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Understand How Calibrin®-Z Controls T-2 Toxin in Broilers

January 19, 2024
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Poultry producers mostly know T-2 toxin by the horrific lesions seen on the beaks of poultry. The fast-acting T-2 toxin has a major impact on the growth and performance of poultry and livestock. Luckily it is not the most common trichothecenes toxin produced by Fusarium molds, deoxynivalenol would fill that spot, but T-2 is considered the most toxic of the trichothecenes. This poison can be inhaled or adsorbed through the skin or the gastro-intestinal tract and causes multiple problems in poultry and livestock. A short list of problems includes decreased gain and feed efficiency, decreased egg production and hatchability, decreased immune function, and increased mortality. It has been shown to have a synergistic negative effect with other mycotoxins in the diet or when administered in conjunction with lipopolysaccharide (aka LPS). This may be one reason why there is additional negative impact when there is a co-challenge with T-2 and gram-negative bacteria. Control of other dietary mycotoxins or LPS concurrently with T-2 may be important in any attempt to decrease its effects.   

 

T-2 toxin decreases the productivity of poultry and livestock by inhibiting protein synthesis at the cellular level and causing cell death. In eukaryotic cell’s DNA, RNA, and protein, synthesis is inhibited by T-2 toxin. It also induces apoptosis or programmed cell death.

A major concern in poultry is how T-2 affects the gastrointestinal tract starting with lesions of the beak and gizzard and going through the entire gut. These lesions will affect feed intake, gain, and feed efficiency. But T-2 can affect all aspects of production and reproduction, so egg production and hatchability also need to be considered. In early research looking at the effects of T-2 on hatchability, 2 ppm of T-2 toxin was fed to laying hens, egg production decreased by 3.8 percent, fertility of the eggs that were laid decreased by 1.7 percent, and hatchability of fertile eggs decreased by 5.6 percent. This is a substantial loss of hatched chicks because of the toxin in the feed.

T-2 is quickly adsorbed. And it can be adsorbed through the lungs, the skin, or through the gastrointestinal tract when ingested in the feed. Approximately 90% of T-2 is adsorbed into the body within 30 minutes of ingestion, but it does have a short half-life of less than 20 minutes. T-2 producing Fusarium molds can occur in feedstuffs either during a warm and moist growing season or during storage under high moisture, especially if stored grains have damage such as broken or cracked kernels. The best option for producers is to use feedstuffs free of all toxins, however, the reality is that this is not always possible. In those cases where feedstuffs are being fed that may contain T-2 it would be beneficial to have a fast-acting toxin binder in the diets.    

Because T-2 is so damaging and so rapidly absorbed, the toxin binder that is used needs to work and work fast. Calibrin®–Z, available in select international markets, adsorbed ~70% of T-2 toxin within 1 minute in research looking at speed-of-binding in vitro. This was approximately 24 times faster than the other products used in the trial. Additionally, Calibrin-Z had previously been shown to bind other mycotoxins and LPS in vitro and in vivo, which may be important during a T-2 challenge. A test to determine the binding ability of Calibrin-Z in vitro was conducted to look at seven common fungal biotoxins where the binder-to-toxin ratio was as if there was 1 kg of Calibrin-Z per metric ton of feed vs. observed concentrations of mycotoxins in feed. In vitro data showed that Calibrin-Z could bind LPS, but it has also been seen in vivo when Calibrin-Z was being fed to laying hens.

Calibrin-Z Mitigates the Effects of T-2 Toxin in Broiler Chicks

Recently, research was conducted at a large university in Brazil to determine the effects of Calibrin-Z on broiler chickens challenged with dietary T-2 Toxin. For this experiment, a total of 180 one-day-old male Cobb 500 broiler chicks were used. At the beginning of the trial the birds had an average body weight of 47 grams, with the average initial weight for each bird being equal. They were fed three different treatments 1) Unchallenged Control; 2) Challenged Control with 2 ppm T-2 Toxin; and 3) 2 ppm T-2 Toxin with 0.5% dietary Calibrin-Z. They were fed the treatment diets for 21 days. No aflatoxins, deoxynivalenol, diacetoxyscirpenol, fumonisins, ochratoxin A, T- 2 Toxin or zearalenone were detected in the feed ingredients that were tested before mixing the diets. The T-2 Toxin that was added to the feed for the challenged treatments was produced by Fusarium sporotrichioides fungi, and was 82% T-2 Toxin, 18% HT-2 Toxin.  There were 6 pens that were randomly assigned to each treatment and there were 10 chicks in each pen. Chicks had free access to a constant supply of food and water. The diet was corn-soybean meal-based and formulated according to requirements in the Cobb Broiler Management Guide.  

The intent of the study was to determine the effect of T-2 toxin on growth performance of broilers and how the addition of Calibrin-Z helped to mitigate any negative effects. Calibrin-Z is a unique calcium montmorillonite that has been shown to bind toxins, both fungal and bacterial, as well as lipopolysaccharides (LPS).

Feeding Calibrin-Z to the birds challenged with T-2 toxin increased body weight by 5% compared to the birds that were fed diets with T-2 toxin and no Calibrin-Z. This improvement returned body weight to that of the unchallenged control birds. In this experiment, there was no effect of feeding T-2 on feed intake with birds on all three treatments having equal feed intake.

Because there was no difference in feed intake the feed conversion ratio followed the same pattern seen in body weight. Feeding Calibrin-Z to birds challenged with T-2 toxin improved feed conversion by 8 points, with values of 1.50 for Calibrin-Z fed birds compared to 1.58 for birds that only received T-2 toxin in the feed.

While aflatoxin is the mycotoxin best known for its negative effects on the liver, T-2 toxin can also have bad effects. In this study, relative liver weight was higher in the challenged control, 3.33%, compared to the unchallenged control, 2.84%, but feeding Calibrin-Z again mitigated the negative effect of the T-2 toxin, with challenged birds fed Calibrin-Z having a relative liver weight of 2.96%.   

T-2 Toxin is a fast-acting mycotoxin that has a tremendous impact on animal performance. It acts in a synergistic way with challenges from other mycotoxins and lipopolysaccharide. To decrease its impact, you need a fast-acting toxin binder to control T-2 quickly while also controlling other potential problems. Calibrin-Z is a fast-acting multi-toxin binder that has proven results.   

