Public pressure to reduce or eliminate antimicrobials as ingredients of feed for poultry and other agricultural animals is mounting, primarily due to the fear of multidrug-resistant bacteria in clinical infections in both animals and humans. Exploration of the occurrence of antibiotic resistance in the gut flora of wildlife avian flocks that presumptively do not receive antimicrobials will determine the rate of resistance in a naïve population. Fecal samples collected from a healthy population of the yellow-headed blackbirds (YHB) (Xanthocephalus xanthocephalus) in North Dakota were cultured to determine what genera and species of gram-negative facultative anaerobic bacteria these wild birds carry in their intestinal flora and to evaluate the antimicrobial susceptibility profiles. Isolates of Escherichia coli were further characterized for the presence of putative virulence factors and for pathogenic potential using the chicken embryo lethality assay (ELA). The ELA was performed in chicken embryos with challenges at both 12 days and 16 days of incubation to determine whether the 16-day-old embryos were better able to fight the infection and subsequent disease and also to determine whether the ELA could distinguish between primary and secondary avian Escherichia coli pathogens. After screening 33 isolates from the 21 fecal samples, only two E. coli isolates were identified. The predominant genus and species of bacterium identified was Pantoea agglomerans. Collectively, 12 of the 33 isolates (36%) exhibited no resistance to any antimicrobial tested. However, several multidrug-resistant isolates of varying genera were identified. Among the antimicrobial resistances observed, the most common was to ampicillin (60%), followed by cephalothin (33%). Neither E. coli isolate belonged to serogroups that are notorious for causing major outbreaks of colibacillosis in poultry, and only one E. coli isolate retained resistance to any antibiotics; nevertheless, the ELA results indicate that at least one of these E. coli may be a primary pathogen of chickens. This study demonstrates that antibiotic resistance occurs in the gut flora of natural populations of YHB despite the absence of antibiotic pressure. In addition, these results indicate that YHB will harbor E. coli isolates that are potentially pathogenic in poultry. However, these E. coli isolates are not a significant reservoir for multiple antibiotic resistances nor are they widespread in the population of YHB surveyed in North Dakota.

The virulence factors of avian pathogenic Escherichia coli (APEC) isolated in Japan were investigated. Serogroups O, serotypes K1 and K5, and genes cvaC, iss, iutA, papA, tsh, and usp, which have been thought to be related to virulence, were examined for their association with E. coli strains isolated from diseased and healthy chickens. The frequently recognized serogroups O1, O2, and O78 were found in 56 of 125 (44.8%) strains of diseased chickens (APEC) versus 13 of 100 (13.0%) strains of healthy chickens (commensal E. coli), a significant difference at risk ratio < 0.01. Although iss, iutA, and tsh were widely distributed in the APEC irrespective of O serogroup, papA, usp, and the K1 serotype were detected in serogroup O2 of APEC. The kfiD gene related to the K5 capsule and VT, LT, and ST genes related to exotoxins were not detected in any strains examined.

Avian leukosis viruses (ALVs) are common in many poultry flocks and can be detected using an enzyme-linked immunosorbent assay or any other test designed to identify p27, the group-specific antigen located in gag. However, endogenous retroviruses expressing p27 are often present and can be confused with exogenous ALVs. A more specific and informative assay involves targeting the variable envelope glycoprotein gene (gp85) that is the basis for dividing ALVs into their different subgroups. We designed polymerase chain reaction (PCR) primers that would specifically detect and amplify viruses from each of the six ALV subgroups: A, B, C, D, E, and J. Subgroup B and D envelopes are related, and our B-specific primers also amplified subgroup D viruses. We also designed a set of common primers to amplify any ALV subgroup virus. To demonstrate the usefulness of these primers, we obtained from the Center for Veterinary Biologics in Iowa culture supernatant from chicken embryo fibroblasts infected with an ALV that was found to be a contaminant in two commercial Marek's disease vaccines. Using our PCR primers, we demonstrate that the contaminant was a subgroup A ALV. We cloned and sequenced a portion of the envelope gene and confirmed that the ALV was a subgroup A virus. Unlike typical subgroup A viruses, the contaminant ALV grew very slowly in cell culture. We also cloned and sequenced a portion of the long terminal repeat (LTR) from the contaminant virus. The LTR was found to be similar to those LTRs found in endogenous ALVs (subgroup E) and very dissimilar to LTRs normally found in subgroup A viruses. The E-like LTR probably explains why the contaminant grew so poorly in cell culture.

