N-Acetylation by hepatic arylamine N-acetyltransferase (NAT, EC 2.3.1.5) is a major route in the metabolism and detoxification of numerous drugs and foreign chemicals. NAT is the target of a common genetic polymorphism of clinical relevance in human populations. We have used our recently isolated rabbit cDNA rnat to clone three human NAT genes from human leukocyte DNA. None of the three genomic coding sequences was interrupted by introns. Two genes, designated NAT1 and NAT2, each possessed open reading frames of 870 bp. Both genes have been assigned to human chromosome 8, pter-q11. Following transfection they were transiently expressed in monkey kidney COS-1 cells. NAT1 and NAT2 gave rise to functional NAT proteins, as judged by their NAT enzyme activity with the arylamine substrate sulfamethazine. Western blots with NAT-specific antisera detected proteins of apparent molecular weight of 33 and 31 kD in NAT1- and NAT2-transfected cultures, respectively. The product of NAT2 had an identical apparent molecular weight as that of NAT detected in human liver cytosol. The deduced amino acid sequence of NAT2 also contained 6 peptide sequences which had previously been determined from tryptic peptides of the polymorphic NAT purified from human liver. These data suggest that NAT2 encodes the polymorphic NAT protein. The third gene, NATP, had multiple deleterious mutations and did not encode a functional NAT protein; it most likely represents a pseudogene.

To investigate hemodynamic and hormonal effects of ghrelin, a novel growth hormone (GH)-releasing peptide, we gave six healthy men an intravenous bolus of human ghrelin (10 microg/kg) or placebo and vice versa 1-2 wk apart in a randomized fashion. Ghrelin elicited a marked increase in circulating GH (15-fold). The elevation of GH lasted longer than 60 min after the bolus injection. Injection of ghrelin significantly decreased mean arterial pressure (-12 mmHg, P < 0.05) without a significant change in heart rate (-4 beats/min, P = 0.39). Ghrelin significantly increased cardiac index (+16%, P < 0.05) and stroke volume index (+22%, P < 0.05). We also examined ghrelin receptor [GH secretagogues receptor (GHS-R)] gene expression in the aortas, the left ventricles, and the left atria of rats by RT-PCR. GHS-R mRNA was detectable in the rat aortas, left ventricles, and left atria, suggesting that ghrelin may cause cardiovascular effects through GH-independent mechanisms. In summary, human ghrelin elicited a potent, long-lasting GH release and had beneficial hemodynamic effects via reducing cardiac afterload and increasing cardiac output without an increase in heart rate.

A major gene responsible for age-related hearing loss (AHL) in C57BL/6J mice was mapped by analyses of a (C57BL/6J x CAST/Ei) x C57BL/6J backcross. AHL, as measured by elevated auditory-evoked brainstem response (ABR) thresholds, segregated among backcross mice as expected for a recessive, primarily single-gene trait. Both qualitative and quantitative linkage analyses gave the same genetic map position for the AHL gene (Ahl on chromosome 10, near D10Mit5. Marker assisted selection was then used to produce congenic lines of C57BL/6J that contain different CAST-derived segments of chromosome 10. ABR test results and cochlear histopathology of aged progenitors of these congenic lines are presented. Ahl is the first gene causing late-onset, non-syndromic hearing loss that has been reported in the mouse.

Although there is a mounting body of evidence that eosinophils are recruited to sites of allergic inflammation by a number of beta-chemokines, particularly eotaxin and RANTES, the receptor that mediates these actions has not been identified. We have now cloned a G protein-coupled receptor, CC CKR3, from human eosinophils which, when stably expressed in AML14.3D10 cells bound eotaxin, MCP-3 and RANTES with Kds of 0.1, 2.7 and 3.1 nM, respectively. CC CKR3 also bound MCP-1 with lower affinity, but did not bind MIP-1 alpha or MIP-1 beta. Eotaxin, RANTES, and to a lessor extent MCP-3, but not the other chemokines, activated CC CKR3 as determined by their ability to stimulate a Ca(2+) -flux. Competition binding studies on primary eosinophils gave binding affinities for the different chemokines which were indistinguishable from those measured with CC CKR3. Since CC CKR3 is prominently expressed in eosinophils we conclude that CC CKR3 is the eosinophil eotaxin receptor. Eosinophils also express a much lower level of a second chemokine receptor, CC CKR1, which appears to be responsible for the effects of MIP-1 alpha.

