Neonchocotyle violantei Neonchocotyle violantei n. sp. (Monogenea, Hexabothriidae) from n. sp. (Monogenea, Hexabothriidae) from Pseudobatos lentiginosus Pseudobatos lentiginosus (Rhinopristiformes, Rhinobatidae) of (Rhinopristiformes, Rhinobatidae) of Yucatán, Gulf of Mexico = Yucatán, Gulf of Mexico = Neonchocotyle violantei Neonchocotyle violantei n. sp. n. sp. (Monogenea, do (Monogenea, Hexabothriidae) do Pseudobatos lentiginosus Pseudobatos lentiginosus (Rhinopristiformes, Rhinobatidae) de Yucatán, Golfo do México (Rhinopristiformes, Rhinobatidae) de Yucatán, Golfo do México

Neonchocotyle violantei n. sp. (Monogenea: Hexabothriidae) infects the gill of the Atlantic guitarfish, Pseudobatos lentiginosus (Rhinopristiformes, Rhinobatidae) from littoral waters of Celestún, Yucatán, Mexico. It is assigned to Neonchocotyle because it has, among other features, an asymmetrical haptor, a seminal receptacle, a smooth oötype, and an egg with two elongate filaments. It differs from Neonchocotyle pastinacae , the only congener, by having a small body (821 long by 315 wide, length to width = 2.6:1), two pairs of microhooks between the haptoral appendix suckers, extracaecal (submarginal) vaginal pores, and 5-9 testes. This is the first record of a species of Neonchocotyle in the Gulf of Mexico, the first monogenean reported from P. lentiginosus in Mexico and the second species of Hexabothriidae reported from Mexico.

An individual of Pseudobatos lentiginosus (= Atlantic guitarfish) was collected by a local fisherman in littoral waters of Celestún, Yucatán, Mexico, adjacent to the Reserva de la Biósfera Ría Celestún Ramsar site no. 1333
guitarfish was collected from the shallow marine platform nearby the mouth of the river and the costal lagoons (Ficha Informativa de los Humedales de Ramsar, Reserva de la Biosfera Ría Celestún. Given the magnitude of biodiversity of this region (CIUDAD DE MÉXICO, 2000), the discovery of a new species of Hexabothriidae is unsurprising; this species is described herein and the genus amended to include previously undocumented characters.

Materials and Methods
One Atlantic guitarfish was collected by a local fisherman using a long line in littoral waters of Celestún, Yucatán (20°51'33"N; 90°24'00"W), Gulf of Mexico, in February 1999. The external body surface of the guitarfish was examined using a magnifying glass. Each gill arch was excised, placed in a Petri dish with seawater, and examined using a stereomicroscope. Monogeneans, dead at the time of collection, were removed from gill lamellae and transferred temporarily to dishes containing seawater. When all worms had been collected, they were fixed with Alcohol-Formalin-Acetic Acid (AFA) at room temperature for at least 12 h, and transferred for storage to 70% ethyl alcohol following Pulido-Flores & Monks (2005). Specimens were stained with Gomori's trichrome, Mayer's carmalum, or Delafield's hematoxylin, dehydrated in an ethanol series, cleared in methyl salicylate, and mounted individually on slides in Canada balsam. Specimens were examined using a compound optical microscope equipped with differential interference contrast (DIC or Nomarski) optics, and drawings were made with the aid of a drawing tube. Measurements were made using an ocular micrometer; all measurements are given in micrometers as the mean followed in parentheses by the range and the number of structures measured. The numeration of the sucker complexes is according to Boeger & Kritsky (1989) and measurements of sclerites follow Vaughan & Christison (2012); only shaft length (from points h-i) and shaft width (from e-f ) are reported herein (see VAUGHAN & CHRISTISON, 2012;figures 1C, 1B, respectively). Measurements of some structures of N. pastinacae were estimated from the figures of Ktari & Maillard (1972) because type materials (MNHN-Paris 52166) from the Muséum National d'Histoire Naturelle de Paris could no be located by the Museum. Names for standard shapes of structures are taken from Clopton (2004). Statistical analysis of variance (ANOVA) was used to assess differences in the sizes of suckers and sclerites, respectively, of the new species. Helminths were identified using existing taxonomic keys and published literature (PRICE, 1942;YAMAGUTI, 1963;KTARI & MAILLARD, 1972;EUZET & MAILLARD, 1974;BOEGER & KRITSKY, 1989;BULLARD & DIPPENAAR, 2003). Specimens were deposited in the Colección Nacional de Helmintos, Instituto de Biología, Universidad Nacional Autónoma de México, Mexico city, Mexico (CNHE) and the Harold W. Manter Laboratory of Parasitology, University of Nebraska State Museum, Lincoln, Nebraska (HWML).

