Vascularized dorsal dartos flap to prevent fistula in tubularized incised plate urethroplasty for primary distal and mid shaft hypospadias

The aim of the present study was to evaluate the importance of neourethral covering using vascularized dorsal flap for preventing fistula in Tabularized incised plate (TIP) urethroplasty. The study included 52 children (aged 2-10 years) who had hypospadias repaired including 44  with distal and 8  with mid shaft hypospadias. In all children, a standard tabularized incised plate urethroplasty was followed by reconstruction of new surrounding urethral tissue. A longitudinal dartos flap was harvested from excessive dorsal preputial and penile hypospadiac skin and transposed to ventral side by a buttonhole maneuver. It was sutured to the glans wings and the neomeatus and to the corpora covernosa over the neo-urethra. Thus the new urethra was completely covered with well-vascularzed subcutaneous tissue. At a mean follow-up of 18 months, the result was successful with no fistula or urethral stenosis, except 2 of the mid penile hypospadias. All patients had good functional and cosmetic results with straight penis and vertical slit shaped meatus at the tip of the penis. The 2 patients developed tiny fistula, which were closed spontaneously after meatal dilatation. In conclusion, urethral covering should be part of TIP urethroplasty. A dorsal well-vascularized dartos flap, button holed ventrally is a good choice for preventing fistula for distal and mid shaft hypospadias.

Effi cacy and safety of single-dose liposomal amphotericin B for visceral leishmaniasis in a rural public hospital in Bangladesh: a feasibility study

Background To rapidly reduce the burden of visceral leishmaniasis for national elimination programmes, an acceptable, safe, and eff ective treatment is needed that can be delivered at primary health-care centres. We aimed to assess the tolerability, safety, and cure rate of single-dose liposomal amphotericin B (AmBisome, Gilead, USA) for visceral leishmaniasis treatment in such a setting in Bangladesh. Methods We enrolled patients who had been diagnosed with visceral leishmaniasis at Muktagacha upazila (subdistrict) hospital, Bangladesh. Eligible participants were at least 5 years old and had a history of fever for more than 2 weeks, splenomegaly, rK39 rapid test positivity, and haemoglobin concentrations of at least 50 g/L. Participants were provided a one-off intravenous infusion of liposomal amphotericin B (10 mg/kg bodyweight). Clinical assessments were done during treatment, before hospital discharge, and on days 30 and 180 after treatment. Cure was defi ned as resolution of fever, decrease in spleen size, and an increase in haemoglobin by 10% compared with baseline or to at least 100 g/L. We estimated effi cacy in terms of initial cure (at day 30) and fi nal cure (at 6 months), and safety in all patients who were enrolled (intention-to-treat analysis). We also assessed effi cacy in all patients who completed treatment and 6 month follow-up after treatment with or without visceral leishmaniasis relapse (per protocol analysis). We assessed acceptability in terms of proportion of patients who consented to treatment. This study was registered with the Australian New Zealand Clinical Trial Registry, number CTRN12612000367842. Findings Between March 5, and Aug 14, 2012, 329 (55%) of 594 cases of suspected visceral leishmaniasis were confi rmed. Of these cases, fi ve patients did not consent to treatment and 24 were ineligible for treatment. In the intention-to-treat analysis, 261 (87%) of 300 patients achieved initial cure and 290 (97%) achieved fi nal cure. In the per-protocol analysis, 260 (88%) of 296 patients achieved initial cure and 289 (98%) achieved fi nal cure. One patient did not start treatment owing to an allergic reaction to liposomal amphotericin B. During treatment or within 2 h afterwards, 79 (26%) patients developed fever, 109 (36%) had fever with rigor, and 56 (19%) had hypotension. No patients needed referral to a tertiary hospital for management of adverse events. Interpretation Treatment of visceral leishmaniasis in a primary health-care facility with single-dose liposomal amphotericin B could safely and eff ectively be adopted by the national visceral leishmaniasis elimination programme in Bangladesh

n vitro study of antiamoebic effect of methanol extract of mature seeds of Carica papaya on trophozoites of Entamoeba histolytica

Antiamoebic activity of methanol extract of mature seeds of Carica papaya was tested in vitro on axenic culture of Entamoeba histolytica using metronidazole as a reference amoebicidal agent. The MIC of seed extract was > 62.5 µg/mL as compared to < 0.8 µg/mL for metronidazole. The present study suggests that the mature seeds of C. papaya have antiamoebic effect but less pronounced than metronidazole

Development of Quantitative Rapid Isothermal Amplification Assay for Leishmania donovani

Quantification of pathogen load, although challenging, is of paramount importance for accurate diagnosis and clinical management of a range of infectious diseases in a point-of-need testing (PONT) scenario such as in resource-limited settings. We formulated a quantification approach to test the standard-curve based absolute quantification ability of isothermal recombinase polymerase amplification (RPA) assay. As a test of principle, a 10-fold dilution series of Leishmania donovani (LD) genomic DNA prepared in nuclease-free-water (NFW), and from culture-spiked-blood (CSB) were tested, and a 15 min assay was performed. A modified algorithm was formulated to derive the detection outcome. The threshold-record times (Tr) in seconds thus obtained were plotted against the initial load of parasite genomes for log-linear regression analysis. The quantitative RPA (Q-RPA) assay was further evaluated against a LD quantitative (q)-PCR assay with DNA extracted from visceral and post-Kala-azar dermal leishmaniasis case specimens and stratified into different ranges of threshold cycle (Ct). The best-fitted regression models were found linear with mean r2/root mean square error (RMSE) values of residual points (in seconds) estimated as 0.996/8.063 and 0.992/7.46 for replicated series of NFW and CSB, respectively. In both series, the lower limit of detection reached less than 0.1 parasite genome equivalent DNA. Absolute agreement between Q-RPA and LD-qPCR was found for test positivity, and strong positive correlations were observed between the Tr and Ct values (r = 0.89; p < 0.0001) as well as between the absolute parasite loads (r = 0.87; p < 0.0001) quantified by respective assays. The findings in this very first Q-RPA assay for leishmaniasis are suggestive of its potential in monitoring LD load in clinical specimens, and the development of rapid Q-RPA assays for other infectious diseases

