Childs play - Newborn screening in the UK

The newborn screening programme is one of the largest in the UK – here we learn how the current challenges of screening 700,000 newborns each year can be met by new technologies in the laboratory.

THERE have been major advances in newborn screening in the UK, particularly over the last 4 years with new and expanded programmes for sickle cell disorders, cystic fibrosis and medium chain acyl CoA dehydrogenase deficiency. This article provides an overview of the current programmes in the UK with particular emphasis on the recent advances, emerging new technologies, current challenges and the pivotal role of the laboratory in the whole newborn screening process.

Newborn screening in the UK has a long history. It began in the late 1950s with locally organised screening programmes for phenylketonuria using ferric chloride solutions (subsequently replaced by the Phenistix test strip) to test babies’ urine for phenylpyruvic acid, a characteristic metabolite of the disorder. In 1969, following developments in the USA led by the late Robert Guthrie, the UK Departments of Health recommended changing to a blood based screen, increased concentration of phenylalanine being a more sensitive and specific indicator of phenylketonuria (PKU) than the urinary metabolites. Universal screening for PKU using dried blood spots collected onto a filter-paper card (the Guthrie card) at around 1 week of age was in place throughout the United Kingdom by the early 1970s. It became possible to screen for a further range of disorders with the advent of radio-immunoassay in the 1970s. Assays for both thyroxine and thyroid stimulating hormone (TSH) in dried blood spots were developed and various programmes abroad used these as markers for congenital hypothyroidism (CHT) in single or two tier systems. In 1980 the UK Departments of Health recommended screening for CHT using TSH only.

Since then screening tests for many other disorders have been developed worldwide and the more recent story is the debate around what has become technically possible on the one hand and what is considered desirable and affordable from a public health perspective on the other. This lead to introduction of screening for muscular dystrophy in Wales and numerous other local initiatives(including cystic fibrosis, sickle cell disorders and  some amino acid disorders), some of which were evaluated and subsequently stopped - e.g. galactosaemia in Scotland. The founding of a National Screening Committee in 1996 has provided a more recent focus for the debate and has since enabled a collective and coordinated approach to screening in the UK.

The national newborn screening programme is one of the largest screening programmes in the UK with approximately 700, 000 newborns screened each year. All screening tests are performed from the same blood sample using discs (usually 3mm diameter) punched from dried blood spots collected on to the request card (Fig 1). The programme is hugely successful with local audits demonstrating more than 99% coverage. The laboratory service has gone through a period of consolidation over recent years - in the 1990s there were 26 screening laboratories, there are now 16 (13 in England), most with a workload in the region of 50 - 70,000 babies screened per annum (Fig 2).

Phenylketonuria
Guthrie’s original screening method for phenylketonuria used a bacterial growth-inhibition test. Alternative methods soon came into use, including fluorimetry with continuous flow equipment and chromatography, initially paper chromatography and later commercially produced thin-layer plates, with ninhydrin visualisation. Chromatography allowed detection of other amino acid disorders such as maple syrup urine disease, homocystinuria (severe variants only), and tyrosinaemia - an early example of what would now be termed “multiplex analysis”. More recently, UK laboratories have switched to tandem mass spectrometry (MS-MS) for the measurement of phenylalanine.

Congenital hypothyroidism
The development of non-radioactive immunoassays for newborn screening, particularly using the automated DELFIA system with microtitre plates (Perkin Elmer) for TSH brought greater ease of use. Integrated systems greatly reduce manual handling of the sample, reagents, and the resultant numerical data. Increasing analytical sensitivity of TSH assays has raised interpretational problems, however. The clinical significance of the slightly increased concentrations now detectable is often unclear and there are therefore issues around what cut-off should be used to define a presumptive positive result. The increasing number of premature and very-low birth-weight babies, who may give a false negative result, is also raising difficulties and there is an active debate over the need to retest such babies at a later date.

Cystic fibrosis (CF)
Screening for CF using increased immunoreactive trypsin (IRT) as a marker was first used in the UK in 1980 and several regional schemes were introduced over the next two decades. The discovery of the CFTR gene, mutations of which are responsible for classical infantile cystic fibrosis and a range of milder variants presenting later in life, led to the introduction of CFTR mutation analysis on the same dried blood spot as a second tier in the screening test. This maximises the proportion of cases detected and minimises the requirements for repeat blood samples. A complex protocol combining measurements of IRT with mutation analysis on those spots with the highest (0.5%) IRT concentrations was adopted and CF screening should be in place across the UK by the end of 20071. Measurement of IRT is by immunoassay using the same platform as for the TSH assays. Mutation analysis is a two stage process with four “common” mutations as the first stage followed by a panel of 30 mutations. Introduction of DNA analysis into the protocol has highlighted problems of disease definition and detects a small percentage of unaffected carriers.

