Magadi Tilapia Research Articles

The genome organization of the Lake Magadi tilapia, Oreochromis Alcolapia grahami, a cichlid extremophile

The genome of vertebrates is made of a mosaic of long stretches of DNA, called isochores, which are compositionally uniform, and belong to a few families of GC-poor (L1 and L2) and GC-rich (H1, H2, and H3) components. Poikilotherms tend to have GC-poor genomes, while endotherms comprise both GC-poor and GC-rich isochores. The thermal theory claimed that temperature and natural selection played an active role in favoring GC-rich genomic regions, yet empirical evidence was difficult to obtain. Early work based on cesium chloride ultracentrifugation gradients showed that the Lake Magadi tilapia, a hot-water adapted fish species, displayed GC-rich regions that were absent from a close relative that lives in colder water. The goal of this study was to revisit the original study using full genome sequencing. We found that the original GC-rich regions are indeed present, that they are interspersed in the genome. Indeed, when comparing Lake Magadi tilapia with the temperate Nile tilapia, we found that 59.3 % of the genome of Lake Magadi tilapia had a base composition higher than 40 %GC, as opposed to 55.3 % of the genome of the Nile tilapia having a base composition higher than 40 % GC. We also found that their genomes comprised similar amounts of repetitive elements (20 % and 19.5 %, respectively) indicating that the shifts in base composition might not be due to repetitive elements. Further work on repetitive element analyses, protein coding genes and additional hot-water adapted fishes will provide clues as to the origin of GC-rich isochores in Lake Magadi tilapia.

Fasting in the ureotelic Lake Magadi tilapia, Alcolapia grahami, does not reduce its high metabolic demand, increasing its vulnerability to siltation events.

Lake Magadi, Kenya, is one of the most extreme aquatic environments on Earth (pH~10, anoxic to hyperoxic, high temperatures). Recently, increased water demand and siltation have threatened the viable hot springs near the margins of the lake where Alcolapia grahami, the only fish surviving in the lake, live. These Lake Magadi tilapia largely depend on nitrogen-rich cyanobacteria for food and are 100% ureotelic. Their exceptionally high aerobic metabolic rate, together with their emaciated appearance, suggests that they are energy-limited. Therefore, we hypothesized that during food deprivation, Magadi tilapia would economize their energy expenditure and reduce metabolic rate, aerobic performance and urea-N excretion. Surprisingly, during a 5-day fasting period, routine metabolic rates increased and swimming performance (critical swimming speed) was not affected. Urea-N excretion remained stable despite the lack of their N-rich food source. Their nitrogen use switched to endogenous sources as liver and muscle protein levels decreased after a 5-day fast, indicating proteolysis. Additionally, fish relied on carbohydrates with lowered muscle glycogen levels, but there were no signs indicating use of lipid stores. Gene expression of gill and gut urea transporters were transiently reduced as were gill rhesus glycoprotein Rhbg and Rhcg-2. The reduction in gill glutamine synthetase expression concomitant with the reduction in Rh glycoprotein gene expression indicates reduced nitrogen/ammonia metabolism, most likely decreased protein synthesis. Additionally, fish showed reduced plasma total CO2, osmolality and Na+ (but not Cl−) levels, possibly related to reduced drinking rates and metabolic acidosis. Our work shows that Lake Magadi tilapia have the capacity to survive short periods of starvation which could occur when siltation linked to flash floods covers their main food source, but their seemingly hardwired high metabolic rates would compromise long-term survival.

Gyrodactylus magadiensis n. sp. (Monogenea, Gyrodactylidae) parasitising the gills of Alcolapia grahami (Perciformes, Cichlidae), a fish inhabiting the extreme environment of Lake Magadi, Kenya.

