﻿Tapeworms (Platyhelminthes, Cestoda) from marine chondrichthyans of the Southwestern Atlantic Ocean, and the sub-Antarctic and Antarctic islands: a checklist

﻿Abstract A parasite-host list of cestodes parasitizing chondrichthyans in the Southwest Atlantic off Argentina and surrounding waters of Antarctica is compiled based on the available literature. The list is based on published descriptions and redescriptions of species, and newly collected worms during the current study. A total of 57 valid species belonging to 28 genera of the orders Cathetocephalidea, Diphyllidea, Gyrocotylidea, Lecanicephalidea, Onchoproteocephalidea, Phyllobothriidea, Rhinebothriidea, “Tetraphyllidea”, and Trypanorhyncha is listed. Information on hosts, localities, specimens in collections and comments on tapeworms are also included. A host-parasite list including chimaeras (1 order, 1 genus), batoids (4 orders, 10 genera), and sharks (3 orders, 5 genera) is provided. Tapeworm diversity, distribution range, and host associations are discussed. The cestodes orders Phyllobothriidea and Rhinebothriidea exhibit the highest species richness, with 13 and 12 species, respectively. Onchoproteocephalideans and rhinebothriideans have the broadest geographic distribution in the study area. Regarding hosts, arhynchobatid skates are the group most frequently associated with cestodes. However, further collecting efforts are necessary to understand whether this data reflect the real diversity and host association of these parasites or is a result of a bias in sampling.


Introduction
According to Froese and Pauly (2022), more than 500 species of fishes have been registered along the Southwestern Atlantic off Argentina and the sub-Antarctic and Antarctic islands (including South Georgia, Elephant, and Joinville islands), including 100 chondrichthyan species (Table 1) (Gabbanelli et al. 2018;Concha et al. 2019;Froese and Pauly 2022). Since these cartilaginous fishes are the definitive hosts of a great diversity of adult cestodes (Caira and Jensen 2014), it is not uncommon to find a large variety of taxa of tapeworms along the Southwestern Atlantic and the Southern seas.
Complete and accurate species lists are essential for many biological disciplines such as ecology, conservation, and biogeography. Particularly, comprehensive fish cestodes datasets are necessary if we consider the usefulness of these parasites as biological tags for stock identification of their elasmobranch hosts in the context of overfishing and habitat degradation have profoundly altered the populations of marine elasmobranch (Dulvy et al. 2014;Irigoitia et al. 2017;Irigoitia et al. 2022). To date, cestodes from the Southwestern Atlantic, sub-Antarctic, and Antarctic regions were listed only in a few articles. These included a list of fishes and their tapeworms from South America (Alves et al. 2017), a compilation of marine invertebrates from the Argentine Sea focusing on taxonomic information at the generic level only (Bigatti and Signorelli 2018), and a few works about cestodes of Antarctic fishes (Rocka and Zdzitowiecki 1998;Rocka 2003Rocka , 2017. The analysis of the endoparasites in Antarctic fishes showed significantly higher values of diversity indices compared to the sub-Antarctic ichthyofauna (Muñoz and Cartes 2020); it would be interesting to consider the diversity of cestodes in a wider context, especially including the Southwestern Atlantic and southern latitudes off Antarctica in a single study. However, no complete work about cestodes from chondrichthyans, with detailed distributional ranges and host associations in this particular area of the Southern Hemisphere has been compiled so far.
In order to facilitate further studies, the main goal of this work is to elaborate a complete checklist of cestodes in chondrichthyan hosts based on summarizing references. The study area includes the Southwestern Atlantic Ocean off Argentina, Río de la Plata estuary, and the surrounding waters of South Georgia and the El ephant and Joinville islands by surrounding waters of South Georgia, El ephant and Joinville islands. This list includes information on localities, specimens in collections, and comments about the parasites and their hosts reported in previous works. We have also incorporated information about cestodes described in the last years, which included numerous new records and new localities (Menoret et al. 2017;Franzese and Ivanov 2018, 2020a, b, 2021Menoret and Ivanov 2021;Franzese et al. 2022; this study).

