Genetic characterization of Escherichia coli O157: H7/- strains carrying the stx2 gene but not producing Shiga toxin 2.
Nine Escherichia coli O157: H7/- strains isolated primarily from non-clinical sources in Thailand and Japan carried the stx(2) gene but did not produce Stx2 toxin in a reversed passive latex agglutination (RPLA) assay. A strain (EDL933) bearing a stx(2) phage (933W) was compared to a strain (Thai-12) that was Stx2-negative but contained the stx(2) gene. To study the lack of Stx2 production, the Thai-12 stx(2) gene and its upstream nucleotide sequence were analyzed. The Thai-12 stx(2) coding region was intact and Stx2 was expressed from a cloned stx(2) gene using a plasmid vector and detected using RPLA. A lacZ fusion analysis found the Thai-12 stx(2) promoter non-functional. Because the stx(2) gene is downstream of the late promoter in the stx(2) phage genome, the antitermination activity of Q protein is essential for strong stx(2) transcription. Thai-12 had the q gene highly homologous to that of Phi21 phage but not to the 933W phage. High-level expression of exogenous q genes demonstrated Q antitermination activity was weak in Thai-12. Replication of stx(2) phage was not observed in Stx2-negative strains. The q-stx(2) gene sequence of Thai-12 was well conserved in all Stx2-negative strains. A PCR assay to detect the Thai-12 q-stx(2) sequence demonstrated that 30% of O157 strains from marketed Malaysian beef carried this sequence and they produced little or no Stx2. These results suggest that stx(2)-positive O157 strains that produce little or no Stx2 may be widely distributed in the Asian environment.
Koitabashi
Tsutomu
T
Graduate School of Medicine, Kyoto University, Japan.
Vuddhakul
Varaporn
V
Radu
Son
S
Morigaki
Tadaaki
T
Asai
Norio
N
Nakaguchi
Yoshitsugu
Y
Nishibuchi
Mitsuaki
M
D000818:Animals; D010582:Bacteriophage lambda / Q000235:genetics; D001483:Base Sequence; D002417:Cattle; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D016521:Electrophoresis, Gel, Pulsed-Field; D004927:Escherichia coli Infections / Q000382:microbiology*; D019453:Escherichia coli O157 / Q000235:genetics* / Q000378:metabolism*; D005516:Food Microbiology; D006801:Humans; D007841:Latex Fixation Tests; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D022641:Shiga Toxin 2 / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics
D004269:DNA, Bacterial; C066119:Q protein, Bacteriophage lambda; D022641:Shiga Toxin 2; D014764:Viral Proteins