It is well known that the risk of contamination is particularly high at early specimen preparation stages when the tissues are still fresh and the body fluids are liquid. Contamination may be avoided by means of appropriate prevention measures, careful cleansing and disinfection of working surfaces and instruments, as well as appropriate disposal of tissues remaining after the procedure (Smith and Holladay, 2001). Little information is available on the possibility of plastinated specimens being infected by microbial species after all plastination stages are completed and the specimens are released for educational purposes.
S10 plastination silicone: structure vs susceptibility to infection
Studies on the chemical activity used in S10 cold-temperature plastination technique suggest that the polyalkyl siloxane within Biodur S10 is likely to be a hydroxyl-terminated polydimethylsiloxane or very similar silicone monomer (Holladay et al., 2001). Polydimethylsiloxanes (PDMS) are macromolecular organosilicon polymers with -Si-O- bonding pattern repeated in the molecular chains. The polysiloxane structure is summarized using the formula -[R2Si-O]- where R is a methyl (alkyl) group; thus, the compounds are classified as alkyl siloxanes (Mojsiewicz-Pieńkowska and Łukasiak, 2003). Silicones, particularly methylsilicone rubbers, are used for production of biomedical materials such as breast implants (Dunn et al., 1992), nasal implants (Deva et al., 1998), ophthalmological implants (Teferra, 2017) and vocal cord implants (Echternach et al., 2008). Polydimethylsiloxanes were also reported as some of the polymer candidates for the low cost, mass production of bio-microelectromechanical system devices (Bio-MEMS) (Mata et al., 2005). Infections of silicone implant materials have been observed and reported in the literature (Wixtrom et al., 2012).
Microbial growth on plastinated specimen surfaces
In our study, no bacterial flora were detected on the surfaces of plastinated specimens. However, other authors have reported cases of fungal microflora growth observed on S10-plastinated specimens. Infections were manifested by the presence of numerous, rapidly growing, white, green, and black spots on the specimen surfaces. Similar features of fungal infections could even be observed on the wooden material of shelves used for specimen storage. Different fungal species were reported for different types of surfaces: Penicillium janthinellum was observed on kidney, cerebellum and brain stem specimens, Penicillium corylophilum was detected on abdominal sagittal section, stomach, and hand specimens; Aspergillus niger was detected on rotator cuff muscle specimens, Aspergillus flavus was detected on heart specimens, and Aspergillus fumigatus was detected on abdominal transverse section specimens. Notably, fungal flora was detected only within the superficial specimen layers. Careful analysis of deeper tissues revealed no presence of microbial cultures (Prinz et al., 1999).
Microfloral cultures on plastinated specimens: causes and prevention
Rapid increase in humidity is reported as the main cause behind plastinated specimens becoming infected with fungal microflora (Prinz et al., 1999). No precise guidelines regarding the humidity and temperature conditions for the storage of such specimens are available in the literature. At our department, plastinated specimens are stored in facilities at constant temperature of 21° C and humidity of 45%; the conditions are subject to continuous monitoring. Since no microbial growth could be observed after 5 years of storage on any of the plastinated specimens, the storage conditions can be considered optimal. In a similar manner, air-conditioned, low-humidity storage conditions have been recommended by other authors as means to prevent fungal contamination of specimens (Prinz et al., 1999). If, however, such a contamination occurs, detailed guidelines describing the procedure of eradicating the infection from the surfaces of plastinated specimens are available in the literature (Prinz et al., 1999).
In our opinion, the instruction to “store in a cool and dry place” is not enough to effectively prevent contamination of plastinates. Temperature and humidity in the storage rooms should be subject to regular monitoring.
Characteristics of microbial species detected on tested surfaces
Micrococcus
The Micrococcus species detected on the anatomical models comprise the natural microflora of skin and mucosal membranes of humans and other mammals (Carr and Kloos, l977). Together with genera Staphylococcus and Planococcus, the Micrococcus species comprise the family of Micrococcaceae which belongs to a group of 17 Gram-positive cocci (Bergey and Holt, 1994). They are aerobic, non-spore-forming bacteria; some may present with cilia (Herbert et al., 1988). They grow in the temperature range of 25°-37° C. All strains are capable of growing in the presence of 5% NaCl while some are capable of growing even in the presence of 10-15% NaCl (Bergey and Holt, 1994). Micrococcus species may pose a threat to human health only when one’s immunity is impaired. Cases of bacteremia caused by Micrococcus species have been reported as complications of immunodeficiency in some patients (von Eiff et al., 1996).
Staphylococcus
Staphylococcus epidermidis is one of the leading species found in the microbiota of skin and mucosal membranes in humans (Scharschmidt and Fischbach, 2013). They belong to the group of coagulase-negative staphylococci (Becker et al., 2014). S. epidermidis are Gram-positive (Bojar and Holland, 2002). Commensal S. epidermidis are permanent skin residents throughout human life (Grice and Segre, 2011). In order to survive on the skin surface, S. epidermidis have developed a number of mechanisms to detect and defend against, or to bypass, the human immune system (Kocianova et al., 2005).
The human body is known to provide habitats for different bacterial strains in different body compartments (Costello et al., 2009). When in the dissecting room, students come in contact with many surfaces, potentially leaving behind bacteria typically dwelling on the skin of their hands. When microorganisms are transferred from a human body onto a man-made surface, their presence on that surface is strongly dependent on the contact with humans (Davis et al., 2012). However, atypical strains may also be transferred from man-made surfaces into human systems (Ferier et al., 2010).
Considering the characteristics of the bacteria of Micrococcus species, one may conclude that their presence on the surfaces of training room equipment, bone specimens, and anatomical models is closely correlated with these surfaces coming into contact with human skin. Since the swab collections were taken immediately after completion of classes, and before the rooms were cleaned, the counts of bacteria present on the surfaces were high enough to be detected by the test method.
In the case of S. epidermidis, the structure of the bacterial cell wall, consisting of multiple layers of peptidoglycan (murein) ensures high cell stability so that the bacteria are resistant against desiccation, osmotic shock, and mechanical factors (Bojar and Holland, 2002). Thus, they are capable of surviving for significant periods after being transferred from hands onto external surfaces, allowing us to detect their presence on the door handle and the table top. Other authors have reported on a similar mechanism responsible for transmission of various microfloral species to and from man-made surfaces in closed facilities. For example, Meadow et al. (2014) reported that numerous Lactobacillus strains typical for intestines and vagina were detected on chairs in the lecture rooms of the University of Oregon, USA. Streptococcus species typical for human skin and oral cavity, as well as some Streptococci observed in humans with certain pathological conditions were detected on lecture room desktops. Bacteria typical for human skin were also detected on the room floor, in addition to other species which are typically present in soil rather than human bodies. Sphingomonas and Alicyclobacillus species were detected on lecture room walls (Meadow et al., 2014).
Other authors who studied the microflora present on the desktops at Connecticut schools, grades 7 to 12, observed an absolute prevalence of bacteria and fungi from the genera Streptococcus (≥37%) and Candida (≥ 38%), respectively (Kwan et al., 2018).
Microbial analysis of air within the Louvre Museum in Paris, revealed the presence of 103/104 Escherichia coli/Aspergillus fumigatus genome equivalents per m3 (Gaüzere et al., 2014).
