Very virulent infectious bursal disease virus infection caused changes in cloacal temperature and clinical manifestations in pigeons (Columba livia domestica) and is transmitted to sentinel chickens.

In this study, changes in cloacal temperature and clinical manifestations due to very virulent infectious bursal disease virus (vvIBDV) infection in pigeons (Columba livia domestica) and transmission to chickens were demonstrated. Thirty pigeons (3-6 weeks old) and thirty chickens (3 weeks old) divided into 4 groups (I-IV) were used for this study. Group I comprised of 10 uninoculated pigeons only; II comprised of 10 inoculated pigeons and 10 sentinel chickens; III comprised of 10 sentinel pigeons and 10 inoculated chickens, while IV comprised of 10 uninoculated chickens only. Pigeons in group II and chickens in group III were each inoculated with 0.20 mL (titre of 109.76CID50/mL) of vvIBDV (Nigerian strain). Cloacal temperature was monitored and clinical manifestations scored post-inoculation (pi). Results indicated significant (P < 0.05) pyrexia at 2 days pi (dpi), mild clinical signs and no mortality in inoculated pigeons. Significant (P < 0.05) pyrexia at 2-4 dpi, severe clinical signs and mortality (50%; 60%) were observed in inoculated and sentinel chickens. IBDV antigen and antibody were detected in pigeons and chickens. Pigeons showed response to vvIBDV infection thus suggesting susceptibility of pigeons to IBD. Sentinel chickens presented clinical manifestations of IBD and this suggests transmission from pigeons to chickens. This study therefore documents pyrexia and clinical manifestations due to vvIBDV infection in pigeons and successful transmission of the virus between pigeons and chickens.

Glans inflation morphology and female cloaca copulatory interactions of the male American alligator phallus†.

The phallic glans of the American alligator (Alligator mississippiensis) is the distal termination of the semen-conducting sulcus spermaticus and during copulation has the closest, most intimate mechanical interactions with the female urodeum, the middle cloacal chamber that contains the opening to the vaginal passages and oviducts. However, the details of this interface leading to insemination and gamete uptake are unclear. Here, we: (1) histologically characterize the underlying tissue types and morphologically quantify the shape changes associated with glans inflation into the copulatory conformation, (2) digitally reconstruct from MRI the 3D shape of functional tissue compartments, and (3) diffusible iodine-based contrast-enhanced computed tomography image the copulatory fit between male phallus and female cloaca. We discuss these results in relation to tissue type material properties, the transfer on intromittent forces, establishing potential copulatory lock, inflated glans volume scaling with body mass/length, the mechanics of semen targeting and insemination, and potential female cryptic choice impacting multiple clutch paternity. In part, this study further clarifies the phallic morphological variation observed among crocodylians and begins to investigate the role(s) these divergent male forms play during copulation interacting with female cloacal forms to increase reproductive success.

Evidence from Tarentola mauritanica (Gekkota: Phyllodactylidae) helps validate thermography as a tool to infer internal body temperatures of lizards.

Infrared (IR) thermal imaging has become an increasingly popular tool to measure body temperature of animals. The high-resolution data it provides with short lag and minimum disturbance makes it an appealing tool when studying reptile thermal ecology. However, due to the common phenomenon of regional heterothermy and surface-to-core temperature gradients, it is essential to select the appropriate body part to measure and provide calibrations to accurately infer internal body temperatures. This work follows from a previous study on lacertid lizards to assess the reliability of thermography-measured body temperatures, from several body locations, as a proxy for internal body temperature in lizards. This study focuses on the Moorish gecko, Tarentola mauritanica, due to its distant phylogenetic relationship and its different ecology and morphology from the previously tested species. A total of 60 adult geckos of both sexes and of a range of sizes were tested in thermal gradients and subjected to a sequence of randomly assorted treatments of heating and cooling. The temperatures of the animals were periodically measured with a thermal camera at six different body parts and, immediately after, the cloacal temperature was then measured with a thermocouple probe. Body parts' temperatures, obtained thermographically, were regressed against cloacal temperature using OLS regression and the pairwise correlations were tested using Spearman coefficients. Relationships among all body parts and between all body parts and the cloaca were strong in all cases (R2 > 0.87, Spearman Correlation > 0.95). The observed pattern was very similar to those previously obtained from lacertid lizards. Ultimately, the eye proved to provide the best overall proxy for internal temperature, when accounting for both the slope and intercept of the regression. Hence, this study provides further support for the establishment of the eye as the standard location to infer internal body temperatures of lizards through thermography.

