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Preventing Polyomavirus Infections

What data says a vaccine could work?

During polyomavirus outbreaks in mixed psittacine bird collections, infected survivors, and some birds that are exposed to them, have been shown to develop anti-polyomavirus neutralizing antibodies.1-3 Some young adult birds without antibodies will develop antibodies when housed adjacent to previously infected breeding adults, confirming that an antibody response does occur following natural exposure to the virus.1, 2, 4, 5 The detection of virus neutralizing antibodies in flocks of birds in which individuals are clinically normal suggests that many infections are subclinical (infected birds do not develop disease).1-3, 6 Collectively, these findings suggest that some exposed birds are able to mount an effective immune response. If a natural immunity to disease occurs, then it should be possible to induce a similar protective immunologic response through vaccination.

Will a vaccine protect psittacine birds from infection? Yes!

Experimental studies have indicated that an antibody response can be induced through vaccination, and that the resulting immunologic response is protective. In one study using blue and gold macaw chicks, an inactivated avian polyomavirus vaccine elicited polyomavirus neutralizing antibodies in all the vaccinates. The induced immunologic response protected the vaccinated chicks from subsequent challenge with live virus (Table 1 and 2).7 In other studies, an inactivated avian polyomavirus vaccine has been shown to protect Amazon parrots, cockatoos, African grey parrots and chickens from infection. (Ritchie, et al unpublished)

Is the vaccine safe? Yes!

In several studies, it was determined that avian polyomavirus could not be safely mixed with oil as an adjuvant for use in psittacine birds; however, vaccination trials have indicated that other adjuvants including Acemannan, Equimmune, aluminum hydroxide and Permulum can be used safely in companion birds (Table 3).8 (Ritchie, et al, unpublished data)

The safety and immunogenicity of avian polyomavirus vaccines, administered either intramuscularly or subcutaneously, were evaluated in a group of 233 mixed species Psittaciformes. The group of birds used for this study was considered the "typical" species in which the vaccine would be used and included lesser sulfur-crested cockatoos, sulfur-crested cockatoos, umbrella cockatoos, citron cockatoos, Moluccan cockatoos, Goffin's cockatoos, triton cockatoos, bare-eyed cockatoos, Major Mitchell's cockatoos, blue and gold macaws, red-fronted macaws, green-winged macaws, military macaws, scarlet macaws, African grey parrots, hawk-headed parrots, Amazon parrots, eclectus parrots, sun conures and Pionus parrots. Vaccinates ranged in age from 12-week-old neonates to > 5-year-old adults.

Vaccination stimulated a marked virus neutralizing antibody response, particularly in birds that already had antibodies prior to vaccination. The vaccine used in this study elicited polyomavirus neutralizing antibodies by 2 weeks after the second vaccination in 93% of the birds that did not have antibodies (seronegative) prior to vaccination, and 63% of the birds that already had antibodies (seropositive) prior to vaccination. Seventy-six percent of all the vaccinates had at least a 4-fold increase in virus neutralizing antibody titer at this time. Birds that were seronegative ( 8) at the beginning of the study had a greater increase in virus neutralizing antibody titer than did birds with a high titer ( 64) at the beginning of the study. The pre-and post-vaccination virus neutralizing antibody titers in vaccinates that were seronegative on day 0 are presented in Figure 1. The pre-and post-vaccination virus neutralizing antibody titers in vaccinates that were seropositive on day 0 are presented in Figure 2.6

Birds that were seronegative (titer 8) at the start of the study were statistically more likely (p = 0.000016) to respond to vaccination (4-fold or greater increase in titer) than birds that were seropositive (titer 16). Birds that had a low titer (16 or 32) were statistically more likely (p = 0.071) to develop a 4-fold or greater increase in antibody titer than birds that had a high prevaccination titer ( 64). A vaccinate's species, the presence or absence of an adjuvant, and the route of vaccination did not have a statistically significant effect on seroconversion. It is of particular interest to the avicultural community that the tested vaccine induced an antibody response in a wide range of psittacine species.6

Serious reactions were not observed in any vaccinate, and the appetites and attitudes of all the birds remained normal throughout the study. In 233 vaccinates representing species of macaws, cockatoos, conures and parrots, gross reactions were limited to small scab formation at the subcutaneous injection site in 3 African grey parrots. These minor lesions resolved uneventfully, without therapy. Some cockatoos and macaws experienced a heavy molt of up to 10 days duration that started 3 to 5 days after the second vaccination. It could not be determined if this molt occurred in response to vaccination, the stress associated with handling or was induced by climatic or other external factors. The molt was uneventful and appeared to have no adverse affect on the vaccinates.6

It was assumed that the birds in this study which were seropositive prior to vaccination (63% of total birds) had been previously infected, survived the infection and were possibly resistant to polyomavirus-induced disease. If this hypothesis was correct, the most important group in which post-vaccination seroconversion had to occur was those that were seronegative prior to vaccination. The fact that 93% of the birds that were seronegative prior to vaccination seroconverted by 2 weeks after the second vaccination suggests that the inactivated vaccine used in this study cold be safely used to produce a population of birds with virus neutralizing antibodies to avian polyomavirus.6

Will the vaccine negatively affect birds that already have been infected? No!

