York State Newborn Screening Program Research Articles

Multi-Laboratory Evaluation of Prototype Dried Blood Spot Quality Control Materials for Creatine Kinase-MM Newborn Screening Assays

Pilot studies to detect newborns with Duchenne Muscular Dystrophy (DMD) by newborn bloodspot screening (NBS) have been conducted under the New York State Newborn Screening Program (NYS) and are currently in progress as part of the Early Check Program at Research Triangle Institute (RTI) International. The Newborn Screening Quality Assurance Program (NSQAP) at the U.S. Centers for Disease Control and Prevention (CDC) produced a set of seven prototype dried blood spot (DBS) reference materials spiked with varying levels of creatine kinase MM isoform (CK-MM). These DBS were evaluated over a 3-week period by CDC, NYS, and RTI, all using the same CK-MM isoform-specific fluoroimmunoassay. Results from each laboratory were highly correlated with the relative proportion of CK-MM added to each of the six spiked pools. Based on reference ranges established by NYS and RTI for their pilot studies, these contrived DBS collectively spanned the CK-MM ranges found in typical newborns and the elevated ranges associated with DMD. This set allows quality assessment over the wide range of fluctuating CK-MM levels in typical and DMD-affected newborns.

Utility of Newborn Dried Blood Spots to Ascertain Seroprevalence of SARS-CoV-2 Antibodies Among Individuals Giving Birth in New York State, November 2019 to November 2021

Serosurveys can be used to monitor population-level dynamics of COVID-19 and vaccination. Dried blood spots (DBSs) collected from infants contain maternal IgG antibodies and are useful for serosurveys of individuals recently giving birth. To examine SARS-CoV-2 antibody prevalence in pregnant individuals in New York State, identify associations between SARS-CoV-2 antibody status and maternal and infant characteristics, and detect COVID-19 vaccination among this population. A population-based, repeated cross-sectional study was conducted to detect SARS-CoV-2 nucleocapsid (N) and spike (S) IgG antibodies. Deidentified DBS samples and data submitted to the New York State Newborn Screening Program between November 1, 2019, and November 30, 2021, were analyzed. Prenatal exposure to SARS-CoV-2 antibodies. The presence of IgG antibodies to SARS-CoV-2 N and S antigens was measured using a microsphere immunoassay. Data were analyzed by geographic region and compared with reported COVID-19 cases and vaccinations among reproductive-aged females (15-44 years of age). Data were stratified by infant birth weight, gestational age, maternal age, and multiple birth status. Dried blood spot samples from 415 293 infants (median [IQR] age, 1.04 [1.00-1.20] days; 210 805 [51.1%] male) were analyzed for SARS-CoV-2 antibodies. The first known antibody-positive infant in New York State was born on March 29, 2020. SARS-CoV-2 seroprevalence reflected statewide and regional COVID-19 cases among reproductive-aged females in the prevaccine period. From February through November 2021, S seroprevalence was strongly correlated with cumulative vaccinations in each New York State region and in the state overall (rs = 0.92-1.00, P ≤ .001). S and N seroprevalences were significantly lower in newborns with very low birth weight (720 [14.8%] for S and 138 [2.8%] for N, P < .001) and low birth weight (5160 [19.3%] for S and 1233 [4.6%] for N, P = .009) compared with newborns with normal birth weight (77 116 [20.1%] for S and 19 872 [5.2%] for N). Lower N and higher S seroprevalences were observed in multiple births (odds ratio [OR], 0.84; 95% CI, 0.75-0.94; P = .002 for N and OR, 1.24; 95% CI, 1.18-1.31; P < .001 for S) vs single births and for maternal age older than 30 years (OR, 0.87; 95% CI, 0.80-0.94; P < .001 for N and OR, 1.17; 95% CI, 1.11-1.23; P < .001 for S) vs younger than 20 years. In this study, seroprevalence in newborn DBS samples reflected COVID-19 case fluctuations and vaccinations among reproductive-aged women during the study period. These results demonstrate the utility of using newborn DBS testing to estimate SARS-CoV-2 seroprevalence in pregnant individuals.