Effects of heat stress on pullet cloacal and body temperature.

One measure of the thermal status of poultry is cloacal temperature measured with a cloacal thermometer; however, this method requires handling the bird, is invasive, and can be stressful. Infrared thermography is an alternative means for assessing bird thermal status. The objective of this study was to investigate the body temperature response of pullets subjected to different environmental air temperatures during the growing phase and to evaluate the relationship between the cloacal temperature and the body parts surface temperature. A total of 648 chicks (Lohmann LSL Lite) were used in 2 different phases, phase I (day 1 through 6 wk of age) and phase II (week 7 through 17). During phase I, chicks were reared at 1 of 3 different thermal environments: thermal comfort (35°C-19°C), mild heat stress (38°C-22°C), or mild cold stress (28°C-17°C). In phase II, pullets were randomly redistributed to 1 of 4 daytime temperature treatments: 20°C; 25°C; 30°C; and 35°C, all with night time temperature of 20°C. Cloacal temperature and body surface temperature for 8 parts (head, eye, comb, chest, back, wing, leg, head area, and body area) were obtained weekly from 4 to 2 birds per treatment, respectively, during phase II. There were no effects for the interactions between the 2 experimental phases for cloacal and body parts surface temperature. There was a strong correlation (P < 0.001) between cloacal temperature and each body part temperature; cloacal temperature followed a quadratic response to environmental air temperature treatments. Pullets subjected to 35°C/20°C and 30°C/20°C had the highest body parts temperatures compared with the other 2 treatments (P < 0.05). The leg surface temperature was greatest in all treatments, and the chest the lowest. Regression between cloacal and body parts temperature had a 95% predictive accuracy of better than 0.4°C, suggesting a useful alternative to direct cloacal temperature measurement.

Efeito da manipulação térmica durante a incubação sobre as variáveis hematológicas, bioquímica sérica e morfometria da bolsa cloacal de codornas japonesas submetidas ao estresse crônico por calor / Effect of thermal manipulation during incubation on the hematological variables, serum biochemistry and morphometry of cloacal bursa of Japanese quails submitted to chronic heat stress

Objetivou-se avaliar os efeitos da manipulação da temperatura de incubação sobre a resposta imune de codornas desafiadas termicamente após eclosão. Para isso, foram utilizados 540 ovos, distribuídos em três incubadoras, com temperatura de 37,8°C e umidade de 60%. A partir do sexto dia de incubação até a eclosão, as temperaturas foram ajustadas em 37,8°C (padrão), 38,5°C (intermediária) e 39,5°C (alta). Após a eclosão as codornas foram pesadas e distribuídas, em delineamento inteiramente ao acaso, com três temperaturas de incubação (37,8, 38,5 e 39,5°C) e duas temperaturas de ambiente (estresse e termoneutro). Aos 10, 20, 30 e 40 dias, quatro codornas por tratamento foram eutanasiadas para coleta da bolsa cloacal, do fígado e do coração, para se determinar o peso absoluto (g), o peso relativo (%) e a área dos folículos bursais. Sangue foi coletado para realização do hemograma, do leucograma e da bioquímica sérica. Os dados foram analisados e as diferenças entre as médias foram determinadas pelo teste de Tukey a 5%. O estresse térmico por calor, a partir dos 20 dias, promove redução no peso absoluto do fígado, do coração, da bolsa cloacal e na área dos folículos bursais, além de heterofilia, linfopenia e aumento da relação heterófilo/linfócito. Em conclusão, o estresse térmico por calor após 10 dias de idade pode causar imunossupressão.(AU)

