Definitive identification of the species in the Burkholderia cepacia comple
x by routine clinical microbiology methods is difficult, Phenotypic tests t
o identify B, multivorans and B. vietnamiensis have been established; more
recent work indicates B, stabilis may also be identified by growth characte
ristics and biochemical tests, However, attempts to identify genomovars I a
nd III have, thus far, proved unsuccessful, Previously, we demonstrated the
utility of two primer pairs, directed to the rRNA operon, to specifically
identify the B, cepacia complex in a PCR, One of these primer pairs, G1-G2,
only amplified a DNA fragment from genomovars I and III and B, stabilis in
a PCR with genomic DNA isolated from prototypical strains representing the
five genomovars. Sequence analysis of the rRNA operon for all the genomova
rs indicated that this primer pair targeted a region shared by these isolat
es. Further analysis revealed a region of heterogeneity between genomovar I
II and B. stabilis internal to the amplified product of G1-G2, Primers desi
gned to target this region were tested with prototypical strains following
an initial amplification with the G1-G2 primer pair, New primers specific f
or the prototypical genomovar III and B, stabilis were designated SPR3 and
SPR I, respectively. Analysis of 93 isolates representing 18 genomovar I, 1
3 B. multivorans, 36 genomovar III, 11 B, stabilis, and 15 B. vietnamiensis
isolates was performed. DNA from all isolates of genomovars I and III and
B. stabilis was amplified by G1-G2, Genomovar III isolates yielded a produc
t with SPR3/G1 while B, stabilis amplified with SPR4-G1, Genomovar I isolat
es were amplified by either SPR3-G1 or SPR4-G1, but not both. B, multivoran
s yielded a product with SPR3-G1 but not G1-G2, and B, vietnamiensis isolat
es were negative in all PCRs, Thus using an algorithm with G1-G2, SPR3-G1,
and SPR4-G1 primers in a PCR analysis, genomovar III isolates can be separa
ted from B, stabilis and the identity of B, multivorans and B, vietnamiensi
s can be confirmed.