoplankton. Phytoplankton is an efficient indicator of changes
in nutrient loads, but is also effective in evaluating responses to
many other environmental stressors, due to its fast population
response to changes in water quality, hydrology or climate
(Domingues et al., 2008). According to the European Directive
(EC, 2000), phytoplankton metrics that are fundamental in defining
and classifying the ecological status of surface waters are biomass
(as chl a), community changes (composition and species abun-
dance), and increase in the frequency and intensity of blooms.
Phytoplankton has been considered as a water quality key-
element in many studies for coastal ecosystems. A synthetic
approach for water quality assessment in the Atlantic Ecoregion
(Basque country), involved the development of an integrated index
based on chlorophyll a, phytoplankton total abundance (when
exceeding 107 cells/L), and abundance of harmful phytoplankton
species (Borja et al., 2004; Revilla et al., 2009; Vincent et al., 2002).
For the North Sea and the Atlantic Ecoregions (British coastal
waters), Devlin et al. (2007) developed an Integrated Phytoplank-
ton Index taking into account phytoplankton biomass (as chl a), the
frequency of elevated phytoplankton counts (individual species

1470-160X/$ – see front matter ß 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.ecolind.2010.01.005
 S. Spatharis, G. Tsirtsis / Ecological Indicators 10 (2010) 840–847 841

