Diagnostic utility of egg yolk for the detection of avian metapneumovirus antibodies in laying hens

Surveillance and diagnosis of avian metapneumovirus (AMPV) infection typically involve measurement of serum antibodies. In the current study, eggs instead of serum samples were used for the detection of AMPV antibodies in egg-laying chicken hens by enzyme-linked immunosorbent assay (ELISA). AMPV-free commercial layer hens were experimentally challenged with AMPV strain SC1509 through intravenous or oculonasal administration. Antibody levels were determined by ELISA. AMPV antibodies were detected in egg yolks from challenged hens by 7 days postinoculation (dpi), with the peak titer at 16 dpi. Antibody levels in eggs laid at 28 dpi correlated well (r = 0.93) with sera taken 28 dpi from the same hens. In a field trial of the yolk ELISA, six broiler breeder farms were surveyed, and all tested positive for AMPV antibodies in hen eggs, although positivity varied from farm to farm. Abnormal discolored eggs collected from outbreak farms had significantly higher titers of AMPV yolk antibodies than normal eggs from the same farm, unlike clinically healthy farms, where normal and abnormal eggs had similar antibody titers. These results indicate that diagnosis of AMPV infection by yolk ELISA to detect anti-AMPV antibodies may be a suitable alternative to serologic testing.     

The spread of Salmonella gallinarum 9R vaccine strain under field conditions.

A live vaccine based on an attenuated Salmonella gallinarum 9R strain is in use in a Salmonella enteritidis control program in commercial layer flocks in The Netherlands. In a field study, the potential spread of the vaccine strain from vaccinated flocks to nonvaccinated flocks has been studied after both the primary and the booster injection at four different rearing farms and at one layer farm. The vaccinated and the nonvaccinated flocks were monitored at regular intervals by bacteriologic and serologic examination. In this field study, no evidence was found for the fecal spread of the vaccine strain.     

Use of long-term vaccination with a killed vaccine to prevent fecal shedding of Mycobacterium avium subsp paratuberculosis in dairy herds

OBJECTIVES: To determine whether vaccination with a killed vaccine prevents fecal shedding of Mycobacterium avium subsp paratuberculosis, to compare effectiveness of a culture and cull program in vaccinated and nonvaccinated herds, and to compare paratuberculosis-related preventive management in vaccinated and nonvaccinated herds. SAMPLE POPULATION: 58 commercial Dutch dairy herds. DESIGN: Cross-sectional study (study A) in vaccinated (n = 25) and nonvaccinated (29) herds of dairy cows. Longitudinal study (study B) in vaccinated (n = 2) and nonvaccinated (2) herds of dairy cows. PROCEDURE: In study A, fecal samples were obtained from adult cows in herds with and without a history of vaccination with a killed vaccine. Management measures were evaluated. In study B, fecal samples were obtained 4 times at 6-month intervals from cows older than 6 months. Cows that had positive test results were removed from the herd directly after the outcome of the culture. RESULTS: In study A, differences were not detected among the 25 herds that were vaccinated; culture results were positive for M avium subsp paratuberculosis in 4.4% of herds. In 29 herds that had not been vaccinated, culture results were positive in 6.7%. In study B, the percentage of positive results on culture decreased from 10.9% and 5.7% to 3.5% and 0%, respectively in the 2 vaccinated herds. In the 2 nonvaccinated herds, percentages decreased from 6.1% and 16.5% to 0% and 2.3%, respectively. Management practices were different between herds that were vaccinated and herds that were not; owners of herds that were not vaccinated followed more preventive management procedures and practiced less feeding of raw milk to calves. CONCLUSIONS AND CLINICAL RELEVANCE: Vaccination of calves with a killed vaccine does not prevent transmission of M avium subsp paratuberculosis; therefore, hygienic practices remain essential in herd management.     

