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Live vaccines for three species of Eimeria
by Glenn Anderson and Wayne Jorgensen
March 2004
RIRDC Publication No 03/143 RIRDC Project No DAQ-259J
1. Attenuated lines of E. mitis, E. brunetti and E. praecox to be developed for incorporation in an efficacious live vaccine protective against all seven species of Eimeria in Australian chickens.
2. A trial technique to evaluate coccidiostat resistance to be developed.
Project outcomes are summarised below under the two objective headings.
1. Attenuated lines of E. mitis, E. brunetti and E. praecox to be developed for incorporation in an efficacious live vaccine protective against all seven species of Eimeria in Australian chickens.
1a. Source of vaccine
and challenge isolates of E.
mitis, E. brunetti and E.
praecox.
Parasites used in live
coccidiosis vaccines must have high susceptibility to anti-coccidial compounds
(coccidiostats). Parent strains used for vaccine development were therefore
collected from small, noncommercial flocks that did not routinely use coccidiostats
and in which clinical coccidiosis had not been a problem. Isolates of E.
mitis, E. brunetti and E.
praecox were collected from flocks
of this type during the phase 2 project (RIRDC Project DAQ-215J). In addition,
isolates of E. mitis, E.
brunetti and E.
praecox were collected from commercial
operations for use as challenge strains to test the immunity induced by
the vaccine lines. All isolates were purified by a series of single oocyst
passages (i.e. inoculating a single oocyst into a chicken and allowing
it to reproduce). Purified isolates were stored in liquid nitrogen.
1b. Modification
of the prepatent period of one strain of each of these species by selecting
for precocious development.
The prepatent period of
the parasites was modified by serial passaging of parasites through susceptible
chickens, in each case using the oocysts recovered earliest from the previous
passage. Two isolates of E.
mitis were passaged in this way.
The prepatent period of the Jorgensen strain was successfully reduced 20
hours over nine passages and a concomitant reduction in virulence was achieved.
The strain was later evaluated for use as a vaccine.
Two strains of E. brunetti underwent selection for precocious development (Bowden strain (9 passages) and Monarto strain (13 passages)). Neither was suitable for use as a vaccine strain due to high pathogencity. The Bowden parent strain, however, was found during drug sensitivity trials to have very low pathogenicity. After discussions with the project commercial partners and the RIRDC Research Manager, the Bowden parent strain was subsequentally evaluated for use as a vaccine.
Eimeria praecox is the least pathogenic of the Eimeria species present in Australia. Two strains (Jorgensen and Kelly) were evaluated for virulence while being tested for drug sensitivity. Due to its very low pathogenicity, the Jorgensen strain was chosen for further evaluation without passaging for precocious development.
1c. Characterisation
of strains in terms of drug sensitivity, reproductive potential, pathogenicity
and protection to homologous and heterologous challenge.
The Jorgensen and Kelly
strains of E. mitis,
the Monarto and Bowden strains of E.
brunetti, and the Jorgensen and
Kelly strains of E. praecox
were characterised for drug sensitivity.
The Jorgensen strain of E.
mitis (precocious), the Bowden
strain of E. brunetti and
the Jorgensen strain of E.
praecox were characterised for
pathogenicity and protection against heterologous challenge. The Jorgensen
strain of E. mitis (precocious)
was also characterised for reproductive potential and protection against
homologous challenge. The Bowden strain of E.
brunetti (precocious) also underwent
characterisation for protection against homologous challenge. All characterisation
trials were cage trials with randomised block designs.
Drug sensitivity: The Bowden strain of E. brunetti and the Jorgensen strain of E. praecox were found to be susceptible to manufacturers’ recommended treatment doses of Toltrazuril, Amprolium and Sulphaquinoxaline. The Jorgensen strain of E. mitis was found to be susceptible to recommended treatment doses of Toltrazuril and Amprolium.
Pathogenicity: Doses of 20 000 oocysts of Jorgensen strain E. mitis or 15 000 oocysts of Jorgensen strain E. praecox or Bowden strain E. brunetti caused no significant increase in feed conversion ratios or decrease in bodyweight gain of experimental birds, indicating that the proposed vaccine strains had very low pathogenicity.
Homologous challenge: The Jorgensen strain of E. mitis provided protection against challenge with the parent strain. Homologous challenge trials were not performed on the Bowden strain of E. brunetti or the Jorgensen strain of E. praecox as the vaccine strains are unmodified parent strains. It was demonstrated, however, that the precocious line of the Bowden strain of E. brunetti provided protection against challenge with the parent strain.
Heterologous challenge: Birds vaccinated with the Jorgensen strain of E. mitis, Bowden strain of E. brunetti or Jorgensen strain of E. praecox were protected against challenge with two field strains of the same species.
Reproductive potential: The attenuated line of the Jorgensen strain of E. mitis demonstrated a highly reduced reproductive potential compared with the parent strain, which is an indicator of the success of the attenuation process. Reproductive potential trials were not performed on the Bowden strain of E. brunetti or the Jorgensen strain of E. praecox as they were unmodified parent strains.
2. A trial technique
to evaluate coccidiostat resistance to be developed.
A randomised block design
was developed to determine the ability of in-feed coccidiostats (Narasin
and Monensin) to protect birds against the harmful effects of infection
with various parasite stocks.
Parasites used in the new trial format were Eimeriavax 4m (the new vaccine containing E. necatrix, E. tenella, E. acervulina and E. maxima developed in Stages 1 and 2 RIRDC projects), field isolates from a commercial layer farm and a broiler farm, and the Kelly strain of E. praecox, which was not significantly affected by Toltrazuril in the standard drug sensitivity trial. There was evidence of some resistance to both Narasin and Monensin.
Conclusion
The candidate vaccine
strains selected for field evaluation were drug susceptible, had low pathogenicity
and were highly protective against challenge with two virulent field isolates
(heterologous challenge). The strains have been transferred to Eimeria
Pty Ltd for incorporation in
new custom vaccines. A new project to develop the diagnostic tools necessary
to support use of the live vaccines in the marketplace and ensure their
continued efficacy will be supported by RIRDC from 2004.
 
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