Daniel G. Blackburn

Structure, function, and evolution of the oviducts of squamate reptiles, with special reference to viviparity and placentation

In lizards and snakes, the oviducts function in fertilization, sperm storage, egg transport, eggshell deposition, maintenance of the early embryo, and expulsion of the egg or fetus. In viviparous forms they also contribute to placentae responsible for gas exchange and nutrient provision to the fetus. Dissections of species of 30 genera coupled with data from the literature indicate that squamate oviducts vary interspecifically in seven macroscopic features, including the extent and nature of regional differentiation, vascular supply, topographic asymmetry, number of oviducts, vaginal pouches, and relationship to the cloaca. The uterus, infundibulum, and vagina differ histologically in their epithelia, glands, and myometrial layers. Season cyclicity occurs in all three oviductal regions, most prominently in the uterus, and is under endocrinological control. Regional and cytological specializations reflect the diverse functions performed by the oviduct. Definitive evidence for oviductal albumen production and egg resorption is lacking. In viviparous squamates, three uterine specializations may facilitate maternal-fetal gas exchange: an attenuated epithelium, reduced uterine glands (and a reduced shell membrane), and increased vascularization. Contrary to previous reports, pregnant uteri show no epithelial erosion or capillary exposure. Specializations for nutrient provision to the fetus include mucosal hypertrophy, enlarged glandular epithelia, and multicellular glands whose secretions are absorbed by the chorioallantois. Comparisons with other amniotes indicate that squamates inherited the oviduct as an organ with capabilities for egg uptake and transport, fertilization, eggshell deposition, and oviposition. Other features have evolved convergently among squamates: infundibular sperm receptacles, unilateral oviduct loss, uterine gestation, placentation, and specializations for placentotrophy. Cladistic analysis indicates that oviductal features associated with deposition of tertiary egg investments in reptiles reflect evolutionary convergence as well as secondary simplification, rather than a unidirectional trend towards increased specialization.

Reptilian viviparity: past research, future directions, and appropriate models☆

Squamate reptiles represent an ideal group for studies of viviparity, because they have evolved this reproductive pattern frequently, relatively recently, and at low taxonomic levels. A phylogenetic approach shows particular promise in helping us interpret anatomical, physiological, and ecological diversity. This review summarizes four major categories of active investigation: (1) reproductive anatomy and physiology; (2) placental structure and function; (3) reproductive endocrinology; and (4) reproductive and physiological ecology. Evolutionary reconstructions suggest that on many occasions viviparity has evolved concomitantly with functional placentation, through reduction of the shell membrane and hormonal modifications that prolong gestation. Studies of placentotrophic clades as well as reproductively bimodal species offer great potential for explaining the evolution of viviparity and placentation. However, live-bearing squamates are reproductively diverse, and appear to have solved physiological problems associated with viviparity by a variety of mechanisms. Consequently, studies on one or a few squamate species appear increasingly unlikely to yield all-inclusive explanations. Future studies and analyses should abandon assumptions of universal physiological mechanisms and a single historical sequence, in favor of the documentation of diversity in phylogenetic and quantitative terms.

SQUAMATE REPTILES AS MODEL ORGANISMS FOR THE EVOLUTION OF VIVIPARITY

ABSTRACT For over a century, research has been conducted on squamates in order to reveal how viviparity has evolved in mammals and other vertebrates. The recent proliferation of studies has yielded much information on anatomical, physiological, ecological, and evolutionary aspects, allowing a reassessment of squamates as model organisms for the study of viviparity. Strong support for the “squamate model” comes from phylogenetic analyses that have shown that squamates have evolved viviparity with great frequency (> 108 origins), at low taxonomic levels, and in geologically recent times. However, available data also indicate that viviparity has evolved by different chronologies and mechanisms in squamates, fishes, and mammals. Further, generalizations about squamates are difficult to make, given the diverse mechanisms by which they achieve viviparity. Thus, similarities between squamates must be demonstrated empirically, and generalizations should be based on quantitative, phylogenetic analyses of multiple lineages. Explanations for similarities between squamate clades can invoke such concepts as evolutionary constraints, exaptations, and selection pressures, and should distinguish between adaptations, correlated attributes, and features that predate viviparity. However, homocentric assumptions of an orthogenetic transformation towards the eutherian condition should be abandoned, along with untested assumptions that viviparity squamates and mammals is similar. The value of the squamate model ultimately may lie in insights it provides into physiological problems rather than in universality of specific mechanisms that have evolved to meet those problems.

