BEHAVIOR OF THE CATARINA PUPFISH (CYPRINODONTIDAE: MEGUPSILON APORUS), A SEVERELY IMPERILED SPECIES

Abstract

We provide the first description of behavior in the Catarina pupfish (Megupsilon aporus). Aggressive, courtship, and spawning behaviors resemble those of other North American cyprinodontids. However, M. aporus differs from others in the group in absence of breeding territoriality in males. Male M. aporus often perform opercular rotation during aggressive displays and jaw-nudging during courtship, behaviors that, among other North American cyprinodontids, are absent or known only in Floridichthys. Some unusual features of behavior (lack of territoriality) and morphology (dwarfism; absence of pelvic fins) in M. aporus might have been shaped by interaction with a cohabitant, the Potosi pupfish Cyprinodon alvarezi. [PUBLICATION ABSTRACT]

Headnote

ABSTRACT-We provide the first description of behavior in the Catarina pupfish (Megupsilon aporus). Aggressive, courtship, and spawning behaviors resemble those of other North American cyprinodontids. However, M. aporus differs from others in the group in absence of breeding territoriality in males. Male M. aporus often perform opercular rotation during aggressive displays and jaw-nudging during courtship, behaviors that, among other North American cyprinodontids, are absent or known only in Floridichthys. Some unusual features of behavior (lack of territoriality) and morphology (dwarfism; absence of pelvic fins) in M. aporus might have been shaped by interaction with a cohabitant, the Potosi pupfish Cyprinodon alvarezi.

RESUMEN-Proporcionamos la primera descripcíon del comportamiento en el cachorrito enano del Potosí (Megupsilon aporus). El comportamiento de agresi ón, cortejo y desove se asemeja a el de otros cyprinodóntidos norteamericanos. Sin embargo, M. aporus difiere de los demás al carecer la territorialidad en machos reproductores. Machos de M. aporus frecuentemente muestran una rotaci ón opercular durante exhibiciones agresivas y un empuj ón mandibular durante el cortejo, comportamientos que, entre los cyprinodóntidos norteamericanos, están ausentes o son conocidos solamente en Floridichthys. Algunas de las características inusuales del comportamiento (falta de la territorialidad) y morfología (enanismo; ausencia de aletas pélvicas) de M. aporus podrían haber sido conformadas por la interaccíon con un cohabitante, Cyprinodon alvarezi.

The monotypic genus Megupsilon, which appears to be >7,000,000 years old (Echelle, 2008), is extinct in the wild and virtually unknown except for comments on its demise (Contreras and Lozano, 1996) and studies of its systematic relationships (Parenti, 1981; Parker and Kornfield, 1995; Costa, 1997; Echelle et al., 2005). The genus, represented by M. aporus (Catarina pupfish), once occupied a small spring system near El Potosí, Nuevo Léon, Mexico, where it co-occurred with another endemic pupfish, Cyprinodon alvarezi (Potosi pupfish). By 1994, both species were extinct in the wild as a result of overmining of groundwater and drying of habitats (Contreras and Lozano, 1996).

The continued existence of M. aporus is extremely precarious. Until recently, two lineages initiated separately with wild-caught individuals >20 years ago were being maintained by private aquarists and university personnel in Mexico, Germany, and the United States (A. Valdez, pers. comm.). In the summer of 2012, the two lineages consisted of only one population each, one of which died out in December 2012 (D. Schmidt and M. Schneider, pers. comm.). The surviving population comprises about 20 individuals held at the Children's Aquarium at Fair Park, Dallas, Texas (S. Walker and C. Yancey, pers. comm.).

