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Environmental Assessment for the Use of Marek's Disease-Newcastle Disease Vaccine, Serotypes 2 and 3, Live Virus, Live Marek's Disease Vector

For Public Release

August 9, 2010

Prepared by:

Canadian Centre for Veterinary Biologics
Terrestrial Animal Health Division
Canadian Food Inspection Agency

The information in this environmental assessment was current at the time of its preparation. It is possible that the situation may have changed since that time. If you have any questions, please consult the Canadian Centre for Veterinary Biologics.


Table of Contents

  • Summary
  • 1. Introduction
    • 1.1 Proposed Action
    • 1.2 Background
  • 2. Purpose and Need for Proposed Action
    • 2.1 Significance
    • 2.2 Rationale
  • 3. Alternatives
  • 4. Molecular and Biological Characteristics of Parental and Recombinant Organisms
    • 4.1 Identification, Sources, and Strains of Parental Organisms
    • 4.2 Source, Description and Function of Foreign Genetic Material
    • 4.3 Method of Accomplishing Genetic Modification
    • 4.4 Genetic and Phenotypic Stability of the Vaccine Organism
    • 4.5 Horizontal Gene Transfer and Potential for Recombination
    • 4.6 Host Range / Specificity, Tissue Tropism and Shed / Spread Capabilities
    • 4.7 Comparison of the Modified Organisms to Parental Properties
    • 4.8 Route of Administration / Transmission
  • 5. Human Safety
    • 5.1 Previous Safe Use
    • 5.2 Probability of Human Exposure
    • 5.3 Possible Outcomes of Human Exposure
    • 5.4 Pathogenicity of Parent Micro-Organisms in Humans
    • 5.5 Effect of Gene Manipulation on Pathogenicity in Humans
    • 5.6 Risk Associated with Widespread Use of the Vaccine
  • 6. Animal Safety
    • 6.1 Previous Safe Use
    • 6.2 Fate of the Vaccine in Target and Non-Target Species
    • 6.3 Potential of Shed and/or Spread from Vaccinate to Contact Target and Non-Target Animals
    • 6.4 Acquisition to Virulence Resulting from Back Passage in Animals
    • 6.5 Effect of Overdose in Target and Potential Non-Target Species
    • 6.6 The Extent of the Host Range and the Degree of Mobility of the Vector
  • 7. Affected Environment
    • 7.1 Extent of Release Into the Environment
    • 7.2 Persistence of the Vector in the Environment / Cumulative Impacts
    • 7.3 Extent of Exposure to Non-Target Species
    • 7.4 Behaviour of Parent Micro-Organisms and Vector in Non-Target Species
  • 8. Environmental Consequences
    • 8.1 Risks and Benefits
    • 8.2 Relative Safety Compared to Other Vaccines
  • 9. Mitigative Measures
    • 9.1 Worker Safety
    • 9.2 Handling Vaccinated or Exposed Animals
  • 10. Monitoring
    • 10.1 General
    • 10.2 Human
    • 10.3 Animal
  • 11. Consultations and Contacts
  • 12. Conclusions and Actions
  • 13. References

Summary

Marek's Disease-Newcastle Disease Vaccine, Serotypes 2 and 3, Live Virus, Live Marek's Disease Vector consists of a live chicken herpesvirus (Marek's Disease Serotype 2) as well as a live turkey herpesvirus modified by the introduction of a gene sequence from the Newcastle disease virus. This vaccine is indicated for use in ovo at 18 days of embryonation for the prevention of very virulent Marek's disease and Newcastle disease in chickens.

The vaccine was evaluated by the Canadian Centre for Veterinary Biologics (CCVB) of the Canadian Food Inspection Agency (CFIA). As part of the requirements for licensing the product in Canada, an environmental assessment was conducted. This environmental assessment is a public document which contains information on the molecular and biological characteristics of the live recombinant organism, target animal and non-target animal safety, human safety, environmental considerations and risk mitigation measures.

This environmental assessment is based on information provided by the manufacturer (Intervet, Inc., Millsboro, Delaware, U.S. Veterinary Biologics Establishment License No. 165A), as well as information independently obtained by the CCVB reviewer.