Evolutionary origins of viviparity in the reptilia. II: Serpentes, amphisbaenia and ichthyosauria

Superimposition of reproductive mode data from the literature over phylogenetic classification systems reveals that viviparity (live-bearing reproduction) has evolved on at least 35 independent occasions among the Serpentes, once in the Amphisbaenia, and once in the Ichthyosauria. Of the ophidian origins of the live-bearing mode, at least fourteen have occurred in the Colubridae, twelve in the Viperidae, three in the Hydrophiidae (used in the sense of Smith et al., 1977), and one in each of the following groups: Boidae, Acrochordidae, Tropidophiidae, Uropeltidae, Typhlopidae, and Elapidae. Previous analysis has distinguished and defined 45 origins of viviparity among the lizards. Here, ten additional saurian origins are recognized on the basis of unpublished and recently published evidence, three in the Iguanidae, two in the Scincidae, and one in each of the following groups: Agamidae, Chamaeleontidae, Anguidae, Xenosauridae, and Anniellidae. As phylogenetic relationships are clarified, further origins seem likely to be detected, particularly in the Colubridae, Hydrophiidae, Scincidae, and Iguanidae. At present, however, at least 92 origins of viviparity can be recognized within the class Reptilia. Reptilian viviparity has arisen on multiple occasions in each of the six major biogeographic regions, with a majority of the origins having occurred in the Old World. Nearly 19% of the extant reptile species are probably live-bearers, including more than 20% of the snakes and over 19% of the lizards. About 71 % of the viviparous species belong to either the Scincidae, Colubridae, Viperidae, or Iguanidae. The discontinuous distribution of the origins of viviparity among the reptilian families supports the hypothesis that selective pressures, preadaptations, and constraints vary at high taxonomic levels.

Ecology and life history of the viviparous lizard Mabuya bistriata (Scincidae) in the Brazilian Amazon

The viviparous lizard Mabuya bistriata was studied in two lowland tropical forest sites in Amazonian Brazil with additional data taken on museum specimens. These diurnal lizards are active primarily during midto late morning on fallen logs or low on tree trunks. They are heliothermic averaging 32.9 ? 0.98 C in body temperatures. Prey include orthopterans, spiders, eruciform larvae, termites, and other invertebrates. Prey differences between the two sites most likely reflect differences in prey availability associated with tropical seasonality in rainfall.

Evolution of vertebrate viviparity and specializations for fetal nutrition: A quantitative and qualitative analysis

Phylogenetic analyses indicate that viviparity (live‐bearing reproduction) has originated independently in more than 150 vertebrate lineages, including a minimum of 115 clades of extant squamate reptiles. Other evolutionary origins of viviparity include 13 origins among bony fishes, nine among chondrichthyans, eight in amphibians, one in Paleozoic placoderms, six among extinct reptiles, and one in mammals. The origins of viviparity range geologically from the mid‐Paleozoic through the Mesozoic to the Pleistocene. Substantial matrotrophy (maternal provision of nutrients to embryos during pregnancy) has arisen at least 33 times in these viviparous clades, with most (26) of these origins having occurred among fishes and amphibians. Convergent evolution in patterns of matrotrophy is widespread, as reflected by multiple independent origins of placentotrophy, histotrophy, oophagy, and embryophagy. Specializations for nutrient transfer to embryos are discontinuously distributed, reflecting the roles of phylogenetic inertia, exaptation (preadaptation), and constraint. Ancestral features that function in gas exchange and nutrition repeatedly and convergently have been co‐opted for nutrient transfer, often through minor modification of their components and changes in the timing of their expression (heterochrony). Studies on functional and evolutionary morphology continue to play a central role in our attempts to understand viviparity and mechanisms of fetal nutrition. J. Morphol. 276:961–990, 2015.

Histology of the late-stage placentae in the matrotrophic skink Chalcides chalcides (Lacertilia; Scincidae)

