Polymorphic microsatellites isolated from the cave fish Astyanax fasciatus

Six polymorphic microsatellite loci from a library of the tetra Astyanax fasciatus from Mexico were isolated. Amplification and heterozygosity were tested in four cave and four surface populations. These loci were developed for population genetic study to detect gene flow between cave and surface populations.     

Studies on the effects of Ca2++ and Co++ on the swimming behavior of the blind Mexican cave fish

Summary The hypothesis that the blind cave fish (Astyanax hubbsi) adjusts the level of stimulation to its lateral line system (LLS) by varying its own velocity was examined. When the sensitivity of the LLS sense organs was reduced by lowering the Ca²⁺ concentration in the water or by adding Co²⁺ the fish compensated for this by swimming at a higher velocity.Abbreviation LLS lateral line system     

Predation of a cave fish (Poecilia mexicana, Poeciliidae) by a giant water-bug (Belostoma, Belostomatidae) in a Mexican sulphur cave

Abstract.  1. Caves are often assumed to be predator-free environments for cave fishes. This has been proposed to be a potential benefit of colonising these otherwise harsh environments. In order to test this hypothesis, the predator–prey interaction of a belostomatid (predator) and a cave fish (prey) occurring in the Cueva del Azufre (Tabasco, Mexico) was investigated with two separate experiments.
2. In one experiment, individual Belostoma were given a chance to prey on a cave fish, the cave form of the Atlantic molly ( Poecilia mexicana ), to estimate feeding rates and size-specific prey preferences of the predator. In the other experiment, population density of Belostoma was estimated using a mark–recapture analysis in one of the cave chambers.
3. Belostomatids were found to heavily prey on cave mollies and to exhibit a prey preference for large fish. The mark–recapture analysis revealed a high population density of the heteropterans in the cave.
4. The absence of predators in caves is not a general habitat feature for cave fishes. None the less predation regimes differ strikingly between epigean and hypogean habitats. The prey preference of Belostoma indicates that cave-dwelling P. mexicana experience size-specific predation pressure comparable with surface populations, which may have implications for life-history evolution in this cave fish.     

Comparative ultrastructural investigations of the pineal organ of the blind cave fish,Anoptichthys jordani,and its ancestor,the eyed river fish,Astyanax mexicanus

A comparative ultrastructural study has been made of the pineal organ in specimens of two closely related populations of the characid fish, Astyanaz mexicanus. The specimens of one population are living in the river, under natural light conditions. The specimens of the other population, originally described as Anoptichthys jordani, are living in a completely dark cave. In specimens of both populations the pineal organ consists of a spindle shaped end-vesicle, connected to the diencephalic roof by a slender stalk. The pineal tissue is compact and consists predominantly of glia-like supporting cells and sensory cells resembling the photoreceptor cells of the lateral vertebrate eye. Phagocytotic microglia-like cells can be found in close contact with the outer segments of the sensory cells. Nerve cells are located in the neighbourhood of neuropil formations, in which synaptic contacts are established between sensory cells and nerve cells. From these nerve cells fibers are emerging, forming the pineal tract that runs down the pineal stalk towards the diencephalon. On the basis of the ultrastructure described by other authors it is concluded that the pineal organ in specimens of the river population of Astyanax mexicanus resembles the pineal organ of other fish species. In specimens of the river population, reared under normal light-dark conditions for 3, 9 or 18 months, conspicuous morphological changes have not been detected in the presumably light-sensitive outer segments of the sensory cells or in other parts of the pineal tissue. In specimens of the cave populations, reared under identical conditions, an age-dependent, gradual regression of the regular outer segment organization of the pineal sensory cells takes place. In other parts of the pineal tissue, only small morphological changes can be observed. In specimens of the cave population, reared in constant darkness, the regression of the pineal outer segment organization begins earlier and is obvious. It is postulated that the gradual age-dependent regression of the regular organization of the outer segments in the pineal organ of cave specimens of Astyanax mexicanus is genetically determined and indicates a regressive evolution of the pineal light sensitivity. The expression of the regressive traits is dependent on the environmental light conditions.     

