Introduction

Harbours are among the most altered coastal areas. They usually represent polluted areas with low hydrodynamism, reduced oxygen in the water column, and high concentrations of pollutants in the sediment. Anthropogenic discharges into harbours and shallow bays, where residence times are extended due to partial enclosure, can have severe effects on local pelagic and benthic communities (Danulat et al. 2002). Ceuta harbour is one of the most important harbours in the Strait of Gibraltar. It has two opposing entrances connected by a channel (Fig. 1) which increases water renewal across the harbour. As a result, moderate oxygen levels are maintained in the water column and sediment heterogeneity is increased (Guerra-García 2001). Consequently, this area is a suitable site for analysing the relationships between macrofaunal assemblages and sediment variables, for elucidating the main factors affecting the spatial distribution of the soft-bottom fauna, and for evaluating possible positive effects of an increased water renewal on macrofaunal communities in harbours. Previous studies (Guerra-García and García-Gómez 2004a, 2004b) have dealt with the crustaceans and molluscs of Ceuta harbour. The present paper focuses on its polychaete assemblages.

Fig. 1
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Location of the harbour of Ceuta, North Africa. The 21 sampling stations are indicated (E1–E21)

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Polychaetes are among the most frequent and abundant metazoans in marine benthic environments (Fauchald and Jumars 1979). They are widely distributed geographically and occupy a variety of marine and estuarine habitat types (Belan 2003). Polychaetes often comprise over one third of the total number of macrobenthic species (Fauchald and Jumars 1979). In marine sediments they show high species richness and diversity as well as high biomass and density, up to 80% of the total benthos abundance (Belan 2003). Polychaetes have been found to be useful indicators of organic pollution, and many species have a high level of tolerance to adverse effects such as pollution and natural perturbations (Levin et al. 1996; Borja et al. 2000; Inglis and Kross 2000; Samuelson 2001). While polychaete communities associated with macrophytes have rarely been studied with respect to environmental variables (Sánchez-Moyano et al. 2002), relevant studies of soft-bottom polychaete assemblages are abundant (Nicolaidou and Papadopoulou 1989; Lardicci et al 1993; Pardal et al. 1993; Méndez 2002; Belan 2003). Environmental factors such as water movement, dissolved oxygen, granulometry of sediment and organic matter content have been demonstrated to play an important role in the distribution of soft-bottom polychaetes (Lardicci et al. 1993; Méndez 2002).

Usually, polychaete species richness and diversity inside harbours are low because of high pollution levels and a lack of oxygen in the water column (Estacio et al. 1997; Dhainaut-Courtois et al. 2000). The presence of two opposing entrances in Ceuta harbour, increasing water renewal and sediment heterogeneity, could have strong positive effects on polychaete assemblages inside the harbour. To test this hypothesis, we have studied and compared the physico-chemical parameters of sediments and associated polychaete assemblages at stations inside and outside the harbour.

Methods

Study area

The harbour at Ceuta is located in northern Africa, Strait of Gibraltar (Fig. 1). Inside the harbour there is a high variation in sediment characteristics (Guerra-García 2001). The presence of the San Felipe Channel promotes water renewal across the middle of the harbour and, consequently, increases sediment grain size. Nevertheless, there are also more enclosed areas where water renewal is reduced, and the mud content of sediments, silting, suspended solids and organic matter in the water column are high (Guerra-García 2001; Guerra-García and García-Gómez 2001). The harbour of Ceuta is characterised by an intense shipping traffic, and frequent loading and dumping is involved in shipping operations. There are two urban effluent outfalls originating from the city of Ceuta, but no river empties into the harbour. In contrast to some harbours in southern Spain, such as Algeciras Port (Estacio et al. 1997), there is no significant industrial activity adjacent to Ceuta harbour. Therefore, the contamination of the harbour is mainly derived from shipping activities and sewage disposal outfalls.

Sample collection

The sampling was carried out in June 1999. A total of 21 stations (15 inside and 6 outside the harbour) were chosen to encompass the broadest range of environmental conditions. The exact location of the stations was determined by the absence of rocky outcrops. Station E10, located in the San Felipe Channel, was considered as an external station. Sediments were collected with a van Veen grab of 0.05 m2. Four grab samples were collected at each station. Three of them were allocated to study the polychaete fauna (0.15 m2), and the fourth was used for sediment analysis.

Processing of biological samples

The sediment samples were sieved (mesh size of 0.5 mm) and the retained fractions were fixed in 4% neutral formalin stained with Rose Bengal. Organisms were sorted out by eye, identified to species level if possible, and counted.

