Antibiotics as Growth Promoters Are Ending, So What's the Fuss?

Over the years research produced improvements in growth as high as 20% and improvements in feed efficiency as high as 15%. At today's broiler numbers (7.5 billion) these improvements were valued at $500 million in broilers alone in the US. Improvement in gain and feed efficiency in broilers is now only 1 or 3%. Such improvements now are probably valued at less than $100 million. Why is there this backward slide in the benefits achieved from antibiotics as growth promoters?

Antibiotics improve growth only in the presence of microbes. If animals are raised in germ-free environments animals grow faster, but antibiotics do not enhance performance. Microbial suppression of growth is not directly due to the microbes, but due to the immune system stimulation, resulting in a suppression of food intake and a depression in growth. Direct injection of the immune hormones released during the immune response will depress growth in a similar manner as the exposure to microbes. However, in today's consolidated animal environments, the use of antibiotics to reduce the level of immune stimulants became the most practical means of enhancing animal performance.

Genetic selection of animals for increased growth and improved feed efficiency has probably played a major role in lowered efficiency from the feeding of antibiotics. Continued genetic selection for growth and efficiency in immune-stimulating environments theoretically selects for animals less likely to have immune responses that induce reduced performance. This can be visualized by considering the ecosystem of the chicken house. When the top 20% performing broilers are repeatedly selected out of a population of primary breeding stock, the birds most likely to thrive in performance are those with the poorest immune response. Since antibiotics enhance growth by reducing the presence of immune stimulants, repeated selection of birds for enhanced growth would result in less growth-promoting effects of antibiotics. We have conducted studies with primary breeding poultry strains that suggested selection for growth and feed efficiency did modify the immune response. Birds selected for improved performance traits have poorer macrophage function and decreased cell-mediated immunity.

While it is becoming evident that genetic selection in immune-stimulating environments will probably eliminate the use of antibiotics as growth promoters in the poultry industry, it is less clear if the immune modification from generation to generation is sustainable. How much immunity is enough? The answer is not yet available.

• Alicyclobacillus
• Mycotoxin Research Plans
• CLA Revisited

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We characterized two isolates (YSG1 and YSG2) from an apple juice packaged by a hot-fill process. YSG1 grew at pH values ranging from 2.5–5.5 and temperatures from 20–55ºC. YSG2 grew at pH 2.0–6.0 and temperatures from 40–70ºC. The growth of YSG1 in shelf-stable juices was investigated at temperatures of 20, 25, 30, and 37ºC. YSG2 was not tested because its minimum growth temperature is 40ºC. YSG1 did not grow in any of the juices tested at 20 or 25ºC, but grew at 30 and 37ºC in several apple-based, punch-flavored, berry and strawberry juices.

During 48 h of storage, the numbers of YSG1 increased >2 log10 CFU/ml in the apple-based juices and 1 log₁₀ CFU/ml in the punch, berry and strawberry juices. The greatest numbers of YSG were detected in juices incubated at 37ºC. Considering that Alicyclobacillus can survive heat-treatment processes and grow at 30ºC indicates that it should be viewed as a potential spoilage organism, particularly of apple juices. A survey of apple juice and apple juice blends would provide additional data on the prevalence and significance of this spoilage organism.

Well-known examples of major mycotoxins include aflatoxin (sterigmatocystin), ochratoxin A, trichothecenes (DON) and fumonisin that are produced by various aspergilli, penicillia and fusaria. My research is focused on understanding how fungal sporulation and biosynthesis of these mycotoxins are regulated in Aspergillus and Fusarium. This should provide us new approaches to control fungal infection, sporulation and mycotoxin contamination.