and total cell counts), and seasonal progression of functional
groups. Regarding the coastal waters of the Eastern Mediterranean
Ecoregion, one of the most oligotrophic marine environments
around the world (Krom et al., 1991), chlorophyll a is the only
parameter taken into consideration so far for WFD implementation
(Simboura et al., 2005). The limits set by these authors are below
0.10 mg/L of chl a for High water quality, 0.10–0.40 for Good, 0.40–
0.60 for Moderate, 0.60–2.21 for Poor and above 2.21 for Bad water
quality. Chlorophyll concentrations represent a simple and
integrative measure of the phytoplankton community response
to nutrient enrichment or succession (Devlin et al., 2007; Harding,
1994). However, community structure (i.e. the distribution of
individuals to species) conveys different information by also
considering heterotrophic species that are not represented in
chlorophyll measurements (Domingues et al., 2008). Moreover,
previous studies have demonstrated that an increase in chloro-
phyll a due to eutrophication is always accompanied by changes in
phytoplankton community structure in terms of total abundance,
species richness, and evenness (Tsirtsis and Karydis, 1998; Tsirtsis
et al., 2008). For phytoplankton abundance, a four-level
classification system was developed in the past for Eastern
Mediterranean coastal waters (Kitsiou and Karydis, 2000). The
limits set were <4160 cells/L for oligotrophy, 4160–31,399 cells/L
for lower mesotrophy, 31,400–188,333 cells/L for higher meso-
trophy, and >188,334 cells/L for eutrophication. However this
scale has not been adjusted so far to the five-level system required
by the WFD.
Changes in community structure are often quantified through a
number of indices which constitute popular tools in studies
associated with community ecology and disturbance (Washington,
1984). The most important advantages provided by indices are the
ability for direct comparisons between communities that have few
or no species in common and the easiness of their application and
interpretation (Magurran, 2004). Alternative methods to taxo-
nomic-based indicators related to body size, abundance distribu-
tion among functional groups, functional diversity, and
productivity descriptors have been found suitable for other
biological quality elements (Mouillot et al., 2006). However, for
phytoplankton, productivity and species-abundance data are
considered so far the most effective means to explore changes
in community structure (Mouillot et al., 2006). In the framework of
new policies for water quality assessment, investigators should
retain a parsimonious approach, assessing the suitability of
existing indices rather than developing new ones (Borja and
Dauer, 2008; Diaz et al., 2004). In addition, the evaluation of water
quality status should ideally incorporate suitable multimetric
indices (Borja and Dauer, 2008; Diaz et al., 2004; Domingues et al.,
2008) taking into consideration as many of the fundamental
attributes of phytoplankton as possible (i.e. biomass, community
structure, and frequency of blooms).
An important shortcoming in the development process of water
quality assessment schemes is the inherent noise or variability of
field data associated with seasonality, hydrodynamic circulation,
grazing, and patchiness (Karydis, 1996) or stochastic processes
(Mouillot and Lepretre, 2000). This variability masks and distorts
the information associated with a particular stressor or process of
interest (Karydis, 1992) and therefore is considered as undesirable,
especially in water quality assessment schemes which are often
using statistical distributions such as the lognormal and log series
(Tsirtsis et al., 2008) along a wide spectrum of productivity, from
oligotrophy to eutrophication, characteristic of Eastern Mediter-
ranean coastal waters.
The present study aims to assess the sensitivity of 22 ecological
indices of phytoplankton diversity to efficiently detect eutrophic
trends in Eastern Mediterranean coastal waters, and thus their
applicability in the Water Framework Directive. The development
and use of simulated phytoplankton communities covering a wide
and continuous eutrophication spectrum enabled (a) the unbiased
examination of the sensitivity (monotonicity and linearity) of the
indices and (b) the development of coastal water quality
classification scales using selected indices. Moreover a new,
integrated index of water quality was developed taking into
account three of the metrics proposed by the European Directive
for phytoplankton, which is chlorophyll a, abundance, and
community structure. The proposed classification schemes are
finally evaluated with an extensive validation dataset from coastal
areas of the Aegean Sea.
2. Methodology
2.1. Field data
Available datasets from coastal areas of the Aegean Sea, Eastern
Mediterranean were used to develop the proposed water quality
classification schemes and assess their efficiency. Detailed
information on the datasets, as well as previous assessment of
the water quality of the coastal areas under consideration, is
provided in Spatharis et al. (2008). For each dataset, information
on chlorophyll a, nutrients, and species abundances existed on a
monthly basis covering a full annual cycle and a total number of
816 samples was available for analysis. The areas under consider-
ation cover a wide range of biomass (chl a in the range 0.01–
8.80 mg/L and phytoplankton abundance from 960 to
9,905,980 cells/L), mainly affected by anthropogenic activities
(agriculture, urbanization, tourism) in the coastal zone. Summary
statistics for chlorophyll a and nutrient concentrations are given in
Table 1. More eutrophic areas according to previous assessments
based on chl a and/or nutrient concentrations are the Inner
Saronikos (Ignatiades et al., 1992; Karydis, 1996), the Kalloni Gulf
(Spatharis et al., 2007a,b), the Mytilene port (Tsirtsis, 1995) and
the Rhodos ports (Stefanou et al., 2000), whereas outer Saronikos
(Ignatiades et al., 1992; Karydis, 1996) and the Gera Gulf
(Arhonditsis et al., 2000) are considered as mesotrophic. Mytilene
strait (Tsirtsis, 1995), Rhodos coastal (Stefanou et al., 2000) and
Rhodos offshore (Vounatsou and Karydis, 1991) are characteristic
of oligotrophy. Rhodos offshore waters in particular, located north-
western of the Island of Rhodos, Greece, are typical of the pristine
undisturbed marine environment of the Eastern Mediterranean
(Vounatsou and Karydis, 1991). Therefore, this site has been
extensively used in the past to express reference conditions when
developing water quality assessment scales for the Aegean Sea
Table 1
Dataset information and mean annual values of chlorophyll a (mg/L) and dissolved
nutrient concentrations (mM) for the coastal areas in the Aegean Sea.
Site Stations Samples Chl a DIP DIN

based on annual means. An alternative approach in the assessment Rhodos offshore R1–R5 145 0.10 0.002 0.064
of water quality can be based on the use of simulated communities Rhodos coastal RH1–2 & 6–10 84 0.10 0.002 0.083
Mytilene strait M2 67 0.33 0.002 0.122
(Boyle et al., 1990; Mouillot and Wilson, 2002) that retain the main
Rhodos ports RH3–RH5 36 0.44 0.002 0.293
structural characteristics of natural communities. The elimination Gera Gulf GG3–GG8 89 0.84 0.007 0.053
of noise in the simulated communities reveals the signal, and Outer Saronikos Gulf S3–S9 168 0.63 0.004 0.148
therefore supports the quantification of the community response Mytilene port M1 39 1.01 0.003 0.166
Kalloni Gulf K3–K8 140 1.19 0.003 0.301
to environmental stressors. Previous studies have demonstrated
Inner Saronikos Gulf S1, S2 48 1.65 0.020 0.354
that phytoplankton communities can be successfully modeled
842 S. Spatharis, G. Tsirtsis / Ecological Indicators 10 (2010) 840–847

