Introduction

The meiofauna is composed of small organisms that live in or on sediments, animals and plants. They are small invertebrates, in most cases no larger than 1 mm, and can measure up to 45 µm.

Mare (1942) used the term "Meiofauna" which is derived from the Greek meios "smaller", i.e., organisms smaller than macrofauna, but larger than microfauna. Thus, grouping together all metazoans that are retained in a 38 µm sieve, he concludes by defining meiofauna.

Nematodes are considered to be the most abundant metazoans in marine sediments with densities of thousands of organisms per square meter (Gerlach, 1971). In fact, their ecology is related not only to their remarkable quantitative occurrence in the benthos, but also in their fundamental role within the trophic chains of aquatic ecosystems, the incidence they have on marine currents and their function in stabilizing trophic effects on the shores (Platt and Warwick, 1980).

Marine currents cause a strong mixing that tends to homogenize the physical and chemical characteristics of the water, both horizontally and vertically (Cruz, 1980), being important in controlling the distribution of benthic communities and in the direct effects on the growth and physiology of organisms.

Heip (1982) considers that nematodes and copepods are the most abundant in almost all habitats, making them ideal for ecological and statistical analyses. The second place in numerical importance is occupied by harpacticoid copepods.

According to Vincx & Heip (1991), nematodes are particularly remarkable for their persistence, being found in all environmental conditions.

Benthic meiofauna can also be defined as a complex of organisms adapted to interstitial life between sediment particles. In recent years there has been an increase in studies aimed at using meiofauna as an indicator of anthropogenic impact (Warwick, 1993).

Vincx (1996), after several works related to the meiofauna, categorizes twenty-three higher taxa among which we have: Nematoda, Turbellaria, Oligochaeta, Polychaeta, Copépoda, Ostrácoda, Mystacocarida, Halacaroidea, Hydrozoa, Nemertina, Entoprocta, Gastropoda, Aplacófora, Brachiopoda, Holothuroidea, Tunicata, Priapúlida, Sipuncúlida.

Cruz (1998), points out that the term meiofauna is used to identify organisms smaller than the macrofauna, but larger than the microfauna, the latter term is now restricted only to protozoa, although some groups due to their size are integrating the meiofauna and are studied as part of this classification.

The habitat versatility of nematodes occupying any niche that provides an available source of organic carbon in marine, freshwater and terrestrial waters Bongers et. al., (1999).

It is important to note that benthic infauna communities, especially meiofauna, are affected in various ways within a disturbed zone. Dissolved oxygen concentration and fresh organic matter content are the main abiotic factors controlling bioturbation in eutrophic sediments. Therefore, the fauna may vary depending on the type, intensity and frequency of disturbance. In general, in coastal areas the biomass of meiofauna is lower than that of macrofauna (Gutiérrez, 2000).

The benthos is one of the most important groups in the flow of matter and energy in the ecosystem (Neira, 2001). Therefore, currents are very important in the distribution of these communities, along with the type of sediment. However, due to their higher turnover rates, meiofauna play a proportionally greater role in energy transfer (Neira, 2001).

It should be noted that due to their great diversity, meiofaunal organisms have been used as an excellent indicator of water quality in environmental biomonitoring, and the identification of their numerous species is a great scientific contribution (Ruiz et. al., 2005).

The objective of this research is to identify the biodiversity and interannual variation of the meiofauna by analyzing the response of the communities in relation to their structure, based on diversity and density.

The composition and quality patterns at the higher taxa level were evaluated by means of statistical tools currently in use and detailed in the methodology.

Materials and methods

During February and August 2004, 2007 and 2013, on the beaches of San Pedro de Manglaralto at the southern limit of the National Center for Aquaculture and Marine Research (CENAIM) latitude 01° 56' 30" S - longitude 80° 43' 30" W and in Salinas, latitude 02°12' 06" S - longitude 80° 57' 24" W located on the Santa Elena Peninsula, coast of Ecuador.

February was considered the wet season and August the dry season (Figure 1).

Figure 1.
Location of sampling stations: Station 1. San Pedro; Station 2. Salinas.

