Lutein belongs to the group of carotenoids, which is divided into carotenes and xanthophylls. Lutein, chemically expressed as “3,3’-dihydroxy-α-carotene”, is a xanthophyll always accompanied by its isomer zeaxanthin. It is synthesized out of two α-carotenes through hydroxylation.

Lutein provides benefits for animals and humans

Lutein has antioxidant protective properties

Under normal conditions, the cells in the animal and human organism control ROS (reactive oxygen species) levels. Usually, there is a balance between the generation of ROS and their elimination by scavenging systems. However, the high performance levels in modern animal production can easily lead to high ROS levels, translated into oxidative stress and leading to cell damage. Cell damage contributes to the generation of cancer and early aging in humans. In animals, the negative impact of oxidative stress can be responsible for lower performance and inferiority of meat and eggs.

Antioxidants stop ROS by taking up their energy

Through the uptake of energy, molecules can get into an excited state. One example is singlet excited oxygen, a highly reactive form of oxygen able to destroy proteins, lipids, and DNA. Carotenoids can intervene in this process: by exchanging electrons, the singlet excited oxygen gets neutralized, and the carotenoid gets into this excited state with higher energy. Once able to release this energy as heat into the environment, the carotenoid gets back to its normal state and can once again start acting as an antioxidant.

In this way, carotenoids, including lutein, ‘quench’ the energy of excited molecules and prevent the adverse effects of ROS (reactive oxidative substances).

Antioxidant properties profitably used

The antioxidant character of lutein plays an important role in the treatment or prophylaxis of macular degeneration in humans (Landrum & Bone, 2001). There is also evidence that lutein can be used to improve the visual and retinal function in dogs (Wang et al., 2016). In the eye, lutein and zeaxanthin, occurring in the retina and the macula, neutralize free radicals produced due to the ultraviolet light and thereby prevent damage to the macula.

Further possible applications are against cardiovascular diseases (Dwyer et al., 2001)  and various types of cancer (e.g., breast cancer, Gong et al., 2018).

Lutein is important in infant nutrition

Lutein and its isomer zeaxanthin are the two primary carotenoids found in human milk (Giordano and Quadro, 2018). Stringham and co-workers (2019) postulate that lutein plays an important role in children’s visual and cognitive development/optimization. They report that a lutein supplementation of the mother can lead to a higher concentration of this substance in the milk and, consequently, in the child’s plasma (Sherry et al., 2014). In dairy cows, an increased level of lutein in the milk can also be observed (Xu et al., 2014), suggesting that lutein could also be essential in calf development.

Lutein stimulates the immune system

Another benefit of lutein is its positive influence on the immune system.

On the one hand, lutein stimulates the production of antibodies. In dogs, Guimarães Alarça et al. (2016) could show an increase of CD4+ and CD8+ T-lymphocyte subtypes. Kim et al. (2000) demonstrated the increase of lymphocytes and cells expressing CD5, CD4, CD8, and major histocompatibility complex class II (MHC II) molecules. Bédécarrats and Leeson (2006) provoked a higher secondary antibody response to infectious bronchitis vaccination in laying hens.

Besides, lutein acts as an anti-inflammatory agent, as shown in vitro by Chao et al. (2015) and in broiler chickens by Moraes and team (2016).

Lutein improves the attractivity of poultry products

In the marketing of poultry products, appearance and color are of central importance for evaluating quality. Egg yolk coloration is to a large extent a matter of regional preferences, however it is clear that an egg with a yolk that does not have the typical color is classified as inferior by the consumer. In areas with traditional corn growing, a white-skinned chicken is not commercially viable. Even when pullets are bought, the shanks and beaks should be yellow.

The use of xanthophylls like lutein and zeaxanthin enables producers to safely control the color of the egg yolk and of the broiler skin. It also leads to a healthy color of the shanks and beaks of the birds.

Lutein in a nutshell

Lutein is a true all-rounder: a substance that delivers benefits across the board. In plants, it helps fruits and petals become attractive for insects and other animals. It positively influences the animal, acting as an antioxidant, promoting infant development, and stimulating the immune system. As a pigment, it makes poultry and poultry products look more attractive to the consumer. Through its presence in eggs and milk, lutein provides clear and clean benefits to both animals and humans.

References

Bédécarrats, G.Y. and S. Leeson. “Dietary lutein influences immune response in laying hens.”  J. Appl. Poult. Res. 15 (2006): 183–189.

https://doi.org/10.1093/japr/15.2.183

Chao, Shih-Chun, Tommaso Vagaggini, Chan-Wei Nien, Sheng-Chieh Huang, and Hung-Yu Lin. “Effects of Lutein and Zeaxanthin on LPS-Induced Secretion of IL-8 by Uveal Melanocytes and Relevant Signal Pathways.” Journal of Ophtalmology, vol. 2015 Article ID 152854 (2015): 7 pages. https://doi.org/10.1155/2015/152854

Dwyer, James H., Mohamad Navab, Kathleen M. Dwyer, Kholood Hassan, Ping Sun, Anne Shircore, Susan Hama-Levy, Greg Hough, Xuping Wang, Thomas Drake, C. Noel Bairey Merz, and Alan M. Fogelman. “Oxygenated Carotenoid Lutein and Progression of Early Atherosclerosis.” Circulation (American Heart Association) 103, no. 24 (2001): 2922-2927.

https://doi.org/10.1161/01.CIR.103.24.2922

Gong, Xiaoming, Joshua R. Smith, Haley M. Swanson, and Lewis P. Rubin. “Carotenoid Lutein Selectively Inhibits Breast Cancer Cell Growth and Potentiates the Effect of Chemotherapeutic Agents through ROS-Mediated Mechanisms.” Molecules 23 no. 4(2018): 905.

http://dx.doi.org/10.3390/molecules23040905

Guimarães Alarça, Laís, Fabiane Yukiko Murakami, Ananda Portella Félix, Everton Luis Krabbe, Simone Gisele de Oliveira, Sebastião Aparecido Borges da Silva. “Dietary lutein supplementation on diet digestibility and blood parameters of dogs.” Cienc. Rural 46 no.12 (2016)

http://dx.doi.org/10.1590/0103-8478cr20151493

Kim, Hong Wook, Boon Chew, Teri Ann S Wong, Jean Soon Park, Bor-Chun Weng, Katherine M Byrne, Michael G Hayek, and Gregory A. Reinhart. “Dietary lutein stimulates immune response in the canine.” Veterinary Immunology and Immunopathology 74 no. 3-4 (2000): 315-327.

https://doi.org/10.1016/S0165-2427(00)00180-X

Landrum, J. T. and R.A. Bone. “Lutein, zeaxanthin, and the macular pigment.” Archives of Biochemistry and Biophysics 385 no. 1 (2001): 28–40.

https://doi.org/10.1006/abbi.2000.2171.

Moraes, M. L., A. M. L. Ribeiro, E. Santin, and K. C. Klasing. “Immunology, health, and disease: effects of conjugated linoleic acid and lutein on the growth performance and immune response of broiler chickens.” Poultry Science 95 (2016): 237–246.

http://dx.doi.org/10.3382/ps/pev325

Ochoa Becerra, Mario, Luis Mojica Contrerasa, Ming Hsieh Loa, Juan Mateos Díaz, Gustavo Castillo Herrera. “Lutein as a functional food ingredient: Stability and bioavailability.” Journal of Functional Foods 66 (2020): 103771.

https://doi.org/10.1016/j.jff.2019.103771

Sherry, Christina L.,  Jeffery S. Oliver, Lisa M. Renzi, and Barbara J. Marriage. “Lutein supplementation increases breast milk and plasma lutein concentrations in lactating women and infant plasma  concentrations but does not affect other carotenoids.” J. Nutr. 144 (2014): 1256–1263.

http://dx.doi.org/10.3945/jn.114.192914

Stringham, James M., Elizabeth J Johnson, and B Randy Hammond. “Lutein across the lifespan: From childhood cognitive performance to the aging eye and brain.” Curr Dev Nutr 3 (2019): nzz066.

http://dx.doi.org/10.1093/cdn/nzz066

Wang, Wei, Jerome Hernandez, Cecil Moore, Janet Jackson, and Kristina Narfström. “Antioxidant supplementation increases retinal responses and decreases refractive error changes in dogs.” J. Nutr. Sci. 5 e18 (2016): 7 pages

http://dx.doi.org/10.1017/jns.2016.5

Xu, C.Z., H. F. Wang, J. Y. Yang, J. H. Wang, Z. Y. Duan, C. Wang, J. X. Liu , and Y. Lao. “ Effects of feeding lutein on production performance, antioxidative status, and milk quality of high-yielding dairy cows.” J. Dairy Sci. 97;  American Dairy Science Association (2014):7144–7150

http://dx.doi.org/10.3168/jds.2014-8276

Strong demand by consumers; restaurant chains and wholesalers for antibiotic-free (ABF) meat; the threat of antimicrobial resistance; and stringent regulations on the use of antibiotics – there are many good reasons for poultry producers to strive for antibiotic-free production systems. Crucially, to successfully produce poultry meat without antibiotics requires a paradigm shift that starts right at the parent stock level, with the antibiotic-free production of hatching eggs.

Broiler breeders’ gut health is linked to progeny’s performance

Broiler breeders’ performance is measured in terms of how many saleable day old chicks (DOCs) per hen they produce. However, within a sustainable ABF production system (also known as No Antibiotics Ever or NAE), this parameter is not seen in isolation. Breeder hens’ nutritional and health status not only affect the number of DOCs they can produce, but also the transfer of nutrients, antibodies, microbiota and even contaminants, e.g. mycotoxins, to the egg – and therefore, their progeny’s long-term health and performance.

This starts with egg formation, which requires several metabolic processes in the hen to function perfectly. If the hen’s intestinal integrity is compromised, for example due to mycotoxins, she will absorb fewer nutrients, which in turn affects egg formation. Mycotoxicosis has particularly insidious effects for egg formation as it can damage the liver whose biosynthetic activities strongly impact on the egg’s internal (yolk) and external (eggshell) quality.

Chick embryos depend on the maternal antibodies and nutrients deposited in the yolk, including vitamin D3, carotenoids, and fatty acids, to develop normally. Eggshell quality, among other things, affects the embryo’s access to oxygen, which is especially important when it develops body tissues.

Hens’ ability to form healthy eggs depends on their diet and health. Research indicates that, via the impact on egg formation, broiler breeders’ feeding program quantifiably influences their progeny’s immune system and intestinal health. There is indeed a direct relationship between parent and offspring’s gut health because the chick’s microbiome is in part also inherited from the hen. The impact on DOC quality is thus one of many dimensions to consider when calibrating one’s broiler breeders feeding approach.

The challenge of feeding an ABF broiler breeder

Just as their offspring, breeder hens are genetically predisposed for rapid growth and muscle development. From rearing right through to the laying period, poultry nutritionists need to carefully balance their diets and moderate weight gain in order for hens to reach their reproductive potential.

