| Part I:Congenital Cytomegalovirus Infection in Severe Fetal MalformationsBackground:Human cytomegalovirus (CMV) is a frequent cause of intrauterine infection and fetal abnormalities. Congenital CMV infection depends highly on maternal infection status. In developed world, the prevalence of CMV IgG in pregnant women is just 36.9-68.3%. Nearly half of the pregnant women are susceptible to primary CMV infection, making it a significant cause of fetal abnormalities. Estimated 9.9-15.4% of adverse pregnancy outcomes could be attributed to CMV infection. In our previous study, however,98.7% child-bearing women in Jiangsu, China, are CMV IgG seropositive, indicating less chance of getting primary infection during pregnancy. Relevant data based on large sample size are lacking, and the contribution of congenital CMV infection in severe fetal malformations remains elusive in developing countries.Objectives:In this study, we aimed to investigate CMV infection in severe fetal malformations and the maternal infection status, and to determine the role of congenital CMV infection in congenital malformations.Methods:During December 2007 to December 2014,436 fetuses with severe malformations and terminated pregnancy were enrolled in Drum Tower Hospital, Medical School of Nanjing University. All fetuses underwent routine autopsy examinations. Postmortem kidney, lung, liver, skin, heart and placenta specimens, cord blood and amniotic fluid were collected and used to detect CMV infection: transplacental CMV transmission results in viral replication mainly in the renal tubular epithelium, thus CMV DNA was first detected in fetal kidneys, then other tissues by real-time PCR; CMV IgM was determined in cord plasma. CMV IgM, IgG, and IgG avidity index (AI) were measured in maternal plasma by ELISA.Results:A total of 436 singleton pregnant women aged 28.4 ± 4.6 and their 436 fetuses (237 males) at gestational age of 26.8 ± 5.2 weeks were enrolled. CMV DNA was positive in kidneys and other tissues of seven fetuses. Of 238 fetuses with plasma available, one was positive for CMV IgM and its kidney, lung, placenta and cord blood were also positive for CMV DNA. Thus, the confirmed congenital CMV infection rate was 1.60% (7/436) in the cohort in the present study. Histopathological characteristics could be seen in the seven fetuses, suggesting possible CMV infection. Hematoxylin-eosin staining showed intranuclear and intracytoplasmic CMV inclusion bodies in three kidneys, two livers, one lung and one pancreas. Other lesions possibly related to infection were observed, including hemosiderin deposits in the ventricular and periventricular zone, microcalcifications in the kidneys, and diffuse villitis. Immunohistochemistry consistently showed nuclear and cytoplasmic distribution of viral proteins in three kidneys and livers. CMV infection was found in 5 (6.1%) of 82 fetuses with central nervous system anomalies,1 (11.1%) of 9 fetuses with abdominal anomalies,1 (0.59%) of 168 fetuses with multiple congenital malformations, and none of fetuses with other anomalies (177). Of 293 pregnant women with plasma available,279 (95.2%) were CMV IgG positive only,6 (2.1%) were positive for both IgG and IgM, and 8 (2.7%) others were negative for both CMV IgG and IgM. The IgG AI was determined in all 285 (97.3%) CMV IgG positive women. Three (1.0%) women had a low AI (<30%), indicating primary infection; 186 (63.5%) had a medium AI (30-50%); and 96 (32.8%) had a high AI (>50%), indicating latent infection. Of 5 mothers with infected fetuses 1 (20%) was CMV IgG and IgM positive, while 5 (1.7%) of 288 mothers with uninfected fetuses were positive respectively(P= 0.099). Low AI values were more frequent in mothers with CMV infected fetuses than those with uninfected fetuses (20% vs 0.7%, P<0.001).Conclusions:Congenital CMV infection in fetuses with severe congenital malformations is rare. Overall, there is no close association between CMV infection and severe fetal malformations in China. If any, CMV may play a role in central nervous system anomalies. Maternal screening