Study on Hemoglobinopathies and G6PD deficiency in Terai Districts of

Nepal
Summary:
Haemoglobinopathies are a group of recessively inherited genetic conditions due to a
hemoglobin disorder, the molecule in red blood cell that delivers oxygen throughout the
body. So far, more than 1,000 mutations have been identified that result in either
hemoglobin result in either a change in the structure and quality of the haemoglobin
(haemoglobin variants) or reduction in the quantity of haemoglobin produced (thalassemia)
Hemoglobin disorders mainly fall into two main categories: sickle-cell disease (SCD) and
thalassemia. Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common
enzymopathy worldwide, affecting estimated 400 million people. The major morbidity
associated with G6PD deficiency is hemolytic anemia, which in some individuals may be
life-threatening.
According to central bureau of Statistics (CBS) 2011, there are around 13.31 million
populations in Terai district of Nepal. There are numerous hospital based and community
based reported the prevalence of Hemoglobinopathies and G6PD among Nepalese
population especially among Tharu residing in western and far western regions. A very
recent study conducted by NHRC, reported prevalence of 11.3% SCD among 20,000 Tharu
population aged 1-29 years in Bardiya district. Similarly, studies have reported the
prevalence of G6PD deficiency was 3.5% -7.2% and is prevalent among different ethnic
groups in Nepal. Despite a good number of research performed in this area, exact number of
Tharu with haemoglobinopathies and G6PD is yet to be determined. Hence, thus this study
aims to estimate prevalence of Hemoglobinopathies (Thalassemia and Sickle Cell disease)
and G6PD and its associated clinical symptoms among 1-29years of population residing in
Terai district of Nepal.
The study will be conducted in 20 districts of Terai region with total of 7200 participants
including 3600 Tharu Population and 3600 other ethnic groups. Estimation of
hemoglobinopathies and G6PD deficiencies will be done through the various lab
investigations.
Introduction
Hemoglobin is oxygen-binding protein which consists of an iron-containing heme ring and
four globin chains: 2 alphas (α) globin chains and 2 beta (β) globin chains. There are different
types of normal hemoglobin in human blood. The percent prevalence of each hemoglobin type
depends on the stage of development. During pregnancy the fetus primarily produces fetal
hemoglobin (HbF) which comprises two alpha and two gamma chains (alpha2 gamma2). At
birth, HbF accounts for approximately 80 percent of hemoglobin and HbA accounts for 20
percent. Hemoglobin A (HbA) is the most common type of adult form hemoglobin which has
two alpha and two beta chains (alpha2 beta2). Hemoglobin A2 (HbA2) is less common adult
form hemoglobin. It comprises two alpha and two delta chains (alpha2 delta2) [1-3].

Haemoglobinopathies are a group of recessively inherited genetic conditions due to a

hemoglobin disorder, the molecule in red blood cell that delivers oxygen throughout the body.
More than 1,000 mutations have been identified that result in either hemoglobin result in
either a change in the structure and quality of the haemoglobin (haemoglobin variants) or
reduction in the quantity of haemoglobin produced (thalassemia) [4, 5].

Hemoglobin disorders mainly fall into two main categories: sickle-cell disease (SCD) and
thalassemia. Alpha thalassemia is a type of thalassemia due to deficient or absent synthesis of
alpha globin chains, leading to more beta globin chains which are controlled by two genes on
each chromosome 16. Deletion of one or more of these genes causes deficient production of
alpha globin chains. In Alpha thalassemia silent carrier status, there is a single gene deletion
which is asymptomatic with normal hematologic findings. In alpha thalassemia trait (minor)
there is two-gene deletion which results in microcytosis but usually no anemia. The three-
gene deletion significantly affect in excess production of hemoglobin H (HbH). Alpha
thalassemia intermedia (HbH) disease, results in hemolysis, microcytic anemia and
splenomegaly.

The Hemoglobin Bart's (Hb Bart's) is result of four-gene deletion which has four gamma
chains (gamma4). Fatal hydrops fetalis is a consequence of Alpha thalassemia major with Hb
Bart's [6].

