Striving for gut eubiosis with fatty acids
The concept of a “Eubiotic” approach is to overcome digestive disorders by ensuring a healthy gastro-intestinal microflora. Pathogenic bacteria need to be suppressed and lactic acid-producing bacteria to be promoted rather than killing off the total intestinal microflora with antibiotics. In broilers, short and medium chain fatty acids can support the healthy state of the gastro-intestinal tract.
The use of AGP’s in feed minimised disease outbreaks to a significant extent. Since they have been banned in Europe and other countries, enteric diseases have increased in poultry flocks, such as necrotic enteritis caused by Clostridium perfringens. This gram-positive anaerobic spore-forming bacterium is found commonly in soil and sewage and is a normal member of the microbial community in the gastro-intestinal tract of animals and humans, more particularly in the hind-gut. It is suggested that colonisation of poultry by C. perfringens occurs at an early stage in life and can be transmitted within the integrated broiler chicken operation, starting from the hatchery.
Conditions that promote excessive growth of C. perfringens in the chicken’s upper intestine are the typical digestive disorders caused by stress, excessive use of antibiotics, and a sub-acute status of coccidiosis. Especially Eimeria species that colonise the small intestine, such as Eimeria maxima and Eimeria acervulina, are known to be predisposed to necrotic enteritis. The use of high protein diets or diets high in non-starch polysaccharides derived from grains such as wheat and barley also strongly influence the incidence of necrotic enteritis in broilers.The disease occurs mostly in broiler chicks starting at the age of two weeks. The acute form of the disease leads to increased mortality in the broiler flocks. This can account for 1% mortality per day for several consecutive days during the last weeks of the rearing period. In the sub clinical form, damage to the intestinal mucosa caused by C. perfringens leads to decreased digestion and absorption, reduced weight gain and increased feed conversion ratio. According to our experience, a sub clinical status of necrotic enteritis is in the first place compromising feed intake leading to depressed growth and lower feed efficiency.
Necrotic enteritis despite in-feed antibiotics
Sub clinical necrotic enteritis is also reported in countries where the AGP’s are still in use. From our own experience (broiler trial, Australia, 2009), we noticed that virginiamycin at 40 ppm in the starter period gave a numerical advantage. However, there was no (further) effect on animal performances using virginiamycin at 20 ppm after the second week of broiler production when Clostridium perfringens usually starts colonising the small intestines (Table 1). Therefore, it can be suggested that, during the period when Clostridium perfringens becomes invasive, virginiamycin at low dosage is no longer effective.
Organic acids
Alternatives to AGPs have been widely developed. Organic acids seem to attribute to animal health in the absence of AGPs. The use of organic acids and salts is a long standing concept, used for more than three decades. Their antimicrobial action is mostly due to the pH changes of the environment in which the microorganisms occur.
All microorganisms need optimum conditions for their growth, including an optimum pH level. Bacteria are known to prefer a pH near to neutral values (pH 6.5-7.5), yeast prefer lower pH values and moulds have the widest range of acceptable pH. Various experiments have revealed the efficacy of organic acids, such as formic- and propionic acid, to successfully replace AGP’s without a significant reduction in performance. Formic-, propionic-, lactic-, acetic-, fumaric- and citric acids emerge as prospective growth promoters but more consistent and cost-effective performance is obtained through acid combinations. The use of organic acids to acidify swine diets is a common practice to reduce post-weaning diarrhoea in piglets and the mode of action is quite well understood. The use of organic acids and their salts in the poultry industry is still in an early stage. A proper application of organic acids in poultry production must improve protein digestion and nutrient retention, resulting in a better feed efficiency and increased growth and ultimately a more cost-effective production of meat and eggs. Creating a healthy intestinal microflora (eubiosis) is the focal point on poultry nutrition to overcome problems with necrotic enteritis caused by Clostridium perfringens.
