Comparative responses to severe hypoxia and subsequent recovery in closely related amphipod populations (Gammarus minus) from cave and surface habitats

The locomotory and ventilatory activities, oxygen consumption, and the intermediary and energy metabolism modifications of a spring and a cave population of the aquatic amphipod crustacean Gammarus minus were investigated in normoxia, severe hypoxia ( < 0.03 kPa) and subsequent recovery. The aims of this study were to compare (1) the reactions of both populations to these experimental conditions, (2) these results with those obtained on the hypogean amphipod Niphargus, and (3) the degree of adaptation to hypoxia showed by both populations of G. minus. Despite their different origins, both populations of G. minus presented identical responses in all experimental conditions. The lethal time for 50% of the population was about 6 h, and the oxygen consumption about 44 μmol O₂/g dw per h in normoxic conditions. The metabolic effects of severe hypoxia and subsequent recovery were significant compared to normoxic conditions, but also similar between both populations for alanine, arginine phosphate, ATP, glycogen and lactate levels. This study (i) underlines the statement that a high resistance to lack of oxygen is not universally found in subterranean organisms, but is more related to oxygen availability and/or to the energetic state of each subterranean ecosystem, and (ii) highlight the diversity of adaptive responses to an environmental constraint expressed by hypogean crustaceans. This revised version was published online in July 2006 with corrections to the Cover Date.     

Comparative study of electron transport system activity and oxygen consumption of amphipods from caves and surface habitats

1. Respiratory electron transport system (ETS) activity and oxygen consumption were measured at 10 °C for the hypogean amphipods Niphargus stygius (from two epikarst locations) and N. krameri (from a sinking river in a cave) and the epigean amphipod Gammarus fossarum (inhabiting a spring and a river). 2. ETS activity and oxygen consumption were compared between caves and the two surface locations, and between hypogean and epigean amphipods. ETS activities were found not to differ between animals from different locations, or between epigean and hypogean amphipods. As expected, the oxygen consumption of N. krameri and N. stygius was lower than that of G. fossarum. 3. The high ratio between ETS activity and oxygen consumption (ETS/R ratio) showed that N. krameri and N. stygius possess high metabolic potential that can be exploited for energy recovery and rapid restoration of body reserves immediately following the appearance of favourable conditions (food and/or oxygen). In contrast, G. fossarum exhibited a low ETS/R ratio, indicating exploitation of a great proportion of the metabolic potential for standard metabolism. 4. Feeding in N. stygius increased oxygen consumption but not ETS activity, and thus fed N. stygius had a much lower ETS/R ratio than starved ones. 5. A relatively high metabolic potential (i.e. high ETS/R ratio) may be an adaptation to hypogean life in amphipods, in that it improves survival under poor food and/or oxygen conditions.     

Behavioural, ventilatory and respiratory responses of epigean and hypogean crustaceans to different temperatures

Impact of temperature (from -2 to 28 degrees C) on survival, oxygen consumption, locomotory and ventilatory activities was measured in two aquatic subterranean crustaceans (Niphargus rhenorhodanensis and Niphargus virei) and in a morphologically close surface-dwelling crustacean (Gammarus fossarum). The hypogean N. virei presented all characteristics of a stenothermal organism: it showed small thermal plasticity and optimised its performance on a narrow range of temperature. In contrast, the epigean G. fossarum and more surprisingly the hypogean N. rhenorodanensis can be both characterized as eurythermal organisms: they exhibited important survival times and conserved their performance optimum throughout a large range of temperature. Such differences of survival and performance patterns in two hypogean organisms were unexpected since they both live in very thermally buffered biotopes. Our data suggest fresh hypotheses about the role of glaciations in the history and adaptation of hypogean crustaceans.     