Beta thalassemia is another type of thalassemia. It is result of lacking or missing of beta

globin chains. This leads to excess alpha chains which are controlled by single gene on each
chromosome. In beta thalassemia trait (minor) there is defect in one gene which is usually
asymptomatic and results in microcytosis and mild anemia. Thalassemia major is also known
as Cooley anemia. In thalassemia major there is synthesis of both genes is severely reduced or
absent. At birth person with the beta thalassemia major usually asymptomatic due to presence
of HbF but symptoms start to develop by age of six months. If the production of beta chains is
less severely reduced, the person has beta thalassemia intermedia. Person with beta
thalassemia intermedia experience less severe symptoms and do not require lifelong
transfusions to survive past 20 years of age [6].

Sickle cell disease is a multisystem disorder that is caused by a single gene mutation.   The
most important haemoglobin variants, is sickle cell, Hb S. Sickle hemoglobin (HbS) is caused
by a mutation in the hemoglobin (HBB) gene causing a single amino acid substitution at
position 6 of the beta globin molecule (B 6 Glu- Val, valine for glutamic acid replacement)
leading to a structural variant of normal adult hemoglobin (HbA), namely sickle hemoglobin
(HbS)[7, 8]. SCD occurs when both parents pass unusual haemoglobin genes to their baby.
SCD genotypes include sickle cell trait (SA), sickle cell anemia/disease(SS), HbSC disease
(SC), sickle beta-thalassemia/ disease (β0/β+), HbSE disease (SE), S with other Hb variants:
D, O-Arab, other SF, Hb S/HPFH. Mutations in both β-globin subunits result in disease based
on a homozygous or heterozygous expression. In the case of sickle cell anemia (HbSS),
mutations are homozygous with production of HbS. This leads to chronic haemolytic disorder
that is marked by tendency of hemoglobin molecules within red cells to polymerize together
and deform the red cell into a sickle (or crescent) shape resulting in characteristic vaso-
occlusive (VOC) events and accelerated haemolysis [8]. HbS is the most common
pathological hemoglobin variant worldwide. Other diseases classed under sickle cell disease
(SCD), for example HbSE, HbSC and HbSβ-thalassemia are heterozygous expression. Despite
of being a monogenic disease, SCD has clinical heterogeneity. Individuals with same
genotype have different clinical aspects. Without treatment, which is rarely available in low-
income, high burden countries, the vast majority of children born with SCA die before the age
of 5 years [9].

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzymopathy

worldwide, affecting estimated 400 million people [10]. The major morbidity associated with
G6PD deficiency is hemolytic anemia, which in some individuals may be life-threatening.
Hemolysis can be elicited by infection, hyperglycemia, certain foods, and certain medications
[11]. G6PD deficiency is an X-linked genetic disorder with 187 known allelic mutations [12].
G6PD, a critical enzyme in the pentose phosphate pathway, exhibits diminished activity in
these patients, leading to inadequate production of protective intracellular thiols during
oxidative stress. While the deficiency is ubiquitous across cell types, erythrocytes are
particularly vulnerable to oxidative stress in the G6PD-deficient state. The World Health
Organization (WHO) has classified G6PD deficiency in category I-V according to the
magnitude of the enzyme deficiency and hemolysis severity [11, 13].

According to WHO approximately 5% of the world’s population carries trait genes for
hemoglobin disorders, mainly, sickle-cell disease and thalassemia [14]. It is estimated that
each year over 300 000 babies with severe forms of these diseases are born worldwide; over
80 per cent of these births occur in low and middle income countries [15]. The percentage of
people who are carriers of the gene is as high as 25% in some regions. These conditions are
most prevalent in tropical regions; however population migration has spread these diseases to
most countries [16]. Thalassaemias are the most common in Asia, the Mediterranean basin,
and the Middle East. The global frequency of the HbS gene is particularly prevalent in
tropical areas, and its highest prevalence occurs in Middle East, Mediterranean regions,
Southeast Asia, and sub-Saharan Africa especially Nigeria [9, 17]. It has been recognized that
sickle cell trait has its highest in areas that are hyper endemic for malaria. It suggests that
carrier state of HbS afforded selective protection against lethal forms of malaria [18-20].
Similarly, Sub-Saharan Africa has the highest average estimated prevalence (7.5%) of (G6PD)
followed by Middle East (6.0%) and Asia (4.7%) [21]. The affected population in malaria-
endemic countries is estimated to be 220 million males and 133 million females [22, 23] .
Rationale of the study:

Haemoglobinopathies are the common monogenic diseases causing one of the

world’s major health problems. Genetic disorders of hemoglobin are more frequently
found in developing countries especially in country of Mediterranean region and, parts of Asia
and Africa but now have spread globally. It is believed that the global spread of these
inheritable diseases might be due to international migration occurring in different
locations of the world [5, 24, 25]. One of the reasons for increasing number of
haemoglobinopathies related diseases is high in number of consanguineous marriage in
some countries and poor public health measures to control this issue [26, 27]. Studies from
various parts of the world reports that sickle cell is prevalent in variety of ethnic
group and tribal population living across the world [27, 28].

According to central bureau of Statistics (CBS) 2011, there are around 13.31 million
populations in Terai district of Nepal [29]. Numerous hospital based and community based
study has reported the prevalence of Hemoglobinopathies and G6PD among Nepalese
population especially among Tharu residing in western and far western regions. A very recent
study conducted by NHRC, reported prevalence of 11.3% SCD among 20,000 Tharu
population aged 1-29 years in Bardiya district [30]. Similarly, studies have reported the
prevalence of G6PD deficiency was 3.5% -7.2% and is prevalent among different ethnic
groups in Nepal [31, 32].

Despite a good number of research performed in this area, precisely exact number of Tharu
with haemoglobinopathies and G6PD is yet to be determined thus making it a matter of
utmost importance for the management of this disorder. Similarly; besides Tharu population,
studies have shown that heamoglobinopatthies and G6PD are also prevalent among other
ethnic groups residing in Terai region. It is known that Terai people share common socio-
economic conditions as well as belongs to malaria endemic area. Hence, it is utmost
importance to determine the exact prevalence of heamoglobin disorder and G6PD among
population of Terai district of Nepal. Thus this study aims to estimate prevalence of
Hemoglobinopathies (Thalassemia and Sickle Cell disease) and G6PD and its associated
clinical symptoms among 1-29years of population residing in Terai district of Nepal.
Objectives

General Objectives

 To determine the prevalence of hemoglobinopathies and G6PD (Thalassemia and

Sickle Cell disease) among population of Terai district of Nepal.

Specific Objectives

 To determine the prevalence of hemoglobinopathies and G6PD in 1-29 years age

group among population of Terai district of Nepal.
 To compare the hemoglobinopathies and G6PD among Tharu and other than Tharu
population.
 To determine the correlation of associated socio demographic factors.
 To find the correlation of the associated clinical symptoms to hemoglobinopathies.

Methodology

Study design:

This study will be a cross-sectional study to determine the prevalence and compare the
hemoglobin disorder among the Tharu and other than Tharu population of Terai districts of
Nepal.

Study population

The study population includes men and women aged 1–29 years who had been living at their
place of residence for at least six months. People with the following characteristics will not be
included:

 Aged less than 1 year.

 Aged more than 29 years
 Too frail to participate in the study.
 History of blood transfusion history in the past three months
 Unable or unwilling to give informed consent
 Unable or unwilling to give informed assent of their children.
Sample Size Calculation:

Sample Size will be calculated by using sample size calculator as recommended by WHO
(sample size calculator STEPS) to generate reliable estimates that it is sufficient for analysis at
national level.

1st Step: Minimum sample size needed (the sampling domain)

Minimum sample size will be calculated by using following formula:

Where:

Z = level of confidence measure and represents the number of standard errors away from the
mean. This describes the uncertainty in the sample mean or prevalence as an estimate of the
population mean (normal deviate if alpha equals 0.05, Z = 1.96, for 95% confidence level).

P = Prevalence of hemoglobinopathy in Nepalese Population, 36.58%= 37% [33].

q = 1-P

d = margin of error

=1.96^2*0.37*0.63/0.05^2= 358.2~358

The calculated sample size n=358, without taking into account the non-response and design
effect.

2nd Step: adjusting for design effect and non-response

In calculation of sample size, to achieve a more robust estimate, the sample size will be
adjusted for non-response (20%) and design effect of 2.

n = 358/0.80 * 2 = 895~900

3rd Step: Sample size at the national level: Furthermore, since the data is supposed to be
analyzed by sex wise and 2 age category (0-15, 15-30), the sample size is multiplied by 4
n = 900 * 4=3600 (sample size)

We will recruit same number of other than Tharu population as comparison group to
determine/compare the prevalence of hemoglobinopathies.

