Frog Development Research Articles

Musashi-1-A Stemness RBP for Cancer Therapy?

Simple SummaryMusashi–1 (MSI1) is an RNA–binding protein that promotes stemness properties. It was initially discovered as a regulator of neuronal development and oocyte maturation in flies and frogs. Due to its specific expression pattern with high levels during development and in a variety of cancers, MSI1 evolved as an interesting target for cancer therapy. In cancer cells, the protein mainly promotes an undifferentiated state enhancing cancer growth and therapy resistance. In this review, we summarize previous findings from development of other organisms, outline MSI1′s expression and function in different cancer entities and highlight the development of MSI1–directed inhibitors.The RNA–binding protein Musashi–1 (MSI1) promotes stemness during development and cancer. By controlling target mRNA turnover and translation, MSI1 is implicated in the regulation of cancer hallmarks such as cell cycle or Notch signaling. Thereby, the protein enhanced cancer growth and therapy resistance to standard regimes. Due to its specific expression pattern and diverse functions, MSI1 represents an interesting target for cancer therapy in the future. In this review we summarize previous findings on MSI1′s implications in developmental processes of other organisms. We revisit MSI1′s expression in a set of solid cancers, describe mechanistic details and implications in MSI1 associated cancer hallmark pathways and highlight current research in drug development identifying the first MSI1–directed inhibitors with anti–tumor activity.

Cooperation behavior of fore- And hindlimbs during jumping in Rana dybowskii and Xenopus laevis.

Frogs are characterized by their outstanding jumping ability, depending on the rapid extension of hindlimbs to propel their bodies into air. A typical jumping cycle could be broken into four phases: preparation, takeoff, flight, and landing. Considerable research has been performed to discuss the function of hindlimbs of frogs during takeoff phase, whereas the literature of limbs' motion in jumping between different species was limited. To profile the evolution of locomotion in anurans, it is necessary to investigate on the motion of fore‐ and hindlimbs of frogs within different taxa. In this work, we put forward a detailed description of jumping behavior of two frog species, Rana dybowskii and Xenopus laevis. High‐speed cameras were used to explore the movement of different joints in fore‐ and hindlimbs of these two animals, and kinematic analysis was operated to identify both homologous behaviors and significant differences between them. We found that the Rana dybowskii's fore‐ and hindlimbs had good cooperation during jumping, while the Xenopus laevis' uncooperative behavior in limbs may give a functional explanation for the deficiency in terrestrial jumping; besides, the R. dybowskii's landing followed the “hands‐belly‐feet slap” strategy, and Xenopus laevis had clumsy landing with “belly‐flops” sequence. The result gained here clarifies the cooperation behavior of anuran limbs and may supply a new insight into our understanding of the anuran's evolution.

Carryover effects minimized the positive effects of treated wastewater on anuran development

Constructed wetlands (CWs) are a potential solution for wastewater treatment due to their capacity to support native species and provide tertiary wastewater treatment. However, CWs can expose wildlife communities to excess nutrients and harmful contaminants, affecting their development, morphology, and behavior. To examine how wastewater CWs may affect wildlife, we raised Southern leopard frogs, Lithobates sphenocephalus, in wastewater from conventional secondary lagoon and tertiary CW treatments for comparison with pondwater along with the presence and absence of a common plant invader to these systems – common duckweed (Lemna minor) - and monitored their juvenile development for potential carryover effects into the terrestrial environment. The tertiary CW treatment did not change demographic or morphological outcomes relative to conventional wastewater treatment in our study. Individuals emerging from both wastewater treatments demonstrated lower terrestrial survival rates than those emerging from pondwater throughout the experiment though experiment-wide survival rates were equivalent among treatments. Individuals from wastewater treatments transformed at larger sizes relative to those in pondwater, but this advantage was minimized in the terrestrial environment. Individuals that developed with duckweed had consistent but marginally better performance in both environments. Our results suggest a potential trade-off between short-term benefits of development in treated effluent and long-term consequences on overall fitness. Overall, we demonstrate that CWs for the purpose of wastewater treatment may not be suitable replicates for wildlife habitat and could have consequences for local population dynamics.

