Helicobacter pylori, the causative agent of type B gastritis, peptic ulcers, gastric adenocarcinoma and MALT lymphoma, uses the Cag type IV secretion system to induce a strong proinflammatory response in the gastric mucosa and to inject its effector protein CagA into gastric cells. CagA translocation results in altered host cell gene expression profiles and cytoskeletal rearrangements, and it is considered as a major bacterial virulence trait. Recently, it has been shown that binding of the type IV secretion apparatus to integrin receptors on target cells is a crucial step in the translocation process. Several bacterial proteins, including the Cag-specific components CagL and CagI, have been involved in this interaction. Here, we have examined the localization and interactions of CagI in the bacterial cell. Since the cagI gene overlaps and is co-transcribed with the cagL gene, the role of CagI for type IV secretion system function has been difficult to assess, and conflicting results have been reported regarding its involvement in the proinflammatory response. Using a marker-free gene deletion approach and genetic complementation, we show now that CagI is an essential component of the Cag type IV secretion apparatus for both CagA translocation and interleukin-8 induction. CagI is distributed over soluble and membrane-associated pools and seems to be partly surface-exposed. Deletion of several genes encoding essential Cag components has an impact on protein levels of CagI and CagL, suggesting that both proteins require partial assembly of the secretion apparatus. Finally, we show by co-immunoprecipitation that CagI and CagL interact with each other. Taken together, our results indicate that CagI and CagL form a functional complex which is formed at a late stage of secretion apparatus assembly.

A metal-organic framework (MOF) for reversible alteration of guest molecule adsorption, here carbon dioxide, upon photochemical or thermal treatment has been discovered. An azobenzene functional group, which can switch its conformation upon light irradiation or heat treatment, has been introduced to the organic linker of a MOF. The resulting MOF adsorbs different amount of CO2 after UV or heat treatment. This remarkable stimuli-responsive adsorption effect has been demonstrated through experiments.

Trichinellosis outbreaks occur occasionally in Vietnam following the consumption of undercooked pork. Diagnosing trichinella can be problematic because fever and myalgia are nonspecific, and diagnosis may be delayed. We describe 5 Vietnamese patients in whom trichinellosis was diagnosed after several weeks of illness.

We describe a mathematical model and Monte-Carlo (MC) simulation of viral evolution during acute infection. We consider both synchronous and asynchronous processes of viral infection of new target cells. The model enables an assessment of the expected sequence diversity in new HIV-1 infections originating from a single transmitted viral strain, estimation of the most recent common ancestor (MRCA) of the transmitted viral lineage, and estimation of the time to coalesce back to the MRCA. We also calculate the probability of the MRCA being the transmitted virus or an evolved variant. Excluding insertions and deletions, we assume HIV-1 evolves by base substitution without selection pressure during the earliest phase of HIV-1 infection prior to the immune response. Unlike phylogenetic methods that follow a lineage backwards to coalescence, we compare the observed data to a model of the diversification of a viral population forward in time. To illustrate the application of these methods, we provide detailed comparisons of the model and simulations results to 306 envelope sequences obtained from 8 newly infected subjects at a single time point. The data from 6/8 patients were in good agreement with model predictions, and hence compatible with a single-strain infection evolving under no selection pressure. The diversity of the samples from the other two patients was too great to be explained by the model, suggesting multiple HIV-1-strains were transmitted. The model can also be applied to longitudinal patient data to estimate within-host viral evolutionary parameters.

The precise identification of the HIV-1 envelope glycoprotein (Env) responsible for productive clinical infection could be instrumental in elucidating the molecular basis of HIV-1 transmission and in designing effective vaccines. Here, we developed a mathematical model of random viral evolution and, together with phylogenetic tree construction, used it to analyze 3,449 complete env sequences derived by single genome amplification from 102 subjects with acute HIV-1 (clade B) infection. Viral env genes evolving from individual transmitted or founder viruses generally exhibited a Poisson distribution of mutations and star-like phylogeny, which coalesced to an inferred consensus sequence at or near the estimated time of virus transmission. Overall, 78 of 102 subjects had evidence of productive clinical infection by a single virus, and 24 others had evidence of productive clinical infection by a minimum of two to five viruses. Phenotypic analysis of transmitted or early founder Envs revealed a consistent pattern of CCR5 dependence, masking of coreceptor binding regions, and equivalent or modestly enhanced resistance to the fusion inhibitor T1249 and broadly neutralizing antibodies compared with Envs from chronically infected subjects. Low multiplicity infection and limited viral evolution preceding peak viremia suggest a finite window of potential vulnerability of HIV-1 to vaccine-elicited immune responses, although phenotypic properties of transmitted Envs pose a formidable defense.

