Discrepancy of Cytogenetic Analysis in Western and Eastern Taiwan Pediatrics and Neonatology (2013) 54, 161e165 Available online at www.sciencedirect.com journal homepage: http://www.pediatr-neonatol.com ORIGINAL ARTICLE Discrepancy of Cytogenetic Analysis in Western and Eastern Taiwan Yu-Hsun Chang a,b, Pui-Yi Chen c, Tzu-Ying Li c, Chung-Nan Yeh c, Yi-Shian Li c, Shao-Yin Chu a,d,e,*, Ming-Liang Lee e a Department of Pediatrics, Buddhist Tzu Chi General Hospital, Hualien, Taiwan b Institute of Medical Science, Tzu Chi University, Hualien, Taiwan c Laboratory of Cytogenetics, Genetic Counseling Center, Buddhist Tzu Chi General Hospital, Hualien, Taiwan d Department of Medical Education, Buddhist Tzu Chi General Hospital, Hualien, Taiwan e School of Medicine, Tzu Chi University, Hualien, Taiwan Received Nov 15, 2011; received in revised form Mar 5, 2012; accepted Mar 15, 2012 Key Words amniocentesis; amniocyte karyotyping; cytogenetic analysis * Corresponding author. 707, Sec. 3, E-mail address: chu_chu@tzuchi.c 1875-9572/$36 Copyright ª 2012, Taiw http://dx.doi.org/10.1016/j.pedneo.2 Objective: This study aimed at investigating the results of second-trimester amniocyte karyo- typing in western and eastern Taiwan, and identifying any regional differences in the preva- lence of fetal chromosomal anomalies. Methods: From 2004 to 2009, pregnant women who underwent amniocentesis in their second trimester at three hospitals in western Taiwan and at four hospitals in eastern Taiwan were included. All the cytogenetic analyses of cultured amniocytes were performed in the cytoge- netics laboratory of the Genetic Counseling Center of Hualien Buddhist Tzu Chi General Hospital. We used the chi-square test, Student t test, and ManneWhitney U test to evaluate the variants of clinical indications, amniocyte karyotyping results, and prevalence and types of chromosomal anomalies in western and eastern Taiwan. Results: During the study period, 3573 samples, 1990 (55.7%) from western Taiwan and 1583 (44.3%) from eastern Taiwan, were collected and analyzed. The main indication for amniocyte karyotyping was advanced maternal age (69.0% in western Taiwan, 67.1% in eastern Taiwan). The detection rates of chromosomal anomalies by amniocyte karyotyping in eastern Taiwan (45/1582, 2.8%) did not differ significantly from that in western Taiwan (42/1989, 2.1%) (p Z 1.58). Mothers who had abnormal ultrasound findings and histories of familial hereditary diseases or chromosomal anomalies had higher detection rates of chromosomal anomalies (9.3% and 7.2%, respectively). The detection rate of autosomal anomalies was higher in eastern Taiwan (93.3% vs. 78.6%, p Z 0.046), but the detection rate of sex-linked chromosomal anom- alies was higher in western Taiwan (21.4% vs. 6.7%, p Z 0.046). Chung-Yang Rd., Hualien, Taiwan 970, ROC. om.tw (S.-Y. Chu). an Pediatric Association. Published by Elsevier Taiwan LLC. All rights reserved. 012.11.013 mailto:chu_chu@tzuchi.com.tw http://dx.doi.org/10.1016/j.pedneo.2012.11.013 www.sciencedirect.com/science/journal/18759572 http://www.pediatr-neonatol.com http://dx.doi.org/10.1016/j.pedneo.2012.11.013 http://dx.doi.org/10.1016/j.pedneo.2012.11.013 162 Y.-H. Chang et al Conclusion: We demonstrated regional differences in second-trimester amniocyte karyotyping results and established a database of common chromosomal anomalies that could be useful for genetic counseling, especially in eastern Taiwan. Copyright ª 2012, Taiwan Pediatric Association. Published by Elsevier Taiwan LLC. All rights reserved. 1. Introduction Second-trimester amniocentesis for amniocyte karyotyping is a common procedure for prenatal diagnosis of chromo- somal anomalies.1 Indications for amniocyte karyotyping are advanced maternal age, abnormal maternal serum screening results, abnormal prenatal ultrasound findings, and known family history of chromosomal anomalies or hereditary diseases.2,3 In recent years, Taiwanese women have married later and become pregnant at older ages. After the implementation of national health insurance, antenatal care usage also increased.4 Therefore, second- trimester amniocentesis for advanced maternal age and abnormal maternal serum screening test are routinely diagnostic recommendations at Taiwan. Karyotyping and risk calculation results not only offer the opportunity for pregnant women to have comprehensive genetic coun- seling, but also significantly decrease the prevalence of newborn babies affected by chromosomal anomalies. We report the analysis of amniocyte karyotyping results of 3573 fetal amniocyte specimens from a cytogenetic laboratory in Taiwan. We investigated the indications for amniocyte karyotyping, prevalence of fetal chromosomal anomalies, and regional differences between western and eastern Taiwan. 