Different stages of a breeder’s life cycle come with different objectives – for example, good flock uniformity in the rearing period versus egg size and hatchability in the laying phase – and thus different requirements in terms of calories, amino acids, vitamins, and minerals. What remains constant is that the actual nutrient intake depends on intestinal health, determining both the breeders’ performance and, via the impact on egg characteristics, its progeny’s performance.

The feeding regimes adopted to avoid hens becoming overweight can have a negative effect on their gut flora. Without antibiotics as a tool to maintain or recover optimal gut function, even mild intestinal disorders can quickly become chronical impairments that negatively impact breeders’ productivity. In ABF production systems, intestinal health therefore needs to be a central focus for the feeding strategy.

Can phytomolecules improve broiler breeders’ performance?

Among the plethora of feed additives, phytomolecules, or secondary plant compounds, stand out as a class of active ingredients that may help to improve gut health and thereby reduce the use of antibiotics.  Synthesized by plants as a defense mechanism against pathogens, phytomolecules combine digestive, antimicrobial and antioxidant properties.

Some studies have shown that phytomolecules-based products can increase broilers’ body weight gain and improve laying hens’ laying rate, egg mass and egg weight. Both broilers and laying hens responded to the inclusion of phytomolecules in their diet with inclusion rate-dependent improvements in feed conversion. To evaluate if phytomolecules could similarly improve broiler breeders’ performance, two trials were conducted.

Study I: Effect of phytomolecules on laying performance during peak production

The first study was set up on a farm in Thailand. In total, 40000 Cobb broiler breeders (85% female, 15% male) were divided into two groups with 8500 hens (one house) in the control and 25500 (three houses) in the trial group. Both groups were fed standard feed. The trial group additionally received a phytomolecules-based liquid complementary feed (Activo® Liquid, EW Nutrition GmbH) via the waterline from week 24 to week 32 at a rate of 200ml/1000L during 5 days per week.

Activo® Liquid was found to have a positive influence on laying performance (Figure 1). The average laying rate increased by 7.2% during the trial period, resulting in almost 3 additional hatching eggs per hen housed. A further indication of the beneficial influence that this particular combination of phytomolecules had on gut health was a 0.2% lower mortality.

Figure 1: Laying rate (%) of breeder hens during first 9 weeks of production

Study II: Effect of phytomolecules on laying performance after peak production

For a second study, conducted in the Czech Republic, 800 female and 80 male Hubbard breeders (JA57 and M77, respectively) were divided into 2 groups with 5 replicate pens and 80 female and 8 male breeders per pen. The experiment started after the peak-production period, at 34 weeks of age and ended at 62 weeks of age. All animals received a standard mash diet. For one group a phytogenic premix (Activo^®, EW Nutrition GmbH) was added to the diet at a rate of 100g/MT.

The results indicate that Activo^® helped maintain the breeder hens’ egg laying performance close to the breed’s genetic potential (Figure 2). In the course of the experiment, Activo® supplemented birds produced 3.6 more eggs than control birds, while consuming a similar amount of feed. As a result, feed consumption per egg produced was lower for birds receiving phytomolecules than for the control birds (169.9 versus 173.6g/d, respectively).

As hatchability was not influenced by the dietary treatment in this study (P>0.5), the 3.6 extra eggs resulted in 2.9 extra day old chicks per hen produced, during the post-peak period alone.
The microencapsulated, selected phytomolecules contained in Activo^® are likely to have improved gut health and feed digestibility, and thereby enhanced the animals’ feed efficiency.

Figure 2: Laying rate (%) of breeder hens week 35 till 62

Chicken or egg? Antibiotic-free poultry production looks at the bigger picture

To successfully produce antibiotic-free poultry meat requires a systematic re-think of each component of the production process. Broiler breeders’ lay the foundation for their progeny’s health and performance via the egg. Breeder hens need to be in optimal health to consistently deliver optimal eggs. Without recourse to antibiotics for maintaining or recovering intestinal functionality, an effective ABF production needs to make gut health central to its feeding approach.

The trials reviewed demonstrate that selected phytomolecules quantifiably boost breeders’ laying performance, increasing the number of hatching eggs and DOCs, while reducing mortality and feed consumption per egg produced. As part of an intelligent antibiotic reduction strategy, the right phytogenic products can be potent tools to help poultry producers achieve their NAE objectives.

by S. Regragui Mazili, T. van Gerwe and M. Caballero

References

Calini, F., and F. Sirri. “Breeder Nutrition and Offspring Performance.” Revista Brasileira De Ciência Avícola 9, no. 2 (2007): 77-83. doi:10.1590/s1516-635×2007000200001.

Ding, Jinmei, Ronghua Dai, Lingyu Yang, Chuan He, Ke Xu, Shuyun Liu, Wenjing Zhao, et al. “Inheritance and Establishment of Gut Microbiota in Chickens.” Frontiers in Microbiology 8 (October 10, 2017): 1967.

Kuttappan, Vivek A., Eduardo A. Vicuña, Juan D. Latorre, Amanda D. Wolfenden, Guillermo I. Téllez, Billy M. Hargis, and Lisa R. Bielke. “Evaluation of Gastrointestinal Leakage in Multiple Enteric Inflammation Models in Chickens.” Frontiers in Veterinary Science 2 (December 14, 2015): 66.

Moraes, Vera M. B., Edgar O. Oviedo-Rondón, Nadja S. M. Leandro, Michael J. Wineland, Ramon D. Malheiros, and Pamela Eusebio-Balcazar. “Broiler Breeder Trace Mineral Nutrition and Feeding Practices on Embryo Progeny Development.” Avian Biology Research 4, no. 3 (2011): 122–32.

Oviedo-Rondon, Edgar O., Nadja S. M. Leandro, Rizwana Ali, Matthew Koci, Vera M. B. Moraes, and John Brake. “Broiler Breeder Feeding Programs and Trace Minerals on Maternal Antibody Transfer and Broiler Humoral Immune response1.” The Journal of Applied Poultry Research 22, no. 3 (October 1, 2013): 499–510.

Most grains used as feed raw materials are susceptible to mycotoxin contamination. These toxic secondary metabolites are produced by fungi before or after harvest and cause severe economic losses all along agricultural value chains. For livestock, negative consequences include acute effects such as impaired liver and kidney function, vomiting, or anorexia, as well as chronic effects such as immunosuppression, growth retardation, and reproductive problems. Mycotoxin management is, therefore, of the utmost priority for animal producers worldwide.

But how is it that mycotoxins cause such damage in the first place? This article delves into the complex processes that take place when mycotoxins come into contact with the gastrointestinal tract (GIT). The intestinal epithelium is the first tissue to be exposed to mycotoxins, and often at higher concentrations than other tissues. A deeper understanding of how mycotoxins affect the GIT allows us to appreciate the cascading effects on animal health and performance, why such damage already occurs at contamination levels well below official safety thresholds – and what we can do about it.

The intestinal epithelium: the busy triage site for nutrients and harmful substances

When mycotoxins are ingested, they encounter the GIT’s intestinal epithelium (Figure 1). This single layer of cells lining the intestinal lumen serves two conflicting functions: firstly, it must be permeable enough to allow the absorption of nutrients. On the other hand, it constitutes the primary physiological barrier against harmful agents such as viruses, microorganisms, and toxins.

Within the intestinal epithelium, several types of highly specialized cells are involved in epithelial regeneration, nutrient absorption, innate defense, transport of immunoglobulins, and immune surveillance. The selective barrier function is maintained due to the formation of complex networks of proteins that link adjacent cells and seal the intercellular space. Besides, the intestinal epithelium is covered with mucus produced by goblet cells, which isolates its surface, preventing the adhesion of pathogens to the enterocytes (intestinal absorptive cells).

Due to its dual involvement in digestive and immune processes, the intestinal epithelium plays a pivotal role in the animal’s overall health. Importantly, the epithelium is directly exposed to the entire load of ingested mycotoxins. Hence their effects can be problematic even at low concentrations.

Figure 1: The intestinal epithelium

Problematic effects of mycotoxins on the intestinal epithelium

Most mycotoxins are absorbed in the proximal part of the gastrointestinal tract (Table 1). This absorption can be high, as in the case of aflatoxins (~90%), but also very limited, as in the case of fumonisins (<1%); moreover, it depends on the species. Importantly, a significant portion of unabsorbed toxins remains within the lumen of the gastrointestinal tract.

Some of the mycotoxins that enter the intestinal lumen can be bio-transformed into less toxic compounds by the action of certain bacteria. This action, however, predominantly happens in the large intestine – therefore, no detoxification takes place before absorption in the upper parts of the GIT. Part of the absorbed mycotoxins can also re-enter the intestine, reaching the cells from the basolateral side via the bloodstream. Furthermore, they re-enter through enterohepatic circulation (the circulation of substances between the liver and small intestine). Both actions increase the gastrointestinal tract’s overall exposure to the toxins.

Table 1: Rate and absorption sites of different mycotoxins

Adapted from: Biehl et al., 1993; Bouhet & Oswald, 2007; Devreese et al., 2015; Ringot et al., 2006

The damaging impact of mycotoxins on the intestinal epithelium initially occurs through:

• A decrease in protein synthesis, which reduces barrier and immune function (Van de Walle et al., 2010)
• Increased oxidative stress at the cellular level, which leads to lipid peroxidation, affecting cell membranes (Da Silva et al., 2018)
• Changes in gene expression and the production of chemical messengers (cytokines), with effects on the immune system and cellular growth and differentiation (Ghareeb et al., 2015)
• The induction of programmed cell death (apoptosis), affecting the reposition of immune and absorptive cells (Obremski & Poniatowska-Broniek, 2015)

Importantly, studies based on realistic mycotoxin challenges (e.g., Burel et al., 2013) show that the mycotoxin levels necessary to trigger these processes are lower than the levels reported as safe by EFSA, the Food Safety Agency of the European Union. The ultimate consequences range from diminished nutrient absorption to inflammatory responses and pathogenic disorders in the animal (Figure 2).

Figure 2: Mycotoxins’ impact on the GIT and consequences for monogastric animals

1. Alteration of the intestinal barrier‘s morphology and functionality

The mycotoxins DON, fumonisin, and T2 induce a reduction in the rate of epithelial cell proliferation and differentiation. This causes a decrease in the height and the surface of the intestinal villi, which in turn leads to a reduction in nutrient absorption. Additionally, some nutrient transporters are inhibited by the action of mycotoxins such as DON and T2, for example, negatively affecting the transport of glucose.

Several studies indicate that mycotoxins such as aflatoxin B1, DON, fumonisin B1, ochratoxin A, and T2, can increase the permeability of the intestinal epithelium of poultry and swine (e.g. Pinton & Oswald, 2014). This is mostly a consequence of the inhibition of protein synthesis. As a result, there is an increase in the passage of antigens into the bloodstream (e.g., bacteria, viruses, and toxins). This increases the animal’s susceptibility to infectious enteric diseases. Moreover, the damage that mycotoxins cause to the intestinal barrier entails that they are also being absorbed at a higher rate.

