Biological Journal ojlhe Linnean Society (1996), 58: 371-383. With 4 figures
Habitat and phylogeny influence salinity
discrimination in crocodilians: implications for
osmoregulatory physiology and historical
biogeography
KATE JACKSON*, DAVID G. BUTLER AND DANIEL R. BROOKS
Department
I$<OO~OQ,
University of Toronto, Toronto, Ontario, Canada, M5S IAI
Received 3 M v 1995, acceptedfor publication 1% September 1995
Crocodylids are better adapted than alligatorids, through a suite of morphological specializations, for life
in hyperosmotic environments. The presence of such specializations even in freshwater crocodylids has
been interpreted as evidence for a marine phase in crocodylid evolution, consistent with the transosceanic migration hypothesis of crocodilian biogeography. The ability to discriminate fresh water from
hyperosmotic sea water, and to avoid drinking the latter, is known to be an important osmoregulatory
mechanism for estuarine crocodylids. This study was undertaken to determine whether the ability to
discriminate between hyper- and hypo-osmotic salinities is determined by habitat, as it is in other
normally freshwater reptiles, or whether, like morphological adaptations associated with estuarine lie,
it has a phylogenetic basis. Two species of freshwater alligatorid were found to drink fresh water and
hyperosmotic sea water indiscriminately, while an estuarine population of a normally freshwater
alligatorid species drank only fresh water. This indicated that salinity discrimination is determined at
least in part by habitat. However, all three crocodylid species tested drank fresh water but not
hyperosmotic sea water, suggesting that, in crocodilians, the ability to distinguish between fresh water
and sea water is influenced by phylogeny as well as by habitat. The implications of this result are
discussed in the context of two alternate hypotheses for the historical biogeography of the Crocodilia.
01996 l'hc Linncan Society of London
ADDITIONAL KEY W O R D S -C r o c d i a - Eusuchia
marine adaptation - biogeography.
-
osmoregulation
-
evolution - physiology -
CONTENTS
Introduction . . . . . . . . . . . . . . . . . . . .
Lack of salinity discrimination in Caiman crocodilur . . . . . . .
Material and methods . . . . . . . . . . . . . . .
Results and discussion . . . . . . . . . . . . . . .
Influence of habitat on salinity discrimination in Alhgafor rnksirsiphis
Material and methods . . . . . . . . . . . . . . .
Results and discussion . . . . . . . . . . . . . . .
Influence ofphylogeny on salinity discrimination in crocodilians . .
Material and methods . . . . . . . . . . . . . . .
Results and discussion . . . . . . . . . . . . . . .
General discussion . . . . . . . . . . . . . . . . . .
Acknowledgements . . . . . . . . . . . . . . . . . .
References . . . . . . . . . . . . . . . . . . . . .
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*Current address: Museum of Comparative Zoology, Harvard University, Cambridge, MA 02 138, USA
0024-4066/96/080371
+ 13 $18.00/0
37 1
0 I996 The Linnean Society of London
372
K. JACKSON n A L .
INTRODUCTION
Reptiles inhabiting marine and estuarine environments face the challenge of
maintaining a constant plasma osmolality while living in a hyperosmotic medium.
They employ a variety of osmoregulatory strategies, including behavioural
modifications, such as avoiding drinking hyperosmotic sea water, and morphological
adaptations associated with osmoregulation such as salt-secretingglands and reduced
integumental permeability.