 As the animal health business of Oil-Dri® Corporation of America, Amlan products are backed by Oil-Dri’s 80-plus years of mineral science expertise. Oil-Dri and Amlan are vertically integrated and own every step of the production process to consistently deliver safe, high-quality animal health products around the world. Calibrin-Z, a calcium montmorillonite clay, is sold as a broad-spectrum toxin binder. To understand how Calibrin-Z can work in your production system, contact your local Amlan representative.

 

References:  

Chi, M. S., C. J. Mirocha, H. J. Kurtz, G. Weaver, F. Bates, and W. Shimoda. 1977. Effects of T-2 Toxin on Reproductive Performance and Health of Laying Hens. Poultry Sci. 56:628 – 637.  

Tai, J.-H. and J. J. Pestka. 1988. Synergistic interaction between the trichothecene T-2 toxin and Salmonella typhimurium lipopolysaccharide in C3H/HeN and C3H/HeJ mice. Toxicol Lett 44:191–200.  

Mycotoxins: Risks in Plant, Animal, and Human Systems. 2003. Task Force Report No. 139 Council for Agricultural Science and Technology. Ames, Iowa, USA.

Article Showcases Phylox® in Watt Poultry International

December 21, 2023
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Watt Poultry® International published an article earlier this year on Amlan international’s product, Phylox®. Phylox is a natural alternative to anticoccidial drugs that works well alone, as part of a rotation program, or as part of a bio-shuttle program to control coccidiosis. The natural ingredients of phylox targets multiple Eimeria species with several modes of action, while also promoting a healthy intestinal barrier and improving intestinal immunity. If you missed this article, here is your chance to learn more about Phylox. Follow the link to learn more.

Reduce Salmonella Prevalence and Load with NeutraPath®

December 21, 2023
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Salmonellosis is a foodborne pathogen that causes illness and death worldwide. A blended feed additive has been shown to have good effects in vitro on a wide range of Gram-negative and Gram-positive bacteria and in vivo to mitigate the effects of Clostridium perfringens in broilers and E. coli in swine. Thus, it was decided to investigate the ability of the proprietary blend of essential oils, medium-chain fatty acids, and an activated toxin-adsorbing mineral (NeutraPath® available in select international markets) to control Salmonella. In vitro and in vivo research with Salmonella typhimurium and Salmonella heidelberg showed the blend could reduce prevalence and bacterial load of Salmonella in broiler chickens. Based on these in vitro and in vivo data, feeding this blend could be a potential new method to help control Salmonella in broiler chickens and aid in control of pathogens at the farm level. 

 

A Foodborne Pathogen 

Salmonella is a very common pathogenic bacteria that can be passed from animals to humans. The typical symptoms of salmonellosis in humans are diarrhea, fever, and stomach cramps, with the occasional vomiting. Generally, this is mild and doesn’t require medical intervention, but it can be deadly, especially in young children. Worldwide, Salmonella is one of four main causes of diarrheal diseases, with diarrheal disease being the 2nd leading cause of death in children under five. In the United States approximately 1,350,000,000 people are infected annually, ~26,500 people are hospitalized, and ~420 die each year. The very young, the very old, pregnant women, and people with compromised immune systems are generally affected the most.  

According to the World Health Organization, Salmonella, a hardy bacteria, can survive in a dry environment for several weeks but several weeks turns into several months if it is in water.  

 

Sometimes salmonellosis can result from coming in direct contact with animals that carry the bacteria, typically reptiles or birds. There are two species of Salmonella, bongori and enterica. Salmonella bongori is normally associated with cold-blooded animals but can infect humans. Selling tiny turtles (those with shells less than 4 inches long) has been prohibited in the U.S. since 1975 because of their association with salmonellosis in children. Salmonella enterica has more serovars with approximately 80 that can infect humans and animals. With an increase in the popularity of raising your own chickens the CDC (Centers for Disease Control) has issued repeated reminders about the safe handling of chickens, including a reminder “don’t kiss your chickens” because of serious outbreaks of salmonellosis, especially among children, linked to raising chickens in the backyard.

But eating or mishandling raw or undercooked contaminated food is the source of most cases of salmonellosis in humans. Live poultry often don’t show signs of carrying Salmonella even if their intestines contain the pathogenic bacteria. There are multiple ways that birds can be exposed to Salmonella. Exposure can be through contaminated feed, from wild birds or rodents, or from a contaminated barn. It is even possible that poultry may be contaminated before the egg that they hatched from was laid. Hens can have bacteria in the ovary or oviduct and the egg can be contaminated before the shell forms around the egg, meaning that even clean, washed eggs could be contaminated. The Poultry Industry has been working diligently to control this problem and this source of contamination of broiler chickens has declined in recent years.  

 

Even though a majority of the foodborne illnesses due to Salmonella originate from non-poultry sources, twenty-three percent of the Salmonella outbreaks in the U.S. are linked to poultry consumption (16.8% from chicken, 6.6% from turkey) with another 6.3% coming from eggs. Salmonella in the intestine of poultry can lead to contamination of poultry meat during processing. Proper cooking will kill the bacteria, but improper handling may spread bacteria around the kitchen and raw vegetables, undercooked meat or uncooked foods containing eggs (i.e., cookie dough) may still be contaminated with live bacteria. Together the CDC, FDA (Food and Drug Administration), and USDA (United States Department of Agriculture) have a goal of reducing Salmonella illnesses by 25% by 2030. In order to do this, they need to decrease Salmonella infections from all products regulated by the Food Safety and Inspection Service division of the USDA by 25%. One way to help reach this goal is to help minimize the amount of Salmonella that comes into the processing plant making it less likely that contamination of poultry meat will occur.  

Poultry producers need help to accomplish this goal. Research has shown that a proprietary blend of essential oils, medium-chain fatty acids, and an activated toxin-adsorbing mineral (NeutraPath®, Amlan International, available in select international markets) may be of assistance. The blend has anti-virulence effects because of its ability to bind quorum sensing molecules, exotoxins, and endotoxins associated with bacteria. It also has direct bacteriostatic/ bactericidal effects against both Gram-positive and Gram-negative bacteria. Its efficacy has been proven over years of in vitro and in vivo trials at multiple research sites, against multiple bacteria, in multiple animal species.  

