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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.

 

Seeing is Believing! Calibrin®-Z Binding Effect

December 7, 2023
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(Figure 1) Platinum octaethylporphyrin molecules seen adsorbed by Calibrin-Z using Cryogenic Transmission Electron Microscopy. Platinum (pink) seen under microscopy indicates where the organic compound was bound in the interconnected pores of the unique clay mineral. 

Providing Economic Value for More Than a Decade  

For more than 16 years, Calibrin®-Z (available in select international markets) has helped poultry producers mitigate the damage that mycotoxins cause to their livestock, their sustainability, and their bottom line. Calibrin-Z’s ability to bind mycotoxins, fungal toxins that negatively affect the health and performance of livestock, has been shown both in experimental settings and on the farm. The implication from this research was that the unique clay mineral that comprises Calibrin-Z was blocking the negative effects of mycotoxins on animal performance by adsorbing the toxins in the pores of the clay.  The way the binding occurred was known but had not been seen directly. 

Until Now!  

Scientists from Oil-Dri worked with university scientists to enable us to see organic molecules binding to Calibrin-Z (Figure 1). To do this they used Cryogenic Transmission Electron Microscopy.  This type of microscopy is used to look at biological and materials structures at an almost atomic level. The material of interest is flash-frozen to keep from damaging the structure of the organic material that is being observed. 

Octaethylporphyrin, is an organic molecule that was chosen to represent the mycotoxins that Calibrin-Z normally adsorbs. It has a general size and planar orientation similar to that of mycotoxins. Platinum is not an element that is typically found in the clay mineral that makes up Calibrin-Z and can be seen using cryogenic transmission electron microscopy. This combination of factors makes it an excellent marker to use to visualize Calibrin-Z’s binding sites. When the organic portion of the platinum octaethylporphyrin molecule is adsorbed onto the Calibrin-Z binding sites you can see the platinum with a cryogenic transmission electron microscope. The platinum in the picture taken under the microscope was interspersed between the layers and on the outer surface of the Calibrin-Z particles. This shows that the organic compound was bound in the interconnected pores as was anticipated.   

Selection and Quality 

Calibrin-Z’s natural ability to adsorb biotoxins is based on the clay mineral used in its manufacture. The source of the clay mineral was chosen after years of testing and comparisons of a multitude of different potential sites. This source was selected based off its innate ability to bind toxins, the ability to improve that binding with processing, and its benign chemical profile. With vertical integration, mine to market traceability, and decades of reserves, this unique clay mineral is the foundation of Amlan International’s animal health products.   

Calibrin-Z is composed mainly of calcium montmorillonite with opal lepispheres. The opal lepispheres are intimately interwoven within the nano-scale layers of montmorillonite. They help Calibrin-Z maintain its structure during a proprietary processing step that expands the number of biotoxins Calibrin-Z adsorbs. The unique structure of Calibrin-Z is vital to its toxin binding capacity.

 

Absorption and Adsorption 

A kilogram of Calibrin -Z has approximately the same surface area as 60 soccer fields.  This is because over 99% of Calibrin-Z’s total surface area is inside the particle. Calibrin-Z’s internal network of interconnected channels and pores is ~50% of its total volume. When Calibrin-Z is fed to livestock or poultry, fluid in the intestine rapidly absorbs into the mineral’s pores through capillary action. Biotoxins in the fluid move inside via the networks of capillary channels. From a molecular perspective it is as if they are traveling on a superhighway. Biotoxins adsorb once they reach the binding sites on the pores’ surfaces.  

The biotoxin molecules are attracted onto the pore surfaces via adsorption, this is both chemisorption and physisorption.  Biotoxins will structurally coordinate themselves onto charged surfaces and bind via ion-dipole and electrostatic interactions. While mycotoxins tend to be smaller and can enter the pores of Calibrin-Z and bind there, bacterial toxins tend to be larger but may also bind. Theoretically, there are special physical properties that allow the molecular conformation of the bacterial toxin to become distorted, which allows them to adsorb onto macro-surfaces within the pore spaces. Someday we may be able to use microscopy to see that, too. 

Because of its structure, the clay mineral that Calibrin-Z is made from is naturally hydrophilic and will bind to polar molecules. But Calibrin-Z undergoes a proprietary processing method  that causes dehydroxylation of the clay mineral’s crystal structure. During this process the opal lepispheres spread between the layers maintain its channels and binding sites. Thus, Calibrin-Z continues to bind polar molecules, such as the mycotoxin aflatoxin, but processing also allows it to have the ability to bind non-polar mycotoxins such as zearalenone. The ability to mitigate the effects of multiple mycotoxins has been shown using both in vitro and in vivo research.  

