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Evidence of transient HIV infections was found in 8 subjects at high-risk for HIV infection among 47 longitudinally studied over 2-5 (average approximately 3.5) years, whereas only two subjects developed progressive infection. All of these subjects developed serum antibodies (Ab) to conformational epitopes of HIV gp41 (termed "early HIV Ab"), but the 8 transiently infected subjects lost this Ab within 4-18 months, and did not seroconvert to positivity in denatured antigen EIA or Western Blot (WB). However, the two progressively infected subjects eventually seroconverted in the EIA and WB tests within one to two months after the appearance of "early HIV Ab". HIV env and nef sequences were directly PCR amplified from the peripheral blood mononuclear cells (PBMCs) of two of the eight transiently infected subjects during the time of "early HIV Ab"-postivity, and these showed significant sequence divergence from the HIV strains in the laboratory, indicating that they were not laboratory contaminants. Genome identity typing ("paternity-typing") of PBMC samples obtained at the time of "early HIV Ab"-positivity, and later when Ab was absent from each of the 8 subjects, showed that blood samples were not mixed-up. This provides further evidence that transient or occult infection with HIV does occur, and perhaps at a greater frequency than do progressive infections.

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Uranyl nitrate inhibited root growth of Allium cepa at > or = 25 microM concentration. Fluorimetric analysis of metal uptake indicated the entry and accumulation of uranium into the root cell. Uranyl nitrate was neither clastogenic nor aneugenic as it failed to induce micronuclei significantly, but between 25 and 100 microM concentration, it increased significantly the frequency of sister chromatid exchange over that of control, implying its genotoxicity that possibly interfered with DNA replication and/or repair process.

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The functional significance of naturally occurring variants of human hepatitis B virus (HBV) remains largely unknown. Previously, we reported an immature secretion phenotype caused by a highly frequent mutation at amino acid 97 of the HBV core (capsid) protein (HBcAg). This phenotype is characterized by a nonselective and excessive secretion of virions containing an immature genome of single-stranded viral DNA. To extend our study of virion secretion to other naturally occurring variants, we have characterized mutations at HBcAg codons 5, 38, and 60 via site-directed mutagenesis. Although the phenotype of the mutation at codon 38 is nearly identical to that for the wild-type virus, our study reveals that a single mutation at codon 5 or 60 exhibits a new extracellular phenotype with significantly reduced virion secretion yet maintains normal intracellular viral DNA replication. A complementation study indicates that the mutant core protein alone is sufficient for the "low-secretion" phenotype. Furthermore, the low-secretion phenotype of the codon 5 mutant appears to be induced by the loss of a parental proline residue, rather than by the gain of a new amino acid. Our study underscores the core protein as another crucial determinant in virion secretion, in addition to the known envelope proteins. Our present results suggest that a very precise structure of both alpha-helical and nonhelical loop regions of the entire HBcAg molecule is important for virion secretion. The low-secretion variants may contribute to the phenomenon of gradually decreasing viremia in chronic carriers during the late phase of persistent infection.

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A very frequent missense mutation at codon 97 of human hepatitis B virus (HBV) core antigen (HBcAg) has been found in chronic carriers worldwide. Functional characterization of this mutant revealed one intracellular and two extracellular phenotypes in contrast to wild-type HBV: (i) a 6- to 12-fold decrease in the level of the full-length relaxed circular DNA, a 4- to 5-fold decrease in the plus-strand DNA, and an approximately 1.8-fold decrease in the minus-strand and overall DNA levels in the intracellular viral core particles; (ii) a 5.7-fold increase in the immature secretion of Dane particles, containing minus-strand, single-stranded virion DNA; and (iii) a significant reduction of nonenveloped core particles in the medium. The steady-state levels of mutant and wild-type core proteins expressed from the same vector appeared to be similar. Using a complementation assay and gradient centrifugation analysis, we demonstrated that this mutant core protein alone is necessary and sufficient for immature secretion. The decreased level of intracellular HBV DNA is caused by both the cis defect of the mutant genome and the trans defect of the mutant core protein. We have dissected further the relationship between the intracellular and extracellular phenotypes of mutant F97L. The pleiotropic effects of the HBcAg codon 97 mutation were observed consistently in several different experimental settings. The mechanism and biological significance of these findings are discussed.

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The Vibrio cholerae rpoH gene coding for the heat-shock sigma factor, sigma 32, has been cloned and shown to functionally complement Escherichia coli rpoH mutants. The nt sequence of the gene has been determined and the deduced aa sequence is more than 80% homologous to the E. coli rpoH gene product. Downstream of the V. cholerae rpoH gene, an unidentified dehydrogenase gene (udhA) is present on the opposite strand facing rpoH. The predicted secondary structure of the 5'-proximal region of V. cholerae rpoH mRNA is apparently different from the conserved secondary structures of the rpoH mRNA reported for several bacterial species. The 'RpoH box', a stretch of 9 aa (QRKLFFNLR) unique to sigma 32 factors, and the 'downstream box' sequence complementary to a part of the 16S rRNA, have been detected.

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Sixteen heat shock proteins (Hsps) have been identified in the hypertoxinogenic strain 569B of Vibrio cholerae which are synthesized in response to small and large elevations of temperature. The induction of the Hsps is necessary for the cells to survive the deleterious effects of heat. There is no difference in the pattern of induction of the Hsps in V. cholerae strains varying in levels of toxinogenicity. One of the major low-molecular-mass Hsps, a 16-kDa protein, is preferentially degraded following shift down of temperature. This protein is induced at a much lower level at high temperatures in cells maintained in the laboratory for a prolonged period. The only Hsp located in the outer membrane of V. cholerae cells is a 23-kDa protein. Western immunoblot analysis with human immune sera collected from convalescent cholera patients revealed that this protein is markedly immunogenic. The human immune serum also reacted with the 69- and 16-kDa major Hsps and the 88-, 66-, and 46-kDa Hsps but not with the 61-kDa major Hsp identified as the groEL gene product. All major Hsps reacted with rabbit anti-V. cholerae sera. Ethanol stress leads to the induction of four of the major Hsps and three additional proteins.

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