Osteopilus Septentrionalis Research Articles

Physiological and behavioral responses to novel saline conditions in an invasive treefrog.

Salinity can be an environmental stressor for anurans, as their highly permeable skin makes them prone to osmotic stress when exposed to saline conditions. However, certain anuran species have colonized areas near saltwater habitats, suggesting an ability to acclimate to saline conditions. Here, we evaluated physiological and behavioral responses to saline conditions in adult Cuban treefrogs (Osteopilus septentrionalis), an invasive anuran found throughout Florida. To examine their response to salinity, adult frogs were maintained in two treatments simulating a freshwater (0.5 ppt) or brackish (8.0 ppt) environment for 6 weeks. To assess their physiological response to this potential stressor, all frogs were submerged in a brackish solution to quantify individual weight change every 2 weeks. We found that frogs maintained in brackish solution lost more weight at Weeks 2 and 6 when compared to Week 0, suggesting that salinity may be an environmental stressor for Cuban treefrogs. Yet, the weight change at Week 4 was similar to the pre-exposure period, which may indicate that constant exposure to salinity may alter their physiological response to saline conditions. To supplement the physiological analyses, we investigated avoidance behavior towardsaline conditions by offering individuals a choice between freshwater or brackish environments. Our results showed that Cuban treefrogs chose freshwater environments more frequently and may thus avoid saline ones. This study reveals that salinity may induce plastic and avoidance responses in Cuban treefrogs, potentially allowing them to expand their range into areas typically stressful for most anurans.

Pterygodermatites (Mesopectines) whartoni (Nematoda: Rictulariidae) encysted larvae in invasive Cuban treefrogs (Osteopilus septentrionalis) from Florida, United States.

Species of Pterygodermatites are spirurid nematodes that have expanded their geographic distribution worldwide. They infect a variety of mammalian definitive hosts with few reports of potential paratenic infections in amphibian and reptile hosts. In this study, we report Pterygodermatites sp. larvae identified in free-ranging, invasive Cuban treefrogs (Osteopilus septentrionalis), from central Florida, United States. Encysted larvae were recovered from the skeletal muscle and/or the coelomic cavity of three frogs; molecular characterization of the small subunit (18S) ribosomal RNA and cytochrome oxidase I genes of the parasites matched reported sequences of Pterygodermatites (Mesopectines) whartoni (Tubangui, 1931). This is a parasite native to Southeastern Asia and to the best of the authors' knowledge, it is the first report of the species in the New World. The recovery of invasive Pterygodermatites from invasive Cuban treefrogs in North America highlights the growing concern regarding the potential impact non-native parasites and invasive species may have on native wildlife populations.

Carbon nitride dots do not impair the growth, development, and telomere length of tadpoles

Carbon nanoparticles, or carbon dots, can have many beneficial uses. However, we must consider whether they may have any potential negative side effects on wildlife or the ecosystem when these particles end up in wastewater. Early development stages of amphibians are particularly sensitive to contaminants, and exposure to carbon dots could disrupt their development and cause morbidity or death. Past studies have investigated short-term exposure to certain types of nanoparticles, but if these particles get into wastewater exposure may not be short term. Therefore, we tested whether chronic exposure to different concentrations of carbon dots affects the growth, metamorphosis, and telomere length of Cuban tree frog (Osteopilus septentrionalis) tadpoles. We exposed 12 groups of five tadpoles each to different concentrations of carbon dots and a control for three months and tracked survival, growth and metamorphosis. We used carbon nitride dots approximately 2 nm in size at concentrations of 0.01 mg/ml and 0.02 mg/ml, known to interrupt development in zebrafish embryos. After three months, we measured telomere length from tissue samples. We found no difference in tadpole survivorship, growth, development rate, or telomere length among any of the groups, suggesting that carbon dots at these concentrations do not disrupt tadpole development.

The Cuban Treefrog, Osteopilus septentrionalis (Duméril & Bibron), is established in Jamaica

The Cuban Treefrog Osteopilus septentrionalis is native to the Bahamas, the Cayman Islands, and Cuba and has been introduced throughout the Caribbean islands and the U.S., including the states of Florida, Georgia, Hawaii, and Louisiana (IUCN SSC Amphibian Specialist Group 2021). Genetic studies also suggested that the species may have naturally dispersed to Florida (Heinicke et al. 2011). Although not previously recorded from Jamaica, I shared the presence of the species in Jamaica with the IUCN during the review of the status of frogs from that island (IUCN SSC Amphibian Specialist Group 2021). However, because of its potential confusion with native hylids, it is important to better document its presence on Jamaica. Here, I report on details surrounding the establishment of the species on Jamaica and its current distribution on that island.

