Heterophil function and resistance to staphylococcal challenge in broiler chickens naturally infected with avian leukosis virus subgroup J

Avian leukosis virus subgroup J has a high tropism for myeloid lineage cells and frequently induces neoplastic transformation of myelocytes. The impact of congenital avian leukosis virus subgroup J infection on the function of circulating heterophils and susceptibility to staphylococcal infection was investigated. Six-week-old broiler chickens negative for exogenous avian leukosis viruses or congenitally infected with avian leukosis virus subgroup J were inoculated intravenously with 10(6) colony-forming units of Staphylococcus aureus, and pre- and postinoculation heterophil function was assessed. All chickens developed a leukocytosis with heterophilia after inoculation, but total leukocyte and heterophil counts were significantly higher in leukosis-negative chickens than in virus-infected chickens. Tenosynovitis was more severe in leukosis-negative chickens, and 2/10 (20%) of the virus-infected chickens had no histologic evidence of tenosynovitis. Osteomyelitis in the tibiotarsus or tarsometatarsus developed in 5/10 (50%) of the chickens in each group. S. aureus was recovered from the hock joint of 6/10 (60%) of the chickens in each group. Heterophils from all chickens exhibited similar phagocytic ability pre- and postinoculation. Heterophils from virus-infected chickens exhibited less bactericidal ability preinoculation than did heterophils from leukosis-negative chickens. However, postinoculation bactericidal ability was similar in both groups. Avian leukosis virus subgroup J provirus was present in heterophils isolated from congenitally infected chickens. Heterophils isolated from broiler chickens congenitally infected with avian leukosis virus subgroup J exhibit no significant functional deficits, and infected and uninfected chickens exhibit similar susceptibility to staphylococcal infection.     

Reduced erythrocyte deformability as a possible contributing factor to pulmonary hypertension and ascites in broiler chickens.

Deformability of erythrocytes and hematological parameters in white leghorn and broiler chickens were measured at age 7, 14, 21, 28, and 35 days. For deformability testing, a simple vertical apparatus containing a polycarbonate membrane with 5-microns pores was used. This technique assesses erythrocyte deformability by measuring the filtration time of an erythrocyte suspension through the pores. There was a significant difference in filtration time between the leghorns and broilers at all sampling times. These results indicate that reduced erythrocyte deformability in broilers may be one of the predisposing factors that increase resistance to blood flow and alter the rheology of blood in the microcirculation in the lung. Increased resistance to flow may result in pulmonary hypertension, heart failure, and ascites in broiler chickens.     

Pathogenesis of ascites in broilers raised at low altitude: aetiological considerations based on echocardiographic findings

This study reports novel insight into the aetiology of pulmonary hypertension and ascites in broiler chickens. The scope of measurements was focused on anatomical and functional parameters, and blood flow patterns in leghorns (resistant to ascites), fast-growing broilers (susceptible to ascites), broilers developing ascites, and ascitic broilers evaluated in vivo using echocardiography, and further examined in the context of postmortem findings. Both, in vivo observed features and postmortem findings, showed clear differences between broilers and leghorns, and between normal and ascitic broilers. Abnormalities in the heart chamber geometry and blood flow patterns were detected upon echocardiographic examination in all ascitic broilers. Right and left atrio-ventricular (AV) valve regurgitation were common findings in ascitic broilers and some apparently normal broilers, with left AV valve insufficiency being a predominant feature with respect to degree and frequency of occurrence. Blood flow disturbances were not detected in leghorns. Left ventricular fractional shortening (functional parameter) was considerably reduced (P < 0.01) in ascitic birds (mean: 21.7 +/- 2.0 SE) in comparison with normal broilers (mean: 39.1 +/- 3.6 SE), or leghorns (mean: 43.3 +/- 2.4 SE). The presented findings indicate that pathological and functional changes in the left ventricle and atrium play a significant role in the pathogenesis of ascites in broilers. Severe dilation of the left atrium and pulmonary veins seen on postmortem examination, as well as regurgitant blood flow in the left atrium, demonstrated by Doppler study in ascitic birds, provide evidence that chronically elevated pressure in the left atrium is involved in the aetiology of pulmonary hypertension and ascites in fast-growing broilers.     