Dunson (1980, 1985, 1986) and Mazzotti & Dunson (1989) studied the
physiological basis of putative marine adaptations in reptiles using snakes and turtles
as models. Dunson & Mazzotti (1989)recognized a number of conditions which they
interpreted as a gradient of evolutionary specializations. In their model, the
presumed plesiomorphic condition is represented by aquatic (freshwater)snakes and
turtles (e.g. Niodia, Chebdra). The second stage is represented by estuarine
populations of the same species, which differ from freshwater populations in being
able to tolerate limited exposure to hyperosmotic sea water by selectively drinking
only hypo-osmotic water (Dunson, 1980, 1986). Reptilian nephrons lack loops of
Henle, and are therefore not capable of producing a hypertonic urine. Like marine
birds, therefore, some reptiles possess salt-secreting glands capable of secreting a
hypertonic NaCl solution extrarenally. The third stage of marine adaptation is
marked by the appearance of salt-secreting glands of low secretory capacity (volume
and concentrations) (e.g. Acrochordus, Malachys), which allow a constant plasma
osmolality to be maintained when used in conjunction with selective drinking of only
hypo-osmotic water punson, 1985).The fourth and final stage is represented by the
truly marine reptiles, the sea snakes (Hydrophiinae, Laticaudinae), sea turtles
(Cheloniidae, Dermochelyidae), and the marine iguana (Ambbrhychus). In these
species the salt-secreting glands are well developed and allow the maintenance of a
constant plasma osmolality even when hyperosmotic sea water and osmoconforming prey such as jellyfish are ingested. Salt glands have been independently
derived several times in reptiles. Salt glands are sub-lingual in snakes, lachrymal in
turtles, and nasal in lizards (Peaker & Linzell, 1975). Crocodylids but not alligatorids
have lingual salt glands of low secretory capacity (Taplin & Grigg, 1981;Taplin et al.,
1982).
Crocodilians include species in which some populations inhabit brackish or
estuarine habitats. Crocodylus acutus and C. porosus (Crocodylidae) are primarily
estuarine, while several other normally freshwater crocodylid species (e.g. C.
cahphractus, C.johmtoni, C. moreletti, C. nibticus, and C. palustris) have some estuarine
populations. Estuarine populations of Allkator rnissk-apiensis, Caiman crocodilus and Ca.
latirostni (Alligatoridae)are also known to exist (reviewed by Taplin, 1988). Estuarine
crocodylids are more common than estuarine alligatorids, perhaps because they
possess morphological specializations, independent of habitat, which confer an
advantage to them over alligatorids in adapting to hyperosmotic conditions. The
lingual glands of crocodylids, for example, secrete an hyperosmotic solution of NaCl
in response to stimulation with methacholine chloride, while those of alligatorids
secrete an iso-osmotic solution (Taplin & Grigg, 1981;Taplin, Grigg & Beard, 1985).
Additionally, crocodylids possess a heavily keratinized buccal epithelium, so that
osmotic water loss under hyperosmotic conditions is presumably less than for
alligatorids, which lack protection for the highly permeable buccal epithelium
(Taplin & Grigg, 1989).
PHYLOGENY AND SALINITY DISCRIMINATION IN CROCODILIANS
373
Opportunistic drinking of fresh water or hypo-osmotic sea water is thought to be
an important mechanism allowing crocodilians inhabiting estuarine areas to
maintain a constant plasma osmolality in a fluctuating hyperosmotic environment.
The estuarine crocodylids, C. acutus and C. porosus, as well as estuarine populations of
C. johnstoni, will not drink hyperosmotic sea water even when severely dehydrated
(Mazzotti & Dunson, 1984; Taplin, 1984, 1988; Taplin, Grigg & Beard, 1993).
Moreover, these species can distinguish precisely between brackish water of hyperand hypo-osmotic salinities (Mazzotti & Dunson, 1984; Taplin, 1984).
No experimental data exist on the ability of alligatorids to selectively avoid
drinking hyperosmotic sea water. Lauren (1985) found that juvenile A. mksirszpiensir
died after three weeks of continuous exposure to salinities of 15ppt or greater, and
Mazzotti & Dunson (1984) observed that the mortality rate for A. rnk.&sipiensir was
higher than that of C. acutus when both species were exposed to the same regime of
alternating hyper- and hypo-osmotic salinities. Bentley and Schmidt-Nielsen (1965)
observed in the course of an experiment on skin permeability in the freshwater
alligatorid Ca. crocodilus that 20% of their experimental animals died 18-24 hours
after being placed in a 33 ppt NaCl solution, apparently as a result of drinking the
medium.
