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This study developed potential blood-based biomarker tests for diagnosing and differentiating schizophrenia (SZ), bipolar disorder type I (BD), and normal control (NC) subjects using mRNA gene expression signatures. A total of 90 subjects ( n = 30 each for the three groups of subjects) provided blood samples at two visits. The Affymetrix exon microarray was used to profile the expression of over 1.4 million probesets. We selected potential biomarker panels using the temporal stability of the probesets and also back-tested them at two different visits for each subject. The 18-gene biomarker panels, using logistic regression modeling, correctly differentiated the three groups of subjects with high accuracy across the two different clinical visits (83-88% accuracy). The results are also consistent with the actual data and the "leave-one-out" analyses, indicating that the models should be predictive when applied to independent data cohorts. Many of the SZ and BD subjects were taking antipsychotic and mood stabilizer medications at the time of blood draw, raising the possibility that these drugs could have affected some of the differential transcription signatures. Using an independent Illumina data set of gene expression data from antipsychotic medication-free SZ subjects, the 18-gene biomarker panels produced a receiver operating characteristic curve accuracy greater than 0.866 in patients that were less than 30 years of age and medication free. We confirmed select transcripts by quantitative PCR and the nCounter® System. The episodic nature of psychiatric disorders might lead to highly variable results depending on when blood is collected in relation to the severity of the disease/symptoms. We have found stable trait gene panel markers for lifelong psychiatric disorders that may have diagnostic utility in younger undiagnosed subjects where there is a critical unmet need. The study requires replication in subjects for ultimate proof of the utility of the differential diagnosis.

Menstrual toxic shock syndrome (mTSS) is a rare, recognizable, and treatable disease that has been associated with tampon use epidemiologically. It involves a confluence of microbial risk factors (Staphylococcus aureus strains that produce the superantigen-TSST-1), as well as environmental characteristics of the vaginal ecosystem during menstruation and host susceptibility factors. This paper describes a series of experiments using the well-characterized model of porcine vaginal mucosa ex-vivo to assess the effect of these factors associated with tampon use on the permeability of the mucosa. The flux of radiolabeled TSST-1 and tritiated water ((3)H2O) through porcine vaginal mucosa was determined at various temperatures, after mechanical disruption of the epithelial surface by tape stripping, after treatment with surfactants or other compounds, and in the presence of microbial virulence factors. Elevated temperatures (42, 47 and 52°C) did not significantly increase flux of (3)H2O. Stripping of the epithelial layers significantly increased the flux of labeled toxin in a dose-dependent manner. Addition of benzalkonium chloride (0.1 and 0.5%) and glycerol (4%) significantly increased the flux of (3)H2O but sodium lauryl sulfate at any concentration tested did not. The flux of the labeled toxin was significantly increased in the presence of benzalkonium chloride but not Pluronic® L92 and Tween 20 and significantly increased with addition of α-hemolysin but not endotoxin. These results show that the permeability of porcine vagina ex-vivo to labeled toxin or water can be used to evaluate changes to the vaginal environment and modifications in tampon materials, and thus aid in risk assessment., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Objective: To investigate the influence of menses on the vaginal microbiota and determine whether tampons that differ in material composition influence these bacterial communities in different ways., Design: A single-centre trial with randomised, complete block design., Setting: Procter & Gamble facility., Sample: Seven self-declared healthy, female volunteers of reproductive age., Methods: Volunteers used a pad and two types of tampons during the study, one product exclusively each month for three sequential menstrual cycles. During menses and once each mid-cycle, vaginal bacterial community composition was characterised by cultivation-independent methods based on pyrosequencing of V1-V2 variable regions of 16S ribosomal RNA genes., Main Outcome Measures: Changes in the species composition, abundance and diversity in vaginal bacterial communities over time and between treatments., Results: The vaginal microbiotas of all seven women were dominated by Lactobacillus spp. at mid-cycle, and the compositions of those communities were largely consistent between cycles. Community dynamic patterns during menses varied considerably and were more or less individualised. In three of the seven women the community diversity during pad use was significantly different from at least one tampon cycle., Conclusions: Changes in the composition of the vaginal microbiota during menses were common, but the magnitude of change varied between women. Despite these changes, most communities were capable of resuming a composition similar to previous mid-cycle sampling times following menstruation. Overall we conclude that the two tampons tested do not significantly impact the vaginal microbiota in different ways; however, larger studies should be performed to confirm these findings., (© 2013 The Authors BJOG An International Journal of Obstetrics and Gynaecology © 2013 RCOG.)

