Low T-cell Receptor Excision Circles Research Articles

Predictors for low T-cell receptor excision circles (TREC) in infants without severe combined immunodeficiency (SCID) or thymic defects

Predictors for low T-cell receptor excision circles (TREC) in infants without severe combined immunodeficiency (SCID) or thymic defects

Abnormal Newborn Screening for Severe Combined Immunodeficiency due to CD4 T cell lymphopenia associated with Heterozygous FOXN1 Variants in the Forkhead Domain

FOXN1 performs a pivotal role in the development of thymic epithelium. Homozygous FOXN1 deficiency results in T-B+NK+ SCID while FOXN1 haploinsufficiency is associated with T cell lymphopenia and nail dystrophy. We report two patients with variants of uncertain significance (VUS) in FOXN1 who presented with lymphopenia detected on newborn screening.Patient A is a 17-month-old female born to non-consanguineous parents who presented with lowT-cell receptor excision circles (TRECs). At 2 weeks old, her lymphocyte subset panel revealed low T cells, but normal B and NK cells (Table 1). Follow up at 3 months revealed uptrending T cells and significantly decreased B cells. She had no concerning clinical features or infections at that time. By 6 months, her T cells were again downtrending, but her B cells had normalized. She required antibiotics for acute otitis media, which quickly resolved. At 15 months, T cell lymphopenia persisted (Figure 2). Her infection history was reassuring, developmental milestones met and physical exam notable only for spooned nails.Patient B is a 4-month-old female also born to non-consanguineous parents presenting with low TRECs. Initial workup at 5 days revealed T cell lymphopenia, but normal B and NK cells (Table 1). At 2 months, CD4 lymphopenia persisted though CD8T cells normalized (Figure 2). She had a self-limiting upper respiratory infection, but otherwise unremarkable infection history. Development and physical exam were normal.A targeted 429-gene panel for inborn errors of immunity revealed heterozygous FOXN1 VUSs in both Patient A (c.890T > G, p.Ile297Ser) and Patient B (c.890T > C, p.Ile297Thr). Patient A's variant was not present in population databases. Patient B's variant has been observed (gnomAD frequency of 0.007%) but not present in population databases for the patient's ethnicities. Surprisingly, both are located at the same position within the Forkhead domain (Figure 1). Algorithms to predict the effect of missense changes on FOXN1 protein structure and function (SIFT, PolyPhen-2, Align-GVGD) all predicted these variants as likely to be disruptive. Further understanding of the phenotypes associated with heterozygous FOXN1 variants of uncertain significance is crucial to guiding diagnostics, treatment, and prognosis. Table 1Lymphocyte panel results at original presentation (Patient A: 2 weeks old, Patient B: 5 days old). Reference ranges obtained from J Allergy Clin Immunol, Volume 112, Number 5, pp 973–980, November 2003VariantCD4+ CD45RA+ (Naïve CD4+ T Cells, cells/μL)CD4+ CD45RO+ (Memory CD4+ T cells, cells/μL)CD8+ T Cells (cells/μL)CD19+ B Cells (cells/μL)CD16+ CD56+ (NK Cells, cells/μL)Patient Ac.890T>G (p.Ile297Ser)633100518423700Patient Bc.890T>C (p.Ile297Thr)569126122610311Ref range (0–3 months)1200–370060–900560–1700300–2000170–1700 [Display omitted] [Display omitted]

Immunologic, Perinatal, and Postnatal Characteristics of Infants with Idiopathic T Cell Lymphopenia