Newborn Screening by Sequence and the Road Ahead

Newborn screening (NBS)3 programs are an anomaly in medicine—a universally applied, state-mandated evaluation of healthy individuals. The program had its origin in the presymptomatic diagnosis of phenylketonuria (1), but as technologies for the use of dried blood spots (DBS) have improved, there has been a rapid expansion of the number of disorders evaluated, as well as an increase in the governmental mandates for testing (2). DNA-based diagnosis is already a feature of NBS. The first integrated use of genetic testing was for the confirmation of hemoglobinopathies (3). Genetic testing is now used as a primary or confirmatory technique in nearly every NBS system in the US. In addition to the analysis of sequence, DNA-based testing can also be used to detect the normal maturation of cell-mediated immunity through the detection of T-cell receptor excision circles (TRECs) (4). Clearly, additional tests will be added over time. The expanded use of DNA-based testing in NBS requires a reliable extraction method that delivers genomic DNA of sufficient quality and quantity for downstream applications. In this issue of Clinical Chemistry , Saavedra-Matiz and colleagues report details of the DNA-extraction protocol used in the New York State Newborn Screening Program (5). The source of the DNA sample is DBS on 903 filter papers, which are commonly used in NBS. Therefore, changes in the collection technique (i.e., phlebotomy), the volume of blood collected, or the type of card used for collection would not be required for most screening programs. The key advantages of this protocol are its extremely low cost and simplified use of automation. The minimization of hands-on work reduces labor costs and the chances for the introduction of error. The authors' system is easily scalable to the number of samples that might be reasonably anticipated in a state …

Cost-Effective and Scalable DNA Extraction Method from Dried Blood Spots

Dried blood spot (DBS) samples have been widely used in newborn screening (NBS) for the early identification of disease to facilitate the presymptomatic treatment of congenital diseases in newborns. As molecular genetics knowledge and technology progresses, there is an increased demand on NBS programs for molecular testing and a need to establish reliable, low-cost methods to perform those analyses. Here we report a flexible, cost-efficient, high-throughput DNA extraction method from DBS adaptable to small- and large-scale screening settings. Genomic DNA (g.DNA) was extracted from single 3-mm diameter DBS by the sequential use of red cell lysis, detergent-alkaline, and acid-neutralizing buffers routinely used in whole blood and plant tissue DNA extractions. We performed PCR amplification of several genomic regions using standard PCR conditions and detection methods (agarose gel, melting-curve analysis, TaqMan-based assays). Amplicons were confirmed by BigDye® Terminator cycle sequencing and compared with reference sequences. High-quality g.DNA was extracted from hundreds of DBS, as proven by mutation detection of several human genes on multiple platforms. Manual and automated extraction protocols were validated. Quantification of g.DNA by Oligreen® fluorescent nucleic acid stain demonstrated a normal population distribution closely corresponding with white blood cell counts detected in newborn populations. High-quality, amplifiable g.DNA is extractable from DBSs. Our method is adaptable, reliable, and scalable to low- and high-throughput NBS at low cost ($0.10/sample). This method is routinely used for molecular testing in the New York State NBS program.

Does Galactocerebrosidase Activity Predict Krabbe Phenotype?

This study sought to determine whether galactocerebrosidase activity is predictive of Krabbe onset age, or of survival from onset when controlling for age at onset of signs. We analyzed data on 55 symptomatic patients from the Hunter James Kelly Research Institute's World-Wide Registry. They were tested for galactocerebrosidase activity at Jefferson Medical College (Philadelphia, PA), using survival models in a path model context. Higher galactocerebrosidase activity was predictive of later symptom onset times (P = 0.0011), but did not predict survival after symptom onset (P = 0.9064) when controlling for the logarithm of age at onset. No child with early infantile (aged 0-6 months) phenotype demonstrated galactocerebrosidase activity >0.1 nmol/hour/mg protein. Survival times within a given phenotype did not vary with galactocerebrosidase activity. Although low galactocerebrosidase activity does not predict phenotype, higher activity in the abnormal range (>0.1 nmol/hour/mg protein in this sample) was not identified in the early infantile variant. Galactocerebrosidase activity may be important to consider when predicting phenotype in the newborn screening population. Our findings provide empiric evidence that the upper end (0.15 nmol/hour/mg protein) of the high-risk galactocerebrosidase group in the New York State newborn screening program is conservatively appropriate.

