| Objective:Through to the maternal and infant patients with hepatitis b serological, genetic testing and epidemiological investigation, explore the relationship between the HBV serum markers, HBV-DNA levels, the gene mutation and mother-to-child transmission intrauterine infection, so as to guide the clinical effective blocking mother-to-child transmission.Methods:1 According to the experiment design standard to collect 374 cases of pregnant women in the sixth hospital of Shijiazhuang and the second hospital of Hebei medical university, signed informed consent, collected related information, filled out the survey table;2 To collect the above to be included in the blood of pregnant women, newborn within 24 h cord blood or venous blood,-20℃ cryopreserved after separating the serum for serological and corresponding quantitative test of HBV-DNA. And after the baby was born in 1 month, 6 months and 1 year, each period blood collected once, then the same process;3 The specimens was collected and saved by the and sent to hebei centers for disease control and prevention under the condition of-80℃ freezing preservation, did the second detection for the unknown specimens of serological and corresponding quantitative test of HBV-DNA;4 Referenced the all known standard of HBV whole gene and gene sequence of each fragment in NCBI, designed and compounded of specificity of the amplification and sequencing primers in the relevant literature;5 The serological and HBV-DNA quantitative results was statistical processed by the SPSS19.0 software, then selected the experimental group and the control group. Count data by chi-square test, measurement data with mean ±standard deviation test, with P<0.05 for the difference, and suggested the statistical significance;6 After extraction of the HBV-DNA, PCR amplification, the purpose segment was selected and sequenced according to the electrophoresis results;7 To analyze the homology of the nucleotide and amino acid after the sequenced sequences were spliced by DNAStar software, built the system treeand analyze HBV gene mutation and characteristics.Result:1 HBs Ag, HBe Ag and HBc Ab test positive in pregnant women were accounted for 48.66%(148/374) in 374 cases of hepatitis b patients, HBs Ag, HBe Ab and HBc Ab test positive were 39.57%(148/374), and the rest were 11.77%(44/374); HBs Ag, HBe Ag and HBc Ab test positive patients with HBV-DNA positive detection rate was 92.8%(169/182), HBs Ag, HBe Ab and HBc Ab test positive was 47.3%(70/148), the two groups of patients with mean lg HBV-DNA were 7.26±1.56 copies/ml and 4.89±0.95 copies/ml. There were significant differences between the two group HBV-DNA positive detection rate, so as the numerical content.375 cases of newborns at birth time HBs Ag positive detection rate was 7.47%(28/375), HBV-DNA positive rate was 2.40%(9/375), 6 cases of newborn were positive for HBs Ag and HBV-DNA, the number is 31 cases of newborn HBV infection, the rate was 8.27%(31/375); HBs Ag positive rate was 4.00%(15/375), HBV-DNA positive rate was 1.07%(4/375) in newborns after 1 month peripheral blood of HBV markers and HBV-DNA reviews, including 2 cases of newborns were positive for HBs Ag and HBV-DNA, therefore the infection rate of newborns in 1 month was 4.53%(17/375); The difference was statistically significant(chi-square=4.36, P<0.05) between of newborns 1 month of birth and at birth rate. The review of baby birth in 6 months peripheral blood HBV markers and HBV-DNA were 14 cases, 4 cases, and there was no evident difference after six months.2 S gene nucleotide homology comparison results showed that the homology between A68, A7, A7B0 and the reference sequence with AF100309 were 99.7%, 99.3%, 99.3% respectively, belong to genotype B, the other specime of group with the reference sequence AF286594 were greater than 98.5%, belong to the same genotype C type; In addition, compared mother with newborns, A7 and A7B0, A41 and A41B0, A31 and A31B0, A72 and A72B0, A90 and A90B0, A135 and A135B0 the homology were respectively 100%, 98.2%, 100%, 99.9%, 99.8%, 99.9%, the child was born after 1 month, 6 months or 1 year, gene had a high homology and no obvious change. Common genetic variants were: A7 and A7B0 common variants 345G'A, 793G'A, 796G'T, 799G'A, A41B0, a89 and A106 common variants 783G'A, 592C'T, A41B0 and A198 common variants 508G'A, A31, A198 and A219 common variants 499T'C, A41B0 and A41 common variants 289A'T, A41 302G'A, 241 A'G, 320C'G, A68 482A'G, 724C'T, A41B0 438T'C, 776A'C, 759G'C. Corresponding amino acid variants are: A7 and A7B0 common variants Y64 C, I213 M, A41B0, a89 and A106 common variants N210 S, A41 S50 G, A68 V110 I, A41B0 S95 L, L208 I, A202 G.3 Pre-S nucleotide homology comparison results were over 96%, common variants included: A41, A31, A135 and A72 common variants 2860G'A caused by amino acid variants Q4 K, A7 and A7B0 common variants 3061C'T caused by amino acid variants P72 L, A7, A41 and A41B0 common variants 3081T'G caused by amino acid variants G79 W, there were also a lot of variants such as A7 3059T'A, A41B0 3062A'C, A7 and A7B0 3092G'A did not. The child was born after 1 month, 6 months or 1 year, gene had a high homology and no obvious change.4 HBV X nucleotide homology comparison results were over 96% between same genotype specimens. Common variants included: A135 variants 2143A'G, A173 and A226 common variants 2146C'T, A31, A135 common variants 2167A'G, A7, A222 and A226 common variants 2182G'T, A31, A72, A173, A198 and A219 common.5 HBV C nucleotide homology comparison results were over 96%. There were no obvious difference between the experimental group and the control group with the standard sequence AF286594 、 AF100309 in homology. Common variants included: 1915G'A, 1936T'C, 2011G'A, 3205T'C, but only one caused amino acid change D61 Y. The child was born after 1 month, 6 months or 1 year, gene had a high homology and no obvious change.6 HBV P nucleotide homology comparison results were over 96% between same genotype specimens. The homology of experimental group and control group with the standard sequence AF286594 were greater than 97%, part reached 100%, but the homology of the AF100309 were less than 93.7%. Common variants included: A106, A198 and A219 common variants 855T'A caused by amino acid variants M573 L, A106 and a129 common variants 876 C'T caused by amino acid variants Y579 I, A41, A41B0, A90 common variants 1092 C'T caused by amino acid variants Y651 H, and A41, A41B0, A90 common variants 1479C'A, A41, A41B0 common variants 1497T'G did not cause the variation of amino acid. the child was born after 1 month, 6 months or 1 year, gene had a high homology and no obvious change.Conclusion:1 The rate HBs Ag, HBe Ag and HBc Ab test positive pregnant women with HBV-DNA positive detection is significantly higher than the HBs Ag, HBe Ab and HBc Ab test positive and others;2 The higher HBV-DNA levels in pregnant women, the greater risk of mother-to-child transmission intrauterine infection.The serum HBs Ag and HBV-DNA carrying case of newborns within 24 h can not be as a standard of judging mother-to-child transmission intrauterine infection.3 It may be the cause of mother-to-child transmission intrauterine infection that 302G'A, 345G'A of HBV S gene lead to the variation of amino acid S50 G, Y64 C.4 There are some mutations in Pre-S, X, C and P genes of mother-to-child transmission intrauterine infection specimens and the control group, the mutation significance is uncertain, still need to do further research.5 Besides the A41 mother-to-child gene homology is 97.2%, there is no HBV gene mutation in a short time. |
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