Cyanomargarita gen. nov. (Nostocales, Cyanobacteria): convergent evolution resulting in a cryptic genus.

Two populations of Rivularia-like cyanobacteria were isolated from ecologically distinct and biogeographically distant sites. One population was from an unpolluted stream in the Kola Peninsula of Russia, whereas the other was from a wet wall in the Grand Staircase-Escalante National Monument, a desert park-land in Utah. Though both were virtually indistinguishable from Rivularia in field and cultured material, they were both phylogenetically distant from Rivularia and the Rivulariaceae based on both 16S rRNA and rbcLX phylogenies. We here name the new cryptic genus Cyanomargarita gen. nov., with type species C. melechinii sp. nov., and additional species C. calcarea sp. nov. We also name a new family for these taxa, the Cyanomargaritaceae.

Variation in timing of ossification affects inferred heterochrony of cranial bones in Lissamphibia.

The evolutionary origin of Lissamphibia likely involved heterochrony, as demonstrated by the biphasic lifestyles of most extant orders, differences between Anura (with tadpole-to-froglet metamorphosis) and Urodela (which lack strongly defined metamorphosis), and the appearance of direct development among separate lineages of frogs. Patterns in the timing of appearance of skeletal elements (i.e., ossification sequence data) represent a possible source of information for understanding the origin of Lissamphibia, and with the advent of analytical methods to directly optimize these data onto known phylogenies, there has been a renewed interest in assessing the role of changes in these developmental events. However, little attention has been given to the potential impact of variation in ossification sequence data--this is particularly surprising given that different criteria for collecting these data have been employed. Herein, new and previously published ossification data are compiled and all pairs of data for same-species comparisons are selected. Analyses are run to assess the impact of using data that were collected by different methodologies: (1) wild- versus lab-raised animals; (2) different criteria for recognizing timing of ossification; and (3) randomly selecting ossification sequences for species from which multiple studies have been published, but for which the data were collected by different criteria. Parsimov-based genetic inference is utilized to map ossification sequence data onto an existing phylogeny to reconstruct ancestral sequences of ossification and infer instances of heterochrony. All analyses succeeded in optimizing sequence data on internal nodes and instances of heterochrony were identified. However, among all analyses little congruence was found in reconstructed ancestral sequences or among inferred instances of heterochrony. These results indicate a high degree of variation in timing of ossification, and suggest a cautionary note about use of these data, particularly given that in most instances issues associated with the original sources of data (e.g., wild- vs. lab-raised animals; or criteria for identification of earliest ossification) are not addressed. Potential sources of variation in the original data are discussed and may explain the incongruence observed here.

Ossification sequence heterochrony among amphibians.

Heterochrony is an important mechanism in the evolution of amphibians. Although studies have centered on the relationship between size and shape and the rates of development, ossification sequence heterochrony also may have been important. Rigorous, phylogenetic methods for assessing sequence heterochrony are relatively new, and a comprehensive study of the relative timing of ossification of skeletal elements has not been used to identify instances of sequence heterochrony across Amphibia. In this study, a new version of the program Parsimov-based genetic inference (PGi) was used to identify shifts in ossification sequences across all extant orders of amphibians, for all major structural units of the skeleton. PGi identified a number of heterochronic sequence shifts in all analyses, the most interesting of which seem to be tied to differences in metamorphic patterns among major clades. Early ossification of the vomer, premaxilla, and dentary is retained by Apateon caducus and members of Gymnophiona and Urodela, which lack the strongly biphasic development seen in anurans. In contrast, bones associated with the jaws and face were identified as shifting late in the ancestor of Anura. The bones that do not shift late, and thereby occupy the earliest positions in the anuran cranial sequence, are those in regions of the skull that undergo the least restructuring throughout anuran metamorphosis. Additionally, within Anura, bones of the hind limb and pelvic girdle were also identified as shifting early in the sequence of ossification, which may be a result of functional constraints imposed by the drastic metamorphosis of most anurans.

Formation and ossification of limb elements in Trachemys scripta and a discussion of autopodial elements in turtles.