Physico-chemical analysis

Sediments from each station were mixed and stored at −20°C in pre-cleaned glass jars until analysis, and then freeze-dried. Granulometry was performed according to Buchanan and Kain (1984). The percentage of sand was used as a granulometric indicator in the environmental matrix. The organic content was analysed by two methods: (1) by ashing sediment samples (three replicates of 2 g each) to 500°C for 6 h and re-weighing (Estacio et al. 1997); and (2) by oxidation using K2Cr2O7 (Loring and Rantala 1977).

To estimate the percentage of the lipid fraction, freeze-dried and homogenised sediments from each stations (approx. 5 g) were extracted in a 500 ml Soxhelt extractor for 24 h using a mixture of dichloromethane/methanol (9:1, v:v). The elemental sulphur was removed using copper powder (Hostettler and Kvenvolden 1994). The extracts were reduced in volume on a rotary evaporator and concentrated by gentle nitrogen “blow down”. The lipid fraction was determined by gravimetry and expressed as a percentage. The total nitrogen (ppm) was assessed via Kjeldahl digestion. The concentrations of P, Al, As, B, Ba, Ca, Cd, Co, Cr, Cu, Fe, K, Li, Mg, Mn, Na, Ni, Pb, S, Sr and Zn were measured using inductively coupled phasma spectrometry ICP-OES (Therme Jarrel Ash, IRIS Advantage) after digestion with aqua regia (HNO3:HCl; 1:3, v:v) in teflon-lined, high pressure decomposition vessels.

Statistical analysis

The total number of species, the Shannon-Wiener diversity index (Shannon and Weaver 1963) and Pielou’s evenness index (Pielou 1966) were calculated for each station. Possible differences between internal and external stations were tested using one-way ANOVA, after verifying normality (Kolmogorov-Smirnov test) and homogeneity of variances (Barlett test). A standard product-moment correlation analysis was conducted to reduce the number of variables considered. The Principal Component Analysis was used for the ordination of stations based on the physico-chemical data. Environmental data were log (x+1) transformed (Estacio et al. 1997; Guerra-García and García-Gómez 2001). The affinities among stations based on polychaete species were established by MDS (non-metric multidimensional scaling) and cluster analysis using UPGMA (unweighted pair-group method using arithmetic averages). To test the ordination, the stress coefficient of Kruskal was employed (Kruskal and Wish 1978). The relationships among environmental measures and polychaete assemblages were studied by a canonical correspondence analysis (CCA). The abundance data of polychaetes were transformed by the fourth root, and the Bray-Curtis similarity index was used (Sánchez-Moyano and García-Gómez 1998). Relationships between multivariate biological structure and environmental variables were also examined using the BIO-ENV procedure (Clarke and Ainsworth 1993). Percentage of similarity analysis (SIMPER) (Clarke 1993) was used to determine the species involved in grouping of the different stations. Multivariate analyses were carried out using the PRIMER package (Clarke and Gorley 2001) and the PC-ORD programme (McCune and Mefford 1997). For univariate analyses, the BMDP was used (Dixon 1983).

Results

Environmental variables

The standard product-moment correlation analysis of environmental data was useful to reduce the 27 variables measured (Table 1) to 11 variables. The %lipids, B, Ba, K, Na, S and OMc (organic matter: calcination method) showed a strong correlation (r>0.85, P<0.001) with the OMo (organic matter: oxidation method). Cu correlated (r>0.85, P<0.001) with Zn, Pb and As. Cr correlated (r>0.9, P<0.001) with Co, Mg and Ni. The correlation between Al and Li was significant (r>0.9, P<0.001), and Ca and Sr were also correlated (r>0.89, P<0.001). Therefore, the reduced matrix included the depth, % sand, OMo, N, P, Al, Ca, Cr, Cu, Fe and Mn.

Table 1 Values of sediment variables at the 21 sampling stations (E1–E21). OM Organic matter (c calcination method, o oxidation method); Lip lipids; P total phosphorous; N total nitrogen
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The first axis of the PCA ordination (Fig. 2) explained 54.03% of the total variance and correlated positively with the percentage of sands, and negatively with the organic matter and the concentrations of P, N, Al, Cr, Cu and Mn. The second axis, which explained 15.2%, was negatively correlated with depth and Ca. Consequently, the first axis ordinates the stations along a pollution gradient which is correlated with the type of sediment (the higher levels of pollutants in sediments were associated with lower sand content and, consequently, higher silt and clay content). The stations outside the harbour were located together with the inner stations affected by the water renewal across the San Felipe Channel in the PCA output, all characterised by high sand content, whereas stations E1, E2, E5, E6, E7, E15 and E16 were characterised by finer sediments with higher organic matter and heavy metal content. The second axis ordinates the stations mainly according to a depth gradient.

