Allele-Specific Recombinase Polymerase Amplification to Detect Sickle Cell Disease in Low-Resource Settings

Abstract To address the global burden of sickle cell disease and the need for novel therapies, the American Society of Hematology partnered with the US Food and Drug Administration to engage the work of 7 panels of clinicians, investigators, and patients to develop consensus recommendations for clinical trial end points. The panels conducted their work through literature reviews, assessment of available evidence, and expert judgment focusing on end points related to patient-reported outcome, pain (non–patient-reported outcomes), the brain, end-organ considerations, biomarkers, measurement of cure, and low-resource settings. This article presents the findings and recommendations of the end-organ considerations, measurement of cure, and low-resource settings panels as well as relevant findings and recommendations from the biomarkers panel.

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Prevalence of sickle cell disease among children of tribal population in India: Feasibility of screening at community level in low‐resource settings

Pediatric Blood & Cancer ◽

10.1002/pbc.28911 ◽

2021 ◽

Author(s):

Bontha V. Babu ◽

Parikipandla Sridevi ◽

Shaily Surti ◽

Mano R. Ranjit ◽

Deepa Bhat ◽

...

Keyword(s):

Sickle Cell Disease ◽

Sickle Cell ◽

Community Level ◽

Tribal Population ◽

Cell Disease ◽

Low Resource Settings ◽

Low Resource

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Point-of-care screening for sickle cell disease in low-resource settings: A multi-center evaluation of HemoTypeSC, a novel rapid test

American Journal of Hematology ◽

10.1002/ajh.25305 ◽

2018 ◽

Vol 94 (1) ◽

pp. 39-45 ◽

Cited By ~ 16

Author(s):

Cindy Steele ◽

Annette Sinski ◽

Jacqueline Asibey ◽

Marie-Dominique Hardy-Dessources ◽

Gisèle Elana ◽

...

Keyword(s):

Sickle Cell Disease ◽

Sickle Cell ◽

Point Of Care ◽

Rapid Test ◽

Cell Disease ◽

Low Resource Settings ◽

Low Resource

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Rapid detection of multidrug-resistant tuberculosis based on allele-specific recombinase polymerase amplification and colorimetric detection

PLoS ONE ◽

10.1371/journal.pone.0253235 ◽

2021 ◽

Vol 16 (6) ◽

pp. e0253235

Author(s):

Nuntita Singpanomchai ◽

Yukihiro Akeda ◽

Kazunori Tomono ◽

Aki Tamaru ◽

Pitak Santanirand ◽

...

Keyword(s):

Multidrug Resistant ◽

Sybr Green ◽

Sybr Green I ◽

Recombinase Polymerase Amplification ◽

Low Resource Settings ◽

Low Resource ◽

Resistant Tuberculosis ◽

Multidrug Resistant Tuberculosis ◽

Mdr Tb ◽

Allele Specific

Multidrug-resistant tuberculosis (MDR-TB) poses a serious threat to TB control. Early diagnosis and proper treatment are essential factors to limit the spread of the disease. The existing molecular tests for MDR-TB usually require specific instruments, steady power supply, and routine maintenance, which might be obstacles for low-resource settings. This study aimed to develop allele-specific isothermal recombinase polymerase amplification (allele-specific RPA) to simultaneously detect the most common mutations in the rpoB gene at codons 516, 526, and 531, which are associated with rifampicin resistance, and in the katG gene at codon 315, which is related to isoniazid resistance. Allele-specific primers targeting four major mutations, rpoB516, rpoB526, rpoB531, and katG315, were constructed and used in individual RPA reactions. The RPA amplicons were endpoints detected by the naked eye immediately after applying SYBR Green I. The optimised RPA assay was evaluated with the Mycobacterium tuberculosis wild-type strain H37Rv and 141 clinical M. tuberculosis isolates. The results revealed that allele-specific RPA combined with SYBR Green I detection (AS-RPA/SYBR) detected these four major mutations with 100% sensitivity and specificity relative to DNA sequencing. The limits of detection for these particular mutations with AS-RPA/SYBR were 5 ng. As a result of the outstanding performance of AS-RPA/SYBR, including its easy setup, speed, lack of a specific instrument requirement, and lack of cross-reaction with other bacteria, this technique may be integrated for the molecular diagnosis of MDR-TB, especially in low-resource settings.

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Klotho Polymorphisms and Priapism In Sickle Cell Disease.

Blood ◽

10.1182/blood.v116.21.1653.1653 ◽

2010 ◽

Vol 116 (21) ◽

pp. 1653-1653 ◽

Cited By ~ 1

Author(s):

Claudia R Lustosa Souza ◽

Marily M Azevedo Shimmoto ◽

Perla Vicari ◽

Martha Mariana A S Arruda ◽

Marina Roizenblatt ◽

...

Keyword(s):

Sickle Cell Disease ◽

Sickle Cell ◽

Vascular Function ◽

Conflicts Of Interest ◽

Significant Proportion ◽

Cell Disease ◽

Specific Pcr ◽

Allele Specific ◽

Male Patients ◽

Allele Specific Pcr

Abstract Abstract 1653 Background: Sickle cell disease (SCD) is a monogenic disorder with phenotypic heterogeneity, possibly determined by polymorphisms (SNPs) in genes whose products modify the pathophysiology of the disease. Priapism is one of the most common vaso-occlusive complications of SCD, and it occurred in more than 30% of males. The Klotho (KL) gene appears to be associated with vascular function and nitric oxide biology and the presence of SNPs could affect its function. Association between KL and priapism in SCD patients was suggested by Nolan et al. in 2004. However, other authors could not confirm this finding (Elliot et al., 2007). Objective: We decided to evaluate the relevance of SNPs rs2249358, rs211234 and rs9536314 to the occurrence of priapism in patients with SCD followed at Outpatient clinic at Escola Paulista de Medicina/UNIFESP. Methods: Forty male patients with SCD were enrolled, 39 (97.5%) with sickle cell anemia (SS) and one (2.5%) SC hemoglobinopathy. The manifestation of priapism was identified through analyses of medical records. The SNP rs2249358 was identified by PCR followed by restriction with XbaI. The other SNPs, rs211234 and rs9536314, were analyzed by allele specific PCR. Statistical analysis: t test, Chi2 or Fisher. This study was approved by Ethical Committee, and all patients agreed in participate. Results: The median age of the patient was 28.5 years-old (20-68 y.o.). Fourteen out of 40 patients had priapism (35%), each one with SS disease. The group of patients with priapism were older (32.5 y.o., 25–68 y.o.) than the group without this manifestation (27.5 y.o., 20–56 y.o.) (p=0.03). There was no statistical difference in the distribution of the SNPs rs211234 and rs9536314 between the two groups of patients (p=0.51 and p= 0.09, respectively). Regarding the distribution of SNP rs2249358, the group with priapism presented 8 individuals (57.1%) with GG genotype, 5 (35.7%) with AA and 1 (7.17%) with AG, whereas in the group without priapism, the distribution was different: 5 (19.2%) with GG, 7 (26.9%) with AA and 14 (53.8%) with AG genotype (p=0.0212). When we compare the presence of at least one A allele (AA or A-) with the G allele in homozygosis (GG), we observed that the A allele has a protector effect (OR: 0.1786; CI: 0.04232–0.7535) (p=0.031). Conclusions: In a relatively small group of patients with SCD, it was observed a significant proportion of individuals with priapism, which reinforces the importance of this manifestation. We also observed correlation between SNP rs2249358 of KL gene and priapism, as suggested previously. Disclosures: No relevant conflicts of interest to declare.

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