Selection of physiological and metabolic adaptations to food deprivation in the Pyrenean newt Calotriton asper during cave colonisation

Food restriction is one of the major and most common constraints that subterranean animals face in their biotope. Cave-dwelling organisms thus have to cope with fasting periods that can extend from a month to a year. However, adaptive fasting resistance previously found in subterranean fauna has only been highlighted by direct comparisons with phylogenetically distant epigean organisms, which could severely impact conclusions. Here we report physiological and metabolic responses to 42 days of fasting followed by 10 days of refeeding in two populations (one subterranean and one epigean) of Calotriton asper. In the fed state (control), the hypogean population exhibited a hypometabolism together with higher glycogen (+ 25% in liver and muscles) and triglyceride stores (+ 50% in muscles). During the fasting period, cave individuals exhibited a 20% decrease in VO₂ whereas epigean individuals experienced no significant change. In addition, the energetic reserves always remained higher in the hypogean population. According to phylogenic and biogeographic data, cave colonization by this species dates back to less than 10,000 years, suggesting a rapid selection of adaptive traits related to fasting. This study strongly suggests that cave colonization induces a decrease in metabolism together with a higher capacity to accumulate energy reserves and therefore to withstand unpredictable fasting periods.     

Oxygen supply and the adaptations of animals in groundwater

1. The first part of this review focuses on the oxygen status of natural groundwater systems (mainly porous aquifers) and hyporheic zones of streams. The second part examines the sensitivity of groundwater organisms, especially crustaceans, to low oxygen concentrations (< 3.0 mg L^?1 O₂). 2. Dissolved oxygen (DO) in groundwater is spatially heterogeneous at macro- (km), meso- (m) and micro- (cm) scales. This heterogeneity, an essential feature of the groundwater environment, reflects changes in sediment composition and structure, groundwater flow velocity, organic matter content, and the abundance and activity of micro-organisms. Dissolved oxygen also exhibits strong temporal changes in the hyporheic zone of streams as well as in the recharge area of aquifers, but these fluctuations should be strongly attenuated with increasing distance from the stream and the recharge zone. 3. Dissolved oxygen gradients along flow paths in groundwater systems and hyporheic zones vary over several orders of magnitude (e.g. declines of 9 × 10^?5 to 1.5 ×10^?2 mg L^?1 O₂ m^?1 in confined aquifers and 2 × 10^?2 to 1 mg L^?1 O₂ m^?1 in parafluvial water). Several factors explain this strong variation. Where the water table is close to the surface, oxygen is likely to be consumed rapidly in the first few metres below the water table because of incomplete degradation of soil-generated labile dissolved organic carbon (DOC) in the vadose zone. Where the water table is far from the surface, strong oxygen depletion in the vicinity of the water table does not occur, DO being then gradually consumed as groundwater flows down the hydraulic gradient. In unconfined groundwater systems, oxygen consumption along flow paths may be compensated by down-gradient replenishment of DO, resulting either from the ingress of atmospheric oxygen or water recharge through the vadose zone. In confined groundwater systems, where replenishment of oxygen is impossible, the removal time of DO varies from a few years to more than 10 000 years, depending mainly on the organic carbon content of the sediment. Comparison of the hyporheic zones between systems also revealed strong differences in the removal time and length of underground pathways for DO. This strong variability among systems seems related to differences in contact time of water with sediment. 4. Although groundwater macro-crustaceans are much more resistant to hypoxia than epigean species, they cannot survive severe hypoxia (DO < 0.01 mg L^?1 O₂) for very long (lethal time for 50% of the population ranged from 46.7 to 61.7 h). In severe hypoxia, none of the hypogean crustaceans examined utilized a high-ATP yielding metabolic pathway. High survival times are mainly a result of the combination of three mechanisms: a high storage of fermentable fuels (glycogen and phosphagen), a low metabolic rate in normoxia, and a further reduction in metabolic rate by reducing locomotion and ventilation. It is suggested here that the low metabolic rate of many hypogean species may be an adaptation to low oxygen and not necessarily result from an impoverished food supply. 5. An interesting physiological feature of hypogean crustaceans is their ability to recover from anaerobic stress and, more specifically, rapidly to resynthesize glycogen stores during post-hypoxic recovery. A high storage and rapid restoration of fermentable fuels (without feeding) allows groundwater crustaceans to exploit a moving mosaic of suboxic (< 0.3 mg L?1 O₂), dysoxic (0.3–3.0 mg L^?1 O₂) and oxic (> 3 mg L^?1 O₂) patches. 6. It is concluded that although hypogean animals are probably unsuited for life in extensively or permanently suboxic groundwater, they can be found in small or temporarily suboxic patches. Indeed, their adaptations to hypoxia are clearly suited for life in groundwater characterized by spatially heterogeneous or highly dynamic DO concentrations. Their capacity to survive severe hypoxia for a few days and to recover rapidly would explain partly why ecological field studies often reveal the occurrence of interstitial taxa in groundwater with a wide range of DO.     

