Chapter 2 Squamate Scenarios
2. Squamate Scenarios
The order Squamata, so called because of the dermal covering of overlapping horny scales, comprises the great majority of living reptiles. Although the scaly covering is characteristic of nearly all the members of the order, the most essential differences distinguishing them from other reptiles are, as usual, found in the skeleton, and especially in the skull.
Samuel Williston, 1914
The cocal is a wind battered coconut palm grove on Trinidad’s east coast, inhabited by fishermen, subsistence gardeners, and coconut gatherers. A road parallels the trees allowing easy access to the palm grove, beaches, and adjacent wetlands. Dead palm fronds cover the sandy soil, providing protection to its many life forms and adding a layer of complexity to the physical structure of the habitat. The tiny Streaked Gecko (Gonatodes vittatus) is abundant in the leaf litter and its eggs can often be seen in small crevices on the trunks of the palms. It shares this habitat with an even smaller relative but one rare in the cocal, the Mole’s Gecko (Sphaerodactylus molei). Both these geckos forage for tiny insects. The large, common Green Iguana (Iguana iguana) occasionally climbs the palms to bask, but it is more often seen grazing on vegetation in the nearby forests and orchards. On the landward edge of the beach large, Green-headed Ameivas or Jungle Runners (Ameiva atrigularis) chase the smaller and related Whiptail Lizard (Cnemidophorus lemniscatus). In moist soil under the dead palm fronds, the unusual Black and White Worm Lizard (Amphisbaena fuliginosa) forages slowly for beetle larvae, while the very active Greater Windward Skink (Copeoglossum aurae) hunts insects in and out of the patches of sunlight on the palm grove floor. A few small streams run under the road and form pools just before they drain into the Caribbean. Here, the Banded Rear-fanged Water Snake (Helicops angulatus) hunts frogs and fish after the sun has set. A few kilometers to the west is Nariva Swamp, a wetland inhabited by the largest living boid snake, the highly aquatic Green Anaconda (Eunectes murinus). It exceeds eight meters, weighs more than 100 kg, and gives birth to live young. The slender, meter-long Vine Snakes (Oxybelis aeneus) wait in ambush for geckos and anoles on the branches of small trees and shrubs scattered at the edges of the palm grove and around the gardens.
Despite living quite different lives, all of these reptiles are squamates, and have distinct evolutionary histories that converge on a point in deep time on an ancient continent where they share a common ancestor. Trinidad is less than 5000 km2, and while geographers consider it an island of the West Indies, it is, in fact a fragment of the South American continent. The flora and fauna of Trinidad demonstrate that the island has been connected to mainland South America throughout much of its history. In fact, very few West Indian species actually inhabit the island. Despite Trinidad’s small size, 70 species of squamate reptiles are found here and, on a small scale, they represent the global diversity of squamates.
Today, more than 8200 lizards, amphisbaenas, and snakes compose the order Squamata. Squamates inhabit all continents except Antarctica and, while most occur in the tropics and subtropics, some have adapted to high latitudes and altitudes. They shared a common ancestor because they have in excess of 50 shared derived characteristics, including: scales that periodically shed the outer layer of keratin, discrete Jacobson’s organs or vomeronasal organs for chemical sensing, hemipenes (a divided penis with retractable right and left halves), saccular ovaries (ovaries that contain an open chamber surrounded by many blood vessels), and the numerous skull characters referenced in Williston’s introductory quote. They also share a considerable amount of DNA, powerful evidence they shared common ancestor as well. Squamates together with their sister group, the Rhynchocephalia (beak-heads), form the Lepidosauria, an ancient group of reptiles that pre-dates the dinosaurs. Rhynchocephalia is known from fossils and two living species, the tuataras of the genus Sphenodon. Tuataras are lizard–like in body form, but they have non-replaceable teeth, a relative solid skull and, while males lack a penis, they have paired pockets for storing the sperm transfer organ in the tail’s base like lizards and snakes. The two living species of beak-heads occur on small rocky islands off the coast of New Zealand where they live unusual lives when compared to the squamates.
Life is Plastic, Evolution Tinkers
Life is restricted to the biosphere, a 42 km thick region comprised of the planet’s surface and the area just above and below it. Microorganisms have been found high in the upper atmosphere, in the deep ocean and, in the solid rock of the Earth’s crust. The thin layer of upper atmospheric gases can support ballooning spiders and the occasional bird only for a short period of time. Imagine any place within the biosphere and life is there. Life has adapted to virtually all environments from temperatures and moisture extremes to every imaginable chemical or pH extreme. Life is plastic – that is it is flexible and adaptable.
Evolution tinkers with what exists and produces novel life forms. Genes, developmental processes, and enzymes are modified generation after generation to construct modified chemistry, anatomy, and organisms from ancestral systems. Reshuffling genes and the proteins they produce, controlling them in alternate ways, and changing the time at which they are expressed or suppressed is how evolution spawns new organisms. Changes in body plans, the rearrangement of organs, or the increase, decrease, or elimination of structures results in the rich biodiversity of Earth.
The idea that one gene produces one protein, and one protein produces one action or trait was popular. Today, we know genes multi-task. Genes and the information they contain are continuous from parent to offspring, species to species, genus to genus, and phylum to phylum, and from the members of one kingdom of life to another. As genes move through time they are duplicated, altered, and co-opted for new tasks. Novel structures and novel organisms are generated as cells flip genetic switches and natural selection acts on them.
Snakes are tetrapods (literally “four feet”) despite the fact that all living species lack feet. They are tetrapods because they are descended from lizards, and organisms are organized based upon ancestry not similarity of appearance. Evolution does not start from scratch; it tinkers, using the flexibility of the existing genome to develop novelty. Snakes are novel tetrapods. The technological developments of the past 50 years have allowed science to gain a greater understanding of evolution and how it works. Our knowledge of snakes has greatly increased in the first decade of the 20th century as a result of this technology.
New Tools
Systematic biology has developed two relatively new tools for sorting out relationships between species or groups of species, DNA technology and cladistics. Both tools have increased in popularity over the last 20 years as the technologies they rely on have become less expensive and more readily available. Despite being discovered more than 100 years ago, it is only within the past 50 years that DNA’s importance has become well understood. And more recent advances in computer and molecular biological technologies now allow science to read and apply the information in DNA.
All animals rely on DNA (deoxyribonucleic acid) to pass on the instructions for making new offspring. Forensic science crime dramas and science fiction have brought attention to DNA so that virtually all people in the developed world have heard about the molecule. The chemical messages in DNA are the instructions on how to make a protein, or a message about regulating another piece of DNA. Proteins make up about 70% of the dry weight of cells, and are complex folded chains of smaller molecules called amino acids. Proteins control much of what happens in cells as well as providing structure. A unit of DNA that codes for making a protein is a gene, and humans have about 25,000 genes.
The DNA code is made up of four base units known as nucleotides that are symbolized with the letters ATCG. Unit A always pairs with T, and unit C always pairs with G. These small molecules are in a long double chain twisted into the elegant double helix shaped macromolecule―DNA. The code for how to make a protein or regulate another gene is held in the unique sequence of the nucleotides, just as the meaning of a word lies within the arrangement of letters and the meaning of a sentence lies within the arrangements of words on a page. A sequence of three nucleotides codes for a sequence of messenger RNA units, a second information molecule that in turn codes for one of 20 amino acids that compose a protein or part of a protein in living organisms.