Sickle cell disease incidence among newborns in New York State by maternal race/ethnicity and nativity

Sickle cell disease is estimated to occur in 1:300-400 African-American births, with higher rates among immigrants from Africa and the Caribbean, and is less common among Hispanic births. This study determined sickle cell disease incidence among New York State newborns stratified by maternal race/ethnicity and nativity. Newborns with confirmed sickle cell disease born to New York State residents were identified by the New York State newborn screening program for the years 2000-2008 and matched to birth records to obtain birth and maternal information. Annual incidence rates were computed and bivariate analyses were conducted to examine associations with maternal race/ethnicity and nativity. From 2000 to 2008, 1,911 New York State newborns were diagnosed with sickle cell disease and matched to the birth certificate files. One in every 1,146 live births was diagnosed with sickle cell disease. Newborns of non-Hispanic black mothers accounted for 86% of sickle cell disease cases whereas newborns of Hispanic mothers accounted for 12% of cases. The estimated incidence was 1:230 live births for non-Hispanic black mothers, 1:2,320 births for Hispanic mothers, and 1:41,647 births for non-Hispanic white mothers. Newborns of foreign-born non-Hispanic black mothers had a twofold higher incidence of sickle cell disease than those born to US-born non-Hispanic black mothers (P < 0.001). This study provides the first US estimates of sickle cell disease incidence by maternal nativity. Women born outside the United States account for the majority of children with sickle cell disease born in New York State. Such findings identify at-risk populations and inform outreach activities that promote ongoing, high-quality medical management to affected children.

Developmental and Functional Outcomes in Children with a Positive Newborn Screen for Krabbe Disease: A Pilot Study of a Phone-Based Interview Surveillance Technique

To assess the utility of a telephone-based interview system in providing ongoing monitoring of the developmental and functional status of children with both positive newborn screens for Krabbe disease and low galactocerebrosidase activity on confirmatory testing, and to determine whether this approach provides improved compliance with follow-up compared with formal neuropsychological testing. Infants with low galactocerebrosidase activity (as detected by the New York State newborn screening program) were eligible for this longitudinal prospective cohort study. Consenting families were interviewed by telephone at infant ages of 4, 8, 12, 18, and 24 months. Designated instruments were the Ages and Stages Questionnaires, the Clinical Linguistic and Auditory Milestone Scale, the Gross Motor Quotient, the Warner Initial Developmental Evaluation of Adaptive and Functional Skills 50, and the WeeFIM II 0-3 instrument. Assessments with the Bayley Scales of Infant and Toddler Development, Third Edition (Bayley III) were scheduled at age 12 and 24 months. Seventeen patients were enrolled; 16 were assessed at age 12 and 18 months, and 15 were assessed at age 24 months. Scores were within the normal range on all tests of developmental and functional status, with the exception of expressive language. Only 7 patients completed the Bayley Scales of Infant and Toddler Development, Third Edition assessments; all their scores were in the normal range. This telephone-based technique allows close monitoring of the developmental and functional status of children with a positive newborn screen for this neurometabolic disease, with special attention to detecting plateauing or regression of developmental milestones. Compliance is improved compared with formal neuropsychological testing.

Consented testing of newborns and childbearing women for human immunodeficiency virus through a newborn metabolic screening program

