Age-related changes in antioxidant defenses of the Mediterranean centipede Scolopendra cingulata (Chilopoda).

The activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), glutathione reductase (GR), and glutathione S-transferase (GST), as well as the concentrations of sulfhydryl (SH) groups and glutathione (GSH) were analyzed in five age classes of the Mediterranean centipede Scolopendra cingulata as follows: embryo, adolescens, maturus junior, maturus, and maturus senior. The data obtained showed the presence of SOD, CAT, GSH-Px, GR, GST, and SH groups in embryos. The transition from embryo to adolescens was accompanied by an increase in the activities of all studied enzymes, in response to the increased production of ROS due to the increased metabolic activity of the centipede associated with growth and development. Our results show that trends in antioxidant enzyme (AOE) activities were not uniform among adult age classes, suggesting that maturus junior, maturus, and maturus senior differentially respond and/or have different susceptibility to ROS. On the other hand, GSH concentration in embryos was undetectable, highest in adolescens and decreased in the latter part of life. Pearson correlation analysis in embryos showed that the activities of the AOEs were strongly and positively correlated with each other but negatively correlated with GSH and SH groups. At later age classes, SOD, CAT, GSH-Px, GR, GSH, and SH groups were no longer significantly correlated with GST. In the discriminant analysis, the variables that separated the age classes were GR, GST, SH groups, and body length. Body length was directly related to the age of individuals, clearly indicating that development/aging affects the regulation of antioxidant defense in this species.

Isotope data from amino acids indicate Darwin's ground sloth was not an herbivore.

Fossil sloths are regarded as obligate herbivores for reasons including peculiarities of their craniodental morphology and that all living sloths feed exclusively on plants. We challenge this view based on isotopic analyses of nitrogen of specific amino acids, which show that Darwin's ground sloth Mylodon darwinii was an opportunistic omnivore. This direct evidence of omnivory in an ancient sloth requires reevaluation of the ecological structure of South American Cenozoic mammalian communities, as sloths represented a major component of these ecosystems across the past 34 Myr. Furthermore, by analyzing modern mammals with known diets, we provide a basis for reliable interpretation of nitrogen isotopes of amino acids of fossils. We argue that a widely used equation to determine trophic position is unnecessary, and that the relative isotopic values of the amino acids glutamate and phenylalanine alone permit reliable reconstructions of trophic positions of extant and extinct mammals.

Inhibitory action of thymol on fecal microbial activity in Tamandua tetradactyla and its effect on glucocorticoid metabolite measurement.

Faecal glucocorticoid measurement is a potentially important tool for improving wildlife conservation, but its use is still limited by methodological issues including the need to avoid modifications of steroids by faecal microorganisms during storage. The freezing of faeces is recommended as a means of avoiding such alterations, but this is costly under non-controlled environmental conditions. The present study was designed to determine whether the application of thymol reduced the proliferation of microorganisms in the faeces of Tamandua tetradactyla and whether it influenced faecal glucocorticoid metabolite (FGM) measurements. Tamandua tetradactyla faeces were individually collected after defaecation, divided into fractions (5.5 g each) and kept in sealed glass Petri dishes at 22 ±â€¯2 °C. A thymol solution (550 µL; 5 mg g-1 feces; 80% ethanol) or an 80% ethanol solution (550 µL, control) was added before storage of faeces. Negative controls for FGM consisted of samples without thymol or ethanol solutions. All samples were evaluated at 0, 24, 48 and 72 h post-defaecation. Thymol was first incubated with a glucocorticoid standard in a faeces-free tube or in a faecal sample in order to determine whether it interfered with FGM measurements. Data showed that thymol did not affect FGM measurements. Post-defaecation time caused a significant reduction in FGM measurements in the negative control, an increment at 48 h in the control, and no change in FGM measurements in thymol treatment. FGM measurements were significantly different between groups (negative control > control - treatment). Thymol caused a significant reduction of up to three orders of magnitude in total coliforms, total aerobic and anaerobic heterotrophic mesophilic bacteria, mold and yeast per gram of faeces at 24, 48 and 72 h. The reduction in microbial activity presumably contributed to the stability of FGM over time. Spore-forming bacteria (SFB) in faeces were not reduced by thymol. We propose thymol as an alternative to freezing since it stabilizes FGMs for at least 3 days after collection in the faeces of Tamandua tetradactyla.