Evaluation of scratches as an essential element in the development of avian cellulitis in broiler chickens.

Currently, the published cellulitis models do not adequately address the actual pathogenesis as seen in the commercial broiler industry. In this model, small dermal scratches were made on the skin of broiler chickens, which were then placed on litter seeded with avian cellulitis-associated Escherichia coli. The research confirms scratches are required for the induction of avian cellulitis. The research also confirms that "type I" cellulitis lesions or those previously thought to be due to hatchery-borne infections can be induced with scratches. The described methods provide a realistic model for cellulitis development that will improve the reliability of prophylactic and therapeutic-regimen efficacy testing data, thereby providing information more directly useful to the commercial broiler industry.     

Subclinical infectious bursal disease in an integrated broiler production operation.

A field study was designed to determine the prevalence of subclinical infectious bursal disease (IBD) in broiler chickens from a commercial poultry company. Bursae of Fabricius (BF) from two vaccinated and three nonvaccinated broiler flocks were evaluated histologically, and antibody profiles of these broiler and matched parent breeder flocks were established. Lesions of IBD, including lymphoid necrosis, stromal edema, and infiltrates of heterophils and macrophages, were first detected in BF at 24 days of age in both vaccinated and nonvaccinated chickens. At 41 days, all BF had lesions characteristic of IBD, including severe lymphoid depletion, proliferation of epithelial cells, and mild fibroplasia. Although mean maternal antibody levels (measured by enzyme-linked immunosorbent assay) in broilers were apparently protective through day 12, IBD antibodies decreased to nonprotective levels (below 1,000) by day 16 or 20. Titers began to increase by day 28 or 32 because of field exposure. Sentinel birds, placed with broiler flocks, also developed IBD antibody titers. Broiler breeders had low and nonuniform antibody titers. Prevalence of field IBD exposure was high, and existing vaccination programs were not effective.     

Efficacy of coarse-spray administration of a reovirus vaccine in young chickens.

Coarse-spray (CS) administration of a commercial S1133 reovirus vaccine in chickens for prevention of clinical viral tenosynovitis (VT) infection was evaluated. In Expt. 1, one-day-old specific-pathogen-free (SPF) white leghorns were vaccinated with a combination of reovirus, Newcastle disease (ND), and infectious bronchitis (IB) vaccines by CS and infectious bursal disease vaccine by the subcutaneous (SQ) route. In Expt. 2, one-day-old commercial broilers were vaccinated by CS with reovirus vaccine and Marek's disease (MD) vaccine by SQ. In Expt. 3, one-day-old commercial broilers received reovirus vaccine in combination with ND-IB vaccines at 1 day of age by CS and MD vaccine by SQ. Some birds received an initial or second vaccination at 7 days of age by CS or the drinking-water (DW) route. Birds vaccinated by CS at 1 day of age with reovirus vaccine did not produce circulating virus-neutralizing antibody against reovirus, although they had resistance to VT infection. In contrast, initial or booster vaccination at 7 days of age by CS or DW resulted in an antibody response and greater resistance to challenge than did CS vaccination at 1 day of age. There was no difference in efficacy between CS and DW routes at 7 days of age. The reovirus vaccine did not interfere with other vaccines as measured by serologic (ND-IB-IBD) or challenge (MD) studies.     

Experimental induction of the cutaneous form of Marek's disease in broilers

Straight-run chickens of the Ross broiler hybrid were experimentally infected with the skin homogenates of sound broilers, broilers conditionally edible, and broilers confiscated for alternations in skin. The chickens infected with the homogenates of healthy skin were negative in the Marek's disease test. In 17% of the chickens infected with the skin from the conditionally edible broilers, macroscopically observable cutaneous lesions were induced and 60% of the infected chickens had microscopic changes in the skin; visceral organs were always affected by the infection. In the chickens infected with the skin of a confiscated bird, the long storage exerted its unfavourable effect and the infected birds were negative in the Marek's disease test. Our results indicate that the active form of Marek's disease was induced.     

