Ultrastructure of the reproductive system of the black swamp snake (Seminatrix pygaea). VI. Anterior testicular ducts and their nomenclature

In this study, the anterior testicular ducts of the North American natricine snake Seminatrix pygaea are described using light and electron microscopy. From the seminiferous tubules, the rete testis passes into the epididymal sheath, a structure along the medial border of the testis heavily invested with collagen fibers. The rete testis consists of simple, nonciliated cuboidal epithelium (principal cells). The intratesticular ducts of the rete testis are narrow (50–70 μm) at their junction with the seminiferous tubules, widen (80–100 μm) as they extend extratesticularly, and divide into smaller branches as they anastomose with the next tubules, the ductuli efferentes. The ductuli efferentes are lined by simple cuboidal epithelium but possess nonciliated principal cells as well as ciliated cells. These are the only ducts in the male reproductive system with ciliated cells. The ductuli efferentes are narrow (25–45 μm), divide into numerous branches, and are highly convoluted. The ductus epididymis is the largest duct in diameter (240–330 μm), and the diameter widens and the epithelium thins posteriorly. The ductus epididymis is lined by nonciliated, columnar principal cells and basal cells. No regional differences in the ductus epididymis are apparent. Ultrastructural evidence suggests that all of the nonciliated principal cells in each of the anterior testicular ducts function in both absorption and secretion. Absorption occurs via small endocytic vesicles, some of which appear coated. Secretion is by a constitutive pathway in which small vesicles and a flocculent material are released via a merocrine process or through the formation of apocrine blebs. The secretory product is a glycoprotein. Overall, the characteristics of the anterior testicular ducts of this snake are concordant with those of other amniotes, and the traditional names used for snakes are changed to conform with those used for other sauropsids and mammals. J. Morphol., 2010. © 2009 Wiley‐Liss, Inc.     

The comparative morphology of the larynx in snakes

Young, B.A. 2000. The comparative morphology of the larynx in snakes. —Acta Zoologica (Stockholm) 81 : 177–193 The larynx and glottal tube were examined in 10 specimens each of 22 snake species. Qualitative analysis through dissection and clearing and staining revealed distinct morphological variation in the cartilage and intrinsic musculature of the larynx. Quantitative analysis of nine morphometric characters revealed laryngeal sexual dimorphism in three species, significant interspecific differences in the coefficients for every morphometric feature regressed on body size, no significant intraspecific differences in the log transformed means, and poor discrimination of species using principal component analysis. The observed variations in laryngeal morphology did not correlate with the phylogeny of the species, or with habitat preferences.     

The anatomy of the upper cretaceous snake Najash rionegrina Apesteguía & Zaher, 2006, and the evolution of limblessness in snakes

Najash rionegrina Apesteguía & Zaher, 2006 , a terrestrial fossil snake from the Upper Cretaceous of Argentina, represents the first known snake with a sacrum associated with robust, well‐developed hind limbs. Najash rionegrina documents an important gap in the evolutionary development towards limblessness, because its phylogenetic affinities suggest that it is the sister group of all modern snakes, including the limbed Tethyan snakes Pachyrhachis, Haasiophis, and Eupodophis. The latter three limbed marine fossil snakes are shown to be more derived morphologically, because they lack a sacrum, but have articulated lymphapophyses, and their appendicular skeleton is enclosed by the rib cage, as in modern snakes.     

Semen collection and evaluation in free‐ranging Brazilian rattlesnakes (Crotalus durissus terrificus)

Two hundred‐ninety species of reptiles are estimated to need urgent action for conservation, with at least 113 threatened species worldwide. The International Union for Conservation of Nature and Natural Resources (IUCN) Red List of Threatened Species includes 80 species of snakes, with six native Brazilian species, a number likely to be an underestimation. Some authors believe that assisted reproduction would be an important tool to improve reproduction in captivity of some reptiles. An efficient technique for semen collection and evaluation is an important step in development of protocols for cryopreservation of semen or artificial insemination in snakes, contributing to the conservation of endangered species. Although these techniques are important, some basic semen parameters are described for four of the ～2,900 snake species in the world. The Brazilian rattlesnake (Crotalus durissus terrificus) was chosen as a model for semen collection in snakes because it is found quite often in Sao Paulo State. Semen was collected once from each animal by the same investigator during the mating season of this species in Brazil. After antiseptic cleansing of the skin around the cloaca, the snakes were injected subcutaneously with a dose of 15 mg/kg of 1% solution of lidocaine around the cloaca. Semen was collected with ventral massages after cloacal relaxation and directly from genital papilla inside the cloaca. A total of 28 ejaculates from 39 animals were obtained, representing collection efficiency of 71.80%. Semen volume and concentration in Brazilian rattlesnakes ranged from 3–70 µl and from 0.94–2.23 × 10⁹ spermatozoa/ml, respectively. Zoo Biol 0:1–6, 2007. © 2007 Wiley‐Liss, Inc.     

