There are various genera of bacteria that fall under the category of probiotics - Bacillus, Lactobacillus, Paenibacillus, Enterococcus, Aeromonas, Alteromonas, Arthrobacter, Bifidobacterium, Microbacterium, Clostridium, Phaeobacter, Pseudoalteromonas, Pseudomonas, Rhodosporidium, Roseobacter, Streptomyces and Vibrio. But the most commonly used across human and animal applications is genus Lactobacillus.

Figure 1. Cheese consumption in the 11th century. Source: Tacuinum Sanitatis, a medieval handbook on health and well-being.

Lactobacillus is a genus that includes Gram-positive, rod-shaped bacteria that are most commonly known for their role in carbohydrate fermentation and the ability to produce lactic acid as a result of this process. Lactobacillus species don’t have the ability to form endospores like their cousin Bacillus. For this reason, they are rarely found as free-living bacteria. These microorganisms first gained popularity as cultures necessary for producing fermented milk products, like cheese and yogurt, and as probiotic bacteria for humans.

The gut microbiome in humans and other animals has been gaining increasing attention in the last couple of decades due to its significant effect on health. A variety of species of gut bacteria directly and indirectly affect early development, the ability to process certain foods, immune response, hormonal balance, weight, mood, and a wide variety of other processes in the body. As our knowledge of probiotic organisms has increased, so has our use of these probiotics in areas outside the human health and milk industry.

Today, these bacteria are also widely used in agriculture and aquaculture and play a very important role in animal health and sustainable production of food. Aside from Bacillus, Lactobacillus species are the most studied and widely used probiotic bacteria mainly due to their great effectiveness in promoting natural digestive and immune pathways in animals.

From Discovery to Today

Using microorganisms for health benefits has a long history – one of the first reported cases goes back to the Roman historian Plinius in 76 B.C.[1], who recommended the use of fermented milk as a remedy for gastroenteritis. One of the first Lactobacilllus species that was found was L. acidophilus, which was isolated from the human digestive tract in 1900. It was first identified as Bacillus acidophilus, but after the discovery of the Lactobacillus species and novel identification methods, its taxonomy was revised and the species renamed to Lactobacillus acidophilus.

Figure 2. Scanning electron micrograph (SEM) of Lactobacillus delbrueckii subsp. bulgaricus. Source: SciencePhotoLibrary.

Studying the fermentation process for yogurt production, Bulgarian researcher Stamen Grigorov discovered Bacillus bulgaricus in 1905 (now known as Lactobacillus delbrueckii subsp. bulgaricus). Inspired by his research, Ilya Metchnikoff, Russian zoologist, bacteriologist and immunologist, published a paper two years later in which he declared that one of the main reasons for the longevity of Bulgarian farmers was due to their daily consumption of yogurt. He suggested that cultures present in Bulgarian yoghurt may have the ability to inhibit the aging process caused by putrefactive bacteria in the colon.

The genus Lactobacillus was officially discovered in 1921 by American bacteriologists E.B. Fred, W.H. Peterson, and J.A. Anderson. The genus was characterized by the ability to metabolize certain sugars and other carbon-based compounds. It was very hard to properly determine the exact species before PCR (Polymerase Chain Reaction) was discovered in 1983, so many different Lactobacillus species were put under the same name. With the discovery of new gene sequencing techniques, a whole new world of Lactobacillus species was discovered and their application became more specialized and efficient.

Lactobacillus in aquaculture

Figure 3. Aside from increasing the danger of antibiotic-resistant bacteria, antibiotic application also depletes the gut microbiota in aquaculture species. Source: The Rise and Fall of Antibiotics in Aquaculture, Microbial Communities in Aquacultur… — Figure 3. Aside from increasing the danger of antibiotic-resistant bacteria, antibiotic application also depletes the gut microbiota in aquaculture species. Source: The Rise and Fall of Antibiotics in Aquaculture, Microbial Communities in Aquaculture Ecosystems.

The aquaculture industry has significantly expanded during the last two decades, and it is predicted that it will continue to grow in the coming years. In order to meet the increasing demands of the market, the aquaculture industry has started to rely on high stocking densities and aggressive feed input. These practices often result in a lot of accumulated waste, poor water quality, and increased stress levels in animals, creating a very favorable environment for pathogens to spread rapidly. In order to prevent the incidence of diseases and huge economic losses that go with them, aquaculture producers have used a number of techniques, including (until recently) antibiotic usage. Many countries have recognized the negative effects of antibiotics and banned their use in aquaculture, and many others are following their footsteps.

However, the ban of antibiotics leaves a very big gap in the toolbox of aquaculture producers, now in need of an alternative that will be at least as effective as antibiotics without the negative effects.

For this reason, many scientists and aquaculture experts have turned to probiotics.

There are many genera of probiotic bacteria, and they include a variety of specific and non-specific species. Lactobacillus is one of the most widely represented genera in animal health applications because of the many beneficial effects they have on animal production.