Newcastle disease virus (NDV) belongs to the Paramyxoviridae family and has had a great impact on the poultry industry. In the past, NDV strains were generally pathogenic only to fowls, but goose paramyxovirus (goose-origin APVM-1) is highly infectious to waterfowl, and, thus, there have been frequent outbreaks in China since 1997. In this study three pairs of specific primers were designed to detect the virulence of different NDV and goose-origin APVM-1 isolates and to differentiate between NDV and goose-origin APVM-1 using multiplex reverse transcription–polymerase chain reaction (RT-PCR). Pathogenicity tests were performed to confirm multiplex RT-PCR results. Data from our study indicate that multiplex RT-PCR is a convenient, low-cost, and effective technique for rapid identification. Twenty-six viral isolates of NDV and four goose-origin APVM-1 were analyzed, and we found that the VII genotype of NDV is the most prevalent type in South China and that it is not closely related to viral strains common to Europe.

A longitudinal survey to detect enteric viruses in intestinal contents collected from turkeys in eight commercial operations and one research facility was performed using molecular detection methods. Intestinal contents were collected from turkeys prior to placement, with each flock resampled at 2, 4, 6, 8, 10, and 12 wk of age. The samples were screened for astrovirus, rotavirus, reovirus, and turkey coronavirus (TCoV) by a reverse transcriptase and polymerase chain reaction (RT-PCR), and for groups 1 and 2 adenovirus by PCR. Rotavirus was the only virus detected prior to placement (7 of 16 samples examined). All of the commercial flocks were positive for rotavirus and astrovirus from 2 until 6 wk of age, and most were intermittently positive until 12 wk of age, when the birds were processed. Of the 96 samples collected from birds on the farms, 89.5% were positive for astrovirus, and 67.7% were positive for rotavirus. All flocks were negative for TCoV, reovirus, and group 1 adenovirus at all time points, and positive for group 2 adenovirus (hemorrhagic enteritis virus) at 6 wk of age. All the flocks monitored were considered healthy or normal by field personnel. Turkeys placed on research facilities that had been empty for months and thoroughly cleaned had higher body weights and lower feed conversion rates at 5 wk of age when compared to turkeys placed on commercial farms. Intestinal samples collected at 1, 2, and 3 wk of age from these turkeys were free of enteric viruses. This report demonstrates that astroviruses and rotaviruses may be present within a turkey flock through the life of the flock. Comparison of infected birds with one group of turkeys that were negative for enteric viruses by the methods used here suggests that astrovirus and/or rotavirus may affect production. The full impact on flock performance needs to be further determined.

Recent studies have revealed the presence of astroviruses and rotavirus in numerous poorly performing and healthy chicken and turkey flocks in the United States. The phylogenetic analysis of the sequence data produced during these studies has identified four groups of avian astroviruses circulating in the United States: turkey astrovirus types 1 and 2 (TAstV-1 and TAstV-2), avian nephritis virus (ANV), and a chicken-origin astrovirus (CAstV). As the molecular epidemiology of poultry enteric disease is poorly understood, the development of updated diagnostic assays is crucial to the continued surveillance and management of enteric disease in affected as well as healthy flocks. This report details the development of a multiplex reverse transcriptase–polymerase chain reaction (RT-PCR) assay specific for astroviruses and avian rotavirus in turkey-origin and chicken-origin samples. The assay consists of two multiplex tests, one for turkey-origin samples and one for chicken-origin samples. The turkey sample test differentially identifies TAstV-1, TAstV-2, ANV, and avian rotavirus. The test for chicken-origin samples differentially identifies CAstV, ANV, and avian rotavirus. Assay sensitivity varied by target sequence between approximately 10 copies for avian rotavirus alone and approximately 2 × 10⁶ copies for TAstV-2 in the presence of a heterologous competitor RNA sequence. Each test was shown to be specific for the intended target by testing for cross-reaction with other common avian enteric viruses. The specificity was further shown by testing 109 chicken specimens and 32 turkey specimens from commercial flocks with the appropriate test and sequencing the RT-PCR amplicons to confirm amplification of the correct target.