Sperm aggregations in female Agkistrodon piscivorus (Reptilia:Squamata): a histological and ultrastructural investigation

Upon copulation in female Agkistrodon piscivorus, sperm migrate up the oviduct to sperm storage tubules (SSTs) in the posterior infundibulum. The epithelium of the SSTs is composed of ciliated and secretory cells and differs ultrastructurally from that of the epithelium lining the lumen of the posterior infundibulum. Sperm pass through an area composed primarily of ciliated cells at the opening of each gland before aligning themselves in parallel arrays with their nuclei facing an area composed primarily of secretory cells at the base of the tubules. Sperm are also found embedded inter- and intracellularly in the SSTs. The secretory vacuoles in the SSTs become highly electron dense after the start of the fall mating season along with the synthesis of lipid droplets. Histochemical analysis reveals that the alteration in secretory material density is caused by the production of neutral carbohydrates. Some sperm remain in aggregates in the nonglandular section of the posterior uterus until the time of ovulation. However, ultrastructural evidence indicates these sperm degrade before ovulation. Therefore, sperm in posterior aggregates have no role in fertilization of ovulated ova. The data presented here support the hypothesis that infundibular sperm storage is the mode that snakes utilize to sequester viable sperm until ovulation.     

Higher-level relationships of snakes inferred from four nuclear and mitochondrial genes

Higher-level snake relationships are inferred from sequence analyses of one nuclear gene (C-mos) and three mitochondrial genes (12S rRNA, 16S rRNA and cytochrome b). Extant snakes belong to two lineages: the fossorial Scolecophidia, which feed on small prey on a frequent basis, and the ecologically diverse Alethinophidia ('typical' snakes), which feed on large prey on an infrequent basis. The vast majority of Alethinophidia, if not all of them, belong to two clades, corresponding to two distinct prey neutralization modes: unimodal constriction for the Henophidia (locomotor and feeding systems coupled) and injection of toxic saliva, in addition (or not) to diverse alternate modes of constriction, for the Caenophidia (locomotor and feeding systems uncoupled). Within Alethinophidia, non-macrostomatan (small gape) Aniliidae (genus Anilius) and macrostomatan (large gape) Tropidophiidae (genera Trachyboa and Tropidophis), both from the Neotropics, are closest relatives. Although our data are insufficient to robustly infer the ancestral mode of life of snakes, we find evidence of plasticity in the basic ecological and trophic modes of snakes. Consequently, the macrostomatan condition should not be treated a priori as a derived character state devoid of homoplasy.     

Skeletal heterochrony is associated with the anatomical specializations of snakes among squamate reptiles

Snakes possess a derived anatomy, characterized by limb reduction and reorganization of the skull and internal organs. To understand the origin of snakes from an ontogenetic point of view, we conducted comprehensive investigations on the timing of skeletal elements, based on published and new data, and reconstructed the evolution of the ossification sequence among squamates. We included for the first time Varanus, a critical taxon in phylogenetic context. There is comprehensive delay in the onset of ossification of most skeletal elements in snakes when compared to reference developmental events through evolution. We hypothesize that progressing deceleration accompanied limb reduction and reorganization of the snake skull. Molecular and morphological studies have suggested close relationship of snakes to either amphisbaenians, scincids, geckos, iguanids, or varanids. Likewise, alternative hypotheses on habitat for stem snakes have been postulated. Our comprehensive heterochrony analyses detected developmental shifts in ossification for each hypothesis of snake origin. Moreover, we show that reconstruction of ancestral developmental sequences is a valuable tool to understand ontogenetic mechanisms associated with major evolutionary changes and test homology hypotheses. The “supratemporal” of snakes could be homolog to squamosal of other squamates, which starts ossification early to become relatively large in snakes.     