**How do Lactobacillus species improve the health of fish and crustaceans?**

Just like other probiotics, Lactobacillus species support beneficial effects on aquaculture species in a number of ways, both directly and indirectly. The six main functions of Lactobacillus species in aquaculture include:

Stimulation of gastrointestinal tract (GI) development - Lactobacillus bacteria can colonize the gut of fish and crustaceans at early phases of development (larval stage) and ensure their proper growth by preventing the establishment of potentially harmful bacteria. This is very important, as pathogen infections in larvae are often detrimental to their development and result in permanent damage, as well as weakened immune system. Lactobacillus species also aid nutrient absorption, increasing feed efficiency and availability of necessary elements for the development of larvae and young fish and crustaceans.
Improvement of digestive function and weight gain - Lactobacillus species increase utilization of carbon sources with the help of their digestive enzymes, which increase nutrient availability, as well as feed to yield conversion. These bacteria also increase the tightness of intestinal mucosa and stimulate the activity of gut wall cells, so they improve the natural processes within their host’s gut. This results in better feed utilization, increased growth rates, survival rates and weight gain in fish and shrimps.
Stimulation of the host’s immune response - Probiotic bacteria like Lactobacillus were shown to have a beneficial effect on host’s immune system by increasing the total number of lymphocytes, as well as activity of lysozymes and other enzymes important for immune defense. They can stimulate non-specific immune responses and increase activity of the host’s monocytes, macrophages, natural killer cells, and other agents that deal with unwanted residents of the gut microbiome. For example, L. rhamnosus has the ability to increase respiratory burst in rainbow trout, which means that it helps the fish to quickly release reactive oxygen compounds like hydrogen peroxide and superoxide anion. These compounds play an important role in cell signaling and defense against pathogens.
Increasing reproduction - Reproductive capacity of aquaculture species is heavily influenced by the amount of available nutrients – fats, proteins, vitamins, fatty acids, etc. These elements are necessary for every part of the reproduction process, and proper development of larvae. Lactobacillus probiotics can increase the fecundity of fish by synthesizing some of those necessary elements we mentioned (like vitamin B complex), ensuring better carbon source utilization, and stimulating natural immunity of aquaculture species. For example, researchers studied a probiotic which contains Lactobacillus acidophilus, L. casei, Bifidobacterium thermophilum, and Enterococcus faescium, and its effect on fecundity of female green swordtail. The results were clear – the number of fecundated fish increased almost 50% in the group that received probiotics compared to the number of fecundated fish in control (fish that weren’t treated with the probiotic product).
Protection from pathogenic bacteria - The occurrence of diseases is very high in intensive agriculture and causes serious economic losses. To battle this issue, many aquaculture producers previously relied on antibiotics, but due to the great danger of antibiotic-resistant bacteria, their use became severely restricted in many countries. Probiotics show great potential to completely fill this gap and provide a more sustainable way of getting high yields and producing healthy aquatic animals, without increasing the danger of microbial resistance to antibiotics and harming the environment.
GI is one of the most common places in animal bodies for establishment of pathogenic bacteria. Probiotics have the ability to colonize the gut quickly and thus reduce the surface area where pathogens can adhere and establish their colonies. By taking up space and nutrients from harmful bacteria, Lactobacillus species can outcompete them and thus prevent disease development. Applying probiotics with Lactobacillus species was shown to modulate the gut microbiome and stimulate the activity of enzymes important for immune responses. Some Lactobacillus species can help increase immunity to specific pathogens, for example, L. plantarum increases disease resistance to Vibrio harveyi in Pacific white shrimps, while L. acidophilus increases defensive responses of giant fresh river prawn against pathogenic Vibrio species.
Improving water quality - Water quality is one of the key factors for healthy and successful production in aquaculture. Accumulation of organic waste, ammonia, nitrites and nitrates is a constant problem, and a high concentration of these compounds in water can cause great damage to fish and crustaceans, and increase their mortality. Simply changing the contaminated water with fresh water can be costly in some areas, and also have a negative effect on the environment, considering that the waste water is often released without processing into streams, rivers, and seas. This is why sustainable producers opt for bioremediation, and probiotic bacteria are used to kick-start this process. These bacteria are live organisms that have the ability to produce enzymes and metabolize harmful compounds, and as such can provide beneficial and long-lasting effects on water quality and animal health. Aside from the famous bioremediatory species like Bacillus, Nitrosomonas and Nitrobacter, certain Lactobacillus species like L. plantarum and L. casei also show the ability to reduce the amount of harmful waste in the water and improve growth of shrimps.

Most aquaculture probiotics are comprised of a blend of species to achieve the best effects; as a result, Lactobacillus species are found in a combination with Bacillus, Enterococcus, Lactococcus, Nitrobacter and other probiotic bacteria. The combination of species and their individual quantity in the product depends on the aquaculture species and desired effects.

Conclusion

Many members of genus Lactobacillus provide various benefits to aquaculture species naturally – from increasing nutrient availability and stimulating digestive functions, to strengthening immune responses and fighting off pathogens. Besides keeping fish and shellfish healthy and well-nourished, these incredible bacteria can also increase their fecundity and survival rates of larvae. Their most prominent advantage is that they are live organisms that can establish and reproduce on their own, providing protection over long periods of time, and thus decreasing the cost of production.

[1] Bottazzi V. Food and feed production with microorganisms. Biotechnology 1983;5:315–63.