Objective: In this program a postpartum woman could consent to receive her newborn’s human immunodeficiency virus test result from the New York State Newborn Screening Program. Study Design: By state regulation each postpartum woman was counseled and offered her newborn’s human immunodeficiency virus test result. With the mother’s consent, newborn human immunodeficiency virus antibody test results from the Newborn Screening Program were sent to the baby’s pediatrician; otherwise, test results were blinded. Data were analyzed for births from August 1, 1996, to January 31, 1997. Results: Overall, 92.5% of women offered newborn human immunodeficiency virus testing consented to receive the result. Among 444 human immunodeficiency virus–positive women offered newborn testing, consented testing resulted in a 21.4% increase in knowledge of human immunodeficiency virus status from 72.3% (n = 321) at delivery to 93.7% (n = 416) after newborn testing; 6.3% (n = 28) of human immunodeficiency virus–positive women delivered of infants who did not consent apparently remained unaware of their human immunodeficiency virus status. Conclusion: Combined prenatal and consented newborn testing identified 94% of human immunodeficiency virus–positive mothers and exposed newborns, allowing early entry into care. Such testing may provide an opportunity for women not previously tested for the human immunodeficiency virus to learn their status but is not a substitute for universal prenatal human immunodeficiency virus counseling and consented human immunodeficiency virus testing. (Am J Obstet Gynecol 2000;183:245-51.)

Cost Efficacy of A New Filter Paper In New York State Newborn Screening Program (NYSNSP) 1364

Cost Efficacy of A New Filter Paper In New York State Newborn Screening Program (NYSNSP) 1364

Multivariate discrimination for phenylketonuria (PKU) and non-PKU hyperphenylalaninemia after analysis of newborns’ dried blood-spot specimens for six amino acids by ion-exchange chromatography

Ion-exchange HPLC was developed for testing dried blood-spot specimens from newborns. The method is suitable for quantitative confirmatory testing of abnormal specimens detected in the New York State Newborn Screening Program. Positive specimens were initially identified among all New York State newborns with semiquantitative bacterial inhibition assays (BIA) for aminoacidopathies, including phenylketonuria (PKU) and non-PKU hyperphenylalaninemia (HP), maple syrup urine disease, and homocystinuria. A selection of 1346 specimens from routine BIA screening, including 131 newborns with PKU or persistent HP, were tested by HPLC. Of 179 BIA results that were falsely positive, 98 (55%) were also falsely positive by HPLC in which the Phe/Tyr ratio was the discriminator and the threshold was set to attain 100% sensitivity. Investigation of three multivariate discriminatory methods revealed that linear discriminant analysis excluded all but 35 (20%) of the BIA false-positives.

Rapid, efficient method for multiplex amplification from filter paper.

Guthrie cards derived from the New York State Newborn Screening Program were utilized to develop a rapid, economical method for amplifying multiple genes to detect mutations that impact public health. These specimens are untraceable to the donor because identifiers are removed and discarded; therefore, these pilot studies were carried out anonymously. The sample preparation requires minimal manipulation, is amenable to automation, and is useful in laboratories which routinely process large numbers of samples, such as those in typical newborn screening laboratories. Multiple gene fragments may be amplified from a 1 mm punch which contains less than 1 microl of whole blood. The blood spots used in these studies contain sufficient material for up to 25 amplification reactions which multiplex at least four different gene fragments each. Since sufficient material remains on the card after the routine testing is complete, this simple assay can greatly expand the efficacy of current newborn screening programs by permitting DNA diagnosis of some disorders when indicated, particularly those in which genotype-phenotype correlations are useful. In addition to newborn screening specimens, this method is also applicable to whole blood from adults after phlebotomy and from lymphoblastoid cell lines. Use of filter paper for DNA analysis is particularly useful for shipped specimens or for population studies whose subjects are refractory to phlebotomy.

An audit of infants ‘unscreened’ by the neonatal screening system in Scotland during 1991

Introduction: The aim of this study was to assess the number of infants born in Scotland from whom a newborn screening specimen did not arrive at the screening laboratory. Methods: In a systematic 1 in 8 sample, records on 8379 babies registered as born during 1991, were supplied by the Office of the Registrar General (Scotland). These were matched with records from the neonatal screening laboratory database. For all unmatched records, local health board offices were phoned for evidence of a negative result. Results: Specimens from 8324 of these babies were received and tested by the screening laboratory. Of the remaining 55 babies, 34 had died within 24 h of birth, five had returned home (three to England and Wales and two to outside the United Kingdom), four refused for parental objection (three were Jehovah's Witnesses), and 12 were unscreened. Thus, only 0.14% (12 of 8379 of registered infants (95% Confidence Interval 0.22, 0.06)) were unscreened by the Inborn Errors Screening Program in Scotland during 1991. Discussion: From this study, an estimated 96 infants were unscreened for newborn screening (95% Confidence Interval 39, 143) of 67032 infants registered as born in Scotland during 1991. At this rate, a case of congenital hypothyroidism or phenylketonuria would be missed only every 47 and 78 years, respectively, as a result of having been unscreened by the program. Improvement is expected from regular audit now underway. The rate of 0.14% for Scotland is much lower than the rate of 0.82% estimated for unscreened infants in the New York State Newborn Screening Program.