Sampling of meiofauna was applied with scientific criteria, since samples can only be used if they are representative of the population or group to be investigated, due to their morphodynamic variability and the difficulty of comparing them in different geographical areas.

The methodology used is divided into: Field Methodology and Laboratory Methodology.

The field methodology consists of: Sample collection, sealing - labeling, preservation and fixation of collected sediment samples.

The laboratory methodology consists of: Decanting, Washing, Centrifuging, Counting and Identification of the meiofauna.

Meiofaunal Counting and Identification

Stained organisms were placed with distilled water in a 10x10 counting chamber under the stereoscope (S9 D); each grid was traversed and meiofaunal species were determined under the 40x eyepiece. For more detailed species identification, a microscope (Leica DM500) was used under the 10x, 40x and 100x eyepieces. Once the samples were analyzed, they were again preserved in 4% formalin. Total density was expressed as number of individuals per 10cm² (ind/10cm) for interpretation purposes.

For the identification of the specimens we used the keys of the works of Bongers and Ferris (1999); Calles (1999); Calles (2001); Calles, Vincx, Cornejo and Calderón (2005); Ruiz, Domínguez, Marín and Miño (2006).

Statistical analysis of data.

A Check Date was performed to verify that there were no repeated data in terms of samples and groups. We worked with mean data, standard deviation, maximums and minimums, and 95% confidence intervals.

To estimate the specific species richness, the Margalef (D Mg) index (1951) was used, which mentions that an index with values lower than 2.00 represents a low species richness and, on the contrary, values close to 5.00 or higher reflect a high species richness.

The biological diversity of the meiofauna was calculated using the Shannon index. It is usually represented as H' and is expressed as a positive number, which in most natural ecosystems varies between 0.5 and 5, although its normal value is between 2 and 3; values below 2 are considered low and above 3 are high. It has no upper limit or in any case it is given by the base of the logarithm used (Shannon 1948).

The analysis of individual variables and abundance was performed with the STATISTICA 7.0 program using the Mann Whitney Test (Z statistic Mann Whitney; p<0.001), where "p" represented the statistically significant difference between the medians. The data obtained were used to compare two sample means taken from the same population, thus verifying whether two sample means are equal or not.

To determine if the effect of each factor in the design was statistically significant, a permutational analysis of variance (PERMANOVA) was applied, using Euclidean distance and 999 permutations to generate the sampling distribution.

The SPSS (Statistical Package for the Social Sciences) program with the Statistica 7.0 add-on was used for the multi-variable analysis.

Simple correlation similarity matrices were constructed, the analysis was carried out by the proximity scaling method, and stress levels or also called stress coefficients were obtained, which have to be aligned to 0 for the model to be adequate for the analysis and for the fit to be considered statistically adequate.

The Tucker model must be close to 1 to be certain that the model has a good fit. This algorithm was designed as an alternative for obtaining least squares estimators, decomposing the data into three markers associated with each source of variation analyzed.

From these matrices, the pattern of similarity between groups was analyzed, for which a numerical ordination by non-metric multidimensional scaling (MDS) was constructed. The differences in the multi-variable structure, based on the density of each taxa, were determined.

Results

Biodiversity of meiofauna

Specific species richness was estimated with the Margalef index (D Mg), where it was found that species richness is similar in both stations. However, there is no equity in the distribution of each species and this is due to the dominance of a single group.

According to the Margalef index, there is a functional relationship between the number of species and the total number of individuals, in San Pedro August 2007 was reported as the most diverse month with 2.30 bits/org, i.e. for every 2 individuals, an additional species will be found. The same occurred in Salinas where August 2004 was the most diverse month with 2.09 bits/org.

Based on the values recorded in this index, the meiofaunal community in San Pedro and Salinas reflected an average species richness in these months, since they presented a very small proportion of different species for the number of individuals of the same group found in the sample, this is due to the fact that the dominant group in both stations was the phylum Nematoda (Figure 1).

Figure 1.-
Diversity values for H'(Margalef) expressed in bits/org at the San Pedro and Salinas stations.

