Wild-caught Frogs Research Articles

Behavioural preference for low levels of UV-B radiation in two neotropical frog species from Costa Rica

Abstract:Tropical frogs experience damaging effects from exposure to UV-B radiation, and some diurnally active, conspicuous species exhibit avoidance behaviours to high levels of UV-B. To determine if similar behaviours are present in other diurnal frogs, we experimentally compared UV-B avoidance in two common species of neotropical diurnal frogs –Oophaga pumilio, an aposematic poison frog andCraugastor bransfordii, a cryptic leaf-litter frog – in response to different levels of UV-B. Wild-caught frogs were tested in experimental arenas fitted with filters that permitted two different levels of UV-B (low: 14% vs. high: 84% UV-B). Both species spent significantly more time under the low UV-B filter, suggesting that behavioural preferences for lower levels of UV-B are common to different diurnal species. Furthermore, maleO. pumiliosignificantly preferred lower levels of UV-B, whereas females did not exhibit a preference for lower UV-B, which may suggest differences in UV-B exposure or sensitivity and/or alternative mechanism(s) to avoid UV-B between sexes. Although limited in scope, the findings of our study suggest that UV-B avoidance may be a behavioural adaptation common to all diurnal frogs.

Isolation of Brucella inopinata-Like Bacteria from White's and Denny's Tree Frogs.

Brucella inopinata strain BO1 and B. sp. strain BO2 isolated from human patients, respectively, are genetically different from classical Brucella species. We isolated bacteria of the genus Brucella from two species of wild-caught tropical frogs kept in the facilities in Japan: White's tree frog, which inhabits Oceania, and Denny's tree frog, which inhabits Southeast Asia. Phylogenetic analyses based on 16S rRNA and recA gene sequences and multilocus sequence analysis showed that two isolates of Brucella spp. showed significant similarity to BO1, BO2, and the isolates from other wild-caught frogs. These results suggest that a variety of frog species are susceptible to a novel clade of Brucella bacteria, including B. inopinata.

Consistency of Published Results on the Pathogen Batrachochytrium dendrobatidis in Madagascar: Formal Comment on Kolby et al. Rapid Response to Evaluate the Presence of Amphibian Chytrid Fungus (Batrachochytrium dendrobatidis) and Ranavirus in Wild Amphibian Populations in Madagascar.

Consistency of Published Results on the Pathogen Batrachochytrium dendrobatidis in Madagascar: Formal Comment on Kolby et al. Rapid Response to Evaluate the Presence of Amphibian Chytrid Fungus (Batrachochytrium dendrobatidis) and Ranavirus in Wild Amphibian Populations in Madagascar.

Effects of herbicides and the chytrid fungus Batrachochytrium dendrobatidis on the health of post-metamorphic northern leopard frogs (Lithobates pipiens)

Effects of exposure to contaminants such as pesticides along with exposure to pathogens have been listed as two major contributors to the global crisis of declining amphibian populations. These two factors have also been linked in explanations of the causes of these population declines. We conducted a combined exposure experiment to test the hypothesis that exposure to two agricultural herbicides would increase the susceptibility of post-metamorphic northern leopard frogs (Lithobates pipiens) to the amphibian fungal pathogen Batrachochytrium dendrobatidis (Bd). We assessed the independent and interactive effects of these exposures on the health and survival of the frogs. Wild-caught frogs underwent a 21-day exposure to a nominal concentration of either 2.1μg/L atrazine (Aatrex® Liquid 480) or 100μga.e./L glyphosate (Roundup® Original), followed by Bd, and then were observed until 94 days post-initial exposure to the herbicides. Actual levels of atrazine were between 4.28±0.04μg/L and 1.70±0.26μg/L while glyphosate degraded from 100μga.e./L to approximately 7μga.e./L within 6 days of initial exposure to the herbicides. Compared to controls, the glyphosate formulation reduced the snout-vent length of frogs during the pesticide exposure (at Day 21), and the atrazine formulation reduced gain in mass up to Day 94. No treatment affected survival, splenosomatic or hepatosomatic indices, the densities and sizes of hepatic and splenic melanomacrophage aggregates, the density and size of hepatic granulomas, proportions of circulating leucocytes, the ratio of neutrophils to lymphocytes, or the ratio of leucocytes to erythrocytes. Histological assessment of samples collected at Day 94 revealed no evidence of Bd infection in any Bd-exposed frogs, while real-time PCR detected only one case of light infection in a single atrazine- and Bd-exposed frog. Frogs exposed to Bd shed their skin significantly more frequently than Bd-unexposed frogs, which may have helped them resist or clear infection, and could explain why no interaction between the herbicides and Bd was detected. The results suggest that these frogs were resistant to Bd infection and that pre-exposure to the herbicides did not alter this resistance. The effects seen on the growth following herbicide exposure is a concern, as reduced growth can lower the reproductive success and survival of the amphibians.

