Silks have a long history of biomedical use as sutures. Silk can be purified, chemically modified to attach RGD sequences and processed into highly porous scaffolds for tissue engineering. We report biocompatibility studies of silk films (with or without covalently bound RGD) that were seeded with bone-marrow derived mesenchymal stem cells (MSC) and (a) cultured in vitro with human MSC or (b) seeded with autologous rat MSC and implanted in vivo. Controls for in vitro studies included tissue culture plastic (TCP; negative control), TCP with lipopolysaccharide (LPS) in the cell culture medium (positive control), and collagen films; controls for in vivo studies included collagen, PLA and TCP. After 9 h of culture, the expression of the pro-inflammatory Interleukin 1 beta (IL-1beta) and inflammatory cyclooxygenase 2 (COX-2) in human MSC were comparable for silk, collagen and TCP. After 30 and 96 h, gene expression of IL-1beta and COX-2 in MSC returned to the baseline (pre-seeding) levels. These data were corroborated by measuring IL-1beta and prostaglandin E2 levels in culture medium. The rate of cell proliferation was higher on silk films than either on collagen or TCP. In vivo, films made of silk, collagen or PLA were seeded with rat MSCs, implanted intramuscularly in rats and harvested after 6 weeks. Histological and immunohistochemical evaluation of silk explants revealed the presence of circumferentially oriented fibroblasts, few blood vessels, macrophages at the implant-host interface, and the absence of giant cells. Inflammatory tissue reaction was more conspicuous around collagen films and even more around PLA films when compared to silk. These data suggest that (a) purified degradable silk is biocompatible and (b) the in vitro cell culture model (hMSC seeded and cultured on biomaterial films) gave inflammatory responses that were comparable to those observed in vivo.

The prefrontal cortex is essential for the temporal integration of sensory information in behavioural and linguistic sequences. Such information is commonly encoded in more than one sense modality, notably sight and sound. Connections from sensory cortices to the prefrontal cortex support its integrative function. Here we present the first evidence that prefrontal cortex cells associate visual and auditory stimuli across time. We gave monkeys the task of remembering a tone of a certain pitch for 10 s and then choosing the colour associated with it. In this task, prefrontal cortex cells responded selectively to tones, and most of them also responded to colours according to the task rule. Thus, their reaction to a tone was correlated with their subsequent reaction to the associated colour. This correlation faltered in trials ending in behavioural error. We conclude that prefrontal cortex neurons are part of integrative networks that represent behaviourally meaningful cross-modal associations. The orderly and timely activation of neurons in such networks is crucial for the temporal transfer of information in the structuring of behaviour, reasoning and language.

The Q gene is largely responsible for the widespread cultivation of wheat because it confers the free-threshing character. It also pleiotropically influences many other domestication-related traits such as glume shape and tenacity, rachis fragility, spike length, plant height, and spike emergence time. We isolated the Q gene and verified its identity by analysis of knockout mutants and transformation. The Q gene has a high degree of similarity to members of the AP2 family of transcription factors. The Q allele is more abundantly transcribed than q, and the two alleles differ for a single amino acid. An isoleucine at position 329 in the Q protein leads to an abundance of homodimer formation in yeast cells, whereas a valine in the q protein appears to limit homodimer formation. Ectopic expression analysis allowed us to observe both silencing and overexpression effects of Q. Rachis fragility, glume shape, and glume tenacity mimicked the q phenotype in transgenic plants exhibiting post-transcriptional silencing of the transgene and the endogenous Q gene. Variation in spike compactness and plant height were associated with the level of transgene transcription due to the dosage effects of Q. The q allele is the more primitive, and the mutation that gave rise to Q occurred only once leading to the world's cultivated wheats.

