Enhancing TB Case Detection: Experience in Offering Upfront Xpert MTB/RIF Testing to Pediatric Presumptive TB and DR TB Cases for Early Rapid Diagnosis of Drug Sensitive and Drug Resistant TB

Background

Diagnosis of pulmonary tuberculosis (PTB) in children is challenging due to difficulties in obtaining good quality sputum specimens as well as the paucibacillary nature of disease. Globally a large proportion of pediatric tuberculosis (TB) cases are diagnosed based only on clinical findings. Xpert MTB/RIF, a highly sensitive and specific rapid tool, offers a promising solution in addressing these challenges. This study presents the results from pediatric groups taking part in a large demonstration study wherein Xpert MTB/RIF testing replaced smear microscopy for all presumptive PTB cases in public health facilities across India.

Methods

The study covered a population of 8.8 million across 18 programmatic sub-district level tuberculosis units (TU), with one Xpert MTB/RIF platform established at each study TU. Pediatric presumptive PTB cases (both TB and Drug Resistant TB (DR-TB)) accessing any public health facilities in study area were prospectively enrolled and tested on Xpert MTB/RIF following a standardized diagnostic algorithm.

Results

4,600 pediatric presumptive pulmonary TB cases were enrolled. 590 (12.8%, CI 11.8–13.8) pediatric PTB were diagnosed. Overall 10.4% (CI 9.5–11.2) of presumptive PTB cases had positive results by Xpert MTB/RIF, compared with 4.8% (CI 4.2–5.4) who had smear-positive results. Upfront Xpert MTB/RIF testing of presumptive PTB and presumptive DR-TB cases resulted in diagnosis of 79 and 12 rifampicin resistance cases, respectively. Positive predictive value (PPV) for rifampicin resistance detection was high (98%, CI 90.1–99.9), with no statistically significant variation with respect to past history of treatment.

Conclusion

Upfront access to Xpert MTB/RIF testing in pediatric presumptive PTB cases was associated with a two-fold increase in bacteriologically-confirmed PTB, and increased detection of rifampicin-resistant TB cases under routine operational conditions across India. These results suggest that routine Xpert MTB/RIF testing is a promising solution to present-day challenges in the diagnosis of PTB in pediatric patients.     

Purification and characterization of trypsin inhibitor from seeds of faba bean (Vicia faba L.)

A trypsin inhibitor from seeds of faba bean (Vicia faba L.) was purified to near homogeneity as judged by native-PAGE with about 11 % recovery using ammonium sulphate fractionation, ion-exchange chromatography on DEAE-cellulose and gel filtration through Sephadex G-100. The inhibitor had a molecular weight of 18 kD as determined by SDS-PAGE and Sephadex G-100. The inhibitor inhibited trypsin and chymotrypsin to the extent of 48 and 12 %, respectively. The inhibtion was of non-competitive type with dissociation constant for the enzyme inhibitor complex in the region of 0.07 mg·ml⁻¹. The inhibtor was stable between pH 4 and 5. It completely lost its activity when heated at 125 °C for 1 h or at 100 °C for 2 h. The inhibitor also lost its activity on exposure to 2-mercaptoethanol. Based on these properties, it could be concluded that Vicia faba trypsin inhibitor belongs to Bowman-Birk type of inhibitors, as it has molecular weight lower than generally observed for Kunitz type inhibitors.     

A protocol for in vitro mass propagation of asiatic hybrids of lily through liquid stationary culture

Summary A simple, rapid and cost-effective in vitro scheme has been proposed for mass propagating two cultivars of Asiatic lily hybrids. An average of seven bulblets was formed after 17 d when 1×1 cm² bulb scale segments (explants) were cultured on Murashige and Skoog (MS) medium with 3% sucrose and 0.5 μM α-naphthaleneacetic acid (NAA). On MS medium containing 0.5 μM NAA and 6 or 9% sucrose, depending on the cultivar, large numbers of bulblets of increased size (3.5–5.0 cm in circumference) were formed under a 16/8 h photoperiod. A continuous system of mass propagation of bulblets was achieved through in vitro scale formation (secondary explants) on MS medium supplemented with 23 μM kinetin and 0.5 μM NAA, as well as scale proliferation on MS basal liquid stationary medium. Upon transplantation all bulblets sprouted, of which 40% flowered in the first season. Under ideal conditions, ca. 9.68×10⁵ bulblets can be produced from a single scale segment in 1 yr by following the systematic propagation steps proposed here.     

Regulation of alpha 1 (I)-collagen gene expression in response to cell adhesion in Swiss 3T3 fibroblasts

Nonadhesive conditions cause Swiss 3T3 fibroblasts to enter a quiescent state that is reversed upon reattachment to a surface. Previously, we demonstrated that adhesion in serum-free conditions is sufficient to activate suspension-arrested cells out of Go, with the induction of the growth-associated genes, c-fos, c-myc, and actin. In this study, we have employed this system to identify programs of gene expression that respond primarily to the adhesive state of the cell, rather than the growth state. We show that cells in different adhesive states can be distinguished by their patterns of protein synthesis. Analysis of one adhesion-responsive protein led to its identification as pro-alpha 1 (I)-collagen. Pro-alpha 1 (I)-collagen synthesis and mRNA levels are decreased up to 6-fold in suspension-arrested fibroblasts, but are enhanced up to 5-fold as cells approach confluence. This suggests that the reduced expression in suspension-arrested cells is not simply a result of quiescence. In addition, reattachment of suspended cells in serum-free conditions caused a 7-fold induction of collagen mRNA levels and a greater than 20-fold rise in the rate of procollagen synthesis. The expression of c-myc was induced during adhesion in serum-free medium as well as by serum addition to suspension-arrested cells. However, alpha 1 (I)-collagen gene expression was unaffected by serum in the absence of adhesion. These results indicate that collagen gene expression is directly responsive to cell adhesion, independent of the growth state.     

