LED Fluorescence Microscopy for the Diagnosis of Pulmonary Tuberculosis: A Multi-Country Cross-Sectional Evaluation

Background

The diagnosis of tuberculosis (TB) in resource-limited settings relies on Ziehl-Neelsen (ZN) smear microscopy. LED fluorescence microscopy (LED-FM) has many potential advantages over ZN smear microscopy, but requires evaluation in the field. The aim of this study was to assess the sensitivity/specificity of LED-FM for the diagnosis of pulmonary TB and whether its performance varies with the timing of specimen collection.

Methods and Findings

Adults with cough ≥2 wk were enrolled consecutively in Ethiopia, Nepal, Nigeria, and Yemen. Sputum specimens were examined by ZN smear microscopy and LED-FM and compared with culture as the reference standard. Specimens were collected using a spot-morning-spot (SMS) or spot-spot-morning (SSM) scheme to explore whether the collection of the first two smears at the health care facility (i.e., “on the spot”) the first day of consultation followed by a morning sample the next day (SSM) would identify similar numbers of smear-positive patients as smears collected via the SMS scheme (i.e., one on-the-spot-smear the first day, followed by a morning specimen collected at home and a second on-the-spot sample the second day). In total, 529 (21.6%) culture-positive and 1,826 (74.6%) culture-negative patients were enrolled, of which 1,156 (49%) submitted SSM specimens and 1,199 (51%) submitted SMS specimens. Single LED-FM smears had higher sensitivity but lower specificity than single ZN smears. Using two LED-FM or two ZN smears per patient was 72.8% (385/529, 95% CI 68.8%–76.5%) and 65.8% (348/529, 95% CI 61.6%–69.8%) sensitive (p<0.001) and 90.9% (1,660/1,826, 95% CI 89.5%–92.2%) and 98% (1,790/1,826, 95% CI 97.3%–98.6%) specific (p<0.001). Using three LED-FM or three ZN smears per patient was 77% (408/529, 95% CI 73.3%–80.6%) and 70.5% (373/529, 95% CI 66.4%–74.4%, p<0.001) sensitive and 88.1% (95% CI 86.5%–89.6%) and 96.5% (95% CI 96.8%–98.2%, p<0.001) specific. The sensitivity/specificity of ZN smear microscopy and LED-FM did not vary between SMS and SSM.

Conclusions

LED-FM had higher sensitivity but, in this study, lower specificity than ZN smear microscopy for diagnosis of pulmonary TB. Performance was independent of the scheme used for collecting specimens. The introduction of LED-FM needs to be accompanied by appropriate training, quality management, and monitoring of performance in the field.

Trial Registration

Current Controlled Trials ISRCTN53339491 Please see later in the article for the Editors'' Summary     

Options for active case detection of visceral leishmaniasis in endemic districts of India, Nepal and Bangladesh, comparing yield, feasibility and costs

Background

The VL elimination strategy requires cost-effective tools for case detection and management. This intervention study tests the yield, feasibility and cost of 4 different active case detection (ACD) strategies (camp, index case, incentive and blanket approach) in VL endemic districts of India, Nepal and Bangladesh.

Methodology/Principal Findings

First, VL screening (fever more than 14 days, splenomegaly, rK39 test) was performed in camps. This was followed by house to house screening (blanket approach). An analysis of secondary VL cases in the neighborhood of index cases was simulated (index case approach). A second screening round was repeated 4–6 months later. In another sub-district in India and Nepal, health workers received incentives for detecting new VL cases over a 4 month period (incentive approach). This was followed by house screening for undetected cases. A total of 28 new VL cases were identified by blanket approach in the 1^st screening round, and used as ACD gold standard. Of these, the camp approach identified 22 (sensitivity 78.6%), index case approach identified 12 (sensitivity – 42.9%), and incentive approach identified 23 new VL cases out of 29 cases detected by the house screening (sensitivity – 79.3%). The effort required to detect a new VL case varied (blanket approach – 1092 households, incentive approach – 978 households; index case approach – 788 households had to be screened). The cost per new case detected varied (camp approach $21 – $661; index case approach $149 – $200; incentive based approach $50 – $543; blanket screening $112 – $629). The 2^nd screening round yielded 20 new VL cases. Sixty and nine new PKDL cases were detected in the first and second round respectively.

