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Field Reports |
* California Animal Health and Food Safety Laboratory System–Turlock Branch; and Fresno Branch, University of California, Davis, PO Box 1522, Turlock 95381
2 Corresponding author: brcharlton{at}ucdavis.edu
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SUMMARY |
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 TOP SUMMARY DESCRIPTION OF PROBLEMMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONS AND APPLICATIONSREFERENCES AND NOTES |
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Key Words: infectious bursal disease • poor performance • broiler
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DESCRIPTION OF PROBLEM |
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TOPSUMMARYDESCRIPTION OF PROBLEMMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONS AND APPLICATIONSREFERENCES AND NOTES |
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Case History
At the beginning of 2005, several submissions were received from a broiler ranch in California with a history of nonuniformity. The ranch manager suspected the involvement of RSS. The ranch consisted of 6 houses, with a total capacity of approximately 120,000 birds. Previous single-case diagnostic submissions revealed problems with bacterial septicemia caused by Escherichia coli infection. Reovirus was occasionally isolated from swollen hock joints. Intranuclear viral inclusion bodies were observed in the liver of one 23-d-old bird, and adenovirus was isolated from the same liver. On the basis of these observations, an in-depth laboratory investigation was initiated to determine the cause of the poor performance and to identify potential disease agents.
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MATERIALS AND METHODS |
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TOPSUMMARYDESCRIPTION OF PROBLEMMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONS AND APPLICATIONSREFERENCES AND NOTES |
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Sampling and Laboratory Investigations
Focus Testing.
The intensive sampling of this investigation necessitated focused sample collection and testing for economic reasons while maintaining the California Department of Food and Agriculture state surveillance program.
Samples.
Each week during the grow-out period, 10 birds per house were selected for laboratory examination. In the first 3 wk, the birds selected for testing were the smaller, weaker birds. From 4 to 6 wk, 2 separate groups of birds per house were sampled. The first group consisted of 5 birds that appeared to be healthy, and the second group consisted of 5 chickens growing poorly. Bursa and liver tissue samples were taken in 2 separate jars for histopathology.
Pathology.
All the birds were examined for gross lesions. Histological examination focused on liver and bursal tissues because of a previous diagnosis of inclusion body hepatitis and bursal abnormalities. This focus on select tissues was also an attempt to contain field investigation costs and reduce the laboratory workload. Liver and bursal tissue samples were fixed in 10% phosphate-buffered formalin, dehydrated in ethanol, cleared in xylene, embedded in paraffin, sectioned, and stained with hematoxylin and eosin by routine procedure.
Bacteriology.
Aerobic cultures of 2 livers and 2 air sacs were taken from each group once a week. Isolation of bacteria was done by culturing samples on blood and MacConkey agar plates and incubating them for 24 and 48 h at 37°C and 10% CO2. Identification of the bacteria was carried out by using standard techniques. Further identification was performed with a commercial biochemical test system [10]. Two intestinal pools representing 5 ceca per pool were cultured for Salmonella each week. Briefly, each tissue pool was inoculated into 9 mL of selenite broth and incubated for 20 to 24 h at 37°C. After incubation, the broth was streaked on Hekton and MacConkey agar plates and incubated aerobically at 37°C for 24 to 48 h. Salmonella suspect colonies were tested with appropriate biochemical tests for final identification, and then grouped and serotyped.
Virology.
Liver, cecal tonsil, pancreas, and tendon organ pools were collected and virus isolation was performed by standard protocols [11].
Bursal Reverse Transcription-Polymerase Chain Reaction and Restriction Fragment Length Polymorphism
On the basis of the severe lymphocytic depletion noted in the bursas by histopathology, bursal tissue pools were selected from the 6 houses for reverse transcription-polymerase chain reaction (RT-PCR) and restriction fragment length polymorphism. Two samples per house were chosen for testing. Each sample represented 5 bursal tissues from 5 birds. One sample was selected before the occurrence of histological bursal lesions at 5 to 16 d of age, and the other sample was selected after the occurrence of lesions at 22 to 30 d of age. The selected bursal samples were tested at the Ohio Agricultural Research and Development Center laboratory, Ohio State University, by the RT-PCR-restriction fragment length polymorphism method, as described previously [12].
