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Rural Industries Research & Development Corporation
Summary of full report
MOLECULAR EPIDEMIOLOGY
OF RHODOCOCCUS EQUI
By Dr G F Browning and Ms A C Morton
December 2000
RIRDC Project No UM-29A
The significance of the virulence associated plasmid in Rhodococcus equi isolated from foals on Australian stud farms was analysed by using polymerase chain reaction to examine 210 isolates from clinical cases for the presence of the vapA gene, which is carried on the virulence plasmid. All but 3 isolates contained the vapA gene.
Relationships between Rhodococcus equi isolates from clinical cases of pneumonia on Australian stud farms were compared to each other and to isolates from other countries using pulsed field gel electrophoresis of genomic DNA digested with the restriction endonuclease AsnI. Forty four strains were found among 210 Australian isolates, with five of these accounting for over half the isolates, and the 22 strains isolated more than once for 90% of the isolates. The average genotypic diversity on each farm and in each year was found to be less than the genotypic diversity of the isolates taken as a whole, with 11% of the total diversity being due to differences between farms. A small number of strains on each farm were found to have caused at least half the clinical cases of disease, and these varied between farms, and to a lesser extent, years. Most strains were found on more than one farm, and some very similar restriction patterns were found among isolates from different continents, indicating that strains can be very widespread. Multiple strains were isolated in five of the six cases where more than one isolate from a single foal was examined, indicating that disease may commonly be caused by simultaneous infection with multiple strains. No significant differences were found between strains in the clinical disease caused. It was therefore concluded that there are a number of different strains of R. equi which can be virulent, and these strains tend to be widespread, but farms tend to have particular strains associated with them. There was no evidence that particular strains were associated with more severe disease.
R. equi were grown
under a variety of restrictive growth conditions, including iron, calcium
and magnesium restriction and higher carbon dioxide concentrations. The
proteins produced by R. equi under these conditions were compared
using Coomassie blue staining and Western blotting with sera from clinical
cases, but there was no consistently present immunoreactive protein unique
to the restrictive conditions which might be an appropriate basis for a
serological test.
Current approaches to identifying
an appropriate antigen for an improved serological test are based on expressing
different genes present on the virulence plasmid in Escherichia coli, then
examining these recombinant proteins for reactivity with sera from clinical
cases. This has led to the identification of at least one recombinant protein
with potential as a diagnostic antigen.
It is clear that future work
needs to examine the ecology of virulent R. equi on stud farms,
as past studies may have been predominantly examining affects on populations
which were not carrying the virulence plasmid. Furthermore ongoing work
needs to concentrate on the important antigens which may be present on
the virulence plasmid. We were unable to find any evidence that particular
strains were more virulent, although there was a clear difference between
the dominant strain causing disease on some studs. On the balance of probability
it seems unlikely that differences in mortality rate, or the incidence
of disease, on different studs can be attributed predominantly to the strain
present on that farm.
 
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