Experimental Risk Assessment of Recombinant Newcastle Disease Viruses Used as Vaccines

Patti Miller from the USDA ARS Southeast Poultry Research Laboratory reported that recombinant Newcastle Disease Viruses (NDV) used as live vaccines were assessed for:

1. the potential for recombinant NDV-vectored vaccines (rNDV) containing the Avian Influenza virus (AIV) H5 gene to recombine with low pathogenicity H5, H6 and H9 AIV strains, and originate a virus with increased virulence
2. the risk of rNDV vaccines, containing an attenuated fusion (F) protein cleavage site, to revert back to a virulent virus phenotype, and
3. to assess the ability of rNDV vaccines to infect and spread in non-target species.

Co-infection of 14-day-old embryonating chicken eggs (ECE) with rNDV-LaSota/AI-H5 and LPAI strains, rA/whooper/Mongolia/2005 (H5), A/duck/Penn/97 (H6) or A/ruddy_turnstone/NJ/02 (H9) resulted in the death of 90 per cent of the embryos within 72 hours post-inoculation (PI).

Infection of 14-day-old ECEs with wild type NDV strains (LaSota and Australia) or with rNDV strains [LaSota (rLaSota) or ZJ1 (rZJ1*Lento)] (900 eggs/virus) resulted in the death of 65 (7.2 per cent), 10 (1.1 per cent), 23 (2.5 per cent) and six (0.6 per cent) embryos, respectively, within 72 hours post-inoculation.

Sequencing of the F gene cleavage site, under these experimental conditions, revealed all rNDV are stable without reversion to a virulent phenotype. Experiments to assess the ability of rNDV to infect and spread in pigeons and sparrows have shown that rLaSota, and rLaSota/AIV-H5 are able to infect and spread in these species. All virus-inoculated pigeons shed the virus in oral and/or cloacal secretions and at least one out of four contact birds shed the virus for two or more days.

Miller and colleagues concluded their findings suggest that the system is suitable to assess the risk associated with recombinant NDV vaccines.

Wide-Range of Protection Achieved by Vaccination with a Recombinant HVT-NDV Vaccine against Different Genotypes of Newcastle Disease Virus

Newcastle Disease (ND) is one of the major diseases causing great losses for the poultry industry, according to Vilmos Palya from Ceva Santé Animale in Hungary. In the past decades, there has been a major shift in the genotypes of NDV strains that have been identified as prevalent in poultry.

Vaccination plays an important role in the prevention of the disease. The aim of this study was to test the efficacy of a recombinant turkey herpesvirus (HVT) vaccine (Vectormune® ND, Ceva) expressing the F-gene of a genotype I NDV strain against velogenic NDV strains representing several genotypes from different geographical areas of the world.

Day-old SPF chicks had been vaccinated with the recombinant vaccine and challenged at four weeks of age with one of the eight velogenic NDV strains as follows:

1. Herts 33/56 reference strain (genotype IV)
2. a recent isolate from Malaysia (genotype VII)
3. a field isolate from Mexico (genotype V)
4. a genetically unusual Peruvian isolate (genotype VIId)
5. an isolate from the Philippines belonging to the genotype causing epizootics in Asia (genotype VIIa)
6. a South-African strain RB Daagstan ND/01/ZA (genotype VIII)
7. a recent Chinese isolate (genotype VIId) and
8. a pneumo-neurotropic isolate from Saudi-Arabia (genotype VIId).

Evaluation of protection was performed on the basis of absence of clinical signs and reduction of challenge virus shedding.

Vaccination with this recombinant vaccine prevented clinical signs and reduced significantly both oro-nasal and cloacal virus shedding of all challenge virus strains used. Dr Palya presented detailed results at the meeting.

Induction of Local Respiratory Immunity Following Vaccination of Chickens with an HVT-NDV-Recombinant

New generation herpes virus of turkeys (HVT)-based vaccines provide an interesting alternative to currently used live vaccines due to their increased safety and the possibility of early application in the face of maternal immunity, according to Dr Silke Rautenschlein of the University of Veterinary Medicine in Hanover, Germany.

For protection against NDV, induction of local immune reactions would be desirable to interfere with virus replication at the site entry. However, not much is known about the ability of HVT-NDV vaccines to induce local respiratory immunity, she said.

The objective of their study was to compare the induction of local immune reactions and protection against challenge between an HVT-NDV and a live NDV vaccine as well as the combination of both in chickens.

None of the vaccines induced any clinical signs or lesions.

As expected, the live NDV-vaccine alone induced a specific local and systemic humoral immune response as well as up-regulation of T-cell sub-populations in the upper respiratory tract.

Interestingly, the HVT-NDV vaccine induced humoral as well as cell-mediated immune reactions locally in the respiratory tract, and these local reactions were enhanced if the recombinant was combined with the live vaccine strain.

Dr Rautenschlein concluded the study provides circumstantial evidence for local protective mechanisms induced by the HVT-NDV vaccine, which is supported by the fast clearance of a lentogenic NDV challenge virus in the vaccinated groups.