Because of Calibrin-Z’s proprietary heat treatment the toxins that it adorbs include a broad-spectrum of polar and non-polar toxins. Therefore, Calibrin-Z has shown high adsorption properties for mycotoxins, enterotoxins, and endotoxins.  

 

Examples of Biotoxins Bound by Calibrin-Z 

 Natural and Reliable to Use 

Calibrin-Z is shown to be a reliable and effective biotoxin binder. When added to animal feed at up to 5X the recommended dose it showed no negative effects. In fact, there was often a numerical improvement in gain, feed intake, or feed conversion when Calibrin-Z was added to an unchallenged diet. This indicates that there was no significant negative effect of Calibrin products on nutrient utilization.  

The unique surface chemistry and structural properties of the calcium montmorillonite in Calibrin-Z, added to its proprietary thermal-processing method, are what provide its optimal toxin binding capacity. This is what sets Calibrin-Z apart from other clay-based products. We have long known this because of its structure, how it works in vitro and how for more than a decade it has improved the performance of livestock and poultry. And now we, and you, are able to see it with our own eyes.  

To learn more about broad spectrum biotoxin binder Calibrin-Z, and how you can add it to your poultry and livestock feed, visit amlan.com 

Rapidly Adsorb Aflatoxins and Improve Poultry Performance with Calibrin-A

December 15, 2022
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Calibrin-A White Broilers

Mitigating the effects of mycotoxin-contaminated feed is a goal of every poultry producer in order to keep birds healthy and reduce the negative effects of mycotoxicosis on performance. Aflatoxin is a common threat to poultry productivity, targeting and damaging the liver and causing mortality in severe cases. Subclinical cases can reduce feed intake, weight gain and efficiency, which negatively impact the cost of production and, ultimately, profits.

Aflatoxin is a polar or hydrophilic (water-loving) mycotoxin, which means it and other polar mycotoxins, like ergotamine, ergovaline and cyclopiazonic acid (CPA), are attracted to hydrophilic surfaces. Calibrin®-A (available in select international markets) is a mineral-based feed additive that rapidly adsorbs these polar mycotoxins due to its hydrophilic binding sites. If mycotoxin diagnostic tests (e.g., BioInsights) detect feed is contaminated with polar mycotoxins, Calibrin-A is an effective solution for reducing performance loss.

The Calibrin-A Difference

Calibrin-A contains one ingredient — our selectively-source calcium montmorillonite with opal CT lepispheres. We select our calcium montmorillonite from a specific location within our mine to ensure product consistency, quality and reliability for customers. We’re very specific about where we source our mineral, because of its natural physical and chemical properties. We also use proprietary mineral processing methods that are tailored for each product. These unique properties are what create the difference between Calibrin-A and other clay binders in the market. Calibrin-A naturally adsorbs polar mycotoxins and is designed to have a high particle count and increased access to hydrophilic binding sites. The combination of natural mineral characteristics and processing techniques creates a highly effective, fast-acting feed additive for binding polar mycotoxins.

Rapid Polar Mycotoxin Adsorption Is Key

Eliminating the fast uptake of mycotoxins into the digestive system is key to preventing the negative health and performance effects of mycotoxicosis. Mycotoxins quickly metabolize in the intestines and liver and can circulate in the blood for days or weeks. While certain toxins enter the body more quickly than others, the negative effects consistently result in decreased performance and unrealized economic potential. Calibrin-A rapidly adsorbs polar mycotoxins, reducing their bioavailability in the body and mitigating performance loss (Figure 1).

Figure 1: Calibrin-A rapidly adsorbs aflatoxin. Source: Trilogy Analytical Laboratory, USA.

Calibrin-A Improves Performance of Aflatoxin-Fed Broilers

The impact rapid aflatoxin adsorption by Calibrin-A has on bird productivity was shown in two broiler studies. In research conducted at the University of Missouri (Columbia, MO), Calibrin-A abated the detrimental effects of aflatoxin-contaminated feed on broiler health and performance. The study compared a control diet to diets containing 2 ppm of aflatoxin, with or without 0.5% Calibrin-A, fed to day-old Ross 308 chicks for 21 days.

As expected, aflatoxin in the feed caused decreased (P < 0.05) feed intake, weight gain and feed efficiency, and increased (P > 0.05) relative liver weight compared to control birds. The liver is the main target of aflatoxin if it enters the body from the intestine. Aflatoxin will cause the liver to swell and it can become “fatty” with a yellow appearance. The swollen liver and decreased weight gain causes increased relative liver weight.