The following study was undertaken to obtain experimental data on drinking of
hyperosmotic sea water by a freshwater alligatorid, Ca. crocodilus, for comparison with
existing data from estuarine crocodylids, and to obtain data from representative
alligatorid and crocodylid species from freshwater and estuarine habitats, to
determine whether the reported difference in capacity for salinity discrimination
represented (1) a difference between freshwater and estuarine populations, similar to
those observed in other normally freshwater reptiles in which some populations are
estuarine, or (2) a difference between crocodylids and alligatorids analogous to
morphological differences associated with marine adaptation between these two
families.
LACK OF SALINITY DISCRIMINATION IN C4LkU.N CROCODILUS
Mah'al and methods
Captive-raised juvenile (100-300 g) Caiman crocodilus (n = 9) were imported from
Venezuela, housed in a tank of dechlorinated tap water with a land/water choice, at
30'C (air temperature), and fed live minnows ad libitum. Blood was sampled in order
to determine which sea water dilutions were hyper- and hypo-osmotic to caiman
plasma. Blood samples of 0.2mL were withdrawn from the caudal vein and
centrifuged (4000 X g)at 5OC for 10 min. Plasma was collected from caimans before
and after dehydration (10% of initial body mass), and stored at -80' until analysis.
Na+ and K + concentrations were measured by flame photometry (Instrument
Laboratories, Model 943), and osmolality by freezing point depression (Advanced
Instruments micro-osmometer, Model 3MO).
Two experiments were conducted to determine whether dehydrated caimans
would drink sea water. The first experiment measured the amount of water ingested
by dehydrated caimans at different sea water dilutions. Sea water solutions in both
experiments were made using Instant Ocean sea salt (Aquarium Systems, Mentor,
Ohio 44060, USA). Six unfed caimans were selected from a group of nine and
K. JACKSON ETA,
374
weighed. They were then dehydrated in a current of air (3OoC), re-weighed and
placed in a 50-gdon plastic tank containing fresh water 10 cm in depth. After 15
min, they were re-weighed to determine, by difference, the amount of water ingested.
This procedure was repeated for 5, 10, 15, 20, and 30ppt sea water to determine
whether or not increases in salinity would affect drinking by dehydrated caimans.
Integumentary osmotic uptake of water in this species is known to be in the order of
1.1 pL cmp2hr-' (Bentley & Schmidt-Nielsen, 1965). In a 200 g animal immersed in
fresh water for 15 min this represents a gain of only 95pL (Surface area
= 11.7Ma~sO.~~;
Dunson & Mazzotti, 1988), so this was not an important factor in
measuring drinking by increased mass. Between experiments, the test caimans were
returned to the fresh water holding tank for a period of at least 7 days to allow time
for rehydration. The second experiment was used to determine the amount of water
ingested by dehydrated caimans during a longer period of exposure to hyperosmotic
sea water. Body mass of nine unfed caimans was measured before and after
dehydration, following a 15-min exposure, and finally, a 75-min exposure to 20 ppt
sea water.
Results and discussion
Following dehydration there was a significant increase in plasma Na concentration (15%)and osmolality (13%)but no significant change in plasma K + (Table 1).
Blood haematocrit increased by 42%, indicating that the vascular compartment had
become smaller in response to dehydration. Caiman plasma was found to be
hyperosmotic to 10ppt sea water and hypo-osmotic to 15ppt sea water, both before
and after dehydration.
At all sea water concentrations tested, dehydrated caimans drank a significant
volume of water, regaining 2-5% of their initial body mass (Fig. 1). No overall
statistical difference was found in the amount drunk of the different sea water
dilutions (ANOVA, P > 0.05), and a Scheffe test showed no significant difference in
amount drunk between any two dilutions (P> 0.05). When the 15-minute
observation period was increased to 75 min (Fig. 2) caimans transferred to 20 ppt sea
water continued to drink water and to increase in mass so that the final mass was
99.6% of the original. This indicated that the caimans continued to drink
hyperosmotic sea water when given access to it for more that 15 min.