A Pilot Study Investigating Plasma Protein Electrophoresis in One Anuran and Six Urodelan Species.

As threats to amphibian health increase, there is a growing need for diagnostic tools to assess and monitor their health status. Plasma protein electrophoresis has proven to be useful in other nonmammalian species. It enables quantification of protein fractions in plasma that may be altered in various disease processes, and is therefore useful in narrowing down differential diagnoses and detecting inflammation, in combination with other modalities such as biochemical and hematologic testing. The amphibian electrophoretogram must be defined before baseline reference intervals are obtained across species. Agarose gel electrophoresis was performed on plasma samples collected from presumed clinically normal individuals of one anuran and six urodelans: Osteopilus septentrionalis (n=2), Gyrinophilus porphyriticus (n=1), Notophthalmus viridescens (n=1), Eurycea guttolineata (n=2), Amphiuma tridactylum (n=2), Cryptobranchus alleganiensis (n=5), and Siren lacertina (n=6). The electrophoretograms varied in number of fractions between each species; however, the number of fractions was consistent within a species. An albumin migrating fraction was consistently observed in all species. A prealbumin migrating fraction was identified in species that primarily use organs other than skin for respiration. This study provides preliminary examples of a normal plasma protein electrophoretogram for seven amphibian species. Further studies quantifying reference intervals and identification of protein fractions will help establish protein electrophoresis as a useful tool in amphibian health investigations.

Lack of vitamin B12 impairs innate and adaptive immunity of Cuban tree frog (Osteopilus septentrionalis) tadpoles.

Vitamin B12 is a micronutrient required by a variety of organisms for healthy cellular functioning. Despite the systemic effects observed in cases of B12 deficiency, relatively little is known about how vitamin B12 affects immune health, especially in amphibians, which are declining at unprecedented rates. In this study, we tested how supplementing an algae diet with B12 affects the innate and adaptive immunity of Cuban tree frog (Osteopilus septentrionalis) tadpoles. We found that innate immunity, as measured by a bacterial killing assay, was significantly more robust in B12-supplemented tadpoles than control tadpoles, but no significant differences were found in natural antibody production or hematocrit between groups. Adaptive immunity, as measured by Aeromonas hydrophila-specific IgY antibodies, was significantly greater in tadpoles challenged with A. hydrophila and supplemented with B12 than in control tadpoles, those only challenged with A. hydrophila, and those only given B12. Our results suggest that vitamin B12 is an important factor in maintaining a functional immune system in tadpoles, which may also be true for all vertebrates.

Ontogeny of immunity and potential implications for co-infection.

Immunity changes through ontogeny and can mediate facilitative and inhibitory interactions among co-infecting parasite species. In amphibians, most immune memory is not carried through metamorphosis, leading to variation in the complexity of immune responses across life stages. To test if the ontogeny of host immunity might drive interactions among co-infecting parasites, we simultaneously exposed Cuban treefrogs (Osteopilus septentrionalis) to a fungus (Batrachochytrium dendrobaditis, Bd) and a nematode (Aplectana hamatospicula) at tadpole, metamorphic and post-metamorphic life stages. We measured metrics of host immunity, host health and parasite abundance. We predicted facilitative interactions between co-infecting parasites as the different immune responses hosts mount to combat these infectious are energetically challenging to mount simultaneously. We found ontogenetic differences in IgY levels and cellular immunity but no evidence that metamorphic frogs were more immunosuppressed than tadpoles. There was also little evidence that these parasites facilitated one another and no evidence that A. hamatospicula infection altered host immunity or health. However, Bd, which is known to be immunosuppressive, decreased immunity in metamorphic frogs. This made metamorphic frogs both less resistant and less tolerant of Bd infection than the other life stages. These findings indicate that changes in immunity altered host responses to parasite exposures throughout ontogeny. This article is part of the theme issue 'Amphibian immunity: stress, disease and ecoimmunology'.