Adding Calibrin‑A to the diet of birds fed aflatoxin improved weight gain and feed efficiency (Figure 2), and reduced the mycotoxin-induced increase in relative liver weight (Figure 3). Mortality rate of the control and Calibrin-A-fed birds (2.5%) was lower than the aflatoxin-fed birds (10%).

Figure 2: Calibrin-A improved weight gain and feed efficiency in broilers fed aflatoxin-contaminated feed (P < 0.05).

Figure 3: Calibrin-A reduced the mycotoxin-induced increase in relative liver weight (P > 0.05).

In a study conducted at SAMITEC (Santa Maria, Brazil), four groups of male broiler chicks (6 reps x 10 chicks each) received a diet with or without aflatoxin (2.8 ppm) and with or without 0.5% Calibrin-A (CON, AFL, CON + Calibrin-A, AFL + Calibrin-A). Calibrin-A improved (P < 0.05) the feed intake and body weight of birds fed aflatoxin (Figures 4 and 5) and reduced (P < 0.05) the average liver weight of birds fed aflatoxin (Figure 6). Additionally, adding Calibrin-A to the control diet (no aflatoxin) at 10 times the recommended dose had no negative effects on growth performance (Figures 4 and 5), indicating that Calibrin-A does not significantly interfere with nutrient use.

In order to achieve statistical significance between the challenged and non-challenged birds, a much higher concentration of aflatoxin was used in both studies than would typically be seen in poultry diets. Because birds were challenged with a high amount of aflatoxin, Calibrin-A was also included at a higher dose than typically recommended. The ratio of Calibrin-A to aflatoxin in the feed was 2500:1 (5000 ppm Calibrin-A and 2 ppm aflatoxin) for the University of Missouri study, for example, which is equivalent to the recommended inclusion rate of Calibrin-A (500 ppm) and 0.2 ppm of aflatoxin in the feed. This amount of aflatoxin is still higher than the concentration typically found in poultry feed.

Figure 4: Calibrin-A increased feed intake in birds fed aflatoxin (P < 0.05).

Figure 5: Calibrin-A increased the average weight of birds fed aflatoxin to a weight similar to control birds (P < 0.05).

Figure 6: Calibrin-A decreased the average liver weight of birds fed aflatoxin (P < 0.05).

These studies demonstrate the performance and health benefits of feeding Calibrin-A to rapidly adsorb polar mycotoxins like aflatoxin. Single-ingredient, mineral-based Calibrin-A is an effective solution to mitigating the risk of mycotoxicosis from aflatoxin-contaminated feed. To learn more about Calibrin-A or to try Calibrin-A for yourself, contact your local sales representative.

 

 

Poultry Science Study Shows NeutraPath® Targets Salmonella Isolate Using Multiple Methods

April 28, 2022
NeutraPath® logo with packaged poultry in background.

Source: Xue H, Wang D, Hargis BM, Tellez-Isaias G. Research Note: Virulence gene downregulation and reduced intestinal colonization of Salmonella enterica serovar Typhimurium PHL2020 isolate in broilers by a natural antimicrobial (NeutraPath™). Poultry Science. 2022 Mar 7:101822. https://doi.org/10.1016/j.psj.2022.101822.

Reducing intestinal Salmonella colonization in poultry is a key strategy in controlling Salmonella contamination of poultry products and, in turn, lowering the incidence of salmonellosis in people. Subtherapeutic levels of antibiotic growth promoters (AGP) can help control enteric pathogens like Salmonella, but restrictions in AGP use have created the need for antibiotic-free methods of reducing enteric pathogens in poultry.

A natural mineral-based feed additive that has previously shown action against Salmonella prevalence is NeutraPath® — a select blend of essential oils, fatty acids and a thermally processed enterosorbent mineral. A recent Poultry Science study investigated the antimicrobial effects of NeutraPath against Salmonella enterica serovar Typhimurium strain PHL2020 (ST-PHL2020) and the effects of NeutraPath on ST-PHL2020 virulence gene expression.

The study showed that NeutraPath exhibited a potent antimicrobial effect against ST-PHL2020 and reduced its intestinal colonization. NeutraPath also modulated ST-PHL2020 virulence network development by downregulating mRNA expression of key virulence genes and blocking expression of downstream effectors involved in Salmonella invasion. Together, the results show that NeutraPath has the potential to reduce ST-PHL2020 intestinal colonization in broilers and downregulate key ST-PHL2020 virulence genes.