A new species of Gyrodactylus von Nordmann, 1832 is described from the gills of Alcolapia grahami, a tilapian fish endemic to Lake Magadi. This alkaline soda lake in the Rift Valley in Kenya is an extreme environment with pH as high as 11, temperatures up to 42 °C, and diurnal fluctuation between hyperoxia and virtual anoxia. Nevertheless, gyrodactylid monogeneans able to survive these hostile conditions were detected from the gills the Magadi tilapia. The worms were studied using light microscopy, isolated sclerites observed using scanning electron microscopy, and molecular techniques used to genetically characterize the specimens. The gyrodactylid was described as Gyrodactylus magadiensis n. sp. and could be distinguished from other Gyrodactylus species infecting African cichlid fish based on the comparatively long and narrow hamuli, a ventral bar with small rounded anterolateral processes and a tongue-shaped posterior membrane, and marginal hooks with slender sickles which are angled forward, a trapezoid to square toe, rounded heel, a long bridge prior to reaching marginal sickle shaft, and a long lateral edge of the toe. The species is also distinct from all other Gyrodactylus taxa based on the ITS region of rDNA (ITS1–5.8s–ITS2), strongly supporting the designation of a new species. These findings represent the second record of Gyrodactylus from Kenya, with the description of G. magadiensis bringing the total number of Gyrodactylus species described from African cichlids to 18.

Some like it hotter than others

![Graphic][1] If you want to meet a truly tough fish, buy a ticket to Lake Magadi, Kenya. Here, the Magadi tilapia swim happily in the hot, alkaline waters of the lake, where oxygen levels soar and plummet daily and water chemistry pushes the boundaries of acid–base regulation. Few fish can

Mammalian metabolic rates in the hottest fish on earth.

The Magadi tilapia, Alcolapia grahami, a small cichlid fish of Lake Magadi, Kenya lives in one of the most challenging aquatic environments on earth, characterized by very high alkalinity, unusual water chemistry, and extreme O2, ROS, and temperature regimes. In contrast to most fishes which live at temperatures substantially lower than the 36–40 °C of mammals and birds, an isolated population (South West Hot Springs, SWHS) of Magadi tilapia thrives in fast-flowing hotsprings with daytime highs of 43 °C and night-time lows of 32 °C. Another population (Fish Springs Lagoon, FSL) lives in a lagoon with fairly stable daily temperatures (33–36 °C). The upper critical temperatures (Ctmax) of both populations are very high; moreover the SWHS tilapia exhibit the highest Ctmax (45.6 °C) ever recorded for a fish. Routine rates of O2 consumption (MO2) measured on site, together with MO2 and swimming performance at 25, 32, and 39 °C in the laboratory, showed that the SWHS tilapia exhibited the greatest metabolic performance ever recorded in a fish. These rates were in the basal range of a small mammal of comparable size, and were all far higher than in the FSL fish. The SWHS tilapia represents a bellwether organism for global warming.

Magadi tilapia ecological specialization: filling the early gap in the speciation continuum.

Cichlid fish are well known for their high speciation rates, which are usually accompanied by spectacular and rapid diversification in eco-morphological and secondary sexual traits. This is best illustrated by the famous repeated explosive radiations in the African Great Lakes Tanganyika, Malawi and Victoria, each lake harbouring several hundreds of mostly endemic species. Correspondingly, cichlids diversified very rapidly in many other lakes across their range. Although the larger radiations, unparalleled in vertebrates, are certainly the most intriguing, they are also the most intricate and difficult to address because of their complex nature. This is where smaller, simpler systems may prove to be the most useful. In this issue of Molecular Ecology, Kavembe et al. (2016) report very recent genetic diversification accompanied by ecological specialization in cichlids of the small and ecologically extreme Lake Magadi, in Kenya. Combining geometric morphometrics, stable isotope analysis, population genomics using RADSeq data and coalescent-based modelling techniques, the authors characterize the eco-morphological differences between genetically distinct populations of Magadi tilapia (Alcolapia grahami), which are consistent with the different environmental conditions they experience, and infer their history of divergence. The simplicity of the focal system and the use of a multidisciplinary approach make this work particularly important for our understanding of the early stages of speciation, in both cichlids and other organisms.

