Amniocentesis

Amniocentesis, commonly known as an amniotic fluid test, is a medical procedure used to diagnose chromosomal abnormalities and fetal infections during pregnancy, as well as to determine sex. A little quantity of amniotic fluid containing fetal tissues is collected from the amniotic sac surrounding a growing fetus during this operation. Following that, the fetal DNA is analyzed for genetic abnormalities.

The most frequent cause for an amniocentesis is to see if a fetus has specific genetic disorders or chromosomal abnormalities, such as Down syndrome. Amniocentesis or another procedure known as chorionic villus sampling (CVS) can be used to diagnose certain issues during pregnancy. These prenatal screenings can be beneficial to expectant parents since they allow them to assess the fetal health status and the feasibility of therapy.

When a woman is between 15 and 20 weeks pregnant, she undergoes an amniocentesis. Women who decide to have this test are generally those who are at greater risk for genetic and chromosomal disorders, in part because the procedure is invasive and entails a slight risk of miscarriage. Because this procedure can be used for prenatal sex determination, it is illegal in several countries.

Physiology of amniotic Fluid

In the first trimester of pregnancy, maternal and fetal compartments are critical for the production of amniotic fluid. In the early stages of pregnancy, fetal skin is not keratinized, allowing easy passage of water and other tiny molecules and solutes through the amnion and chorion. Amniotic fluid, like maternal and fetal extracellular fluid, serves as a nonsterile aqueous electrolyte solution.

During the second trimester of pregnancy, the diffusion mechanism stops when fetal skin keratinizes, making it impervious to water and other solutes. Transvaginal ultrasonography detects urine in the embryonic bladder at 9 weeks of gestation, while transabdominal sonography detects urine at 11 weeks. Fetal urine is the main component of amniotic fluid during this period. It is hypotonic (80–140 mOsm/litre) and contains higher quantities of urea, uric acid, and creatinine as the fetal kidneys grow. A fetus at term excretes 500 to 700 mL of urine every day.

At 12 weeks of gestation, the average amniotic fluid volume is 60 ml. The average volume at 16 weeks is 175 ml. The amount of amniotic fluid changes after 20 weeks. The amount of amniotic fluid grows continuously during pregnancy, reaching a peak of 400–1200 ml around 34–38 weeks. In the third trimester, the net rise in amniotic fluid is just 5–10 mL/day. After 38 weeks, fluid volume decreases by around 125 ml/week, reaching an average amount of 800 ml at 40 weeks.

Amniocentesis vs Chorionic Villus Sampling(CVS)

Prenatal diagnostics allows for the identification of a wide range of chromosomal abnormalities, gene disorders, X-linked diseases, neural tube anomalies, and infections before the fetus is born. Amniocentesis, chorionic villus sampling, and fetal blood sampling or cordocentesis are examples of invasive prenatal diagnostic techniques.

Amniocentesis:

Amniocentesis is an invasive procedure. A needle is used to remove amniotic fluid from the uterine cavity in this method. A qualified obstetrician performs this surgery transabdominally and under ultrasound supervision. This was done for the first time in 1956 by Fuchs and Riis for the identification of genetic disorders in the fetus.

It is carried out for both diagnostic and therapeutic objectives. It is used for diagnostic purposes, such as chromosomal, biochemical, histopathological, and microbiological analysis. This is done as a therapeutic technique in individuals with polyhydramnios to lower the amount of amniotic fluid.

Fetal exfoliated cells, transudates, urine, and other secretions are found in amniotic fluid. It is possible to perform it between 15 and 20 weeks of pregnancy. Depending on the weeks of gestation, amniocentesis can be performed in the early or late weeks of pregnancy. Early amniocentesis is conducted between 11 and 14 weeks of pregnancy.

Due to the lack of membrane fusion, there is a higher risk of procedure-related problems than with other invasive treatments. The operation is more likely to be conducted in the latter weeks of pregnancy since the cell culture failure rate is greater when performed earlier. Amnionic fluid leakage, fetal death, and talipes equinovarus are all more likely. The couple is counseled on the procedure's indications, risks, advantages, and limits.

Chorionic Villus Sampling(CVS):

It is a prenatal invasive procedure. It is done under ultrasound guidance. In this procedure, ultrasonography is used to guide the catheter or needle into the chorion frondosum. It is done abdominally and is followed by aspiration of tissue (chorionic villi) for genetic or chromosomal analysis with a syringe containing tissue culture media. It is done in the first trimester for prenatal diagnosis between 10-13 weeks.

Depending on the position of the uterus and bladder, gestational age of the patient, and placental localization it can be performed transabdominally or transcervically. Safer and early termination of pregnancy is possible as karyotype results are available within 7-10 days.

It is indicated in chromosomal and genetic disorders. The samples collected are sent for DNA analysis. It is not performed in vaginal bleeding, in cases of cervical abnormalities, and severe infections. The major complications involved in this procedure are limb reduction defects, the presence of chromosomal abnormalities present in the extraembryonic tissue which are not found in the fetal tissue, intrauterine infections, membrane rupture, and fetal loss.