These results (Figs 1, 2) show that caimans drink water of all salinities tested when
dehydrated by 10% of body mass. That the caimans regained 2-5% of their initial
body mass during 15 min of exposure to water over the range of salinities, is
+
TABLE1. Osmolality, Na+ and K+ concentrations, and blood haematocrit in caiman plasma
before and after dehydration by 10% of body mass. *Indicates a significant increase (P<0.05,
paired &test,corrected for multiple comparisons)
Plasma
Before dehydration
After dehydration
% change
n
Na+ (mM)
K+ (mM)
Osmolality
( m o m kg')
Blood
haematocrit (%)
6
6
145.7i1.4
167.1k1.6
+15%*
4.10i0.25
4.38i0.30
no change
28921.6
327i2.4
+13%*
16.2k0.7
23.0i1.3
+42%*
PHYLOGENY AND SAIJNITY DISCRIMINATION IN CROCODILIANS
h
T
5-
"
375
0
5
10
15
20
30
Salinity (ppt)
Figure 1. Mass of water ingested by Caiman crocodilus (n = 6) following dehydration. Values are
mean f SEM. Differences in amount drunk at different salinities are not significant (P> 0.05, ANOVA,
Scheffe test).
interpreted as an indication of drinking, as opposed to integumental uptake by
dehydrated animals, because (1) the exposure period was short (1 5 min) in order to
minimize the possible effects of osmotic uptake, and (2) the animals increased in mass
during exposure to water of hyperosmotic salinities in which the osmotic gradient
should have led to a decrease rather than an increase in mass if diffusion across the
integument had been an important factor.
Figure 2. Body mass in grams (mean f SEM) of Ca. uocodihs (n = 9), and mass of 20ppt sea water
ingested as a percentage of initial body mass (mean f SEM): initial mass (prior to dehydration),
dehydrated mass (following dehydration), mass after 15 min exposure to sea water, and after 75 min
exposure. * Indicates significant increase in mass (P< 0.001, rmANOVA).
water ingested; (m) body
mass.
376
K. JACKSON E T A .
Results from the first experiment (Fig. 1) indicated a slight, though statistically
nonsignificant, decrease in the amount drunk at the two highest salinities (20ppt and
30ppt). It seemed possible that this result might indicate that although the caimans
were drinking at all salinities, they stopped drinking after an initial mouthful at
strongly hyperosmotic salinities. However, when the experiment was repeated using
20ppt sea water, this time weighing the caimans twice and extending the period of
exposure to 75 min, the mean increase in mass after 15 min was 6.5% of initial body
mass (Fig. 2). This is the largest mass increase recorded at any salinity, supporting the
view that the amount drunk does not decrease at the highest salinities. When reweighed after 75 min, the caimans were found to have further increased their mass
(Fig. 2). This result is evidence against the idea that the caimans stopped drinking
after an initial mouthful at hyperosmotic salinities.
These data provide quantitative evidence of the inability of the freshwater
alligatorid, Ca. crocodiZus to osmoregulate by selectively drinking only water of hypoosmotic salinities. These results are strikingly different from those obtained in studies
of the estuarine crocodylids, C. acutus (Mazzotti & Dunson, 1984) and C. porosus
(Taplin, 1984). These species will not drink sea water of hyperosmotic salinities and
are capable of distinguishing precisely between hyper- and hypo-osmotic sea water.
What our experimental data for a freshwater alligatorid, and those of other
researchers from estuarine crocodylids fail to reveal, however, is whether this
difference in capacity for behavioural osmoregulation represents: ( 1) a phylogenetic
constraint on the capacity of alligatorids to adapt to estuarine conditions (e.g.
inability of alligatorids to taste salt), (2) a behavioural modification within the
potential to evolve independently in any crocodilian population exposed to
fluctuating salinities, or (3) a behaviour learned by individual crocodilians exposed to
fluctuating salinities (i.e. without a genetic basis).