Freshwater amphipods as biomonitors of metal pollution in surface and interstitial aquatic systems

1. The relationships between levels of metals (Zn, Cu) in the environment (water, sediment) and in the tissues of three species of amphipod crustaceans were investigated in superficial and interstitial ecosystems of the Rivers Rhone and Ain, France. 2. Epigean and hypogean amphipods differed in their ability to accumulate Zn and Cu. At all sites, Gammarus fossarum (epigean) showed lower Zn or Cu body concentrations than Niphargus of, rhenorhodanensis or Niphargopsis casparyi (hypogean). Differences were also observed between the two interstitial taxa. 3. A significant difference among sites was observed for metal concentrations in sediment. In contrast, no significant inter-site differences were found for metal concentrations in organisms. 4. No significant correlation was found between metal concentrations in the three taxa and quantities of metals in the water. A positive correlation was observed between Zn and Cu body content in Gammarus fossarum and metal level in the sediment. No relationships were found between interstitial amphipods and metal concentrations in the sediment. 5. Differences in accumulation rates among the three amphipod species suggest their potential use as biological indicators for monitoring metal pollution.     

Global diversity of amphipods (Amphipoda; Crustacea) in freshwater

Amphipods are brooding peracaridan crustaceans whose young undergo direct development, with no independent larval dispersal stage. Most species are epibenthic, benthic, or subterranean. There are some 1,870 amphipod species and subspecies recognized from fresh or inland waters worldwide at the end of 2005. This accounts for 20% of the total known amphipod diversity. The actual diversity may still be several-fold. Amphipods are most abundant in cool and temperate environments; they are particularly diversified in subterranean environments and in running waters (fragmented habitats), and in temperate ancient lakes, but are notably rare in the tropics. Of the described freshwater taxa 70% are Palearctic, 13% Nearctic, 7% Neotropical, 6% Australasian and 3% Afrotropical. Approximately 45% of the taxa are subterranean; subterranean diversity is highest in the karst landscapes of Central and Southern Europe (e.g., Niphargidae), North America (Crangonyctidae), and Australia (Paramelitidae). The majority of Palearctic epigean amphipods are in the superfamily Gammaroidea, whereas talitroid amphipods (Hyalella) account for all Neotropic and much of the Nearctic epigean fauna. Major concentrations of endemic species diversity occur in Southern Europe, Lake Baikal, the Ponto-Caspian basin, Southern Australia (including Tasmania), and the south-eastern USA. Endemic family diversity is similarly centered in the Western Palearctic and Lake Baikal. Freshwater amphipods are greatly polyphyletic, continental invasions have taken place repeatedly in different time frames and regions of the world. In the recent decades, human mediated invasions of Ponto-Caspian amphipods have had great impacts on European fluvial ecosystems. Guest editors: E. V. Balian, C. Lévêque, H. Segers and K. Martens Freshwater Animal Diversity Assessment     

The Laterosensory Canal System in Epigean and Subterranean Ituglanis (Siluriformes: Trichomycteridae), With Comments About Troglomorphism and the Phylogeny of the Genus

The laterosensory system is a mechanosensory modality involved in many aspects of fish biology and behavior. Laterosensory perception may be crucial for individual survival, especially in habitats where other sensory modalities are generally useless, such as the permanently aphotic subterranean environment. In the present study, we describe the laterosensory canal system of epigean and subterranean species of the genus Ituglanis (Siluriformes: Trichomycteridae). With seven independent colonizations of the subterranean environment in a limited geographical range coupled with a high diversity of epigean forms, the genus is an excellent model for the study of morphological specialization to hypogean life. The comparison between epigean and subterranean species reveals a trend toward reduction of the laterosensory canal system in the subterranean species, coupled with higher intraspecific variability and asymmetry. This trend is mirrored in other subterranean fishes and in species living in different confined spaces, like the interstitial environment. Therefore, we propose that the reduction of the laterosensory canal system should be regarded as a troglomorphic (= cave‐related) character for subterranean fishes. We also comment about the patterns of the laterosensory canal system in trichomycterids and use the diversity of this system among species of Ituglanis to infer phylogenetic relationships within the genus. J. Morphol. 278:4–28, 2017. ©© 2016 Wiley Periodicals,Inc.     

