A new genus and species of cyclopoid (Crustacea, Copepoda, Cyclopinidae) from a coastal system in the Gulf of Mexico

Abstract A new, monotypic genus of the interstitial marine cyclopoid copepod family Cyclopinidae G.O. Sars, 1913 is described from male and female specimens collected at Laguna de Términos, a large coastal lagoon system in the southern Gulf of Mexico. Mexiclopina campechana gen. et sp. n. cannot be adequately placed in any extant genus within the family. It differs from other cyclopinid genera in having a unique combination of characters including: 1) absence of modified brush-like seta on the mandibular exopod; 2) maxillule exopod with stout setal elements and brush-like setae absent; 3) basis of mandible with one seta; 4) presence of a modified seta on endopod of fourth leg; 5) fifth leg exopod unsegmented, armed with three elements in the female and five in the male; 6) intercoxal sclerite of first swimming leg with two medial spiniform processes on distal margin. The new genus is monotypic and appears to be most closely related to Cyclopina Claus, 1863 and Heptnerina Ivanenko & Defaye, 2004; the new species was compared with species of Cyclopina and it resembles Cyclopina americana Herbst, 1982 and Cyclopina caissara Lotufo, 1994. This is the second record of a species of Cyclopinidae in Mexico and the first in the Gulf of Mexico; the number of cyclopinid species recorded from the Americas is now 13.


Introduction
The cyclopoid copepod family Cyclopinidae G.O. Sars, 1913 is one of the most diverse and successful among the benthic marine poecilostomatoid/cyclopoid copepods. It contains 12 valid genera (Boxshall and Halsey 2004;. Members of this family occupy a wide range of habitats, having been reported from shallow coastal environments (Reid 1990;Lotufo 1994;Karanovic 2008), anchialine caves (Jaume and Boxshall 1996a), and deep-sea hydrothermal vents (Ivanenko and Defaye 2004). Its knowledge in the Americas is still developing, but it is clear that its diversity has been more studied in South America than in the other subcontinents (Nicholls 1939;Lotufo and Rocha 1991;Lotufo 1994;Rocha and Botelho1998). Only one species of the diverse and widespread genus Cyclopina, C. caissara Lotufo, 1994 has been recorded in Mexico and Central America (Reid 1990;Gómez and Martínez-Arbizu 2004) and nine in South America, mainly in Brazil (Lotufo and Rocha 1991;Lotufo 1994;Rocha and Botelho 1998). Overall, the knowledge of the cyclopinid copepod diversity in this kind of habitats is still lagging and certainly deserves further taxonomic research, particularly in the Northwestern Atlantic region.
Laguna de Términos, in the Mexican state of Campeche, in the southern Gulf of Mexico (between 18°26' and 18°44'N; 91°13' and 91°54'W) is one of the largest lagoon estuarine ecosystems of the gulf; it has a significant ecological and economic importance in southeastern Mexico because of its permanent connection to the sea and high productivity and diverse fish fauna (Yáñez-Arancibia and Day 1982;Ramos-Miranda et al. 2006). Copepods have been investigated but only those of the plankton community (Salas-Marmolejo 1981). As part of a study to know the helminth fauna of this coastal system and the role played by invertebrates and vertebrates as intermediate, "transport", "carrier", paratenic or definitive hosts, night samples were obtained at shallow areas of the lagoon where a mixture of plankton and epibenthic or interstitial copepods was likely to be collected. Our samples contained a new genus and species of the family Cyclopinidae which is herein fully described and illustrated based on male and female specimens.

