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A Probiotic is More Than Just a High Cell Count

At first glance, developing a probiotic product seems straightforward. Suitable bacterial strains are selected, cultured, combined, and formulated into a dosage form. The product is then tested to ensure it contains the required number of viable microorganisms.

In practice, however, this process is significantly more complex.

Probiotic bacteria differ in their growth conditions, stability, sensitivity to oxygen, metabolic activity, and behavior in mixed cultures. They are affected by production conditions, drying, storage, packaging, and the composition of the product matrix.

That is why reliable analysis does not begin with the finished product; it should be incorporated as early as the development stage.

vermicon supports probiotic manufacturers, pharmaceutical companies, CDMOs, and research institutions throughout the entire development process. The range of services extends from the identification and characterization of individual bacteria, through the analysis of complex mixtures, to stability testing and routine quality control.

Unambiguously Identifying The Right Bacteria

At the start of any product development, the question arises: Which microorganisms will actually be used?

That may sound obvious. In practice, however, closely related bacterial species are difficult to distinguish from one another using traditional microbiological methods. Identification becomes particularly challenging with complex mixtures or with bacteria that are difficult to culture under the selected conditions.

A non-specific total cell count cannotanswer this question. It merely indicates how many cells or particles are detected; it does not reveal which species are actually present.

vermicon therefore uses specific molecular biological methods. With fluorescence in situ hybridization(FISH), bacteria can be directly identified based on characteristic regions of their ribosomal RNA. The target cells are made visible using fluorescently labeled gene probes.

This allows individual species or defined groups of bacteria to be identified even within complex probiotic mixtures.

This provides an important foundation for development. After all, only when the composition of a mixture is known with certainty can subsequent changes be reliably evaluated.

Developing Targeted Mixed Cultures

Many probiotics consist not of a single strain, but of several bacterial species. Simply mixing the desired microorganisms together is not sufficient.

The individual populations can develop differently during production and storage. One species may be particularly stable, while another may lose viability more quickly. Some bacteria are more sensitive to oxygen, moisture, or temperature. Others may be protected or harmed by components of the matrix.

Analytically separating the individual species is also challenging. Cultivation methods can reach their limits when different strains exhibit similar colony morphologies or have different growth requirements. While non-specific flow cytometry quickly provides a total cell count, it does not automatically distinguish between the bacterial species present.

The Flow VIT® technology developed by vermicon combines FISH with flow cytometry. This allows viable cells to be specifically identified and quantified. Even multiple target populations within a complex mixture can be detected separately.

For product development, this means that the manufacturer not only sees the total number of cells present but also recognizes how the individual components of a formulation behave.

Viability is Not The Same as Cultivability

The traditional method for determining probiotic cell counts is often based on cultivation. This method measures how many bacteria can form colonies under defined conditions.

This method is well-established but does not always fully reflect the biological state of a population.

Bacteria can be viable without immediately forming a colony under the selected laboratory conditions. Conversely, aggregations can result in multiple cells being counted as a single colony-forming unit. In mixed products, there is the additional challenge that individual species cannot always be reliably distinguished from one another.

A more nuanced approach is therefore advisable for the development of a probiotic.

Stability Must Be Tested for Each Species

A probiotic must remain effective not only on the day of manufacture. The declared bacteria must be maintained in sufficient numbers and in a suitable physiological state throughout the intended shelf life.

For a multi-species product, a total cell count alone is not sufficient for this purpose.

A stable total count can mask the fact that a robust species remains virtually unchanged, while a more sensitive target species declines sharply. The total count still appears plausible, even though the product’s composition has changed significantly.

The specific quantification of individual species therefore enables a significantly more precise stability assessment.

vermicon can assist manufacturers in planning and conducting stability studies. This allows for the investigation of how individual target populations develop under various storage conditions, over different time periods, and in different product matrices. Viability can be tracked throughout the entire shelf life, and the composition of the final product can be validated.

Such data not only aids in quality control; it can also indicate as early as the development phase whether a formulation, manufacturing process, or packaging should be adjusted.

The Product Matrix Plays a Key Role

Probiotic bacteria are rarely found in an analytically simple environment. Capsules, powders, foods, suspensions, or pharmaceutical formulations contain excipients, proteins, fats, plant components, or other particles.