Detection of EHV-1 and EHV-4 DNA in unweaned Thoroughbred foals from vaccinated mares on a large stud farm

REASONS FOR PERFORMING STUDY: A silent cycle of equine herpesvirus 1 infection has been described following epidemiological studies in unvaccinated mares and foals. In 1997, an inactivated whole virus EHV-1 and EHV-4 vaccine was released commercially in Australia and used on many stud farms. However, it was not known what effect vaccination might have on the cycle of infection of EHV-1. OBJECTIVE: To investigate whether EHV-1 and EHV-4 could be detected in young foals from vaccinated mares. METHODS: Nasal and blood samples were tested by PCR and ELISA after collection from 237 unvaccinated, unweaned foals and vaccinated and nonvaccinated mares during the breeding season of 2000. RESULTS: EHV-1 and EHV-4 DNA was detected in nasal swab samples from foals as young as age 11 days. CONCLUSIONS: These results confirm that EHV-1 and EHV-4 circulate in vaccinated populations of mares and their unweaned, unvaccinated foals. POTENTIAL RELEVANCE: The evidence that the cycle of EHV-1 and EHV-4 infection is continuing and that very young foals are becoming infected should assist stud farms in their management of the threat posed by these viruses.     

Longitudinal field studies of Avian Metapneumovirus and Turkey Hemorrhagic Enteritis Virus in turkeys suffering from colibacillosis associated mortality

The aim of this study was to evaluate if the exposure to Avian metapneumovirus (aMPV) and/or to Turkey hemorrhagic enteritis virus (THEV) was significant for the induction of episodes of colibacillosis in aMPV and THEV vaccinated turkeys. Colibacillosis-associated mortality was recorded and longitudinal virological studies performed in three consecutive turkey flocks reared in the same farm. aMPV and THEV diagnostic swabs and blood samples were made once a week up to 14 weeks of age. Swabs were processed by molecular techniques for viruses detection and antibody titres were evaluated. Field subtype B aMPVs were detected in all flocks at different ages of life always associated with respiratory signs and increase of colibacillosis-associated mortality. THEV has been consistently detected in all flocks since the 9th week of age. Vaccination with a single dose of the THEV commercial inactivated vaccine available in Italy seems does not protect the birds from the infection. Sequence comparison of the hexon protein of one of the THEV strains detected, and strains isolated worldwide, revealed high similarity between them. These results are consistent with the notion that the hexon protein, being the major antigenic component of the virus, is highly conserved between the strains. Results showed that field aMPV infection is directly correlated to colibacillosis-associated mortality. Less clear appears the role of THEV because the endemicity of aMPV makes difficult to evaluate its role in predisposing colibacillosis in absence of aMPV. It would be interesting to further investigate this issue through experimental trials in secure isolation conditions.     

Vaccination against Salmonella enteritidis in Dutch commercial layer flocks with a vaccine based on a live Salmonella gallinarum 9R strain: evaluation of efficacy, safety, and performance of serologic Salmonella tests

This study describes a field trial in which 80 commercial layer flocks, with an increased risk of Salmonella enteritidis (SE) infection and placed on farms with a certified Standardized Biosecurity Programme (SBP) or a request for a SBP certificate, were vaccinated with a vaccine based on a live attenuated Salmonella gallinarum (SG) 9R strain. An evaluation is presented of the efficacy of the vaccine against SE infections, the effect on the performance of serologic Salmonella tests, and the spread of the vaccine strain to the egg content. For the efficacy study, assessment of the flock level occurrence of SE infections in the vaccinated group of 80 flocks was compared with that of a nonvaccinated group of 1854 flocks hatched in the same period. This control group was examined according to the compulsory control programme in The Netherlands. An evaluation was done of the performance of serologic Salmonella tests and the spread of the vaccine strain to the inner egg content of five of the vaccinated flocks. Findings demonstrated the flock level occurrence of SE infections in the vaccinated group (2/80 = 2.5%) to be significantly (P = 0.01) lower than that of the nonvaccinated group (214/1854 = 11.5%). Vaccination resulted in 59.0% positive test results in lipopolysaccharide BD enzyme-linked immunosorbent assay (ELISA) for detecting antibodies against Salmonella serogroups B and D and 0% positive test results in the rapid plate agglutination test for detecting antibodies against S. pullorum (SP)/SG. The mean specificities of two blocking ELISAs (gm- and i-double antibody sandwich ELISAs) based on the flagellar antigen of SE and Salmonella typhimurium (ST) on the same sera were 99.6% and 96.1%, respectively. The vaccine strain could not be isolated from any of the 450 pools of 10 eggs. On the basis of these results, we concluded that vaccination with a vaccine based on an attenuated SG 9R strain contributes to the reduction of SE infections in commercial layer flocks. Furthermore, serologic monitoring of SE, ST, and SP/SG can still be carried out on flocks vaccinated with an attenuated SG 9R strain. Additionally, we found no indication of the spread of the vaccine strain to the egg content.     

