Metabarcoding data may provide abundance information
As expected, and in accordance with other studies investigating the quantitative power of metabarcoding in spiders (Kennedy et al., 2020) or other taxa (Thomas et al., 2016; Deagle et al., 2018; Lamb et al., 2019; Krehenwinkel et al., 2017; Schenck, Geisen, Kleinbölting, & Traunspurger, 2019), our results indicate that the relative read abundance (RRA) of a taxon is positively related to its proportion in both weight and abundance. However, the strength of this relation is not constant across spider families, which dissipates the possibility of using a unique correction factor to derive abundance information from read counts for all taxa obtained in metabarcoding analyses. Indeed, similar studies have also found these differences in the factor linking RRA and individual abundance across spider families (Kennedy et al., 2020) or RRA and mass across different taxa in other animal groups (Thomas et al., 2016).
Interestingly, the observations of some of the families in the plots relating RRA to proportion in mass were consistently above or below the 1:1 line. Upon detailed inspection, the spider families mostly above the identity line corresponded to families with small juvenile individuals (between 0.9 and 2.4 mg in mass), while families mostly below the identity line corresponded to those with large juvenile individuals (between 5.5 and 18.7 mg). Families with an intermediate juvenile mass (2.5 to 3 mg) were not clearly above or below the 1:1 line. This suggests that taxa with small or large juvenile sizes might be respectively over or underrepresented by their reads counts with respect to their real weight proportion in the sample in metabarcoding analyses. Additional studies with spiders and other taxa would help determine if this is a consistent trend and, if so, if it is applicable to other groups apart from spiders.
We agree with previous studies stating that, albeit with caution and with a certain degree of uncertainty, using the RRA with the corresponding correction factors as a surrogate of the occurrence of a taxon in the sample still provides more precise information of the community composition than using presence/absence data (Lamb et al., 2019; Kennedy et al., 2020). However, these correction factors need to be developed individually for different taxa, for example using mock communities included as quantitative controls during metabarcoding (Lamb et al., 2019).
CONCLUSIONS
Our study suggests that incorporating immature stages of spiders in bioinventorying initiatives has a relevant effect on diversity estimates, because a considerable proportion of the species present as juveniles is not found among adults. This impact goes beyond simply modifying species richnesses, as it also alters the level of similarity among communities and how they compare to each other. These findings do not question the information provided by adult-based inventories, but add a novel yet relevant layer of knowledge previously overlooked that may influence some of the interpretations derived from biological inventories. Adding juvenile information to rapid biodiversity assessment protocols provides more accurate data regarding comparisons of community composition. Metabarcoding analysis of all stages present in a sample enables more effective monitoring strategies, and ultimately better-informed conservation decisions.
The proportion of reads obtained from metabarcoding for certain spider families was positively related to their proportion in weight and abundance in the sample, suggesting that metabarcoding data are to a certain extent quantitative. The strength of this relation, however, was not constant across families, as already reported in former studies. Nonetheless, the use of read counts appropriately transformed with taxon-specific correction factors as a proxy of the occurrence of a taxon in the sample could still provide more accurate information about the community composition than simple presence/absence data.
ACKNOWLEDGEMENTS
Collections were conducted under the corresponding permits kindly provided by the following individuals and institutions: Miguel M. de la Hoz (Picos de Europa), Elena Villagrasa (Ordesa), Maria Merced Aniz Montes (Aigüestortes), Angel Rodriguez Martin (Monfragüe), Angel Gómez Manzaneque (Cabañeros), and Blanca Ramos Losada (Sierra Nevada). We are grateful to all the people that contributed to the samplings as well as to all the park rangers that supported us in the field.
This work was funded by the Organismo Autónomo de Parques Nacionales (OAPN #485/2012) (to MA). MD was supported by APIF PhD fellowship from the University of Barcelona. Additional support was provided by 2017SGR73 (to MA) from the Catalan Government.
AUTHOR CONTRIBUTIONS
MA, JMO and MD designed the study. AE and MD performed the laboratory work. OW, MD and JMO performed the data analyses. MD, JMO and MA drafted the manuscript with contributions of all authors. All authors have revised and approved the final manuscript.
DATA ACCESSIBILITY
- Fasta file with sequences of adult specimens: Dryad, https://doi.org/10.5061/dryad.g79cnp5q0.
- Fasta file with sequences of immature specimens: Dryad, https://doi.org/10.5061/dryad.z08kprrcv.
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SUPPORTING INFORMATION
Additional Supporting Information may be found in the online version of this article:
Table S1 Information on the adult sequenced specimens used in this study.
Table S2 Final dataset containing all the MOTUs of immature stages of spiders obtained by metabarcoding.
Figure S1 Rarefaction curves showing the number of MOTUs per replicate against an increasing number of reads.
FIGURES