Regressive evolution in the Mexican cave tetra, Astyanax mexicanus

The evolutionary forces driving the reduction of eyes and pigmentation in cave-adapted animals are unknown; Darwin famously questioned the role of natural selection in eye loss in cave fishes: "As it is difficult to imagine that eyes, although useless, could be in any way injurious to animals living in darkness, I attribute their loss wholly to disuse"[1]. We studied the genetics of eye and pigmentation regression in the Mexican cave tetra, Astyanax mexicanus, by mapping and quantitative trait loci (QTL) analysis. We also mapped QTL for the putatively constructive traits of jaw size, tooth number, and numbers of taste buds. The data suggest that eyes and pigmentation regressed through different mechanisms. Cave alleles at every eye or lens QTL we detected caused size reductions, consistent with evolution by natural selection but not with drift. QTL polarities for melanophore number were mixed, however, consistent with genetic drift. Arguments against a role for selection in the regression of cave-fish eyes cited the insignificant cost of their development [2, 3], but we argue that the energetic cost of their maintenance is sufficiently high for eyes to be detrimental in the cave environment. Regression can be caused either by selection or drift.     

Eye reduction and phylogenetic age in ophidiiform cave fish1

The West Indian Ophidiiform fish are characterized by the occurrence of several troglobitic species. They are Ogilbia (=Typhliasina pearsei (Yucatan), Lucifuga subterruneus, L. dentatus and L. simile (Cuba), and L. spelaeotes (Babama Islands). The surface relatives, e.g. Ogilbia spp., prefer lightpoor zones. This is shown by a large lens and a retina almost exclusively consistin of rods. L. dentatus, L. subterraneus, and O. pearsei possess extremely reduced eyes. The eye of L. selaeotes has morphologically an intermediate position. In correlation to varying eye size a different number of rods may be developed. Furthermore in some specimens the lens starts degenerating. Contrar to the extremely reduced cave forms all specimens of L. spelaeotes can still perceive light with tleir eyes. The different degrees of reduction in the West Indian Ophidiiform cave fish probably reflect varying phylogenetic ages. A possible date of origin of L. spelaeotes could be at the beginning or the end of the last glaciation. The eyes of neonate individuals of the live-bearing L. dentatus and O. pearsei possess a lens and a retina rudiment. No fundamental structural and histological differences between both species were found at this stage.     

Life on the edge: hydrogen sulfide and the fish communities of a Mexican cave and surrounding waters

Most eucaryotic organisms classified as living in an extreme habitat are invertebrates. Here we report of a fish living in a Mexican cave (Cueva del Azufre) that is rich in highly toxic H₂S. We compared the water chemistry and fish communities of the cave and several nearby surface streams. Our study revealed high concentrations of H₂S in the cave and its outflow (El Azufre). The concentrations of H₂S reach more than 300 μM inside the cave, which are acutely toxic for most fishes. In both sulfidic habitats, the diversity of fishes was heavily reduced, and Poecilia mexicana was the dominant species indicating that the presence of H₂S has an all-or-none effect, permitting only few species to survive in sulfidic habitats. Compared to habitats without H₂S, P. mexicana from the cave and the outflow have a significantly lower body condition. Although there are microhabitats with varying concentrations of H₂S within the cave, we could not find a higher fish density in areas with lower concentrations of H₂S. We discuss that P. mexicana is one of the few extremophile vertebrates. Our study supports the idea that extreme habitats lead to an impoverished species diversity.     

Influence of light and darkness on the ultrastructure of the pineal organ in the blind cave fish,Astyanax mexicanus

Ultrastructural changes of the pineal organ were investigated in the blind cave fish, Astyanax mexicanus, kept under continous artificial light (5000 lux), in continuous darkness, and under natural light conditions. The pineal end-vesicle of the fish kept under natural photoperiod consisted of photoreceptor cells and supporting cells mixed with a few ganglion cells. The photoreceptor cells possessed well-developed outer segments with regularly arranged lamellar membranes. The supporting cells contained a number of lipid droplets and large globular cisternae filled with fine granules. In the fish kept under continuous light or in darkness, the pineal end-vesicle displayed a dilated lumen, and the outer segments of the receptors showed signs of degeneration. Furthermore, alterations of cell organelles were observed in the photoreceptor and supporting cells.     

Developmental constraints in regressive evolution: studies of the expression of the γs-crystallin gene in the developing lens of cave-dwelling Astyanax fasciatus (Cuvier, 1819) (Teleostei, Characidae) by in situ hybridization

The early morphogenesis of the lens and the expression of the γs-crystallin gene was studied in epigean Astyanax fasciatus and its cave-dwelling derivative. At early stages, the lens of the cave fish develops in a way that is similar to the epigean form. Later, the developmental timing is delayed and growth ceases in the cave-fish lens. With the beginning of cytodifferentiation, the development of the lens breaks down. Crystallin lens fibres are not produced at any time and the γs-crystallin gene, which is transcribed during a limited period in the lens of epigean fishes, is not active in cave specimens. This study confirms earlier immunofluorescence observations that demonstrated the lack of crystallin proteins in the cave-fish lens, but is in contrast to results on the blind mole rat, which showed a persistence of functioning crystallins in the degenerated lens of this species. The significance of developmental constraints in regressive evolution is discussed.     