Sampling strategy for hemoglobinopathies and G6PD deficiency survey

1. A total of 7200 persons (900 in each of eight categories) will be required, of which
3600 will be from Tharu group and 3600 are from other than Tharu.
2. Primary sampling units (PSU) are wards as defined in census 2011.
3. 32 persons (8 person x 4 categories) will be selected in each PSU from each of Tharu
and other than Tharu.
4. 225 (rounded to 226) PSU will be required which is split into 113 each for Tharu and
other than Tharu.
5. In the 20 Terai district this number 113 is proportionately distributed, according to
share of Tharu Households and other than Tharu Households numbers.
6. In proportional allocation Dhanusha district got no PSU for Tharu HH, so it will be
assigned one PSU.
7. PPS sample of 114 PSU will be taken by taking number of Tharu households as
measure of size (MOS). In these PSUs only Tharu Households will be listed and
sampled.
8. Again, PPS sample of 113 PSU will be taken by taking number of other than Tharu
households as measure of size (MOS). In these PSUs only non Tharu Households will
be listed and sampled.
9. Some PSU will be common in (7) and (8) in those PSU both Tharu and other than
Tharu households will be listed and sampled.

Data collection Tool and Technique

Socio-demographic information and family history of the participant will be taken by using
structured questionnaire by the trained enumerators which will be followed by phlebotomy of
3ml of blood by the experienced Staff nurses/Health Assistants in a 5 ml EDTA vial. All the
collected samples will be maintained in cold chain boxes until further processed. For the
diagnosis of heamoglobinopathies, a preliminary screening by complete blood count along
with reticulocyte count which is followed by Hb electrophoresis or HPLC (High Performance
Liquid Chromatography). Diagnosis of G6PD deficiency will be carried out using rapid assay
kit (………Company) and the procedure provided in the manual with the kit will be followed.

The positive samples of hemoglobinopathies (Thalassemia and Sickle Cell disease) and G6PD
will be stored at -800 C for further research purposes in future.

Participants will be reported for their test outcomes. Patients who diagnosed positives for
hemoglobinopathies (Thalassemia and Sickle Cell disease) and G6PD will be further advised
to visit health care center/ hospitals for further action and counseling.

Validity and Reliability of the Tool

Questionnaire will be designed with expert consultation. Labeling of the blood samples will
be done based on bar coding system to minimize the primary error of entry. For blood
collection, adequate training will be conducted to enumerators and the recruitment will be
done based on the experience of the phlebotomist. Blood testing instrument will be calibrated
in specific time interval. Quality control documentation of CBC and Hb electrophoresis/
HPLC equipment’s will be maintained on a daily basis before running the samples. Cross
validation of the samples will be done from the internal laboratories and instrument will be
checked regularly for their consistency and adequacy.

Potential Bias

Certain instrumental error may cause some cases to be under-reported or misreported.

Supervision and Monitoring

Field level supervision and monitoring will be done by investigators team from NHRC at
different phases of study like at initial phase, mid phase and end phase for the blood sample
collection, transportation and different test at different levels.

Data management and Analysis

Collected data will be stored in the Nepal Health Research Council office and will be handled
at the same place. Blood sample will be sent to the Local level facility or the central level
based on the capacity and performance for HPLC or Hb electrophoresis. Once the analysis
done the report will be collected by NHRC and then will be analyzed using bio statistical
software.
Ethical Considerations:

This study will receive ethical approval from Nepal Health Research Council (NHRC). Every
participant will be explained of procedure of the study, confidentiality of the information,
right to refuse in the study and written assent/ consent will be obtained before conducting
study. Participants will be informed regarding their right to withdraw from the study at any
time without penalty and issues concerning confidentiality and consent will be upheld in
accordance with ethical research standards. Data obtained from the research participants will
not be used for any other purpose than the research.

Expected Outcome of the Research

This study will helps to determine the prevalence of hemoglobinopathies (Thalassemia and
Sickle Cell disease) and G6PD among population of Terai district of Nepal. This study will
also help to compare the hemoglobinopathies and G6PD among Tharu and non-Tharu
population.