1. Introduction

1.1 Proposed Action

The Canadian Centre for Veterinary Biologics (CCVB), Terrestrial Animal Health Division, Canadian Food Inspection Agency (CFIA) is responsible for regulating the use of veterinary biologics in Canada under the legal authority provided under the Health of Animals Act and Regulations.

Any veterinary biologic manufactured, sold or represented for use in Canada must comply with the requirements specified by the CFIA regarding the safety, purity, potency, and efficacy of the product. Intervet Inc. (Millsboro, Delaware, U.S.) through Intervet Canada Corp. (Kirkland, Que.) has submitted the following vaccine for licensing in Canada:

  • Marek's Disease-Newcastle Disease Vaccine, Serotypes 2 and 3, Live Virus, Live Marek's Disease Vector (Trade name: INNOVAX-ND-SB), USDA Product Code 17H1.R2, CCVB File 800VV/M10.0/I6.2

This environmental assessment was prepared by CCVB as part of the overall assessment for licensing the above vaccine in Canada.

1.2 Background

Marek's Disease-Newcastle Disease Vaccine, Serotypes 2 and 3, Live Virus, Live Marek's Disease Vector is manufactured by Intervet, Inc., Millsboro, Delaware (U.S. Veterinary Biologics Establishment License No. 165A), and is currently licensed for sale in the U.S. This recombinant vaccine consists of a live chicken herpesvirus (Marek's disease serotype 2), as well as a live turkey herpesvirus modified by the insertion of genetic material from the Newcastle disease virus. The vaccine is a stand-alone product. CCVB has previously licensed a different vaccine based on a recombinant turkey herpesvirus construct containing portions of another virus for use in chickens.

Newcastle Disease (ND) is a highly contagious avian viral disease present in many parts of the world. Pathogenicity of the virus varies with the strain, from asymptomatic infection with lentogenic strains to 100 percent mortality in velogenic viscerotropic Newcastle disease strains (vvND or Exotic Newcastle Disease) (King, 2006).

Marek's disease (MD) is a viral oncogenic (neoplastic) disease of poultry found worldwide. It is considered to be ubiquitous and is presumed present in all flocks except those maintained under stringent pathogen-free conditions (Powell, 1986). The virus is extremely difficult to eradicate from flocks for several reasons. The virus spreads very quickly between individuals (whether vaccinated or not), who become infected for long periods of time, shedding the organism into the environment where it can persist for months (Fadly, 2006).

2. Purpose and Need for Proposed Action

2.1 Significance

The label indication for INNOVAX-ND-SB Marek's Disease-Newcastle Disease Vaccine is for the vaccination of 18-day-old chicken embryos as an aid in the prevention of very virulent MD and ND.

2.2 Rationale

The CCVB evaluates veterinary biologic product submissions for licensure under the Health of Animals Act and Regulations. The general criteria for licensing are as follows: a) the product must be pure, safe, potent and efficacious; b) the product must be licensed in the country of origin; c) vaccine components must be relevant to Canadian disease conditions; and d) the product must be produced and tested in accordance with generally accepted "good manufacturing practices." This U.S. origin vaccine meets these general criteria and presented no unacceptable importation risk, and therefore was evaluated for licensing by CCVB.

Since this product is a biotechnology-derived vaccine of Class II category, CCVB is required to perform an environmental assessment before its potential release into the environment. This environmental assessment has been conducted to determine whether the release of this product presents safety risks to animals, public health or the environment.

3. Alternatives

The two alternative options being considered are as follows: a) to issue a Permit to Import Veterinary Biologics to Intervet Canada Corp. for the importation of INNOVAX-ND-SB Marek's Disease-Newcastle Disease Vaccine, Serotypes 2 and 3, Live Virus, Live Marek's Disease Vector if all licensing requirements are satisfactory; or b) not to issue a Permit to Import Veterinary Biologics if licensing requirements are not met.

4. Molecular and Biological Characteristics of Parental and Recombinant Organisms

4.1 Identification, Sources, and Strains of Parental Organisms

The proposed vaccine qualifies as a Class II Veterinary Biologic (live expression vectors that carry one or more foreign genes that code for immunizing antigens and/or immune stimulants). The recombinant vaccine construct consists of a protein-expressing gene sequence from Newcastle disease virus (NDV) inserted into an avirulent herpesvirus of turkeys (HVT) viral vector. A live non-oncogenic Marek's disease virus, serotype 2 (MDV-2) is also included in the vaccine.