The objective of this study was to evaluate the effects of manipulation of the incubation temperature on the immune response of quails challenged thermally after hatching. For this, 540 eggs were distributed in three incubators, with temperature of 37.8°C and 60% humidity. From the 6th day of incubation to hatching the temperatures were adjusted to 37.8°C (standard), 38.5°C (intermediate) and 39.5°C (high). After hatching the quails were weighed and distributed in a completely randomized design with three incubation temperatures (37.8, 38.5 and 39.5°C) and two ambient temperatures (stress and thermoneutral). At 10, 20, 30 and 40 days four quail per treatment were euthanized to collect the cloacal burse, liver and heart to determine the absolute weight (g), relative weight (%) and area of the bursal follicles. Blood was sampled for determination of hemogram, leukogram and serum biochemistry. The data were analyzed and the differences between the means were determined by the Tukey test at 5%. Heat stress from 20 days onwards promotes a reduction in the absolute weight of the liver, heart, cloacal sac and in the area of the follicles. In addition, there was heterofilia, lymphopenia and increased heterophile/lymphocyte ratio. In conclusion, heat stress after 10 days of age can cause immunosuppression.(AU)

Gut microbiota differences during metamorphosis in sick and healthy giant spiny frogs (Paa spinosa) tadpoles.

Gut microbiota plays important roles in host nutrition, immunity, development and health; therefore, disruption of the gut microbiota is closely associated with development of diseases in the host. In amphibians, metamorphosis is associated not only with extensive changes in the gut microbiota, but also with high mortality. Therefore, we hypothesized that unsuccessful restructuring of the gut microbiota during metamorphosis was an important factor that caused the fatalities. To test this hypothesis, we investigated the gut microbiota of apparently sick and healthy giant spiny frog tadpoles during metamorphosis, using high-throughput sequencing of the 16S rRNA gene. Our results showed that most dominant phyla differed significantly among developmental stages of sick and healthy Paa spinosa tadpoles. The differences in the dominant genera in sick and healthy tadpoles were the highest at the stage of degeneration of cloacal tube (TDCT). After the metamorphosis, the composition of the gut microbiota was more alike between healthy and sick tadpoles at the stage of forelimb emergence than at TDCT. These results imply that failed restructuring of the gut microbiota during metamorphosis caused the death of P. spinosa tadpoles. These results provided an important reference to prevent the high actual of P. spinosa tadpoles during metamorphosis. SIGNIFICANCE AND IMPACT OF THE STUDY: We investigated the gut microbiota of apparently sick and healthy giant spiny frog (Paa spinosa) tadpoles during metamorphosis, using high-throughput sequencing of the 16S rRNA gene. Our results showed that the differences in the dominant genera in sick and healthy tadpoles were the highest at the stage of degeneration of cloacal tube. After the metamorphosis, the composition of the gut microbiota was alike between healthy and sick tadpoles. These results imply that failed restructuring of the gut microbiota during metamorphosis caused the death of P. spinosa tadpoles.

COMPARISON OF SURFACE, ESOPHAGEAL, AND CLOACAL TEMPERATURES IN DIFFERENT REPTILE SPECIES.

The objective of this study was to compare surface, esophageal, and cloacal temperatures in awake iguanas, chameleons, and snakes at two different ambient temperatures and in alligators at one ambient temperature. Surface, esophageal, and cloacal temperatures were measured in all animals twice, with exception of the alligators, where temperatures were taken once. The first set of temperature readings was done at lower environmental temperature. An analysis of variance (ANOVA) with a mixed-effect model was used to analyze the temperature difference between esophageal and cloacal temperature against each other, and against surface temperature. Significance was set at P < 0.05. In all animals, surface, esophageal, and cloacal temperatures increased significantly (P < 0.0001) with increased ambient temperature. Esophageal and cloacal temperature were not significantly different from each other in iguanas and chameleons at lower environmental temperature or in snakes and chameleons at high ambient temperature. In snakes, esophageal temperature 26.1 ± 1.6°C was significantly higher than cloacal temperature 25.2 ± 0.9°C (P = 0.0016) at lower ambient temperature. In alligators no difference between esophageal and cloacal temperature was observed at the given ambient temperature. Surface temperature was significantly lower than esophageal and cloacal temperature in all species investigated, except iguanas at lower ambient temperature. The results of this study suggest that in healthy awake iguanas and chameleons at any ambient temperature and in snakes at high ambient temperature, cloacal temperatures are reflective of esophageal temperature in the species evaluated. In alligators, rectal temperature reflected esophageal temperature at the given ambient temperature. Surface temperature in contrast was underestimating esophageal temperature in all species investigated.

