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Prenatal hemoglobinopathy screening & prevention in India: A cross-sectional study
For correspondence: Mr Rajat Kumar Agarwal, Department of Clinical Research, Sankalp India Foundation, Bengaluru 560 034, Karnataka, India e-mail: rajat@sankalpindia.net
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Received: ,
Accepted: ,
Abstract
Background & objectives
India carries a significant burden of hemoglobinopathies, with beta-thalassemia and sickle cell disease (SCD) carrier rates ranging from 3-40 per cent by region. Despite the importance of universal screening, widespread prevention is challenging due to socio-cultural stigma, low awareness, and poor follow up. Sankalp’s thalassemia prevention program used targeted prenatal screening rather than mass or cascade screening. The aim of this study was to assess the efficacy and feasibility of a targeted antenatal screening strategy for the prevention and management of severe hemoglobinopathies in India.
Methods
A cross-sectional study was conducted in 36 districts across five Indian States between April 2023 and March 2024. Pregnant women within 20 wk gestation attending public hospitals were screened for hemoglobinopathies using high-performance liquid chromatography (HPLC). If a woman was identified as a carrier, her partner was tested. At-risk couples received molecular confirmation, followed by prenatal testing and counselling to support informed decisions.
Results
Of the total 34,647 enrolled women, 33,270 (96%) were screened, identifying 2,268 (6.8%) carriers, 903 (2.7%) beta thalassemia, 1,176 (3.5%) SCD, and 189 (0.6%) other hemoglobinopathies. Partners of 1,959 women (86.4%) were tested, identifying 339 at-risk couples. Testing was unnecessary in five cases due to benign mutations. Fetal testing was completed in 246 pregnancies, identifying 65 affected foetuses. Among these, 44 families (68%) chose to discontinue pregnancy. One affected birth was prevented for every 787 families enrolled.
Interpretations & Conclusions
The Sankalp program for hemoglobinopathies prevention demonstrates that targeted antenatal screening within public healthcare is a feasible, effective, and scalable strategy for hemoglobinopathy prevention in socio-economically disadvantaged populations.
Keywords
Antenatal screening
hemoglobinopathy
prevention
sickle cell disease
thalassemia
India has an average prevalence of beta thalassemia carriers of 3-4 per cent1,2, while the sickle cell disease (SCD) carrier rate ranges widely from 0 to 40 per cent3, depending on the geographical area. The need for a universal screening programme for thalassemia and SCD is well recognised3-7, but is hampered by substantial socio-cultural barriers1. The management of clinically significant hemoglobinopathies in India has been estimated to require 9.24 million donor blood units, with an 86 per cent increase in cost coverage reaching over 19 per cent of the current National Health Budget by the year 20267. Several prevention strategies have been tried across the world8, each with its own strengths and limitations.
Screening and counselling high school children and following up beta thalassemia carriers after a period of 20 years showed that the majority of screened individuals did not recollect being carriers, and none of the 41 carrier individuals who were married had their partners tested before marriage9. Screening university students in the 17-25 yr age group, who may be more receptive, however, has not been shown to be effective since only a small percentage of them would enter university, and hence a large proportion of the population, particularly in rural areas, would not be covered10. Socio-cultural stigmatisation is a well-documented barrier to premarital screening in India11. In Fact, 99 per cent of prospective carrier couples married even after knowing their high-risk status12. While acknowledging the success of screening pregnant women in antenatal clinics in some private hospitals in India, this may not apply to public hospitals, where in 2015 only 15-20 per cent of low socio-economic status pregnancies were registered10.The National Family Health Survey (NFHS-5), 2019-21 for India reported that mothers who had an antenatal check-up in the first trimester were 70 per cent, up from 59 per cent in 2015-1613. Many important questions related to antenatal screening acceptance, fathers’ testing compliance, at-risk foetus testing, and pregnancy interruption rates remain to be addressed.
This study was undertaken to evaluate the efficacy and feasibility of a targeted antenatal screening approach, integrated into routine public healthcare services, for the prevention of thalassemia, sickle cell disease (SCD), and other clinically significant hemoglobinopathies by implementing a targeted screening approach, rather than mass or cascade screening from affected cases or known carriers. Focused on preventing births affected by thalassemia, SCD, and other haemoglobin variants, it primarily targeted families from lower socio-economic backgrounds seeking antenatal care at public hospitals and clinics across 36 districts in five States. This approach reduces stigma, as hemoglobinopathy screening is part of routine antenatal checkups, making parents more receptive to counselling.