Sickle cell disorders (SCD)
Local programmes for SCD newborn screening began in London and Birmingham in the early 1970s. Electrophoresis (citrate agar and cellulose acetate) were the early methods but these have been superseded by HPLC and isoelectric focussing (IEF). The national Sickle Cell and Thalassaemia (SC&T) Programme linking newborn screening for sickle cell disorders with antenatal screening for sickle cell and thalassaemia was set up in 20012 following ministerial commitment in the NHS plan (2000). From 2006 all newborn babies in England are now offered screening - the national policy is for a two tier method using either HPLC or IEF as the initial screen with the other as the confirmatory test. Both of these technologies also detect other haemoglobin variants (of variable clinical significance) and carrier status. Huge numbers of carriers are detected - several fold (around 20 times) greater than the number with disease states. A particular issue for this programme is the impact of transfusion on the test result and therefore all babies who have had a transfusion currently need a repeat test after 4 months. An option to introduce DNA testing as a second tier for transfused babies would obviate this.

Medium-chain acyl-CoA dehydrogenase deficiency (MCADD) and Tandem Mass Spectrometry (MS-MS)
Over the last decade the development of MS-MS with electrospray ionisation has made it possible to screen for a wide range of inherited metabolic disorders (IMD) using dried blood spots3. Experience in large-scale service has largely been limited to measuring amino acids and acylcarnitines, the latter detecting many organic acidaemias and most disorders of fatty acid oxidation. Several UK screening laboratories which were using MS-MS for diagnostic testing for IMD also began to use this technology for PKU screening as it offers improved sensitivity, speed and potential for greater automation. However, the UK has been slow to adopt MS-MS screening for other disorders, which is being pursued much more vigorously in the USA and elsewhere4.

Following three reviews commissioned by the UK Health Technology Assessment Programme and after some years of deliberation a pilot study of MS-MS screening for a single disorder, medium-chain acyl-CoA dehydrogenase deficiency, was launched in 2004. This condition is relatively common in the UK, approximately 1 in 10,000 births and may be detected by an increased concentration of octanoylcarnitine in blood. Affected individuals are at risk of severe metabolic decompensation if subjected to prolonged fasting, often associated with infection, which may result in sudden death or permanent neurological damage. Treatment is simple and effective. The pilot study was undertaken in six laboratories and covered approximately half the babies born in the UK over 2 years5. The results, reviewed in 2006, convinced the National Screening Committee to recommend screening and roll out of MCADD screening across England is planned over the next two years.

The limited use of MS-MS screening by not adding further well-characterised disorders to the panel is an issue of ongoing debate by both laboratory-based screeners and the clinicians they serve.

The UK National Screening Committee (NSC) advises Ministers, the devolved National Assemblies and the Scottish Parliament on screening matters, including the case for implementing new programmes. Compared to many other countries, the UK newborn screening policy is very conservative and before any new programmes can be introduced evidence needs to be carefully considered against agreed principles stipulated by the NSC6

There are two programmes concerning newborn screening the Sickle Cell and Thalassaemia Programme (SC&T) and the Newborn dried blood spot Programme which covers all other disorders. The UK National Screening Programme Centre (UKNSPC) was established in 2002 as a focus for newborn screening with a remit to develop standards, guidelines, provide information and function in a coordinating role. The screening labs function as a network with links to the UKNSPC. They also have their own professional organisation the UKNSLN7 which undertakes some shared initiatives with the Metabolic Biochemistry Network8.

In 2005 policies and standards were published as a basis for data collection and measurement of performance, in particular timeliness of processes and completeness of coverage. Data for 2005/6 showed that the ability to measure standards varies due to inadequate IT systems in the screening laboratories.

Registers are an important part of the screening process; there are some historical registers for PKU and CHT but these require to be updated/further developed - registers are important for assessment of disorder specific detection rates on a national basis, to enable long term follow up of outcome and are a resource for research.
 
The laboratory is at the centre of the screening pathway and plays a wide role with extensive pre and post analytical functions with complex communication routes. The role extends beyond the traditional analytical functions with involvement in sample collection, follow up of presumptive positives and data collection. Good quality blood-spot samples, taken at the correct age and transported rapidly to the laboratory, are prerequisites for an effective service and the laboratory provides feed-back and education to help maintain standards. For most babies who are well and in the community there is no requesting physician to receive and take action or interpret the analytical results - the screening director has the responsibility of arriving at a presumptive diagnosis and arranging appropriate clinical referral for diagnostic assessment and treatment or, in the case of unaffected carriers detected (for CF or SCD) for genetic counselling. Although there are established protocols the off-protocol situations (e.g. sick baby in hospital) require special handling in order to provide  an effective service to the babies and their families.  Some laboratories have screening  nurse-specialists with  roles including liaison  and education to ensure good sample quality, facilitating referrals for presumptive positive cases and contacting families directly in cases where rapid feed-back of results is required.

The number of different organisations and professional groups involved in the screening process is huge and increasing as the programmes have expanded. The central functions of the lab are critical to ensuring governance of the whole process.

Screening crosses many organisational boundaries and effective performance management requires a multi agency/multi-disciplinary approach with structures and clear lines of responsibility. Numerous reorganisations of the NHS has made this difficult and this is an outstanding issue in some parts of the country. It is envisaged that eventually screening systems will link directly into the national NHS IT system, making it much easier to ensure that - declines excepted - every baby has been screened.