The reproducibility and validity of self-reported menopausal status were evaluated among the 121,700 female US registered nurses aged 30-55 years in 1976 who are participants in the Nurses' Health Study, a prospective cohort study of diseases in women. When questioned in 1978, 6,591 of the women who were premenopausal in 1976 reported that their menses had ceased and provided their age at menopause. Two years later, in 1980, 98.8 per cent of the 6,591 women again reported that they were postmenopausal and again gave the reasons for menopause (natural or surgical). Among those women who reported surgical menopause during the 1976-1978 follow-up interval, age at menopause was reported to within one year on both the 1978 and 1980 questionnaires by 95 per cent of women. Among those reporting natural menopause during the 1976-1978 period, 82 per cent of women reported their age at menopause to within one year on the two follow-up questionnaires. A random sample of 255 women reporting surgical menopause between 1982 and 1984 was identified and medical records were obtained for 200. For all but two women, there was complete agreement between self-report and medical record for details of hysterectomy and extent of ovarian surgery. The reproducibility of self-reported age at menopause was assessed among 31,405 women who were menopausal in 1976. Reported age at menopause on consecutive questionnaires showed increasing within-person variance with increasing duration since menopause.(ABSTRACT TRUNCATED AT 250 WORDS)

BACKGROUND

Hematopoietic stem cell transplantation remains the only treatment for most patients with severe combined immunodeficiencies (SCIDs) or other primary immunodeficiencies (non-SCID PIDs).

OBJECTIVE

To analyze the long-term outcome of patients with SCID and non-SCID PID from European centers treated between 1968 and 2005.

METHODS

The product-limit method estimated cumulative survival; the log-rank test compared survival between groups. A Cox proportional-hazard model evaluated the impact of independent predictors on patient survival.

RESULTS

In patients with SCID, survival with genoidentical donors (n = 25) from 2000 to 2005 was 90%. Survival using a mismatched relative (n = 96) has improved (66%), similar to that using an unrelated donor (n = 46; 69%; P = .005). Transplantation after year 1995, a younger age, B(+) phenotype, genoidentical and phenoidentical donors, absence of respiratory impairment, or viral infection before transplantation were associated with better prognosis on multivariate analysis. For non-SCID PID, in contrast with patients with SCID, we confirm that, in the 2000 to 2005 period, using an unrelated donor (n = 124) gave a 3-year survival rate similar to a genoidentical donor (n = 73), 79% for both. Survival was 76% in phenoidentical transplants (n = 23) and worse in mismatched related donor transplants (n = 47; 46%; P = .016).

CONCLUSIONS

This is the largest cohort study of such patients with the longest follow-up. Specific issues arise for different patient groups. Patients with B-SCID have worse survival than other patients with SCID, despite improvements in each group. For non-SCID PID, survival is worse than SCID, although more conditions are now treated. Individual disease categories now need to be analyzed so that disease-specific prognosis may be better understood and the best treatments planned.

Mammalian cells harbor three highly homologous and widely expressed members of the ras family (H-ras, N-ras, and K-ras), but it remains unclear whether they play specific or overlapping cellular roles. To gain insight into such functional roles, here we generated and analyzed H-ras null mutant mice, which were then also bred with N-ras knockout animals to ascertain the viability and properties of potential double null mutations in both loci. Mating among heterozygous H-ras(+/-) mice produced H-ras(-/-) offspring with a normal Mendelian pattern of inheritance, indicating that the loss of H-ras did not interfere with embryonic and fetal viability in the uterus. Homozygous mutant H-ras(-/-) mice reached sexual maturity at the same age as their littermates, and both males and females were fertile. Characterization of lymphocyte subsets in the spleen and thymus showed no significant differences between wild-type and H-ras(-/-) mice. Analysis of neuronal markers in the brains of knockout and wild-type H-ras mice showed that disruption of this locus did not impair or alter neuronal development. Breeding between our H-ras mutant animals and previously available N-ras null mutants gave rise to viable double knockout (H-ras(-/-)/N-ras(-/-)) offspring expressing only K-ras genes which grew normally, were fertile, and did not show any obvious phenotype. Interestingly, however, lower-than-expected numbers of adult, double knockout animals were consistently obtained in Mendelian crosses between heterozygous N-ras/H-ras mice. Our results indicate that, as for N-ras, H-ras gene function is dispensable for normal mouse development, growth, fertility, and neuronal development. Additionally, of the three ras genes, K-ras appears to be not only essential but also sufficient for normal mouse development.