Mycoplasma synoviae (MS) is an important pathogen of domestic poultry and is prevalent in commercial layers. During the last decade Escherichia coli peritonitis became a major cause of layer mortality. The possible role of MS in the E. coli peritonitis syndrome of laying hens was studied. Four groups of 64 mycoplasma-free commercial layers at the onset of lay (about 80% daily production) were challenged with a virulent MS strain or a virulent avian E. coli strain or both. The four experimental groups were identified as follows: negative control, E. coli, MS, and MS plus E. coli. A typical E. coli peritonitis mortality was reproduced and included one, three, zero, and five birds in the negative control, E. coli, MS, and MS plus E. coli groups, respectively. Only the increased mortality in the MS plus E. coli group had statistical significance. Four weeks postchallenge 10 clinically normal birds from each of the four experimental groups were necropsied. All of the examined birds in the two MS-challenged groups demonstrated severe tracheal lesions. Body cavity lesions were detected in two and four birds in the MS and MS plus E. coli groups, respectively. The results demonstrate a possible pathogenesis mechanism of respiratory origin with regard to the layer E. coli peritonitis syndrome, show the MS pathological effect in layers, and indicate that a virulent MS strain can act as a complicating factor in the layer E. coli peritonitis syndrome.

Understanding impacts of disease on wild bird populations requires knowing not only mortality rate following infection, but also the proportion of the population that is infected. Greater sage-grouse (Centrocercus urophasianus) in western North America are known to have a high mortality rate following infection with West Nile virus (WNv), but actual infection rates in wild populations remain unknown. We used rates of WNv-related mortality and seroprevalence from radiomarked females to estimate infection rates in a wild greater sage-grouse population in the Powder River basin (PRB) of Montana and Wyoming from 2003 to 2005. Minimum WNv-related mortality rates ranged from 2.4% to 13.3% among years and maximum possible rates ranged from 8.2% to 28.9%. All live-captured birds in 2003 and 2004 tested seronegative. In spring 2005 and spring 2006, 10.3% and 1.8% respectively, of newly captured females tested seropositive for neutralizing antibodies to WNv. These are the first documented cases of sage-grouse surviving infection with WNv. Low to moderate WNv-related mortality in summer followed by low seroprevalence the following spring in all years indicates that annual infection rates were between 4% and 29%. This suggests that most sage-grouse in the PRB have not yet been exposed and remain susceptible. Impacts of WNv in the PRB in the near future will likely depend more on annual variation in temperature and changes in vector distribution than on the spread of resistance. Until the epizootiology of WNv in sagebrush-steppe ecosystems is better understood, we suggest that management to reduce impacts of WNv focus on eliminating man-made water sources that support breeding mosquitoes known to vector the virus. Our findings also underscore problems with using seroprevalence as a surrogate for infection rate and for identifying competent hosts in highly susceptible species.

In general, avian influenza (AI) vaccines protect chickens from morbidity and mortality and reduce, but do not completely prevent, replication of wild AI viruses in the respiratory and intestinal tracts of vaccinated chickens. Therefore, surveillance programs based on serological testing must be developed to differentiate vaccinated flocks infected with wild strains of AI virus from noninfected vaccinated flocks in order to evaluate the success of vaccination in a control program and allow continuation of national and international commerce of poultry and poultry products. In this study, chickens were immunized with a commercial recombinant fowlpox virus vaccine containing an H5 hemagglutinin gene from A/turkey/Ireland/83 (H5N8) avian influenza (AI) virus (rFP-H5) and evaluated for correlation of immunological response by hemagglutination inhibition (HI) or agar gel immunodiffusion (AGID) tests and determination of protection following challenge with a high pathogenicity AI (HPAI) virus. In two different trials, chickens immunized with the rFP-H5 vaccine did not develop AGID antibodies because the vaccine lacks AI nucleoprotein and matrix genes, but 0%–100% had HI antibodies, depending on the AI virus strain used in the HI test, the HI antigen inactivation procedure, and whether the birds had been preimmunized against fowlpox virus. The most consistent and highest HI titers were observed when using A/turkey/Ireland/83 (H5N8) HPAI virus strain as the β-propiolactone (BPL)–inactivated HI test antigen, which matched the hemagglutinin gene insert in the rFP-H5 vaccine. In addition, higher HI titers were observed if ether or a combination of ether and BPL-inactivated virus was used in place of the BPL-inactivated virus. The rFP-H5 vaccinated chickens survived HPAI challenge and antibodies were detected by both AGID and HI tests. In conclusion, we demonstrated that the rFP-H5 vaccine allowed easy serological differentiation of infected from noninfected birds in vaccinated populations of chickens when using standard AGID and HI tests.