Key role of Chlamydophila psittaci on Belgian turkey farms in association with other respiratory pathogens

Two hundred turkey sera from eight Belgian and two French farms were tested for the presence of antibodies against avian pneumovirus (APV), Ornithobacterium rhinotracheale (ORT), Mycoplasma gallisepticum, Mycoplasma meleagridis and Chlamydophila psittaci. At slaughter, C. psittaci, APV and ORT antibodies were detected in 94, 34 and 6.5% of the turkeys, respectively. No antibodies against M. gallisepticum or M. meleagridis were present. Additionally, turkeys on three Belgian farms were examined from production onset until slaughter using both serology and antigen or gene detection. All farms experienced two C. psittaci infection waves, at 3-6 and 8-12 weeks of age. Each first infection wave was closely followed by an ORT infection starting at the age of 6-8 weeks, which was still detectable when the second C. psittaci infection waves started. Animals on farm A were not vaccinated against APV leading to an APV subtype B outbreak accompanying the first C. psittaci infection wave. Despite subtype A APV vaccination on farms B and C, the second C. psittaci infection waves were accompanied (farm B) or followed (farm C) by a subtype B APV infection. On all farms respiratory signs always appeared together with a proven C. psittaci, APV and/or ORT infection. This study suggests an association between C. psittaci, APV and ORT, and indicates the multi-factorial aetiology of respiratory infections in commercial turkeys. All three pathogens should be considered when developing prevention strategies for respiratory disease.     

Antibody response of pigs to inactivated monovalent and bivalent vaccines for porcine parvovirus and pseudorabies virus

Groups of pigs vaccinated with an inactivated bivalent vaccine containing porcine parvovirus (PPV) and pseudorabies virus (PRV) developed geometric mean titers (GMT) of humoral antibody for each of the viruses as high or slightly higher than those of other groups of pigs that were vaccinated with inactivated monovalent vaccines containing one or the other of the same viruses. An increase in GMT after challenge exposure of vaccinated pigs to live virus indicated that vaccination did not prevent virus replication. However, an indication that replication was less extensive in vaccinated pigs was provided by the following. Although neither vaccinated nor nonvaccinated (control) pigs had clinical signs after exposure to the live PPV, the effect of vaccination was evident by the fact that GMT were higher in nonvaccinated pigs after exposure than they were in vaccinated pigs. Conversely, all pigs exposed to live PRV had clinical signs, but these signs varied between mild-to-moderate and transient for vaccinated pigs to severe and fatal for nonvaccinated pigs.     

A field trial to assess the therapeutic and prophylactic effect of a foot rot vaccine in sheep

An 18-week field trial was conducted on a sheep ranch to evaluate the therapeutic and prophylactic effect of a commercial foot rot vaccine. Two hundred sheep were included in the study, 100 with detectable foot rot lesions and 100 without. Approximately 50 sheep from each group were selected randomly and vaccinated twice against foot rot at a 6-week interval in the late spring 1984; the remaining sheep acted as nonvaccinated controls. Therapeutic effect of the vaccine was demonstrated by a cure rate of 53% in vaccinated, foot rot-affected sheep vs a cure rate of 19% in nonvaccinated, foot rot-affected sheep. Prophylactic effect of the vaccine was demonstrated by a foot rot prevalence at the end of the 18-week period of 9% for vaccinated sheep vs 53% for nonvaccinated sheep. Associations of foot rot lesions and vaccination with body condition were found to be significant, as was the association between foot rot lesions and mortality.     

