This document discusses screening, diagnosis, and counseling for hemoglobinopathies. It covers: - The importance of screening to detect conditions early and identify carriers. - Target populations and timing for screening, including newborn, childhood, adolescence, premarital, antenatal. - Techniques for analysis including HPLC, IEF, CE, and DNA/protein methods. - Benefits of newborn screening like preventing complications. - Options for carrier couples like prenatal diagnosis, IVF, adoption. - Counseling challenges for conditions like sickle cell disease in India. - Various state and national programs and initiatives for controlling hemoglobinopathies.

1. Dr Ebin Roshan Paul
Screening, Prenatal diagnosis and
Counselling of Hemoglobinopathies

2. Screening for Hemoglobinopathies
 Why screening important ?
 1. Early detection and intervention of Thal.Major, severe
form of Thal.Intermedia, SCD will help to reduce the
mortality and morbidity of condition, also improves the
QoL.
 2. Detection of carrier status will help to prevent birth of
an affected child in future.

3. Target population, Timing and
strategies
 No single strategy can meet the need of population.
 Population selection and timing of test is important  will
determine options for future.

5. Newborn screening
 DBS spot card used / EDTA - heel prick/cord blood
 4 good quality spots
 Ideally within 24 hours of collection
 With proper labelling  place of birth and MR number
 prior transfusion if any
 family of origin and demographics
 antenatal result if present

6. Techniques for Analysis
Primary Screening on DBS card
 1. HPLC
 2. Isoelectric Focusing (IEF)
 3. Capillary Electrophoresis (CE)
 A second line test needed on the same sample to validate
findings – protein sequence analysis ( mass spectrometry) or
DNA analysis

7.  Screening is not a diagnostic test and not have 100%
sensitivity and specificity.
 HPLC and IEF are the preferred methods for newborn
screening on the basis of cost-benefit analysis.

9. Benefits of NB screening
 Lethal complications can occur in presymptomatic age.
 Early detection and comprehensive care mortality and
morbidity.
 Prophylactic penicillin and PCV vaccination for SCD to
prevent pneumococcal sepsis.

11. Childhood (6months-6years)
 Universal screening for severe anaemia ( <7mg/dl )
 Thal.Major and SCD can be identified, Thal.Intermedia not
picked up
 Cost effective and since preschool children, approached
through Anganwadi.

12. Adolescence
 Prior intense education programme before screening
 Benefits: time to adapt with information, future partner
selection choices, helps to remove stigmatization.
 Those who cannot avoid marriage with carrier  prenatal
diagnostics and other choices.
 Pilot project in Uttrakhand showed high acceptance and
retention of information by adolescence.

13. Premarital population
 Not applicable to all society due to fear and stigmatisation.
 Options provided are either avoiding the marriage or go
with prenatal diagnosis before each pregnancy.
 Religious beliefs and local customs can also influence in
decision.

14. Prenatal Diagnosis
 Carrier couples
 First started by globin chain synthesis analysis in fetal blood
around 1970s by placental aspiration.
 Biochemical and molecular analysis used for PND.
 RDB and ARMS for mutation detection.

15.  Fetal DNA sampling methods  CVS (>11wks)
 Amniocentesis (>16 wks)
 Cordocentesis (18-20wks)
 Pre-requisite are  Pre-test counselling and written
informed consent of couple
 Carrier status of both parents
 Blood group of mother to prevent Rh
incompatibility if present.

16. Choice for “at risk couple”
 Prenatal testing is a choice ( mutation studies of parents and
then analysing the fetus affected or not, then giving
counselling and option of termination of affected fetus)
 When not feasible due to ethical and cultural issues
 not to have children
 adoption
 in vitro fertilisation ( PGD ) and AI

17. Limitations of PND
 No test are absolute ( 2% error )
 Decision of termination of an already established
pregnancy is usually difficult and create emotional stress
 Post test counselling is very important for emotional stress
in future.

19. Antenatal population
 Universal screening should be offered with CBC in first
antenatal visit in all levels.
 MCV(<80), MCH(<27), relative high RBC count, normal RDW
 HbA2 and HbF analysis advised
 If carrier status noted, then husbands evaluated for “at risk
couples” and if detected PND based on Gestational age.

20. Cascade screening
 Siblings and extended family of patients and carriers
 Uttrakhand project found this more effective in screening
for carriers and detect “couple at risk”.
 Some parents unwilling to communicate diagnosis and
misinformed relatives may stay away from affected family
so need proper counselling.

22. Genetic counselling
 An educational process that seeks to assist affected and/or
other at-risk individuals to understand the nature of the
genetic disorder, its transmission and the options available
in management and family planning.
 (Kelly, 1986; Harper, 2004).

23.  Counselling should be in simple/local language, removing
tensions, misconceptions, social stigma.
 Taking a proper family history and pedigree.
 Couples should be reassured of getting healthy baby after
prenatal intervention (at risk couples).
 Information about procedure and its risk associated (pre-test
counselling)

24.  Those who cannot choose termination due to religious and cultural
issues should be given options of PGD, artificial insemination with non
carrier, adoption, not to have children.
 Nature of condition, predicted phenotype, severity, treatment options
should be discussed (post-test counselling).
 Implication of carrier status and recurrence chances in subsequent
pregnancy (25:50:25) should be discussed.

25.  Screening and risk assessment of family/relatives and HLA
matching if planning transplantation.
 In “multi level consanguinity” risk assessment is difficult and
chances of having other recessive or dominant condition to co
exist is high.
 Usual screening for hemoglobinopathies may miss detecting
these conditions. Should be included in counselling.

