Combined Rapid (TUBEX) Test for Typhoid-Paratyphoid A Fever Based on Strong Anti-O12 Response: Design and Critical Assessment of Sensitivity

Rapid diagnostics can be accurate but, often, those based on antibody detection for infectious diseases are unwittingly underrated for various reasons. Herein, we described the development of a combined rapid test for two clinically-indistinguishable bacterial diseases, typhoid and paratyphoid A fever, the latter fast emerging as a global threat. By using monoclonal antibodies (mAbs) to bacterial antigens of known chemical structures as probes, we were able to dissect the antibody response in patients at the level of monosaccharides. Thus, a mAb specific for a common lipopolysaccharide antigen (O12) found in both the causative organisms was employed to semi-quantify the amounts of anti-O12 antibodies present in both types of patients in an epitope-inhibition particle-based (TUBEX) immunoassay. This colorimetric assay detected not only anti-O12 antibodies that were abundantly produced, but also, by steric hindrance, antibodies to an adjoining epitope (O9 or O2 in the typhoid or paratyphoid bacillus, respectively). Sensitivity and, particularly, reaction intensities, were significantly better than those obtained using an anti-O9 or anti-O2 mAb-probe in the examination of paired sera from 22 culture-confirmed typhoid patients (sensitivity, 81.8% vs 75.0%) or single sera from 36 culture-confirmed paratyphoid patients (52.8% vs 28.6), respectively. Importantly, sensitivity was better (97.1% for typhoid, 75.0% for paratyphoid) if allowance was made for the absence of relevant antibodies in certain specimens as determined by an independent, objective assay (ELISA) — such specimens might have been storage-denatured (especially the older paratyphoid samples) or procured from non-responders. Benchmarking against ELISA, which revealed high concordance between the two tests, was useful and more appropriate than comparing with culture methods as traditionally done, since antibody tests and culture target slightly different stages of these diseases. Paired sera analysis was insightful, revealing 64% of typhoid patients who had no change in antibody titer over 4–16 days, and 14% with no IgM-IgG class-switching

Moderate and heavy metabolic stress interval training improve arterial stiffness and heart rate dynamics in humans

Traditional continuous aerobic exercise training attenuates age-related increases of arterial stiffness, however, training studies have not determined whether metabolic stress impacts these favourable effects. Twenty untrained healthy participants (n = 11 heavy metabolic stress interval training, n = 9 moderate metabolic stress interval training) completed 6 weeks of moderate or heavy intensity interval training matched for total work and exercise duration. Carotid artery stiffness, blood pressure contour analysis, and linear and non-linear heart rate variability were assessed before and following training. Overall, carotid arterial stiffness was reduced (p  0.05). This study demonstrates the effectiveness of interval training at improving arterial stiffness and autonomic function, however, the metabolic stress was not a mediator of this effect. In addition, these changes were also independent of improvements in aerobic capacity, which were only induced by training that involved a high metabolic stress

Menstruation angina: a case report

<p>Abstract</p> <p>Introduction</p> <p>Menstruation is commonly associated with migraine and irritable bowel but is rarely correlated with angina or myocardial ischaemia. Only a small number of cases have been reported suggesting a link between menstruation and myocardial ischaemic events.</p> <p>Case presentation</p> <p>A case of menstruation angina is reported in order to raise awareness of this association. A 47-year-old South Asian woman presented with recurrent chest pains in a monthly fashion coinciding with her menstruations. Each presentation was associated with troponin elevation. Angioplasty failed to resolve her symptoms but she eventually responded to hormonal therapy.</p> <p>Conclusions</p> <p>The possibility of menstruation angina should always be taken into account in any female patients from puberty to menopause presenting with recurrent chest pains. This can allow an earlier introduction of hormonal therapy to arrest further myocardial damage.</p

TrpC3 Regulates Hypertrophy-Associated Gene Expression without Affecting Myocyte Beating or Cell Size

Pathological cardiac hypertrophy is associated with an increased risk of heart failure and cardiovascular mortality. Calcium (Ca2+) -regulated gene expression is essential for the induction of hypertrophy, but it is not known how myocytes distinguish between the Ca2+ signals that regulate contraction and those that lead to cardiac hypertrophy. We used in vitro neonatal rat ventricular myocytes to perform an RNA interference (RNAi) screen for ion channels that mediate Ca2+-dependent gene expression in response to hypertrophic stimuli. We identified several ion channels that are linked to hypertrophic gene expression, including transient receptor potential C3 (TrpC3). RNAi-mediated knockdown of TrpC3 decreases expression of hypertrophy-associated genes such as the A- and B-type natriuretic peptides (ANP and BNP) in response to numerous hypertrophic stimuli, while TrpC3 overexpression increases BNP expression. Furthermore, stimuli that induce hypertrophy dramatically increase TrpC3 mRNA levels. Importantly, whereas TrpC3-knockdown strongly reduces gene expression associated with hypertrophy, it has a negligible effect on cell size and on myocyte beating. These results suggest that Ca2+ influx through TrpC3 channels increases transcription of genes associated with hypertrophy but does not regulate the signaling pathways that control cell size or contraction. Thus TrpC3 may represent an important therapeutic target for the treatment of cardiac hypertrophy and heart failure