The effect of food restriction on the regulation of gonadotropin-releasing hormone in male house finches (Haemorhous mexicanus).

Seasonal activation of the vertebrate hypothalamic-pituitary-gonadal (HPG) axis and gonadal development is initiated by gonadotropin-releasing hormone-I (GnRH) release from the hypothalamus. In photoperiodic species, the consistent annual change in photoperiod is the primary environmental signal affecting GnRH cell activity, including changes in the synthesis and secretion of this neuropeptide. Non-photoperiodic environmental cues such as energy availability also influence HPG axis activity, but the mechanisms mediating this influence, in particular on the GnRH system, are unclear. Understanding how the neuroendocrine system integrates environmental information is critical in determining the plasticity and adaptability of physiological responses to changing environments. The primary objective of this study was to investigate GnRH-mediated changes in HPG axis activity and gonadal development in response to energy availability in a wild bird. We hypothesized that negative energy balance inhibits HPG axis activity by affecting GnRH secretion. Moderate food restriction for several weeks in male house finches, Haemorhous mexicanus, decreased body condition and inhibited photoinduced testicular growth compared to birds fed ad libitum. Food restriction did not affect plasma luteinizing hormone (LH; a correlate of GnRH release) or plasma testosterone, but it enhanced the plasma LH response to an injection of the glutamatergic agonist, N-methyl-D-aspartate (NMDA). Thus, food restriction may decrease photoinduced HPG axis activation by acting centrally, in particular by attenuating the release of accumulated GnRH stores.

Expression of digestive enzyme and intestinal transporter genes during chronic heat stress in the thermally manipulated broiler chicken.

Heat stress has a serious impact on nutrient digestion and absorption in broiler chickens. This study aimed to investigate the effects of chronic heat stress (CHS) on the mRNA expression of digestive enzymes and nutrient transporter genes in thermally manipulated (TM) broiler chickens. The evaluated genes encompassed pancreatic lipase, trypsin, amylase, maltase, and alkaline phosphatase as well as certain glucose transporter (GLUT2, SGLT1), amino acid transporter (y+LAT1, CAT1), and fatty acid transporter (FABP1, CD36) genes in the jejunal mucosa. Thermal manipulation was carried out at 39°C and 65% relative humidity for 18 h daily from embryonic days (ED) 10-18, while CHS was induced by raising the temperature to 35°C for 7 D throughout post-hatch days 28 to 35. After 0, 1, 3, 5, and 7 D of CHS, the pancreas and jejunal mucosa were collected from the control and TM groups to evaluate the mRNA expression by relative-quantitative real-time qRT-qPCR. Thermal manipulation significantly decreased the cloacal temperature (Tc) and the hatchling weight, and improved weight gain in broilers during post-hatch life and CHS. In addition, TM decreased the mortality rate during CHS. During CHS, the mRNA expression levels of SGLT1, GLUT2, FABP1, and trypsin were significantly decreased after 1 D in control chickens, and this lower expression persisted until day 7, after which it further decreased. In contrast, in TM chickens, SGLT1, GLUT2, and FABP1 expression decreased after 3, 5, and 7 D of CHS, respectively, while no significant change in trypsin expression was observed throughout the CHS period. Moreover, it was found that TM significantly modulated the mRNA expression dynamics of CD36, alkaline phosphatase, y+LAT1, CAT1, lipase, amylase, and maltase during CHS exposure. The findings of this study suggest that, in broiler chickens, TM has a long-lasting impact on nutrient digestion and absorption capabilities as well as Tc, mortality rates, and BW during CHS.

Physiological responses to central and peripheral injection of polyinosinic-polycytidylic acid in chicks.