In vivo research in chickens showed that the blend decreased the effects of Clostridium perfringens. A summary of the studies showed that it improved mortality, gain, and feed conversion in challenged broilers. In weaning pigs, it was shown to decrease the impact of enterotoxigenic E-coli (F-18+). In the pigs challenged with E. coli the blend improved feed efficiency and decreased frequency of diarrhea. When the fecal microbiome was examined, there was a higher relative abundance of Lactobacillaceae and a lower relative abundance of Enterobacteriaceae. Enterobacteriaceae is a family of Gram-negative bacteria that includes both E. coli and Salmonella. These positive results led researchers to investigate its effects on Salmonella, with research being conducted both in vitro and in vivo.  

 

Salmonella Research 

 

In vitro tests were used to determine the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of NeutraPath against S. heidelberg. Three different concentrations (1, 3 and 5 mg/ml) of NeutraPath with a control of 0 mg/ml were added to samples of an S. heidelberg strain. The MIC of NeutraPath for S. heidelberg was found to be 5 mg/ml. To determine the MBC Salmonella cultures were incubated at 37ºC for 16 hours without agitation. After incubation, bacterial counts were measured by serial dilution. A 30 μl aliquot of each dilution was plated onto lysogeny broth and incubated overnight. The MBC of NeutraPath for S. heidelberg was determined to be 4 μg/ml. These assays demonstrated that the blend has strong in vitro inhibitory and bactericidal activities against this key pathogen.  

Because of the results seen in vitro researchers conducted an in vivo experiment. The researchers wanted to know how the blend would affect the Salmonella prevalence in birds previously challenged with S. heidelberg and fed for a short period of time before sampling.  

After feeding the treatments for seven days, pre-moistened boot-sock swab sampling showed that there was S. heidelberg contamination in 100% of the pens. Cecal digesta samples and cloacal swabs were also collected from 10 of the broilers that had been directly challenged at hatch to determine Salmonella prevalence. The prevalence of Salmonella decreased by 40% in the cecal digesta (83.3% vs. 50.0%) and by 55% (60.0% vs. 26.7%) in the cloacal swabs when the blend was added to the diet for 7 days prior to testing. 

 

Comparable studies were done at second location with another group of researchers to establish that the results from the initial studies could be repeated in other Salmonella serovars. This time studies looked at the effects of the blend on Salmonella typhimurium.  

 

First an in vitro study digestion was used to simulate the crop, proventriculus and intestinal section of the gastrointestinal tract. Each “section” had pH and enzymatic conditions that would correspond to that area of the gut. Adding the antimicrobial blend inhibited the growth of S. typhimurium, reducing the total colony forming units recovered in each section. This positive result meant that an in vivo study was warranted. This time the broilers were started at one day of age. Thirty male broiler chicks were placed on two treatments. The treatments were a challenged control with non-treated feed, or birds fed that diet + 0.25% of the blend. At nine days-of-age chickens were given an oral dose of 106 CFU (Colony Forming Units) of live S. typhimurium. Twenty-four hours after the challenge ceca and cecal tonsils were removed so that they could be evaluated for Salmonella recovery. Both the number of positive samples and the amount of Salmonella bacteria found in those positive samples decreased in the treated birds. Feeding the blend decreased the ceca that tested positive for S. typhimurium by 41.7%, 100% of the tested ceca were positive in birds fed the untreated control compared to only 58.3% of the ceca in the birds that were fed the blend. In the birds that tested positive the total S. typhimurium bacterial load recovered in the ceca also dropped by 1.84 log10 CFU/g compared to the untreated control.

Conclusion 

Around the world foodborne salmonellosis continues to be a problem. Regulators are targeting aggressive goals for salmonella reduction in poultry meat. A proprietary blend of essential oils, medium-chain fatty acids, and an activated toxin-adsorbing mineral has been shown to work against a variety of bacteria. Research has shown efficacy against C. perfringens, E. coli, and S. typhimurium and S. heidelberg. Based on the current in vitro and in vivo research adding the blended product has the potential to reduce Salmonella colonization in broiler chickens. This is a viable option for use in poultry health programs including those controlling Salmonella contamination. To learn more about NeutraPath or to contact a local representative, visit amlan.com.

 

The Role of Quorum Sensing in Necrotic Enteritis Development

July 11, 2023
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The fast onset of necrotic enteritis and the devastating production losses it inflicts make it one of the most challenging diseases for the poultry industry, particularly for antibiotic-free producers. Clostridium perfringens, the cause of necrotic enteritis, possesses a number of virulence factors that allow it to mount a fast, efficient attack on the host including potent toxins and cell-to-cell communication (quorum sensing). However, natural mineral-based products that can disrupt quorum sensing and reduce the virulence of C. perfringens are available.

The Pathogenesis of Necrotic Enteritis

C. perfringens is an anerobic, spore-forming pathogen found in the normal microbiota of poultry, as well as the poultry house. Necrotic enteritis occurs when predisposing conditions, such as a change in diet, immune status or intestinal pathophysiology, promote an overgrowth of C. perfringens. Coccidiosis can also increase the incidence of necrotic enteritis, as the damage that Eimeria spp. cause to intestinal epithelial cells promotes the invasion of C. perfringens (as well as other pathogens).

C. perfringens’ Rapid Infection Rate

C. perfringens is one of the fastest growing bacterial pathogens. Under optimal conditions it can replicate every 8 to 10 minutes — outgrowing other resident bacteria to achieve intestinal colonization.1 As well as a rapid growth rate, C. perfringens infection involves multiple steps, which likely occur simultaneously, including colonization, replication, nutrient procurement, evasion of host immune defenses, host tissue damage and transmission.2

Exotoxin Roles in Necrotic Enteritis Development

Multiple exotoxins can be produced by C. perfringens, including alpha-toxin and necrotic enteritis toxin B-like toxin (NetB). Alpha-toxin is cytotoxic to endothelial cells, red blood cells, white blood cells and platelets, while NetB toxin forms pores in cell membranes that allow electrolytes to rupture cells, causing cell death and necrotic lesions in the small intestinal mucosa.3 These two toxins are known to have a role in necrotic enteritis development.

