Dr . Muhammad RafiqueAssist. Prof. Paediatrics

College of MedicineK K U Abha K S A

PRENATAL DIAGNOSIS

It introduces option for management. From interruption of abnormal pregnancies to prepare for specialized peri-natal care.

PRENATAL Dx.------CONT.

Methods of prenatal diagnosis:

.Maternal blood sampling. .Fetal ultrasonography. .Fetal X-ray .Fetal MRI .Amniocentesis .Chorionic villus sampling

MATERNAL BLOOD SAMPLING

• Elevated alpha fetoproteins:• Twins , • intestinal obstruction , • neural tube defects.

• Decreased alpha fetoproteins:• Trisomy 21/ Down synd.

• Elevated chorionic-gonadotropins:• Trisomy 21

MATERNAL BLOOD SAMPLING—CONT.

• Deceased chorionogonadotropin: .Trisomy 13, .Trisomy 18• Decreased unconjugated estriol: .Trisomy 21

FETAL ULTRASONOGRAPHY

Maternal trans-abdominal USG can visualize all major fetal organs at 16-18 weeks of gestation and can diagnose disorders like;

Hydrops fetalis , hydronephrosis , neural tube defects , intestinal obstruction , CHD , diaphragmatic hernia , limb reduction anomalies and growth assessment etc.

CORDOCENTESIS

Fetal blood can be directly sampled in late gestation through USG guided per-cutaneous umbilical cord blood sampling & can detect;

.fetal anemia . Thrombocytopenia .thalassemia . Fetal oxygenation .acid base disorders .cells for DNA .fetal hypo-albuminemia

CHORIONIC VILLUS SAMPLING

CVS at 10-12 weeks provides good cyto- trophoblast (dividing cells) DNA for molecular analysis that can be rapidly karyotyped.

Detection possible for: Thalassemia, hemophilia A, PKU, CAH., alpa-1 antitrypsin deficiency, SCD etc. Trisomy 21,18,13,turner&klinefelter synd.

AMNIOCENTESIS

Amniotic fluid can be sampled at 10-14 weeks gestation for fetal cells -can be cultured For • Cyto-genetics • Molecular analysis• Metabolic analysis.• Alpha-fetoproteins measurement.Note –complication of miscarriage <1%.

Amniocentesis

INDICATIONS FOR CVS/AMNIOCENTESIS

• Maternal age above 35 years.• Previous child with chromosomal abnormality. • Either parent a translocation carrier.• F/Hx. of genetic disease, Dx. by biochemical or

DNA analysis.• Parental request for sex determination because

of F/Hx. of X-linked disorder.• Maternal blood sample show chromosomal abn.• As a part of work up for fetal anomalies by USG.

GENETIC TESTING

Analysis of genetic material to obtain information related to individual’s health status using either chr. analysis or DNA

based testing.Two types. 1-Linkage analysis 2-Direct mutation analysis

LINKAGE ANALYSIS

It is used if• Responsible gene is mapped but not yet identified.• Impractical to find specific mutation due to

large size & No. of different mutation in a gene e.g. CF.

DIRECT MUTATION TESTING

• It is preferred to detect gene mutation because genome sequence is elucidated and

technologies improved.• It avoids pitfalls of linkage testing by detection of

gene mutation.• In some disorders all individual have same

mutation. e.g. SCD.• In some, people have different mutation e.g. CF.

PREDICTIVE GENETIC TESTING

• Testing in individuals who are at risk to develop a genetic disorder e.g. due to F/Hx. before developing s/symptoms , especially which disorders are age dependent and s/s increase with increasing age , e.g. Huntington disease, cancer.

• Note-development of disease is not necessary in every mutation containing person.

GENETIC COUNSELLINIG

• Process by which a person is informed about possibility of developing a disorder or transmitting it to his offspring and chances of his offspring having similar disorder.

• Important steps are ; .History .pedigree

construction .physical examination . diagnosis, .counseling .follow up

GENETIC OUNSELLING STEPS

• Offer possible help to manage disorder.• Sex detection & pre-natal Dx.by amniocentesis

to decide parents to have/not the baby.• Explain outcome statistically for each pregnancy.• Provide good knowledge about disorder’s

etiology, Dx. and Mx. in discussion.• Provide new information whenever available.

GENETIC COUNSELLING STEPS

Timing of counseling; • Not too early (because family still concerned about welfare of affected individual & emotionally upset). • Not too late (because an other at risk fetus is conceived ).

GENETIC OUNSELLING STEPS

• Careful F/Hx. and pedigree construction• Document pre-natal, pregnancy & delivery Hx.• Review available informations about disorder.• Careful physical exam. of affected individual

and apparently unaffected family individuals.• Establish accurate Dx. by available tests.• Mode of inheritance establish and explain.

