BIOLOGY INVESTIGATORY PROJECT

SESSION – 2022- 23
Submitted by – Yashwardhan
Class – XIIth (Science)
Roll no. (CBSE) –
“MY INVESTIGATORY PROJECT ON
AMNIOSENTESIS”

UNDER THE GUIDANCE OF

Mrs. Aditi Yadav,
PGT (BIOLOGY)
 Contents
1.
Certificate

2. Declaration

3. Acknowledgement

4. Aim

5. What is amniocentesis?
Who is a candidate for
6.
amniocentesis?
What can be detected through an
7.
amniocentesis?
8. Procedure

9. After procedure

10. Rapid test

11.
Full karyotype
12. Result

13.
Complications of amniocentesis
14. Conclusion

15. A case study from a Media

journal

Certificate
 Declaration
I hereby declare that the project work
entitled “Amniocentesis”, submitted to
Department of Biology, Kendriya Vidyalaya,
RK Puram, Sector – 04 is prepared by me.

Yashwardhan
Class: XIIth (Science)
 Acknowledgement
I would like to express a deep sense of thanks and
gratitude to my project guide Mrs. Aditi Yadav ma’am
for guiding me immensely through the course of the
project. He always evinced keen interest in my project.
His constructive advice & constant motivation have
been responsible for the successful completion of his
project.
My sincere thank goes to our principal sir for his co-
ordination in extending every possible support for the
completion of this project.
I must thanks to my classmates for their timely help
and support for completion of this project.
Last but not the least; I would like to thank all those
who had helped directly or indirectly towards the
completion of this project.

YASHWARDHAN
Class- XIIth (Science)
 Aim:-
To study the
Amniocentesis in
humans and
showing the Merits
and De – Merits of
the process.
 What is amniocentesis?

 A procedure performed usually in the beginning of

pregnancy to detect chromosomal abnormalities in the
foetus.
 Usually done when a women is between 16 to 22
weeks of pregnancy.
 During this procedure a sample of amniotic fluid is
taken from the amnion sac (amnion) surrounding the
unborn baby and its DNA is examined for genetic
abnormalities.
 The amniotic fluid- has skin cells of the developing
body, as well as his/her waste products like urine.
Each cell from the baby in fluid contains their
complete set of DNA (genetic information). Analyzing
these cells helps the doctors access the foetus health
and detects any potential problems.
 The entire amniocentesis appointment lasts
approximately 45 minutes –most of which involves a
detailed ultra sound examination.
 Who is candidate for
amniocentesis?
 Any pregnant woman who, at her due date will be 40 years of
age or older.
 Any women whose combines results are high risk (blood tests
and/ or foetal ultrasound).
 Any women whose ultrasound results indicate a potential
foetal chromosomal abnormally.
 Any woman who has family history or whose partner has a
family history of one or more incidents of chromosomal
anomalies or genetic disorders with a high recurrence risk.
 It is also possible for women under the age of 40, with no
history of genetic disorder to have an amniocentesis; it is a
matter of personal choice.
What can be detected through an
amniocentesis?
 Nearly all chromosome disorder, including Down’s syndrome
as well as sex chromosome abnormalities.
 Several hundred genetic disorders, such as cystic fibrosis and
sickle cell disease. The test is not used to look for all of them,
but if your baby is at increased risk for one or more of these
disorders, in most cases amniocentesis can usually tell you
whether he/she has the disease.
 Neural tube defects such as spina-bifida.

