Ecological niche of Neanderthals from Spy Cave revealed by nitrogen isotopes of individual amino acids in collagen Yuichi I. Naito a, * , Yoshito Chikaraishi b , Doroth  ee G. Drucker a , Naohiko Ohkouchi b , Patrick Semal c , Christoph Wißing a , Herv  e Bocherens a, d a Fachbereich Geowissenschaften, Pal€ aobiologie (Biogeologie), Universit€ at Tübingen, H€ olderlinstraße 12, 72074 Tübingen, Germany b Institute of Biogeosciences, Japan Agency for Marine-Earth Science and Technology, 2-15 Natsushima-cho, Yokosuka 237-0061, Japan c Department of Paleontology, Royal Belgian Institute of Natural Sciences, 1000 Brussels, Belgium d Senckenberg Center for Human Evolution and Paleoenvironment (HEP), Universit€ at Tübingen, H€ olderlinstraße 12, 72074 Tübingen, Germany article info Article history: Received 6 November 2014 Accepted 31 January 2016 Available online xxx Keywords: Neanderthal Nitrogen isotope Amino acid Ecological niche Subsistence strategy Plant diet abstract This study provides a refined view on the diet and ecological niche of Neanderthals. The traditional view is that Neanderthals obtained most of their dietary protein from terrestrial animals, especially from large herbivores that roamed the open landscapes. Evidence based on the conventional carbon and nitrogen isotopic composition of bulk collagen has supported this view, although recent findings based on plant remains in the tooth calculus, microwear analyses, and small game and marine animal remains from archaeological sites have raised some questions regarding this assumption. However, the lack of a protein source other than meat in the Neanderthal diet may be due to methodological difficulties in defining the isotopic composition of plants. Based on the nitrogen isotopic composition of glutamic acid and phenylalanine in collagen for Neanderthals from Spy Cave (Belgium), we show that i) there was an inter- individual dietary heterogeneity even within one archaeological site that has not been evident in bulk collagen isotopic compositions, ii) they occupied an ecological niche different from those of hyenas, and iii) they could rely on plants for up to ~20% of their protein source. These results are consistent with the evidence found of plant consumption by the Spy Neanderthals, suggesting a broader subsistence strategy than previously considered. © 2016 Elsevier Ltd. All rights reserved. 1. Introduction Dietary subsistence is a major component of the biology of a human population. Neanderthals became extinct around 40,000 years ago (Higham et al., 2014), and their subsistence strategies have attracted a lot of scientific attention since this information is crucial to evaluating their cognitive abilities and the possible reasons for their extinction. Several hypotheses regarding the demise of the Neanderthals at the time when early anatomically modern humans entered Europe involve dietary differences be- tween the two types of hominins (Bocherens and Drucker, 2006; Froehle and Churchill, 2009; Hoffecker, 2009). Numerous studies, based on different approaches such as zooarchaeology and tooth wear patterns, have provided convincing evidence for a Neanderthal diet comprised largely of meat from large terrestrial herbivores (Lalueza Fox and P erez-P erez, 1993; Gardeisen, 1999; Boyle, 2000; Ready, 2010; Gaudzinski, 2014; Fiorenza et al., 2015). In this context, the first isotopic study of Neanderthal bone collagen published over two decades ago showing an iso- topic composition close to those of animal predators such as wolves and hyenas was not surprising (Bocherens et al., 1991). Since then, many Neanderthal specimens have been analysed, and in all cases where collagen was well-preserved, a similar pattern was found (Fizet et al., 1995; Bocherens et al., 1999, 2005, 2013; Richards et al., 2008; Richards and Trinkaus, 2009). On the other hand, recent findings of plant remains trapped in a tooth calculus testify to the consumption of plants by Neanderthals (Henry et al., 2011, 2014; Hardy et al., 2012). Studies of dental microwear support a more diverse diet in some Neanderthal groups, which has been linked with increasing tree-cover (El Zaatari et al., 2011). However, as none of the palaeodietary ap- proaches is able to accurately quantify the proportions of meat or * Corresponding author. E-mail address: ynaito@sss.fukushima-u.ac.jp (Y.I. Naito). Contents lists available at ScienceDirect Journal of Human Evolution journal homepage: www.elsevier.com/locate/jhevol http://dx.doi.org/10.1016/j.jhevol.2016.01.009 0047-2484/© 2016 Elsevier Ltd. All rights reserved. Journal of Human Evolution 93 (2016) 82e90