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Poultry Producers’ Important Role in Reducing the Global Salmonellosis Challenge

March 23, 2022
Microscopic salmonella with Varium logo text graphic.

Salmonella is one of the most prevalent foodborne zoonotic pathogens worldwide. However, by using strategies that reduce the contamination of poultry products at the farm and processing plant levels, poultry producers and processors can play an important role in reducing the incidence of salmonellosis and the emergence of antimicrobial-resistant Salmonella strains.

Poultry-Related Salmonellosis

Salmonellosis is a common human foodborne illness and one of four key global causes of diarrheal diseases in people according to the World Health Organization. Poultry-related salmonellosis is typically caused by Salmonella spp. passing from poultry to people through contaminated eggs and meat. Poultry are often asymptomatic carriers, and their intestinal tracts serve as pathogen reservoirs, potentially leading to contamination of food products.

Salmonella Transmission

To enter the human food chain, Salmonella must first colonize the bird’s intestinal tract. After colonization, Salmonella can spread via horizontal transmission (bird to bird), contaminating the environment and the carcass during slaughter. Salmonella colonization of the cecum can also result in vertical transmission (parent to progeny) through contamination of the yolk, albumen and eggshell membranes.

Reducing Salmonella Contamination

Salmonella can contaminate meat products during processing, causing contaminated poultry carcasses to serve as a source of infection in consumers. Innovative technology provides processors with methods to reduce contamination at the poultry plant; however, control of Salmonella at the farm level is also an important step in reducing the risk of salmonellosis in people.

Antimicrobial-Resistant Salmonella Strains

Antimicrobial-resistant pathogens, which include strains of Salmonella, are a major concern for public health care worldwide. The U.S. Centers for Disease Control and Prevention (CDC) reported that over a three-year period, an average of 16% of all nontyphoidal Salmonella were resistant to at least one essential antibiotic.

The concern over antimicrobial resistance (in all pathogens, not just Salmonella) has led to a global effort to reduce the use of in-feed antibiotics in poultry production in an effort to slow the emergence of antimicrobial-resistant pathogens. This presents a challenge for poultry producers since they are still being urged to control Salmonella in the poultry barn to reduce contamination of meat during processing.

Reduce Salmonella with a Non-Pharmaceutical Solution

A natural feed additive that producers can use to help limit Salmonella in poultry is Varium® — a patented mineral-based product sold in Amlan’s international markets. Varium enhances multiple aspects of the intestinal environment, creating production results consistent with those observed with antibiotic growth promoter use. The patented technology in Varium includes a synergistic formulation of three ingredients with distinct modes of action: Varium reduces levels of pathogenic bacteria and their toxins in the intestinal lumen, acts as an enterocyte energy source, and stimulates the intestinal immune system to help birds naturally defend against pathogens.

Varium has been shown to agglutinate (adsorb) Salmonella spp., which can help prevent colonization of the intestinal wall and subsequent proliferation (Figure 1).

First Salmonella Close-Up Stage 8 Info Graphic | Amlan International
Second Salmonella Close-Up Stage 8 Info Graphic | Amlan International
Figure 1: Agglutination (adsorption) of Salmonella spp. by Varium. The scanning electron microscopy images were taken at 4 μ (top) and 20 μ (bottom). Images courtesy of the University of Georgia, Athens, GA.

Supporting the in vitro agglutination results, Varium also reduced Salmonella colonization in vivo in a 28-day broiler trial conducted at Imunova Análises Biológicas (Curitiba, Brazil). In this study, broilers challenged with Salmonella enterica serovar Enteritidis and supplemented with Varium had a 5-log reduction in cecal Salmonella levels on day 14, compared to the challenged control, and reduced overall Salmonella levels (Figure 2).

Salmonella and Public Health Concerns info graphic.
Figure 2. Compared to the challenged control, treatment with Varium rapidly reduced the bacterial load in the cecum as indicated by the Salmonella most probable number (MPN). Different letters indicate a significant difference between groups on day 14, and a main treatment effect of P = 0.0526 was also observed.

Salmonellosis and antimicrobial-resistant Salmonella strains are important global public health concerns. However, with the assistance of natural mineral-based feed additives like Varium, poultry producers can help reduce the Salmonella risks for consumers at the farming stage. To learn more about Varium, click here.

The Distinctive Properties of Our Biotoxin Binder Calibrin®-Z

March 22, 2022
Calibrin-Z binding with Amlan logo info graphic.