Conclusions/Significance

ACD in the VL elimination campaign has a high yield of new cases at programme costs which vary according to the screening method chosen. Countries need the right mix of approaches according to the epidemiological profile, affordability and organizational feasibility.     

An annotated checklist of the orchids of Nepal

An annotated checklist comprised of 458 taxa of orchids known from Nepal is provided, including 104 genera, 437 species, 16 varieties, 3 subspecies and 2 forma and 18 endemic species. In Nepal, orchid species are distributed from 60–5200 m a.s.l. In the checklist, notes on altitudinal ranges, habit, habitat, global distribution, phenology, etc. are presented.     

Exogenous application of L-phenylalanine and ferulic acid enhance phenylalanine ammonia lyase activity and accumulation of phenolic acids in pea (Pisum sativum) to offer protection against Erysiphe pisi

Phenylalanine ammonia lyase (PAL) activity was measured using HPLC in pea leaves following exogenous application of L-phenylalanine and ferulic acid. Treatment with different concentrations (50, 100, 150 ppm) of L-phenylalanine caused increased activity of PAL activity in comparison to control. In pea leaves treated with 50 ppm L-phenylalanine, maximum PAL activity was observed after 72 h of treatment. Application of ferulic acid first reduced PAL activity at lower concentration (50 ppm) but it further increased at higher concentrations of the compound (100 and 150 ppm) in pea leaves compared to control. Minimum PAL activity was 0.19 nM cinnamic acid/min/g fresh wt after 24 h at 50 ppm and then increased with time. Treatment with both compounds significantly increased the accumulation of phenolic acids and salicylic acid and reduced conidial germination of Erysiphe pisi on pea leaves. They were equally effective at 100 and 150 ppm in reducing conidial germination. Conidial germination on L-phenylalanine-treated leaves was 26% after 24 h and that on ferulic acid treated leaves 34% compared to control (46%). Foliar application of different concentrations of L-phenylalanine increased the level of ferulic acid in the leaves of pea plants. Maximum enzyme activity in terms of the accumulation of cinnamic acid (79.3 and 83.5 μg/g fresh wt) was observed following the application of L-phenylalanine after 24 and 48 h respectively. At 50 ppm, cinnamic acid accumulation in pea leaves was 35.6 and 39.4 μg/g fresh wt and 74.3 and 86.5 μg/g fresh wt at 100 ppm.     

Availability and Readability of Emergency Preparedness Materials for Deaf and Hard-of-Hearing and Older Adult Populations: Issues and Assessments

A major public health challenge is to communicate effectively with vulnerable populations about preparing for disasters and other health emergencies. People who are Deaf or Hard of Hearing (Deaf/HH) and older adults are particularly vulnerable during health emergencies and require communications that are accessible and understandable. Although health literacy studies indicate that the readability of health communication materials often exceeds people’s literacy levels, we could find no research about the readability of emergency preparedness materials (EPM) intended for Deaf/HH and older adult populations. The objective of this study was to explore issues related to EPM for Deaf/HH and older adult populations, to assess the availability and readability of materials for these populations, and to recommend improvements. In two California counties, we interviewed staff at 14 community-based organizations (CBOs) serving Deaf/HH clients and 20 CBOs serving older adults selected from a stratified, random sample of 227 CBOs. We collected 40 EPM from 10 CBOs and 2 public health departments and 40 EPM from 14 local and national websites with EPM for the public. We used computerized assessments to test the U.S. grade reading levels of the 16 eligible CBO and health department EPM, and the 18 eligible website materials. Results showed that less than half of CBOs had EPM for their clients. All EPM intended for clients of Deaf/HH-serving CBOs tested above the recommended 4^th grade reading level, and 91% of the materials intended for clients of older adult-serving CBOs scored above the recommended 6^th grade level. EPM for these populations should be widely available through CBOs and public health departments, adhere to health literacy principles, and be accessible in alternative formats including American Sign Language. Developers should engage the intended users of EPM as co-designers and testers. This study adds to the limited literature about EPM for these populations.     