2. Materials and Methods 2.1. Sample collection From 2004 to 2009, pregnant women who underwent amniocentesis in their second trimester at three hospitals (Taipei Branch, Taichung Branch, and Dalin Branch of Buddhist Tzu Chi General Hospital) in western Taiwan and at four hospitals (Hualien, Yuli Branch, and Kuanshan Branch of Buddhist Tzu Chi General Hospital, and Menno- nite Christian Hospital) in eastern Taiwan were included. The Ethics Review Board of the hospital reviewed and approved the study protocol. All cytogenetic analyses of cultured amniocytes were performed in the cytogenetics laboratory of the Genetic Counseling Center of Hualien Buddhist Tzu Chi General Hospital. We retrospectively reviewed and analyzed these results. The indications for amniocentesis included advanced maternal age, suspected Down syndrome, suspected trisomy 18, suspected neural tube defect, familial heredi- tary disease or chromosomal anomaly, abnormal ultrasound findings, elective choice, and others. Mothers aged 35 years or more at the time of delivery were defined as having advanced maternal age. Suspected Down syndrome, sus- pected trisomy 18, and suspected neural tube defect were classified according to maternal serum screening results.5e7 Amniocentesis performed due to maternal anxiety or for personal purposes was defined as elective. 2.2. Cytogenetic analysis The high-resolution banding technique for R-bands by BrdU using Giemsa staining was performed using a standard in situ protocol.8,9 Fifteen colonies were analyzed routinely for each karyotype report. 2.3. Statistical analysis For evaluating the demographic data, we obtained the sample size (n), mean, standard deviation (SD), median, and range. We used the chi-square test to evaluate the independence of variances and Student t test to evaluate the mean differences of the variances such as maternal age. We also used the nonparametric ManneWhitney U test to evaluate the differences of the variances without normal distributions in gravidity, spontaneous abortion, and artifi- cial abortion. We used SPSS 16.0 to perform these analyses. 3. Results From 2004 to 2009, we analyzed 3573 amniocyte cultures, 1990 (55.7%) from western Taiwan and 1583 (44.3%) from eastern Taiwan. Two samples failed karyotyping (0.056%) and were excluded from the analysis. Maternal nationality was not significantly different in western and eastern Taiwan (p Z 0.699). Pregnant women who underwent amniocentesis in eastern Taiwan were younger and had greater gravidity, spontaneous abortions, and artificial abortions (Table 1). The main indication for amniocyte karyotyping was advanced maternal age (69.0% in western Taiwan, 67.1% in eastern Taiwan). Suspected Down syndrome was the second indication and was almost equal in both western and eastern Taiwan (15.5% vs. 15.3%). The indications of amniocyte karyotyping, owing to suspected trisomy 18, suspected neural tube defect, familial hereditary disease, or chromosomal anomaly, and abnormal ultrasound findings were more frequent in eastern Taiwan, but advanced maternal age and elective karyotyping were more frequent in western Taiwan (Table 2). Foreign-born mothers had a greater abnormal karyotype rate (7/134, 5.2%) than Taiwan-born mothers (80/3437, 2.3%) (p Z 0.033). Mothers who had elective amniocyte karyotyping had significantly lower abnormal karyotype rates (1/374, 0.3%) than others (86/3197, 2.7%) (p Z 0.004). Mothers younger than 34 years did not have Table 1 Demographic data of pregnant women who received amniocyte karyotyping in western and eastern Taiwan. Western Taiwan Eastern Taiwan Maternal