2. Impaired immune function in the intestine

The intestine is a very active immune site, where several immuno-regulatory mechanisms simultaneously defend the body from harmful agents. Immune cells are affected by mycotoxins through the initiation of apoptosis, the inhibition or stimulation of cytokines, and the induction of oxidative stress. Studies demonstrate that aflatoxin, DON, fumonisin, T2, and zearalenone interact with the intestinal immune system in such a manner that the animal’s susceptibility to viral and bacterial infections increases (e.g., Burel et al., 2013). Moreover, by increasing their fecal elimination, the horizontal transmission of pathogens is extended.

For poultry production, one of the most severe enteric problems of bacterial origin is necrotic enteritis, which is caused by Clostridium perfringens toxins. Any agent capable of disrupting the gastrointestinal epithelium – e.g. mycotoxins such as DON, T2, and ochratoxin – promotes the development of necrotic enteritis. The inhibition of the intestinal immune system caused by mycotoxins such as aflatoxin, DON, and T2 also collaborates with the development of this disease.

3. Alteration of the intestinal microflora

The gastrointestinal tract is home to a diverse community of bacteria, fungi, protozoa, and viruses, which lines the walls of the distal part of the intestine. This microbiota prevents the growth of pathogenic bacteria through competitive exclusion and the secretion of natural antimicrobial compounds, volatile fatty acids, and organic acids.

Recent studies on the effect of various mycotoxins on the intestinal microbiota show that DON and other trichothecenes favor the colonization of coliform bacteria in pigs. DON and ochratoxin A also induce a greater invasion of Salmonella and their translocation to the bloodstream and vital organs in birds and pigs – even at non-cytotoxic concentrations. It is known that fumonisin B1 may induce changes in the balance of sphingolipids at the cellular level, including for gastrointestinal cells. This facilitates the adhesion of pathogenic bacteria, increases in their populations, and prolongs infections, as has been shown for the case of E. coli.

From the perspective of human health, the colonization of the intestine of food-producing animals by pathogenic strains of E. coli and Salmonella is of particular concern. Mycotoxin exposure may well increase the transmission of these pathogens, posing a risk for human health.

4. Interaction with bacterial toxins

When mycotoxins induce changes in the intestinal microbiota, this can lead to an increase in the endotoxin concentration in the intestinal lumen. Endotoxins or lipopolysaccharides (LPS) are fragments of Gram-negative bacteria’s cell walls. They are released during bacterial cell death, growth, and division. Hence endotoxins are always present in the intestine, even in healthy animals. Endotoxins promote the release of several cytokines that induce an enhanced immune response, causing inflammation, thus reducing feed consumption and animal performance, damage to vital organs, sepsis, and death of the animals in some cases.

The synergy between mycotoxins and endotoxins can result in an overstimulation of the immune system. The interaction between endotoxins and estrogenic agents such as zearalenone, for example, generates chronic inflammation and autoimmune disorders because immune cells have estrogen receptors, which are stimulated by the mycotoxin. The combination of DON at low concentrations and endotoxins in the intestine, on the other hand, has been shown to engender a decrease in transepithelial resistance and to alter the balance of the microbiota.

What to do? Proactive toxin risk management

To prevent the detrimental consequences of mycotoxins on animal health and performance, proactive solutions are needed that support the intestinal epithelium’s digestive and immune functionality and help maintain a balanced microbiome in the GIT. Moreover, it is crucial for any anti-mycotoxin product to feature both anti-mycotoxin and anti-bacterial toxin properties and that it supports the organs most targeted by mycotoxins, e.g., the liver. EW Nutrition’s Mastersorb® Gold premix is based on the synergistic combination of natural clay minerals, yeast cell walls, and phytomolecules. Its efficacy has been extensively tested, including as a means for dealing with E. coli endotoxins.

Mastersorb® Gold: anti-mycotoxin activity stabilizes performance and strengthens liver health

A field trial conducted in Germany on male Ross 308 broilers showed that for broilers receiving a diet contaminated with DON and zearalenone, adding 1kg of Mastersorb® Gold per ton of feed to their diet led to significant performance enhancements. Not only did they recuperate the mycotoxin-induced weight loss (6% increase relative to the group receiving only the challenge), but they gained weight relative to the control group (which received neither the challenge nor Mastersorb® Gold). Feed conversion also improved by 3% relative to the group challenged with mycotoxins.

A scientific study of crossbred female pigs showed that Mastersorb® Gold significantly reduced the deleterious effects of fumonisin contamination in the feed. The decrease in weight gain and the decline of feed conversion could be mitigated by 6.7% and 13 FCR points, respectively (Figure 3). Also, the sphinganine/sphingosine (Sa/So) ratio, a biomarker for fumonisin presence in the blood serum, could be decreased by 22.5%.

Figure 3: Mastersorb® Gold boosts performance for pigs fed a fumonisin-contaminated diet

Another study of crossbred female piglets, carried out at a German university, investigated whether Mastersorb® Gold could support performance as well as liver health under a naturally occurring challenge of ZEA (~ 370ppb) and DON (~ 5000ppb).  Mastersorb® Gold significantly improved weight gain and feed conversion in piglets receiving the mycotoxin-contaminated diet: daily body weight gain was 75g higher than that of a group receiving only the challenge, and the FCR improved by 24% (1.7 vs. 2.25 for the group without Mastersorb® Gold). Moreover, Mastersorb® Gold significantly improved the liver weight (total and relative) and the piglets’ AST levels (aspartate aminotransferase, an enzyme indicating liver damage). A tendency to improve spleen weight and GGT levels (gamma-glutamyl transferase, another enzyme indicative of liver issues) was also evident, all of which indicate that Mastersorb® Gold effectively counteracts the harmful impact of mycotoxin contamination on liver functionality.

In-vitro studies demonstrate Mastersorb® Gold’s effectiveness against mycotoxins as well as bacterial toxins

Animal feed is often contaminated with two or more mycotoxins, making it important for an anti-mycotoxin agent to be effective against a wide range of different mycotoxins. Besides, to prevent mycotoxins damaging the GIT, an effective product should ideally adsorb most mycotoxins in the first part of the animal’s intestine (under acidic conditions). In-vitro experiments at an independent research facility in Brazil showed that an application of 0.2% Mastersorb® Gold binds all tested mycotoxins at rates from 95 to 97% at a pH level of 3, using realistic challenges of 1000ppb (Aflatoxin B1 and ZEA) and 2500ppb (Fumonisin B1 and DON). The binding results achieved for Fumonisin and DON, which are often considered outright “nonbinding,” under challenging close to neutral conditions (pH 6), are particularly encouraging.

Figure 4: Mastersorb® Gold binding capacity against different mycotoxins (%)

Concerning its efficacy against endotoxins, an in vitro study conducted at Utrecht University, among other studies, has shown Mastersorb® Gold to be a strong tool against the LPS released by E. coli. For the test, four premium mycotoxin binders were suspended in a phosphate buffer solution to concentrations of 0.25% and 1%. E. coli LPS were suspended to a final concentration in each sample of 50ng/ml. Against this particularly high challenge, Mastersorb® Gold achieved a binding rate of 75% at an inclusion rate of 1%: clearly outperforming competing products, which at best showed a binding rate of 10%.

Conclusion

A healthy gastrointestinal tract is crucial to animals’ overall health: it ensures that nutrients are optimally absorbed, it provides effective protection against pathogens through its immune function, and it is key to maintaining a well-balanced microflora. Even at levels considered safe by the European Union, mycotoxins can compromise different intestinal functions such as absorption, permeability, immunity, and microbiota balance, resulting in lower productivity and susceptibility to disease.

To safeguard animal performance, it is important to continually strive for low levels of contamination in feed raw materials –  and to stop the unavoidable mycotoxin loads from damaging the intestinal epithelium through the use of an effective anti-mycotoxin agent, which also supports animals against endotoxins and boosts liver function. Research shows that Mastersorb Gold is a powerful tool for proactive producers seeking stronger animal health, welfare, and productivity.

By Marisabel Caballero and Sabria Regragui Mazili

Sub-therapeutic doses of antibiotic growth promoters (AGPs) were used for more than 50 years in poultry production to achieve performance targets – until growing concerns arose regarding antibiotic resistance (Kabir, 2009) and decreasing efficacy of antibiotics for medical purposes (Dibner & Richards, 2005).

Isolates of ESBL-producing E.coli from animals, farmworkers, and the environment were found to have identical multidrug resistance patterns (A. Nuangmek et al., 2018). There is also evidence that AMR strains of microorganisms spread from farm animal to animal workers and beyond. Global AMR fatalities are increasing and might reach 10 million by 2050 (Mulders et al., 2010, Trung et al., 2017, Huijbers et al., 2014).

In light of this, certain AGPs have already been banned, and there is a strong possibility of future restrictions on their use worldwide. Bans are effective: the MARAN report 2018 shows that lower antibiotics usage following the EU ban on AGPs has reduced resistant E.coli in broilers. Another positive consideration is the market opportunities that exist for antibiotic residue-free food.

However, the key element that poultry producers need to get right for antibiotic reduction to be successful is respiratory health management. This article looks at why respiratory health is a particular challenge – and how phytogenic solutions can help.

A closer look at the chickens’ respiratory system

The respiratory tract is equipped with a functional mucociliary apparatus consisting of a protective mucous layer, airway surface liquid layer, and cilia on the surface of the ciliated cells. This apparatus produces mucus, which traps the inhaled particles and pathogens and propels them out of the airways. This mechanism, called the mucociliary clearance, is the primary innate defense mechanism of the respiratory system.

High stocking density combined with stressful environmental factors can negatively influence birds’ immune systems (Heckert et al., 2002; Muniz et al., 2006), making them more susceptible to respiratory disease. When a bird suffers from respiratory disease, which is nowadays usually complicated by a co-infection or secondary bacterial infection, there is an excess production of mucus that results in ciliostasis and, therefore, in an impaired mucociliary clearance. The excess mucus in the tract obstructs the airways by forming plagues and plugs, resulting in dyspnea (hypoxia) and allowing the invasive bacteria to adhere and colonize the respiratory system.

The build-up of mucus in the respiratory tract severely reduces oxygen intake, causing breathlessness, reduced feed intake, and a drop in the birds’ energy levels, which negatively impacts weight gain and egg production. Respiratory problems can result from infection with bacteria, viruses, and fungi, or exposure to allergens. The resultant irritation and inflammation of the respiratory tract leads to sneezing, wheezing, and coughing – and, therefore, the infection rapidly spreads within the flock.

Relatively high stocking density is the norm in poultry production

Low or no antibiotics: how to manage respiratory disease?