 Amniocentesis is the only way to obtain information about

foetal lung maturity. Amniocentesis is occasionally used late
in pregnancy to assess whether the baby’s lungs are mature
enough for the baby to breathe on his own.
 PROCEDURE
1. An ultrasound is used as a guide to determine a safe
location for the needle to insert through the abdominal wall
to enter the amniotic sac. So the fluid may be safely
removed.
2.  A sample of amniotic fluid is collected through the needle.
The procedure takes about 45 minutes, although the
collection of fluid takes less than five minutes.
3. The amniotic fluid, which contains cells shed by the foetus,
is sent to the laboratory for analysis.
 After procedure
 Advised to rest for 24 hours
 No strenuous work or exercise for 72 hours
 No air travel for 72 hours
 Seek urgent medical attention if-
o Feeling shivery
o High fever of 38 degree Celsius or above
o Persistent lower back pain and abdominal pain
 After the amniocentesis procedure, the sample of amniotic
fluid (the fluid that surrounds the unborn baby in the womb)
will be taken to a laboratory for testing.
 There are two different type of tests
o a rapid test
o A full karyotype
 Rapid Test
 A rapid test looks for abnormalities on specific
chromosomes (the parts of the body’s cells that carry
genes). We can get its reports within24-48 hours. A rapid
test can identify a number of chromosomal conditions that
cause physical and mental abnormalities. These are:
o Down’s syndrome- symptoms are as follows
1. Flat facial features.
2. Small head and ears.
3. Short neck.
4. Bulging tongue.
5. Eyes that slant upward.
6. Atypically shaped ears.
7. Poor muscle tone.

o Edward’s syndrome-- symptoms are as follows

1. Physical Abnormalities. Such as small and abnormally
shaped head, abnormally small jaw and mouth, overlapping
fingers and underdeveloped fingernails. Scrunched fists.
Low-set ears. ...
2. Heart problems.
3. Developmental disorders.

o Patau’s syndrome-- symptoms are as follows

1. Cleft lip and palate.
2. an abnormally small eye or eyes (microphthalmia)
3. absence of 1 or both eyes (anophthalmia)
4. reduced distance between the eyes (hypotelorism)
5. Problems with the development of the nasal passages.

 The results of a rapid test should be ready after three working days.
This test is almost 100% accurate, but its only tests for the three
conditions listed above.
 Full Karyotype
 Each cell in the body contains 23 pairs of chromosomes. A full
karyotype checks all of these.
 The cells in the sample of amniotic fluid are grown for up to 10
days. In a laboratory before being examined under a
microscope to check for:
o The no. Of chromosomes
o The appearance of chromosome
 Results from full karyotype will usually be ready in 2 or 3
weeks. In about 1 in every 100 tests, the results may not be
clear. This could be due to the mother’s blood contaminating
the sample of amniotic fluid, which may have prevented cells
from growing properly.

Down’s syndrome Edward’s syndrome Patau’s syndrome

 Result
 After amniocentesis, the sample of amniotic fluid is sent to a
lab for analysis.
 Results usually take 10 days to three weeks depending on
the lab
 It takes that long to get result because the cells have to
grow in the lab in order to analyse them.
 In the lab, genetic and chemical test are done.
 For genetic tests, proteins, minerals and other compounds
in the amniotic fluid are analysed.
 Amniocentesis results will either be positive or negative.

Positive Test result Negative Test result

 If the test result is positive it  For most women who have
means the baby has the amniocentesis, results of
disorder that was being the procedure will be
tested for. One should be negative. That is, there
aware that there is no cure baby will not have any
for the majority of disorders that were tested
chromosomal conditions, so for.
she needs to consider her
options carefully.

 Options may include :  It is possible to have a

1. Continuing with her negative result from
pregnancy while getting amniocentesis but the
information and advice baby may still be born with
about the condition. So, the condition tested for or
she is prepared for another chromosomal
carrying for her baby. condition. This is because
2. Ending her pregnancy. a normal test result
3. Options for ending her doesn’t exclude every
pregnancy will depend on chromosomal disorder.
how many weeks
pregnant she is when
making the decision.
Complications of amniocentesis:

 It is important to be aware of the possible complications

during or after amniocentesis. These are outlined below:

Miscarriage:
o There is a small risk that amniocentesis can cause a
miscarriage (the loss of the pregnancy). The risk is estimated to
be around 1 in 100.

Club foot:
o Amniocentesis may cause club foot in baby. This is when the
baby is born with a deformed ankle and foot. However, the likely
hood of this happening is higher if you have amniocentesis
before week 15 of pregnancy.

Rhesus Disease:
o Rhesus disease is a condition where proteins in a pregnant
woman’s blood attack her baby’s blood cells.
o Rhesus disease is only possible if the mother’s blood is RH-
negative and the baby’s blood is RH-positive. If this is the case,
amniocentesis could trigger RH disease if the mother’s blood is
exposed to the baby’s blood during the procedure.