nationality n % n % p Taiwanese 1917 96.3 1521 96.1 0.699 Foreigner 73 3.7 62 3.9 Maternal age Mean SD Mean SD 34.3 3.7 33.8 4.5 <0.001 Pregnancy history Median Range Mean rank Median Range Mean rank Gravidity 2 1e9 1616.5 2 1e11 1931.5 <0.001 Spontaneous abortion 0 0e5 1589.5 0 0e5 1651.3 <0.001 Artificial abortion 0 0e8 1552.6 0 0e9 1683.7 <0.001 n Z sample size; SD Z standard deviation. Amniocyte karyotyping in eastern Taiwan 163 a significantly lower abnormal karyotype rate (39/1399, 2.8%) than those aged 34 years or older (48/2171, 2.2%) (p Z 0.275). The detection rate of chromosomal anomalies by amniocyte karyotyping in eastern Taiwan (45/1582, 2.8%) did not differ significantly from that of western Taiwan (42/1989, 2.1%) (p Z 1.58) (Table 3). Mothers who had abnormal ultrasound findings and histories of familial hereditary disease or chromosomal anomaly had higher detection rates of chromosomal anomalies (9.3% and 7.2%, respectively) (Table 2). The types of chromosomal anoma- lies are listed in Table 4. The detection rate of autosomal anomalies was higher in eastern Taiwan (42/45. 93.3%) than in western Taiwan (33/42, 78.6%) (p Z 0.046). Detection rates of sex-linked chromosomal anomalies were higher in western Taiwan (9/42, 21.4%) than in eastern Taiwan (3/45, 6.7%) (p Z 0.046). De novo abnormal chromosomal detec- tion rate was 8% (7/87). 4. Discussion This study showed differences in amniocyte karyotyping results between western and eastern Taiwan. The Table 2 Indications and detection rate of chromosomal anoma Indications Western Taiwan Eastern Taiwan n % n % Advanced age pregnancy 1373 69.0 1025 64. Suspected Down syndrome 309 15.5 239 15. Suspected trisomy 18 1 0.1 44 2. Suspected neural tube defect 3 0.2 34 2. Familial hereditary disease or chromosomal anomaly 33 1.7 36 2. Abnormal antepartum sonography 47 2.4 50 3. Elective 224 11.3 150 9. Others 0 0.0 5 0. Total 1990 1583 Chi-square test: * p < 0.001, yp Z 0.177. n Z sample size; N/A Z n detection rate of chromosomal anomalies in our center was similar to those of previous studies undertaken in Taiwan.10,11 According to previous studies, newborns of foreign-born mothers had better neonatal outcomes in Taiwan.12,13 Surprisingly, these mothers had higher rates of chromosomal anomalies in our study. However, it is well known that advanced maternal age is a risk factor for chromosomal anomalies. However, we found that it was not associated with higher risk of chromosomal anomalies in our study. Besides the socioeconomic and biological aspects of sample collection, selection bias could also explain the paradox. The first three most common reasons for amniocyte karyotyping were advanced maternal age (57.29e58.85%), abnormal maternal serum screening results (21.34e26.17%), and abnormal ultrasound findings (4.50e8.86%) in previous studies. In our study, more mothers with advanced age underwent amniocyte kar- yotyping both in western (69.0%) and in eastern Taiwan (64.8%), but fewer mothers had abnormal maternal serum screening results and abnormal ultrasound findings in both western (15.8%, 2.4%, respectively) and eastern (20.0%, lies by amniocyte karyotyping for various indications. * Detection rate of chromosomal anomaliesy Total Western Taiwan Eastern Taiwan Total n % n % n % n % 8 2398 67.1 31 2.3 23 2.2 54 2.3 1 548 15.3 5 1.6 9 3.8 14 2.6 8 45 1.3 0 0.0 1 2.3 1 2.2 1 37 1.0 0 0.0 2 5.9 2 5.4 3 69 1.9 3 9.1 2 5.6 5 7.2 2 97 2.7 2 4.3 7 14.0 9 9.3 5 374 10.5 1 0.4 0 0.0 1 0.3 3 5 0.1 0 N/A 1 20.0 1 20.0 3573 42 2.1 45 2.8 87 2.4 ot available. Table 3 Results of amniocyte karyotyping in western and eastern Taiwan. Western Taiwan Eastern Taiwan Total n % n % n % Normal female 781 39.3 601 38.0 1382 38.7 Normal female variant 149 7.5 137 8.7 286 8.0 Normal male 881 44.3 668 42.2 1549 43.4 Normal male variant 136 6.8 131 8.3 267 7.5 Autosomal anomalies 33 1.7 42 2.7 75 2.1 Sex chromosomal anomalies 9 0.5 3 0.2 12 0.3 Total 1989 1582 3571 Chi-square test: p Z 0.048. n Z sample size. 164 Y.