Predation of the Cuban Treefrog (Osteopilus sepentrionalis) and three nonnative lizard species by Eastern Screech-Owls (Megascops asio) in Manatee County, Florida, USA

Predation of the Cuban Treefrog (Osteopilus sepentrionalis) and three nonnative lizard species by Eastern Screech-Owls (Megascops asio) in Manatee County, Florida, USA

Cuban Treefrog (Osteopilus septentrionalis) in Florida

Florida is the global epicenter for introduced nonnative reptiles and amphibians. These include well-known species such as Burmese pythons and green iguanas as well as dozens of other species of snakes, lizards, and frogs. There are four species of nonnative frogs currently established in Florida. The purpose of this publication, a UF/IFAS numbered Organism ID., is to summarize the status, biology, and impacts of one of those frogs, the Cuban treefrog. The publication also presents strategies to manage Cuban treefrogs to help alleviate human conflicts and benefit native species. The main target audience is homeowners who suspect they have Cuban treefrogs on their property and/or need technical assistance dealing with these often-problematic frogs. Environmental educators, natural resource managers, and professional scientists will also find the information herein of interest.

New Annotated Records of Helminth Parasites with Light Microscopy II. Acanthocephala (Polymorphida)

We give an account of 8 species of polymorphid acanthocephalans in 3 families collected from 14 various host species in different geographical locations. Each species account is annotated and morphologically presented using light microscopy showing characteristic diagnostic features. None of these accounts has been previously published. All geographical records and many host associations are new. This treatment includes Centrorhynchus spinosus (Kaiser, 1893) adults from Strix aluco Linn. In Iran and cystacanths from Osteopilus septentrionalis (Duméril & Bibron) in Florida; Sphaerirostris lancea (Westrumb, 1821) from Vanellus vanellus (Linn.) in Ukraine; Sphaerirostris picae (Rudolphi, 1819) Golvan, 1956 from various hosts in Ukraine, Iran, Bulgaria and Egypt (Centrorhynchidae); Plagiorhynchus (Prosthorhynchus) cylindraceus (Goeze, 1782) Schmidt & Kuntz, 1966 from Turdus merula Linn. in Iran and Sturnus vulgaris Linn. in Wisconsin; Plagiorhynchus (Prosthorhynchus) nicobarensis Soota & Kansal, 1970 from S. vulgaris in Iran (Plagiorhynchidae); Andracantha gravida (Algret, 1941) Schmidt, 1975 from Gavia immer (Brunnich) in Florida; Arhythmorhynchus brevis Van Cleave, 1916 from Casmerodius albus (Linn.) in Florida, and Southwellina hispida (Van Cleave, 1925) Witenberg, 1932 (= Polymorphus ardeae Belopolskaya, 1958) from Ardea cinerea Linn. in Macedonia and from Nycticorax violaceus (Linn.) in Florida.

Pseudacris regilla metamorphs acquire resistance to Batrachochytrium dendrobatidis after exposure to the killed fungus.

The pathogenic fungus Batrachochytrium dendrobatidis (Bd) is associated with drastic global amphibian declines. Prophylactic exposure to killed zoospores and the soluble chemicals they produce (Bd metabolites) can induce acquired resistance to Bd in adult Cuban treefrogs Osteopilus septentrionalis. Here, we exposed metamorphic frogs of a second species, the Pacific chorus frog Pseudacris regilla, to one of 2 prophylactic treatments prior to live Bd exposures: killed Bd zoospores with metabolites, killed zoospores alone, or a water control. Prior exposure to killed Bd zoospores with metabolites reduced Bd infection intensity in metamorphic Pacific chorus frogs by 60.4% compared to control frogs. Interestingly, Bd intensity in metamorphs previously exposed to killed zoospores alone did not differ in magnitude relative to the control metamorphs, nor to those treated with killed zoospores plus metabolites. Previous work indicated that Bd metabolites alone can induce acquired resistance in tadpoles, and so these findings together indicate that it is possible that the soluble Bd metabolites may contain immunomodulatory components that drive this resistance phenotype. Our results expand the generality of this prophylaxis work by identifying a second amphibian species (Pacific chorus frog) and an additional amphibian life stage (metamorphic frog) that can acquire resistance to Bd after metabolite exposure. This work increases hopes that a Bd-metabolite prophylaxis might be widely effective across amphibian species and life stages.