Reduced serologic response to Newcastle disease virus in broiler chickens exposed to a Chinese field strain of subgroup J avian leukosis virus

In this study, a Chinese field strain of subgroup J avian leukosis virus (ALV-J), NX0101, was studied for its immunosuppressive effects in both commercial broilers and SPF white Leghorn chickens infected at 1 day of age. Our data demonstrated that NX0101 induced much more significant body and immune organ weight loss in the infected commercial broiler chickens in an earlier age than that in the SPF white Leghorn chickens. At the same time antibody responses to vaccinations of Newcastle disease virus (NDV) and infectious bursa disease virus (IBDV) in the NX0101-infected chickens were also evaluated and compared between the commercial broiler chickens and the SPF white Leghorn chickens. Compared with the control group of chickens, the hemagglutination inhibition (HI) antibody response to NDV vaccines was significantly reduced in the NX0101-infected commercial broiler chickens from as early as 20 days after vaccination. However, no significant difference in HI antibody response was seen when HI titers reached their peaks in the NX0101-inoculated and control SPF white Leghorn chickens, except it declined significantly faster in infected birds. Neither of these two types of chickens showed significant decrease of antibody response to IBDV vaccination. Herein, we conclude that this NX0101 strain of ALV-J could selectively suppress humoral immune reactions to NDV, especially in broilers. But challenge experiments were not conducted and, therefore, it cannot be known if decreased antibody levels correlated with decreased protection against NDV in this case.     

Effect of a commercial competitive exclusion culture on reduction of colonization of an antibiotic-resistant pathogenic Escherichia coli in day-old broiler chickens

One-day-of-age broiler chickens were administered a commercial competitive exclusion (CE) product and then challenged by three different methods with an Escherichia coli O78:K80 that was pathogenic for poultry and resistant to six antibiotics. Three challenge methods were used on 2-day-old broilers: direct challenge, precolonized seeder, and instant seeder. Direct challenge was accomplished by administering the challenge E. coli per os. The precolonized seeder challenge had two chicks that had received the challenge E. coli 24 hr previously, whereas the instant seeder challenge had two chicks given the challenge E. coli per os with immediate placement with the experimental birds. One oral dose of the commercial CE product significantly reduced the colonization of the small intestine, large intestine, and ceca by the highly antimicrobial resistant poultry pathogenic E. coli O78:K80 at 7 and 14 days postchallenge by all three challenge methods. The overall mean reductions in colonization were 3.0 log10 for the large intestine, 3.0 log10 for the small intestine, and 4.0 log10 for the cecum. The most severe challenge method, on the basis of the least amount of reduction of colonization of the challenge E. coli by the CE, was by the direct oral gavage at 2 days of age.     

Enteropathogenicity of Dutch and German avian reoviruses in SPF white leghorn chickens and broilers

The enteropathogenicity of avian reoviruses (ARVs), isolated from chickens affected with malabsorption syndrome (MAS) from The Netherlands and Germany was studied. In the first trial seven different ARVs isolated from MAS cases were inoculated in 1-day-old specific pathogenic free (SPF) white leghorns. The pathogenicity was compared with 2 ARVs isolated from cases of tenosynovitis, namely reference strain S1133 and a Dutch strain. Although a difference in the severity of the clinical disease was observed, all reoviruses could induce vacuolar degeneration and sloughing of the epithelium of the small intestine at day 2 post inoculation (PI) till day 7 PI. Two Dutch and one German ARV derived from MAS causing the most severe intestinal lesions at day 2 PI, were further studied in the second trial using SPF broilers. These reoviruses did not cause weight gain depression in the broilers although lesions in the small intestine were present from day 1 up to day 4 PI and were more severe than in the white leghorn chickens. In one of the inoculated groups apical denuded villi were already present at day 1 PI. At day 7 PI the small intestine of the infected broilers appeared to be normal. Reovirus antigen was detected in the cytoplasm of the enterocytes at the tip and middle section of the affected villi both in layers and in broilers. To study the role of intestinal CD4+ and CD8+ T-cells and macrophages/monocytes in the pathogenesis of ARV, the numbers of these cells of the jejunal villi of one infected and the control broiler groups were compared. CD4+ T-cells were detected in low numbers and only in the infected broiler group at day 14 PI. The numbers of CD8+ T-cells and macrophages/monocytes were significantly higher in the infected broiler group than in the control broiler group at day 7 and 14 PI and at day 7 PI respectively. Our study indicates that the reovirus alone cannot induce intestinal lesions as found in MAS chickens. Moreover, CD8+ T-cells may play a major role in the pathogenesis and or reovirus clearance in the small intestine.     