Three hundred 1-day-old Japanese quail (Coturnix coturnix japonica) were divided into two groups of 150 each. One group was maintained on quail mash alone, whereas Fusarium verticillioides culture material (FCM) was added to quail mash in the second group from 5 days of age and supplied 150 mg FB₁/kg mash. At day 21, each group was further subdivided into two groups, yielding four groups with 75 birds apiece, which served as the control (group CX), the Salmonella Gallinarum alone group (group CS), the FB₁ alone group (group FX), and the group fed FB₁ and infected with Salmonella Gallinarum (group FS). An oral challenge with Salmonella Gallinarum organisms (2 × 10⁴ colony-forming units [cfu]/ml) was given to groups CS and FS at 21 days of age. Three quail each, were necropsied on day 21 (0 day interval) from groups CX and FX, whereas at subsequent intervals, i.e., 1, 2, 3, 5, 7, 10, 14, and 21 days postinfection (DPI), they were sacrificed from all four groups (CX, CS, FX, and FS) to study the agglutinin response to Salmonella Gallinarum and pathologic changes. The agglutinin titers to Salmonella Gallinarum in the combination group (FS) were generally lower when compared with those in group CS. A reduction in the size of spleen along with depletion of white pulp, thinning of cardiomyocytes, lymphoid cell depletion from bursal follicles, and renal tubular nephrosis were characteristic pathologic changes in group FX. In contrast, there was mild to severe enlargement of spleen accompanied by necrosis and reticuloendothelial cell hyperplasia, pericarditis, myocarditis, and focal interstitial nephritis in groups CS. Similar but more severe lesions were observed in the combination group (FS). In addition, the flabby texture of heart, hydropericardium, and ascites were mainly observed in group FS. It is concluded that continuous presence of fumonisins at 150 mg/kg diet increases the severity of Salmonella Gallinarum infection in young Japanese quail.

Marek's disease virus (MDV) is an oncogenic cell-associated herpesvirus that causes T-cell lymphoma in chickens. Lymphoproliferative neoplasms in Marek's disease (MD) occur in various organs and tissues, including the viscera, peripheral nerves, skin, gonads, and musculatures. MDV is restrictively produced in the feather follicle epithelial (FFE) cells, and it gains access to the external environment via infected cells or as infectious enveloped cell-free virus particles. The goals of the present study were to 1) determine whether the MDV-induced skin lesions are neoplastic in nature or inflammatory reactions to viral infection, 2) determine whether physical presence of feather follicles (FF) is necessary for skin tumor development, and 3) study the role of skin epithelial cells not associated with feathers or FF in the replication and dissemination of infectious virus particles. Scaleless chickens that produce only a few scattered feathers and no sculate scales along the anterior metatarsi were used as a unique model to study the pathogenesis of dermal lesions. Histologic and immunohistochemical analysis revealed that the cutaneous lesions were tumorous as was manifested by massive accumulation of lymphoblasts and extensive activation of meq oncoprotein, the hallmark of MDV oncogenesis, within the skin lesions. Neoplastic cutaneous lesions in the scaleless chickens indicate that feather follicles are not necessary for skin tumor development. Finally, our preliminary data indicate that inoculation with supernatant fluid from homogenized and sonicated skin samples of MDV-infected scaleless chickens induces MD in susceptible birds, suggesting that skin epithelial cells not associated with FF also harbor infectious viral particles.

