[Deficiencies in diagnosing HIV-infection during pregnancy in Brazil, 1998]. / Falhas na identificação da infecção pelo HIV durante a gravidez em São Paulo, SP, 1998.

OBJECTIVE: The increasing number of women with AIDS in the state of São Paulo has lead to the implementation of a series of measures to reduce mother-to-child HIV transmission. The objective of this study was to evaluate these measures' deficiencies regarding coverage and quality of prenatal care in some HIV reference services in the state of São Paulo. METHODS: All HIV-positive women, aged 18 years or more, who gave birth in 1998 were interviewed when they came for a visit with an infectologist or a pediatrician in three cities (São Paulo, Santos and São José do Rio Preto) of the state of São Paulo. A structured questionnaire was applied. Prenatal care and time of their HIV infection diagnosis (before, during or after pregnancy) were assessed. RESULTS: Of 116 women interviewed, 109/116 (94%) had attended a prenatal care service during pregnancy, 64% had their first visit in the first trimester and 80% had 3 or more visits during pregnancy. The mean age of those who attended a prenatal service was 29.1 years, higher than those who did not attend any service (24.3 years). The HIV-positive status was known by 45%, 38% and 17% of the women before pregnancy, during pregnancy and after delivery, respectively. HIV testing was offered to 82% who did not know their serologic status, and among these, only 56% were informed about the importance of getting tested. The basic health care units (UBS) were less efficient in conveying information to the mothers about their children's infection risk (p=0.037) and their treatment needs (p=0.014). CONCLUSIONS: The main deficiencies identified were lack of HIV testing during pregnancy and inadequate information. Though basic health care units are the most important source of care for this population, its contribution to the understanding of risks and treatment needs was the most unsatisfactory.

Tandem duplication. A novel type of triplet repeat instability.

Triplet repeat sequence (TRS) inserts containing (CTG.CAG)(n) (17-175 units in length) were tandemly duplicated when propagated in plasmids in Escherichia coli. The products of this novel type of TRS genetic instability are tracts of as many as 34 multiple units, which contain the entire TRS as well as 129 base pairs of nonrepetitive flanking sequence. The duplication process required the presence of two or more TRS-containing units. Close proximity (170 base pairs) of the TRS to the R6K gamma origin of replication of the pUTminiTn5Cm-derived constructs stimulated the tandem duplication process. These events are proposed to occur due to secondary structure formation, stalling of DNA synthesis, and slippage-mediated misalignment of the complementary strands relative to each other during DNA replication. This mechanism may account for the TRS-associated duplications in protein kinase and metalloprotease genes in neuroblastomas and melanomas, as well as the massive repeat expansions in type II triplet repeat neurological diseases.

DNA polymerase III proofreading mutants enhance the expansion and deletion of triplet repeat sequences in Escherichia coli.

The influence of mutations in the 3' to 5' exonucleolytic proofreading epsilon-subunit of Escherichia coli DNA polymerase III on the genetic instabilities of the CGG.CCG and the CTG.CAG repeats that cause human hereditary neurological diseases was investigated. The dnaQ49(ts) and the mutD5 mutations destabilize the CGG.CCG repeats. The distributions of the deletion products indicate that slipped structures containing a small number of repeats in the loop mediate the deletion process. The CTG.CAG repeats were destabilized by the dnaQ49(ts) mutation by a process mediated by long hairpin loop structures (>/=5 repeats). The mutD5 mutator strain stabilized the (CTG.CAG)(175) tract, which contained two interruptions. Since the mutD5 mutator strain has a saturated mismatch repair system, the stabilization is probably an indirect effect of the nonfunctional mismatch repair system in these strains. Shorter uninterrupted tracts expand readily in the mutD5 strain, presumably due to the greater stability of long CTG.CAG tracts (>100 repeats) in this strain. When parallel studies were conducted in minimal medium, where the mutD5 strain is defective in exonucleolytic proofreading but has a functional MMR system, both CTG.CAG and CGG.CCG repeats were destabilized, showing that the proofreading activity is essential for maintaining the integrity of TRS tracts. Thus, we conclude that the expansion and deletion of triplet repeats are enhanced by mutations that reduce the fidelity of replication.

Small slipped register genetic instabilities in Escherichia coli in triplet repeat sequences associated with hereditary neurological diseases.

Genetic instability investigations on three triplet repeat sequences (TRS) involved in human hereditary neurological diseases (CTG.CAG, CGG.CCG, and GAA.TTC) revealed a high frequency of small expansions or deletions in 3-base pair registers in Escherichia coli. The presence of G to A polymorphisms in the CTG.CAG sequences served as reporters for the size and location of these instabilities. For the other two repeat sequences, length determinations confirmed the conclusions found for CTG.CAG. These studies were conducted in strains deficient in methyl-directed mismatch repair or nucleotide excision repair in order to investigate the involvement of these postreplicative processes in the genetic instabilities of these TRS. The observation that small and large instabilities for (CTG.CAG)175 fall into distinct size classes (1-8 repeats and approximate multiples of 41 repeats, respectively) leads to the conclusion that more than one DNA instability process is involved. The slippage of the complementary strands of the TRS is probably responsible for the small deletions and expansions in methyl-directed mismatch repair-deficient and nucleotide excision repair-deficient cells. A model is proposed to explain the observed instabilities via strand misalignment, incision, or excision, followed by DNA synthesis and ligation. This slippage-repair mechanism may be responsible for the small expansions in type 1 hereditary neurological diseases involving polyglutamine expansions. Furthermore, these observations may relate to the high frequency of small deletions versus a lower frequency of large instabilities observed in lymphoblastoid cells from myotonic dystrophy patients.

The radiosensitivity of human malignant melanomas evaluated by cytokinesis-block micronucleus assay.

Cytokinesis-block micronucleus assay (CB-MNA) was applied for comparison of radiation sensitivity of 25 human malignant melanomas in primary culture. Cells obtained from tumor specimens were irradiated (0-4.Gy) on dishes, incubated with cytochalasin B (2 micrograms/ml) to block cytokinesis, stained in situ and micronuclei (MN) scored in binucleate cells (BNC). Proportions of BNC in nonirradiated controls after fixed time of incubation (96 h) ranged from 2.3 to 38% indicating great differences (C.V. = 74%) in proliferative activity among tumors evaluated. No correlation was observed between proliferative activity and susceptibility of cells to induction of MN by radiation. The great inter-tumor heterogeneity was observed in respect of radiation sensitivity expressed either as normalized (Net) frequency (Fq) of BNC with MN or as number of MN per BNC. Both endpoints differed widely at 2 Gy and 4 Gy as well (Net FqBNC with MN = 0.28-25.4% or 1.5-45% and MN/BNC = 0.004-0.309 or 0.013-0.593 respectively at 2 Gy and 4 Gy) with coefficients of variation ranging from 44 to 57%. Extreme difference in MN frequency was also observed between one primary tumor and its metastasis indicating intra-tumor heterogeneity. Our results suggest that CB-MNA may contribute some clinically useful information for discriminating tumors that will eventually respond to radiotherapy and those that will probably not. However, studies aimed at comparison of MN induction in vitro with clinical radioresponsiveness of malignant melanomas are urgently required.