Cells have DNA in two locations. Most DNA is found in the nucleus where it is organized into chromosomes and often notated as cDNA. A smaller piece of DNA occurs within the mitochondria (mtDNA). It is the mtDNA that has been widely used in comparing species and groups of species. The usefulness of mtDNA is due to two attributes; it is always inherited from the maternal parent and it mutates rapidly. This means that it does not undergo the recombination events that occur in cDNA, it is present in multiple copies that are readily accessible to researchers, and it can be used to test relatively recent events. On the other hand, cDNA is more conservative and mutates much slower, making nuclear genes more useful for understanding more ancient events.
The sequence of the A-T-C-G nucleotides in genes can be used to compare species. The information makes it possible to determine how closely two species are related, just as forensic science uses the sequence to determine who is related at a natural disaster or crime scene. When genes are sequenced for different species the information is stored in GenBank in the USA, the European Molecular Biology Laboratory in Germany, or The DNA Data Bank of Japan. These data bases are available on the Internet and can be accessed by the general public. Anyone can learn to obtain the sequences for any gene for any species that has been researched. Currently, these databases catalog more than 100 billion bases from more than 165,000 species, a number that grows daily as scientists around the world continue sequencing DNA, including some squamates. Vertebrates are thought to have a similar number of genes. We know that genome sizes vary considerably, and therefore some species have increased or decreased the number of repetitive segments, altered the intron sizes, or modified other parts of the non-coding DNA. Genome size can be measured by total weight of the chromosomes, expressed in picograms (pg) which is one trillionth of a gram. Using this method, the Marbled Lung Fish (Protopterus aethiopicus) has a genome that weighs 132.83 pg, the heaviest vertebrate genome known. And on the other end of the range, with the lightest known vertebrate genome a full 380 times smaller, is the Green Puffer Fish (Terraodon fluiatilis) at 0.35 pg. Genomes can also be measured using the number of base pairs of nucleotides, expressed in billions of base pairs or gigabases (gb). Among reptiles, the largest average genome size is found in crocodilians with ~3 gb while the smallest average genomes are in birds with 1.4 gb. The reptile with the largest number of base pairs is the tuatara with 5 gb.
Chris Organ and colleagues summarized what we currently know about reptile genome evolution, and reported genome size appears consistent with a model of continuous gradual evolution, while genomic compartmentalization (the number of macro- and micro-chromosomes) suggests early rapid change. Snakes tend to have an average number of chromosomes between 34 and 37, with 12 to 18 macro-chromosomes and 17 to 20 micro-chromosomes. The haploid genome size rages from 2.0 to 2.9 gb in the species examined, with boids (Boidae) having smaller genomes and sea snakes (Hydrophiinae) having larger genomes. For a comparison of some squamate genomes see Appendix 1.
DNA is an accurate method of determining relatedness. However, when applying it to animals researchers often use relatively small amounts, ranging from a few hundred to several thousand base pairs from one or a relatively few genes. While this may provide a basic idea of relatedness, large data sets that include whole genomes will soon be available to untangle relationships at a much greater level of resolution.
DNA sequences can also be used to produce estimates of the time when two species or groups of species last shared an ancestor. This methodology is dependent upon the DNA clock. Comparison of DNA sequences became possible in the 1970’s when DNA hybridization techniques were used to compare mutation rates. However, it was not until the late 1990’s when DNA sequencing technology, and the ability to clone bits of DNA, allowed the rapid collection of data and the comparison of nucleotide sequences and sequence patterns. The DNA clock works on the assumption that neutral mutations or substitutions in the code are not selected for as long as they don’t change the amino acid at the given position in the protein. Mutations accumulate at a rate constant enough to estimate divergence times if independent information is available to calibrate the rates (fossils, biogeography, or time-stamped ancient DNA). Rapidly evolving mtDNA and cDNA are used for rapidly ticking clocks for the fine resolution of events over short time periods. Nuclear genes that are slowly changing are used for dating events deeper in time. Molecular clocks have always been controversial, but they continue to develop and produce useful estimates for divergence events and they remain the only way to estimate ages of organisms that lack a fossil record. Under the best circumstances, DNA clocks should provide dates that support the fossil record, but it seems improbable that those dates will always match since paleontology cannot be expected to recover the first or the last member of a species.
Phylogenetic systematics or cladistics, the second and lesser known tool for studying relatedness, was developed in the 1950’s (first published in English in 1966) by Willi Hennig, a German entomologist. Cladistics is based on determining which organisms share common derived character states. All vipers all share two things; front fangs on a rotating maxillary bone and a common ancestor. That ancestor and all of its descendants (the vipers) form a clade that is said to be monophyletic, or descendent from a single common ancestor. Some of the vipers, the pit-vipers, also all share heat–sensing pits on the loreal scale. Within the pitvipers are the rattlesnakes, they have folding front fangs, a heat sensing pit and a rattle. Rattlesnakes and their ancestor form a clade nested within the pitvipers, which is nested within the vipers, which is nested with the snakes. Snakes and their ancestor are nested within lizards. The goal of cladistics is to have a classification system that reflects ancestry. A clade is a group of species and their shared common ancestor.
Where problems arise is in the coding of characters and the selection of out groups. In order to build an evolutionary tree for a group of organisms it is necessary to collect characteristics of each organism and code them with 0, 1, or 2. Folding front fangs could be coded as (0) absent or (1) present. Likewise, heat sensing pits may be coded (0) absent (1) on the loreal scale, or (2) on the labial scale. The information is then placed in matrix that can be analyzed by a computer program that compares the data of the group under study to an outgroup (an organism not a member of the clade, but thought to be relatively closely related). The data can then be used to produce a cladogram, or a phylogenetic tree, which visually represents evolutionary patterns and relatedness of a group of organisms.
DNA and cladistics studies have become popular with the increased availability of computers and many people have now looked at squamates using the anatomy of living and fossil forms as well as DNA sequences from living forms. Cladistic relationships backed by morphological and genetic studies are powerfully supported. Unfortunately, there are sometimes disagreements. When conflicts between morphology and DNA occur, John Wiens wrote,
This question must be addressed on a case-by-case basis. In many cases, the morphological hypotheses were only weakly supported, and the incongruence may simply reflect the morphological data being inconclusive. For some groups, the most widely used morphology- based phylogenies were not based on actual analysis…In other cases, there may be a relatively clear source of error in the molecular data…
He goes on to note that while conflicts happen, congruence between morphology and molecular studies occur much more frequently. The DNA clock can be used to provide an independent test of the placement of fossil taxa. For instance, if a fossil predates a lineage by a significant amount of time, it likely represents a different, potentially new, lineage. Today most studies using DNA rely on model-based analysis that uses what is known about the evolution of DNA sequences to produce a tree that best fits the model of evolution. The most widely used model methodologies are the Maximum Likelihood and Bayesian analysis.