Materials & Methods
This cross-sectional study was undertaken by Sankalp Thalassemia Prevention Programme, Sankalp India Foundation, Bengaluru, Inda. This study was conducted in accordance with the ethical standards of the Institutional Research Committee and the 1964 Declaration of Helsinki.
Core features of the prevention approach
High antenatal workload hospitals – the sites for the prevention clinics
Sankalp India Foundation established dedicated 87 clinics in high antenatal workload hospitals, securing MOUs to define roles and responsibilities. Each clinic had a full-time coordinator as the main point of contact. New centres were introduced with support from hospital authorities, obstetrics, and local counsellors. Coordinators underwent a two-day experiential training to understand management of hemoglobinopathies for effective counselling. The programme expanded from five centres in three States to 36 districts across five States, including Andhra Pradesh1, Karnataka3, Gujarat5, Maharashtra12, and Madhya Pradesh14, with varied setups at district and sub-district levels.
Steps involved in antenatal screening
Women attending routine antenatal check-ups were referred for hemoglobinopathy screening after thorough counselling. Initially, only those between 8-14 wk were referred, but this was expanded to include women up to 20 wk. After registration in the StopThal system (integrated software) and obtaining consent, 2 ml of venous blood was collected, barcoded, and stored at 2-6°C until shipped to the central laboratory. Logistics service providers ensured timely transport, with some locations using a hub-and-spoke model.
If the mother was a carrier, the father was also tested. If both parents were carriers, an ultrasound was requested to determine gestational age, and molecular testing was performed on both samples. Foetal testing was offered only for pregnancies at risk of severe hemoglobinopathies. If the family agreed, foetal testing appointments were scheduled, and transport support was provided. Foetal samples were then sent to central laboratories for analysis.
Results were shared with the family, and counselling was provided to support informed decision-making, including options for termination if the foetus was affected. Throughout the programme, detailed records were maintained for follow ups, and a priority-driven dashboard was used to monitor progress, including missed follow ups, risk alerts, and delays. Families with a known history of hemoglobinopathies were also referred for foetal screening.
Fetal sampling
The organisation established a network of foetal medicine clinics for localised sample collection and genetic counselling, while centralising testing through a set of empanelled laboratories.
Paperless knowledge management – StopThal
All programme data was stored on the cloud-based StopThal platform, accessible to coordinators via password-protected accounts. It was linked to the central laboratory, enabling electronic exchange of clinical information and reports. The platform generated unique family codes, maintainedclinical records, consent copies, and automatic status updates throughout the screening process.
Quality management
Meetings, policies, incidents, and trainings were tracked on the quality management platform. Electronic systems ensured immediate data integrity checks, flagged missing critical data, and provided summaries to primary users and supervisors, ensuring high data quality.
Screening for hemoglobinopathies
The laboratory used a high-performance liquid chromatography (HPLC) upfront approach for hemoglobinopathies screening. This approach is highly sensitive, time- and cost-effective14. Samples were tested using HPLC on Bio-Rad Variant-II and D-10 equipment (BioRad, Switzerland). For inconclusive results, red cell indices were assessed using Sysmex XN-350 (Sysmex, Japan), and gene mutations were tested when necessary. If HPLC results were ambiguous, capillary electrophoresis (CE) on Sebia platforms was used for retesting.
Mutation studies were performed for couples at-risk based on HPLC findings. A high-throughput laboratory employed quantitative real time PCR (qPCR), Sanger sequencing, Gap-PCR (Gap Polymerase Chain Reaction), MLPA (Multiplex Ligation-dependent Probe Amplification), and NGS (Next-Generation Sequencing) in a stepwise, cost-effective approach to identify mutations quickly.
Centralised NABL-accredited labs ensured high-quality testing, using gold-standard methods, advanced technology, and stringent quality control. The lab provided advisory services, consolidating reports and offering next-step guidance, with categorical advice sent to the StopThal system.
Resourcing
The programme provided free antenatal screening to patients. Host hospitals supplied storage refrigerators, staff for sample collection, and coordinator space at no cost. Alternately the programme provided these resources where needed. Hospitals incurred no direct expenses, with sample collection tubes, syringes, and transport boxes provided by the programme. Foetal sample collection expenses were reimbursed, and programme resources were funded through fundraising efforts.
Programme management
Agile methodology15 was extensively used to ensure highly responsive and iterative planning, execution, and evaluation. Agile allowed the programme to respond quickly and measurably. Pre-scheduled weekly team review and monthly stakeholder reporting and review were organised. Occasionally, surprise visits were performed in centres, especially those whose performance was considered unsatisfactory.
