Lessons learnt in the screening and diagnosis of haemoglobinopathies.

For this paper, cases reported formally and anecdotally to the authors in their screening and diagnostic roles have been selected to demonstrate areas where errors have occurred, and caution should be exercised. The cases demonstrate that it is vital that the performance and limitations of the techniques used, along with the phenotypic presentation of cases where haemoglobin variants and/or thalassaemias are coinherited are understood by those performing result interpretation. Those who deliver the service as well as those who receive reports and give results and counselling should be aware of the complexity of the topic.

Significant haemoglobinopathies: A guideline for screening and diagnosis: A British Society for Haematology Guideline: A British Society for Haematology Guideline.

Antenatal screening/testing of pregnant women should be carried out according to the guidelines of the National Health Service (NHS) Sickle Cell and Thalassaemia Screening Programme. Newborn screening and, when necessary, follow-up testing and referral, should be carried out according to the guidelines of the NHS Sickle Cell and Thalassaemia Screening Programme. All babies under 1 year of age arriving in the United Kingdom should be offered screening for sickle cell disease (SCD). Preoperative screening for SCD should be carried out in patients from ethnic groups in which there is a significant prevalence of the condition. Emergency screening with a sickle solubility test must always be followed by definitive analysis. Laboratories performing antenatal screening should utilise methods that are capable of detecting significant variants and are capable of quantitating haemoglobins A2 and F at the cut-off points required by the national antenatal screening programme. The laboratory must ensure a provisional report is available for antenatal patients within three working days from sample receipt.

Reliability of the current newborn screening action value for beta thalassaemia disease detection in England: A prospective study.

OBJECTIVE: Beta thalassaemias are a group of hereditary red cell disorders resulting in a reduced or absent production of the main adult haemoglobin, adult haemoglobin. In England, the NHS Sickle Cell & Thalassaemia Screening Programme recommends reporting newborn beta thalassaemia disease as an incidental finding when detected whilst screening for sickle cell disease. The current action value to initiate further investigations is 1.5% adult haemoglobin, using high-performance liquid chromatography or capillary electrophoresis. We examined the reliability of this action value. METHODS: A 44-month country-wide prospective study using data from 13 newborn screening laboratories in England. RESULTS: There were 81 cases reported with an adult haemoglobin of 1.5% or less at first-line screen, of which nine were lost to follow-up. The six false-positive results were all of 32 weeks' gestation or less. Of the 66 true-positives, 36 had confirmatory molecular results (11 of these cases also have results from tandem mass spectrometry), 19 had clinical confirmation and 11 had the results of both parents available which were consistent with the screening result. There was one false-negative, a confirmed beta thalassaemia major case with an adult haemoglobin of 1.7%, above the action value at first-line screen but known to be at risk from parental results and therefore referred into clinical care by the laboratory. CONCLUSIONS: This study demonstrates a positive predictive value of 91.7%, with a specificity of 99.9% and a sensitivity of 98.5%. These results confirm the reliability of the current action value.

Newborn Screening for Sickle Cell Disease in Europe.

The history of newborn screening (NBS) for sickle cell disease (SCD) in Europe goes back almost 40 years. However, most European countries have not established it to date. The European screening map is surprisingly heterogenous. The first countries to introduce sickle cell screening on a national scale were France and England. The French West Indies started to screen their newborns for SCD as early as 1983/84. To this day, all countries of the United Kingdom of Great Britain and Northern Ireland have added SCD as a target disease to their NBS programs. The Netherlands, Spain and Malta also have national programs. Belgium screens regionally in the Brussels and Liège regions, Ireland has been running a pilot for many years that has become quasi-official. However, the Belgian and Irish programs are not publicly funded. Italy and Germany have completed several pilot studies but are still in the preparatory phase of national NBS programs for SCD, although both countries have well-established concepts for metabolic and endocrine disorders. This article will give a brief overview of the situation in Europe and put a focus on the programs of the two pioneers of the continent, England and France.

Newborn screening for sickling and other haemoglobin disorders using tandem mass spectrometry: A pilot study of methodology in laboratories in England.

OBJECTIVE: To determine (i) if electrospray mass spectrometry-mass spectrometry with the SpOtOn Diagnostics Ltd reagent kit for sickle cell screening could be integrated into the English newborn screening programme, under routine screening conditions, and provide mass spectrometry-mass spectrometry results which match existing methods, and (ii) if common action values could be set for all manufacturers in the study, for all assessed haemoglobins, to indicate which samples require further investigation. METHODS: Anonymised residual blood spots were analysed using the SpOtOn reagent kit as per manufacturer's instructions, in parallel with existing techniques at four laboratories. Mass spectrometry-mass spectrometry instrumentation at Laboratories A and B was AB Sciex (Warrington, UK) AP4000, and at Laboratories C and D, Waters Micromass (Manchester, UK), Xevo TQMS and Premier, respectively. RESULTS: There were 23,898 results accepted from the four laboratories. Excellent specificity at 100% sensitivity was observed for haemoglobin S, haemoglobin C, haemoglobin E and haemoglobin OArab. A common action value was not possible for Hb C, but action values were set by manufacturer. The two haemoglobin DPunjab cases at Laboratory D were not detected using the common action value. Conversely, false-positive results with haemoglobin DPunjab were a problem at the remaining three laboratories. CONCLUSIONS: This multicentre study demonstrates that it is possible to implement mass spectrometry-mass spectrometry into an established screening programme while maintaining consistency with existing methods for haemoglobinopathy screening. However, one of the instruments investigated cannot be recommended for use with this application.