4.2 Source, Description and Function of Foreign Genetic Material

A gene sequence from NDV was selected from those likely to stimulate protective immunity in chickens. Details of the actual sequence are on file at CCVB.

4.3 Method of Accomplishing Genetic Modification

Details of the methods used in the construction of the recombinant organism are on file at CCVB. The master seed virus was tested for extraneous agents, purity and safety according to tests described in the U.S. Code of Federal Regulations (9 CFR). These data have been reviewed and are on file at CCVB.

4.4 Genetic and Phenotypic Stability of the Vaccine Organism

In vitro testing using Southern blot analysis revealed genetic stability of the recombinant organism through five passages in a cell line. Immunofluorescence assay and a Southern blot were also used to determine the phenotypic and the genotypic stability of the recombinant organism after in vivo testing. No gross genetic or phenotypic alterations were detected after five back passages in the target species, the chicken.

4.5 Horizontal Gene Transfer and Potential for Recombination

This vaccine combines the recombinant organism with another live herpesvirus, MDV-2, which shows significant homology in its genome to HVT (Alfonso et al., 2000; Fukuchi et al., 1984; Gibbs et al., 1984). There exists a theoretical low risk of gene transfer and recombination between the two viruses, as reports can be found in the literature of experimental recombination between herpesviruses within co-infected cells in vitro as well as in vivo. Overall, the risk of recombination is considered to be no higher for this product than for other multivalent vaccines.

Virulence is one characteristic of a virus that may change (increase or decrease) if recombination occurs. Both herpesvirus strains in the vaccine are considered avirulent, but there exists a theoretical low risk of a novel recombinant organism being produced whose virulence is increased, as seen in studies with other herpesviruses (Henderson et al., 1990; Katz et al., 1990). It must be noted that this MDV-2 strain and a different parental strain of HVT have been combined in existing multivalent vaccines which have been used for years with no indication of acquiring virulence. The risk of acquisition of virulence of either organism via recombination is considered to be very low for this product.

4.6 Host Range / Specificity, Tissue Tropism and Shed / Spread Capabilities

In vivo studies conducted by the manufacturer determined the host range, tissue tropism, and shed / spread capabilities of the recombinant organism. Replication of the recombinant organism was detected in chickens and turkeys only, and the virus did not replicate in other avian species such as pigeons and ducks. Dissemination of the recombinant organism in vivo was also shown to be no different from that of the parental HVT. No transmission of the recombinant organism was seen to occur from vaccinated chickens to other in-contact, unvaccinated chickens or ducks. It did spread to in-contact turkeys, however, which would be expected due to the fact that turkeys are a host species of the parental virus.

Since there have been reports of NDV causing mild conjunctivitis in mammals and humans, another study was performed which showed that the recombinant organism did not replicate nor cause any clinical signs when used to inoculate rabbits via the intraocular route.

4.7 Comparison of the Modified Organisms to Parental Properties

The host range, tissue tropism, and shed/spread capabilities of the recombinant organism are expected to be similar to the parental HVT vaccine strain.

4.8 Route of Administration / Transmission

Avian herpesviruses are mainly spread in shed feather dander, but wild-type HVT has been shown to not spread readily between chickens infected early in life (Cho and Kenzy, 1975). Like its parental organism, the recombinant organism did not spread from vaccinated chickens to in-contact unvaccinated chickens, and though wild-type HVT is known to persist in the environment, in vitro studies performed by the manufacturer showed environmental persistence of the recombinant organism as a cell-associated virus to be restricted to less than seven days.

5. Human Safety

5.1 Previous Safe Use

Though the recombinant organism has not been used previously in Canada, it has been licensed for use in the U.S. since 2007. Parental HVT has been safely used for decades in vaccines to control MD. The NDV gene insert is not expected to be a safety concern since it consists only of a protein-expressing gene sequence of the complete virus. Diluents and preservatives in this product have all been used previously in other products with no safety concerns.

5.2 Probability of Human Exposure

Human exposure to the vaccine itself (as a cell-associated virus) is likely to be limited to veterinarians, poultry operators and animal technicians during vaccination. Consistent with what is observed with other recombinant HVT vaccines, the organism may be shed from vaccinated animals to a minimal degree; thus exposure of these personnel to mature virus in shed feather follicle epithelium is also possible.

