The longitudinal association of eating behaviour and ADHD symptoms in school age children: a follow-up study in the RHEA cohort.

Previous evidence suggests a link between attention deficit hyperactivity disorder (ADHD) symptoms and disordered eating behaviours; however, the direction of the causal association remains unclear. Building on our previous research, we aimed to examine the longitudinal association between eating behaviours at 4 years, ADHD symptoms at 6 years of age, and the role of body mass index (BMI). We included children from the RHEA mother-child cohort in Greece, followed up at 4 and 6 years (n = 926). Parents completed the Children's Eating Behaviour Questionnaire (CEBQ) to assess children's eating behaviour at 4 years and the ADHD Test (ADHDT) and Child Behaviour Checklist for ages 6-18 (CBCL/6-18) to evaluate ADHD symptoms at 4 and 6 years, respectively, as well as measures of BMI. Longitudinal structural equation modeling (SEM) was carried out to evaluate the associations of all variables between 4 and 6 years. Food responsiveness at 4 years was positively associated with hyperactivity at age 6, whereas emotional overeating was negatively associated with hyperactivity. There was no evidence of an association between eating behaviours of preschoolers and BMI at 6 years, or BMI at 4 years and later ADHD symptoms and vice versa. Findings suggest that food responsiveness is an early marker of ADHD symptoms at 6 years of age. In contrast to our hypothesis there was no significant association between ADHD at age 4 and BMI at age 6.

Enzyme annotation in UniProtKB using Rhea.

MOTIVATION: To provide high quality computationally tractable enzyme annotation in UniProtKB using Rhea, a comprehensive expert-curated knowledgebase of biochemical reactions which describes reaction participants using the ChEBI (Chemical Entities of Biological Interest) ontology. RESULTS: We replaced existing textual descriptions of biochemical reactions in UniProtKB with their equivalents from Rhea, which is now the standard for annotation of enzymatic reactions in UniProtKB. We developed improved search and query facilities for the UniProt website, REST API and SPARQL endpoint that leverage the chemical structure data, nomenclature and classification that Rhea and ChEBI provide. AVAILABILITY AND IMPLEMENTATION: UniProtKB at https://www.uniprot.org; UniProt REST API at https://www.uniprot.org/help/api; UniProt SPARQL endpoint at https://sparql.uniprot.org/; Rhea at https://www.rhea-db.org.

Ontogenetic shape changes in the pelvis of the Greater Rhea (Aves, Palaeognathae) and their relationships with cursorial locomotion: a geometric morphometric approach.

Knowledge of the ontogenetic pattern of morphological features is essential to improve biological interpretations. The study of morphological features of the pelvic girdle and hind limb apparatus throughout growth is an excellent approach to understand how the skeletal morphology and muscles are interrelated during growth in a bird with a specialized mode of locomotion. The Greater Rhea (Rhea americana) is a large cursorial palaeognathous bird with long legs and powerful musculature. The postnatal shape changes of the pelvis of this bird were studied with geometric morphometric techniques, using landmarks and semilandmarks. In addition, regression analyses were used to explore the association between pelvic shape changes with muscle and body mass. The pelvises of 16 specimens of Rhea americana from 1 month old to adulthood were studied in dorsal and lateral views. Noticeable differences in pelvic shape were noted between ages, particularly in lateral view. In young birds, the pre- and post-acetabular ilium was subequal in length, whereas in adults the pre-acetabular ilium became shorter. In dorsal view, the main shape changes observed were the progressive thinning of both ilium portions and the elongation of the vertex craniolateralis ilii from chicks to adulthood. In this view, the only clear differentiation was between young and adult birds. Shape differences were influenced by body mass and pelvic muscles; the post-acetabular muscle mass explained the highest percentage of the variation. The specialized locomotion of Greater Rhea is reflected in their pelvic musculoskeletal system, in which the change to a longer post-acetabular ilium correlates with the growth of the powerful post-acetabular muscles. The actions of these muscles provide the necessary strength to support the body mass, minimize the body swinging movements and propel the body forward during locomotion. Bone morphology is affected by the forces produced by body mass and the muscle activity, demonstrating the presence of common growth mechanisms, which are primordial and gave rise to a functional and properly proportioned adult.

