title: Comparative toxicity and virulence of Escherichia coli clones expressing variant and chimeric Shiga-like toxin type II operons.
abstract: Shiga-like toxin (SLT)-producing strains of Escherichia coli are known to cause diarrhea, hemorrhagic colitis, and hemolytic-uremic syndrome in humans. The SLTs, particularly those related to type II (SLT-II), are a diverse family of toxins which may have differing in vitro or in vivo properties. To examine the impact of naturally occurring SLT-II sequence variation on the capacity of a given E. coli strain to cause disease, operons encoding four different SLT-II-related toxins, designated SLT-II/O111, SLT-II/OX3a, SLT-II/OX3b, and SLT-II/O48, were cloned in the same orientation in pBluescript. French pressure cell lysates of E. coli DH5 alpha derivatives carrying these plasmids differed markedly in cytotoxicity for Vero cells, with 50% cytotoxic doses ranging from 20 to 328,000/ml. The strains also differed in oral virulence for streptomycin-treated mice, as judged by survival rate and/or median survival time, but virulence did not necessarily correlate with in vitro cytotoxicity. The SLT-II type associated with the lowest oral virulence was SLT-II/O111. Both the overall survival rate and the median survival time of mice challenged with clones producing this toxin were significantly greater than that for mice challenged with a clone producing the closely related SLT-II/OX3a. Experiments with clones carrying chimeric O111/OX3a SLT-II operons indicated that the reduced virulence was associated with an Arg-176-->Gly substitution in the mature A subunit. Clones producing SLT-II/O48 and SLT-II/OX3b had similarly high cytotoxicities for Vero cells, but the latter was more virulent when fed to streptomycin-treated mice, as judged by median survival time. Experiments with clones carrying chimeric O48/OX3b SLT-II operons indicated that the increased virulence was a function of the A subunit of SLT-II/OX3b, which differs from the A subunit of SLT-II/O48 by only two amino acids (Met-4-->Thr and Gly-102-->Asp, respectively). These findings raise the possibility that naturally occurring SLT-II sequence variations may impact directly on the capacity of a given SLT-producing E. coli strain to cause disease.
authors.affiliation: Department of Microbiology, Women's and Children's Hospital, North Adelaide, South Australia.
authors.forename: A W
authors.initials: AW
authors.lastname: Paton
authors.forename: A J
authors.initials: AJ
authors.lastname: Bourne
authors.forename: P A
authors.initials: PA
authors.lastname: Manning
authors.forename: J C
authors.initials: JC
authors.lastname: Paton
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001427:Bacterial Toxins / Q000633:toxicity*; D001483:Base Sequence; D002522:Chlorocebus aethiops; D004768:Enterotoxins / Q000633:toxicity*; D004926:Escherichia coli / Q000472:pathogenicity*; D005798:Genes, Bacterial; D006367:HeLa Cells; D006801:Humans; D007668:Kidney / Q000473:pathology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015345:Oligonucleotide Probes / Q000737:chemistry; D009876:Operon; D011993:Recombinant Fusion Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D022641:Shiga Toxin 2; D013329:Structure-Activity Relationship; D014709:Vero Cells
chemical_list: D001427:Bacterial Toxins; D004768:Enterotoxins; D015345:Oligonucleotide Probes; D011993:Recombinant Fusion Proteins; D022641:Shiga Toxin 2