Molecular Subtyping Of Toxigenic Vibrio Cholerae O139 Causing Epidemic Cholera In India And Bangladesh, 1992-1993

Abstract

Since October 1992, > 150,000 cases of cholera have been reported from India and Bangladesh; the great majority of Vibrio cholerae isolates belong to the newly established serogroup O139. To better understand the interaction of genetic and epidemiologic factors responsible for their sudden appearance and rapid spread, representative toxigenic V. cholerae O139 isolates were molecularly characterized and compared with a set of toxigenic V. cholerae O1 and non-O1/non-O139 strains. DNA sequences of the cholera toxin B subunit gene and multilocus enzyme electrophoresis markers of V. cholerae O139 strains were identical to those of V. cholerae O1 isolates of the seventh pandemic. Two distinct ribotypes and four pulsed-field gel electrophoretic patterns were observed for O139 strains. V. cholerae O139 strains were very similar to V. cholerae O1 strains of the seventh pandemic but clearly different from the toxigenic V. cholerae strains of serogroups other than O1 and O139.

Transition State Structure for the Hydrolysis of NAD+Catalyzed by Diphtheria Toxin

“…[5][6][7][8][9][10][11][12][13][14][15][16][17][18] O]glucose was purified by sequential phosphorylation with hexokinase, anion-exchange chromatography, dephosphorylation with glucose 6-phosphatase, and repurification under the same conditions. Hexokinase and glucose 6-phosphatase were dialyzed against 50 mM potassium phosphate (pH 7.5) before use to remove unlabeled glucose.…”

“…Hexokinase and glucose 6-phosphatase were dialyzed against 50 mM potassium phosphate (pH 7.5) before use to remove unlabeled glucose. [5][6][7][8][9][10][11][12][13][14][15][16][17][18] O]Glucose (55 μmol in 2 mL) was phosphorylated to glucose 6-phosphate with 2 u of hexokinase, 60 mM ATP, and 60 mM MgCl 2 in 50 mM potassium phosphate (pH 7.5) for 2 h at room temperature. The reaction mixture was reduced to approximately half-volume under vacuum, applied to a 10 mL column of DEAE-Sephadex A-25 which had been equilibrated with H 2 O, then washed with 10 × 1 mL of H 2 O.…”

“…The reaction mixture was reduced to approximately half-volume under vacuum, applied to a 10 mL column of DEAE-Sephadex A-25 which had been equilibrated with H 2 O, then washed with 10 × 1 mL of H 2 O. [5][6][7][8][9][10][11][12][13][14][15][16][17][18] O]Glucose 6-phosphate was eluted with 10 × 1 mL of 1 M NH 4 OAc. The fractions containing product were identified using the reducing sugar assay, 33 pooled, frozen, and lyophilized twice.…”

“…The [5][6][7][8][9][10][11][12][13][14][15][16][17][18] O]glucose 6-phosphate was dephosphorlyated with 0.1 u of glucose 6-phosphatase in 50 mM potassium phosphate (pH 7.5) for 2 h at room temperature. Anion exchange chromatography was performed as above, with [5][6][7][8][9][10][11][12][13][14][15][16][17][18] 14 C]NAD +34 were synthesized as described previously, 31 with several modifications. The synthesis of nicotinic acid adenine dinucleotide (NaAD + ) was done in two steps to allow incorporation of the 14 C-label from [8][9][10][11][12][13][14] C]ATP.…”

“…The double primary, [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15] N,1′-14 C] KIE = 1.062 ± 0.010 (n = 2), was measured and found to be, within experimental error, equal to the product of the individual KIEs, 1.065. This relationship was also observed for the solvolytic hydrolysis of NAD + , where the measured [1-15 N,1′-14 C] KIE = 1.034 ± 0.002 (n = 6) was equal to the product of the individual KIEs, 1.036.…”