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The Effects of Soft Tissue Removal Methods on Porcine Skeletal Remains: a Comparative Analysis

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The Effects of Soft Tissue Removal Methods on Porcine Skeletal Remains: a Comparative Analysis February 2021 New Florida Journal of Anthropology Tarbet Hust, E.S. and Snow, M.H. The Effects of Soft Tissue Removal Methods on Porcine Skeletal Remains: A Comparative Analysis Emily S. Tarbet Hust1, Meradeth H Snow2 1SNA International, 2University of Montana Abstract Sus scrofa domesticus limbs were obtained as a human proxy to study the effects of five distinct materials used in published methods of flesh removal: Dermestes lardarius beetles (also referred to as dermestids), distilled-water boil, bleach boil, enzyme-based detergent simmer, and ammonia simmer. Each method was evaluated based on a set of specific criteria, focusing on time efficiency, macroscopic damage, and the effects on DNA preservation and potential for future analysis. While the dermestid beetles had the longest time-expectancy and were the most labor-intensive method, they caused minimal damage to the bone surface and did not appear to affect the DNA preservation. Heated maceration methods sped up the process considerably, but that often led to decreased DNA quantity and minimal to severe amounts of macroscopic damage. The ammonia simmer method was the only method tested that was found in zoological literature but did not appear to have any published use within the forensic field, operating occasionally instead as a degreasing agent. While the ammonia method required the most safety precautions, the method was efficient both in time and tissue removal, and left amplifiable DNA, perhaps indicating a potential future in more forensic contexts. In contrast, the enzyme-based detergent method, often praised in published literature, performed poorly in multiple categories of evaluation. Each method proved to have different advantages and disadvantages, with no method performing the best or worst in every evaluated criterion. The results of this re- search highlight how differently each method performs and how easily skeletal material, and the DNA within, can be affected by maceration techniques. Method selection can severely impact later analysis and the choice should be made with consideration and awareness of the potential risks and desired results. Keywords: Forensic Anthropology; Forensic Science; Maceration; Defleshing; mtDNA Introduction nally, forensic anthropologists al- so use Maceration, or the removal of soft these techniques during casework to re- tissue from skeletal remains, is a process duce fleshed or decomposing individuals familiar to members of a wide array of to skeletal remains for analysis. Due to scientific fields and contexts. Flesh re- the wide array of fields and goals behind moval is used in museums to prepare the removal of soft tissue from skeletal bones for display and for comparative re- remains, there are an equally wide array search, hunters may prepare remains for of developed methods to perform the showcasing, and body donation research task. facilities use recently skeletonized re- However, no consistent standard mains of modern individuals to refine and exists for forensic applications of tissue update the markers used in biological removal, which leads to a wide range of profile construction and analysis. Additio- protocols and varied guidance in the am- Tarbet Hust, E.S. and M. Snow (2021) The Effects of Soft Tissue Removal Methods on Porcine Skeletal Remains: A Comparative Analysis. New Florida Journal of Anthropology 1(2), 30-46 DOI 10.32473/nfja.v1i2.124117 30 February 2021 New Florida Journal of Anthropology Tarbet Hust, E.S. and Snow, M.H. ount of additives, temperatures, and tim- scored and documented in detail. The re- ing. While the interest in method efficacy sults in this study were then used to cre- has increased in recent years (Ajayi et ate a method selection flowchart, empha al., 2016; Ecklund, 2007; Frank et al., sizing the different advantages and dis- 2015; Mann & Berryman, 2012), there re- advantages of the tested methods. Be- mains a lack of cohesive agreement on cause the goals and intentions behind which additives may perform best in sim- soft tissue removal tend to vary widely ilar scenarios or which methods have the between fields and contexts, the results most negative impact on skeletal remains are designed with a specific emphasis on and DNA viability. While many of these the viability and practicality of the method methods and additives are used outside within a forensic context, but the general of the forensic sciences, safety and con- information can be relevant to any field sistency in forensic analysis is of the ut- familiar with the task. most importance. The bulk of the litera- ture reflects primarily on macroscopic Materials and Methods damage that affects trauma analysis, as Research Sample well as methods that may complicate The research sample consisted of DNA extraction (Ecklund, 2007; Fenton 17 domesticated pig (Sus scrofa domes- et