Identity and density of parasite exposures alter the outcome of coinfections: Implications for management

Abstract Although research has focused on density‐dependent responses of hosts to single‐parasite infections, hosts are exposed to numerous parasites simultaneously under natural conditions and if these exposures lead to infections, they can threaten host populations and ecosystem stability. Moreover, spatiotemporal variation in abundance of co‐occurring parasites might influence host infection intensity. If interactions are consistent between different coinfecting parasites, then these patterns could give managers another tool to control disease spread and even predict problematic disease emergences. We investigated how parasite density and identity alter within‐host coinfection dynamics. To test this, we simultaneously exposed Cuban treefrogs Osteopilus septentrionalis as a model amphibian species to all pairwise combinations of three problematic parasites that commonly coinfect amphibians: the fungus Batrachochytrium dendrobatidis (Bd), the nematode Aplectana hamatospicula and Ranavirus. Hosts were exposed to one parasite at a fixed dose and another parasite at a range of five doses. Higher doses of Bd decreased Ranaviral and A. hamatospicula loads, but Bd load was not influenced by the dose of either parasite. Ranaviral load was negatively associated with A. hamatospicula dose, but A. hamatospicula load was not affected by Ranaviral dose. We found that all the pairwise coinfections were dependent on parasite density and that pairwise interactions were highly asymmetric—strong in one direction and weak in the other—consistent with interactions dominating food webs. Synthesis and applications: We also revealed that the exposure dose of A. hamatospicula was positively associated with host tolerance to Bd infection and negatively associated with Ranaviral load in hosts. Ranavirus and Bd cause mass die‐offs in amphibians, but A. hamatospicula does not. Therefore, in systems where these parasites coexist, maintaining or increasing densities of A. hamatospicula could reduce the negative effects of Bd and Ranavirus infections. Additionally, if these asymmetric and density‐dependent patterns from community ecology are applicable to other amphibian coinfections or coinfections in other systems, this should allow conservation organizations and resource managers to predict outbreaks and manage host declines associated with deadly parasites by modifying the abundance of coinfecting parasites that might be easier to manage.

Muscle preactivation and the limits of muscle power output during jumping in the Cuban tree frog Osteopilus septentrionalis.

Previous studies of jumping in frogs have found power outputs in excess of what is possible from direct application of muscle power and concluded that jumping requires the storage and release of elastic strain energy. Of course, the muscles must produce the work required and their power output should be consistent with known muscle properties if the total duration of muscle activity is known. Using the Cuban tree frog, Osteopilus septentrionalis, I measured jumping performance from kinematics and used EMG measurements of three major jumping muscles to determine the duration of muscle activity. Using the total mass of all the hindlimb muscles, muscle mass-specific work output up to 60 J kg-1 was recorded. Distributed over the duration of the jump, both average and peak muscle mass-specific power output increased approximately linearly with the work done, reaching values of over 750 and 2000 W kg-1, respectively. However, the muscles were activated before the jump started. Both preactivation duration and EMG amplitude increased with increasing amounts of work performed. Assuming the muscles could produce work from EMG onset until toe-off, the average muscle mass-specific power over this longer interval also increased with work done, but only up to a work output of 36 J kg-1. The mean power above this value of work was 281 W kg-1, which is approximately 65% of the estimated maximum isotonic power. Several reasons are put forward for suggesting this power output, although within the known properties of the muscles, is nevertheless an impressive achievement.

Infection Experiments Indicate That Common Florida Anurans and Lizards May Serve as Intermediate Hosts for the Invasive Pentastome Parasite, Raillietiella orientalis

Raillietiella orientalis, an invasive pentastome parasite, inhabits the lungs of snakes as adults and is rapidly spreading in Florida and Australia. The species that serve as intermediate hosts are currently unknown. We conducted laboratory infection experiments with insects (Blaberus discoidalis), lizards (Anolis sagrei), and anurans (Anaxyrus terrestris, Lithobates sphenocephalus, Osteopilus septentrionalis) to develop an understanding of which species may serve as intermediate hosts and to determine the fitness consequences of infection by R. orientalis. Lizards and insects, but not anurans, were readily infected by consuming food that was contaminated with pentastome eggs. Anolis sagrei and L. sphenocephalus were both infected after eating a single roach that was infected with R. orientalis larvae. Comparison with uninfected control animals revealed that pentastome infection did not significantly affect survival or growth in roaches, lizards, or anurans. The life cycle of R. orientalis in Florida is likely to involve a sequence of three hosts, with eggs hatching in coprophagous insects that infect lizards and anurans that infect the definitive host (snakes) after they are consumed. Our results indicate that the native species that serve as intermediate hosts are unlikely to experience major negative consequences from R. orientalis, unlike the native snake species that serve as the definitive hosts. The diversity of species that can serve as intermediate hosts and the potential for vehicular rafting by infected roaches and anoles indicate that the rapid geographic range expansion of R. orientalis will probably continue.