Body Size Is Related to Temperature Preference in Hyla chrysoscelis Tadpoles

Temperature plays a critical role in growth and development of anurans. Here, we investigated thermal preference of tadpoles from a wide-ranging anuran, Hyla chrysoscelis, by establishing an ex situ thermal gradient in a laboratory setting. We collected 64 tadpoles with a mean (±SD) body length of 12.2 ± 2.1 mm (range, 7–15 mm). Mean temperature preference of tadpoles ranged from 12.9 to 30.0°C, and we found a positive relationship between temperature preference and body length. Although body length is only one component of amphibian development, its apparent link with temperature preference is an exciting area for further research in this species as well as other species. Furthermore, given the broad distribution of H. chrysoscelis, there is an opportunity to investigate how thermal preferences could limit population edges and how populations may respond to shifts in environmental temperatures.

Temperature-mediated trade-off between development and performance in larval wood frogs (Rana sylvatica).

Countergradient variation has been detected in diverse taxa. In a common manifestation, individuals from colder environments develop faster than conspecifics from warmer environments when placed in a common garden. Where such a pattern exists, it implies a trade-off: Individuals from warmer environments have intrinsic rates of development lower than those demonstrated by other individuals of the same species. We explored a trade-off between development rate and locomotor performance in the wood frog (Rana sylvatica), an amphibian for which countergradient variation has been well documented. We reared wood frogs from 10 populations under two temperature regimes, bracketing the temperatures observed in local natural ponds. Individuals reared under warmer conditions developed more rapidly but exhibited burst speeds 20% lower than individuals reared under colder conditions. The slope of the reaction norm was consistent across the 10 populations and thus, we found no evidence of countergradient variation in performance. Burst speed assays of wild-caught tadpoles from the same populations showed a similar but nonsignificant trend, with greater variability among ponds. Overall, our findings support the existence of a development-performance trade-off that may be of broad importance and which may help explain the widespread occurrence of countergradient variation.

Pattern of Gonadal Sex Differentiation in the Rice Field Frog Hoplobatrachus rugulosus (Anura: Dicroglossidae).

Sex differentiation during gonadal development is diversified among anuran amphibian species. In this study, the anuran experimental species Hoplobatrachus rugulosus was examined. The pattern of gonadal sex differentiation was observed by morphological and histological approaches. The gonad was observed morphologically at Gosner stage 33, while distinct testis and ovary were evident from 3-4 weeks after metamorphosis ended. Histological analysis showed that genital ridge formation began at stage 25 and ovarian differentiation began at stage 36. The developing ovary appeared with numerous primary oogonia, which developed into oocytes, while the medulla regressed to form an ovarian cavity. During metamorphosis, only an ovary was observed. Testicular differentiation seemed to begin later, during the first week after metamorphosis, and occurred via an intersex condition. The intersex gonads contained developing testicular tissue with both normal and atretic oocytes. The fully developed testis was first identified at 6 weeks after metamorphosis. Comparing the times of gonadal differentiation and somatic development revealed that the ovary exhibited a basic rate of differentiation while the testis exhibited a retarded one. These results establish that males of this species develop later than do females, and the testis develops through an intersex gonad, as is evident from its seminiferous cord formation, the presence of testis-ova, and atretic oocytes in the tissue. Thus, the pattern of gonadal sex differentiation in H. rugulosus is an undifferentiated type, in which only female gonads are observed during metamorphosis and intersex and male gonads are observed later. These results are crucial for further research on the sexual development of anurans.

Differential effect of natural and pigment‐supplemented diets on larval development and phenotype of anurans

AbstractDiet has major effects on development, growth, health, and performance of animals. Photosynthetic pigments, in particular, are key components of the diet of many organisms that can play important roles in their development and performance. Compared to other taxa, we know relatively little about how these pigments impact amphibian larval development. We conducted diet supplementation experiments on tadpoles of two amphibian species, in which we offered various naturally co‐occurring aquatic macrophytes or diet enriched or not with supplement pigments. We evaluated the effect of these diets on developmental period, size, mass, color, and jumping performance of metamorphs exposed or not to risk of simulated pond drying during the larval period. Diet based on Myriophyllum alterniflorum resulted in high mortality rates of Mediterranean tree frog larvae (Hyla meridionalis), this macrophyte did not affect survival of Western spadefoot toads (Pelobates cultripes), but individuals took longer to reach metamorphosis. Regardless of the macrophyte consumed, spadefoot toad tadpoles speeded up metamorphosis when exposed to reduced water volume, whereas tree frog tadpoles did not show significant responses to simulated pond drying. The three supplemented diets tested—β‐carotene, lutein, and chlorophyll—had positive effects on size and mass of tree frogs; supplemented diet based on β‐carotene improved dorsal coloration of tree frogs and had positive effects on size and mass at metamorphosis of spadefoot toads. Tadpoles fed with supplements did not exhibit significant response to water reduction. Our study highlights the importance of larval diet on phenotypically plastic responses and post‐metamorphic traits in anurans.