In terms of diversity, the Shannon-Weaver index (H'), which expresses differences between communities based on the number of species present and their relative abundance, the meiofauna showed significant heterogeneity in its values between seasons where a succession of dominance was evident among the genera; Ceramonema33%, Rhynconema 22% and Daptonema 19% belonging to the most abundant group (phylum Nematoda). August 2007 was reported as the most diverse month with 0.39 bits/org in San Pedro; and August 2004 with 0.60 bits/org in Salinas.

It can be observed that this range provided a low value of diversity, which concludes that it is not a very diverse population, since there is a high uniformity in a single group. When comparing the values for this index, they denote very low diversity in the sampling stations; investigations such as those of Calles A, K. (2001), Calles, A., Vincx, M., Cornejo, P., and Calderón, J. (2005), obtained values similar to those recorded in this study, which indicates the low biological diversity of this type of ecosystems dominated by Nematoda (Figure 2).

Figure 2.-
Diversity values for H'(Shannon and Weaver) expressed in bits/org at the San Pedro and Salinas stations.

Variations in meiofaunal densities, abundance and composition.

The highest total density of meiofauna was recorded in Salinas in August 2013 replicate 1 with 1004 ind/10cm², while the lowest total density was also obtained in Salinas in August 2007 replicate 1 with 249 ind/10cm².

Using the Mann Whitney Test with a significant level (Z statistic Mann Whitney; p<0.001), the temporal variations of the total meiofauna registered a (Z=1.96; p<0.001***). No significant differences were obtained with respect to total meiofaunal density and nematode density between the two stations (Figure 3).

Figure 3.-
Total abundance of meiofaunal groups for San Pedro and Salinas.

At the San Pedro and Salinas stations, 9 groups were recorded, where the phylum Nematoda was the most abundant, representing 85% of the total meiofauna with a population of 18344 ind/10cm², followed by Turbellaria (9%), Polychaeta (3%) and Rotifera (3%).

Due to their low representativeness the remaining 5 groups: Harpacticoid copepods, Nauplii, Ostracoda, Isopoda and Halacaroidea were grouped as "Others" for percentage analysis and graphical representation (Table 1).

Table 1

Total relative abundance values for the meiofaunal groups

The phylum Nematoda was the most abundant group representing 85% of the total meiofauna with a population of 18344 ind/10cm², followed by Turbellaria (9%), Polychaeta (3%) and Rotifera (3%).

Regarding the percentage contribution of each taxon to the total abundance of San Pedro, Nematodes represented 83% of all samples analyzed, their highest contribution was recorded in February 2004 (replicate 1) with 91%, Turbellaria contributed 10% followed by Polychaeta (3%) Rotifera (4%) and Others (0.2%).

In Salinas, Nematodes reported their highest percentage representing 87% of all samples analyzed, their highest contribution was recorded in August 2013 (replicate 3) with 92%, Turbellaria contributed with 8% followed by Polychaeta (3%) Rotifera (2%) and Others (0.6%) (Figure. 4).

Figure 4.
Plotted values of total relative frequency of meiofaunal groups for San Pedro and Salinas.

In San Pedro, the abundance of the total meiofauna was recorded between 577 ± 183 ind/10cm² with a range of 307 to 876 ind/10cm². Nematoda abundance ranged from 480 ± 166 ind/10cm² with a range of 215 to 735 ind/10cm². Turbellaria was recorded as the second most dominant group with an abundance between 57 ± 22 ind/10cm² with a range of 18 to 91 ind/10cm² followed by Polychaeta with an abundance between 19 ± 12 ind/10cm² with a range of 2 to 40 ind/10cm² and Rotifera with an abundance between 20 ± 8 ind/10cm² with a range of 8 to 35 ind/10cm².

In Salinas, the abundance of the total meiofauna was recorded between 625 ± 238 with a range of 279 to 1004 ind/10cm². Nematoda abundance ranged from 539 ± 244 ind/10cm² with a range of 173 to 918 ind/10cm². Turbellaria was recorded as the second most dominant group with an abundance between 48 ± 21 ind/10cm² with a range of 20 to 85 ind/10cm² followed by Polychaeta with an abundance between 19 ± 9 ind/10cm² with a range of 7 to 35 ind/10cm² and Rotifera with an abundance between 15 ± 5 ind/10cm² with a range of 6 to 24 ind/10cm² (Table 2).