Injury from needle:

o During amniocentesis, the placenta (the organ that links a
mother‘s blood to the baby’s blood) may be punctured by the
needle. Sometimes this is necessary to access the amniotic
fluid. If this happens, the puncture wound usually heals
without any problems developing.
o Ultrasound (where high frequency sound waves are used to
create images) is now commonly used to guide the needle. This
significantly reduces the risk of injury from the needle.

Infection:
o In very rare cases, an infection may develop if the procedure
introduces bacteria in the amniotic sac (the sac surrounding the
foetus that contains amniotic fluids). This can cause:
  A high temperature (fever) of 38-39 degree Celsius or
above
 Tenderness of abdomen (tummy)
 Contractions (when your abdomen tightens then relaxes).
 One should seek medical attention if she has any of these
symptoms. The risk of developing a serious infection from
amniocentesis is estimated to be less than 1 in 1000.

FACT – There is a conditional ban on Amniocentesis under India’s

Abortion Acts { such as MTP Act 1971 and Pre-Natal Diagnostic
Techniques Act (PCPNDT) in 1994 }. As this was a technology for
detection of abnormalities in foetus but in India this was used as
weapon to influence FEMALE FEOTICIDE. This technology could
determine the gender of foetus at early stage of Pregnancy. People
force the pregnant victim to whether abort the foetus or kill it after
birth, if she is a girl child.
 Conclusion

Amniocentesis is a safe procedure that can provide

helpful information about the health of foetus. It
may be offered to be a woman who is at high risk of
having a baby with a genetic disease. The risk of
miscarriage of amniocentesis is about 0.5% or 1 in
200.

It is the patient’s decision whether the benefits of

procedure out-weigh the risks.
Amniocentesis in HIV Pregnant Women: 16
Years of Experience – A case study
Abstract
The iatrogenic risk of HIV vertical transmission, calculated in initial epidemiologic studies, seemed to
counterindicate invasive prenatal diagnosis (PND) procedures. The implementation of highly active
antiretroviral therapy (HAART) represented a turning point in PND management, owing to a rapid and
effective reduction of maternal viral load (VL). In the present study, we identified cases of vertical
transmission in HIV-infected pregnant women who did amniocentesis in the second trimester of pregnancy
(n = 27), from 1996 to 2011. We divided our sample into Group A—women under HAART when submitted
to amniocentesis (n = 20) and Group B—women without antiretroviral therapy before amniocentesis (n = 7).
We had 1 case of vertical transmission in Group B. Preconceptional or early first trimester HIV serology is
essential to avoid performing an amniocentesis without antiretroviral therapy or viral suppression. When
there is an indication for amniocentesis in an HIV-infected pregnant woman, it should be done if the patient
is on HAART and, if possible, when VL is undetectable. Nowadays, with combined first trimester screening
test to select pregnancies with high risk of aneuploidies, advanced maternal age is a less frequent indication
to perform PND invasive procedures, representing an outstanding gain in prenatal diagnosis of this
population.

