Environmental microbiology: Perspectives for legal and occupational medicine
Introduction
Environmental matrices (water, soil and air) are characterized by the presence of microorganisms, living beings with size ranging from few millimeters to less than 100 nm. This biological component is rarely organized as single species populations but can be described as a network of microbiological groups, with specific biodiversity, interacting each other in a never-ending fight for nutrients [1]. These biological interactions get involved bacteria, protozoa, fungi, plants/algae and metazoa, in addition to the specific viral particles (as obligate intracellular parasites). Only considering bacteria, it has been estimated that few grams of soil are likely to harbor many tens of thousands of different species [2]. Between eukaryotes, fungi represent one of the largest and most diversified kingdom: they strongly contribute the ecological networks and are reactive to environmental modifications [3], [4]. Similar considerations can be extended to other eukaryotic microorganisms present in the soil [5]. Also water samples have a strong biodiversity, both at a prokaryotic and eukaryotic level, with the presence of beta diversity fluctuations linked to environmental mutations [6], [7], [8], [9]. Considering the air matrix, specific microbiotas characterize indoor environments in strictly correlation with growing conditions found on solid surfaces and environmental determinants [10], [11].
Microbiological populations colonize not only the external environment but also human body districts that correspond to highly stable ecological niches. Studies of human microbiome have revealed that individuals harbor a great variety of microorganism collections in and on their bodies [12]. It is estimated that the number of bacteria in the body is actually of the same magnitude as the number of human cells [13]. The human body is an ecosystem made up of complex microorganism communities that inhabit the hairs, nostrils and respiratory tracts, urogenital and gastro-intestinal tracts, skin and other districts. There is endogenous or resident microflora, that resides over and in the body permanently and exogenous or transient microflora, which depends on the environment which an individual is exposed to. Human microbiome shows spatial and temporal complexity: it differs by individual, body site, age, geographical location, health status and lifestyle (diet, living together with other people, having pets, sexual behavior, smoking habits, kissing, etc) [14], [15], [16], [17], [18], [19], [20], [21], [22].
An example of human microflora can be found in the gastro-intestinal system. This human microbiota is characterized by several hundreds of identified species and its biodiversity is strongly connected with dietary and health conditions [23], [24]. Similarly, the oral district has a complex community of microorganisms, playing an important function in protecting against colonization of extrinsic bacteria which could affect systemic health [25], [26]. This defensive action is played at the skin and vaginal level too, by a stable microbiota where few species modify the environment for reducing the probability of infection by pathogens [27], [28].
The microbiological colonization of a difficult environment is helped by the synergic action of many different species in creating a biofilm, a surface-associated heterogeneous structure comprising different populations of microorganisms surrounded by a self-produced matrix that allows their attachment to inert or organic surfaces [29]. The biofilm stabilizes the environment, facilitating microorganism proliferation and consolidating biodiversity. Microbial species with biofilm forming capacities initialize the colonization process, which is then completed by the presence of other microrganisms, leading to a spatial structuring and a balance between cooperation and competition [30], [31], [32]. The oral microflora has been described as a community organized in a complex dental biofilm, with a biodiversity degree affected by hygienic actions or dietary modifications [33]. Bacterial composition on dental biofilm can have strong medical implication. Similarly, the biofilm formation on water environments (natural or artificial) need to be monitored in order to prevent Legionella proliferation, a bacterium associated with a severe pneumonia [34].
The consideration that biofilm represents a major mode of microbial life underlines the importance of laboratory tools able to unmask the biodiversity degree present in a specimen, and not only to study the presence of a single pathogen or biomarker. This new approach is crucial both for preventive actions in several medical fields, and for the reconstruction of environmental modifications and human manipulations. Legal and occupational medicine are both characterized by a strong interest in the analysis of environmental matrices and their microbiological component. The number of microbiological publications in occupational medicine and legal medicine in 2017 is comparable and correspond to 3–4% of total papers published in these two fields (Fig. 1). Recent evolution of molecular tools and the availability of automated procedures has widened the potentiality of microbiological analysis in environmental samples. At the same time occupational and legal medicine practitioners need validated procedures in order to effectively support events reconstruction during litigations.
The present paper presents possible applications of microorganism population analysis for legal medicine (a medical field where the identification of sample contamination and modification is crucial), and for occupational medicine (a typical preventive medical field). In the paragraph on legal medicine investigations a specific description of microbial population analysis is presented for bioterrorism, human microbiota on the crime scene, body fluid identification, microbes and geolocation, thanatomicrobiome and body decomposition. In the paragraph on occupational medicine the importance of the analysis of microbial population is highlighted through recent data health risk and organic dust, indoor microbial contamination, biofilm formation and Legionella; this latter point is crucial issues for many working environments, including hospitals. Finally, a specific description of technical tools for microbial characterization will underline how the new high throughput approaches can lead to an effective analysis of environmental biodiversity.
Section snippets
Microbiota and legal medicine investigations
When a crime is committed, it is crucial to uncover and gather all the relevant evidences to help identify what happened and who was involved. The study of microbiomes has enormous potential in legal medicine because microorganisms (predominantly bacteria, fungi, viruses and microeukarya) are ubiquitous and particular communities of microbes are often associated with specific processes or environments. Microorganisms have been recognized as physical evidence since the early 20th century,
Environmental microbiota and occupational medicine
Safety evaluation of working environment requires the identification of physical, chemical, biological and psychosocial hazards [108]. Between them, the biological hazard deserves a particular attention due to the complexity of the microorganisms that can populate working sites. Both the degree of microbiological biodiversity and the concentration of singular species are two variables that should be considered during safety assessment (Fig. 2). Complexity of interactions between multiple
Technological evolution
Since the discovery of microscopic living beings, an intense technological effort has been dedicated to the improvement of protocols for the discrimination and the identification of microorganisms. The usage of staining methods (and of liquid/solid growing media) has allowed the emergence of microbial taxonomy and more complex considerations on biodiversity [145]. At the end of 20th century, with the development of nucleic acid techniques, scientists have become able to quantify the degree of
Conclusions
Occupational and legal medicine are fields closely related: this affinity is so strong that has raised important jurisprudential questions and deserve more attention from national and international institutions [171]. At a scientific and technical level, this closeness is clearly highlighted by the common focus on environmental matrices. In particular, microbiome analysis can be a valuable approach for depicting the interactions of human being and environment. The recent availability of new
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