Vaginal tampons are Class II medical devices used by women to manage menstruation. The purpose of this study was to investigate intravaginal temperature changes with simulated and actual menstrual tampon use. Tampons (with varying absorbent compositions) embedded with a thermocouple sensor were used to study temperature effects in vitro in a model of the vagina (condom placed in a hollow glass tube, jacketed in a 37 degrees C water bath, and dosed with human menses to fluid saturation) and clinically during menstrual tampon wear under controlled conditions (up to 8 h in a stationary, supine position). Elevations in the temperature of the tampon core occurred upon menses fluid acquisition both in vitro and clinically. Temperature profile characteristics varied from a transient spike with commercial cotton-rayon blend tampons of two different absorbencies to a small but sustained rise (> or =6 h) with a carboxymethyl cellulose (CMC)-containing prototype. On the basis of the results from this study, fluid absorption by tampons generates an exothermic event whose characteristics vary with tampon design and composition. We speculate the small, sustained increased in tampon temperature noted during this study may enhance the production of a bacterial exotoxin associated with tampons composed of CMC.

Vulvovaginal candidiasis (VVC) is an insidious infection that afflicts a large proportion of women of all ages, and 5 to 8% of affected women experience recurrent VVC (RVVC). The aim of this study was to explore the possible importance of vaginal bacterial communities in reducing the risk of RVVC. The species composition and diversity of microbial communities were evaluated for 42 women with and without frequent VVC based on profiles of terminal restriction fragment polymorphisms of 16S rRNA genes and phylogenetic analysis of cloned 16S rRNA gene sequences from the numerically dominant microbial populations. The data showed that there were no significant differences between the vaginal microbial communities of women in the two groups (likelihood score, 5.948; bootstrap P value, 0.26). Moreover, no novel bacteria were found in the communities of women with frequent VVC. The vaginal communities of most women in both groups (38/42; 90%) were dominated by species of Lactobacillus. The results of this study failed to provide evidence for the existence of altered or unusual vaginal bacterial communities in women who have frequent VVC compared to women who do not have frequent VVC. The findings suggest that commensal vaginal bacterial species may not be able to prevent VVC.

The hormones progesterone and estrogen and, more precisely, their sophisticated interdependent fluctuations over the course of the female human lifespan, have long been known to play a dominant role in the physiological development and homeostasis of the human female. What is only recently coming to light, however, is that the fluctuation of these two hormones also plays a crucial role in neurological and psychological development and function which impacts brain function, cognition, emotional status, sensory processing, appetite, and more. The ability of reproductive hormones to impact psychoneurological processes involves the interplay of several body systems, lending credibility to the view of premenstrual syndrome (PMS) as a disorder founded in real biochemical disturbances. The effects of the menstrual cycle on cognitive, emotional, and sensory function in the female of childbearing age are reviewed. In addition, recent evidence is discussed which confirms the biological basis of PMS as a real disorder of primarily autoimmune origin.

Background: Transcript abundance (TA) measurement in whole blood frequently is conducted to identify potential biomarkers for disease risk and to predict or monitor drug response. Potential biomarkers discovered in this way must be validated by quantitative technology. In this study we assessed the use of standardized reverse transcription PCR (StaRT-PCR) to validate potential biomarkers discovered through whole blood TA profiling., Methods: For each of 15 healthy volunteers, 6 blood samples were obtained, including 3 samples at each of 2 separate visits. Total variation in TA for each gene was partitioned into replicate, sample, visit, study participant, and residual components., Results: Variation originating from technical processing was <5% of total combined variation and was primarily preanalytical. Interindividual biological sample variation was larger than technical variation. For 12 of 19 tests, the distribution of measured values was gaussian (Shapiro-Wilks test)., Conclusion: For control or diseased population groups with variation rates as low as those observed in this control group, 17 individuals per group would be required to detect 1 SD change with 80% power with a 2-sided alpha = 0.05 statistical test for mean differences.

Over the last decade, the introduction of microarray technology has had a profound impact on gene expression research. The publication of studies with dissimilar or altogether contradictory results, obtained using different microarray platforms to analyze identical RNA samples, has raised concerns about the reliability of this technology. The MicroArray Quality Control (MAQC) project was initiated to address these concerns, as well as other performance and data analysis issues. Expression data on four titration pools from two distinct reference RNA samples were generated at multiple test sites using a variety of microarray-based and alternative technology platforms. Here we describe the experimental design and probe mapping efforts behind the MAQC project. We show intraplatform consistency across test sites as well as a high level of interplatform concordance in terms of genes identified as differentially expressed. This study provides a resource that represents an important first step toward establishing a framework for the use of microarrays in clinical and regulatory settings.