Megupsilon aporus and its sister group, Cyprinodon, belong to a monophyletic group containing four other genera, Cualac, Jordanella, Floridichthys, and Garmanella (Parenti, 1981; Costa, 1997; Echelle et al., 2005). All except the wide-ranging New World genus Cyprinodon (ca. 50 species) contain only one or two species confined to North America. Megupsilon aporus is one of the smallest pupfishes. Maximum reported standard lengths in males and females are 26 and 36 mm, respectively (Miller and Walters, 1972), making it comparable in size to the Devils Hole pupfish C. diabolis, a dwarfed species, which, in its natural habitat (Devils Hole), reaches a maximum standard length of 35 mm (Miller, 1948). Both species also are unusual in lacking a pelvic girdle and pelvic fins (Miller 1948; Miller and Walters, 1972). Dwarfism in C. diabolis disappears in populations maintained in artificial pools outside of Devils Hole (Wilcox and Martin, 2006), but absence of pelvic fins persists (Martin et al., 2012). Both traits are retained in captive M. aporus. Megupsilon aporus differs from Cyprinodon in having a sexually dimorphic karyotype. Females resemble Cyprinodon in having 2N = 48 acrocentric chromosomes, whereas males have 2N = 47 with a large, metacentric Y derived from fusion of two acrocentrics (Uyeno and Miller, 1971).

There is no published description of behavior of M. aporus. In contrast, detailed descriptions of behavioral repertoires in Cyprinodon are available from the survey of 13 species (and 19 hybrid combinations) by Liu (1969) and several studies focusing on individual species (Barlow, 1961; Cox, 1966; Kaill, 1967; Arnold, 1972; Echelle, 1973). Similar descriptions also are available for one (F. carpio) of the two species of Floridichthys (Foster, 1967; Kaill, 1967), and the one species of Jordanella (Mertz and Barlow, 1966; Kaill, 1967). Published behavioral information for the remaining two monotypic genera is limited to brief descriptions of spawning in Garmanella (Richter, 1975; Loiselle, 1981) and a comment on breeding territoriality in males of Cualac (Seegers, 2000). Choice of mates and breeding-system dynamics in Cyprinodon have been studied extensively (e.g., Kodric-Brown, 1981; Kodric- Brown and Strecker, 2001; Leiser and Itzkowitz, 2004), and choice of mates and parental care in Jordanella have been examined in considerable detail (e.g., Hale and St. Mary, 2007).

In this report, we describe breeding and aggressive behaviors of M. aporus, with an emphasis on qualitative comparisons with the sister group (Cyprinodon). Liu (1969) noted in his discussion of Cyprinodon, that nuptial males of an undescribed El Potosi pupfish were nonterritorial and that they exhibited jaw-nudging during courtship and opercular rotation in agonistic interactions. Miller and Walters (1972) described this species as M. aporus and cited personal communication with RKL for the occurrence of jaw-nudging and opercular rotation, but with no description. Haas (1979) reported that male M. aporus x C. alvarezi hybrids exhibited jaw-nudging (=jaw protrusion). Here we expand on observations by Liu (1969) and present photo-documentation of various components of agonistic and breeding behaviors.

MATERIALS AND METHODS-This report is based primarily on data gathered by RKL at the University of California, Los Angeles. This included observations on staged and unstaged male-female and male-male encounters. Unstaged encounters occurred in an 80-L aquarium (76 by 31 cm; 37 cm deep) used to maintain the stock of 18 fish that were studied. Staged encounters occurred in 56-L aquaria (62 by 32 cm; 31 cm deep). All aquaria had a 16-h photoperiod provided by one or two 25 or 40-watt incandescent light bulbs mounted in the hood of the aquarium. For staged courtship and mating encounters, a male and a female were removed from the stock aquarium and placed in separate compartments of an aquarium divided into halves by a barrier (plastic window screening) that allowed interaction with no physical contact. After [double dagger]24 h, the female was placed into the compartment of the male and RKL immediately began recording behavior. Six such encounters were recorded. Eight similarly staged encounters were performed for recordings of male-male agonistic interactions. Records consisted of written notes supplemented with 35-mm photography (Tri-X or Ektachrome film; Pentax SLR camera, Pentax-Ricoh Imaging Company, Tokyo, Japan) and 16-mm ciné photography (highspeed Royal-X Pan film, ASA 1250; Bealieu 16-mm reflex camera, Beaulieu Company, Romorantin-Lanthenay, France). Additional observations were made during brief visits to the Dallas Aquarium in 2008 and 2011 when RKL and AAE, respectively, made observations and photographed agonistic behavior in the refuge population (total ca. 200 fish in six aquaria in 2008; 20 fish in three aquaria in 2011). For field observations, four ichthyologists (R. Brown, R. Haas, V. Walters, and RKL) spent >1 h observing and photographing fish at El Potosí in July 1972.

