Stilbonematinae

Scharhauser, Florian, Zimmermann, Judith, Gruber-Vodicka, Harald R., Ott, Jörg A. & Leisch, Nikolaus, 2020, Morphology of obligate ectosymbionts reveals Paralaxus gen. nov.: A new circumtropical genus of marine stilbonematine nematodes, Zoologica Scripta 49 (3), pp. 379-394 : 390

4.1 | COI-based barcoding is not suitable for Stilbonematinae

The COI gene is the most common marker gene for PCRbased animal %meta) barcoding, despite the observation that highly diverse base composition has led to unreliable amplification results in many animal phyla including nematodes % Bhadury et al., 2006; Deagle, Jarman, Coissac, Pompanon, & Taberlet, 2014; Derycke et al., 2010). By utilizing single nematode metagenomics, we could overcome such amplification problems and show that the COI is a valuable phylogenetic marker gene to distinguish stilbonematine genera and species. However, analyses of the metagenome-derived full-length COI sequences revealed that within Stilbonematinae the COI gene has a highly diverse base composition, prohibiting the design of specific primers on any region of the COI gene. Our mismatch analyses for commonly used primer sets showed multiple mismatches as well as possible alternate binding sites. One may overcome priming mismatches by using very low stringency PCR conditions, at the cost of specificity. Armenteros, Ruiz-Abierno, et al. %2014) were able to amplify COI fragments from stilbonematine DNA extracts but morphological species descriptions and 18S-based results did not conform to the phylogenetic placement based on the COI. One possible explanation could be that the PCR picked up minimal contaminations of other Stilbonematinae with lower mismatch counts, instead of the gene of the target organism with high numbers of mismatches %Figure 5). Together, these factors show that the COI gene is not well suited for PCR-based barcoding in Stilbonematinae .