Introducing Lactobacillus part 2: Probiotics, fermentation, and the microbiome

LAB as probiotics

Lactobacillus bacteria are part of the Lactic Acid Bacteria (LAB) group – a mixed group of lactic acid-producing bacteria which require a fermentable carbohydrate for growth.

LAB are very well known amongst lovers of yoghurt and probiotic devotees. Long before the gut and skin microbiome were considered as a target for food and cosmetics, people enjoyed eating live yoghurt and using it on their bodies and faces to moisten and brighten skin, (also using live yoghurt to treat conditions such as vaginal thrush).

Traditional uses of live yoghurt, along with our increased understanding of the human microbiome, has fuelled interest in live skin care products. Shelf-life and challenge testing for product safety and regulation, all provide barriers making it difficult  for commercial brands to utilise live microbes in their cosmetics.

That said, some pioneering companies, such as Mother Dirt, are selling live skin care. While not using LAB, Mother Dirt’s website states there are 3+ billion colony forming units (CFUs) of ammonia-oxidizing bacteria (AOB) in every bottle.

Lactobacilli, meanwhile, are being formulated in encapsulated forms into anhydrous serums, and micro-encapsulated freeze-dried forms for use in oil-and-water cosmetic creams [1]. Some cosmetic developers are using LAB strains isolated from skin. However, because probiotics used in foods are well known and easily sourced, the LAB species associated with probiotic types of cosmetics may not necessarily be the strains found in human skin microbiota. More commonly, in place of living microbes, probiotic cosmetics list ingredients as a genus name such as Lactobacillus, Bacillus, Bifidobacterium, Streptococcus or Micrococcus followed by term such as ferment, lysate, filtrate, extract etc.

LAB and fermentation

Many popular fermented foods are made using LAB, either on their own or with other microbes:

Yoghurt contains the LAB species Lactobacillus delbrueckii subsp. bulgaricus which works together with Streptococcus thermophilus bacteria to convert milk into the final product. Other lactobacilli and bifidobacteria spp. may be found in some yoghurts.

The mildly fizzy alcoholic beverage, Kumis, is made by fermenting while churning raw unpasteurized mare’s milk with LAB. Similarly, (but without churning) Kefir is made by fermenting cow or goat milk with LAB and yeast.

Moving away from dairy products, sauerkraut is LAB-fermented finely shredded cabbage. Similarly, Korean kimchi is cabbage (or other vegetables) fermented normally in the presence of fish sauce, by LAB (including Lactobacillus kimchii, Leuconostoc mesenteroides, Leuconostoc pseudomesenteroides, Leuconostoc lactis, Lactobacillus brevis, and Lactobacillus plantarum (now classified as Lactiplantibacillus plantarum). Interestingly, Latilactobacillus sakei (mentioned in part 1, because some strains produce bacteriocins), is the dominant LAB in over-ripened kimchi [2].

Natural pickles also depend on LAB. In the case of pickles, it is the LAB in the natural microflora of whatever is being pickled, i.e., the fruit, vegetable, etc., that ensures the sugars leached out by the brine, are fermented through to acid. The combination of salt and acid prevents spoilage microbes (including fungi) and pathogens from growing, and so preserves the pickled food for safe consumption.

In the battle for supremacy amongst microbes, LAB create pHs that only they and other acidophiles can enjoy. Lactobacillus spp. are also used in meat and meat products. For example, several species of Lactobacillus are added as starter cultures in salami production. They also act as competitive microbes in the microbiota in meat products, extending their shelf life by competing with pathogens.

On the negative side, Lactobacillus spp. can cause souring and colour changes in meat. At pH values around 5.0, the growth of heterofermentative Lactobacillus spp. dominate. Souring at this pH is due to the production of organic acids, such as acetic acid, along with lactic acid, as well as the production of gas by heterofermentative spp. Some Lactobacillus spp. are able to form hydrogen peroxide (H2O2) in the presence of oxygen, which can cause meat to appear grey-green, yellow, or white due to myoglobin becoming oxidised.

Fermentation, whether by accident or by design, can make food and drink more pleasurable. This is illustrated nicely by chocolate, which relies on natural indigenous microbes, including LAB, to develop its flavour. This occurs when the contents of the freshly opened Theobroma Cacao pods are spilled out and left to ferment naturally.

Fermentation can also release nutrients from foods. The wild fermentation stage leading to the mildly alcoholic Tanzanian drink ‘Mbege’, (made from bananas, millet, (Eleusine coracana) and sometimes quinine bark), releases nutrients, making the millet more nutritious and much more palatable [3].  

Extending storage and improving flavour has had humans intentionally fermenting foods since at least Neolithic times. The ancient practice of using some of a previous batch as the ‘starter culture’ for subsequent batches, continues today. For example, although traditional sourdough bread can be made spontaneously using the natural LAB microbes in flour, sourdough bread made with ‘starter culture’ (retained from previous batches) is preferred because it results in sourdough bread with more predictable qualities.