Post-kala-azar Dermal Leishmaniasis with Mucosal Involvement: An Unusual Case Presentation including Successful Treatment with Miltefosine

Post-kala-azar dermal leishmaniasis (PKDL) is a dermatologic manifestation that usually occurs after visceral leishmaniasis (VL) caused by Leishmania donovani . It is characterized by hypopigmented patches, a macular or maculopapular rash and nodular skin lesions on the body surface. Involvement of the mucosae is very rare and unusual in PKDL. We report a case of PKDL that presented with polymorphic skin lesions, along with involvement of peri-oral mucosa and tongue from an endemic area for kala-azar in Bangladesh. In the absence of a definite past history of kala-azar, a clinical suspicion for PKDL was confirmed by positive rapid serological tests against two recombinant (rK39 and rK28) leishmanial antigens, demonstration of Leishmania donovani (LD) body in the slit skin smear, and isolation of promastigotes by culture from a nodular lesion. The patient was treated with oral Miltefosine for three consecutive months and showed significant clinical improvement as demonstrated by a negative slit skin smear at two months after initiation of therapy. We report this case as an unusual presentation of mucosal involvement in PKDL and subsequent treatment success with Miltefosine

Evaluation of Rapid Extraction Methods Coupled with a Recombinase Polymerase Amplification Assay for Point-of-Need Diagnosis of Post-Kala-Azar Dermal Leishmaniasis

To detect Post-kala-azar leishmaniasis (PKDL) cases, several molecular methods with promising diagnostic efficacy have been developed that involve complicated and expensive DNA extraction methods, thus limiting their application in resource-poor settings. As an alternative, we evaluated two rapid DNA extraction methods and determined their impact on the detection of the parasite DNA using our newly developed recombinase polymerase amplification (RPA) assay. Skin samples were collected from suspected PKDL cases following their diagnosis through national guidelines. The extracted DNA from three skin biopsy samples using three different extraction methods was subjected to RPA and qPCR. The qPCR and RPA assays exhibited highest sensitivities when reference DNA extraction method using Qiagen (Q) kit was followed. In contrast, the sensitivity of the RPA assay dropped to 76.7% and 63.3%, respectively, when the boil & spin (B&S) and SpeedXtract (SE) rapid extraction methods were performed. Despite this compromised sensitivity, the B&S-RPA technique yielded an excellent agreement with both Q-qPCR (k = 0.828) and Q-RPA (k = 0.831) techniques. As expected, the reference DNA extraction method was found to be superior in terms of diagnostic efficacy. Finally, to apply the rapid DNA extraction methods in resource-constrained settings, further methodological refinement is warranted to improve DNA yield and purity through rigorous experiments

Evaluation of Loopamp™ Leishmania Detection Kit and Leishmania Antigen ELISA for Post-Elimination Detection and Management of Visceral Leishmaniasis in Bangladesh

With reduced prevalence of visceral leishmaniasis (VL) in the Indian subcontinent (ISC), direct and field deployable diagnostic tests are needed to implement an effective diagnostic and surveillance algorithm for post-elimination VL control. In this regard, here we investigated the diagnostic efficacies of a loop-mediated isothermal amplification (LAMP) assay (Loopamp™ Leishmania Detection Kit, Eiken Chemical CO., Ltd, Japan), a real-time quantitative PCR assay (qPCR) and the Leishmania antigen ELISA (CLIN-TECH, UK) with different sampling techniques and evaluated their prospect to incorporate into post-elimination VL control strategies. Eighty clinically and rK39 rapid diagnostic test confirmed VL cases and 80 endemic healthy controls were enrolled in the study. Peripheral blood and dried blood spots (DBS) were collected from all the participants at the time of diagnosis. DNA was extracted from whole blood (WB) and DBS via silica columns (QIAGEN) and boil & spin (B&S) methods and tested with qPCR and Loopamp. Urine was collected from all participants at the time of diagnosis and was directly subjected to the Leishmania antigen ELISA. 41 patients were followed up and urine samples were collected at day 30 and day 180 after treatment and ELISA was performed. The sensitivities of the Loopamp-WB(B&S) and Loopamp-WB(QIA) were 96.2% (95% CI 89·43-99·22) and 95% (95% CI 87·69-98·62) respectively. The sensitivity of Loopamp- DBS(QIA) was 85% (95% CI 75·26- 92·00). The sensitivities of the qPCR-WB(QIA) and qPCR-DBS(QIA) were 93.8% (95% CI 86·01-97·94) and 72.5% (95% CI 61·38-81·90) respectively. The specificity of all molecular assays was 100%. The sensitivity and specificity of the Leishmania antigen ELISA were 97.5% (95% CI 91·47-99·70) and 91.95% (95% CI 84·12-96·70) respectively. The Leishmania antigen ELISA depicted clinical cure at day 180 in all the followed-up cases. Efficacy and sustainability identify the Loopamp-WB(B&S) and the Leishmania antigen ELISA as promising and minimally invasive VL diagnostic tools to support VL diagnostic and surveillance activities respectively in the post-elimination era