Periodontal diseases are chronic inflammatory conditions that affect the supporting structures of teeth, including the gingiva, periodontal ligament, cementum, and alveolar bone [[1], [2], [3]]. Periodontal disease has been traditionally divided into two categories, gingivitis and periodontitis, based on whether attachment loss occurred [[4], [5], [6], [7]]. The term gingivitis (or dental plaque-induced gingival diseases) is used to designate a reversible stage of periodontal disease since gingivitis is an inflammatory lesion resulting of general increase in plaque mass around the gingival margin without periodontal ligament and alveolar bone loss [3]. By contrast, periodontitis is a chronic and irreversible inflammatory disease that results from the accumulation of subgingival biofilm (dental plaque), microbial dysbiosis (i.e. imbalanced oral microbiome) with overgrowth of pathogenic bacteria in the gingival crevicular fluid, dental rots and epithelium, and the immune host response leading to destruction of periodontium, which subsequently progresses to alveolar bone loss, resulting in mobility and potentially loss of the affected tooth [3,[8], [9], [10], [11], [12], [13]]. Today, it is estimated that 20–50 % of the global population is affected by periodontitis [14] with severe form estimated to affect around 19.0 % of the global adult population, representing the 6th most prevalent condition in the world [15,16].

Molecular studies have shown that the human oral microbiome can consist of up 700 differing taxa, [17] with over 400 of these taxa being identified in the periodontal pockets alone [18]. The traditional 1990's theory of microbial etiology of periodontitis recognized the role of the so-called ‘red complex’, which comprises Gram-negative anaerobes Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia as the main bacteria responsible for active periodontitis [19]. However, this model has been reconsidered in 2012 and 2016 under the lens of a synergistic and dysbiotic microbial community rather than by selected ‘periopathogens' (i.e., the red complex) and by the concept of Porphyromonas gingivalis as the ‘key-stone pathogen’ responsible of biofilm community of pathogens, respectively. More recently, the IMPEDE model developed in 2020 seeks to explain the nexus between periodontal inflammation and change in the polymicrobial biofilm in the gingival sulcus (for a review [19]).

Dysbiosis in the oral microbiota may be influenced by various non-modifiable and modifiable risks. The former includes host determinants (e.g., immune function, genetics) and menopause, while the latter include smoking, diabetes mellitus, and pregnancy [14,[20], [21], [22]]. Additionally, comorbidities of periodontitis with systemic disorders like cardiovascular disease, rheumatoid arthritis, inflammatory bowel disease are well-established in clinical practice [[23], [24], [25]]; therefore, periodontitis has not only a central role within the realm of oral health, but it represents a major public health issue [26,27].

Among the environment determinants, the role of diet and nutrition towards the expression periodontal diseases is largely overlooked since dietary advice in the dental setting predominantly focuses on the prevention of dental caries [28]. However, balanced macro- and micronutrient intake is essential in periodontal health [29]. It has been demonstrated that microbial dysbiosis and periodontal inflammation is not merely due to inappropriate oral hygiene but also includes host environment factors like diets rich in fermentable and/or processed carbohydrates, high-fat dairy products, and saturated fat in meat that promote inflammatory reaction and progression in periodontal diseases [28,[30], [31], [32], [33]], while rich in Omega-3 fatty acids, vitamins, minerals and vegetables may increase periodontal health [28,34,35].

In human archaeological remains, periodontitis is traditionally analyzed by metric measurements with >2 mm threshold of root exposure as indicative of periodontitis [e.g., [[36], [37], [38], [40], [41]]] against the combination of metric and qualitative alveolar bone evaluation [[42], [43], [44], [45], [46], [47], [48]] or exclusive examination of alveolar bone texture [e.g., [39], [[49], [50], [51], [52], [53], [54]]]. A combined approach of vertical (localized) and horizontal (generalized) bone loss evaluation was applied by Raitapuro-Murray et al. [55]. However, macroscopic evaluation of periodontitis has notable limitations, particularly due to postmortem damage, and individual variation in bone preservation, which can obscure or mimic pathological features and lead to potential misinterpretations. To address these challenges, combining macroscopic analysis with complementary analytical techniques enhances both diagnostic accuracy and the depth of interpretative. In this regard, the application of omic approaches such as meta-genomics and proteomics, to dental calculus have only recently provided a new tool to gain insights into periodontitis and to access genetic and protein information of a whole microbial community directly [17,[56], [57], [58], [59], [60]]. Palaeoproteomics of dental calculus is an effective method for investigating both the etiology of and host responses to ancient periodontal disease since subgingival dental calculus formation promote periodontitis [30,61,62]. Proteins are functional agents, and their expression differs in response to pathological conditions. These pieces of evidence, revealing information on oral pathologic processes, could not be obtained solely by DNA analysis, which could only reveal the presence of certain taxa in analyzed specimens. The potential of palaeoproteomics analysis of dental calculus for studying relationship of diet, health, immune system and oral diseases in the past has not been fully exploited [e.g., [57], [[63], [64], [65]]], despite successful applications in studies aiming at dietary reconstruction in ancient Eurasia [e.g., [[66], [67], [68], [69], [70], [71], [72], [73], [74], [75], [76]]].

In this study, we aim to reconstruct periodontal health in pre-Roman groups from the 1st millennium BCE Italy, a period characterized by social stratification, differential access to resources and intensification of agriculture. This phenomenon was of considerable historical and economic interest [77] and is evidenced by archaeological findings, including the development of complex urban centers, differential burial types and mortuary accoutrements, and definition of rural landscapes shaped by intensive agriculture, with land use focused on staple crops production and livestock husbandry [[78], [79], [80], [81]], as well as by waterworks and discovery of iron tools like ploughshares, sickles, and hoes that points to the technological means for exploiting of arable fields [82]. To explore the biological consequences of these changes, we integrate macroscopic analysis of interdental septa - as indicators of periodontal health - with palaeoproteomic profiling of dental calculus. This combined approach tests the hypothesis that social and economic dynamics of this period are reflected in a proinflammatory diet. Increased reliance on carbohydrate-rich staples as a result of agricultural intensification, as well as greater consumption of animal proteins, often associated with the dietary privileges of emerging elites, may have disrupted oral eubiosis, promoting the proliferation of pathogenic taxa such as Porphyromonas gingivalis.