Field efficacy of different vaccines against infectious bursal disease in broiler flocks

A field study was performed to determine the efficacy of three commercially available vaccines against infectious bursal disease (IBD) in commercial broilers raised in a high IBD virus (IBDV) risk area. Live attenuated intermediate and intermediate plus vaccines were used in four flocks. Birds were vaccinated orally at the estimated vaccination time. Three broiler flocks were vaccinated subcutaneously with a turkey herpesvirus (HVT)-IBD vector vaccine at one day old. Evaluation of the efficacy of different vaccines was focused on humoral immune response, bursa/body weight (B/Bw) ratio, molecular detection of IBDV in ileocaecal tonsils and bursa of Fabricius, and production parameters. The serological results showed that although the uptake of all three vaccine strains was confirmed in the lymphoid organs, no significant antibody response to vaccination was detected in flocks vaccinated with intermediate and intermediate plus vaccines. A significant increase in antibody titres detected in flocks vaccinated with the vector vaccine indicated its ability to induce an immune response in birds with a high level of maternally derived antibodies. Observations obtained in this field trial did not confirm the expected reduction of the B/Bw ratio in flocks vaccinated with less attenuated vaccines. No significant differences were observed between birds vaccinated with the vector vaccine and those immunised with the intermediate plus vaccine. Very virulent IBDV was confirmed in the flock vaccinated with the intermediate vaccine. The infection induced reduced B/Bw and moderate mortality but did not affect the production parameters. Field infection was not detected in broilers vaccinated with the intermediate plus vaccine and the vector vaccine.     

Immunomodular effect of fusion gene DNA vaccine of avian metapneumoviruses

The objective of this work was to develop and evaluate the immunomodular effect of a DNA vaccine based on the fusion (F) gene of avian metapneumoviruses (aMPV) and to study its protection against field virus challenge, as this will help to better control the disease in turkeys. In this study, the F protein of the Egyptian isolate (Giza-turkey rhinotracheitis-4) of the B-subtype of aMPV isolated in 2009 was expressed from a DNA plasmid in Vero cells. After 1 i.m. injection of turkey poults with this plasmid, the antibody response was detected by ELISA. The turkey poults inoculated with locally prepared DNA aMPV vaccine had highly significant phagocytic activity, as measured by phagocytic percent and index of macrophage activation, in comparison to those inoculated with inactivated and live attenuated vaccines and with the noninfected control group. Intratracheal challenge of turkey poults at 21 d postvaccination by a dose of 100 uL of field Egyptian Giza-turkey rhinotracheitis-4 virus of a titer 6 log₁₀ tissue culture infective dose 50 resulted in 100% protection in poults that received locally prepared DNA aMPV vaccine, whereas those that received commercial aMPV vaccines experienced 80 and 90% protection; typical clinical signs of aMPV infection were seen in control nonvaccinated poults. Therefore, a high success rate was noted when using F gene DNA plasmid vaccine by the induction of a potent immunomodular effect for both cell-mediated and humoral immune response. The use of the F gene DNA plasmid vaccine developed in this study provided 100% protection in vaccinated poults, which can help in controlling aMPV infections in turkeys.     

Characterisation of an antigenically unusual virus responsible for two outbreaks of Newcastle disease in the Republic of Ireland in 1990.

Antigenic characterisation of two highly virulent virus isolates from outbreaks of Newcastle disease on two closely connected farms in County Monaghan, Republic of Ireland, in 1990 showed the viruses to be indistinguishable but unlike other Newcastle disease viruses so far tested. However, they appeared to be antigenically closest to avirulent viruses isolated from waterfowl from several countries and from chickens in Northern Ireland in 1986. Despite the antigenic differences, chickens vaccinated with a live commercial Hitchner B1 vaccine were protected against intramuscular challenge with one of the virulent isolates.     