INEUENCE OF HABITAT ON SALINITY DISCRIMINATION IN ALLIGATOR MISSISSIPI~SIS
Matmial and methodr
This experiment assessed the effect of habitat in determining the capacity for
salinity discrimination in crocodilians, by comparing Alligator rnissiripimk from
freshwater and estuarine populations. Experiments on freshwater A. rnississipimis
were performed on captive-bred hatchlings (52-68g) (n = 20) at the St. Augustine
Alligator Farm in Florida. Estuarine A. rnissiripiensis juveniles (248-372g) (n = 3)
were collected from a freshwater pond on Sapelo Island, one of a string of barrier
islands off the coast of Georgia. Sapelo Island is approximately 20 km long by 6 km
wide and is separated from the shore by a salt marsh 10km wide, although it formed
part of the mainland as recently as 5-10000 years ago (Martof, 1963). Alligators
from this population live in the freshwater ponds and salt marshes of the island.
Alligators ranging in size from large juveniles ( > 1 kg) to adults have been observed
on the beach facing the Atlantic Ocean, and large individuals are often seen several
kilometres from shore. Captured alligators were housed temporarily in fibreglass
tanks (approx. 60cm3), one alligator per tank, in an outbuilding where they were
exposed to outdoor temperatures but protected from rain and direct sunlight.
In preparation for the experiment, alligators were dehydrated by 10% of initial
body mass, to stimulate thirst, by keeping them out of the water for 24-40 hours.
PHnOGENY AND SALINITY DISCRIMINATION IN CROCODILIANS
377
Once dehydrated, the animals were weighed and transferred to a tank containing
either fresh water or 30 ppt sea water 10cm in depth. After 10-1 5 min they were
removed from the water, blotted dry, and reweighed in order to determine, by
difference, the amount of water ingested. For the freshwater alligators, in which the
sample size was large, the animals were separated into two groups of ten. One group
was exposed to sea water and the other to fresh water. For the estuarine alligators,
the sample size was much smaller. AU the dehydrated animals were therefore
exposed initially to 30 ppt sea water and then, if they did not drink, transferred to
fresh water, since response (drinkingversus not drinking) to sea water would be more
informative than response to fresh water. Moreover, animals which did not drink
30ppt sea water in the course of 15 min exposure were still dehydrated by 10% of
initial body mass, and could therefore be transferred to fresh water afterward and
used to test response to fresh water. Sea water solutions were prepared as described
above for Ca. crocodilus.
Results and discussion
Freshwater A. rnksksipiensk increased significantly in mass following exposure to
fresh water and to 30ppt sea water (t-test, P < 0.001) (Tables 2, 3). There was no
significant difference in amount of water drunk in fresh water versus sea water (t-test,
P > 0.05). By contrast, the estuarine A. rnksissajhnsis increased in mass significantly
more in fresh water than in sea water (paired t-test, P < 0.025) (Tables 2,3). This
result supports the hypothesis that the ability to selectively drink only water of hypoosmotic salinities is determined at least in part by habitat. Grigg (pers. comm.) has
observed that estuarine populations of Ca. lahosbis also avoid drinking hyperosmotic
sea water.
Other estuarine populations of normally freshwater reptiles follow the same
TABLE
2. Drinking of fresh water by dehydrated crocodilians. Values are mean +/- SEM.
*Indicatesa significant increase in mass
Species
Ca. mcodilus
A . mississipensis
A. rnississipiensis
c. pmsus
C. siamiensis
0. t&z.spLS
Origin
n
6
freshwater, captive
20
freshwater, captive
estuarine population 3
6
freshwater, captive
20
freshwater, captive
3
freshwater, captive
Dehydrated mass (9)
Water ingested (9)
Drinking?
116.5k8.01
57.7k0.46
273.0k18
140.5k7.07
56.6k0.5
65.5k1.5
4.7*0.71*
1.0*0.22*
19.7*3.3*
2.9*0.48*
1.75*0.28*
1.83*0.6*
Yes
Yes
Yes
Yes
Yes
Yes
TABLE
3. Drinking of 30 ppt sea water by dehydrated crocodilians. Values are mean +/- SEM.
*Indicatesa significant increase in mass
Species
Ca. modilus
A . mississipiensis
A. mississipamis
c. p m s w
C. siamiensis
0. tetrosprc
Origin
n
freshwater, captive
6
20
freshwater, captive
estuarine population 3
6
freshwater, captive
20
freshwater, captive
3
freshwater, captive
Dehydrated mass (g)
Water ingested (g)
Drinking?