(Ignatiades et al., 1992; Karydis, 1992, 1994, 1996; Karydis and
Tsirtsis, 1996; Kitsiou and Karydis, 2000; Tsirtsis and Karydis,
1998; Simboura et al., 2005; Stefanou et al., 2000).
2.2. Development of simulated phytoplankton communities
Statistical distributions including the lognormal and log series,
allocating individuals in a community to species, can successfully
model phytoplankton communities (Tsirtsis et al., 2008). In the
current study, simulated phytoplankton communities were
generated using the log series distribution (Fisher et al., 1943).
The log series is characterized by two parameters, x and a, which
can be calculated when the ratio S/N is known (Magurran, 2004),
where S the species richness and N the total abundance of a sample.
When the two parameters of the log series are known, the number
of species in a community corresponding to abundance classes
(octaves) of doubling intervals can be easily calculated (Tsirtsis et
al., 2008). In the current study, a relationship was firstly
established between S and N from the field data (816 samples),
that is S = 6.241  log(N)  12.642; this relationship was found
statistically significant (R2 = 0.480, P < 0.01) by applying linear
regression analysis. Then given N (total abundance), S can be
calculated and thereafter x and a from the ratio S/N. Finally,
knowing the shape of the log series distribution (defined by x and
a), the number of species corresponding to each abundance class
can be calculated and total cells N can be allocated to species S. The
important role of dominant species in phytoplankton communities
was accounted by assigning a percentage of the total abundance to
the most dominant species. This percentage was calculated from
the field data. It was found that the abundance of the most
dominant species (N1) is related with total abundance (N) by the
equation log(N1) = 1.123  log(N)  0.968. This relation was sta-

Table 2
The 22 indices applied in the current study including 13 diversity, 7 evenness and 2 dominance indices.

Index type Index Formula Reference

Diversity indices Margalef S1 Margalef (1958)

D¼
ln N
Gleason S Ludwig and Reynolds (1988)
D¼
ln N
Menhinick S Menhinick (1964)
D ¼ pffiffiffiffi
N
PS
Simpson’s n  ðni  1Þ Ludwig and Reynolds (1988)
D ¼ i¼1 i
n  ðn  1Þ
Shannon H0 X S
ni n Shannon and Weaver (1949)
H0 ¼   ln i
i¼1
n n
Shannon D0 D0 ¼ S  ðS  d Þ
0
Camargo (2008)
Hill N0 N0 = S Ludwig and Reynolds (1988)
Hill N1 N1 = exp(H0 ) Ludwig and Reynolds (1988)
Hill N2 1 Ludwig and Reynolds (1988)
N2 ¼
Simpson0 s D
Odum S  1000 Odum et al. (1960)
O¼
N
Kothe Smax  Si Pielou (1975)
D¼
Smax
  !
Hurlbert N XS Hurlbert (1971)
PIE ¼  1 p2i
N1 i¼1
qffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi
PS
McIntosh 2 McIntosh (1967)
n i¼1 ni
M¼ pffiffiffi
n n

Evenness indices Evenness E1 H0 Pielou (1975)

E1 ¼
ln S
Evenness E2 expðH0 Þ Sheldon (1969)
E2 ¼
S
Evenness E3 expðH0 Þ  1 Ludwig and Reynolds (1988)
E3 ¼
S1
Evenness E4 1=Simpson0 s D Ludwig and Reynolds (1988)
E4 ¼
expðH0 Þ
Evenness E5 ð1=Simpson0 s DÞ  1 Ludwig and Reynolds (1988)
E5 ¼
expðH0 Þ  1
0
Evenness E6 E6 ¼ 1  d Camargo (2008)
0
Redundancy H  H0 Patten (1962)
R ¼ 0 max
Hmax  H0min

Dominance indices Berger–Parker n1 Berger and Parker (1970)