Eco‐morphological differentiation in Lake Magadi tilapia, an extremophile cichlid fish living in hot, alkaline and hypersaline lakes in East Africa

Ecological diversification through divergent selection is thought to be a major force during the process of adaptive radiations. However, the large sizes and complexity of most radiations such as those of the cichlids in the African Great Lakes make it impossible to infer the exact evolutionary history of any population divergence event. The genus Alcolapia, a small cichlid lineage endemic to Lakes Magadi and Natron in East Africa, exhibits phenotypes similar to some of those found in cichlids of the radiations of the African Great Lakes. The simplicity within Alcolapia makes it an excellent model system to investigate ecological diversification and speciation. We used an integrated approach including population genomics based on RAD-seq data, geometric morphometrics and stable isotope analyses to investigate the eco-morphological diversification of tilapia in Lake Magadi and its satellite lake Little Magadi. Additionally, we reconstructed the demographic history of the species using coalescent simulations based on the joint site frequency spectrum. The population in Little Magadi has a characteristically upturned mouth--possibly an adaptation to feeding on prey from the water surface. Eco-morphological differences between populations within Lake Magadi are more subtle, but are consistent with known ecological differences between its lagoons such as high concentrations of nitrogen attributable to extensive guano deposits in Rest of Magadi relative to Fish Springs Lagoon. All populations diverged simultaneously only about 1100 generations ago. Differences in levels of gene flow between populations and the effective population sizes have likely resulted in the inferred heterogeneous patterns of genome-wide differentiation.

Genomics of Adaptation to Multiple Concurrent Stresses: Insights from Comparative Transcriptomics of a Cichlid Fish from One of Earth's Most Extreme Environments, the Hypersaline Soda Lake Magadi in Kenya, East Africa.

The Magadi tilapia (Alcolapia grahami) is a cichlid fish that inhabits one of the Earth's most extreme aquatic environments, with high pH (~10), salinity (~60% of seawater), high temperatures (~40 °C), and fluctuating oxygen regimes. The Magadi tilapia evolved several unique behavioral, physiological, and anatomical adaptations, some of which are constituent and thus retained in freshwater conditions. We conducted a transcriptomic analysis on A. grahami to study the evolutionary basis of tolerance to multiple stressors. To identify the adaptive regulatory changes associated with stress responses, we massively sequenced gill transcriptomes (RNAseq) from wild and freshwater-acclimated specimens of A. grahami. As a control, corresponding transcriptome data from Oreochromis leucostictus, a closely related freshwater species, were generated. We found expression differences in a large number of genes with known functions related to osmoregulation, energy metabolism, ion transport, and chemical detoxification. Over-representation of metabolism-related gene ontology terms in wild individuals compared to laboratory-acclimated specimens suggested that freshwater conditions greatly decrease the metabolic requirements of this species. Twenty-five genes with diverse physiological functions related to responses to water stress showed signs of divergent natural selection between the Magadi tilapia and its freshwater relative, which shared a most recent common ancestor only about four million years ago. The complete set of genes responsible for urea excretion was identified in the gill transcriptome of A. grahami, making it the only fish species to have a functional ornithine-urea cycle pathway in the gills--a major innovation for increasing nitrogenous waste efficiency.

Air‐breathing fishes

Air‐breathing fishes

Air breathing in Magadi tilapia Alcolapia grahami, under normoxic and hyperoxic conditions, and the association with sunlight and reactive oxygen species

Observations of the Magadi tilapia Alcolapia grahami in hot, highly alkaline Lake Magadi revealed that they air breathe not only during hypoxia, as described previously, but also during normoxia and hyperoxia. Air breathing under these latter conditions occurred within distinct groupings of fish (pods) and involved only a small proportion of the population. Air breathing properties (duration and frequency) were quantified from video footage. Air breathing within the population followed a diel pattern with the maximum extent of pod formation occurring in early afternoon. High levels of reactive oxygen species (ROS) in the water may be an irritant that encourages the air-breathing behaviour. The diel pattern of air breathing in the field and in experiments followed the diel pattern of ROS concentrations in the water which are amongst the highest reported in the literature (maximum daytime values of 2.53 – 8.10 μM H₂O₂). Interlamellar cell masses (ILCM) occurred between the gill lamellae of fish from the lagoon with highest ROS and highest oxygen levels, while fish from a normoxic lagoon with one third the ROS had little or no ILCM. This is the first record of air breathing in a facultative air-breathing fish in hyperoxic conditions and the first record of an ILCM in a cichlid species.