To establish the safety of a vaccine intended for widespread use, a flock of birds was used in which the virus neutralizing antibody titers to avian polyomavirus were not determined prior to the study. From previous studies, it was expected that many of the vaccinates would have prevaccination virus neutralizing titers. In fact, 63% of the birds used in this study were considered to have been previously exposed to avian polyomavirus because of the detection of virus neutralizing antibodies prior to vaccination.6

In another study, psittacine birds that were seropositive prior to vaccination developed no severe, adverse reactions after vaccination, even when they were vaccinated up to 5 times in a 49 day period (Table 4 and 5). The use of seropositive birds was considered important in both of these studies to confirm that an inactivated avian polyomavirus vaccine would not cause any adverse reactions in birds that were seropositive prior to vaccination.8

Given the prevalence of polyomavirus infections in companion birds, as indicated by the detection of virus neutralizing antibodies, it is noteworthy that an inactivated vaccine intended for commercial release did not cause adverse reactions in vaccinates that were seropositive before vaccination. A safe, effective, inactivated avian polyomavirus vaccine has been licensed by the University of Georgia to a manufacturer and is currently being tested to meet USDA requirements for registration. When this vaccine is available, the guidelines for use provided by the manufacturer should be carefully followed.

Reducing the chances of a polyomavirus outbreak!

Until this vaccine is available, sound hygienic practices, maintaining closed aviaries, preventing visitors from entering avian nurseries and attempting to identify and isolate birds that are shedding the virus using viral-specific DNA probe (Avian Research Associates, 100 Techne Center, Suite 101, Milford, OH. 45150, 1-513-248-4700) are the best methods for controlling this virus.

Polyomavirus virions are small, nonenveloped particles that are resistant to severe environmental conditions, a variety of disinfectants, organic solvents, to freezing and thawing and to heating to 56 C for two hours.9-11 A polyomavirus that infects primates (SV-40), has been shown to be inactivated by some products containing ethanol and resistant to others containing the same active ingredient.4, 12, 13 The environmental stability of avian polyomavirus causes a considerable problem in the aviary or hospital because persistently infected adult budgerigars have been shown to shed virus in their feather dust or excrement, and clinically affected nonbudgerigar psittacines have been shown to shed virus in their excrement. Transmission of the virus can be reduced by strict adherence to hygiene and appropriate use of disinfectants.

Cleaning of virus contaminated objects by manual removal of any organic debris (feces, food, blood, feather dust, etc) followed by the use of appropriate disinfectants is required to prevent or contain a polyomavirus outbreak. It should be noted that the presence of organic debris may reduce the efficacy of many disinfectants, increasing the contact time needed for a disinfectant to inactivate a pathogen. Chlorine-and iodine-containing disinfectants are highly reactive oxidizing agents that are easily inactivated through contact with organic debris.14 Thus, it is imperative that items that come in contact with birds be cleaned before they are disinfected.

The results of one study found that of the tested disinfectants that were effective against avian polyomavirus, Clorox was the least expensive (Table 6 ). However, this compound does produce fumes that can be irritating to mucus membranes and must be used in areas with sufficient ventilation. In addition, Clorox is irritating to skin and is corrosive to metals.15 Stabilized chlorine dioxide was found to inactivate polyomavirus despite its apparent safety for exposed humans and animals. Some studies suggest that in many applications, chlorine dioxide may be a superior disinfectant to chlorine (Clorox).16 At working dilutions, this disinfectant is considered safe for humans and animals and is used by many municipalities as the principle agent to eliminate potential pathogens from drinking water. In Europe, chlorine dioxide is used to treat drinking water because, unlike chlorine, it does not form carcinogenic trihalomethanes, chlorophenols or chloramines.17, 18 Nolvasan was found to be ineffective against avian polyomavirus, which may explain why nurseries that use this product to soak syringes between feedings can still experience polyomavirus outbreaks.

