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UNIVERSITY OF MEDICINE AND PHARMACY CRAIOVA

FACULTY OF GENERAL MEDICINE

Ph.D. THESIS
Beta- thalassemia trait - epidemiological and clinical aspects in children
in Constanta County

Scientific Supervisor,
Professor of Pediatrics DUMITRU BULUCEA, Ph.D.

Ph.D. Candidate,
FRECUS CORINA ELENA

2010

SUMMARY

I.
II.

INTRODUCTION
STATE OF KNOWLEDGE

1. History of diagnosis and treatment in beta-thalassemic syndromes


2. History of beta-thalassemic syndromes in Romania
3. Ethnic distribution
4. The influence of malaria
5. Inherited thalassemia
6. Classification of the thalassemias
7. Human Hemoglobins : composition and genetics
8. Molecular diagnosis of beta-thalassemic syndromes
9. Relation between genotip and phenotip of beta-thalassemia
10. Molecular genetic tests of beta-thalassemia
11. Pathophysiology of the beta-thalassemic syndromes
12. Diagnosis of the beta-thalassemic syndromes
13. Treatment
14. Genetic counseling and prenatal diagnosis
III.
A.
B.
C.
D.
E.
F.
G.
H.
I.
J.

PERSONAL CONTRIBUTION
Introduction
Objectives
Materials
Methods
Results and discussions
Conclusive discussions
Clinical cases
Conclusions
References
Annexes