Mortality and morbidity in wild Taiwanese pangolin (Manis pentadactyla pentadactyla).

Globally, pangolins are threatened by poaching and illegal trade. Taiwan presents a contrary situation, where the wild pangolin population has stabilized and even begun to increase in the last two decades. This paper illustrates the factors responsible for causing mortality and morbidity in the wild Taiwanese pangolin (Manis pentadactyla pentadactyla) based on radio-tracking data of wild pangolins and records of sick or injured pangolins admitted to a Taiwanese wildlife rehabilitation center. Despite being proficient burrowers, results from radio-tracking show that Taiwanese pangolins are highly susceptible to getting trapped in tree hollows or ground burrows. Data from Pingtung Rescue Center for Endangered Wild Animals showed that trauma (73.0%) was the major reason for morbidity in the Taiwanese pangolin with trauma from gin traps being the leading cause (77.8%), especially during the dry season, followed by tail injuries caused by dog attacks (20.4%). Despite these threats, Taiwan has had substantial success in rehabilitating and releasing injured pangolins, primarily due to the close collaboration of Taiwanese wildlife rehabilitation centers over the last twenty years.

Effect of increasing taurine and methionine supplementation on urinary taurine excretion in a model insectivore, the giant anteater (Myrmecophaga tridactyla).

The giant anteater (Mymercophaga tridactyla) is a highly specialized insectivore for which nutrient requirements are not clearly established, making diet formulation challenging for this species. Multiple clinical reports suggest anteaters have an obligate dietary taurine (TAU) requirement. Sulphur amino acid (SAA) metabolism in adult anteaters was evaluated using noninvasive methods to measure TAU synthesis potential from dietary methionine (MET) and a basal diet containing on a dry matter (DM) basis 1.7 mg TAU/kg DM and 6.9 g MET/kg DM. Urinary equilibrium times for TAU excretion were determined by feeding the basal diet with or without 1.5 g/kg DM supplemental TAU (crossover design; n = 4). Effects of supplemental dietary TAU (1.7, 2.0, 2.4, 2.7, 3.0, 3.3 g/kg DM) or MET (6.9, 9.0, 11.2 g/kg DM) on urinary TAU were evaluated (randomized block trials; n = 5 or 4 respectively). All urinary values (TAU, MET, unbound inorganic sulphate) were normalized to creatinine (CRT). Results indicate urinary TAU equilibrium in anteaters requires at least 2 weeks of feeding. Urinary ratio of TAU to CRT (TAU:CRT) increased as dietary TAU content increased from 1.7 to 3.0 g/kg DM, consistent with renal homoeostatic modulation of TAU excretion. Our data indicate that TAU needs were met by TAU in the basal diet or by de novo synthesis. Supplemental MET resulted in ~five- to eightfold increases in urinary TAU:CRT excretion, further supporting existence of mechanisms for TAU synthesis from dietary SAA in anteaters. Adult anteaters appear able to synthesize TAU when diets contain adequate SAA, but dietary TAU may be critical if protein intakes are low or of poor quality. This study may provide guidance on choice of domestic canids vs. felids as suitable physiologic models for improved nutrition in giant anteaters, and also outlines a noninvasive method for assessing TAU status/metabolism that may be useful across species.

Chaetophractus villosus as a sentinel organism: Baseline values of mitotic index, chromosome aberrations and sister chromatid exchanges.