One of my immediate investigations is specifically aimed for two Fusarium mycotoxins, trichothecenes and fumonisins. Fumonisins are polyketide mycotoxins that frequently contaminate maize, and thus maize-based foods and feeds throughout the world. They cause liver and kidney toxicity in animals, in addition to leukoencephalomalacia in horses and pulmonary edema in pigs. They also cause cancer in rats and mice. Fumonisins are produced by various Fusarium species. However, production by Gibberella fujikuroi mating population A (MP-A) (anamorph, Fusarium moniliforme, syn. F. verticilloides) is of particular concern because this species is prevalent in maize where it causes root, stalk and ear disease, and where it is frequently present in healthy tissue.

Trichothecenes are produced by a variety of different Fusarium species. They are immunotoxic and potent inhibitors of protein synthesis, which can predispose animals to other diseases. Dietary exposure to trichothecenes can lead to increased susceptibility to other microbial infection. Pigs and other monogastric animals, also humans, appear to have the greatest susceptibility to these toxins. One of the most prevalent trichothecene-producing fungi is Fusarium graminearum; it has been found to contaminate corn, wheat, barley, rye, oats and rice.

These Fusarium species and mycotoxins have been a health hazard to humans and domestic animals for a long time, but until the past 30 years their effects have been largely overlooked. Recently, genes involved in trichothecenes and fumonisin biosynthesis have been identified, though it is unknown what upstream genetic elements are associated with their control. One of my research objectives is to identify upstream genetic elements that function in Fusarium sporulation and mycotoxin biosynthesis.

As a model system, I have employed a filamentous fungus, Aspergillus nidulans, to study fungal growth and mycotoxin biosynthesis. This fungus provides a sophisticated experimental system for genetic and molecular analyses for multicellular development, as well as mycotoxin biosynthesis. It is known there are two antagonistic signaling pathways: growth and sporulation. A model has been proposed in which asexual sporulation and mycotoxin biosynthesis in A. nidulans are linked by a common need to slow growth by inactivating a growth-signaling pathway mediated by a heterotrimeric G-protein. I have found that this regulatory mechanism is present in other aflatoxin-producing fungi and could provide a means of controlling aflatoxin contamination. I strongly believe that controlling a G-protein signaling pathway for growth or development-specific signaling could be a way of eliminating aflatoxins (and other mycotoxins) from the field.

In next 12 to 18 months, the above-proposed model will be tested in major Fusarium species, F. graminearum and G. fujikuroi. Initially the major genetic elements (G-protein subunits and regulator of G-protein signaling) will be identified using reverse genetics approaches. Once these major genetic elements are identified and functionally characterized, further investigation will be done to learn the signal pathways in Fusarium species.

A postdoctoral research associate, Dr. Jeong-Ah Seo, will join my laboratory in November 2000. She was trained in Fusarium mycotoxin research for her Ph.D. and has continued to work on fumonisin biosynthetic genes in the Western Regional USDA laboratory in Peoria, IL. Soon a couple of graduate research assistants will come to my laboratory, too. My research team hopes to make good progress in Fusarium sporulation and mycotoxin research.

The anticancer effect in animals and weight modification properties of CLA were first reported by FRI researchers in the 1980s. Gradually several laboratories in the U.S. and other countries have become interested, and CLA is now being studied extensively.

Mike Pariza from FRI reported new work with 80 obese people, half taking 3 grams of CLA supplement daily, the other half on a placebo. Everyone was put on a diet and encouraged to exercise. Weight loss was about the same for both groups, an average of 5 pounds. However, during the trial, some subjects gained lean muscle. Those on CLA and those on placebo gained weight differently. Those taking the placebo gained at a ratio of 75% fat and 25% lean muscle. The CLA group gained weight that was about 50–50 fat and lean.

While it is apparent CLA causes the body to favor lean muscle over fat, Pariza stated "It's nothing of the magnitude of an anabolic steroid." An interesting additional observation was that some of the CLA dieters found it easier to stay on the diet and had reductions in blood pressure and LDL cholesterol.

One report from Norway cited small but statistically significant reductions in body fat gain. Another Norwegian report showed that subjects on CLA lost statistically significant amounts of weight without being on a diet. Once again weight losses were small, a 160-pound person losing 2–3 pounds over 12 weeks.