Methods
Night zooplankton samples were obtained on February 13, 2015 with three hand nets (two of 100 and one of 200 µm) in shallow areas (depth: 60-120 cm) of the lagoonal system, particularly at Isla Tortuga (18°44'29.3"N; 91°29'44.6"W). Water temperature was 25 °C, salinity 28psu, and pH slightly alkaline (7.5). Trawls followed a parallel course with respect to the coastline. Samples were placed in a bucket with 5 liters of water; copepods were isolated alive 5 hours after collection, they were later on fixed in 4% formaldehyde buffered with borax (30 g/l of formaldehyde at 40%) and kept in a 5% glycerin/ 70% ethanol solution. More than 35 male and female specimens were taxonomically examined in the laboratory; specimens were processed, dissected and examined following Reid (2003). Dissected specimens/appendages were mounted in semi-permanent slides with glycerine sealed with Entellan®, a commercial, fast drying mounting medium and sealant. Drawings were prepared at 1000× magnification with the aid of a camera lucida mounted on a standard Olympus CX31 compound microscope. Some specimens were prepared for SEM examination with a TOPCON SM-510 microscope at facilities of ECOSUR in Tapachula, Mexico. The process included dehydration of specimens in progressively higher ethanol solutions (60, 70, 80, 96, 100%), critical point drying, and gold-palladium coating (20 nm) following usual methods. This hitherto unknown genus and species was described and illustrated following the current standards for the taxonomic study of the group (Gómez and Martínez Arbizu 2004;Karanovic 2008). The type specimens were deposited in the collection of zooplankton held at El Colegio de la Frontera Sur (ECO-CH-Z), in Chetumal, Mexico and in the National Museum of Natural History, Smithsonian Institution, Washington, D.C. (USNM). Original zooplankton samples containing more non-type specimens remain in the helminth collection of the Universidad Autónoma Metropolitana-Xochimilco, Mexico (CHUX), maintained by the co-author (RJAA). Etymology. The genus name is composed by the prefix 'Mexi' in reference to Mexico, the country from which it was collected and the suffix 'clopina' to show its affinity with the genus Cyclopina.
Anal somite with ventral and dorsal surfaces smooth, posterior margin ornamented with row of minute spinules along ventral margin at point of insertion of caudal rami. Anal operculum smooth. Caudal ramus (Fig. 1D) length/width ratio range: 1.17-1.20. Dorsal and ventral surface of caudal rami smooth except for row of spinules along posterior margin at insertion of caudal setae (Fig. 1D). Inner margin of caudal rami smooth. Rami with six setae; seta I absent; seta II inserted midway of outer margin; seta III shorter than seta VI, both lightly plumose; seta IV about 3.2 times as long as seta III, with heteronomous ornamentation, with spinules on proximal outer margin and with plumose distal half; proximal inner margin with few spinules, distal third plumose; seta V longest, about 1.5 times as long as seta IV, naked proximally, with few rigid spinules proximally and lightly plumose distally along both margins; dorsal seta VII as long as seta II, about twice as long as ramus (Fig. 1D). Rostrum wide, tapering distally into pointed tip.
Antenna (Fig.1C): 4-segmented, fused coxa and basis cylindrical, with long lightly setulose basal seta on outer margin and slender, short inner exopodal seta (exp in Fig. 1C). Endopod 3-segmented. First endopodal segment cylindrical, about twice as long as succeeding second segment, with long medial seta reaching distal margin of second endopodal segment; segment ornamented with patch of spinules around insertion of seta. Second endopodal segment with patch of spinules on proximal position. Setation formula of endopodal segments 1-3 as: 1, 5, 6.
Maxilliped (Figs 2D; 5B): slender, 6-segmented, precoxa and coxa fused forming syncoxal segment with three endites; proximal endite with single seta, second with 3 unequal setae, third endite ornamented with cluster of cuticular scales, armed with two long, subequal stout setae. Basis expanded distally, medial margin ornamented with row of long, stiff setules, and with two subdistal setae. Endopod 4-segmented; first and second segments naked, third segment with one lightly plumose seta, fourth segment with four elements including two short plumose and two long, stout simple setae.