1. The purpose of the present study was to determine if intracerebroventricular (ICV) and intraperitoneal (IP) injection of polyinosinic-polycytidylic acid (poly I:C), a viral mimetic that binds to toll-like receptor-3 (TLR3), affects food intake, voluntary activity, cloacal temperature, plasma corticosterone (CORT) and glucose concentrations, and crop emptying rate in chicks (Gallus gallus). 2. Both ICV and IP injection of poly I:C significantly decreased food intake. 3. IP but not ICV injection of poly I:C significantly suppressed voluntary activity, whereas ICV injection decreased time spent sitting. Both ICV and IP injection of poly I:C significantly increased plasma CORT and glucose concentration. Neither ICV nor IP injection of poly I:C significantly affected cloacal temperature. 4. In addition, ICV injection of poly I:C significantly reduced crop emptying rate, whereas IP injection had no effect. 5. These results suggested that central TLR3 is related to anorexia, stress response and retardation of crop emptying while peripheral TLR3 is related to anorexia, change in behaviour and stress responses during viral infection in chicks.

Aerobic performance in tinamous is limited by their small heart. A novel hypothesis in the evolution of avian flight.

Some biomechanical studies from fossil specimens suggest that sustained flapping flight of birds could have appeared in their Mesozoic ancestors. We challenge this idea because a suitable musculoskeletal anatomy is not the only requirement for sustained flapping flight. We propose the "heart to fly" hypothesis that states that sustained flapping flight in modern birds required an enlargement of the heart for the aerobic performance of the flight muscles and test it experimentally by studying tinamous, the living birds with the smallest hearts. The small ventricular size of tinamous reduces cardiac output without limiting perfusion pressures, but when challenged to fly, the heart is unable to support aerobic metabolism (quick exhaustion, larger lactates and post-exercise oxygen consumption and compromised thermoregulation). At the same time, cardiac growth shows a crocodilian-like pattern and is correlated with differential gene expression in MAPK kinases. We integrate this physiological evidence in a new evolutionary scenario in which the ground-up, short and not sustained flapping flight displayed by tinamous represents an intermediate step in the evolution of the aerobic sustained flapping flight of modern birds.

Semen characteristics and biochemical composition of cloacal foam of male Japanese quails (Coturnix coturnix Japonica) fed diet incorporated with selenium.

An attempt was made to investigate the effect of dietary selenium (Se) on physical and cloacal gland size, foam production, biochemical composition of foam and semen biochemical characteristics of male Japanese quail (Coturnix coturnix Japonica). Two hundred twenty-five (225)-day-old male Japanese quail were randomly distributed to three dietary treatment groups for a period of 20 weeks. Each treatment comprised of three replicates, each containing 25 chicks. Three experimental diets were supplemented with 0, 0.5 and 1.0 mg Se/kg (T1 , T2 and T3, respectively), and diet T1 was considered as control. Sodium selenite was used as the source of selenium. All the birds were provided with feed and water ad libitum. Cloacal foam characteristics, that is cloacal gland index and foam weight, were significantly higher in T2 group. However, body weight, frequency of foam discharge and testes weight (left and right) did not differ significantly (p > 0.05). Physical characteristics of semen, that is semen volume and sperm concentration, did not differ (p > 0.05) among the Se-treated groups. The sperm motility, live-dead count and abnormality improved significantly (p < 0.05) in 0.5 mg/Se-supplemented group compared to 0 or 1.0 mg/Se-supplemented groups. Similarly, fertility and hatchability percentages were higher (p < 0.05) in 0.5 mg/Se-supplemented group than in control or 1.0 mg/Se-supplemented counterparts. The biochemical characteristics of foam in terms of total protein, acid phosphatase (ACP) and nitric oxide did not differ (p > 0.05), while the concentration of glucose was higher (p < 0.05) in 0.5 mg/Se-supplemented diet. On the other hand, alkaline phosphatase (ALP), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were lower (p < 0.05) in 0.5 mg/Se-supplemented group compared to control or 1.0 mg/Se-supplemented groups. From this study, it was concluded that supplementation of 0.5 mg Se/kg diet was beneficial for foam variables, biochemical composition of foam, semen characteristics and fertility in male Japanese quail.

Assessing the reliability of thermography to infer internal body temperatures of lizards.