Effect of drinking water chlorination on Campylobacter spp. colonization of broilers

The main source for Campylobacter spp. transmission from the environment to broiler chickens is still unclear. One implicated reservoir for the organism has been untreated broiler drinking water. This study was conducted with broilers first using experimental conditions (isolation units) and second under commercial conditions. We compared the rate of intestinal colonization in chickens provided 2 to 5 parts per million (ppm) chlorinated drinking water in relation to the frequency of colonization in chickens given unsupplemented drinking water. No significant difference (P > 0.05) was detected in isolation frequency or level of Campylobacter spp. colonization in birds provided chlorinated drinking water and control birds provided water without supplemental chlorine. In the isolation unit experiments, 86.3% (69/80) of the control and 85.0% (68/80) of the treated birds were colonized at levels corresponding to an average of 10(5.2) and 10(5.1) log colony-forming units (cfu) Campylobacter spp./g of cecal contents, respectively. Additionally, two sets of paired 20,000 bird broiler houses, with and without chlorination (2-5 ppm chlorine), were monitored in a commercial field trial. Effectiveness of chlorination was judged by prevalence of Campylobacter spp. in fecal droppings (960 samples) taken from the flocks in treated and control houses. Birds from the control houses were 35.5% (175/493) Campylobacter spp. positive, while 45.8% (214/467) of the samples from the houses having chlorinated drinking water yielded the organism. Chlorination of flock drinking water at the levels tested in this study was not effective in decreasing colonization by Campylobacter spp. under commercial production practices presently used in the United States.     

Ornithobacterium rhinotracheale, a primary pathogen in broilers

Field strains of Ornithobacterium rhinotracheale were tested on their virulence in different chicken breeds. Ornithobacterium rhinotracheale was able to induce lesions after aerosol challenge without a previous priming with virus, and thus O. rhinotracheale was proven to be a primary pathogen. The virulence of Dutch strains, isolated between 1995 and 1998, did not increase, but the Dutch isolates and a South African strain were more pathogenic compared with an American strain of O. rhinotracheale. White specific-pathogen-free leghorns were less susceptible to O. rhinotracheale infection than broilers, whereas there was no difference in susceptibility between commercial broilers and specific-pathogen-free broilers.     

Isolation of Campylobacter from circulating blood of commercial broilers

Campylobacter spp. are present in organs and tissues of broiler chickens but the dissemination route is unclear. The aim of the current study was to determine Campylobacter prevalence within circulating blood of commercial broilers. Broilers were acquired from 19 flocks originating from three commercial poultry processing companies. Using aseptic blood collection techniques, 5 ml of circulating blood was collected from each bird and the sample analyzed for Campylobacter. The Campylobacter colonization status of each bird was determined by aseptically sampling and analyzing the ceca. Campylobacter was recovered from 58% (11/19) of flocks sampled. From the 248 total birds sampled, 12% and 46% of the birds had Campylobacter in the blood and ceca, respectively. This study documents Campylobacter prevalence in the circulating blood of commercially raised broilers. Campylobacter presence in the circulatory system may indicate the path used by the organism for rapid dissemination to organs and tissues. From a processing viewpoint, Campylobacter presence in circulating blood of market-age broilers may increase the likelihood of cross-contamination between birds during slaughter.     

Acute airsacculitis in untreated and cyclophosphamide-pretreated broiler chickens inoculated with Escherichia coli or Escherichia coli cell-free culture filtrate.