-H. Chang et al 3.2%, respectively) Taiwan than those reported in the previous studies. Among women with abnormal serum screening results, abnormal karyotypes were found in 1.6% (5/313) in western and 3.8% (12/317) in eastern Taiwan. The detection rate of chromosomal anomalies by amniocyte karyotyping for women with abnormal ultrasound findings was significantly higher in eastern Taiwan (14.0%) than in Table 4 Types of chromosomal anomalies in western and easte Western Taiwa n % Autosomal anomalies Numerical anomalies Trisomy 21 13 31 Trisomy 18 1 2 Numerical and structural anomalies 1 2 Structural anomalies Translocationdreciprocal 4 9 TranslocationdRobertsonian 3 7 Inversion 1 2 Addition 2 4 Derivative 1 2 Duplication 1 2 Insertion 0 0 Marker 1 2 Mosaicism Numerical 3 7 Structural 2 4 Subtotal* 33 78 Sex chromosomal anomalies Numerical anomalies 45X 1 2 45X mosaicism 3 7 47XXY 3 7 47XYY 0 0 Mosaicism 2 4 Subtotal* 9 21 Total 42 Chi-square test: *p Z 0.046. n Z sample size. western Taiwan (4.3%) and in previously reported data (1e8.86%).10,11 Therefore, detailed antepartum sonography and family history review are important to identify the mothers at risk for chromosomal anomalies. We found that the detection rate for autosomal anom- alies was higher in eastern Taiwan and that of sex chro- mosomal anomalies was higher in western Taiwan, according to second-trimester amniocyte karyotyping. This finding was comparable to the data in the population-based birth registry of Taiwanese newborns in 2002. Advanced maternal age was the most important risk factor of chro- mosomal anomalies.14 Since there was a discrepancy between different areas, other than socioeconomic status, personal habits, environmental factors, and occupational factors, a possible explanation was that there were still pregnant women, especially aboriginal women, at risk of chromosomal anomalies who did not receive antepartum genetic screening examinations in eastern Taiwan.4,15 There were some limitations to this study. We did not have information about some important risk factors such as socioeconomic status, ethnicity, smoking, alcohol consumption, and occupation. Although we included seven hospitals around Taiwan, selection bias could not be avoi- ded completely. rn Taiwan. n Eastern Taiwan Total n % n % .0 16 35.6 29 33.3 .4 3 6.7 4 4.6 .4 2 4.4 3 3.4 .5 7 15.6 11 12.6 .1 1 2.2 4 4.6 .4 8 17.8 9 10.3 .8 1 2.2 3 3.4 .4 2 4.4 3 3.4 .4 0 0.0 1 1.1 .0 1 2.2 1 1.1 .4 0 0.0 1 1.1 .1 0 0.0 3 3.4 .8 1 2.2 3 3.4 .6 42 93.3 75 86.2 .4 0 0.0 1 1.1 .1 1 2.2 4 4.6 .1 1 2.2 4 4.6 .0 1 2.2 1 1.1 .8 0 0.0 2 2.3 .4 3 6.7 12 13.8 45 87 Amniocyte karyotyping in eastern Taiwan 165 5. Conclusion In this 6-year retrospective analysis, we demonstrated regional differences in second-trimester amniocyte kar- yotyping results and established a database of common chromosomal anomalies that could be useful for genetic counseling, especially for women in eastern Taiwan. Acknowledgments The authors thank Professor Jye-Siung Fang, Dr Jiang-Chang Yun and Syue-Jhih Li, and Institutes include; Mennonite Christian Hospital, Yuli Veterans Hospital, Hualien Armed Forces General Hospital, Department of OBS/GYN, Taipei Branch, Taichung Branch, and Yuli Branch, Buddhist Tzu Chi General Hospital, for sending amniocyte samples and for their supervision. References 1. Schonberg SA. Cytogenetic analysis in prenatal diagnosis. West J Med 1993;159:360e5. 2. Wieacker P, Steinhard J. The prenatal diagnosis of genetic diseases. Dtsch Arztebl Int 2010;107:857e62. 3. Kazerouni NN, Currier B, Malm L, Riggle S, Hodgkinson C, Smith S, et al. Triple-marker prenatal screening program for chromosomal defects. Obstet Gynecol 2009;114:50e8. 4. Chen CS, Liu TC, Chen LM. National health insurance and the antenatal care use: a case in Taiwan. Health Policy 2003;64: 99e112. 5. Haddow JE, Palomaki GE, Knight GJ, Williams J, Pulkkinen A, Canick JA, et al. 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Chang YH, Chen PC, Hsieh CJ, Jeng SF, Liao HF, Su YN, et al. Perinatal and infant health outcomes among neonates born to aboriginal parents in Taiwan. Acta Paediatr Taiwan 2007;48: 135e40. Discrepancy of Cytogenetic Analysis in Western and Eastern Taiwan 1. Introduction 2. Materials and Methods 2.1. Sample collection 2.2. Cytogenetic analysis 2.3. Statistical analysis 3. Results 4. Discussion 5. Conclusion Acknowledgments References