Characterisation of microbial attack on archaeological bone

Abstract

As part of an EU funded project to investigate the factors influencing bone preservation in the archaeological record, more than 250 bones from 41 archaeological sites in five countries spanning four climatic regions were studied for diagenetic alteration. Sites were selected to cover a range of environmental conditions and archaeological contexts. Microscopic and physical (mercury intrusion porosimetry) analyses of these bones revealed that the majority (68%) had suffered microbial attack. Furthermore, significant differences were found between animal and human bone in both the state of preservation and the type of microbial attack present. These differences in preservation might result from differences in early taphonomy of the bones.

Introduction

Bone is used extensively as a source of information in archaeological research as it is usually the only animal or human tissue preserved, though often it is altered or destroyed as well. With the development of biomolecular analysis, the significance of understanding bone preservation has increased. One of the earliest types of alteration is microbial attack. Yoshino et al. [42]observed evidence of microbial alteration in bone 5 years post-mortem, indeed Haversian canals were (still) filled with bacteria. Bell et al. [5]found evidence of microbial attack in bone in a forensic study, as soon as 3 months post mortem, though this sample was recovered from a predator scat.

Biological alteration in bone is in most cases caused by fungi [31], bacteria [2], [14], [23], or cyanobacteria in marine environments [4]. Wedl first described microbial attack on mineralised tissue in 1864, finding tunnels of approximately 8 μm in diameter in sections of teeth exposed to untreated well water and in fossil reptile teeth. This type of tunnelling is defined as Wedl or centrifugal tunnelling by Hackett [14]. These tunnels range in diameter from 5 to 10 μm and under electron microscopy [31]appear empty with well-defined calcified walls, implying that collagen and mineral are both resorbed by the fungi.

Three types of microscopical focal destruction (mfd)—described by Hackett as ‘linear longitudinal’, ‘budded’ and ‘lamellate’—are generally assumed to be caused by bacteria. These mfd can be distinguished histologically by morphology; size, shape, the presence of a hypermineralised rim and the presence of a lamellate content. Under light microscopy these mfd show a granulous or fibrillar content, but higher magnification shows that their interior actually consists of small pores [2], [23], [40]. Mercury intrusion porosimetry (HgIP) shows an increased volume in pores with diameters in the 0.1–10 μm region, with the largest increase observed at approximately 0.6 and 1.2 μm [35], [40], consistent with the average diameter for bacteria (0.5 μm [18]). Indeed, Baud and Lacotte [2]argue that the small pores that are visible at electron microscope level are the mineralised remains of bacteria, in which case they would not form the interconnected porosity observed by Turner-Walker et al. [40].

We report here some results from a European study, which attempted to identify the key contributory factors to bone deterioration. Biological alteration of archaeological bone is a widespread phenomenon apparently not limited to a single burial environment [21]. Its effect on bone preservation is important as it can accelerate degradation by increasing the bone porosity [35]and simultaneously reduces chances of success for biomolecular research through loss or contamination of target molecules [9]. Two methodological approaches, mercury intrusion porosimetry (HgIP) and histology, are combined for the first time in this project. While histology has been widely used previously to characterise microbial attack (e.g. [3], [11], [14]), HgIP makes it possible to analyse a larger sample of bone and provides quantitative results [40].