Diurnal activity in the Cuban Treefrog, Osteopilus septentrionalis (Anura: Hylidae), associated with an unusually high prey density, in western Cuba

Diurnal activity in the Cuban Treefrog, Osteopilus septentrionalis (Anura: Hylidae), associated with an unusually high prey density, in western Cuba

Metabolites from the fungal pathogen Batrachochytrium dendrobatidis (bd) reduce Bd load in Cuban treefrog tadpoles

Abstract Batrachochytrium dendrobatidis (Bd) has been associated with massive amphibian population declines worldwide. Wildlife vaccination campaigns have proven effective for mitigating damage from other pathogens, and there is evidence that adult frogs can acquire resistance to Bd when exposed to killed Bd zoospores and the metabolites they produced. Here, we investigated whether Cuban treefrogs tadpoles Osteopilus septentrionalis can gain protection from Bd through exposure to a prophylaxis treatment composed of killed zoospores or soluble Bd metabolites. We used a 2 × 2 factorial design, crossing the presence or absence of killed zoospores with the presence or absence of Bd metabolites. All hosts were subsequently exposed to live Bd to evaluate susceptibility. Exposure to killed zoospores did not induce a protective response. However, tadpoles exposed to Bd metabolites had significantly lower Bd intensity and prevalence than tadpoles that were not exposed to metabolites. The metabolites Bd produce pose no risk of Bd infection and therefore make an epidemiologically safe prophylaxis treatment, protecting tadpoles against Bd. This work provides a promising potential for protecting amphibians in the wild as a disease management strategy for controlling Bd‐associated declines.

Characterizing Tissue Enzyme Activities in the Marine Toad (Rhinella marina)

Abstract Plasma biochemistry tests are routine diagnostic tools in veterinary medicine that require an understanding of the enzyme activity profiles of individual organs for accurate interpretation. Tissue enzyme activity has been found to vary among and within taxa and has not been described in the marine toad (Rhinella marina). This study measured enzyme activities in the plasma, liver, heart, skeletal muscle, gastrointestinal system, kidney, and pancreas of five marine toads, to evaluate the tissue specificity of analytes commonly included on plasma biochemical analysis. In the marine toad, alanine transaminase (ALT) and aspartate transaminase activities were not specific to one organ, with minimal ALT activity measured in any organ tissue. Alkaline phosphatase activity was significantly higher in heart tissues and gamma-glutamyltransferase (GGT) activity was significantly higher in pancreatic tissues than in all other organ tissues, and plasma. GGT activity in the kidney was also significantly higher than that found in plasma, but not all other tissues. Amylase and lipase were highest in the plasma. Creatine phosphokinase and lactate dehydrogenase (LDH) were specific to skeletal muscle, with significant LDH activity also found in cardiac tissue. This information will be vital to the interpretation of plasma biochemistry analysis in the marine toad and shows similarities to tissue enzyme activity in the Cuban treefrog (Osteopilus septentrionalis), suggesting extrapolation to other anurans is possible.

Rat Lungworm (Angiostrongylus cantonensis) in the Invasive Cuban Treefrog (Osteopilus septentrionalis) in Central Florida, USA.

Cuban treefrogs, Osteopilus septentrionalis, were grossly examined for parasites and parasite species confirmed by PCR. Angiostrongylus cantonensis larvae were recovered from the hind leg muscle of O. septentrionalis. This is the first report of the zoonotic rat lungworm in the Cuban treefrog and new geographic location (Volusia County) in Florida, US.