Indications of Amniocentesis

Diagnostic indications:

It is used for chromosomal analysis: Karyotyping and DNA analysis (to diagnose Sex-linked disorders, inborn errors of metabolism, and neural tube defects).
It is done in cases of advanced maternal age (Age > 35years), abnormal biochemical screening markers (maternal alpha-fetoprotein, human chorionic gonadotropin, unconjugated estriol) in 1st or 2nd trimester, ultrasound detection of an abnormality or soft tissue markers (nuchal translucency, nasal bone hypoplasia, nuchal pad edema), family or personal history of chromosomal abnormalities in previous pregnancies, abnormal parental karyotype.
Diagnosis of fetal infections TORCH infections such as CMV parvovirus Toxoplasma Gondii.
Assessment of severity of Rh isoimmunisation: To assess bilirubin levels in amniotic fluid and to assess the grade of severity of alloimmunization.
Diagnosis of chorioamnionitis.
Biochemical analysis for alpha protein level and acetylcholinesterase level estimation.

Therapeutic indications:

Therapeutic role in relieving maternal discomfort and instillation of intra-amniotic drugs.
Decompression amniocentesis in twin oligohydramnios-polyhydramnios sequence (TOPS): as it decreases the volume of amniotic fluid in the polyhydramnios sac, amniotic fluid pressures are decreased in both the sacs resulting in increased placental thickness thus improving uteroplacental circulation and improving fetal outcome.
Amnioinfusion in oligohydramnios to prevent fetal lung hypoplasia, and umbilical cord compression during labor.
Fetal blood transfusion in fetuses having severe hemolysis.

Contraindications of Amniocentesis

Hepatitis B and HIV infections can be transmitted from maternal circulation to the fetus during the procedure.
Decreased amniotic fluid (oligohydramnios).
Oral anticoagulation therapy must be stopped 48-72 hours before the procedure and patients may be shifted to low molecular weight heparin.

Equipments used in amniocentesis

The procedure is done under continuous ultrasound guidance. The following equipments are required:

Ultrasonography machine
Sterile swabs and drapes
Syringe 2 ml, 10 ml
Needle 20 gauge to 22 gauge
Containers for collection and sample transport
5% Povidine iodine solution

Precautions before undergoing amniocentesis

Couples should undergo genetic counselling.
Written consent should be taken.
The risks, advantages, indications, treatments, and difficulties related to the mother and fetus must be well discussed to the couple. The procedure's fetal and maternal hazards should also be thoroughly discussed.
The number of days necessary to acquire results, failure to culture cells, and the type of cytogenetic test done on the given sample should all be described.
Anti-D must be given to Rh-negative women.
The technique should be properly documented.
Prior to the surgery, ultrasound is performed to document the number of fetuses, viability of the fetus, placental position, gestational age, site of cord insertion, and any noticeable fetal abnormality.
Both the operator and assistant must scrub with antiseptic and use sterile gloves. The exposed abdominal surface area must be cleaned with povidine iodine with sterile gauze and antiseptic solutions.
A sterile plastic cover must be placed over the ultrasound probe. The gel should be retained on the inner surface since it aids in the transmission of ultrasonic waves. To reduce the danger and spread of contamination, sterile gel should be utilized.

Technique of amniocentesis

A detailed ultrasound is performed after adequate genetic counseling and informed written consent to determine gestational age, placental location, gross congenital abnormalities, maximum vertical pocket (MVP), fetal position, fetal movements, and amniotic fluid volume.

Povidine iodine is used to prepare the skin, then sterile ultrasonography gel is applied. After validating the criteria and completing the preparation, amniocentesis is performed aseptically. The procedure is carried out with the use of ultrasonography.

A 20-gauge to 22-gauge spinal needle is used to enter the amniotic cavity under continuous ultrasound guidance. The needle is directed under proper visualization in the clear region of amniotic fluid. It must be ensured that fetal parts, umbilical cord or placenta are not present in the region of needle insertion.

Transplacental puncture is often avoided in cases of alloimmunization or infections to the mother, such as human immunodeficiency virus (HIV), hepatitis B virus (HBV), or hepatitis C virus (HCV) (HCV).

To avoid tenting of the amniotic membrane, a firm entry into the amniotic cavity is advised. Amniotic fluid is progressively aspirated when entrance into the cavity is verified. The first 1 ml to 2 ml of amniotic fluid is removed since it contains the most maternal cells. For karyotype testing, approximately 18 ml to 20 ml of amniotic fluid is required, and 2 ml to 5 ml is required for enzyme deficiency testing. After sufficient amniotic fluid has been gathered, the needle is withdrawn.

Complications of amniocentesis

Both maternal and fetal complications can occur with amniocentesis.