Table 2

Abundances, Maximums and Minimums by meiofaunal group (Ind/10cm²), San Pedro and Salinas.

Variation in seasonal patterns in meiofaunal communities.

There were significant differences between stations and groups of meiofauna, which are detailed below, analyzing the most representative groups. The highest abundance of meiobenthos was recorded in August 2013, the dry season in San Pedro with 2428 ind/10cm² and in Salinas with 2742 ind/10cm², a time when strongly marked negative anomalies were recorded throughout the Pacific Ocean basin where the Sea Surface Temperature (SST) maintained negative values, being higher in region 3 (west of Galapagos) that reached up to -0.7 ° C. In the region, negative anomalies were maintained, even increasing from -0.2 to -0.4°, while a slight increase in temperature for the season was recorded along the coast.

The lowest abundance in San Pedro was reported in August 2007 with 1089 ind/10cm² and in Salinas in February 2004 with 877 ind/10cm². Nematoda was the most abundant group of the total meiofauna in both stations, the highest densities were recorded in August 2013 replicate 1 with 735 ind/10cm² for San Pedro and 918 ind/10cm² for Salinas. A steady decline was recorded from August 2004 to August 2007.

The lowest abundance was reported in August 2007 replicate 1 for San Pedro with 215 ind/10cm² and 173 ind/10cm² for Salinas (Figure 5)

Figure 5.-
Variation of total temporal density (Ind/10cm²) Phylum nematoda.

Multivariate analysis of meiofaunal communities.

It was verified by means of the similarity analysis whose Stress-I index (effort coefficient) was 0.13112 cof (Optimal Scaling Factor) that the data obtained comply with the objective of minimizing the quadratic error, the Tucker index was also calculated whose value was 0.99137 cof, which was considered as a very high value close to 1.

The estimates of the Tucker model (TLI) were obtained by applying the iterative process of the algorithm, with a 99.84% fit, taking into account the highest absolute values of each group within the stations.

Table 3

Stress measures and Tucker's congruence coefficient Groups of the total meiofauna

Based on the abundance values of the meiofaunal groups present in the stations, a multidimensional scaling analysis was performed to identify similarities in the community structure.

The MDS applied to the abundance values revealed that there was greater variation between the densities of the groups within the same station than between the stations themselves; therefore, it was not considered that the difference in densities was due to the seasonality of the sampling.

Likewise, the samples also showed patterns of similarity among the most abundant groups of the meiofauna, so that, based on the results obtained in the multivariate analysis, it was possible to group those most similar to each other in terms of the presence of individuals.

Three prominent clusters were generated. The first cluster (a) headed by Nematoda, followed by Turbellaria, Polychaeta and Rotifera reflects the dominance of these groups representing high similarity, the second cluster (b) shows the groups showing medium similarity and the third cluster (c) represents the groups with low similarity (Figure 6).

In some ways, as observed in the analysis of the total meiofaunal community, the spatial distribution of stations appears to reflect a geographic pattern, i.e., geographically close stations. However, the closeness of the stations in the MDS is due to other factors such as a higher density of individuals by more representative groups, since the sampled stations are not geographically close to each other.

Figure 6.
Similarity ordering using the non-metric multidimensional method. Groups of the total meiofauna.

The phylum Nematoda proved to be the most dominant group; this always appears as the most important group of the meiofauna, a condition previously reported by Coull (1988); free-living nematodes usually make up 90% of all meiofauna (Mc-Intyre, 1971).

The dominance of nematodes over the other groups was due to the presence of areas with a greater amount of organic matter, a characteristic reported by Calles (1999), who recorded for the beaches of San Pedro and Salinas, a similar condition regarding the high dominance of nematodes over other groups, such as Copepoda and Polychaeta, due to the direct influence of organic matter, where SPM (Suspended Particulate Matter) values of 16.57 mg/l for Salinas and 83.40 mg/l for San Pedro were reported; POM (Particulate Organic Matter) values of 4.47 mg/l for Salinas and 8.20 mg/l for San Pedro.