al., 2003; Frank et al., 2015; King & ticus) limbs from the North Dakota State Birch, 2015; Lynn & Fairgrieve, 2009). University agricultural program. The pig Heated maceration methods have limbs were primarily the hock portion of long been the preferred and more inves- the leg and contained fragments of two tigated method due to their shorter dura- long bones, either the tibia and fibula or tion and less manually laborious nature radius and ulna depending on whether it than unheated methods. Simmering and was a hind or front limb. The pigs were all boiling techniques utilizing a variety of from the same environment and were additives have become common practice butchered in the same manner at approx- (Couse & Connor, 2015). The additives imately the same age, all being just under range from household cleaning products a year old. such as dish and laundry detergent to di- The 17 pig hocks were assigned a rectly adding chemicals such as sodium maceration method at random, with three perborate or carbonate, while still other hocks being assigned to each category methods utilize heated water with no ad- and the remaining two being reserved for ditional additives (Lee et al., 2010; Uhre manual soft tissue removal to allow for a et al., 2015). DNA comparison with samples that did The following research focuses on not undergo one of the tested methods. several commonly used soft tissue re- Control DNA samples were not taken moval methods, involving four heated prior to the maceration of the pig hocks to maceration methods and one method uti- avoid additional variables and considera- lizing Dermestes lardarius beetles, here- tions in regard to specific element and re- after referred to as dermestids. The gion of bone (Antinick & Foran, 2018). In amount of additives used, the duration to lieu of a true control sample, consistency completion each maceration method was attempted by ensuring all pigs were took, and the effects the method had on the same age from the same environ- the resulting skeletonized material were ment and butchered at the same time Tarbet Hust, E.S. and M. Snow (2021) The Effects of Soft Tissue Removal Methods on Porcine Skeletal Remains: A Comparative Analysis. New Florida Journal of Anthropology 1(2), 30-46 DOI 10.32473/nfja.v1i2.124117 31 February 2021 New Florida Journal of Anthropology Tarbet Hust, E.S. and Snow, M.H. with the same technique to limit back- in skeletal preparation within a zoological ground variation as much as possible museum. In forensic uses, ammonia (Arismendi et al., 2004). While this is not tends to be used as a degreaser after the a true control sample, whereby samples application of a primary method (Lee et from each pig hock would have been col- al., 2010), but in zoological preparation lected prior to treatment, the two un- solutions of up to 50% ammonia can be treated pig hocks can be used for com- used in a simmer for the entire process parison by providing an estimation and (Hoffmeister and Lee, 1963). The re- expectation of the DNA preservation and searcher’s own experience following zo- degradation resulting from the tested ological museum protocols involved methods. soaking delicate remains in a room tem- perature ammonia and water mixture for Tested Maceration Methods several days to remove residual tissue Five soft tissue removal methods, and grease that the dermestid colony consisting of four heated maceration leaves behind. Personal experience with methods and one method utilizing a col- use of ammonia and published uses in ony of dermestids, were selected for zoological contexts were combined to study based on methods found in previ- create the method performed in this ously published literature and researcher study. experience (Table 1). All the heated Two methods, distilled water and maceration method materials, including 6.25% bleach solution, were kept at a low start-up equipment, are widely accessi- boil, while the 12.25% household ammo- ble and relatively low cost. Cost could be nia and the 10% enzyme-based deter- additionally lowered if tap water, as op- gent methods were kept at a lower sim- posed to the distilled water used in these mering temperature for the duration of trials, and off-brand additives were used, the test. A laundry detergent with an ac- and materials were purchased in bulk cessible ingredient list was utilized to quantities. confirm that both lipase and protease en- All heated maceration methods zymes were present in the selected de- were performed in a small, ventilated tergent (Uhre et al., 2015). Method tem- room with as many variables kept con- peratures were based on those used in stant across methods as possible. Pig selected references (Table 1). The tests hocks subject to heated maceration were monitored at half-hour checkpoints methods did not undergo any manual for temperature checks and photos to en- processing (Couse & Connor, 2015). The sure consistency of temperature through- heated methods were performed using out the duration of the method and to an 18qt stockpot containing a two-gallon document the bone surface.
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