A single dose (5 x 108 organisms) of attenuated A-B' Vibrio cholerae classical Inaba recombinant vaccine strain CVD 103-HgR or placebo was administered to 24 healthy young Thai adults in a randomized, placebo-controlled, double-blind trial of safety and immunogenicity. None of the volunteers experienced untoward reactions. The vaccine strain was recovered from 2 of 12 vaccinees. The vibriocidal antibody response (the best immunological correlate of protection) was good: 11 of 12 vaccinees (92%) manifested significant serotype-homologous Inaba antibody rises with a peak reciprocal geometric mean titer (RGMT) postvaccination of 3,417; 9 of 12 exhibited significant serotype-heterologous Ogawa antibody rises (prevaccination RGMT, 180; peak RGMT, 2,874). Nine of 12 vaccinees had significant rises in serum antitoxin. None of the controls exhibited rises in vibriocidal or antitoxic antibody. This preliminary study further confirms the safety and immunogenicity of CVD 103-HgR live oral cholera vaccine and paves the way for larger community zation with a combined B subunit/whole cell vaccine. J. Infect.

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Heat-stable enterotoxigenic Escherichia coli was identified by nucleotide hybridization with RNA transcripts ot the gene encoding heat-stable A-2 enterotoxin. Radiolabeled enterotoxin gene RNA transcripts are easier to prepare and avoid the preparation of cloned DNA probes that can be nonspecific if they contain cloning vector DNA. Enterotoxigenic Escherichia coli (ETEC) is usually identified by testing isolates for enterotoxin production in bioassays or serological tests (2, 7, 11, 14, 15). Alternatively, ETEC can be identified by detecting the genes encoding these enterotoxins by DNA hybridization, a method which has been particularly useful in detecting ETEC in large numbers of specimens (3-6). Specific DNA fragments of plasmids containing the cloned enterotoxin genes have been used in DNA hybridization assays to identify ETEC (4, 5). Probes produced by endonuclease digestion must be electrophoretically separated from plasmid-cloning vector DNA to obtain a specific probe and must also be nick translated (9). Nick translation is relatively easy to perform with large DNA fragments, such as the 850-base-pair fragment used as the heat-labile enterotoxin (LT) gene probe, but is more difficult to perform on small DNA probes used to detect the genes encoding heatstable enterotoxin (STA-1 [154 base pairs] or STA-2 [216 base pairs]). Ninety-nine percent of E. coli that hybridized with the LT probe produced LT, as measured by the Y-1 adrenal cell assay, but only 72% of E. coli that hybridized with the STA-2 probe produced ST, as measured by the suckling mouse assay (5). The lack of specificity of the cloned enterotoxin gene probes was due to probes that contained cloning vector DNA. Niriety-eight percent of E. coli that hybridized with the ST oligo probe produced ST (6). To develop a more convenient probe to identify genes encoding STA-2, we clotied the genes encoding STA-2 into pSP64 and used RNA transcripts to detect ST+ ETEC by colony hybridization. Plasmid DNA was isolated from E. coli C600(pSLM004) and was digested with EcoRI, HindIII, and HpaII (Bethesda Research Laboratories, Inc., Gaithersburg, Md.) as previously described (10). The endontclease digestion fragments were separated by electrophoresis on a 1.5% low-meltingtemperature agarose gel (FMC Corp., Marine Colloids Div., Rockland, Maine) in Tris-borate buffer, and a 216-base-pair DNA fragment encoding STA-2 was éfluted by heat, phenol extracted, and ethanol precipitated (8). This DNA fragment was blunt-end ligated with pSP64 (Promega Biotec, Madison, Wis.