Unsurprisingly, respiratory diseases in poultry are a major cause of mortality and economic loss in the poultry industry. For Complicated Chronic Respiratory Disease (CCRD), for instance, although the clinical manifestations are usually slow to develop, Mycoplasma gallisepticum (MG), in combination with E. coli, can cause severe airsacculitis. Beside feed and egg production reduction, these problems are of high economic significance since respiratory tract lesions can cause high morbidity, high mortality, and significant carcass condemnation and downgrading.

Producers need to pre-empt the spread of respiratory pathogens, react quickly to alleviate respiratory distress and maintain the mucociliary apparatus’ functionality. Traditionally, treatment options are based on antiviral, anti-inflammatory, and antibiotic drugs. Can the poultry industry limit losses from respiratory infections without excessive recourse to antibiotics?

Indeed, a sudden reduction in antibiotic usage comes with a risk of impaired performance, increased mortality, and impaired animal health and welfare. The impact has been quantified as a 5% loss in broiler meat production per sq. meter (Gaucher et al., 2015). Effective antibiotics reduction requires a combination of innovative products and suitable consultancy services to manage poultry gut health, nutrition, flock management, biosecurity, and, particularly, respiratory health.

Non-antibiotic alternatives to control diseases and promote broiler growth, such as organic acids (Vieira et al., 2008), probiotics (Mountzouris et al., 2010), prebiotics (Patterson & Burkholder, 2003), and essential oils (Basmacioğlu Malayoğlu et al., 2010) have been the subject of much research in recent years.

Phytogenic solutions: proven efficacy

Essential oils, which are extracted from plant parts, such as flowers, buds, seeds, leaves, twigs, bark, wood, fruits, and roots, have a particularly well-established track record of medicinal applications. Efforts have centered on phytomolecules, the biologically active secondary metabolites that account for the properties of essential oils (Hernández et al., 2004; Jafari et al., 2011).

Studying these properties is challenging: essential oils are very complex natural mixtures of compounds whose chemical compositions and concentrations are variable. For example, the concentrations of the two predominant phytogenic components of thyme essential oils, thymol and carvacrol, have been reported to range from as low as 3% to 60% of the whole essential oil (Lawrence and Reynolds, 1984).

Another well-researched example is eucalyptus oil. The essential oils of eucalyptus species show antibacterial, anti-inflammatory, diaphoretic, antiseptic, analgesic effects (Cimanga et al., 2002) and antioxidant properties (Lee and Shibamoto, 2001; Damjanović Vratnica et al., 2011). The oils are mainly composed of terpenes and terpene derivatives in addition to some other non-terpene components (Edris, 2007). The principal constituent found in eucalyptus is 1,8-cineole (eucalyptol); however, other chemotypes such as α-phellandrene, ρ-cymene, γ-terpinene, ethanone, and spathulenol, among others, have been documented (Akin et al., 2010).

Close-up of eucalyptus leaf oil glands and
the molecular structure of eucalyptol C₁₀H₁₈O (red = oxygen; dark grey = carbon; light grey = hydrogen)

Antimicrobial activity

In modern intensive broiler production, bacterial diseases such as salmonellosis, colibacillosis, mycoplasmosis, or clostridia pose serious problems for the respiratory system and other areas. Analyses of the antibacterial properties of essential oils have been carried out by multiple research units (Ouwehand et al., 2010; Pilau et al., 2011; Solorzano- Santos and Miranda-Novales, 2012; Mahboubi et al., 2013; Nazzaro et al., 2013; Petrova et al., 2013).

Phenols, alcohols, ketones, and aldehydes are clearly associated with antibacterial activity; the exact mechanisms of action, however, are not yet fully understood (Nazzaro et al., 2013). Essential oils’ antimicrobial activity is not attributable to a unique mechanism but instead results from a cascade of reactions involving the entire bacterial cell (Nazzaro et al., 2013). However, it is accepted that antimicrobial activity depends on the lipophilic character of the components.

The components permeate the cell membranes and mitochondria of the microorganisms and inhibit, among others, the membrane-bound electron flow and thus the energy metabolism. This leads to a collapse of the proton pump and draining of the ATP (adenosine triphosphate) pool. High concentrations may also lead to lysis of the cell membranes and denaturation of cytoplasmic proteins (Nazzaro et al., 2013; Gopi et al., 2014).

According to current knowledge, lavender, thyme, and eucalyptus oil, as well as the phytomolecules they contain, show enhanced effects when combined with other essential oils or synthetic antibiotics (Sadlon and Lamson, 2010; Bassole and Juliani, 2012; Sienkiewicz, 2012; de Rapper et al., 2013; Zengin and Baysal, 2014).

Minimum inhibitory concentration (MIC) of some essential oil components against microorganisms in vitro

Immune system boost I: improved production of antibodies

Some essential oils were found to influence the avian immune system positively, since they promote the production of immunoglobulins, enhance the lymphocytic activity, and boost interferon-γ release (Awaad et al., 2010; Faramarzi et al., 2013; Gopi et al., 2014; Krishan and Narang, 2014). Placha et al. (2014) showed that the addition of 0.5g of thyme oil per kg of feed significantly increased IgA levels.

Awaad et al. (2010) experimented on birds vaccinated with the inactivated H5N2 avian influenza vaccine. The experiment revealed that adding eucalyptus and peppermint essential oils to the water at a rate of 0.25 ml per liter resulted in an enhanced cell-mediated and humoral immune response.

Saleh et al. (2014), who applied thyme and ginger oils in quantities of 100mg and 200mg per kg of feed, respectively, observed an improvement in chickens’ immunological blood profile through increased antibody production. Rehman et al. (2013) stated that the use of herbal products containing eucalyptus oil and menthol in broilers showed consistently higher antibody titers against NDV (Newcastle disease virus), compared to untreated broilers.

Immune system boost II: better vaccine responses and anti-inflammatory effects

Essential oils are also used as immunomodulators during periods when birds are exposed to stress, acting protectively and regeneratively. Importantly, the oils alleviate the stress caused by vaccination (Barbour et al., 2011; Faramarzi et al., 2013; Gopi et al., 2014). The study by Kongkathip et al. (2010) confirmed the antiviral activity of turmeric essential oil.

In recent years studies have been carried out on the use of essential oils in conjunction with vaccination programs, including those against infectious bronchitis (IB), Newcastle disease, and Gumboro disease. The results of the experiments show that essential oils promote the production of antibodies, thus enhancing the efficacy of vaccination (Awaad et al., 2010; Barbour et al., 2010; Barbour et al., 2011; Faramarzi et al., 2013).

Essential oils contain compounds that are known to possess strong anti-inflammatory properties, mainly terpenoids, and flavonoids, which suppress the metabolism of inflammatory prostaglandins (Krishan and Narang, 2014). Also, other compounds found in essential oils have anti-inflammatory, pain-relieving, or edema-reducing properties, for example, linalool from lavender oil, or 1,8-cineole, the main component of eucalyptus oil (Peana et al., 2003).

Immune system boost III: antioxidant effects and radical scavenging

An imbalance in the rate of production of free radicals or removal by the antioxidant defense mechanisms leads to a phenomenon referred to as oxidative stress. A mixture of Oregano (carvacrol, cinnamaldehyde, and capsicum oleoresin) was found to beneficially affect the intestinal microflora, absorption, digestion, weight gain and also to have an antioxidant effect on chickens (Bassett, 2000).

Zeng et al. (2015) indicated the positive effect of essential oils on the production of digestive secretions and nutrient absorption. They reduce pathogenic stress in the gut, exert antioxidant properties, and reinforce the animal’s immune status.

Inside the cell, essential oils can serve as powerful scavenger preventing mutations and oxidation (Bakkali et al., 2008). Studies have demonstrated the concentration-dependent free radical scavenging ability of oils from eucalyptus species (Kaur et al., 2010; Marzoug et al., 2011; Olayinka et al., 2012). Some authors attribute the strong antioxidant capacity of essential oils to their phenolic constituents and synergistic effect between tannins, rutin, thymol, and carvacrol, and probably 1, 8-cineole. Moderate DPPH radical scavenging activity reported by Edris(2007), El-Moein et al. (2012), and Kaur et al. (2011).

Vázquez et al. (2012) have demonstrated the potential of the phenolic compounds in eucalyptus bark as a source of antioxidant compounds. The study showed that eucalyptus had ferric reducing antioxidant power in the ranges 0.91 to 2.58 g gallic acid equivalent (GAE) per 100 g oven-dried bark and 4.70 to 11.96 mmol ascorbic acid equivalent (AAE) per 100 g oven-dried bark, respectively (see also Shahwar et al., 2012). Moreover, Eyles et al. (2004) were able to show superoxide dismutase (SOD)-like activity for different compounds and fractions isolated from wood extracts.

Last but not least: positive effects on the respiratory system

In poultry production houses, especially in summer, high temperatures and low humidity increase the amount of air dust. Under such conditions, respiratory tract disorders in broiler chickens, including the deposition of particulates, become more frequent and more severe.

Clinical signs of respiratory disease in chickens include coughing, sneezing, and rales

Thyme oil, thanks to the phytomolecules thymol and carvacrol, supports the treatment of respiratory disorders. These substances smooth tightened muscles and stimulate the respiratory system. An additional advantage lies in their expectorant and spasmolytic properties (Edris, 2007).

These properties are also seen in essential oils such as eucalyptus and peppermint, which contain eucalyptol and menthol. They thin out the mucus and facilitate its removal from the airways. As a result, the airways are cleared and breathing during inflammation becomes easier (Durmic and Blache, 2012).

Another positive effect of the terpenoid compounds used in commercial preparations for poultry is that they disinfect the bronchi, preventing respiratory infections (Awaad et al., 2010; Barbour et al., 2011; Mahboubi et al., 2013). Barbour and Danker (2005) reported that the essential oils of eucalyptus and peppermint improved the homogeneity of immune responses and performance in MG/H9N2-infected broilers.

Grippozon: the phytogenic solution for respiratory health

Grippozon is a liquid composition with a high content of essential oils, which are combined to systematically prevent and ease respiratory diseases. The formulation is derived from the research on essential oils’ effectiveness against respiratory pathogens that are common in animal farming. Grippozon exhibits a synergistic action of all its components to optimally support animal health. It contains a high concentration of active components; both their quantity and quality are guaranteed to deliver results.

Application of Grippozon

Grippozon application can be flexibly adapted to most common housing systems. It is fully water-soluble for use in the drinking line and it is also possible to nebulize a diluted solution in air.

The dose recommendation in drinking water usually amounts to 100ml to 200ml per 1000 liters of drinking water (Grippozon administration has not been reported to affect water consumption). The active substances in Grippozon adhere to mouth mucosa and become volatile in the breathing air later on. Therefore Grippozon can enter the respiratory system indirectly as well. The volatile compounds also spread into the whole barn air and, thus, indirectly via breathing into the respiratory system (and farmers notice the smell of essential oils when Grippozon is applied through in the waterline)

Grippozon can also be used as a spray at a rate of 200ml/10 liters of water for 2000 birds, twice daily on 2-3 days a week. This produces a very effective nebulization effect and offers faster respiratory relief to birds.

