Enhanced Case Detection and Improved Diagnosis of PKDL in a Kala-azar-Endemic Area of Bangladesh

Objectives

To support the Bangladesh National Kala-azar Elimination Programme (NKEP), we investigated the feasibility of using trained village volunteers for detecting post-kala-azar dermal leishmaniasis (PKDL) cases, using polymerase chain reaction (PCR) for confirmation of diagnosis and treatment compliance by PKDL patients in Kanthal union of Trishal sub-district, Mymensingh, Bangladesh.

Methods

In this cross-sectional study, Field Research Assistants (FRAs) conducted census in the study area, and the research team trained village volunteers on how to look for PKDL suspects. The trained village volunteers (TVVs) visited each household in the study area for PKDL suspects and referred the suspected PKDL cases to the study clinic. The suspected cases underwent physical examinations by a qualified doctor and rK39 strip testing by the FRAs and, if positive, slit skin examination (SSE), culture, and PCR of skin specimens and peripheral buffy coat were done. Those with evidence of Leishmania donovani (LD) were referred for treatment. All the cases were followed for one year.

Results

The total population of the study area was 29,226 from 6,566 households. The TVVs referred 52 PKDL suspects. Probable PKDL was diagnosed in 18 of the 52 PKDL suspect cases, and PKDL was confirmed in 9 of the 18 probable PKDL cases. The prevalence of probable PKDL was 6.2 per 10,000 people in the study area. Thirteen PKDL suspects self-reported from outside the study area, and probable and confirmed PKDL was diagnosed in 10 of the 13 suspects and in 5 of 10 probable PKDL cases respectively. All probable PKDL cases had hypopigmented macules. The median time for PKDL development was 36 months (IQR, 24–48). Evidence of the LD parasite was documented by SSE and PCR in 3.6% and 64.3% of the cases, respectively. PCR positivity was associated with gender and severity of disease. Those who were untreated had an increased risk (odds ratio = 3.33, 95%CI 1.29–8.59) of having persistent skin lesions compared to those who were treated. Patients'' treatment-seeking behavior and treatment compliance were poor.

Conclusion

Improved detection of PKDL cases by TVVs is feasible and useful. The NKEP should promote PCR for the diagnosis of PKDL and should find ways for improving treatment compliance by patients.     

The phage display of Bacillus subtilis Lipase A significantly enhances catalytic activity due to altered nanoscale distribution in colloidal solution

Screening libraries of mutant proteins by phage display is now relatively common. However, one unknown factor is how the bacteriophage scaffold itself influences the properties of the displayed protein. This communication evaluates the effect of solution parameters on the catalytic activity of phage displayed Bacillus subtilis Lipase A (BSLA), compared to the free enzyme in solution. While the pH- and temperature-activity profiles of BSLA were not intrinsically affected by phage display, the nanoscale distribution of BSLA within the micellar assay buffer was. This lead to a pronounced increase of activity of phage–BSLA relative to the free enzyme, owing to the accumulation of phage–BSLA at the substrate-rich micelles. Considering this result obtained for BSLA, caution is warranted and similar effects should be considered when selecting other enzymes/proteins by phage display, as the activity of the displayed protein may differ from that of the free protein.     

An expression microarray approach for the identification of metastable epialleles in the mouse genome

Genetic loci displaying environmentally responsive epigenetic marks, termed metastable epialleles, offer a solution to the paradox presented by genetically identical yet phenotypically distinct individuals. The murine viable yellow agouti (A^vy) metastable epiallele exhibits stochastic DNA methylation and histone modifications associated with coat color variation in isogenic individuals. The distribution of A^vy variable expressivity shifts following maternal nutritional and environmental exposures. To characterize additional murine metastable epialleles, we utilized genome-wide expression arrays (N = 10 male individuals, 3 tissues per individual) and identified candidates displaying large variability in gene expression among individuals (V_i = inter-individual variance), concomitant with a low variability in gene expression across tissues from the three germ layers (V_t = inter-tissue variance), two features characteristic of the A^vy metastable epiallele. The CpG island in the promoter of Dnajb1 and two contraoriented ERV class II repeats in Glcci1 were validated to display underlying stochasticity in methylation patterns common to metastable epialleles. Furthermore, liver DNA methylation in mice exposed in utero to 50 mg bisphenol A (BPA)/kg diet (N = 91) or a control diet (N = 79) confirmed environmental lability at validated candidate genes. Significant effects of exposure on mean CpG methylation were observed at the Glcci1 Repeat 1 locus (p < 0.0001). Significant effects of BPA also were observed at the first and fifth CpG sites studied in Glcci1 Repeat 2 (p < 0.0001 and p = 0.004, respectively). BPA did not affect methylation in the promoter of Dnajb1 (p = 0.59). The characterization of metastable epialleles in humans is crucial for the development of novel screening and therapeutic targets for human disease prevention.Key words: epigenetics, metastable epiallele, viable yellow agouti, environmental epigenomics     

Advances in Chemistry and Bioactivity of the Genus Chisocheton Blume

Chisocheton is one of the genera of the family Meliaceae and consists of ca. 53 species; the distribution of most of those are confined to the Indo‐Malay region. Species of broader geographic distribution have undergone extensive phytochemical investigations. Previous phytochemical investigations of this genus resulted in the isolation of mainly limonoids, apotirucallane, tirucallane, and dammarane triterpenes. Reported bioactivities of the isolated compounds include cytotoxic, anti‐inflammatory, antifungal, antimalarial, antimycobacterial, antifeedant, and lipid droplet inhibitory activities. Aside from chemistry and biological activities, this review also deals briefly with botany, distribution, and uses of various species of this genus.     