Exotic Newcastle Disease and Avian Influenza Real-Time RT-PCR.
All the birds submitted for this study were monitored for exotic Newcastle disease and avian influenza (AI) in accordance with the California Department of Food and Agriculture surveillance program. Each week, 5 cloacal swabs from 5 birds and 5 pharyngeal swabs from the same 5 birds were collected. The cloacal and pharyngeal swabs were pooled separately and sent to California Animal Health and Food Safety-Davis to be tested for NDV and AI by real-time RT-PCR using standard procedures as described previously [13, 14].
Serology.
The serum samples were tested for AI, NDV, infectious bronchitis virus, CAV, and IBDV diseases by enzyme-linked immunosorbent assays [15] and for Mycoplasma gallisepticum and Mycoplasma synovia by plate agglutination tests [16]. Tests were performed according to the manufacturer’s instructions.
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RESULTS AND DISCUSSION |
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TOPSUMMARYDESCRIPTION OF PROBLEMMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONS AND APPLICATIONSREFERENCES AND NOTES |
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In this in-depth diagnostic investigation, we did not observe any nonuniformity or gastrointestinal disease. On the basis of personal observations and in concurrence with the farm manager, we noticed no variation in bird size in any of the 6 houses during weekly visits to the farm. Contrary to expectations, the average weight at the end of the run and the weight gains recorded on wk 5 and 6 gave no indication of poor performance. The condemnations reported from the processing plant were primarily attributable to E. coli septicemia. The reported condemnation rates in houses 1 through 5 were nearly the same and averaged 1.26%. In house 6, the recorded condemnation rate was much higher (6.19%), primarily because of cellulitis.
The necropsy examination of the 60 birds each week revealed gross lesions typical of E. coli septicemia occurring in the first 2 wk. Swollen hock joints and valgus deformation were also sporadically detected at the end of the study. No evidence of enteritis or intestinal parasites was found macroscopically or microscopically.
The in-depth diagnostic investigation did reveal a significant bursal disease problem. The severe lymphocytic depletion, atrophy, and stromal fibroplasia noted histologically in all the bursas tested from the 10 birds in wk 3, 4, 5, and 6 suggested that the birds were exposed to an IBDV between 15 to 22 d of age. This fact was supported by the detection by RT-PCR of an IBDV strain in samples taken at 22 to 30 d of age. All bursal tissues from the 5- to 16-d-old birds tested negative for IBDV by RT-PCR. The IBDV detected in the 6 houses all belonged to molecular group 6, and all had the same sequence as the T1 strain isolated from Georgia [17]. They were identical across the hypervariable VP2 region except for a valine at position 42. This is amino acid number 252 in the VP2 protein.
The T1 strain is designated as a hot strain, causing marked atrophy of the bursa within 3 to 4 d of infection. It also breaks through the maternal immunity to IBDV produced by conventional variant and classic vaccine strains [19]. The IBDV vaccine used on this particular ranch belonged to molecular group 4. This could explain why the birds were not fully protected from infection with the T1-like strain.
Chicken anemia virus may also explain the lymphocytic depletion [20] because the birds appearing healthy showed no histological difference when compared with those appearing poorly. Unfortunately, CAV molecular diagnostic tests or histological examinations of the thymus were not undertaken in this study.