Previous in vitro and in vivo animal studies showed that O(2) and CO(2) concentrations can affect virulence of pathogenic bacteria such as Staphylococcus aureus. The objective of this work was to measure O(2) and CO(2) levels in the vaginal environment during tampon wear using newly available sensor technology. Measurements by two vaginal sensors showed a decrease in vaginal O(2) levels after tampon insertion. These decreases were independent of the type of tampons used and the time of measurement (mid-cycle or during menstruation). These results are not in agreement with a previous study that concluded that oxygenation of the vaginal environment during tampon use occurred via delivery of a bolus of O(2) during the insertion process. Our measurements of gas levels in menses showed the presence of both O(2) and CO(2) in menses. The tampons inserted into the vagina contained O(2) and CO(2) levels consistent with atmospheric conditions. Over time during tampon use, levels of O(2) in the tampon decreased and levels of CO(2) increased. Tampon absorbent capacity, menses loading, and wear time influenced the kinetics of these changes. Colonization with S. aureus had no effect on the gas profiles during menstruation. Taken collectively, these findings have important implications on the current understanding of gaseous changes in the vaginal environment during menstruation and the potential role(s) they may play in affecting bacterial virulence factor production.

Gene probes derived from the insertion sequence IS986, which have previously been shown to differentiate isolates of Mycobacterium tuberculosis for epidemiological analysis, are also capable of distinguishing two groups of BCG vaccine strains. Most BCG strains have a single copy of IS986, at the same chromosomal site, while the Brazilian, Japanese and USSR strains have an additional copy at a different, common location. These results correlate with the results of previous antigenic analysis and may reflect a different clonal origin of the two groups of BCG strains.

Some physical factors influencing the performance of the tuberculin Tine Test have been investigated. Conditions under which tuberculin is removed from the tines were examined, by direct observation of elution with water, and by insertion into a number of materials, including human and animal skin. Uptake of tuberculin on to tines was also studied and quantitative estimates were obtained by dipping the tines in tuberculin mixed with 32P; it was found that the amount of tuberculin adhering to the tines was approximately doubled by applying a second coat after the first had dried. Tines coated with this mixture were applied to human skin from a cadaver, and after removal it was noted that approximately half the tuberculin orginally coating the tines had been deposited in the skin; the quantity inserted by doubly-coated tines was approximately double that deposited by singly-coated ones.

The chemical properties, stereochemical relationships and solution conformation, as assessed in part by proton NMR spectroscopy, for vitamin D3, its major metabolites [including 1alpha,25-(OH)2D3, its hormonally active form] and a number of A-ring and side chain analogues are evaluated and discussed in relation to their biological activity. In particular the relative ability of many of these seco-steroids to compete both with 25-OHD3 for its chick serum binding protein and 1alpha,25-(OH)2-D3 for its chick intestinal cytosol-chromatin receptor system was quantitated, in vitro. Further, the relative effectiveness of all these metabolites and analogues to mediate in vivo intestinal calcium absorption and bone calcium mobilization was determined. Collectively these chemical and biological studies constitute a "systems analysis" of the various steroid structural parameters both required and tolerated by the multi-stepped endocrine system associated with the biological actions of vitamin D.

Four collaborating BCG production laboratories had each prepared vaccine from four different BCG strains. In this laboratory the morphology of colonies cultured from samples of the 16 vaccine lots thus available was examined, and it was found that changes monitored in earlier small-scale experimental reconstructions had also occurred during some of the full-scale production procedures, in accordance with prediction. In particular, a minority population carried by the Danish BCG strain had replaced the original majority when this strain had been employed as seed for the production of vaccine by the British procedure. Similarly, a minority carried by the Japanese strain had replaced the original majority when this strain was introduced as seed into the production procedures in the United Kingdom and France. Serial subcultures made in this laboratory showed, in accordance with previous experience, that the changes that had occurred in the Japanese strain could be completely reversed by serial subculture as a surface pellicle on Sauton medium.