Newborn screening (NBS) measures T cell receptor excision circle (TREC) counts in infants to identify infants with potential primary immunodeficiency. Low TREC counts are associated with T cell lymphopenia (TCL). Infants are considered to have idiopathic TCL if no cause for the lymphopenia is found. By convention, infants whose T cell counts normalize within 12 months of birth are considered to have transient TCL. This study aimed to characterize and differentiate infants with transient and persistent idiopathic TCL across demographic, immunologic, perinatal, and postnatal factors. There were 53 infants referred to an academic medical center for idiopathic TCL between September 2010 and August 2021, following abnormal NBS at a gestational age of at least 37 weeks. They were evaluated for CD3+, CD4+, and CD8+ T cell counts. Demographic, immunologic, perinatal, and postnatal data were collected from the electronic health record. Chi-square analysis was performed for mode of delivery, neonatal intensive care unit (NICU) stay, and hospitalization within 12 months of birth. Descriptive statistics were calculated and Mann-Whitney tests performed for assessing initial TREC counts, initial T cell counts, and birth weight by gestational age percentile. Of the 53 infants evaluated, 28 had transient while 25 had persistent idiopathic TCL. The transient cohort was 60.7% male and 28.6% African-American, and 35.7% had Medicaid. The persistent cohort was 60.0% male and 32.0% African-American, and 56.0% had Medicaid. The cohorts did not differ by mode of delivery (χ2(1,N = 52) = 3.71, p = 0.054), NICU stay (χ2(1,N = 51) = 0.11, p = 0.735), or postnatal hospitalization (χ2(1, N = 49) = 0.82, p = 0.365). Persistent lymphopenic infants had significantly lower median CD3+ (1176 vs. 2144 cells/µL; p < 0.001), CD4+ (866 vs. 1460 cells/µL; p < 0.001), and CD8+ (293 vs. 539 cells/µL; p = 0.003) counts than transient lymphopenic infants, but did not differ by median TREC count (73 vs. 52 TRECs/µL; p = 0.219) or birth weight percentile (41st vs. 20th percentile; p = 0.363). In our cohort, persistent idiopathic TCL infants had lower initial T cell counts than transient infants. None of the other factors differed between the groups. Future research should evaluate for differences across other relevant factors, including maternal health factors and other relevant perinatal and postnatal factors.

The diagnosis of severe combined immunodeficiency (SCID): The Primary Immune Deficiency Treatment Consortium (PIDTC) 2022 Definitions

Severe combined immunodeficiency (SCID) results from defects in the differentiation of hematopoietic stem cells into mature T lymphocytes, with additional lymphoid lineages affected in particular genotypes. In 2014, the Primary Immune Deficiency Treatment Consortium published criteria for diagnosing SCID, which are now revised to incorporate contemporary approaches. Patients with typical SCID must have less than 0.05 × 109 autologous T cells/L on repetitive testing, with either pathogenic variant(s) in a SCID-associated gene, very low/undetectable T-cell receptor excision circles or less than 20% of CD4 T cells expressing naive markers, and/or transplacental maternally engrafted T cells. Patients with less profoundly impaired autologous T-cell differentiation are designated as having leaky/atypical SCID, with 2 or more of these: low T-cell numbers, oligoclonal T cells, low T-cell receptor excision circles, and less than 20% of CD4 T cells expressing naive markers. These patients must also have either pathogenic variant(s) in a SCID-associated gene or reduced T-cell proliferation to certain mitogens. Omenn syndrome requires a generalized erythematous rash, absent transplacentally acquired maternal engraftment, and 2 or more of these: eosinophilia, elevated IgE, lymphadenopathy, hepatosplenomegaly. Thymic stromal defects and other causes of secondary T-cell deficiency are excluded from the definition of SCID. Application of these revised Primary Immune Deficiency Treatment Consortium 2022 Definitions permits precise categorization of patients with T-cell defects but does not imply a preferred treatment strategy.

FOXI3 haploinsufficiency contributes to low T-cell receptor excision circles and T-cell lymphopenia

Newborn screening can identify neonatal T-cell lymphopenia through detection of a low number of copies of T-cell receptor excision circles in dried blood spots collected at birth. After a positive screening result, further diagnostic testing is required to determine whether the subject has severe combined immunodeficiency or other causes of T-cell lymphopenia. Even after thorough evaluation, approximately 15% of children with a positive result of newborn screening for T-cell receptor excision circles remain genetically undiagnosed. Identifying the underlying genetic etiology is necessary to guide subsequent clinical management and family planning. We sought to elucidate the genetic basis of patients with T-cell lymphopenia without an apparent genetic diagnosis. We used clinical genomic testing as well as functional and immunologic assays to identify and elucidate the genetic and mechanistic basis of T-cell lymphopenia. We report 2 unrelated individuals with nonsevere T-cell lymphopenia and abnormal T-cell receptor excision circles who harbor heterozygous loss-of-function variants in forkhead box I3 transcription factor (FOXI3). Our findings support the notion that haploinsufficiency of FOXI3 results in T-cell lymphopenia with variable expressivity and that FOXI3 may be a key modulator of thymus development.

Can we identify WHIM in infancy? Opportunities with the public newborn screening process.