Radiographic diagnosis of metabolic bone disease in captive bred mountain chicken frogs (Leptodactylus fallax)

Asymptomatic captive bred and wild-caught mountain chicken frogs (Leptodactylus fallax) were radiographed for evidence of metabolic bone disease (MBD). All 22 captive bred frogs had multiple folding fractures of long bones, decreased bone density, and cortical thinning, whereas none of the 11 wild-caught frogs had any radiographic evidence of MBD. These findings suggest that the nutritional requirements of L. fallax need to be examined for captive management purposes.

Survey of Spirometra erinaceieuropaei spargana infection in the frog Rana nigromaculata of the Hunan Province of China

The prevalence of spargana infection in frogs ( Rana nigromaculata) was investigated in China's central Hunan Province, from March 2007 to October 2009. 59 of 292 (20.2%) wild-caught frogs were found to be infected with plerocercoids (spargana) of the genus Spirometra. Spargana were recovered from the skeletal muscle of the hind limb. The infection rate ranged from 4.5% to 27.4%, and the infection intensity was 1–15 spargana per frog. To identify the species identity of the collected spargana, a portion of the mitochondrial cytochrome c oxidase subunit 1 ( cox1) gene was amplified, sequenced, and analyzed. Sequence variations for cox1 among all the examined spargana were 0.0–3.1%, with 14 variable sites being identified in sequences obtained (3.1%, 14/446), representing 6 different cox1 sequences. Phylogenetic analysis showed that all the spargana isolates in Hunan province represented Spirometra erinaceieuropaei. This is the first report of S. erinaceieuropaei infection in frogs in Hunan province, China.

Batrachochytrium dendrobatidis and Chytridiomycosis in Anuran Amphibians of Colombia

In order to investigate the possible presence of the chytrid fungus Batrachochytrium dendrobatidis (Chytridiomycota: Chytridiales) in frogs (Amphibia: Anura) of Colombia, we made a retrospective examination of formalin-fixed specimens preserved in natural history collections. Using the staining technique of hematoxylin and eosin to identify B. dendrobatidis in histological slices, we found evidence of the fungus in 3 of the 53 frog species examined from a total of 672 specimens collected in 17 departments within Colombia between 1968 and 2006. The infected specimens were found dead or dying in recent years in high elevation sites, suggesting that chytridiomycosis (the disease caused by the fungus) may represent a significant threat to Colombian amphibians. We conclude that a more extensive search for B. dendrobatidis in museum specimens and wild-caught frogs should be undertaken as soon as possible, using both histological and molecular genetic techniques, in order to further characterize the geographic and taxonomic extent of infections of B. dendrobatidis.

Evidence for Biosynthesis of Pseudophrynamine Alkaloids by an Australian Myobatrachid Frog (Pseudophryne) and for Sequestration of Dietary Pumiliotoxins

Australian myobatrachid frogs of the genus Pseudophryne have only two classes of alkaloids in skin extracts, pseudophrynamines (PSs) and pumiliotoxins (PTXs). The former are unique to such Australian frogs, while the PTXs occur worldwide in all other genera of frogs/toads that contain lipophilic alkaloids. The major alkaloid of wild-caught frogs from one population of Pseudophryne semimarmorata was PTX 267C, while PSs were only minor or trace alkaloids. Captive-raised frogs from the same parental stock had no PTXs, but had larger amounts of PSs. A PTX fed to captive-raised frogs accumulated into skin along with dihydro and hydroxy metabolites. Thus, Pseudophryne frogs appear to biosynthesize PSs, but to sequester into skin dietary PTXs. In addition, biosynthesis of PSs appears reduced when high levels of dietary PTXs have accumulated into skin. This is the first evidence indicating that certain frogs are capable of synthesizing rather than merely sequestering alkaloids. A wide range of PSs, including many with molecular weights >500, were detected using both GC-mass spectral and LC-mass spectral analysis.