Astrocytes contribute to the formation and function of synapses and are found throughout the brain, where they show intracellular store-mediated Ca(2+) signals. Here, using a membrane-tethered, genetically encoded calcium indicator (Lck-GCaMP3), we report the serendipitous discovery of a new type of Ca(2+) signal in rat hippocampal astrocyte-neuron cocultures. We found that Ca(2+) fluxes mediated by transient receptor potential A1 (TRPA1) channels gave rise to frequent and highly localized 'spotty' Ca(2+) microdomains near the membrane that contributed appreciably to resting Ca(2+) in astrocytes. Mechanistic evaluations in brain slices showed that decreases in astrocyte resting Ca(2+) concentrations mediated by TRPA1 channels decreased interneuron inhibitory synapse efficacy by reducing GABA transport by GAT-3, thus elevating extracellular GABA. Our data show how a transmembrane Ca(2+) source (TRPA1) targets a transporter (GAT-3) in astrocytes to regulate inhibitory synapses.

1. The concentrations of the oxidized and reduced substrates of the ;malic' enzyme (EC 1.1.1.40) and isocitrate dehydrogenase (EC 1.1.1.42) were measured in freeze-clamped rat livers. By assuming that the reactants of these dehydrogenase systems are at equilibrium in the cytoplasm the [free NADP(+)]/[free NADPH] ratio was calculated. The justification of the assumption is discussed. 2. The values of this ratio obtained under different nutritional conditions (well-fed, 48hr.-starved, fed with a low-carbohydrate diet, fed with a high-sucrose diet) were all of the same order of magnitude although characteristic changes occurred on varying the diet. The value of the ratio fell on starvation and on feeding with the low-carbohydrate diet and rose slightly on feeding with the high-sucrose diet. 3. The mean values of the ratio were calculated to be between 0.001 and 0.015, which is about 100000 times lower than the values of the cytoplasmic [free NAD(+)]/[free NADH] ratio. 4. The differences in the redox state of the two nicotinamide-adenine dinucleotide couples can be explained on a simple physicochemical basis. The differences are the result of equilibria that are determined by the equilibrium constants of a number of highly active readily reversible dehydrogenases and transaminases and the concentrations of the substrates and products of these enzymes. 5. The decisive feature is the fact that the NAD and NADP couples share substrates. This sharing provides a link between the redox states of the two couples. 6. The application of the method of calculation to data published by Kraupp, Adler-Kastner, Niessner & Plank (1967), Goldberg, Passonneau & Lowry (1966) and Kauffman, Brown, Passonneau & Lowry (1968) shows that the redox states of the NAD and NADP couples in cardiac-muscle cytoplasm and in mouse-brain cytoplasm are of the same order as those in rat liver. 7. The determination of the equilibrium constant at 38 degrees , pH7.0 and I 0.25 (required for the calculation of the [free NADP(+)]/[free NADPH] ratio), gave a value of 3.44x10(-2)m for the ;malic' enzyme (with CO(2) rather than HCO(3) (-) as the reactant) and a value of 1.98x10(-2)m(-1) for glutathione reductase.

In the Escherichia coli genome, five putative open reading frame (ORF) clusters, mdlAB, ybjYZ, yddA, yojHI, and yhiH, have been assumed to be possible genes for ABC drug efflux transporters (I. T. Paulsen, M. K. Sliwinski, and M. H. Saier, Jr., J. Mol. Biol. 277:573-592, 1998). We cloned all of these ORFs in multicopy plasmids and investigated the drug resistance of drug-supersensitive host cells lacking constitutive multidrug efflux transporter genes acrAB. Among them, only ybjYZ gave significant erythromycin resistance and significantly decreased the accumulation of [(14)C]erythromycin. Therefore, ybjYZ was renamed macAB (macrolide-specific ABC-type efflux carrier). Plasmids carrying both the macA and -B genes conferred resistance against macrolides composed of 14- and 15-membered lactones but no or weak resistance against 16-membered ones. Neither of the two genes produced resistance alone. The DNA sequence suggests that MacB is an integral membrane protein with four transmembrane segments and one nucleotide-binding domain, while MacA belongs to a membrane fusion protein (MFP) family with a signal-like sequence at its N terminus. The expression of the histidine-tagged proteins confirmed that MacB is an integral membrane protein and MacA is a peripheral membrane protein. In addition, MacAB required TolC for its function in a way similar to that of most of the MFP-dependent transporters in E. coli. MacB is thus a novel ABC-type macrolide efflux transporter which functions by cooperating with the MFP MacA and the multifunctional outer membrane channel TolC. This is the first case of an experimentally identified ABC antibiotic efflux transporter in gram-negative organisms.