Materials and methods
After an exhaustive bibliographical search, an annotated and revised parasite-host checklist was generated for the adult cestodes from marine chondrichthyans reported between 35°S-63°S. The geographical area considered covers the Southwestern Atlantic Ocean off Argentina (from 35°S southward), Río de la Plata estuary, and surrounding waters off South Georgia, Elephant, and Joinville islands. The cestode species are arranged according to taxonomic categories and are presented alphabetically, followed by data on their hosts, including valid species name, order, family, and synonymous species name used in literature (if available) in parentheses. The information for localities includes location, coordinates in degrees and minutes (if available in the literature), province, and country (where applicable) only for the type locality. The type-host and the type locality refer to data included in the original descriptions of cestodes species.  1  3  0  Triakidae  2  4  3  Echinorhiniformes  Echinorhinidae  1  1  0  Hexanchiformes  Hexanchidae  3  3  1  Lamniformes  Alopiidae  1  1  0  Carchariidae  1  1  0  Cetorhinidae  1  1  0  Lamnidae  3  3  0  Squaliformes  Dalatiidae  2  2  0  Etmopteridae  2  6  0  Somniosidae  3  4  0  Squalidae  1  3  0  Squatiniformes  Squatinidae  1  3  1  Subtotals  6  16  27  45  6  Totals  11  28  46  100  33 Other hosts and other localities only refer to the records within the study area, including those in the original descriptions as well as those mentioned in redescriptions, other papers and newly collected materials sampled during the present study. Specimens in collections include type material from original descriptions, voucher specimens from redescriptions and new voucher specimens prepared during the present study. Information about the new voucher specimens is in bold. For the preparation of the figures, estimated coordinates were assigned to those records that lacked such information in the original publication.
Based on the information from the parasite-host checklist, the host-parasite data were subdivided into two inventories, one for batoids and chimaeras and another for sharks. The host species are arranged according to taxonomic categories and presented alphabetically, followed by the data on their parasites.
New vouchers of cestodes were obtained from the spiral intestines of chondrichthyans that had been caught by commercial trawlers between 2009 and 2017. The spiral intestines were fixed in 10% formalin and transferred to 70% ethanol for storage in the Laboratorio de Sistemática y Biología de Parásitos de Organismos Acuáticos (SIBIPOA) of Instituto de Biodiversidad y Biología Experimental y Aplicada (IBBEA, CONICET-UBA). Cestodes were hydrated in a graded ethanol series, stained with Harris' hematoxylin, dehydrated in a graded ethanol series, cleared in methyl salicylate, and mounted in Canada balsam (Menoret and Ivanov 2021;Franzese et al. 2022).
The accession numbers of the available molecular sequences were taken from the Gen-Bank database, considering only those specimens whose identification is not doubtful.

Acanthobothrium stefaniae Franzese & Ivanov, 2018
Type host. Discopyge tschudii Heckel (Torpediniformes: Narcinidae).  Brooks et al. (1981) reported two specimens of Acanthobothrium sp. from M. goodei at Río de la Plata, which could be a different species. They pointed out that one of these specimens could correspond to the same species reported by Ostrowski de Núñez (1971) in Z. brevirostris. Beer et al. (2019) reported Acanthobothrium sp. from B. cousseauae and B. magellanica off Malvinas Islands. The deposited material only corresponds to the specimens studied by Brooks et al. (1981).

Locality. Malvinas Islands Shelf, Southwestern Atlantic Ocean.
Reference. Beer et al. (2019). Comments. These specimens were studied by Beer et al. (2019) at a molecular rather than morphological level, without reaching an identification at the specific level.  Brooks et al. (1981) reported ten specimens of Phyllobothrium sp. from M. goodei at the Río de la Plata estuary. The deposited material only corresponds to the specimens studied by Brooks et al. (1981).

Rockacestus siedleckii
Comments. Rocka (2017) established the name Rajicestus for cestodes from Antarctic and sub-Antarctic skates described originally in Wojciechowska (1991b) as members of Phyllobothrium. Unfortunately, no generic diagnosis or type species was designated; therefore, the name Rajicestus is unavailable.  Beer et al. (2019) studied these specimens at the molecular rather than the morphological level, without reaching generic or specific identification. Caira et al. (2021) noted that the specimens of Phyllobothriidea gen. sp. found by Beer et al. (2019) could correspond to the genus Rockacestus; however, further molecular and morphological studies are necessary to identify them at the specific level. Beer et al. (2019) also pointed out the presence of Phyllobothriidea gen. sp. parasitizing D. chilensis. Nevertheless, the distribution of D. chilesis is restricted to the Pacific Ocean; therefore, this record is based on a misidentification of the host (Concha et al. 2019 References. Carvajal and Dailey (1975), Franzese et al. (2022). Reference. Ivanov and Campbell (2002).