Earth’s topography was quite different in the Permian, 299–251 MYA [Figure 2–1]. Tectonic movements were pushing the existing land masses closer together. At the start of the Permian ice and tundra covered some of the land, but tropical swamps persisted in continental interiors. With time, the climate moderated, ice sheets receded and a seasonally wet and dry regime covered much of the interior landmass. With a trend toward drier conditions, the Carboniferous coal swamp forests, with their giant horsetails, club mosses, and the largest insects to ever live, declined and were replaced with shrub-sized ferns, seed ferns, and scale trees. Gymnosperms, such as conifers and ginkgoes, became the predominate vegetation. Permian invertebrates, fish, amphibians and reptiles were abundant and richly diverse; ecosystems were complex with coral reefs, estuaries, and forests present throughout most of the Permian. An estimated 67% of marine life was sessile, and sponges, corals, and brachiopods were abundant. Permian reptiles included the large sailbacked pelycosaurs, the early mammal-like dinocephalians, and the herbivorous pareisaurs. Terrestrial ecosystems had four or five levels of carnivores in their food webs, making them as complex as modern mammal communities. In addition to the large Permian reptiles, there were smaller reptiles, for the most part poorly known, but adapted for life above ground, on the ground, below ground, as well as in the water.
INSERT FIGURE 2-1.
Figure 2–1. The Geological Time Scale. The MYA (millions of years ago) column is the date when that period or epoch ended.
As the Permian came to a close all of the Earth’s land masses were loosely joined together in the C-shaped super-continent Alfred Wegner named Pangea [Figure 2–2]. A single super-ocean, Panthalassa, surrounded the land and its embayment, the interior curve of the C, which became known as the Tethys Sea.
INSERT FIGURE 2-2.
Figure 2–2. Map of the position of Permian landmasses and oceans. Redrawn and modified from Wignall and Twitchett, 2002.
The Permian-Triassic extinction has been called the Great Dying and the Mother of all Extinctions. The loss of life forms did not happen overnight, the extinctions are hypothesized to have lasted over the course of a million years, but a catastrophic event seems to have initiated the crisis. Michael Benton and Richard Twitchett estimated that 95% of the Permian species were lost and asked the question, what combination of events could have such a devastating impact on life? They examined the evidence and while an extra-terrestrial impact is possible, they consider massive volcanic activity more probable. Others have argued that an impact event may well have triggered the eruptions. Extensive volcanic eruptions in the Siberian Traps of eastern Russia produced 2 million km3 of basalt lava that covered 1.6 million km2 of eastern Russia to depths of 400–300 m, and the flows of liquid rock have now been dated, and they are thought to have continued for 600,000 years, beginning about 251.2 MYA. Pyroclastic ash and other debris were expelled into the upper atmosphere. This may have first cooled the atmosphere and then triggered runaway global warming by melting frozen gas hydrates that released methane. The methane accelerated atmospheric warming and raised the temperature about 6º C. In addition to events in the Siberian Traps, Permian extinctions have been linked to other events occurring at roughly the same time. The Emeishan mantle plume erupted in what is now southwest China. Clouds of toxic gases, including carbon dioxide, combined with the volcanic ash and further altered the climate with periods of cooling and warming. Acidic rains and a free-oxygen crisis dramatically altered life on earth.
Hypotheses for the cause of the extinction event as well as evidence to support and refute them continue to accumulate. The Zechstein Sea, an ancient hyper-saline ocean that covered central Europe in the Permian, is now thought to have caused or contributed to, the event. Ludwig Weißflog and colleagues proposed this body of water released 3.7 million of tons of chlorine-based gases each year.
Rocks laid down at the extinction event were scattered in China, Siberia, and South Africa, and it was in South Africa that Rob ert Broom recovered a partial skull of a lizard-like creature he named Paliguana. Broom was educated at Glasgow University moved to South Africa where he practiced medicine in rural areas until 1928 when he took a post as zoologist-geologist at Victoria College. Later he became curator of paleontology at the Transvaal Museum in Pretoria and is best known for his work on early hominid fossils and the excavations at the fossil-rich Karoo Beds. However, despite his other work, Broom’s discovery of Paliguana remains significant.
Unfortunately, Paliguana is poorly preserved and cannot be assigned to the Rhynchocephalia, Squamata, or even Lepidosauria. Susan Evans considers the specimen a basal lepidosauromorph, a group that includes the pseudolizards, beak-heads, and the squamates. Paliguana is significant in that it provides evidence for the survival of lizard-like reptiles beyond the Permian extinction. There are DNA clock dates that place the Squamata/ Rhynchocephalia–Lepidosauromorph split and the Squamata–Rhynchocephalia split in the Permian.
So, what would it take to survive extreme temperature fluctuations, acid precipitation, toxic clouds of gas, and a biosphere in collapse? Characteristics like a low metabolism, a highly flexible diet, the ability to spend long periods underground or underwater without food, and the ability to shut down the cellular demand for oxygen would dramatically increase an animal’s chances of survival. Also, laying eggs with the ability to extend their incubation time, slow embryonic development or place the embryo into a hibernation-like state would increase the probability of survival of offspring through harsh environmental times. The ability to alter the sex ratio of the population would also be useful, increasing the number of males or females should there be a shortage of one sex or the other. Asexual reproduction could also be advantageous as would the ability to store sperm. Collectively modern reptiles have all these traits but most of them are in lineages that can only be traced to the Cretaceous (146–65.5 MYA) in the fossil record.
The beak-heads, order Rhynchocephalia, have a global distribution in the fossil record from the Triassic to the Cretaceous. Almost 30 known genera from the Mesozoic lived many different lifestyles and used diverse habitats. Dietary preference varied widely and, as suggested by tooth structure, herbivores, insectivores, and carnivores were all present. Some, like the tuatara, had teeth for shearing food, cutting it like a pair of scissors. Beak-heads were diverse; most were terrestrial, some were aquatic, and some had long bodies with reduced limbs. The clade shares an enlarged tooth row on a palatine bone that parallels the maxillary teeth, a trait not found in any other tetrapod. Beak-heads also have a lower temporal bar which may be complete or partial; a characteristic usually not found in lizards.
At least one species is thought to have been venomous. Victor Reynosa described Sphenovipera jimmysjoyi from the Middle Jurassic of Tamaulipas, México. This unusual beak-head had fangs on its lower jaw bone with deep grooves on the front, inner surface of the tooth. Additionally, the rear portion of its lower jaw was shortened, allowing jaw protrusion and subsequent gape increase that would have made consumption of large prey possible. Reynosa hypothesized that Sphenovipera held its prey in the mouth, and chewed to deliver its venom.
Beak-heads played an important role in ancient ecosystems until the late Cretaceous. Sebastián Apesteguia and Fernando Novas found skeletal remains of Priosphenodon, a meter long relative of the tuatara, to be the most common tetrapod in the Patagonian fossil beds from the Late Cretaceous. This suggests that the beak-heads were not completely replaced by the lizards until after the Cretaceous extinction event that destroyed the dinosaurs about 65 MYA.
In 1831 John Gray, curator at the British Museum of Natural History, described the tuatara on the basis of a single skull. He considered it to be an agamid lizard and named it Sphaenodon, for its chisel-shaped front teeth. Nine years later he obtained a complete specimen and, not recognizing it as Sphaenodon, he described it a second time. Still considering the reptile an agamid lizard, the species was named Hatteria punctata. Gray’s successor at the museum, Albert Günther, recognized the tuatara’s unique anatomy as one distinct from lizards and established the order Rhynchocephalia (from the Greek rhynchos meaning snout and cephlo meaning head) for the tuatara and the extinct members of the clade.