Indinavir (IDV) (also called CRIXIVAN, MK-639, or L-735,524) is a potent and selective inhibitor of the human immunodeficiency virus type 1 (HIV-1) protease. During early clinical trials, in which patients initiated therapy with suboptimal dosages of IDV, we monitored the emergence of viral resistance to the inhibitor by genotypic and phenotypic characterization of primary HIV-1 isolates. Development of resistance coincided with variable patterns of multiple substitutions among at least 11 protease amino acid residues. No single substitution was present in all resistant isolates, indicating that resistance evolves through multiple genetic pathways. Despite this complexity, all of 29 resistant isolates tested exhibited alteration of residues M-46 (to I or L) and/or V-82 (to A, F, or T), suggesting that screening of these residues may be useful in predicting the emergence of resistance. We also extended our previous finding that IDV-resistant viral variants exhibit various patterns of cross-resistance to a diverse panel of HIV-1 protease inhibitors. Finally, we noted an association between the number of protease amino acid substitutions and the observed level of IDV resistance. No single substitution or pair of substitutions tested gave rise to measurable viral resistance to IDV. The evolution of this resistance was found to be cumulative, indicating the need for ongoing viral replication in this process. These observations strongly suggest that therapy should be initiated with the most efficacious regimen available, both to suppress viral spread and to inhibit the replication that is required for the evolution of resistance.

Spinal anaesthesia developed in the late 1800s with the work of Wynter, Quincke and Corning. However, it was the German surgeon, Karl August Bier in 1898, who probably gave the first spinal anaesthetic. Bier also gained first-hand experience of the disabling headache related to dural puncture. He correctly surmised that the headache was related to excessive loss of cerebrospinal fluid (CSF). In the last 50 yr, the development of fine-gauge spinal needles and needle tip modification, has enabled a significant reduction in the incidence of post-dural puncture headache. Though it is clear that reducing the size of the dural perforation reduces the loss of CSF, there are many areas regarding the pathogenesis, treatment and prevention of post-dural puncture headache that remain contentious. How does the microscopic pattern of collagen alignment in the spinal dura affect the dimensions of the dural perforation? How do needle design, size and orientation influence leakage of CSF through the dural perforation? Can pharmacological methods reduce the symptoms of post-dural puncture headache? By which mechanism does the epidural blood patch cure headache? Is there a role for the prophylactic epidural blood patch? Do epidural saline, dextran, opioids and tissue glues reduce the rate of CSF loss? This review considers these contentious aspects of post-dural puncture headache.

A collection of yeast temperature-sensitive mutants was screened by an enzymatic assay to find a mutant defective in the acetylation of histone H4. The assay used a fractionated cell extract and measured acetylation of a peptide corresponding to amino acids 1-28 of H4. There are at least two activities in this fraction that acetylate the peptide. A mutation, hat1-1, that eliminates one of the activities was identified and mapped to a locus near the centromere of chromosome XVI. The HAT1 gene was cloned and found to encode a protein of 374 amino acids. Analysis of the peptide used in the assay demonstrated that the HAT1 enzyme acetylates lysine 12 of histone H4. hat1 mutants have no obvious growth defects or phenotypes other than the enzyme defect itself. The HAT1 protein expressed in Escherichia coli gave histone acetyltransferase activity in vitro, demonstrating that HAT1 is the structural gene for the enzyme.