Leg 5 (Figs 4D; 6D, E) with coxobasis subrectangular, armed with outer seta. Exopod unsegmented, ornamented with few spinules on inner margin and group of minute spinules on outer margin (Figs 4D, 6E). Exopod armed with five elements, two long, inner setae, one small medial spine, one medial seta and outer flanged spine with serrate hyaline frill; as in female, latter element (arrowed in Fig. 6E) blunt, about 2.5 times as long as inner spine.
Leg 6 represented by flat, rounded plate bearing two slender setae and an inner spiniform process (Figs 4D; 6D).
Etymology. The species is named after the state of Campeche in southeast Mexico. Gender is feminine.
Habitat. The lagoon has a length of 70 km and 30 km at its widest sector. It has extense coverage of seagrass beds (mainly Thalassia testudinum), mangrove areas and zones with no vegetation. It is a shallow system, (average depth = 2.5 m). The lagoon receives freshwater input from several rivers. Most of its bottom is covered by sediments of sand, silt and clay with a high content of calcium carbonate mainly in the vicinity of Boca de Puerto Real (between 50 and 70%).
Remarks. Based on the first examination of these specimens, they were tentatively identified as a species of Cyclopina Claus, 1863 by the combined display of the following features: 10-segmented female antennule with sixth antennulary segment being longest, antenna with single exopodal seta; female fifth leg exopod with three armature elements, the apical seta flanked by two spines; leg 1 with 3-segmented endopod; and   caudal seta I absent (cf. Vervoort 1964;Jaume and Boxshall 1997;Boxshall and Halsey 2004). However, when Jaume and Boxshall's (1996b) key to the cyclopinid genera was run, our specimens could not be adequately placed in a genus and it did not fit in the generic diagnoses of other related cyclopinids Boxshall 1996a, b, 1997;Martínez Arbizu 1997a, b;Humes 1999;Ivanenko and Defaye 2004). Also, based on our morphological comparison with Cyclopina esilis Brian, 1938, the best described species of Cyclopina (Jaume and Boxshall 1996a), it was clear that despite their affinities, the new genus and Cyclopina diverge in several important characters. In addition, the monotypic genus Heptnerina (Ivanenko and Defaye 2004) shares some characters with the new genus (i.e., swimming legs segmentation, number of female antennulary segments, armature of male and female fifth legs, segmentation of mandible palp) but differ in some others, as explained below. Overall, the genus Mexiclopina gen. n. differs from the other cyclopinid genera in having a unique combination of characters including: 1) absence of modified brush-like seta on the 4th mandibular exopodal segment; 2) maxillule exopod with stout setal elements and no brush-like setae; 3) presence of modified seta on the fourth leg endopod; 4) fifth leg exopod armed with three elements in the female and five in the male; 5) outer exopodal spine of leg 5 blunt in both sexes; 6) male sixth leg with two outer slender setae and inner spiniform process; 7) intercoxal sclerite of first swimming leg with two medial spiniform processes on distal margin. The new genus diverges from Heptnerina in the lack of an endopodal lobe in leg 5, in the presence of a single antennary exopodal seta vs. two setae present in H. confusa (Ivanenko and Defaye 2004, fig. 3A), and the lack of a modified seta on the maxillule exopodal lobe and also in the mandible exopod (Ivanenko and Defaye 2004, figs. 3A, C). The new genus differs from Cyclopina in the lack of a brush-like seta on the mandible exopod (Jaume and Boxshall 1996a;Lotufo 1994); this character is distinctive of the genus and it is present in the type species, C. gracilis Claus, 1863. Remarkably, in the new genus the intercoxal sclerite of leg 1 has a distinctive feature not previously observed in Cyclopina; it has two medial spiniform processes on the posterior margin (Fig. 3F), similar acute processes in leg 1 are present in Troglocyclopina balearica Boxshall, 1996 (Jaume andBoxshall 1996b), but are absent in Heptnerina (Ivanenko and Defaye 2004). The new genus clearly diverges from Troglocyclopina Jaume & Boxshall, 1996 in having six setae instead of five on the distal segment of endopod of leg 1 (Jaume and Boxshall 1996b, figs. 4A) but also in the presence of two exopodal setae on the antenna (Jaume and Boxshall 1996b, fig. 3A) vs. a single exopodal seta in Mexiclopina.