Tuataras (Sphenodon) are the only living beak-heads and therefore may, like the squamates, provide clues to what the nature of the ancestral lepidosaur. The diversity of beak-heads makes generalizations about their lifestyles difficult, and the tuataras living today are quite derived. But still, they are still the closest living relatives to the Mesozoic forms and the only window science has into beak-head life history beyond what is inferred from the fossil record. Tuataras live in burrows that may be self-created or co-opted from seabirds. Tuataras use chemical cues to detect food, and they feed on insects associated with seabird nests such as crickets, beetles, moths, and wetas. They also eat seabird eggs and chicks, rats, and carrion. Males lack a penis, and sperm transfer is similar to the way it is in birds, with a cloacal kiss. Females can store sperm for 10–12 months. A shallow nest holds the eggs, and incubate for 13–16 months, during winter development stops. Adults, which hibernate through the winter months are estimated to live a century. Some estimates of longevity are as high as 300 years, but the evidence is lacking. The oldest known wild individual is an 88-year-old female, and a captive male is thought to be 111 years old.
Vicky Schaerlaeken and colleagues recently analyzed the tuatara for prey handling abilities. They looked at the size of the external adductor muscle that closes the jaw and examined film of feeding behavior. Their goals included an attempt to understand the role of the lower temporal bar in tuatara feeding. What they found was an adductor muscle smaller than that found in lizards of a comparable size, a weaker bite force than that delivered by a similarly sized lizard, and longer prey handling times. The lower temporal bar appears to stabilize the skull when feeding and the tuatara shows a shearing or scissor-like action when biting prey. This is in contrast to the bite and puncture feeding observed in most lizards.
Tuataras have a cold-adapted metabolism. Michael Thompson and Charles Daugherty measured the metabolism of 142 adults and juveniles at temperatures between 10–15º C on Stephens Island, New Zealand. They compared this data to similar data for modern lizards and wrote,
The metabolic rate of tuatara at 13º C is very similar to that of a lizard at 13º C, the difference being that 13º C is the body temperature at which tuataras are active, but most lizards are not.
They described tuataras as cryophilic (cold-loving), observed activity at temperatures as low as 5.4º C, and found no evidence that they thermoregulate. Tuataras are thermoconformers; their body temperature conforms to that of the environment. The Maori of New Zealand believe the tuatara warms itself using the body heat of seabirds using the same burrow, but this has not been documented and is not supported by the thermal studies done to date.
The generalized diet and a cold-tolerant metabolism, as well as the lineage's 220 MY survival record, are strong evidence for the highly adaptable nature of the Rhynchocephalia. Not only did their ancestors survive the Permian extinction, but also the Cretaceous extinction, events that marked the end for many other lineages.
The beak-heads share a number of common traits with some lizards, including using the tongue to capture food, using the tongue and hyoid to move food into the throat, and using visual cues for hunting from ambush. Additionally, both groups have relatively low metabolisms. However, beak-heads have a lower jaw that swivels on the rear of the skull while squamates have a lower jaw that swivels from the quadrate bone. The quadrate bone then pivots on the back of the skull and gives the squamates a lower jaw that is suspended from the skull with more flexibility and mobility than is seen in the beak-heads. Also, the lower temporal bar present in beak-heads, but absent in lizards, give them a weaker bite force than that found in squamates. The discovery of the Triassic lizard Tikiguania estesi from India suggested not only that squamates separated from beak-heads before the Pangean supercontinent started to break up, but that stem lizards had already split into the lineages that include all other lizards and snakes.
Lizards have been traditionally divided into two large clades. The soft-tongued lizard clade has about 1200 living species, while the hard-tongued clade has about 6000 species, including the snakes. The soft-tongued lizards have muscular tongues that can be used to collect food. The soft-tongued iguanian clade contains a few large, herbivorous iguanas and numerous, smaller insectivorous species: the anoles, the spiny lizards, agamid lizards, and the true chameleons. Many of these lizards feed on ants, wasps, and beetles that are visually located. The lizard lies in ambush seizing prey with its tongue once the invertebrate comes within range. Rarely do you see soft-tongued lizards flicking their tongues to sample molecules in their environment. Food recognition and prey capture used in the iguanians is at least superficially similar to that of the beak-heads; therefore, the iguanians may have retained these potentially ancestral traits.
Ants, wasps, bees, and many beetles have noxious chemical defenses such as formic acid, venom, and quinones used to repel predators. It seems likely that these defenses may be the result, at least in part, of predation pressure from beak-heads and lizards. The Aculeata (ants, bees, and wasps) are dated to the Triassic (about 200 MYA) using trace fossils, and the beetles are somewhat older and may have originated in the Permian (about 285 MYA). Therefore, the beak-heads and lizards have had a long history of co-evolution with these abundant, noxious insects.
Iguanians are very territorial and cryptic in coloration and behavior if they move out of their habitat they tend to stand out and expose themselves to a greater risk of predation. Chameleons rely on slow, deliberate body movements while ambushing bugs from branches, and combined with their color changing ability avoid detection by predators. Soft-tongued lizards have a relatively rigid skull, as well as a lower jaw suspended from a quadrate but their skull is solid, and they lack the bite force possible in other squamates.
The scleroglossans, or hard-tongued lizards, grasp prey with their jaws. Their tongues are not as soft or muscular as the soft-tongued lizards, but are more keratinized (making the tongue harder) and used to collect information. As a result, they rely more on chemosensory information to identify and locate prey during active foraging. Often, they are observed extending their tongue (tongue flicking) to sample the environment for molecules. The hard-tongued species tend to avoid ants, wasps, and beetles and take prey that is less obvious, hidden in leaf litter, burrows, or crevices. The hard-tongued lizards have the advantage of finding resources that were unavailable to the iguanians. By freeing the tongue from use during feeding, the hard-tongued lizards were able to use the organ to chemically locate resources. As a group the hard-tongued lizards (we will ignore snakes for the moment) feed on prey with greater nutritional value (termites, grasshoppers, insect larvae, and spiders), and lack the noxious chemicals, but are more difficult to find.
Hard-tongued lizards have the suspended lower jaw, but they also have a flexible skull with a joint located behind the eyes (in snakes this joint is in front of the eyes); the ability to flex their snout up and down, which increases their bite force and their ability to manipulate prey.
All of this makes for a great story supported by fossils, anatomical, ecological, and behavioral evidence. But, life is complex and while science looks for the simplest answer, nature does not always follow the simplest route.
The first two attempts to understand squamate relationships using DNA were published in 2004. Nicolas Vidal and S. Blair Hedges used parts of two nuclear genes (C-mos and RAG-1) from 19 families. Their outgroups included turtle and tuatara sequences and, as might be expected; the tuatara was the sister to the squamates. The unusual blind snake-lizards (dibamids) formed the most basal clade. Geckos and the legless flap-footed geckos were the next most basal clade. Skinks, teiids, and lacertid lizards followed, along with their sister, the mostly legless worm lizards (amphisbaenids). Anguimorph lizards like the monitor lizards, the gila monster, and the traditional anguids (includes many reduced limb and legless species) were next, and they formed the sister to the snakes. Snakes were not considered to be the closest relatives of the monitor lizards, or the legless worm lizards, or blind snake lizards. Loss of limbs clearly evolved more than once in lizard history. More significantly, the authors considered the placement of snakes to indicate a terrestrial origin as opposed to a marine origin because they did not associate with the monitor lizards. They wrote,
The exclusion here of snakes from varanoids undermines the marine hypothesis of snake origins because it breaks the proposed transition from marine squamate reptiles (mosasauroids) to early marine snakes (pachyophiids)…
A second study, also published in 2004 by Ted Townsend, and colleagues. They used parts of nuclear and mitochondrial genes composing about 4600 base pairs to examine the relationships between 69 species of squamates. Their study produced similar results but showed the hard–tongued lizards, the scleroglossans, did not form a clade. Snakes took a sister position to the anguid lizards and limbless squamates again all had independent origins.