Pronounced genetic differentiation of small, isolated and fragmented tilapia populations inhabiting the Magadi Soda Lake in Kenya

Lake Magadi, an alkaline hypersaline lake in Kenya, is one of the most extreme water bodies known. Although its water temperatures often exceed 40°C, a particular lineage of ‘dwarf’ tilapia, Alcolapia grahami, has evolved remarkable adaptations to survive in this hostile environment. Magadi tilapia exists in small fragmented populations in isolated lagoons within Lake Magadi and its satellite Lake, Little Magadi. In spite of the potential this tilapia holds for understanding evolutionary processes in stressful environments, few genetic studies have focused on this species. We examined the genetic diversity and spatial genetic relationships of Magadi tilapia populations using microsatellite and mitochondrial markers. High levels of genetic variation were found to be supporting the hypothesis that A. grahami populations represent remnants of a much larger fish population that inhabited paleo-lake Orolonga. In contrast to previous studies, we found a well-supported genetic structure of A. grahami consisting of three differentiated genetic clusters (a) Little Magadi, (b) Fish Spring Lagoon and (c) Rest of Magadi. Given the importance of this species to the Magadi ecosystem and its potential evolutionary significance, the three genetic clusters should be considered as separate gene pools and conservation strategies aimed at protecting the species based on these clusters are recommended.

Morphological evaluation of spermatogenesis in Lake Magadi tilapia (Alcolapia grahami): A fish living on the edge

Spermatogenesis in Lake Magadi tilapia (Alcolapia grahami), a cichlid fish endemic to the highly alkaline and saline Lake Magadi in Kenya, was evaluated using light and transmission electron microscopy. Spermatogenesis, typified by its three major phases (spermatocytogenesis, meiosis and spermiogenesis), was demonstrated by the presence of maturational spermatogenic cells namely spermatogonia, spermatocytes, spermatids and spermatozoa. Primary spermatogonia, the largest of all the germ cells, underwent a series of mitotic divisions producing primary spermatocytes, which then entered two consecutive meiotic divisions to produce secondary spermatocytes and spermatids. Spermatids, in turn, passed through three structurally distinct developmental stages typical of type-I spermiogenesis to yield typical primitive anacrosomal spermatozoa of the externally fertilizing type (aquasperm). The spermatozoon of this fish exhibited a spheroidal head with the nucleus containing highly electron-dense chromatin globules, a midpiece containing ten ovoid mitochondria arranged in two rows and a flagellum formed by the typical 9 + 2 microtubule axoneme. In addition, the midpiece, with no cytoplasmic sheath, appeared to end blindly distally in a lobe-like pattern around the flagellum; a feature that was unique and considered adaptive for the spermatozoon of this species to the harsh external environment. These observations show that the testis of A. grahami often undergoes active spermatogenesis despite the harsh environmental conditions to which it is exposed on a daily basis within the lake. Further, the spermiogenic features and spermatozoal ultrastructure appear to be characteristic of Cichlidae and, therefore, may be of phylogenetic significance.

Rh proteins and NH4+-activated Na+-ATPase in the Magadi tilapia (Alcolapia grahami), a 100% ureotelic teleost fish

The small cichlid fish Alcolapia grahami lives in Lake Magadi, Kenya, one of the most extreme aquatic environments on Earth (pH ~10, carbonate alkalinity ~300 mequiv l(-1)). The Magadi tilapia is the only 100% ureotelic teleost; it normally excretes no ammonia. This is interpreted as an evolutionary adaptation to overcome the near impossibility of sustaining an NH3 diffusion gradient across the gills against the high external pH. In standard ammoniotelic teleosts, branchial ammonia excretion is facilitated by Rh glycoproteins, and cortisol plays a role in upregulating these carriers, together with other components of a transport metabolon, so as to actively excrete ammonia during high environmental ammonia (HEA) exposure. In Magadi tilapia, we show that at least three Rh proteins (Rhag, Rhbg and Rhcg2) are expressed at the mRNA level in various tissues, and are recognized in the gills by specific antibodies. During HEA exposure, plasma ammonia levels and urea excretion rates increase markedly, and mRNA expression for the branchial urea transporter mtUT is elevated. Plasma cortisol increases and branchial mRNAs for Rhbg, Rhcg2 and Na(+),K(+)-ATPase are all upregulated. Enzymatic activity of the latter is activated preferentially by NH4(+) (versus K(+)), suggesting it can function as an NH4(+)-transporter. Model calculations suggest that active ammonia excretion against the gradient may become possible through a combination of Rh protein and NH4(+)-activated Na(+)-ATPase function.

