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Palaeoproteomic analysis of skeletal remains from Grotta Guattari (Italy) (advance online)

D. ; https://orcid.org/0000-0002-8239-6381 MylopotamitakiMax Planck Institute for Evolutionary Anthropology , Leipzig , GermanyS. SilvestriniDepartment of Cultural Heritage , University of Bologna , Via Degli Ariani 1 , Ravenna 48121 , ItalyG. TrocheGlobe Institute , University of Copenhagen , Copenhagen , DenmarkFerracci A.Department of History, Culture and Society , University of Rome Tor Vergata , Rome , ItalyM. GattaDepartment of History, Culture and Society , University of Rome Tor Vergata , Rome , ItalyM. RolfoDepartment of History, Culture and Society , University of Rome Tor Vergata , Rome , ItalyE. SalviatiDepartment of Pharmacy , University of Salerno , Fisciano , SA , ItalyP. CampigliaDepartment of Pharmacy , University of Salerno , Fisciano , SA , ItalyF. HarkingNovo Nordisk Foundation Center for Protein Research , University of Copenhagen , Copenhagen , DenmarkJ. OlsenNovo Nordisk Foundation Center for Protein Research , University of Copenhagen , Copenhagen , DenmarkM. RomandiniDepartment of Cultural Heritage , University of Bologna , Via Degli Ariani 1 , Ravenna 48121 , ItalyJ. HublinS. BenazziDepartment of Cultural Heritage , University of Bologna , Via Degli Ariani 1 , Ravenna 48121 , ItalyF. WelkerGlobe Institute , University of Copenhagen , Copenhagen , Denmark

2026en

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Litsenziya: cc-by-nc

ABI

Annotatsiya

Excavations at Grotta Guattari (Italy; 66–65 ka) in the previously unexplored Antro del Laghetto area uncovered numerous morphologically unidentified skeletal remains from both the cave’s internal (aquatic) and external (dry) sectors, providing an opportunity to investigate how contrasting depositional environments influence protein preservation. Taxonomic identifications were attempted using MALDI-ToF (n = 200) and LC-MS/MS (n = 490). LC-MS/MS provided limited taxonomic identifications for specimens from aquatic environments, whereas MALDI-ToF generated no reliable taxonomic identifications for the same environment. However, it should be noted that the proteomic analysis was performed on randomly selected bone specimens from different locations throughout the cave, so direct performance comparisons between MALDI-ToF and LC-MS/MS cannot be made. The protein aggregation capture (PAC) extraction method failed to generate taxonomic results, whereas a consecutive acid and ammonium bicarbonate (Ambic) extraction combined with LC-MS/MS yielded validated genus-level assignments for 142 specimens. Among these, four specimens were assigned to the genus Homo following additional validation, while acknowledging the limitations imposed by false discovery rate filtering and the absence of a Neanderthal proteome in the reference database. These findings highlight the importance of tailored proteomic workflows for highly degraded archaeological assemblages.

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