Newborn screening (NBS) for severe combined immunodeficiency (SCID) utilizing T-cell receptor excision circles (TRECs) has been implemented in all 50 states as of December 2018 and has been transformative for the clinical care of SCID patients. Though having high sensitivity for SCID, NBS-SCID has low specificity, therefore is able to detect other causes of lymphopenia in newborns including many inborn errors of immunity (IEIs). In a recent study, three of six newborns later diagnosed with Warts, Hypogammaglobulinemia, Infections, and Myelokathexis (WHIM) syndrome were found to have a low TRECs and lymphopenia at birth. This presents an opportunity to increase the detection and diagnosis of WHIM syndrome by NBS-SCID with immunological follow-up along with a combination of flow cytometry for immune cell subsets, absolute neutrophil count, and genetic testing, extending beyond the conventional bone marrow studies. Coupled with emerging technologies such as next-generation sequencing, transcriptomics and proteomics, dried blood spots used in NBS-SCID will promote earlier detection, diagnosis, and therefore treatment of IEIs such as WHIM syndrome.

Premature Infants Have Normal Maturation of the T Cell Receptor Repertoire at Term.

Premature infants are known to have immature immune systems compared to term infants; however, the impacts of ex utero immune development are not well characterized. Our previous retrospective clinical review showed prolonged T cell lymphopenia in a subset of extremely premature infants, suggesting that they may have lasting abnormalities in their T cell compartments. We used T cell receptor (TCR) repertoire sequencing to analyze the composition of the T cell compartment in premature and term infants in our NICU. We collected twenty-eight samples from individual subjects and analyzed the number of clonotypes, repertoire diversity, CDR3 length, and V gene usage between groups based on gestational age at birth and postmenstrual age at the time of sample collection. Further, we examined the TCR repertoire in infants with severe bronchopulmonary dysplasia (BPD) and those with abnormal T cell receptor excision circle (TREC) assays. Former extremely premature infants who were corrected to term postmenstrual age had TCR repertoire diversity that was more similar to term born infants than extremely premature infants, supporting normal maturation of the repertoire. Infants with severe BPD did not appear to have increased abnormalities in repertoire diversity. Decreased TCR repertoire diversity was associated with repeatedly abnormal TREC screening, although the diversity was within the normal range for subjects without low TRECs. This study suggests that extremely premature infants demonstrate normal maturation of the T cell repertoire ex utero. Further work is needed to better characterize postnatal T cell development and function in this population.

Long-Term Follow-Up of Newborns with 22q11 Deletion Syndrome and Low TRECs

BackgroundPopulation-based neonatal screening using T-cell receptor excision circles (TRECs) identifies infants with profound T lymphopenia, as seen in cases of severe combined immunodeficiency, and in a subgroup of infants with 22q11 deletion syndrome (22q11DS).PurposeTo investigate the long-term prognostic value of low levels of TRECs in newborns with 22q11DS.MethodsSubjects with 22q11DS and low TRECs at birth (22q11Low, N=10), matched subjects with 22q11DS and normal TRECs (22q11Normal, N=10), and matched healthy controls (HC, N=10) were identified. At follow-up (median age 16 years), clinical and immunological characterizations, covering lymphocyte subsets, immunoglobulins, TRECs, T-cell receptor repertoires, and relative telomere length (RTL) measurements were performed.ResultsAt follow-up, the 22q11Low group had lower numbers of naïve T-helper cells, naïve T-regulatory cells, naïve cytotoxic T cells, and persistently lower TRECs compared to healthy controls. Receptor repertoires showed skewed V-gene usage for naïve T-helper cells, whereas for naïve cytotoxic T cells, shorter RTL and a trend towards higher clonality were found. Multivariate discriminant analysis revealed a clear distinction between the three groups and a skewing towards Th17 differentiation of T-helper cells, particularly in the 22q11Low individuals. Perturbations of B-cell subsets were found in both the 22q11Low and 22q11Normal group compared to the HC group, with larger proportions of naïve B cells and lower levels of memory B cells, including switched memory B cells.ConclusionsThis long-term follow-up study shows that 22q11Low individuals have persistent immunologic aberrations and increased risk for immune dysregulation, indicating the necessity of lifelong monitoring.Clinical ImplicationsThis study elucidates the natural history of childhood immune function in newborns with 22q11DS and low TRECs, which may facilitate the development of programs for long-term monitoring and therapeutic choices.

Implementation of Universal Newborn Screening for Severe Combined Immunodeficiency in Singapore While Continuing Routine Bacille-Calmette-Guerin Vaccination Given at Birth.