Results
From April 2023 to March 2024, a total of 34,647 women were enrolled, mostly within the first 20 wk of pregnancy, across 36 districts in five States (Madhya Pradesh - 15, Maharashtra - 12, Karnataka - 3, Gujarat - 5, and Andhra Pradesh - 1). Figure 1 shows the distribution of enrolled women by week of pregnancy. Testing could not be completed for 1377 (4%) women. Among these families, 1182 (86%) samples were rejected by the laboratory because of sample deterioration, improper labelling, etc., and the remaining 195 (14%) samples were lost through the process of collection and transport.
- Distribution of women based on pregnancy week at enrolment.
Of the 33,270 (96%) women for whom testing was completed, 903 (2.7%) women were found to be carriers of beta thalassemia, 1,176 (3.5%) were found to be carriers of SCD, while 189 (0.6%) were found to be carriers of other hemoglobinopathies resulting in a total carrier rate of 6.8 per cent. Table I is the full list of all hemoglobinopathies identified.
| Total women screened | Beta thalassemia carrier, n (%) | SCD carrier, n (%) | Other hemoglobinopathies carriers, n (%) | Total carriers, n (%) | |
|---|---|---|---|---|---|
| Overall | 33,270 | 903 (2.7) | 1176 (3.5) | 189 (0.6) | 2,268 (6.6) |
| Andhra Pradesh | 9,651 | 215 (2.2) | 32 (0.3) | 29 (0.3) | 276 (2.9) |
| Gujarat | 929 | 40 (4.3) | 52 (5.6) | 6 (0.6) | 98 (10.5) |
| Karnataka | 5,167 | 135 (2.6) | 12 (0.2) | 31 (0.6) | 178 (3.4) |
| Madhya Pradesh | 4,412 | 96 (2.2) | 355 (8) | 36 (0.8) | 487 (11) |
| Maharashtra | 13,111 | 417 (3.2) | 725 (5.5) | 87 (0.7) | 1,229 (9.4) |
Of the 2,268 women who were found to be carriers of hemoglobinopathies, we were successful in recalling 1,959 (86.4%) Partners for testing. Once the couple was confirmed to be at risk, gene mutation studies were performed. Table II lists the mutations identified.
| Hemoglobinopathy | Mutation | HVGS name | Distribution (%) |
|---|---|---|---|
| Carrier - Sickle Cell Disease | CD 6 GAG>GTG [Glu>Val] | HBB:c.[20A>T;112T>G] | 67.6 |
| Carrier - Beta Thalassemia | IVS I-5 (G>C) | HBB:c.92+5G>C | 18.9 |
| CD 15(TGG>TAG) | Hbb:c.47G>A | 2.7 | |
| CD 41/42 (-CTTT) (CD 41/42 (-TTCT), CD 41/42 (-TCTT)) | HBB:c.126_129delCTTT | 1.7 | |
| IVS I (-1) AGG>ACG (Arg>Thr) (Hb Kairouan) - Hb Monroe | HBB:c.92G>C | 1.2 | |
| CD 8/9 (+G) | HBB:c.27dupG | 0.6 | |
| 619 bp deletion (Asian Indian) | NG_000007.3:g.71609_72227del619 | 0.4 | |
| Poly A (T>C) AATAAA>AACAAA | HBB:c.*110T>C | 0.3 | |
| IVS II-837 (T>G) | HBB:c.316-14T>G | 0.3 | |
| CD 16 GGC>GG- | CD 16 GGC>GG- | 0.3 | |
| IVS I-1 G>A | HBB:c.92+1G>A | 0.2 | |
| CD 30 (G>A) or IVS I (-1) AGG>AAG (Arg>Lys) | HBB:c.92G>A | 0.2 | |
| IVS I-6 (T>C) | HBB:c.92+6T>C | 0.2 | |
| CD 10 GCC>GCA [Ala>Ala] | HBB:c.33C>A | 0.2 | |
| IVS I-1 (G>T) | HBB:c.92+1G>T | 0.1 | |
| Dutch I 12.6kb Beta thal | NG_000007.3:g.68071_80682del12612 | 0.1 | |
| CD 39 CAG>TAG [Gln>STOP] | HBB:c.118C>T | 0.1 | |
| CD 37 (TGG>TAG) | HBB:c.113G>A | 0.1 | |
| -28 (A>G) | HBB:c.-78A>G | 0.1 | |
| CD 121 GAA>TAA (120aa) | HBB:c.364G>T | 0.1 | |
| CAP +1 (A>C) | HBB:c.-50A>C | 0.1 | |
| -101 (C>T) | HBB:c.-151C>T | 0.1 | |
| Carrier - Delta-Beta Thalassemia | Indian inversion-deletion rearrangement(Aγδβ) type | - | 0.2 |
| IndianGγAγ (δβ)°-thal | NC_000011.10:g.5214461_5247124del | 0.6 | |
| Carrier - Hb D-Iran | CD 22 GAA>CAA | HBB:c.67G>C | 0.2 |
| Carrier - Hb D-Punjab | CD 121 GAA>CAA [Glu>Gln] Hb D-Punjab | HBB:c.364G>C | 1.4 |
| Carrier - Hb E | CD 26 GAG>AAG [Glu>Lys] | HBB:c.79G>A | 1.1 |
| Carrier - Hereditary persistence of foetal haemoglobin (HPFH) | Indian HPFH-3 | NG_000007.3:g.71609_72227del619 | 0.2 |
| Carrier – Others | Deletion in the HBB gene cluster | 0.8 | |