Grippozon is an impactful tool for managing respiratory problems. Thanks to its effective mucolytic and relaxant activity, Grippozon gives symptomatic relief to the birds during high-stress periods of respiratory diseases. Mucus in the trachea works as media for the proliferation of bacteria and viruses, so by thinning the mucus, Grippozon slows down the proliferation of bacteria and the spread of disease. Grippozon helps in improving air quality and air intake. It can also be used to stimulate the immune response during vaccination.

Authors:
Ruturaj Patil – Product Manager Phytogenic Liquids
Kowsigaraj Palanisamy – Global Validation Trial Manager

References available on request

Author: Sabria Regragui Mazili, Science Communication Team, EW Nutrition

As people across the globe are re-discovering home cooking, grocery items such as milk, flour, and eggs are in higher demand than ever. Legendary chef Michel Roux once said that “Eggs are more than an element of cooking… The little egg is life.” Let us look at the complex system that allows us to enjoy this superfood – and how egg producers tackle the supply chain disruptions caused by the pandemic.

The coronavirus pandemic has led to increased demand for eggs and, therefore, to temporary shortages

Eggs: from farm to table

The story of our breakfast eggs starts with another set of eggs. The hens and cocks on so-called parent stock or layer breeder farms produce fertilized eggs (1). These “hatching eggs” are transferred to specialized hatcheries where the chicks hatch after 21 days (2). The female chicks, who will go on to become the hens that lay our eggs, are sent to so-called rearing or pullet farms, where they grow up (3). The young hens are then transferred to the layer farms (4).

When they are about 20 weeks old, the hens start to lay – and join the 7 billion chickens that produce more than 1.3 trillion eggs for us every year. During their 14-month laying period, hens will lay 360+ eggs.

Newly laid eggs need to be collected, and checked for quality (grading). About 50-65% of the hens’ output can be sold as “shell eggs,” i.e. whole eggs, to outlets servicing the hospitality industry and end consumers (5). These eggs need to be stamped and then packaged in the egg cartons we all know. “Breaker eggs,” i.e. irregularly sized or broken eggs, do not go to waste: they are moved to processing plants that produce liquid egg and egg powder products for food industry clients such as industrial bakeries and ice cream manufacturers (6).

How many steps it ultimately takes to get eggs from the farm to your plate depends on the complexity of the marketing channel. Farms might do their branding on-site; others work with specialized companies for packaging and branding. Some farms have direct relationships with retailers; others sell to wholesale outlets and distributors. The logistics for getting eggs quickly and safely from A to B involve dedicated egg distribution centers and fleets of refrigerated trucks.

Did you know?

An egg’s color depends on the layer hen’s breed. In the beginning, all eggs are white; those that end up being brown (or, in the case of certain chicken breeds, green or blue) have pigments deposited on them as the eggs pass through the hen’s oviduct.
The reason brown eggs are usually more expensive than white ones is that brown egg layer breeds tend to be heavier and consumer more feed. The eggshell color does not influence the egg’s taste or nutritional value.

Infographic: The egg supply chain, from farm to table

Coronavirus and food: disruptive effects on supply

The impact of the COVID-19 pandemic is felt along each step of this complex process. Disruptions in the supply chain for feed ingredients that mostly come from China, such as amino acids and vitamins, has led to increased price volatility of these ingredients, affecting farmers’ production costs.

Labor availability is a significant concern, for example with respect to the numerous transitions between locations. According to a recent Rabobank report on the situation in Europe, even though food transportation services are exempted from lockdown measures, the high incidence of COVID-19 cases are contributing to driver shortages and higher transport costs. Importantly, everyone working on the “front line” of food supply chains – drivers, as well as farmers, processors, distributors, or retail employees – are doing so at considerable personal risk.

… and effects on demand

Despite these disruptions, animal producers are under pressure to increase production as food consumption patterns are changing in light of COVID-19. In Argentina, for example, the national egg producers association just reported that household consumption of eggs rose by 40% since the country went into lockdown on March 20.

In the US, egg orders from some retailers have increased by up to six times their normal levels, according to market research firm Urner Barry, while wholesale egg prices have risen 180% throughout March. Yet, hens can’t lay more than they already do; to increase flocks takes time and significant investment in additional facilities.

To ease egg supply shortages, the US Food and Drug Administration has temporarily relaxed certain packaging and labeling requirements for shell eggs sold in retail markets. In Canada, egg supplies usually destined for restaurants are being re-directed to meet increased consumer demand.

Egg producers are finding solutions

This would not work in Germany, where, according to egg and poultry analyst Margit Beck, many of the eggs used in the ailing foodservice sector are cage eggs*, for which there is little end consumer demand. Cage egg prices are effectively in free fall, while crisis-induced consumer reflections on purchasing behaviors appear to strengthen demand for free-range and organic eggs. Smaller farmers in Germany and Austria report that they capitalize on this trend through increased direct sales to customers at farm shops and markets.

The sight of (temporarily) empty supermarket shelves might be disconcerting – but, positively, it ought to inspire us to appreciate the incredible work that goes into putting eggs on our tables. Clapping for health care workers has become a new tradition during this pandemic. It would only be fitting to also give a hand for the numerous people along food supply chains whose commitment keeps us all going.

* Note: Conventional battery cages have been banned in Germany since 2010; eggs classified as cage eggs come from laying hens kept in small-group housing systems or in so-called “enriched cages”.

How lipopolysaccharides cause disease

LPS are rather large and structured chemical molecules with a weight of over 100,000 D. They are highly thermostable; boiling in water at 100°C for 30 minutes does not destabilize their structure. LPS consist of three chemically distinct sections: a) the innermost part, lipid A, consisting mostly of fatty acids; b) the core, which contains an oligosaccharide; and c) the outer section, a chain of polysaccharides called O-antigen (Figure 1).

Figure 1: Structure of an LPS

The toxicity of LPS is mainly caused by lipid A; however, both lipid A and O-antigen stimulate the immune system. This happens when the LPS pass the mucosa and enter the bloodstream or when they attack the leukocytes.

The intestinal mucosa is the physical immune barrier that protects the microvilli from external agents (bacteria, free LPS viruses, etc.). Despite its strength (the thickness, for example, amounts to ≈830 µm in the colon and ≈123 µm in the jejunum), vulnerable points exist (cf. Zachary 2017).

LPS can easily come into contact with the cells of the lamina propria (a layer of connective tissue underneath the epithelium) through the microfold (M) cells of the Peyer’s patches (which consist of gut-associated lymphoid tissue). The M cells are not covered by mucus and thus exposed.

Secondly, LPS can also pass through the mucosa, where they become entangled in this gelatinous structure. There, they come into contact with the lymphocytes or can reach the regional lymph nodes through the afferent lymphatic vessels.

Thirdly, LPS might affect the tight junctions, the multiprotein complexes that keep the enterocytes (cells that form the intestinal villi) cohesive. By destabilizing the protein structures and triggering enzymatic reactions that chemically degrade them, LPS can break the tight junctions, reaching the first capillaries and, consequently, the bloodstream.

The presence of endotoxins in the blood, endotoxemia, can trigger problematic immune responses in animals. An innate immune stimulation leads to an increase in the concentration of pro-inflammatory cytokines in the blood and, consequently, to an induced febrile response in the animal: heat production increases, while the available metabolic energy decreases.  As a result, performance suffers, and in the worst-case scenario, septic shock sets in. Furthermore, when LPS compromise intestinal integrity, the risk of secondary infections increases, and production performance may decline.

LPS’ modes of action

How does all of this happen? The physiological consequences of endotoxemia are quite complex. Simplified, the immune system response to LPS in the blood takes three forms:

• The stimulation of TLR4 (toll-like receptor 4) induces monocytes and macrophages to secrete critical pro-inflammatory cytokines, primarily interleukin (IL) IL-1β, IL-6, IL-8, and tumor necrotic factor (TNF) α and β. TLR4 is a structure on the cell membrane of mainly macrophages and leukocytes, which is activated by the LPS-binding protein (LBP).
• The complement cascade constitutes about 10% of plasma proteins and determines the chemotaxis and activation of leukocytes. It can form a membrane attack complex (MAC), which perforates the membranes of pathogenic cells, enabling lysis.
• The Hagemann factor, also known as coagulation factor XII: once stimulated by LPS, it initiates the formation of fibrin (through the intrinsic coagulation pathway), which might lead to thrombosis. The Hagemann factor directly stimulates the transformation of prekallikrein to kallikrein (enzymes involved in regulating blood pressure).

Figure 2: How LPS leads to endotoxemia – 3 modes of action

These three modes of action of inflammatory stimulation lead to important physiological reactions:

• Pro-inflammatory cytokines (see above) modulate the functional expression of other immune cell types during the inflammatory response;
• Metabolites of arachidonic acid (prostaglandins, leukotrienes, and lipoxins), intra- and extracellular messengers that influence the coagulation cascade;
• Synthesis in the blood of bradykinin, a peptide responsible for the typical symptoms of inflammation, such as swelling, redness, heat and pain;
• PAF (platelet-activating factor), which creates inflammatory effects through narrowing of the blood vessels and constriction of the airways, but also through the degranulation of leukocytes.

The symptoms of endotoxemia are:  hypotension, metabolic acidosis, hemoconcentration, intestinal hemorrhage, fever, activations of neutrophils and endothelial cells, and predisposition to thrombosis.

In case of a progression to septic shock, the following sequence takes place:

1) Reduction in blood pressure and increased heart rate (hemodynamic alterations)

2) Abnormalities in body temperature

3) Progressive hypoperfusion at the level of the microvascular system

4) Hypoxic damage to susceptible cells

Up to here, symptoms follow a (severe) endotoxemia pathogenesis. A septic shock furthermore entails:

5) Quantitative changes in blood levels of leukocytes and platelets

6) Disseminated intravascular coagulation (see Hageman factor)

7) Multi-organ failure

8) Death of animal

If an animal is continously challenged with endotoxins, experiences septic shock, or comes close to it, it risks developing LPS tolerance, also known as CARS (compensatory anti-inflammatory response syndrome). This syndrome essentially depresses the immune system to control its activity. The anti-inflammatory prerogative of CARS is not to interfere directly with the elimination of pathogens but to regulate the “excessive” inflammatory reaction in a hemostatic way. However, this regulation can be extremely dangerous as the syndrome involves a lack of homeostasis control, and an excessive depression of the immune system leaves the organism exposed to the actual pathogens.