C. perfringens Growth Depends on Host Nutrients

C. perfringens relies on nutrients from the host to live and multiply — a process which results in the destruction of host tissues (formation of necrotic lesions). C. perfringens lacks enzymes needed for amino acid biosynthesis and subsequent protein synthesis, so enzymes and toxins are released to degrade structural proteins from the host.4 The host amino acids and/or peptides are then taken in by C. perfringens for use in its own protein synthesis. 4

To produce energy, C. perfringens degrades large sugar compounds from the host and ferments them, producing gas that enhances the anerobic environment.4 C. perfringens also produces hyaluronidases that increase connective tissue permeability and help C. perfringens spread into deeper tissues.4

Quorum Sensing Controls Exotoxin and Enzyme Production

C. perfringens uses quorum sensing (cell-to-cell communication) to coordinate exotoxin and enzyme production to occur when its population reaches a density that supports the most efficient use of its metabolic resources.5 For example, to determine the optimum time to start producing NetB, the accessory gene regulator-like (Agr-like) quorum-sensing system sends out signals that are recognized by the VirR/VirS two-component regulatory system.5 Once the VirR/VirS system detects that the C. perfringens population has reached the threshold density, it switches on the expression of NetB and other virulence and related metabolism genes.2

Quorum Quenching Reduces Pathogen Virulence

Quorum quenching is an approach that can disrupt the quorum-sensing system of pathogenic bacteria, preventing cell-to-cell communication and the expression of quorum-sensing-controlled genes that produce toxins and other virulence factors. Additionally, quorum-quenching products should reduce the chance of antibiotic resistance, since they are modifying bacteria behavior rather than killing them.

Natural Quorum-Sensing Control

One product that has displayed quorum-quenching properties is the mineral-based feed additive Calibrin®-Z (available in select international markets). This all-natural single-ingredient product binds bacterial pathogens and the toxins they produce, as well as multiple mycotoxins, to help protect the intestinal barrier against enteric disease. Other natural mineral-based products can also help manage necrotic enteritis; read this article to learn more.

An in vitro study found that Calibrin-Z separated out quorum-sensing molecules by adsorption or catalytically broke them down into small fragments. By reducing the concentration of quorum-sensing biochemicals, Calibrin-Z can potentially disrupt the ability of pathogenic bacteria (including C. perfringens) to produce toxins, since quorum sensing controls this function. Calibrin-Z was also shown to effectively bind alpha-toxin and NetB toxin, further reducing the virulence of C. perfringens.

 

The global reduction in the use of in-feed antibiotics has compelled producers to rely on other management methods to maintain a healthy intestinal environment in poultry and reduce the risk of necrotic enteritis. The use of best-practice management strategies and inclusion of mineral-based feed additives that reduce the virulence of C. perfringens can assist in promoting intestinal health and maximizing production efficiency. To learn more about necrotic enteritis and natural mineral-based methods to control it, contact your local Amlan representative.

 

References

  1. Kiu R, Hall LJ. An update on the human and animal enteric pathogen Clostridium perfringens. Emerg Microbes Infect. 2018;7:141.
  2. Prescott JF, Parreira VR, Mehdizadeh Gohari I, Lepp D, Gong J. The pathogenesis of necrotic enteritis in chickens: what we know and what we need to know: a review. Avian Pathol. 2016;45:288–94.
  3. Chi, F. A Viable Adjunct or Alternative to Antibiotics: Meta-Analysis of Broiler Research Shows Natural Growth Promoter Delivers Feed Efficiency Equal to Antibiotics. Amlan International.
  4. Shimizu T, Ohtani K, Hirakawa H, Ohshima K, Yamashita A, Shiba T, Ogasawara N, Hattori M, Kuhara S, Hayashi H. Complete genome sequence of Clostridium perfringens, an anaerobic flesh-eater. Proc Natl Acad Sci U S A. 2002;99:996–1001.
  5. Yu Q, Lepp D, Mehdizadeh Gohari I, Wu T, Zhou H, Yin X, Yu H, Prescott JF, Nie SP, Xie MY, Gong J. The Agr-Like Quorum Sensing System Is Required for Pathogenesis of Necrotic Enteritis Caused by Clostridium perfringens in Poultry. Infect Immun. 2017;85:e00975-16.

 

Natural Mineral-Based Neutralization of E. coli Toxins

April 10, 2023
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Bacterial pathogens such as Escherichia coli (E. coli) are a leading cause of enteric disease and economic loss for animal protein producers. Found readily in the environment of poultry and swine, E. coli can produce powerful toxins (exotoxins and endotoxins, see Figure 1) that damage the integrity and function of the intestinal barrier. These toxins cause inflammation, diarrhea and malabsorption that result in less gain, higher feed conversion and reduced performance. However, multiple studies have confirmed that natural mineral-based feed additives can provide an antibiotic-free method of reducing the negative production effects of E. coli toxins.

Figure 1: E. coli exotoxins are produced by the cell and secreted into the environment. Endotoxin (lipopolysaccharide, LPS) is released from the cell wall of E. coli or other gram-negative bacteria after the bacteria dies.

 

Powerful Toxins

Multiple E. coli strains produce potent exotoxins as part of their virulence package. These powerful toxins are major causes of enteric disease, particularly in neonatal and recently weaned animals. The toxins damage the intestinal lining, resulting in diarrhea, dehydration and even death.

