Newcastle Disease Vaccine, Inactivated
(Ph Eur monograph 0870)
Newcastle disease vaccine (inactivated) [also known as avian
paramyxovirus 1 vaccine (inactivated) for vaccines intended for some species] consists of
an emulsion or a suspension of a suitable strain of Newcastle disease virus (avian
paramyxovirus 1) that has been inactivated in such a manner that immunogenic activity is
The virus is propagated in embryonated eggs from healthy
flocks or in suitable cell cultures (5.2.4).
The test for inactivation is carried out in embryonated eggs
or suitable cell cultures and the quantity of inactivated virus used is equivalent to not
less than ten doses of vaccine. No live virus is detected.
The vaccine may contain an adjuvant.
CHOICE OF VACCINE COMPOSITION
The vaccine is shown to be satisfactory with respect to safety
(5.2.6) and immunogenicity (5.2.7) for each species and category of birds for which it is
intended. The following tests may be used during demonstration of immunogenicity.
For domestic fowl, the test with virulent challenge (test B)
described under Potency is suitable for demonstrating immunogenicity. For other species of
birds (for example, pigeons or turkeys), test C described under Potency is suitable for
Batch potency test
For vaccines used in domestic fowl
Carry out test A described under Potency but, if the nature of
the product does not allow valid results to be obtained with test A or if the batch does
not comply with test A, carry out test B. A test using fewer than twenty birds per group
and a shorter observation period after challenge may be used if this has been shown to
give a valid potency test.
For vaccines used in species other than domestic
Carry out a suitable validated test for which a satisfactory
correlation has been established with test C described under Potency, the criteria for
acceptance being set with reference to a batch that has given satisfactory results in the
latter test. A test in chickens from a flock free from specified pathogens (5.2.2) and
consisting of a measure of the serological response to graded amounts of vaccine (for
example, 1/25, 1/50 and 1/100 dose with serum sampling 17 to 21 days later) may be used.
When injected into animals free from antibodies against
Newcastle disease virus, the vaccine stimulates the production of such antibodies.
If the vaccine is intended for use in domestic fowl, vaccinate
ten chickens from a flock free from specified pathogens (5.2.2). If the vaccine is not for
use in domestic fowl, use ten birds of one of the species for which the vaccine is
intended, free from antibodies against Newcastle disease virus. Inject twice the
vaccinating dose by a recommended route into each of ten birds, 14 to 28 days old. Observe
the birds for 21 days. No abnormal local or systemic reaction occurs.
Inject two-fifths of a dose into the allantoic cavity of each
of ten embryonated hen eggs, 9 to 11 days old, from flocks free from specified pathogens
(5.2.2) (SPF eggs) and incubate. Observe for 6 days and pool separately the allantoic
fluid from eggs containing live embryos, and that from eggs containing dead embryos,
excluding those dying within 24 h of the injection. Examine embryos that die within 24 h
of injection for the presence of Newcastle disease virus: the vaccine does not comply with
the test if Newcastle disease virus is found.
Inject into the allantoic cavity of each of ten SPF eggs, 9 to
11 days old, 0.2 ml of the pooled allantoic fluid from the live embryos and, into each of
ten similar eggs, 0.2 ml of the pooled fluid from the dead embryos and incubate for 5 to 6
days. Test the allantoic fluid from each egg for the presence of haemagglutinins using
The vaccine complies with the test if there is no evidence of
haemagglutinating activity and if not more than 20 per cent of the embryos die at either
stage. If more than 20 per cent of the embryos die at one of the stages, repeat that
stage; the vaccine complies with the test if there is no evidence of haemagglutinating
activity and not more than 20 per cent of the embryos die at that stage.
Antibiotics may be used in the test to control extraneous
Inject a double dose by a recommended route into each of ten
chickens, 14 to 28 days old and from a flock free from specified pathogens (5.2.2). After
3 weeks, inject one dose by the same route. Collect serum samples from each chicken 2
weeks later and carry out tests for antibodies to the following agents by the methods
prescribed for chicken flocks free from specified pathogens (5.2.2): avian
encephalomyelitis virus, avian infectious bronchitis virus, avian leucosis viruses,
egg-drop syndrome virus, avian bursal disease virus, avian infectious laryngotracheitis
virus, influenza A virus, Marek's disease virus. The vaccine does not stimulate the
formation of antibodies against these agents.