BACKGROUND

Circulating C-reactive protein (CRP) is associated with the metabolic syndrome and might be causally linked to it. Our aim was to generate estimates of the association between plasma CRP and metabolic syndrome phenotypes that were free from confounding and reverse causation, to assess the causal role of this protein.

METHODS

We examined associations between serum CRP concentration and metabolic syndrome phenotypes in the British Women's Heart and Health Study. We then compared these estimates with those derived from a mendelian randomised framework with common CRP gene haplotypes to generate unconfounded and unbiased estimates of any causal associations.

RESULTS

In a sample of British women, body-mass index (BMI), systolic blood pressure, waist-to-hip ratio, serum concentrations of HDL cholesterol and triglycerides, and insulin resistance were all associated with plasma CRP concentration. CRP haplotypes were associated with plasma CRP concentration (p<0.0001). With instrumental variable analyses, there was no association between plasma CRP concentration and any of the metabolic syndrome phenotypes analysed. There was strong evidence that linear regression and mendelian randomisation based estimation gave conflicting results for the CRP-BMI association (p=0.0002), and some evidence of conflicting results for the association of CRP with the score for insulin resistance (p=0.0139), triglycerides (p=0.0313), and HDL cholesterol (p=0.0688).

CONCLUSIONS

Disparity between estimates of the association between plasma CRP and phenotypes comprising the metabolic syndrome derived from conventional analyses and those from a mendelian randomisation approach suggests that there is no causal association between CRP and the metabolic syndrome phenotypes.

While X-ray crystallography provides atomic resolution structures of proteins and small viruses, electron microscopy provides complementary structural information on the organization of larger assemblies at lower resolution. A novel combination of these two techniques has bridged this resolution gap and revealed the various structural components forming the capsid of human type 2 adenovirus. An image reconstruction of the intact virus, derived from cryo-electron micrographs, was deconvolved with an approximate contrast transfer function to mitigate microscope distortions. A model capsid was calculated from 240 copies of the crystallographic structure of the major capsid protein and filtered to the correct resolution. Subtraction of the calculated capsid from the corrected reconstruction gave a three-dimensional difference map revealing the minor proteins that stabilize the virion. Elongated density penetrating the hexon capsid at the facet edges was ascribed to polypeptide IIIa, a component required for virion assembly. Density on the inner surface of the capsid, connecting the ring of peripentonal hexons, was assigned as polypeptide VI, a component that binds DNA. Identification of the regions of hexon that contact the penton base suggests a structural mechanism for previously proposed events during cell entry.

Animal venom components are of considerable interest to researchers across a wide variety of disciplines, including molecular biology, biochemistry, medicine, and evolutionary genetics. The three-finger family of snake venom peptides is a particularly interesting and biochemically complex group of venom peptides, because they are encoded by a large multigene family and display a diverse array of functional activities. In addition, understanding how this complex and highly varied multigene family evolved is an interesting question to researchers investigating the biochemical diversity of these peptides and their impact on human health. Therefore, the purpose of our study was to investigate the long-term evolutionary patterns exhibited by these snake venom toxins to understand the mechanisms by which they diversified into a large, biochemically diverse, multigene family. Our results show a much greater diversity of family members than was previously known, including a number of subfamilies that did not fall within any previously identified groups with characterized activities. In addition, we found that the long-term evolutionary processes that gave rise to the diversity of three-finger toxins are consistent with the birth-and-death model of multigene family evolution. It is anticipated that this "three-finger toxin toolkit" will prove to be useful in providing a clearer picture of the diversity of investigational ligands or potential therapeutics available within this important family.