Since literature suggests that HVT may replicate in these avian species for extended and possibly indefinite periods, as it is a herpesvirus which is capable of persistent infection (Calnek and Witter, 1991; Cho 1974), there is a chance that individuals working in abattoirs will also be exposed to the recombinant virus.

Exposure to humans through the consumption of meat from vaccinated birds will be reduced by the fact that the recombinant HVT + NDV virus is localized to lymphocytes associated with visceral organs and feather follicles, and not the tissues humans predominantly consume as meat. Moreover, even if trace amounts of the recombinant virus were present in chicken meat, studies have shown that the vast majority of ingested nucleic acid is efficiently degraded in the human digestive tract (Jonas et al., 2001).

Human exposure is thus possible, though it is not expected to have any adverse health effects.

5.3 Possible Outcomes of Human Exposure

There are no reports of HVT causing infection in any species other than chickens and turkeys. Even in these species, the virus may replicate, but it does not cause clinical disease. Though Newcastle disease is considered zoonotic, causing transient conjunctivitis in exposed humans, the NDV component in this vaccine consists of a gene insert, which is unlikely to cause any adverse health effects in exposed humans. Exposure, including inadvertent injection, of humans to this vaccine is expected to cause no serious adverse effects.

5.4 Pathogenicity of Parent Micro-Organisms in Humans

As above, the parental HVT organism is non-pathogenic to humans, and while NDV is mildly zoonotic, this vaccine includes an NDV gene coding for one protein, and is unlikely to be pathogenic to humans.

5.5 Effect of Gene Manipulation on Pathogenicity in Humans

The NDV gene insert is not expected to result in any change in pathogenicity of the HVT backbone.

5.6 Risk Associated with Widespread use of the Vaccine

No significant public heath issues are expected to result from widespread use of the vaccine.

6. Animal Safety

6.1 Previous Safe Use

The recombinant organism was found to be safe in experimental studies in chickens (label-indicated target species) at vaccinal and 10-times vaccinal doses, as well as in several non-target species, both avian (turkey, pigeon, duck) and mammalian (rabbit).

INNOVAX-ND-SB was administered to over three million chickens in the field safety trials conducted by the manufacturer, and the vaccine has been licensed in the U.S. since 2007. Parental HVT has been safely used for decades in vaccines to control MD in chickens.

6.2 Fate of the Vaccine in Target and Non-Target Species

There are no reports of wild-type HVT causing clinical disease in any species. Back passage studies in chickens indicate that the recombinant organism is unlikely to acquire virulence.

Studies conducted in target and non-target species demonstrate that the host range and tropism of the recombinant organism vaccine strain were not altered from the parental HVT strain. These studies also showed that the recombinant organism can persist for 21 days (end of experimental period) in chickens and turkeys, and literature suggests that it may replicate in these species for extended and possibly indefinite periods, as it is a herpesvirus vector which is capable of persistent infection (Calnek and Witter, 1991; Cho 1974).

No studies have been conducted to investigate the long-term safety of this product in the target species (chickens) or in birds of non-target yet host species (turkeys).

6.3 Potential of Shed and/or Spread from Vaccinate to Contact Target and Non-Target Animals

In the literature, HVT has been shown to be shed sporadically in the feather dander of vaccinated chickens (Zygraich and Huygelan, 1972; Cho, 1974). In manufacturer's studies, the recombinant organism did not spread to any in-contact avian species other than turkeys, the host species of the parental organism.

6.4 Acquisition to Virulence Resulting from Back Passage in Animals

Testing indicated that the master seed virus is genetically and phenotypically stable up to passage MSV+5, and is free of extraneous agents as per 9 Code of Federal Regulations 113.300. Backpassage studies in the target species, chickens, have not shown any acquisition of virulence of the recombinant organism.

6.5 Effect of Overdose in Target and Potential Non-Target Species

Manufacturer's studies reported no gross lesions or detrimental effects on hatchability/survivability in chickens injected (in ovo) with 10 times the vaccinal dose.

6.6 The Extent of the Host Range and the Degree of Mobility of the Vector

HVT has a very narrow host range, restricted to turkeys and chickens. The recombinant organism is shed in a manner similar to its parental organism, and while it is not horizontally transmitted between chickens, it can spread from vaccinated chickens to in-contact turkeys.