Accurate identification of the transmitted virus and sequences evolving from it could be instrumental in elucidating human immunodeficiency virus type 1 (HIV-1) transmission and in developing vaccines, drugs or microbicides to prevent infection. Here we describe an experimental approach to analyze HIV-1 env genes as intact genetic units from plasma virion RNA by single genome amplification (SGA) followed by direct sequencing of uncloned DNA amplicons. We show that this strategy precludes in vitro artifacts caused by Taq-induced nucleotide substitutions and template switching, provides an accurate representation of the env quasispecies in vivo, and has an overall error rate (including nucleotide misincorporation and insertion-deletion) of less than 8 x 10(-5). Applying this method to the analysis of plasma virus from 12 Zambian subjects who were sampled within 3 months of seroconversion, we show that transmitted or early founder viruses can be identified and that molecular pathways and rates of early env diversification can be defined. Specifically, we show that 8 of the 12 subjects were infected by a single virus, while 4 others acquired more than one virus; that the rate of virus evolution in one subject during an 80 day period spanning seroconversion was 1.7 x 10(-5) substitutions per site per day; and that evidence of strong immunologic selection can be seen in Env and overlapping Rev sequences based on nonrandom accumulation of nonsynonymous mutations. We also compared the SGA approach with more conventional bulk PCR amplification methods on the same patient samples and found that the latter is associated with excessive rates of Taq-induced recombination, nucleotide misincorporation, template resampling and cloning bias. These findings indicate that HIV-1 env genes, other viral genes, and even full-length viral genomes responsible for productive clinical infection can be identified by SGA analysis of plasma virus sampled at intervals typical of large scale vaccine trials and that pathways of viral diversification and immune escape can be determined accurately.

OBJECTIVES: To outline current practice regarding the prehospital use of subcutaneous epinephrine, and systematically review the existing literature to determine the level of support for its use in the elderly. Many health care personnel are reluctant to administer subcutaneous epinephrine for potentially life-threatening conditions such as asthma and anaphylaxis in older patients. This sytematic review examined the following focused question:"For older patients not known to have coronary artery disease, does administration of subcutaneous epinephrine carry a significant enough risk of cardiovascular side effects to mandate age as a relative contraindication to self-administration or emergency medical services administration in the prehospital setting?" METHODS: The MEDLINE and Health Star databases were searched to identify studies evaluating the use of subcutaneous epinephrine in the treatment of asthma and anaphylaxis. Bibliographies from included studies, known reviews, and textbooks were examined to identify additional studies. The strength of evidence presented in each study was assessed in accordance with the classification system proposed by the American Heart Association's Emergency Cardiovascular Care Committee. RESULTS: The review of the literature revealed only three case reports (level VII evidence) that record adverse reactions of epinephrine when used for anaphylaxis and allergy, while several level III and V studies found no adverse effects when used for asthma. No controlled trials documenting adverse effects were found. CONCLUSIONS: The authors did not find significant evidence to contraindicate the use of subcutaneous epinephrine in older patients who are not known to have coronary artery disease, who present with either asthma or allergic reactions.

OBJECTIVES: To characterize reproductive hormone levels, symptoms, and attitudes related to menopause among healthy, menstruating white and African-American women aged 44 to 49 years. DESIGN: Pilot study; cross-sectional survey. SETTING: Community-based convenience sample of women in the Philadelphia metropolitan area. PARTICIPANTS: Thirty-three African-American and 35 white women. MEASUREMENTS: The survey instrument collected demographic data, medical and reproductive history, health practices and behaviors. It included previously validated function, depression, and quality-of-life instruments, and a Menopause Attitude Scale that included two factors, attitudes toward the menopause and attitudes toward medical therapy. Anthropometric measurements were taken at enrollment, and reproductive hormones and daily symptom logs were followed over two menstrual cycles. MAIN RESULTS: The two groups were comparable in mean age (African-American 46.2 years, white 46.9 years). Serum levels of estradiol, follicle-stimulating hormone, dihydroepiandrosterone-sulfate, and progesterone were comparable. Symptoms were similar in type and frequency. However, the African-American women had significantly more positive attitudes toward menopause, were more likely to rely on family for information about menopause, and were less likely to have been recommended hormone replacement therapy by their physicians. A majority of women in each group expressed satisfaction with the care they had received. CONCLUSIONS: Perimenopausal African-American and white women have different expectations of menopause and the role of medical care in menopause. This bears directly on women's acceptance of hormone replacement therapy. Conclusions are limited by the small sample size and convenience nature of the study population: further work with larger samples is needed to confirm these apparent differences.

Plasma transthyretin (TTR, formerly called prealbumin) is a 55-kd protein that participates in the plasma transport of both thyroxine and retinol (vitamin A). TTR concentrations are disproportionately high in human ventricular CSF, suggesting that TTR is either selectively transported across or synthesized de novo within the blood-CSF barrier. To address this question, we adopted a molecular genetic approach; after isolating a cDNA clone encoding human TTR, we previously demonstrated specific TTR messenger RNA (mRNA) synthesis in rat choroid plexus. We have now extended these investigations to the human brain. Northern analysis of postmortem brain homogenates revealed abundant TTR mRNA in choroid plexus, but not in cerebellum or cerebral cortex. Choroid plexus mRNA was readily translated into TTR preprotein in an in vitro translation system. An immunocytochemical survey of human postmortem brain sections revealed the presence of TTR protein specifically and uniquely in the cytoplasm of choroid plexus epithelial cells; these results were corroborated at the mRNA level by an extensive survey of whole rat-brain sections by in situ hybridization. Therefore, within the mammalian CNS, TTR is the first known protein synthesized solely by the choroid plexus, suggesting a special role for TTR in the brain or CSF. Whether this function differs from its established plasma transport functions is presently unknown.