Sampling strategies and methodological procedures are a theme of discussion in all areas of Science, considering their potential impacts on results, interpretations, and conclusions. In the case of ecological studies based on recent foraminifera, most of papers vary on collecting single samples or replicates, on counting living and/or dead assemblages, and on the depth of the sediment layer collected. In this study we discussed the implications of the following questions, especially in the intertidal zone of a tropical estuary: (i) Do collecting a single sample or replicates affect results and interpretations? (ii) Are there differences in results and interpretations generated from counting living (stained) or dead assemblages?; and (iii) Do varying the sampling depth of sediment layers influence the results and interpretations? We defined three collection points along the mangrove margins associated to the Itapanhaú River (Bertioga, State of São Paulo, Brazil). In each point, we drilled three microcores with the depth of 20 cm, laterally distant 30 cm from each other. In each microcore sediment samples (3 mL) were extracted in the layer intervals of 0–1, 4–5, 9–10, 14–15, and 19–20 cm. Foraminifera from all the samples were compared regarding absolute abundances, species composition, diversity, evenness and, communities similarities. We observed that replicates produce better results than single samples, both for the living and dead assemblages. Pseudoreplication simulation produced divergent results. Replicates make possible the application of more accurate inferential statistical analysis because the spatial variations of the species are captured. There were significant differences in ecological indicators and respective interpretations between analyzes based on counting living and dead assemblages so that they should not be considered directly comparable to each other. Counting living foraminifera requires higher effort for collecting and picking tests to reach the statistical minimum recommended. Regarding sampling depth, the first centimeter seems to be sufficient to find the vast majority of living foraminifera, mainly to estimate species richness. However, thicker sediments result in a longer time interval integrated into the sample, so the time-scale and variables that control foraminifera may change for interpretation. Our findings are consistent with previous studies carried out in subtropical estuaries, also expanding methodological recommendations to tropical estuaries. Future studies should explore the best distances among replicates.
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