Toxoplasma gondii Clp family protein: TgClpB1 plays a crucial role in thermotolerance.

BACKGROUND: In 2003, Paul Hebert proposed DNA barcoding, based on the first half of a standardized gene, the Cytochrome c Oxidase subunit I (COI), to identify animals. Subsequently, two large-scale projects enabled the sequencing of more than 1.3 million putative species worldwide. Two decades ago, we decided to adopt this approach as a tool to investigate the freshwater zooplankton diversity of Mexican aquatic systems. Several copepod species have been described by us with the aid of this marker, mainly of the family Diaptomidae, particularly of the species-rich genera Leptodiaptomus and Mastigodiaptomus. We also re-described topotypes of the widespread M. albuquerquensis and documented the invasion of exotic cyclopid species.

METHODS: In Mexico, we have sequenced the COI of 1,725 free-living freshwater copepods, including 925 diaptomid calanoids and 811 cyclopid cyclopoids, representing up to 43.7% of the total specimens sequenced for North America. To delineate the putative species diversity, we used the Barcode Index Number (BIN) and the Assemble Species by Automatic Partitioning method and compared both. For Leptodiaptomus we prepared a Maximum Likelihood (ML) tree, for a detailed analysis.

RESULTS: Our results suggest that central-southeastern Mexico may represent a potential radiation center for speciose diaptomid genera like Mastigodiaptomus (15 species), Leptodiaptomus (eight species), and Arctodiaptomus, which likely constitutes a regional species complex yet to be described. A comparison of Mexican data with that from North America (NA) showed that the only truly widespread copepod species, distributed from Arctic latitudes to the central Mexican plateau, is Leptodiaptomus sicilis, while all others have more restricted distributions. From the total specimens sequenced in NA, the BIN count revealed 89 Molecular Operational Taxonomic Units (MOTUs), but only 47 of them have been identified to species level. In some cases, diaptomid haplotype variants have received different BINs for a single specimen. The taxonomic impediment appears to be more pronounced in Cyclopoida, with only 32% of the total 235 BINs identified to species level. Despite these limitations, the use of MOTUs from these baselines is valuable for biomonitoring changes in freshwater ecosystems. We found that in some cases, mostly where singletons represented a BIN, the Assemble Species by Automatic Partitioning (ASAP) method provided a better representation of MOTUs. Conversely, when haplotypes of different species, such as those found in the Leptodiaptomus novamexicanus complex, are closely similar, ASAP fails, but ML can distinguish them. Therefore, it is urgent to apply an integrative taxonomy approach to propose the most convincing hypotheses regarding these issues. This publicly available online copepod baseline represents a useful tool for exploring and understanding species distributions, detecting possible new species and translocations, and revealing centers of speciation in NA.