IntroductionSevere Combined Immunodeficiency (SCID) is generally fatal if untreated; it predisposes to severe infections, including disseminated Bacille-Calmette-Guerin (BCG) disease from BCG vaccination at birth. However, delaying BCG vaccination can be detrimental to the population in tuberculosis-endemic regions. Early diagnosis of SCID through newborn screening followed by pre-emptive treatment with anti-mycobacterial therapy may be an alternative strategy to delaying routine BCG vaccination. We report the results of the first year of newborn SCID screening in Singapore while continuing routine BCG vaccination at birth.MethodNewborn screening using a T-cell receptor excision circle (TREC) assay was performed in dried blood spots received between 10 October 2019 to 9 October 2020 using the Enlite Neonatal TREC kit. Patients with low TREC had lymphocyte subset analysis and full blood count performed to determine the severity of lymphopenia and likelihood of SCID to guide further management.ResultsOf the 35888 newborns screened in 1 year, no SCID cases were detected, while 13 cases of non-SCID T-cell lymphopenia (TCL) were picked up. Using a threshold for normal TREC to be >18 copies/μL, the retest rate was 0.1% and referral rate to immunologist was 0.04%. Initial low TREC correlated with low absolute lymphocyte counts (ALC), and subsequent normal ALC corresponded with increases in TREC, thus patients with normal first CD3+ T cell counts were considered to have transient idiopathic TCL instead of false positive results. 7/13 (54%) had secondary TCL (from sepsis, Trisomy 21 with hydrops and stoma losses or chylothorax, extreme prematurity, or partial DiGeorge Syndrome) and 6/13 (46%) had idiopathic TCL. No cases of SCID were diagnosed clinically in Singapore during this period and for 10 months after, indicating that no cases were missed by the screening program. 8/9 (89%) of term infants with abnormal TREC results received BCG vaccination within the first 6 days of life when TREC and ALC were low. No patients developed BCG complications after a median follow-up of 17 months.ConclusionNewborn screening for SCID can be implemented while continuing routine BCG vaccination at birth. Patients with transient TCL and no underlying primary immunodeficiency are able to tolerate BCG vaccination.

Conditions Other than Severe Combined Immune Deficiency Found on SCID Newborn Screening

Severe combined immune deficiency (SCID) is caused by a wide variety of genetic variants that result in absent or diminished levels of mature and functional T cells. SCID Newborn Screening programs that detect T cell receptor excision circles have resulted in early intervention and improvement in survival rates in infants with SCID but have also identified other conditions with low T cells. These conditions trigger what could be thought of as false positive tests for SCID, but for the most part they do reflect low T cells in conditions other than SCID. The scope of this review is entities other than severe combined immune deficiency that cause low T cell receptor excision circles (TRECs) and are thus identified on newborn screening. Secondary immune deficiencies from prematurity, neonatal medical conditions, hypomorphic or incomplete variants of SCID, transient T cell lymphopenia of infancy, and primary immune deficiencies that do not qualify as SCID are all elaborated. Identification and treatment options for these conditions leading to low TRECs are discussed.

First Year of TREC-Based National SCID Screening in Sweden.

Screening for severe combined immunodeficiency (SCID) was introduced into the Swedish newborn screening program in August 2019 and here we report the results of the first year. T cell receptor excision circles (TRECs), kappa-deleting element excision circles (KRECs), and actin beta (ACTB) levels were quantitated by multiplex qPCR from dried blood spots (DBS) of 115,786 newborns and children up to two years of age, as an approximation of the number of recently formed T and B cells and sample quality, respectively. Based on low TREC levels, 73 children were referred for clinical assessment which led to the diagnosis of T cell lymphopenia in 21 children. Of these, three were diagnosed with SCID. The screening performance for SCID as the outcome was sensitivity 100%, specificity 99.94%, positive predictive value (PPV) 4.11%, and negative predictive value (NPV) 100%. For the outcome T cell lymphopenia, PPV was 28.77%, and specificity was 99.95%. Based on the first year of screening, the incidence of SCID in the Swedish population was estimated to be 1:38,500 newborns.

TREC Screening for WHIM Syndrome.