Experiences with the marsupial frogs: reminiscences of a developmental biologist.

This article provides a brief account of the career of Eugenia M. del Pino. Casual events and serendipity played important roles in modeling her career as a developmental biologist. In collaboration with colleagues and students, she analyzed the biology and development of the marsupial frog Gastrotheca riobambae (family: Hemiphractidae) in comparison with Xenopus laevis and tropical frogs. The emphasis was placed on oogenesis and the early stages of development. Topics include the mono- and multi-nucleated modes of oogenesis. She described two modes of gastrulation in frogs, gastrulation modes one and two, according to the timing of notochord elongation. She was able to establish a pioneer laboratory for the comparative analysis of frog development in Ibero America at the Pontifical Catholic University of Ecuador, in Quito. Her contributions to society include her influence in the establishment of the National Academy of Sciences of Ecuador, and efforts toward the conservation of the Galápagos Archipelago. She is part of a pioneer group of professors that placed Biology as an academic discipline in Ecuador. The experiences of her career reveal that we all face difficulties in our jobs. However, nothing is impossible when we follow a passion. Her work reveals that the key to success is to turn obstacles into opportunities.

Effects of Environmental Contaminants at Great Bay National Wildlife Refuge on Anuran Development, Gonadal Histology, and Reproductive Steroidogenesis: A Comparison of In Situ and Laboratory Exposures.

Previous monitoring at Great Bay National Wildlife Refuge (NWR), Newington, New Hampshire documented high prevalence of amphibian malformations at sites contaminated with potential endocrine active compounds. In the present study, a combination of in situ and laboratory experiments were used to determine whether contaminants present in the sites affect amphibian growth and reproductive development. Wood frog (Rana sylvatica) tadpoles were exposed in situ at four sites (Ferry Way, Beaver Pond, Lower Peverly, and Stubbs Pond) at Great Bay NWR and northern leopard frog (Rana pipiens) tadpoles were exposed in the lab to sediments collected from three sites (Beaver Pond, Ferry Way, Stubbs Pond) at Great Bay NWR as well as a positive (estradiol) and negative control. High mortality was observed at Stubbs Pond and extended larval period at Beaver Pond in the in situ exposure. Only three malformations were noted in the lab experiment, whereas there was a 63% prevalence of rounded femurs in Beaver Pond metamorphs in the in situ exposure. Only 2.4% (5 of 207) of R. sylvatica metamorphs exhibited abnormal reproductive development, whereas intersex metamorphs occurred in treatments and controls in the lab experiment at rates as high as 26%. Reproductive development was more advanced and estradiol to androgen ratios reduced in male metamorphs from Beaver Pond in both the in situ and lab exposures. DDT, PCBs, and PAHs were detected in sediments at Great Bay NWR at concentrations that exceed regulatory or guidance values, with concentrations of PAHs being highest at Lower Peverly Pond and DDT highest at Stubbs Pond. The effects on anuran development may be attributable to the primary contaminants-DDT and PCBs-acting on the thyroid and gonadal axes.

Evolution of the Unique Anuran Pelvic and Hind limb Skeleton in Relation to Microhabitat, Locomotor Mode, and Jump Performance.