For many years lizard thermal ecology studies have relied on the use of contact thermometry to obtain internal body temperature (Tb) of the animals. However, with progressing technology, an interest grew in using new, less invasive methods, such as InfraRed (IR) pyrometry and thermography, to infer Tb of reptiles. Nonetheless few studies have tested the reliability of these new tools. The present study tested the use of IR cameras as a non-invasive tool to infer Tb of lizards, using three differently body-sized lacertid species (Podarcis virescens, Lacerta schreiberi and Timon lepidus). Given the occurrence of regional heterothermy, we pairwise compared thermography readings of six body parts (snout, eye, head, dorsal, hind limb, tail base) to cloacal temperature (measured by a thermometer-associated thermocouple probe) commonly employed to measure Tb in field and lab studies. The results showed moderate to strong correlations (R2=0.84-0.99) between all body parts and cloacal temperature. However, despite the readings on the tail base showed the strongest correlation in all three species, it was the eye where the absolute values and pattern of temperature change most consistently followed the cloacal measurements. Hence, we concluded that the eye would be the body location whose IR camera readings more closely approximate that of the animal's internal environment. Alternatively, other body parts can be used, provided that a careful calibration is carried out. We provide guidelines for future research using thermography to infer Tb of lizards.

Relationships of eggshell, air cell, and cloacal temperatures of embryonated broiler hatching eggs during incubation.

The relationships of eggshell, air cell, and embryo cloacal temperatures in Ross × Ross 708 broiler hatching eggs were determined. Twenty eggs were weighed and set on each of 3 tray levels of a single incubator. Eggshell temperature (EST) of the eggs were recorded once in the morning (AM) and afternoon (PM) between 0 and 19 d of incubation (DOI) using an infrared thermometer (IRT). All eggs were candled and a transponder was implanted in the air cell of eggs containing live embryos (12 per tray level) at 12 DOI. At 19 DOI, transponders were implanted in the cloaca of live embryos from those same eggs. Air cell temperature (ACT) and EST readings were recorded once in the AM and PM between 12 and 19 DOI, and ACT and cloaca temperature (CLT) readings were recorded every 6 h between 19 and 21 DOI. The EST and ACT readings between 13 and 19 DOI were positively correlated. However, their respective mean temperatures between 13 and 19 DOI differed. The EST and ACT were not significantly influenced by tray level. Nevertheless, a main effect due to location (eggshell vs. air cell), and an interaction between DOI and time of day (AM and PM) in the 13 to 19 DOI interval were observed. Furthermore, an interaction was observed between location (air cell and cloaca) and the 6 h sequential time periods in the 19 to 21 DOI interval. However, across the entire 19 to 21 DOI interval, mean ACT and CLT were not significantly different, and were positively correlated. These data suggest that ACT readings are higher than those of EST during the last half of incubation, and that between 13 and 19 DOI, ACT readings may have the potential for use as a minimally invasive method by which to more accurately estimate the true core body temperature of broiler embryos. The effects of this method on hatchability and post-hatch performance need determination to better establish its practicality.

Lectin staining of the uterovaginal junction and sperm-storage tubule epithelia in broiler hens.

Mammalian sperm bind to terminal carbohydrates associated with glycoconjugates on the apical surface of oviduct epithelial cells in the caudal region of the oviduct and undergo cellular and molecular modifications associated with capacitation prior to ovulation. In contrast, chicken sperm are stored for up to 23 d in sperm-storage tubules (SST) localized in the uterovaginal junction (UVJ). Little is known of the cellular and molecular mechanisms that regulate sperm storage in and release from the SST. The purpose of this study was to identify glycoconjugates associated with the SST epithelial cell surface using lectins. Virgin hens and hens of higher and lower fertility in egg production for 6 to 16 wk were used in this study. Sections of UVJ mucosa containing SST were stained with fluorescent conjugated lectins and examined by confocal microscopy. Carbohydrate moieties associated with the UVJ and SST epithelia differed in their lectin binding patterns. No differences in the lectin binding patterns within the 2 epithelia were discernible between the virgin and younger and older hens. Minor differences were observed between the higher and lower fertility hens. Only lectins specific for galactose and N-acetylgalactosamine moieties were localized to the luminal surface of the SST. While resident sperm may be closely apposed to the SST epithelial cell apical microvilli, it is unlikely that sperm binding to the microvilli via terminal carbohydrates associated with glycoconjugates is a requisite for prolonged storage. However, the possibility of SST epithelial cell communication with resident sperm via shedding microvillous vesicles characterized by surface glycoconjugates with terminal galactose and N-acetylgalactosamine moieties is currently being investigated.