Sentinel species are useful tools for studying the deleterious effects of xenobiotics on wildlife. The large hairy armadillo (Chaetophractus villosus) is the most abundant and widely distributed mammal in Argentina. It is a long-lived, omnivorous, burrowing species, with fairly restricted home ranges. To evaluate the level of spontaneous genetic damage in this mammal, we determined the baseline values of several genotoxicity biomarkers. The study included 20 C. villosus adults of both sexes from eight pristine localities within its geographic distribution range. Genotoxicity analysis was performed on 72-h lymphocyte cultures, using mitomycin C as positive control. We obtained the baseline values of mitotic index (MI=10.52±0.30 metaphases/total cells, n=20), chromosome aberrations (CA=0.13±0.22, n=20), sister chromatid exchanges (SCE)=6.55±0.26, n=6) and replication index (RI=1.66, n=6). MI and CA did not show significant differences (P>0.05) among localities or between sexes. No significant differences in MI, CA, SCE, and RI (P>0.05) were found between values from the pristine localities and historical data. There were significant differences in CA, SCE, and RI (P<0.05) between lymphocyte cultures from pristine localities and those exposed to mitomycin C. We propose the large hairy armadillo as a sentinel organism for environmental biomonitoring of genotoxic chemicals due to its abundance, easy manipulation, well-known biology, the fact that it is usually exposed to different mixtures and concentrations of environmental contaminants, and the baseline values of genetic damage characterized by MI, CA, SCE and RI as biomarkers.

Fecal estrogen, progestagen and glucocorticoid metabolites during the estrous cycle and pregnancy in the giant anteater (Myrmecophaga tridactyla): evidence for delayed implantation.

BACKGROUND: Declining numbers of wild giant anteaters highlight the importance of sustainable captive populations. Unfortunately, captive reproductive management is limited by the lack of external physical indicators of female reproductive status and the aggressive behavior of males. We examined the endocrinology of the estrous cycle and pregnancy, and whether delayed implantation is a gestational strategy for giant anteaters as described for other xenarthrans. METHODS: Feces were collected from seven captive females 3-5 times weekly and mating was recorded. Concentrations of estrogen (estrone-glucuronide, E1, and estradiol-17ß, E2), progestagen (20-oxo-progestagens, P4), and glucocorticoid (GC) metabolites were examined in fecal extracts by enzyme immunoassay. RESULTS: Estrous cycles for nulliparous females (6 cycles, n = 2) compared to the multiparous female (6 cycles, n = 1) were shorter (47.3 +/- 4.3 days versus 62.5 +/- 2.6 days) with relatively lower luteal phase concentrations of P4 (49.4 +/- 2.9 ng/g versus 136.8 +/- 1.8 ng/g). The four remaining females had unclear ovarian activity: two females exhibited apparent luteal activity but unclear fluctuations in estrogens, while the other two females had parallel fecal P4 and estrogens concentrations. Pregnancy ranged 171-183 days with females returning to estrus post-partum as early as 60 days (n = 3, 1.8-4 years of age at mating). Delayed implantation was indicated by a biphasic elevation in fecal P4 metabolites: the initial 4-fold increase occurred for 81-105 days and was followed by a 26-fold secondary rise in P4 metabolites lasting 66-94 days prior to parturition. Fecal GC was correlated with fecal estrogens and greatest during estrus, late pregnancy, and six days prior to parturition (estrous cycle GC, 14.4-62.8 ng/g; pregnancy GC, 13.6-232.7 ng/g). CONCLUSIONS: Estrous cycles of giant anteaters occurred year-round, but were shorter and more intermittent in younger nulliparous animals compared to a multiparous female. A pronounced elevation in fecal P4, estrogen, and GC occurred during late gestation after an initial post-mating delay providing evidence for delayed implantation. Adrenocorticoid activity indicated impending parturition. Differences in estrous cycle characteristics with age and the protracted but variable gestation length must be considered to improve reproductive success and neonatal survival in giant anteaters.

The fetomaternal interface in the placenta of three species of armadillos (Eutheria, Xenarthra, Dasypodidae).