Aptamer-Assisted Detection of the Altered Expression of Estrogen Receptor Alpha in Human Breast Cancer

An increase in the expression of estrogen receptors (ER) and the expanded population of ER-positive cells are two common phenotypes of breast cancer. Detection of the aberrantly expressed ERα in breast cancer is carried out using ERα-antibodies and radiolabelled ligands to make decisions about cancer treatment and targeted therapy. Capitalizing on the beneficial advantages of aptamer over the conventional antibody or radiolabelled ligand, we have identified a DNA aptamer that selectively binds and facilitates the detection of ERα in human breast cancer tissue sections. The aptamer is identified using the high throughput sequencing assisted SELEX screening. Biophysical characterization confirms the binding and formation of a thermodynamically stable complex between the identified DNA aptamer (ERaptD4) and ERα (Ka = 1.55±0.298×10⁸ M^-1; ΔH = 4.32×10⁴±801.1 cal/mol; ΔS = -108 cal/mol/deg). Interestingly, the specificity measurements suggest that the ERaptD4 internalizes into ERα-positive breast cancer cells in a target-selective manner and localizes specifically in the nuclear region. To harness these characteristics of ERaptD4 for detection of ERα expression in breast cancer samples, we performed the aptamer-assisted histochemical analysis of ERα in tissue samples from breast cancer patients. The results were validated by performing the immunohistochemistry on same samples with an ERα-antibody. We found that the two methods agree strongly in assay output (kappa value = 0.930, p-value <0.05 for strong ERα positive and the ERα negative samples; kappa value = 0.823, p-value <0.05 for the weak/moderate ER+ve samples, n = 20). Further, the aptamer stain the ERα-positive cells in breast tissues without cross-reacting to ERα-deficient fibroblasts, adipocytes, or the inflammatory cells. Our results demonstrate a significant consistency in the aptamer-assisted detection of ERα in strong ERα positive, moderate ERα positive and ERα negative breast cancer tissues. We anticipate that the ERaptD4 aptamer targeting ERα may potentially be used for an efficient grading of ERα expression in cancer tissues.     

alpha-Glucosidase inhibitory pentacyclic triterpenes from the stem bark of Fagara tessmannii (Rutaceae)

In addition to fatty acids, a mixture of sterols (beta-sitosterol, stigmasterol, campesterol and stigmastanol), lupeol, arctigenin methylether, sesamin, vanillic acid (1), 2,6-dimethoxy-1,4-benzoquinone (2), betulinic acid and two pentacyclic triterpene acetates were isolated from Fagara tessmannii Engl. They were identified as 3beta-acetoxy-16beta-hydroxybetulinic acid (3a) and 3beta,16beta-diacetoxybetulinic acid (3b), and their structures were established using 1 and 2D NMR spectra and by comparison with published data. Two derivatives of the compounds were prepared. Some isolated compounds were evaluated for their antifungal and antibacterial activities. Compounds 1 and 3a showed significant inhibition of alpha-glucosidase.     

Microtitre plate-based antibacterial assay incorporating resazurin as an indicator of cell growth, and its application in the in vitro antibacterial screening of phytochemicals

The resazurin assay utilising microtitre-plate, described by Drummond and Waigh in 2000, has been modified to achieve more accuracy in the determination of the minimum inhibitory concentration (MIC) values of natural products, including crude extracts, chromatographic fractions or purified compounds against various bacterial strains. This modified resazurin method is simple, sensitive, rapid, robust and reliable, and could be used successfully to assess antibacterial properties of natural products.     

Length–weight relationships of three carangid fish species Alepes vari (Cuvier, 1833), Uraspis uraspis (Günther, 1860) and Carangiodes oblongus (Cuvier, 1833) from the Bay of Bengal coast,Bangladesh

Length–weight relationships (LWRs) were determined for three Carangid fish, [Alepes vari (Cuvier, 1833), Uraspis uraspis (Günther, 1860) and Carangiodes oblongus (Cuvier, 1833)] inhabiting in the Bay of Bengal coast, Bangladesh. A total of 147 individuals of three species were sampled from four different sampling points of the Bay of Bengal coast, Bangladesh and examined from April 2017 to October 2017. Fishermen operated beach seine nets (mesh size BSN, 30 mm) and drift gillnet (10.5 cm) twice in each sampling campaign to catch the studied species. The calculated value for parameter “b” of LWRs were 3.003, 2.945 and 2.980 for A. vari, U.uraspis and C. oblongus, respectively.