The vaccine complies with the test for sterility prescribed in
the monograph on Vaccines for veterinary use (0062).
For vaccines for use in domestic fowl carry out
test A. If an unsatisfactory result is obtained with test A but a satisfactory result is
obtained with test B, the vaccine complies with the requirements. For vaccines for use
only in other species, such as pigeons, carry out test C.
intramuscularly into each of not fewer than ten chickens, 21 to 28 days old and from a
flock free from specified pathogens (5.2.2), a volume of the vaccine equivalent to 1/50 of
a dose. 17 to 21 days later, collect serum samples from each vaccinated chicken and from
each of not fewer than five control chickens of the same age and from the same source.
Measure the antibody levels in the sera by the haemagglutination-inhibition (HI) test
using the technique described below or an equivalent technique with the same numbers of
haemagglutinating units and red blood cells. The test system used must include negative
and positive control sera, the latter having an HI titre of 5.0 log2 to 6.0 log2.
The vaccine complies with the test if the mean HI titre of the
vaccinated group is equal to or greater than 4.0 log2 and that of the
unvaccinated group is 2.0 log2 or less. If the HI titres are not satisfactory,
carry out test B.
Inactivate the test sera by heating at 56C for 30 min. Add
0.025 ml to the first row of wells in a microtitre plate. Add 0.025 ml of a buffered 9 g/l
solution of sodium chloride R at pH 7.2 to pH 7.4 to the rest of the wells. Prepare
twofold dilutions of the sera across the plate. To each well add 0.025 ml of a suspension
containing 4 haemagglutinating units of inactivated Newcastle disease virus. Incubate the
plate at 4C for 1 h. Add 0.025 ml of a 1 per cent V/V suspension of red blood
cells collected from chickens, 3 to 4 weeks old and free from antibodies against Newcastle
disease virus. Incubate the plate at 4C for 1 h. The HI titre is equal to the highest
dilution that produces complete inhibition.
chickens 21 to 28 days old and from a flock free from specified pathogens (5.2.2). For
vaccination, use not fewer than three groups, each of not fewer than twenty chickens; keep
a group of ten chickens as controls. Choose a number of different volumes of the vaccine
corresponding to the number of groups: for example, volumes equivalent to 1/25, 1/50 and
1/100 of a dose. Allocate a different volume to each vaccination group. Inject
intramuscularly into each chicken the volume of vaccine allocated to its group. 17 to 21
days later, challenge all the chickens by intramuscular injection of 6 log10
embryo LD50 of the Herts (Weybridge 33/56) strain of Newcastle disease virus.
Observe the chickens for 21 days. Calculate the PD50 by standard statistical
methods from the number of chickens that survive in each vaccinated group without showing
any clinical evidence of Newcastle disease during the 21 days. The vaccine complies with
the test if the smallest dose stated on the label corresponds to not less than 50 PD50
and the lower confidence limit is not less than 35 PD50 per dose. If the lower
confidence limit is less than 35 PD50 per dose, repeat the test; the vaccine
must be shown to contain not less than 50 PD50 in the repeat test. The test is
not valid unless all the control birds die within 6 days of challenge.
not fewer than twenty birds of the target species, free from antibodies against avian
paramyxovirus 1, in accordance with the recommendations for use. Maintain as unvaccinated
controls a group of not fewer than ten birds of the same age, from the same source and
free from antibodies against avian paramyxovirus 1. The test is invalid if serum samples
obtained at the time of the first vaccination show the presence of antibodies against
avian paramyxovirus 1 in either vaccinates or controls or if tests carried out at the time
of challenge show such antibodies in controls. 4 weeks after the last vaccination,
challenge each bird intramuscularly with a sufficient quantity of virulent avian
paramyxovirus 1. The test is not valid if fewer than 90 per cent of the control birds die
or show serious signs of Newcastle disease virus infection. The vaccine complies with the
test if not fewer than 90 per cent of the vaccinated birds survive and show no serious
signs of avian paramyxovirus 1 infection.