154.8k11.5
56.1k0.34
273.0k18
140.5k7.07
58.3i0.46
65.5i1.5
4.0*0.93*
1.1*0.13*
0.3k0.33
0.49k0.17
O.bO.25
0.17Al.17
Yes
Yes
No
No
No
No
K. JACKSON E'TAL.
378
pattern. In the turtle, Chebdra, and the aquatic snake, Niodia, individuals from
freshwater populations drink all salinities, whereas those from estuarine populations
selectively avoid drinking hyperosmotic sea water (Dunson, 1980, 1986).
INFLUENCE OF PHYLOGENY ON SALINITY DISCRIMINATION IN CROCODILIANS
Material and methods
In order to evaluate the possible role of phylogeny in determining the capacity for
salinity discrimination in crocodilians, an attempt was made to obtain salinity
discrimination data from a large number of crocodilian taxa so as to reveal
phylogenetic patterns. Experiments were therefore undertaken to test for salinity
preference in an additional three crocodylid species, C. porosus (Australasian), C.
siamimis (Asian), and Osteolaemus tetraspis (African). Although the animals used were
captive-bred, freshwater-raised hatchlings, the wild population from which their
ancestors originated is not known. However, C. siamiemis is known only from
freshwater habitats (Taplin, 1988). Data from Osteolaemus were especially interesting
from a phylogenetic perspective, since this genus is the sister group of Crocodylus (Fig.
3). Although it is known that estuarine C. porosus and C. acutus hatchlings are able to
distinguish precisely between hyper- and hypo-osmotic salinities (Taplin, 1984;
Mazzotti & Dunson, 1984), we included C. porosus in our study and used captiveborn, freshwater-raised hatchlings which had never previously been exposed to sea
water, so as to rule out the possibility that avoidance of drinking hyperosmotic sea
(freshwater)
\
(estuarine)
\
/
Figure 3. The phylogenetic distribution of salinity preference among crocodilian species examined. (F/S)
indicates animals which drank both fresh water and sea water, and (F/-) indicates those which drank fresh
water but not sea water.
PHYLOGENY AND SALINITY DISCRIMINATION IN CROCODTLIANS
379
water is a behaviour learned by individual crocodilians in response to exposure to sea
water.
Experiments on C. siamiensiE (n = 20), and 0. tetras@ (n = 3) were carried out at
the St. Augustine Alligator Farm, using captive-born, freshwater-raised hatchlings
(61-78 g, 0. tetraspts; 54-69 g, C. siamiasis. Freshwater-raised C. porosus hatchlings
(12 7- 1 7 1 g) (n = 7) were obtained from the Long Kuan Hung Crocodile Farm in
Singapore, and experiments were performed under laboratory conditions at the
University of Toronto. The experimental procedure was the same as that described
above for the estuarine and freshwater alligatorids. For C. s i u m k i s , in which the
sample size was relatively large (n = 20), the animals were divided into two groups
of ten. One group was exposed to fresh water and the other to 30 ppt sea water. The
sample sizes of 0. tetrapis (n = 3) and C. porosus (n = 6 ) were smaller. Animals were
therefore first exposed to 30 ppt sea water and then, if they did not drink, transferred
to fresh water, as described above for estuarine A. misksipiensis.
Results and discussion
All three crocodylid species drank fresh water but not 30 ppt sea water (Tables 2,
3). C. porosus and 0. tetraspis increased in mass significantly more in fresh water than
in sea water (paired t-tested; P C 0.005 C. porosus, P < 0.05 0. tetrmpis). The mass
increase in the freshwater group of C. s i a m k i s was significantly greater than that for
the sea water group (t-test, P < 0.005).
