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Most researchers assume minimal impact of pretreatment on strontium isotope ratios ( 87 Sr/ 86 Sr) for bones and teeth, and methods vary tremendously. We compared 14 pretreatment methods, including no prep other than powdering enamel, ashing, soaking in water, an oxidizing agent (bleach or hydrogen peroxide) or acetic acid (0.1 M, 1.0 M, and 1.0 M buffered with calcium acetate), and a combination of these steps. We prepared and analyzed aliquots of powdered molar enamel from three proboscideans (one modern captive Indian elephant, Elephas maximus indicus ; one Pleistocene mastodon, Mammut americanum ; and one Miocene gomphothere, Afrochoerodon kisumuensis ). Each pretreatment was performed in triplicate and we measured 87 Sr/ 86 Sr, Sr concentration, and uranium (U) concentration, using the same lab space and instrumentation for all samples. Variability in 87 Sr/ 86 Sr and Sr and U concentrations was considerable across pretreatments. Mean 87 Sr/ 86 Sr across methods ranged from 0.70999 to 0.71029 for the modern tooth, 0.71458 to 0.71502 for the Pleistocene tooth, and 0.70804 to 0.70817 for the Miocene tooth. The modern tooth contained the least Sr and negligible U. The Pleistocene tooth contained slightly more Sr and measurable amounts of U, and the Miocene tooth had approximately 5x more Sr and U than the Pleistocene tooth. For all three teeth, variance in 87 Sr/ 86 Sr, Sr concentrations, and U concentrations among replicates was statistically indistinguishable across pretreatments, but there were apparent differences among pretreatments for the modern and Pleistocene teeth. Both contained relatively little Sr, and it is possible that small amounts of exogenous Sr from reagents, building materials or dust affected some replicates for some pretreatments. For the modern tooth, median 87 Sr/ 86 Sr varied considerably (but statistically insignificantly) across pretreatments. For the Pleistocene tooth, variability in median 87 Sr/ 86 Sr was also considerable; some pretreatments were statistically distinct but there were no obvious patterns among methods. For the Miocene tooth, variability in median 87 Sr/ 86 Sr was much smaller, but there were significant differences among pretreatments. Most pretreatments yielded 87 Sr/ 86 Sr and Sr concentrations comparable to, or lower than, untreated powder, suggesting selective removal of exogenous material with high 87 Sr/ 86 Sr. Further evaluation of the mechanisms driving isotopic variability both within and among pretreatment methods is warranted. Researchers should clearly report their methods and avoid combining data obtained using different methods. Small differences in 87 Sr/ 86 Sr could impact data interpretations, especially in areas where isotopic variability is low. 

We present eight new radiocarbon dates for endemic and invasive rodents from Trouing Jérémie #5, a paleontologically-rich sink hole on the Tiburon Peninsula, Haiti. This includes new dates for two species that have been previously directly dated ( Isolobodon portoricensis and Brotomys voratus) as well as three endemic rodents which have no previous direct radiometric dates ( Plagiodontia velozi, Hexolobodon phenax, and Rhizoplagiodontia lemkei). The radiocarbon date for P. velozi, the largest of these species, is from the very early Holocene (10,995 ± 190 calendar years before present; Cal BP). Specimens of medium-bodied species, H. phenax and R. lemkei date to the mid-Holocene, while specimens of smaller-bodied I. portoricencis and B. voratus have dates falling in the Late-Holocene. These dates confirm that several of the extinct rodent species coexisted with the first humans, who arrived on the island ca. 6000 years ago. In contrast, murid Rattus specimens date to the last few centuries. Rats arrived with Europeans in the late 15th or early 16th Century and a radiocarbon date of ca. 500 Cal BP for one individual suggests that they likely spread quickly across the island. Collectively, these dates establish that vertebrate accumulations at Trouing Jérémie #5 span the Holocene. Remains from this site may provide a useful time sequence for future work examining ecological change across the Holocene as well as regional extirpation patterns. 

