Extended Data Fig. 1: Overview of samples, study design and quality measures to validate the authenticity of the palaeofaeces.

a, Schematic of gene-catalogue and genome-reconstruction pipelines. b, Samples used in this study, archaeological sites and ¹⁴C dating. Data were obtained from this study (Mexico) and previous studies: Fiji³¹, Peru⁴, Madagascar¹³, Tanzania⁵⁷, USA^4,55, Denmark⁵⁶ and Spain⁵⁶. Map data are from Google Maps (2021 Google, INEGI). c, Scanning electron microscopy images of dietary remains in the palaeofaeces. Zape1, maize pollen grains (more than 191,000 grains per gram) (top) and agave phytoliths (middle and bottom). Zape2, U. maydis spores (hundreds of millions per gram). Zape3, Chenopod or amaranth foliage and/or buds (smaller pollen) and squash (larger pollen with spines). UT30.3, druse phytoliths, annular xylem vessel secondary wall thickenings and epidermis of Cactaceae. A complete description is provided in Supplementary Information section 2. Reproducibility and independently repeated experiments are described in the Methods. d, Principal component analysis of the species composition of palaeofaeces, soil samples and publicly available archaeological sediment samples^70,71. Species were identified by MetaPhlAn2²⁰. e, Prediction of source of microbial communities by SourceTracker2⁷² using the species abundance matrix from MetaPhlAn2 as input. Archaeological sediment samples included three soil samples collected in this study, seven Holocene human-associated sediments from CoproID⁷¹ and 40 Pleistocene sediment samples⁷⁰. f, The percentage of reads aligned to the MetaPhlAn2 database per sample (HMP, n = 146; Mexican, n = 22; Fijian, n = 174; palaeofaeces, n = 8; soil, n = 3) (Supplementary Information section 4). g, aDNA damage levels of Firmicutes and Bacteroidetes genomes for medium-quality and high-quality pre-filtered and filtered bins (two-tailed Wilcoxon rank-sum test; pre-filtered bins Bacteroidetes, n = 69 MAGs; pre-filtered bins Firmicutes, n = 359 MAGs; filtered bins Bacteroidetes, n = 24 MAGs; filtered bins Firmicutes, n = 161 MAGs) (Supplementary Information section 5). 5p, 5′ end; 3p, 3′ end. h, Abundances of VANISH²¹ and BloSSUM²² families as identified by MetaPhlAn2²⁰ (palaeofaeces n = 8; non-industrial n = 370; industrial n = 418). In f–h, data are presented as box plots (middle line, median; lower hinge, first quartile; upper hinge, third quartile; upper whisker extends from the hinge to the largest value no further than 1.5× the interquartile range from the hinge; lower whisker extends from the hinge to the smallest value at most 1.5× the interquartile range from the hinge; data beyond the end of the whiskers are individually plotted outlying points).