M. Drancourt et al., 16S ribosomal DNA sequence analysis of a large collection of environmentaland clinical unidentifiable bacterial isolates, J CLIN MICR, 38(10), 2000, pp. 3623-3630
Some bacteria are difficult to identify with phenotypic identification sche
mes commonly used outside reference laboratories. 16S ribosomal DNA (rDNA)-
based identification of bacteria potentially offers a useful alternative wh
en phenotypic characterization methods fail. However, as yet, the usefulnes
s of 16S rDNA sequence analysis in the identification of conventionally uni
dentifiable isolates has not been evaluated with a large collection of isol
ates. In this study, we evaluated the utility of 16S rDNA sequencing as a m
eans to identify a collection of 177 such isolates obtained from environmen
tal, veterinary, and clinical sources. For 159 isolates (89.8%) there was a
t least one sequence in GenBank that yielded a similarity score of greater
than or equal to 97%, and for 139 isolates (78.5%) there was at least one s
equence in GenBank that yielded a similarity score of greater than or equal
to 99%. These similarity score values were used to defined identification
at the genus and species levels, respectively, For isolates identified to t
he species level, conventional identification failed to produce accurate re
sults because of inappropriate biochemical profile determination in 76 isol
ates (58.7%), Gram staining in 16 isolates (11.6%), oxidase and catalase ac
tivity determination in 5 isolates (3.6%) and growth requirement determinat
ion in 2 isolates (1.5%), Eighteen isolates (10.2%) remained unidentifiable
by 16S rDNA sequence analysis but were probably prototype isolates of new
species. These isolates originated mainly from environmental sources (P = 0
.07), The 16S rDNA approach failed to identify Enterobacter and Pantoea iso
lates to the species level (P = 0.04; odds ratio = 0.32 [95% confidence int
erval, 0.10 to 1.14]). Elsewhere, the usefulness of 16S rDNA sequencing was
compromised by the presence of 16S rDNA sequences with > 1% undetermined p
ositions in the databases. Unlike phenotypic identification, which can be m
odified by the variability of expression of characters, 16S rDNA sequencing
provides unambiguous data even for rare isolates, which are reproducible i
n and between laboratories. The increase in accurate new 16S rDNA sequences
and the development of alternative genes for molecular identification of c
ertain taxa should further improve the usefulness of molecular identificati
on of bacteria.