26. Beta Thalassemia
 Thal.Intermedia  wide range of spectrum ranging from
features of T.minor to T.major
 To assess the predictable phenotype and severity
evaluation of α γ δ genes also necessary.
 Ratio of α to non α chains (β/δ/γ) correlates with the
clinical phenotype severity. (α : non α chain)

27.  α thal in β homozygous condition can have mild clinical
condition than β homozygous alone ( excess α chain ) and
decreases the severity.
 Coexisting HPFH with β thal homozygous can have a milder
phenotype. Free α will go and bind with γ chain.
 “δ/β double heterozygous” can have HbA2 normal and
resemble like α thal heterozygous.
 “α/β double heterozygous” can have normal RBC indices
and elevated HbA2 level.

28. HbE disease (EE)
 Counselling  asymptomatic, no anaemia, no haemolysis,
no SM
 Parental screening is mandatory for heterozygous status
(AE) and beta thalassemia status
 Main differential diagnosis at birth is Hb Eβ0 but it will
symptomatic after the disappearance of HbF.

29. Hb E trait (AE)
 1) Counsel baby will not have any health problems. Repeat
test advised at 6 month to confirm.
 2) In future before marriage partner has to be screened for
hemoglobinopathies.
 3) Parental screening is advised, to check beta thalassemia
status and assess future risk of having a HbE/Beta
thalassemia baby.

30. HbE-Beta Thal / HbE- β+ or β0
 Wide range of phenotype from Thal major (severe)  mild
forms of Thal Intermedia  Thal minor (mild)
 Majority will be moderate severity. Hb 6-7 g/dl. CF similar
to Thal Intermedia. Usually not need transfusion. Iron
overload may occur in future.
 Coinheritance of α thal with HbE-β thalassemia result less
severity and milder phenotype. Reduces the α chain and
reduces the α : non α chain imbalance.

31.  Coinheritance of determinants that increase HbF
expression can also decrease the severity of HbE-β
thalassemia.
 Association of milder forms of β+ thalassemia (with
production of some β chain) are also have milder forms of
HbE-β thalassemia.

32. Hb SE
 Compound heterozygous SE.
 Mild chronic haemolytic anaemia, vaso-occlusive crisis are
rare. Usually presents at second decade of life.
 Few cases reported all over world, 8 cases from south
India.

33. HbS-Beta Thal ( HbS / β+ or β0 )
 compound heterozygous (βs/βthal)
 Only few cases reported tribal population India.
 Tribal areas: Asymptomatic or mild with a few vaso-occlusive
crisis. Most were having β0 mutation with associated α
thalassemia prevalence. (Mukherjee 2010)
 Non tribal areas: noted severe β+ mutation and clinically more
severe than the tribal area. (Mukherjee 2010)

34. SCD – counselling dilemmas
India
 Situation in India is different. SCD is prevalent mainly
scheduled caste and tribe population who are in low SES,
living in illiteracy, poverty and have local religious beliefs.
 Need extensive counselling in local language, visual aids to
create awareness and need for testing.
 Unlike β Thal the clinical severity is much less predictable in
SCD. India has a very variable range of clinical presentation.
Very few follow up data.

35.  Presence of the Arab-Indian haplotype, higher HbF levels
and the presence of associated α-thalassemia mitigate the
severity of the disease in some tribal groups and it is not
always predictable. (Mukherjee 2000,2002)
 Most often cases parents opt termination in homozygous
fetus there are chance that some may have a milder
phenotype and can live up to a reasonable age with
disease. (Colah 2005)

36. Fight against
Hemoglobinopathies

37. Various programmes and
Initiatives
 Jai Vigyan programme : by ICMR covering 6 states
(Maharashtra, Gujarat, Karnataka, West Bengal, Punjab,
Assam). Beta thal carrier frequency 0 - 9.3%. Assam HbE
prevalence 41-66%.
 West Bengal Thalassaemia Control Programme (WBTCP) :
2 nodal centres and 21 centres. Beta Thal prevalence 4 to
10%. Started prenatal analysis.

38.  Sankalp India Foundation: Bangalore based NGO blood
donation, chelation aid, thalassemia relief, Bone marrow
transplant aid.
 Gujarat TCP by Red Cross Society: screening in college and
schools. Started PND centres.
 Maharashtra: 4 district started screening, prevention and
treatment for Thalassemia and SCD.

39.  Punjab : “Project Rainbow” ( Center, State, NGO,
Corporate, patients-parents groups ) for care of
Thalassemia patients.
 “ThalCare” : Web based platform by Sankalp Foundation
for thalassemia management. Used here
 “Thalaman”: software developed to store and assimilate
data online.

40.  “Thalassochip” : diagnostic tool for beta thalassemia based
on aaryed primer extension ( APEX). Help to identify beta
thal mutation and Hb variants on a low operating cost.

41.  Sickle cell anaemia control programme (SCACP)
Gujarat : started in 2006, mainly targeting tribal
population. All antenatal are given screening on
“Mamta Divas” – a special immunisation day, if found
carriers then father is screened and given counselling.
Colour coded cards are given for people based on
status.

42. Road map ahead.....
 Need of education and awareness generation in rural and
urban areas of various states.
 Most of data from hospital and selected population not
reflect actual disease burden. Several states and ethnic
groups, community not screened and “micro mapping”
needed in every state.

43.  Adequate centres, trained staffs and counsellors need.
Providing hands on training at regular intervals.
 Establishment of more day care centres as patient load
increasing.
 Encouragement of bone marrow transplant in low risk
patients and affordable families. Establishment in
government set up.

44.  Joint and sincere effort from Center, State, Various NGO’s ,
Corporate houses, patient support groups backed by
political support for the implementation of the National
Control Programme.