Ninety commercial broiler chickens were divided into three equal groups; 30 were injected with brain-heart-infusion broth into the cranial thoracic air sacs (controls), 30 were similarly inoculated with a culture of Escherichia coli, and 30 were similarly inoculated with E. coli cell-free culture filtrate. Birds were examined from 0 to 6 hours post-inoculation. E. coli-inoculated and cell-free culture filtrate-inoculated chickens reacted similarly, with exudation of heterophils into the air sac. Microscopically, heterophils were present in low numbers perivascularly 0.5 hour after inoculation and became more numerous by 3 hours post-inoculation. By 6 hours post-inoculation, there was severe swelling of air sac epithelial cells and thickening of the air sac by proteinaceous fluid and heterophils. Ultrastructurally, air sac epithelial cells were swollen and vacuolated, and interdigitating processes were separated. Histologically and ultrastructurally, all features in control chickens were normal, with only rare heterophils in the air sac interstitium. In E. coli-inoculated and cell-free culture filtrate-inoculated chickens, cell counts (predominantly heterophils) in air sac lavage fluids increased markedly at 3 and 6 hours, with only slight increases in counts from lavages of controls. Heteropenia was observed in E. coli-inoculated chickens, whereas heterophilia was observed in cell-free filtrate chickens and controls. Ninety additional chickens were pretreated with cyclophosphamide, subdivided into three equal groups, and inoculated and examined similarly as above. Cyclophosphamide pretreatment reduced inflammatory changes in air sacs, lowered cell numbers in lavage fluids, and abolished hematologic changes; however, it did not prevent epithelial cell changes. These results indicate that cell-free culture filtrate of E. coli induces changes similar to those induced by cultures of E. coli.     

A study on pathogenesis of sudden death syndrome in broiler chickens

Sudden death syndrome (SDS) in fast growing broiler chickens has been recognized as a patho-physiological entity for four decades, but its pathogenesis still remains unknown. More recent investigations provided evidence that link SDS to cardiac arrhythmia, but the mechanism triggering arrhythmogenesis and factors responsible for fatal outcome are poorly understood. In order to understand the chain of events leading to SDS in broilers, the present study focused on putative mechanisms that trigger arrhythmia and mechanisms that predispose the myocardium to fatal arrhythmia. Susceptibility of broilers to cardiac arrhythmia under stress conditions was evaluated using a simulated stress test with epinephrine. Detailed histopathological evaluation of the broiler heart was undertaken to identify structural features that may predispose the myocardium to fatal arrhythmia. The simulated stress challenge revealed that many broilers are highly susceptible to stress induced cardiac arrhythmia. In some broilers the stress challenge induced severe ventricular arrhythmia, and the life threatening nature of this arrhythmia was evidenced by the fact that several birds showing the most severe arrhythmic responses, died suddenly within several days after the stress challenge. Examination of hearts of broilers that died of SDS revealed microscopic lesions in the cardiomyocytes, and widespread changes in the sub-endocardial and mural His-Purkinje system (HPS). Immune staining for Caspase-3 confirmed that numerous Purkinje cells in the left ventricular myocardium from broiler chickens that died of SDS were undergoing apoptosis. The observed lesions suggest that the electrical stability of the myocardium was compromised. Taken together, our findings indicate that stress is a most likely trigger of cardiac arrhythmia in broilers, whereas the pathological changes seen in the myocardium and in the HPS in fast growing broilers provide a very conducive milieu for sustained ventricular arrhythmia. In cases where the electrical stability of the myocardium is compromised, even an episodic arrhythmic event may readily degenerate to catastrophic ventricular fibrillation and sudden death. We conclude that the combination of stress and changes in the cardiomyocytes and HPS are the key requisite features in the pathogenesis of SDS.     

肉鸡肿头综合征的病理形态学变化

对黑龙江省某鸡场发生肿头综合征（ Ｓ Ｈ Ｓ） 的４ 例肉鸡进行了病理学观察。肉眼所见为病鸡的眼周围、肉髯、肉冠的肿胀,包括肉髯、肉冠在内的头部皮下的蜂织炎,头盖骨气室内的干酪样物贮留。病理组织学所见主要为头部皮下组织、头盖骨气室及鼻粘膜下的纤维素性、化脓性肉芽肿性炎,并见有间质性心肌炎及支气管粘膜上皮组织纤毛脱落等变化。从上述病例的头部病灶中分离到了大肠杆菌。     