Substance use and sexual behaviours of Japanese men who have sex with men: A nationwide internet survey conducted in Japan

BACKGROUND: Japanese men who have sex with men (MSM), especially those living in large metropolitan areas such as Tokyo and Osaka, are facing a growing HIV/AIDS epidemic. Although the Internet is used as a new venue for meeting sex partners, it can also serve as a useful research tool for investigating the risk behaviours of Japanese MSM. This Internet survey explored the extent of substance use and its association with sexual risk behaviours among Japanese MSM. METHODS: Between 28 February 2003 and 16 May 2003 MSM were recruited through 57 Japanese gay-oriented Web sites, gay magazines, and Internet mailing lists. Participants completed a structured questionnaire anonymously through the Internet. RESULTS: In total, 2,062 Japanese MSM completed the questionnaire. The average age of participants was 29.0 years and 70.5% identified as gay, 20.8% as bisexual, and 8.7% as other. Overall, 34.5% reported never using a substance, 45% reported ever using one type of substance (lifetime reported single substance users), and 19.6% had used more than 1 type of substance (lifetime reported multiple substance users) in their lifetimes. The substances most commonly used were amyl nitrite (63.2%), 5-methoxy-N, N-diisopropyltryptamine (5MEO-DIPT) (9.3%), and marijuana (5.7%). In the multivariate analysis, unprotected anal intercourse, having had 6 or more sexual partners, visiting a sex club/gay venue in the previous 6 months, a lower education level, and being 30 to 39 years of age were associated with both lifetime single and lifetime multiple substance use. Lifetime reported multiple substance use was also correlated with having a casual sex partner, having symptoms of depression, being diagnosed as HIV-positive, and greater HIV/AIDS-related knowledge. CONCLUSION: This is the first Internet-based research focused on the sexual and substance use behaviours of MSM in Asia. Our findings suggest a compelling need for prevention interventions to reduce HIV risk-related substance use behaviours among Japanese MSM. The results also suggest that the Internet is potentially a useful tool for collecting behavioural data and promoting prevention interventions among this population

Assessment and comparative analysis of a rapid diagnostic test (Tubex®) for the diagnosis of typhoid fever among hospitalized children in rural Tanzania

Background: Typhoid fever remains a significant health problem in many developing countries. A rapid test with a performance comparable to that of blood culture would be highly useful. A rapid diagnostic test for typhoid fever, Tubex&reg;, is commercially available that uses particle separation to detect immunoglobulin M directed towards Salmonella Typhi O9 lipopolysaccharide in sera.Methods: We assessed the sensitivity and specificity of the Tubex test among Tanzanian children hospitalized with febrile illness using blood culture as gold standard. Evaluation was done considering blood culture confirmed S. Typhi with non-typhi salmonella (NTS) and non - salmonella isolates as controls as well as with non-salmonella isolates only.Results: Of 139 samples tested with Tubex, 33 were positive for S. Typhi in blood culture, 49 were culture-confirmed NTS infections, and 57 were other non-salmonella infections. Thirteen hemolyzed samples were excluded. Using all non - S. Typhi isolates as controls, we showed a sensitivity of 79% and a specificity of 89%. When the analysis was repeated excluding NTS from the pool of controls we showed a sensitivity of 79% and a specificity of 97%. There was no significant difference in the test performance using the two different control groups (p &gt; 0.05).Conclusion: This first evaluation of the Tubex test in an African setting showed a similar performance to those seen in some Asian settings. Comparison with the earlier results of a Widal test using the same samples showed no significant difference (p &gt; 0.05) for any of the performance indicators, irrespective of the applied control group

Induction of Erythroid Differentiation in Human Erythroleukemia Cells by Depletion of Malic Enzyme 2