Loci for autosomal dominant "zonular pulverulent" cataract have been mapped to chromosomes 1q (CZP1) and 13q (CZP3). Here we report genetic refinement of the CZP3 locus and identify underlying mutations in the gene for gap-junction protein alpha-3 (GJA3), or connexin46 (Cx46). Linkage analysis gave a significantly positive two-point LOD score (Z) at marker D13S175 (maximum Z [Zmax]=>7.0; maximum recombination frequency [thetamax] =0). Haplotyping indicated that CZP3 probably lies in the genetic interval D13S1236-D13S175-D13S1316-cen-13pter, close to GJA3. Sequencing of a genomic clone isolated from the CZP3 candidate region identified an open reading frame coding for a protein of 435 amino acids (47,435 D) that shared approximately 88% homology with rat Cx46. Mutation analysis of GJA3 in two families with CZP3 detected distinct sequence changes that were not present in a panel of 105 normal, unrelated individuals. In family B, an A->>G transition resulted in an asparagine-to-serine substitution at codon 63 (N63S) and introduced a novel MwoI restriction site. In family E, insertion of a C at nucleotide 1137 (1137insC) introduced a novel BstXI site, causing a frameshift at codon 380. Restriction analysis confirmed that the novel MwoI and BstXI sites cosegregated with the disease in families B and E, respectively. This study identifies GJA3 as the sixth member of the connexin gene family to be implicated in human disease, and it highlights the physiological importance of gap-junction communication in the development of a transparent eye lens.

Incidence studies of Parkinson's disease (PD) are important for both health-care planning and epidemiological research. This report reviews the methods and results of previous incidence studies of PD and makes recommendations for future studies. Original articles that described the incidence of PD were located using several strategies. The methods were summarised, and the results of studies with similar methodologies were compared on a standardised population. Twenty-five incidence studies were included. Each used different methods to identify incident patients, although most screened both primary care and hospital records. Only eight studies were prospective, and only two of these had any follow-up. The diagnostic criteria for PD varied (11 studies used two or more cardinal motor features, four used the UK Brain Bank criteria), as did the exclusion criteria and the definition of an incident case. In 16 studies, attempts were made to confirm the diagnosis by examination of patients by a specialist as part of the study. None of the studies used identical methods, but five were sufficiently similar to merit comparison. Four of these gave a similar incidence (16-19/100000/year), but one from Italy had a much lower incidence (8.4/100000), the reason for which was unclear. Five studies found significantly greater incidence in men. This review highlights the difficulties in performing good quality incidence studies of PD. Further incidence studies using standardised methods are required. A set of minimal scientific criteria has been devised to improve the quality and consistency of future studies.

OBJECTIVE

To determine the effect of handwashing on the risk of respiratory infection.

METHODS

We searched PubMed, CAB Abstracts, Embase, Web of Science, and the Cochrane library for articles published before June 2004 in all languages. We had searched reference lists of all primary and review articles. Studies were included in the review if they reported the impact of an intervention to promote hand cleansing on respiratory infections. Studies relating to hospital-acquired infections, long-term care facilities, immuno-compromised and elderly people were excluded. We independently evaluated all studies, and inclusion decisions were reached by consensus. From a primary list of 410 articles, eight interventional studies met the eligibility criteria.

RESULTS

All eight eligible studies reported that handwashing lowered risks of respiratory infection, with risk reductions ranging from 6% to 44% [pooled value 24% (95% CI 6-40%)]. Pooling the results of only the seven homogenous studies gave a relative risk of 1.19 (95% CI 1.12%-1.26%), implying that hand cleansing can cut the risk of respiratory infection by 16% (95% CI 11-21%).

CONCLUSIONS

Handwashing is associated with lowered respiratory infection. However, studies were of poor quality, none related to developing countries, and only one to severe disease. Rigorous trials of the impact of handwashing on acute respiratory tract infection morbidity and mortality are urgently needed, especially in developing countries.