In vivo priming heterophil innate immune functions and increasing resistance to Salmonella enteritidis infection in neonatal chickens by immune stimulatory CpG oligodeoxynucleotides

Oligodeoxynucleotides (ODN) containing CpG dinucleotides (CpG-ODN) mimic bacterial DNA and stimulate the innate immune system of vertebrates. Here, we investigated the effects of intraperitoneal (ip) administered CpG-ODN on the innate immune functions of chicken heterophils. Our results demonstrated CpG-ODN-dependent priming of chicken heterophil degranulation and oxidative burst. Heterophils from chickens treated with CpG-ODN exhibited significantly higher (p<0.05) degranulation activity compared to PBS and control ODN (ODN containing no CpG motif) treated groups when stimulated with opsonized Salmonella enterica serovar enteritidis. Similarly, oxidative burst activity, which generates bactericidal reactive oxygen species, was significantly higher (p<0.05) in heterophils from the CpG-ODN treated group than from PBS and control ODN groups when stimulated with formalin-killed S. enteritidis. The priming effects of CpG-ODN on heterophil immune functions continued at least 4 days post-treatment. In the infection study, newly hatched chickens were treated with CpG-ODN, control ODN or PBS for 24h then challenged with oral inoculation of S. enteritidis. A significant reduction (p<0.05) in colonization by S. enteritidis was observed in chickens treated with CpG-ODN. Our study provides evidence that immunostimulatory CpG-ODN potentiates the innate immune responses of heterophils and enhances resistance to infectious pathogens in neonatal chickens.     

Influence of intensive and extensive breeding on lactic acid bacteria isolated from Gallus gallus domesticus ceca

In the present study, lactic acid bacteria (LAB) from the cecum of chickens bred either under intensive (commercial broilers) or extensive (free-range) conditions were isolated, identified and some of their probiotic characteristics determined. The LAB identified by 16S-23S rRNA PCR-ARDRA were mainly of Lactobacillus species and to a lesser extent of Enterococcus spp. for all animals. Free-range chickens showed a higher presence of Lactobacillus acidophilus while Lactobacillus reuteri and Lactobacillus johnsonii were more frequently recovered from commercial broilers. Lactobacillus crispatus was found only in commercial broilers, Lactobacillus vaginalis and Lactobacillus agilis only in free-range chickens and Lactobacillus salivarius in both types. Enterococcus isolates from ceca of commercial broilers showed a higher resistance to antimicrobial drugs. Lactobacillus isolates from free-range chickens presented a higher frequency of in vitro antagonistic activity against selected pathogens than from commercial broilers. All LAB isolates had predominantly non-hydrophobic surfaces, but with variations depending on age of the chickens and breeding conditions. Animal breeding caused variation on composition, antimicrobial susceptibility, antagonistic activity and surface hydrophobicity of LAB from chicken cecum. LAB isolates from ceca of free-range chickens have potential as probiotic agents, which may be used in the future as replacing the use of antimicrobials as growth promoters.     

Sequential spread of Campylobacter infection in a multipen broiler house

Generally, colonization with Campylobacter jejuni is first detected in broilers 2-3 wk after hatching. Once introduced into a flock, this infection spreads very rapidly. The sources and routes of transmission of C. jejuni in broilers remain debatable. In this study, the spread of infection was monitored in a commercial multipen broiler house in which birds were contained in discrete groups and sampled sequentially. Colonization was monitored in two broiler flocks up to slaughter. Serotyping and fla typing methods were applied to differentiate all the C. jejuni strains isolated. In flock 1, colonization was first detected at 32 days of age in birds located at the rear of the house. By 40 days, nearly all the birds were infected with the same strain (fla type 1.9). However, at 46 days of age, a second strain (fla type 3.7) was detected in some of the birds. These birds were also located toward the rear of the house. In flock 2, infection was detected at 5 wk of age. This infection was once again first detected in birds located at the rear of the house. In this flock, only a single fla type (1.1) was isolated throughout. A survey of the broiler house relative to the location of first point of infection indicated the use of an entrance door unprotected by boot dips. However, securing this door during the second flock study did not prevent infection.