In a 2005 follow-up, Vidal and Hedges used 6192 base pairs from nine nuclear protein-coding genes. However, they sampled only 19 taxa representing the major squamate lineages and used the tuatara to represent the beak-heads (Rhynchocephalia). This study produced estimated dates of divergence, the point in time when the various groups last shared an ancestor. Beak-heads and squamates were estimated to have shared ancestor 240 MYA (or within the range of 251–221 MYA), a date close to the Permian extinction event when many ecological niches would have become available.
The next oldest lineage was the blind snake-lizards (Dibamidae) they formed the sister to the geckos and the flap-footed, legless geckos. These three clades are the only squamates having paired egg teeth or caruncles; the structure hatchlings use to escape the eggshell, all other squamates have a single egg tooth. They are estimated to have shared an ancestor 225 MYA (240–207 MYA).
A slightly younger lineage is the skinks. Today, about 1000 species of skinks are global in distribution, they have a small bone under each scale giving them hard bodies, and they are placed in a clade with the African plated and girdled-tailed lizards (Cordylidae) and the Western Hemisphere night lizards (Xantusiidae). Skinks are basal in this clade and were estimated to have shared an ancestor 192 MYA (209–176 MYA).
Snakes show up next as the most basal to two clades of lizards, the iguanians, and the anguimorphs. All three are estimated to have shared an ancestor 179 MYA (194–169 MYA). A conflict with the date of this clade’s origin is evident. Vidal and Hedges’ estimated date of 179 MYA in the Jurassic is later, about 44 MY later than that known fossil iguanid from the Triassic of India, Tikiguania estesi. It may be the date based on DNA is off but is also possible that the fossil will not turnout to be a true iguanian.
Again, the DNA did not support the hard-tongued lizard clade; snakes are the sister to the soft-tongued iguanians and the anguimorphs. And, the Townsend et al. hypothesis that the soft-tongued lizards are not carrying the ancestral feeding behavior of the beak-heads, but evolved it independently was supported.
A third DNA study using the nuclear gene RAG-1 was done by Andrew Hugall and colleagues. They examined 88 tetrapod species, including 37 squamates and the tuatara, and used a relaxed clock method to estimate the age of various clades. The shared ancestor of beak-heads and squamates was 268 to 275 MYA, a date prior to the Permian extinction event. Geckos and their relatives are thought to be the oldest squamate lineage last sharing an ancestor with the other squamates about 185 MYA. The blind snake-lizards were the sister to the skinks and their relatives and shared an ancestor with the other squamates only slightly later than the gecko ancestor. The Lacerta clade is about 165 MY old, and they form the sister to a clade that includes the snakes as the sister group to anguimorphs and iguanians. In this study, the Lacerata is the sister to the clade that includes the snakes, anguimorphs, and iguanians, and they shared an ancestor about 160 MYA.
Vidal and Hedges came back in 2009 with a revised timetree for the squamates [Figure 2–3] suggesting squamates arose toward the end of the Triassic instead of the early Triassic. Those blind snake-lizards, the dibamids, had a Triassic ancestor; all of the other modern clades had origins in the Jurassic or Cretaceous. Unexpectedly, most of the worm lizards (the amphisbaenians) did not evolve until the early Cenozoic, about 40–50 MYA; the exception being the Florida Worm Lizard (Rhineura floridana) and its fossil relatives the rhineurids that originated in the mid-Cretaceous, about 125 MYA. The clade containing the snakes, anguimorphs, and iguanians― the Toxicofera had an origin in the Jurassic, (197–161 MYA). The Toxicofera clade will play an important role in the future discussion. However, like all previous studies, the snake-iguanian-anguimorph trichotomy remained unresolved. Figure 2–4 illustrates representatives of the major squamate clades. For a summary of the major clades of living squamates and their distributions see Appendix 2.
INSERT FIGURE 2-3.
Figure 2–3. Squamate phylogeny hypothesized by Vidal and Hedges (2009) with a time scale at the bottom. Letters correspond to photos of representatives of major clades shown if Figure 2–4.
A morphological analysis of squamates, done by Jack Conrad in 2008 runs contrary to the DNA evidence and supports some of the previous studies that used morphology. The hard-tongued lizard (Scleroglossa) clade has a single ancestor exclusive of the soft-tongued iguanian clade, and perhaps more interesting, this study found the mostly limbless blind snake lizards (dibamids), worm lizards, and snakes to be each other’s closest relatives. Conrad’s analysis includes them in a larger clade with the skinks, a group of lizards that contains many species that show reduced limbs and elongated bodies. Similar arrangements have been produced in other studies and it seems to result from the lack of limbs that polarizes the legless together in an artificial group.
INSERT FIGURE 2-4.
Figure 2–4. Representatives of major squamate clades. [A] Amphisbaenia, Black and White Worm Lizard, Amphisbaena fuliginosa [B] Lacertidae, Long-tailed Grass Lizard, Takydromus sexlineatus [C] Teiidae, Canyon Spotted Whiptail, Aspidoscelis burti [D] Helodermatidae, Gila Monster, Heloderma suspectum [E] Anguidae, Island Glass Lizard, Ophisaurus compressus [F] Varanidae, Nile Monitor Lizard, Varanus niloticus [G] Iguania, Spiny-tailed Iguana, Ctenosaura similus [H] Serpentes, Hognosed snake, Heterodon platirhinos [I] Xantusidae, Desert Night Lizard, Xantusia vigilis [J] Cordylidae, Sungazer, Cordylus giganteus [K] Scincidae, Broadheaded Skink, Plestiodon laticeps [L] Gekkonidae, Mourning Gecko, Lepidodactylus lugubris [M] Pygopodidae, Burton’s Flap-foot, Lialis burtoni [N] Dibamidae, the New Guinea Blind Snake-lizard, Dibamus novaeguineae from Sulawesi.
Search the literature on squamate phylogeny and you will find studies that use anatomy, others that use DNA and a few that use anatomy and DNA and when the studies are examined for the closest relatives to the snakes virtually all of the major groups of lizards have been considered the sister to the snakes at one time or another. The advantage to DNA is that it circumvents the problem of homoplasies, characters that look like they are identical through descent from a common ancestor but are in reality the product of convergent evolution.
In 2009 Michael Lee combined nuclear DNA, mitochondrial DNA, and morphological data in an analysis that puts snakes, iguanians, and anguimorph lizards in a clade that is in agreement with the relationships recovered by Vidal and Hedges in 2007 when they used nine nuclear genes and recognized the Toxicofera. Lee’s analysis placed snakes and fossil Cretaceous marine lizards in the Toxicofera. This study constrained the morphological and mtDNA data to choose between alternate inter-relationships among the iguanians, snakes and anguimorphs and united snakes with the anguimorphs lizards.
Given that the blind snake-lizards (Dibamidae) [Figure 2–5] show up as the most basal squamates in many of the molecular studies raises the question, did the ancestral squamate have reduced legs? As characters are coded for more anatomy, more DNA base pairs are sequenced, and as whole genomes are read and compared the evolutionary trees will be more reflective of history, in the meantime, the studies are very entertaining.