Novel Use of PIT Tags in Sea Cucumbers: Promising Results with the Commercial Species Cucumaria frondosa.

The lack of a reliable and innocuous mark-recapture method has limited studies that would provide essential information for the management of commercial sea cucumbers. Tagging sea cucumbers is notoriously difficult because of their plastic nature and autolysis capacities. The markers that have so far been tested, mainly on or through the body wall, were either lost rapidly or had major drawbacks (e.g. suitable only for batch identification, requiring complex analysis, causing infections, necrosis, behavioural changes and mortality). The present study explored the efficacy of passive integrated transponder (PIT) tags for individually marking sea cucumbers by assessing retention rates and long-term side effects of tags inserted in previously unstudied tissues/organs. Individuals of the species Cucumaria frondosa were tagged in the body wall, aquapharyngeal bulb and at the base of the oral tentacles. They were monitored closely for evidence of stress, infection, change in feeding and spawning behaviour and tag retention rate. Implanting the tag in an oral tentacle to reach the hydrovascular system of the aquapharyngeal bulb achieved the best retention rates in full-size individuals: from a maximum of 92% after 30 days to 68% at the end of the experimental period (300 days). Efficacy was lower in smaller individuals (84% after 30 d and 42% after 300 d). Following a slight increase in cloacal movements for 15 h post tagging, no side effect was noted in sea cucumbers tagged in the aquapharyngeal bulb via the tentacles. Feeding and spawning behaviours were not affected and no signs of infections or abnormal cell development in the vicinity of the tags were observed. This study indicates that marking sea cucumbers with 8.2 mm long PIT tags implanted via the oral tentacle is an effective technique, yielding relatively high retention rates over long periods without any detectable physiological or behavioural effects.

Ameliorative effects of melatonin administration and photoperiods on diurnal fluctuations in cloacal temperature of Marshall broiler chickens during the hot dry season.

Experiments were performed with the aim of determining the effect of melatonin administration on diurnal fluctuations in cloacal temperature (CT) of Marshall broiler chickens during the hot dry season. Birds in group I (12L:12D cycle) were raised under natural photoperiod of 12-h light and 12-h darkness, without melatonin supplementation, while those in group II (LL) were kept under 24-h continuous lighting, without melatonin administration. Broiler chickens in group III (LL + melatonin) were raised under 24-h continuous lighting, with melatonin supplementation at 0.5 mg/kg per os. The cloacal temperatures of 15 labeled broiler chickens from each group were measured at 6:00, 13:00, and 19:00 h, 7 days apart, from days 14-42. Temperature-humidity index was highest at day 14 of the study, with the value of 36.72 ± 0.82 °C but lowest at day 28 with the value of 30.91 ± 0.80 °C (P < 0.0001). The overall mean hourly cloacal temperature value of 41.51 ± 0.03 °C obtained in the 12L:12D cycle birds was significantly higher (P < 0.001) than the value of 41.16 ± 0.03 °C recorded in the melatonin-treated group but lower than that of 41.65 ± 0.03 °C obtained in the LL birds. Mortality due to hyperthermia commenced at day 28 in both 12L:12D cycle and LL broiler chickens but was delayed till day 42 in LL + MEL broiler chickens. In conclusion, melatonin administration alleviated the deleterious effects of heat stress on broiler chickens by maintaining their cloacal temperature at relatively low values.

Cloacal gland, endocrine testicular, and adrenocortical photoresponsiveness in male Japanese quail exposed to short days.