Mutations in the CEBPA gene are present in 7%-10% of human patients with acute myeloid leukemia (AML). However, no genetic models exist that demonstrate their etiological relevance. To mimic the most common mutations affecting CEBPA-that is, those leading to loss of the 42 kDa C/EBPalpha isoform (p42) while retaining the 30kDa isoform (p30)-we modified the mouse Cebpa locus to express only p30. p30 supported the formation of granulocyte-macrophage progenitors. However, p42 was required for control of myeloid progenitor proliferation, and p42-deficient mice developed AML with complete penetrance. p42-deficient leukemia could be transferred by a Mac1+c-Kit+ population that gave rise only to myeloid cells in recipient mice. Expression profiling of this population against normal Mac1+c-Kit+ progenitors revealed a signature shared with MLL-AF9-transformed AML.

Soon after Shannon defined the concept of redundancy it was suggested that it gave insight into mechanisms of sensory processing, perception, intelligence and inference. Can we now judge whether there is anything in this idea, and can we see where it should direct our thinking? This paper argues that the original hypothesis was wrong in over-emphasizing the role of compressive coding and economy in neuron numbers, but right in drawing attention to the importance of redundancy. Furthermore there is a clear direction in which it now points, namely to the overwhelming importance of probabilities and statistics in neuroscience. The brain has to decide upon actions in a competitive, chance-driven world, and to do this well it must know about and exploit the non-random probabilities and interdependences of objects and events signalled by sensory messages. These are particularly relevant for Bayesian calculations of the optimum course of action. Instead of thinking of neural representations as transformations of stimulus energies, we should regard them as approximate estimates of the probable truths of hypotheses about the current environment, for these are the quantities required by a probabilistic brain working on Bayesian principles.

Nonfermenting bacteria are ubiquitous environmental opportunists that cause infections in humans, especially compromised patients. Due to their limited biochemical reactivity and different morphotypes, misidentification by classical phenotypic means occurs frequently. Therefore, we evaluated the use of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) for species identification. By using 248 nonfermenting culture collection strains composed of 37 genera most relevant to human infections, a reference database was established for MALDI-TOF MS-based species identification according to the manufacturer's recommendations for microflex measurement and MALDI BioTyper software (Bruker Daltonik GmbH, Leipzig, Germany), i.e., by using a mass range of 2,000 to 20,000 Da and a new pattern-matching algorithm. To evaluate the database, 80 blind-coded clinical nonfermenting bacterial strains were analyzed. As a reference method for species designation, partial 16S rRNA gene sequencing was applied. By 16S rRNA gene sequencing, 57 of the 80 isolates produced a unique species identification >>or=99% sequence similarity); 11 further isolates gave ambiguous results at this threshold and were rated as identified to the genus level only. Ten isolates were identified to the genus level >>or=97% similarity); and two isolates had similarity values below this threshold, were counted as not identified, and were excluded from further analysis. MALDI-TOF MS identified 67 of the 78 isolates (85.9%) included, in agreement with the results of the reference method; 9 were misidentified and 2 were unidentified. The identities of 10 randomly selected strains were 100% correct when three different mass spectrometers and four different cultivation media were used. Thus, MALDI-TOF MS-based species identification of nonfermenting bacteria provided accurate and reproducible results within 10 min without any substantial costs for consumables.

