Over the past decade, research has made it clear that having a balanced and diverse skin microbiota is essential for skin health and appearance. Many products available on the market have microbiome-friendly claims, or aim to address imbalances in the skin microbiome. Cosmetic microbiology testing is instrumental in substantiating such claims.

In this article, we will examine the role of the human skin microbiota, and various tests used by contract research organizations to substantiate microbiota-friendly, anti-microbial and other microbiome related product claims.

The skin is the largest organ of the human body, accounting for around 15% of the total adult body weight. It is composed of 3 distinct layers: the epidermis, the dermis, and subcutaneous tissues. Together, these layers carry out several vital functions that include thermoregulation, prevention of excess water loss, and protection against external threats.

In addition, recent published data shows that the surface of the skin may also provide an optimal environment for the growth of microorganisms. Collectively, they comprise the skin microbiotame, which is intricately linked to human health and disease.

The skin microbiota is generally divided into two groups: resident and transient microorganisms.

• Resident microbes, also known as commensal microbes, are part of a precise group of microorganisms routinely located on the skin surface. They are often considered harmless, and even offer beneficial effects to the host. Commensal microorganisms often protect the human host against invading pathogens, either through direct antimicrobial activity, or by triggering protective immune responses in the skin.

• Transient microbes, on the other hand, are generally harmful and pathogenic. They usually colonize the skin after injury or trauma, and may lead to infections and inflammation.

Under normal homeostatic conditions with proper hygiene, intact skin barrier function and normal resident flora, the skin microbiota is healthy. On the other hand, any imbalance in the microbiota may lead to skin diseases.

The skin surface hosts a diverse population of microbes, and different anatomical areas are home to different bacterial communities. Depending on the temperature, moisture and lipid content of the skin, three main skin environments can be defined:

• Dry areas: arms, palms, hands;

• Moist areas: armpits, between the fingers, behind the knees (popliteal fossa), the groin (inguinal folds), navel, etc.;

• Sebaceous areas: face, cheeks, external auditory canal, back of the scalp, upper chest and back.

While microorganisms were initially considered harmful to the skin, research today suggests that a proper balance of the skin microbiome is required to maintain skin health. Different products and methods are constantly introduced to the market to assist with maintaining a balanced skin microbiome. Consequently, research labs have developed a number of microbiome related tests to guide product research and confirm product claims.

Tests to check whether a substance is “microbiome-friendly” allow to substantiate manufacturer claims that a compound or formulation does not create an imbalance in the bacterial composition of the skin microbiome. When putting their products on the market, brands want to make sure that their formulas will have the potential expected effect, while not causing a disruption or dysbiosis.

Tests offered to substantiate microbiome-friendly claims include assays that measure the effect of tested compounds on bacterial growth. Such effect can be stimulating (promoting bacterial growth), bacteriostatic (stopping bacteria from reproducing) or bactericidal (killing bacteria). The viability assay shows the number of viable cells after exposure to the compound over a period of time.

Such assays can be carried out on single bacterial strains or on a bacterial mix. For example, a mix of Staphylococcus aureus, Staphylococcus epidermidis, Cutibacterium acnes and Corynebacterium xerosis is considered representative of the skin microbiome. The bacterial mix can be amended to suit specific product research needs. If the experiment also calls for quantifying specific bacterial strains in the mix, conventional microbiological tests can be used to provide that additional information.

In addition to these conventional methods, there are modern microbiome assays using 3D skin models, such as Reconstructed Human Epidermis (RHE). Research labs with expertise in tissue engineering can offer to produce customize RHE in-house for product efficacy and “microbiote-friendly” testing.

Microbiome tests using reconstructed human epidermis can measure the effect of tested products on bacterial adhesion to the skin model. As with other assays, these tests can be performed with single bacterial strains or bacterial mixes. In addition to the quantitative analysis of bacterial adhesion, electronic microscopy can also be used to provide qualitative data.

As we described earlier, the skin, while being an intricate home for many microorganisms, also has a major role in detecting invading pathogens. Indeed, under certain conditions that lead to dysbiosis or a downregulation in the immune response, microorganisms are able to circumvent normal host defense mechanism and become pathogenic. In that case, certain components of the skin (keratinocytes, fibroblasts and sebocytes) act as responders against pathogenic microorganisms and trigger cutaneous innate immune response.

Such immune response can present as acne or atopic dermatitis. Products used to treat these conditions may have anti-microbial and anti-inflammatory claims. To substantiate them, researchers design assays to evaluate the potential anti-inflammatory effect of some compounds in keratinocytes, fibroblasts or sebocytes infected by certain bacteria (e.g. Staphylococcus aureus, Staphylococcus epidermidis, Cutibacterium acnes and Pseudomonas Aeroginosa).

Moreover, using RT-qPCR technology (quantitative reverse transcription polymerase chain reaction), the effects of the tested compounds may also be analyzed using a dedicated PCR array, whereby genes have been selected for their importance in anti-bacterial activity and inflammation.  Such assays can be designed for 2D and 3D models.

In this same context, contract research organizations are looking deeper into certain commensal yeasts of the skin microbiome. These include the Malassezia species, lipid-dependent microorganisms that can contribute to seborrheic dermatitis, dandruff formation, rosacea, odor and other conditions. Understanding the lipid requirements of the Malassezia species in these pathogenic states may provide insights into their biology and enable customers to test compounds with potential anti-Malasezia fungi effect.