Effect of fasting on hypogean (<Emphasis Type="Italic">Niphargus stygius</Emphasis>) and epigean (<Emphasis Type="Italic">Gammarus fossarum</Emphasis>) amphipods: a laboratory study

Two amphipods, the hypogean Niphargus stygius and epigean Gammarus fossarum, were analyzed for fatty acid (FA) composition, electron transport system (ETS) activity and respiration (R) during a laboratory fasting experiment. In agreement with ETS and R measurements (and the ETS/R ratio), the hypogean N. stygius utilized FA more slowly than the epigean G. fossarum. Inter-specific differences in the utilization of certain FA during fasting were also revealed. While N. stygius tended to preserve all of its FA during the experimental fasting period, G. fossarum showed a tendency to utilize MUFA (monounsaturated FA) and SAFA (saturated FA) and preferentially retain PUFA (polyunsaturated FA). The significant correlations between ETS activity and composition of specific FA during fasting can be linked to R. During the fasting, both ETS activity and respiration rate of G. fossarum decreased, however, ETS/R ratio increased. In contrast, N. stygius did not show significant changes in these parameters. This is the first report, which connects ETS activity with changes in concentrations of specific FA during fasting. Such evolutionary adaptations of hypogean species enables them to better survive chronically low and/or discontinuous food supplies compared to epigean species, which live in environments where food shortages are much less frequent.     

Morphology and reproduction of the cavefish Trichomycterus chaberti and the related epigean Trichomycterus cf. barbouri

Hypogean and epigean populations of Trichomycterus catfishes inhabit streams from different environments (cave, headwater, canyon and valley) in the Torotoro National Park in the Andes, Bolivia. A significant reduction in the diameter of the eyes and in the surface area of the mesencephalon was observed in subterranean populations, along with an increase in the surface area of the telencephalon. Contrary to expectations, the barbel did not appear to be longer in hypogean populations. The observed pattern of modification of the other variables (pigmentation, eye asymmetry, surface area of the cerebellum and rhombencephalon, fecundity and egg diameter) corresponded to a gradient of values from valley to canyon, headwater and subterranean populations. This result argues not for a simple distinction between epigean and hypogean populations but for an adaptation to an environmental gradient of constraints in which caves correspond to an extreme situation.     

Temporal Organization in Locomotor Activity of the Hypogean Loach, Nemacheilus Evezardi, and its Epigean Ancestor

Locomotor activity rhythm in the hypogean population of Nemacheilus evezardi was recorded first under light-to-dark (LD) 12 : 12 h cycle and then DD. The results were compared with that of its epigean counterpart held under comparable regimes. In LD 12 : 12, while hypogean loach exhibited a distinct bimodality in its locomotor activity rhythm, it was altogether absent in the case of epigean population. In hypogean loach, dark-to-light transition peak in LD was observed to free-run under DD. The same was not discernible in case of epigean loach. The locomotor activity rhythm in epigean fish was noticed to free-run in DD either from the dawn peak or dusk peak in LD. It is hypothesized that the hypogean fish still possesses a functional oscillator underlying its overt circadian rhythm in locomotor activity. The ecophysiological significance of these findings is yet to be fully understood.     