GENETIC OUNSELLING STEPS

• Therapy and referral; Some pt. need specialist care e.g . Turner

synd. need endocrinologist evaluation. • Prevention of complications should be priority• Support groups; Provide information about support group that

provide funds for research on specific disorder• Nondirective counseling;

INDICATIONS OF GENETIC COUNSELLING• Advanced parental age; .Maternal >35 years .Paternal >50years• Previous child with F/Hx. of .congenital abnormality .Dysmorphology .Isolated birth defects .metabolic disorder .Chromosomal abnormality .single gene defect

INDICATIONS OF GENETIC OUNSELLING

• Adult onset disease/pre-symptomatic testing ; .cancer .Huntington disease• Consanguinity• Teratogenic exposure• Repeated miscarriage or infertility

INDICATIONS OF GENETIC COUNSELLING

• Pregnancy screening abnormality .maternal high alpha fetoproteins .fetal USG .fetal karyotyping• Heterozygote screening based on ethnic risk; .sickle cell anemia .thalassemia• Follow up to abnormal genetic testing

MANAGEMENT &TREATMENT OF GENETIC DISORDERS

• GD mostly chronic.• Few are amenable to curative therapies.• Provide information about GD, G.counseling ,

anticipatory guidance and appropriate medical surveillance.

• Surgical Mx. is available for many congenital anomalies that are predisposition to tumours.

MANAGEMENT &TREATMENT OF GENETIC DISORDERS

• PHYSIOLOGICAL THERAPIES;• Attempts to ameliorate phenotype of a GD.• Underlying GD not altered by treatment.• Used in inborn error of metabolism e.g. • dietary manipulation in PKU, avoiding phenylalanine diet. .Enzyme supplement for methyl-malonic-acidemia. .Bisphosphonate for osteogenesis imperfecta.

MANAGEMENT & TREATMENT OF GENETIC DISORDERS

PHARMALOGICAL THERAPIES;• Use of pharmaceuticals to remove ammonia in

urea cycle defect.• It directly target defective cellular pathway

that is altered by abnormal /missing gene.Replacement therapy ; e.g. .cystic fibrosis .Gauscher disease

MANAGEMENT & TREATMENT OF GENETIC DISORDERS

Organ transplant;• E.g. transplant of liver in inborn error of

metabolism.• Transplant of bone marrow in many

haemotological disorders.Gene therapy;• Less invasive mean of achieving cure of a GD.

PREVENTION OF GENETIC DISORDERS

• Some GD has gene defect + environmental factors impact e.g. multifactorial disorders.

• Prevention possible by .avoiding inciting environmental factor e.g.avoid maternal phenytoin use to avoid fetal CHD. .By instituting treatment that reduce risk e.g. maternal use of folic acid to avoid fetal neural- tube defect.

NEWBORN SCREENING TESTS

1-Chromosomal analysis; Body cells are cultured ,arrested in mitosis in

metaphase .Chromosome first evaluated microscopically then images are captured by video camera and stored on a computer . Homologous chr. are paired , arranged into Karyotype. Chromosomes are exposed to a hypotonic solution, fixed and stained with

“ trypsin Giemsa”which produces dark and light bands for detail study.

NEWBORN SCREENING TESTS

2-FISH (fluorescence in situ hybridization); - Used to identify , presence, absence, or

rearrangement of specific DNA segment.

- It involve unique DNA sequence labeled with a fluorescent dye , which is exposed to single stranded DNA on a microscopic slide.

NEWBORN SCREENING TESTS

FISH---cont. -Probe, pairs with its complementary DNA

sequence and can be visualized by fluorescence microscopy.

- It is particularly useful for detecting very small deletion which not detectable by band analysis.

NEWBORN SCREENING TESTS

3-Spectral karyotyping (SKY) & multicolour FISH;• 24 different chr. Painting probes are used for chr.

and labeled with different combination of 5 fluorescent dyes(which omit at different wavelength).

• 23 pair of chr. have unique spectra of wavelength fluorescence.

• Especially used to identify complex chr. Re-arrangement found in tumours.

NEWBORN SCREENING TESTS

4-COMPARITIVE GENOMIC HYBRIDIZATION(CGH) -FISH based method to measure differences in a particular DNA sequence / chr. segment between two different DNA samples. -Pt’s DNA labeled with green fuorescent dye &

normal reference DNA with red . -Equal amount of two mixed &used for FISH.

NEWBORN SCREENING TESTS

4-COMPARITIVE GENOMIC HYBRIDIZATION(CGH) -Ratio of green to red fluorescence measured

along each chr. -Region of amplification of pt’s DNA shows

excess green fluorescence & region of loss of pt’s DNA shows excess of red fluorescence.

-If green and red same 1:1,chromosome would appear yellow.