July | August 2011 Feature title: Breakthrough in Natural Growth Promotion

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F: Growth Promotion
acid catabolites), decrease fat digestibility, stimulate rapid turnover of absorptive epithelial cells, require an increased rate of mucus secretion by intestinal goblet cells and stimulate immune system development and inflammatory responses. All of these effects lead to a loss of net energy to the microflora, resulting in decreased growth performance. Therefore, it is not only highly important to control possible harmful bacteria, but also to keep the well balanced bacterial population within the gut.

F: Growth Promotion
This cell wall layer is significantly thinner in gram-negative bacteria compared to gram-positive bacteria. However, gramnegative bacteria are surrounded by an additional outer membrane, which provides the bacteria with an inherent resistance to hydrophobic antibiotics Figure 1: Inhibition of fish pathogens by an antimicrobial mixture (acid mixture, plus a and detergents. phytochemical) and an antimicrobial mixture Often organic acids and the Biomin Perforizer were combined with other naturally derived products such as essential oils in However, the mode of action of different an attempt to use possible synergisms phytochemicals varies a lot. to more powerful combat pathogenic So, for example, cinnamaldehyde which bacteria. is a phytochemical derived from cinnamon Essential oils in general serve as antibark oil, has a more complex mode of oxidants, stimulate the immune system, action as it targets the FtsZ protein. This suppress harmful microorganisms on one protein plays a major role in the cell division side but stimulate beneficial microbes on of potentially harmful bacteria. the other, they regulate the activity of FtsZ polymerizes into filaments, which certain enzymes, are known to protect assemble at the place within the cell, where the gut villi and to interfere with the DNA the cell division takes place. There they replication of bacterial cells, therefore havform into a polymeric structure known as ing anti-bacterial effects.

Breakthrough in

Naturally combating bacteria

Already a long time ago organic acids were identified to be able to alter the gastro-intestinal microflora by reducing in particular acid-intolerant bacterial species such as E. Coli, Salmonella and Campylobacter resulting in increased growth performance. However, when combating bacteria the structural differences of grampositive and gram-negative bacteria have to be considered. The cytoplasm of the cell is surrounded by the cytoplasmic membrane. The cytoplasmic membrane is covered by a thick cell wall layer.

Natural Growth Promotion

by Angela Riemensperger PhD and Gonalo Santos, Biomin Holding GmbH, Austria
he growth of aquaculture production has been increasing at a fast rate and this trend is expected to continue for some time into the future. This growth is also accompanied by more intensive farming conditions that may lead to a higher incidence of fish diseases. Antibiotic growth promoters (AGPs) have been used in such challenging condi-

"There is a growing awareness regarding the misuse of antibiotic growth promoters (AGPs), which leads to the need to find more sustainable alternatives for responsible aquaculture production"
tions to keep pathogenic bacteria under control. There is a growing awareness regarding the misuse of antibiotic growth promoters (AGPs), which leads to the need to find more sustainable alternatives for responsible aquaculture production. Among these alternatives several feed additives, including organic acids, may be promising substitutes to the AGPs in aquaculture.

Organic acids are known to have positive effects on growth performance and animal health. For terrestrial animals, mainly pigs and poultry, these effects are already well documented. Also in fish there is mounting evidence that organic acids improve growth performance and health of fish. Several studies reported the effects of organic acids in different species including carnivorous fish such as Atlantic salmon, rainbow trout, and Artic charr, also herbivorous fish such as tilapia, and omnivorous fish such as carp and catfish. These studies mainly reported an improvement in performance parameters such as lower feed conversion ratios, increased weight gain and improved nutrient digestibility. These effects are mediated by an antimicrobial effect of organic acids. The antimicrobial effect of organic acids is also shown by the reduced mortality of tilapia fed with acids and challenged with Vibrio anguillarum. Interestingly, for arctic charr it was also reported that fish fed diets with organic acids had a slower gastric emptying rate and this could enhance the antimicrobial potential of the acid salts. The presence of bacteria represents not only a possible threat to the animals health resulting in economic loses, it is also resulting in a loss of energy and nutrients. This

is, as possibly necessary immune responses require energy and the host is competing with the bacteria for nutrients. Therefore, it needs to be taken care that bacteria are effectively controlled. In recent years, great effort was put into enhancing the antimicrobial effects seen when using organic acids in aquaculture feeds. The successful control of pathogenic bacteria can hereby be enhanced by combining organic acid with other natural derived products, such as a phytochemical and the new Biomin Per4izer, a permeabilising substance, which increases the permeability of the outer membrane of gram-negative bacteria. This is a totally new strategy as in this way the antimicrobial effect is boosted considerably.

Naturally ahead

Biotronic Top3
the

Effects of the presence of pathogenic bacteria on growth performance

On one side, animals live in symbiosis with different bacteria, while some bacteria are also known to cause diseases. The so-called beneficial bacteria can protect the host from pathogenic bacteria, regulate the development of the gut or produce vitamins and hormones for the host. However, the presence of possibly harmful bacteria in the gut always represents a possible threat to growth performance. In general, bacteria within the gastrointestinal tract compete with the host animal for nutrients. Furthermore, bacteria secrete toxic compounds (i.e. toxic amino

breakthrough
Inc re weig ased Im ht gain pr conv oved fee Ma ersion d x econimized bene omical t

The BIOMIN perforizing substance damages the outer membrane of Gram-negative bacteria thus boosting the synergistic effect of organic acids and the phytochemical.