Corneal lipid deposition in Cuban tree frogs (Osteopilus septentrionalis) and its relationship to serum lipids: an experimental study.

To evaluate the association between corneal lipid infiltration (corneal arcus) and dietary cholesterol in Cuban tree frogs (Osteopilus septentrionalis), 47 wild-caught frogs were fed diets of either regular or high-cholesterol crickets containing 0.7% and 1.7% cholesterol dry matter, respectively. Serum total cholesterol and triglycerides were measured when the frogs were euthanized after 17 mo. In a subsample of frogs, serum lipoproteins were characterized using high-performance liquid chromatography. The first case of corneal lipid deposition occurred in a female frog after 13 mo on the high-cholesterol diet. In the subsequent 4 mo, 5/11 males and 11/35 females developed the disease. Four of these affected frogs were females on the regular diet. Frogs with corneal lipid deposition had elevated serum total cholesterol (27.3 +/- 19.8 mmol/L) and low-density lipoproteins (LDL, 17.8 +/- 18.9 mmol/L) compared with unaffected captive frogs (16.5 +/- 20.4 and 9.0 +/- 7.6 mmol/L, respectively). Corneal lipid deposition was more prevalent in frogs on the high-cholesterol diet, and this group had higher serum total cholesterol (34.1 +/- 15.2 mmol/L in females, 22.8 +/- 14.8 mmol/L in males) than did frogs on the diet of regular crickets (12.3 +/- 8.7 mmol/L in females, 10.4 +/- 3.1 mmol/L in males). Captive frogs on both diets had higher serum total cholesterol than did wild frogs (3.1 +/- 2.1 mmo/L in females, 5.3 +/- 2.6 mmo/L in males). This additional serum cholesterol was primarily carried on very low-density lipoproteins (VLDL) and LDL rather than high-density lipoproteins (HDL), as indicated by the significantly higher ratio of VLDL cholesterol and LDL cholesterol over HDL cholesterol in captive frogs compared with wild frogs. Elevation in this ratio was significantly higher in captive females than in captive males. There was no evidence that increased serum cholesterol in captive females was directly related to the process of vitellogenesis.

Interrelationship between food availability, fat body, and ovarian cycles in the frog,Rana tigrina, with a discussion on the role of fat body in anuran reproduction

Long-term experiments were conducted to study the progression of vitellogenic cycles in Rana tigrina (an annual breeder) having different foraging backgrounds and held under conditions of weekly or daily food supply and in presence or absence of abdominal fat bodies. They were autopsied in June to assess fecundity. In nature an adult R. tigrina produces on an average 4,000 eggs/100 g body mass (b.m.) And spawns in June-July following monsoon rains. Weekly feeding from July to next breeding season, June resulted in a significant decrease in both fecundity (1700 eggs/100 g body b.m.) And mean size of eggs, compared to well-fed or wild-caught frogs. The abdominal fat bodies were barely seen in frogs fed weekly throughout, whereas in frogs fed weekly from July-December but daily from January onwards, the fat bodies became noticeable (1% of b.m.) And number and mean size of eggs increased significantly over those fed weekly throughout. Frogs captured in January possessed enlarged fat bodies (5% of b.m.), depicting a good foraging history. Maintenance of these frogs on a weekly feeding regimen led to an exhaustion of fat stores. They produced less number of eggs (2, 000/100 g b.m.) As compared to wild frogs but of normal size, whereas daily feeding slowed down a depletion of fat body mass and also significantly increased fecundity (3,000/100 g b.m.) Over the weekly fed individuals. Sham operation or fat body ablation in October or February had no significant effect on total fecundity per se (3,000-3,500 eggs/100 g b.m.) Compared to that of wild-caught frogs. However, eggs were significantly smaller due to fat body ablation despite daily feeding. The study shows that food abundance/fat bodies influence egg size and number in R. tigrina and that a direct or indirect functional relationship exists between fat body and ovarian cycles that are characteristically inverse to each other. J. Exp. Zool. 286:487-493, 2000.