Floor pen study to evaluate the serological response of broiler breeders after vaccination with ts-11 strain Mycoplasma gallisepticum vaccine

To determine the Mycoplasma gallisepticum (MG) rapid serum plate agglutination (RSPA) test response of broiler breeders after ts-11 strain vaccination, 55 Cobb pullets derived from a nonvaccinated, MG-negative, commercial, broiler breeder grandparent flock were monitored from 8 to 20 wk of age (over a 12-wk trial period). To evaluate the effect of lateral spread of the ts-11 vaccine strain on RSPA test results from commingled and adjacently penned birds, treatment groups included (A) birds vaccinated with ts-11strain MG at 8 wk of age, (B) commingled nonvaccinates in the same pen as the vaccinated birds, (C) nonvaccinates in a second pen separated from the first pen by a distance of 2 m, and (D) birds vaccinated with ts-11 strain MG at 8 wk of age and kept in a separate room. Rapid serum plate agglutination tests were performed once a week for 6 wk and then every 2 wk for 6 more wk, postvaccination. A polymerase chain reaction (PCR) assay specific fbr ts-11 strain MG was used to confirm vaccination, and a second PCR specific for non-ts-11 strain MG was used to confirm the absence of field infection. Seroconversion was first detected by the RSPA test 2 wk postvaccination and attained maximum positive rates of 58% at 12 wk postvaccination in treatment A and 60% at 8 wk postvaccination in treatment D. Seroconversion rates in nonvaccinated, commingled pullets was 10% at 5 wk and 30% at 12 wk after the vaccination of pen mates. The ts-11-specific PCR detected the vaccine strain in 80%-100% of the vaccinated birds 2 wk after vaccination. One of 15 nonvaccinated birds penned 2 m from vaccinated birds yielded ts-11 by PCR assay 12 wk after vaccination, which indicates that the spread of ts-11 over short distances may be possible in situations in which there is a common caretaker. PCR on tracheal swabs taken 12 wk postvaccination detected ts-l1 in 50% and 60% of the vaccinated birds in treatments A and D, respectively; in 30% of the commingled nonvaccinates; and in 6.6% of the separately penned nonvaccinates. In contrast, choanal swabs collected from vaccinated birds at 12 wk were 21% and 40% PCR positive for ts-11 strain MG, while those from nonvaccinates were negative. All samples were PCR negative for field strain MG. The pattern of seroconversion as measured by RSPA test in small groups of broiler breeders was different from that previously reported for leghorns. Lateral spread of the ts-11 strain to commingled pen mates occurred rapidly, causing RSPA seroconversion patterns that mimicked those of the vaccinated pen mates.     

Immunogenicity and efficacy study of a commercial Borrelia burgdorferi bacterin.

The immunogenicity and efficacy of a commercial Borrelia burgdorferi bacterin was evaluated for stimulation of the host immune response and protection against clinical disease associated with experimentally induced borreliosis in dogs. A total of 30 vaccinated and 24 control dogs were used in 3 separate studies. The vaccine was given IM as two 1-ml doses separated by a 3-week interval. Two weeks or 5 months following the last vaccination, the dogs were challenge inoculated with 7 daily doses of a virulent preparation of a B burgdorferi field isolate through intraperitoneal, subcutaneous, and intradermal routes with or without glucocorticoid administration at the same time. The development of B burgdorferi spirochetemia and clinical disease in the dogs after challenge exposure was studied. Serum samples were obtained from the dogs at various times during the study for serum neutralizing antibody determination and protein immunoblot antibody assay against various geographic isolates of B burgdorferi. Challenge exposure induced limb/joint disorder, fever, anorexia, signs of depression, and B burgdorferi spirochetemia in the nonvaccinated control dogs. The vaccine was found to elicit cross-reactive serum neutralizing and protein immunoblot antibody responses in dogs to various isolates of B burgdorferi and to protect the vaccinated dogs against experimentally induced borreliosis.     