Regional Risk Assessment of a Brazilian Rain Forest Reserve

The objective of this study was to identify subareas inside and near an Atlantic Rain Forest reserve, the Parque Estadual Turístico do Alto Ribeira (PETAR), most likely to be affected by land use in the vicinity of the area. In addition, the study aimed to compare risks per stressor source (agriculture, human settlements and mining) to both epigean (surface) and hypogean (subterranean) aquatic fauna. The methodological approach included the relative vulnerability of endpoints to the stressors (pesticides, metals, nutrients, and particles) and ranking of stressor sources and habitats (epigean and hypogean streams) based on their relative distribution in 14 subareas within the catchment areas of the main rivers that cross PETAR: Pilões, Betari and Iporanga. Four subareas presented high risk for both epigean and hypogean fauna. Three of those areas were located inside the Betari catchment area, where most of the settlements and abandoned lead mines are located. The fourth area was situated in the headwaters of the Pilões River, where agricultural activities are intense. Agriculture and human settlements were the activities most likely to cause impacts on aquatic ecosystems. Uses of risk assessment results include management of the PETAR and communication to stakeholders by the Park Administration.     

Metabolic responses and arginine kinase expression under hypoxic stress of the kuruma prawn Marsupenaeus japonicus

In response to hypoxia at PO(2) 1.3-1.7 mg/L for 6 h, the kuruma prawn Marsupenaeus (Penaeus) japonicus showed a dramatic decrease in phosphoarginine storage in muscle, with normal levels restored during 4-h post-hypoxic recovery. Large stores of muscle glycogen only decreased between 4 and 6 h during hypoxia, but greatly diminished during recovery. Muscle ATP levels and energy charge decreased only slightly under hypoxia. Lactate levels increased slightly during hypoxia and promptly returned to control levels during recovery. These data indicate that phosphoarginine works in muscle as an ATP buffer during hypoxia and glycogen is utilized as an energy source during recovery. Under hypoxia, up- and down-regulated proteins were identified after 2D electrophoresis and partial sequences were obtained after protease digestion. Fructose bisphosphate aldolase was down-regulated during hypoxia, suggesting the suppression of glycolysis under hypoxia. Several partial sequences from three protein spots up-regulated under hypoxia were all assigned to arginine kinase, suggesting the existence of several isoforms of arginine kinase in the muscle of M. japonicus. This arginine kinase up-regulation under hypoxia may indicate a provision for oxygen re-supply after anaerobiosis. This is consistent with the prompt replenishment of phosphoarginine stores during recovery from hypoxia.     

Anaerobic metabolism in the lugworm Arenicola Marina L.: The transition from aerobic to anaerobic metabolism

After onset of anaerobic conditions Arenicola Marina does not switch immediately to typical anaerobiosis. The adaptation occurs in three steps.

• 1.1. In the first 3 hr the phosphagen stores are mobilized, glycogen is degraded to alanine, aspartate is metabolized to succinate and volatile fatty acids (transition period).
• 2.2. Starting approximately after 3 hr, the route of glycogen degradation at the phosphoenolpyruvate-branchpoint is changed gradually from metabolizing via pyruvate kinase to phosphoenolpyruvate carboxykinase. At the same time the metabolic rate is reduced significantly (switching period).
• 3.3. After more than 12 hr the energy supply occurs exclusively via the known pathways typical for long-term anaerobiosis.

     

Dynamics of surface water/groundwater ecotones in a karstic aquifer

SUMMARY.

• 1 Interactions between surface waters and groundwaters were analysed by studying degrees of similarity between a surface stream and groundwater in a karstic system.
• 2 Exchange of water and solutes (‘abiotic interactions’) and living organisms (‘biotic interactions’) were quantified by characteristic parameters measured at the outlet of the karstic system during hydrological changes across the year as well as during a flood.
• 3 The ratio of bicarbonate to calcium described abiotic interactions. In the seasonal study, biotic interactions were estimated from the proportion of epigean organisms in the communities sampled at the outlet. These interactions during the flood were estimated by comparing numbers of Cladocera, epigean organisms, with numbers of Niphargus virei, hypogean organisms.
• 4 The location of the ecotone created by interactions between the surface stream and the karstic system showed marked spatial fluctuations according to the prevailing hydrology. Interactions were strong during high-water periods whereas they were negligible during low-water. Similar fluctuations occurred within a few hours during the flood.
• 5 Results are compared with those from fluvial systems to discuss the importance of the dynamics of these surface/underground ecotones in the functioning of lotie ecosystems.

