Studies of infectious laryngotracheitis vaccines: immunity in broilers

Broiler chickens were vaccinated at 18 days of age against infectious laryngotracheitis (ILT) using chicken-embryo-origin (CEO) and tissue-culture-origin (TCO) vaccines, each vaccine given either by drinking water, spray, or eyedrop. Controls were not vaccinated. The broilers were challenged 3 weeks later with virulent ILT virus (USDA challenge strain). Serum samples taken before challenge were analyzed by a virus neutralization (VN) test to determine titers due to vaccination. Both vaccines, regardless of route of administration, produced low VN titers, geometric mean titer (GMT) being less than 4.0 in all vaccinated groups. When administered by the same route, the CEO vaccine produced higher titers than the TCO vaccine. Titers following drinking-water or eyedrop administration of vaccines were higher than titers following spray vaccination. There was an inverse relationship between pre-challenge VN titers of groups of birds and the percentage of birds in the groups dying from ILT virus challenge. The drinking-water route of vaccination provided the most protection, while the spray provided the least.     

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.     

Protection of chickens after live and inactivated virus vaccination against challenge with nephropathogenic infectious bronchitis virus PA/Wolgemuth/98

Protection provided by live and inactivated virus vaccination against challenge with the virulent nephropathogenic infectious bronchitis virus (NIBV) strain PA/Wolgemuth/98 was assessed. Vaccinations with combinations of live attenuated strains Massachusetts (Mass) + Connecticut (Conn) or Mass + Arkansas (Ark) were given by eyedrop to 2-wk-old specific-pathogen-free leghorn chickens. After live infectious bronchitis virus (IBV) vaccination, some chickens at 6 wk of age received an injection of either an oil emulsion vaccine containing inactivated IBV strains Mass + Ark or an autogenous vaccine prepared from NIBV PA/Wolgemuth/98. Challenge with PA/Wolgemuth/98 was given via eyedrop at 10 wk of age. Serum IBV enzyme-linked immunosorbent assay antibody geometric mean titers (GMTs) after vaccination with the combinations of live attenuated strains were low, ranging from 184 to 1,354, prior to NIBV challenge at 10 wk of age. Both inactivated vaccines induced an anamnestic response of similar magnitudes with serum GMTs of 6,232-12,241. Assessment of protection following NIBV challenge was based on several criteria virus reisolation from trachea and kidney and renal microscopic pathology and IBV-specific antigen immunohistochemistry (IHC). Live attenuated virus vaccination alone with combinations of strains Mass + Conn or Mass + Ark did not protect the respiratory tract and kidney of chickens after PA/Wolgemuth/98 challenge. Chickens given a live combination vaccination of Mass + Conn and boosted with an inactivated Mass + Ark vaccine were also susceptible to NIBV challenge on the basis of virus isolation from trachea and kidney butshowed protection on the basis of renal microscopic pathology and IHC. Live IBV-primed chickens vaccinated with an autogenous inactivated PA/Wolgemuth/98 vaccine had the highest protection against homologous virulent NIBV challenge on the basis of virus isolation.     

Protective immunity to infectious bronchitis in broilers vaccinated against Marek's disease either in ovo or at hatch and against infectious bronchitis at hatch

Two experiments were conducted using commercial broiler chickens to determine if Marek's disease (MD) vaccines HVT/SB-1 and HVT plus CVI-988 given either in ovo or at hatch adversely affected the efficacy of infectious bronchitis (IB) vaccines (Ark and Mass serotypes) given by eyedrop on the day of hatch. Nonvaccinated negative controls and controls that received only IB vaccines were included in each study. Birds were challenged with either infectious bronchitis virus (IBV) Mass-41 or IBV Ark-99 on either day 26 or 27 of age. Protection was assessed 5 days post-IBV challenged by virus isolation from the trachea. The day of hatch mean antibody titer to IBV was 12,668 +/- 4704 and 2503 +/- 3243 by enzyme-linked immunosorbent assay in experiments 1 and 2, respectively. In each study, nonvaccinated controls had a significantly higher (P < or = 0.05) incidence (88%-100%) of IBV challenge virus isolation than did controls vaccinated for IB but not for MD. Analysis of data from both studies showed that protection to IB in groups that received only IB vaccines at hatch ranged from 55.0% to 77.3%, whereas protection to IB in groups receiving both MD and IB vaccines ranged from 50.0% to 95.5%. In both experiments and within IBV challenge serotype, broilers given MD vaccines (in ovo or at hatch) and IB vaccines at hatch had protection rates to IBV challenges that were not significantly less (P < or = 0.05) than IB protection rates of groups that received only IB vaccines at hatch. Analysis of these data shows that administration of high-titered MD vaccines either in ovo or at hatch did not affect the efficacy of an IB vaccination (serotypes Ark and Mass) given by eyedrop at hatch.     