B¼
n
McNaughton n þ n2 McNaughton (1967)
a¼ 1
n
The terms used in the formulas are given below:
S = the number of species in a sample or a population.
N = the number of individuals in a population or community.
Ni = the number of individuals in species i of a population or community.
n = the number of individuals in a sample from a population.
ni = the number of individuals in a species i of a sample from a population.
pi = ni/n = the fraction of a sample of individuals belonging to species i.
Smax = the maximum number of species in a sample.
n1, n2 = the number of individuals in the two most abundant species. PK
0 j pi  p j j
H0max ¼ ln S (Pielou, 1975), H0min ¼ N1 ln ðNSþ1Þ!
N!
(Pielou, 1975), and d ¼ i 6¼ j S (Camargo, 2008).
 S. Spatharis, G. Tsirtsis / Ecological Indicators 10 (2010) 840–847
tistically significant by applying regression analysis (R2 = 0.957,
P < 0.01). More details about the modeling procedure have been
given elsewhere (Tsirtsis et al., 2008).
In the current study the total abundance of the phytoplankton
communities was set in the range of 103 to 107 cells/L characteris-
tic of the coastal waters of the Aegean. Twenty-two ecological
indices expressing diversity, evenness, and dominance were
calculated for the simulated communities using the cell number
as a proxy, and their fit to the corresponding indices’ values of field
communities was statistically evaluated.
2.3. Ecological Indices
Ecological indices are characterized as diversity, evenness, or
dominance indices, according to their mathematical formula
weighting more to the species richness or evenness components
of community structure. In the current study 13 diversity, 7
evenness, and 2 dominance indices commonly applied in
community ecology and aquatic studies (Karydis and Tsirtsis,
1996; Washington, 1984) have been considered (Table 2).
2.4. Sensitivity analysis of ecological indices and development of
water quality classification scales
The sensitivity analysis of the 22 indices to detect structural
changes of phytoplankton communities due to eutrophication and
the development of coastal water quality classification schemes,
were performed with the following step-by-step procedure:
1. A new five-level scale was developed for phytoplankton
abundance according to the requirements of the WFD, based
on an existing four-level scale (Oligotrophic, Lower Mesotro-
phic, Upper Mesotrophic, Eutrophic), proposed by Kitsiou and
Karydis (2000). This scale was developed using three standard
datasets from the Aegean coastal waters, already described
above, characteristic of eutrophication (stations S1 and S2 in
Saronikos Gulf), mesotrophy (stations S3 to S9 in Saronikos
Gulf), and oligotrophy (stations R1 to R5 offshore of Rhodos
Island).
2. 22 ecological indices were calculated on simulated phytoplank-
ton communities of 103 to 107 cells/L, which is the observed
range in Aegean coastal waters as mentioned above. For each
index, six values were calculated by the model corresponding to
the limits of abundance set in step 1. Thereafter the limits of
each index were tested for monotonicity (consistent increase or
decrease along the eutrophication spectrum). The monotonic
behavior is considered as a prerequisite for an efficient water
quality index, since the indices are used in a quantitative
manner (Washington, 1984) and their limits have to follow a
rank order. Indices that have shown non-monotonic behavior
were considered inefficient for water quality assessment and
were therefore excluded from further analysis.
3. The monotonic indices were then tested for linearity along the
studied eutrophication spectrum. Linearity was set as an
objective, considering that the linear behavior maximizes the
distances between the limits, resulting to more equally spaced
water quality scales. Linearity of the indices was tested by
applying linear regression analysis and the significance of the
departures from linearity was considered by calculating the
studentized residuals (Zar, 1998).
4. Selected ecological indices that have shown monotonic and
linear behavior were further used for the development of five-
level water quality classification schemes for the WFD
implementation. The Ecological Quality Ratios (EQRs) of the
various quality levels were calculated for each index. Finally, a
new Integrated Phytoplankton Index was developed from the
843
average EQR values of selected indices in order to account for
most of the phytoplankton metrics defined by the WFD (chl a,
abundance, diversity).
3. Results
The limits for phytoplankton abundance in the existing four-
level scale for eutrophication proposed by Kitsiou and Karydis
(2000) were <4160 cells/L for oligotrophy, 4160–31,399 cells/L for
lower mesotrophy, 31,400–188,333 cells/L for higher mesotrophy,
and >188,334 cells/L for eutrophication. Trying to develop a new
five-level scale for WFD by modifying the above scale, a
correspondence was set at first between the characterizations
according to the eutrophication level and the WFD characteriza-
tions (High, Good, Moderate, Poor, and Bad). This was achieved by
considering the mean chlorophyll a concentration of each group of
stations used for the scaling by Kitsiou and Karydis (2000)
according to the existing chlorophyll a scale for Eastern
Mediterranean by Simboura et al. (2005). It was found that the
characterizations oligotrophy, lower mesotrophy, higher meso-
trophy, and eutrophication correspond to the High, Good,
Moderate and Poor water qualities of the WFD. For oligotrophy
or High water quality in particular, both authors used the same
dataset (Rhodos offshore) to define reference conditions, also being
used in the current study. Trying to develop a five-level quality
scale for abundance complying with the WFD, a limit for Bad water
quality was defined in the present study, always using the same
dataset. The upper outlying values of the whole dataset were
defined (49 values), that is values greater than the upper quartile of
the dataset plus 1.5 times the interquartile range (Tukey, 1977).
These upper outlying values were considered as characteristic of
the Bad water quality of WFD (or dystrophy according to the
eutrophication level notation), and their median (710,470 cells/L)
was set as the limit between Poor and Bad water qualities.
Simulated communities in the range of abundance of 103 to
7
10 cells/L were generated by the log series distribution. The rank-
abundance plot of a selected number of simulated communities of
12, 17, 23 and 31 species, being the minimum, lower quartile,
median, and upper quartile respectively of species richness in the
816 field samples, is plotted in Fig. 1. The important role of the
dominant species in phytoplankton communities is revealed,
especially when abundance is high. Moreover, evenness is affected
by the total abundance and species richness, being higher for
communities characteristic of oligotrophy and showing a decreas-
ing trend with eutrophication (as expressed by the increase in cell
number). The goodness-of-fit of the simulated communities to
field data (816 samples) was then tested. Twenty-two indices,
expressing diversity, evenness, and dominance were calculated for
both simulated and field communities of the same abundance, and
Fig. 1. Rank-abundance plots of simulated phytoplankton communities of 12, 17, 23
and 31 species, that is the minimum, lower quartile, median, and upper quartile
respectively of species richness in the 816 field samples.
844 S. Spatharis, G. Tsirtsis / Ecological Indicators 10 (2010) 840–847

