Integrative taxonomy in helminth analysis: protocols and limitations in the twenty-first century.

The term integrative taxonomy was coined in 2005 for the identification of microorganisms using morphological, molecular, pathological and ecological components. Since then, more than 200 scientific articles have been published using this term in the helminthology field in different geographical regions of the world, leading to accurate identifications of specimens and expanding the known biodiversity by describing novel species. Importantly, the responsible use of this approach has led to balanced results in which molecular data do not overshadow morphological, pathological or ecological information. By having different perspectives, new taxonomic situations have arrived including the definition of cryptic diversity, species complex and genotypes. In the present work, we present the main procedures and principles for conducting an integrative taxonomy study of parasitic helminth including collection and preservation of specimens, fixation and staining of worms for light microscopy and scanning electron microscopy, and histopathological, ecological and DNA analysis. This guide is aimed at researchers in their endeavor to analyze helminth parasites collected from wild or domestic animals with examples provided, but the protocols presented herein may be extrapolated for the analysis of helminths collected from other hosts (e.g. human or fish). One important aspect is that the recommendations presented herein derive from the authors' experiences, which will hopefully be of use to the reader.