Summary At the time of description, the morphology of Ptychaphelenchus eucalypticola Hodda, 2009 indicated it could be assigned to either the Aphelenchoididae Skarbilovich, 1947 (Paramonov, 1953) or the Parasitaphelenchidae Ruehm, 1956 (Siddiqi, 1980) within the Aphelenchoidoidea Skarbilovich, 1947 (Siddiqi, 1980). Although P. eucalypticola was, tentatively, and remains assigned to the Aphelenchoididae, its relationships with other aphelenchoids have not been reassessed, and no molecular data were previously available for this species. We re-collected P. eucalypticola from its type host and locality, Eucalyptus macrorhyncha F. Muell. ex Benth., from Mount Ainslie, ACT, Australia. We performed Bayesian inference and maximum likelihood analyses of a concatenated 18S + 28S rDNA gene sequence dataset to determine the position of P. eucalypticola within the Aphelenchoidoidea, followed by 18S and 28S single-gene analyses to further assess relationships between this species and an expanded set of close relatives. All analyses indicated P. eucalypticola is correctly assigned to the Aphelenchoididae, in a clade comprising all species of Ficophagus Davies & Bartholomaeus, 2015 and some species presently assigned to Aphelenchoides Fisher, 1894, sister to Martininema Davies & Bartholomaeus, 2015 and additional species of Aphelenchoides. Our 18S single-gene analyses did not resolve the position of P. eucalypticola relative to Aphelenchoides and Ficophagus; however, our 28S single-gene analyses indicated a sister relationship between P. eucalypticola and Ficophagus. This sister relationship is plausible as the former species shares many characteristics with species of the latter genus; however, there are sufficient morphological differences to consider P. eucalypticola as representative of a distinct lineage within the Aphelenchoidoidea.
Difficulties inherent in the morphological identification of cyst nematodes of the genus Heterodera Schmidt, 1871, an important lineage of plant parasites, has led to broad adoption of molecular methods for diagnosing and differentiating species. The pool of publicly available sequence data has grown significantly over the past few decades, and over half of all known species of Heterodera have been characterized using one or more molecular markers commonly employed in DNA barcoding (18S, internal transcribed spacer [ITS], 28S, coxI). But how reliable are these data and how useful are these four markers for differentiating species? We downloaded all 18S, ITS, 28S, and coxI gene sequences available on the National Center for Biotechnology Information (NCBI) database, GenBank, for all species of Heterodera for which data were available. Using a combination of sequence comparison and tree-based phylogenetic methods, we evaluated this dataset for erroneous or otherwise problematic sequences and examined the utility of each molecular marker for the delineation of species. Although we find the rate of obviously erroneous sequences to be low, all four molecular markers failed to differentiate between at least one species pair. Our results suggest that while a combination of multiple markers is best for species identification, the coxI marker shows the most utility for species differentiation and should be favored over 18S, ITS, and 28S, where resources are limited. Presently, less than half the valid species of Heterodera have a sequence of coxI available, and only a third have more than one sequence of this marker.
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