Anurans (frogs and toads) have a unique pelvic and hind limb skeleton among tetrapods. Although their distinct body plan is primarily associated with saltation, anuran species vary in their primary locomotor mode (e.g., walkers, hoppers, jumpers, and swimmers) and are found in a wide array of microhabitats (e.g., burrowing, terrestrial, arboreal, and aquatic) with varying functional demands. Given their largely conserved body plan, morphological adaptation to these diverse niches likely results from more fine-scale morphological change. Our study determines how shape differences in Anura's unique pelvic and hind limb skeletal structures vary with microhabitat, locomotor mode, and jumping ability. Using microCT scans of preserved specimens from museum collections, we added 3D landmarks to the pelvic and hind limb skeleton of 230 anuran species. In addition, we compiled microhabitat and locomotor data from the literature for these species that span 52 of the 55 families of frogs and ∼210 million years of anuran evolution. Using this robust dataset, we examine the relationship between pelvic and hind limb morphology and phylogenetic history, allometry, microhabitat, and locomotor mode. We find pelvic and hind limb changes associated with shifts in microhabitat ("ecomorphs") and locomotor mode ("locomorphs") and directly relate those morphological changes to the jumping ability of individual species. We also reveal how individual bones vary in evolutionary rate and their association with phylogeny, body size, microhabitat, and locomotor mode. Our findings uncover previously undocumented morphological variation related to anuran ecological and locomotor diversification and link that variation to differences in jumping ability among species.

Effect of temperature fluctuations on larval function of the Baja California chorus frog, Pseudacris hypochondriaca

Environmental temperature has an extensive impact on the phenotypes of all animals.Temperature fluctuations, often experienced naturally across daily or longer cycles, may cause profound effects on physiological phenotypes of animals, particularly during development and in ectotherms like amphibians. This study tested this hypothesis by evaluating the effects of temperature fluctuations during the larval development of the eurythermal Baja California chorus frog (Pseudacris hypochondriaca), a species native to southern California and northern Baja California. Eggs were collected from ponds in San Marcos, California within 24 hours of fertilization and separated into four temperature treatments: 15°C constant, and fluctuating treatments over 24 hours of 12.5°C – 17.5°C, 10°C – 20°C, and 15°C – 25°C. Oxygen consumption rate (V̇o2), critical thermal maximum (CTMAX), and swim speed were measured for tadpoles at 15°C during hindlimb toe development. Relative to mass‐specific V̇o2 in 15°C constant tadpoles, mass‐specific V̇o2 measured at 15°C showed a slight increase in 12.5°C – 17.5°C tadpoles, a larger increase in 10°C – 20°C tadpoles, but no difference in 15°C – 25°C tadpoles. Relative to CTMAX in 15°C constant tadpoles, CTMAX was similar in 12.5°C – 17.5°C tadpoles, slightly higher in 10°C – 20°C tadpoles, and greatly increased in 15°C – 25°C tadpoles. Swim speed measured at 15°C was similar across all temperature treatments. Overall, greater fluctuations had stronger effects on larval V̇o2 and CTMAX, and tadpoles tend to acclimate to higher temperatures during daily cycles. Acclimation in thermal tolerance and metabolic rates of this eurythermal amphibian indicates the animals will likely show plasticity in some traits in response to fluctuating temperatures. Future studies will examine these traits across a range of acute measurement temperatures to fully understand the functional implications of fluctuating temperatures.

Exposure during embryonic development to Roundup® Power 2.0 affects lateralization, level of activity and growth, but not defensive behaviour of marsh frog tadpoles

As glyphosate-based herbicides, sold under the commercial name Roundup®, represent the most used herbicides in the world, contamination of the freshwater environment by glyphosate has become a widespread issue. In Italy, glyphosate was detected in half of the surface waters monitoring sites and its concentrations were higher than environmental quality standards in 24.5% of them. It can last from days to months in water, leading to exposure for aquatic organisms and specifically to amphibians’ larvae that develop in shallow water bodies with proven effects to development and behaviour. In this study, we tested the effects of a 96 h exposure during embryonic development of marsh frog’s tadpoles to three ecologically relevant Roundup® Power 2.0 concentrations. As expected, given the low concentrations tested, no mortality was observed. Morphological measurements highlighted a reduction in the total length in tadpoles exposed to 7.6 mg a.e./L, while an increase was observed at lower concentrations of 0.7 and 3.1 mg a.e./L compared to control group. Tadpoles raised in 7.6 mg a.e./L also showed a smaller tail membrane than those raised in the control solution. Regarding behaviour, we tested tadpoles in two different sessions (Gosner stages 25 and 28/29) for lateralization, antipredator response and basal activity. Lower intensity of lateralization was detected in tadpoles raised at the highest Roundup® concentration in the first session of observation, while no significant difference among treatments was observed in the second one. In both sessions, effects of Roundup® Power 2.0 embryonic exposure on antipredator response, measured as the proportional change in activity after the injection of tadpole-fed predator (Anax imperator) cue, were not detected. Tadpoles exposed during embryonic development to Roundup® exhibited lower basal activity than the control group, with the strongest reduction for the 7.6 mg a.e./L treatment. Our results reinforce the concern of Roundup® contamination impact on amphibians.

