G. R. Huff, W .E. Huff, and N. C. Rath
USDA Agricultural Research Service
Poultry Production and Product Safety Research Unit
Center of Excellence for Poultry Science
University of Arkansas
Fayetteville, Arkansas
U.S.A.
Introduction
Turkey osteomyelitis complex (TOC) is the name given to a syndrome defined by the U. S. Food Safety Inspection Service (FSIS) to describe normal-appearing processed turkey carcasses which contain lesions including arthritis/synovitis, soft-tissue abscesses, and osteomyelitis of the proximal tibia. The presence of a green liver is used as an indicator to remove suspect turkeys from the processing line. These turkeys are then subjected to a federally mandated standard 10-cut procedure to detect the suspected bone, joint, and soft-tissue lesions (Cook, 1988). While it has been assumed that most turkeys with TOC lesions also have green livers, lesions have been detected in birds without green livers both experimentally and in commercially grown birds (Clark et al., 1991; Bayyari et al., 1994). Also, at least half of the birds that are examined due to the presence of a green liver have no other lesions, and are thus downgraded unnecessarily (Barnes et al., 1990; Clark et al. 1991; Mutalib et al., 1996). Because of the deficiencies of the present inspection system, we have endeavored to determine the etiology of this condition in order to enable its prevention and control. Previous research has indicated that the lesions are caused by various opportunistic organisms, mainly Staphylococcus aureus and Escherichia coli (Clark et al, 1991; Bayyari et al., 1994; Droual et al.,1996), suggesting that TOC incidence may be influenced more by deficiencies in the host immune response rather than by the virulence of any one organism. This syndrome is primarily a disease of adolescent male turkeys indicating that sex hormone effects on immunity may be involved.
We have reported that birds with TOC lesions have higher levels of cell-wall deficient bacteria in their livers and decreased immunity compared to normal turkeys. We believe that differences in the response to stress of individuals within flocks of male turkeys, possibly owing to divergent selection for fast growth, may be responsible for a susceptibility to infection with common bacteria normally found in their environment. We have established an experimental model of TOC which demonstrates that treatment with 2 mg/kg body weight of a synthetic glucocorticoid, dexamethasone (DEX), can decrease resistance to bacterial infection in turkeys. This model suggests that the stress response of the turkey may be involved in the immunosuppression which results in TOC lesions.
Review of USDA-ARS TOC research
Our basic question has been: Why do a small proportion (~0.2%) of otherwise healthy-appearing turkeys develop these opportunistic infections? Understanding of how and why TOC lesions develop may lead to methods to prevent the lesions. The following studies were designed to answer this question and to investigate the possibilities of using nutritional supplementation to decrease TOC incidence.
Field Study
In our first study (Bayyari et al., 1994) we examined 100 turkeys each week throughout a 15 week grow out in order to determine the incidence and cause of TOC. We randomly sampled 50 turkeys from each of 2 farms that previously had a relatively high incidence of TOC. We found that one farm had a much higher incidence of TOC and that farm also had more early and weekly mortality, air sacculitis, and seroconversion to Mycoplasma meleagridis and Newcastle disease virus. This study suggested that TOC may be related to chronic respiratory infection.
We found that while S. aureus was isolated from some of the lesions, we were just as likely to isolate other opportunistic bacteria, such as E coli and Enterococcus species. We attempted to use more intensive methods of culturing such as decreasing the amount of oxygen, using longer incubation, and utilizing hypertonic and serum-supplemented media. In previous studies bacteria were isolated from an average of 40% of the liver, bone, and joint lesions cultured. Using extended incubation and hypertonic medium, we were able to isolate a slow-growing, pleomorphic microbe from many of the lesions which had appeared sterile using traditional methods. After trying to identify this organism as an actinomyces sp., we came to the realization that what we were isolating were the L-forms or cell-wall deficient forms of the common, opportunistic bacterial species we were also finding in the lesions. L-form bacteria are known to grow inside both red and white blood cells, and it is thought they may be more prevalent in animals with a low immune response. L-form isolation was about 3X higher from green livers than from normal livers, so we considered that maybe there may be a deficiency in the ability of the green liver turkeys to kill phagocytosed bacteria.
Immune Studies and Genetic Differences
We studied the immune systems of birds with and without TOC and saw no significant differences in phagocytosis or bacterial killing by phagocytes. There was a significant decrease in the function of the T-cell mediated immune response of the TOC birds, and an increase in complement levels, and heterophil/lymphocyte (H/L) ratio, which is an indicator of stress (Bayyari et al.,1997a).
We also studied differences in the immune response of different genetic strains of turkeys. We found that birds selected for faster growth had a decrease in some responses and an increase in others, as compared to their parent stock. We also found differences between the two most widely used commercial strains of turkeys, in particular, a decreased toe-web response in the commercial line known to have a higher incidence of TOC (Bayyari et al., 1997b, 1997c).