Sex recognition in surface- and cave-dwelling Atlantic molly females (<Emphasis Type="Italic">Poecilia mexicana</Emphasis>, Poeciliidae,Teleostei): influence of visual and non-visual cues

Cave fishes need to rely on non-visual senses, such as the sense of smell or the lateral line to communicate in darkness. In the present study, we investigated sex identification by females of a cave-dwelling livebearing fish, Poecilia mexicana (cave molly), as well as its surface-dwelling relatives. Unlike many other cave fishes, cave mollies still possess functional eyes. Three different modes of presentation of the stimulus fish (a male and an equally sized female) were used: (i) the stimulus fish were presented behind wire-mesh in light, allowing the focal female to perceive multiple cues, (ii) the experiment was carried out under infrared conditions, such that only non-visual cues could be perceived and (iii) the stimulus fish were presented in light behind transparent Plexiglas, allowing for the use of visual cues only. Females of all populations examined preferred to associate with the stimulus female in at least one of the treatments, but only when visible light was provided, suggesting that far-range sex recognition is limited or even absent in the cave molly under naturally dark conditions.     

Spatial parameters encoded in the spatial map of the blind Mexican cave fish, Astyanax fasciatus

Although much is now known about the mechanisms that insects, birds and mammals use to orient within familiar areas, our knowledge of such mechanisms in fish is scant. I used the transformational approach to test whether the blind Mexican cave fish can encode shape and size in an internal representation of space. These fish are excellent study animals, as they swim at high velocities (presumably to enhance lateral line organ stimulation) when faced with unfamiliar landmarks or environments. As they are blind, potentially confounding cues from visual global landmarks are unavailable. The fish learnt a square configuration of four landmarks and so must have been be able to encode spatial relationships between the elements within this configuration. After learning landmark arrays, the cave fish showed significant dishabituation (swimming velocity was increased) when exposed to landmark transformations. The fish must therefore have been comparing the environment that they perceived with an internal representation of the environment that they had learnt. The results show that blind Mexican cave fish can encode size (absolute distance between landmarks) and possibly also shape within their spatial maps.     

The function of wall-following behaviors in the Mexican blind cavefish and a sighted relative,the Mexican tetra (<Emphasis Type="Italic">Astyanax</Emphasis>)

Mexican blind cavefish exhibit an unconditioned wall-following behavior in response to novel environments. Similar behaviors have been observed in a wide variety of animals, but the biological significance and evolutionary history of this behavior are largely unknown. In this study, the behaviors of Mexican blind cavefish (Astyanax sp.) and sighted Mexican tetra (Astyanax mexicanus) were videotaped after fish were introduced into a novel environment under dark (infrared) or well-lit conditions. Under dark conditions, both sighted and blind morphs exhibited wall-following behaviors with subtle but significant differences. Blind morphs swam more nearly parallel to the wall, exhibited greater wall-following continuity and reached higher levels of sustained swimming speeds more quickly than sighted morphs. In contrast, sighted morphs in the light remained motionless near the wall for long periods of time or moved slowly around the center of the tank without entraining to the walls. These results are consistent with the idea that wall-following is a shared, primitive trait that serves an exploratory function under dark conditions to compensate for the absence of vision. This behavior has become more honed in blind morphs for exploratory purposes—in large part due to the enhanced, active-flow sensing abilities of the lateral line.     

Hepatic respiratory compensation and haematological changes in the cave cyprinid, Phreatichthys andruzzii

In several ectotherms, including all members of the Osteichthyes studied so far, the spleen is capable of storing and releasing erythrocytes according to the animal's respiratory needs. The tropical cave cyprinid Phreatichthys andruzzii uses its liver rather than the spleen as the site of accumulation in the respiratory compensation process, like the amphibian Rana esculenta. The reversible process of erythrocyte accumulation in the liver is very evident in animals anaesthetized with chlorobutanol; MS-222, an anaesthetic widely used in lower vertebrates alters all the haematological parameters and is not suitable for studies on blood and respiration. The hepatic respiratory compensation mechanism is as efficient as the splenic one: in animals kept at 18 °C for 24 h the mean liver weight percentage was 70% higher than in specimens kept at 28 °C (2.43% of the body weight compared to 1.39%, in groups of six specimens) while mean red blood cell counts fell from 2.49 to 1.60 · 10¹² per l, in agreement with the haematocrit value and haemoglobin concentration; mean corpuscular volume remained constant (at about 177 fl). Accepted: 15 April 1997     