Proprietary mineral technology is the foundation for Amlan’s innovative value-added products for animal protein producers. In this article, we take an in-depth look into the mineral technology used in our all-natural feed additive Calibrin®-Z and its unique properties that are the Amlan difference.

Consistent, Controlled Mineral Supply

The physical and chemical properties of a mineral can differ depending on where it is mined. That is why — to ensure consistent quality — Amlan only uses a single-source mineral in our products. Amlan is vertically integrated as the animal health business of Oil-Dri® Corporation of America, allowing Amlan and Oil-Dri to control every step of the production process and reliably deliver safe, high-quality products.

Calibrin-Z: Our All-Natural Broad-Spectrum Biotoxin Control Product

Calibrin-Z protects poultry and livestock health and performance by binding intestinal pathogens, bacterial exotoxins and endotoxins and polar and nonpolar mycotoxins. It is composed of a single ingredient — our proprietary mineral technology, thermally processed to create the specific physical and chemical properties that give Calibrin-Z its powerful mode of action.

A Network of Interconnected Pores

The distinctive properties of Calibrin-Z include a high surface area and extensive porosity. More than 99% of Calibrin-Z’s total surface area is internal due to the product’s structural properties. This means that targeted molecules can migrate via interconnected networks of capillary channels towards internal binding sites. These physical features provide Calibrin-Z with a high adsorption capacity for binding a broad range of mycotoxins, bacterial pathogens and their toxins.

Layers Within Layers

The mineral in Calibrin-Z is a particular type of phyllosilicate (“phyllo” meaning sheet) and is primarily calcium montmorillonite with amorphous opal-CT lepispheres and other minor and trace minerals.

Phyllosilicates consist of silicon, oxygen, magnesium and water molecules, and either aluminum or iron atoms. The aluminum, iron or magnesium atoms form octahedron structures, whereas the silicon forms tetrahedrons. These formations give the mineral a nano-scale structure of a 2:1 layer of octahedrons between tetrahedrons. Between the 2:1 layers are interlayers of water molecules and cations (Figure 1). Various positively charged sites in the mineral structure — interlayer cations and broken edge octahedral units — provide the adsorption sites.

Structure of Mineral in Calibrin-Z Binding Info Graphic | Amlan International
Figure 1: A progressive view of the structure of the mineral in Calibrin-Z down to the nano-scale layers.

Proprietary Thermal Processing

Typical montmorillonites have water molecules between the mineral layers that make the pores and surfaces hydrophilic for adsorbing hydrophilic (polar) molecules (e.g., aflatoxins) but do not bind hydrophobic (nonpolar) molecules (e.g., zearalenone and fumonisin). However, the montmorillonite used in Calibrin-Z undergoes proprietary thermal processing that uses an optimized temperature and time to allow adsorption of hydrophilic and hydrophobic toxins (Figure 2).

Thermal Processing of Calibrin-Z Info Graphic | Amlan International
Figure 2: Thermal processing of Calibrin-Z allows binding of hydrophilic and hydrophobic molecules.

Thermal processing eliminates most of the water molecules from the mineral in Calibrin-Z, making it more hydrophobic. The process is carefully controlled since excessive heat that completely dries the mineral — removal of the interlayer water molecules — would destroy Calibrin-Z’s binding capabilities. The naturally occurring opal-CT lepispheres help maintain the layered sheet structure of the mineral during processing and provide Calibrin-Z’s high binding capacity (Figure 3). Amlan’s proprietary processing method also avoids the use of harmful chemicals typically used by other companies preserving a natural composition.

Structure and Processing of Calibrin-Z Info Graphic | Amlan International
Figure 3: Naturally occurring opal-CT lepispheres maintain Calibrin-Z’s structure during thermal processing (removal of a controlled amount of interlayer water molecules).

A Variety of Binding Mechanisms

Calibrin-Z’s binding forces include hydrophobic interactions, chelation, electrostatic attractions, hydrogen bonding and van der Waals forces. Thermal processing allows an interaction between both polar hydrophilic molecules and non-polar hydrophobic molecules and the inter-mineral layer. This is the method used to adsorb mycotoxins to Calibrin-Z.

Bacterial exotoxin binding to Calibrin-Z occurs through molecular ion exchange mechanisms. For example, a part of the Clostridium perfringens alpha-toxin electrostatically anchors (tethers) to either the positively charged broken-edge sites (exposed alumina octahedra) or the positively charged interlayer cations of Calibrin-Z.

