The Evolution of Lactation in Mammalian Species.

Lactation is a defining characteristic of all mammals, and, indeed, mammals draw their name from mammae, or mammary glands. The evolution of mammary glands has been the subject of debate since Charles Darwin. The purpose of this brief review is not to examine all past theories of mammary evolution but to consider the evolution of the mammary gland in relation to (1) modern paleobiology, giving special attention to the mammaliaforms which had many mammalian features, including delayed tooth development suggestive of milk intake. (2) Comparative aspects of mammary development in monotremes, marsupials, and eutherians, which reveal the close developmental relation of mammary glands to other skin glands and hair follicles. (3) The evolution of caseins, which are now known to derive from secretory calcium-binding phosphoproteins, which have a long history in regulating biomineralization. (4) The evolution of lipid secretion, and especially the evolutionary incorporation of immune system components (such as xanthine oxidoreductase) into the fat globule membrane. (5) The evolution of lactose synthesis, and especially the synthesis of the wide array of oligosaccharides found in some milks, including monotremes, marsupials, caniform carnivores, and humans.

Chemical structures of oligosaccharides in milks of the American black bear (Ursus americanus americanus) and cheetah (Acinonyx jubatus).

The milk oligosaccharides were studied for two species of the Carnivora: the American black bear (Ursus americanus, family Ursidae, Caniformia), and the cheetah, (Acinonyx jubatus, family Felidae, Feliformia). Lactose was the most dominant saccharide in cheetah milk, while this was a minor saccharide and milk oligosaccharides predominated over lactose in American black bear milk. The structures of 8 neutral saccharides from American black bear milk were found to be Gal(ß1-4)Glc (lactose), Fuc(α1-2)Gal(ß1-4)Glc (2'-fucosyllactose), Gal(α1-3)Gal(ß1-4)Glc (isoglobotriose), Gal(α1-3)[Fuc(α1-2)]Gal(ß1-4)Glc (B-tetrasaccharide), Gal(α1-3)[Fuc(α1-2)]Gal(ß1-4)[Fuc(α1-3)]Glc (B-pentasaccharide), Fuc(α1-2)Gal(ß1-4)[Fuc(α1-3)]GlcNAc(ß1-3)Gal(ß1-4)Glc (difucosyl lacto-N-neotetraose), Gal(α1-3)Gal(ß1-4)[Fuc(α1-3)]GlcNAc(ß1-3)Gal(ß1-4)Glc (monogalactosyl monofucosyl lacto-N-neotetraose) and Gal(α1-3)Gal(ß1-4)GlcNAc(ß1-3)Gal(ß1-4)Glc (Galili pentasaccharide). Structures of 5 acidic saccharides were also identified in black bear milk: Neu5Ac(α2-3)Gal(ß1-4)Glc (3'-sialyllactose), Neu5Ac(α2-6)Gal(ß1-4)GlcNAc(ß1-3)[Fuc(α1-2)Gal(ß1-4)GlcNAc(ß1-6)]Gal(ß1-4)Glc (monosialyl monofucosyl lacto-N-neohexaose), Neu5Ac(α2-6)Gal(ß1-4)GlcNAc(ß1-3)[Gal(α1-3)Gal(ß1-4)GlcNAc(ß1-6)]Gal(ß1-4)Glc (monosialyl monogalactosyl lacto-N-neohexaose), Neu5Ac(α2-6)Gal(ß1-4)GlcNAc(ß1-3){Gal(α1-3)Gal(ß1-4)[Fuc(α1-3)]GlcNAc(ß1-6)}Gal(ß1-4)Glc (monosialyl monogalactosyl monofucosyl lacto-N-neohexaose), and Neu5Ac(α2-6)Gal(ß1-4)GlcNAc(ß1-3){Gal(α1-3)[Fuc(α1-2)]Gal(ß1-4)[Fuc(α1-3)]GlcNAc(ß1-6)}Gal(ß1-4)Glc (monosialyl monogalactosyl difucosyl lacto-N-neohexaose). A notable feature of some of these milk oligosaccharides is the presence of B-antigen (Gal(α1-3)[Fuc(α1-2)]Gal), α-Gal epitope (Gal(α1-3)Gal(ß1-4)Glc(NAc)) and Lewis x (Gal(ß1-4)[Fuc(α1-3)]GlcNAc) structures within oligosaccharides. By comparison to American black bear milk, cheetah milk had a much smaller array of oligosaccharides. Two cheetah milks contained Gal(α1-3)Gal(ß1-4)Glc (isoglobotriose), while another cheetah milk did not, but contained Gal(ß1-6)Gal(ß1-4)Glc (6'-galactosyllactose) and Gal(ß1-3)Gal(ß1-4)Glc (3'-galactosyllactose). Two cheetah milks contained Gal(ß1-4)GlcNAc(ß1-3)[Gal(ß1-4)GlcNAc(ß1-6)]Gal(ß1-4)Glc (lacto-N-neohexaose), and one cheetah milk contained Gal(ß1-4)Glc-3'-O-sulfate. Neu5Ac(α2-8)Neu5Ac(α2-3)Gal(ß1-4)Glc (disialyllactose) was the only sialyl oligosaccharide identified in cheetah milk. The heterogeneity of milk oligosaccharides was found between both species with respect of the presence/absence of B-antigen and Lewis x. The variety of milk oligosaccharides was much greater in the American black bear than in the cheetah. The ratio of milk oligosaccharides-to-lactose was lower in cheetah (1:1-1:2) than American black bear (21:1) which is likely a reflection of the requirement for a dietary supply of N-acetyl neuraminic acid (sialic acid), in altricial ursids compared to more precocial felids, given the role of these oligosaccharides in the synthesis of brain gangliosides and the polysialic chains on neural cell adhesion.