Introduction
The widespread use of highly active antiretroviral therapy (HAART) during the last decade has significantly
reduced the rates of HIV mortality and disease progression. The rate of vertical transmission in HIV-infected
pregnant women on HAART is around 1-2%, being almost zero when associated with an elective caesarean
delivery and avoidance of breastfeeding. Simultaneously, there has been an increase in pregnancy rates
among HIV-infected women, raising new problems and issues in prenatal diagnosis (PND), such as those
concerning invasive procedures to diagnose chromosomal abnormalities (amniocentesis and chorionic villus
sampling). The increase in the mean maternal age is a challenge in prenatal diagnosis, particularly in HIV-
infected pregnant women.
In the past, invasive procedures as amniocentesis were generally discouraged in HIV-infected pregnant
women, due to increased risk of vertical transmission. The puncture of the uterine wall or placenta and
lesions of the foetal skin or umbilical cord may all increase the foetal exposure to maternal virus [ 4].
Amniocentesis itself has potential morbidity, such as rupture of membranes, chorioamnionitis, or placental
abruption, with consequent foetal loss or vertical transmission if gestation goes on.
Studies that analyze vertical transmission rates after amniocentesis have been scant, and existing data are
limited. First studies report an increase in vertical transmission after procedures undertaken during the
second or third trimesters of pregnancy. However, during the pre-HAART era, amniocentesis was not
performed in most of the centres, even if there was a medical/obstetrical indication, and therefore these
studies include very small and heterogeneous samples. Since 2003, the reported risk of vertical transmission
has markedly diminished because of the wide spread of antiretroviral therapy. Some centres, thus, started to
offer amniocentesis during second trimester to HIV-infected pregnant women, whenever a strong indication
(genetic or infectious) exists. These studies have reported no cases of vertical transmission after invasive
procedures among HIV-infected women treated with HAART.
According to the British Guidelines, for women who have started HAART but whose viral load is not yet
undetectable, it may be advisable to delay the amniocentesis until the maternal viral load is undetectable if at
all possible. In women not already taking HAART, administration of antiretroviral therapy to cover the
procedure is advised.
The aim of our study was to identify cases of vertical transmission in HIV-infected pregnant women who did
second trimester amniocentesis in our hospital.

Methods
 The researchers analysed amniocentesis (n = 27) performed in our institution from the observational cohort
of HIV-infected pregnant women. The sample was obtained from the database, which included all HIV-
infected pregnant woman who gave birth between 1996 and 2011 (n= 804). All clinical files were reviewed
and data were collected in order to obtain demographic characteristics of the sample, risk factors associated
with HIV infection (such as drug abuse), obstetrical variables such as parity, mode of delivery, obstetrical
complications, indication of amniocentesis and gestational age when it was accomplished, HIV subtype and
transmission category, antiretroviral regimen, viral load close to amniocentesis and close to labour, foetal
cerotype, newborn data such as weight, antiretroviral prophylaxis regimen, and HIV DNA PCR. Our sample
was divided into two subgroups: women under HAART when submitted to amniocentesis (Group A, n =
20) and women without antiretroviral therapy before amniocentesis (Group B, n = 7) (Figure 1). SPSS
Version 17.0 was used to obtain statistical analysis of both groups and to compare differences in
transmission rates among groups. The results were analyzed statistically using Chi-square. A P value below
0.05 was considered to indicate statistical significance.

Figure 1. Summary of the study. HAART: highly active antiretroviral therapy; ARV: antiretroviral therapy.
This study was approved by the Ethical Committee of our hospital.

3. Results
Between 1996 and 2011, amniocentesis was performed in 3.36% of our cohort ().

3.1. Demographics

The mean maternal age of our study group was 37.7 years, and most of them were Caucasian and multifarious
(Table 1). Sexual transmission of HIV was the main way of infection, and HIV 1 was the most frequent subtype (n =
21). In Group A, 2 women had double infection (HIV-1 and HIV-2) and other 2 were HIV-2 infected. The remaining
women were HIV-1 infected. In Group B, 2 women had HIV-2 infection and 5 were HIV 1-infected.
Table 1
Demographic characteristics.

Group A Group B
(/N
Demographics
(/N %) %)
Age (years)    
Mean 37.7     
   <35 4/20% 1/14.3%
   ≥35 16/80% 6/85.7%

Race    

Caucasian  15/75% 4/57.1%

African  4/20% 3/42.9%
Indian  1/5% —
Parity    
Nullifarous  5/25% 2/28.6%

   
Category of transmission

15/75% 7/100%
Sexual 
Substance abuse  4/20% —
Transfusion  1/5% —

   
Subtype of HIV

16/80% 5/71.4%
HIV 1 
HIV 2  2/10% 2/28.6%
Double infection (HIV-1 and HIV-
2/10% —
2) 

3.2. Characteristics of Amniocentesis

Advanced maternal age was the most frequent indication for amniocentesis. Gestational age at the time of
amniocentesis was, in most cases, between 16 and 19 weeks (Table 2). Two cases of chromosomal abnormalities, both
trisomy 21, were diagnosed. These two amniocentesis were performed, one for advanced maternal age and the other
for augmented nuchal translucency.
Table 2
Characteristics of amniocentesis.