Molecular conformation change mechanisms are also possible binding methods. Large exotoxins can distort their molecular structures or conformations to adsorb themselves onto macro-surfaces within the pore spaces.

Compatible With Nutrient Availability

While Calibrin-Z excels at binding biotoxins, its binding abilities do not interfere with the absorption of important nutrients in the diet. It is possible that some minor quantity of nutrients could temporarily be absorbed into Calibrin-Z’s pores. However, this is via weak thermodynamic and kinetic interactions that are readily reversible. Therefore, nutrients can travel in to and out of Calibrin-Z particles based on concentration gradients in the gastrointestinal tract.

A 42-day swine study conducted by SAMITEC in Brazil, examined the performance of pigs fed a common basal diet (Control) and Calibrin-Z included at 5 kg/MT, a level that is 10 times the recommended dose. Even at this very high inclusion rate, Calibrin-Z had no adverse effects on nutrient availability, supporting equivalent weight gain and feed conversion.

Body Weight and Calibrin-Z Info Graphic | Amlan International

The proprietary mineral technology used in Calibrin-Z is what sets it apart from other companies’ mineral-based products. For more information about Calibrin-Z and how it can help protect your animals from the deleterious effects of biotoxins, contact us at info@amlan.com.

Peer-Reviewed Study Shows Gut Health Benefits for Weaned Pigs Fed NeutraPath®

March 7, 2022
NeutraPath® for production efficacy in pigs logo graphic.

Source: Yijie He, Cynthia Jinno, Chong Li, Sara L Johnston, Hongyu Xue, Yanhong Liu, Peng Ji. Effects of a blend of essential oils, medium-chain fatty acids, and a toxin-adsorbing mineral on diarrhea and gut microbiome of weanling pigs experimentally infected with a pathogenic Escherichia coli. Journal of Animal Science. 100 (1), January 2022: skab365, https://doi.org/10.1093/jas/skab365.

 

Poor production efficiency and reduced resistance to bacterial disease are common issues for pigs during the weaning period. Traditionally, prophylactic antibiotics were used to help pigs through this stressful time; however, increasing regulations and consumer demands have reduced or eliminated the use of antibiotics in some swine production facilities. Therefore, natural alternatives are needed to help support pigs’ health and performance through this transition period.

In a paper published in the Journal of Animal Science, the authors (from UC-Davis and Amlan International) investigated the use of the Amlan product NeutraPath® as a natural support during weaning without antibiotics. NeutraPath is a formulated blend of functional feed additives and a toxin-binding mineral that has previously shown potent bactericidal and bacteriostatic effects in vitro, and efficacy against bacterial infections in broiler chickens. He et al., examined the effects of NeutraPath on growth, diarrhea incidence and the gut microbiome of enterotoxigenic E. coli-challenged weaned pigs.

The study found that NeutraPath reduced the frequency of diarrhea in ETEC-challenged pigs, indicating a greater resistance to disease. Additionally, changes in the fecal microbiome and ileal mucosa microbiota composition suggested NeutraPath may help maintain desirable balance in the intestinal microbial ecosystem. NeutraPath-fed pigs also had better performance during the recovery period. Collectively, the results demonstrated that NeutraPath has potential to play a key role in supporting pig health and performance during the weaning period as antibiotic use is decreased.

Read the paper here: https://doi.org/10.1093/jas/skab365.

Scientific Expertise and a State-of-the-Art Facility: The Foundation for Developing Amlan’s Natural and Efficacious Products

February 18, 2021
Still image from INNOVATION NATION interview with Dr. Hongyu Xue MD, PhD, at Research Laboratory.

With cutting-edge equipment and state-of-the-art facilities, Amlan’s talented Life Sciences team creates value-adding mineral-based products for poultry and livestock producers. The Richard M. Jaffee Center for Applied Microbiology houses the specialized equipment used by the Life Sciences team who draw upon their extensive research experience and knowledge base to create novel, natural solutions that improve poultry and livestock health and production efficiency.

Life Science Experts

The Amlan research team is led by Life Sciences Director Hongyu Xue MD, PhD, an experienced scientist in clinical and animal nutrition, who brings a unique point of view to Amlan’s research by leveraging his expertise in human medicine. Dr. Xue has a background in academic medicine and research that cross-links gastroenterology, immunology, microbiology and nutrition. Working with Dr. Xue is a multi-disciplinary team of scientists with expertise across microbiology, animal nutrition, chemistry and material science areas. Several members of the Life Sciences team were recently featured on an episode of “Built in America: INNOVATION NATION” on the Fox Business Network, where they showcased some of their novel research.