This matrix can influence the measurement. It can generate fluorescent background signals, bind cells, complicate sample preparation, or interfere with specific detection methods.

A method that works with a pure culture is therefore not automatically suitable for the finished product.

vermicon develops and optimizes sample preparation procedures and detection protocols for specific products and matrices. The goal is not a theoretically viable analysis, but a robust method that delivers reproducible results under real-world conditions.

This methodological adaptation is a particularly important part of product development, especially for new formulations.

Examining Bacteria Directly in Their Spatial Structure

Sequencing methods provide valuable information about the composition of microbial communities. However, this process involves extracting the biological material, resulting in the loss of the original spatial organization.

FISH offers a different approach. The bacteria are visualized directly within the sample. This makes it possible to determine whether target organisms are present individually, form aggregates, adhere to surfaces, or are spatially associated with other populations.

This information can be relevant in the development of probiotic products. For example, cell aggregates can influence cultivation and cell count determination. The distribution of bacteria in complex matrices or model systems can also be directly examined.

vermicon thus combines molecular identification with visualization. The bacteria are not only detected via an extracted signal but are also observed in their actual microbiological context.

Detecting Contamination Early

During the development and production of probiotic products, it is not enough to simply verify the presence of the desired bacteria. It is equally important to determine whether undesirable microorganisms have entered the process.

Contamination can originate from raw materials, production equipment, the environment, packaging, or the manufacturing process. It can alter stability, compromise the reliability of cell count determinations, or jeopardize product quality.

vermicon therefore also offers specific tests for monitoring microbial quality and controlling contamination. In the event of unusual findings, microorganisms can be identified and potential causes narrowed down.

The advantage of specific analysis is that it goes beyond simply detecting a nonspecific increase in bacterial count. It allows for a targeted investigation to determine which organism is responsible for the change.

From Outsourced Services to In-House Routine Analysis

Not every manufacturer needs the same type of support.

In an early development phase, it may make sense to first have specific issues addressed as a laboratory project at vermicon. This may include feasibility studies, method development, formulation comparisons, stability studies, or the characterization of complex mixtures.

Once a product enters routine production, the analytical work can be transferred to the company’s own laboratory.

Standardized Flow VIT® test kits are available for numerous Lactobacillus and Bifidobacterium species, as well as for Streptococcus thermophilus. The kits are compatible with common flow cytometers and can be integrated into existing laboratory workflows. The workflow is designed to reduce manual effort and ensure reproducible results.

This allows vermicon to serve both as a development partner and as a provider of a long-term in-house analytical solution.

Customized Solutions for New Strains and New Products

The probiotics market continues to evolve. New strains, more complex combinations, new dosage forms, and live biotherapeutics present new challenges for analytical testing.

Standard methods do not always address these issues.

vermicon therefore also develops customer-specific detection solutions. New gene probes can be designed for specific target organisms, tested for specificity, and adapted to the respective sample matrix. Methods can be optimized for individual products and integrated into existing development or quality processes.

This is particularly relevant when a manufacturer is developing a new strain, investigating a specific combination of bacteria, or when existing methods fail to yield clear results.

Quality Control is The Result of Good Development

Of course, quality control remains a central component of probiotic manufacturing. Manufacturers must ensure that their products have the intended composition, contain a sufficient number of viable bacteria, and remain stable throughout their shelf life.

However, robust quality control does not begin only at the end of production.

It begins with the unambiguous identification of the microorganisms used. It continues with the development of suitable detection methods, the analysis of mixed cultures, the assessment of viability and activity, and the analysis of matrix effects and stability.

This is precisely where vermicon’s more comprehensive approach comes into play.

The company does not merely provide individual test results. It helps manufacturers gain a better microbiological understanding of their bacteria, processes, and products. With nearly 30 years of experience, in-house technology development, and high-performance services, vermicon supports projects from the initial research phase through routine product monitoring.

Anyone developing a probiotic product should therefore not wait until the first batch has been produced to discuss analytical testing.

The sooner the right microbiological questions are asked, the more effectively development risks can be identified, formulations optimized, and reliable testing procedures established.

In this process, vermicon is not just a quality control laboratory, but a microbiological development partner.