Characteristics of Group A Group B

amniocentesis (/N %) (/N %)

Indication
   

Advanced maternal age  15/75% 5/71.4%

Increased NT  2/10% 1/14.3%
CMV Infection  — 1/14.3%
Is immunization Rh  1/5% —
Maternal genetic disease (cystic
1/5% —
fibrosis) 
Malformation  1/5% —

GA (weeks)    

13–15  — 1/14.3%

16–19  4/57.1%
13/65%
19–20  1/5% —

≥20  2/28.6%
6/30%
Chromosomal abnormalities    
Trisomy 21  1/5% 1/14.3%

Viral load at the time of procedure (copies/ (copies/

mL) mL)

 <50
11/55% —
50–999  5/25% —
1000–10000  1/5% —
 >10000 3/15% —
Unknown  — 7/100%

NT: nuchal translucency; GA: gestational age.

In Group A, viral load (VL) at amniocentesis was undetectable in 11 cases (2 of these patients were HIV-2 infected),
whereas in Group B viral load was unknown in all women. However, in 2 women of Group B, who started prenatal
care only after amniocentesis, viral load was quantified subsequently one HIV-2-infected woman had undetectable
viral load 4 weeks after the procedure and the other had 5.790 copies/mL 9 weeks after amniocentesis.

3.3. Pregnancy Outcome

The most frequent obstetrical complications in our study group were fetal growth restriction (FGR), preterm labour,
and chronic hypertension (Table 3).
Table 3
Pregnancy outcome.

Grou
Pregnancy Group A
pB
outcome
(/N %)
(/N %)
Obstetrical complications  

Chronic hypertension  1/5% 1/14.3%

Foetal growth restriction  4/20% —
Gestational diabetes  1/10% 1/14.3%
Preterm labour/PPROM  4/20% —
Hydramnios  2/10% —
Infection (CMV)  — 1/14.3%
Isoimmunization Rh  1/5% 

Mode of delivery  

Vaginal  4/20% 2/28.6%

Instrumental  1/5% —
Elective caesarian  14/70% 5/71.4%
Caesarian during labour  1/5% —

Viral load close to labour (copies/mL)

   
<50 13/65% 3/42.9%
50–999  5/25% 
1000–10000  1/5%  
>100001/5% Unknown  — 4/57.1%

PPROM: preterm premature rupture of membranes; GA: gestational age.

Elective cesarean after 37 weeks was the predominant mode of delivery in both groups. In Group A, VL determined
close to labor was undetectable in 13 women, and in Group B it was undetectable in 4 women, the remaining 3 being
unknown.

Among the 27 newborns, only one case of HIV 1 infection was diagnosed, owing to Group B. It occurred in a patient
with a diagnosis of HIV infection at 30 weeks of gestation (in 1998), who had already done amniocentesis at 16 weeks
for primary CMV maternal infection. Primary care physician sent this patient at 9 weeks of gestation to our
emergency department due to a rash, which was interpreted as an exanthema subitum. She was referred to our PND
center and laboratory tests of CMV, Toxoplasmosis, Rubella, and Parvovirus B19 were requested. Since the patient
had been referred from primary care, HIV serology had already been requested, and according to the patient, with
pending results. Therefore, it was not requested in our tertiary care center. CMV results were positive for primary
infection and the amniocentesis was planned. However, this patient was absent from medical care until 30 weeks,
when she returned with a diagnosis of HIV infection (requested by her primary care physician), presumably due to a
disease denial. Spontaneous labour began at 38 weeks and she had a vaginal delivery. She had done neither
combination antiretroviral drug therapy nor AZT prophylaxis during pregnancy, as well as adequate prenatal care.
AZT intravenous perfusion was done during labour.
Among the 777 women who were not submitted to invasive procedures, there were 4 cases of vertical transmission
(0.5%). When we compared this transmission rate with the transmission rate of 1/27–3.7% (all HIV pregnant women
who underwent amniocentesis), no statistical significant difference was found (4/777 versus 1/27; P value =
0.4083; X^2 -test). Comparing the difference in transmission rate between Groups A and B, we obtained a P value of
0.5756 (0/20 versus 1/7; X^2 -test).