Farm animal research on endotoxemia pathogenesis

Lipopolysaccharides are difficult to quantify in the intestine of a live animal. One way to evaluate a possible endotoxemia is to analyze biomarkers present in the bloodstream. The most important one is the LPS themselves, which can be detected in a blood sample taken from the animal via ELISA. Other biomarkers include pro-inflammatory interleukins, such as TNF α and β, IL-6 or IL-8, and fibrin and fibrinogen (though they are not specific to endotoxemia). It is vital to carry out a blood sample analysis to deduce a possible endotoxemia from symptoms and performance losses in the animal.

How the metabolic effects of endotoxemia depress performance

One of the biggest issues caused by endotoxemia is that animals reduce their feed intake and show a poor feed conversion rate (FCR). Why does this happen? The productive performance of farm animals (producing milk, eggs, or meat) requires energy. An animal also requires a certain baseline amount of energy for maintenance, that is, for all activities related to its survival. As a result of inflammation and all those physiological reactions mentioned above, endotoxemia leads to a feverish state. Maintenance needs to continue; hence, the energy required for producing heat will be diverted from the energy usually spent on producing milk, egg, meat, etc., and performance suffers.

The inflammation response can result in mitochondrial injury to the intestinal cells, which alter the cellular energy metabolism. This is reflected in changes to the levels in adenosine triphosphate (ATP), the energy “currency” of living cells. A study by Li et al. (2015) observed a respective reduction of 15% and 55% in the ATP levels of the jejunum and ileum of LPS-challenged broilers, compared to the unchallenged control group. This illustrates the extent to which animals lose energy while they experience (more or less severe) endotoxemia.

Figure 3: Reduction in ATP level in Jejunum and Ileum in broilers (adapted from Li et al., 2015)

A piglet study by Huntley, Nyachoti, and Patience (2017) took this idea further (Figure 4):  3 groups of 10 Yorkshire x Landrace pigs, weighing between 11 and 25 kg, were studied in metabolic cages and in respiratory chambers. This methodology allows for simultaneous measurement of oxygen consumption, CO₂ production, energy expenditure, physical activity, and feed/water intake. The study found that LPS-challenged pigs retained 15% less of the available metabolizable energy and showed 25% less nutrient deposition. These results show concrete metabolic consequences caused by the febrile response to endotoxemia we discussed above.

Figure 4: Retained Energy as % of ME intake and nutrient deposition of pigs in metabolic cages (adapted from Huntley, Nyachoti, and Patience, 2017)

Control treatment (CON) = Pigs fed by a basal diet
Immune system stimulation treatment (ISS) = Pigs given LPS (E. coli serotype 055:B5) injection

A loss of energy retained due to a reduction in available metabolizable energy leads to losses in performance as the amount of energy available for muscle production and fat storage will be lower. Furthermore, the decrease in feed intake creates a further energy deficit concerning production needs.

A trial carried out at the University of Illinois examined the effects of repeated injections of 400 μg E. coli LPS on chick performance from 11 to 22 days after hatching. The chicks were fed casein-based diets with graded levels of arginine. LPS administration reduced weight gain (P<0.05) and feed intake, and these effects tended to be worse at higher levels of arginine supplementation (Figure 5). The researchers hypothesize that, in response to endotoxin and elevated cytokine levels, macrophages use more arginine to produce nitric oxide, diverting it from protein production for muscle development.

Figure 5: Effects of LPS on feed intake and body weight gain in chicks fed graded level of arginine (based on Webel, Johnson, and Baker, 1998)

NC = negative control

This data on poultry complements the results for swine, again showing that endotoxin-induced energy losses quantifiably depress animal performance even in milder disease cases.

The way forward: Endotoxin mitigation

Animals suffering from endotoxemia are subject to severe metabolic dysfunctions. If they do not perish from septic shock, they are still likely to show performance losses. Moreover, they at great risk of immunosuppression caused by the immune system “overdrive.” Effective endotoxin mitigating agents can help to prevent these scenarios.

EW Nutrition’s Mastersorb Gold is not only a leading anti-mycotoxin agent; thanks to its specific components, it effectively binds bacterial toxins. An in vitro study conducted at the Hogeschool Utrecht laboratory (part of Utrecht University) evaluated the binding capacity of Mastersorb Gold on LPS compared to three different competitor products. All products were tested at two different inclusion rates. At an inclusion rate of 0.25%, only Mastersorb Gold reduced the toxin load on the solution by 37%. At 1% inclusion, Mastersorb Gold bound 75% of the toxin, while only one competitor product demonstrated any binding (10%).

Lipopolysaccharides are a constant challenge for animal production. The quantity of Gram-negative bacteria in an animal intestine is considerable; therefore, the danger of immune system over-stimulation through endotoxins cannot be taken lightly. Producers need to prioritize the maintenance of intestinal eubiosis in production animals proactively; for instance, through targeted gut health-enhancing additives based on phytomolecules and, possibly, organic acids.

Most importantly, the detrimental impact of LPS can be mitigated by using a high-performance agent such as Mastersorb Gold. To limit losses from an energy point of view yields positive results in terms of production levels and the prevention of secondary infections, preserving animal health and farms’ economic viability.

By Claudio Campanelli, EW Nutrition

References

Adib-Conquy, Minou, and Jean-Marc Cavaillon. “Compensatory Anti-Inflammatory Response Syndrome.” Thrombosis and Haemostasis 101, no. 01 (2009): 36–47. https://doi.org/10.1160/th08-07-0421.

Huntley, Nichole F., C. Martin Nyachoti, and John F. Patience. “Immune System Stimulation Increases Nursery Pig Maintenance Energy Requirements.” Iowa State University Animal Industry Report 14, no. 1 (2017). https://doi.org/10.31274/ans_air-180814-344.

Li, Jiaolong, Yongqing Hou, Dan Yi, Jun Zhang, Lei Wang, Hongyi Qiu, Binying Ding, and Joshua Gong. “Effects of Tributyrin on Intestinal Energy Status, Antioxidative Capacity and Immune Response to Lipopolysaccharide Challenge in Broilers.” Asian-Australasian Journal of Animal Sciences 28, no. 12 (2015): 1784–93. https://doi.org/10.5713/ajas.15.0286.

Mani, Venkatesh, James H Hollis, and Nicholas K Gabler. “Dietary Oil Composition Differentially Modulates Intestinal Endotoxin Transport and Postprandial Endotoxemia.” Nutrition & Metabolism 10, no. 1 (2013): 6. https://doi.org/10.1186/1743-7075-10-6.

Webel, D.M., R.W. Johnson, and D.H. Baker. “Lipopolysaccharide-Induced Reductions in Body Weight Gain and Feed Intake Do Not Reduce the Efficiency of Arginine Utilization for Whole-Body Protein Accretion in the Chick.” Poultry Science 77, no. 12 (1998): 1893–98. https://doi.org/10.1093/ps/77.12.1893.

Zachary, James F. “Chapter 4 – Mechanisms of Microbial Infections.” Essay. In Pathologic Basis of Veterinary Disease, 132–241. St Louis, MO: Mosby, 2017. https://doi.org/10.1016/B978-0-323-35775-3.00004-7.

The large majority of poultry specialists in Europe consider phytomolecules as one of the key elements in diets for broilers, broiler breeders, and layers when birds are raised without antibiotics. A quick glance at the market will reveal more commercial products than can possibly be imagined. There are three basic elements you should bear in mind when making your choice:

1. Most phytomolecules are volatile. As such, unprotected products will soon evaporate if left exposed to the open air – as it happens, for instance, with feed prepared in commercial farms. Microencapsulation is therefore essential.
2. There are countless phytomolecules. Consequently, finding the right mix for the task required is essential, as not all mixtures will get you the desired result. When designing a phytomolecule mix, the manufacturer must have the necessary knowledge and experience to achieve the desired result.
3. Phytomolecules are powerful. This is to say that you cannot just keep adding higher quantities to achieve a better result. Finding the exact inclusion rates for the right purpose is a difficult balancing exercise.

In fact, the right protection, the right mix and the right inclusion rates must be combined to ensure that the animals do not refuse the feed (worst case scenario) or just fail to benefit from the inclusion of phytomolecules.

Among the feed additives, phytomolecules (or secondary plant compounds) stand out as a class of active ingredients that may help to improve gut health and thereby reduce the use of antibiotics.  Synthesized by plants as a defense mechanism against pathogens, phytomolecules promote the digestion of feed ingredients (Zhai et al. 2018), prevent loss of gut integrity during enteric challenges (Liu et al. 2018), and have antimicrobial properties that hinder the growth of potential pathogens (Chowdhury, 2018). Phytomolecules can prevent the overgrowth of opportunistic pathogens, thereby reducing the frequency of occurrence of diseases such as necrotic enteritis and dysbacteriosis and thus improve performance data such as daily weight gain and feed efficiency.

Beyond the phytomolecules’ proven effects, what works best in supporting the health and welfare of your animals is, in fact, a holistic program (such as those offered by EW Nutrition) that consists of an effective combination of innovative products and consultancy services in the fields of gut health, nutrition, AMR monitoring, and biosecurity management.

*This article is available in Dutch.

by Ajay Bhoyar, Global Technical Manager, EW Nutrition

Anyone working with today’s fast-growing broiler chicken knows that it is a sensitive creature – and so is its gut health. Thanks to continuous improvements in terms of genetics and breeding, nutrition and feeding, as well as general management strategies, broiler production has tremendously upped performance and efficiency over the past decades. It is estimated that, between 1957 and 2005, the broiler growth rate increased by over 400%, while the feed conversion ratio dropped by 50%.

These impressive improvements, however, have come at the cost of intense pressure on the birds’ digestive system, which needs to process large quantities of feed in little time. To achieve optimal growth, a broiler’s gastrointestinal tract (GIT) needs to be in perfect health, all the time. Unsurprisingly, enteric diseases such as necrotic enteritis, which severely damages the intestinal mucosa, hamper the intestines’ capacity to absorb nutrients and induce an inflammatory immune response.

The modern broiler’s gut – a high-performing, but sensitive system

However, in a system as high performing as the modern broiler’s GIT, much less can lead to problems. From when they are day-old chicks up to slaughter, broilers go through several challenging phases during which they are more likely to show impaired gut functionality, e.g. after vaccinations or feed changes. Good management practices go a long way towards eliminating unnecessary stressors for the animals, but some challenging periods are unavoidable.

The transition from starter to grower diets is a classic situation when nutrients are very likely to not be well digested and build up in the gut, fueling the proliferation of harmful microbes. Immunosuppressive stress in combination with an immature intestinal microflora results in disturbances to the bacterial microbiota. At “best”, this entails temporarily reduce nutrient absorption, in the worst case the birds will suffer serious intestinal diseases.