Genetic identification of the ciliates from greater rheas (Rhea americana) and lesser rheas (Rhea pennata) as Balantioides coli.

The ciliate species Balantioides coli can be cross-transmitted between humans and several animal species. Usually harmless, sometimes it can be pathogenic and cause the death of the host. In birds, B. coli has been confirmed in ostriches by genetic analysis, but the identification from South American greater rheas (Rhea americana) and lesser rheas (Rhea pennata pennata) is tentative. Since these species are reared for commercial purposes and for reintroduction into the wild, it is necessary to elucidate whether the ciliate from rheas is B. coli to minimize health risks for humans and for other domestic and wild animals. Individual parasite cells are collected from Argentinean isolates of reared greater rheas and of wild and reared lesser rheas, and their ITS region was PCR amplified; the cloning products were sequenced and compared with sequences available in public databases. The results have shown that several sequence types are expressed at the same time in the parasite cells, and all correspond to B. coli, confirming the possibility of cross-transmission of the parasite between wild and reared South American rheas and several mammal species and humans.

Effect of maternal environment on yolk immunoreactive corticosterone and its influence on adrenocortical and behavioral activity in chicks of Greater Rhea (Rhea americana).

Maternal corticosterone in avian eggs may modify offspring phenotype in order to increase survival in poor environments. In the Greater Rhea (Rhea americana), we previously found that yolk immunoreactive corticosterone is influenced by the quality of the maternal environment: eggs laid by females of the intensive rearing system (IRS), living in poor captive conditions, had higher yolk immunoreactive corticosterone than those produced by females of the semi-extensive rearing system (SRS), living in better conditions. Here, we evaluate if these different hormone levels are associated with the production of different phenotypes. We collected eggs from the IRS and SRS for hormonal quantification and artificial incubation. Then, half of the chicks selected from each environment were exposed to a capture and restraint protocol, and the rest remained undisturbed and were used as controls. In the IRS, we found that higher yolk immunoreactive corticosterone was associated with the production of chicks that had reduced hatchability, lower hatchling mass and higher baseline fecal glucocorticoid metabolites (FGM) than those produced by SRS females. Moreover, after capture and restraint, IRS chicks did not modify their FGM nor their behaviors compared to their controls, while SRS chicks increased their FGM and spent more time ambulating and less time pecking, compared to their controls. These results indicate that yolk immunoreactive corticosterone could modify offspring phenotype. Although future studies are needed to elucidate their implications for fitness, our results suggest that yolk corticosterone could be mediating an adaptive maternal effect that allows individuals to better cope with poor conditions.

Anatomopathological changes, quantification and distribution of Libyostrongylus spp. in regions of the proventriculus and ventriculus of naturally- and experimentally-infected ostriches.

Nematodes of the genus Libyostrongylus parasitize ostriches, causing high mortality rates. These nematodes are found in the proventriculus and ventriculus of ostriches, but little is known about their distribution and the possible anatomopathological changes they cause in the various regions of these organs. This paper describes the distribution and quantification of Libyostrongylus and pathological changes found in regions of the proventriculus and ventriculus of ostriches with high and low levels of both natural and experimental infection. Ostriches were necropsied and tissue samples from the distinct regions of both organs were analysed based on nematode counts and histopathology after staining with haematoxylin and eosin, Masson's trichrome or Alcian blue/PAS. The cranial and glandular regions of the proventriculus were the most parasitized. The ventriculus contained more nematodes in the caudal region. No macro- or microscopic pathological changes were observed in either of these organs of experimentally-infected birds. However, naturally-infected birds with high levels of infection presented proventriculus with macroscopic lesions and heterophilic infiltrates surrounding nematodes. In the glandular region of this organ, nematodes were located in the adenomeres of the secretory ducts, causing altered architecture and erosions and ulcerative lesions with damaged epithelium. Nematode eggs were found in the koilin layer of the middle and caudal regions of the ventriculus only of these birds. The pH of the regions assessed by Alcian blue/PAS staining changed from acidic in the proventriculus to more alkaline in the caudal region of the ventriculus. These data add knowledge to the biology of Libyostrongylus. RESEARCH HIGHLIGHTS The most parasitized areas were the cranial and glandular regions of the proventriculus. Naturally-infected birds with high levels of infection presented macro lesions in the proventriculus and damaged epithelium. Nematode eggs were found in the ventriculus. The proventriculus had an acidic pH, which turned alkaline towards the ventriculus.

