During the last part of egg incubation in king penguins, the male can preserve undigested food in the stomach for several weeks. This ensures survival of the newly hatched chick, in cases where the return of the foraging female from the sea is delayed. In accordance with the characterization of stress-induced bacteria, we demonstrate the occurrence of strong antimicrobial activities in preserved stomach contents. We isolated and fully characterized two isoforms of a novel 38-residue antimicrobial peptide (AMP), spheniscin, belonging to the ␤-defensin subfamily. Spheniscin concentration was found to strongly increase during the period of food storage. Using a synthetic version of one of two spheniscin isoforms, we established that this peptide has a broad activity spectrum, affecting the growth of both pathogenic bacteria and fungi. Altogether, our data suggest that spheniscins and other, not yet identified, antimicrobial substances may play a role in the long term preservation of stored food in the stomach of king penguins.Lack of food resources induced by climatic changes may impair the breeding success of wild animals. For king penguins, the female usually comes back from foraging at sea to feed the chick at hatching. However, in relation to the circumpolar wave, thought to be linked with the El Niñ o Southern Oscillation phenomenon (1), the return of the female may be delayed due to the necessity of foraging at a greater distance than usual. The survival of the chick can still be ensured by the male, as it can provide food preserved in its stomach during the final 3 weeks of incubation (2, 3). A remaining unanswered question is how can food be preserved from attack by microorganisms, when the rich 38°C buffered growth medium (3, 4) and a long retention time in the stomach would be expected to favor microbial colonization (5, 6). The maintained mass and energetic value of the food throughout the fast (3) and the stressed-induced characteristics of the stomach bacteria (4) altogether suggest the existence of a protection mechanism against food degradation by microorganisms.Like other mucosal surfaces, the gastrointestinal tract surface interacts directly with the external environment and therefore has to be protected from damage and invasion by ingested or indigenous microorganisms. One established facet of epithelial host defense is the synthesis and secretion of AMPs 1 (7,8). The role of these AMPs is not limited to epithelial protection through innate immune responses since they can attract human blood cells, alert the adaptive immune system and induce gastrointestinal secretions (8 -10). It has also been assumed that some AMPs influence the resident microflora in the small intestine lumen (11,12). An additional role could then be the participation of AMPs in the control of microbial proliferation, contributing to the preservation of retained food.In the present study, we found numerous, strongly active antimicrobial substances in the stomach contents of male penguins that efficiently conserve food during th...

Recently two ␤-defensins, named spheniscins, have been isolated from the stomach content of the king penguin (Aptenodytes patagonicus), which is capable of preserving food for several weeks during egg incubation (Thouzeau, C., Le Maho, Y., Froget, G., Sabatier, L., Le Bohec, C., Hoffmann, J. A., and Bulet, P. (2003) J. Biol. Chem. 278, 51053-51058). It has been proposed that, in combination with other antimicrobial peptides, spheniscins may be involved in this long term preservation of food in the bird's stomach. To draw some structure/ function features, the three-dimensional structure in aqueous solution of the most abundant spheniscin (Sphe-2) was determined by two-dimensional NMR and molecular modeling techniques. The overall fold of Sphe-2 includes a three-stranded antiparallel ␤-sheet stabilized by three disulfide bridges with a pairing typical of ␤-defensins. In addition, the N-terminal segment shows helical features on most structures. Sphe-2 is highly cationic, and its surface displays a hydrophobic patch. Comparative modeling revealed that this patch is preserved in avian defensins. The activity of Sphe-2 against a pathogenic Gram-positive strain was retained in vitro in the conditions of osmolarity found in penguin stomach content and also in different salt concentrations and compositions up to those reported for seawater. Comparison with structurally related mammalian ␤-defensins showed that the hydrophobic patch is not preserved in mammalian ␤-defensins and that the high cationicity of Sphe-2 is presumably the critical factor for its retained activity in high salt concentrations. Such peculiarities, in addition to a broad activity spectrum, suggest that penguin defensins may represent interesting probes for the design of highly efficient antibiotics to fight off pathogens that develop in relatively salt-rich body fluids.During the final stage of egg incubation in king penguins (Aptenodytes patagonicus), the male can preserve undigested food in the stomach for several weeks (1). This ensures survival of the newly hatched chick in the event that the return of the foraging female from the sea is delayed. In accordance with the characterization of stress-induced bacteria (2), a previous study has demonstrated that numerous antimicrobial activities exist in preserved stomach contents (3). Two antimicrobial peptides have been isolated and fully characterized, namely spheniscin-1 and -2 (Sphe-1/pBD-1 and Sphe-2/pBD-2).1 The two forms of spheniscins differ by a single residue, His 14 , in Sphe-1 versus Arg 14 in Sphe-2. A data bank search revealed that spheniscins are members of the well known defensin family.Defensins are small (3-5 kDa) cationic antimicrobial peptides that are part of the innate immunity of vertebrates (4, 5), invertebrates (6), and plants (7). In vertebrates, defensins can be divided into ␣-, ␤-, and -defensin subfamilies. The unusual -defensin, which is a circular peptide, was initially isolated from rhesus macaque (8). Recently a pseudogene coding for a homologue, named retrocyclin, wa...

Male king penguins are able to store undigested food in their stomach for up to 3 weeks during their incubation fast, which evidently implies some modification of their digestive process. Using small electronic recorders, we studied the change in gastric pH, motility and temperature during the first week of food storage. The pH could be maintained at values as high as 6 throughout the incubation fast, a pH that is unfavourable for avian gastric proteinase activity. Gastric motility was never completely inhibited but could be markedly reduced. Stomach temperature was maintained at around 38°C.The fact that stomach temperature of incubating birds did not show a daily rhythmic fluctuation as seen in nonbreeding birds could be due to temperature constraints on embryo development. Thus the present study demonstrates substantial adjustments of pH and gastric motility in incubating king penguins, which may contribute to the inhibition of digestive gastric processes.

Energetic adaptation to fasting in the cold has been investigated in a nocturnal raptor, the barn owl (Tyto alba), during winter. Metabolic rate and body temperature (Tb) were monitored in captive birds, (1) after acute exposure to different ambient temperatures (Ta), and (2) during a prolonged fast in the cold (4 degrees C), to take into account the three characteristic phases of body fuel utilization that occur during a long-term but reversible fast. In postabsorptive birds, metabolic rate in the thermoneutral zone was 4. 1+/-0.1 W kg-1 and increased linearly below a lower critical temperature of 23 degrees C. Metabolic rate was 70% above basal at +4 degrees C Ta. Wet thermal conductance was 0.22 W kg-1 degrees C-1. During fasting in the cold, the mass-specific resting metabolic rate decreased by 16% during the first day (phase I) and remained constant thereafter. The amplitude of the daily rhythm in Tb was only moderately increased during phase II, with a slight lowering (0. 6 degrees C) in minimal diurnal Tb, but rose markedly in phase III with a larger drop (1.4 degrees C) in minimal diurnal Tb. Refeeding the birds ended phase III and reversed the observed changes. These results indicate that diurnal hypothermia may be used in long-term fasting barn owls and could be triggered by a threshold of body lipid depletion, according to the shift from lipid to protein fuel metabolism occurring at the phase II/phase III transition. The high cost of regulatory thermogenesis and the limited use of hypothermia during fasting may contribute to the high mortality of barn owls during winter.

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