A concurrent infection of chickens with infectious laryngotracheitis virus (ILTV), a herpesvirus, and fowlpox virus (FWPV), an avipoxvirus, is described. Two techniques, an immunohistochemistry (IHC) technique and a multiplex polymerase chain reaction (PCR), were used to examine 11 tissue samples from chickens clinically diagnosed as FWPV-infected, but only IHC was used to examine six tissue–paraffin blocks prepared from turkeys suspected of having FWPV infection. By multiplex PCR, both FWPV and ILTV were detected from three chicken samples (FI-90, FI-93, and FI-94); both FWPV and ILTV were detected from only two samples (FI-93 and FI-94) by IHC. All chicken samples were positive for FWPV by both PCR and IHC. Viral DNA from these samples was further confirmed by restriction enzyme analysis. When turkey samples were analyzed by the double-stain IHC, all six samples showed the presence of FWPV antigens, but no ILTV antigens. The double IHC technique, using monoclonal antibodies against FWPV and ILTV, was successful in simultaneous demonstration of specific FWPV and ILTV antigens colocalized in infected tissue samples as well as within individual cells. This paper emphasizes the importance of reliable tests that detect specifically the presence of ILTV and FWPV in infected tissue samples. The multiplex PCR assay holds potential to be versatile, rapid, and more sensitive (100%) than IHC (67%) for the simultaneous detection of two different avian viruses. Furthermore, the presence of mixed infection should always be kept in mind in the virologic analysis of respiratory sickness of poultry.

The effects of viral strain, viral dose, and age of bird at inoculation on subgroup J avian leukosis virus (ALV J) persistence, neutralizing antibody (VNAb) response, and tumors were studied in commercial meat-type chickens. Chickens were inoculated on the fifth day of embryonation (5 ED) or on day of hatch (DOH) with either 100 or 10,000 50% tissue-culture infective dose (TCID₅₀) of one of three ALV J strains, namely ADOL Hc1, ADOL 6803, or ADOL 4817. At 1, 3, 7, 11, 15, 19, 23, 27, and 32 wk posthatch, chickens were examined for ALV J viremia and VNAb against the inoculated strain of ALV J. A high incidence (83%–100%) of ALV J persistence was observed in all treatment groups. Development of VNAb did not always lead to viremia-free status; even though 18% of the chickens developed VNAb, only 4% were able to clear viremia. The viral strain, dose, and age of bird at inoculation seemed to have an effect on the incidence of VNAb; however, the differences were statistically significant in only some treatment groups. Chickens infected with ADOL 6803 had higher incidence of VNAb than chickens infected with ADOL Hc1 and ADOL 4817 (P < 0.05 in groups 5 ED at 100 TCID₅₀ and DOH at 10,000 TCID₅₀). There was a trend in all groups inoculated with 100 TCID₅₀ to have higher incidence of VNAb than that of groups inoculated with 10,000 TCID₅₀ (ADOL 6803 at 5 ED and ADOL 4817 at DOH [P < 0.05]; ADOL Hc1 at DOH [P < 0.08]). In most treatment groups (ADOL Hc1 at 100 and 10,000 TCID₅₀, ADOL 6803 at 10,000 TCID₅₀, and ADOL 4817 at 100 TCID₅₀), chickens inoculated at DOH had higher incidence of VNAb than that of chickens inoculated at 5 ED (ADOL 6803 at 10,000 TCID₅₀ [P < 0.05], ADOL Hc1 at 100 TCID₅₀ [P < 0.08]). Incidence of ALV J-induced tumors and tumor spectrum were influenced by viral strain, age at inoculation, and VNAb response.

Live attenuated subtype B avian metapneumovirus (aMPV) vaccines from two different commercial sources (Vac 1 and Vac 2) were used to vaccinate two groups of day-old specific-pathogen-free chicks. The chicks were challenged at 21 or 49 days of age with a virulent subtype B virus. Parameters compared were persistence of vaccine viruses, their ability to induce humoral antibody responses, and the protection they offered against virulent challenge. Vac 1 virus was detectable for at least 14 days after vaccination, and Vac 2 virus was detectable for at least 7 days after vaccination. The serologic response after Vac 1 by using a subtype B-based enzyme-linked immunosorbent assay was significantly higher than that induced by Vac 2, even though the titer of the recommended dose had a higher titer than Vac 1. After challenge at 21 or 49 days of age, both vaccines gave 100% protection against clinical disease. For the recovery of challenge virus, one of 10 chicks given the Vac 2 was positive by virus isolation or reverse transcriptase-polymerase chain reaction (RT-PCR) on each occasion except on day 10 after challenge at 49 days. With groups vaccinated with Vac 1, no challenge virus was detected except on day 5 after challenge at 49 days, when one of 10 chickens was positive only by RT-PCR. This study has shown that commercial aMPV vaccines from different sources can provide identical protection in terms of clinical disease after virulent challenge and in their ability to induce humoral antibodies, even though the titer of their recommended doses is widely different.