Key words: beta-thalassemia trait, children, epidemiology, clinical and


hematologic aspects

INTRODUCTION
The thalassemias are a heterogeneous group of inherited anemias caused by
mutations affecting the synthesis of hemoglobin. Beta thalassemia is most
common in persons of Mediterranean, African, and Southeast Asian descent.
Thalassemia trait affects 5 to 30 percent of persons in these ethnic groups
Scientists and public health officials predict that thalassemia will become a
worldwide issue in the next century.
Beta-thalassemic syndromes are frequent in Romania, and the south-east part of
our country seems to be the most affected by this kind of genetic abnormality.
The paper Beta- thalassemia trait - epidemiological and clinical aspects in
children in Constanta consists in 197 pages: 66 pages about the state of
knowledge in beta-thalassemic syndromes and 131 pages of personal
contribution.
The first part of the thesis, structured in 14 chapters, focuses on the state
of the art date regarding beta-thalassemic syndromes: Historical data,
Epidemiology, Classification of the thalassemias, Hemoglobin composition and
genetics, Physiopathology, Clinical and laboratory findings in beta-talassemic
syndromes, Treatment. The information is presented using 17 figures and 13
tables.
The main historical data about beta-thalassemic syndromes are
systematically displayed, in terms of international and Romanian chronology. The
historical aspects are followed by the theoretical part of the thesis describing the
main data regarding thalassemia: histological aspects, hemoglobin genetics and
classification, clinical findings, laboratory data and therapeutic intervention.
The second part of the paper the personal contribution- represents two
thirds of the thesis, containing 98 tables and 65 figures. There are eight chapters:
Objectives, Materials, Methods, Results and discussions, Conclusive discussions,
Conclusions, References and Annexes.
The main goals of the work were:
- to make an epidemiological study regarding thalassemia trait in children
from Constanta county
- a better understanding of clinical and hematological aspects of thalassemia
trait in children
- comparative
clinical and laboratory study between ethnical groups
diagnosed with beta thalassemia trait
- comparative laboratory study between children with thalassemia trait and
children diagnosed with iron deficiency anemia
Material and methods An observational retrospective and prospective
study was done on 204 patients diagnosed with thalassemia trait in the Pediatric
Department of the County Hospital of Constanta from 1999-2008.
I determined the frequency and the prevalence of the thalassemia trait of
the study group, the geographical distribution, clinical findings and hematologic
parameters analyses. The data were introduced into electronic system (Microsoft
Access 2007) and were statistically analyzed.
The cases were divided in two groups: the retrospective study (41 cases)
and the prospective study (163 cases). According with hemoglobin electrophoresis
I made a statistic comparative hematologic study between three groups of
children: L1 group (HbA2>3.5%), L2 group (HbA2>3.5% and HbF elevated), L3
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group (HbA2<3.5 % and HbF elevated). There were used some mathematical
formulas in order to compare thalassemia trait with iron deficiency anemia.
Discussions and conclusions:
The performed study confirms that the beta-thalassemic syndromes are
microcytic hipochromic anemias of normal or hypersideremic type with high level
presence worldwide and also present in our country.
The cumulative incidence of heterozygote beta-thalassemia during the
study performed (1999-2008) in Constanta County was 0.32%. The prospective
study incidence was four times higher (0.49%) compared to the retrospective
study.
The achieved prevalence for Constanta City was double (0.28%) than the
rest of the County (0.13%). Ethnic prevalence study revealed that the highest
prevalence was obtained in Greeks (1.43%), followed by Romani people (1.25%)
and Turks (0.29%); the lowest prevalence was obtained for the Romanians.
The average diagnosis age for the entire group of children was 8.38+/-5.85
limited from 9 months to 18 years, having its peak frequency at the 7 to 16 years
group (31.86%). The statistic analysis of the average diagnosis age of the
prospective group compared to the retrospective group showed no significant
statistic differences between the two groups (p=0.3086).
The clinical signs in heterozygote beta-thalassemia was mostly unspecific,
the pallor (rarely associated with scleral subicterus: 18.62%) being the only
constantly evidenced sign. Mild splenomegaly was diagnosed in 15.19% of the
cases, especially in L2 group of children, in which hemoglobin electrophoresis
showed HbA2>3.5% and slightly increased HbF (36.36%).
The minor thalassemia associated pathology was diverse: un important
number of children showed low weight (65 cases), associated iron deficiency
anemia (40 cases), common rickets (22 cases), allergic rhinitis (23 cases),
infantile recurrent wheezing (12 cases), atopic dermatitis (16 cases), asthma (7
cases). Particularly, we discovered within our group of children one case of Wilson
disease, one case of Silver-Russel syndrome and one with diabetes mellitus.
In the performed study we obtained p<0.0001 (extremely statistic
significant) for patients diagnosed with asthma/infantile recurrent wheezing (19
cases) compared to the number of asthma cases from all hospitalized cases.
Hemoglobin-disease detection should be performed for patients having asthma
associated with microcytar hypocromic anemia.
Hemoglobin level was slightly lower for the studied group (average
hemoglobin level was 10.55+/-0.97); the majority of the cases (85 cases; 41.67%)
had the Hb between 10 and 11g/dl; not to be neglected is the percentage of
patients with Hb>11g/dl (38.80%) with high occurrence in children over 3 years old
and teenagers; normal hemoglobin does not exclude heterozygote thalassemia.
The average number of erythrocytes was 5.57+/-0.56; the majority of the
cases (164; 80.39%) had a number of erythrocytes more than 5 mil/cmm; most of
the cases of minor thalassemia associated with iron deficiency had the number of
erythrocytes between 4.5-5mil/cmm.
The average of MCV value was 61.38+3.99; most of the cases (100
patients; 49.02%) showed values of MCV between 60-65 fl.
The average of MCH value was 20.02+/-1.76; the majority of the cases
(140 patients; 73.53%) had the MCH value between 18-22 pg.
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The RDW values were normal in the majority of the cases (137; 67.15%).
Increased RDW values were encountered especially in cases of iron deficiency
anemia associated with minor thalassemia, as well as in the L3 group of patients
(with hemoglobin electrophoresis showing normal HbA2 and slightly increased
HbF, interpreted as thalassemia).
The correct interpretation of the blood film in a hypochromic anemia can
suggest the diagnosis of heterozygote beta-talassemia; thus, changes of the
erythocitary series as hypochromia, microcytosis, anisocytosis, poikylocitosis,
target-erythrocytes, macrocytosis, erythrocytes with azurophiles granules and
tear drop shaped erythrocytes, schistocytosis, ovalocytosis and spherocytosis
being present with a frequency that decreases according to the order above, along
with a slightly lower hemoglobin.
Depending on the hemoglobin electrophoresis results, the L1 group
(increased HbA2) was best represented (78.92%), followed by the L2 group with
both HbA2 and HbF increased (16.18%) and the L3 group (4.9%) with normal
HbA2 and HbF present ( thalassemia).
By statistic comparison analysis of the three groups, we achieved the
following results: between groups L1 and L2 there is a significant statistic
difference regarding two parameters: hemoglobin and the number of erythrocytes
(p<0.005); between groups L1 and L3 there are the most significant statistic
differences for the parameters: Hb, MCV, MCH, RDW (p<0.005); between groups
L2 and L3 there is a highly significant statistic difference only for RDW (p<0.005).
The majority of heterozygote thalassemia associated with iron deficiency
cases were diagnosed between 6 months and 3 years and had Hb between 9 an
10 g/dl; we obtained extremely significant statistic differences (p<0.001) for Hb,
MCV, RDW and the number of erythrocytes and highly significant statistic
differences for MCH (p<0.01) between these patients and those without
associated iron deficiency. Furthermore, for the heterozygote thalassemia with
associated iron deficiency group, we obtained a much lower percentage of
accurate diagnosis (by applying differential formulas) than the group without iron
deficiency.
The best indexes that showed the distinction between iron deficiency
anemia and minor thalassemia (as Youden criteria) were Mentzer and Ehsani
indexes (Youden index=95.11), followed by England index (Youden index=93.23),
Shein-Lal index (Youden index=82.70) and Srivastava index (Youden
index=81.95). The two indexes (Mentzer and Ehsani) together can provide a
better distinction between the two types of anemia, hypochromic and microcytic.
Monitoring hematological parameters such as RDWi (Youden index=93.6) and
RBC (Youden index=87.96) proved to be useful as well since these parameters
obtained maximum Youden indexes in other studies.
The data of the present study allow issuing the hypothesis of the existence
of an iceberg phenomenon, assuming that the real incidence and prevalence in
Dobrogea is much higher and that these epidemiologic indexes will increase
during the next years, pending that medical and research activity, both at primary
assistance level (family medicine) and secondary and third level assistance
(hospital), will further pursue the discovery of heterozygosity, especially by
activities within the outbreak center.
Considering that the beta-thalassemic syndromes represent a public health
issue, both by their frequency and by the social and medical problems they arise,
it is of outmost importance that primary prophylaxis should be performed in a
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correct manner, at least by timely postnatal identification of heterozygotes, useful


