It is a recommendation of the draft OIE norms for CBPP vaccine that the production of CBPP vaccine be based on a seed lot system and that the seed lot must have been shown to be safe for cattle when administered parenterally (subcutaneously), and be capable of conferring an immunity which lasts for at least one year. The seed lot prepared in such a manner must not be submitted to more than 3 additional passages to produce the final vaccine.
I. Materials and Methods
Animals
Cattle which should be at least 24 months old and should come from an area free from CBPP are required for the test of CBPP vaccine potency. They should be representative of the breed(s) to be vaccinated. The selected cattle are indelibly marked and held in quarantine for at least 30 days before start of experiment. They are bled at the time of purchase and just before vaccination and their sera shown to be free from antibody to CBPP by the CFT or ELISA tests. Any animal that shows signs of disease during the quarantine period shall not be used for the test of potency unless the cause is determined and recognised as having no effect on the experiments.
CBPP vaccine
Reconstituted and diluted in sterile distilled water or PBS or 1M MgSO4 solution so that 1 ml of the vaccine contains at least 107 live mycoplasmas (i.e.the minimum number of mycoplasmas that should be contained in one field dose of CBPP vaccine).
Experimental design
Many procedures have been described for the performance of potency testing of CBPP vaccine. Generally the cattle to be used for the test of potency are divided into three groups. One group constitutes the vaccinates (to be inoculated with the vaccine), the second group will comprise the unvaccinated controls (to be inoculated with the reconstitution fluid) while the third group of cattle will be the intubates (i.e. those animals which are artificially infected with virulent mycoplasma and will serve as the source of virulent contact challenge. Twelve months after vaccination the animals that were immunised, the controls and the intubates are mixed and confined in an infectious disease containment facility for three months (challenge period). Throughout the experimental period the following activities are carried out:
temperature of each animal is recorded daily
blood samples are collected from each animal on day 15, 30 and thereafter every month following vaccination
observed clinical signs of disease are recorded
animals that die are subjected to complete post mortem examination and attempts should be made to isolate M. mycoides subsp. mycoides from the lung and the tracheobronchial lymph nodes. Any artificially infected animal that dies should be replaced with another infected animal in order to maintain the ratio of at least 1 disease donor animal to 3 challenge cattle.
At the end of the challenge all the remaining cattle are killed and examined post mortem to establish presence or absence of CBPP lesions in the lung. Lung tissue and thoracic lymph nodes should also be cultured for mycoplasma. Isolated mycoplasma should be identified.
all the sera collected since the commencement of the experiment are tested for CBPP antibody by CF or ELISA tests.
II. Interpretation of the result
A seed lot passes the test for potency in cattle if a vaccine prepared using the seed lot protects at least 80% of the vaccinated animals. In addition, at least 80% of the vaccinated animals:
should not show CBPP clinical signs
should not show CBPP lesions on post mortem
should not have M. mycoides subsp. mycoides either in the lung or in the tracheobronchial lymph nodes
At least 80% of the control animals should develop signs of CBPP and at post mortem they should show lesions typical of CBPP.
A scoring system that has been used for the assessment of the potency of CBPP vaccine in cattle is given below.
Ref: Hudson, J.R. and Turner, A.W. (1963); Scoring of the status of CBPP in immunised animals following challenge. Australian Vet. Journal., 39, 373.