Stress: an experimental model of Turkey Osteomyelitis Complex
We have developed an experimental model for reproducing TOC lesions. We can increase the incidence of TOC to about 30% by treating 5-week- old birds with 3 DEX injections and inoculating low numbers of E. coli into the airsac (Huff et al, 1998). Our original hypothesis was that the stress of moving birds at 5 weeks of age to grow-out houses may increase the incidence of colisepticemia, resulting in latent infections in birds unable to completely kill these bacteria. Later stresses may then allow reactivation of these unapparent infections resulting in TOC lesions. Recent studies have shown that this may not be entirely the case. A second series of DEX injections produces a much higher incidence of TOC lesions (70-80%) whether or not E. coli is inoculated, and S. aureus is isolated from most of these lesions both with and without E. coli isolation (Huff et al.,1999b). This indicates that there may be a cumulative effect of various stressors throughout turkey production which results in decreased resistance to bacterial infection.
The immunosuppressive effects of stress may be greater in male birds than in females. We have found that female turkeys are more resistant to the DEX-E.coli challenge than males (Huff et al., 1999a). Turkey osteomyelitis complex is a disease that primarily affects male turkeys. We believe that the combined effects of disease, management, and environmental stressors may decrease the immune response in genetically susceptible male turkeys.
In summary, we have established an experimental model for the reproduction of all of the lesions describing the TOC syndrome, including green liver, arthritis/synovitis extending into the tendons and producing purulent and caseous lesions in the muscles of the leg and thigh, and osteomyelitis of the proximal tibia. These lesions are identical to those seen in commercial processed turkey carcasses, and in older birds they occur without the external signs of septicemia/toxemia. In this model they are associated with respiratory infection, whether or not the birds are inoculated with bacteria. While the E.coli inoculum can be cultured from the majority of lesions, S. aureus is also isolated from many birds, and is the predominant isolate in the blood, air sac and TOC lesions of birds given only DEX treatment. Stress affecting the immune system appears to be the cause of TOC.
Nutritional Immunomodulation and TOC
A number of vitamins have been indicated for improving the stress response and immunity. We have studied the effects of vitamin C and vitamin E in our TOC model, however, in our experiments there was no benefit of vitamin C or E supplementation on TOC incidence. Dietary supplementation with yeast products containing beta-glucans also had no effect on TOC incidence.
Vitamin D3 Supplementation
We did have positive effects when we evaluated the ability of supplemental vitamin D3 to protect male turkeys from the immunosuppressive effect of DEX treatment (Huff et al.,2000). We gave poults supplemental vitamin D3 (High D, I.D. Russell Company, 2064 IU/l ad libitum in drinking water) during the first 5 days of brooding and again at a higher level (4128 IU/l) before, during, and after each stressful event, which we defined as weekly weighing and DEX challenges. Vitamin D3 supplementation had no effect on disease resistance, hematology values, or body and organ weights of birds which were treated with DEX at 5 weeks of age and necropsied 2 weeks later. However, vitamin D3 supplementation significantly decreased mortality, TOC incidence, green liver incidence, and the recovery of bacteria from the liver and airsac of 14-week-old male turkeys which had been treated with DEX at 5 weeks and again at 12 weeks of age (Figure 5).
Airsacculitis scores were 4-fold higher in DEX-treated birds not given supplemental vitamin D3 (P = 0.0002) (Figure 1). DEX injection increased the percentage of heterophils and decreased the percentage of lymphocytes in the peripheral blood of birds not given supplemental vitamin D3, however 2X DEX-treated birds given supplemental vitamin D3 were protected from these changes (Figure 2). Vitamin D3 supplementation also significantly protected against DEX-induced loss of body weight in 2X DEX-treated turkeys (P = 0.0005) (Figure 3).
Differences in the genotype of the vitamin D receptor, which apparently regulates bone density, have been shown to affect susceptibility to a number of infectious diseases in humans (Hill, 1998), and this may be related to the etiology of TOC. The ability of vitamin D3 supplementation to affect changes in the physiology and disease resistance of stressed birds emphasizes the current conceptual view that vitamin D3 is not only important to bone health, but is a prohormone that affects all vital systems (DeLuca and Zierold, 1998) and that supplementation during stressful periods may help to modulate the immune response and increase resistance to infection with opportunistic bacteria.
A Link between genetics and stress-induced immunosupression?
There has been a lot of research which has shown that selecting birds for fast growth can make them more susceptible to disease. In turkeys, the association between fast growth and decreased disease resistance has come primarily through the study of four closed genetic turkey lines developed at the Ohio Agricultural Research and Development Center (OARDC) at The Ohio State University. These lines include a randombred control line (RBC1) and its subline (Egg line) selected exclusively for increased egg production over a 250-d period and another randombred control (RBC2) and its subline selected for increased 16-wk BW (F Line). Recently we compared the Egg line, F line, and a commercial turkey line (Comm) for their responses to Escherichia coli challenge following dexamethasone injection (Dex) or E. coli challenge preceding transport stress (Transport). At 14 wk of age, the Dex group was treated with 3 injections of 2mg Dex/kg BW followed by airsac challenge with 100 cfu of E. coli.