Fig. 2. Monotonic and non-monotonic behavior of selected ecological indices along the cell number gradient (in log scale). Calculations were based on simulated species-
abundance data generated with the log series model.

Spearman rank coefficient was used as a measure of correlation.
Significant correlations (P < 0.01) were found for all indices, the
Spearman coefficients being in the range from 0.245 for evenness
E4 to 0.979 for Odum index.
Six values were calculated for each index on simulated
phytoplankton communities; the minimum and maximum values
(for 103 and 107 cells/L), and the values corresponding to the limits
of the five-level scale defined for phytoplankton abundance above.
The monotonic behavior of each index was checked by plotting its
variability in the range of 103 to 107 cells/L, after standardization
on a 0–100 scale, for comparative reasons. Most of the indices (14
out of 22) have shown monotonic behavior, that is a consistent
trend of increase as Margalef’s index, or decrease as Menhinick’s
and evenness E1 indices (Fig. 2). The indices of Simpson, Shannon
H0 and D0 , Hill N1 and N2, Hurlbert, and McIntosh were non-
monotonic presenting a characteristic hump-shape curve. As
observed for Simpson and Hill N1 (Fig. 3), these indices increase
when the abundance is low (below 104 cells/L) and then they
gradually decrease reaching their minima at very high abundances.
Evenness E4 presents the opposite trend, showing a gradual decrease
and then a rise for abundances greater than 8  105 cells/L (Fig. 2).
The above-mentioned nine non-monotonic indices were considered
unreliable to be used for water quality assessment and were
excluded from further analysis.
The monotonic indices selected in the previous step were
further tested for linearity by applying linear regression analysis.

Fig. 3. Test of linearity of selected monotonic indices along the water quality levels (H, High; G, Good; M, Moderate; P, Poor; B, Bad). Menhinick and evenness E2 indices do not
present significant departures from linearity, whereas Odum and Redundancy deviate more than 2 standard deviations at one and two points, respectively.
 S. Spatharis, G. Tsirtsis / Ecological Indicators 10 (2010) 840–847 845

