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      Cryptic speciation of the zoogonid digenean Diphterostomum flavum n. sp. demonstrated by morphological and molecular data Translated title: Spéciation cryptique du Digène Zoogonidae Diphterostomum flavum n. sp. démontrée par des données morphologiques et moléculaires

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          Abstract

          Diphterostomum brusinae (Zoogonidae) is a digenean species that has been recorded worldwide parasitizing marine fishes. Several species have been synonymized with D. brusinae because they lack conspicuous morphological differences. However, due to the breadth of its geographic distribution and the variety of hosts involved in the life cycles, it is likely to be an assemblage of cryptic species. Diphterostomum flavum n. sp. is described here as a morphologically cryptic relative of D. brusinae, in the fish Pinguipes brasilianus (Pinguipedidae) off the Patagonian coast, Southwestern Atlantic Ocean, and its life cycle is elucidated through morphology and molecular analysis. This species uses the gastropod Buccinanops deformis (Nassariidae) as first and second intermediate host with metacercariae encysting within sporocysts. They also, however, use the polychaete Kinbergonuphis dorsalis (Onuphidae) as second intermediate host. No morphological differences were found between adults of D. flavum n. sp. and D. brusinae; however, the number of penetration glands of the cercariae, a diagnostic feature, differed (9 vs. 3 pairs), as well as the ITS2 sequences for the two species. This work provides morphological and molecular evidence of cryptic diversification among species described as D. brusinae, in which the only clear differences are in larval morphology and host spectrum. The strict specificity to the snail acting as the first intermediate host and the variety of fishes with different feeding habits acting as definitive hosts support the likely existence of multiple cryptic species around the world.

          Translated abstract

          Diphterostomum brusinae (Zoogonidae) est une espèce de digène, parasite des poissons marins, qui a été signalée dans le monde entier. Plusieurs espèces ont été synonymisées avec D. brusinae parce qu’elles ne montrent pas de différences morphologiques visibles. Cependant, en raison de l’étendue de sa répartition géographique et de la variété d’hôtes impliqués dans les cycles de vie, il est probable qu’il s’agisse d’un assemblage d’espèces cryptiques. Diphterostomum flavum n. sp. est décrit ici comme une espèce morphologiquement cryptique de D. brusinae, chez le poisson Pinguipes brasilianus (Pinguipedidae) au large de la côte de la Patagonie, dans le sud-ouest de l’océan Atlantique, et son cycle de vie est élucidé par la morphologie et l’analyse moléculaire. Cette espèce utilise le gastéropode Buccinanops deformis (Nassariidae) comme premier et deuxième hôte intermédiaire avec des métacercaires s’enkystant dans les sporocystes. Cependant, elle utilise également le polychète Kinbergonuphis dorsalis (Onuphidae) comme deuxième hôte intermédiaire. Aucune différence morphologique n’a été trouvée entre les adultes de D. flavum n. sp. et D. brusinae, mais le nombre de glandes de pénétration des cercaires, une caractéristique diagnostique, diffère (9 paires contre 3), ainsi que les séquences ITS2 pour les deux espèces. Ce travail fournit des preuves morphologiques et moléculaires de la diversification cryptique parmi les espèces décrites comme D. brusinae, dans lesquelles les seules différences claires sont dans la morphologie des larves et le spectre de l’hôte. La spécificité stricte au mollusque servant de premier hôte intermédiaire et la variété de poissons avec différentes habitudes alimentaires servant d’hôtes définitifs soutiennent l’existence probable de plusieurs espèces cryptiques dans le monde.

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          Molecular-genetic analyses reveal cryptic species of trematodes in the intertidal gastropod, Batillaria cumingi (Crosse).

          Cryptic species of the digeneans, Cercaria batillariae (Heterophyidae) and an undescribed philophthalmid, were detected using polymerase chain reaction-based restriction fragment-length polymorphism methodology and sequence analyses. These digeneans were all collected from the same species of gastropod first intermediate host, Batillaria cumingi (=Batillaria attramentaria). The mitochondrial cytochrome oxidase subunit 1 gene (approximately 800bp) and nuclear internal transcribed spacer 1 gene (approximately 400bp) were used for species level discrimination. Restriction fragment-length polymorphism analyses of cytochrome oxidase subunit 1 gene showed that C. batillariae included 10 distinguishable fragment patterns, and the philophthalmid included five patterns. On the basis of subsequent sequence analyses, the restriction fragment length polymorphism patterns of C. batillariae were grouped into eight phylogenetically distinct lineages and those of the philophthalmid into three phylogenetically distinct lineages. There was no evidence of gene flow among the different lineages due to the lack of heterozygosity within the observed internal transcribed spacer 1 gene fragment patterns. This suggests that all of these lineages are different species. Most of these species were widespread, but some exhibited restricted geographic distributions. We discuss the implications of these findings for host specificity of these trematodes. These results demonstrate the utility of genetic analysis to distinguish species of morphologically similar trematodes. Hence, trematode species diversity may often be underestimated when species identifications are limited to morphological features.
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            A comparison between mitochondrial DNA and the ribosomal internal transcribed regions in prospecting for cryptic species of platyhelminth parasites.