Three sets of primers were designed for PCR detection and differentiation of Campylobacter jejuni and Campylobacter coli. The first PCR assay was designed to coidentify C. jejuni and C. coli based on their 16S rRNA gene sequences. The second PCR assay, based on the hippuricase gene sequence, identified all tested reference strains of C. jejuni and also strains of that species which lack detectable hippuricase activity. The third PCR assay, based on the sequence of a cloned (putative) aspartokinase gene and the downstream open reading frame, identified all tested reference strains of C. coli. The assays will find immediate application in the rapid identification to species level of isolates. The assays combine with a protocol for purification of total DNA from fecal samples to allow reproducible PCR identification of campylobacters directly from stools. Of 20 clinical samples from which campylobacters had been cultured, we detected C. jejuni in 17, C. coli in 2, and coinfection of C. jejuni and Campylobacter hyointestinalis in 1. These results were concordant with culture and phenotypic identification to species level. Strain typing by PCR-restriction fragment length polymorphism of the flagellin (flaA) gene detected identical flaA types in fecal DNA and the corresponding campylobacter isolate. Twenty-five Campylobacter-negative stool samples gave no reaction with the PCR assays. These PCR assays can rapidly define the occurrence, species incidence, and flaA genotypes of enteropathogenic campylobacters.

We previously demonstrated that human pericytes, which encircle capillaries and microvessels, give rise in culture to genuine mesenchymal stem cells (MSCs). This raised the question as to whether all MSC are derived from pericytes. Pericytes and other cells defined on differential expression of CD34, CD31, and CD146 were sorted from the stromal vascular fraction of human white adipose tissue. Besides pericytes, CD34+ CD31- CD146- CD45- cells, which reside in the outmost layer of blood vessels, the tunica adventitia, natively expressed MSC markers and gave rise in culture to clonogenic multipotent progenitors identical to standard bone marrow-derived MSC. Despite common MSC features and developmental properties, adventitial cells and pericytes retain distinct phenotypes and genotypes through culture. However, in the presence of growth factors involved in vascular remodeling, adventitial cells acquire a pericytes-like phenotype. In conclusion, we demonstrate the co-existence of 2 separate perivascular MSC progenitors: pericytes in capillaries and microvessels and adventitial cells around larger vessels.

OBJECTIVE

(1) To describe the relative importance of gestational weight gain, postpartum exercise, food intake and breastfeeding to weight change from early pregnancy to 1 y postpartum; and (2) to identify subgroups of women at greatest risk for major weight gain surrounding childbearing.

METHODS

A prospective cohort study of women who registered for obstetrical care in a hospital and primary care clinic system serving a 10 county area of upstate New York.

METHODS

A total of 540 healthy adult women who gave birth to full-term singleton infants.

METHODS

Sociodemographic characteristics, exercise, food-related behaviors and breastfeeding were assessed using the medical record and a mailed questionnaire. Body weight was measured at prenatal visits and 1 y postpartum. Weight retained and major weight gain (4.55 kg) at 1 y postpartum were the main outcomes.

METHODS

Linear and logistic regression analyses were conducted.

RESULTS

Women were on average 1.51+/-5.95 kg heavier at 1 y postpartum than they were in early pregnancy. Nearly 25% of women experienced a major weight gain of 4.55 kg or more at 1 y postpartum. Gestational weight gain, exercise frequency, change in food intake and breastfeeding were each significantly related to postpartum weight retention. With the exception of breastfeeding, all of these factors were also associated with major weight gain. Women under 20 y or over 40 y at delivery, and single women retained significantly more weight. Lower income women with gestational weight gains above the Institute of Medicine (IOM) range retained 3.73 kg more than lower income women who gained within the range. They were also 4.7 times more likely to experience major weight gain with childbearing. The impact of exceeding the IOM gestational weight gain guidelines was three times greater in lower income women than it was in higher income women.

CONCLUSIONS

Gestational weight gain, postpartum exercise frequency, and food intake are significantly associated with weight change from early pregnancy to 1 y postpartum and major weight gain with childbearing. Lower income women who gain more weight in pregnancy than the IOM recommends are at high risk for major weight gain with childbearing.

Cell viability and in vitro cytotoxicity assay methods were developed using a combination of dyes, 4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate (WST-1), neutral red (NR) and crystal violet (CV), with HeLa cells as a bioindicatior. As WST-1 produces a highly water soluble and non-cytotoxic formazan dye, ti allows each assay to be carried out in one culture dish. The combined cell viability assay using WST-1, NR and CV gave an absorbance that correlated linearly with the number of cells over the range 1000 to 50,000 cells/well. The combined assay was applied to the evaluation of IC50 values for sodium dodecyl sulfate as a model toxicant, which yielded similar values to those obtained with each assay independently.