Morphology and evolution of the snake cornea

To investigate whether the thickness of the cornea in snakes correlates with overall anatomy, habitat or daily activity pattern, we measured corneal thickness using optical coherence tomography scanning in 44 species from 14 families (214 specimens) in the collection at the Natural History Museum (Denmark). Specifically, we analyzed whether the thickness of the cornea varies among species in absolute terms and relative to morphometrics, such as body length, spectacle diameter, and spectacle thickness. Furthermore, we examined whether corneal thickness reflects adaptation to different habitats and/or daily activity patterns. The snakes were defined as arboreal (n = 8), terrestrial (n = 22), fossorial (n = 7), and aquatic (n = 7); 14 species were classified as diurnal and 30 as nocturnal. We reveal that the interspecific variation in corneal thickness is largely explained by differences in body size, but find a tendency towards thicker corneas in diurnal (313 ± 227 μm) compared to nocturnal species (205 ± 169 μm). Furthermore, arboreal snakes had the thickest corneas and fossorial snakes the thinnest. Our study shows that body length, habitat, and daily activity pattern could explain the interspecific variation in corneal morphology among snakes. This study provides a quantitative analysis of the evolution of the corneal morphology in snakes, and it presents baseline values of corneal thickness of multiple snake species. We speculate that the cornea likely plays a role in snake vision, despite the fact that results from previous studies suggest that the cornea in snakes is not relevant for vision (Sivak, Vision Research, 1977, 17, 293–298).     

Snake phylogeny based on osteology,soft anatomy and ecology

Relationships between the major lineages of snakes are assessed based on a phylogenetic analysis of the most extensive phenotypic data set to date (212 osteological, 48 soft anatomical, and three ecological characters). The marine, limbed Cretaceous snakes Pachyrhachis and Haasiophis emerge as the most primitive snakes: characters proposed to unite them with advanced snakes (macrostomatans) are based on unlikely interpretations of contentious elements or are highly variable within snakes. Other basal snakes include madtsoiids and Dinilysia--both large, presumably non-burrowing forms. The inferred relationships within extant snakes are broadly similar to currently accepted views, with scolecophidians (blindsnakes) being the most basal living forms, followed by anilioids (pipesnakes), booids and booid-like groups, acrochordids (filesnakes), and finally colubroids. Important new conclusions include strong support for the monophyly of large constricting snakes (erycines, boines. pythonines), and moderate support for the non-monophyly of the trophidophiids' (dwarf boas). These phylogenetic results are obtained whether varanoid lizards, or amphisbaenians and dibamids, are assumed to be the nearest relatives (outgroups) of snakes, and whether multistate characters are treated as ordered or unordered. Identification of large marine forms, and large surface-active terrestrial forms, as the most primitive snakes contradicts with the widespread view that snakes arose via minute, burrowing ancestors. Furthermore, these basal fossil snakes all have long flexible jaw elements adapted for ingesting large prey ('macrostomy'), suggesting that large gape was primitive for snakes and secondarily reduced in the most basal living foms (scolecophidians and anilioids) in connection with burrowing. This challenges the widespread view that snake evolution has involved progressive, directional elaboration of the jaw apparatus to feed on larger prey.     

Seasonal variation in the oviduct of female Agkistrodon piscivorus (Reptilia:Squamata): An ultrastructural investigation

The annual oviductal cycle of the Cottonmouth, Agkistrodon piscivorus, is described using electron microscopy. This is only the second such study on a snake and the first on a viperid species. Specimens were collected in reproductive and nonreproductive condition throughout the year and five ultrastructurally unique regions were recognized: the anterior infundibulum, posterior infundibulum, glandular uterus, nonglandular uterus, and vagina. Except for the anterior infundibulum and vagina, which exhibit no seasonal variation in ultrastructure, the oviduct becomes highly secretory at the start of vitellogenesis. This includes the entire luminal border of the uterus, the tubular glands of the glandular uterus, and the luminal border and sperm storage tubules of the posterior infundibulum. The secretory materials produced in the oviduct vary among regions of the oviduct, and also can vary among time periods in the same region of the oviduct. Variation is especially evident in the sperm storage tubules. Secretory activity in the sperm storage tubules ceases after ovulation, but the tubular glands of the glandular uterus remain secretory until parturition, at which time secretory activity in the varying sections of the oviduct decreases dramatically. After parturition, the oviduct remains in a dormant state until the next reproductive season. The seasonal variation in oviducal morphology mirrors the temperate primitive reproductive cycle known for some pitvipers. Uterine glands of A. piscivorous are more similar in secretory activity to those of an oviparous lizard than a viviparous colubrid snake, suggesting variation in uterine gland morphology between snakes of different families. J. Morphol., 2008. © 2008 Wiley‐Liss, Inc.