Groups of eight chickens were challenged with 10-fold dilutions of one of two strains of Mycoplasma synoviae (MS); each challenge group contained two noninfected sentinels. Both strains were highly efficient in colonizing the respiratory tract with challenge doses as low as 76 and 24 color-changing units/bird. Infection spread rapidly (within 7 days) to sentinels, while uninfected control chickens separated from infected chickens by two empty pens remained uninfected for the 56-day experimental period.

Although sentinels and birds challenged with the lowest doses had weaker or slightly slower antibody responses in some cases as measured by serum plate agglutination, enzyme-linked immunosorbent assay (ELISA), and hemagglutination inhibition (HI), they generally exhibited a typical antibody response. Agglutination reactions tended to be weak, but a high percentage of tests (generally >30% from day 14 postchallenge) were positive. ELISA results were variable, and in some cases reactor rates were low (generally <20%), even though the chickens were colonized in the upper respiratory tract. The HI test was reliable in detecting infected groups; usually >50% were positive from 14 days postchallenge. Mean HI titers were higher when using hemagglutination antigens prepared from the homologous MS strain as compared with antigen prepared from the heterologous strain or with standard antigen prepared from WVU 1853.

To evaluate the possibility of virus transmission through feathers of call ducks, we performed two experiments, intranasal infection study and transmission study, using the Japanese H5N1 highly pathogenic avian influenza virus (HPAIV) A/chicken/Yamaguchi/7/2004 (Ck/Yama/7/04). In Experiment 1, 1-day-old, 2-wk-old, and 4-wk-old birds were inoculated intranasally with Ck/Yama/7/04. Birds in all age groups exhibited necrosis and/or viral antigens in the feather epithelium. Nonpurulent encephalitis and focal necrosis of the pancreas and heart also were common to inoculated birds. In Experiment 2, nine 2-wk-old birds that were orally inoculated with feathers of an infected call duck exhibited the nonpurulent encephalitis, necrosis of the feather epithelium, and focal necrosis of the pancreas and heart, accompanied by viral antigens. These lesions were similar to those in intranasal infection. Some birds were positive for the virus isolation from cloacal swabs and hemagglutination inhibition antibody. The infection was confirmed in seven of nine birds. This study confirmed that the Japanese HPAIV can replicate in the feather epithelium, causing necrosis in call ducks through the natural infection route. It also suggests that feathers of call ducks infected with Ck/Yama/7/04 can be a potential source of infection for unaffected birds in nature.

Bursal samples collected from different field outbreaks in commercially reared chicken flocks from India that were suspected of very virulent (vv) infectious bursal disease (vvIBD) were tested. Two vaccine strains that are commonly being used in India also were included to ascertain their relatedness with the field isolates. When tested with real-time reverse transcriptase–polymerase chain reaction (RT-PCR), 14 of the 15 samples were found to be positive for vvIBD virus (vvIBDV) genetic sequences as determined by the vv232 and vv256 vvIBDV-specific probes. A melting temperature of 50 C and above was characteristic of vvIBDV strains. The vaccine strain infectious bursal disease intermediate (IBDI)-plus (IBDI ) had a higher melting temperature compared with IBDI, suggesting more relatedness to the vvIBDV strains. The real-time RT-PCR technique can be a useful tool in differentiating classic and vvIBDV strains and thereby assist in adopting more effective control strategies. Sequencing of the VP2 hypervariable region of these isolates further confirmed the results of real-time RT-PCR. All the suspected vvIBDV samples were found to share unique amino acid substitutions at positions 222 A, 256 I, 294 I, and 299 S characteristic of the very virulent strains. More sequence differences occurred at the nucleotide level among the vvIBDVs. They shared exactly the same amino acid sequence among themselves and also with the Bangladesh isolate BD-3-99 and some of the Nigerian isolates. They differed by one amino acid from earlier published Indian, Asian, and European vvIBDV VP2 sequences. The nucleotide sequence of IBDI vaccine showed more similarities with vvIBDV sequences; hence, it may be of more value in the control of these very virulent strains.