BACKGROUND: Placental characters vary among Xenarthra, one of four supraordinal clades of Eutheria. Armadillos are known for villous, haemochorial placentas similar to humans. Only the nine-banded armadillo has been well studied so far. METHODS: Placentas of three species of armadillos were investigated by means of histology, immunohistochemistry including proliferation marker, and transmission and scanning electron microscopy. RESULTS: The gross anatomy differed: Euphractus sexcinctus and Chaetophractus villosus had extended, zonary placentas, whereas Chaetophractus vellerosus had a disk. All taxa had complex villous areas within the maternal blood sinuses of the endometrium. Immunohistochemistry indicated the validity of former interpretations that the endothelium of the sinuses was largely intact. Tips of the villi and the columns entering the maternal tissue possessed trophoblast cell clusters with proliferation activity. Elsewhere, the feto-maternal barrier was syncytial haemochorial with fetal vessels near the surface. CONCLUSIONS: Differences among armadillos occurred in regard to the extension of the placenta, whereas the fine structure was similar. Parallels to the human suggest that armadillos are likely to be useful animal models for human placentation.

Energy intake for maintenance in a mammal with a low basal metabolism, the giant anteater (Myrmecophaga tridactyla).

Giant anteaters (Myrmecophaga tridactyla) are among those mammals for which a particularly low metabolism has been reported. In order to verify presumably low requirements for energy, we used eight anteaters (two males, six females; aged 1-14 years; body mass between 46 and 64 kg) in a total of 64 individual trials, in which a variety of intake levels was achieved on various diets. Digestible energy (DE) intake was quantified by measuring food intake and faecal excretion and analysing representative samples for gross energy, and animals were weighed regularly. Maintenance DE requirements were calculated by regression analysis for the DE intake that corresponded to zero weight change. Differences between individuals were significant. Older anteaters (n = 3 animals aged 12-15 years in 29 trials) had lower relative requirements than younger ones (n = 5 animals aged 1-7 years in 35 trials); thus, giant anteaters resemble other mammals in which similar age-specific differences in energy requirements are known. However, estimated maintenance requirements were 347 kJ DE/kg(0.75)/day in the anteaters, which is low compared to the 460-580 kJ DE/kg(0.75)/day maintenance requirements of domestic dogs. The lack of knowledge that metabolic requirements are below the mammalian average could make species particularly susceptible to overfeeding, if amounts considered adequate for average mammals were provided. Non-scientific reports on comparatively fast growth rates and high body masses in captive giant anteaters as compared to free-ranging animals suggest that body mass development and feeding regimes in captivity should be further assessed.

Seasonal reproduction in male pichis Zaedyus pichiy (Xenarthra: Dasypodidae) estimated by fecal androgen metabolites and testicular histology.

Poaching poses a threat to a wide variety of wildlife, and basic information about the biology of hunted species needs to be collected before their populations decline to the extent that requires drastic human intervention. As the survival of a species is related to its ability to reproduce, data on its reproductive cycle are necessary for the development of management strategies. The hypothesis was tested that the reproductive season of pichis (Zaedyus pichiy), small hibernating armadillos that inhabit arid environments in Argentina and Chile, is limited to spring months. Gonadal competence of semi-captive and wild-caught male pichis of Mendoza Province, Argentina was studied, by measuring fecal immunoreactive testosterone concentrations and evaluating spermatogenic activity. Results suggest that Z. pichiy is a seasonal breeder that regulates reproduction through photoperiodic cues. Gonadal competence was limited to a period of 3-5 months in spring and early summer and was reflected in enlarged testes, increased spermatogenesis, and significantly elevated fecal immunoreactive testosterone concentrations. The reproductive season for males from southern Mendoza was almost 6 weeks shorter than in the north. This fact, along with significant morphological differences between both groups, suggests that northern and southern pichis belong to two distinct populations. It is concluded that prolonged breeding seasons and more favorable environmental conditions in northern Mendoza favor a prolongation of the reproductive season that may allow pichis to breed later in the year, thus maximizing reproductive opportunities.