for CMV may have minimal value in identifying fetal malformations in China. Thus, it is unnecessary to routinely perform maternal antibody screening for CMV during pregnancy.Part II:Seroepidemiological Study of Hepatitis B Virus Infection among Children Aged 7 Months-12 Years in Jiangsu ProvinceBackground:The key strategy to control Hepatitis B virus (HBV) infection is universal hepatitis B immunization in infants and preschool children. China has integrated hepatitis B vaccine into the national routine immunization program and provided all neonates with free vaccine since 2002. A national serosurvey in 2006 showed that the hepatitis B surface antigen (HBsAg) prevalence of children aged 1-4 years decreased to 1.0%, however, it was higher than the children in other highly endemic areas, including Taiwan (0.5%) and Singapore (0.3%).Objectives:The present study aimed to estimate the latest HBV seroprevalence and immunity to HBV in children born during January 2002 to May 2014, and to evaluate the impact of routine immunization against HBV in Jiangsu Province, China.Methods:A cross-sectional study was conducted in eight hospitals during October 2014 to December 2014. A total of 3442 children (M:F= 2072:1370) aged 7 months to 12 years (5.5 ± 3.6) were enrolled. All serum specimens were measured for the HBV serologic markers by ELISA and quantitative microparticle enzyme immunoassay. If HBsAg positive, or anti-HBc positive and HBsAg negative, the samples were detected for HBV DNA by real-time PCR assay. For HBV DNA positive samples, S region was amplified by nested-PCR, after which genotypes were analyzed and sequences were determined to identify HBV vaccine escape mutants.Results:The prevalence of HBsAg was 0.35%(12/3442,95% CI:0.19-0.63%) among children aged 7 months to 12 years; varied in different age groups,0.16%, 0.36%,0.13% and 1.00% in 7 months-3 years,>3-6,>6-9, and>9-12 years (P= 0.031), respectively. No significant differences were observed between males and females (0.24% vs 0.51%, P= 0.240) or urban and rural areas (0.35% vs 0.35%, P= 1.000). The portion of anti-HBc positive and HBsAg negative, indicating acute self-resolved infection, was 0.99% (34/3442,95% CI:0.70-1.39%). Different incidence between males (0.87%) and females (1.17%) was not statistically significant (P= 0.385). Self-resolved infection was 1.10%,0.73%,1.20% and 0.84% {P= 0.752) in the four age groups, respectively, indicating no increase infection rate by age in the cohort. Of the 3442 children,2542 (73.85%,95% CI:72.34-75.31%) were positive for anti-HBs (≥10mIU/ml), and 535 (15.54%,95% CI:14.35-16.80%) were between 2-9.9 mIU/ml. Overall, immunity to HBV was demonstrated in 3077 (89.39%,95% CI:88.30-90.39%) children of the cohort. Geometric mean concentrations (GMC) of anti-HBs by age were 94.8,40.7,31.7, and 25.8mIU/ml (P = 0.001), respectively. GMC in children aged 7 months to 3 years were higher than those who aged>3-6 years (P= 0.053),>6-9 years (P= 0.007), and>9-12 years (P = 0.003). The positivity of anti-HBs was higher in the urban areas than rural (78.2% vs 71.7%, P <0.001) and similar between male and female (74.0% vs 73.6%, P= 0.765). HBV DNA level was 6.2 log IU/ml (3.3-8.1 log IU/ml) among the 12 HBsAg-positive children, of whom 8 (66.7%) were genotype C and 4 (33.3%) were genotype B. No mutation was detected in S gene sequence. HBV DNA was undetermined in all the 34 children with positive anti-HBc and negative HBsAg, indicating no occult HBV infection.Conclusions:Since hepatitis B vaccine was integrated into routine immunization program in Jiangsu Province, HBsAg prevalence among children decreases significantly to a similar level to that in Taiwan and Singapore; the majority of children have immunity to HBV, indicating hepatitis B vaccination has had a dramatic impact on preventing HBV infection. There is no increased risk of chronic infection and acute self-resolved infection with the increasing age. No mutation of S gene is associated with vaccine failure in the cohort. |
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