Since its introduction in 1959, Ellman's reagent (5,5'-dithio-bis(2-nitrobenzoic acid)) has been the favorite reagent for spectrophotometric measurement of protein sulfhydryls. Meanwhile however, evidence has accumulated that many protein sulfhydryls give an incomplete reaction with Ellman's reagent, even during prolonged assay times. In the present study, the kinetic problem was solved by including cystamine as a "mediator" between the protein sulfhydryl and Ellman's reagent, as previously applied in an enzymatic thiol assay [9]. As an alternative, 4,4'-dithiodipyridine (DTDP) was used in place of Ellman's reagent. Due to its small size, amphiphilic nature, and lack of charge, DTDP quickly reacts with poorly accessible protein sulfhydryls, without any catalysis by cystamine. The DTDP method and the Ellman/cystamine method were both optimized for maximal sensitivity, minimal sample consumption (detection limit 0.2 nmol mL(-1), determination limit 0.6 nmol mL(-1)), and minimal assay time (5 min). In validation experiments, both methods gave identical results and the measured sulfhydryls/protein matched the expected values. Electronic supplementary material to this paper can be obtained by using the Springer Link server located at http://dx.doi.org/10.1007/s00216-002-1347-2.

BACKGROUND

Although it is agreed that a major polyploidy event, gamma, occurred within the eudicots, the phylogenetic placement of the event remains unclear.

RESULTS

To determine when this polyploidization occurred relative to speciation events in angiosperm history, we employed a phylogenomic approach to investigate the timing of gene set duplications located on syntenic gamma blocks. We populated 769 putative gene families with large sets of homologs obtained from public transcriptomes of basal angiosperms, magnoliids, asterids, and more than 91.8 gigabases of new next-generation transcriptome sequences of non-grass monocots and basal eudicots. The overwhelming majority (95%) of well-resolved gamma duplications was placed before the separation of rosids and asterids and after the split of monocots and eudicots, providing strong evidence that the gamma polyploidy event occurred early in eudicot evolution. Further, the majority of gene duplications was placed after the divergence of the Ranunculales and core eudicots, indicating that the gamma appears to be restricted to core eudicots. Molecular dating estimates indicate that the duplication events were intensely concentrated around 117 million years ago.

CONCLUSIONS

The rapid radiation of core eudicot lineages that gave rise to nearly 75% of angiosperm species appears to have occurred coincidentally or shortly following the gamma triplication event. Reconciliation of gene trees with a species phylogeny can elucidate the timing of major events in genome evolution, even when genome sequences are only available for a subset of species represented in the gene trees. Comprehensive transcriptome datasets are valuable complements to genome sequences for high-resolution phylogenomic analysis.

BACKGROUND

The adaptive radiations of cichlid fishes in East Africa are well known for their spectacular diversity and their astonishingly fast rates of speciation. About 80% of all 2,500 cichlid species in East Africa, and virtually all cichlid species from Lakes Victoria (approximately 500 species) and Malawi (approximately 1,000 species) are haplochromines. Here, we present the most extensive phylogenetic and phylogeographic analysis so far that includes about 100 species and is based on about 2,000 bp of the mitochondrial DNA.

RESULTS

Our analyses revealed that all haplochromine lineages are ultimately derived from Lake Tanganyika endemics. We find that the three most ancestral lineages of the haplochromines sensu lato are relatively species poor, albeit widely distributed in Africa, whereas a fourth newly defined lineage - the 'modern haplochromines' - contains an unparalleled diversity that makes up more than 7% of the worlds' approximately 25,000 teleost species. The modern haplochromines' ancestor, most likely a riverine generalist, repeatedly gave rise to similar ecomorphs now found in several of the species flocks. Also, the Tanganyikan Tropheini are derived from that riverine ancestor suggesting that they successfully re-colonized Lake Tanganyika and speciated in parallel to an already established cichlid adaptive radiation. In contrast to most other known examples of adaptive radiations, these generalist ancestors were derived from highly diverse and specialized endemics from Lake Tanganyika. A reconstruction of life-history traits revealed that in an ancestral lineage leading to the modern haplochromines the characteristic egg-spots on anal fins of male individuals evolved.

CONCLUSIONS

We conclude that Lake Tanganyika is the geographic and genetic cradle of all haplochromine lineages. In the ancestors of the replicate adaptive radiations of the 'modern haplochromines', behavioral (maternal mouthbrooding), morphological (egg-spots) and sexually selected (color polymorphism) key-innovations arose. These might be - together with the ecological opportunity that the habitat diversity of the large lakes provides - responsible for their evolutionary success and their propensity for explosive speciation.

