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Letter
Yersinia pestis
Genotyping
To the Editor: Drancourt et al. (1) report the
development of an original genotyping system for Yersinia pestis
based on intergenic spacer sequencing. However, the approach appears to
rely upon the characterization of polymorphisms due to tandem repeat variation.
Eight spacers are used in the report, 7 of which contain tandem repeats,
and the sequence variability used to produce the typing data and the strain
clustering result from variation in the number of tandem repeats (and
incorrect data analysis produces a dendrogram with 34 branches from only
19 different isolate types). Three of the spacers and associated polymorphisms
were previously reported. Spacers YP3 and YP5 are, respectively, ms38
and ms56 (2); spacer YP10 is M61 (3).
YP3 is later used to investigate ancient DNA samples, and 3 amplification
products are described in detail. The sequences are compared to modern
sequences by BLAST analysis, which is not relevant for tandem repeats.
Instead, the Figure shows how internal variation
within the array can be coded to facilitate interpretation. In this collection,
Orientalis strains are "abcdeeef," whereas Antiqua strains from
Africa are "abcdeef." All these different codes can be deduced
one from the other by simple duplication and deletion events, with no
need to invoke point mutations. The codes for all 3 ancient samples are
identical to the Orientalis code "abcdeeef."
In conclusion, the data presented by Drancourt et al. do not appear to
support their claim. They did not invent a new genotyping method but used
the well-known multiple locus variable analysis (MLVA) number of tandem
repeats approach. The finding that the "genotype Orientalis was involved
in all three pandemics" is not valid since the Orientalis type is
defined by a biochemical assay, resulting in all known Orientalis strains
from a 93-bp glycerol-3-phosphate dehydrogenase microdeletion (4,5),
which was not investigated here.
Gilles Vergnaud*
*Centre d'Etudes du Bouchet, Vert le Petit, France
Suggested citation
for this article:
Vergnaud G. Yersinia pestis genotyping [letter]. Emerg Infect Dis
[serial on the Internet]. 2005 Aug [date cited]. Available from
http://www.cdc.gov/ncidod/EID/vol11no08/04-0942_05-0568.htm
References
- Drancourt M, Roux V, Dang LV, Tran-Hung L, Castex
D, Chenal-Francisque V, et al. Genotyping,
Orientalis-like Yersinia pestis, and plague pandemics. Emerg
Infect Dis. 2004;10:1585–92.
- Le Flèche P, Hauck Y, Onteniente L, Prieur A, Denoeud F, Ramisse V,
et al. A
tandem repeats database for bacterial genomes: application to the genotyping
of Yersinia pestis and Bacillus anthracis. BMC Microbiol.
2001;1:2.
- Klevytska AM, Price LB, Schupp JM, Worsham PL, Wong J, Keim P. Identification
and characterization of variable-number tandem repeats in the Yersinia
pestis genome. J Clin Microbiol. 2001;39:3179–85.
- Motin VL, Georgescu AM, Elliott JM, Hu P, Worsham PL, Ott LL, et al.
Genetic
variability of Yersinia pestis isolates as predicted by PCR-based
IS100 genotyping and analysis of structural genes encoding glycerol-3-phosphate
dehydrogenase (glpD). J Bacteriol. 2002;184:1019–27.
- Pourcel C, Andre-Mazeaud F, Neubauer H, Ramisse F, Vergnaud G. Tandem
repeats analysis for the high resolution phylogenetic analysis of Yersinia
pestis. BMC Microbiol. 2004;4:22.
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In Response: We thank Dr. Vergnaud for his
response (1). Since the time of our publication (2),
2 articles related to our paper were either submitted or published. One
(3) reported identification of Yersinia pestis–specific
genes in teeth from patients who died during the Justinian plague; another
proposed identification of Y. pestis strains by using variable
numbers of tandem repeats analysis (VNTR) (4). The authors
concluded that isolates could easily be compared in their database by
using 7 markers. As opposed to work with cultures where ample, high-quality
DNA template is available, successful amplifications with 7 different
primer sets cannot be achieved by using DNA extracted from ancient teeth
(5). By comparing genome sequences, we evaluated short
intergenic spacers that were more divergent. Divergences included mutations,
deletions, and duplications (VNTR). Phylogenetically, an entire repeat
unit has the same weight as that of a single nucleotide polymorphism.
By sequencing, we have identified all events (single nucleotide polymorphism
and VNTR). Sequencing is more versatile for use in strain identification
(5), allows distinction at the species level, and can
be applied directly on clinical and forensic samples.
The discovery of a unique sequence is critical to authenticate results
in such controversial areas as paleomicrobiology (5).
Fortunately, we have identified a unique sequence that contains several
mutations. These mutations do not exclude this strain from being Y.
pestis (see Figure). Additionally, we doubt
that our conclusions would have been accepted had we simply used the VNTR,
demonstrating only an amplicon of the right size on a gel.
In conclusion, our results have been validated by others. The sequence
is original and, therefore, authentic. Dr. Vergnaud agrees that the results
we presented did represent a sequence associated with the Orientalis biovar.
This finding may end the controversy.
Didier Raoult,*
Michel Drancourt,* Pierre Edouard Fournier,* and Hiro Ogata*
*Centre National de Reference, Marseille, France
Suggested citation
for this article:
Raoult D, Drancourt M, Fournier PE, Ogata H. Yersinia pestis genotyping
[in response]. Emerg Infect Dis [serial on the Internet]. 2005 Aug [date
cited]. Available from http://www.cdc.gov/ncidod/EID/vol11no08/04-0942_05-0568.htm
References
- Vergnaud G. Yersinia pestis genotyping. Emerg
Infect Dis. 2005;11:1317–8.
- Drancourt M, Roux V, Dang LV, Tran-Hung L, Castex D, Chenal-Francisque
V, et al. Genotyping,
Orientalis-like Yersinia pestis, and plague pandemics. Emerg
Infect Dis. 2004;10:1585–92.
- Weichmann I, Grupe G. Detection
of Yersinia pestis DNA in two early medieval skeletal finds from
Aschheim (Upper Bavaria, 6th century AD). Am J Phys Anthropol. 2005;126:48–55.
- Pourcel C, Andre-Mazeaud F, Neubauer H, Ramisse F, Vergnaud G. Tandem
repeats analysis for the high resolution phylogenetic analysis of Yersinia
pestis. BMC Microbiol. 2004;4:22.
- Drancourt M, Raoult D. Paleomicrobiology:
current issues and perspectives. Nat Rev Microbiol. 2005;3:23–35.
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