4-O-Acetyl-sialic acid (Neu4,5Ac2) in acidic milk oligosaccharides of the platypus (Ornithorhynchus anatinus) and its evolutionary significance.

Monotremes (echidnas and platypus) retain an ancestral form of reproduction: egg-laying followed by secretion of milk onto skin and hair in a mammary patch, in the absence of nipples. Offspring are highly immature at hatching and depend on oligosaccharide-rich milk for many months. The primary saccharide in long-beaked echidna milk is an acidic trisaccharide Neu4,5Ac2(α2-3)Gal(ß1-4)Glc (4-O-acetyl 3'-sialyllactose), but acidic oligosaccharides have not been characterized in platypus milk. In this study, acidic oligosaccharides purified from the carbohydrate fraction of platypus milk were characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and (1)H-nuclear magnetic resonance spectroscopy. All identified structures, except Neu5Ac(α2-3)Gal(ß1-4)Glc (3'-sialyllactose) contained Neu4,5Ac2 (4-O-acetyl-sialic acid). These include the trisaccharide 4-O-acetyl 3'-sialyllactose, the pentasaccharide Neu4,5Ac2(α2-3)Gal(ß1-4)GlcNAc(ß1-3)Gal(ß1-4)Glc (4-O-acetyl-3'-sialyllacto-N-tetraose d) and the hexasaccharide Neu4,5Ac2(α2-3)Gal(ß1-4)[Fuc(α1-3)]GlcNAc(ß1-3)Gal(ß1-4)Glc (4-O-acetyl-3'-sialyllacto-N-fucopentaose III). At least seven different octa- to deca-oligosaccharides each contained a lacto-N-neohexaose core (LNnH) and one or two Neu4,5Ac2 and one to three fucose residues. We conclude that platypus milk contains a diverse (≥ 20) array of neutral and acidic oligosaccharides based primarily on lactose, lacto-N-neotetraose (LNnT) and LNnH structural cores and shares with echidna milk the unique feature that all identified acidic oligosaccharides (other than 3'-sialyllactose) contain the 4-O-acetyl-sialic acid moiety. We propose that 4-O-acetylation of sialic acid moieties protects acidic milk oligosaccharides secreted onto integumental surfaces from bacterial hydrolysis via steric interference with bacterial sialidases. This may be of evolutionary significance since taxa ancestral to monotremes and other mammals are thought to have secreted milk, or a milk-like fluid containing oligosaccharides, onto skin surfaces.

Proximate composition of milk of the bongo (Tragelaphus eurycerus) in comparison to other African bovids and to hand-rearing formulas.