RESULTS-The distribution of some morphological and behavioral traits in M. aporus and Cyprinodon are presented in Table 1. The nape and sides of the body in nuptial male M. aporus have a blue iridescence that is shared with Cyprinodon and no other North American cyprinodontid; it is less intense in Megusilon than in most species of Cyprinodon. Nuptial male M. aporus have a black, vertical eye-bar that occurs in some, but not all, species of Cyprinodon (Liu, 1969; Echelle, 1973). Males differ from male Cyprinodon by lacking a terminal black band on the caudal fin and by having a large, black, lateral blotch just anterior to the caudal peduncle and extending onto the anal fin (Fig. 1a; Miller and Walters, 1972). Females are golden olivaceous with a weak, usually broken, midlateral band and no indication of the dorsal-fin ocellus seen in females of most Cyprinodon. During spawning, females become pale and lose the midlateral band. Juveniles and non-breeding males resemble females in color pattern, including the well-developed lateral band. In male-male interactions, color pattern of submissive males resembles the pattern of females, with a faint midlateral band (Fig. 1b).

In communal aquaria, sexes formed loose, mixed aggregations or they were distributed somewhat evenly throughout the aquarium. Males were more aggressive toward males than toward females. Males in communal aquaria and in staged encounters displayed mutual opercular rotation, biting, charging, or a combination of these, usually accompanied by lateral or frontal display, or agonistic looping behavior, or some combination of these agonistic behaviors. Some of these behaviors were directed toward male C. alvarezi when held in the same aquarium (Fig. 1a). In opercular rotation, opercles are rotated outward and the branchial basket expands downward with separation of the branchiostegal rays; the body is tilted slightly head-downward with median fins rigidly fanned out (Figs. 1a-c). No vibration of fins or body was detected in lateral display, although this is common in Cyprinodon. Tail-beating (a male slaps his caudal fin at the opponent) occasionally occurred, and there was one instance of mouthfighting (males tumble about with jaws locked together).

Male M. aporus showed no evidence of breeding territoriality in communal aquaria at the University of California, Los Angeles, or at Children's Aquarium at Fair Park. In two instances, a stock-aquarium male briefly defended an area with low-intensity aggression. More typically, the male was aggressive toward other fish in a small space surrounding his body. The breeding system consisted only of consort-pairing, in which a male followed a female around while courting and making spawning attempts. Pairs sometimes formed when females approached a male while he was courting or spawning with another female.

Courtship occurred in all six of the staged male-female encounters, and spawning occurred in five. In early courtship, the male approached a female and followed, or he performed courtship looping (Fig. 2) or, less often, agonistic looping. In courtship looping, the male, with lowered dorsal and anal fins, swims rapidly around the female one or more times in a tight, horizontal circle with a radius (possibly constrained by size of the aquarium) two or three times the length of the female. Agonistic looping, which is more often directed at males, is similar except median fins are held erect and opercles are rotated as in aggressive displays. Looping often is followed by contact behavior in which the male performs jawnudging (Figs. 2b and 2c) or attempts to contact the female laterally in a spawning attempt. In jaw-nudging, the male repeatedly protracts and retracts his premaxillaries, sometimes, at least, causing his snout to push against the head of the female; it has not been determined whether actual contact is common or not. Often, the male engages in jaw-nudging while moving back and forth along the nape of the female; these back and forth movements resemble various forms of contact behavior termed nuzzling in Cyprinodon (Liu, 1969).