INSERT FIGURE 2-5.
Figure 2–5. The blind snake-lizards (Dibamidae) may be relicts of the oldest living squamate lineage. Top: The head and anterior body of Greer’s Blind Snake-lizard, Dibamus greeri, a Southeast Asian species. Bottom: the tail, remnant rear legs, and hemipenes (between the rear legs).
Biologists have long considered sex a problem, why should it take two organisms to make one? Asexual reproduction is much simpler, one organism producing its own offspring is efficient and the offspring receives all of the genetic material from the parent. Sexual reproduction is both inefficient and messy. Two animals need to agree to get close enough to each other so that reproductive cells and the genetic material they contain can be passed from one individual to another. Sex involves time and energy not required in asexual reproduction, cells are frequently wasted, and sex risks breaking up co-adapted gene complexes. However, the fact is most organisms from bacteria to animals recombine DNA, therefore it seems likely sex has some real benefits to survival, that is long-term survival of genes. The answer to the benefits of sex question is in the recombination of genes making offspring more fit to survive changing environments and diseases. Sex generates the raw material for evolution.
Chromosomes were discovered independently by Walter Sutton in the USA and Theodor Boveri in Germany early in the 20th century; soon afterward sex chromosomes were discovered by Thomas Morgan working with fruit flies. Since humans and other mammals were discovered to have their sex determined by chromosomes (packages of DNA) received from the male parent. Male fruit flies and male mammals possess a small Y and a large X chromosome and the XY system has become the traditional explanation for sex determination in biology textbooks. A sperm carrying a small Y chromosome usually produces males, while a sperm carrying a large X chromosome usually produces a female. The XY combinations are normally males and XXs are normally females.
The discovery of the SRY gene in 1985 further narrowed the factors that determined sex in mammals, the SRY gene is normally on the Y chromosome, but sometimes jumps to the X chromosome making an XX male a rare possibility. Nature is complex, and in squamates, the females often have the structurally distinct sex chromosomes, so they are said to be ZW and the males are ZZ, but some squamate species have the XY sex chromosome system. When structurally distinct sex chromosomes are present they can be detected by studying cells and karyotypes. However, sex chromosomes may not be structurally distinct, they may be the same size and in some cases, a single gene may determine sex, or initiate a cascade of events that trigger the development of gonads and secondary sexual traits.
The default gender in vertebrate embryos is female. Embryos only become male if a genetic switch has been turned on. The evolution of genetic sex determination mechanisms across tetrapods has yet to be satisfactorily explained, but Jeremiah J. Smith and S. Randal Voss have found a region of a salamander chromosome that corresponds to sex chromosomal regions of birds and mammals suggesting the sex chromosomes of birds (the Z) and the sex chromosome of mammals (the X) were recruited from a common ancestral chromosome present in amphibians. Evolution tinkers.
The Bearded Dragon, Pogona vitticeps [Figure 2–6], has a micro-chromosome that carries the gene that determines an individual’s sex; in such cases identification of genetic sex determination (GSD) may require more advanced molecular techniques. Sex determination in reptiles has a surprising variety of mechanisms that involve both genetics (DNA) and the embryo’s environment. Sex determination can be by genes (with or without sex chromosomes) as in most mammals; some reptiles use parthenogenesis (asexual reproduction); and some squamates use the temperature of the embryo during a critical period of development to determine sex (temperature-dependent sex determination, TSD) in at least three different patterns. And, perhaps most surprisingly some lizards (and possibly other reptiles) that use GSD can have the genetic mechanism overridden by environmental conditions (TSD). So, while an animal may have the genes to be a male, it can become an egg-producing female because of the temperature of the embryo during a critical period of development.
INSERT FIGURE 2-6.
Figure 2–6. A hatching Bearded Dragon, Pogona vitticeps.
Females of the live-bearing Ocellated Skink (Niveoscincus ocellatus) from high elevations in Tasmania can determine the sex of their offspring by the amount of time they spend basking. Erik Waspstra and colleagues demonstrated this with captive skinks as well as wild caught gravid females. Females with limited basking opportunities (4 hours per day) produced more male offspring, while females with more basking opportunities (10 hours per day) produced more females.
Madeleine Charnier first discovered TSD in the Red–headed Rock Lizard (Agama agama) in 1966, since then TSD has been found in fish, turtles, crocodiles and alligators, tuataras, and other lizards. Eric Charnov and James Bull proposed a hypothesis to explain TSD over genetic (chromosomal) sex determination: under some environmental conditions, greater reproductive fitness goes to female offspring, while under other conditions male offspring are more reproductively fit. Natural selection can be expected to favor females that deposit their eggs in nests that will be warm, cool, or have a range of temperatures depending upon the sex with the highest reproductive value for a given environment. Given this mechanism, selection of a nesting site by the female becomes quite important, nests in shade, sun, or partial sun will determine the sex ratio of the offspring. And for species giving birth to live young, basking time and body temperature become important.
Experiments done by David Crews and James Bull as well as other colleagues over the past 20 years using the Leopard Gecko (Eublepharis macularis) [Figure 2–7] demonstrated incubation temperature not only determines sex but influences adult physiology, behavior, anatomy, and brain development. Recent experiments found incubation temperatures influence mate choice and aggressive behavior in the Leopard Gecko. Eggs incubated at 32.5º C produced mostly males and some females with adults of both sexes showing considerable aggression; while eggs incubated at 30.0º C are mostly females but also produce some males and both sexes are less aggressive. Using a simple Y-maze, males were given a choice of females incubated at the two different temperatures. Male leopard geckos incubated at 32.5º C preferred females incubated at 30º C and males incubated at 30º C preferred females incubated at 34º C. They concluded that mate choice is due to an environmental influence on brain development that leads to different perceptions of the opposite sex. Mate choice is best understood in the context of the individual’s life history.
INSERT FIGURE 2-7.
Figure 2–7. The Leopard Gecko, Eublepharis macularis. This lizard has been used extensively in experiments on temperature-dependent sex determination and as a model for how the developmental environment of the embryo impacts the adult.
The Jacky Dragon, Amphibolurus muricatus, is a short-lived agamid lizard inhabiting dry forests of southeastern Australia and was used by Daniel Warner and Richard Shine to test the Charnov-Bull hypothesis. They manipulated embryos with steroid hormones to produce both sexes at a variety of temperatures. Normally, females are produced at 23–26º C, while males are produced at 30–33º C, and temperatures of 27–30º C produce offspring of both sexes. They incubated eggs under each temperature regime and applied a steroid hormone to half of them to override the thermal effects. The hormone blocked the conversion of testosterone (male hormone) to oestradiol (female hormone), so that males were produced at cooler temperatures and females at warmer temperatures. Previous work had shown that the hormone treatment had no effect on morphology or survival. They found warm incubation resulted in accelerated development of females. Eggs hatched early in the season had the highest lifetime fitness (successful reproduction) but no increase in fitness was detected in intermediate or cool treatment females. Males incubated at intermediate temperatures had the highest fitness; those from the extreme temperature treatments had the lowest fitness. The Charnov-Bull hypothesis was supported by empirical evidence for the first time in reptiles.
