Gut bacteria that produce urease, the enzyme hydrolysing urea, contribute to nitrogen balance in diverse vertebrates, although the presence of this system of urea-nitrogen recycling in Amphibia is as yet unknown. Our studies of the wood frog (), a terrestrial species that accrues urea in winter, documented robust urease activity by enteric symbionts and hence potential to recoup nitrogen from the urea it produces. Ureolytic capacity in hibernating (non-feeding) frogs, whose guts hosted an approximately 33% smaller bacterial population, exceeded that of active (feeding) frogs, possibly due to an inductive effect of high urea on urease expression and/or remodelling of the microbial community. Furthermore, experimentally augmenting the host's plasma urea increased bacterial urease activity. Bacterial inventories constructed using 16S rRNA sequencing revealed that the assemblages hosted by hibernating and active frogs were equally diverse but markedly differed in community membership and structure. Hibernating frogs hosted a greater relative abundance and richer diversity of genera that possess urease-encoding genes and/or have member taxa that reportedly hydrolyse urea. Bacterial hydrolysis of host-synthesized urea probably permits conservation and repurposing of valuable nitrogen not only in hibernating but, given urea's universal role in amphibian osmoregulation, also in virtually all Amphibia.
A low-cost, graduate student–led, metacognition-based study skills course taught alongside introductory biology is described. The course improved scores on lecture exams, study habits, and specific study techniques (outlines/concept maps). Implications of the results for implementation of first-year interventions in STEM are discussed.
Nitric oxide (NO) is a gaseous free radical that in diverse organisms performs many signaling and protective functions, such as vasoregulation, inhibition of apoptosis, antioxidation, and metabolic suppression. Increased availability of NO may be especially important during life-history periods when organisms contend with multiple stresses. We investigated dynamics of the NO metabolites, nitrite (NO) and nitrate (NO), in the blood plasma, heart, liver, and skeletal muscle of the wood frog (Rana sylvatica), an amphibian that endures chronic cold, freezing, hypoxia, dehydration, and extended aphagia during hibernation. We found elevated concentrations of NO and/or NO in the plasma (up to 4.1-fold), heart (3.1-fold), and liver (up to 4.1-fold) of frogs subjected to experimental hypoxia (24 h, 4 °C), and in the liver (up to 3.8-fold) of experimentally frozen frogs (48 h, - 2.5 °C), suggesting that increased NO availability aids in survival of these stresses. During a 38-week period of simulated hibernation, NO and/or NO increased in the plasma (up to 10.4-fold), heart (up to 3.3-fold), and liver (5.0-fold) during an initial 5-week winter-acclimatization regimen and generally remained elevated thereafter. In hibernation, plasma NO was higher in frogs indigenous to Interior Alaska than in conspecifics from a temperate locale (southern Ohio), suggesting that NO availability is matched to the severity of environmental conditions prevailing in winter. The comparatively high NO availability in R. sylvatica, a stress-tolerant species, together with published values for other species, suggest that the NO protection system is of general importance in the stress adaptation of vertebrates.
Although seasonal increases in cold hardiness are well documented for temperate and polar ectotherms, relatively little is known about supplemental increases in cold hardiness during winter. Because many animals are exposed to considerable thermal variation in winter, they may benefit from a quick enhancement of cold tolerance prior to extreme low temperature. Hatchling painted turtles (Chrysemys picta) overwintering in their natal nests experience substantial thermal variation in winter, and recently, it was found that brief subzero chilling of winter-acclimated hatchlings decreases subsequent chilling-induced mortality, increases blood concentrations of glucose and lactate, and protects the brain from cryoinjury. Here, we further characterize that phenomenon, termed 'cold conditioning', by exposing winter-acclimated hatchling turtles to -3.5, -7.0, or -10.5 °C gradually or repeatedly via daily thermal fluctuations over the course of 5 days and assessing their survival of a subsequent cold shock to a discriminating temperature of -12.7 °C. To better understand the physiological response to cold conditioning, we measured changes in glucose and lactate concentrations in the liver, blood, and brain. Cold conditioning significantly increased cold-shock survival, from 9% in reference turtles up to 74% in cold-conditioned turtles, and ecologically relevant daily thermal fluctuations were at least as effective at conferring cryoprotection as was gradual cold conditioning. Cold conditioning increased glucose concentrations, up to 25 μmol g, and lactate concentrations, up to 30 μmol g, in the liver, blood, and brain. Turtles that were cold conditioned with daily thermal fluctuations accumulated more glucose in the liver, blood, and brain, and had lower brain lactate, than those gradually cold conditioned. Given the thermal variation to which hatchling painted turtles are exposed in winter, we suggest that the supplemental protection conferred by cold conditioning, especially that induced by daily thermal fluctuations, may be important for their overwinter survival. Investigation into the duration of the cold-conditioning induced protection and its occurrence in natural field conditions is needed to better understand its ecological significance. We suggest that future work exploring the underlying mechanisms of cold conditioning should focus on non-colligative effects of glucose, expression of small Hsps, changes in membrane structure, and ion homeostasis.
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