Results
Family Hexabothriidae Price, 1942(sensu BOEGER & KRITSKY, 1989 Genus Neonchocotyle Ktari & Maillard, 1972 Type species Neonchocotyle pastinacae Ktari & Maillard, 1972 KRITSKY, 1989). Body elongate narrowly eliptoid to narrowly deltoid; tegument lacking papillae. Haptor asymmetrical, wider than body ( Figures 1A), with 3 paired sucker-sclerite complexes; longitudinal haptoral axis forming an angle of approximately 45º from midline of body; see correct orientation in Figure 1A). Sucker-sclerite complexes similar in form; each sclerite comprising a C-shaped shaft embedded in sucker wall with distal end having pointed tip. Dorsal haptoral appendix armed with 2 suckers; hamuli present or absent; microhooks present or absent. Two anterior glands present or absent; if present one on each side, having ducts continuing to pores at anterior margin of body. Ceca bifurcate, extending posteriorly from anterior end of body; lateral branches having diverticula extending lateral; ceca becoming confluent posterior to testes, extending posterior to form single blind-ending branch extending into haptor. Number of testes 8 to approximately 30, slightly longer than wide, intercecal, oval to spherical, seminal vesicle present. Male copulatory organ enveloped by a pouch, with terminal end sclerotized or not; divided into proximal and distal portions. Proximal region of copulatory organ muscular; distal region narrow, surrounded by pars prostatica. Two vaginae present, parallel, having proximal and distal portions; proximal region connecting to vitelline reservoir, glandular and muscular distal region connecting to vaginal pore. Vaginal pores ventral, associated with encircling muscle, lateral to proximal portion of male copulatory organ, posterior to cecal bifurcation. Ovary tubular, deeply lobed, with ascending and descending branches; ascending branch dextral, sinuous, reaching to region of oviduct, descending branch coiled, connected posteriorly to oötype. Oötype, dorsal to ovary, ventral to vas deferens, leading to uterus. Seminal receptacle present. Transverse vitelline ducts dorsal, forming Y-shaped reservoir; proximal region of vaginae connected to anterior branches of reservoir and posterior region of reservoir join to oviduct (see KTARI & MAILLARD, 1972, Figure 12 Type-locality. Celestún, Yucatán, Mexico (20°51'33"N; 90°24'00"W).