The courted female initiates spawning by tilting headdownward and nipping the substrate. Nipping is a ritualized, somewhat exaggerated feeding movement in which she delivers a single bite at the spawning substrate. She then settles to the bottom (or a vertical surface) as the male sidles alongside, forming his body into an Sshape and pressing the female onto the substrate (Fig. 2d). During sidling, the male wraps his anal fin around the venter of the female in the region of her anal fin. After wrapping, the pair vibrates briefly and the act culminates with a quick jerk and straightening of the body of the female. Vibration and jerking presumably coincide with release of sperm and eggs. Number of eggs released with each spawn has not been determined, but it is likely to be a single egg, as in the closely related genera Cyprinodon (Mire and Millet, 1994) and Jordanella (Mertz and Barlow, 1966). The pair then separates and additional rounds of courtship and spawning might ensue, or they might remain in lateral contact and go directly to spawning. Spawning has been observed on sand, on the bare bottom of an aquarium, in spawning mops of yarn (floating or on the bottom), and on vertical surfaces (e.g., the previously mentioned plastic screen). Spawning on vertical surfaces is most likely to occur when the surface has a coat of algae (J. Cokendolpher, pers. comm.).

Behavioral observation of M. aporus in its natural habitat was difficult because of its wariness and small size, presence of dense submergent and emergent vegetation in the preferred, shallow-water habitat (Miller and Walters, 1972), and many village children in the water. Male M. aporus were in reproductive coloration. No courtship or spawning was noted, but male-male interactions, including agonistic looping, were detected. Male C. alvarezi were defending territories and spawning in more open waters of the pool. The only interaction between the two species was chasing of male M. aporus by the much larger males of C. alvarezi.

DISCUSSION-The literature has been contradictory regarding presence-absence of breeding territoriality in males of M. aporus. Miller and Walters (1972:7) commented that the smallness of males relative to females ''may be correlated with the absence of territorial behavior.'' However, in a comment on C. alvarezi, Miller et al. (2005:296) mentioned that, ''competition for territories with . . . Megupsilon aporus may have been intense,'' and that, ''it was seen threatening M. aporus males defending territories.'' We have seen no evidence of a territorial breeding system in male M. aporus, but they are aggressive toward other fish, conspecific or not.

Absence of breeding territoriality in males appears unique to M. aporus among North American cyprinodontids. Such territoriality has been described for Cyprinodon, Jordanella, and Floridichthys, and its presence in Cualac is mentioned by Seegers (2000) and seen both in the field (M. Salazar, pers. comm.) and in captive populations (A. Valdez, pers. comm.). Territoriality in male Garmanella is mentioned briefly in the hobbyist literature (Loiselle, 1981). Cyprinodon diabolis appears unique for its genus in not showing territoriality in its native habitat (Kodric- Brown, 1981). However, territoriality in males occurs in artificial refuges established outside of Devils Hole (Wilcox and Martin, 2006). Similar variation in presence- absence of territoriality occurs in the cyprinodontid genus Aphanius, the Old World ecological equivalent of Cyprinodon. Most species show territoriality, but it is absent in A. fasciatus (Grech and Schembri, 1993) and in some, but not all, populations of A. anatoliae (Grimm, 1981); breeding in absence of territoriality is based on consort pairing, as in M. aporus.

With one exception (opercular rotation), aggressive behaviors observed in M. aporus have been described for other North American cyprinodontids that have been studied in detail (Cyprinodon, Floridichthys, and Jordanella). This includes tail-beating, an element of especially intense aggression described for a variety of Cyprinodon (Liu, 1969; Echelle, 1973), Floridichthys (Kaill, 1967), and Jordanella (Mertz and Barlow, 1966; Kaill, 1967), as well as more distantly related cyprinodontoids such as Aphanius (Foster, 1967) and Fundulus (Able and Hata, 1984). Mouthfighting, an especially conspicuous and intense expression of aggressive behavior, is described for several species of Cyprinodon (Newman, 1907; Liu, 1969; Echelle, 1973). It is not reported for other cyprinodontids, although it does occur in the cyprinodontoid families Aplocheilidae (Foster, 1967) and Fundulidae (Newman, 1907; Able and Hata, 1984).

The behavior most similar, in North American cyprinodontids, to opercular rotation in M. aporus is opercular flickering described by Kaill (1967) for male Floridichthys. However, the latter is a repetitious on-offrotation of the operculum that causes a repeated flash of color and it might act more for distant sexual advertisement rather than close-range aggression. Behaviors more similar to opercular rotation in M. aporus occur in certain other cyprinodontoids (Foster, 1967; Able and Hata, 1984) and a variety of distantly related fishes. These behaviors, variously termed lowering of branchiostegals, raised opercula, gill-cover erection, and branchiostegal flaring, usually are associated with aggression and have arisen independently in multiple groups.