Short-term therapy of atherosclerosis with low dose indomethacin: an experimental study

The effects of low dose indomethacin therapy in primary prevention of diet-induced atherosclerosis of rhesus monkeys was studied. The parameters studied were serum cholesterol concentration, thromboxane A2 (T x B2), prostacyclin (6-keto-PGF1 alpha) in serum/plasma, and the extent and intensity of coronary atherosclerosis. Although indomethacin did not affect serum cholesterol, it reduced serum T x B2 significantly (P less than 0.01). Plasma 6-keto-PGF1 alpha was not restored to the pretreatment level. A significant protective role of the drug was noted as far as coronary atherosclerosis is concerned (P less than 0.01).     

In vitro vegetative propagation of Leucaena leucocephala (Lam.) de Wit

A method for in vitro clonal multiplication of Leucaena leucocephala cv K-8 is described. On MS with BAP (3×10^–6M), at the optimum temperature of 30°C, shoots from seedling and adult trees multiplied at a rate of 6–7 fold every three weeks. The addition of adenine or glutamine reduced precocious leaf drop. All shoots rooted on MS with IAA (5×10^–6M). Micropropagated plants have been successfully transferred to soil.     

Lithium Treatment Aggregates the Adverse Effects on Erythrocytes Subjected to Arsenic Exposure

The present study was designed to investigate the effects of lithium treatment on red blood cells which were given arsenic exposure. Long-term lithium therapy is being extensively used for the treatment of bipolar disorders. Arsenic is a group I carcinogen and a major toxic pollutant in drinking water that affects millions of people worldwide. Male SD rats were segregated into four groups, viz. normal control, lithium treated, arsenic treated, and lithium + arsenic treated. Lithium was supplemented as lithium carbonate at a dose level of 1.1 g/kg diet for a period of 8 weeks. Arsenic was given in the form of sodium arsenite at a dose level of 100 ppm in drinking water, ad libitum, for the same period. Lysates of red blood cells were used to investigate the effects of lithium and arsenic treatments on anti-oxidant enzymes, reduced glutathione (GSH), and lipid peroxidation (LPO) levels. Various hematological parameters, activities of Na⁺ K⁺ ATPase and delta-aminolevulinic acid dehydratase (δ-ALAD) were also assessed. A significant reduction was observed in the activities of antioxidant enzymes, GSH levels, total erythrocyte counts, Na⁺ K⁺ ATPase, and ALAD enzyme activities in lysates of red blood cells when exposed either to lithium or arsenic. In addition, a significant increase in the levels of malondialdehyde (MDA), lymphocytes, neutrophils, and total leukocytes was also observed following lithium as well as arsenic treatments. However, when arsenic-treated rats were subjected to lithium treatment, a pronounced alteration was noticed in all the above parameters. Therefore, we conclude that lithium supplementation to the arsenic-treated rats enhances the adverse effects on red blood cells and therefore use of lithium may not be medicated to patients who are vulnerable to arsenic exposure through drinking water. It can also be inferred that adverse effects of lithium therapy may get aggravated in patients thriving in the arsenic-contaminated area.

     

Effect of lithium on hepatic drug-metabolizing enzymes of protein-deficient rats

Protein deficiency was produced by feeding synthetic 8%-protein diet. Lithium carbonate at the dose level of 1.1g/kg diet was administered to normal and protein-deficient rats for a period of one mo. A significant inhibition in the levels of cytochrome (cyt) P₄₅₀, cyt b₅, glutathione (GSH), glutathione S-transferase (GST) and glutathione peroxidase (GPx), but an increase in γ-glutamyl transpeptidase (γ-GT), was observed in low-protein LP-fed rats. Lithium treatment to normal rats caused no significant change in the activities of cyt P₄₅₀, cyt b₅, GST, and GSH levels, whereas there was elevation in the activities of γ-GT and GPx and suppression in glutathione reductase (GRd) activity. Lithium administration to LP-fed rats resulted in significant increases in the hepatic γ-GT and GPx activities.     

<Emphasis Type="Italic">In Vitro</Emphasis> and <Emphasis Type="Italic">Ex Vivo</Emphasis> Evaluations of Lipid Anti-Cancer Nanoformulations: Insights and Assessment of Bioavailability Enhancement

Lipid-based nanoformulations have been extensively investigated for improving oral efficacy of plethora of drugs. Chemotherapeutic agents remain a preferred option for effective management of cancer; however, most chemotherapeutic agents suffer from limitation of poor oral bioavailability that is associated with their physicochemical properties. Drug delivery via lipid-based nanosystems possesses strong rational and potential for improving oral bioavailability of such anti-cancer molecules through various mechanisms, viz. improving their gut solubilisation owing to micellization, improving mucosal permeation, improving lymphatic uptake, inhibiting intestinal metabolism and/or inhibiting P-glycoprotein efflux of molecules in the gastrointestinal tract. Various in vitro characterization techniques have been reported in literature that aid in getting insights into mechanisms of lipid-based nanodevices in improving oral efficacy of anti-cancer drugs. The review focuses on different characterization techniques that can be employed for evaluation of lipid-based nanosystems and their role in effective anti-cancer drug delivery.