The comparison of freshwater and estuarine populations of A. m i s k i p h i s
indicated that for alligatorids at least, the capacity to discriminate between fresh
water and hyper-osmotic sea water, and to avoid drinking the latter, is an adaptation
found only in populations inhabiting areas where they are exposed to sea water.
However, the results of the experiments on the freshwater crocodylids, C.siamiensis
and 0. tetraspis, and on freshwater-raised hatchlings of the estuarine species, C.
porosus, suggest that phylogeny is also involved. All crocodylids, whether freshwater
or estuarine and with or without previous experience of hyperosmotic sea water,
discriminate between fresh water and hyperosmotic sea water and will not drink the
latter. Although previous studies have not tested freshwater crocodylids, it has
previously been shown that estuarine C. acutus (Mazzotti & Dunson, 1984) as well as
estuarine populations of the normally freshwater C. johnstoni (Taplin et al., 1993)
selectively avoid drinking hyperosmotic sea water.
GENERAL DISCUSSION
All crocodylid species, whether estuarine or freshwater, drank only fresh water.
Among the alligatorids, however, only those from an estuarine population
distinguished between salinities. These results are presented in Figure 3 superimposed on a phylogenetic tree depicting the evolutionary relationships of the taxa
involved (Norell,j d e Benton & Clark, 1988). In crocodilians, therefore, the capacity
for salinity discrimination has a strong phylogenetic component, analogous to
morphological adaptations associated with estuarine life, such as lingual saltsecreting glands and a heavily keratinized buccal epithelium, which are present in
crocodylids and absent from alligatorids. A search for the physiological mechanism
380
K.JACKSON ET AL
underlying salinity discrimination in crocodylids may shed new light on the
evolutionary significance of this adaptation. It would be useful, for example, to know
whether crocodylids and estuarine alligators use the same mechanism to distinguish
between fresh water and sea water.
Dserences in osmoregulatory physiology between crocodylid and alligatorid
crocodilians are of particular interest in the context of two conflicting hypotheses to
explain the current global distribution of crocodilians. The trans-oceanic migration
hypothesis (Densmore, 1983),explains the distribution of living crocodilian species as
the result of a post-Pliocene trans-oceanic migration on the part of a marine-adapted
ancestral crocodylid. This hypothesis depends on an upper Cretaceous/early
Tertiary divergence between crocodylid and alligatorid lineages, which is more
recent than indicated by the fossil record (Buffetaut, 1979; Sill, 1968; Steel, 1973),
but supported by molecular clock calculations based on haemoglobin sequence data
(Densmore, 1983). Lingual salt-secreting glands have been interpreted as crocodylid
synapomorphies associated with adaptation to marine conditions on the part of a
crocodylid ancestor, and consistent with the trans-oceanic migration hypothesis
(Taplin et al., 1985; Taplin & Grigg, 1989), as has the heavily keratinized (and
presumably less permeable) buccal epithelium of crocodylids and gavialids and the
non-keratinized buccal epithelium of alligatorids. The presence of salt-secreting
glands in freshwater crocodiles is thus considered vestigial.
An alternative hypothesis explains the distribution of living crocodilians as the
result of speciation and upstream migration by a widely distributed, estuarine
ancestral group. Systematic evidence from the co-evolving digenean parasites of
crocodilians indicates a cosmopolitan distribution in the early Cretaceous, which
supports the fossil data suggesting an ancient origin (Brooks, 1979; Brooks &
O’Grady, 1989; Brooks & MacLennan, 1993).However, the parasite data also show
a mixture of freshwater (e.g. digeneans of the family Proterodiplostomidae) and
estuarine-derivedparasite groups (e.g. digeneans of the subfamily Acanthostominae),
and could therefore be consistent with either a freshwater or an estuarine origin.
The phylogenetic significance of salinity discrimination in crocodilians can be
interpreted in four different ways, depending on whether the capacity for salinity
discrimination is assumed to be the plesiomorphic or the derived condition, and on
whether the Crocodilia had a cosmopolitan distribution in the Cretaceous (as
suggested by the parasite data) or diverged more recently (as suggested by the
molecular data). The first possibility (Fig. 4A) is that lack of salinity preference is
plesiomorphic, and that the ability to distinguish between salinities has evolved
independently in alligatorid populations exposed to sea water, and in an ancestral
crocodylid. This interpretation is consistent with both the trans-oceanic migration
hypothesis and the estuarine origin hypothesis depending on the time scale involved.
