[Variability of human cytomegalovirus UL144 gene in low-passage clinical isolates analyzed by HMA-SSCP].

BACKGROUND: Human cytomegalovirus (HCMV) infection is an important infectious agent that results in neonatal disease and congenital deformity. HCMV infection may affect in many organs. The different symptoms and tissue tropism of HCMV infection perhaps resulted from the genetic polymorphism of HCMV. HCMV UL144 open reading frames encode a homologue of the tumor necrosis factor receptor. It seems important to study the strain-specific variability of UL144 sequence in low-passage clinical isolates and to discuss if the variability related to the clinical HCMV infection. METHODS: HCMV-UL144 gene was amplified by PCR assay in 65 low-passage clinical isolates and urine from 7 healthy children who were HCMV-DNA positive by quantitative PCR. All the positive PCR products were analyzed by Heteroduplex mobility assay and single-stranded conformation polymorphism (HMA-SSCP) and 32 of them were sequenced. RESULTS: Fifty-five isolates and 5 urine specimens were HCMV-UL144 positive by PCR. Sequencing and HMA-SSCP analysis showed that significant strain-specific variability was present in the UL144 ORFs. Comparing UL144 sequences and the corresponding symptoms showed that genotype 2 did not exist in megacolon isolates. And genotype 1 and 3 were the major types among microcephaly and jaundice isolates respectively. CONCLUSION: HCMV-UL144 existed in almost all the low passage isolates. HMA-SSCP assay is an easy and effective method to detect the genetype of HCMV-UL144 sequence. The characteristic of sequences in different isolates showed that UL144 gene may play an important role in HCMV infection.

[Polymorphisms of HCMV US28 gene of the clinical isolates from children].

BACKGROUND: To study the polymorphism of human cytomegalovirus US28 gene in children and investigate the relationship between the polymorphism and pathogenesis. METHODS: The FQ-PCR was carried out to determine the DNA quantity of clinical isolate and then the segmental PCR and HMA-SSCP were performed to test the mutation of US28 gene. The typical isolates from different diseases were selected to clone and sequence, then the results were analyzed. RESULTS: The nucleic acid mutation is frequent among the sequence of US28, those mutations focus on the two ends of US28, but most of them are sense mutation. The important functional groups of US28 are highly conserved. The amino acid mutation of some isolates resulted in the change of secondary structure, but the phylogenetic tree analysis did not show any clear association between the pathogenesis and the distribution of clinical isolates. The comparison of US28 sequences from AIDS patients with the sequences from children in our study showed that both sequences have their own specific high mutation points. CONCLUSION: There is polymorphism among the HCMV-US28 gene of clinical isolates from children. There observed no clear relationship was between the pathogenesis and the distribution of clinical isolates.

Sequence variability of human cytomegalovirus UL144 open reading frame in low-passage clinical isolates.

OBJECTIVE: To explore the relationship between human cytomegalovirus (HCMV) UL144 sequence variability and clinical disease. METHODS: HCMV UL144 open reading frame (ORF) was amplified by PCR assay in 72 low-passage isolates [65 congenitally infective children and 7 healthy children who were HCMV-DNA positive by quantitative PCR (qPCR)]. All positive PCR products were analyzed by heteroduplex mobility assay and single-stranded conformation polymorphism (HMA-SSCP) and 32 of them were sequenced. RESULTS: Fifty-five patient isolates and five healthy children isolates were HCMV-UL144 positive by PCR. Sequencing and HMA-SSCP analysis showed that significant strain-specific variability was present in the UL144 ORF. Phylogenetic analysis indicated that the nucleotide sequences could be separated into 3 major genotypes. Comparing between UL144 sequences and the corresponding symptoms showed that genotype 2 did not exist in megacolon isolates. And genotype 1 and 3 were the major types among microcephaly and jaundice isolates respectively. CONCLUSIONS: HCMV-UL144 existed in most of low passage isolates and sequences were hypervariable. The UL144 ORF and its predicted product with the high level of sequence variability in different kinds of isolates suggest that UL144 ORF might play a role in HCMV infectivity and subsequent diseases.

[Effect of toxoplasma infection during pregnancy to the development of fetus and infant].

OBJECTIVE: To investigate the effect of Toxoplasma (Tox) infection during pregnancy to the development of fetus and infant. METHODS: Enzyme linked immunoadsorbent assay (ELISA) was employed to detect Tox-IgM in peripheral blood of 3,908 pregnant women. To Tox-IgM positive cases, the umbilical blood of their neonates were sampled to detect Tox-IgM and abortion tissues were sampled to detect Tox-DNA with polymerase chain reaction. Infection group included 95 cases whose Tox-IgM or DNA were positive, and control group consisted of 119 cases without infection. Different supervision methods were used to periodically observe the intelligence development of infants 3 and 12 months after birth in these two groups. After initial intervention, verbal intelligence quotients (VIQs), performance intelligence quotients (PIQs) and full scale intelligence quotients (FSIQs) were assessed when the infants were 1 and 4 years old of these two groups. RESULTS: The incidences of abortion, stillbirth, pre-term delivery, FGR and malformation in infection group were 12 cases (12.6%), 5 cases (5.3%), 4 cases (4.2%), 4 cases (4.2%) and 3 cases (3.2%), while the incidences of these abnormalities in control group were 3 cases (2.7%), 1 cases (0.9%), 2 cases (1.8%), 2 cases (1.8%) and 1 cases (0.9%). Comparing with these two groups, there was significant difference (P < 0.01). Relative risk (RR) of these 5 abnormalities was 4.7, 5.9, 2.3, 2.3 and 3.6 respectively. Mental development index (MDI) and psychomotor development index (PDI) of infants in infection group were 93 +/- 13 and 101 +/- 16. They were significantly lower than that of control group, which were 107 +/- 17 and 111 +/- 13. There was significant difference between these two groups (P < 0.05). After intervention in 1 and 4 years, their VIQs, PIQs and FSIQs were apparently improved (P < 0.05). CONCLUSION: Congenital Tox infection is closely correlated with abnormal pregnancy outcomes and infantile mental retardation. Early intervention can improve intelligence development.