Changes in the Danish BCG strain under certain regimens of subculture have been shown in preceding studies to be associated with selection of a minority population. Three Czechoslovakian BCG strains, all originally derived from the Danish strain but thereafter and in distinction from it maintained on potato media, have now been investigated. Changes in the immunizing potency of two of these strains have been attributed by other workers to employment of the richer potato media in place of Sauton medium as used for maintenance of the parent Danish strain. However, results from the present study suggest rather that selection of a pre-existing minority genotype or of a new mutant occurred. This proposal is supported by the finding that the third strain has maintained characteristics similar to those of the Danish parent despite many previous transfers on potato media.Another BCG strain investigated was the Japanese which, like the three Czechoslovakian strains, had been previously maintained on potato media. This strain has been shown in the present study to resemble the Danish strain in supporting a minority population yielding non-spreading colonies. Czechoslovakian vaccine prepared with seed culture supplied from Tokyo has retained characteristics similar to those of the Japanese parent. Although a majority population yielding spreading colonies appears so far to have been retained in both centres, it is considered that selection of the minority could still occur in the course of routine transfer.

A rapid and efficient method for the separation of egg yolk phospholipids by normal-phase high-performance liquid chromatography has been developed. The separation is accomplished on an Ultrasil-NH2 column using hexane, isopropanol, methanol and water mixtures with direct ultraviolet detection at 206 nm. Phospholipids from a methanol extract of fresh lyophilized egg yolks were analyzed providing complete separation of (in order of elution) neutral lipids, phosphatidylcholine, sphingomyelin, lysophosphatidylcholine and phosphatidylethanolamine. Identification of eluting species was accomplished by comparative retention times of standard samples by thin-layer chromatographic analyses of collected fractions. Furthermore, a mixture of naturally occurring phospholipid standards from bovine and egg sources has been separated by this method. In addition to separating the individual classes of phospholipids, in some instances, separation of molecular species within a class was achieved as in the case of cerebrosides, sphingomyelin and partially with phosphatidylethanolamine.

Samples of Bacille Calmette-Guérin (BCG) vaccines from four collaborating production laboratories, each of which had prepared vaccine from four different daughter strains of BCG, had previously been monitored for changes in colony morphology and the present study was undertaken to determine whether the changes observed were reflected in the patterns of protein secretion and lipid content. In the samples examined there was evidence for a correlation between colony morphology and the presence or absence of mycoside B. As the components of BCG that determine virulence and protective immunity are unknown, care must be taken to ensure constancy of the strains during the manufacture of vaccines.

Under appropriate conditions of growth colonies showing fine wrinkling (rugosity) of their surface and characteristic of certain BCG strains can be distinguished from colonies with a smoother non-rugose morphology that are characteristic of some other BCG strains. This study has been concerned with the influence of nutritional and other media constituents on the evolution of these colonies. In a preliminary investigation nutritional components such as enzymic digest of casein, asparagine and salts were systematically eliminated from an agar medium, separately and together, and in the presence or absence of glycerol. From this and supplementary experiments, it appeared that the nutritional element with most effect on the size and morphology of colonies is the carbohydrate. Small inocula of BCG will normally grow on simple agar media only in the presence of enrichments such as blood or albumin, or if charcoal has been added. However, in a second investigation it was found that individual colonies would develop from such inocula placed 2 cm or more away from a concentrated inoculum that had been seeded onto the medium approximately 7-14 days earlier. It appears that a diffusible agent is produced from multiplying bacilli within a colony which counteracts toxic factors in the medium and may also assist nutrition within the colony. To examine the effects of carbohydrate on the evolution of colonies, in a third investigation glycerol was added in increasing quantities to an agar medium enriched with increasing quantities of bovine albumin and/or 5% of blood, and suspensions of French (Pasteur) and British (Glaxo) BCG vaccines were seeded onto it. Individual colonies cultured from these two strains have a rugose and non-rugose morphology respectively, and to highlight the effects in question, the evolution over a prolonged period of concentrated 'drop-colonies' seeded onto the medium from each strain was studied. There was a very marked difference between the two strains in the evolution of such 'drop-colonies', and it appeared that the lateral spread of fine rugosity from those of the Pasteur strain represented an enhanced ability of small numbers of bacilli to take up the nutrient.