Virulent Newcastle Disease Virus in Tanzania

Virulent Newcastle Disease virus (vNDV) is endemic in Africa and is a major cause of mortality of poultry in many countries. David L. Suarez of the USDA ARS Southeast Poultry Research Laboratory explained that, although it is one of the main disease threats in the East Africa, little work has been done to characterise the viruses circulating in the region.

As part of a surveillance project of live bird markets in Tanzania, cloacal and oropharyngeal swabs were taken from apparently healthy birds for virus isolation.

From these samples, a number of virulent Newcastle disease viruses were isolated and characterised.

Full genome sequencing showed that all the viruses clustered together into a potential new genotype of the Class II classification scheme, although considerable variation - up to six per cent - was observed between viruses in the country.

Representative isolates were further characterised in vivo to determine the intracerebral pathogenicity index.

Because of the genetic differences of these viruses to the common vaccine strains, a vaccine study was also performed with both LaSota and I2 vaccine. The I2 vaccine is a thermostable vaccine that is also widely available and used in Tanzania.

Results of the vaccination study were pending at the time of the meeting.

However, Dr Suarez reported that the characterisation of viruses from this surveillance project provides the first complete genome sequence of viruses from this region and provides information on vaccine protection using locally available vaccine strains. He added that the characterisation of these viruses will provide key information for the better control of the virus in the region.

Evaluation of a Turkey Specific Real Time-PCR to Measure Newcastle HVT Vaccine Replication When Applied in-ovo or Subcutaneously

The Embrex Inovoject® system has been used for more than 20 years around the globe, primarily in broiler chickens, reported Steven Clark of Zoetis. Recently, he said, devices have been introduced into turkey hatcheries in Brazil and US. As well, vectored ND vaccines have recently been trialed by the turkey industry at the hatchery.

Hatchery administration may provide earlier protection than traditional in-field application; in-ovo administration may provide even earlier immunity than traditional day-of-hatch. Vectored ND vaccines - either pox or HVT - may provide long-term protection following only one administration.

Clark and colleagues developed a diagnostic tool specific for turkeys, which allows for the quantification of the replication rate of HVT in turkeys vaccinated with such vaccines. They based the new tool on the previously developed chicken specific HVT quantitative PCR (qPCR) commonly used by their company's Diagnostic Services team to test field samples.

Specific primers were developed for the turkey iron binding protein gene (TRFE, ovotransferrin precursor gene) using GenBank available sequences. The target product was amplified from turkey DNA sample, cloned and sequenced to confirm the presence of the TRFE gene fragment. The qPCR conditions were optimised for dual target reaction, allowing for comparison of HVT replication rates across samples.

Results from field samples comparing HVT replication in turkeys vaccinated in-ovo or subcutaneously were discussed by Dr Clark at the meeting.

Atypical Turkey Newcastle Disease in the Upper Midwest. Part I. Field Experiences

Mild or lentogenic strains of APMV-1 (Newcastle Disease virus) are capable of causing significant respiratory disease in susceptible turkey populations, reported Andre F. Ziegler of the University of Minnesota. Between 2008 and 2012, a lentogenic APMV-1 virus infection was diagnosed in 150 commercial turkey cases in the upper Midwest. Initially, starting in a defined geographic area, this disease was eventually identified in multiple areas in the upper Midwest.

Protective vaccination regimens varied, he said, but predominating protocols were seemingly ineffective in preventing respiratory disease and mortality in affected turkey poults. Cumulative mortality associated with respiratory disease in these cases was as high as 50 per cent but more typically 10 to 20 per cent of affected flocks.

Multiple co-pathogens were typically associated with flocks exhibiting severe disease. Sequencing of the viral genome, performed at the USDA-NVSL, identified two lineages of atypical APMV-1 agents over this time, Dr Ziegler added.

Atypical Turkey Newcastle Disease in the Upper Midwest. Part II. Virus Characterisation and Vaccinal Protection

In a related presentation, Mary Lea Killian of the National Veterinary Services Laboratories in Ames, Iowa, also reported that since 2008, atypical APMV-1 viruses have been circulating in turkey flocks in the Upper Midwest region of the United States.

Characterisation of the viruses by OIE standard methods classifies these atypical viruses as low virulent (ICPI<0.7). The isolates do not have the multiple basic amino acid motif to be characterised as virulent NDV but they do have phenylalanine at position 117, a characteristic normally found only in virulent NDV.

Analysis of the amino acid sequence of the fusion protein indicates the viruses are more closely related to wild bird APMV-1 viruses than virulent NDV viruses.

Commercial flocks infected with these viruses show increased mortality, however, co-infection with other bacterial respiratory pathogens is common. In addition, commercial flocks are vaccinated with one or more commercial vaccine products.

Molecular and biological characterisation of the viruses were discussed by Dr Killian, as were vaccine protection studies performed with different vaccine combinations.

November 2013