Table 3 Table 5
Assessment of the linearity of the 14 monotonic ecological indices: regression Water quality assessment of coastal Aegean waters (5, High; 4, Good; 3, Moderate;
coefficient (R2), significance level, number of unusual residuals and their departure 2, Low; 1, Bad) based on chlorophyll a (Simboura et al., 2005), phytoplankton
from linearity in terms of standard deviations. abundance, two ecological indices’ (Menhinick and evenness E2) scales, and the
proposed Integrated Phytoplankton Index (IPI).
Index R2 Sign. level Studentized Deviation
residuals Site Stations Chl a Cell no. Menhinick E2 IPI

Odum 0.704* 0.0367 1 3.89 Rhodos offshore R1–R5 5 4 5 4 5

Kothe 0.925** 0.0022 1 6.48 Rhodos coastal RH1–2 & 6–10 4 4 5 4 5
Gleason 0.896** 0.0042 2 2.12, 2.06 Mytilene strait M2 4 3 4 4 4
Margalef 0.905** 0.0035 1 2.25 Rhodos ports RH3–RH5 3 3 4 3 4
Menhinick 0.961** 0.0006 0 – Gera Gulf GG3–GG8 2 3 4 4 3
Hill N0 0.996** 0.0000 1 2.37 Outer Saronikos Gulf S3–S9 2 2 3 3 3
Evenness E1 0.984** 0.0001 2 2.28, 2.98 Mytilene port M1 2 2 3 3 2
Evenness E2 0.959** 0.0006 0 – Kalloni Gulf K3–K8 2 1 2 2 2
Evenness E3 0.953** 0.0008 0 – Inner Saronikos Gulf S1, S2 2 2 3 3 2
Evenness E5 0.952** 0.0009 1 3.17
Evenness E6 0.960** 0.0005 0 –
Redundancy 0.984** 0.0001 2 2.28, 2.98
Berger–Parker 0.938** 0.0015 1 4.96
index can be easily calculated as the mean of the EQRs of the three
McNaughton 0.928** 0.0020 1 3.05
metrics in each quality class, since these ratios are standardized in
the 0–1 range. The scales developed for selected indices
(Menhinick, E2, IPI) for the implementation of the Water
Significant departures from the linear behavior were checked by Framework Directive in Eastern Mediterranean, along with the
considering the studentized residuals that is observations which existing scales (a) of chlorophyll a for the Aegean waters (Simboura
deviate significantly from the linear model fitted. These residuals et al., 2005) and (b) of abundance by (Kitsiou and Karydis, 2000) as
are expressed in numbers of standard deviation and in case they modified in the current study, are presented in Table 4.
exceed two, they can be considered as outliers and be removed A summary of the water quality classification of several coastal
from the analysis. Statistically significant linear behavior at sites in the Aegean according to the already existing and newly
P < 0.05 was observed for all indices (Table 3). Moreover, developed quality metrics is presented in Table 5. Although there
Menhinick, and evenness indices E2, E3, and E6 have shown no is an overall agreement in the characterization of each site, some
significant residuals, therefore their behavior can be considered as differences were observed, spanning one quality level at most. The
fully linear. The diversity indices of Hill N0, Margalef, Odum and classifications ranged between High and Low, none of the coastal
Kothe, the evenness index E5, and the dominance indices Berger– areas classified in the Bad water quality, except from Kalloni
Parker and McNaughton presented significant departures from the according to the abundance criterion. Considering the differences
linear behavior with one value deviating more than 3 standard between the various metrics, it seems that scales based on
deviations. The rest of the indices, such as Gleason’s, evenness E1, ecological indices are more optimistic compared to the chlorophyll
and Redundancy have shown remarkable departures from linearity a and abundance scales. This optimism is further reflected to the
with one or two values deviating more than 2 standard deviations. proposed synthetic IPI index, if it is compared with the already
A graphical presentation of the residuals for selected indices is established chlorophyll a scale of Simboura et al. (2005) for coastal
shown in Fig. 3. Aegean waters.
Considering both the criteria of monotonicity and linearity, as
well as the goodness-of-fit of the simulated to the field data, it 4. Discussion
seems that the most efficient ecological indices for the develop-
ment of water quality classification schemes based on phyto- Many water quality assessment schemes have been developed
plankton data are the diversity index of Menhinick and the on phytoplankton for the implementation of the WFD in European
evenness indices E2, E3, and E6. For these indices the Ecological regional seas (Borja et al., 2004; Devlin et al., 2007; Revilla et al.,
Quality Ratios (EQRs) can be also calculated by standardizing their 2009; Sagert et al., 2005), using biomass (chl a and abundance),
values to fit the 0–1 range. In order to synthesize the information composition, and the frequency and intensity of blooms. For
on the water quality characterization based on chlorophyll a, Eastern Mediterranean, chlorophyll a is the only metric used so far
phytoplankton abundance, and community structure, an Integrat- for the WFD (Simboura et al., 2005). In the current study a five-
ed Phytoplankton Index (IPI) is proposed by accounting the above level classification system was developed on phytoplankton
metrics with equal weights. Diversity is expressed in terms of the abundance, as a modification of an existing four-level scale
Menhinick index, a mathematically simple index calculated from (Kitsiou and Karydis, 2000). The characterization of water quality
species richness and total abundance. The proposed integrated with the abundance scale seems more conservative compared to