The Transport group was given the same E. coli challenge at 1x104 cfu/bird without Dex treatment, and was subjected to transport stress, including 12 h of holding time in a transport vehicle, 8 d after the challenge.
Surviving birds were all necropsied two weeks post infection. Mortalities and necropsied birds were scored for TOC using the FSIS 10 cut procedure and all knee joints and any joints with visible lesions were cultured for E.coli.
Mortality was significantly higher in the Comm line as compared to the Egg line and was intermediate in the F line (Figure 4). There was no TOC in Egg line birds. Percentage of TOC and knee inflammation (arthritis) was numerically higher in the large bodied lines as compared to the Egg line (Figures 5 and 6). The challenge strain of E.coli was isolated from significantly more F line and Comm line knee cultures as compared to the Egg line.
This study suggests that intense selection for performance may compromise immunity by altering the response to stress and supports a study from Poland (Kowalski et al. 2002) which compared two commercial turkey lines and found that the lighter and slower-growing BUT-9 line was more resistant to stress than the faster-growing Big-6 line and suggested that the lighter line might be more suitable for commercial poultry production because of its better response to stress. It also implies that faster growing birds may harbor opportunistic bacteria within their tissues to a greater extent than slower-growing birds, suggesting an opportunity for increasing food safety by improvement of the stress response of commercial turkeys.
Summary
These studies are providing evidence of the link between TOC and modulation of the stress response. It appears there may be a cumulative effect of the various stressors involved in turkey production, which can have a profound effect on the immune system of some male birds, leading to decreased disease resistance. In addition, these studies dramatically illustrate the effects that stress can have on production as well as condemnation problems such as air sacculitis and cellulitis, which are often seen in our model. We hope this research will encourage producers to reassess production practices with an eye on reducing stress whenever possible. Poultry research has been focused on improving production values through the use of nutrition and genetics, and the control of infectious disease through vaccination.
Decreasing infections by opportunistic pathogens, such as air sacculitis and TOC, will require an equal effort toward the design of creative management practices that reduce stress, and an enforcement of the good management practices already in place. Further study may lead to a means to genetically select birds with an improved ability to tolerate the stresses inherent in commercial poultry production, in nutritional, behavioral, and environmental means of modulating the stress response.
What can we do?
We believe that TOC is related to stress. For your birds, stress can be anything or everything from the discomfort caused by high or low environmental temperatures, wet litter, or running out of feed to the fear elicited by moving to a grow out house or being unable to avoid the aggression of bigger birds. And while some individuals may thrive in the competition inherent in living in a very large population, others will be at a disadvantage due to size or temperament.
What can be done about it? If the effects of stress are so complex, why even bother trying? Probably the easiest and most important thing one can do is to just think. Think about how the birds can be stressed, and then do all that you can to fix the situation. A lot of things just can't be changed, but many can be improved by a little consideration. We have listed some of the stressors that can be faced by poults throughout a grow out below. Our challenge is to try to think of more ways in which one can improve the welfare and performance of the flocks. What is it in a particular situation that can produce unexpected changes in the environment, unexpected surprises for the birds? What are they afraid of? A turkey grower in Kansas, after reading this list, realized that running the housekeeping tiller in his brooding house was really scaring his poults. He believes that since he stopped tilling, his birds have had less osteomyelitis. Coincidence? Maybe. But we feel that if you add together all of the little things you can do to make birds more comfortable and less fearful, you will not only make a big difference in your birds' welfare, but also in their health.
For turkey growers, one of the greatest stressors on the poults is moving birds from one house to another in a multi-stage system. We need to find alternatives to these types of management practices that can really impact health and disease. We are searching for ways to improve the birds' stress response through genetics and nutrition. But until real solutions are found, decreasing stress is up to the farmer. Think about ways to decrease the fear and reduce the changes faced by the birds.
Try vitamin D? Grow a slower bird? These both may help. But perhaps the best thing the farmer can do to reduce stress and improve health and welfare in his flocks is to try to maintain a sense of consistency as best as possible. Make careful observations and be aware of ways the farmer can reduce fear and uncertainty. And continue to think about how sometimes big changes and sometimes small changes in our lives and our environment can profoundly affect the way we feel.
- Hatching, Catching, Rough handling
- Beak and toe trimming, Vaccination
- Transportation, Open field stress
- Heat stress, Cold stress
- Ammonia, Dust, Endotoxin, Tilling machines
- Disease
- Fear of humans, Social hierarchy, Overcrowding
- Genetic selection for fast growth, Lighting schedules
- Running out of feed, poor nutrition or feed quality
- Running out of water, poor water quality
- Wet litter
- Moving from brooder houses to grow-out houses
- Catching and transportation to processing plant
References are available on request
From Proceedings of the "Midwest Poultry Federation Convention", St. Paul, Minnesota, U.S.A.