T cell receptor excision circle (TREC) quantification is a recent addition to newborn screening (NBS) programs and is intended to identify infants with severe combined immunodeficiencies (SCID). However, other primary immunodeficiency diseases (PID) have also been identified as the result of TREC screening. We recently reported a newborn with a low TREC level on day 1 of life who was diagnosed with WHIM (warts, hypogammaglobulinemia, infections, myelokathexis) syndrome, a non-SCID primary immunodeficiency caused by mutations in the chemokine receptor CXCR4. We have now retrospectively reviewed the birth and clinical histories of all known WHIM infants born after the implementation of NBS for SCID. We identified six infants with confirmed WHIM syndrome who also had TREC quantification on NBS.Three of the six WHIM infants had low TREC levels on NBS. All sixpatients were lymphopenic but only one infant had a T cell count below 1,500 cells/μL. The most commonclinical manifestation was viral bronchiolitis requiring hospitalization. One infant died of complications related toTetralogy of Fallot, a known WHIM phenotype. The results suggest that WHIM syndrome should be considered in the differential diagnosis of newborns with low NBS TREC levels. Not applicable.

When Screening for Severe Combined Immunodeficiency (SCID) with T Cell Receptor Excision Circles Is Not SCID: a Case-Based Review.

Newborn screening efforts focusing on the quantification of T cell receptor excision circles (TRECs), as a biomarker for abnormal thymic production of T cells, have allowed for the identification and definitive treatment of severe combined immunodeficiency (SCID) in asymptomatic neonates. With the adoption of TREC quantification in Guthrie cards across the USA and abroad, typical, and atypical SCID constitutes only ~ 10% of cases identified with abnormal TRECs associated with T cell lymphopenia. Several other non-SCID-related conditions may be identified by newborn screening in a term infant. Thus, it is important for physicians to recognize that other factors, such as prematurity, are often associated with low TRECs initially, but often improve with age. This paper focuses on a challenge that immunologists face: the diagnostic evaluation and management of cases in which abnormal TRECs are associated with variants of T cell lymphopenia in the absence of a genetically defined form of typical or atypical SCID. Various syndromes associated with T cell impairment, secondary forms of T cell lymphopenia, and idiopathic T cell lymphopenia are identified using this screening approach. Yet there is no consensus or guidelines to assist in the evaluation and management of these newborns, despite representing 90% of the patients identified, resulting in significant work for the clinical teams until a diagnosis is made. Using a case-based approach, we review pearls relevant to the evaluation of these newborns, as well as the management dilemmas for the families and team related to the resolution of genetic ambiguities.

Dynamic of plasma IL-22 level is an indicator of thymic output after allogeneic hematopoietic cell transplantation

AimsInterleukin-22 (IL-22) promotes thymus recovery and improves T-cell recovery in preclinical allogeneic hematopoietic cell transplant models. However, the correlation between IL-22 and thymus recovery is unknown in human transplant. Materials and methodsIn this study, plasma IL-22 levels of transplanted humans were analyzed peri-transplant. Thymic output was assessed by detecting blood signal joint T-cell receptor excision circles (TRECs). Flow cytometry was applied to measure T-cell subsets. Key findingsPlasma IL-22 level positively correlated with blood TRECs level at days 14 and 28 posttransplant. Multiple linear regression analysis showed plasma IL-22 level, occurrence of acute graft-versus-host disease (aGVHD) and age were significantly associated with blood TRECs level at day 28 after allotransplant. An increase of plasma IL-22 level during day 14 and day 28 correlated with faster recovery of blood TRECs and naïve T-cell levels in allotransplant recipients. Recipients with high TRECs levels at day 28 had lower incidence of aGVHD comparing with those who with low TRECs levels according to a median split of their TRECs levels, an effect also seen in the high IL-22 level and low IL-22 level cohorts. Other factors such as age and infection had impacts on plasma IL-22 level in allotransplants. SignificanceOur findings suggest that dynamic change of plasma IL-22 level is an indicator of thymic output and occurrence of aGVHD. Monitoring plasma IL-22 level might help to assess recovery of thymus function in human allotransplants.

T-Cell Receptor Excision Circles in Newborns With Congenital Heart Disease

BT Davey, RW Elder, MM Cloutier. J Pediatr. 2019;213:96–102.e2 The authors of this study sought to inform their hypothesis that patients with congenital heart disease (CHD) had lower T-cell receptor excision circles (TRECs) than the general population, and low TREC levels in CHD patients are associated with infections leading to hospitalization. Five hundred and seventy-five patients with a diagnosis of structural CHD born between October 2011 and April 2016 were enrolled in this study. They were recruited from the two major Connecticut children’s hospitals, Connecticut Children’s Medical Center and Yale-New Haven Children’s Hospital, which …

TREC and KREC profiling as a representative of thymus and bone marrow output in patients with various inborn errors of immunity.