The birds from houses 1, 2, and 3 were derived from older breeder flocks and showed a deficiency of CAV and IBDV maternal antibodies in the first week. On average, only 80% were serologically positive for IBDV, and 60% were positive for CAV. During the first week, all of the birds in houses 4, 5, and 6 had maternal antibodies against IBDV, and 80% were serologically positive for CAV titers. The IBDV titers dropped by the second week, and almost all birds were negative for IBDV by the third week. Infectious bursal disease virus titers then increased steadily until the end of the grow-out period. Most of the examined birds had maternal antibodies against CAV on the first week, which turned seronegative between wk 2 and 4, and then seroconverted by the last 2 wk of the study. The same pattern was observed for the reovirus antibody titers. Infectious bronchitis virus and NDV titers were as expected for the vaccination program. The flocks remained serologically negative for M. gallisepticum, M. synovia, and AI.
The significance of the reovirus isolated from pancreas, cecal tonsil, and tendon pools is thought to be minimal. These isolations occurred repeatedly throughout the entire grow-out period from both healthy and possibly underweight birds. Avian reoviruses have a wide spectrum of pathogenic capability, especially in chickens. The diagnosis of a disease process is dependent on the demonstration of lesions and the detection of the virus in clinical samples [21]. Because the histological focus of this study was on bursal and liver tissues, the significance of reovirus from the tendon is unknown. It has been reported that reovirus occurs widely in both normal and diseased chickens [22].
Adenovirus was isolated from liver, pancreas, cecal tonsil, and tendon pool samples from birds collected on wk 3, 4, 5, and 6. No indication of intranuclear inclusion bodies or other histological lesions was found in the livers of the tested birds, and no attempt was made to further characterize the isolated adenoviruses. Therefore, the significance of the isolated adenoviruses is also unknown but is thought to be minimal. Adenoviruses (FAV1 to FAV12) are widely distributed in poultry flocks [23] and the exact role that these viruses play as etiological agents of specific diseases in chickens remains obscure. Some strains of FAV4 can cause infectious hydropericardium in chickens [24]. The sporadic occurrence of classic inclusion body hepatitis in combination with the isolation of different serotypes of adenovirus has also been reported [25–27].
Severe cryptosporidiosis was recorded from the bursas in the last 3 wk of the grow-out period. The importance of finding Cryptosporidium in the bursas during the last 3 wk of the study is not clear. Several reports suggest that the performance of broilers can be adversely affected by cryptosporidial infections [28]. This infection may also be a consequence of immunosuppression caused by IBDV infection, a possible CAV infection, or both.
The high number of condemnations caused by cellulitis reported from house 6 can most likely be attributed to the higher ratio of males to females in this specific house, with relatively poor feathering of males leading to increased susceptibility to skin scratches. Although no general underweight problem was observed during this study, the bursal damage was significant and may have led to immunosuppression, making the birds more susceptible to bacterial infections, such as E. coli and Salmonella. It also may have contributed to the increased level of condemnations seen at the processing plant. The overall flock performance was acceptable and met expectations, but it is conceivable that the performance could have been even better if the birds had been protected against IBDV field challenge. It should also be considered that if the birds had been challenged with a respiratory infection, in addition to suffering immunosuppression caused by IBDV infection, the flock may not have reached the level of performance that was actually achieved. No major management changes were undertaken at the company site during this investigation. At the end of study it was questionable whether the farm manager was affected by the flocks being under observation and having weekly visits, leading to management improvement.
Poor performance can be a poorly defined condition that requires cooperation from the producer, a responsible veterinarian, and a diagnostic laboratory. The multiple systematic sampling for diagnostic investigation used in this study provided valuable information that could not have been obtained from single or sporadic submissions. The single identification of an IBDV T1-like strain would have few implications. The occurrence of the same IBDV strain in multiple birds from multiple houses at a similar age increases the significance of the virus as an important factor to the producer’s complaint. Further investigation and manipulation of the IBDV vaccination program may assist the producer in obtaining more uniform production.
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CONCLUSIONS AND APPLICATIONS |
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TOPSUMMARYDESCRIPTION OF PROBLEMMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONS AND APPLICATIONSREFERENCES AND NOTES |
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FOOTNOTES |
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REFERENCES AND NOTES |
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TOPSUMMARYDESCRIPTION OF PROBLEMMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONS AND APPLICATIONSREFERENCES AND NOTES |
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