Genus incertae sedis and other forms with uncertain family allocations
Comments. Ivanov and Brooks (2002) redescribed C. australis based on the material studied originally by Ostrowski de Núñez (1973), who considered this species a subspecies of C. verticillatum.
Comments. Ruhnke (2011) considers P. myliobatidis as a possible member of the order Rhinebothriidea.

Geographical distribution of the cestode orders
The tapeworm orders reviewed in this study show different geographical ranges. These distributions are represented in Fig. 1A for the Phyllobothriidea, Fig. 1B for the Onchoproteocephalidea, Fig. 2A for the Rhinebothriidea, Fig. 2B for "Tetraphyllidea" and Gyrocotylidea, Fig. 3A for the Diphyllidea and Lecanicephalidea, and Fig. 3B for Trypanorhyncha. Geographical sites of the order Cathetocephalidea could not be rep-  resented since the only existing record reports Cathetocephalus australis in Argentina, without specifying the locality or coordinate. The orders with the broadest geographic distributions are Onchoproteocephalidea (Fig. 1B) and Rhinebothriidea ( Fig. 2A), with representatives in the Río de la Plata estuary, along the Argentine Sea, and the southern islands. In addition, the phyllobothriideans (Fig. 1A) show a similar distribution, although without records between the 40°S-47°S latitudes, in the central region of the Argentine Sea. On the other hand, the cestodes with the narrowest distribution are those of the order Gyrocotylidea (Fig. 2B), being recorded only in two locations in the Buenos Aires Province.

Cestode diversity
Cestodes as parasites of chondrichthyans have been mostly recorded in the Northern Hemisphere (Caira et al. 2022). However, reports in southern latitudes have remarkably increased in the last decades due to focused sampling efforts in the area (Menoret and Ivanov 2012a, b, 2014 Ivanov 2018Ivanov , 2020aIvanov , b, 2021Palm et al. 2019;Oosthuizen et al. 2021;Franzese et al. 2022;Van Der Spuy et al. 2022). The present annotated checklist comprises 57 valid cestode species of 28 genera in nine orders, registered in the Río de la Plata estuary, Southwestern Atlantic off Argentina and the surrounding waters off Antarctica. The orders Phyllobothriidea and Rhinebothriidea show the highest richness at the specific level, with 13 and 12 valid species, respectively; they are followed by the order Trypanorhyncha, with a total of eight species. In addition, the list includes cestodes without an identification up to the generic or the specific level, as in the case of the Onchoproteocephalidea (i.e., Acanthobothrium sp.), Phyllobothriidea (i.e., Genus sp., Guidus sp., Phyllobothrium sp.), Rhinebothriidea (i.e., Genus sp., Echeneibothrium sp., Pseudanthobothrium sp.) and Trypanorhyncha (i.e., Grillotia sp.) (Ostrowski de Núñez 1971;Brooks et al. 1981;Beer et al. 2019). Several of the comments made in this work are intended to aid in future morphological and molecular studies addressing the lower taxonomic resolution of these entities.
In view of the high degree of specificity of adult cestodes to their marine hosts (Reyda and Marques 2011;Caira and Jensen 2017) and that only 33% (33/100) of marine chondrichthyans in the study area have been sampled for cestodes (Table 1), this fauna is probably underestimated. We can speculate that more than 60 species of cestodes have not yet been discovered in this area. Future taxonomic surveys will be essential to increase the knowledge of the diversity of these parasites in the region.