Evidence that some squamates have both temperature-dependent sex determination and genotypic sex determination mechanisms have started to accumulate, with the research to date suggesting that under extreme conditions species with GSD may have their genes overridden by TSD. Adrian Quinn and colleagues incubated bearded dragon (Pogona vitticeps) eggs at high temperatures and obtained genetic males that were physically females. The Australian Eastern Three-lined Skink, (Bassiana duperreyi) showed a similar mechanism, with genetic females converted into males by cool nest temperatures.
Nest temperatures, and therefore the site selected for nesting by a female squamate, may have a direct impact on the sex ratio. Using the short-lived Jacky Dragon already discussed, Daniel Warner and Richard Shine radio-tracked females to see where they would place their nests. The females consistently chose nest sites with low canopy cover that received high levels of sunlight. These sites had a higher mean temperature, a higher maximum temperature, and greater temperature fluctuations than did randomly chosen nearby sites. Early in the season, these nesting sites produce mostly female offspring, and females nesting later in the season produced mostly male offspring. Given this mechanism, there would be a shift in the sex ratio of the offspring during the year. Females that hatch early have less competition and more pre-winter growth which equals a higher reproductive output.
But more surprises regarding sex determination were yet to come. Rajkumar Radder and colleagues have added yet another mechanism for sex determination in squamates. Again using the Australian Eastern Three-lined Skink they discovered large eggs tend to produce females, and small eggs tend to produce males. And, if you remove some yolk after the egg was laid the embryo is likely to switch to being a male, even if it has female sex chromosomes. Injecting extra yolk, the egg will likely produce a female. Therefore, females may influence the sex of their offspring by the way they allocate yolk during egg production.
Experiments by Nicola Mitchell and colleagues with both species of tuataras found females are produced at lower temperatures (18–22º C) and males at higher temperatures (22–24º C). Exposure to these temperatures when the embryo is between 25–55% developed determines the sex. The tuatara pattern of females produced at cooler temperatures and males at higher temperatures (FM) is rare in other reptiles. In lizards (as well as turtles and crocodilians) with TDS females are most often produced at both low and high-temperature extremes and males are produced at moderate temperatures (the FMF pattern), or males are produced at low temperatures and females at high temperatures (the MF pattern). Exposure of tuatara embryos to temperatures of 25º C or above have lethal consequences, and therefore it seems unlikely female tuataras will eventually be shown to be produced at the extreme temperatures and males at moderate temperatures.
Martina Pokorná and Lukáš Kratochvíl compiled information from more than 423 studies on sex determination in squamates and used a comparative phylogenetic analysis to reconstruct the evolution of squamate sex determination. Previous researchers had suggested that genetic sex determination (GSD) was the ancestral squamate condition, and temperature-dependent sex determination mechanisms had evolved separately in various groups. Pokorná and Kratochvíl’s analysis suggested the reverse, TSD was the ancestral squamate trait and that various lizard lineages have transitioned to GSD, they found little support for the reversal of this trend, lineages that evolved GSD did not have descendants return to TSD. They asked the question, are sex chromosomes an evolutionary trap? That is, once sex chromosomes have evolved they cannot be lost. However to date, snakes have all been found to use GSD with the ZW chromosome system and under some circumstances, females use parthenogenesis― virgin birth. Given the plasticity found in sex determination mechanisms in many vertebrate lineages, it may only be a matter of time until snakes too are shown to have a greater diversity of mechanisms.
Squamate reproduction made headlines around the world in late 2006 when it was discovered that a female Komodo Dragon (Varanus komodoensis) living in the Chester Zoo laid 25 eggs 11 of which hatched. The female Komodo Dragon had never encountered a male while living at the zoo so when she laid eggs the zoo keepers realized she was using parthenogenesis to reproduce. DNA tests confirmed the offspring contained only the mother’s DNA. Another captive bred female in the London Zoo laid four viable eggs out of 22. This female had not been in the presence of a male in 2.5 years. Of interest, is that the second female subsequently mated with a male and laid four eggs, one of which hatched. Using genetic fingerprinting, offspring from the first clutch contained only the females DNA. The offspring produced after mating contained both parents’ DNA. The previous year, parthenogenesis had been reported in a related species, a captive Yellow-spotted Monitor, Varanus panoptes.
Parthenogenesis is widespread in lizard and snake lineages and it is often described as an evolutionary dead end for species that use it to the exclusion of sexual reproduction. However, it is, in fact, facultative in many species and a long-term survival strategy for an individual’s genes. The Komodo Dragon discovery and many observations on other lizards, as well as snakes isolated in captivity, suggested female squamates are capable of parthenogenesis when sperm are unavailable. Zoos and other captive situations produce this condition, but in nature, it occurs when a female wanders out of its normal habitat. Or, there is a reduced number of males in the population.
Parthenogenesis is accomplished during egg production. Meiosis produces eggs that have one member from each pair of chromosomes (n), the extra chromosomes are reduced by the cell splitting into a large egg and a smaller polar body carrying the extra chromosomes. Females produce viable eggs without sperm when the polar body nucleus acts like a sperm and fuses with the egg’s nucleus to produce a 2n cell capable of growing into an embryo.
Female squamates that have a ZW or an XY sex-chromosome combination can produce asexual offspring that have a ZZ or WW (or an XX or YY) sex chromosome combination. The ZZs (or XYs) are males; the WWs (and YYs) are thought not to survive development. An isolated ZW female could produce sons that could later produce sperm to fertilize her eggs or the eggs of another female that may have wandered into the same isolated location. This may be the reason why parthenogenesis occurs most frequently in species with the ZW system. An XX female could produce only XX female offspring that may, in turn, be able to reproduce asexually or sexually should it encounter a male.
So what determines whether a species or population has a XY/XX or a ZW/WW sex chromosome system? John H. Werren and co-workers have proposed these systems are produced when there is a single dominant gene that determines the sex of the individual and when it is more costly to produce one sex over another. If male offspring reduce the fitness of its brood or parents (more costly males) a dominant Mm male and a recessive mm female system evolved. Should the female offspring reduce the overall fitness (be more costly) a dominant Ff female, and a recessive ff male system will evolve. In species with XY systems, males are more costly to produce. The Y chromosome is smaller and degenerates to reduce recombination of genes between the Y and the X, there are several reasons for this and it appears to be a way to regulate the dosage of genes an individual receives. The ZW sex chromosome system evolves when females are more costly to fitness.
Multiple paternity is also a widespread phenomenon in tuatara and squamate reproduction. Examine any clutch of eggs or litter of offspring from a sexually reproducing squamate and it is likely that the offspring have two or more fathers. The benefits of multiple paternity are often described as favoring the female’s genes. After all, 100% of the offspring are carrying part of the mother’s genome, while only a fraction of the offspring are carrying the genetic material of any one male. However, Tobias Uller and Mats Olsson argue that there is no evidence that high levels of multiple matings are producing benefits to females and that evidence for indirect genetic benefits from multiple paternity are weak. Instead, they suggest multiple paternity results from females frequently encountering males; that female squamate are not particularly choosy about picking a mate, and the cost of repeated matings is low to the female and benefits the male. But, they cite studies, with the European Adder (Vipera berus) and the Sand Lizard (Lacerta agilis) where there is evidence for indirect genetic benefits to females because of a positive correlation between multiple matings and increased offspring viability. In a multiple paternity study of two aquatic homalopsid snakes from Southeast Asia Harold K. Voris and colleagues found litters to have three to five male parents. Males can be more abundant than females at some locations and they also suggest multiple paternity results from females frequently encountering males, the low costs of mating to females, or the cost involved in resisting mating with multiple males.