Fowl cholera immunity induced by various vaccines in broiler minibreeder chickens determined by enzyme-linked immunosorbent assay

Broiler minibreeder hens were vaccinated for protection against fowl cholera at 12 and 21 weeks of age using several vaccination schemes, which included a live Pasteurella multocida (CU strain) vaccine, two commercial polyvalent fowl cholera oil-based bacterins, and two experimentally prepared polyvalent oil-based bacterins. Some treatment groups received only live or killed vaccines, whereas others received a live vaccine at 12 weeks followed by a killed product at 21 weeks. At 42 weeks of age, all birds that received the live CU vaccine twice or once followed by a bacterin survived challenge. Birds that received killed vaccines only were significantly less protected but still showed a respectable survival rate of 86%. All unvaccinated controls died within 72 hr after challenge. At 72 weeks of age, overall protection was lower than that at 42 weeks, regardless of vaccination treatment. Antibody titers were usually higher in birds that received bacterins than in those receiving live vaccines, yet overall protection was still greater in those birds that received the live cholera vaccine twice.     

Detection of infectious bursal disease vaccine viruses in lymphoid tissues after in ovo vaccination of specific-pathogen-free embryos.

Control of infectious bursal disease virus (IBDV) by vaccination is important for poultry production worldwide. Two vaccines, an IBDV immune complex (ICX) vaccine and an IBDV-2512 vaccine, were administered at 100 mean embryo infectious dose to specific-pathogen-free 18-day-old broiler embryos in ovo. At 3, 6, 9, 15, and 21 days post in ovo vaccination (PIOV), bursa, spleen, and thymus tissues were collected and analyzed for virus protein by antigen capture chemiluminescent enzyme-linked immunosorbent assay (ELISA). Chicks were bled and antibody titers were determined by the antibody ELISA. At 21 days PIOV, chickens were challenged with a 1:500 dilution of an antigenic standard IBDV strain. At 28 days PIOV, birds were euthanatized and bursa weight:body weight ratios were determined. Embryos vaccinated with either vaccine exhibited 92% hatchability; however, within 1 wk of hatch, birds vaccinated with IBDV-2512 showed 56% mortality, whereas those given IBDV-ICX had only 3.2% mortality. Both IBDV-ICX and IBDV-2512 vaccines were detected in bursa, spleen, and thymus at day 3 PIOV. A 5-day delay in virus replication was observed with IBDV-ICX vaccine. By day 15 PIOV, the IBDV-ICX was no longer detectable in the bursa and spleen but persisted in the thymus. The IBDV-2512 vaccine persisted in the spleen and thymus on day 15 PIOV. By day 21 PIOV, neither vaccine virus was detected in any lymphoid organ. This assay can be useful in the early detection of vaccine virus in the tissues of chickens vaccinated via the in ovo route. Both vaccines caused bursal atrophy at all times PIOV. The IBDV-2512 caused splenomegaly at day 6 PIOV, whereas splenomegaly was not seen in IBDV-ICX-vaccinated birds until day 9 PIOV. Thymus atrophy was observed in IBDV-2512-vaccinated chicks from day 3 PIOV, whereas this occurred on day 15 PIOV in IBDV-ICX-vaccinated birds. Bursa weight: body weight ratios in IBDV-ICX-vaccinated unchallenged and vaccinated challenged birds were not different (P < 0.05).     