Plan for utilization of research findings

The overall findings of the research will be shared with the concerned authorized body and
Ministry of Health and Population (MoHP) about prevalence of Thalassemia and Sickle Cell
disease as well as G6PD and its associated complication. The distribution of
hemoglobinopathy related diseases in Terai districts will be helpful to apply public health
measures effectively for control and prevention of these diseases. Findings from G6PD
deficiency in Terai districts especially in malaria endemic region further supports MoHP to
and EDCD (Epidemiology and Disease control division) to strengthen  clear programmatic
and policy-making implications for screening G6PD before malaria treatment. The report
further will be helpful to finalize the protocol of Thalassemia and Sickle Cell disease
management.

Estimated Budget Breakdown

We will develop together (as per Executive Chief Instructions)

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Annex-1

Table 1: Number of PSU for sampling of Tharu and other than Tharu in each district

Final PSU for other than

District Final PSU for Tharu Tharu
Jhapa 1 9
Morang 4 10
Sunsari 6 7
Saptari 5 5
Siraha 2 6
Dhanusa 1 7
Mahottari 1 6
Sarlahi 1 7
Rautahat 2 5
Bara 5 5
Parsa 3 4
Chitawan 4 6
Nawalparasi 7 6
Rupandehi 6 8
Kapilbastu 5 4
Dang 11 4
Banke 5 4
Bardiya 15 2
Kailali 22 5
Kanchanpur 8 3
Total 114 113

Annex-II

Complete blood Count

CBC with reticulocyte count of the collected blood samples will be done by the automated
analyzer majorly for the quantitative determination of Hb types, MCV, MCH, and RDW
as MCH<77pg, MCV<27 fl and HBA2 of >3.5% are indicative of hemoglobin disorders
(Thalassemia and Sickle Cell disease).

The phlebotomy and CBC process flowchart

3ml of blood will be collected by the median cubital venipuncture of the research
participants by trained Health assistants/pediatric staff nurses adhering to the WHO
guidelines for phlebotomy.
 All the collected samples will be labeled, handled, stored and transported properly to
maintain the integrity of hemoglobin as high heat and humidity can change the levels of
hemoglobin A and S

The samples will be stored immediately in cold chain boxes and transported to the local
facilitator for the storage and processing of the blood samples.

The blood samples will be run for complete blood count and reticulocyte in an automated
five parts hemolytic analyzer within four hours of collection of blood samples.

Annex-III

Flow Chart of Technique for the study

Visit to the selected site for the feasibility study

Pilot study to be performed in 100 -150 samples in a similar setting

Introduction about the study to the participant

Obtaining ethical consent (verbal &written) from the participant and ethical assent from the
participants under 18 years

Obtaining socio demographic and clinical family history of the same disease from the
participant a day before blood collection

Determining the height, weight, and blood pressure of the respondents

Collecting Blood sample from the selected participant as defined above

 Transportation of the Blood samples will be transported to the Local level facility
for CBC.

Complete Blood Count test of selected blood sample at local level facility

G6PD test and High Performance Liquid Chromatography (HPLC) or Hb electrophoresis

test will be done at the Center

Annex-IV
Laboratory diagnosis of hemoglobinopathies (Thalassemia and Sickle Cell disease)
 Annex V

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Annex VI

For Bar Coding

R e s e a r c h P a r tic ip a n t
N um ber└ ─ ┴ ─ ┘ └ ─ ┴ ─ ┴ ─ ┘ House Hold Number└─┴─┴─┘└─┴─┘
Survey information

Location and Date Response code

1 Interviewer ID ……………… I1

2 Province ……………….. I2

3 District ………………. I3

4 Local level ……………… I4

5 ward no. ………………. I5

6 Date of interview ………………. I6

7 Time of interview ……………… I7

Consent , Interview language and Name Response Code

Consent has been read and obtained Yes 1 I8
8
No 2 If no, End
Interview language English
9 Nepali I9

10 I10
First Name ………………..

11 Family name ………………. I11

12 Contact number where possible I12
Enter 88 if refused and Enter 99 if not
available (must be in 10 digit)
…………………
 Demographic information
Question Response code
C1
Male 1
13 Sex Female 2
Others 3
C2
What is your date of birth? └─┴─┘ └─┴─┘
└─┴─┴─┴─┘
14
d ye
Don’t know ……………. 77 d mm ar
C3
15 How old are you? └─┴─┘ Years