Canine parainfluenza-Bordetella bronchiseptica vaccine immunogenicity

The immunogenicity and safety of 3 serials of a canine parainfluenza (CPI) virus-Bordetella bronchiseptica vaccine was evaluated. Each serial was used to vaccinate 10 dogs with single doses given intranasally. The 30 vaccinated and 10 nonvaccinated controls dogs were challenge exposed with aerosols of virulent CPI virus and B bronchiseptica at 18 days and at 21 days, respectively, after vaccination. After challenge exposure, none of the 30 vaccinated dogs had clinical signs of disease; however, 9 of the 10 nonvaccinated dogs developed coughing problems. The CPI virus was isolated from nasal swab specimens obtained from nonvaccinated dogs on an average of 5.1 days after challenge exposure, but was not isolated from any of the specimens obtained from the vaccinated dogs. Bordetella bronchiseptica was isolated from nasal swab specimens obtained from both vaccinated and nonvaccinated dogs up to 18 days after challenge exposure. The erythrocyte sedimentation rates and total leukocyte counts for control dogs were generally increased, in contrast to those for the vaccinated groups. Dogs showed a primary serologic response to CPI virus and B bronchiseptica after vaccination and an anamnestic response to the bacterium after challenge exposure. Adverse local or systemic reactions attributable to the bivalent vaccine were not observed in the vaccinated dogs.     

Effect of in ovo administered reovirus vaccines on immune responses of specific-pathogen-free chickens

We investigated the effect of in ovo administered reovirus vaccines on the immune responses of specific-pathogen-free chickens. T-cell mitogenic responses to concanavalin A were numerically lower at 9 and 12 days of age and significantly lower at 6 days of age in birds vaccinated with a commercial reovirus vaccine compared with unvaccinated birds or birds vaccinated with an experimental reovirus-antibody complex vaccine. There were no significant differences in proportions of subpopulations of helper (CD4+CD8-) or cytotoxic (CD4-CD8+) T cells except at 12 days of age, when the percentages of CD4-CD8+ cells in the two vaccinated groups were statistically higher than in the nonvaccinated group. B-cell populations were not different among vaccine groups except at 9 days of age, when the vaccinated groups had the highest level of B cells. This commercial reovirus vaccine should not be given in ovo to embryos having little or no maternal antibody, otherwise immunosuppression may occur in the chicks. The addition of the antibody complex to the vaccine prevented this T-cell immunosuppression.     

Avian metapneumovirus subtype A in China and subtypes A and B in Nigeria

In order to detect and characterize avian metapneumovirus, organs or swabs were collected from 697 chicken and 110 turkeys from commercial farms in Southwestern Nigeria and from 107 chickens from live bird markets in Southeastern China. In Nigeria, 15% and 6% of the chicken and turkey samples, respectively, and 39% of the chicken samples from China, were positive for aMPV genome by PCR. The sequence of a 400 nt fragment of the attachment protein gene (G gene) revealed the presence of aMPV subtype A in both Nigeria and Southeastern China. Essentially identical subtype A viruses were found in both countries and were also previously reported from Brazil and the United Kingdom, suggesting a link between these countries or a common source of this subtype. In Nigeria, subtype B was also found, which may be a reflection of chicken importations from most major poultry-producing countries in Europe and Asia. In order to justify countermeasures, further studies are warranted to better understand the metapneumoviruses and their impact on poultry production.     

Evaluation of the antibody response in pigs vaccinated against Streptococcus suis capsular type 2 using a double-antibody sandwich enzyme-linked immunosorbent assay.

A double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) was standardized for the detection of specific antibodies following vaccination with Streptococcus suis capsular type 2 bacterins. No statistically significant increase of antibody titers was detected in vaccinated piglets compared to the nonvaccinated control group, even if a minority of piglets demonstrated an important postvaccinal response. Three of four vaccinated sows showed a low antibody response to vaccine and specific immunity was detected in piglets of only one litter of these three sows. Passive protection studies showed that none of the sera from vaccinated piglets were protective for mice whereas serum obtained from hyperimmunized pigs gave protection.