State of the art knowledge in adrenocortical and behavioral responses to environmental challenges in a threatened South American ratite: Implications to in situ and ex-situ conservation.

The Greater Rhea (Rhea americana) is an endemic ratite to South America, whose wild populations have undergone a remarkable decrease due to habitat degradation and fragmentation by the expansion of the agricultural frontier, poaching and predation by dogs. Anthropogenic perturbations in wild environments, as well as the management in captivity, can generate different stress responses in this species, thus, the monitoring of adrenocortical and behavioral activities are considered primary assessment tools with both conservation and welfare implications. In this review we analyze and integrate the different measurements of glucocorticoids (in plasma, feces, and yolk) carried out in different captive and wild populations, taking into account the diverse predictable and unpredictable conditions to which the Greater Rhea responds in each of those environments. In addition, the translocation of this bird is presented as an application of stress physiology in field ecology for conservation purposes, in which we evaluated how this species responds when it is released into a novel environment. Our results indicate that this ratite has a striking high sensitivity of the hypothalamic-pituitary-adrenal (HPA) axis compared to that of other bird species and shows a wide variety of adrenocortical responses depending on the environment in which it lives. This suggests that its HPA axis has a phenotypic plasticity that enables the rhea to cope with the environmental challenges. In this sense, we propose that one of the routes of this plasticity could be mediated by the maternal transfer of steroid hormones to the egg. Finally, we discuss the importance of integrating the monitoring of the adrenocortical response along with the environmental variables that define the life history of the species, in management and conservation programs ex-situ and in situ.

Redescription of Procyrnea uncinipenis (Molin, 1860) (Nematoda: Habronematidae) based on material from Rhea americana (L.) (Aves: Rheidae).

A detailed description of Procyrnea uncinipenis (Molin, 1860) (Habronematidae) is provided based on light- and scanning electron microscopy of newly collected material ex Rhea americana (L.) from a private conservation breeding park located in Cachoeiro de Itapemirim (State of Espírito Santo, Brazil). Of the four host specimens analysed, two were infected with P. uncinipenis; a total of 441 nematodes were collected in the proventriculus and gizzard. The specimens represented large nematodes, sexually dimorphic, with females larger than males, reddish in vivo, with two well developed lateral pseudolabia and two interlabia, one dorsal and one ventral, long oesophagus, divided into a muscular and glandular portion, didelphic ovaries, opisthodelphic uteri, two phasmids and a rounded chitinous structure near the caudal end. Males possess posterior extremity curved in spiral ventrally, with bilateral caudal alae and unequal in size and shape spicules. This study adds new morphological data, thus contributing to the knowledge on P. uncinipenis, a common parasite of R. americana.

Effect of several packaging conditions on the microbiological, physicochemical and sensory properties of ostrich steaks during refrigerated storage.

A total of 365 ostrich steaks were packaged in air (AIR), vacuum (VAC), MAP1 (70% O2 + 30% CO2), MAP2 (30% O2 + 30% N2 + 40% CO2), MAP3 (20% O2 + 30% N2 + 50% CO2), MAP4 (50% N2 + 50% CO2), MAP5 (20% N2 + 80% CO2) or MAP6 (100% CO2). Microbial counts (10 groups), pH, Aw and sensory properties (nine-point hedonic scale) were determined on days 0, 1, 3, 7 and 15 of storage (4 °C). On day 0, microbial counts (log10 cfu/g) ranged from undetectable levels (Brochothrix thermosphacta, enterococci) to 3.21 ± 0.63 (total aerobic counts -TAC-). The highest and the lowest microbial loads throughout storage were observed in AIR and MAP6, respectively. On day 15 TAC as high as 9.96 ± 0.20 log10 cfu/g were found in AIR. The shelf-life (time until overall acceptability score fell below 5) was 3 days (MAP1, MAP2), 7 days (MAP3, AIR) or 15 days (MAP4, MAP5, MAP6). Only for VAC the shelf-life limit extended beyond 15 days.

A new species of Procyrnea (Nematoda: Habronematidae) parasitic in Rhea pennata (Aves: Rheidae) from Patagonia, Argentina, with a key to species of the genus.