Remarks and Discussion
Neonchocotyle was proposed by Ktari & Maillard (1972) for Neonchocotyle pastinacae that infects gill of common stingray, Dasyatis pastinaca, from the Gulf of Tunis. Bullard & Dippenaar (2003), building upon information summarized by Boeger & Kritsky (1989), furnished a key that distinguished Neonchocotyle from all other hexabothriids by the presence of an asymmetrical haptor, appendix originating from dorsal surface of haptor, sclerites not equally sized, and prostatic region present. These characters, except for the inequality in the size of the sclerites, are present in the specimens that we collected in Mexico, so we have assigned the new species to this genus. However, several features present on the new species had not been described for the genus, although some are present in the description and figures of Ktari & Maillard (1972). Thus, an emendation of the genus was required. Boeger & Kritsky (1989), who were only able to study of paratype from the personal collection of Claude Maillard, remarked that they could not confirm the origin of the haptoral appendix in the specimen of N. pastinacae. Specimens of that species were not available to us for study; however, the haptoral appendix of the new species originates from the dorsal haptoral surface, as seen in Callorhynchocotyle Suriano & Incorvaia, 1982and Epicotyle Euzet & Maillard, 1974(SURIANO & INCORVAIA, 1982EUZET & MAILLARD, 1974;VAUGHAN & CHRISTISON, 2012). Ktari & Maillard (1972) mention that the vas deferens formed a loop before it joined the male copulatory organ. Boeger & Kritsky (1989) also mention this in their diagnosis of the genus, but neither they nor the author of the original description depicted  this feature in their figures. We could not differentiate a looped section of the vas deferens in our specimens, so we did not mention it in our diagnosis and it was not illustrated.
Despite the increase in the number of studies of the phylogenetic relationships of monogeneans using morphological features (BOEGER & KRITSKY, 1989, 1993CHISHOLM et al., 1995;WHITTINGTON, 2004), there still exist confusion between characters (morphological features) and the states (derived forms) of those characters. All of the states of a particular character are homologous. Thus, all of the states should be referred to by the same name regardless of differences in appearance (derived condition). An example is the use of vasa efferentia to refer to ducts that carry sperm from the testis. The structure that carries sperm from testis in every species of monogenean (the vas efferens) is homologous a should be referred to by the same name even though its derived form is a dilated, glandular-wall (BULLARD & DIPPENAAR, 2003) or it is a thin non-glandular wall (GLENNON et al., 2005). Theses are forms (thing walled, glandular, or looped) of the same homologous structure, so they should be referred to by the same name (vas efferens). It is confusing to change the name of the structure depending on its derived condition.
Neonchocotyle violantei n. sp. is the second species of the genus. One remarkable difference between N. violantei and N. pastinacae is body size. The body of N. violantei n. sp. is smaller than that of N. pastinacae (821 long x 315 wide vs. 6580 x 1041, respectively). Furthermore, the new species can be distinguish from N. pastinacae by having two anterior glands on each side of the body, submarginal (extracaecal) vaginal pores ( Figure 6B) (intracaecal in N. pastinacae ( Figure 6A)), and the number of testes. Two pairs of microhooks, similar to those depicted for Squalonchocotyle sp. by Bychowsky (1957) are present in the new species (not described for N. pastinacae). The microhooks of the new species are narrowly fusiform in ventral view, shallowly cresentic in lateral view; this form was described as "buttonhole" by Bychowsky (1957; see their figure 290); all microhooks could be distinguished as sclerotized structures. Hamuli were not found in the new species but are present in N. pastinacae. We could not discern any holes in the area where the hamuli are located in any of our specimens which might indicate that they had been lost and it is unlikely that they would have been lost in all 12 specimens.
Not direct comparisons can be made between the sizes of the haptors of the two species because Ktari & Maillard (1972) did not mention at which point they measured the haptor. However, the haptor of the new species is smaller than the measurements given for N. pastinacae (205 x 754 vs. 874 x 1,466 wide). Other differences in sizes included the pharynx (37 x 38 vs. 110 x 75), the pouch of the male copulatory organ (181 long vs. 536), the ovary (302 vs. 582), and the eggs (113 vs. 135).
The number of testes is different in each species. Ktari & Maillard (1972) did not give the number of testes for N. pastinacae, but in their figure (KTARI & MAILLARD, 1972; their figure 1) the number of the testes appears to be more than 30; the mean number of testes of the new species, N. violantei, is between 8 and 9. The terminal end of the male copulatory organ of the new species is sclerotized; scleritization was no mentioned by Ktari & Maillard (1972) for N. pastinacae, and its presence or absence in that species could not be confirmed. Ktari & Maillard (1972; their figures 1, 12) described and depicted that the proximal portion of the vaginae are connected to the anterior branches of the vitelline reservoir, and that the transverse vitelline ducts connect to the Y-shaped vitelline reservoir. The posterior region of the reservoir joins to the oviduct. The transverse ducts are dorsal and were obscured in our specimens.
Neonchochotyle violantei n. sp. is the third helminth reported from Pseudobatos lentiginosus in the Gulf of Mexico. Rodríguez-Ibarra et al. (2011) reported Paramonilicaecum-type metacercariae (Digenea) from Laguna de Tampamacho on the northern Gulf coast of Veracruz, Mexico, and Vardo-Zalik & Campbell (2011) described Acanthobothrium lentiginosum Vardo-Zalik & Campbell, 2011 from the south Texas coast, Gulf of Mexico. The present report is from the opposite (eastern) side of the Gulf of Mexico, suggesting that much remains to be learned about the helminths of P. lentiginosus.
Key to the genera of Hexabothriidae Price, 1942 All currently publish keys to the Hexabothriidae are incomplete because they do not include all currently known taxa for many reasons. Of the must often uses keys, that of Yamaguti (1963) is outdated because it does not include the taxa described since that time and his systematic arrangement differs from current hypotheses. The more recent key of Bullard & Dippenaar (2003) is consistent with current hypotheses but lack taxa because there was insufficient morphological information or because they were described more recently. The key of Yamaguti (1963) lacks eight more recently described genera and the other two keys lacked two genera, one for lack of information and one because it was described more recently than those publications (BOEGER & KRITSKY, 1989;BULLARD & DIPPENAAR, 2003). Two genera that often are excluded from studies, Pseudohexabothrium Brinkmann, 1952 and Pristonchocotyle, are included in our key. The key presented herein is not base on a phylogenetic hypothesis for the family, but it includes all genera known to date.
Pristonchocotyle was excluded in the systematic treatments of Boeger & Kritsky (1989) and Bullard & Dippenaar (2003), perhaps because the description of its type species P. intermedia Watson & Thorson, 1976 was incomplete. For example, in the figures of that paper (WATSON & THORSON, 1976) it is not clear whether the haptor is symmetrical or asymmetrical and we were not able to study the slides and review this character. Watson & Thorson (1976, p. 633) mention that the, "Anterior hook measures 154 to 224 (189) long by 29 to 46 (39) wide, remaining hooks appear in a row […]", but the drawing of the holotype shows only three sucker-sclerite complexes and they are not labeled, so the normal position of each cannot be determined. The haptoral appendix is shown on the upper margin of the drawing of the haptor (WATSON & THORSON, 1976; their figure 17) although it was not specified in the description. It is not clear in their drawings whether it arises laterally or dorsolateral. Ogawa's (1991) description of Pristonchocotyle papuensis Ogawa, 1991 reported that the haptoral appendix originates dorsally and clearly depicted the arrangement of the haptoral sucker-sclerite complexes. This and other information in the description allowed the genus to be included in the key. The only other genus having vaginae that join medially (X-shaped) is Rhinobatonchocotyle but it differs from Pristonchocotyle by lacking a seminal receptacle.