Photoperiod is the most important "noise-free" seasonal environmental cue for synchronizing physiological states (such as reproductive activity) in birds. However, in photoperiodic birds such as Japanese quail, the effect of photoperiod on adrenocortical activity remains unclear, particularly in males with differences in cloacal gland photoresponsiveness. At 8 wk of age, birds (n = 55) were either assigned to a short photoperiod (8L:16D; SD) or maintained under long photoperiod (16L:8D; LD; control). After 5 wk of SD exposure, males were classified as nonresponsive (SD-NR; with foam production) or responsive (SD-R; with no foam production) to short days, depending on the cloacal gland volume was above or below 1,000 mm(3). At 14 wk of age, droppings were collected during 3 consecutive days to determine corticosterone (CMs) and androgen metabolites (AMs) by enzyme immunoassays. Male Japanese quail under LD showed significantly higher concentrations of CMs (300 ± 10 ng/g) and AMs (1,257 ± 115 ng/g) than birds kept under SD. Under short days, SD-NR and SD-R showed differences (P < 0.0001), both in CM (153 ± 8 ng/g and 98 ± 6 ng/g, respectively) and AM concentrations (1,294 ± 309 ng/g and 275 ± 53 ng/g, respectively). Interestingly, although SD-NR males exhibited no cloacal gland arrest (according to cloacal gland volume and foam production) and similar testicular activity (AM values) to LD males, they showed lower concentrations of CMs compared with males that remained on LD (P < 0.05). Our findings suggest length of photoperiod affected hypothalamic-pituitary-adrenal activity; however, that was not the only factor involved, because birds subjected to shorter days but whose hypothalamic-pituitary-gonadal axis failed to respond had intermediate CM values. Further research is required to elucidate the underlying mechanisms of this interesting finding.

Distinct neuroendocrine mechanisms control neural activity underlying sex differences in sexual motivation and performance.

Sexual behavior can be usefully parsed into an appetitive and a consummatory component. Both appetitive and consummatory male-typical sexual behaviors (respectively, ASB and CSB) are activated in male Japanese quail by testosterone (T) acting in the medial preoptic nucleus (POM), but never observed in females. This sex difference is based on a demasculinization (= organizational effect) by estradiol during embryonic life for CSB, but a differential activation by T in adulthood for ASB. Males expressing rhythmic cloacal sphincter movements (RCSMs; a form of ASB) or allowed to copulate display increased Fos expression in POM. We investigated Fos brain responses in females exposed to behavioral tests after various endocrine treatments. T-treated females displayed RCSM, but never copulated when exposed to another female. Accordingly they showed an increased Fos expression in POM after ASB but not CSB tests. Females treated with the aromatase inhibitor Vorozole in ovo and T in adulthood displayed both male-typical ASB and CSB, and Fos expression in POM was increased after both types of tests. Thus, the neural circuit mediating ASB is present or can develop in both sexes, but is inactive in females unless they are exposed to exogenous T. In contrast, the neural mechanism mediating CSB is not normally present in females, but can be preserved by blocking the embryonic production of estrogens. Overall these data confirm the difference in endocrine controls and probably neural mechanisms supporting ASB and CSB in quail, and highlight the complexity of mechanisms underlying sexual differentiation of behavior.

Aquatic eggs are fertilised by multiple males not engaged in amplexus in a stream-breeding frog.

In anuran amphibians, multiple males amplexing a single female to fertilise her eggs has been found for less than 25 species, whereas matings without amplexus are known for less than five species. Here we provide a new example of simultaneous polyandry with multiple males not engaged in amplexus, in Feirana taihangnicus, a stream-dwelling, explosive breeder endemic to central China. Laboratory experiments showed that when one female was kept with one male in a vessel with elevated, flat stones, the female stood on her head with her swollen cloaca against the undersurface of the stone substrate to lay eggs (clutch size ranged from 371 to 533, n=7 females). Then, 10-102 min after oviposition began, the male stood on his head and released sperm over the eggs distributed as a single layer on the stone surfaces. It took about 3h for the female to finish oviposition and for the male to finish fertilisation. On average, 96% of eggs were fertilised. In natural oviposition habitats, stream sections with slow flowing, we observed that 1 up to 15 males (8.7±6.6, n=6 cases), none in amplexus, participated in fertilising the eggs deposited by a single female. Evolutionary implications of this unusual reproductive strategy remain to be explored.