Given the extremely high prevalence of persistent polyomavirus infections, and the frequency with which budgerigars can shed the virus,19 it must be considered extremely dangerous and inadvisable to maintain young, nonbudgerigar psittacines in the same air space with budgerigars. The potential for intraspecies transmission of polyomavirus may be a particular problem in pet retailers that maintain both large and small psittacine birds.

A DNA probe based test is extremely valuable for identifying birds that are shedding virus in their excrement during an outbreak. Birds that are shedding the virus can be separated from others in a nursery to prevent further transmission. Testing cloacal swabs of a bird at the time of death to determine if it is shedding virus will help determine if its environment is contaminated. If the environment is contaminated, then there is a potential for viral amplification in a susceptible population. If an infected bird dies soon after infection, it may not be shedding virus at the time of death, and thus the bird's environment may not be contaminated with virus. Birds that are clinically ill, are found to be shedding polyomavirus or are in direct contact with birds that are clinically ill or shedding polyomavirus should be isolated (placed in a separate geographic location) from birds that are clinically normal and not shedding virus.20-22

Nonbudgerigar psittacine birds that are shedding avian polyomavirus should be isolated, not euthanatized. Birds that are shedding avian polyomavirus are likely to be of no further concern when an effective vaccine becomes available for widespread use. Nonbudgerigar psittacines that are shedding avian polyomavirus can be best managed by maintaining them in restricted environments in which they do not directly or indirectly (i.e., through contaminated excrement, secretions, bedding or enclosures) expose other nonvaccinated birds, particularly neonates, to the virus. Breeding birds shedding polyomavirus should be separated from the remainder of the collection, and offspring from these birds should be raised separately from offspring of birds that are not shedding the virus.22

Testing budgerigars for the presence of virus neutralizing antibodies and culling (destroying) positive birds has been suggested as a method to establish specific-pathogen-free flocks of budgerigars.22, 23 Additionally, depopulation of budgerigar aviaries experiencing outbreaks followed by restocking with seronegative birds has been suggested as a method of controlling enzootic infections in this species.22-24 However, given the seroprevalence of avian polyomavirus in nonbudgerigar psittacine birds (up to 63% in some flocks)6 culling all the birds with an antibody titer creates an unacceptable carnage and is not practical, nor recommended procedure for controlling polyomavirus infections. Additionally, neonates from a specific-pathogen-free flock that were shipped to another location would be expected to be particularly susceptible to infection. Vaccination will provide a better method to control avian polyomavirus infections in nonbudgerigar psittacines than will any type of testing procedures.6-8

Acknowledgments: Major sustained contributions that have made this work possible have been provided by the Cowan Avian Health Foundation, the International Avian Research Foundation, Midwest Avian Research Exposition, International Aviculturists Society, Terry Clyne, Richard and Luanne Porter, Kathleen Sazbo, Knick Enterprises, Avian Research Fund, Gateway Parrot Club, South Jersey Bird Club, Kentuckian Bird Society, Central Indiana Cage Bird Club, Hookbill Hobbyists of Southern California, Greater Brandon Avian Society, and Zeigler Bros, Inc. Hundreds of aviculturists, bird clubs and veterinarians have also made significant contributions. This research could not have been completed without the technical assistance of Michelle Weatherly, Richard and Luanne Porter, Bill Bennett, Brett Blanchard, Mary Ervin, Kathy Murphy, Sherri and Aaron Jones, Will Pace, Don Sanders, Debbie Seaman, Marcus Valentine, Cynthia Webb and Diane Wolff.

Table contents not shown secondary to formatting difficulties. The reader is referred to the Proceedings for detailed information.

Table 1--Virus-neutralizing antibody titers in a group of blue and gold macaw chicks on days 10, 20 and 30 after vaccination with ß-propiolactone-inactivated avian polyomavirus.

Table 2--Virus neutralizing antibody titers in a group of blue and gold macaw chicks overtime after viral challenge with viable avian polyomavirus on days 37, 39 and 52.

Table 3. Reciprocal virus neutralizing antibody titers in 24 psittacine birds vaccinated with one of five antigen+ plus adjuvant combinations by the subcutaneous route.

Table 4. Reciprocal virus neutralizing antibody titers in 21 psittacine birds vaccinated with antigen* plus Acemannan or Equimune adjuvant by the intramuscular route.

Table 5. Reciprocal virus neutralizing antibody titers in 24 birds with antigen+ only or one of four antigen-adjuvant combinations by the subcutaneous route. Birds are divided into those that were seronegative or seropositive before vaccination.

Table 6. Activity of disinfectants at recommended dilutions against avian polyomavirus.

References

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