Fig. 2
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PCA analysis of the physico-chemical variables measured in the sediment of each station. Circles Internal stations, squares external stations

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Polychaete fauna

The 21 stations provided a total of 56 species. Table 2 shows the total abundance (ind/0.15 m2, sum of the three van Veen replicates) of each species at each different station. Many of the species were rare and occurred only at one or two stations. The most abundant species, Capitella capitata, Cirriformia tentaculata, Exogone verugera, Nereis falsa, Potamilla reniformis and Pseudomalacoceros tridentata, were mainly found at internal stations. The species richness and Shannon-Wiener diversity did not differ significantly between internal and external stations (Table 3), while the evenness index was slightly higher at the external stations, due to lower polychaete abundances (Fig. 3). The highest number of species were recorded at stations E4, E10 (channel) and E14 (inside the harbour), the highest diversity values at stations E4 and E9 (inside), and the highest evenness values at stations E17 and E18 (outside). It is remarkable that even the stations with extremely high levels of organic matter (particularly lipids) and heavy metals, such as the internal stations E5, E6, E7 and E16, are characterised by similar values of species richness and similar diversity indexes as the unpolluted external stations (Table 1, Fig. 3).

Table 2 Total abundance (ind/0.15 m2, sum of the three van Veen replicates) of each species at each different station
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Table 3 Mean values, standard deviations (SD) and ranges of the number of species, diversity (H’), evenness (J) per 0.15 m2 for internal (n=15) versus external (n=6) stations (ns not significant, * P<0.05)
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Fig. 3
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Species richness, Shannon-Wiener diversity index (H’), Pielou Evenness index (J) and abundance (ind/0.15 m2) at the 21 sampling stations

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When the multivariate approach was used, the external stations were clearly separated from the internal stations (Fig. 4). This indicates that, although species richness and diversity values are similar inside and outside the harbour, the species composition allows for a discrimination between internal and external stations, even better than environmental parameters (Fig. 2). The external stations E10 (Channel) and E17 (San Amaro) are also separated from the rest of the external stations.

Fig. 4
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Cluster classification and MDS ordination of the stations according to the abundance of polychaete species. Inner and outer stations are separated by doted line. Circles Internal stations, squares external stations

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Table 4 shows the average abundance of the most relevant species, listed in order of decreasing contribution to the average dissimilarity. The spionid Pseudomalacoceros tridentata was the species which contributed most to the dissimilarity between internal/external stations. It was present or abundant at all internal stations, but absent from all external stations. Capitella capitata, Potamilla reniformis and Nereis falsa followed the same general pattern, although they were present at some external stations. Cirriformia tentaculata and Platynereis dumerilii were not found outside the harbour. On the other hand, Jasmineira elegans, Parapionosyllis labronia, Glycera capitata and Anaitides maderiensis were only found outside the harbour. Hyalinoecia bilineata was abundant at station E10 (San Felipe Channel) and was also present at stations E4 and E9, which are affected by the water renewal across the harbour.

Table 4 Average abundances of the most relevant species of the stations located at the internal (INT) and external (EXT) sites. Species are listed in order of decreasing contribution to the average dissimilarity (Av. Dis.) between the two groups up to about 70% of accumulated total dissimilarity (Cum. Dis.%). The ratio indicates Dis./SD. The total average dissimilarity between groups is 88.02%
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Polychaete assemblages and environmental measures

The canonical correspondence analysis (CCA) confirmed the ordination of the stations previously obtained by the Cluster and MDS analysis. The external stations are distributed along the positive part of axis 1 (Fig. 5). Axis 1 seems to separate the stations according to a pollution gradient mainly determined by phosphorous and heavy metals such as Cu (and consequently Zn, Pb and As, highly correlated with Cu). Species such as Jasmineira elegans, Parapionosyllis labronica, Glycera capitata, Schistomeringos neglecta and Anaitides maderiensis, distributed in the external stations, are located at the positive end of axis 1, while Pseudomalacoceros tirdentata, Potamilla reniformis, Capitella capitata and Platynereis dumerilii are grouped at the negative end of axis 1, associated with the highest concentrations of P, Cu and organic matter in the sediment. Axis 2 is mainly correlated with the organic matter. This axis separates station E10 from the remaining stations based on the presence of Hyalinoecia bilineata. According to the BIO-ENV, the best combination of variables to explain the biological data (r=0.42) was obtained from the combination of five variables from the reduced matrix (Cu, P, Ca, Mn, %sand).