Other remarkable features of the new genus include: 1) the short, stout distal setae of the exopodal segment of the maxillule (asterisks in Fig. 2B); these setae are long, flexible in Cyclopina (Lotufo 1994;Jaume and Boxshall 1996a;Karanovic 2008) and Heptnerina (Ivanenko and Defaye 2004); 2) the female P6, represented by short plate armed with two slender setae; it is similar to that known in species of Cyclopina but differs from Heptnerina (Ivanenko and Defaye 2004, fig. 1E), with three unequal setae; and 3) the modified, short spiniform outer seta of the third endopodal segment of leg 4 (asterisk in Fig. 3E), not described in any other cyclopinid.
Because of the close morphological resemblance of the new species with Cyclopina, we performed a comparison with the most closely related species of this genus. Only a few species of Cyclopina have a female leg5 with the inner spine of the exopodal segment less than half the length of the outer spine, the latter being longer than the segment itself (Jaume and Boxshall 1996a). This group of species include C. kieferi Schäfer, 1936, from Europe, C. esilis Brian, 1938 from Mediterranean anchialine caves, C. americana Herbst, 1982, from North Carolina, USA, C. caissara Lotufo, 1994from Brazil (Lotufo 1994 and from the Mexican Pacific (Gómez and Martínez Arbizu 2004), and C. amita from Australia (Karanovic 2008). The new species shares this feature with this group of species but it can be easily distinguished from C. caissara by the segmentation of the antennules, the new species having 10 segments, like most other known species of Cyclopina, whereas C. caissara has a 12-segmented antennule both in specimens from Brazil (Lotufo 1994, fig. 37) and from Mexico (Gómez and Martínez Arbizu 2004, fig. 3A). Also, the length/width ratio of the caudal rami differs between these two species, being slightly longer in C. caissara (ratio=1.3-1.5; Lotufo 1994; Gómez and Martínez Arbizu 2004) vs. 1.17-1.2 in the new species. The shape and size of the outermost terminal flanged spine of the male fifth leg differ in these species, being broad and blunt in the new species vs. slender and pointed in C. caissara (Lotufo 1994, fig. 52). Also, the female fifth leg differs in the size and proportions of these spines; the outer spine is more than 4 times as long as the inner one in C. caissara (Lotufo 1994, fig. 49), whereas in the new species this element is only about twice longer than the inner spine. In C. caissara the armature of the female sixth leg consists only of two elements, the inner one corresponding to a thick stout serrate seta (Lotufo 1994, fig. 50;Gómez and Martínez Arbizu 2004, fig. 1C), thus differing from the slender seta present in homologous position in the new species (Fig. 5A).
The new species differs from C. esilis in the display of a long terminal seta on the exopod of mandibular palp; it is the longest and is slightly broader than the rest of exopodal setae; contrastingly, this seta is remarkably short and modified, umbrella-like, in C. esilis (Jaume and Boxshall 1996a, fig . 2B). In addition, both species can be readily distinguished by the proportions of the caudal rami, being 2.6-3.3 times longer than wide, relatively elongate in C. esilis (Jaume and Boxshall 1996a, fig. 1F,G), vs. short and subquadrate (length/width ratio 1.2) in the new species.