African bovids represent a highly diverse group with divergent neonatal care strategies. The extent to which their milks reflect this diversity is poorly understood. We analyzed milk of the bongo (Tragelaphus eurycerus) to compare its composition to milks of other African bovids and to evaluate bongo milk replacement formulas. Milk samples from three individuals (0 through 300 days postpartum, n = 28) were assayed for dry matter (total solids), crude fat, crude protein, total sugar, ash, calcium, and phosphorus; gross energy was assayed on a subset of samples and compared to calculated values. Nutrient composition changed very little over the lactation period except for day 0 (colostrum) and the last sample (day 300). Bongo milk (days 6-286) contained (mean ± SEM): 28.1 ± 0.7% dry matter (71.9 ± 0.7% water), 12.3 ± 0.6% fat, 10.6 ± 0.3% crude protein, 3.6 ± 0.1% sugar, 1.05 ± 0.03% ash, 0.26 ± 0.01% calcium, 0.16 ± 0.01% phosphorus, and a GE of 1.88 ± 0.06 kcal/g. The protein content of bongo milk accounts for 33% of energy. High protein energy appears to be typical of Tragelaphines and of African bovids that utilize a "hider" system of postnatal care. The stability of milk composition until day 300 suggests complete weaning may not occur until 9 months rather than at 6 months of age, as commonly assumed. None of the milk replacement formulas previously used for bongos was well matched to bongo milk composition; therefore, a new milk replacement formula is proposed.

Can an ancestral condition for milk oligosaccharides be determined? Evidence from the Tasmanian echidna (Tachyglossus aculeatus setosus).

The monotreme pattern of egg-incubation followed by extended lactation represents the ancestral mammalian reproductive condition, suggesting that monotreme milk may include saccharides of an ancestral type. Saccharides were characterized from milk of the Tasmanian echidna Tachyglossus aculeatus setosus. Oligosaccharides in pooled milk from late lactation were purified by gel filtration and high-performance liquid chromatography using a porous graphitized carbon column and characterized by (1)H NMR spectroscopy; oligosaccharides in smaller samples from early and mid-lactation were separated by ultra-performance liquid chromatography and characterized by negative electrospray ionization mass spectrometry (ESI-MS) and tandem collision mass spectroscopy (MS/MS) product ion patterns. Eight saccharides were identified by (1)H NMR: lactose, 2'-fucosyllactose, difucosyllactose (DFL), B-tetrasaccharide, B-pentasaccharide, lacto-N-fucopentaose III (LNFP3), 4-O-acetyl-3'-sialyllactose [Neu4,5Ac(α2-3)Gal(ß1-4)Glc] and 4-O-acetyl-3'-sialyl-3-fucosyllactose [Neu4,5Ac(α2-3)Gal(ß1-4)[Fuc(α1-3)]Glc]. Six of these (all except DFL and LNFP3) were present in early and mid-lactation per ESI-MS, although some at trace levels. Four additional oligosaccharides examined by ESI-MS and MS/MS are proposed to be 3'-sialyllactose [Neu5Ac(α2-3)Gal(ß1-4)Glc], di-O-acetyl-3'-sialyllactose [Neu4,5,UAc3(α2-3)Gal(ß1-4)Glc where U = 7, 8 or 9], 4-O-acetyl-3'-sialyllactose sulfate [Neu4,5Ac(α2-3)Gal(ß1-4)GlcS, where position of the sulfate (S) is unknown] and an unidentified 800 Da oligosaccharide containing a 4-O-acetyl-3'-sialyllactose core. 4-O-acetyl-3'-sialyllactose was the predominant saccharide at all lactation stages. 4-O-Acetylation is known to protect sialyllactose from bacterial sialidases and may be critical to prevent microbial degradation on the mammary areolae and/or in the hatchling digestive tract so that sialyllactose can be available for enterocyte uptake. The ability to defend against microbial invasion was probably of great functional importance in the early evolution of milk saccharides.

Evo-devo of the mammary gland.