Procyrnea choique n. sp. is described from the lesser rhea, Rhea pennata d´Orbigny (Aves: Rheidae), from the Protected Natural Area Península Valdés, Chubut province, Argentina. The new species resembles P. ficheuri, P. murrayi, P. excisiformis, P. dollfusi, P. haliasturi, P. anterovulvata, P. graculae, P. brevicaudata, P. uncinipenis, P. javaensis, P. ameerae, P. ornata, P. aegotheles, P. spiralis, P. ruschii and P. aptera, mainly in the absence of lateral alae; but differs from its congeners by having a left spicule without barbs, the position of the vulva which is post-equatorial, the absence of lateral ridges, absent median precloacal papilla, asymmetrical caudal alae, males more than 6 mm long, left spicule 1 mm long and spicule ratio 1:3. Our results extend the taxonomy of Procyrnea Chabaud, 1958 and comprise the first report of a habronematid from R. pennata. A key to species of Procyrnea is presented.

RETROSPECTIVE STUDY OF MORTALITY IN CAPTIVE STRUTHIONIFORMES IN A FRENCH ZOO (1974-2015).

The husbandry and medical records, and necropsy reports, of 1,002 captive Struthioniformes that died at the Réserve Africaine de Sigean (France) from 1974 to 2015 were examined. The goal of this study was to determine the most common causes of mortality in ostriches ( Struthio camelus), emus ( Dromaius novaehollandiae), and American rheas ( Rhea americana), in order to highlight common causes of mortality, as well as the influence of age, gender, and rearing methods on mortality. The most prevalent cause of mortality was pathology of digestive origin in the neonates of all three species, especially yolk-sac infections, which accounted for 41% of all deaths in this captive neonate population and was especially prevalent in hand-reared neonates. Other causes included musculoskeletal disorders in emus (particularly hand-reared) and rheas; trauma in neonate ostriches, mainly due to crushing by parents; leg deformities in emu chicks and juveniles; general sepsis in hand-reared ratite chicks due to a chlamydiosis outbreaks (1989-1990); trauma by conspecifics in subadult ostriches and emus; stress myopathy in subadult rheas, particularly after introduction to a new enclosure; evisceration inflicted by herbivorous enclosure mates on adult male ostriches; fatal peritonitis following salpingitis in adult female ostriches; and death associated with ocular disorder in adult male emus. Although this study was conducted at one institution, and its results reflect this zoo's management and zootechnical practices, its findings could well have implications for management practices involving other captive Struthioniforme populations.

Morphology of the nematode Deletrocephalus dimidiatus Diesing, 1851 from the rhea, Rhea americana Linnaeus, 1758, together with a key to species of Deletrocephalinae.

The family Deletrocephalidae consists of three species, from two genera: Deletrocephalus dimidiatus, D. cesarpintoi and Paradeletrocephalus minor, which differ from one another in terms of the buccal capsule structure, the dorsal ray from the male copulatory bursa and the length of the vagina in the female. All these species are parasites of the gastrointestinal tract of the rhea, Rhea americana. Only D. dimidiatus was reported to be a parasite of the other rhea species, Darwin's rhea, R. pennata. Currently, there are no studies on the ultrastructure and biology of these parasites or their pathogenicity in these birds. Thus, the present study aimed to examine the ultrastructure and add more details of the morphology of D. dimidiatus from R. americana. Four adult rheas were necropsied, and the gastrointestinal tract was collected and examined for the presence of parasites. Nematodes were analysed by optical and scanning electron microscopy (SEM). All birds were parasitized by D. dimidiatus. Some morphological and morphometric characters were observed that differ from reports of other studies of this species by other authors. In addition, new details were added, well described by SEM, and a key to the species of the subfamily Deletrocephalinae is given.

Influence of the rearing system on yolk corticosterone concentration in captive Greater Rheas (Rhea americana).