Though sequences of formation and ossification of bony elements have been described for many taxa, controversy surrounds the formation of limb elements in turtles. Three hypotheses for patterns of formation of autopodial elements have been proposed, differing primarily in the origin of Distal Carpal/Tarsal 3, the digital arch, and Centrale 4. Patterns of formation and ossification of limb elements are described for Trachemys scripta. These patterns are compared to similar data for representatives of four families of turtles (Cheloniidae, Chelydridae, Emydidae, and Trionychidae). Hypotheses of limb formation are compared in the context of new and published data. Three species (Trachemys scripta, Chrysemys picta, and Chelydra serpentina) suggest that Distal Carpal 3 forms by branching from the ulnare, whereas Distal Carpal 3 may branch from Distal Carpal 4 in Macrochelys temminckii and Chelonia mydas; data from Graptemys nigrinoda, Apalone spinifera, and Eretmochelys imbricata did not provide evidence for the origin of Distal Carpal 3. Centrale 4 was not observed to branch from the ulnare and apparently arises by de-novo condensation. Distal Carpal 4 did not branch from Centrale 4 in any species. Until the developmental origins of Distal Carpal 3 and Centrale 4 are understood, interspecific variation in the origin of these elements remains, and may explain some of the observed differences. Trends of ossification in the fore- and hind limb autopodium also are summarized. Homology of elements in pedal Digit V is discussed, and we suggest that the hooked proximal element of this digit be recognized as Distal Tarsal 5.

Formation of the chondrocranium of Trachemys scripta (Reptilia: Testudines: Emydidae) and a comparison with other described turtle taxa.

Few descriptions of the formation of the chelonian chondrocranium exist. Herein, developmental stages critical to the formation of the chondrocranium of the Red-eared Slider, Trachemys scripta (Testudines: Emydidae), are described and illustrated, with particular attention given to ontogenetic changes that take place in the orbitotemporal region of the skull. Morphological descriptions are based on cleared and double-stained and serially-sectioned embryos. These specimens allowed for a detailed evaluation of the developmental morphology of the trabeculae, interorbital septum, pilae metoptica, taeniae marginalis, acrochordal cartilage, pilae antotica, parachordal cartilages, and crista sellaris. Additionally, the formation of the chondrocranium of T. scripta is compared to those of Chrysemys picta (Emydidae) and Caretta caretta (Chelonidae). Overall, the patterns of formation and remodeling of the chondrocranium are quite similar among these species, with the most conspicuous differences observed in remodeling of the posterior orbital cartilages (specifically, the pila metoptica). Reorganization of these cartilages is discussed briefly in the context of associated extraocular muscles for T. scripta and C. caretta. A prominent intertrabecula is reported in T. scripta, supporting previous observations of this structure in emydid turtles.

Patterns of chondrification and ossification in the skull of Graptemys pseudogeographica, the false map turtle (Emydidae).

Patterns of ossification and chondrification are well-described for several species of turtles, but details of the chondrocranial anatomy are known for only a handful of species. Cleared and double-stained embryos of Graptemys pseudogeographica were used to examine the fully formed chondrocranium and the formation, chondrification, and ossification of the cranium. The chondrocranium of G. pseudogeographica possesses an unusually large, irregularly shaped foramen epiphaniale that is joined with the fenestra olfactoria. As in other emydids, and many turtles generally, the taenia marginalis is present only as a small projection and the taenia medialis is lacking in mature stages of embryonic development. Ossification data for G. pseudogeographica are consistent with those of other Testudines in that the dentary and maxilla (dermal elements of the upper and lower jaws) ossify early, whereas the articular (an endochondral bone of the lower jaw) ossifies relatively late. Additionally, comparative ossification shows that the vomer is quite variable in its relative timing of ossification across Testudines.

Skeletal development of Macrochelys temminckii (Reptilia: Testudines: Chelydridae).