Frogs fine-tune springy muscle and tendon for leaps

Long before the advent of gunpowder, humans had harnessed the power of catapults in warfare to destructive effect. Yet, biology had already beaten us to capitalising on the power of springs. ‘Fast and powerful movements like the jump of the flea or the strike of a mantis shrimp smasher are possible because they use elastic energy storage mechanisms’, says Elizabeth Mendoza from the University of California, Irvine, USA. Even soft-bodied frogs get in on the action, storing energy in the plantaris longus muscle–tendon unit – linking the back of the knee to the heel – ready for release in powerful leaps. Yet in 2011, when Tom Roberts, Emily Abbott and Manny Azizi investigated the source of energy powering the jumps of three frog and toad species, they were surprised that the animals’ explosive launches were driven by plantaris longus muscle contractions of about the same power (∼300 W kg−1), even though Cuban tree frogs (Osteopilus septentrionalis) leapt with a take-off power of ∼1100 W kg−1 while cane toads (Bufo marinus) achieved take-offs with a power of ∼200 W kg−1. Realising that the animals must be fine-tuning the energy storage capacity of the springy plantaris longus muscle–tendon unit, resulting in vastly different propulsion, Mendoza and Azizi scrutinised the legs of cane toads, Cuban tree frogs and bullfrogs (Rana catesbeiana) to determine how they differed. ‘All of our animals came from the pet trade’, says Mendoza, who collected samples of the animals’ plantaris longus muscle–tendon units to measure the amount of force exerted by the muscles as they contracted. ‘There are many challenges with this technique’, says Mendoza, such as keeping the muscle alive, making sure the nerve branch was not crushed or pulled too hard, and keeping the muscle–tendon unit securely in place in the force measuring rig. Then, she and Azizi investigated the structure of the frogs’ plantaris longus muscles, cutting the muscle open to reveal the featherlike arrangement of the fibres within to measure their stacking angle relative to the direction in which the muscle pulls (the pennation angle). Finally, the duo calculated how much work the muscle did during a contraction and the overall stiffness of the muscle–tendon unit to find out how much energy it could store.After months of painstaking analysis, it was clear that the plantaris longus muscle–tendon unit of the Cuban tree frog stood out from the crowd. ‘It was modified for increased elastic energy storage’, says Mendoza, explaining that the structure is stiffer than that of the cane toad and bullfrog muscle–tendon unit, allowing the Cuban tree frogs to store more elastic energy to power their mighty leaps. In addition, when Mendoza and Azizi measured the force generated by the tree frog's muscle (∼30 N g−1), it was almost double that produced by the cane toad and bullfrog (∼17 N g−1).But how was the tree frog's muscle able to generate so much force when it was so much smaller (∼0.3 g) than the bullfrog and cane toad's muscles (∼1.3 and ∼1 g, respectively)? Scrutinising the muscle structures, the duo realised that the muscle fibres in the Cuban tree frog plantaris longus were stacked at a much steeper angle (∼25 deg compared with the cane toad and bullfrog's 20 deg pennation angle), allowing the tree frogs to pack more fibres into the muscle to generate more force.‘A tuned increase in muscle force and elastic structure stiffness increases elastic energy storage in Cuban tree frogs’, says Mendoza, and Azizi adds, ‘In terms of jump distance, Cuban tree frogs are the best, followed by bullfrogs and then cane toads’. So, the Cuban tree frog's springier legs tie in nicely with their exceptional vaulting abilities.

Tuned muscle and spring properties increase elastic energy storage.

Elastic recoil drives some of the fastest and most powerful biological movements. For effective use of elastic recoil, the tuning of muscle and spring force capacity is essential. Although studies of invertebrate organisms that use elastic recoil show evidence of increased force capacity in their energy loading muscle, changes in the fundamental properties of such muscles have yet to be documented in vertebrates. Here, we used three species of frogs (Cuban tree frogs, bullfrogs and cane toads) that differ in jumping power to investigate functional shifts in muscle-spring tuning in systems using latch-mediated spring actuation (LaMSA). We hypothesized that variation in jumping performance would result from increased force capacity in muscles and relatively stiffer elastic structures, resulting in greater energy storage. To test this, we characterized the force-length property of the plantaris longus muscle-tendon unit (MTU), and quantified the maximal amount of energy stored in elastic structures for each species. We found that the plantaris longus MTU of Cuban tree frogs produced higher mass-specific energy and mass-specific forces than the other two species. Moreover, we found that the plantaris longus MTU of Cuban tree frogs had higher pennation angles than the other species, suggesting that muscle architecture was modified to increase force capacity through packing of more muscle fibers. Finally, we found that the elastic structures were relatively stiffer in Cuban tree frogs. These results provide a mechanistic link between the tuned properties of LaMSA components, energy storage capacity and whole-system performance.