The commercial controlled fermentation for wine, yogurt, cheese, sauerkraut, pickles, beer, cider, kimchi, kefir, and other fermented foods begins with a starter culture. The selective pressures of passaging of microbes through countless generations, to fermenting the food substrates, has led to certain LAB species such as the already mentioned Lactobacillus kimchii, being associated with a particular fermented food or drink. In this way, humans have favoured species of LAB in much the same way as they favoured and domesticated cereals and livestock.

LAB in the gut microbiome

The human gut microbiota is naturally transmitted from mother to infant through gestation, vaginal birth and breastmilk. 

The infant gut of breast-fed babies is first dominated by varieties of Bifidobacterium, that metabolise human milk oligosaccharides, whereas formula-fed infants begin with a more diverse gut community [4]. By around three weeks, the infant gut is colonised and, regardless of mode of birth, the diversity of the gut microbiota gradually increases.

As LAB-fermented foods have been consumed by young and old since pre-history and LAB have been found in breast milk [5], it is unsurprising to find LAB species amongst the prominent gut microbes. Firmicutes (such as Clostridium, Enterococcus, Lactobacillus, and Ruminococcus), Bacteroidetes (such as Bacteroides and Prevotella) and, to a lesser extent Proteobacteria and Actinobacteria dominate the gut microbiome.

The populations of microbes in the established adult gut are influenced by local conditions so they vary according to the region of the gut, and their numbers normally ebb and flow in harmony with changes in the diet. However, whether ingested LAB succeed in becoming members of the gut microbiome is still contentious.

Pasolli, et al., [6] have recently shown that there are generally low numbers of LAB species in adult stool samples (indicating that the levels of LAB species in the gut microbiota are low) and the amount of LAB in stool samples is influenced by age, lifestyle, and location. The most common gut LAB are Streptococcus thermophilus and Lactococcus lactis, both of which are also found in breast milk, as well as in fermented dairy products.

In the 2017 Scientific America article, ‘Do Probiotics Really Work?’, the author suggests that strains of Bifidobacterium spp. or Lactobacillus spp. found in many yogurts and probiotic products may not be the same kind that can survive the highly acidic human stomach and go on to colonize the gut [5]. In contrast, the work by Pasolli, et al., [6] supports the claim that ingested LAB can survive to colonise the gut. Their recent genome-based study of the differences between food and gut LAB microbes linked LAB from food with LAB living in the gut microbiome. They showed that some LAB strains in fermented foods are closely related to LAB strains in the gut, strongly suggesting that these strains of LAB did survive the highly acidic human stomach to colonize the gut. In the doubting, 2017 Scientific America article, it was reported that out of seven randomized placebo-controlled trials involving probiotics, just one study found a statistically significant change in the levels of Bifidobacterium longum in faecal samples. This study had 34 healthy volunteers consuming daily for two weeks, either a drink containing 10 billion live Bifidobacterium longum bacteria or a placebo of maltodextrin. In 7 subjects, these significant changes were still detectable 5 months after the study finished – indicating that live Bifidobacterium longum bacteria from the probiotic containing food, had become established in the gut.

Lactobacillales in the skin microbiome

A recent skin microbiome study showed Lactobacillales microbes were present at significant amounts in cheek samples. However, when looking deeper at the genus level, the relative abundance of Lactobacillus fell to much smaller concentrations. Species identified included Lactiplantibacillus plantarum and Latilactobacillus sakei.

Lactiplantibacillus plantarum is able to use a range of carbon sources so adapts more readily to different environments. As it happily grows at skin pH and at body temperature, preferring to replicate in aerobic conditions, its presence in the skin is not unexpected though it is more commonly found in saliva and in the gut. Latilactobacillus sakei produce sakacin P – which inhibits growth of several pathogenic bacteria – so it may contribute in protecting the skin from infection.

Conclusion

The multitude of different Lactic acid producing bacteria means taxonomy is complicated but  without doubt, their usefulness for preserving and improving food quality, is unquestioned. Although only present in small numbers, LAB play an important role in the secret life of skin. They help control local pH, produce biofilms and for block sites where invading microbes would like to adhere. They can also modulate the host’s immune response making LAB fermented foods especially interesting as probiotics. Bacteriocins (antimicrobial proteins) produced by some strains of LAB play a part in the germ-warfare, which is constantly played out by skin and gut microbiota. Commercially, LAB bacteriocins such as Nisin are used in foods and can be used in cosmetics. These bacteria have accompanied humans for mutual benefit for eons (even into outer space). Because they are versatile and easy to culture, they are part of biotechnologies’ biological tool kit and so will be our companions long into the future.

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References

‘Revealing the secret life of skin ‐ with the microbiome you never walk alone.’

International Journal of Cosmetic Science. Volume42, Issue2. p116-126. https://doi.org/10.1111/ics.12594

10.1038/scientificamerican0717-26