Eubiotic effect
Organic acids can demonstrate eubiotic effects in different ways. Dissociated organic acids release protons that result in lowering the pH. Most of pathogenic microorganism (salmonella, E.coli) stop growing at a pH lower than 4.5, except for lactic acid producing bacteria (Lactobacilli, Streptococci, Bifidobacter). The organic acid anions, when dissociated, are charged and not lipid permeable. However, some of them have a destructive effect on the outer membrane of gram-negative bacteria such as Salmonella spp and E.coli. These gram-negative bacteria have an outer cell membrane consisting of a lipopolysaccharide (LPS) surface stabilised by calcium and magnesium ions. This membrane provides an effective barrier against a variety of antimicrobials. Molecules that can disrupt the integrity of this membrane are termed permeabilisers. Typically sorbic-, lactic- and citric acids are strong chelators and interfere with these metals from the cell membrane and increase the permeability of the LPS-layer of these gram-negative bacteria, allowing a better diffusion of the other organic acids or even provoking leakages. It is therefore recommended to use those permeabilisers along with organic acids as this will increase their effectiveness. Combined use of organic acids with lactic acid is typical. In addition to its antimicrobial property due to the lowering of the pH, lactic acid indeed also functions as a permeabiliser of the gram-negative bacterial outer membrane and may act as a potentiator of the effects of other organic acids.Undissociated organic acids have a lipophilic character and have the ability to pass through the cell membrane of gram-negative bacteria and to enter the microbial cell. There they will dissociate and lower the pH of the plasma. The cell must eliminate the H+across the cell membrane to restore the pH gradient (proton motif force). This requires a lot of energy and will lead to the death of the cell.
At a low pH, more of the organic acid will be in the undissociated form. Consequently, antimicrobial activity of organic acids is indisputable at low pH (gastric environment), but uncertain at pH above 6 (intestinal environment).
Antibacterial fatty acids
The antibacterial activity of Short Chain Fatty Acids (SCFA, <8 carbon atoms) in an acidic environment against gram-negative pathogens has been known for a long time. The most prominent natural fats rich in Medium Chain Fatty Acids (MCFA, C8 – C14) are coconut oil and palm kernel oil with less than 10% C8 (caprylic acid) and C10 (capric acid) but high proportions of C12 (lauric acid) and C14 (myristic acid). The acid dissociation constant (pKa) of MCFAs are close to 5 and are therefore more suitable to be active in the intestines than SCFAs.
Previous studies showed that lauric acid (C12) is the most inhibitory saturated MCFA against gram-positive organisms and has the highest antimicrobial activity against C. perfringens (minimum inhibitory concentration (MIC) of 0.1 – 0.2 mg/ml), followed by myristic and capric acid.
The mode of action of MCFA is not fully understood. In their undissociated form they may diffuse into the bacterial cells and dissociate within the protoplasm, thereby leading to intracellular acidification. Whether they diffuse freely across the cell membrane, as undissociated SCFAs are doing, or whether they are transported by carrier proteins as is the case with long-chain fatty acids (> C14) is less clear. Another possible mechanism of action may be a physical or functional alteration of the gastrointestinal colonisation site of pathogens in chicks; however, no studies are available showing the effects of MCFAs on gastrointestinal morphology. But MCFA may also alter the outer membranes of bacteria as they are used for incorporation in the membrane as phospholipids leading to membrane fluidity and even leakages. As such they may as well prevent colonisation or they may have a direct inhibitory effect on the expression of virulence factors necessary for colonisation. MCFAs have been shown to decrease salmonella invasion in intestinal epithelial cells.
Combining SFCA and MFCA
It is expected to see an added effect when combining SCFAs and their salts with MCFAs. This was confirmed in a broiler trial in an independent research institute in Poland (Figure 1) under practical conditions with feeds containing high levels of cereal grains, enzymes and coccidiostats, where this combination resulted in significant performance improvements. Moreover, another broiler trial in Australia shows that this combination can perfectly be used as an alternative to antimicrobial growth promoters (Figure 2). These trials show that the application of a combination of SCFAs and MCFAs (enriched in C12) increases final bodyweight up to 100 grammes and decreases FCR up to 3-5%. It may be suggested that this is the result of a lower subclinical status of necrotic enteritis. Depending on market prices, this leads to a potential profitability of €5-10 cents per kg of live weight broiler. For Europe, producing 12.1 million tonnes of broiler meat per year – and where the ban of AGP’s is almost complete – this means a potential increase in profitability of more than €605 million per year.
References are available from the authors.
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