Influence of male competition on male mating behaviour in the cave molly, Poecilia mexicana

In many species, male mating behaviour is correlated with male body size, with large males often being preferred by females. Small surface-dwelling Poecilia mexicana males compensate for this disadvantage by being more sexually active and using sneaky copulations. In a cave-dwelling population, however, small males do not show this behaviour. Do small males alter their behaviour in the presence of a large rival? Here, we investigated the influence of male competition on male mating behaviour in the cave form. Two males of different sizes were mated with a female either alone or together with the other male. No aggressive interactions were observed between either fish. There was no statistically significant difference in the frequency of sexual behaviours between the two treatments. In both treatments, large males were more sexually active than small males. Thus, small cave molly males do not switch to an alternative mating behaviour in the presence of a larger rival. Possibly, the extreme environmental conditions in the cave (e.g. low oxygen content and high levels of hydrogen sulphide) favour saving energetic costs, resulting in the absence of alternative mating behaviour in small males.     

Oxygen consumption of Astyanax fasciatus (Characidae,Pisces): a comparison of epigean and hypogean populations

Synopsis The standard and routine oxygen consumptions of Astyanax fasciatus from one surface population (Rio Teapao) and three cave populations (Chica, Micos and Pachon caves: sAnoptichthys jordani, the Micosfish and Anoptichthys antrobius) were determined individually over 24 hours by the use of a flow-through respirometer and polarographic oxygen electrodes. The phylogenetically oldest Pachon fish had a significantly lower standard metabolic rate (0.230 ± 0.036 mg O₂ g^-1 h^-1) than the epigean Teapao fish, the hybrid Chica fish and the phylogenetically younger Micos fish (0.314 ± 0.081 mg O₂g-^-1h^-1, 0.284 ± 0.048 mg O₂g^-1h^-1, 0.277 ± 0.063 mg O₂g^-1h^-1). No significant differences could be determined among the latter three populations. A significant difference in routine metabolic rate existed only between the Pachon fish (0.309 ± 0.0.56 mg O₂g^-1h^-1) and the Teapao fish (0.415 ± 0.071 mg O₂g^-1h^-1). The Chica fish (0.356 ± 0.084 mg O₂g^-1h^-1) and the Micos fish (0.355 ± 0.080 mg O₂ g^-1h^-1) could not be separated from either the Teapao or the Pachon fish, but a decreasing trend from the surface population through the Chica and the Micos to the Pachon population was obvious. During a starvation period of 29 days the metabolic rate of epigean Teapao and hypogean Pachon fish decreased significantly by 32.5% and 34.8% (standard oxygen consumption rate) and 27.5% and 28.2% (routine oxygen consumption rate), respectively. Body mass loss during the starvation period was 16.3% for the Teapao fish and 9.5% for the Pachon fish.     

Sperm production in an extremophile fish,the cave molly (<Emphasis Type="Italic">Poecilia mexicana</Emphasis>, Poeciliidae,Teleostei)

A prominent trade-off in life history theory and evolution balances the costs of reproduction with those of basic somatic needs. Hence, reproductive efforts may be reduced in environments where additional energy is required for somatic maintenance. Here, we investigated male sperm stores in Atlantic mollies (Poecilia mexicana) from a sulfidic cave and several sulfidic and non-sulfidic surface habitats. We found significant differences among populations in the number of sperm stripped per male, which was also correlated with differences in gonad weights. The largest sperm stores were detected in males from non-sulfidic surface creeks, while males from a partially sulfidic surface system had lower sperm counts, and males from completely sulfidic systems, surface as well as subterranean, had even fewer available sperm. We conclude that the extreme environmental conditions in sulfidic habitats appear to constrain male sperm production, since hydrogen sulfide as a naturally occurring toxin requires energy-demanding adaptations. Furthermore, we examined sperm counts of lab-reared cave and surface mollies in response to energy limitation. Males from stock populations were placed under high and low food treatments for a 2-week period and then stripped of sperm. Sperm counts of surface mollies tended to be reduced by low food availability, whereas sperm counts of cave mollies did not significantly vary between food treatments, which likely points towards a higher starvation resistance in cave mollies.