Transepithelial potential in the Magadi tilapia, a fish living in extreme alkalinity

We investigated the transepithelial potential (TEP) and its responses to changes in the external medium in Alcolapia grahami, a small cichlid fish living in Lake Magadi, Kenya. Magadi water is extremely alkaline (pH = 9.92) and otherwise unusual: titratable alkalinity (290 mequiv L(-1), i.e. HCO(3) (-) and CO(3) (2-)) rather than Cl(-) (112 mmol L(-1)) represents the major anion matching Na(+) = 356 mmol L(-1), with very low concentrations of Ca(2+) and Mg(2+) (<1 mmol L(-1)). Immediately after fish capture, TEP was +4 mV (inside positive), but stabilized at +7 mV at 10-30 h post-capture when experiments were performed in Magadi water. Transfer to 250% Magadi water increased the TEP to +9.5 mV, and transfer to fresh water and deionized water decreased the TEP to -13 and -28 mV, respectively, effects which were not due to changes in pH or osmolality. The very negative TEP in deionized water was attenuated in a linear fashion by log elevations in [Ca(2+)]. Extreme cold (1 vs. 28°C) reduced the positive TEP in Magadi water by 60%, suggesting blockade of an electrogenic component, but did not alter the negative TEP in dilute solution. When fish were transferred to 350 mmol L(-1) solutions of NaHCO(3), NaCl, NaNO(3), or choline Cl, only the 350 mmol L(-1) NaHCO(3) solution sustained the TEP unchanged at +7 mV; in all others, the TEP fell. Furthermore, after transfer to 50, 10, and 2% dilutions of 350 mmol L(-1) NaHCO(3), the TEPs remained identical to those in comparable dilutions of Magadi water, whereas this did not occur with comparable dilutions of 350 mmol L(-1) NaCl-i.e. the fish behaves electrically as if living in an NaHCO(3) solution equimolar to Magadi water. We conclude that the TEP is largely a Na(+) diffusion potential attenuated by some permeability to anions. In Magadi water, the net electrochemical forces driving Na(+) inwards (+9.9 mV) and Cl(-) outwards (+3.4 mV) are small relative to the strong gradient driving HCO(3) (-) inwards (-82.7 mV). Estimated permeability ratios are P (Cl)/P (Na) = 0.51-0.68 and [Formula: see text] = 0.10-0.33. The low permeability to HCO(3) (-) is unusual, and reflects a unique adaptation to life in extreme alkalinity. Cl(-) is distributed close to Nernst equilibrium in Magadi water, so there is no need for lower P (Cl). The higher P (Na) likely facilitates Na(+) efflux through the paracellular pathway. The positive electrogenic component is probably due to active HCO(3) (-) excretion.

Studies of Some Molecular Properties of the Vacuolar H+-ATPase in Rainbow Trout (Oncorhynchus mykiss)

A cloned cDNA fragment of the rainbow trout gill vacuolar H +-ATPase (H+ VATPase; proton pump) B subunit was used as a probe to examine: i) its interspecific distribution among marine and freshwater species and ii) its expression during a variety of acid-base and ionic disturbances. Northern blots of gill total RNA, performed under conditions of high stringency, revealed cross hybridisation between the trout probe and 11 of 16 species that were examined. Cross hybridisation was not observed in the pacific hagfish (E. stoutit), Lake Magadi tilapia (Oroechromis alcalicus Grahami), bigfin eelpout (Lycodes cortezianus), blackfin poacher (Bathyagonus nigripinnus) or freshwater American eel (Anguilla rostrata). Acute (3 h) exposure of trout to external hypercapnia (PwCO2 = ∼7 mm Hg) was associated with a transient increase after 1 h in gill H +-ATPaSe mRNA levels. Thus, the increase in gill H+-ATPase activity that is known to accompany hypercapnic acidosis in trout may reflect, at least in part, its transcriptional or post- transcriptional regulation. Plasma Cortisol levels were elevated in the hypercapnic fish (45±15 to 83±4 ng mL-1) and because 30 Cortisol was previously implicated as a regulator of H+-ATPase activity, mRNA levels were quantified in fish subjected to chronic Cortisol elevation. An increase in plasma Cortisol concentration from 90±10 (sham implants) to 300±60 ng mL-1 (Cortisol implants) for 4 days was associated with an approximate doubling of gill H+-ATPase steady-state mRNA levels. Exposure of trout for 72 h to ion-poor water caused a persistent reduction in the concentration of gill H+-ATPase steady-state mRNA. The functional significance of this response is unclear but may reflect a reduced rate OfNa+ uptake across the gill. These results are discussed with reference to the physiological role of the branchial H+-ATPase in both acid-base and ionic regulation.