Phytomolecules – the intelligent alternative to antibiotics

To safeguard performance during stressful periods, poultry producers need to anticipate them and proactively provide effective gut health support. For many years, this support came in the form of antibiotic growth promoters (AGP): administered prophylactically, they were effective at keeping harmful enteric bacteria in check. However, due to grave concerns about the development of antimicrobial resistance, non-therapeutic antibiotics use has been banned in many countries. Alternatives need to focus on improving feed digestibility and strengthening gut health, attacking the root causes of why the intestinal microflora would become unbalanced in the first place.

Phytomolecules are secondary metabolites active in the defense mechanisms of plants. Studies have found that certain phytomolecules stimulate digestive enzyme activities and stabilize the gut microflora, “leading to improved feed utilization and less exposure to growth-depressing disorders associated with digestion and metabolism” (Zhai et al., 2018). With other trials showing positive effects on broilers’ growth performance and feed conversion, the research indicates that phytomolecules might also specifically support chickens during challenging phases.

The effect of phytomolecules on broilers during a challenging phase

A study was conducted over a period of 49 days on a commercial broiler farm of an AGP-free integration operation in Japan. The farm reported gut health challenges in the second and third week of the fattening period due to vaccinations and changes to the animals’ diets. The trial included 15504 Ross 308 broilers, divided into two groups. The negative control group included a total of 7242 birds, kept in another house.

All the birds were fed the standard feed of the farm. The trial group (8262 birds) received Activo Liquid, which contains a synergistic combination of phytomolecules, administered directly through the drinking water. Activo Liquid was given at an inclusion rate of 200ml per 1000L of water (3.3 US fl oz per gallon of stock solution, diluted at 1:128), from day 8 until day 25, for 8 hours a day.

The results are summarized in Figure 1:

Figure 1: Improved broiler performance for Activo Liquid group (day 49)

The Activo Liquid group clearly showed performance improvements compared to the control group. Livability augmented by 1.5%, while the feed conversion rate improved by 3.2%. This resulted in a more than 5% higher score in terms of the performance index.

Challenging times? Tackle them using phytomolecules

Poultry producers take great care to eliminate unnecessary sources of stress for their birds. Nonetheless, during their lifecycle, broiler chickens face challenging periods during which the balance of the intestinal microflora can easily become disturbed, with consequences ranging from decreased nutrient absorption to full-blown enteric disease.

The trial reviewed here showed that, after receiving Activo Liquid, broilers raised without AGPs showed encouraging performance improvements during a challenging phase of feed changes and vaccinations. Likely thanks to the activation of digestive enzymes and a stabilization of the gut flora, the broilers showed improved livability and feed conversion, thus delivering a much more robust performance during a critical phase of their lives. In times where the non-therapeutic use of antibiotics is no longer an option, phytomolecules allow poultry farmers to effectively support their animals during challenging times.

References

Photo Source: Aviagen

Adedokun, Sunday A., and Opeyemi C. Olojede. “Optimizing Gastrointestinal Integrity in Poultry: The Role of Nutrients and Feed Additives.” Frontiers in Veterinary Science 5 (January 31, 2019): 348.

Jamroz, D., T. Wertelecki, M. Houszka, and C. Kamel. “Influence of Diet Type on the Inclusion of Plant Origin Active Substances on Morphological and Histochemical Characteristics of the Stomach and Jejunum Walls in Chicken.” Journal of Animal Physiology and Animal Nutrition 90, no. 5-6 (March 23, 2006): 255–68. 

Tavárez, Marcos A., and Fausto Solis De Los Santos. “Impact of Genetics and Breeding on Broiler Production Performance: a Look into the Past, Present, and Future of the Industry.” Animal Frontiers 6, no. 4 (October 1, 2016): 37–41.

Zhai, Hengxiao, Hong Liu, Shikui Wang, Jinlong Wu, and Anna-Maria Kluenter. “Potential of Essential Oils for Poultry and Pigs.” Animal Nutrition 4, no. 2 (June 2018): 179–86.

Zuidhof, M. J., B. L. Schneider, V. L. Carney, D. R. Korver, and F. E. Robinson. “Growth, Efficiency, and Yield of Commercial Broilers from 1957, 1978, and 20051.” Poultry Science 93, no. 12 (December 2014): 2970–82. 

Diseases caused by E. coli entail use of antibiotics in animal production

E. coli infections are a major problem in pig production. Especially young animals with an incompletely developed immune system are often unable to cope with the cavalcade of pathogens. In poultry, E. coli are responsible for oedema, but also for respiratory diseases. In young piglets, E. coli cause diarrhoea , oedema, endotoxic shock and death. In order to cure the animals, antibiotics often must be applied. Besides this curative application, antibiotics were and in many countries still are used prophylactically and as growth promoters.

The excessive use of antibiotics, however, leads to the occurrence of antimicrobial resistance (AMR): due to mutations, resistance genes are created which enable enterobacteria such as Salmonella, Klebsiella and E. coli to produce enzymes (ß-lactamases) in order to withstand ß-lactam antibiotics. In case of an antibiotic treatment, the resistant bacteria survive whereas the other bacteria die.
The major problem here is that these resistance genes can be transferred to other bacteria. Harmless bacteria can thus transfer resistance genes to dangerous pathogens, which then cannot be combatted with antibiotics anymore. In this article we explore in detail how AMR happens and how phytomolecules, which have antimicrobial properties, could be a key tool to reduce the need for antibiotics in animal production.

How ß-lactam antibiotics work

The group of ß-lactam antibiotics consists of penicillins, cephalosporins,  monobactams, and carbapenems. These antibiotics are characterised by their lactam ring (Figure 1).

Figure 1: An antibiotic with a ß-lactam ring (in orange)

If bacteria are growing, the cell wall also has to grow. For this purpose existing conjunctions are cracked and new components are inserted. In order for the cell wall to remain a solid barrier, the new components must be interconnected by crosslinks. For the creation of these crosslinks an enzyme is essential, the transpeptidase (figure 2).

Figure 2: building up a stable cell wall with the help of transpeptidase

Due to their structure, ß-lactam-antibiotics also fit as binding partner for transpeptidase. They bind to the enzyme and block it (Kohanski et al., 2010). The crosslinks cannot be created and the stabilization of the cell wall is prevented. Disturbance of cell wall stability leads to the death of the bacterial cell, hence ß-lactam antibiotics act bactericidal.

Figure 3: blocked by ß-lactam antibiotics, transpeptidase cannot serve as enzyme for building the cell wall

The challenge: E. coli producing ß-lactamases

Resistant bacteria, which are able to produce ß-lactamases – enzymes that destroy the ß-lactam ring – prevent their own destruction. Divers point mutations within the ß-lactamase genes lead to the occurrence of “extended-spectrum-beta-lactamases“ (ESBL). ESBL are able to inactivate most of the ß-Lactam-antibiotics.

Another mutation leads to so-called AmpC (aminopenicillin and cephalosporin) ß-lactamases. They enable the E. coli to express a resistance against penicillins, cephalosporins of the second and third generation as well as against cephamycins.

Phytomolecules – an alternative?

One approach to reduce the use of antibiotics is the utilization of phytomolecules. These secondary metabolites are produced by plants to protect themselves from moulds, yeasts, bacteria and other harmful organisms.

The use of plants and their extracts in human and veterinary medicine is well-established for centuries. Besides digestive and antioxidant characteristics they are well known for their bacteriostatic and bactericidal effects.

Consisting of a high number of chemical compounds, they attack at diverse points and their antimicrobial effect is not caused by only one single specific mechanism. This is crucial because it is therefore very unlikely that bacteria can develop resistances to phytomolecules like they do to antibiotics.

How phytomolecules work

Mostly, phytomolecules act at the cell wall and the cytoplasm membrane level. Sometimes they change the whole morphology of the cell. This mode of action has been studied extensively for thymol and carvacrol, the major components of the oils of thyme and oregano.

They are able to incorporate into the bacterial membrane and to disrupt its integrity. This increases the permeability of the cell membrane for ions and other small molecules such as the energy carrier ATP (Adenosin-tri-phosphate). It leads to the decrease of the electrochemical gradient above the cell membrane and to the loss of energy equivalents of the cell.

A special challenge: gram-negative bacteria

Gram-negative bacteria such as E. coli and Salmonella pose a special challenge. The presence of lipopolysaccharides in the outer membrane (OM) provides the gram-negative bacteria with a hydrophilic surface (Nikaido, 2003; Nazarro et al., 2013) (see also blue infobox).

The cell wall therefore only allows the passage of small hydrophilic solutes and is a barrier against macromolecules and hydrophobic compounds such as hydrophobic antibiotics and toxic drugs. The bypassing of the OM therefore is a prerequisite for any solute to exert bactericidal activity toward gram-negative bacteria (Helander et al., 1998).

Based on their trial results Helander et al. (1998)  (1998) concluded that trans-cinnamaldehyde and partly also thymol and carvacrol gain access to the periplasm and to the deeper parts of the cell. Nikaido (1996) also concluded that OM-traversing porin proteins allow the penetration of lipophilic probes at significant rates.

Evaluating phytomolecules I – in vitro trial, Scotland

A trial conducted in Scotland evaluated the effects of Activo Liquid, a mixture of selected phytomolecules and citric acid,  on ESBL-producing E. coli as well as on E. coli that generate AmpC.

Material and methods

For the trial two strains for each group were isolated from the field, a non-resistant strain of E. coli served as control. Suspensions of the strains with 1×10⁴ CFU/ml were incubated for 6-7 h at 37°C (98.6°F) together with diverse concentrations of Activo Liquid or with cefotaxime, a cephalosporin. The cefotaxime group saved as a control for differentiating resistant and non-resistant E. coli.

The suspensions were put on LB agar plates and bacteria colonies were counted after further 18-22h incubation at 37°C.

Results

The antimicrobial efficacy of the blend of phytomolecules depended on the concentration at which they were used (see table 1). A bacteriostatic effect could be shown at dilutions up to 0.1 %, a bactericidal effect at higher concentrations.

Table 1: Effect of phytomolecules against resistant E. coli producing ESBL and AmpC in poultry

Evaluating phytomolecules II – in vitro trial, Germany

A further trial was conducted in Germany (Vaxxinova, Münster), confirming the preceding results.

Material and methods

Four ESBL producing E. coli all isolated from farms and a non-resistant reference strain as control were tested concerning their sensitivity against Activo Liquid. Every bacteria strain (Conc.:1×10⁴ CFU/ml) was subjected to a bacterial inhibition assay in an appropriate medium at 37°C for 6-7 hours.

Results

In this trial Activo Liquid also showed a dose-dependent efficacy, with no or just a bacteriostatic effect up to a concentration of 0.1 %, but bactericidal effects at a concentration of ≥0.2 % (table 2).

Table 2: Effect of phytomolecules against resistant ESBL producing E. coli in pig and in poultry

Phytomolecules: a promising outlook

E. coli infections have devastating effects on animals, from diarrhea to edema, enterotoxic shock and even death. Antibiotic treatments have long been the only practicable answer. However, their excessive use ̶ for instance, the metaphylactic application to thousands of animals in a flock ̶ has led to the development of resistant strains. There is evidence that a reduction of antibiotic use reduces the occurrence of resistances (Dutil et al., 2010).