Few descriptions of the development and sequence of chondrification and ossification of the entire skeleton of turtles exist, particularly compared to other groups of reptiles. In this study, the embryonic skeleton and its ontogenesis are described for the Alligator Snapping Turtle, Macrochelys temminckii (Chelydridae). Morphological descriptions utilize cleared and double-stained embryonic specimens and form the basis of comparison of the ontogenesis of the skeleton between this species and its extant sister taxon, Chelydra serpentina. The embryonic chondrocranium, as well as the sequences of formation and ossification of the entire skeleton, are compared between these closely related species, and afford a unique opportunity to examine differences in their patterns of skeletal formation. In M. temminckii, the first elements to ossify (Stage 17) are associated with the dermatocranium and upper jaw, followed by elements of the palate, lower jaw, and long bones of the limbs. In both species the majority of endochondral braincase elements (prootic, opisthotic, supraoccipital, and exoccipital) ossify after the majority of dermal elements of the skull. The sequences of formation of the chondral primordia of the limb elements, as well as ossification of autopodial elements, are generally congruent between these species.

Osteology and skeletal development of Phyllomedusa vaillanti (Anura: Hylidae: Phyllomedusinae) and a comparison of this arboreal species with a terrestrial member of the genus.

Few descriptions of skeletal development and morphology exist for neobatrachians, despite their abundance and diversity. Herein, the adult morphologies of Phyllomedusa vaillanti and P. atelopoides are described and compared and the ontogeny of the larval skeleton of P. vaillanti is described and compared with those of Hyla lanciformis (the only hylid for which a detailed cranial and postcranial osteological ontogenesis has been described) and P. trinitatis (the only other member of this genus for which the larval skeleton has been described). These descriptions and comparisons are made on the basis of cleared and double-stained, dry skeletal, and alcohol-preserved specimens. In P. vaillanti, the first elements that ossify are the neural arches of the presacral vertebrae (Gosner Stage 34), followed by the parasphenoid, occipital condyles, exoccipitals, and prootics at Stage 38; many elements of the postcranial skeleton ossify contemporaneously with the first cranial elements. Major modifications of the chondrocranium begin at Stage 44. In adults, the skulls of P. vaillanti and P. atelopoides do not seem atypical of hylid frogs, and their elements are gracile and unornamented. Although P. atelopoides is a terrestrial species, the morphology of its hands and feet does not seem to differ dramatically from that of other phyllomedusines, which are arboreal; however, the relative lengths of the appendages and vertebral column are shorter and more robust than those of all other Phyllomedusa.

Osteology and skeletal development of Apalone spinifera (Reptilia: Testudines: Trionychidae).

Despite considerable attention that other groups of reptiles have received, few descriptions of the development and sequences of chondrification and ossification of the entire skeleton of turtles exist. Herein, the adult skeleton of the spiny softshell turtle, Apalone spinifera (Testudines: Trionychidae), is described; this description forms a basis of comparison for the embryonic skeleton and its ontogenesis. Descriptions are made on the basis of cleared and double-stained embryos and dry skeletal postembryonic specimens. The embryonic chondrocranium of A. spinifera is described and compared to those of Emys orbicularis and Caretta caretta, the sequence of chondrification of fore- and hindlimbs are compared with published descriptions of Chelydra serpentina and Chrysemys picta, and the sequence of ossification of elements is compared with those of C. serpentina, Lacerta vivipara, and Alligator mississippiensis. In A. spinifera, the first elements that ossify (Stage 17) are associated with the dermatocranium and mandible, followed by elements of the dermal skull table, lower jaw, and dermal elements of the plastron. In A. spinifera, the sequence of chondrification of limb elements is similar to that of C. serpentina; however, the sequence of ossification varies greatly among Apalone, Chelydra, Lacerta, and Alligator.

Skeletal development in the Chinese soft-shelled turtle Pelodiscus sinensis (Testudines: Trionychidae).