Evaluation of the effectiveness of two infectious bronchitis virus vaccine programs for preventing disease caused by a California IBV field isolate

Infectious bronchitis virus CA99 serotype was isolated from several broiler flocks in Northern California. The virus caused late-onset respiratory disease and increased airsacculitis condemnation in affected flocks despite the use of an established infectious bronchitis virus vaccination program. An experimental study compared Holland/Arkansas and Massachusetts/Arkansas vaccination protocols to determine the efficacy of commercial infectious bronchitis virus vaccines in reducing respiratory disease and airsacculitis lesions found at processing that were associated with a CA99 field isolate. All vaccination groups were given Massachusetts/Connecticut strains of infectious bronchitis virus vaccines at age 1 day followed by vaccination with either Holland/ Arkansas or Massachusetts/Arkansas vaccine strains at 18 days of age. Birds were challenged at age 31 days with a CA99 field isolate. Gross pathology, histopathology, and virus isolation were evaluated. Chickens vaccinated with Holland/Arkansas had marginally better protection against CA99 challenge than chickens vaccinated with Massachusetts/Arkansas, although differences were not statistically significant.     

Efficacy in turkeys of spray vaccination with a temperature-sensitive mutant of Bordetella avium (Art Vax)

Broad-breasted white turkeys were vaccinated with a temperature-sensitive mutant of Bordetella avium (Art Vax) at 2 and 15 days of age and challenged at 22 days of age by contact with infected birds. Necropsy was performed at 35 days of age. Two vaccination protocols (eyedrop/oral and spray cabinet/spray bottle) and two challenge isolates (Arkansas 105 and North Carolina [NC] isolates) were used. Neither the spray nor the eyedrop/oral methods of vaccination prevented infection of the anterior trachea with either of the virulent challenge strains. The spray and eyedrop/oral methods of vaccination were equally effective in reducing the severity of gross lesions in the trachea. The vaccine reduced the severity of gross lesions in the tracheas of turkeys challenged with the NC isolate to a level approximately equal to that observed in unchallenged vaccinated controls, but the vaccine only moderately reduced the severity of lesions in birds challenged with the 105 isolate.     

Studies of infectious laryngotracheitis vaccines: immunity in layers

Ten-week-old layer chickens obtained from a commercial source were eye-drop vaccinated with chicken-embryo-origin (CEO) or tissue-culture-origin (TCO) vaccines for infectious laryngotracheitis (ILT). Controls were not vaccinated. Approximately one-third of the layers were challenged with virulent ILT virus at 21, 40, or 60 weeks of age. Serum samples taken from the layers before challenge were used in a virus neutralization (VN) test to determine vaccination titers at those three ages. Both vaccines induced low VN titers (geometric mean titer [GMT] less than 6). At 21 weeks of age, the titers produced by the two vaccines were not significantly different, but at 40 and 60 weeks of age the VN GMT of the CEO-vaccinated group was significantly greater than that of the TCO-vaccinated group. The VN GMTs did not drop over time in either group and actually rose between 21 and 60 weeks of age in the CEO group. Both vaccines protected layers against severe challenge with virulent ILT virus, neither being significantly better than the other under these experimental conditions. Unvaccinated sentinel chickens were maintained in contact with the vaccinated layers during three intervals between 1 day and 6 weeks post-vaccination. Diagnostic tests performed on the sentinels to detect lateral spread of vaccine virus from vaccinated to unvaccinated chickens showed scattered positive results.     

Virulence of five live vaccines against avian infectious laryngotracheitis and their immunogenicity and spread after eyedrop or spray application

Five live virus vaccines against avian infectious laryngotracheitis were studied with regard to safety, immunogenicity and route of administration. Significant differences in virulence between the vaccine strains were found. Reduced virulence was accompanied by a reduction of immunogenicity and capacity to spread. After eyedrop application, a low virulent vaccine induced 90-100% flock immunity for the first 10 weeks after vaccination (PV), followed by a slow decline to 50% at 31 weeks PV, whereas flock immunity induced with the more virulent types remained at about 90% till the end of the experiments (24 and 48 weeks PV). Aerosol vaccination induced 70-100% flock immunity but vaccine reactions were severe. Application of vaccine in a coarse spray did not result in adverse vaccine reactions but induced a maximal protection rate of only 50%. Microneutralisation titres provided a useful indicator of immunity from the onset of immunity until immunity started to decline. A vaccine virus carrier state was demonstrated by means of sentinel birds.     