We propose a new scenario for mammary evolution based on comparative review of early mammary development among mammals. Mammary development proceeds through homologous phases across taxa, but evolutionary modifications in early development produce different final morphologies. In monotremes, the mammary placode spreads out to form a plate-like mammary bulb from which more than 100 primary sprouts descend into mesenchyme. At their distal ends, secondary sprouts develop, including pilosebaceous anlagen, resulting in a mature structure in which mammary lobules and sebaceous glands empty into the infundibula of hair follicles; these structural triads (mammolobular-pilo-sebaceous units or MPSUs) represent an ancestral condition. In marsupials a flask-like mammary bulb elongates as a sprout, but then hollows out; its secondary sprouts include hair and sebaceous anlagen (MPSUs), but the hairs are shed during nipple formation. In some eutherians (cat, horse, human) MPSUs form at the distal ends of primary sprouts; pilosebaceous components either regress or develop into mature structures. We propose that a preexisting structural triad (the apocrine-pilo-sebaceous unit) was incorporated into the evolving mammary structure, and coupled to additional developmental processes that form the mammary line, placode, bulb and primary sprout. In this scenario only mammary ductal trees and secretory tissue derive from ancestral apocrine-like glands. The mammary gland appears to have coopted signaling pathways and genes for secretory products from even earlier integumentary structures, such as odontode (tooth-like) or odontode-derived structures. We speculate that modifications in signal use (such as PTHrP and BMP4) may contribute to taxonomic differences in MPSU development.

Neutral and acidic milk oligosaccharides of the striped skunk (Mephitidae: Mephitis mephitis).

The biological significance of the tremendous variation in proportions of oligosaccharides and lactose among mammalian milks is poorly understood. We investigated milk oligosaccharides of the striped skunk (Mephitis mephitis) and compared these results to other species of the clade Mustelida. Individual oligosaccharides were identified by proton nuclear magnetic resonance spectroscopy. In the striped skunk, six oligosaccharides were identified: isoglobotriose, 2'-fucosyllactose, A-tetrasaccharide, Galili pentasaccharide, 3'-sialyllactose and monosialyl monogalactosyl lacto-N-neohexaose. Four of these have been found in related Mustelida and the other two in more distantly related carnivorans. The neutral and acidic oligosaccharides derive from three core structures: lactose (Gal(ß1-4)Glc), lacto-N-neotetraose (Gal(ß1-4)GlcNAc(ß1-3)Gal(ß1-4)Glc) and lacto-N-neohexaose (Gal(ß1-4)GlcNAc(ß1-3)[Gal(ß1-4)GlcNAc(ß1-6)]Gal(ß1-4)Glc).

Daughter dearest: Sex-biased calcium in mother's milk among rhesus macaques.

Mother's milk provides building blocks necessary for infant development and growth postnatally. Minerals in milk are particularly important for infant skeletal development and may reflect maternal characteristics that are associated with the capacity to synthesize milk and sex-specific developmental priorities of the infant. Using a large sample of mother-infant dyads assigned to the outdoor breeding colony at the California National Primate Research Center (N=104), we investigated the relationship of milk calcium (Ca) and phosphorus (P) concentrations and the ratio of Ca/P to maternal and infant characteristics and to other milk variables. Ca and P are largely associated with casein micelles, and as expected, both Ca and P were positively correlated with protein concentrations in milk. Neither Ca nor P concentrations were associated with maternal parity. Mothers rearing daughters tended to produce higher mean Ca concentration in milk, and consequently a higher Ca/P ratio, than did mothers rearing sons, even though protein concentration was not elevated. These results suggest that the Ca/P ratio in rhesus milk may have been under separate selective pressure from protein content to facilitate the accelerated rate of skeletal calcification that has been observed in female Macaca mulatta infants.

Milk composition in the Weddell seal Leptonychotes weddellii: evidence for a functional role of milk carbohydrates in pinnipeds.

We propose that secretion of milk sugar has important consequences for the metabolic economies of lactating phocid seals and their pups. Milk was collected from 21 Weddell seals Leptonychotes weddellii in McMurdo Sound, Antarctica, and assayed by standard methods. Milk composition changed over the course of lactation, but at mid- to late lactation (16-40 d postpartum), Weddell seal milk composition was relatively constant at 33.8% +/- 0.82% water, 54.0% +/- 0.80% fat, 10.1% +/- 0.16% crude protein, 0.84% +/- 0.03% sugar, 0.75% +/- 0.02% ash, and 23.3 +/- 0.3 kJ g−1 whole milk (WM). At this stage, milk composition varied among individual seals in all assayed constituents except ash. The concentration of sugar in the aqueous phase of Weddell seal milk ( 24.9 +/- 0.6g sugar L−1 water) was ca. 44%-77% of levels found in terrestrial carnivores, indicating that the low sugar concentration of WM is primarily due to its high fat content, not alteration of the aqueous phase. In early lactation, fasting Weddell seals were estimated to devote 39 g d(-1) glucose to milk sugar synthesis, an amount similar to the estimated demand of the maternal brain. This additional glucose demand must be covered by gluconeogenesis in fasting animals and represents a considerable additional drain on maternal resources. However, provision of sugar to offspring at rates sufficient to meet neonatal substrate requirements appears to be essential for efficient fat and protein deposition and thus may be an important component of the phocid reproductive strategy of rapid growth and early weaning.