Primary immune deficiency (PID) disorders are clinically and molecularly heterogeneous diseases. T cell receptor excision circles (TRECs) and κ (kappa)-deleting excision circles (KRECs) are markers of T and B cell development, respectively. They are useful tools to assess T and B cell function and immune reconstitution and have been used for newborn screening for severe combined immunodeficiency disease (SCID) and agammaglobulinemia, respectively. Their profiles in several genetically confirmed PIDs are still lacking. The objective of this study was to determine TREC and KREC genomic profiling among various molecularly confirmed PIDs. We used real-time-quantitative polymerase chain reaction (RT-qPCR)-based triplex analysis of TRECs, KRECs and β-actin (ACTB) in whole blood genomic DNA isolated from 108 patients with molecularly confirmed PIDs. All agammaglobulinemia patients had low KREC counts. All SCIDs and Omenn syndrome patients secondary to mutations in RAG1, RAG2, DCLRE1C and NHEJ1 had low TREC and KREC counts. JAK3-deficient patients had normal KREC and the TREC count was influenced by the type of mutation. Early-onset ADA patients had low TREC and KREC counts. Four patients with zeta-chain-associated protein kinase 70 (ZAP70) had low TREC. All purine nucleoside phosphorylase (PNP) patients had low TREC. Combined immunodeficiency (CID) patients secondary to AK2, PTPRC, CD247, DCLREC1 and STAT1 had normal TREC and KREC counts. Most patients with ataxia-telangiectasia (AT) patients had low TREC and KREC, while most DOCK8-deficient patients had low TRECs only. Two of five patients with Wiskott-Aldrich syndrome (WAS) had low TREC counts as well as one patient each with bare lymphocyte syndrome (BLS) and chronic granulomatous disease. All patients with Griscelli disease, Chediak-Higashi syndrome, hyper-immunoglobulin (Ig)M syndrome and IFNGR2 had normal TREC and KREC counts. These data suggest that, in addition to classical SCID and agammaglobulinemia, TREC/KREC assay may identify ZAP70 patients and secondary target PIDs, including dedicator of cytokinesis 8 (DOCK8) deficiency, AT and some individuals with WAS and BLS.

Establishing Simultaneous T Cell Receptor Excision Circles (TREC) and K-Deleting Recombination Excision Circles (KREC) Quantification Assays and Laboratory Reference Intervals in Healthy Individuals of Different Age Groups in Hong Kong.

The clinical experience gathered throughout the years has raised awareness of primary immunodeficiency diseases (PIDD). T cell receptor excision circles (TREC) and kappa-deleting recombination excision circles (KREC) assays for thymic and bone marrow outputs measurement have been widely implemented in newborn screening (NBS) programs for Severe Combined Immunodeficiency. The potential applications of combined TREC and KREC assay in PIDD diagnosis and immune reconstitution monitoring in non-neonatal patients have been suggested. Given that ethnicity, gender, and age can contribute to variations in immunity, defining the reference intervals of TREC and KREC levels in the local population is crucial for setting up cut-offs for PIDD diagnosis. In this retrospective study, 479 healthy Chinese sibling donors (240 males and 239 females; age range: 1 month−74 years) from Hong Kong were tested for TREC and KREC levels using a simultaneous quantitative real-time PCR assay. Age-specific 5th–95th percentile reference intervals of TREC and KREC levels (expressed in copies per μL blood and copies per 106 cells) were established in both pediatric and adult age groups. Significant inverse correlations between age and both TREC and KREC levels were observed in the pediatric age group. A significant higher KREC level was observed in females than males after 9–12 years of age but not for TREC. Low TREC or KREC levels were detected in patients diagnosed with mild or severe PIDD. This assay with the established local reference intervals would allow accurate diagnosis of PIDD, and potentially monitoring immune reconstitution following haematopoietic stem cell transplantation or highly active anti-retroviral therapy in the future.