Unmarried C4
Married
Separated 1
Divorced 2
Widowed/Wido 3
wer 4
Cohabitatin 5
g 6
16 What is your marital status? Refused 88
Not able to read and write 1
C5
Able to write and

write(Informal education) 2

17 Less than primary school 3

What is the highest level of Primary school completed 4

Secondary school
education you have completed? completed 5

Higher secondary (10+2)/

PCL 6

Bachelor degree completed 7

Post graduate degree 8
Refused 88
 Dalit 1
2, Tharu, if yes,
Disadvantaged Janajatis G Q.19
C
18 What is your ethnic background? Disadvantaged non-Dalit Terai 6
3
Religious minorities 4
Relatively advantaged Janajatis 5
Upper caste groups 6

Others 7

Refused 88
Which Sub-group of Tharu you C
19 belong to? Rana Tharu 1 7
Kathoriya Tharu 2

Sonha Tharu 3

Dangaura Tharu 4

Paschuhan 5

Rautar Tharu 6

Purbaha Tharu 7

Aarkutwa or Chitwania 8
Tharu

Kochila Tharu 9

Danuwar 10

Lampucchwa Tharu 11

Pahalman Tharu 12
Hindu 1
Buddhist 2
Christian 3
Muslim 4 C
20 Which religion do you believe in? Other (please specify)……. 5 8
 C
Government employee 1 9
Which of the following best Non-government employee 2

21 describes your main work status Self-employed 3

over the past 12 months? Non-paid 4
Student 5
Homemaker 6
Unemployed (able to work) 7
Unemployed (unable to work) 8
Retired 9
Others (please specify) 10
Refused 88

History of Clinical Illness and Medication

Question Response Code

Have you ever visited or admitted to Yes 1

hospital?
22 HC1
No 2 (G Q. 29 )

If yes, how many times have you been

admitted or 1
23 visited to hospital? HC2
2
3
>3
24 During the past had you been diagnosed or Yes 1 HC3
screen for any type of hemoglobin
Disorder? No 2 ( G Q. 29)

Sickle cell anemia

1
Thalassemia
2
If yes, which type of hemoglobin disorder Other (please specify)
25 has you? …………………. 3 HC4

Did anyone in your family members have Yes 1

26 hemoglobin disorder? No 2 HC5

Great Grand Father

Great Grand Mother
Grand Father
Grand Mother
Father
26.1 If yes to whom… Mother HC6
26.2 Which type of hemoglobin disorder had HC7
 Sickle cell Trait/disease/Anemia 1
Thalassemia 2
your family member? Other(please specify)……………3

27 What are the sign and symptoms Pale discoloration of palm or skin 1
have you experienced or Yellowish discoloration of the eye 2
been experiencing? If Sickle cell
anemia and others (select multiple) Difficulty in breathing 3 HC8
History of loss of consciousness or
seizure 4
Leg ulcer(healed or non-healing) 5
FOR MALE: Do you have Involuntary HC1
28 penile Yes 1 2
erection that lasting for more than 30
minute No 2
(Stuttering) or that is recurrent? If sickle
cell anemia and others
Have you ever experienced or visited
29 hospital for severe or chronic Pain? Yes 1
(if, sickle cell anemia and others)
(Exclude Traumatic causes of pain) No 2
29
.1 If Yes, which part of the body Extremities (Hand foot syndrome) 1
you experienced the pain ? Joints 2
HC1
Chest 3 3
Abdomen 4
Back of the trunk 5
Head 6
Others 7
Are you currently receiving or have
received any , treatment for Sickle cell Yes 1
Anemia by a
Health care workers? No 2 HC1
30 4
Yes 1
No 2
If yes Can you mention name of the 8 HC1
Drugs/Medicine currently receiving or
31 have received for Sickle cell Anemia? Don’t Remember 85
Please mention the name of the drug or
32 medicine Cap.Hydroxyurea 1 HC1
NSAID 26
Opioids 3
Folic acid 4
Tricyclic Antidepressants 5
(Amitriptyline/Nortriptyline)
.
Others (please specify) …… 6
Pale discoloration of Skin/eyes 1
Jaundice 2
Hepatomegaly 3
Splenomegaly 4
What are the sign and symptoms have you Fatigue 5 HC1
experienced or experiencing? (If
33 Thalassemia and others) (multiple choice) Other(please specify) 6 7
 Are you currently receiving or have Yes 1 HC1
received any, treatment for Thalassemia by
34 health care workers? No 2 8
Yes 1
If yes Can you mention name of the No 2
Drugs/Medicine currently receiving or
35 have received for thalassemia? Don’t Remember 88
Deferoxamine/Deferasirox
/Deferipron 1
Please the mention the name Luspatercept 2 HC1
36 drugs/medicines Others (please specify) 96 9

