We recently undertook a research project to identify the incidence of AATD in a representative sample of the general population of Ireland. This involved screening 1000 anonymised DNA samples for the presence of the S and Z mutations and was undertaken in collaboration with Dr. Joe McPartlin of the Trinity College Biobank. The gene frequencies revealed for both the S and Z mutation were higher than anticipated based on studies in other European populations.

For the purpose of this study the randomised nationally-based buccal swab collection from the Trinity Biobank was investigated. Ethical approval for the collection was granted by the joint Hospital Ethics Committee of Tallaght and St. James’s Hospitals. The confidentiality of the participant’s data and samples was respected and ensured by irreversible anonymisation. DNA was extracted from buccal swabs according to the Biobank standard protocols. AAT deficient individuals were identified through separate genotyping assays for the S and Z mutations using Real-Time PCR technology and melt curve analysis on a Roche Lightcycler. A Z genotyping assay was performed on 1095 DNA samples provided by the Biobank. This revealed 46 MZ or carrier individuals in the population. The frequency of the Z gene in this population was, therefore, 0.022. An S assay, carried out on 960 biobank samples, revealed 98 MS carriers and 1 SS (homozygote) individual. The frequency of the S gene in this population was, therefore, 0.053. In total, between the two assays, 3 AAT deficient individuals were identified, constituting 2 SZ and 1 SS genotype. A total of 140 AATD carriers were detected, constituting 46 MZ and 98 MS individuals.

Table 1. Number of Deficiency Phenotypes Detected in Population Screened

The percentage of deficiency alleles detected was higher than anticipated from studies in other populations. The allele frequencies for S and Z in Ireland were previously estimated at between 0.02-0.04 and 0.005-0.015 (Luisetti et al, Thorax 2004). The S variant, thought common to the Iberian Peninsula, was detected with unusually high frequency in the Irish population. Our pilot study shows S and Z alleles occur at frequencies of 0.053 and 0.022 respectively in the Irish population. As the random sample was from the 32 counties, and extrapolating from a population of 6 million on the island of Ireland this would suggest there are approximately 2,900 ZZ and 14,000 SZ AAT deficient individuals and over 200,000 MZ carriers on the island of Ireland. Compared to the gene frequencies in our targeted detection programme the Z mutation would appear to be much more clinically significant with a higher penetrance than S in the two populations we have evaluated. The prospect that alpha-1 antitrypsin deficiency is much more common in Ireland than previously thought will help highlight the fact that all COPD, non-responsive asthma and cryptogenic liver disease patients should be tested for AATD. Increased awareness and understanding of AATD is vital to prevent the continuing under-diagnosis of this condition. Early diagnosis of AATD, with appropriate medical follow-up and lifestyle changes, can prevent, or at least postpone, AATD complications.

Adoption of New Sebia Phenotyping Method:

In the last year we have acquired a new piece of equipment for the Targeted Detection Programme. This has allowed us to implement a more accurate method of phenotyping. We can now identify the different alpha-1 antitrypsin phenotypes with an increased sensitivity of detection. The Sebia Hydragel 18 AAT Isofocusing kit is designed for the qualitative detection and identification of the different phenotypes of alpha-1 antitrypsin (AAT) in the electrophoretic patterns of human sera. The procedure includes isoelectricfocusing on agarose gel, performed on the semi-automatic HYDRASYS system, followed by immunofixation with anti-alpha-1 antitrypsin antiserum. The assay is carried out in two stages. Firstly, isoelectrofocusing on agarose gel is used to fractionate the proteins in the serum samples. This is followed by immunofixation with enzyme (peroxidase)-labelled anti-alpha-1 antitrypsin antiserum to identify the various phenotypes of alpha-1 antitrypsin.

To summarise, we have found this new phenotyping method to be highly specific, rapid and simple to perform. It represents a more accurate method of screening for alpha-1 antitrypsin deficiency and improves the identification of not only the most common but also various rare AAT phenotypes.