The effects of short-term treatment with 25-hydroxy-vitamin D3 (25(OH)D3) on intestinal absorption of 47Ca were examined in 18 studies of normal subjects and 16 studies of patients with advanced renal failure. Doses of 25(OH)D3 were 20, 100, 500, or 1000 microgram/day given orally for 7--10 days. There was an increase in 47Ca absorption and urinary calcium in normal subjects receiving 20 microgram/day, while doses of 500 or 1000 microgram/day were required to augment 47Ca absorption in renal failure patients. During treatment, plasma levels of 25(OH)D increased to similar levels in both normal and uremic subjects. A comparison of the dose-response curves found 25(OH)D3 to be 1/125 as potent as 1,25-(OH)2D3 in the normal subjects and 1/400 as potent as 1,25(OH)2D3 in patients with chronic renal failure. Thus, pharmacologic doses of 25(OH)D3 are active in both normal and uremic patients, although relatively greater quantities are necessary in uremia. This difference in relative potency of 1,25(OH)2D3 and 25(OH)D3 may be explained by some conversion of 25(OH)D3 to 1,25(OH)2D3 in normal compared to uremic subjects, while 25(OH)D3 may act in large part via mass action in uremic patients.

To follow an earlier study of the effects on BCG of deep subculture in liquid media, transfers on the surface of Löwenstein-Jensen and Sauton's media have been made. When a vaccine preparation that yields on culture 99+% of spreading-type colonies, with a small minority of non-spreading forms, is transferred on Löwenstein-Jensen medium, the minority population can be readily selected out. Growth of BCG on this medium is not homogeneous, and the selection of either the minority or majority populations appears fortuitous, whereas it was previously demonstrated that deep subculture in Dubos medium tends to favour selection of a minority yielding non-spreading colonies. When such a minority population has been favoured during transfer on Löwenstein-Jensen medium, subsequent subculture in Dubos medium usually accentuates the effect. Subculture as a surface pellicle on Sauton's medium has a powerful effect in reducing the relative size of a minority population yielding non-spreading colonies, and thereafter maintaining 99+% of spreading forms. The relationship between transfers on Löwenstein-Jensen and Sauton's media and those on potato-Sauton medium is briefly discussed.

BCG originated from a virulent bovine strain of the tubercle bacillus after prolonged serial subculture on a potato medium. Since attenuation was achieved, the BCG strain has been distributed to a large number of centres where BCG vaccine is produced. Many of these production laboratories have maintained their BCG lines by continuing serial transfers, but have employed a variety of media for this purpose, and have produced BCG vaccine by a variety of techniques. Distinct differences have developed between some of the daughter strains of BCG, but the mechanism through which these changes have occurred has not been clear. In recent years methods have been developed which have enabled changes taking place within some BCG strains during experimental serial subculture to be monitored. In this survey the relationship of the changes observed to the different techniques employed for the maintenance of BCG lines and for the preparation of vaccine is considered. It is suggested that selection of minority populations within BCG strains noted during experimental studies may provide an analogy with the mechanism through which the original attenuation of the virulent bovine strain was brought about. The relevance of small-scale laboratory investigations to full-scale production procedures is also discussed, and finally some additional measures that might be taken to minimise changes in BCG strains are proposed.

A marked improvement in the ability to consistently distinguish between colonies cultured from BCG strain 1331 (Copenhagen), employed for the routine production of Danish freeze-dried BCG vaccine, and those cultured from BCG strain 1077 (Glaxo), employed for the routine production of British freeze-dried BCG vaccine, was obtained in our laboratory by modifying the constituents of a Dubos-type solid-medium. Several batches of British BCG vaccine had been specially prepared by the manufacturer, using strain 1331 as seed culture, and samples from these batches were cultured on this medium in our laboratory. Two of the batches produced, in place of the spreading colonies normally cultured from strain 1331, a large majority of nonspreading colonies, normally characteristic of strain 1077. To study this very rapid change, modifications of the system of serial transfers employed routinely by the manufacturer were set up in our laboratory, and the relative proportions of the two colony forms cultivated at each stage of transfer were noted. The rapid changes that had occurred during manufacture were reproduced under our experimental conditions, and it was observed that transfer through deep culture in liquid Dubos medium tended to shift the balance between the numbers of spreading and non-spreading colonies in favour of the latter, whilst the reverse was true of the transfers through deep culture in production medium. The ability to monitor changes occurring in BCG strains during serial transfer should facilitate the provision of measures to prevent such changes taking place.

CBA mice rendered immunodeficient by thymectomy, potentially lethal gamma-irradiation and reconstitution with bone marrow cells were used to grow a wide variety of human tumours as subcutaneous implants. Samples of human melanoma obtained at surgery were successfully passaged by transplantation and produced rapidly growing tumours, some of which metastasized to lung, lymph nodes and the para-aortic node; this system was used as a model for the study of immunotherapy of melanoma. Preliminary results show that intratumour injections of C. parvum retard or inhibit the growth of melanoma transplants and, therefore, do not require the involvement of T lymphocytes, whilst BCG has no effect on growth rate.