Adjuvanted Outer Membrane Protein Vaccine Protects Poultry against Infection with Salmonella enteritidis

The immunogenicity of a sonicated extract (SE) and of outer membrane proteins (OMP) of Salmonella enteritidis was tested in birds of about 8 weeks of age. The dose, route of vaccination and the adjuvant used varied in different groups of birds. Two vaccine doses with or without adjuvant were given parenterally or orally 3 weeks apart. OMP vaccines gave significantly higher antibody titres than SE vaccines, as indicated by ELISA. The vaccines adjuvanted with oil produced higher antibody titres than those without any adjuvant. A dose of 1 mg of vaccine produced higher antibody titres than 0.5 mg of vaccine. Adjuvanted vaccine given subcutaneously elicited higher antibody responses than oral vaccines given without adjuvant. The birds were challenged with virulent S. enteritidis organisms at the end of the second week after a booster dose. None of the birds given 1 mg of OMP vaccine subcutaneously shed the organisms when tested by culturing cloacal swabs, although a few birds vaccinated with 0.5 mg of OMP vaccine did so. In general, adjuvanted OMP vaccines gave better protection than SE vaccines.     

Virulence of five live vaccines against Avian Infectious Laryngotracheitis and their immunogenicity and spread after eyedrop or spray application

Summary

Five live virus vaccines against avian infectious laryngotracheitis were studied with regard to safety, immunogenicity and route of administration. Significant differences in virulence between the vaccine strains were found. Reduced virulence was accompanied by a reduction of immunogenicity and capacity to spread.

After eyedrop application, a low virulent vaccine induced 90‐ 100% flock immunity for the first 10 weeks after vaccination (PV), followed by a slow decline to 50% at 31 weeks PV, whereas flock immunity induced with the more virulent types remained at about 90% till the end of the experiments (24 and 48 weeks PV).

Aerosol vaccination induced 70 ‐ 100% flock immunity but vaccine reactions were severe. Application of vaccine in a coarse spray did not result in adverse vaccine reactions but induced a maximal protection rate of only 50%.

Microneutralisation titres provided a useful indicator of immunity from the onset of immunity until immunity started to decline. A vaccine virus carrier state was demonstrated by means of sentinel birds.     

Response to and efficacy of vaccination against eastern equine encephalomyelitis virus in emus

OBJECTIVE: To evaluate humoral immune responses of emus vaccinated with commercially available equine polyvalent or experimental monovalent eastern equine encephalomyelitis (EEE) virus and western equine encephalomyelitis (WEE) virus vaccines and to determine whether vaccinated emus were protected against challenge with EEE virus. DESIGN: Cohort study. ANIMALS: 25 emus. PROCEDURE: Birds were randomly assigned to groups (n = 5/group) and vaccinated with 1 of 2 commercially available polyvalent equine vaccines, a monovalent EEE virus vaccine, or a monovalent WEE virus vaccine or were not vaccinated. Neutralizing antibody responses against EEE and WEE viruses were examined at regular intervals for up to 9 months. All emus vaccinated with the equine vaccines and 2 unvaccinated control birds were challenged with EEE virus. An additional unvaccinated bird was housed with the control birds to assess the possibility of contact transmission. RESULTS: All 4 vaccines induced detectable neutralizing antibody titers, and all birds vaccinated with the equine vaccines were fully protected against an otherwise lethal dose of EEE virus. Unvaccinated challenged birds developed viremia (> 10(9) plaque-forming units/ml of blood) and shed virus in feces, oral secretions, and regurgitated material. The unvaccinated pen-mate became infected in the absence of mosquito vectors, presumably as a result of direct virus transmission between birds. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that emus infected with EEE virus develop a high-titer viremia and suggest that they may serve as important virus reservoirs. Infected emus shed EEE virus in secretions and excretions, making them a direct hazard to pen-mates and attending humans. Commercially available polyvalent equine vaccines protect emus against EEE virus infection.     