Many environmental conditions elevate plasma corticosterone in laying birds, leading to elevated hormone accumulation in the egg. We investigated whether maternal yolk corticosterone levels in Greater Rheas differ between fresh eggs collected from an intensive (IRS) and a semi-extensive (SRS) rearing system. After HPLC validation, yolk corticosterone was measured using a corticosterone (125) I radio-immunoassay kit. Results (mean ± SE) showed that eggs collected from the IRS exhibited a significantly higher corticosterone concentration than eggs from SRS (89.88 ± 8.93 vs. 45.41 ± 5.48 ng/g yolk, respectively). Our findings suggest that rearing conditions under an intensive scheme (e.g., small pens with bare ground, no direct foraging and handling) might be perceived as more stressful for Greater Rhea females than semi-extensive rearing conditions (e.g., low animal density distributed in extensive areas and direct foraging), which would result in the transfer of higher yolk corticosterone levels. A better understanding of environmental conditions and female traits that affect yolk corticosterone deposition provides a background for future studies concerning the roles of maternal corticosterone on offspring development. Zoo Biol. 35:246-250, 2016. © 2016 Wiley Periodicals, Inc.

USE OF COMPUTED TOMOGRAPHY FOR INVESTIGATION OF RHINITIS AND SINUSITIS IN A GREATER RHEA (RHEA AMERICANA).

Sinusitis is a common finding in avian species; however, it presents unique treatment challenges in ratites. A 6-yr-old male greater rhea ( Rhea americana ) presented acutely with right infraorbital sinus swelling and chemosis. Oral antibiotics were prescribed based on bacterial culture results with incomplete resolution of clinical signs. A computed tomography scan of the skull demonstrated a mixed-density soft tissue mass with extensive destruction of the right nasal bone, hard palate, maxilla, and frontal bone. The affected tissue was surgically resected and histopathology was consistent with inspissated necrotic tissue. Aerobic culture of the mass grew a multidrug-resistant Escherichia coli and was treated with amikacin. Ten months postsurgery, no further clinical signs were observed. Advanced diagnostic imaging with computed tomography and surgical therapy facilitated complete resolution of this resistant bacterial sinusitis in a species that is not typically amenable to frequent handling and restraint.

Running, hopping and trotting: tuning step frequency to the resonant frequency of the bouncing system favors larger animals.

A long-lasting challenge in comparative physiology is to understand why the efficiency of the mechanical work done to maintain locomotion increases with body mass. It has been suggested that this is due to a more elastic step in larger animals. Here, we show in running, hopping and trotting animals, and in human running during growth, that the resonant frequency of the bouncing system decreases with increasing body mass and is, surprisingly, independent of species or gait. Step frequency roughly equals the resonant frequency in trotting and running, whereas it is about half the resonant frequency in hopping. The energy loss by elastic hysteresis during loading and unloading the bouncing system from its equilibrium position decreases with increasing body mass. Similarity to a symmetrical bounce increases with increasing body mass and, for a given body mass, seems to be maximal in hopping, intermediate in trotting and minimal in running. We conclude that: (1) tuning step frequency to the resonant frequency of the bouncing system coincides with a lower hysteresis loss in larger, more-compliant animals; (2) the mechanism of gait per se affects similarity with a symmetrical bounce, independent of hysteresis; and (3) the greater efficiency in larger animals may be due, at least in part, to a lower hysteresis loss.

Comparative methane emission by ratites: Differences in food intake and digesta retention level out methane production.

Ratites differ in the anatomy of their digestive organs and their digesta excretion patterns. Ostriches (Struthio camelus) have large fermentation chambers and long digesta retention, emus (Dromaius novaehollandiae) have a short gut and short retention times, and rheas (Rhea americana) are intermediate. A recent study showed that ostriches produce as much methane (CH4) as expected for a similar-sized, non-ruminant mammalian herbivore. We hypothesized that emus and rheas produce less CH4 than ostriches. We individually measured, by chamber respirometry, the amount of O2 consumed as well as CO2 and CH4 emitted from six adult rheas (body mass 23.4±8.3 kg) and two adult emus (33.5 and 32.0 kg) during 23-hour periods on a pelleted lucerne diet. In contrast to previous studies, which classified emus as non-producers, we measured CH4 emissions at 7.39 and 6.25 L/day for emus and 2.87±0.82 L/day for rheas, which is close to values expected for similar-sized non-ruminant mammals for both species. O2 consumption was of a similar magnitude as reported previously. Across ratites, CH4 yield (L/kg dry matter intake) was positively correlated with mean retention time of food particles in the gut, similar to findings within ruminant species. In ratites, this relationship leads to similar body mass-specific CH4 production for a high intake/short retention and a low intake/long retention strategy. Therefore, when investigating CH4 production in herbivorous birds, it is advisable to consider various CH4 measures, not only yield or absolute daily amount alone.