Variation in the composition of milk of Asian elephants (Elephas maximus) throughout lactation.

We investigated milk nutrient composition from three Asian elephant cows over the first 3 years of lactation, including two consecutive lactations in one cow. Body mass gain is presented for three calves during the first year. Milk samples (n = 74) were analyzed for dry matter (DM), fat, crude protein (CP), sugar, ash, calcium (Ca), phosphorus (P), and potassium (K); gross energy (GE) was calculated. Concentrations of most nutrients changed over lactation: DM, fat, CP, Ca, P, and GE were positively correlated to calf age; sugar was negatively correlated to calf age. GE doubled between birth (1 kcal/g) and 2 years of age (2 kcal/g). After accounting for calf age, GE, fat, Ca, and P concentrations differed among the cows. Milk composition also differed between two lactations from the same cow. When milk nutrients were expressed on a mg per kcal basis, the pattern changes: CP, Ca, and P remained relatively constant over lactation on a per energy basis. Calf mass quadrupled over the first year of life; mass gain was linear at 0.9 kg/day. Asian elephant milk composition is variable, both across lactations and between cows, complicating efforts to determine representative values for comparative studies and for the formulation of elephant milk formulas. The fact that CP, Ca, and P were all relatively constant when expressed on a per energy basis may be of biological significance. The increase in nutrient density over lactation undoubtedly limits maternal water loss, reducing the volume of milk necessary to support the calf.

The predominance of type I oligosaccharides is a feature specific to human breast milk.

Human milk and colostrum contain ∼12-13 g/L and ∼22-24 g/L of oligosaccharides, respectively. The chemical structures of >100 human milk oligosaccharides (HMO) have been characterized to date. We determined the concentrations of 10 neutral and 9 acidic colostrum HMO collected during the first 3 d of lactation by using reverse phase HPLC after derivatization with 2-aminopyridine or 1-methyl-3-phenyl-5-pyrazolon. The predominant oligosaccharides were Fuc(α1-2)Gal(ß1-4Glc (2'-FL), Fuc(α1-2)Gal(ß1-3)GlcNAc(ß1-3)Gal(ß1-4)Glc (LNFP I), Fuc(α1-2)Gal(ß1-3)[Fuc(α1-4)]GlcNAc(ß1-3)Gal(ß1-4)Glc (LNDFH I), and Gal(ß1-3)GlcNAc(ß1-3)Gal(ß1-4)Glc (LNT), the concentration of each of which was ∼1-3 g/L. Because these HMO, other than 2'-FL, all contain the Lacto-N-biose type I structure [Gal(ß1-3)GlcNAc], we conclude that HMO containing the type I structure predominate over those containing the N-acetyllactosamine type II structure [Gal(ß1-4)GlcNAc]. This appears to be a feature that is specific to humans, because the milk and colostrum of other species, including apes and monkeys, either contain only type II oligosaccharides or type II predominate over type I. It is possible that type I HMO may have importance as substrates for beneficial bifidobacteria in breast-fed infants. The biological importance of type I HMO predominance warrants further study, both in relation to human health and to human evolution.

Chemical characterization of milk oligosaccharides of the island flying fox (Pteropus hypomelanus) (Chiroptera: Pteropodidae).

Although a considerable amount of information has accumulated about oligosaccharides in the milk and colostrum of representatives of various mammalian orders, nothing is so far known concerning these sugars in the milk of any bat species (order Chiroptera). In this study, we determined that the following oligosaccharides occur in milk of the island flying fox, Pteropus hypomelanus (Chiroptera: Pteropidae): Gal(α1-3)Gal(ß1-4)Glc (isoglobotriose), Gal(ß1-4)GlcNAc(ß1-3)Gal(ß1-4)Glc (lacto-N-neotetraose), Gal(ß1-4)GlcNAc(ß1-3)[Gal(ß1-4)GlcNAc(ß1-6)]Gal(ß1-4)Glc (lacto-N-neohexaose) and Neu5Gc(α2-3)Gal(ß1-4)Glc (3'-NGc-SL). However, lactose was found to be the dominant saccharide in this milk, as in most eutherian mammals. The biologic importance of oligosaccharides in Chiropteran milks warrants further study.