The studies mentioned above were associated with the research conducted by Cantera et. al. (1994), whose results indicated that meiobenthos organisms feed on organic deposits, preferring environments rich in nutrients for their type of feeding, so they tend to dominate in areas with a high concentration of organic matter, whether natural or human.

Rudnick et. at., (1989) and Heip (1995), suggested that there could be two groups of meiobenthos, one that responds immediately to the increase in organic matter and the second that reacts late to the use of old detritus as a food source. Calles (1999) indicated that the low presence of other groups such as copepods and rotifers is due to the fact that these organisms can move freely, so that when the core is introduced, a column of water forms on the sediment, which is used by these organisms to avoid collection.

Anthropogenic activity constituted one of the greatest threats to the habitat of benthic communities. San Pedro and Salinas, the beaches chosen for our study, reported strong anthropogenic pressure, which had a marked impact on the interstitial fauna and on the functioning of the coastal system (Calles 1999).

Gray (1981) reported that beaches with high levels of anthropogenic activity generate greater sediment compaction, making it difficult for organisms to penetrate, decreasing oxygen tension and the amount of meiofauna in the sediment.

The diversity values did not show a marked variability, so San Pedro and Salinas were defined as areas of medium diversity (2.10 - 2.29 bits/org), similar to the ranges recorded by Calles (1999). However, the abundance of organisms varied in both stations; it decreased notably in Salinas during February 2004 because the maximum concentration of individuals was found in few species, based on the numerical distribution of the individuals of the different groups, according to the total number of individuals in the sample analyzed.

In relation to the other groups of meiofauna that were reported, such as Turbellaria, Polychaeta and Rotifera, it was considered that their densities were related to a high granulometric concentration with values ranging from 1.6 mm to 2.1 mm, which indicated, from the granulometric point of view, an increase in the biological richness of the beaches due to the reduced hydrodynamic stress and sedimentary stability, which favored the presence of a large number of species; a particularity that was described in the research carried out by Olafsson et. al, (1991) and Calles et. al., (2005) in which high densities of meiofauna were also recorded in the localities of greater granulometry in relation to those of lower grain size.

Among the factors that were directly related to the interannual variation in the composition of the meiofauna of San Pedro and Salinas in the dry and rainy seasons, temperature and salinity were found, although no significant relationships were found with these variables. Domínguez (2001) and Calles et. al., (2001) reported SST values between 21.4 and 27.5 C where the maximum value was observed in the rainy season and salinity between 32 and 34 PSU which reported its lowest value in the rainy season, presenting a positive correlation: the lower the temperature and the higher the salinity, the higher the densities of total meiofauna and nematodes in the infralittoral zone.

In addition, meiobenthic species, especially nematodes, have been recorded as highly tolerant to brackish water (Platonova and Galtsova, 1985 and Santos et. al., 1996), which contributes to their high density and abundance observed in the study area.

In intertidal habitats, Coull (1999) suggested that there is a tendency for abundance and number of species to decrease as distance from the estuarine or freshwater source decreases. This was not a cause of declining abundance in our investigation since there was no nearby estuarine source directly influencing the sampling area.

Conclusions

Biodiversity in the communities of San Pedro and Salinas was related to areas of medium diversity, registering maximum values of 2.30 and 2.09 bits/org respectively. This revealed that the sites sampled in this study are heterogeneous and of low similarity. Nevertheless, the values of the richness (Margalef) and diversity (Shannon) indices were relatively similar in comparison with previous studies carried out in these localities.

There are significant differences in terms of inter-annual variation, since there was a considerable reduction in the number of groups in the rainy season, such as: Rotifera, Copepoda and Ostracoda, compared to the dry season.

In terms of richness (diversity), the meiofauna recorded low values where Nematoda was the phylum that predominated throughout the study, which presented the highest richness of genera and a high index of dominant species such as Ceramonema, Rhynconema and Daptonema.

The similarity analysis showed that the most statistically representative groups are also the most dominant, headed by Nematoda, followed by Turbellaria and Polychaeta with the genera: Capitella sp. and Sabella sp.

The structure of the meiofauna was not affected by temporal changes in the availability of resources (space and food) and the influence of environmental parameters, so that the significant differences between densities recorded in both the dry and rainy seasons were not significantly reflected by seasonality.

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