3.4. Antiretroviral Therapy

Antiretroviral drug therapy in pregnancy is described in Figure 2. In Group B, therapy regimen was initiated in the
second half of pregnancy, because of late HIV diagnosis (only after amniocentesis). Monotherapy prophylaxis with
AZT was administrated to three pregnant women in Group B.

Figure 2. Antiretroviral therapy during pregnancy. AZT: zidovudine; 3TC: lamivudine; NRTI: nucleoside reverse
transcriptase inhibitors; NNRTI: nonnucleoside reverse transcriptase inhibitors; PI: protease inhibitors.

4. Discussion
The absence of a control group and the small sample dimension raise some limitations to the statistical analysis in the
present work. This is also the case in studies performed by other researchers.
In the particular case of vertical transmission in Group B, we cannot exclude the possibility of a primary HIV
infection considering the rash as the initial manifestation of the disease. Primary HIV infection is a major risk factor
for vertical transmission. Concomitantly, primary infection for CMV may reveal immunosuppression status of the
patient, also contributing to vertical transmission.

It is important to note that the long time span of our study includes a period when AZT monotherapy was considered
to be the gold standard prophylaxis. Nowadays, regimens of HAART are recommended during pregnancy with two
nucleoside reverse transcriptase inhibitors (NRTIs) and one protease inhibitor (PI) [ 10], allowing to achieve a higher
efficacy and sustainability of viral suppression. According to Watts’ review, the patient should have an optimized
antiretroviral treatment and an undetectable viral load before undergoing an invasive procedure as amniocentesis [ 12].
In addition, Lopez and Coll recommended that transplacental amniocentesis should always be avoided [ 13].
Therefore, it is important to propose a viral load determination before amniocentesis for patients with contamination
risk and to postpone the invasive procedure in cases with positive results in order to obtain an undetectable viral load 

In addition, considering that this study involves a long time spam, it is also important to note that physicians have not
always been sensitized to ask for an HIV test before performing PND invasive procedures. Also, advanced maternal
age was the predominant indication for amniocentesis in our cohort. Nowadays, with combined first trimester
screening test to select pregnancies with high risk of aneuploidies, advanced maternal age is a less frequent indication
to perform PND invasive procedures. Among HIV-infected women this important screening test minimizes invasive
procedures, representing an outstanding gain in prenatal diagnosis of this population.

Somigliana et al., in a multicenter study, showed no difference in transmission rate between mothers who underwent
an invasive procedure and the control group (2/60 versus 12/712). In our study, we obtained an overall transmission
rate of 4/777 (0.5%) in HIV pregnant women without invasive procedures, and a transmission rate of 1/27 (3.7%) in
those who had an amniocentesis (4/777 versus 1/27; value = 0.4083; -test). Therefore, due to the small sample
dimension of our study and to the inadequate power for differences in transmission rates, there is no evidence that
amniocentesis is associated with a higher transmission rate. This is also the case in other published series. Moreover,
when we compared transmission rates between our two Groups A and B, we found no statistical significant
differences (0/20–0% versus 1/7–14.3%; value = 0.5756; X^2 - test).

In our opinion, invasive PND techniques should not be precluded to HIV-infected pregnant women at increased risk
for foetal chromosomopathies. However, it is extremely important to adopt a selective attitude in these situations,
informing and clarifying the patient about the risks related to the procedure. Besides the issue of parental
transmission, it is essential to remember the complications inherent to invasive procedures, such as premature rupture
of membranes, chorioamnionitis, and placental abruption, as previously stated.

Conclusions
Preconceptional or early first trimester HIV screening is essential to avoid amniocentesis without antiretroviral
therapy or viral suppression. When amniocentesis is indicated in HIV-infected pregnant women, it should be done
after initiating a combination antiretroviral drug therapy and ideally when viral load is undetectable. The wide spread
of combined prenatal screening tests has been particularly important in these patients, decreasing the need to perform
invasive PND procedures. Although the size of our sample is limited, there was no case of vertical transmission
among pregnant women with prenatal care, who have done amniocentesis on HAART. It would be extremely
important to analyze wider results, in a multicentric study.
 BIBLIOGRAPHY
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