In a study of insulin-resistant diabetes at Purdue University (22 subjects, 8 weeks) 64% showed improvement in their insulin levels while on CLA. Insulin levels in this type of diabetes are often excessive as the body tries to overcome its inability to process glucose.

Pariza thinks CLA may help block fat cells from growing. He said, "It has very little effect on a cell that is already fat, but may be limiting growth in other fat cells."

Dr. Jaehyuk Yu joins FRI as an Assistant Professor My research area is the genetics and biochemistry of mycotoxin biosynthesis. I started mycotoxin research when I joined Dr. F. S. Chu's (former professor at the FRI) laboratory as an M.S. student in 1989. I received my degree with "Immunochemical study of fusarochromanone mycotoxins" in Food Science (Food Microbiology option) in 1991, then transferred to the Genetics department here. I received the Ph.D. in Genetics, in May 1995, with "Genetic studies of secondary metabolism in Aspergillus nidulans: A gene cluster associated with sterigmatocystin biosynthesis and its regulation." Then I did my postdoctoral research at Texas A&M University in "G-protein signaling and its regulation for coordinate control of fungal growth, development and mycotoxin biosynthesis." After 2 years and 9 months postdoctoral research I joined Cereon Genomics LLC, located in Cambridge, MA, as an investigator. There, I worked on fungal and plant genomics. A part of my role was development and utilization of transcript profiling systems in Aspergillus and Arabidopsis. It was a good experience for me working in a company like Cereon. There I learned a lot — not only advanced science, but also leadership and management skills. In 2 years and 3 months there my team and I accomplished several challenging projects. However, something was still missing: having my own focused research program in fungal sporulation and mycotoxin biosynthesis. Fortunately, the UW–Madison was looking for somebody like me! In July 2000, I joined FRI as an assistant professor. A long-term goal of my research is to eliminate/prevent mycotoxin (and fungal) contamination from the field. Well-known examples of major mycotoxins include aflatoxin (sterigmatocystin), trichothecenes (DON), and fumonisin that are produced by various aspergilli and fusaria. My research is focused on understanding what regulatory mechanisms are involved in biosynthesis of these three mycotoxins. Genetics (both forward and reverse), G-protein-mediated signal transduction, and biochemistry of small molecules are examples of my study areas. Hopefully my team will find ways to eventually eliminate fungi and mycotoxins from our foods and feeds. Ever since I left Madison in May 1995, I confirmed that Madison was one of the best places to live. I missed this beautiful place and the excellent University so much. Truly, it is my great pleasure and honor to be a FRI faculty member.  I am originally from the Republic of Korea (South Korea). I went to the Seoul National University, Department of Microbiology for my Bachelor's degree. I've been married almost 11 years to my wife Jeongmee and we have three wonderful children, David (10), Catherine (8), and Angela (4). David and Catherine are Madisonians (born in Madison) and Angela is a Texan. They all love Madison, too.

Assistant Professor  Dr. Jaehyuk Yu

Workshop

Workshop attendees

M. Pariza, F. Chu, R. Weiss, and A. Wong participated in the "Managing Technology from Research to Market" short course on August 10, 2000. The objective of the course was to help participants improve their management of technology research and their ability to move new technology from the laboratory to the market. This course stems from the Asian Partnership Initiative of the UW. Twenty Deputy and Assistant Directors from various Institutes of the Chinese Academy of Sciences attended this workshop (August 7–25, 2000).

FRI at the University of Wisconsin Open House   Members of FRI who helped staff our booth at the College of Agricultural and Life Sciences portion of the recent University of Wisconsin open house. The booth provided information and publications on practical food safety in the home. L to R: Kathy Glass, Ron Weiss, Jean Schoeni, and Eileen Somers.

• Microcystins in Drinking Water
• Foodborne Pathogens Implicated in Autism?

