"Slow VISA," a novel phenotype of vancomycin resistance, found in vitro in heterogeneous vancomycin-intermediate Staphylococcus aureus strain Mu3.
Heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) clinical strain Mu3 spontaneously generates VISA strains at an extremely high frequency (≥1×10(-6)). The generated VISA strains usually grow more slowly than does the parent hVISA strain, but they form colonies on vancomycin-containing agar plates before 48 h of incubation. However, we noticed a curious group of VISA strains, designated "slow VISA" (sVISA), whose colonies appear only after 72 h of incubation. They have extremely prolonged doubling times but have vancomycin MICs of 8 to ∼24 mg/liter when determined after 72 to ∼144 h of incubation. We established strain Mu3-6R-P (6R-P), which has a vancomycin MIC of 16 mg/liter (at 72 h), as a representative sVISA strain. Its cell wall was thickened and autolytic activity was decreased compared to the respective qualities of the parent hVISA strain Mu3. Whole-genome sequencing of 6R-P revealed only one mutation, encoded by rpoB (R512P), which replaced the 512th arginine of the RNA polymerase β-subunit with proline. Its VISA phenotype was unstable, and the strain frequently reverted to hVISA with concomitant losses of pinpoint colony morphology and cell wall thickness and reduced autolytic activity. Sequencing of the rpoB genes of the phenotypic revertant strains revealed mutations affecting the 512th codon, where the proline of 6R-P was replaced with leucine, serine, or histidine. Slow VISA generated in the tissues of an infected patient serves as a temporary shelter for hVISA to survive vancomycin therapy. The sVISA strain spontaneously returns to hVISA when the threat of vancomycin is lifted. The rpoB(R512P) mutation may be regarded as a regulatory mutation that switches the reversible phenotype of sVISA on and off.
Department of Infection Control Science, Graduate School of Medicine, Juntendo University, Tokyo, Japan.
Michie
M
Saito
Department of Bacteriology, Faculty of Medicine, Juntendo University, Tokyo, Japan.
Yuki
Y
Katayama
Department of Bacteriology, Faculty of Medicine, Juntendo University, Tokyo, Japan.
Tomomi
T
Hishinuma
Department of Infection Control Science, Graduate School of Medicine, Juntendo University, Tokyo, Japan.
Akira
A
Iwamoto
Department of Infection Control Science, Graduate School of Medicine, Juntendo University, Tokyo, Japan.
Yoshifumi
Y
Aiba
Department of Bacteriology, Faculty of Medicine, Juntendo University, Tokyo, Japan.
Kyoko
K
Kuwahara-Arai
Department of Bacteriology, Faculty of Medicine, Juntendo University, Tokyo, Japan.
Longzhu
L
Cui
Department of Bacteriology, Faculty of Medicine, Juntendo University, Tokyo, Japan.
Miki
M
Matsuo
Division of Hematology, Department of Medicine, Juntendo University Nerima Hospital, Tokyo, Japan.
Nanae
N
Aritaka
Department of Infection Control Science, Graduate School of Medicine, Juntendo University, Tokyo, Japan Department of Bacteriology, Faculty of Medicine, Juntendo University, Tokyo, Japan khiram06@juntendo.ac.jp.
Keiichi
K
Hiramatsu
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics; D002473:Cell Wall / Q000187:drug effects / Q000235:genetics / Q000382:microbiology; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D016680:Genome, Bacterial / Q000235:genetics; D008826:Microbial Sensitivity Tests / Q000379:methods; D009154:Mutation / Q000235:genetics; D010641:Phenotype; D013211:Staphylococcus aureus / Q000187:drug effects* / Q000235:genetics*; D014640:Vancomycin / Q000494:pharmacology*; D020713:Vancomycin Resistance / Q000235:genetics*
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D014640:Vancomycin; D012321:DNA-Directed RNA Polymerases; C105078:RNA polymerase beta subunit