Effects of two little-studied environmental pollutants on early development in anurans

Despite intensive ecotoxicological research, we still know relatively little about the ecological impacts of many environmental contaminants. Filling these knowledge gaps is particularly important regarding amphibians, because they play significant roles in freshwater and terrestrial ecosystems, and their populations are declining worldwide. In this study, we investigated two pollutants that have been poorly studied in ecotoxicology despite their widespread occurrence in surface waters: the herbicide terbuthylazine and the pharmaceutical drug carbamazepine. We exposed two anuran species throughout their larval development to each of two environmentally relevant concentrations of each pollutant, and recorded mortality and 17 sub-lethal endpoints up to several months after exposure. Mortality was low and unrelated to treatment. In agile frogs (Rana dalmatina), we found that treatment with 0.3 μg/L terbuthylazine decreased tadpole activity and reduced fat bodies in juveniles, whereas treatment with 50 μg/L carbamazepine decreased spleen size and increased spleen pigmentation. In common toads (Bufo bufo), treatment with 0.003 μg/L terbuthylazine increased body mass at metamorphosis, treatment with 0.3 μg/L terbuthylazine increased the size of optic tecta, and treatment with 0.5 μg/L carbamazepine decreased hypothalamus size. Treatment with 50 μg/L carbamazepine reduced the feeding activity of toad tadpoles, decreased their production of anti-predatory bufadienolide toxins, and increased their body mass at metamorphosis; juvenile toads in this treatment group had reduced spleen pigmentation. Neither treatments affected the time to metamorphosis, post-metamorphic body mass, or sex ratios significantly. These results show that environmental levels of both terbuthylazine and carbamazepine can have several sub-lethal effects on anurans, which may be detrimental to individual fitness and population persistence in natural conditions. Our findings further highlight that toxic effects cannot be generalized between chemicals of similar structure, because the terbuthylazine effects we found do not conform with previously reported effects of atrazine, a related and extensively studied herbicide.

Introduction of Developmental Biology at Utkal University, (Odisha, India).

The paper deals with the background and the establishment of a Developmental Biology Laboratory in Utkal University in Odisha state. It describes the process from a humble beginning with limited facilities into a leading research centre, initially for amphibians and later for the endangered olive ridley (Lepidochelys olivacea) turtle. Starting from the biology, reproduction and development in many anurans, the laboratory took up research on regeneration, especially on super-regeneration in tadpoles under the influence of morphogens such as vitamin A (retinoids). Treatment with vitamin A after amputation of the tail inhibited tail regeneration but unexpectedly induced homeotic transformation of tails into limbs in many anurans, starting with the marbled balloon frog Uperodon systoma. This was the first observation of homeotic transformation in any vertebrate. The laboratory continues research on histological and molecular aspects of this phenomenon. In addition, taking advantage of the largest rookery of olive ridley sea turtles in Gahirmatha, in the same state the laboratory has contributed significantly to the biology, breeding patterns, development and especially the temperature-dependent sex determination phenomenon (TSD). This research was extended to biochemical and ultrastructural aspects during development for the first time for any sea turtle. The laboratory has contributed significantly to the conservation of olive ridleys as well as the saltwater crocodile (Crocodylus porosus). Recognition and awards for the laboratory have been received from both national and international bodies.

Metabolic costs of altered growth trajectories across life transitions in amphibians.

