Abstract
The traditional identification of plant-parasitic nematode species by morphology and morphometric studies is very difficult because of high morphological variability that can lead to considerable overlap of many characteristics and their ambiguous interpretation. For this reason, it is essential to implement approaches to ensure accurate species identification. DNA barcoding aids in identification and advances species discovery. This study sought to unravel the use of the mitochondrial marker cytochrome c oxidase subunit 1 (coxI) as barcode for Longidoridae species identification, and as a phylogenetic marker. The results showed that mitochondrial and ribosomal markers could be used as barcoding markers, except for some species from the Xiphinema americanum group. The ITS1 region showed a promising role in barcoding for species identification because of the clear molecular variability among species. Some species presented important molecular variability in coxI. The analysis of the newly provided sequences and the sequences deposited in GenBank showed plausible misidentifications, and the use of voucher species and topotype specimens is a priority for this group of nematodes. The use of coxI and D2 and D3 expansion segments of the 28S rRNA gene did not clarify the phylogeny at the genus level.
Highlights
Conducted with 18S rRNA gene sequences[11, 15, 16] did not provide taxonomic clarity among Longidoridae, since this gene seems to evolve too slowly to be useful as an appropriate marker for phylogenetic studies at the species level
We did not perform a systematic analysis of primer amplification, as we started with the combination COIF/XIPHR2 in the majority of the studied samples; this combination was reported to be efficient in previous studies[21]
All new partial c oxidase subunit 1 (coxI) sequences were obtained using voucher specimens identified by integrative taxonomy, with the combination of morphological characteristics and unequivocal molecular markers from the same individual nematode, viz. the D2–D3 region (Tables 1 and S1) and ITS1 in some cases
Summary
Conducted with 18S rRNA gene sequences[11, 15, 16] did not provide taxonomic clarity among Longidoridae, since this gene seems to evolve too slowly to be useful as an appropriate marker for phylogenetic studies at the species level. Recent studies showed that mtDNA genes evolve much more quickly than rRNA genes, revealing low intra-specific and high inter-specific molecular variability for Longidoridae[12, 16, 18,19,20,21]. It seems to be the most promising marker to relieve taxonomic confusion within this group. The objectives of this research were to evaluate the variability of the mitochondrial marker gene coxI and partial sequence of the 28S rRNA gene within Longidoridae, as well as their usefulness as markers for barcoding and for reconstructing the phylogeny of the group
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