Heat-Labile Enterotoxin

Enterotoxigenic E. coli (ETEC) — a common cause of porcine post-weaning diarrhea (PWD) and a source of diarrhea in poultry — produces the enterotoxin heat-labile toxin (LT). To exert its effects, LT is internalized by binding to surface receptors on the host intestinal mucosa.1 Once inside the cell, LT alters the electrolyte and water balance of the cell, which leads to diarrhea.1 A survey of U.S. ETEC isolates linked to porcine PWD showed that 57.7% had the heat-labile toxin gene.2

Shiga-like Toxin

E. coli types that produce shiga-like toxins are referred to as shiga toxin-producing E. coli (STEC). There are two types of shiga-like toxins (Stx1 and Stx2), and both have multiple subtypes. The mode of action for shiga-like toxin is inhibition of host cell protein synthesis that results in cell death.3 Edema disease (E. coli enterotoxemia) in swine is caused by shiga-like toxin (Stx2) subtype e.4

E. coli Endotoxin

The outer membrane of gram-negative bacteria such as E. coli or Salmonella contains lipopolysaccharides (LPS) which protect the bacteria from the environment. When the bacteria die naturally or as a result of using antibacterial products (e.g., antibiotics, specialized feed additives), endotoxin is released from the cell wall. Endotoxin causes specialized immune cells in the animal’s body to release high levels of cytokines that result in elevated temperature, weakness, tissue necrosis and, at high levels, death.

Management Programs to Control E. coli

With the global push toward antibiotic-free production, maintaining a healthy intestinal environment to reduce the risk of disease from E. coli now requires effective management strategies. Natural feed additives, tight biosecurity, effective sanitation, and good husbandry practices that protect the intestinal environment can all help control the risk of E. coli outbreaks.

Natural Mineral-Based Feed Additives Neutralize E. coli Toxins

The backbone of Amlan’s scientifically proven products is our natural mineral that undergoes proprietary thermal processing to optimize the efficacy of each product. Varium® (available in select international markets) for poultry and NeoPrime® (available in select international markets) for swine are species-specific formulations of three synergistic ingredients that improve feed conversion and intestinal integrity in poultry and maximize post-weaning performance in swine. Calibrin®-Z (available in select international markets) is a low-dose biotoxin binder for all species that actively defends the intestinal barrier against a broad spectrum of toxins produced by bacteria and fungi.

Adsorption of E. coli toxins by Varium and Calibrin-Z

The ability of Varium and Calibrin-Z to adsorb E. coli toxins was assessed in vitro by the United States Department of Agriculture Agricultural Research Service (USDA-ARS), Beltsville, MD. Calibrin-Z or Varium was added separately to each toxin, incubated to allow the toxin to bind, and then centrifuged to separate the free and bound toxins. The USDA researchers had developed a specific ELISA for heat-labile and shiga-like toxins, which was used to analyze the supernatant to quantify the presence of unbound toxins.

Varium and Calibrin-Z were both effective at neutralizing both exotoxins (Figures 2 and 3) and endotoxins. (Figure 4).

 

Figure 2: Varium neutralized E. coli exotoxins in vitro. Source: USDA-ARS, Beltsville, MD.

Figure 3: Calibrin-Z adsorbed E. coli exotoxins in vitro. Source: USDA-ARS, Beltsville, MD.

Figure 4: Varium and Calibrin-Z adsorbed E. coli endotoxin (LPS) in vitro. Calibrin-Z source: Gibraltar Laboratories, Fairfield, NJ. Varium source: USDA-ARS, Beltsville, MD.

NeoPrime Reduces Plasma Endotoxin Concentration

When endotoxin is released from dead gram-negative bacteria, it can damage the intestinal epithelial cells. Weakness in the intestinal barrier can lead to “leaky gut syndrome,” which allows the endotoxin to enter the blood stream. NeoPrime was assessed for its ability to strengthen intestinal integrity by analyzing the blood plasma endotoxin concentration of weanling pigs. Compared to a control that contained two antibiotics and zinc oxide, NeoPrime in multiple diet formulations reduced plasma endotoxin concentration from E. coli or other gram-negative bacteria (Figure 5).

Figure 5: Feeding NeoPrime decreased plasma endotoxins, indicating improved gut barrier function Source: Hunan Agricultural University, Hunan, China.

 

When to Use Our Mineral-Based Feed Additives

The research presented here supports in vivo studies showing that supplementation with Varium, NeoPrime or Calibrin-Z is an effective approach to reducing the effects of enteric disease in poultry and swine. To learn more about these natural mineral-based control methods for improving intestinal health and production efficiency, visit amlan.com or contact us at info@amlan.com.

 

 

References

  1. Duan Q, Xia P, Nandre R, Zhang W, Zhu G. Review of newly identified functions associated with the heat-labile toxin of enterotoxigenic Escherichia coli. Front Cell Infect Microbiol. 2019.
  2. Zhang W, Zhao M, Ruesch L, Omot A, Francis D. Prevalence of virulence genes in Escherichia coli strains recently isolated from young pigs with diarrhea in the US. Veterinary Microbiology. 2007;123:145–152.
  3. Melton-Celsa A. Shiga toxin (Stx) classification, structure, and function. Microbiol Spec. 2014;2.
  4. Baldo V, Salogni C, Giovannini S, D’Incau M, Boniotti MB, Birbes L, Pitozzi A, Formenti N, Grassi A, Pasquali P, Alborali GL. Pathogenicity of shiga toxin type 2e Escherichia coli in pig colibacillosis. Front Vet Sci 2020 Sep 18;7: n.p. doi: 10.3389/fvets.2020.545818.

 

Phylox® Performance Equals Salinomycin or Nicarbazin to Combat a Coccidia Challenge

January 16, 2023
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To an industry looking for anticoccidial alternatives, Amlan International offers Phylox® (available in select international markets), a blend of natural ingredients, Phylox was developed to control coccidiosis without resorting to ionophores or chemicals. Dr. San Ching, a Technical Research Manager at Amlan International was instrumental in the development of Phylox. He had this to say about the development of Amlan’s new product to fight coccidiosis

“We designed, Varium®, which provides excellent results reducing the damage from necrotic enteritis caused by toxins produced by Clostridium perfringens. However, our customers also needed a solution for coccidiosis, which is highly related to necrotic enteritis. Therefore, we developed, Phylox, which works to decrease the effects of the parasitic disease. When we started working on a coccidiosis solution, we soon realized that there was a resistance issue in many of the current alternatives, both for antibiotic ionophores and chemicals. Thus, for Phylox we used natural ingredients that would attack coccidia and the coccidiosis problem in more than one way and avoid the development of resistance. We know that the oocysts’ lifecycle is complicated, but most of it occurs in the gut. We selected ingredients that limit the development of the Eimeria oocysts in multiple ways. Additionally, we wanted to invent a product that can work seamlessly with the vaccines.”

