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The Escherichia coli O4 serotype is among those commonly isolated from urinary tract infections. In order to study the genetics of the O-antigen, the O4 biosynthesis genes from a uropathogenic E. coli have previously been cloned into E. coli K-12. A subclone, GH58, has been identified which reacts with antisera against the O4 serotype. In contrast to the wild-type parental strain, lipopolysaccharide (LPS) from this clone is devoid of rhamnose and does not cross-react with O18 antisera. The recombinant plasmid from GH58, pGH58, was used to transform the rfb deletion strain HU1190. The resultant strain agglutinates in O4 antisera, but produces unpolymerized LPS. Escherichia coli K-12 strains HB101 and RC712 containing pGH58 produce polymerized LPS, indicating that the genetic background of the host can influence the LPS encoded by recombinant molecules. A cosmid, pGH84, has been identified which encompasses the entire pGH58 gene sequences and includes an additional 34 kilobases of DNA. HU1190 containing this cosmid agglutinates in O4 antisera and produces a polymerized LPS. By constructing several deletion subclones of pGH84, we have localized the genes necessary for polymerized LPS to a 5.5 kb ClaI-BamHI fragment. P1 transductants that make polymerized and unpolymerized O4 LPS have also been identified.

The genetic organization and functions of the Shigella dysenteriae 1 rfb gene cluster, which specifies the somatic O antigen in this organism, have been studied in Escherichia coli K-12 by insertion and deletion mutagenesis of pSS9, a pBR322 hybrid containing the Shigella rfb genes. On the basis of the sensitivity/resistance to rough-specific bacteriophage T3 of E. coli K-12 derivatives containing mutant pSS9 plasmids, of the banding patterns and immunoreactivity of LPS isolated from such derivatives and electrophoresed on SDS-polyacrylamide gels, and of the sugar composition of the polysaccharide portion of the LPS determined by chemical analysis, six determinants for O antigen production were identified and localized. At least two determinants are involved in synthesis of TDP-rhamnose and the transfer of a rhamnose residue to the galactose-substituted core. One of these functions is probably TDP-rhamnose synthetase. A third function effects the transfer of a second rhamnose residue to the rha----gal-substituted core. A fourth function, for which evidence was obtained for two determinants (cistrons), is N-acetylglucosamine transferase, whereas a sixth determinant is necessary for extension of the first completed side chain repeat unit to the full O antigen polymer. These results confirmed the previously-determined chemical composition of the S. dysenteriae 1 O antigen and demonstrated that the order of the sugars is glcNAc----rha----rha----gal with gal as the first sugar linked to the core. Evidence was obtained for at least two transcriptional units in the rfb gene cluster and the approximate locations of two promoters are suggested. The detection of new electrophoretic species of LPS that may correspond to LPS biosynthetic intermediates, and the finding on the cell surfaces of structures corresponding to LPS core substituted with one or more O-specific sugars, appear to be novel findings.

Production of the somatic antigen, O-specific polysaccharide of Shigella dysenteriae 1 is determined by the chromosomal rfb gene cluster and the rfp gene located on the 9 kb plasmid pHW400 carried by this organism. When transferred to Escherichia coli K-12, which produces lipopolysaccharide consisting only of core oligosaccharide linked to lipid A, rfp gene-containing plasmids caused modification of the core oligosaccharide leading to the appearance of core molecules with new electrophoretic mobilities. Chemical analysis of the modified core has shown that it is substituted with a galactose residue which is the first sugar of the O-polysaccharide repeat unit.

To test the clinical counterpart of the learned nonuse theory, 25 chronic hemiplegic stroke and head-injured patients with minimal to moderate upper extremity extensor muscle function were required to keep their uninvolved upper extremities within a hand-enclosed sling during waking hours over a 2-week interval. During this forced use period and for 1 year thereafter, changes in force or time-based measures among 21 functional tasks were compared to values at the sixth baseline session, a preintervention time when relearning had plateaued. Significant (P less than 0.05, Friedman's repeated measures followed by Tukey multiple comparison tests) changes were seen in 19 of the 21 tasks with most persisting at the 1-year follow-up. There were no apparent differences between right- and left-sided involvement or between stroke versus head injury clients (Mann-Whitney procedure). Ratings for quality of movement scored from videotapes presented in random order showed no change over time. These data suggest that learned nonuse does occur in select neurological patients and that this behavior can be reversed through application of a forced use paradigm.