Is tissue maturation necessary for flight? Changes in body composition during postnatal development in the big brown bat.

Patterns of offspring development reflect the availability of energy and nutrients, limitations on an individual's capacity to use available resources, and tradeoffs between the use of nutrients to support current metabolic demands and tissue growth. To determine if the long period of offspring dependency in bats is associated with the need for an advanced state of tissue maturation prior to flight, we examined body composition during postnatal growth in the big brown bat, Eptesicus fuscus. Despite their large size at birth (22% of maternal mass), newborn bats are relatively immature, containing 82% body water in fat-free mass. However, the total body water content of newborn bat pups decreases to near-adult levels in advance of weaning, while concentrations of total body fat and protein exceed adult values. In contrast to many other mammals, postnatal growth of bat pups was characterized by relatively stable concentrations of calcium and phosphorus, but declining concentrations of magnesium. These levels remained stable or rebounded in late postnatal development. This casts doubt on the hypothesis that low rates of mineral transfer necessitate an extended lactation period in bats. However, our finding of near-adult body composition at weaning is consistent with the hypothesis that extended lactation in bats is necessary for the young to achieve sufficient tissue maturity to undertake the active flight necessary for independent feeding. In this respect, bats differ from most other mammals but resemble birds that must engage in active flight to achieve nutritional independence.

Chemical characterization of oligosaccharides in the milk of six species of New and Old World monkeys.

Human and great ape milks contain a diverse array of milk oligosaccharides, but little is known about the milk oligosaccharides of other primates, and how they differ among taxa. Neutral and acidic oligosaccharides were isolated from the milk of three species of Old World or catarrhine monkeys (Cercopithecidae: rhesus macaque (Macaca mulatta), toque macaque (Macaca sinica) and Hamadryas baboon (Papio hamadryas)) and three of New World or platyrrhine monkeys (Cebidae: tufted capuchin (Cebus apella) and Bolivian squirrel monkey (Saimiri boliviensis); Atelidae: mantled howler (Alouatta palliata)). The milks of these species contained 6-8% total sugar, most of which was lactose: the estimated ratio of oligosaccharides to lactose in Old World monkeys (1:4 to 1:6) was greater than in New World monkeys (1:12 to 1:23). The chemical structures of the oligosaccharides were determined mainly by (1)H-NMR spectroscopy. Oligosaccharides containing the type II unit (Gal(ß1-4)GlcNAc) were found in the milk of the rhesus macaque, toque macaque, Hamadryas baboon and tufted capuchin, but oligosaccharides containing the type I unit (Gal(ß1-3)GlcNAc), which have been found in human and many great ape milks, were absent from the milk of all species studied. Oligosaccharides containing Lewis x (Gal(ß1-4)[Fuc(α1-3)]GlcNAc) and 3-fucosyl lactose (3-FL, Gal(ß1-4)[Fuc(α1-3)]Glc) were found in the milk of the three cercopithecid monkey species, while 2-fucosyl lactose (5'-FL, Fuc(α1-2)Gal(ß1-4)Glc) was absent from all species studied. All of these milks contained acidic oligosaccharides that had N-acetylneuraminic acid as part of their structures, but did not contain oligosaccharides that had N-glycolylneuraminic acid, in contrast to the milk or colostrum of great apes which contain both types of acidic oligosaccharides. Two GalNAc-containing oligosaccharides, lactose 3'-O-sulfate and lacto-N-novopentaose I (Gal(ß1-3)[Gal(ß1-4)GlcNAc(ß1-6)]Gal(ß1-4)Glc) were found only in the milk of rhesus macaque, hamadryas baboon and tufted capuchin, respectively. Further research is needed to determine the extent to which the milk oligosaccharide patterns observed among these taxa represent wider phylogenetic trends among primates and how much variation occurs among individuals or species.