7. Affected Environment

7.1 Extent of Release into the Environment

The recombinant organism is shed in feather dander of vaccinated chickens. The vast majority of vaccinated chickens will be housed indoors in biosecure facilities, and thus will have little direct exposure to the environment. However, limited release of the vaccine organism may occur when barns are cleaned out, or through the vented air. HVT-contaminated litter and air (dust) appear to be capable of infecting turkeys (Witter and Solomon, 1971).

Potential for occasional limited environmental release through accidental spills, unintended syringe aerosols, or contamination of the vaccination site also exists during routine use of this vaccine.

7.2 Persistence of the Vector in the Environment / Cumulative Impacts

Studies have shown that HVT virus may be recovered from the environment of vaccinated chicken flocks for over eight weeks post-vaccination and possibly longer (Islam and Walkden-Brown, 2007). MD virus shed in chicken feather follicles has been shown to persist in the environment for eight to 12 months (Schat, 1985).

Manufacturer's studies have shown the recombinant organism (as a cell-associated virus) to persist in the environment for no more than seven days. No studies have been conducted to demonstrate the duration (past four weeks) of mature vaccine virus shed in the feather follicles, or the stability/persistence of the virus in the natural environment (e.g. when shed in feather follicle epithelial cells, presence in barn dust and litter, in outdoor manure piles after cleaning of barns, under conditions of sun exposure, etc.).

Herpesviruses are typically inactivated by UV light from the sun (Lytle and Sagripanti, 2005). However, MD viruses in dried feathers and poultry dust have been reported to remain infectious for up to a year (Jurajda and Klimes, 1970; Schat, 1985).

7.3 Extent of Exposure to Non-Target Species

The very limited host range of herpesviruses reduces the risk of spread to non-target mammalian species. Turkeys kept in proximity to chicken or turkey flocks vaccinated with this organism are at possible risk of exposure if they are in contact with bedding or exhaust from the chickens' area, or with the chickens themselves.

Even though studies performed by the manufacturer indicate that the recombinant vaccine virus is non-pathogenic to turkeys, precautionary measures should be followed to reduce the potential for spread of the virus to turkey populations. For this reason, the manufacturer has agreed to add a special warning to the labelling of INNOVAX-ND-SB indicating that the product should not be administered to chickens with probable direct or indirect exposure to turkeys.

7.4 Behaviour of Parent Micro-Organisms and Vector in Non-Target Species

A study conducted by the manufacturer has suggested that the recombinant organism is safe in non-target species, including turkeys, when administered directly. No practical method exists to determine with certainty the behaviour of the organism over a long period of time, should it become established in a natural turkey population.

8. Environmental Consequences

8.1 Risks and Benefits

The vaccine appears to be effective in inducing immunity even when administered to maternal antibody-positive chickens. Overcoming maternal antibody immunity has been a challenge for many of the currently available ND vaccines. In addition, the recombinant HVT + ND virus does not appear to cause the side effects associated with some of the very immunogenic yet virulent live ND vaccines.

The primary risk identified for INNOVAX-ND-SB, as with any vaccine, can be attributed to potential adverse reactions. Manufacturer's safety studies have demonstrated the product to be generally safe in field safety studies conducted in chickens.

An additional, yet theoretical, risk relates to its potential to spread to turkey populations (and potentially other Galliformes) and consequently persist in the environment. It is important to note, however, that no pathogenicity in turkeys is expected for the recombinant HVT + ND virus, even if it does spread to turkeys. In an experiment performed by the manufacturer, turkeys infected with recombinant HVT + ND virus (through either direct inoculation or contact with vaccinated chickens) appeared healthy throughout the duration of the study (up to approximately four weeks of age), akin to turkeys infected with wild-type HVT. The Canadian labelling for INNOVAX-ND-SB also carries a precautionary statement that measures should be taken to avoid contact between vaccinated chickens and turkeys, which should further mitigate the proposed risk of spread to turkeys.

In conclusion, there is a hypothetical risk that turkeys might inadvertently become infected with the vaccine virus, but since there is no evidence to indicate that such an occurrence would be detrimental to turkeys, the benefits of the vaccine in promoting chicken animal health are believed to outweigh the proposed risk of vaccine virus spread to turkeys.