OBJECTIVE

Inflammation plays a key role in cerebral ischemia through activation of microglia and infiltration by leukocytes. Flow cytometry is a well-established method for quantitative and qualitative analysis of inflammatory cells. However, this technique has not been applied to the study of cerebral ischemia inflammation. The aim of this study was to establish a flow cytometric method to measure inflammatory cells in ischemic brain.

METHODS

To perform flow cytometry on brain tissue, we developed 2 cell-isolation methods based on different mechanical dissociation and Percoll gradient separation techniques. The methods were tested on a rat model of permanent middle cerebral artery occlusion. Morphological and immunophenotypic analyses, with the use of anti-CD11b, anti-CD45, and alphabeta T-cell receptor antibodies, were employed to identify and quantify inflammatory cells.

RESULTS

Both methods gave consistent results in terms of yield and reproducibility. The cell suspension contained granulocytes, macrophages, lymphocytes, and neural cells. Morphological and immunophenotypic analyses enabled the identification of a cell-scatter gate (R1a) enriched in inflammatory cells. With both methods, a higher number of events in R1a were recorded in the ischemic hemisphere than in the nonischemic hemisphere (P< or =0.001). CD11b, CD45, and alphabeta T-cell receptor staining confirmed that this augmentation was a reflection of the increase in the number of granulocytes, cells of the monocytic lineage, and lymphocytes.

CONCLUSIONS

Quantitative flow cytometric analysis of ischemic rat brain is feasible and provides a reliable and rapid assay to assess neuroinflammation in experimental models of brain ischemia.

The above- and below-ground parts of rice plants create specific habitats for various microorganisms. In this study, we characterized the phyllosphere and rhizosphere microbiota of rice cultivars using a metaproteogenomic approach to get insight into the physiology of the bacteria and archaea that live in association with rice. The metaproteomic datasets gave rise to a total of about 4600 identified proteins and indicated the presence of one-carbon conversion processes in the rhizosphere as well as in the phyllosphere. Proteins involved in methanogenesis and methanotrophy were found in the rhizosphere, whereas methanol-based methylotrophy linked to the genus Methylobacterium dominated within the protein repertoire of the phyllosphere microbiota. Further, physiological traits of differential importance in phyllosphere versus rhizosphere bacteria included transport processes and stress responses, which were more conspicuous in the phyllosphere samples. In contrast, dinitrogenase reductase was exclusively identified in the rhizosphere, despite the presence of nifH genes also in diverse phyllosphere bacteria.

Mitochondrial DNAs (mtDNAs) from 167 American Indians including 87 Amerind-speakers (Amerinds) and 80 Nadene-speakers (Nadene) were surveyed for sequence variation by detailed restriction analysis. All Native American mtDNAs clustered into one of four distinct lineages, defined by the restriction site variants: HincII site loss at np 13,259, AluI site loss at np 5,176, 9-base pair (9-bp) COII-tRNA(Lys) intergenic deletion and HaeIII site gain at np 663. The HincII np 13,259 and AluI np 5,176 lineages were observed exclusively in Amerinds and were shared by all such tribal groups analyzed, thus demonstrating that North, Central and South American Amerinds originated from a common ancestral genetic stock. The 9-bp deletion and HaeIII np 663 lineages were found in both the Amerinds and Nadene but the Nadene HaeIII np 663 lineage had a unique sublineage defined by an RsaI site loss at np 16,329. The amount of sequence variation accumulated in the Amerind HincII np 13,259 and AluI np 5,176 lineages and that in the Amerind portion of the HaeIII np 663 lineage all gave divergence times in the order of 20,000 years before present. The divergence time for the Nadene portion of the HaeIII np 663 lineage was about 6,000-10,000 years. Hence, the ancestral Nadene migrated from Asia independently and considerably more recently than the progenitors of the Amerinds. The divergence times of both the Amerind and Nadene branches of the COII-tRNA(Lys) deletion lineage were intermediate between the Amerind and Nadene specific lineages, raising the possibility of a third source of mtDNA in American Indians.