Malic enzyme 2 (ME2) is a mitochondrial enzyme that catalyzes the conversion of malate to pyruvate and CO2 and uses NAD as a cofactor. Higher expression of this enzyme correlates with the degree of cell de-differentiation. We found that ME2 is expressed in K562 erythroleukemia cells, in which a number of agents have been found to induce differentiation either along the erythroid or the myeloid lineage. We found that knockdown of ME2 led to diminished proliferation of tumor cells and increased apoptosis in vitro. These findings were accompanied by differentiation of K562 cells along the erythroid lineage, as confirmed by staining for glycophorin A and hemoglobin production. ME2 knockdown also totally abolished growth of K562 cells in nude mice. Increased ROS levels, likely reflecting increased mitochondrial production, and a decreased NADPH/NADP+ ratio were noted but use of a free radical scavenger to decrease inhibition of ROS levels did not reverse the differentiation or apoptotic phenotype, suggesting that ROS production is not causally involved in the resultant phenotype. As might be expected, depletion of ME2 induced an increase in the NAD+/NADH ratio and ATP levels fell significantly. Inhibition of the malate-aspartate shuttle was insufficient to induce K562 differentiation. We also examined several intracellular signaling pathways and expression of transcription factors and intermediate filament proteins whose expression is known to be modulated during erythroid differentiation in K562 cells. We found that silencing of ME2 leads to phospho-ERK1/2 inhibition, phospho-AKT activation, increased GATA-1 expression and diminished vimentin expression. Metabolomic analysis, conducted to gain insight into intermediary metabolic pathways that ME2 knockdown might affect, showed that ME2 depletion resulted in high orotate levels, suggesting potential impairment of pyrimidine metabolism. Collectively our data point to ME2 as a potentially novel metabolic target for leukemia therapy

A novel underuse model shows that inactivity but not ovariectomy determines the deteriorated material properties and geometry of cortical bone in the tibia of adult rats

Our goal in this study was to determine to what extent the physiologic consequences of ovariectomy (OVX) in bones are exacerbated by a lack of daily activity such as walking. We forced 14-week-old female rats to be inactive for 15 weeks with a unique experimental system that prevents standing and walking while allowing other movements. Tibiae, femora, and 4th lumbar vertebrae were analyzed by peripheral quantitative computed tomography (pQCT), microfocused X-ray computed tomography (micro-CT), histology, histomorphometry, Raman spectroscopy, and the three-point bending test. Contrary to our expectation, the exacerbation was very much limited to the cancellous bone parameters. Parameters of femur and tibia cortical bone were affected by the forced inactivity but not by OVX: (1) cross-sectional moment of inertia was significantly smaller in Sham-Inactive rat bones than that of their walking counterparts; (2) the number of sclerostin-positive osteocytes per unit cross-sectional area was larger in Sham-Inactive rat bones than in Sham-Walking rat bones; and (3) material properties such as ultimate stress of inactive rat tibia was lower than that of their walking counterparts. Of note, the additive effect of inactivity and OVX was seen only in a few parameters, such as the cancellous bone mineral density of the lumbar vertebrae and the structural parameters of cancellous bone in the lumbar vertebrae/tibiae. It is concluded that the lack of daily activity is detrimental to the strength and quality of cortical bone in the femur and tibia of rats, while lack of estrogen is not. Our inactive rat model, with the older rats, will aid the study of postmenopausal osteoporosis, the etiology of which may be both hormonal and mechanical

Application of the rainbow trout derived intestinal cell line (RTgutGC) for ecotoxicological studies: molecular and cellular responses following exposure to copper.

There is an acknowledged need for in vitro fish intestinal model to help understand dietary exposure to chemicals in the aquatic environment. The presence and use of such models is however largely restrictive due to technical difficulties in the culturing of enterocytes in general and the availability of appropriate established cell lines in particular. In this study, the rainbow trout (Oncorhynchus mykiss) intestinal derived cell line (RTgutGC) was used as a surrogate for the "gut sac" method. To facilitate comparison, RTgutGC cells were grown as monolayers (double-seeded) on permeable Transwell supports leading to a two-compartment intestinal model consisting of polarised epithelium. This two-compartment model divides the system into an upper apical (lumen) and a lower basolateral (portal blood) compartment. In our studies, these cells stained weakly for mucosubstances, expressed the tight junction protein ZO-1 in addition to E-cadherin and revealed the presence of polarised epithelium in addition to microvilli protrusions. The cells also revealed a comparable transepithelial electrical resistance (TEER) to the in vivo situation. Importantly, the cell line tolerated apical saline (1:1 ratio) thus mimicking the intact organ to allow assessment of uptake of compounds across the intestine. Following an exposure over 72 h, our study demonstrated that the RTgutGC cell line under sub-lethal concentrations of copper sulphate (Cu) and modified saline solutions demonstrated uptake of the metal with saturation levels comparable to short term ex situ gut sac preparations. Gene expression analysis revealed no significant influence of pH or time on mRNA expression levels of key stress related genes (i.e. CYP3A, GST, mtA, Pgp and SOD) in the Transwell model. However, significant positive correlations were found between all genes investigated suggesting a co-operative relationship amongst the genes studied. When the outlined characteristics of the cell line are combined with the division of compartments, the RTgutGC double seeded model represents a potential animal replacement model for ecotoxicological studies. Overall, this model could be used to study the effects and predict aquatic gastrointestinal permeability of metals and other environmentally relevant contaminants in a cost effective and high throughput manner