Fig. 5
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Graph representation of the canonical correspondence analysis (CCA). The species included are those which contributed most to the dissimilarity between internal and external stations according to the SIMPER (Table 4). Circles Internal stations, squares external stations

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Discussion

Since harbours are protected waterways, often with limited water circulation and surrounded by urban and industrial activities, pollutants frequently accumulate on the bottom over time (Reish and Gerlinger 1997). Ceuta harbour is considerably polluted, but provided with a channel which increases the water renewal inside the harbour (Guerra-García 2001). Due to this channel, the polychaete species richness inside Ceuta harbour is unusually high compared with harbours in southern Spain, such as Saladillo harbour, studied by Estacio (1996) and Estacio et al. (1997). This harbour, located in Algeciras Bay, shows similar levels of sediment pollution to Ceuta harbour. However, it has only one entrance and the macrofaunal communities are very poor inside this harbour. Even polychaete species, traditionally considered to be more resistant to pollution than crustaceans and molluscs, occur at considerably lower numbers in Saladillo harbour than in Ceuta harbour.

The positive effect of the channel is particularly evident for some species, such as Hyalinoecia bilineata which was found in high densities at station E10 (Channel) and stations E9 and E4, which are located in the area affected more by the water renewal through the channel. Although the channel mainly affects the central area of the harbour, it is remarkable that even the most enclosed areas are characterised by moderately high values of species richness and diversity. These values are considerably higher than in other harbours, in which the most enclosed areas are characterised by a complete absence of species (Dhainaut-Courtois et al. 2000; Méndez 2002).

In the present study, the univariate analysis was not able to discriminate clearly between internal and external stations, since species richness and diversity values were rather similar inside and outside the harbour. Only the evenness index was significantly higher outside the harbour due to the lower abundances at the external stations. Usually, the outer zones of harbours or bays are characterised by a higher number of species with few individuals (Sánchez-Moyano et al. 2002). The multivariate analysis based on species composition discriminated between internal and external stations much better than the univariate approach. Consequently, the species composition and species abundance differ between inner and outer sites. The presence of the channel seems to contribute to a high species richness in polluted sediments, and the type of pollution seems to determine which species can inhabit the area. The different composition of species inside versus outside the harbour, shown by the Cluster and MDS analysis, was also supported by the SIMPER. Under Ceuta harbour conditions (polluted sediments but high water renewal), the polychaete community, rather than crustaceans and molluscs (Guerra-García 2001), is a useful bioindicator to discriminate between internal and external stations.

In general, the soft-bottom polychaete fauna has proved to be an important tool for characterising the system with respect to granulometry, salinity, and organic matter content of sediments (Raman and Ganapati 1983; Pardal et al. 1993; Méndez 2002). Other authors, however, have pointed out that water depth is the factor which best explains polychaete distribution (Nicolaidou and Papadopoulou 1989). On the other hand, Lardicci et al. (1993) and Mistri et al. (2002) found that water movement and dissolved oxygen in the water column were the main factors determining polychaete assemblages, being more important than sediment grain size, salinity, etc. These results are in agreement with those of the present study, which has demonstrated that increased water renewal promotes the establishment of well-structured diverse polychaete communities.

The present study revealed that the observed distribution of polychaetes is not determined by one of the measured sediment variables per se, but by a combination of variables. Taking into account that internal and external stations can be distinguished better by polychaete assemblages (Fig. 4) than by physico-chemical parameters (Fig. 2), there may be further variables not analysed in the present study which could influence species distribution. For example, recent studies have shown that the proportion of maltenes and asphaltenes in the hydrocarbon fraction of the sediments, not usually measured, might be an important factor affecting macrofaunal assemblages (Guerra-García et al. 2003a).

The polychaete communities of soft-bottom sites seem to be a better bioindicator of environmental gradients than those associated with macrophytes. Recently, Sánchez-Moyano et al. (2002) studied the effect of environmental factors on the spatial variation of epifaunal polychaetes on the alga Halopteris scoparia in Algeciras Bay, Strait of Gibraltar, and found little variation.

Pseudomalacoceros tridentata and Capitella capitata were the dominant polychaetes inside the harbour. Capitella capitata is considered to be one of the global opportunistic species in disturbed marine sediments rich in organic matter (Grassle and Grassle 1974; Pearson and Rosenberg 1978; Grall and Glémarec 1997; Newell et al. 1998). The spionid Pseudomalacoceros tridentata has a great capacity for recolonising disturbed soft-bottom areas and can be found in dense aggregations inside harbour facilities (Guerra-García et al. 2003b). Jasmineira elegans, Parapionosyllis labronica and Glycera capitata were the most “sensitive” species and were found only at the external stations. Glycera capitata has been reported in outer areas of other harbours, while being absent from internal stations (Belan 2003).

Environmental implications of two-entrances harbours

Conventional harbours usually only have one entrance, and when they have two entrances, both are located at the same side (Yin et al. 2000). The existence of two opposing entrances and a connecting channel in Ceuta harbour contributes to increased water movement across the harbour, allowing the establishment of diverse polychaete communities even in heavily polluted sediments. These findings should be taken into consideration for future design, construction and remodelling of harbours.