Newborn bloodspot results: predictive value of screen positive test for thalassaemia major.

AIM: There are limited published data on the performance of the percentage of haemoglobin A (Hb A) as a screening test for beta thalassaemia major in the newborn period. This paper aims to analyse data derived from a national newborn bloodspot screening programme for sickle cell disease on the performance of haemoglobin A (Hb A) as a screening test for beta thalassaemia major in the newborn period. METHODS: Newborn bloodspot sickle cell screening data from 2,288,008 babies were analysed. Data reported to the NHS Sickle Cell and Thalassaemia Screening Programme in England for the period 2005 to 2012 were also reviewed to identify any missed cases (4,599,849 babies). RESULTS: Within the cohort of 2,288,008 births, 170 babies were identified as screen positive for beta thalassaemia major using a cut-point of 1.5% HbA. There were 51 identified through look-back methods and 119 prospectively identified from 4 screening laboratories. Among 119 babies with prospective data, 7 were lost to follow up and 15 were false positive results. Using a cut-off value of 1.5% Hb A as a percentage of the total haemoglobin as a screening test for beta thalassaemia major in the newborn provides an estimated sensitivity of 99% (from the look back arm of the study) with a positive predictive value of 87% (from the prospective arm of the study). Excluding infants born before 32 weeks gestation, the positive predictive value rose to 95%. CONCLUSION: A haemoglobin A value of less than 1.5% is a reliable screening test for beta thalassaemia major in the newborn period.

Positive screening and carrier results for the England-wide universal newborn sickle cell screening programme by ethnicity and area for 2005-07.

AIMS: The overall aim of the new national newborn programme is to identify infants at risk of sickle cell disease to allow early detection and to minimise deaths and complications. METHODS: Universal screening for sickle cell disease was introduced in England between September 2003 and July 2006. The 13 newborn laboratories each screen between 25,000 and 110,000 babies a year using the existing dried bloodspot cards. The specified conditions to be screened for include sickle cell anaemia (Hb SS), Hb SC disease, Hb S/beta thalassaemia, Hb S/D(Punjab) and Hb S/O(Arab). Data are reported on screening results by ethnic group and geographical area. RESULTS: The prevalence of screen positive results across England is 1:2000. There is a 25-fold variation by geographical area. African babies make up 61% of all screen positive results despite representing only 4% of total births. Combined carrier rates vary widely by ethnicity, from 1.85 per 1000 (1:540) in 'White British' to 145 per 1000 (1:7) in 'African' babies. Refusal rates for screening show variation by ethnicity. CONCLUSIONS: These results provide useful information both about the frequency of these conditions and the carrier state and their geographic and ethnic distribution across England. This can be used to refine counselling information and are also useful to target and plan services and public information.

Implementation of the newborn screening programme for sickle cell disease in England: results for 2003-2005.

OBJECTIVES: This paper reports early screening results from the newborn sickle cell disease screening programme recently implemented in England. SETTING: England. Screening is offered at 5-8 days of age as part of the existing bloodspot test and offered to all babies irrespective of ethnicity. METHODS: The laboratory methods recommended are high performance liquid chromatography (HPLC) and iso-electric focusing (IEF). Two methods of analysis must be applied to all screen positive results. The conditions screened for are:- Sickle cell anaemia (Hb SS), Hb SC disease, Hb S/beta-thalassaemia, Hb S/D(Punjab), Hb S/O(Arab), Hb S/HPFH. Carriers identified for the common haemoglobin variants are reported to parents and follow-up counselling is offered. A bespoke laboratory quality assurance programme has been established which has defined standards of satisfactory performance. RESULTS: Provisional figures from the first seven months of screening (up to March 2004) 108,255 infants were screened gave a screen positive rate of 1:900 for these high prevalence areas and a carrier rate of 2.7%. Figures for 2004-2005 show about 250 significant screen positive results for sickle cell disorders and about 6,500 carriers were identified. The birth prevalence for screen positive results from 2004-05 is 1:1500. We estimate that when there is countrywide data, the national birth prevalence will be about 1:2000-1:2,500. CONCLUSION: The results from the national newborn sickle cell screening programme in England-show that the sickle cell disorders are as common as cystic fibrosis (CF) in England, although the distribution of cases is concentrated in London and other urban areas. The findings and approach to implementation adopted in England may be of interest to other Western European countries with increasing rates of sickle cell disease who are considering such programmes and also to other developed countries.

Hb Niigata [beta1(NA1)Val-->Leu] in a Romanian individual resulting from another nucleotide substitution than that found in the Japanese.

We present a new case of Hb Niigata that we named Hb Niigata(C), observed in a woman from Romania, with a mutation different from that described in Japanese (GTG-->CTG instead of GTG-->TTG). This single nucleotide substitution replaces the valine residue for leucine at codon 1 and causes retention of the N-terminal methionine leading to an elongated beta chain. This mutation was without any hematological consequences.