We examined children's and adults' numerical estimation and the representations that gave rise to their estimates. The results were inconsistent with two prominent models of numerical representation: the logarithmic-ruler model, which proposes that people of all ages possess a single, logarithmically spaced representation of numbers, and the accumulator model, which proposes that people of all ages represent numbers as linearly increasing magnitudes with scalar variability. Instead, the data indicated that individual children possess multiple numerical representations; that with increasing age and numerical experience, they rely on appropriate representations increasingly often; and that the numerical context influences their choice of representation. The results, obtained with second graders, fourth graders, sixth graders, and adults who performed two estimation tasks in two numerical contexts, strongly suggest that one cause of children's difficulties with estimation is reliance on logarithmic representations of numerical magnitudes in situations in which accurate estimation requires reliance on linear representations.

CD22 is a surface glycoprotein of B lymphocytes that is rapidly phosphorylated on cytoplasmic tyrosines after antigen receptor cross-linking. Splenic B cells from mice with a disrupted CD22 gene were found to be hyperresponsive to receptor signaling: Heightened calcium fluxes and cell proliferation were obtained at lower ligand concentrations. The mice gave an augmented immune response, had an expanded peritoneal B-1 cell population, and contained increased serum titers of autoantibody. Thus, CD22 is a negative regulator of antigen receptor signaling whose onset of expression at the mature B cell stage may serve to raise the antigen concentration threshold required for B cell triggering.

Algae include a diverse array of photosynthetic eukaryotes excluding land plants. Explaining the origin of algal plastids continues to be a major challenge in evolutionary biology. Current knowledge suggests that plastid primary endosymbiosis, in which a single-celled protist engulfs and "enslaves" a cyanobacterium, likely occurred once and resulted in the primordial alga. This eukaryote then gave rise through vertical evolution to the red, green, and glaucophyte algae. However, some modern algal lineages have a more complicated evolutionary history involving a secondary endosymbiotic event, in which a protist engulfed an existing eukaryotic alga (rather than a cyanobacterium), which was then reduced to a secondary plastid. Secondary endosymbiosis explains the majority of algal biodiversity, yet the number and timing of these events is unresolved. Here we analyzed a five-gene plastid data set to show that a taxonomically diverse group of chlorophyll c(2)-containing protists comprising cryptophyte, haptophyte, and stramenopiles algae (Chromista) share a common plastid that most likely arose from a single, ancient ( approximately 1,260 million years ago) secondary endosymbiosis involving a red alga. This finding is consistent with Chromista monophyly and implicates secondary endosymbiosis as an important force in generating eukaryotic biodiversity.

The product of the abrB gene of Bacillus subtilis is an ambiactive repressor and activator of the transcription of genes expressed during the transition state between vegetative growth and the onset of stationary phase and sporulation. Purified AbrB protein binds specifically in a highly co-operative fashion to fragments of DNA containing the promoters it affects. DNase I footprints of the binding regions in these promoters revealed large protected areas of 50-120 nucleotides or more depending on the promoter. Methylation protection experiments gave protected guanine residues on only one face of the DNA helix. A consensus sequence could be deduced around these guanine residues that was not found around non-protected guanine residues in the footprint region. The results suggested that stationary phase functions and sporulation are repressed during active growth by AbrB and other transition state regulators by binding to the affected promoters in a concentration-dependent manner.

Isolation of Clostridium difficile from fecal specimens has been facilitated by the development of a selective and differential medium, cefoxitin-cycloserinefructose agar (CCFA). We substituted 0.1% sodium taurocholate for the 2.5% egg yolk in CCFA and compared the growth of 15 isolates of C. difficile on the resulting medium with growth on conventional CCFA. The taurocholate-containing medium (TCCFA) quantitatively recovered vegetative forms of C. difficile in the same numbers as CCFA medium. Recovery of spores was a mean 1.7 log(10) higher on TCCFA than on CCFA. Thirty-six of 60 patient stool specimens growing C. difficile gave a heavier growth on TCCFA than on CCFA, and 9 failed to yield C. difficile on CCFA. TCCFA detected spores of 75 colony-forming units per ml from artificially inoculated fecal specimens when conventional stool culturing techniques were used. Fluorescence of colonies of C. difficile was more intense on TCCFA than on CCFA. TCCFA was simpler to prepare and, overall, was more sensitive than CCFA.