Chemical characterization of milk oligosaccharides of an African lion (Panthera leo) and a clouded leopard (Neofelis nebulosa).

The Carnivora include the superfamilies Canoidea and Feloidea. In species of Canoidea other than the domestic dog, Canis lupus, the milk contains only traces of lactose and much larger concentrations of oligosaccharides. In this study, lactose was found to be the dominant saccharide in the milk or colostrum of two species of Feloidea, namely the African lion (Panthera leo) and the clouded leopard (Neofelis nebulosa). In addition to lactose, the following oligosaccharides were characterized in the milk of a lion; Neu5Gc(α2-3)Gal(ß1-4)Glc (3'-NGc-SL), Fuc(α1-2)Gal(ß1-4)Glc (2'-fucosyllactose) and GalNAc(α1-3)[Fuc(α1-2)]Gal(ß1-4)Glc (A-tetrasaccharide). The colostrum of a clouded leopard contained 3'-NGc-SL, Gal(α1-3)Gal(ß1-4)Glc (isoglobotriose) and A-tetrasaccharide. These oligosaccharides differ in some respects from those previously identified in another species of Feloidea, the spotted hyena (Crocuta crocuta). These milks contained 3'-NGc-SL and A-tetrasaccharide, while spotted hyena colostrum did not; however, it contained Neu5Ac(α2-3)Gal(ß1-4)Glc (3'-NAc-SL) and Gal(α1-3)[Fuc(α1-2)]Gal(ß1-4)Glc (B-tetrasaccharide).

Variation in delta13C and delta15N diet-vibrissae trophic discrimination factors in a wild population of California sea otters.

The ability to quantify dietary inputs using stable isotope data depends on accurate estimates of isotopic differences between a consumer (c) and its diet (d), commonly referred to as trophic discrimination factors (TDFs) and denoted by delta(c-d). At present, TDFs are available for only a few mammals and are usually derived in captive settings. The magnitude of TDFs and the degree to which they vary in wild populations is unknown. We determined delta13C and delta15N TDFs for vibrissae (i.e., whiskers), a tissue that is rapidly becoming an informative isotopic substrate for ecologists, of a wild population of sea otters for which individual diet has been quantified through extensive observational study. This is one of the very few studies that report TDFs for free-living wild animals feeding on natural diets. Trophic discrimination factors of 2.2 per thousand +/- 0.7 per thousand for delta13C and 3.5 per thousand +/- 0.6 per thousand for delta15N (mean +/- SD) were similar to those reported for captive carnivores, and variation in individual delta13C TDFs was negatively but significantly related to sea urchin consumption. This pattern may relate to the lipid-rich diet consumed by most sea otters in this population and suggests that it may not be appropriate to lipid-extract prey samples when using the isotopic composition of keratinaceous tissues to examine diet in consumers that frequently consume lipid-rich foods, such as many marine mammals and seabirds. We suggest that inherent variation in TDFs should be included in isotopically based estimates of trophic level, food chain length, and mixing models used to quantify dietary inputs in wild populations; this practice will further define the capabilities and limitations of isotopic approaches in ecological studies.

Using stable isotopes to investigate individual diet specialization in California sea otters (Enhydra lutris nereis).

Differences in diet composition among conspecifics (dietary specialization) have been documented across a broad range of taxonomic groups and habitats, and such variation at the individual level is increasingly recognized as an important component of diversity in trophic interactions. Accurate identification of individual dietary specialization, however, requires longitudinal dietary records that are labor-intensive and cost-prohibitive to obtain for many species. Here we explore the use of stable isotopes (delta13C and delta15N) as a promising technique for detecting and quantifying patterns of individual dietary specialization. Southern sea otters (Enhydra lutris nereis) offer a unique opportunity for testing this approach because (1) they consume a wide variety of prey that span multiple trophic levels, habitats, and ecologically defined functional groups; and (2) individual diet specialization can be validated with existing observational data. We analyzed the isotopic composition of sea otter vibrissae (n = 31) in order to characterize inter- and intra-individual variation in sea otter diets at Monterey Bay, California, USA. At the population level, sea otters showed substantial variation in both delta13C and delta15N values, occupying nearly all of the "isotopic space" created by the diversity of isotopic signatures of potential prey taxa. Most of the variation in sea otter vibrissae was accounted for by differences between individuals, with much less contributed by within-individual variation. A majority of sea otters (approximately 80%) showed relatively little temporal variability in isotopic composition, suggesting that the proportional composition of most individuals' diets is relatively constant over time; a few individuals (approximately 20%) exhibited a high degree of intra-vibrissa isotopic variability, suggesting seasonal shifts in diet composition. These results and our interpretation of them were supported by long-term observational data on the diets of radio-tagged sea otters from the same population (n = 23). Our results demonstrate that stable isotopes can provide an efficient tool for measuring individual- and population-level dietary breadth and may be useful for studying populations where longitudinal data on individuals would otherwise be impossible to acquire. This will be critical for examining the causes and consequences of dietary variation within and among consumer populations, thereby improving our understanding of these important ecological and evolutionary processes at the community level.