High doses of microcystin severely damage liver cells, which can result in internal bleeding, shock, and death. In 1996, 55 dialysis patients in Brazil died after exposure to dialysis water containing cyanobacteria toxins. At lower concentrations, microcystins exert a tumor-promoting effect in experimental animals, and recent research suggests that chronic exposure to microcystins in drinking water may be one factor responsible for the high incidence of primary liver cancer in China. Microcystin levels in Donghu Lake and in a fish pond in Wuhan, China, were found to peak at 0.3 and 0.55 mg/L, respectively, during the late summer to early autumn. Because water from these sources is consumed by local people, the possibility exists for chronic, low-level intoxication for several months during the year.

But microcystin contamination is not just a problem for people using surface waters for drinking. High levels of microcystins were also detected in drinking water pumped from an aquifer near Riga, Latvia. The lakes surrounding Riga have become eutrophic because of the input of domestic and industrial wastes. This lake water is not directly consumed by humans but surface water is pumped to infiltration basins from where it percolates through soil and sand into the groundwater. Apparently microcystins are neither efficiently adsorbed nor degraded during this filtration: ground water pumped out to supply drinking water for the city in October 1996 was found to contain 0.6–1.47 mg microcystin/L.

Relatively high concentrations of microcystins were also detected in some lakes in Alberta, Canada. Chemical analyses showed that average microcystin levels ranged from < 0.05 mg/L to > 1.5 mg/L with the highest concentration reported as 6.17 mg/L. Microcystin levels were greater in lakes with relatively higher phosphorus concentrations. Limiting phosphorus input into lakes may aid in reducing levels of cyanobacterial toxins.

Could this disruption of normal intestinal flora by long-term use of antibiotics be somehow causally related to development of autism? There are numerous reports in the literature of an increased incidence of opportunistic infections in adults and children by Salmonella, Shigella, Clostridium difficile and other bacteria following disruption of protective intestinal microbes by antimicrobials. There is also a suggestive parallel from the data on infant botulism. Clostridium botulinum spores do not germinate, grow, and produce toxin in adults with normal intestinal flora. However, young infants with an inadequate population of intestinal microbes are susceptible to infection with neurotoxic C. botulinum. Therefore, it is hypothesized that regressive-onset autism in some children could result from disruption of normal intestinal flora followed by colonization of the gut by a neurotoxin-secreting bacterium.

If toxins secreted by intestinal bacteria interfere with neuronal activity in the brain, would it then be possible to reduce toxin levels and alleviate symptoms of autism by appropriate antibiotic treatment? A preliminary trial with a group of 12 autistic children was instituted. All had experienced autistic symptoms at approximately 18 months of age following prolonged broad-spectrum antibiotic use, and consequent persistent diarrhea. All children had been developing normally prior to onset of symptoms. During a period of 8 weeks (12 weeks for 1 child), children were given daily doses of 500 mg vancomycin, an antibiotic known to be effective against clostridial species and poorly absorbed from the intestine.

Before, during, and after the trial, children were evaluated by multiple measures of autistic symptoms related to behavior and ability to communicate. Ten children improved significantly in both measures within several days of starting the antibiotic treatment. Another child was judged to have improved by comparing videotapes taken before and after treatment. Unfortunately, after discontinuation of antibiotics, most parents reported significant behavioral deterioration.

These results are tantalizing when one recalls the recent discoveries concerning the role of Helicobacter pylori in ulcer formation and the possibility that Mycobacterium paratuberculosis may be involved in Crohn's disease. It is also a reminder of the potential dangers of indiscriminate use of antibiotics. However, much more research will be necessary before we can definitely say that neurotoxic bacteria colonizing the intestine are a causative agent for some types of autism.

Reference
Sandler RH, Finegold SM, Bolte ER, Buchanan CP, Maxwell AP, Väisänen ML, Nelson MN, Wexler HM. Short-term benefit from oral vancomycin treatment of regressive-onset autism. J. Child Neurol. 15:429–435 (2000).