Cutting-Edge Equipment

The state-of-the-art equipment in the Richard M. Jaffee Center for Applied Microbiology allows the Life Sciences experts to use a variety of research techniques to develop and assess new mineral-based products for the animal health market.

The Life Sciences team enhances Amlan’s unique mineral to develop natural solutions that can control the microbial pathogens that negatively impact the health and productivity of poultry and livestock. To do this, the scientists study these pathogens and their toxins to understand their physiology and mode of action. Some pathogens require anerobic conditions to survive, so these pathogens are cultivated in Amlan’s anerobic chamber — a more efficient and robust method than alternative techniques such as anerobic jar or pouch systems.

Conditions within the chamber can perfectly mimic the anerobic environment of the distal gastrointestinal (GI) tract (e.g., ileum and cecum) which serves as the primary colonization site for a vast variety of common microbial pathogens for poultry and livestock. The anaerobic chamber is of tremendous value to help Amlan’s scientists cultivate and further characterize the target pathogens and select commensal microbial populations colonizing the distal GI tract. Further, the chamber also enables the scientists to screen novel products in development and evaluate their antimicrobial effects for certain anaerobic pathogens. Novel strains of beneficial bacteria (probiotics) can also be isolated, grown and tested in the anerobic chamber. The photo below shows an example of the zone of inhibition surrounding an Amlan-developed probiotic (right) versus a water control (left).

Close-up of petri dish.

Further assessment of the new products can be conducted using equipment such as an ICP-OES (inductively coupled plasma-optical emission spectrometer) that allows the team to examine how different cations released by Amlan’s unique mineral affects bacterial virulence. Some metals (cations) are known to suppress the expression of virulence genes in bacterial pathogens.

The lab also includes a fluorescent microscope that enables microbes to be observed instantly to see a product’s effect on bacterial morphology and viability.

Using real-time polymerase chain reaction (PCR), the Life Sciences team have developed methods to detect toxin genes from microbial pathogens that can have negative effects on poultry and livestock production. Using this information, natural solutions can be developed that disarm these pathogens and help protect birds and animals from disease. For example, real-time PCR is used to investigate the effects of new products on pathogen virulence gene expression. Further, this technology can help determine the copy numbers of specific virulence genes and help make an early diagnosis of specific enteric diseases in poultry and livestock.

The Richard M. Jaffee Center for Applied Microbiology is named after the former chairman of Oil-Dri Corporation of America, doing business as Amlan International. Jaffee’s pioneering spirit and vision for Oil-Dri to conduct research in the life sciences is the influence behind Amlan’s focus on developing value-added next-generation animal health products.

To learn more about the research behind Amlan’s natural and efficacious products, visit https://amlan.com/blog/category/research-studies/.

Varium®: An Effective Alternative to AGPs for Poultry Immunity and Intestinal Integrity

December 4, 2020
Computer-generated illustration of intestinal bacterial activity in poultry.

Protecting gut health, maximizing feed efficiency and increasing growth rates in poultry has traditionally been achieved with antibiotic growth promoters (AGPs). But with the rise of antibiotic-resistant bacteria and consumer demand for antibiotic-free (ABF) protein, the worldwide poultry industry is migrating toward ABF production systems. Poultry producers today need viable, profitable and natural alternatives to AGPs that can help maintain gut health, support efficient feed use and promote growth.

Varium® is a natural performance additive that enhances multiple aspects of the intestinal environment, creating production results consistent with those observed with AGP use. In the intestinal lumen, Varium reduces levels of pathogenic bacteria and their toxins, protecting the intestinal lining from attack. Varium also acts an enterocyte energy source, fostering healthy and strong enterocytes that can better absorb nutrients and support growth. Additionally, Varium stimulates the innate immune system to help birds naturally defend against pathogens. Continue reading to view the research demonstrating the beneficial effects Varium has on poultry immunity and intestinal integrity.

Improved Immune Competence

Birds with a healthy gut have a competent immune system that responds appropriately and is less susceptible to disease-causing bacteria and viruses. In a study conducted with Salmonella-challenged broilers at Imunova Análises Biológicas (Curitiba, Brazil), Varium helped restore immune competence and ultimately favored the development of appropriate defenses against the pathogen. The improved immune competence was demonstrated by the apparent prevention of cytotoxic T cell terminal activation (CD8+CD28– phenotype) which, when it occurs in large numbers, can render the immune system less responsive and competent in fighting against pathogen infections. Varium also restored major histocompatibility complex class II (MHC II) expression, essential for the stimulation of an antigen‑specific immune response, and increased monocyte phagocytic activity compared to the Salmonella-challenged control group. For further details of this study, contact Amlan (info@amlan.com).