Phylox has been shown to be successful in numerous research trials. In 2022, Amlan gave two presentations: A Novel Approach to Coccidiosis Control and A Research-Backed Alternative to Anticoccidial Drugs at the Scientific Forum of IPPE. These presentations represented seven experiments and show Phylox improved growth and immune function in broilers challenged with coccidia. Two additional experiments have been added as a final step in research and development, further proving the value of Phylox for birds facing a coccidia challenge.

In each of the two studies, the coccidia challenge was at day 14 when birds were given an oral dose of 100,000 oocysts of E. acervulina, 50,000 oocysts of E. maxima, and 75,000 oocysts of E. tenella.  The coccidia lifecycle is interesting and complex. Many coccidia species are host specific and do not cross from one host species to another. Several strains of coccidia affect chickens. These strains rise and fall in prominence, in part due to their ability to develop resistance to the chemical or antibiotic that is being used to control them. The three species used in these studies attach to different areas of the gastro-intestinal tract. E. acervuline causes lesions and damage to the upper third of the intestine – the duodenum and upper ileum, E. maxima causes lesions in the middle third of the intestine, and E. tenella cause lesions in the ceca.

 

In the first study, no coccidiosis vaccines or other coccidiosis medications were given. The treatments in the study consisted of both challenged and unchallenged birds with Phylox in the feed and challenged and unchallenged birds without Phylox in the feed. When a coccidia challenge wasn’t given, birds fed Phylox had the same gain and feed conversion as the control birds both for day 14 – 28 and the overall, day 0 – 28, experimental period (Figures 1 &2). This result proves that feeding Phylox doesn’t have a negative affect when a coccidia challenge isn’t present, which is a problem with some other coccidiosis preventatives.

 

Figure 1. Weight gain of birds without and with a coccidia challenge with and without Phylox in the feed.

 

Phylox improved performance in birds challenged with coccidia. Feeding Phylox improved feed conversion in the challenged birds. This was seen in both in the challenge period, day 14 – 28, and for the overall experimental period from day 0 – 28.

 

Figure 2. Feed conversion of birds without and with a coccidia challenge with and without Phylox.

 

Figure 3. Phylox has multiple ways to help prevent coccidiosis.

 

The improvement in feed conversion is because Phylox contains various active ingredients giving it multiple modes of action (Figure 3). This allows Phylox to protect the intestine during a coccidia challenge. First, it interferes with the cell membrane of the coccidia protozoa. Phylox binds to the sterols of the cell membrane, compromising the integrity of the sporozoite cell wall, the cell then dies by apoptosis. Phylox also interrupts the Eimeria lifecycle by preventing oocyst sporulation. By interrupting this step, the oocysts don’t develop from the immature noninfective form to the mature infective form. This disruption protects the intestinal cells from coccidial infection. Other ingredients included in Phylox provide energy to the endothelial cells that line the intestine. The protected intestine is better able to absorb necessary nutrients resulting in improved feed conversion. In addition to improved feed conversion, this protective ability is also shown by the decrease in lesion scores of the challenged birds fed Phylox. The lesion scores formed by each of the coccidia species used in the challenged decreased when birds were fed Phylox when intestines were examined on day 20 (Figure 4).

 

Figure 4. Lesion scores of birds without and with a coccidia challenge with and without Phylox in the feed.

 

 

In an experiment with a similar coccidiosis challenge model, Phylox was compared to using salinomycin or nicarbazin. These feed additives have commonly been used to decrease the effects of coccidiosis.

 

 

Figure 5. Weight gain of coccidia challenged broilers fed salinomycin, nicarbazin, or Phylox.

 

Feeding Phylox resulted in weight gain and feed conversion equal to the salinomycin or nicarbazin treatments (Figure 5 & 6). This is the same result that was seen in the research presented by Dr. Ching at the 2022 IPPE Scientific Forum A Research-Backed Alternative to Anticoccidial Drugs. This result was observed both in the days following the challenge (day 14 – 28), and for the overall experimental period (day 0 – 28). All treatments had better feed conversion than the untreated birds.

 

 

Figure 6. Feed conversion of coccidia challenged broilers fed salinomycin, nicarbazin, or Phylox.

 

Phylox reduced lesion scores for each species of coccidia and as an average of all species compared to the untreated challenge when intestines were examined on day 20. The reduction was equal to that of the ionophore and the chemical coccidiosis treatments. Feeding Phylox also decreased fecal coccidia oocysts counts equal to the decrease seen by feeding salinomycin or nicarbazin. Oocysts were counted in excreta collected from day 19 – 22.

 

 

Figure 7. Lesion score or oocyst counts of coccidia challenged broilers fed salinomycin, nicarbazin, or Phylox.

 

Because Phylox contains various active ingredients it can fill in the gaps that are missing from other coccidiosis control methods. It can be used in No-Antibiotic-Ever programs with no withdrawal requirement. It is effective against multiple Eimeria strains without promoting the emergence of drug-resistant coccidia. And it can be fed with anticoccidial vaccines, preventing disease breakthrough while immunity is being developed by the bird.

Natural Phylox is not only equal to traditional ways of controlling coccidiosis but is equal in economic performance. Phylox can be used to combat coccidiosis in no-antibiotics-ever or traditional production.

Dr. Ching’s presentation of this research at IPPE in the poster session.

Talk to your Amlan representative on how to use Phylox in your production system.

IPSF Presentations Further Support Phylox® Feed as Natural Alternative to Anticoccidial Drugs

August 16, 2022
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Phylox® logo with microbiology background.

Coccidiosis is the most significant disease affecting poultry production, costing the global poultry industry approximately US$3 billion annually. Not only does coccidiosis damage the gut, resulting in reduced efficiency and profitability, but it also contributes to the development of other enteric diseases like necrotic enteritis. Traditionally, antibiotics or synthetic chemicals were used to control coccidiosis; however, consumer concerns over chemicals and drugs in the food chain have reduced their use in recent years. Therefore, poultry producers need a natural, research-backed alternative to chemical coccidiostats and antibiotics — like Phylox® Feed.