History of Blood Transfusion History

Co
Questions Response de
Yes 1
Have you ever had blood Transfused?
37 B1
No 2

3 If YES, For what reason have you

8 been Blood transfused? Sickle Cell Anemia 1 B2
Thalassemia 2
Surgery 3
Others (please specify) ………… 4
Age (years) └─┴─┘
If yes, At what Age you had your first B
39 Blood Transfusion? Don’t know 77 3
4 How Many times have you been Number of B
0 Blood transfused (including recent)? Episode └─┴─┘ 4
Date :- D/M/Y
If yes, When you did recently └─┴─┘└─┴─┘
4 received your Blood └─┴─┘ B5
1 Transfusion? Don’t know 77
1 (Please
If yes, Did you have any
4 Complication during or after Blood Yes specify……….) B6
2 Transfusion? No 2
 Anthropometry measurement
43 Interviewer ID └─┴─┴─┘ M1
Height └─┴─┘ M2a
44 Device IDs for height and weight
Weight └─┴─┘ M2b
in centimeters
45 Height (cm) └─┴─┴─┘. └─┘ M3
46 Weight in kilograms (kg) └─┴─┴─┘.└─┘ M4

Hematological Measurement
Co
de
47 Name of Phlebotomist ________________________ :
B
1
VENOUS Blood
Sample
B
48 Consent obtained Yes .......................1 No ......................... 2 2
B
49 Sample taken Yes .......................1 No ......................... 2 3
Yes, completely .............................................. B
50 Sufficient Volume 1 4
Yes, Partially ..................................................
2
No.....................................................
3
Refused .....................................................
4
Time of Venous
Blood Sample Collection
B
Date of sample taken
51 (Day/Month/Year) /
5

Time of blood collection began (Hour: B

52 minute) : 6
Ho
ur Minute
 Complete Blood Count

Question Response Code

53 Hemoglobin (Hb) mg/dl B7

└─┴─┘.└─┘
54 Mean Corpuscular Volume (MCV)

FL └─┴─┘.└─┘ B8

└─┴─┘.└
55 Mean Corpuscular Hemoglobin (MCH) Pg ─┘ B9
└─┴─┘.└
56 Mean Corpuscular Hemoglobin Concentration Dl ─┘ B10
(MCHC)
5
7 Red Blood Cell Distribution Width (RDW) B11
%└─┴─┘.└─┘
5
8 Platelets n*103 Cells/µL B12

59 Red Blood Cell Count (RBC) n*103 Cells/µL B13

60 White Blood Cell Count (WBC) n*103 Cells/µL B14

61 Neutrophil └─┴─┴─┘% B15

62 Lymphocyte └─┴─┴─┘% B16

63 Monocyte └─┴─┴─┘% B17

64 Eosinophil └─┴─┴─┘% B18

65 Basophil └─┴─┴─┘% B19

High Performance Liquid Chromatography (HPLC)

Question Response Code
Technician ID └─┴─┴─┘ LC1
└─┴─
66 Device ID LC2
┘
└─┴─┘: └─┴─┘
Time of day blood specimen taken (24 hour Hours:
67 clock) minutes LC3
hrs mins
└─┴─┴
─┘ %
68 HbA LC4
 └─┴─┴
69 HbA1 ─┘ %
LC5

└─┴─┴
70 HbA2 ─┘ %
LC6

└─┴─┴
71 HbF ─┘ %
LC7

└─┴─┴
72 HbS ─┘ %
LC8

└─┴─┴
73 HbD ─┘ %
LC9

└─┴─┴
74 HbC ─┘ %
LC1O

└─┴─┴
75 HbE ─┘ %
LC11

└─┴─┴
76 HbG ─┘ %
LC12