Functional analysis of antibody responses of feedlot cattle to bovine respiratory syncytial virus following vaccination with mixed vaccines.

The antibody response of cattle to bovine respiratory syncytial virus (BRSV) immunization was investigated using 4 different commercially available mixed vaccines. Forty, 5-6 month old, beef calves, randomly assigned to groups of 10, were vaccinated on day 0 and 21 with 1 of 3 inactivated vaccines, (3 groups), or a modified live virus (MLV) vaccine. BRSV-specific antibody responses were measured prior to vaccination and on day 35 by using an enzyme linked immunosorbent assay (ELISA), virus neutralization assay (VN), a fusion inhibition assay (FI); and responses were also measured for their ability to facilitate antibody dependent, complement mediated cytotoxicity (ADCMC) of BRSV infected cells. Sera from day 35 were, in addition, analyzed by use of an IgG1, IgG2 isotype specific ELISA. All vaccines induced significant increases in BRSV specific IgG antibody as measured by ELISA, but only one inactivated and the MLV vaccine induced significant increases in VN titers. Fusion inhibiting antibody titers were low or undetected in calves vaccinated with the inactivated vaccines. Vaccination with modified live virus induced significantly higher titers of fusion inhibiting antibodies, which are considered to be most highly correlated with protection. The VN to ELISA and FI to ELISA ratio of the calves that received MLV vaccine were significantly greater than the calves receiving the 3 inactivated vaccines. Vaccination with MLV induced the highest IgG2/IgG1 ratio. This difference was small, and only significant relative to 2 of the inactivated vaccine groups, which were not significantly different from each other. The higher proportion of IgG2 isotype in the MLV sera was not associated with lower ADCMC, a function not attributed to this isotype. The VN and FI titers, but not the ELISA value of the sera, were most predictive of ADCMC. The inactivation processes apparently alter epitopes and affect the induction of functional antibodies.     

Protective efficacy of H5 inactivated vaccines in meat turkey poults after challenge with Egyptian variant highly pathogenic avian influenza H5N1 virus

In contrast to chickens, there is a paucity of information on the potency of H5 vaccines to protect turkeys against the highly pathogenic avian influenza (HPAI) H5N1 virus infections. In this study, 4 groups, 10 turkey poults each, were vaccinated at seven days old with one of H5N2 or H5N1 commercial vaccines or one of two prepared H5N1 vaccines from a local Egyptian variant HPAI H5N1 (EGYvar/H5N1) strain. At 35 days age, all vaccinated and 10 non vaccinated birds were challenged intranasal with 10(6) EID(50)/0.1 ml of EGYvar/H5N1. All vaccines used in this study were immunogenic in turkeys. There was no cross reaction between the commercial vaccines and the Egyptian variant H5N1 antigen as obtained by the hemagglutination inhibition test. Birds vaccinated with H5N2 vaccine were died, while other H5N1 vaccinated groups have had 20-40% mortality. The highest virus excretion was found in non-vaccinated infected and H5N2 vaccinated birds. Eleven peculiar amino acid substitutions in H5 protein of the variant strain were existed neither in the vaccine strains nor in the earliest H5N1 virus introduced into Egypt in 2006. In conclusion, single vaccination at seven days old is inadequate for protection of meat turkeys against variant HPAI H5N1 challenge and multi-dose vaccination at older age is recommended. For the foreseeable future, continuous evaluation of the current vaccines in H5N1 endemic countries in the face of virus evolution is a paramount challenge to mitigate the socio-economic impact of the virus.     

