Increased sensitivity for detecting malaria parasites in human umbilical cord blood using scaled-up DNA preparation

Malaria Journal

Methodology

Spencer D Polley¹  Peter L Chiodini²  Colin J Sutherland²  Maha Hassan³  Fiona Regan³ 

[1] Department of Clinical Parasitology, Hospital for Tropical Diseases, Mortimer Market, University College London Hospitals NHS Foundation Trust, WC1E 6JB, London, UK;Department of Clinical Parasitology, Hospital for Tropical Diseases, Mortimer Market, University College London Hospitals NHS Foundation Trust, WC1E 6JB, London, UK;London School of Hygiene and Tropical Medicine, Keppel Street, WC1E 7HT, London, UK;NHS Blood & Transplant, Colindale Avenue, NW9 5BG, London, UK;

关键词: Malaria;    Cord Blood;    Malaria Parasite;    Umbilical Cord Blood;    Parasite Density;   

DOI  :  10.1186/1475-2875-11-62

 received in 2010-12-08, accepted in 2012-03-05,  发布年份 2012

来源: Springer

【 摘 要 】

BackgroundAll mothers donating umbilical cord blood units to the NHS cord blood bank undergo an assessment for the likelihood of prior exposure to malaria infection. Those deemed at risk due to a history of travel to, or residence in, malaria endemic regions are screened serologically to detect anti-malaria antibodies. A positive result excludes the use of the cord blood for transplant therapy unless a risk assessment can ensure that malaria transmission is extremely unlikely. This paper details the screening of cord blood units from malaria serology positive mothers to detect malaria parasite DNA using a highly sensitive nested PCR.MethodsUninfected blood from a healthy volunteer was spiked with known quantities of malaria parasites and 5 millilitre and 200 microlitre aliquots were subjected to DNA extraction using QIAamp DNA maxi and DNA mini kits respectively. Nested PCR, to detect malarial SSU rRNA sequences, was performed on the purified DNA samples to determine the limit of detection for this assay with both extraction methodologies. Following assay validation, 54 cord blood units donated by mothers who were positive for anti-malaria antibodies were screened by this approach.ResultsWhen DNA was purified from 5 millilitres of blood it was possible to routinely detect as few as 50 malaria parasites per millilitre using nested PCR. This equates to a significant increase in the sensitivity of the current gold standard nucleic acid amplification technique used to detect malaria parasites (routinely performed from > 200 microlitre volumes of blood). None of the 54 donated cord blood units from serology positive mothers tested positive for malaria parasites using this scaled up DNA preparation method.ConclusionSerological testing for malaria parasites may be overly conservative, leading to unnecessary rejection of cord blood donations that lack malaria parasites and which are, therefore, safe for use in stem cell therapy.

【 授权许可】

CC BY   
© Polley et al; licensee BioMed Central Ltd. 2012