     

Is the reduced metabolism of hypogean amphipods solely a result of food limitation?

The epigean amphipod Gammarus lacustris possessed a rate of oxygen uptake (MO2) three times greater that of the hypogean amphipod Stygobromus sp. While much of the difference between these two species could be attributed to different feeding regimes, the hypogean amphipod still exhibited a lower rate of metabolism than the epigean species. This suggests that there can be both a genetic and an environmental (reduced food availability and/or hypoxia) component to the reduced metabolism characteristic of many hypogean animals. The results of this study have re-emphasised the pitfall of making species comparisons without knowledge of the extent of physiological variation within a species. This revised version was published online in July 2006 with corrections to the Cover Date.     

Ecology of Subterranean Fishes: An Overview

A synthesis of ecological data available for subterranean fishes throughout the world is presented, and comparatively analyzed in an evolutionary context. Methods of ecological research are described, and their potential and limitations for the study of hypogean fishes are discussed. Ecology of troglobitic (exclusively subterranean) fishes is discussed with focus on distribution areas, population densities and sizes, use of habitat and movements, life cycle and feeding. When data are available, these species are compared with their epigean relatives. Putative ecological autapomorphies of troglobitic fishes, including habitat change, adaptations to cope with food scarcity, and precocial lifestyles, are interpreted in ecological and evolutionary contexts. Species interactions among subterranean species, including cases of syntopy and predation are briefly analyzed. Non-troglobitic hypogean fishes, with their ecological importance and evolutionary role, are also addressed. Problems of classification of subterranean fishes according to the Schiner-Racovitza system (troglobites, troglophiles and trogloxenes) are discussed, and a scenario of evolution of subterranean populations is presented.     

Electron transport system activity and oxygen consumption of two amphibious isopods,epigean Ligia italica Fabricius and hypogean Titanethes albus (Koch), in air and water

Electron transport system (ETS) activity and oxygen consumption (i.e. respiration, R) were measured in two amphibious isopods, epigean Ligia italica and hypogean Titanethes albus. The measurements were made in air and in water to estimate the exploitation of metabolic potential for actual metabolism in the two media. Oxygen consumption was measured in animals in air before their transfer to water, in water and again after their transfer back to air. Both species showed greater oxygen consumption during submergence in water than in air. Although epigean L. italica had higher oxygen consumption and ETS activity than hypogean T. albus, both species exhibited high and variable ETS/R ratios. A high and variable ETS/R ratio is one of the characteristics of amphibious isopods that enables them to cope with specific environmental conditions in their habitats.     

Utilisation of glycogen,ATP and arginine phosphate in exercise and recovery in terrestrial red crabs,Gecarcoidea natalis

Intermittent locomotion by terrestrial crustaceans may under specific circumstances increase walking distance and may allow partial re-oxidization of anaerobic products, and replenishment of ATP and arginine phosphate. The Christmas Island red crab G. natalis undertakes a substantial breeding migration each year. The leg muscles of G. natalis subjected to bouts of 2.5 min walking and 2.5 min rest were severely anaerobic. Adenylate energy charge and the large arginine phosphate stores were greatly reduced. Walking for 4 min with pauses of only 1 min exacerbated the anaerobiosis and utilised 50% of the endogenous muscle glycogen. Post-exercise, the adenylate energy charge recovered before the arginine phosphate charge and a large and persistent hyperglycaemia accompanied the restoration of glycogen. Arginine phosphate functioned as a large, longer term, energy reservoir-almost as part of the adenylate pool. Gluconeogenesis is yet to be generally substantiated in decapod crustaceans but G. natalis appears to remove lactate slowly and to reincorporate exogenous glucose into muscle glycogen in the same time frame as lactate removal from the haemolymph. The 4:1 exercise/pause regimen facilitated access to energy stores and increased walking distance, and it allowed L-lactate and H(+) efflux from the muscle during pausing. These responses are similar to those of G. natalis in the field, except during the migration when walking was entirely aerobic. Determinations of adenylate, fuel and arginine phosphate reserves and usage during the migration are required together with more detailed behavioral analysis to resolve the dichotomy in metabolic response.