Proteomic Analysis of Signaling Network Regulation in Renal Cell Carcinomas with Differential Hypoxia-Inducible Factor-2α Expression

Background

The loss of von Hippel–Lindau (VHL) protein function leads to highly vascular renal tumors characterized by an aggressive course of disease and refractoriness to chemotherapy and radiotherapy. Loss of VHL in renal tumors also differs from tumors of other organs in that the oncogenic cascade is mediated by an increase in the levels of hypoxia-inducible factor-2α (HIF2α) instead of hypoxia-inducible factor-1α (HIF1α).

Methods and Principal Findings

We used renal carcinoma cell lines that recapitulate the differences between mutant VHL and wild-type VHL genotypes. Utilizing a method relying on extracted peptide intensities as a label-free approach for quantitation by liquid chromatography–mass spectrometry, our proteomics study revealed regulation of key proteins important for cancer cell survival, proliferation and stress-resistance, and implicated differential regulation of signaling networks in VHL-mutant renal cell carcinoma. We also observed upregulation of cellular energy pathway enzymes and the stress-responsive mitochondrial 60-kDa heat shock protein. Finding reliance on glutaminolysis in VHL-mutant renal cell carcinoma was of particular significance, given the generally predominant dependence of tumors on glycolysis. The data have been deposited to the ProteomeXchange with identifier PXD000335.

Conclusions and Significance

Pathway analyses provided corroborative evidence for differential regulation of molecular and cellular functions influencing cancer energetics, metabolism and cell proliferation in renal cell carcinoma with distinct VHL genotype. Collectively, the differentially regulated proteome characterized by this study can potentially guide translational research specifically aimed at effective clinical interventions for advanced VHL-mutant, HIF2α-over-expressing tumors.     

Genetic diversity in Capsicum germplasm based on microsatellite and random amplified microsatellite polymorphism markers

A sound knowledge of the genetic diversity among germplasm is vital for strategic germplasm collection, maintenance, conservation and utilisation. Genomic simple sequence repeats (SSRs) and random amplified microsatellite polymorphism (RAMPO) markers were used to analyse diversity and relationships among 48 pepper (Capsicum spp.) genotypes originating from nine countries. These genotypes covered 4 species including 13 germplasm accessions, 30 improved lines of 4 domesticated species and 5 landraces derived from natural interspecific crosses. Out of 106 SSR markers, 25 polymorphic SSR markers (24 %) detected a total of 76 alleles (average, 3.04; range, 2–5). The average polymorphic information content (PIC) was 0.69 (range, 0.29–0.92). Seventeen RAMPO markers produced 87 polymorphic fragments with average PIC of 0.63 (range, 0.44–0.81). Dendrograms based on SSRs and RAMPOs generated two clusters. All 38 Capsicum annuum genotypes and an interspecific landrace clustered together, whereas nine non-annuum (three Capsicum frutescens, one Capsicum chinense, one Capsicum baccatum and four interspecific landraces) genotypes clustered separately. Genetic variation within non-annuum genotypes was greater than the C. annuum genotypes. Distinctness of interspecific derivative landraces grown in northeast India was validated; natural crossing between sympatric Capsicum species has been proposed as the mechanism of their origin.     

Differential Reprogramming of Defense Network in Capsicum annum L. Plants Against Colletotrichum truncatum Infection by Phyllospheric and Rhizospheric Trichoderma Strains

Journal of Plant Growth Regulation - Induction of defense response in host plants by the Trichoderma spp. has been attributed as one of the major mechanisms leading to inhibition of the pathogenic...