T-cell Receptor Excision Circles in Newborns with Heart Defects

In the fetus, the cardiac neural crest gives rise to both the thymus and the conotruncus of the heart. In newborn screening for severe T-cell lymphopenia neonates with congenital heart defects may be detected. In this study, we investigated the occurrence of T-cell lymphopenia in neonates with or without 22q11.2 deletion syndrome (del) suffering from heart defects. This retrospective cohort study included 125 patients with heart defects. T-cell receptor excision circles (TRECs), a measure for T-cell lymphopenia, were quantified by RT-PCR using stored newborn screening blood spots. Three patient groups were compared: non-conotruncal defects (n = 57), conotruncal defects (n = 42), and 22q11.2 del with conotruncal defects (n = 26). Significantly lower TREC values were detected in patients with 22q11.2 del and conotruncal heart defects compared to those with non-syndromic conotruncal (p < 0.001) and non-conotruncal (p < 0.001) defects. In contrast, no significant difference was found between patients with non-syndromic conotruncal and non-conotruncal heart defects (p = 0.152). Low TREC levels were obtained in neonates treated with heart surgery/intervention within 2 weeks after birth and in those with a fatal outcome (p = 0.02) independent of patient group. A correlation was found between low TREC numbers and oxygen saturation, SpO2 below 95% (p = 0.017). The SpO2 was significantly lower in the non-syndromic conotruncal group compared to non-conotruncal (p < 0.001) and 22q11.2 del group (p = 0.015). No correlation was found between low neonatal TRECs and infections needing hospitalization later in life (p = 0.135). Patients with 22q11.2 del and conotruncal defects have significantly lower TREC levels compared to patients with heart defects without this syndrome.

Multiplex PCR-Based Newborn Screening for Severe T and B-Cell Lymphopenia: The first Pilot Study in Turkey.

Objectives:Severe combined immunodeficiency disease (SCID), non-SCID T-cell lymphopenia, and other primary immunodeficiency diseases with T-cell and B-cell lymphopenia have low the T-cell-receptor-excision circles (TRECs) and κ-deleting-recombination-excision circles (KRECs) levels that can be measured in dried blood spots (DBS) of the newborn. The incidence of SCID and non-SCID T-cell lymphopenia in Western societies has been reported by TREC screening of newborns as 1: 58,000 and 1: 7300, respectively. Since there is no similar study in our country, we aimed to perform the first pilot study of TREC and KREC screening of newborn for SCID and non-SCID T-cell lymphopenia in Turkey.Methods:The heel blood samples of newborns born between 1st October 2015 and 31st December 2016 at two major hospitals in our city were included in this study. TREC and KREC copies were determined by a multiplex quantitative PCR-based method from newborn DBS. Cutoff levels were used as 7 copies per DBS for TRECs and KRECs, 1000 copies for ACTB (internal control). Failed samples or abnormal results in measurements were tested the second time. An immunologist evaluated data of newborns with low TREC and KREC copies clinically and through the laboratory.Results:A total of 1960 DBS were tested. The results of 1856 newborns were evaluated. The low TRECs and/or KRECs levels were detected in 71 newborns (3.8 %). The low TRECs rate was 1.1 %. Preterm newborns have lower levels of TRECs and KRECs than term newborns (both p <0.0001). As a result of immunological research, we did not detect any SCID, but we detected 2 newborns with non-SCID T-cell lymphopenia (1:928). These 2 newborns were found to have frequent and severe infectious diseases or hypogammaglobulinemia in their clinical follow-up, although they did not have absolute lymphopenia.Conclusion:Non-SCID T-cell lymphopenia is common in our country than in western societies. TRECs and KRECs assay should be considered for routine NBS programs in our country. Studies involving more newborns should be conducted to detect SCID.

TREC and KREC in very preterm infants: reference values and effects of maternal and neonatal factors

Objective T-cell receptor excision circles (TREC) and kappa-deleting recombination excision circles (KREC) assays have been used for severe combined immunodeficiencies newborn screening (NBS). We assessed TREC and KREC NBS values in preterm infants and investigated if perinatal characteristics affect their values. Methods We performed a retrospective study collecting data from TREC and KREC NBS database and from mothers’ and infants’ medical charts. Results TREC and KREC values were lower in preterm infants born at 23–31 or 32–36 weeks of gestation than in term infants. Gestational age <28 weeks of gestation, leukopenia, and hypertensive disorders of pregnancy lowered TREC. Hypertensive disorders of pregnancy lowered KREC and intrapartum fever >38 °C increased it. Low TREC and KREC values were not associated to the risk of developing early-onset sepsis and late-onset sepsis. Conclusion TREC and KREC levels are lower in preterm than term infants, but this did not increase the risk of neonatal sepsis.