Chemical characterization of oligosaccharides in chimpanzee, bonobo, gorilla, orangutan, and siamang milk or colostrum.

Neutral and acidic oligosaccharides were isolated from the milk or colostrum of four great ape species (chimpanzee (Pan troglodytes), bonobo (Pan paniscus), gorilla (Gorilla gorilla), and orangutan (Pongo pygmaeus)) and one lesser ape species (siamang (Symphalangus syndactylus)), and their chemical structures were characterized by (1)H-NMR spectroscopy. Oligosaccharides containing the type II unit (Gal(beta1-4)GlcNAc) were found exclusively (gorilla and siamang) or predominately (chimpanzee, bonobo, and orangutan) over those containing the type I unit (Gal(beta1-3)GlcNAc). In comparison, type I oligosaccharides predominate over type II oligosaccharides in human milk, whereas nonprimate milk almost always contains only type II oligosaccharides. The milk or colostrum of the great apes contained oligosaccharides bearing both N-glycolylneuraminic acid and N-acetylneuraminic acid, whereas human milk contains only the latter. Great ape milk, like that of humans, contained fucosylated oligosaccharides whereas siamang milk did not. Since these analyses are based on a limited number of individuals, further research on additional samples of great and lesser ape milk is needed to confirm phylogenetic patterns.

Rhesus macaque milk: magnitude, sources, and consequences of individual variation over lactation.

Lactation represents the greatest postnatal energetic expenditure for mammalian mothers, and a mother's ability to sustain the costs of lactation is influenced by her physical condition. Mothers in good condition may produce infants who weigh more, grow faster, and are more likely to survive than the infants of mothers in poor condition. These effects may be partially mediated through the quantity and quality of milk that mothers produce during lactation. However, we know relatively little about the relationships between maternal condition, milk composition, milk yield, and infant outcomes. Here, we present the first systematic investigation of the magnitude, sources, and consequences of individual variation in milk for an Old World monkey. Rhesus macaques produce dilute milk typical of the primate order, but there was substantial variation among mothers in the composition and amount of milk they produced and thus in the milk energy available to infants. Relative milk yield value (MYV), the grams of milk obtained by mammary evacuation after 3.5-4 h of maternal-infant separation, increased with maternal parity and was positively associated with infant weight. Both milk gross energy (GE) and MYV increased during lactation as infants aged. There was, however, a trade-off; those mothers with greater increases in GE had smaller increases in MYV, and their infants grew more slowly. These results from a well-fed captive population demonstrate that differences between mothers can have important implications for milk synthesis and infant outcome.

Chemical characterization of milk oligosaccharides of a spotted hyena (Crocuta crocuta).

The Carnivora include the superfamilies Canoidea and Feloidea. In species of Canoidea other than Canidae, the milk contains only traces of lactose and much larger concentrations of oligosaccharides. In this study, the following oligosaccharides were characterized in the milk of a spotted hyena, which is a species of Feloidea species: Neu5Ac(alpha2-3)Gal(beta1-4)Glc, Gal(alpha1-3)[Fuc(alpha1-2)]Gal(beta1-4)Glc, Gal(alpha1-3)Gal(beta1-4)Glc and Fuc(alpha1-2)Gal(beta1-4)Glc. Lactose was found to be the predominant saccharide; in this respect, the hyena milk is markedly different from the milks of most species of Canoidea species. The sole presence of 3'-SL in the spotted hyena milk is interesting, because the co-presence of 3'-SL and 6'-SL has been reported in the milk or colostrum of many mammalian species.