We investigated the development of the whole skeleton of the soft-shelled turtle Pelodiscus sinensis, with particular emphasis on the pattern and sequence of ossification. Ossification starts at late Tokita-Kuratani stage (TK) 18 with the maxilla, followed by the dentary and prefrontal. The quadrate is the first endoskeletal ossification and appears at TK stage 22. All adult skull elements have started ossification by TK stage 25. Plastral bones are the first postcranial bones to ossify, whereas the nuchal is the first carapacial bone to ossify, appearing as two unstained anlagen. Extensive examination of ossification sequences among autopodial elements reveals much intraspecific variation. Patterns of ossification of cranial dermal elements are more variable than those of endochondral elements, and dermal elements ossify before endochondral ones. Differences in ossification sequences with Apalone spinifera include: in Pelodiscus sinensis the jugal develops relatively early and before the frontal, whereas it appears later in A. spinifera; the frontal appears shortly before the parietal in A. spinifera whereas in P. sinensis the parietal appears several stages before the frontal. Chelydrids exhibit an early development of the postorbital bone and the palatal elements as compared to trionychids. Integration of the onset of ossification data into an analysis of the sequence of skeletal ossification in cryptodirans using the event-pairing and Parsimov methods reveals heterochronies, some of which reflect the hypothesized phylogeny considered taxa. A functional interpretation of heterochronies is speculative. In the chondrocranium there is no contact between the nasal capsules and planum supraseptale via the sphenethmoid commissurae. The pattern of chondrification of forelimb and hind limb elements is consistent with a primary axis and digital arch. There is no evidence of anterior condensations distal to the radius and tibia. A pattern of quasi- simultaneity is seen in the chondrogenesis of the forelimb and the hind limb.

Effects of temperature regime through premetamorphic ontogeny on shape of the chondrocranium in the American toad, Anaxyrus americanus.

If one considers the substantial amount of information that exists about phenotypic plasticity in amphibians, it is surprising that few studies have examined abiotic factors that influence phenotype through ontogeny. Phenotypic change and stability of morphology are artifacts of organisms that bear significant relevance to evolution within and among taxonomic groups. Here, we examine development as a phenotypically plastic aspect of larval anurans. Fertilized eggs of the American Toad, Anaxyrus (= Bufo) americanus (Holbrook, 1836), were obtained from two pairs of adults, and larvae were reared in four temperature treatments (constant Mean, constant High, constant Low, and Fluctuating regime [Low night-High day]); developmental series were collected from each treatment, representing larvae of this species from Gosner Stages 28-40. Cleared and stained larvae were analyzed with landmark-based geometric morphometric methods to facilitate examination of differences in overall shape change of the larval chondrocranium through ontogeny, as a result of developmental temperature or temperature regime. Changes in shape of the chondrocranium and in amount and direction of phenotypic change through ontogeny were found in response to temperature treatment and temperature regime. Mean chondrocranial shape of the Fluctuating regime was more similar to the consensus shape of the overall data set than were those of all other treatments. Given that differences in amount and direction of shape change were observed among these treatments and throughout ontogeny, one should consider the affects of abiotic factors (such as temperature) when rearing larval anurans for studies of developmental morphology.

Osteology and skeletal development of Pyxicephalus adspersus (Anura: Ranidae: Raninae).

Despite the abundance and diversity of neobatrachians, relatively few descriptions exist of their ontogenesis and skeletal development. Herein, the adult and larval skeleton and the ontogenesis of the skeleton of the African frog, Pyxicephalus adspersus (Ranidae: Raninae), are described on the basis of cleared and double-stained, dry, and alcohol-preserved specimens. In P. adspersus, the first elements that ossify are the neural arches of the pre- and postsacral vertebrae (Gosner Stage 35), followed by the parasphenoid, frontoparietals, and exoccipitals (beginning at Stage 36). Major modifications of the chondrocranium begin at approximately Stage 40. The skull of the adult P. adspersus is exostosed and hyperossified, with many of the dermal bones of the cranium fused and highly ornamented. The osteology of the adult P. adspersus is compared to the skeletal morphologies of Conraua alleni (Raninae), which represents an African ranine for which osteological descriptions exist, and Ceratophrys cornuta (Leptodactylidae), which represents a hyperossified anuran with convergent morphology and behavior. The larval morphology and development of P. adspersus is compared to those of Rana pipiens and R. temporaria, which have been described in relative detail. The sequence of ossification of elements is compared with those of R. pipiens and R. temporaria. This study includes a discussion of the morphology of the elements forming the orbit, as well as a survey of the distribution of several characters related to dentary tusks of other Asian and African ranids. J. Morphol. 240:49-75, 1999. © 1999 Wiley-Liss, Inc.