Of 402 United States owners of side-viewing duodenoscopes surveyed, 222 (55%) responded, reporting 10,435 endoscopic retrograde cholangiopancreatograms. Procedure failed occurred in 30%, complications in 3%, and death in 0.2%. Complications included pancreatitis, cholangitis, pancreatic sepsis, instrumental injury to the gastrointestinal tract, and drug reactions. Pancreatitis was associated with injection into the pancreatic duct, sepsis with injection into an obstructed duct or pseudocyst, and injury with abnormal gastroduodenal anatomy. Experienced workers had a 15% incidence of complications, whereas inexperience gave 4 times the failures (62%) and twice the complications (7%). The causes of complications and their prevention are discussed.

Multiparameter flow cytometry was applied on normal human bone marrow (BM) cells to study the lineage commitment of progenitor cells ie, CD34+ cells. Lineage commitment of the CD34+ cells into the erythroid lineage was assessed by the coexpression of high levels of the CD71 antigen, the myeloid lineage by coexpression of the CD33 antigen and the B-lymphoid lineage by the CD10 antigen. Three color immunofluorescence experiments showed that all CD34+ BM cells that expressed the CD71, CD33, and CD10 antigens, concurrently stained brightly with anti-CD38 monoclonal antibodies (MoAbs). In addition, the CD38 antigen was brightly expressed on early T lymphocytes in human thymus, characterized by CD34, CD5, and CD7 expression. Only 1% of the CD34+ cells, 0.01% of nucleated cells in normal BM, did not express the CD38 antigen. The CD34+, CD38- cell population lacked differentiation markers and were homogeneous primitive blast cells by morphology. In contrast the CD34+, CD38 bright cell populations were heterogeneous in morphology and contained myeloblasts and erythroblasts, as well as lymphoblasts. These features are in agreement with properties expected from putative pluripotent hematopoietic stem cells; indeed, the CD34 antigen density decreased concurrently with increasing CD38 antigen density suggesting an upregulation of the CD38 antigen on differentiation of the CD34+ cells. Further evidence for a strong enrichment of early hematopoietic precursors in the CD34+, CD38- cell fraction was obtained from culture experiments in which CD34+ cell fractions with increasing density of the CD38 antigen were sorted singularly and assayed for blast colony formation. On day 14 of incubation, interleukin-3 (IL-3), IL-6, and GM-CSF, G-CSF, and erythropoietin (Epo) were added in each well. Twenty-five percent of the single sorted cells that expressed CD34 but lacked CD38 antigen gave rise to primitive colonies 28 to 34 days after cell sorting. The ability to form primitive colonies decreased rapidly with increasing density of the CD38 antigen. During 120 days of culture, up to five sequential generations of colonies were obtained after replating of the first-generation primitive colonies. This study provides direct evidence for the existence of a single class of progenitors with extensive proliferative capacity in human BM and provides an experimental approach for their purification, manipulation, and further characterization.

In vivo immune responses and in vitro mitogenic responses to bacterial lipopolysaccharides (LPS) have been compared in strains of C3H mice. C3H/HeJ spleen cultures did not support mitogenic responses to LPS and in vivo these mice produce low IgM responses to LPS. On the basis of these two responses, C3H/HeJ mice have been termed low LPS responders. All other strains of C3H mice tested (C3HeB/FeJ, C3H/DiSn, C3H/Str, CWB, CSW, and C3H/Sf and its H-2 congenics) are high LPS responders supporting large in vitro mitogenic and in vivo immune responses to LPS. The immune response difference between low and high LPS responders is a quantitative one. IgM responses are observed in C3H/HeJ mice in the range of 1.0-10 microg LPS. At lower and higher LPS concentrations, immune responses are not observed. In contrast, high LPS responders elicit LPS immune responses over a much wider dose range (0.1-200 microg). The ability to respond well to LPS is dominant as shown by the response of F(1) hybrid mice of low responder and high responder strains. The linkage relationships of mitogenic and immune responsiveness to LPS have been investigated in backcross (C3H/HeJ x CWB)F(1) x CWB mice. All mice that gave in vivo immune responses to LPS also supported mitogenic responses to LPS. The defect in C3H/HeJ mice that limits mitogenic and immune responsiveness to be due to a single autosomal gene which is not linked to the H-2 histocompatibility or heavy-chain allotype loci.

