Identification and characterization of a newly isolated shiga toxin 2-converting phage from shiga toxin-producing Escherichia coli.
Shiga toxins 1 (Stx1) and 2 (Stx2) are encoded by toxin-converting bacteriophages of Stx-producing Escherichia coli (STEC), and so far two Stx1- and one Stx2-converting phages have been isolated from two STEC strains (A. D. O'Brien, J. W. Newlands, S. F. Miller, R. K. Holmes, H. W. Smith, and S. B. Formal, Science 226:694-696, 1984). In this study, we isolated two Stx2-converting phages, designated Stx2Phi-I and Stx2Phi-II, from two clinical strains of STEC associated with the outbreaks in Japan in 1996 and found that Stx2Phi-I resembled 933W, the previously reported Stx2-converting phage, in its infective properties for E. coli K-12 strain C600 while Stx2Phi-II was distinct from them. The sizes of the plaques of Stx2Phi-I and Stx2Phi-II in C600 were different; the former was larger than the latter. The restriction maps of Stx2Phi-I and Stx2Phi-II were not identical; rather, Stx2Phi-II DNA was approximately 3 kb larger than Stx2Phi-I DNA. Furthermore, Stx2Phi-I and Stx2Phi-II showed different phage immunity, with Stx2Phi-I and 933W belonging to the same group. Infection of C600 by Stx2Phi-I or 933W was affected by environmental osmolarity differently from that by Stx2Phi-II. When C600 was grown under conditions of high osmolarity, the infectivity of Stx2Phi-I and 933W was greatly decreased compared with that of Stx2Phi-II. Examination of the plating efficiency of the three phages for the defined mutations in C600 revealed that the efficiency of Stx2Phi-I and 933W for the fadL mutant decreased to less than 10(-7) compared with that for C600 whereas the efficiency of Stx2Phi-II decreased to 0.1% of that for C600. In contrast, while the plating efficiency of Stx2Phi-II for the lamB mutant decreased to a low level (0.05% of that for C600), the efficiencies of Stx2Phi-I and 933W were not changed. This was confirmed by the phage neutralization experiments with isolated outer membrane fractions from C600, fadL mutant, or lamB mutant or the purified His6-tagged FadL and LamB proteins. Based on the data, we concluded that FadL acts as the receptor for Stx2Phi-I and Stx2Phi-II whereas LamB acts as the receptor only for Stx2Phi-II.
Research Institute, International Medical Center of Japan, Tokyo 162, Japan.
M
M
Watarai
T
T
Sato
M
M
Kobayashi
T
T
Shimizu
S
S
Yamasaki
T
T
Tobe
C
C
Sasakawa
Y
Y
Takeda
D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism*; D002522:Chlorocebus aethiops; D003090:Coliphages / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology; D004926:Escherichia coli / Q000378:metabolism* / Q000821:virology; D029968:Escherichia coli Proteins; D050612:Fatty Acid Transport Proteins; D009994:Osmolar Concentration; D018272:Porins / Q000378:metabolism; D011991:Receptors, Virus / Q000235:genetics / Q000378:metabolism; D022641:Shiga Toxin 2; D014709:Vero Cells
D001425:Bacterial Outer Membrane Proteins; D001427:Bacterial Toxins; D029968:Escherichia coli Proteins; D050612:Fatty Acid Transport Proteins; C113724:OmpC protein; C109237:OmpF protein; D018272:Porins; D011991:Receptors, Virus; D022641:Shiga Toxin 2; C065691:fadL protein, E coli; C016532:maltoporins