| Unknown | Unknown or VUS | 0.2 |
Of the remaining 309 partners for whom the testing could not be done for 95 (31%) as they refused to be tested, 63 (20%) could not be contacted, 62 (20%) families did not have sufficient evidence of adequate counselling or follow up, 47 (15%) women had miscarriage or abortion prior to partner screening, 20 (6.5%) fathers were working in other cities, 14 (14.5%) partners crossed the 24 wk window of pregnancy and 8 (2.6%) had other reasons for not getting tested.
A total of 339 couples were found to be at risk. Of these, 246 (72.6%) could be taken up for foetal testing, 13 (3.8%) had spontaneous abortions, and 5 (1.5%) had hemoglobinopathy combination, which does not justify foetal screening. Foetal screening was organised across a network comprising 11 foetal medicine specialists. Among the remaining 75 who could not proceed to foetal screening, 35 (47%) families accepted the risk, 18 (24%) families missed the window within which termination could be done, 11 (14.6%) families could no longer be contacted and 8 (10.6%) refused screening based on family doctor’s advice and 3 (4%) did not seem to have received sufficient counselling and guidance.
A total of 246 foetal screenswere completed, among which 65 foetuses were found to be affected by severe hemoglobinopathies. Counselling was offered to each family, eventually resulting in 44 families making a decision to discontinue pregnancy, while 21 families accepted an affected foetus. Thus, among the families that concluded the process, 68 per cent of couples with an affected foetus made a choice to end the pregnancy.
Twenty foetuses were affected by thalassemia, 33 by SCD, seven by SCD-Thal, and four were other hemoglobinopathies, together with SCD, and one was other hemoglobinopathies with thalassemia.
A total of 94 couples were enrolled who already had a previous child with hemoglobinopathies. Within these families, a total of 8 (9%) did not opt for foetal testing (5 accepted the risk and 3 could not be contacted), and 3 (3.5%) had spontaneous abortion. Among the five families who accepted the risk, two couples were carriers of both SCD and beta thalassemia, while the other three couples were carriers of SCD only. Among the 83 who opted for foetal screening, 63 (76%) had a foetusthat was not affected, while 20 (24%) had an affected foetus. Within the 20 who had affected foetuses, 1 (5%) accepted the risk and continued pregnancy, while the remaining 19 (95%) opted to discontinue the pregnancy.
Within the 245 families who had no previous births with hemoglobinopathies, 10 (4%) had spontaneous abortions, 67 (27%) did not proceed for foetal screening, while 168 (69%) proceeded to get the foetus tested. Within those who did not proceed for foetal screening, 30 (45%) families accepted the risk, 18 (27%) families missed the window within which termination could be done, 8 (12%) families could no longer be contacted and 8 (12%) refused screening based upon family doctor’s advice and 3 (4.5%) did not seem to have received sufficient counselling and guidance. Among the 168 who proceeded for foetal testing, 118 (72%) had a foetus which was not affected, while 45 (28%) others had an affected foetus. Foetal testing was not justified for the remaining five couples. Within those 45 who had an affected foetus, 25 (56%) opted to discontinue the pregnancy, while the remaining 20 (44%) accepted the risks.
Thus, we were able to prevent one birth with hemoglobinopathies for every 787 families that we enrolled. Within the couples who had no prior birth with hemoglobinopathies, we were able to prevent one birth for every 1,382 families enrolled (Fig. 2).