Three different deletion mutants of simian immunodeficiency virus (SIV) that vary in their levels of attenuation were tested for the ability to protect against mucosal challenge with pathogenic SIV. Four female rhesus monkeys were vaccinated by intravenous inoculation with SIVmac239Delta3, four with SIVmac239Delta3X, and four with SIVmac239Delta4. These three vaccine strains exhibit increasing levels of attenuation: Delta3 < Delta3X <Delta4. The vaccinated monkeys were challenged by vaginal exposure to uncloned, pathogenic SIVmac251 at 61 weeks after the time of vaccination. On the basis of viral RNA loads in plasma, cell-associated virus loads in peripheral blood, and CD4 cell counts, strong protective effects were observed in all three groups of vaccinated monkeys. However, the degree of protection correlated inversely with the level of attenuation; the least-attenuated strain, SIVmac239Delta3, <em>gave</em> the greatest protection. One monkey in the Delta3X group and two in the Delta4 group clearly became superinfected by the challenge virus, but these animals had levels of SIV RNA in plasma that were considerably lower than those of naive animals that were challenged in parallel. Protection against vaginal challenge appears easier to achieve than protection against intravenous challenge, since four other SIVmac239Delta4-vaccinated monkeys showed no protection when challenged intravenously with a much lower inoculum of the same challenge virus stock. Protection against vaginal challenge in the Delta4-vaccinated group occurred in the absence of detectable serum neutralizing activities and appeared to be associated with the development of an early SIV-specific cytotoxic-T-lymphocyte response. Our results demonstrate that mucosal protection can be achieved by systemic immunization with the highly attenuated SIVmac239Delta4 more than 1 year prior to the time of challenge.

Mycobacterium tuberculosis Beijing genotype strains are highly prevalent in Asian countries and in the territory of the former Soviet Union. They are increasingly reported in other areas of the world and are frequently associated with tuberculosis outbreaks and drug resistance. Beijing genotype strains, including W strains, have been characterized by their highly similar multicopy IS6110 restriction fragment length polymorphism (RFLP) patterns, deletion of spacers 1 to 34 in the direct repeat region (Beijing spoligotype), and insertion of IS6110 in the genomic dnaA-dnaN locus. In this study the suitability and comparability of these three genetic markers to identify members of the Beijing lineage were evaluated. In a well-characterized collection of 1,020 M. tuberculosis isolates representative of the IS6110 RFLP genotypes found in The Netherlands, strains of two clades had spoligotypes characteristic of the Beijing lineage. A set of 19 Beijing reference RFLP patterns was selected to retrieve all Beijing strains from the Dutch database. These reference patterns gave a sensitivity of 98.1% and a specificity of 99.7% for identifying Beijing strains (defined by spoligotyping) in an international database of 1,084 strains. The usefulness of the reference patterns was also assessed with large DNA fingerprint databases in two other European countries and for identification strains from the W lineage found in the United States. A standardized definition for the identification of M. tuberculosis strains belonging to the Beijing/W lineage, as described in this work, will facilitate further studies on the spread and characterization of this widespread genotype family of M. tuberculosis strains.