Responsive to Immune System Stimuli

The ability of the immune system to prevent pathogens from establishing a successful infection is vital to keeping birds healthy and productive. The immune response to various stimuli was assessed in two Varium field trials by measuring the antibody titer from two common vaccines and assessing the prevalence of bacteria in the small intestines and digesta.

In a field trial conducted at a commercial farm in Vietnam, broilers were fed a basal diet and coccidiostat with either enramycin (at the manufacturer’s recommended dose) or Varium (0.1%) for the first 28 days. From day 29 to the end of study (either day 35 or 42), the control broilers were fed the basal diet only and the Varium group was fed the basal diet plus Varium (0.1%). Sub-samples from randomly selected birds were obtained and the data analyzed at Nong Lam University, Ho Chi Minh City.

In this trial, the infectious bronchiolitis virus (IBV) antibody titer of Varium-fed broilers was significantly increased on day 15 (P < 0.05 vs. antibiotic-fed control) and similar on day 35 to the antibiotic-fed control. Newcastle disease virus (NDV) titers were also similar on days 15 and 35 in the control and Varium groups. Antibody titers indicate the strength of the acquired immune response to vaccination. These results show that Varium can stimulate an antibody production response to vaccination that is the same as or better than broilers fed AGPs.

In another trial conducted at a university in Pakistan, broilers were fed either an AGP (zinc bacitracin, 0.01%) or Varium (0.10%) for 35 days. Varium fed birds had a higher (P < 0.05) concentration of the beneficial bacteria Lactobacilli and a lower concentration of the pathogenic bacteria Salmonella in the small intestine and digesta. This demonstrates that Varium in the diet was able to maintain a healthier intestinal microbiota. The antibody titer for NDV was also greater for Varium-fed broilers than control birds (Figure 1).

Figure 1: The Newcastle disease virus (NDV) antibody titer (by hemagglutination inhibition assay) was significantly higher (P < 0.05) in Varium-fed broilers than AGP-fed broilers on day 20 (14 days after first vaccination [intraocular and subcutaneous]) and day 35 (14 days after second vaccination [oral booster]). Different letters indicate significant differences (P < 0.05) between groups within day.

Improved Intestinal Integrity

Along with immune competence, the structure and functional integrity of the intestine is also key to reducing the risk of infection. A healthy intestinal tract and competent immune system improve the ability of the bird to block the invasion of pathogens into intestinal epithelial cells and the circulatory system.

In the Imunova Análises Biológicas study, use of a fluorescent marker demonstrated that on days 4 and 8, the increased intestinal permeability observed in the Salmonella-challenged control was mitigated with the addition of Varium to the diet (Figure 2). The reduced permeability confirmed that Varium helped maintain the structural and functional integrity of the intestinal barrier. Varium also effectively reduced excessive migration and infiltration of lymphocytes into the cecal wall, which helped dampen the inflammatory damage and improved intestinal integrity seen in Varium-treated broilers.

Figure 2: Intestinal integrity as measured by the passage of a marker. Salmonella infection resulted in increased passage of a marker from the intestine to blood on days 4 and 8 following bacterial challenge, indicating impaired mucosal integrity. Compared to the Salmonella-infected control, Varium effectively mitigated increased intestinal permeability on days 4 and 8 (P < 0.05 vs. infected control). Different letters indicate significant differences (P < 0.05) between groups within day.

Necrotic Enteritis Scores

The reduction in AGP use has triggered an increase in the occurrence of necrotic enteritis in poultry flocks. Necrotic enteritis is caused by Clostridium perfringens and can cause significant production losses. In the trials conducted in Pakistan and Vietnam, the intestinal lesion score was not different between the Varium and antibiotic-fed groups, indicating that Varium was able to reduce the occurrence of necrotic enteritis to the same extent as the AGP.

Figure 3: Necrotic enteritis lesion score (Day 35) was numerically lower in broilers fed Varium versus broilers fed AGPs.

Varium: Feed Efficiency for Poultry

These trials demonstrated that replacing AGPs in broiler diets with Varium can maintain the immune response and intestinal integrity observed with AGP use and can also potentially improve them. Varium also helped restore the immune response in pathogen-challenged broilers. The direct benefits of the immunity and intestinal integrity results in the field trials was confirmed with growth performance being similar or better than broilers fed AGP. For more information on how Varium improves productivity visit, amlan.com/varium.

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