Climate change is causing increases in temperature and in the frequency of extreme weather events. Under this scenario, organisms should maintain or develop strategies to cope with environmental fluctuations, such as the capacity to modify growth trajectories. However, altering growth can have negative consequences for organisms' fitness. Here, we investigated the metabolic alterations induced by compensatory growth during the larval development of the common frog (Rana temporaria), quantifying changes in oxidative stress, corticosterone levels and telomere length. We induced compensatory growth responses by exposing frog embryos to cold conditions (i.e. a 'false spring' scenario), which cause a delay in hatching. Once hatched, we reared larvae at two different photoperiods (24:0, representing the natural photoperiod of larvae, and 18:6) to test also for the interactive effects of light on growth responses. Larvae experiencing delayed hatching showed fast compensatory responses and reached larger size at metamorphosis. Larvae shortened their developmental period in response to delayed hatching. Non-permanent light conditions resulted in relaxed growth compared with larvae reared under permanent light conditions, which grew at their natural photoperiod and closer to their maximal rates. Growth responses altered the redox status and corticosterone levels of larvae. These physiological changes were developmental stage-dependent and mainly affected by photoperiod conditions. At catch-up, larvae reared at 18:6 light:dark cycles showed higher antioxidant activities and glucocorticoid secretion. On the contrary, larvae reared at 24:0 developed at higher rates without altering their oxidative status, likely an adaptation to grow under very restricting seasonal conditions at early life. At metamorphosis, compensatory responses induced higher cellular antioxidant activities probably caused by enhanced metabolism. Telomere length remained unaltered by experimental treatments but apparently tended to elongate across larval ontogeny, which would be a first evidence of telomere lengthening across metamorphosis. Under the forecasted increase in extreme climatic events, adjusting growth and developmental rates to the dynamics of environmental fluctuations may be essential for survival, but it can carry metabolic costs and affect later performance. Understanding the implications of such costs will be essential to properly estimate the impact of climate change on wild animals.

Increasing our knowledge on direct-developing frogs: The ontogeny of Ischnocnema henselii (Anura: Brachycephalidae)

Due to the difficulty in finding newly fresh spawn in natural environments, few embryonic developmental studies have been carried out in anurans with direct development. Here we provide detailed data on the embryonic ontogeny of Ischnocnema henselii, and compare some morphological aspects related to its developmental mode within and outside of the brachycephaloid clade. Embryonic development in I. henselii is characterized by a unicuspid egg tooth (bicuspid in most other Brachycephaloidea), external gills present, open vent tube, and tail fins with dorsoventral orientation throughout the development. We also provide the first account of skin development in direct-developing frogs, revealing that the maturation of the integument has a typical dorsal to ventral sequence of changes as in most biphasic anurans. The early onset of thyroid development seems to be consistent with the hypothesis that the evolution of direct development in anuran amphibians involved precocious activation of the thyroid axis. A comparative analysis with the still few described embryonic ontogenies for direct-developing species reveals variation in the length of the embryonic period, rate of development, size at hatching and presence/absence of external morphological characters, which suggests heterochronic shifts in the rate of species-specific stage progression. All these details reinforce the idea of the high morphological variability among direct-developing frogs.

Postmetamorphic ontogenetic allometry and the evolution of skull shape in Nest-building frogs Leptodactylus (Anura: Leptodactylidae).

Allometry constitutes an important source of morphological variation. However, its influence in head development in anurans has been poorly explored. By using geometric morphometrics followed by statistical and comparative methods we analyzed patterns of allometric change during cranial postmetamorphic ontogeny in species of Nest-building frogs Leptodactylus (Leptodactylidae). We found that the anuran skull is not a static structure, and allometry plays an important role indefining its shape in this group. Similar to other groups with biphasic life-cycle, and following a general trend in vertebrates, ontogenetic changes mostly involve rearrangement in rostral, otoccipital, and suspensorium regions. Ontogenetic transformations are paralleled by shape changes associated with evolutionary change in size, such that the skulls of species of different intrageneric groups are scaled to each other, and small and large species show patterns of paedomorphic/peramorphic features, respectively. Allometric trajectories producing those phenotypes are highly evolvable though, with shape change direction and magnitude varying widely among clades, and irrespective of changes in absolute body size. These results reinforce the importance of large-scale comparisons of growth patterns to understand the plasticity, evolution, and polarity of morphological changes in different clades.

Early ontogeny and sequence heterochronies in Leiuperinae frogs (Anura: Leptodactylidae).