Maternal Complications:

An estimated 2.6% risk of fetomaternal hemorrhage.
Maternal isoimmunization in Rh-negative cases.
There is minimal chance of the introduction of skin bacteria into the amniotic cavity. The risk of chorioamnionitis and uterine infections is less than 0.1%.
The procedure increases the risk of preterm, preterm premature rupture of membrane, and oligohydramnios.
There is a 2% to 3% risk of vaginal bleeding.
Post-procedure pain and maternal discomfort: Mean pain intensity described is 1.6+/-1.3 when noted on a scale of 0-7.
Amniotic fluid embolism.
Hematoma over maternal skin, intestinal or internal organ injures.

Fetal Complications:

The fetal loss rate associated with amniocentesis on average is 0.11%. The loss is 0.56% within 28 days, 0.09% within 42 days.
Amniotic fluid leak: 1% to 2%, and usually associated with spontaneous sealing of membranes. It may also result in:
Fetal lung hypoplasia
Respiratory distress
Fetal injuries like bleeding from cord, ocular injuries, postural deformities like talipes equinovarus (clubfoot) might occur.

When more than three pricks are used to extract amniotic fluid, the risk of complications increases. If an appropriate fluid sample is not collected in two pricks, the process should be abandoned for 24 hours, following which it can be re-attempted. In experienced hands, people performing more than 300 procedures/year; the risk is less.

The risk of fetal loss is higher in women who are otherwise at a higher risk of miscarriage, such as women carrying fetuses with congenital malformations, fibroids, retroplacental hematoma, morbidly obese women, women with active vaginal infection at the time of the procedure. Amniocentesis was safe and without sequelae in up to 86.0 60 % of patients.

Clinical Significance of Amniocentesis

Following the amniocentesis operation, the amniotic fluid sample will be sent to a laboratory for testing. Once the amniotic fluid has been obtained, it is sent for a standard cell culture report, which is received in 14 days. There are quick chromosomal preparations available that yield results in 1 to 2 days, such as fluorescent in-situ hybridization (FISH) and quantitative fluorescence polymerase chain reaction (QFPCR) (QF-PCR).

The procedure is relatively safe, with fewer complications amongst experienced hands. The location of the placenta is an important factor for amniocentesis. While performing the procedure, one should try to avoid penetration of the placenta.

The anterior and fundal placenta is related with a larger number of complications, such as multiple pricks and blood-stained liquor, but not with an increase in fetal loss rates. Passing the needle through the placenta is related with a small increase in premature birth rates.

There are two different types of tests:

A rapid test: The rapid test checks for chromosomal abnormalities. the rapid test can detect a variety of chromosomal disorders that cause physical and mental abnormalities. This test has a near-perfect accuracy rate. It can detect many chromosomal abnormalities such as Down's syndrome (trisomy 21), Edward's syndrome (trisomy 18), Patau's syndrome (trisomy 13).
A full karyotype: The cells in an amniotic fluid sample are cultured in a laboratory for up to 10 days before being examined under a microscope to determine the number of chromosomes and the appearance of chromosomes. The results of a comprehensive karyotype are normally ready in two to three weeks. Following amniocentesis, the amniotic fluid sample is sent to a lab for examination.

Results usually take 10 days to 3 weeks depending upon the laboratory. In the laboratory, genetic and chemical tests are done. For genetic tests, certain chromosomes and genes are analyzed. For chemical tests, proteins, minerals, and other compounds in the amniotic fluid are analyzed. Amniocentesis results will either be positive or negative.

There is no treatment for the majority of chromosomal disorders, thus the couple should be adequately informed over the continuation of pregnancy.

Conclusion

Amniocentesis is the aspiration of amniotic fluid from the amniotic cavity and is the most common invasive fetal testing procedure. It is usually performed for fetal aneuploidy testing. Amniocentesis is not offered to all pregnant women. It's only offered if there's a higher chance that the fetus could have a genetic condition.

Amniocentesis is a relatively safe and reliable procedure. However, there probably is a slightly increased risk of fetal loss following amniocentesis (approximately 0.5%). Other risks are minimal.

Before the patient undergoes amniocentesis, both couples should undergo genetic counselling, written consent should be taken ,the risks, advantages, indications, treatments, and difficulties related to the mother and fetus must be well discussed to the couple as well as the procedure's fetal and maternal hazards should also be thoroughly discussed.

Obstetrician-gynecologists should do amniocentesis if they are familiar with both the indications for the procedure of second-trimester genetic amniocentesis. Recent socioeconomic trends, such as the greater availability of medical knowledge to the general public and many women's desire to postpone childbirth, will raise public demand for prenatal diagnosis. Obstetrician-gynecologists must be prepared to satisfy these needs.

It is used for both diagnostic and therapeutic purposes. It is used as a prenatal screening test for chromosomal abnormalities such as Down syndrome (trisomy 21), Edward's syndrome (trisomy 18), Patau's syndrome (trisomy 13), and other genetic conditions such as sickle cell disease, thalassaemia, cystic fibrosis, or muscular dystrophy.

The amniotic fluid comprises cells shed by the fetus, as well as enzymes, proteins, hormones, and other chemicals. These cells include genetic information that can be utilized to identify genetic abnormalities as well as open neural tube defects like spina bifida. Based on the family history, testing for inherited gene abnormalities and metabolic diseases may also be performed.