Hammerhead flatworms (Platyhelminthes, Geoplanidae, Bipaliinae): mitochondrial genomes and description of two new species from France, Italy, and Mayotte

Abstract

Background New records of alien land planarians are regularly reported worldwide, and some correspond to undescribed species of unknown geographic origin. The description of new species of land planarians (Geoplanidae) should classically be based on both external morphology and histology of anatomical structures, especially the copulatory organs, ideally with the addition of molecular data. Methods Here, we describe the morphology and reproductive anatomy of a species previously reported as Diversibipalium “black”, and the morphology of a species previously reported as Diversibipalium “blue”. Based on next generation sequencing, we obtained the complete mitogenome of five species of Bipaliinae, including these two species. Results The new species Humbertium covidum n. sp. (syn: Diversibipalium “black” of Justine et al., 2018) is formally described on the basis of morphology, histology and mitogenome, and is assigned to Humbertium on the basis of its reproductive anatomy. The type-locality is Casier, Italy, and other localities are in the Department of Pyrénées-Atlantiques, France; some published or unpublished records suggest that this species might also be present in Russia, China, and Japan. The mitogenomic polymorphism of two geographically distinct specimens (Italy vs France) is described; the cox1 gene displayed 2.25% difference. The new species Diversibipalium mayottensis n. sp. (syn: Diversibipalium “blue” of Justine et al., 2018) is formally described on the basis of external morphology and complete mitogenome and is assigned to Diversibipalium on the basis of an absence of information on its reproductive anatomy. The type- and only known locality is the island of Mayotte in the Mozambique Channel off Africa. Phylogenies of bipaliine geoplanids were constructed on the basis of SSU, LSU, mitochondrial proteins and concatenated sequences of cox1, SSU and LSU. In all four phylogenies, D. mayottensis was the sister-group to all the other bipaliines. With the exception of D. multilineatum which could not be circularised, the complete mitogenomes of B. kewense, B. vagum, B. adventitium, H. covidum and D. mayottensis were colinear. The 16S gene in all bipaliine species was problematic because usual tools were unable to locate its exact position. Conclusion Next generation sequencing, which can provide complete mitochondrial genomes as well as traditionally used genes such as SSU, LSU and cox1, is a powerful tool for delineating and describing species of Bipaliinae when the reproductive structure cannot be studied, which is sometimes the case of asexually reproducing invasive species. The unexpected position of the new species D. mayottensis as sister-group to all other Bipaliinae in all phylogenetic analyses suggests that the species could belong to a new genus, yet to be described.

Hammerhead worms everywhere? Modelling the invasion of bipaliin flatworms in a changing climate

“…Most terrestrial flatworms occur naturally in the tropics and in the temperate Southern Hemisphere, in a large range of habitat types, including from mesophile to xenophile habitats, from alpine herb fields to sandy semi‐desert and from subantarctic rata forest to tropical rain forest (Winsor et al, 1998). However, some species are found outside their native range in large parts of the world such as Europe (Cannon et al, 1999; Čapka & Čejka, 2021; Carbayo et al, 2016; Jones, 2019; Jones et al, 2020; Jones & Sluys, 2016; Justine et al, 2014, 2020, 2022; Justine, Winsor, et al, 2018; Mori et al, 2022), the Americas (Justine et al, 2015, 2019, 2021), Asia (Chaisiri et al, 2019; Hu et al, 2019) and Polynesia (Justine, Lemarcis, et al, 2018; Justine, Winsor, et al, 2018), and some of them occasionally became invasive (Sluys, 2016).…”

The invasive land flatworm Obama nungara in La Réunion, a French island in the Indian Ocean, the first report of the species for Africa

“…The complete mitogenome of specimen JL449 was sequenced in the context of a project in progress. Sequencing was performed following the exact same protocol as described in detail in a recent paper (Justine et al, 2022a), with a k-mer of 125. The sequence corresponding to the cox1 gene was identified using ORF finder on the NCBI portal (https://www.ncbi.nlm.nih.gov/orffinder) and by comparison with a published complete mitogenome (Solà et al, 2015).…”

The invasive land flatworm Obama nungara in La Réunion, a French island in the Indian Ocean, the first report of the species for Africa

“…The cox1 sequence was deposited in GenBank as OM451228. The pool of contigs obtained after assembly was also datamined to look for the presence of alien DNA following Justine et al (2022). The phylogenetic tree (not shown) obtained on this contaminant DNA was based on the partial 5.8S and ITS1, which were aligned with corresponding sequences of slugs (Quinteiro et al, 2005;Rowson et al, 2014), trimmed and computed for a Maximum Likelihood (ML) phylogeny with the General Time Reverse model and 100 bootstrap replicates, all steps performed by MEGAX (Kumar et al, 2018).…”

“…The complete mitogenome of specimen JL449 was sequenced in the context of a project in progress. Sequencing was performed following the exact same protocol as described in detail in a recent paper (Justine et al, 2022), with a k-mer of 125. The sequence corresponding to the cox1 gene was identified using ORF finder on the NCBI portal (https://www.ncbi.nlm.nih.gov/orffinder) and by comparison with a published complete mitogenome (Solà et al, 2015).…”

“…Citizen science is highly effective for collecting information of land flatworms (Justine et al, 2022;Justine et al, 2020a;Justine et al, 2015;Justine et al, 2014Justine et al, , 2018Justine et al, , 2020bMori et al, 2022). One of us (ADM) initiated a citizen science project in La Réunion about land flatworms, supported by the local association ARBRE (Agence de Recherche pour la Biodiversité à La Réunion https://arb-reunion.fr/ ).…”