Serum folate, red cell folate, white blood cell vitamin C, riboflavin saturation index, and serum vitamin A were determined during the first trimester of pregnancy in over 900 cases. For each of these there was a social classes I + II showed the highest levels which differed significantly from other classes, except for serum folate. In 6 mothers who gave birth to infants with neural tube defects, first trimester serum folate, red cell folate, white blood cell vitamin C, and riboflavin values were lower than in controls. In spite of small numbers the differences were significant for red cell folate (P less than 0-001) and white blood cell vitamin C (P less than 0-05). These findings are compatible with the hypothesis that nutritional deficiencies are significant in the causation of congenital defects of the neural tube in man.

OBJECTIVE

To determine the efficacy of a probiotic drink containing Lactobacillus for the prevention of any diarrhoea associated with antibiotic use and that caused by Clostridium difficile.

METHODS

Randomised double blind placebo controlled study.

METHODS

135 hospital patients (mean age 74) taking antibiotics. Exclusions included diarrhoea on admission, bowel pathology that could result in diarrhoea, antibiotic use in the previous four weeks, severe illness, immunosuppression, bowel surgery, artificial heart valves, and history of rheumatic heart disease or infective endocarditis.

METHODS

Consumption of a 100 g (97 ml) drink containing Lactobacillus casei, L bulgaricus, and Streptococcus thermophilus twice a day during a course of antibiotics and for one week after the course finished. The placebo group received a longlife sterile milkshake.

METHODS

METHODS

occurrence of antibiotic associated diarrhoea. Secondary outcome: presence of C difficile toxin and diarrhoea.

RESULTS

7/57 (12%) of the probiotic group developed diarrhoea associated with antibiotic use compared with 19/56 (34%) in the placebo group (P=0.007). Logistic regression to control for other factors gave an odds ratio 0.25 (95% confidence interval 0.07 to 0.85) for use of the probiotic, with low albumin and sodium also increasing the risk of diarrhoea. The absolute risk reduction was 21.6% (6.6% to 36.6%), and the number needed to treat was 5 (3 to 15). No one in the probiotic group and 9/53 (17%) in the placebo group had diarrhoea caused by C difficile (P=0.001). The absolute risk reduction was 17% (7% to 27%), and the number needed to treat was 6 (4 to 14).

CONCLUSIONS

Consumption of a probiotic drink containing L casei, L bulgaricus, and S thermophilus can reduce the incidence of antibiotic associated diarrhoea and C difficile associated diarrhoea. This has the potential to decrease morbidity, healthcare costs, and mortality if used routinely in patients aged over 50.

BACKGROUND

National Research Register N0016106821.

OBJECTIVE

To use time-resolved three-dimensional phase-contrast magnetic resonance (MR) imaging, also called four-dimensional flow MR imaging, to evaluate systolic blood flow patterns in the ascending aorta that may predispose patients with a bicuspid aortic valve (BAV) to aneurysm.

METHODS

The HIPAA-compliant protocol received institutional review board approval, and informed consent was obtained. Four-dimensional flow MR imaging was used to assess blood flow in the thoracic aorta of 53 individuals: 20 patients with a BAV, 25 patients with a tricuspid aortic valve (TAV), and eight healthy volunteers. The Fisher exact test was used to evaluate the significance of flow pattern differences.

RESULTS

Nested helical flow was seen at peak systole in the ascending aorta of 15 of 20 patients with a BAV but in none of the healthy volunteers or patients with a TAV. This flow pattern was seen both in patients with a BAV with a dilated ascending aorta (n = 6) and in those with a normal ascending aorta (n = 9), was seen in the absence of aortic stenosis (n = 5), and was associated with eccentric systolic flow jets in all cases. Fusion of right and left leaflets gave rise to right-handed helical flow and right-anterior flow jets (n = 11), whereas right and noncoronary fusion gave rise to left-handed helical flow with left-posterior flow jets (n = 4).

CONCLUSIONS

Four-dimensional flow MR imaging showed abnormal helical systolic flow in the ascending aorta of patients with a BAV, including those without aneurysm or aortic stenosis. Identification and characterization of eccentric flow jets in these patients may help identify those at risk for development of ascending aortic aneurysm.