The combination in M. aporus of visual display (looping) and contact behaviors during courtship by males also occurs in certain species of Cyprinodon from the Mexican Plateau (Liu, 1969; Turner and Liu, 1977). Contacting in the latter group (apart from sidling and wrapping) normally involves nuzzling the female from below, but some nuzzling occurs from above. Contact by male M. aporus resembles nuzzling in Cyprinodon, but with the addition of jaw-nudging. It usually occurs from above the female along her nape and head region, but it also is performed from alongside the female. Nuzzling with little or no looping is characteristic of courtship by males in western species of Cyprinodon in California, Nevada, and Sonora, whereas looping with no specialized contacting prior to sidling and wrapping characterizes species of the genus from the eastern coast of North America to Texas and New Mexico (Liu, 1969).

Jaw-nudging in M. aporus seems similar in both appearance and function to that described for Flori- dichthys (Kaill, 1967:45; his ''protracting''). It has not been described for any other North American cyprinodontid, but it is reported in a variety of more distantly related cyprinodontoids. In Aphanius, jaw-nudging occurs in A. mento and A. fasciatus. It is reported to be absent in A. cf. cypris (Foster, 1967; P. Loiselle and R. Haas, pers. comm.); this needs re-evaluation because A. cypris currently is treated as a junior synonym of A. mento (Wildekamp, 1995). Jaw-nudging also occurs in some fundulid and valenciid species of killifish (Foster, 1967; Able and Hata, 1984).

The spawning act in M. aporus is similar in general aspect to that of other North American cyprinodontids for which it has been described. With one exception, it involves the nipping [arrow right] sidling [arrow right] S-shaping [arrow right] wrapping [arrow right] vibration [arrow right] jerking sequence. The exception is that nipping in the context of spawning is absent in Jordanella (Mertz and Barlow, 1966; Kaill 1967). Otherwise, the act differs from that of M. aporus in that male Jordanella (Mertz and Barlow, 1966), Floridichthys (Kaill, 1967), and Garmanella (Richter, 1975) wrap both the dorsal fin and anal fin around the female, not just the anal fin. Megupsilon aporus, at least in aquaria, often spawn on vertical surfaces, which, in nature, might have been growths of vegetation. In the same aquarium conditions, Cyprinodon spawned primarily on bottom substrate (Liu, 1969). In the wild, many spawning bouts in Megupsilon have been observed on plants as well as on bottom substrate (D. Springer, pers. comm.).

Interaction with C. alvarezi, the only other native fish in the El Potosí basin, might explain some of the unusual aspects of behavior and morphology in M. aporus. These include lack of breeding territoriality in males, small size, and (perhaps as a developmental correlate) absence of pelvic fins. Despite some partitioning of habitat and dietary differences, the two species did come into regular contact in the relatively restricted confines of their habitat and aggressive interactions between them were observed (Miller and Walters, 1972). Megupsilon aporus ate primarily animal items and was restricted mostly to shallow habitat densely choked with vegetation, whereas C. alvarezi was more omnivorous, and more common in deeper, more open waters. The two species are incompatible in artificial aquaria because M. aporus is eliminated quickly by the larger, more aggressive C. alvarezi (J. Cokendolpher and C. Yancey, pers. comm.). Therefore, the natural system at El Potosí certainly could have fostered considerable coevolution. Conversely, M. aporus likely is a relic of a once more diverse group (Miller and Walters, 1972) and it is possible that its unusual traits were simply characteristic of the group.

We thank A. F. Echelle for editorial comments, B. J. Turner for encouragement and help with the literature, J. Brill and P. V. Loiselle for help with literature and comments on behavior, B. Potvin and S. Walker for access to captive populations at the Dallas Zoo, and J. C. Cokendolpher, B. Cunningham, R. Haas, A. Morales, M. Salazar, D. Springer, A. Valdez, and C. Yancey for comments on behavior, status of captive stocks, or both.

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Submitted 20 October 2011. Accepted 28 November 2012.

Associate Editor was Robert J. Edwards.