            We examined the relative merits of mitochondrial DNA loci and ribosomal DNA internal transcribed spacers for their use in prospecting for cryptic species of platyhelminth parasites. Sequence divergence at ITS1 and ITS2 was compared with divergence at 2 mtDNA loci (NADH dehydrogenase-1 and cytochrome c oxidase I) between closely related species of trematodes and cestodes. Both spacers accumulated substitutions substantially more slowly than mtDNA, which clearly shows a higher level of divergence among species relative to intra-specific variation. Besides a slow rate of substitution, other caveats that may be encountered when using ITS sequences as a prospecting marker are discussed. In particular, we note recent studies that suggest the existence of substantial levels of intra-individual variation in ITS sequences of flatworms. Because it is likely that closely related species share this phenomenon, it may confound the detection of cryptic species, especially if small sample sizes are studied. Although potential limitations of mtDNA are also recognized, the higher rate of evolution and smaller effective population size of this marker increases the probability of detecting diagnostic characters between cryptic species.
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              Revealing the secret lives of cryptic species: Examining the phylogenetic relationships of echinostome parasites in North America.

              The recognition of cryptic parasite species has implications for evolutionary and population-based studies of wildlife and human disease. Echinostome trematodes are a widely distributed, species-rich group of internal parasites that infect a wide array of hosts and are agents of disease in amphibians, mammals, and birds. We utilize genetic markers to understand patterns of morphology, host use, and geographic distribution among several species groups. Parasites from >150 infected host snails (Lymnaea elodes, Helisoma trivolvis and Biomphalaria glabrata) were sequenced at two mitochondrial genes (ND1 and CO1) and one nuclear gene (ITS) to determine whether cryptic species were present at five sites in North and South America. Phylogenetic and network analysis demonstrated the presence of five cryptic Echinostoma lineages, one Hypoderaeum lineage, and three Echinoparyphium lineages. Cryptic life history patterns were observed in two species groups, Echinostoma revolutum and Echinostoma robustum, which utilized both lymnaied and planorbid snail species as first intermediate hosts. Molecular evidence confirms that two species, E. revolutum and E. robustum, have cosmopolitan distributions while other species, E. trivolvis and Echinoparyphium spp., may be more geographically limited. The intra and interspecific variation detected in our study provides a genetic basis for seven species groups of echinostomes which will help accurately identify agents of disease as well as reveal cryptic aspects of trematode biology. Copyright 2010 Elsevier Inc. All rights reserved.
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                Author and article information

                Journal
                Parasite
                Parasite
                parasite
                Parasite
                EDP Sciences
                1252-607X
                1776-1042
                2020
                18 June 2020
                : 27
                : ( publisher-idID: parasite/2020/01 )
                : 44
                Affiliations
                [1 ] Laboratorio de Parasitología, Instituto de Biología de Organismos Marinos (CCT CONICET-CENPAT) Boulevard Brown 2915 U9120ACF Puerto Madryn Argentina
                [2 ] Laboratorio de Parasitología, IIPROSAM – Instituto de Investigaciones en Producción Sanidad y Ambiente, FCEyN, Universidad Nacional de Mar del Plata – CONICET Juan B. Justo 2550 7600 Mar del Plata Argentina
                [3 ] School of Biological Sciences, The University of Queensland Brisbane 4072 Queensland Australia
                [4 ] Laboratorio de Sanidade, ICBAS – Instituto de Ciencias Biomédicas Abel Salazar, Universidade do Porto R. Jorge de Viterbo Ferreira 228 4050-313 Porto Portugal
                [5 ] Laboratorio de Imunidade Inata e Ferro, I3S – Instituto de Investigação e Inovação em Saúde, Universidade do Porto R. Alfredo Allen 4200-135 Porto Portugal
                Author notes
                Article
                parasite200002 10.1051/parasite/2020040
                10.1051/parasite/2020040
                7301638
                32553099
                4cf2b91e-7c4c-4a62-9af1-191b23f8b7a6
                © C. Gilardoni et al., published by EDP Sciences, 2020

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( https://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 30 December 2019
                : 23 May 2020
                Page count
                Figures: 5, Tables: 2, Equations: 0, References: 82, Pages: 11
                Categories
                Research Article

                life cycle,zoogonidae,pinguipes brasilianus,cryptic species,patagonian coast

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