Mexiclopina campechana sp. n. differs from C. americana in body shape, with the third and fourth pedigerous somites strongly produced posteriorly, the process of the fourth somite reaching well beyond the posterior margin of the fifth pedigerous somite (Fig. 1A); in C. americana the posterolateral corners of the fourth pedigerous somite do not reach the posterior margin of the succeeding somite (Herbst 1982, fig. 1). Also, in C. americana the female anal somite is 1.16 times as long as the caudal ramus (Herbst 1982, fig.1), whereas in the new species the anal somite is shorter (0.8 times) than the caudal ramus. The length/width ratio of the caudal rami is also slightly different in both species, 1.2 in M. campechana sp. n., vs. 1.3 in C. americana (Herbst 1982, fig. 2). They differ also in the relative length of the antennulary segments, particularly in the shorter segment 6 in C. americana, which is 26% of the antennule length (Herbst 1982, fig. 3), vs. 21% in the new species from Campeche. In C. americana the antenna lacks the exopodal seta (Herbst 1982, fig. 4), which is present in the new species (Fig.  1C), but in some species like C. amita this seta is also absent (Karanovic 2008). In ventral view the male anal somite of C. americana is long, 1.45 times as long as the caudal rami (Herbst 1982, fig. 10), whereas in the new species it is relatively shorter, 0.7 times as long as the caudal ramus (Figs4A; 5C). In addition, both sexes have a crenulate hyaline frill on the posterior margin of urosomites(Figs. 3A, 5C), whereas these margins are smooth in both sexes in C. americana (Herbst 1982, figs 1;10;11). In C. americana the male fifth leg has four elements on the exopodal segment (Herbst 1982, fig. 13), vs. five in the new species. In addition, the sixth leg of the new species has, like the majority of the species of Cyclopina for which males are known (Karanovic 2008), an inner spine aside the two usual setae; this spine is absent in both C. americana (Herbst 1982, figs 10;11) and C. amita (Karanovic 2008 , fig. 36C). The new species differs from C. amita in the antennule segmentation; this appendage having 11 segments in the Australian species (Karanovic 2008, fig. 34A) vs. 10 segments in M. campechana.
The new species of Mexiclopina shows also some resemblance with C. kieferi, but in this species the external spine of the female fifth leg is 1.2-1.5 times as long as the internal spine (vs. 2.5 in the new species), the caudal rami are clearly longer than the anal somite and have a length/width ratio of 2.6 (Vervoort 1964;Lotufo 1994),thus differing from M. campechana, with an anal somite as long as the caudal rami, which in turn have a 1.2 length/width ratio.
Males are known for only about half the known nominal species of Cyclopina (Karanovic 2008) and the available keys are based on females (Vervoort 1964), thus, characters of this gender have not been fully explored but some of them appear to be potentially important to define species. For instance, the male of C. esilis shares several features with the new species, but the antennulary armature differs. The male antennule of C. esilis has pectinate spines on each of segments 10-13 (Jaume and Boxshall 1996, fig . 4D), whereas these spines are distributed on segments 9-12 in the new species (Fig.  6B). In addition, the male antennule of C. americana has 13 segments (Herbst 1982, fig. 12) vs. 15 in the new species; the last antennular segment is distinctly acute in C. americana (Herbst 1982, fig. 12) and blunt in the new species. Details of the male antennulary armature were not shown in the description of C. americana (Herbst 1982), but this appendage is likely to provide additional differences at the species level.
The male fifth leg of the new species has 5 elements on the exopodal segment, thus diverging from most species of Cyclopina for which males have been described thus far. This feature is shared only with C. esilis, C. caissara, C. kieferi, C. amita, and C. confusa, but the latter has an ornamented anterior surface of the female fifth leg, thus diverging from the smooth condition of the same surface in M. campechana.
The copepod fauna of the Laguna de Términos has been known mainly from plankton surveys (Suárez-Caabro and Gómez-Aguirre 1965;Salas-Marmolejo 1981); relatively little is known from other copepod habitats. The local copepod diversity of interstitial environments may equal or exceed that of their planktonic relatives. The sampling of shallow coastal systems frequently results in the capture of epibenthic or interstitial fauna that is integrated into the water column. This appears to be the case in the new species, belonging to a genus of interstitial forms (Karanovic 2008).