- Summary of outcomes. Source: Image created using Sankalp Integrated Software, StopThal.
Discussion
The Sankalp Programme for Hemoglobinopathies Prevention successfully prevented births with hemoglobinopathies in India’s diverse socio-economic environment, including 13 out of 36 districts with the poorest performance in health and wellness, and only two districts in the top quartile for performance16. This study demonstrated the effectiveness of a multi-tier screening programme, including foetal sampling and genetic testing, in areas with limited healthcare coverage, highlighting its success despite real-world limitations.
Screening for hemoglobinopathies is typically done in the first trimester, but with reliable diagnostics and shorter turnaround times, we enrolled 96 per cent of families up to 20 wk. Since India allows legal termination up to 24 wk, continuing enrolment into the second trimester enables broader participation. Aadhar was used for tracking, while saw revisits and foetal screenings within the same families. Sample rejection and missed time windows caused a four per cent failure in completing screening, but improvements in diagnostics, Partner testing, and foetal screening are being addressed to strengthen prevention efforts.
Hemoglobinopathies are highly diverse, making phenotypic molecular detection complex and challenging17. Using gold standard HPLC for initial screening and combining technologies like qPCR, Sanger sequencing, Gap PCR, MLPA, and NGS for rapid, reliable detection has been key to achieving comprehensive screening.
The carrier rates we observed were 2.7 per cent for Beta-thalassemia, 3.5 per cent for sickle cell, and 0.6 per cent for other hemoglobinopathies, with a combined rate of 6.8 per cent. A unified screening programme is effective, stigma-free, and feasible. However, only 56 per cent of families with no previous affected birth opted for pregnancy termination, highlighting the need for better awareness and counselling.
Foetal screening was not considered justified for five couples who were both carriers of either haemoglobin E (Hb E)18 or haemoglobin D (Hb D)19 which in the homozygous state are typically associated with milder phenotypes with no deleterious effect, or where molecular confirmation for the hemoglobinopathies could not be secured.
Chawla et al20 have reported that over 50 per cent of individuals from the urban area were not willing to undergo pre-marital screening for β thalassemia. Kulkarni et al21, in a study in Bangalore, reported that only 50 per cent of fathers were tested during antenatal visits, but our experience shows higher community receptivity to screening. This study provides valuable insights into operational challenges faced during antenatal screening.
The success so far is driven by Information technology modern diagnostics, logistics, and programme management. Dashboards help coordinators track prevention stages and gestational age, while regional coordinators can intervene when needed. An effective courier service ensures smooth operation across clinics. Quality management is being enhanced to track coordinator performance. Quality checks revealed that many families struggle to recall testing implications, prompting improvements in counselling strategies. The effectiveness of these changes is still being evaluated.
Cost per family enrolled was around ₹ 550 (USD 6.5) at the time of this study and is expected to drop further. Since one birth was prevented for every 787 families enrolled, the cost incurred for every prevention was ₹432,850/-. This cost was just about the cost of managing thalassemia for two years in 200822 and less than half the cost of hematopoietic stem cell transplantation, even in very cost-effective settings23.
At this stage, there still remain several questions, even from the current dataset since the outcomes for families who accepted the risks remain to be assessed. Further to this work is the reporting on the phenotypes and genotypes in different regions and provide more details on the diagnostic workflow. Reasons for failures and factors like location, coordinator skill level, and other determinants are also being evaluated.
Overall, this cross-sectional experience provides real-world evidence on how to operationalise universal prevention programmes using an antenatal screening approach and prevent the birth of babies with severe hemoglobinopathies. The programme demonstrated a SMART (Specific, Measurable, Achievable, Relevant, and Time-bound) approach to prevention. Our experience shows that antenatal screening is a feasible, acceptable, and effective method for thalassemia control in the Indian context.
Acknowledgment
Authors acknowledge National Health Mission of Maharashtra, Madhya Pradesh and Gujarat for their support. Authors also acknowledge all the participating hospitals/institutions. We would like to thank the families who agreed to participate in the programme. Finally, we would like to thank the donors who contributed towards the programme.
Financial support & sponsorship
The study received funding support for screening with the grant from MKH Foundation, Mumbai and Thalassemia and Sickle Cell Society, Hyderabad. The study itself was funded by Sankalp India Foundation.
Conflicts of Interest
None.
Use of Artificial Intelligence (AI)-Assisted Technology for manuscript preparation
The authors confirm that there was no use of AI-assisted technology for assisting in the writing of the manuscript and no images were manipulated using AI.
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