If the marine adaptation by the ancestral crocodylid is recent (after the separation of
the continents), the trans-oceanic migration hypothesis is supported. However, if the
adaptation at the base of the crocodylid lineage occurred prior to the break-up of
Pangaea, it could also be interpreted as consistent with the hypothesis that modern
crocodilians arose from widely distributed estuarine ancestors and that upstream
migration and freshwater adaptation occurred secondarily.
An alternative interpretation (Fig. 4B), is that the ability to distinguish between
salinities is plesiomorphic and has been secondarily lost in freshwater alligatorids.
This is consistent with the estuarine origin hypothesis, though it still does not
necessarily require a marine or estuarine ancestor. At this point, these trees each
PHYLOGENY AND SALINITY DISCRIMINATION IN CROCODILIANS
38 1
require exactly three steps. Obtaining data from more taxa, especially from Gavialti
and from basal alligatorids (e.g. Pul~osuchus)should help to determine which of these
two trees more accurately reflects evolutionary history.
Although these two alternatives each require an equal number of evolutionary
events, it is not known whether or not the derivation of salinity preference from lack
of salinity preference occurs more readily than the opposite, lack of salinity
preference from salinity preference. It is also not known whether it is possible for a
population to secondarily lose its capacity to avoid drinking hyperosmotic sea water.
In contrast to the reverse situation, in which lack of salinity preference confers an
obvious selective disadvantage on an individual inhabiting an estuarine area, there is
no immediately obvious disadvantage associated with the latent capacity for sea
water avoidance in an individual living in fresh water. The same question applies to
morphological specializations associated with marine adaptation. The presence of
lingual salt-secreting glands in crocodylids, for example, has been interpreted as a
crocodylid synapomorphy consistent with the trans-oceanic migration hypothesis
(Taplin & Grigg, 1989).An alternative interpretation is that the presence of lingual
salt-secreting glands is plesiomorphic, reflecting the estuarine ancestry of the group,
A
B
Figure 4. Alternate hypotheses for the evolutionary significance of salinity preference in crocodilians. In
(A) lack of salinity discrimination is the plesiomorphic condition for crocodilians, in (B)it is the derived
condition in freshwater alligatorids. ( F / S ) indicates drinking of both fresh water and sea water; (F/-)
indicates drinking of fresh water but not sea water.
K. JACKSON E7AL..
382
and that their absence from alligatorids represents a secondary loss associated with
upstream migration and speciation. Once again, it is not known whether loss of saltsecreting glands is a likely result of freshwater adaptation by estuarine reptiles. Part
of the problem is that models of marine adaptation in reptiles are based on studies
of snakes and turtles, and assume that the direction of the evolutionary trend is
always from fresh water to sea water (e.g. Dunson & Mazzotti, 1989). Such models
are therefore not necessarily transferable to crocodilians, for which both historical
biogeographical scenarios (trans-oceanic migration and estuarine origin hypotheses)
involve estuarine to fresh water adaptation at some point. Further study of
physiological adaptation to fresh water by marine or estuarine reptiles may provide
answers to some of these questions and a better basis from which to evaluate
alternative reconstructions of the historical biogeography of the Crocodilia.
ACKNOWLEDGEMENTS
The authors gratefully acknowledge the technical and logistical assistance of the
following people and institutions: Dr J. Alberts, C. Durant, M. Price, and G .
Balckom, University of Georgia Marine Institute, Sapelo Island; Mr Lee Bak Kuan
and Lee Peilin, Long Kuan Hung Crocodile Farm, Singapore; Prof T.J. Lam,
National University of Singapore, Dr K. m e t , St. Augustine Alligator Farm, Florida;
and N. White, University of Toronto. This research was supported by NSERC grant
A-2359 to D.G. Butler.
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