Journal List > J Bacteriol Virol > v.40(3) > 1033965

Kim, Park, Kim, Han, Kim, Kim, Lee, Hong, and Lee: Molecular Genetic Typing of Legionella pneumophila Strains Isolated in Jejudo, Korea

Abstract

Twenty two strains of Legionella species isolated from Jejudo, Korea were identified by comparing the rpoB (300 bp), dotA (360 bp), and mip (396 bp) gene sequence analysis. Furthermore, their genotypes were determined by sequence analysis of rpoB/dotA subgroup typing, pulse-field gel electrophoresis (PFGE) and random amplified polymorphic DNA (RAPD) patterns. Of the 22 isolates, 21 strains were identified as L. pneumophila and 1 strain was close to L. erythra (>95% similarity of rpoB and mip). Most of the L. pneumophila strains (90%) belonged to P-I of rpoB/dotA subgroup typing, one strain of each P-III and P-IV. L. pneumophila isolates were further grouped into 4 and 6 different PFGE (P1 to P4) and RAPD (R1 to R6) patterns, respectively. On the basis of these genotypes, which may be useful for future epidemiological studies, existence of diverse L. pneumophilla population in Jejudo, Korea were observed.

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Figure 1.
Genetic relationships of Legionella Jeju isolates (•) among 41 legionella species based on the rpoB and mip sequences. In addition to the sequence similarity, most of the Jeju isolates were clearly identified as L. pneumophila, except one strain (Jeju 8-5). Strains possessing sequences identical to those in the figures are not presented. Bootstrap values higher than 50% are shown.
jbv-40-111f1.tif
Figure 2.
Neighbour-joining trees inferred from (A) the rpoB and (B) the dotA sequences of L. pneumophila Jeju isolates and reference strains (K and JLP) (26). Subgroups (P-I to PVIII and F-I and F-II) are indicated on the right; branch lengths are proportional to the number of nucleotide changes. Bootstrap values higher than 50% are indicated at the corresponding branches. SG, serogroup numbers (SG 1 to 15).
jbv-40-111f2.tifjbv-40-111f2a.tif
Figure 3.
PFGE patterns of SfiI-cleaved genomic DNAs from L. pneumophila Jeju isolates (A) and their schematic four band patterns (PG1 to PG4) (B). Lambda ladder PFGE marker (M) were used as molecular size standards.
jbv-40-111f3.tif
Figure 4.
RAPD typing of L. pneumophila Jeju isolates. Six different band patterns (Type R1 to R6) were observed (B). ATCC 33152, L. pneumophila serogroup 1; M, 100-bp ladder marker; N, negative control.
jbv-40-111f4.tif
Table 1.
Primers to amplify partial fragments of target genes of Legionella isolates
Target gene Primers Size of amplicon (bp) Reference
rpoB RL1 (5_-GAT GAT ATC GAT CAY CTD GG-3) RL2 (5-TTC VGG CGT TTC AAT NGG AC-3 369 28
mip ML1 (5-GATAAG TTG TCT TAT AGC ATT GG-3) ML2 (5-TCTGTC CAT CCT GGG ATA ACT TG-3) 402 29
dotA DL1 (5-TTG ATT TGG TGA AAC TCA ATG G-3) DL2 (5-CAA TCA AAA TCC TGG TGC TTC-3) 434 28
Table 2.
Distribution of L. pneumophila Jeju isolates based on the rpoB and dotA sequences, and RAPD and PFGE patterns
Isolates Subgroup PR type Serogroup
rpoB dotA rpoBa dotAa
Jeju 1-1 P-I P-I P3 R1 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 1-2 P-I P-I P3 R5 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 1-3 P-I P-I ND SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 2 P-I P-I P2 R1 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 3-1 P-I P-I P3 R3 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 3-2 P-I P-I P3 R3 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 4-1 P-I P-I P2 R1 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 4-2 P-I P-I P2 R1 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 4-3 P-I P-I ND SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 5-3 P-I P-I ND SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 6-1 P-I P-I P2 R1 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 6-2 P-I P-I P2 R4 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 6-3 P-III P-III P4 R6 SG 2, SG 6, SG 12 SG 12
Jeju 6-5 P-I P-I P2 R2 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 6-6 P-I P-I P2 R2 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 6-7 P-I P-I P1 R4 SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 7-1 P-I P-I ND SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 8-1 P-IV P-IV ND SG 7, SG 14 (99.3) SG 14 (97.5)
Jeju 8-2 P-I P-I ND SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 8-3 P-I P-I ND SG 1-4 SG 1-1, SG 1-4, SG 9
Jeju 8-4 P-I P-I ND SG 1-4 SG 1-1, SG 1-4, SG 9

a 100% Similarity, ND: Not determined

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