Phylox (available in select international markets) is a blend of bioactive phytochemicals that decreases the negative production and health effects of a coccidia challenge. The ingredients in Phylox contribute to its multiple modes of action which include damaging the coccidia cell structure, boosting anticoccidial immunity and improving general gut health.

Multiple studies have proven Phylox decreases gut damage in Eimeria-challenged broilers, resulting in improved growth and efficiency. Further analysis of some of these trials was presented at the 2022 International Poultry Scientific Forum (IPSF) in Atlanta, Georgia, as a six-trial meta-analysis comparing Phylox to a control diet in Eimeria-challenged broilers. Research was also presented at the 2022 IPSF on how Phylox affects the immune response and gut microbiome of broilers during a coccidia challenge. Summaries of these two IPSF presentations and links to the abstracts are below.

 

A Research-Backed Alternative to Anticoccidial Drugs

A meta-analysis of six in vivo trials that compared the performance and health effects of broilers challenged with experimental coccidiosis and fed either Phylox or a non-supplemented control diet was conducted. Persistent effects of Phylox on growth performance, fecal oocyst shedding and intestinal lesion score were examined.

Phylox improved gut health and performance while also decreasing coccidial lesion scores and oocyst shedding. Less gut damage means improved nutrient use — supporting greater weight gain and feed conversion. Two of the experiments included treatment groups that contained ionophores or anticoccidial chemicals for comparison. In these studies, Phylox delivered gut health results similar to salinomycin, nicarbazin, and Maxiban™ (narasin and nicarbazin), as determined by coccidial lesion scores and feed conversion.

This study supports previous research that shows the potential of Phylox as an alternative to the traditional coccidiostats in poultry production. The results are particularly noteworthy in an era when use of these traditional products continues to be reduced in favor of natural non-antibiotic and non-chemical solutions.

 

A Novel Approach to Coccidiosis Control

This research explored the potential effects of Phylox on host coccidial immunity, the composition and structure of the gut microbiome, and intestinal integrity of broilers challenged with experimental coccidiosis. Peripheral blood mononuclear cell phenotype, ceca-cecal tonsil cytokine mRNA expression, gut microbiome of cecal content and duodenal/jejunum histopathology were examined.

Phylox Maintains Intestinal Immune System Protection

Most of a chicken’s immune tissue is in the gastrointestinal tract, helping to keep pathogenic substances out while allowing nutrients in. IL-10 is an anti-inflammatory cytokine with potent immunosuppressive effects. Coccidia can exploit these immunosuppressive properties, to help them survive in a relatively hostile environment, by increasing IL-10 during a challenge. This decreases the bird’s immune response just when it is needed. However, in this study, Phylox kept IL-10 from increasing during a coccidia challenge, stopping the coccidia from interfering with the bird’s immune response.

Preserved Microbiome Diversity

A disruption to the balance of the gut microbial community is often associated with a loss of microbiota diversity (Mosca et al., 2016). In the Phylox study, Eimeria challenge decreased the α-diversity (the mean species diversity at a local site) of the cecal microbiome, but Phylox returned it to its normal, unchallenged state. This result was for both richness (the total number of species) and evenness (the amount of each species). Additionally, feeding Phylox increased the relative amount of Blautia and L-Ruminococcus — producers of short-chain fatty acids that help the gut and the immune system maintain consistency regardless of challenges.

This study demonstrated that, in addition to damaging Eimeria cell structure and functional integrity, Phylox helps host defense mechanisms by boosting protective immunity against coccidial infection. This multimodal mechanism of action of Phylox contributes to an enhanced resistance to coccidial infection and improved bird productivity.

For more information on these studies or to trial Phylox Feed yourself, contact us at info@amlan.com.

Varium® Rivals Zinc Bacitracin in Maintaining Broiler Intestinal Health

June 28, 2022
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Varium® logo with broilers in the background.

In some countries, the use of zinc bacitracin as an antibiotic growth promoter (AGP) in poultry is restricted due to concern over the increase in multi-drug-resistant bacteria that do not respond to traditional antibiotic treatments. Restrictions in the use of AGPs in animal feeds like zinc bacitracin has spurred the development of natural AGP alternatives that keep birds healthy and growing efficiently.

Natural Performance Promotion

Varium® is a patented natural mineral-based feed additive that promotes efficiency and productivity in poultry. Unlike antibiotics that kill bacteria, the patented technology in Varium includes a synergistic formulation of ingredients that binds pathogenic bacteria and their toxins, provides an energy source for the growth of healthy and strong enterocytes and gently stimulates immunity cells. With multiple modes of action, Varium adds value for producers by replacing the need for multiple feed additives; it can provide the same benefits in one product thereby simplifying diet formulations and reducing costs.

Comparing Varium with Zinc Bacitracin

To demonstrate its effectiveness, Varium was directly compared to zinc bacitracin in a broiler study conducted by a university in Pakistan. In the trial, 180 Ross 308 chicks (10 chicks per pen, 6 pens per treatment) were randomly allocated to one of three treatment groups: control (0.01% zinc bacitracin), Varium 0.1 (0.1%) or Varium 0.15 (0.15%). The broilers were raised under normal production conditions, with the trial ending on day 35. Newcastle disease vaccine was administered to all birds on day 6 (intraocular and subcutaneous) and a booster (oral) was administered on day 21. Newcastle disease titers were measured on days 20 and 35 from 18 birds per treatment. Three birds per pen (18 total per treatment), randomly selected on day 35, and had small intestine morphology and bacterial counts in the small intestine and digesta measured.

Varium Protects Intestinal Health

The study showed that Varium was able to protect intestinal morphology better than zinc bacitracin. In Varium-fed broilers, a significant dose-response effect was observed for intestinal (jejunum) villus height and villi index (villus height to crypt depth) on day 35, with all Varium treatments significantly higher than the zinc bacitracin control (Figure 1). A similar dose response was observed with intestinal (jejunum) crypt depth; all Varium treatments were significantly lower than the control, and Varium 0.15 was significantly lower than Varium 0.1 (Figure 1).

Villus Height, Villi Index, Crypt Depth of broilers information.
Figure 1: Villus height, villi index and crypt depth of broilers fed either 0.01% zinc bacitracin (control) or Varium at 0.1 or 0.15%. Varium demonstrated a dose-response effect that indicated better ability to protect intestinal morphology than zinc bacitracin.