The study of early development in Neotropical Leiuperinae frogs (Anura, Leptodactylidae) has been addressed by several works in recent times. However, a comparative developmental approach under a phylogenetic context was not available. Herein we analyzed the morphological and ontogenetic diversity of embryos belonging to 22 species of the three largest genera in Leiuperinae. We find that in most cases, variations fit with the phylogeny at the inter- and intrageneric levels. Embryo kyphosis and whitish color are synapomorphies for the clade grouping Physalaemus and Engystomops. The presence of a third lower tooth row on the oral disc is plesiomorphic for Leiuperinae, only changing in derived clades. The configurations and developmental trajectories of the lower lip are exceptionally diverse. The developmental sequences optimized on the phylogenetic tree recover an early differentiated first lower tooth row a synapomorphy of Pseudopaludicola and Physalaemus, and an early differentiated second row as synapomorphy of Pleurodema. On the other hand, few features are highly conserved in the subfamily, such as the adhesive glands universally present in a type-C configuration. Our results also suggest that the morphology and ontogeny of embryos is in some cases associated to the environment where they develop. A large body size, poorly developed transient respiratory structures, large yolk provision and delayed development of the digestive tract occur convergently in embryos inhabiting cold, oxygenated environments. Embryos that develop in warmer water bodies in xeric environments show more complex and persistent transient respiratory structures and an early onset of hind limbs development. Our survey highlights that morphology and early development of anurans can be a valuable source of information for phylogenetic studies, and provide fundamental bases to explore and discuss how evolutionary changes can be shaped by environmental conditions.

Comparative and functional analysis of the digital mucus glands and secretions of tree frogs

BackgroundMucus and mucus glands are important features of the amphibian cutis. In tree frogs, the mucus glands and their secretions are crucial components of the adhesive digital pads of these animals. Despite a variety of hypothesised functions of these components in tree frog attachment, the functional morphology of the digital mucus glands and the chemistry of the digital mucus are barely known. Here, we use an interdisciplinary comparative approach to analyse these components, and discuss their roles in tree frog attachment.ResultsUsing synchrotron micro-computer-tomography, we discovered in the arboreal frog Hyla cinerea that the ventral digital mucus glands differ in their morphology from regular anuran mucus glands and form a subdermal gland cluster. We show the presence of this gland cluster also in several other—not exclusively arboreal—anuran families. Using cryo-histochemistry as well as infrared and sum frequency generation spectroscopy on the mucus of two arboreal (H. cinerea and Osteopilus septentrionalis) and of two terrestrial, non-climbing frog species (Pyxicephalus adspersus and Ceratophrys cranwelli), we find neutral and acidic polysaccharides, and indications for proteinaceous and lipid-like mucus components. The mucus chemistry varies only little between dorsal and ventral digital mucus in H. cinerea, ventral digital and abdominal mucus in H. cinerea and O. septentrionalis, and between the ventral abdominal mucus of all four studied species.ConclusionsThe presence of a digital mucus gland cluster in various anuran families, as well as the absence of differences in the mucus chemistry between arboreal and non-arboreal frog species indicate an adaptation towards generic functional requirements as well as to attachment-related requirements. Overall, this study contributes to the understanding of the role of glands and their secretions in tree frog attachment and in bioadhesion in general, as well as the evolution of anurans.

Urban Sprawl Development in Eastern Bandung Region

Eastern Bandung Region including Cileunyi, Jatinangor and Rancaekek is a suburb area of Bandung, which has gone through many changes in land use from agrarian to non-agrarian, also the population density and urban growth rate have increased. This phenomenon can lead to urban sprawl and potentially lead to a negative spatial transformation and inefficient resource allocation. This research objectives are to identify the characteristics of urban sprawl development and the pattern of urban sprawl in Eastern Bandung area in 2002, 2010 and 2017. This study uses quantitative approach with descriptive method also spatial analysis technique, scoring, and descriptive analysis. The spatial data that was used are Google Earth image data in 2002, 2010 and 2017. The results of this study show that the characteristics of urban sprawl development that consist of physical and social dimension is high with a score of 2.46. The pattern of urban sprawl can be identified as the combination of ribbon and leap frog development. Ribbon development can be seen in Cileunyi and Jatinangor, meanwhile leap frog development occurs in Rancaekek. The results of this study hopefully can help improve the understanding about urban sprawl and encourage the policy makers in controlling the spatial transformation wisely.