Arthropod-frog connection: decahydroquinoline and pyrrolizidine alkaloids common to microsympatric myrmicine ants and dendrobatid frogs.

Neotropical poison frogs (Dendrobatidae) contain a wide variety of lipophilic alkaloids, apparently accumulated unchanged into skin glands from dietary sources. Panamanian poison frogs (Dendrobates auratus) raised in a large, screened, outdoor cage and provided for six months with leaf-litter from the frog's natural habitat, accumulated a variety of alkaloids into the skin. These included two isomers of the ant pyrrolizidine 251K; two isomers of the 3,5-disubstituted indolizidine 195B; an alkaloid known to occur in myrmicine ants; another such indolizidine, 211E; two pyrrolidines, 197B and 223N, the former known to occur in myrmicine ants; two tricyclics, 193C and 219I, the former known to occur as precoccinelline in coccinellid beetles; and three spiropyrrolizidines, 222, 236, and 252A, representatives of an alkaloid class known to occur in millipedes. The alkaloids 211E, 197B, and 223N appear likely to derive in part from ants that entered the screened cage. In addition, the frog skin extracts contained trace amounts of four alkaloids, 205D, 207H, 219H, and 231H, of unknown structures and source. Wild-caught frogs from the leaf-litter site contained nearly 40 alkaloids, including most of the above alkaloids. Pumiliotoxins and histrionicotoxins were major alkaloids in wild-caught frogs, but were absent in captive-raised frogs. Ants microsympatric with the poison frog at the leaf-litter site and at an island site nearby in the Bay of Panama were examined for alkaloids. The decahydroquinoline (−)-cis-195A and two isomers of the pyrrolizidine 251K were found to be shared by microsympatric myrmicine ants and poison frogs. The proportions of the two isomers of 251K were the same in ant and frog.

Absence of tetrodotoxins in a captive-raised riparian frog, Atelopus varius

Bufonid frogs of the genus Atelopus contain two classes of skin toxins, namely the steroidal bufadienolides and the water-soluble tetrodotoxins. Frogs of the Panamanian species Atelopus varius have now been raised in captivity and levels in skin extracts of bufadienolides and of tetrodotoxin-like compounds assessed, using inhibition of [ 3H]ouabain binding and inhibition of [ 3H]saxitoxin binding, respectively. Levels of ouabain equivalents, corresponding to bufadienolides, were comparable to those found in wild-caught frogs from the same population in Panama, while tetrodotoxin-like activity was undetectable. The results strongly implicate environmental factors, perhaps symbiotic microorganisms, in the genesis of tetrodotoxins in the skin of frogs of the genus Atelopus, while indicating that the frog itself produces the skin bufadienolides.

Variation in the Incidence of Renal Adenocarcinoma Among Rana pipiens Held Under Tumor-promoting Conditions

In 1975-1976, significant differences were observed in the frequencies of renal tumors in wild-caught frogs from different sites when they were held in the laboratory under tumor-promoting conditions. In 1979-1980, tumors could not be induced in frogs from a site where spontaneous tumors were no longer detectable. It is suggested that because spontaneous tumors have decreased in natural populations, it is less likely that the etiologic agent the Lucke-tumor herpesvirus will be present to promote tumor development under the stress of laboratory conditions.

Levels of batrachotoxin and lack of sensitivity to its action in poison-dart frogs (Phyllobates).

Batrachotoxin is present in remarkably high amounts in the skin of Phyllobates terribilis. Levels of batrachotoxin tend to be reduced when P. terribilis is maintained in captivity, but even after being confined for up to 6 years, these frogs were still at least five times more toxic than other Phyllobates species used by natives for poisoning blowgun darts. Batrachotoxin was not detectable in F1 progeny reared to maturity in captivity. Nerve and muscle preparations from wild-caught frogs and from the nontoxic F1 frogs were both insensitive to batrachotoxin. The regulatory site controlling sodium-channel activation and permeability appears to have been minimally altered to prevent interaction with batrachotoxin, but is still sensitive to other sodium conductance activators (veratridine, grayanotoxin) to which the frogs arenot exposed naturally.