Table 4
Water quality classification schemes and the corresponding Ecological Quality Ratios for Menhinick and evenness E2 indices, and phytoplankton abundance (N). The existing
scale for chlorophyll a by (Simboura et al., 2005) and the proposed Integrated Phytoplankton Index (IPI) are also shown.

Index High Good Moderate Low Bad

Menhinick 0.19–0.15 0.15–0.09 0.09–0.05 0.05–0.03 0.03–0.01

EQRMenhinick 1 0.75 0.38 0.13 0
Evenness E2 0.96–0.77 0.77–0.46 0.46–0.30 0.30–0.21 0.21–0.09
EQ REvenness E2 1 0.75 0.33 0.12 0
N 1000–4160 4160–31,400 31,400–188,334 188,334–710,470 710,470–10,000,000
EQRN 1 0.99 0.96 0.73 0
Chl a <0.10 0.10–0.40 0.40–0.60 0.60–2.21 >2.21
EQRChl a 1 0.96 0.82 0.73 0
IPI 1 0.90 0.72 0.53 0
846 S. Spatharis, G. Tsirtsis / Ecological Indicators 10 (2010) 840–847
chlorophyll a, however the differences span one quality level at
most. This discrepancy is more or less expected since abundance
and chlorophyll a are two different phytoplankton metrics
(Domingues et al., 2008). Chlorophyll a, being a photosynthetic
pigment, is indeed present in all phytoplankton cells, but it only
represents a fraction of the whole biomass. Moreover, heterotro-
phic species, not represented in chlorophyll a measurements
(Domingues et al., 2008), may form important peaks in abundance
that may be associated with other forms of disturbance such as
allochthonous and autochthonous organic inputs rather than
eutrophication. Therefore taking into consideration community
structure as a whole, seems a more representative measure of
water quality than chlorophyll a alone.
In the current study, simulated phytoplankton communities
resembling the field communities and the changes they undergo
due to eutrophication formed the basis for the evaluation of 22
indices. Simulation offers the opportunity to reduce the undesir-
able variability often encountered in field data due to uncontrolled
factors (seasonality, patchiness, hydrodynamic circulation) or
stochastic processes. Moreover it is possible to insert fully
controlled perturbations, assess the response of indices and infer
possible explanations for their behavior (Boyle et al., 1990). The
log series model, which was applied, successfully reproduced the
structure of phytoplankton communities of the Aegean coastal
waters, in terms of diversity, evenness and dominance.
The monotonic behavior of the tested indices was set as a
prerequisite since the water quality categorization in WFD is
performed in a rank order. Some indices showed non-monotonic
behavior, among them the Shannon and Simpson indices, two of
the most used measures of diversity in community ecology (Digby
and Kempton, 1987). These indices showed a hump-shaped
behavior that may be initially related to the increase in species
richness (expressed by the Hill’s N0 index), which is later
overturned for abundances higher than 103 cells/L by the decrease
in the community evenness (expressed through the evenness
indices). This index has been already widely criticized (Boyle et al.,
1990; Karydis and Tsirtsis, 1996), mainly on the basis that its
response is mostly a function of changes in species richness rather
than in the relative abundance of species (Simboura et al., 2005).
The sensitivity of indices to discriminate water quality can be
further enhanced by the linear or close to linear, trend of change
across the studied ecological gradient, since the average distance
between the set of limits is maximized. Among the indices
examined, a small number including the diversity index of
Menhinick and the evenness indices E2, E3, and E6 has passed the
screening procedure for both monotonicity and linearity. These four
indices were considered as the most efficient to assess water quality
for the implementation of the WFD. The scales developed on
Menhinick index of diversity and evenness E2 were used to assess the
water quality of several coastal sites in the Aegean. Between them,
the Menhinick index renders a more optimistic characterization by
classifying some sites of higher quality than the E2 index. The
Menhinick index is a simple index based on species richness and
total abundance, whereas evenness E2 includes the exponent of the
Shannon index carrying the full information on the distribution of
individuals to species (Washington, 1984).