8.2 Relative Safety Compared to other Vaccines

The vaccine is comprised of two avirulent live MD viruses and a gene sequence of ND virus, all of which are non-pathogenic. The vaccine has been shown to be as safe as conventional MD vaccines in target and non-target species. The lack of potential for virulence and the absence of adjuvants typical of killed virus vaccines are additional positive safety features of this vaccine.

9. Mitigative Measures

9.1 Worker Safety

Veterinarians, poultry operators, and animal technicians could be exposed to the live recombinant organism during vaccination. As was discussed in Section 5 above on human safety, such exposure is not expected to be a safety concern. The in ovo route of vaccine administration proposed for INNOVAX-ND-SB should help reduce the incidence of accidental vaccinator self-injection compared to live bird vaccination. Moreover, since the vaccine does not contain any adjuvant, the risk of clinical problems due to accidental self-injection of adjuvant (oil) is removed. Nonetheless, measures should be taken to protect personnel from exposure, as stated on the product insert.

9.2 Handling Vaccinated or Exposed Animals

Since chicks reared in a biosecure facility are not often handled directly by humans, and poultry workers typically employ precautionary biosafety measures, exposure through handling vaccinated chicks is not expected to be great. However, poultry workers could become exposed to the vaccine virus through dust and air inside barns that might be contaminated with virus shed through feather dander. Again, the recombinant virus is not believed to be pathogenic to humans.

10. Monitoring

10.1 General

The vaccine licensing regulations in Canada require manufacturers to report all serious, significant adverse reactions to the CFIA within 15 days of receiving notice from an owner or a veterinarian. Veterinarians may also report suspected adverse reactions directly to the CFIA. If an adverse reaction complaint is received by CCVB, the manufacturer is asked to investigate and prepare a report for the owner's veterinarian and the CFIA. If the problem is resolved to the satisfaction of the veterinarian / client, no further action is usually requested by CCVB. However, if the outcome of the investigation is not satisfactory, CCVB may initiate regulatory action depending on the case, which may include further safety testing, temporarily stopping sales of the product, or product withdrawal from the market.

10.2 Human

No monitoring of the human safety of the product will be carried out.

10.3 Animal

Veterinarians should report any suspected adverse reactions to CCVB as indicated above. Suspected adverse reactions should be reported using Form CFIA/ACIA 2205 – Notification of Suspected Adverse Events to Veterinary Biologics.

11. Consultations and Contacts

Importer

Intervet Canada Corp.
16750, route Transcanadienne
Kirkland, QC H9H 4M7

Manufacturer

Intervet, Inc.
29160 Intervet Lane, Box 318
Millsboro, Delaware USA 19966-0318

12. Conclusions and Actions

Based on our assessment of the available information, CCVB has concluded that the importation and use of Marek's Disease-Newcastle Disease Vaccine, Serotypes 2 and 3, Live Virus, Live Marek's Disease Vector in Canada would not be expected to have any significant adverse effect on the environment, when manufactured and tested as described in the approved Outline of Production, and used according to label directions.

Following this assessment, and the completion of the Canadian veterinary biologics licensing process, the Permit to Import Veterinary Biologics of Intervet Canada Corp. may be amended to allow the importation and distribution of the following product in Canada:

  • Marek's Disease-Newcastle Disease Vaccine, Serotypes 2 and 3, Live Virus, Live Marek's Disease Vector (INNOVAX-ND-SB), USDA Product Code 17H1.R2, CFIA File 800VV/M10.0/I6.2

All serials of this product must be released by the USDA prior to importation into Canada. All conditions described in the Permit to Import Veterinary Biologics must be followed with respect to the importation and sale of this product.

13. References

Alfonso C.L., Tulman E.R., Lu Z., Zsak L., Rock D.L., Kutish G.F. (2001). The Genome of Turkey Herpesvirus. Journal of Virology 75:971-978

Baigent S.J., Smith L.P., Nair V.K., Currie R.J.W. (2006). Vaccinal control of Marek's disease: Current challenges and future strategies to maximize protection. Veterinary Immunology and Immunopathology 112:78-86.

Calnek B.W., Witter R.L. (1991). Marek's Disease. In Diseases of Poultry, edited by B.W. Calnek et al., Iowa State University Press, Iowa. pages 342-385.