Conformational free energy of armadillo metmyoglobin.

The conformational free energy of armadillo metmyoglobin was examined over a pH range of 4.4-8.0 and a guanidinium chloride concentration of 0-2.3 M. For isothermal unfolding at 25 degrees essentially the same value was obtained for the conformational free energy from all the data: 27 +/- 2 kJ/mol. These data suggest that the armadillo has the least stable metmyoglobin of any mammal thus far examined. The cooperativity of the unfolding with respect to denaturant is considerably less than for other mammalian myoglobins. On unfolding only three to four side chains with a pKA of 6 in the unfolded protein are protonated instead of the six found for horse and sperm whale myoglobins.

Classical bile acids in animals, beta-phocaecholic acid in ducks.

1. Bile samples of different animals were analysed and the percentage content of classical bile acids was determined. 2. Herbivorous birds mostly excreted a large proportion of chenodeoxycholic acid. 3. The anteater (Myrmecophaga tridactyla) excreted deoxycholic acid most probably as a primary bile acid. 4. In the bile of ducks (Anas platyrhynchos) a large amount of (23R)3 alpha, 7 alpha, 23-trihydroxy-5 beta-cholan-24-oic acid (beta-phocaecholic acid) was found.

Lactate dehydrogenase in Mycobacterium leprae grown in armadillo liver.

Lactate dehydrogenase (LDH) was detected in extracts of untreated and NaOH-treated Mycobacterium leprae. Since armadillo liver LDH isoenzymes with a similar electrophoretic mobility were shown to be considerably more sensitive to inhibition by oxamate than LDH in M. leprae extracts, it was confirmed that M. leprae grown in armadillo liver has its own LDH. Neither the activity of LDH in M. leprae nor its electrophoretic mobility supported the tentative suggestion that an "anomolous" LDH isoenzyme in infected skin biopsies was in fact an M. leprae-derived enzyme.

Variation of superoxide dismutase levels in extracts of Mycobacterium leprae from armadillo liver.

Recent improvements in the sensitivity of assay methods for superoxide dismutase (SOD) have enabled the detection of this enzyme in 18 cell-free extracts of purified Mycobacterium leprae. By converting back to units of SOD obtained in the cytochrome c-based method previously used in work on this enzyme in mycobacteria, it was shown that extracts of M. leprae had 0.15-3.84 U SOD/mg protein (this study). A mean value of 1.31 U/mg protein was calculated. It was not possible to find any factors which could explain the very high levels in some extracts, although correlation with the period of tissue storage at -80 degrees C suggested that M. leprae in freshly killed tissue would have 1.77 U SOD/mg protein. The possibility of contamination by SODs from host and other organisms was unlikely since on gel electrophoresis extracts of M. leprae with high levels of SOD showed only a single band of activity characteristic of manganese-dependent SOD previously demonstrated.

Isolation and convulsivant effect of the "insulin-like" protein obtained from the exocrine pancreas of Bradypus tridactylus L.

1. A pancreatic insulin-like protein fraction with low electrophoretic mobility showing high molecular weight is present. 2. Isoelectric focussing studies showed that the high molecular weight protein has a pI about 7.4 when used at a pH range between 3.5 and 8.0. 3. The partially purified aldehyde-fuchsin positive high molecular weight protein fraction gave a positive effect for the convulsivant test in mice. 4. A high molecular weight insulin-like protein in pancreatic juice was found. 5. Insulin was found by radioimmunoassay in the basal and post stimulation pancreatic juice.

Autoradiographic demonstration of estrogen uptake by the armadillo pituitary gland.

Adult female armadillos were injected with tritiated estradiol. The pituitary gland was removed and processed for autoradiography. Nuclear uptake and retention of the steroid was found in 28.8% of the cells in the pars distalis and 3.7% of the pituicytes.