Physiological adaptations of the gut in the Lake Magadi tilapia, Alcolapia grahami, an alkaline- and saline-adapted teleost fish

We describe the gut physiology of the Lake Magadi tilapia ( Alcolapia grahami), specifically those aspects associated with feeding and drinking while living in water of unusually high carbonate alkalinity (titratable base=245 mequiv l −1) and pH (9.85). Drinking of this highly alkaline lake water occurs at rates comparable to or higher than those seen in marine teleosts. Eating and drinking take place throughout the day, although drinking predominates during hours of darkness. The intestine directly intersects the esophagus at the anterior end of the stomach forming a ‘T’, and the pyloric sphincter, which comprises both smooth and striated muscle, is open when the stomach is empty and closed when the stomach is full. This unique configuration (a functional trifurcation) allows imbibed alkaline water to bypass the empty stomach, thereby avoiding a reactive mixing with acidic gastric fluids, and minimizes interference with a full stomach. No titratable base was present in the stomach, where the mean pH was 3.55, but the intestine was progressively more alkaline (foregut 6.96, midgut 7.74, hindgut 8.12, rectum 8.42); base levels in the intestinal fluid were comparable to those in lake water. The gut was highly efficient at absorbing water (76.6%), which accompanied the absorption of Na + (78.5%), titratable base (80.8%), and Cl − (71.8%). The majority of Na +, base and water absorption occurred in the foregut by an apparent Na + plus base co-transport system. Overall, more than 70% of the intestinal flux occurred via Na + plus base co-transport, and less than 30% by Na + plus Cl − co-transport, a very different situation from the processes in the intestine of a typical marine teleost.

Ammonia excretion and urea handling by fish gills: present understanding and future research challenges.

In fresh water fishes, ammonia is excreted across the branchial epithelium via passive NH(3) diffusion. This NH(3) is subsequently trapped as NH(4)(+) in an acidic unstirred boundary layer lying next to the gill, which maintains the blood-to-gill water NH(3) partial pressure gradient. Whole animal, in situ, ultrastructural and molecular approaches suggest that boundary layer acidification results from the hydration of CO(2) in the expired gill water, and to a lesser extent H(+) excretion mediated by apical H(+)-ATPases. Boundary layer acidification is insignificant in highly buffered sea water, where ammonia excretion proceeds via NH(3) diffusion, as well as passive NH(4)(+) diffusion due to the greater ionic permeability of marine fish gills. Although Na(+)/H(+) exchangers (NHE) have been isolated in marine fish gills, possible Na(+)/NH(4)(+) exchange via these proteins awaits evaluation using modern electrophysiological and molecular techniques. Although urea excretion (J(Urea)) was thought to be via passive diffusion, it is now clear that branchial urea handling requires specialized urea transporters. Four urea transporters have been cloned in fishes, including the shark kidney urea transporter (shUT), which is a facilitated urea transporter similar to the mammalian renal UT-A2 transporter. Another urea transporter, characterized but not yet cloned, is the basolateral, Na(+) dependent urea antiporter of the dogfish gill, which is essential for urea retention in ureosmotic elasmobranchs. In ureotelic teleosts such as the Lake Magadi tilapia and the gulf toadfish, the cloned mtUT and tUT are facilitated urea transporters involved in J(Urea). A basolateral urea transporter recently cloned from the gill of the Japanese eel (eUT) may actually be important for urea retention during salt water acclimation. A multi-faceted approach, incorporating whole animal, histological, biochemical, pharmacological, and molecular techniques is required to learn more about the location, mechanism of action, and functional significance of urea transporters in fishes.