Taxonomic resolution
The poor taxonomic resolution of several taxa listed in the present study is probably a consequence of the lack of use of multiple tools to develop an integrative taxonomy, such as morphological and molecular studies used as evidence to delineate species boundaries. Some of the previous works cited here lacked modern morphological tools, e.g., scanning (SEM) and transmission electron microscopy (TEM) and the molecular tools necessary for the development of an integrative approach (Ostrowski de Núñez 1971;Brooks et al. 1981). In contrast, the recent work by Beer et al. (2019) recorded in the Argentine Sea numerous specimens of cestodes belonging to different orders but without achieving a specific identification for many of them, using molecular sequences as the only identification tool. The development and use of molecular tools have allowed the detection of cryptic species in some cestode groups (Scholz et al. 2014;Choudhury and Scholz 2020). Of the 57 valid species recorded in this work, only seven have been sequenced, so it is still unknown whether cryptic species will be discovered in this particular region. In addition to molecular sequences, the use of modern morphological tools, such as SEM and TEM, might be helpful in discovering new characters that complement traditional morphological studies, which could contribute to solve species identification problems Mutti et al. 2023). The development of the integrative taxonomy, including the use of all available tools, will allow resolving the poor taxonomic resolution observed in several taxa registered in our study area.
Considering that the major number of cestode species from this checklist are hosted by the myliobatiform Myliobatis goodei (Brooks et al. 1981;Ivanov and Campbell 1998a;Ivanov 2014, 2015;Menoret et al. 2017), it would be interesting to sample M. freminvillei Lesueur, the only species of myliobatid that has not been yet examined for cestodes in the region. On the other hand, only 13% (6/45) of the species of sharks have been reported as hosts in this area (Table 1). Host species with a relatively low occurrence or a particular bathymetric distribution are likely to host an undiscovered and exciting cestode fauna.
More collecting efforts are necessary to conclude if this data reflects the actual biodiversity of cestodes in the different groups of chondrichthyans or is a result of a bias in sampling. Although this list shows the substantial advances in taxonomical surveys in the last decades, expanding the number of sampled hosts is essential to increase the knowledge of the current cestode fauna of chondrichthyans in the region.

Studied area and newly collected material
Five species of cestodes have been recorded in new localities of the Southwestern Atlantic Ocean (Table 3). New material (voucher) identified, processed, and deposited in the MACN parasitological collection corresponds to three onchoproteocephalideans (i.e., Acanthobothrium domingae, A. marplatensis, A. stefaniae) and two rhinebotriideans (i.e., Echeneibothrium williamsi, Notomegarhynchus navonae). One of these records has extended until the Buenos Aires Province the northern limit of the known geographic distribution in the Argentine Sea of E. Williamsi, which, prior to this work, ranged from Santa Cruz Province to Tierra del Fuego Province (Franzese et al. 2022). The remaining new records have added new localities within the province of Buenos Aires for A. domingae, A. marplatensis, A. stefaniae, and N. navonae. Previously, these four species had been reported off Buenos Aires, although in different locations Campbell 1998b, 2002;Ivanov 2018, 2020a).
Several of the original descriptions of cestode species are based on material collected from a single locality. However, this probably reflects the absence of a more exhaustive sampling. The present checklist shows that about half of the species included in this region have additional localities. Among these, Rhinebothrium chilensis and Echeneibothrium williamsi show the highest number with 7 and 6 localities, respectively (Tanzola et al. 1998;Irigoitia et al. 2017). It is likely that as the intensity of sampling increases, new localities will be discovered for several known cestode species.
The localities with the most significant number of cestodes species are Puerto Quequén and Mar del Plata, with 17 and 11 species reported to date, respectively. A strong sampling effort could explain these numbers since both sites are commercial ports from the Buenos Aires Province close to the facilities of the main Argentinean research taxonomic cestodes groups (Luque and Poulin 2007;Randhawa and Poulin 2019).

Conclusions
Some difficulties concerning the understanding of chondrichthyan cestode diversity are: 1) many works have a poor taxonomic resolution or are outdated, with incomplete drawings and without the use of modern tools such as transmission electron microscopy, scanning electron microscopy and molecular approaches; 2) the existence of cryptic species underestimates the actual number of cestodes; 3) less than half of the marine chondrichthyans have been examined for cestodes in the area covered in this work.
A modern taxonomic approach for future characterizations should be made by combining descriptive tools (e.g., TEM and SEM, molecular data, histological sections, and histochemical techniques). It would also be desirable that all the voucher material could be available in public parasitological collections to facilitate its study to the entire community of taxonomists. Regarding sampling effort, it is likely that the higher the number of chondrichthyans examined in parasitological surveys, the higher the number of parasite-host associations will be identified. We have critically compiled as much detailed information as possible including valuable comments, providing a complete list of references and information from the deposited material. We hope this list may help future studies and contributes to correctly estimating the cestode biodiversity that inhabits this underexplored region of the Southern Hemisphere.