For more information visit:

biotronictop3.biomin.net
22.06.11 09:17

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F: Growth Promotion
the antimicrobial mixture of organic acids and the phytochemical. For Vibrio Harveyi a 100% inhibition was already achieved with the antimicrobial mixture. A real synergism between the antimicrobial mixture and the Biomin Per4izer was found for the inhibition of Aeromonas veronii. While the acid mixture and the antimicrobial mixture had inhibiton rates lower than 20%, the addition of the Biomin Per4izer enhanced the inhibition to near 100%.

"Growth performance was improved due to the supplementation of the diets with the acidifier Biotronic Top3. Feed intake was not affected by the treatment"
the Z-ring on the inner membrane in the mid of the cell, which is responsible for the division of the cell. Cinnamaldehyde inhibits not only the formation of FtsZ into filaments, but also inhibits essential processes involved in the Z-ring formation and its function and thus the cell division. This results in a reduction of the bacterial load, within the gastrointestinal tract. Even if the outer membrane of the gram-negative cell is acting as a protective barrier for external agents it is possible to weaken the outer membrane by agents commonly characterised as permeabilizers. However, permeabilizing substances have different modes of action. Some of them remove stabilizing cations from the outer membrane, while others bind to the outer membrane resulting in the loss of barrier function. When permeabilizing substances weaken the outer membrane of gram-negative bacteria, the activity of other antimicrobials is increased by facilitating the entry of external substances, which inhibit or destroy cellular functions, into the cell. This leads to synergistic effects when a permeabilizing substance is added to a mixture of organic acids. However, if a synergism can be found highly depends on the right combination of organic acids and permeabilizing substances, as not with each combination a synergism will be found. found when adding the Biomin Per4izer - a permeabilizing substance to the formula of organic acids and a phytochemical. This is an indication that the permeabilization of the outer membrane of gram-negative bacteria facilitates organic acids and phytochemicals to penetrate the gram-negative cell destroying cellular functions, leading to cellular death. Seen synergistic effects on the inhibition of general pathogenic bacteria draw the attention to the potential effects on the inhibition of pathogenic bacteria specific for fish. Therefore, an in vitro trial was set up, at the Biomin Research Center in Tulln in Austria, to test the effects on the inhibition of bacterial growth of known fish pathogens by an acid mixture combined with a phytochemical and the Biomin Per4izer. Three different pathogens, specific for aquacultured species were tested. This was Yersinia ruckeri, which was shown to cause enteric redmouth disease, a disease which causes significant losses especially in salmonid aquaculture worldwide. The second fish pathogen tested was Vibrio Harveyi a major viulent strain causing vibrosis, which causes mass mortality in larval cultrues and shrimp production. As a third pathogen specific in fish, Aeromonas veronii was chosen. Aeromonas spp. are often isolated from humans suffering from diarrhoea, but also from fish with hemorrhagic septicaemia. Furthermore, the epizooticepizootic ulcerative syndrome in fish, which is causing severe economical losses, is associated with the presence of fish pathogens such as Aeromonas or Vibrio. Results show that inhibition of Yersinia ruckeri was increased by the addition of the Biomin Per4izer to

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Effects on growth performance

As explained earlier these mixture of acids combined with the phytochemical and the Biomin Per4izer - permeabilizing substance can also have important impact as a growth promoter. Therefore, an in vivo trial was set up at the Biomin Research Center in Bangkok in Thailand to test this hypothesis. The fish species used in this trial was red tilapia (Oreochromis mossambicus x Oreochromis niloticus). Groups of 60 fish averaging appr. 45g were stocked in 6 experimental tanks (Volume: 500l). Fish were fed to near satiety 3 times per day for a period of 56 days. Fish were fed a diet based on soybean meal, rice bran and cassava and wheat flower containing 30% crude protein. The control group diet contained no feed additives, whereas the diet of the trial group was supplemented with the natural growth promoter Biotronic Top3 containing an organic acid mixture, a phytochemical and the Biomin Per4izer at an inclusion rate of 1.0 kg/t feed. Growth performance was improved due to the supplementation of the diets with the acidifier Biotronic Top3. Feed intake was not affected by the treatment. However, feed conversion ratio was significantly improved by 5% (P>0.05) in the Biotronic Top3 group compared to the control group. This higher efficiency of ingested feed resulted in significantly higher (P<0.05) final body weight by 5% in the Biotronic Top3 group compared to the control group. Also daily growth coefficient was significantly (P<0.05) improved by 6% in the Biotronic Top3 compared to the control group. In short, the synergy caused by the inclusion of the permeabilizing substance boosts the effect of the antimicrobial ingredients, resulting in economical benefits for the end user. Therefore, it presents an innovative strategy to act against bacteria. The company has launched the natural growth promoter as Biotronic Top3.

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Effects on the inhibition of fish pathogens

Synergistic effects on the inhibition of commonly found bacteria in terrestrial animals such as E. Coli or Salmonella were

20 | INTERNATIONAL AQUAFEED | July-August 2011

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