Cho B.R. (1974). Horizontal transmission of turkey herpesvirus to chickens IV. Maturation in the feather follicle epithelium. Avian Diseases. 19:136-141.

Cho B.R., Kenzy S.G. (1975). Horizontal transmission of turkey herpesvirus to chickens. 3. Transmission in three different lines of chickens. Poultry Science. 54:109-15.

Fadly, A.M. (2006). Marek's Disease. In The Merck Veterinary Manual, 9th edition, edited by C.M. Kahn, S. Line, S.E. Aiello, Merck & Co., Inc., New Jersey.

Fukuchi K., Sudo M., Tanaka A., Nonoyama M. (1985). Map locations of homologous regions between Marek's disease virus and herpesvirus of turkey and the absence of detectable homology in the putative tumour-inducing gene. Journal of Virology 53:994-997.

Gibbs C.P., Nazerian K., Velicer L.F., Kung H. (1984). Extensive homology exists between Marek's disease herpesvirus and its vaccine virus, herpesvirus of turkeys. Proceedings of the National Academy of Sciences 81:3356-3369.

Glaser L.C., Barker I.K., Weseloh D.V.C., Ludwig J., Windingstad R.M., Key D.W., Bollinger T.K. (1999). The 1992 epizootic of Newcastle disease in double-crested cormorants in North America. Journal of Wildlife Diseases 35:319-330.

Islam A., Walkden-Brown S.W. (2007). Quantitative profiling of the shedding rate of the three Marek's disease virus (MDV) serotypes reveals that challenge with virulent MDV markedly increases shedding of vaccinal viruses. The Journal of General Virology 88:2121-2128.

Jonas, D.A., Elmadfa, I., Engel, K.-H., Heller, K. J., Kozianowski, G., König, A., Müller, D., Narbonne, J. F., Wackernagel, W. and Kleiner, J. (2001). Safety considerations of DNA in food. Annals of Nutrition & Metabolism 45(6): 235-254.

Jurajda, V. and Klimes, B. (1970). Presence and survival of Marek's disease agent in dust. Avian Diseases 14(1):188-190.

Karaca G., Anobile J., Downs D., Burnside J., Schmidt C. (2004). Herpesvirus of turkeys: microarray analysis of host gene responses to infection. Virology 318:102-111.

Kinde H., Hullinger P.J., Charlton B., McFarland M., Hietala S.K., Velez V., Case J.T., Garber L., Wainwright S.H., Mikolon A.B., Breitmeyer R.E., Ardans A.A. (2005). The isolation of exotic Newcastle disease (END) virus from non poultry avian species associated with the epidemic of END in chickens in southern California: 2002-2003. Avian Diseases. 49:195-198.

King, D.J. (2006). Newcastle Disease. In The Merck Veterinary Manual, 9th edition, edited by C.M. Kahn, S. Line, S.E. Aiello, Merck & Co., Inc., New Jersey.

Lytle, C. D. and Sagripanti, J.-L. (2005). Predicted inactivation of viruses of relevance to biodefense by solar radiation. Journal of Virology 79(22): 14244-14252.

Powell P.C. (1986). Marek's Disease – A world poultry problem. World's Poultry Science Journal 14:205-218.

Schat K.A. (1985). Characteristics of the virus. In Marek's Disease, edited by L.N. Payne, Martinus Nijhoff Publishing, Boston.

Weingartl H.M., Riva J., Kumthekar P. (2003). Molecular characterization of avian paramyxovirus 1 isolates from cormorants in Canada from 1995 to 2000. Journal of Clinical Microbiology 41:1280-1284.

Witter, R. L. and Solomon, J. J. (1971). Epidemiology of a herpesvirus of turkeys: Possible sources and spread of infection in turkey flocks. Infection and Immunity 4(4): 356-361.

Wobeser G., Frederick A.L., Normal R., Myers D.J., Onderka D., Pybus M.J., Neufeld J.L., Fox G.A., Alexander D.J. (1993). Newcastle disease in wild water birds in western Canada, 1990. Canadian Veterinary Journal 34:353-359.

Zygraich N., Huygelen C. (1972). Inoculation of one-day-old chicks with different strains of turkey herpesvirus. II. Virus replication in tissues of inoculated animals. Avian Diseases. 16:793-798.