Examination of late‐stage placental material of the lizard Chalcides chalcides from the Hubrecht Laboratorium (Utrecht, The Netherlands) reveals several cytological and histological specializations that appear to have been superimposed over a morphological pattern that is typical for squamates. The chorioallantoic placenta is highly vascularized and consists of a single mesometrial placentome and a generalized paraplacentomal region, both of which are epitheliochorial. The placentome is deciduate, and contains deeply interdigitating folds of hypertrophied uterine and chorioallantoic tissue. Chorionic epithelium lining the placentome comprises enlarged, microvilliated cells, a small proportion of which are diplokaryocytes. The placentomal uterine epithelium is not syncytial and consists of enlarged cells bearing microvilli. The yolk sac placenta is a true omphaloplacenta (sensu stricto), being formed by juxtaposition of uterine tissues to an avascular, bilaminar omphalopleure. Epithelium of the omphalopleure is stratified and is hypertrophied into papillae that project into detritus of the uterine lumen. The omphalopleure is separated from the yolk sac proper by a yolk cleft that is not confluent with the exocoelom and is not invaded by the allantois. Neither an omphalallantoic placenta nor a true choriovitelline placenta is present in late gestation. Morphologically, the mature placentae of C. chalcides are among the most specialized to have been described in reptiles, reflecting the substantial maternal‐fetal nutrient transfer that occurs in this species.

Morphogenesis of placental membranes in the viviparous, placentotrophic lizard Chalcides chalcides (Squamata: Scincidae)

In the scincid lizard Chalcides chalcides, females ovulate small ova and supply most of the nutrients for development by placental means. The yolk is enveloped precocially by extraembryonic ectoderm and endoderm during the gastrula stage, establishing a simple bilaminar yolk sac placenta. The shell membrane begins to degenerate at this time, resulting in apposition of extraembryonic and maternal tissues. A true chorioplacenta has developed by the early pharyngula stage, as has a choriovitelline placenta and the first stages of an omphaloplacenta. Although the choriovitelline membrane disappears rapidly, the omphaloplacenta spreads to occupy the entire abembryonic pole. The yolk cleft is not confluent with the exocoelom, and no omphalallantoic placenta develops. By the limb‐bud stage, an allantoplacenta has been established, with a mesometrial placentome composed of interdigitating ridges of chorioallantois and uterine mucosa. The discovery of five distinct placental arrangements in this species, three of which are transitory and two of which have not previously been recorded in reptiles, emphasizes the need for accounts that specify ontogenetic stages and the precise identity and composition of squamate placental membranes. Contrary to previous interpretations, the pattern of extraembryonic membrane development in C. chalcides is evolutionarily conservative, despite the presence of a reduced yolk mass and cytological specializations for nutrient transfer. Our observations indicate that substantial placentotrophy can evolve in squamates without major modifications of morphogenetic patterns. J Morphol 232:35–55, 1997.

Reproduction in Viviparous South American Lizards of the Genus Mabuya

Reproduction in most vertebrates requires that the female construct an egg containing all of the nutrients needed to sustain development and that she deposit that egg in an environment where it can develop and hatch. Like the seed of a plant, a newly-laid egg can be viewed as a compact package of nutrients and energy, housed with detailed genetic intructions on their use, and provided with a protective covering. The success of egg-laying reproduction, i.e., “oviparity,” in terrestrial environments is revealed by its presence in such amniotes as birds, monotremes, turtles, tuataras, crocodilians, and most squamates (i.e., lizards, amphisbaenians, and snakes). Nevertheless, developing eggs that have been laid on land can be subject to the vicissitudes of the terrestrial environment—temperature extremes, thermal fluctuations, dehydration, flooding, and predation by animals, fungi, and bacteria that may be genetically programmed to exploit egg nutrients for their own growth and reproduction.

Nutritional Provision to Embryos in a Predominantly Lecithotrophic Placental Reptile, Thamnophis ordinoides (Squamata: Serpentes)

Quantitative analysis of the composition of eggs and their sibling neonates in the viviparous natricine snake Thamnophis ordinoides revealed that yolk provided the principal source of organic nutrition but that embryos received a substantial allotment of inorganic nutrients from the placentas. The placental provision of water and sodium equaled or exceeded yolk supplies, and placental transport accounted for 23% of neonatal calcium composition. There was no difference between egg and newborn quantities of total phosphorus or total potassium, whereas neonates contained less total magnesium than eggs. The mode of embryonic nutrition in this species is characterized as predominantly lecithotrophic, yet placental nutrient provision contributes significantly to embryonic nourishment. Placental transport of sodium and embryonic uptake of water was greater in recently ovulated eggs that contained relatively low levels of sodium and water respectively. Thus, placental sources compensated for low yolk provision. Placental transport of calcium was independent of yolk calcium content and correlated positively with neonatal calcium content. This pattern of provision, in which placental sources determine neonatal content independent of egg content, has been described as facultative placentotrophy. A similar embryonic nutritional pattern was recognized previously in another predominantly lecithotrophic natricine snake.