method for medium occurrence rate of heterozygotism .
The clinical and laboratory study, based on hematological examinations
(Hb, Ht, erythocytary indexes, blood film examination and so on) of heterozygote
beta-thalassemia cases led to the elaboration of a clinical and hematological
profile of the heterozygote beta-thalassemia for an easier diagnosis at lowest
ages possible, when primary and secondary prophylaxis will have the best
efficiency.

References
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globin gene for identification of beta thalassemia mutation n Romanian
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Ictero- anemiile hemolitice. Ed. Med. Buc., 1986, 91- 116
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CURRICULUM VITAE

Name: FRECUS CORINA ELENA


Date of birth: 25.10.1970
Married, two childrens
Place of birth: Constanta, Romania
Address: 31, Dorobantilor street, Eforie Nord, Romania
Studies:
-High School of Mathematics and Physics Mircea cel Batran,Constanta, 19851989
-Faculty of General Medicine, Ovidius University, Constanta, 1990-1996
1. Professional activity:
- resident doctor -1997-2002 Clinical County Hospital Constanta
- universitary preparator from september 2000 in Ovidius University
Constanta, Faculty of Medicine, Pediatrics Department
- universitary assistant: from December 2002, in Ovidius University Constanta,
Faculty of Medicine
-specialist pediatrician: 2002, in Clinical County Hospital Constanta. Place of
work: Pediatric Department.
- Gastroenterology master , achieved in 2007
- Trainer for a doctor degree Thalassemia trait - epidemiology and clinical
aspects in childrens, UMF Craiova, from 2003, coordinator Professor Dumitru
Bulucea.
2. Scientific activity:
-participations in National Pediatrics Congresses : Sibiu 1998, Constanta 2003,
Galati 2005, Sibiu 2007, Iasi 2009
-participations at Romanian-American Symposiums, Mamaia, Poiana-Brasov,
Cluj Napoca, Durau
-Participation at Pediatric Gastroenterology Congress, Cluj Napoca 2003,
National Gastroenterology Congress in Mamaia 2004, Poiana Brasov 2005,
National Semiology Symposium, Mamaia, 2000

- practical training in Brest feeding Sinaia -2002


- participation at International Congress of Neonatology, 17-18 november 2008,
Roma
- participation at International Congress of Pediatric Research , Hamburg, 2009
I supervised practical lessons in pediatrics and license thesis.
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Books (co-author):
1. Pediatricsvol.2, Ovidius University press, Constanta 2001
Valeria Stroia, Adriana Balasa, Corina Frecus
2. Pediatrics Publishing house Andrei Saguna foundation, 2001
Valeria Stroia, Cristina Fiscuci, Adriana Balasa, Cuzic Viviana, Corina Frecus,
Larisia Mihai
Number of summarized scientific studies published in the country: 42 (co-author)
Number of full scientific studies published in the country: 21
Number of scientific studies to the annual sessions of General Medicine Faculty7
Member of Romanian Pediatric Association: Romanian Pediatric Society
Member of Genetic Society
Dr. Frecus Corina