Introduced predators currently threaten endemic animals on Madagascar through predation, facilitation of human-led hunts, competition, and disease transmission, but the antiquity and past consequences of these introductions are poorly known. We use directly radiocarbon dated bones of introduced dogs ( Canis familiaris ) to test whether dogs could have aided human-led hunts of the island’s extinct megafauna. We compare carbon and nitrogen isotope data from the bone collagen of dogs and endemic “fosa” ( Cryptoprocta spp.) in central and southwestern Madagascar to test for competition between introduced and endemic predators. The distinct isotopic niches of dogs and fosa suggest that any past antagonistic relationship between these predators did not follow from predation or competition for shared prey. Radiocarbon dates confirm that dogs have been present on Madagascar for over a millennium and suggest that they at least briefly co-occurred with the island’s extinct megafauna, which included giant lemurs, elephant birds, and pygmy hippopotamuses. Today, dogs share a mutualism with pastoralists who also occasionally hunt endemic vertebrates, and similar behavior is reflected in deposits at several Malagasy paleontological sites that contain dog and livestock bones along with butchered bones of extinct megafauna and extant lemurs. Dogs on Madagascar have had a wide range of diets during the past millennium, but relatively high stable carbon isotope values suggest few individuals relied primarily on forest bushmeat. Our newly generated data suggest that dogs were part of a suite of animal introductions beginning over a millennium ago that coincided with widespread landscape transformation and megafaunal extinction. 

No endemic Madagascar animal with body mass >10 kg survived a relatively recent wave of extinction on the island. From morphological and isotopic analyses of skeletal “subfossil” remains we can reconstruct some of the biology and behavioral ecology of giant lemurs (primates; up to ∼160 kg) and other extraordinary Malagasy megafauna that survived into the past millennium. Yet, much about the evolutionary biology of these now-extinct species remains unknown, along with persistent phylogenetic uncertainty in some cases. Thankfully, despite the challenges of DNA preservation in tropical and subtropical environments, technical advances have enabled the recovery of ancient DNA from some Malagasy subfossil specimens. Here, we present a nuclear genome sequence (∼2× coverage) for one of the largest extinct lemurs, the koala lemur Megaladapis edwardsi (∼85 kg). To support the testing of key phylogenetic and evolutionary hypotheses, we also generated high-coverage nuclear genomes for two extant lemurs, Eulemur rufifrons and Lepilemur mustelinus , and we aligned these sequences with previously published genomes for three other extant lemurs and 47 nonlemur vertebrates. Our phylogenetic results confirm that Megaladapis is most closely related to the extant Lemuridae (typified in our analysis by E. rufifrons ) to the exclusion of L. mustelinus , which contradicts morphology-based phylogenies. Our evolutionary analyses identified significant convergent evolution between M. edwardsi and an extant folivore (a colobine monkey) and an herbivore (horse) in genes encoding proteins that function in plant toxin biodegradation and nutrient absorption. These results suggest that koala lemurs were highly adapted to a leaf-based diet, which may also explain their convergent craniodental morphology with the small-bodied folivore Lepilemur . 

Orangutans have a 7.6 year average inter-birth interval, the longest of any mammal. Suckling occurs throughout that interval, until the birth of the next offspring, but it is unclear how important milk consumption is during that period, as we cannot assess the actual intake amount. Measurement of stable carbon isotope ratios (d13C), stable nitrogen isotope ratios (d15N) and nitrogen content of feces (%N) provide evidence of the transition between breast milk to solid food. Here we present pilot data on these isotopic ratios from matched fecal samples of mothers and offspring (n=43), collected from wild orangutans in Gunung Palung National Park, Borneo, Indonesia. We found that the youngest infant (2.3 yrs) had the highest d15N values overall, indicative of a higher percentage of animal products (milk) in the diet. Older juveniles (5.8 yrs) did not consistently show higher d15N than their mothers. This may indicate variation in suckling frequency or the amount of breast milk consumed per suckling session. Adolescents (10-13 yrs) showed significantly (GLMM, p< 0.007) lower d15N than samples taken from the same day on their mothers. This is surprising given that they were eating similar diets. We thus compare the isotopic signature of the plants consumed to examine the sources of this variation. We conclude that analysis of fecal samples collected from wild orangutans can be used to assess the relative importance of breast milk in the diet, but caution that isotopic excretion may also be effected by differences in the isotopic content of the diet. Funders: NSF (BCS-1638823, BCS-0936199); National Geographic; USFish/Wildlife (F15AP00812, F13AP00920, 96200-0-G249, 96200-9-G110); Leakey; Disney Wildlife Conservation; Wenner-Gren; Nacey-Maggioncalda; Orangutan Conservancy; Conservation-Food-Health; Woodland Park Zoo; Holloman Price; AZA; Ocean Park Conservation; USAID; Arcus