Ionoregulatory strategies and the role of urea in the Magadi tilapia (Alcolapia grahami)

The unique ureotelic tilapia Alcolapia grahami lives in the highly alkaline and saline waters of Lake Magadi, Kenya (pH ~10.0, alkalinity ~380 mmol·L–1, Na+~350 mmol·L–1, Cl–~110 mmol·L–1, osmolality ~580 mosmol·kg–1). In 100% lake water, the Magadi tilapia maintained plasma Na+, Cl–, and osmolality at levels typical of marine teleosts and drank the medium at 8.01 ± 1.29 mL·kg–1·h–1. Gill chloride cells were predominantly of the sea water type (recessed, with apical pits) but a few freshwater-type chloride cells (surficial, with flat apical exposure) were also present. Whole-body Na+and Cl–concentrations were relatively high and exhibited larger relative changes in response to salinity transfers than did plasma ions. All fish succumbed upon acute transfer to 1% lake water, but tolerated acute transfer to 10% lake water well, and gradual long-term acclimation to both 10 and 1% lake water without change in plasma cortisol. Plasma osmolytes were here maintained at levels typical of freshwater teleosts. Curiously, drinking continued at the same rate in fish adapted to 1% lake water, but chloride cells were now exclusively of the freshwater type. Significant mortality and elevated cortisol occurred after acute transfer to 200% lake water. However, the fish survived well during gradual adaptation to 200% lake water, although plasma cortisol remained chronically elevated. Urea levels accounted for only 2–3% of internal osmolality in 100% lake water but responded to a greater extent than plasma ions during exposure to 10 and 200% lake water, decreasing by 28–42% in the former and increasing by over 500% in the latter relative to simultaneous-control values. Urea thereby played a small but significant role (up to 8% of internal osmolality) in osmoregulation.

Physiological and molecular characterization of urea transport by the gills of the Lake Magadi tilapia (Alcolapia grahami).

The Lake Magadi tilapia (Alcolapia grahami) is an unusual fish, excreting all its nitrogenous waste as urea because of its highly alkaline and buffered aquatic habitat. Here, using both physiological and molecular studies, we describe the mechanism of branchial urea excretion in this species. In vivo, repeated short-interval sampling revealed that urea excretion is continuous. The computed urea permeability of A. grahami gill is 4.74x10(-)(5)+/-0.38x10(-)(5 )cm s(-)(1) (mean +/- s.e.m., N=11), some 10 times higher than passive permeability through a lipid bilayer and some five times higher than that of even the most urea-permeable teleosts studied to date (e.g. the gulf toadfish). Transport of urea was bidirectional, as demonstrated by experiments in which external [urea] was elevated. Furthermore, urea transport was inhibited by classic inhibitors of mammalian and piscine urea transporters in the order thiourea>N-methylurea>acetamide. A 1700 base pair cDNA for a putative Magadi tilapia urea transporter (mtUT) was cloned, sequenced and found to display high homology with urea transporters from mammals, amphibians and other fishes. When cRNA transcribed from mtUT cDNA was injected into Xenopus laevis oocytes, phloretin-inhibitable urea uptake was enhanced 3.4-fold relative to water-injected controls. Northern analysis of gill, red blood cells, liver, muscle and brain using a portion of mtUT as a probe revealed that gill is the only tissue in which mtUT RNA is expressed. Magadi tilapia gill pavement cells exhibited a trafficking of dense-cored vesicles between the well-developed Golgi cisternae and the apical membrane. The absence of this trafficking and the poor development of the Golgi system in a non-ureotelic relative (Oreochromis niloticus) suggest that vesicle trafficking could be related to urea excretion in Alcolapia grahami. Taken together, the above findings suggest that the gills of this alkaline-lake-adapted species excrete urea constitutively via the specific facilitated urea transporter mtUT.

Teleost life in extremity adaptation, evolution and genetics of the lake Magadi tilapia

Teleost life in extremity adaptation, evolution and genetics of the lake Magadi tilapia