Sylvain Ursenbacher and colleagues studied the reproductive success of wild male European Adders (Vipera berus) in low-density populations in Switzerland’s Jura Mountains. Previous studies had shown small "sneaky" males would mate with females while larger males were engaged in combat over the female. They found that the first male to mate with a female was usually the one to fertilize the largest number of eggs. The results showed multiple paternity can occur frequently in natural Adder populations even if the density of adults is low. And, they found male reproductive success significantly increased with body length, only the largest males successfully fathered entire clutches. Additionally, they found no relationship between the number of fathers per clutch and neonate survival. And, females could benefit from multiple matings in populations with high levels of inbreeding or low male fertility. Squamates have a polygynandrous mating system where both sexes engage in multiple matings, even in species that appear to be socially polygynous or monogamous.
So, how do female squamates store sperm? Paola María Sánchez–Martínez and colleagues studied the transitional region between the uterus and the vagina in 12 families of squamates. The mucus-producing tissues of this region are folded to form deep pockets that in at least some of the species form sperm storage structures. While some folds were empty, others were full of sperm with their heads directed in an orderly manner toward the bottom of the fold. Dustin S. Siegel and David Sever summarized the literature on sperm storage tubules in snakes and found them present in five families in the infundibulum of the uterine tube, where the stored sperm would be relatively close to the eggs at ovulation.
Many female squamates have been shown to store sperm for varying periods of time and multiple matings increase the probability a female will have sperm from several males available at ovulation. The sperm can then compete for fertilization in her reproductive tract and her offspring will have a greater diversity of genetic material. How long sperm can survive in the female’s reproductive system is not known, before snakes were known to use parthenogenesis, females were thought to be able to store sperm for decades. That now seems unlikely, given that females are now known to use parthenogenesis when isolated from males.
Environments change and the ability of animals to quickly adapt to changes is undoubtedly important for survival. To date, much of the emphasis of adaptation was thought to be through genes and natural selection. But can factors like weather, food availability, and the presence of predators alter the traits of offspring during development so the offspring from a particular litter are more fit to survive because of their developmental experiences?
Richard Shine and Sharon Downes used the live-bearing Tussock Skink (Pseudemoia pagenstecheri) to test female squamate abilities to assess environmental conditions and alter the traits, or the phenotypes, of their offspring accordingly. Female squamates may do this via the physical conditions the embryos experience during incubation. In captive and semi-wild environments Shine and Downes manipulated the food supply to produce deprived gravid females, restricted basking sites to gravid females, exposed gravid females to the scent of a predatory snake and had control gravid females that had plentiful food, basking sites, and no exposure to the scent of predatory snakes. Offspring were tested for fitness by their size, shape, their ability to run, and their response to predator odors. Offspring from well-fed females were born early, tended to be larger but ran slowly; while smaller young from food-deprived females ran faster. And perhaps most unexpectedly, offspring from females exposed to predatory snake scents showed a more intense response to the scent than offspring from females not exposed to the scents; and they were heavier and had long tails. Also, these offspring did not show the anti-predator raised–tail wagging behavior seen in the offspring whose mother was not exposed to the odor of a predatory snake prior to birth.
Most populations of the common Viviparous Lizard (Lacerta vivipara), as its name suggests, give birth to young, but females in some populations lay eggs with thin shells. Florentino Braña collected eggs from 69 females from an egg-laying population and incubated trios of eggs in close contact. Upon hatching the neonates from each trio were sexed and categorized as a same-sex trio or a hetero sex trio (one male and two females or the reverse). One member from each same-sex trio was selected at random and the individual of the uncommon sex was selected from each hetero trio. Data collected from each lizard included digit length since this trait is sensitive to steroid hormones. Hatchlings incubated next to siblings of the same sex had more body mass, males tended to have fewer ventral scales, females had more. The length of the fourth digit on the hind limb was longer in females incubated with males, a trait normally associated with males. The overall sex ratio did not depart from one male to one female, but the fact that nearby eggs can influence the phenotype of the offspring suggests hormones shared between eggs may impact lifetime fitness.
Hormones circulating in the blood of female lizards and snakes carrying embryos influence the life history of those offspring. Corticosterone is known to influence development, anatomy, and behavior and Kylie Robert and colleagues examined the impact of this hormone on the behavior of fast– and slow– growth Western Terrestrial Garter Snakes (Thamnophis elegans). Previous work had shown that snakes from lakeshore habitats were fast-growth snakes that reached adult sizes of 400 to 700 mm, females produced their first litter at three years of age, the median lifespan was four years, fish were the most common prey and water was always available, these snakes had an average nighttime temperature of 25º C and the population was exposed to high levels of predation from birds. Other snakes from nearby meadow environments had slow-growth. Adult body sizes were 400 to 550 mm, females produced their first litters at five to seven years of age, the median lifespan was eight years, frogs were the most common prey and water availability was ephemeral, predation by birds was low, and the snakes had an average nighttime temperature of about 20º C. Robert and colleagues collected pregnant females from both these populations and found females from the meadow environment (slow-growth snakes) had corticosterone levels six times higher than females from the lake population (fast-growth snakes). At one month of age offspring from fast growth, females had higher corticosterone than those from slow growth females. This was the opposite of the maternal condition. Offspring from fast growth females treated with corticosterone showed less anti-predator behavior (crawling backward to escape), and offspring from corticosterone-treated slow growth females showed less tail lashing behavior. Corticosterone-treated females had a higher rate of stillborn young and young that died within the first month after birth than did non-treated females.
Many squamates lay their eggs in communal nests and Rajkumar Radder and Richard Shine used the Australian Eastern Three-lined Skink (Bassiana duperreyi) to determine what advantages communal nesting may have on the embryos. Field and laboratory data were used to test two ideas: communal nesting was due to the scarcity of nest sites, or communal nesting was an adaptation that that increased fitness. They discovered single clutches and communal clutches were laid in similar sites in the field and that communal nesting was not predictable based upon the available number of nesting sites. In the lab, females laid their eggs next to others rather than choosing an isolated location that was identical. When the eggs were incubated in the lab the tightly packed eggs took up less water, and when the hatchlings from the communal nests were compared with hatchlings that came from single egg nests, they were larger and could run faster than hatchlings from single egg nests. Communal nests may be better able to equalize moisture among a greater number of eggs than those laid singly or in small groups.
Local weather conditions during incubation can impact the phenotypes of squamate offspring. Olivier Lourdais and colleagues used gravid female Aspic Adders (Vipera aspis) collected from the field between 1992 and 2000. The study obtained 173 litters with 817 healthy offspring from this small Palearctic snake; and demonstrated development time, embryo viability, and offspring phenotypes are all affected by the weather. Hot weather in June (early in gestation) increased the ventral scale counts in offspring, while hot weather in mid–July (mid-gestation) shortened development time and birth was early; hot weather in August (late gestation) reduced the number of stillborn young.
Selection of a nesting site by egg-laying females and the time spent basking by live-bearing female squamates, as well as the predators the gravid females are exposed to, impact the young squamate in subtle ways that are not yet completely understood. Add to this, the diversity of sex-determining mechanisms, the high frequency of multiple paternity, the ability to store sperm, the ability of hormones to diffuse from one egg to another, and squamate reproduction is highly plastic and adaptive. Evolution has been tinkering with squamate reproduction for a very long time – probably before the Permian extinction, 251 MYA.