α-Fetoprotein and Hypothyroidism in Infants

To the Editor.— We recently have documented a relationship between serum α-fetoprotein (AFP) and congenital hypothyroidism (CH) in the newborn period. Previous studies in our laboratory and others had reported a relationship between elevated serum AFP and elevated thyroid-stimulating hormone (TSH)/low thyroxine T4 in newborns and infants in the first few months of life.1-4 The New York State Newborn Screening Program for CH routinely screens more than 300 000 specimens per year diagnosing 120 to 140 confirmed cases of CH annually.

Social Work Perspective

The ultimate objective of newborn screening for sickle cell disease should be twofold. The first essential step is the identification of the infants at risk. This has been effectively done in New York state as of 1975 through the New York State Newborn Screening Program. However, identifying these children is not enough. Second is the much more complicated task of providing comprehensive follow-up care for families whose children are affected by the disease, including the much needed psychosocial services. This area continues to be sorely neglected. The increased risk of death due to overwhelming infection in the first 3 years of life for children with sickle cell disease has been noted in the literature. When there is no specialized care, 15% to 20% do not survive. Therefore, it is essential for knowledgeable staff to make contact and begin to develop a trusting relationship as soon as possible with parents of infants born with sickle cell disease. Prophylactic penicillin and pneumococcal vaccination can reduce mortality during the early years. Family involvement with a consistent, available team of health care providers is pivotal in understanding this chronic illness and coping effectively with this extraordinary stress. Our staff is available by telephone for consultations with patients or other medical staff during clinic and emergency room visits and hospitalizations. One element that is clear in our experience at the St Luke's-Roosevelt Hospital Sickle Cell Center in New York City is that adjustment to this chronic illness is a lifelong process. One or two counseling sessions at the time of diagnosis are not sufficient to enable families to fully understand the information given or to realize the impact of having a child with a chronic illness.

Cellulose Acetate/Citrate Agar Electrophoresis of Filter Paper Hemolysates From Heel Stick

In 1965, New York State Public Health Law 2500-a was enacted, mandating the screening of all newborns for phenylketonuria. The law was amended in 1974 to include testing for six other conditions, one of which is homozygous sickle cell disease. Originally, all screening was done in four semiautonomous laboratories located in Erie County (Buffalo), Onondaga County (Syracuse), New York City, and Albany County (Albany). These laboratories were set up to receive, test, and report results of all specimens in their respective regions of the state. Between 1981 and 1985, all screening was centralized at the Wadsworth Laboratories of the New York State Health Department in Albany, where in 1986 the newborn screening laboratory tested more than 288,000 specimens. Hemoglobin is screened with the two-tier procedure of Garrick et al: alkaline electrophoresis on cellulose acetate followed by acid electrophoresis on citrate agar. METHODS/PROCEDURES The average daily volume of the New York State Newborn Screening Program is 1,200 specimens, but the number of specimens can range from 800 to 3,000 per day. Specimens are collected from infant blood obtained by heel stick on Schleicher and Schuell 903 filter paper. Medical personnel are advised to collect the specimen when the infant is three to five days of age and before any planned transfusion is administered. The specimens are air dried and mailed to the laboratory within 24 hours of collection. When received in the laboratory accessioning unit, the specimens are examined to determine their suitability for testing. Specimens are screened for sickle hemoglobin only if there is sufficient blood on the filter paper and the date of collection to date of laboratory receipt is less than 14 day.