Infectious bronchitis (IB) disease progression in vaccinated chickens after challenge was evaluated in a single commercial line of layer chickens presenting two different major histocompatibility complex (MHC) B complex genotypes. MHC B genotypes were determined by DNA sequence-based typing of BF2 alleles. In total, 33 B2/B15 and 47 B2/B21 chickens were vaccinated with an Ark-type IB virus (IBV) attenuated vaccine and challenged with Ark-type IBV field isolate AL/4614/98 14 days later. Additional chickens of both genotypes served as unvaccinated/challenged and unvaccinated/nonchallenged controls. Clinical signs, histopathologic analysis, detection of IBV genomes in tears, and IBV-specific immunoglobulin A (IgA) in tears were used to evaluate disease progression and immune response. The incidence of IBV respiratory signs was significantly higher in B2/21 than in B2/B15 MHC genotype birds. However, neither the severity and duration of respiratory signs nor the severity and incidence of histologic lesions differed significantly with MHC genotype. The levels of IBV-specific IgA in tears of vaccinated and challenged chickens did not differ significantly between MHC genotypes. IBV genomes were present in the tears of vaccinated and challenged birds, and the incidence of detectable IBV genomes did not vary significantly with MHC B genotype. From an applied perspective, these results indicate that vaccinated commercial outbred chickens with these MHC genotypes are equally resistant to IBV.

Turkeys exposed to avian metapneumovirus (aMPV) subtype C showed extensive lymphoid cell infiltrations in the nasal turbinates of the upper respiratory tract. The cellular infiltration occurred after the first virus exposure but not after re-exposure. Quantitation of the relative proportions of mucosal immunoglobulin (Ig)A , IgG , and IgM cells in controls and virus-exposed turkeys revealed that at 7 days after the first virus exposure, when mucosal infiltration was well pronounced, there was a significant increase (P < 0.05) in the numbers of infiltrating IgA but not of IgG and IgM cells. After the second virus exposure, although the overall numbers of mucosal lymphoid cells were similar in the virus-exposed and control turkeys, the relative proportions of IgA and IgG cells were significantly higher in the virus-exposed turkeys (P < 0.05) than in controls. Furthermore, elevated levels of aMPV-specific IgA were detected in the nasal secretions and the bile of virus-exposed birds after the second but not after the first virus exposure. These results suggest, for the first time, the possible involvement of local mucosal immunoglobulins in the pathogenesis of aMPV in turkeys.

Colibacillosis, caused by avian pathogenic Escherichia coli (APEC) is a major problem for the poultry industry resulting in significant losses annually. Previous work in our lab and by others has shown that the increased serum survival gene (iss) is a common trait associated with the virulence of APEC. This gene was first described for its contributions to E. coli serum resistance. However, recently published research has called the contribution of iss to this trait into question. In the present study, the level of serum resistance conferred on an E. coli isolate by iss is examined. Additionally, the contribution of λ bor gene to E. coli serum resistance is studied, as iss is thought to be derived from bor and bor occurs commonly among E. coli. To better understand the iss and bor contributions to serum resistance, a series of iss and bor mutants was generated. An iss deletion (iss⁻) mutant showed a significant drop in its resistance to serum. Similarly, a bor mutant showed a drop in serum resistance but not as drastic as that observed with the iss mutant, suggesting that iss contributes more to serum resistance than bor in this E. coli strain. Also, when iss was reintroduced into the iss⁻ mutant the wild-type level of serum resistance was restored, confirming that the deletion of iss was responsible for the change in resistance seen in the mutant.

Ornithobacteriosis is an infectious disease of avian species that has been reported in almost all countries around the world, except Thailand. The objectives of this study were to determine the seroprevalence of Ornithobacterium rhinotracheale (ORT) and to isolate and identify ORT in broilers and broiler breeders in Thailand. Chicken antibodies had been randomly checked from 17 farms (19 flocks) of broilers and 23 farms (28 flocks) of broiler breeders. The seropositive flocks were 63% and 100% in broilers and broiler breeders, respectively. The sera analysis showed that the individual 280 broiler sera antibody responses were 67.5% negative, 12.9% suspect, and 19.6% positive. The individual antibody responses of 510 broiler breeder sera revealed 12.2% negative, 38.0% suspect, and 49.8% positive samples. The bacteria were isolated and identified by polymerase chain reaction (PCR). Bacterial isolation and identification revealed that nine isolates of the 12 PCR analysis samples showed positive results to PCR analysis. All the positive PCR samples were collected from the broiler breeder farms.

