Prenatal Diagnosis of Cystic Fibrosis by Celocentesis

Abstract

Celocentesis is a new sampling tool for prenatal diagnosis available from 7 weeks in case of couples at risk for genetic diseases. In this study, we reported the feasibility of earlier prenatal diagnosis by celocentesis in four cases of cystic fibrosis and one case of cystic fibrosis and β-thalassemia co-inherited in the same fetus. Celomic fluids were aspired from the celomic cavity between 8⁺² and 9⁺³ weeks of gestation and fetal cells were picked up by micromanipulator. Maternal DNA contamination was tested and target regions of fetal DNA containing parental pathogenetic variants of CFTR and HBB genes were amplified and sequenced. Four of the five fetuses resulted as being affected by cystic fibrosis and, in all cases, the women decided to interrupt the pregnancy. In the other case, the fetus presented a healthy carrier of cystic fibrosis. The results were confirmed in three cases on placental tissue. In one case, no abortive tissue was obtained. In the last case, the woman refused the prenatal diagnosis to confirm the celocentesis data; the pregnancy is ongoing without complications. This procedure provides prenatal diagnosis of monogenic diseases at least four weeks earlier than traditional procedures, reducing the anxiety of patients and providing the option for medical termination of the affected fetus at 8-10 weeks of gestation, which is less traumatic and safer than surgical termination in the second trimester.

Keywords: celocentesis; celomatic fluid; cystic fibrosis; fetal cells; prenatal diagnosis; recessive monogenic diseases; β-Thalassemia.

Figure 1

Workflow for analysis of celomatic fluid. Forty fetal cells are isolated by a micromanipulator and equally distributed in two 0.2 mL Eppendorf tubes. Fetal cells are lysed by alkaline procedure. Initially, a multiplex Polymerase chain reaction (PCR) using CFTR, HBB and STR primers is performed. Subsequently, specific nested PCR for CFTR and HBB target regions and STR are separately performed. Sequencing and SnapShot analysis are performed to evaluate the presence of parental mutations. Fragment analysis is used to evaluate the presence of maternal contamination.