Varium was also able to protect the birds from necrotic enteritis to the same extent as zinc bacitracin. Necrotic enteritis was not found among any of the sampled birds on day 35. Lesion scores (0 to 4 scale) for the entire length of the small intestine were not different between treatments; however, Varium 0.1 had a better effect on intestinal elasticity than the other treatments.

Beneficial Bacteria Increase with Varium

Varium was able to promote colonization of beneficial bacteria while decreasing the population of pathogenic bacteria. Varium 0.1 had significantly more beneficial Lactobacilli than zinc bacitracin and Varium 0.15 had significantly more than all treatments (Figure 2). All Varium treatments decreased the population of Salmonella in the small intestine and digesta compared to zinc bacitracin (Figure 3).

Lactobacilli colonization of the small intestine and digesta information.
Figure 2: Lactobacilli colonization of the small intestine and digesta was improved when the broiler diet was supplemented with Varium.
Salmonella colonization of the small intestine and digesta information.
Figure 3: Salmonella colonization of the small intestine and digesta was significantly reduced when the broiler diet was supplemented with Varium.

A Better Immune Response

Varium also improved the immune response to vaccination. On day 20 and 35, Newcastle disease antibody titer (hemagglutination inhibition test) was significantly higher in all Varium treatments compared to zinc bacitracin. Previous research (contact Amlan for more details, info@amlan.com) has shown that feeding Varium during disease challenge can restore the expression of immune cells that are responsible for stimulation of an antigen-specific immune response and also increase phagocytic activity compared to the control group. This increased immune response, as well as the removal of bacterial toxins that can cause immunosuppression, are thought to be the reasons behind the increase in Newcastle disease antibody titers observed in the present study.

This study confirmed that Varium can be as effective as zinc bacitracin in promoting intestinal health. The doses of Varium at 0.1 and 0.15% performed equal to or better than zinc bacitracin for the parameters tested. For more information on how Varium can improve health, production efficiency and value, visit the Varium product page.

Calibrin®-Z Improves Dairy Cattle Health and Performance in Four Commercial Case Studies

June 7, 2022
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Calibrin Z logo over feeding cows.

If there’s one goal dairy cattle producers strive to achieve, it’s production efficiency. Reproductive efficiency, feed efficiency or overall milk production — dairy producers want their cows performing to their maximum potential. But for a cow to be efficient, she also needs to be healthy.

Dairy farmers around the world face a multitude of potential health challenges in their herds; from heat stress to bacterial disease to mycotoxin-contaminated feed, it’s a lot to manage. It’s unlikely that some of these health challenges can be completely avoided but mitigating them with a natural feed additive like Calibrin®-Z can go a long way to keeping cows healthier and more likely to achieve their production potential.

A Proven Biotoxin Binder

Calibrin-Z is a mineral-based feed additive that binds bacterial pathogens and their toxins, as well as mycotoxins, protecting dairy cattle from a broad spectrum of biotoxins that reduce performance and cause morbidity or mortality. A one-ingredient feed additive, Calibrin-Z is made from our single-source calcium montmorillonite with opal-CT lepispheres that undergoes proprietary thermal processing (tailored to the product) to promote the binding of multiple biotoxins in the intestine of dairy cattle. A healthy gastrointestinal tract means a healthier, more productive cow.

Commercial Dairy Farms Recognize the Benefits of Calibrin-Z

In four commercial-based case studies, Calibrin-Z increased dairy herd performance for several key performance indicators against a wide variety of challenges and in diverse environmental conditions. In Mexico, two dairy farms, approximately 1,200 lactating cows each, were used in a four-month side-by-side study. The two farms shared silage, used the same diet formulation and feed ingredients, and the age, lactation period and condition of the cows were evenly distributed between the two farms. Mycotoxin analysis showed low mycotoxin concentrations during the trial, however, the potential to improve performance of the herd was still evident.

Adding Calibrin-Z to the ration increased milk production, enhanced feed efficiency, decreased somatic cell count and reduced death and abortion loss (Figures 1 and 2).

Milk production before and after Calibrin-Z info graphic.
Figure 1: Milk production was greater in dairy cows fed Calibrin-Z (P < 0.01)

 

Somatic cell count info graphic.
Figure 2: Somatic cell count was lower in dairy cows fed Calibrin-Z (P < 0.001)

 

Calibrin-Z also improved the health of cows from two farms in Mexico that were experiencing other kinds of health challenges. One farm had high levels of mycotoxin contamination (T-2 and DON) and a high incidence of Clostridium in the region. Calibrin-Z was able to improve the general health and production of the herd and reduce death loss. Another farm had medium levels of mycotoxin contamination and was experiencing abnormally high abortions. Heat stress was also a factor on this farm. Calibrin-Z was able to reduce abortion loss, decrease cull numbers and improve the general health of the herd.

The fourth case study was conducted on two dairy farms located in the Yamagata Prefecture, Japan, where data collected over the 88-day study was compared to the previous year’s data. Feed samples showed medium levels of mycotoxin contamination at these farms. Calibrin-Z increased milk production, improved reproductive performance and decreased disease incidence (Figures 3 and 4).

Year over year milk production info graphic.
Figure 3: Calibrin-Z improved milk production in dairy cows compared to the previous year (P < 0.05).

 

Mastitis and enteritis info graphic.
Figure 4: Calibrin-Z reduced the incidence of mastitis and enteritis in dairy cows compared to the previous year.

These commercial case studies demonstrate the benefits Calibrin-Z can bring to a dairy cattle herd, no matter the level of disease or environmental challenges the farm is facing. To start your own Calibrin-Z trial or for more information about the benefits of using biotoxin-binding Calibrin-Z, contact us at info@amlan.com.

The Culture Behind Our Mineral Science

May 3, 2022
The culture behind our mineral science.

We’re innovators of natural mineral-based feed additives that optimize intestinal health and add value for animal protein producers. But that’s not the entire Amlan story. We’re grounded by our family roots, backed by vertically integrated mineral expertise and we bring mineral-based solutions to the animal production industry that are distinctly ours.

About Amlan info graphic.

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