Zoonotic transmission of an H5N1 avian influenza A virus to humans in 2003–present has generated increased public health and scientific interest in the prevalence and variability of influenza A viruses in wild birds and their potential threat to human health. Migratory waterfowl and shorebirds are regarded as the primordial reservoir of all influenza A viral subtypes and have been repeatedly implicated in avian influenza outbreaks in domestic poultry and swine. All of the 16 hemagglutinin and nine neuraminidase influenza subtypes have been isolated from wild birds, but waterfowl of the order Anseriformes are the most commonly infected. Using 9-to-11-day-old embryonating chicken egg culture, virus isolation attempts were conducted on 168 cloacal swabs from various resident, imported, and migratory bird species in Barbados during the months of July to October of 2003 and 2004. Hemagglutination assay and reverse transcription–polymerase chain reaction were used to screen all allantoic fluids for the presence of hemagglutinating agents and influenza A virus. Hemagglutination positive–influenza negative samples were also tested for Newcastle disease virus (NDV), which is also found in waterfowl. Two influenza A viruses and one NDV were isolated from Anseriformes (40/168), with isolation rates of 5.0% (2/40) and 2.5% (1/40), respectively, for influenza A and NDV. Sequence analysis of the influenza A virus isolates showed them to be H4N3 viruses that clustered with other North American avian influenza viruses. This is the first report of the presence of influenza A virus and NDV in wild birds in the English-speaking Caribbean.

A simple, user-friendly, and rapid method to detect the presence of antibodies to egg drop syndrome 76 (EDS) virus in chicken sera based on an immunofiltration (flow-through) test was developed. Purified EDS virus antigen was coated onto nitrocellulose membranes housed in a plastic module with layers of absorbent filter pads underneath. Following addition of serum to be tested and washing, monoclonal antibodies or polyclonal serum to chicken immunoglobulin G (IgG) was used as a bridge antibody to mediate binding between EDS virus-specific IgG and protein A gold conjugate. The appearance of a pink dot indicated the presence of antibodies to EDS virus in the sample tested. The results could be obtained within 5–10 min. The developed immunofiltration test could detect antibodies in the sera of experimentally vaccinated chickens from 2 wk postvaccination. With field sera samples, this test was positive in samples having hemagglutination inhibition titers of 8 and above. This test has the potential to be used as a field-based kit to assess seroconversion in EDS-vaccinated flocks.

Turkey breeder hens showed an increase in mortality beginning at 38 wk of age with no other clinical signs or changes in egg production. While no respiratory signs were observed in live turkeys, those that died consistently had gross lesions of pneumonia. Histopathology of lungs revealed serofibrinous bronchopneumonia, lymphofollicular reaction, and other features suggesting a bacterial etiology. However, except for incidental findings, bacteria were not visualized in the sections examined, and none were isolated in meaningful numbers on routine bacteriologic media. At 42 wk of age the flock showed serologic evidence of infection with Mycoplasma synoviae (MS), and MS was identified by both mycoplasma culture and polymerase chain reaction (PCR) procedures in samples from choanal clefts and tracheas. Results of lung histopathology and PCR tests were consistent with a diagnosis of pneumonia caused by MS.

Three 7-wk-old Bobwhite quail were submitted for necropsy to the Douglas branch of the Georgia Poultry Laboratory Network. Grossly, one bird had multiple white foci in the liver and a mild airsacculitis. In this quail there were multiple hepatic granulomas that contained mats of filamentous bacteria easily seen in hematoxylin- and eosin-stained histologic sections. These bacteria were negative with period acid-Schiff and were not acid fast. Bacteria were gram-positive but were most evident on Warthin-Starry silver-stained sections. The appearance and histochemical characteristics of these bacteria are most consistent with Eubacterium tortuosum.

Listeriosis was diagnosed in a 4-yr-old female cockatiel (Nymphicus hollandicus) that died after exhibiting clinical signs that included a fluffed-up appearance, weakness, and loss of weight of several days duration. Grossly, the bird was moderately emaciated, and the liver and spleen were enlarged. Microscopically, there was mild-to-moderate inflammation associated with rod-shaped, gram-positive bacteria in the liver, spleen, kidneys, adrenal glands, bone marrow, and esophagus. Listeria monocytogenes was isolated from the liver, trachea, and intestine. The isolate was identified as type 1 by agglutination with specific antisera, and it further identified as belonging to serovar group 1/2a, 3a by multiplex polymerase chain reaction assay. Listeria monocytogenes also was detected in affected tissues by immunohistochemistry.