The results of the two in vitro trials in Scotland and Germany demonstrate the bactericidal effects of phytomolecules on E. coli that produce ESBL and AmpC. Using phytomolecules could thus reduce the use of antibiotics and therefore also the occurrence of AMR.

While it is theoretically possible for bacteria to also become resistant against phytomolecules, the probability of this happening is very low: unlike antibiotics, phytomolecules contain hundreds of chemical components with different modes of action. This makes it exceedingly difficult for bacteria to adapt and develop resistance. To tackle the problem of antibiotic-resistant E. coli, antimicrobial phytomolecules therefore offer a promising, sustainable and long-term solution.

By Dr. Inge Heinzl, Editor, EW Nutrition

Literature

Dutil, Lucie, Rebecca Irwin, Rita Finley, Lai King Ng, Brent Avery, Patrick Boerlin, Anne-Marie Bourgault, Linda Cole, Danielle Daignault, Andrea Desruisseau, Walter Demczuk, Linda Hoang, Greg B. Horsman, Johanne Ismail, Frances Jamieson, Anne Maki, Ana Pacagnella, and Dylan R. Pillai. 2010.” Ceftiofur Resistance in Salmonella enterica Serovar Heidelberg from Chicken Meat and Humans, Canada.” Emerg Infect Dis 16 (1): 48-54.

Helander, Ilkka M., Hanna-Leena Alakomi, Kyösti Latva-Kala, Tiina Mattila-Sandholm, Irene Pol, Eddy J. Smid, Leon G. M. Gorris, and Atte von Wright. 1998. “Characterization of the Action of Selected Essential Oil Components on Gram-Negative Bacteria.” J. Agric. Food Chem 46: 3590-595.

Kohanski, Michael A., Daniel J. Dwyer, and James J. Collins. 2010. “How Antibiotics Kill Bacteria: From Targets to Networks.” Nature Reviews 8: 423-435.

Nazarro, Filomena, Florinda Fratianni, Laura De Martino, Raffaele Coppola, and Vincenzo De Feo. 2013. “Effect of Essential Oils on Pathogenic Bacteria.” Pharmaceuticals 6 (12): 1451-1474.

Nikaido, Hiroshi ” Molecular Basis of Bacterial Outer Membrane Permeability Revisited. 2003. ” Microbiology and Molecular Biology Reviews, 67 (4): 593-656.

Rodriguez, Tori. 2015 “Essential Oils Might Be the New Antibiotics.” The Atlantic.

http://www.theatlantic.com/health/archive/2015/01/the-new-antibiotics-might-be-essential-oils/384247/

Rüben, Christiane. 2009. “Antimikrobielle Wirksamkeit von chemischen Einzelkomponenten ätherischer Öle gegenüber ausgewählten Lebensmittelverderbniserregern“. PhD diss, TeHo Hannover.

Antimicrobial resistance (AMR) is a major threat to global public health. It is largely caused by the overuse of antibiotics in human medicine and agriculture. In intensive poultry production most antibiotics are used as antimicrobial growth promoters and/or used as prophylactic and metaphylactic treatments to healthy animals. Reducing such antibiotic interventions is crucial to lowering the incidence of AMR. However, antibiotic reduction often results in undesirable performance losses. Hence alternative solutions are needed to boost poultry performance. Phytomolecules have antimicrobial, digestive, anti-inflammatory and antioxidant properties, which could make them key to closing the performance gap.

Poultry performance depends on intestinal health

Poultry performance is to a large extent a function of intestinal health. The intestines process nutrients, electrolytes and water, produce mucin, secrete immunoglobulins and create a barrier against antigens and pathogens.

In addition, it is an important component of the body’s immune defense system. The intestine has to identify pathogens and reject them, but also has to tolerate harmless and beneficial microorganisms. If the intestines do not function properly this can lead to food intolerance, dysbiosis, infections and diseases. All of these are detrimental to feed conversion and therefore also to animal performance.

Antibiotics reduce the number of microorganisms in the intestinal tract. From a performance point of view this has two benefits: first, the number of pathogens is reduced and therefore also the likelihood of diseases; second, bacteria are eliminated as competitors for the available nutrients. However, the overuse of antibiotics not only engenders AMR: antibiotics also eliminate probiotic bacteria, which negatively impacts the digestive tracts’ microflora.

Products to boost poultry performance may be added to their feed or water. They range from pre- and probiotics to medium chain fatty acids and organic acids to plant extracts or phytomolecules. Especially the latter have the potential to substantially reduce the use of antibiotics in poultry farming.

Phytomolecules are promising tools for antibiotic reduction

Plants produce phytomolecules to fend off pathogens such as moulds, yeasts and bacteria. Their antimicrobial effect is achieved through a variety of complex mechanisms. Terpenoids and phenols, for example, disturb or destroy the pathogens’ cell wall. Other phytomolecules inhibit their growth by influencing their genetic material. Studies on broilers show that certain phytomolecules reduce the adhesion of pathogens such as to the wall of the intestine. Carvacrol and thymol were found to be effective against different species of Salmonella and Clostridium perfringens.

There is even evidence that secondary plant compounds also possess antimicrobial characteristics against antibiotic resistant pathogens. In-vitro trials with cinnamon oil, for example, showed antimicrobial effects against methicillin resistant Staphylococcus aureus, as well as against multiresistant E. coli, Klebsiella pneumoniae and Candida albicans.

Importantly, there are no known cases to date of bacteria developing resistances to phytomolecules. Moreover, phytomolecules increase the production and activity of digestive enzymes, they suppress the metabolism of pro-inflammatory prostaglandins and they act as antioxidants. Their properties thus make them a promising alternative to the non-therapeutic use of antibiotics.

Study design and results

In order to evaluate the effect of phytomolecules on poultry performance, multiple feeding studies were conducted on broilers and laying hens. They were given a phytogenic premix (Activo, EW Nutrition GmbH) that contains standardized  amounts of selected phytomolecules.

To achieve thermal stability during the feed processing and a targeted release in the birds’ gastrointestinal tract, the product is microencapsulated. For each , the studies evaluated both the tolerance of the premix and the efficacy of different dosages.

Study I: Evaluation of the dose dependent efficacy and tolerance of Activo for broilers
Animals:             400 broilers; age: 1-35 days of age
Feed:                  Basal starter and grower diets
Treatments:
– No supplement (negative control)
– 100 mg of Activo /kg of feed
– 1.000 mg of Activo /kg of feed
– 10.000 mg of Activo /kg of feed
Parameters:       weight gain, feed intake, feed conversion ratio, health status, and blood parameters

Results: The trial group given the diet supplemented with 100 mg/kg Activo showed significant improvements in body weight gain during the starter period (+4%) compared to the control group. Additional significant improvements in feed conversion ratio (FCR) in the growing period (+4%) resulted in an overall improvement in FCR of 3%. At a 1.000 mg/kg supplementation, a significant improvement in FCR of 6% was observed over the entire feeding period. Hematological parameters were within the reference range of healthy birds when feeding up to 10,000 Activo/ kg of feed.

Study II: Evaluation of the dose depending efficacy and tolerance of Activo for laying hens

Animals:             200 hens; age: 20 to 43 weeks
Feed:                  basal diet for laying hens
Treatments:
– No supplement (negative control)
– 100 mg of Activo/ kg of feed
– 250 mg of Activo/ kg of feed
– 500 mg of Activo/ kg of feed
– 5.000 mg of Activo/ kg of feed
Parameters:      weight gain, feed intake, feed conversion ratio, health status, and blood parameters

Results: Inclusion levels from 100 mg/kg of Activo onwards improved laying performance, egg mass and egg weight and reduced FCR compared to the control group. Results recorded for hematological parameters were within the reference range of healthy birds when feeding up to 5.000 mg Activo/ kg of feed.

Study III: Evaluation of the dose-dependent effects of Activo for coccidiosis vaccinated broilers

Animals:             960 broiler chickens; age: 42 days
Feed:                  Standard starter and finisher feed
Treatments:
– No supplement (negative control)
– 50 g of Activo /US ton of feed
– 100 g of Activo /US ton of feed
– 150 g of Activo /US ton of feed
– 200 g of Activo /US ton of feed
– 250 g of Activo /US ton of feed
– Antibiotic growth promoter (AGP)(positive control)
Parameters:      weight gain, feed efficiency
Specific:           In order to represent field conditions, the birds were challenged with used, homogenized litter.

Results: A clear dose response for both body weight gain and feed efficiency was observed (see Figure 1): the more phytogenic premix given, the better the birds’ performance. The group with 200g of Activo /US ton of feed showed similar performance levels than the positive control group supplemented with AGP.

Figure 1: Dose-dependent effects of for coccidiosis vaccinated broilers

Study IV:  Evaluation of the dose-dependent effects of Activo for laying hens

Animals:           40 hens; age: week 20 to 43
Feed:                basal diet for laying hens
Treatments:
– No supplement (negative control)
– 100 mg of Activo/ kg of feed
– 250 mg of Activo/ kg of feed
– 500 mg of Activo/ kg of feed
– 5.000 mg of Activo/ kg of feed
Parameters:      weight gain, feed intake, egg production, feed conversion ratio, health status
Duration:         168 days of feeding period

Results: The laying hens showed a higher laying rate when fed with a higher concentration of phytomolecules (Figure 2). Similarly improved results were observed for the feed efficiency. The more phytogenic premix added to their diet the better feed efficiency (Figure 3).

Figure 2: Dose-dependent effects of Activo on laying rate in laying hens

Figure 3: Dose-dependent effects of Activo on feed efficiency in laying hens

In conclusion, all four studies indicate that the inclusion of phytomolecules in broilers’ and laying hens’ diet improves their performance. Increasing levels of a phytogenic premix (Activo) significantly increased the production parameters for both groups. These improvements might bring performance in antibiotic-free poultry production on par with previous performance figures achieved with antimicrobial growth promoters.

The studies also showed that microencapsulated phytogenic premixes are safe when used in dose ranges recommended by the suppliers. No negative effects on animal health could be observed even at a 100 fold / 50 fold of the recommended inclusion rate in diets for broiler or laying hens, respectively. Thanks to their positive influence on intestinal health, phytomolecules thus boost poultry performance in a safe and effective way.

By Technical Team, EW Nutrition

Literature

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Manafi, Milad, Mahdi Hedayati, Saeed Khalaji, and Mohammad Kamely. “Assessment of a Natural, Non-antibiotic Blend on Performance, Blood Biochemistry, Intestinal Microflora, and Morphology of Broilers Challenged with Escherichia Coli.” Revista Brasileira De Zootecnia 45, no. 12 (December 2016): 745-54. doi:10.1590/s1806-92902016001200003.

Photo source: Aviagen