A new synthetic index for water quality assessment in the WFD
is also proposed in the current study, considering that the
evaluation of water quality should be also based on suitable
multimetric indices (Borja et al., 2004; Domingues et al., 2008). For
this multimetric, namely the Integrated Phytoplankton Index (IPI),
three phytoplankton metrics were used that is chlorophyll a,
abundance, and diversity (Menhinick index) with equal weights.
The classification of areas with the IPI was consistent with the
other metrics, but was more optimistic than chlorophyll a or total
abundance, and more conservative than Menhinick’s diversity and
evenness E2 indices. It seems that the structure of phytoplankton
communities is more robust to the changes due to eutrophication
than the chlorophyll a content. Therefore it can be concluded that,
although there is a consistency, each metric carries its own
information and the use of multimetric indices combining various
phytoplankton metrics is appropriate in the framework of
integrated water quality assessment.
The classification of the Aegean coastal waters according to the
proposed metrics (ecological indices and IPI) is generally in
agreement with previous studies on nutrients (Ignatiades et al.,
1992; Primpas et al., 2008; Stefanou et al., 2000; Vounatsou and
Karydis, 1991) and phytoplankton community data (Ignatiades et
al., 1995; Karydis and Tsirtsis, 1996; Kitsiou and Karydis, 2000;
Tsirtsis and Karydis, 1998). The Rhodos offshore waters, which were
used as the reference site in the current study, were characterized as
a typical oligotrophic environment in the past by many authors
(Ignatiades et al., 1992; Karydis, 1992, 1994, 1996; Karydis and
Tsirtsis, 1996; Kitsiou and Karydis, 2000; Tsirtsis and Karydis, 1998;
Simboura et al., 2005; Stefanou et al., 2000), in agreement with the
assessment by the IPI. Coastal sites such as Rhodos inshore (coastal
and ports), the Gulf of Gera, the Strait of Mytilene and the outer
Saronikos have been previously characterized as mesotrophic
according to a three-level classification scheme (Arhonditsis et
al., 2000; Karydis and Coccossis, 1990; Tsirtsis, 1995). The use of a
five-level scheme in the current study allowed a more detailed
discrimination in the field of mesotrophy and the above sites were
characterized as of High, Good, and Moderate quality. Finally, sites
such as the port of Mytilene, and the Inner Saronikos and Kalloni
Gulfs are more heavily affected by anthropogenic activities and have
been previously characterized as eutrophic based on both physico-
chemical and biotic data (Ignatiades et al., 1992; Simboura et al.,
2005; Spatharis et al., 2007a,b), in agreement with the classification
of Poor quality in the current study. Potential dissimilarities of one
quality level at most may be related to the methodologies applied in
the past which were based on a relative local scale, whereas the
proposed metrics are suitable for assessments in the regional scale,
since they were based on a large number of available data from many
coastal areas in the Aegean coastal waters.
The characterization of coastal water quality constitutes a
critical issue for planning and decision making in Integrated
Coastal Zone Management (Karydis and Coccossis, 1990). There-
fore it must be based on robust tools able to synthesize the
information from more than one metrics or quality elements,
reducing in this way possible errors and misleading results. In the
current study an effort has been made for the development of such
qualitative tools for the Eastern Mediterranean Ecoregion. Further
testing of the proposed quality scales, as well as of the proposed
integrated index (IPI), need to be done in order to be established for
water quality assessment in the WFD. Moreover, the application of
the current screening methodology in a eutrophic west European
area would be very interesting (higher chlorophyll and nutrient
levels) and could contribute towards the enhancement of the on-
going research for the implementation of the Water Framework
Directive in European waters.
Acknowledgement
The current study was conducted in the framework of the
Program INTERREG IIIA, Gr-Cy (K.2301.001) co-funded by the
European Union and national funds.
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