Although there are EQA Schemes for newborn screening there are shortfalls which need to be addressed. Similarly, accreditation processes need to be more focussed across the whole screening pathway. 

The newborn screening pathway

Many other western countries (notably the USA) have already introduced screening programmes using MS-MS for a much greater number of disorders than PKU and MCADD. In the USA screening tends to be more laboratory led with primary target disorders and secondary target disorders (based on technology) rather than an individual disorder focussed approach. In the UK the more conservative approach with evaluation of criteria for a specific disorder has undoubtedly contributed to this difference.

There are a number of other stand-alone screening tests that are widely practiced abroad some of which have in the past been the subject of pilot studies in the UK but are currently not part of the NSC recommended  panel. The most notable examples are tests for congenital adrenal hyperplasia (by immunoassay of 17-hydroxyprogesterone), biotinidase deficiency (biotinidase activity) and galactosaemia (enzyme activity or metabolite assay). Some other amino acid disorders (e.g homocystinuria, maple syrup urine disease and tyrosinaemia type 1) could  be incorporated into the MS-MS test panel. International experience shows that most of the organic acidaemias and fatty acid oxidation defects could also be easily incorporated in a single MS-MS screening profile. Addition of some or all of these disorders would emphasise the need, already apparent, for screening laboratories to have close functional links to appropriate specialist diagnostic laboratories and to molecular genetics services.

Preliminary studies9,10 have shown the feasibility of screening for a range of lysosomal storage disorders using newer techniques (MS-MS, microbead multiplex immunoassays) but there are several important issues and fuller evaluation is required.

Immunoassay seems likely to stay, at least in the short term, as the preferred technology for CHT and CF screening. Development in technology for haemoglobins has raised possibilities of both immunoassay and MS-MS as potential methodologies. DNA analysis is also likely to feature in more screening protocols for the future, at least as a second tier or diagnostic stage. Microchip DNA analysis is both feasible and relatively cheap if conducted on a large-enough scale and some screening programmes in the USA are already making use of this technology. However, its adoption in the UK as a primary screening test may be limited by a reluctance to identify subjects with carrier status rather than the disorder itself.

Increased public expectations together with recent improvements in screening test performance by the incorporation of second-tier assays, especially mutation analysis, may lead to more extended screening.

Newborn Screening in the UK has gone through an enormous expansion over the last 3 years and the screening labs are to be congratulated on their huge achievements in responding to the developments, both new programmes and the requests for data from the national programme centre. Approximately 700, 000 babies are now each screened each year for 4 disorders and soon to be five with MCADD.

The challenges ahead are many not least is the need for robust, comprehensive and integrated IT systems for all screening labs with links to the national Child health record systems and the development of effective quality management systems. The new candidates for screening need to be carefully considered and mechanisms for assessment and introduction initiated on a coordinated basis, building on the success of MCADD.
 

References

1. The UK Newborn Screening Programme Centre www.newbornscreening-bloodspot.org.uk

2. Sickle Cell and Thalassaemia Programme  www.sickleandthal.org.uk

3. Wilcken B Recent advances in newborn screening J Inherit Metab Dis 2007; 30:129-133.

4. Pollitt RJ  International Perspectives on Newborn Screening J Inherit Metab Dis 2006; 29: 390-396.

5. Shortland G, Besley G, Bonham J et al Newborn screening for medium-chain acyl-CoA dehydrogenase deficiency (MCADD) : Findings from a multicentre prospective UK Collaborative study ( Abstract) J Inherit Metab Dis 2006 (Suppl 1); 29: 19.

6. National Screening Committee (2003) Second Report of the UK National Screening Committee. (http://www.nsc.nhs.uk); Criteria for appraising the viability, effectiveness and appropriateness of a screening programme. March 2003

7. UK Newborn Screening Laboratory Network  www.newbornsreeening.org

8. Metabolic Biochemistry Network www.MetBio.net

9.Gelb MH, Turecek F, Scott CR, Chamoles NA   Direct multiplex assay of enzymes in dried blood spots by tandem mass spectrometry for the newborn screening of lysosomal storage disorders  J Inherit Metab Dis 2006; 29: 397- 404

10.Wang D, Wood T, Sadilek M, Scott CR, Turecek F and Gelb MH   Tandem Mass Spectrometry  for the direct assay of Enzymes in Dried Blood spots : Application to Newborn Screening for Mucopolysaccharidosis II (Hunter Disease) Clinical Chemistry 2007; 53:137-140 

By Professor Anne Green and Professor Rodney Pollitt. Professor Green was formerly Director of the West Midlands Newborn Screening Service Birmingham Childrens Hospital. She is currently Laboratory Lead for the UK Newborn Screening Programme Centre Great Ormond Street Hospital, London. Professor Pollitt was formerly Director of the Trent Newborn Screening Programme Sheffield Children’s Hospital and Laboratory Advisor for the National CF Programme.