Efficacy of swine influenza A virus vaccines against an H3N2 virus variant

We compared the efficacy of 3 commercial vaccines against swine influenza A virus (SIV) and an experimental homologous vaccine in young pigs that were subsequently challenged with a variant H3N2 SIV, A/Swine/Colorado/00294/2004, selected from a repository of serologically and genetically characterized H3N2 SIV isolates obtained from recent cases of swine respiratory disease. The experimental vaccine was prepared from the challenge virus. Four groups of 8 pigs each were vaccinated intramuscularly at both 4 and 6 wk of age with commercial or homologous vaccine. Two weeks after the 2nd vaccination, those 32 pigs and 8 nonvaccinated pigs were inoculated with the challenge virus by the deep intranasal route. Another 4 pigs served as nonvaccinated, nonchallenged controls. The serum antibody responses differed markedly between groups. After the 1st vaccination, the recipients of the homologous vaccine had hemagglutination inhibition (HI) titers of 1:640 to 1:2560 against the challenge (homologous) virus. In contrast, even after 2nd vaccination, the commercial-vaccine recipients had low titers or no detectable antibody against the challenge (heterologous) virus. After the 2nd vaccination, all the groups had high titers of antibody to the reference H3N2 virus A/Swine/Texas/4199-2/98. Vaccination reduced clinical signs and lung lesion scores; however, virus was isolated 1 to 5 d after challenge from the nasal swabs of most of the pigs vaccinated with a commercial product but from none of the pigs vaccinated with the experimental product. The efficacy of the commercial vaccines may need to be improved to provide sufficient protection against emerging H3N2 variants.     

Immunosuppression and histopathological changes in the bursa of fabricius associated with infectious bursal disease vaccination in chicken

The effect of the infectious bursal disease (IBD) live virus vaccine on the immune response of chicken was evaluated by the assessment of antibody response following vaccination as well as resistance to challenge with virulent virus. Birds were vaccinated at various ages and later challenged with a heterologous vaccine (NDV) or wild-type IBD virus. The BF was examined for histological changes at regular intervals. Antibody levels to NDV were monitored.

Significantly higher mortality rates were observed in birds vaccinated with IBD vaccine than unvaccinated birds (P < 0.01) following challenge, BF from vaccinated birds showed marked lymphocyte depletion and cellular infiltration with mononuclear cells.

Intraocular NDV (NDV-i/o) vaccine given at day old largely prevented the immunodepressive effect of IBD vaccination on NDV vaccine. Groups that received IBD vaccine on day 14 but no NDV i/o suffered higher mortality (41.2%) and showed lower antibody response than those vaccinated on day 1 (0%) or controls which did not receive IBDV (11.8%).     

三个厂家猪瘟活疫苗免疫效果比较试验

在同一条件下对3个厂家生产的猪瘟细胞源活疫苗进行了免疫效果评价试验,并与猪瘟传代细胞苗免疫效果进行比较。结果发现3个厂家生产的猪瘟细胞源活疫苗存在一定差异,2个厂家的免疫效果较好,1个厂家的免疫效果较差。猪瘟传代细胞苗免疫效果优于猪瘟细胞源活疫苗,猪瘟传代细胞苗免疫1次,抗体合格率高,持续时间长,猪瘟细胞源活疫苗要免疫两次才能获得比较好的效果。同时,我们发现高致病性猪蓝耳病活疫苗（JXA1-R株）对猪瘟抗体产生有一定影响。     

Demonstration in live chickens of the carrier state in infectious laryngotracheitis

Two separate groups of nine-week-old specific pathogen free cockerels maintained in isolation were infected with a field strain of infectious laryngotracheitis (ILT) virus, either by intratracheal or combined intranasal and supraconjunctival inoculation. Birds were monitored for virus shedding from five sites on alternate days during the acute phase and three times weekly until week 17. They were then treated with cyclophosphamide on three consecutive days and thereafter swabbed daily. During the acute phase clinical signs were observed and virus was recovered from ocular and nasal sites for up to six to eight days. Initially after the acute phase no virus could be detected. However, from the seventh week after infection intermittent, apparently spontaneous shedding was detected in four of five birds in each group. There was no clear effect of cyclophosphamide treatment on re-excretion patterns, possibly because of the high levels of virus shedding already occurring. Thus, a carrier state for ILT virus has been demonstrated experimentally in live clinically recovered birds.