Detection of Anaerobic Beer Spoilers

Why is the detection of anaerobic beer spoilers so hard?

Anaerobic beer spoilers only became an issue since the technology of beer production was improved in concern of avoiding entry of oxygen into the production process.

The first anaerobic beer spoiling Pectinatus strains were isolated in 1978, Megasphaera was the first time detected in beer and described as a new species in 1986..

Both are responsible for bad smell and extreme off-flavor in beer. They are usually detected in finished products or other liquid samples, most often in low or non-alcohol beers.

Samples contaminated with anaerobic bacteria are often described with terms such as “animal waste”, “sewage” or “baby’s vomit”. But odor analysis alone is not sufficient to identify these anaerobic bacteria – we have also detected Pectinatus in spoilt non-alcohol beer showing no unusual odor at all.

In some cases we have identified Clostridium species as sources of bad smell, so Clostridium butyricum and Clostridium beijerinckii (formerly Clostridium acetobutylicum) from beer, and Clostridium pasteurianum from banana juice. Clostridia are spore formers and therefore resistant to high temperatures which makes them causing severe and long lasting problems.

During routine process control, anaerobic bacteria mostly stay undiscovered, therefore backtracing to their contamination source is difficult to impossible

The detection of anaerobic beer spoiling bacteria usually fails due to false sample handling.

Indeed, microorganisms in a sample often are exposed to a considerable dose of oxygen already during sampling. Often it takes hours from sampling until reaching the anaerobic atmosphere in the laboratory. During this time, the samples and the contained microorganisms are exposed to oxygen from the atmosphere.

Surfaces are sampled with swabs, the swabs are put into reagent tubes and are often filled up with medium only when they are back in the lab.

A common practice for detection of beer spoilers in low concentrations and large volumes is the anaerobic incubation of membrane filters. Clear samples are brought into the lab where they are membrane filtered under normal conditions – which means in an aerobic atmosphere. The filter is then placed into an Agar plate and later incubated in an anaerobic atmosphere.

The described sample processing can only detect such microorganisms which are able to withstand oxygen for a prolonged time and which also are able to proliferate in anaerobic conditions. In a brewery environment, these are Lactobacillus species and the anaerobic yeasts. The strict anaerobic bacteria as Megasphaera or Pectinatus are usually NOT detectable by such methods, at least not when they are present only in low concentrations.

When incubating filtered samples on Agar plates, you also should keep in mind that a high amount of cells does not at all grow up to a visible colony size on membrane filters. Our own tests comparing the growth of yeast cells on agar plates with liquid cultures lead us to a non-growth rate of up to 25% on plates compared to liquid enrichment. This means that only 75% of the present yeast cells did lead to visible and countable colonies on Agar after proper incubation. On membrane filters, the survival and growth rates are even worse than after plating direct on Agar. Therefore the detection limit for microorganisms is worst when using the method membrane filtration followed by incubation of the filter on an Agar plate.

When using the above described methods which are routine in most labs, many anaerobic microorganisms are killed by oxygen from the air before their incubation in the anaerobic atmosphere even starts.

These analyses are therefore not successful in most cases and for the lab do mean only work in vain.

You find our proposed Best Practice method for the detection of anaerobic beer spoilers here.