Influence of breeding season on fecal glucocorticoid levels in captive Greater Rhea (Rhea americana).

Sex hormones and stress-related changes can be seasonally influenced. We investigate whether fecal glucocorticoid metabolite (FGM) levels can differ between male and female captive Greater Rheas during the breeding and non-breeding seasons. Over a 3-year-period, fresh fecal samples from 10 individuals (five of each sex) were collected during the breeding months (October, November, and December) and non-breeding months (April and June). A total of 960 samples were assayed using a commercial radioimmunoassay. Results showed that FGM levels (mean ± SE) were affected by the breeding season in a sex-dependent way. Male Greater Rheas showed significantly higher FGM levels in the breeding months than in the non-breeding months (13.44 ± 0.37 vs. 7.92 ± 0.1 ng/g feces, respectively). By contrast, females did not show FGM seasonal changes throughout the same sampling periods (7.55 ± 0.14 vs. 7.26 ± 0.73 ng/g feces). Moreover, during the breeding season months, males showed higher average FGM levels than females (13.44 ± 0.37 vs. 7.55 ± 0.14 ng/g feces, respectively), and no differences were found between sexes during the non-breeding season (7.92 ± 0.1 vs. 7.26 ± 0.73 ng/g feces, respectively). Our findings suggest that male Greater Rheas have a higher adrenocortical activity during the breeding season, which is probably indirectly related to the increased testosterone levels and agonist interactions that are also observed during that phase. Studies aimed to determine the appropriate sex ratio for captive rearing should be developed to minimize male agonist encounters and therefore improve welfare of the captive group.

Allometric scaling for chemical restraint in greater rheas (Rhea americana) with tiletamine and zolazepam.

BACKGROUND: Chemical restraint is of great importance in the clinical practice of wildlife animals. In such, interspecific allometric scaling proposes pharmacological doses to a wide range of species, based on previously known doses for domestic animals and the target animal's body mass. The objective was to compare chemical restraint responses in the greater rhea (Rhea americana) with conventional doses of tiletamine/zolazepam, found in the literature for the species, and with doses calculated through interspecific allometric scaling extrapolation. From the Federal University of Piauí, six adult greater rheas (Rhea americana), three males and three females, were randomly selected to be subjects in this research. All six animals were submitted to two chemical restraint protocols with tiletamine and zolazepam, per intramuscular injection in the hind limb. The first protocol was composed of doses found on the literature for the species, while the second protocol used doses calculated by interspecific allometric scaling, with the domestic dog as model animal. Heart and respiratory rates, body temperature, eyelid reflex, digital pinch and metatarsal reflex were registered along with latency and ambulation times. RESULTS: The use of interspecific allometric scaling for chemical restraint with the combination tiletamine and zolazepam showed satisfying results, with great similarity to results obtained with conventional doses in Greater rheas. CONCLUSIONS: Literature on chemical restraint and use of tiletamine and zolazepam in rheas is scarce. Chemical restraint is of extreme importance on these animals, due to their aggressive nature and low level of domesticity. This research may further establish the interspecific allometric scaling method as a viable tool for the veterinary physician in formulating anesthetic and chemical restraint protocols for wildlife animals.

Hindlimb bones texture through postnatal ages of Rhea americana (Aves: Palaeognathae).

The bone texture of Rhea americana was evaluated through the examination of a postnatal ontogenetic series. The hind limb bone surfaces of specimens of one, three and five months old, and adults were compared to characterize each stage according to the morphological features generated by their differential ossification. The results suggest a similar process of tissue maturation concerning neognathous birds, although with some differences. A spongy or striated surface with abundant pores in the femur and longitudinal grooves in the tibiotarsus and tarsometatarsus characterizes chicks. Vascularity decreases, and the bone texture gradually changes acquiring a more homogeneous surface, to eventually reach the smooth appearance of adult bones. The establishment of particular textures corresponding to different bones and ontogenetic stages permits the accurate interpretation of remains in ecological, archaeological and paleontological contexts in which bones are fragmented and/or incomplete.