Limited protection from a pathogenic chimeric simian-human immunodeficiency virus challenge following immunization with attenuated simian immunodeficiency virus.

Two live attenuated single-deletion mutant simian immunodeficiency virus (SIV) constructs, SIV239Deltanef and SIVPBj6.6Deltanef, were tested for their abilities to stimulate protective immunity in macaques. During the immunization period the animals were examined for specific immune responses and virus growth. Each construct generated high levels of specific immunity in all of the immunized animals. The SIV239Deltanef construct was found to grow to high levels in all immunized animals, with some animals remaining positive for virus isolation and plasma RNA throughout the immunization period. The SIVPBj6.6Deltanef was effectively controlled by all of the immunized animals, with virus mostly isolated only during the first few months following immunization and plasma RNA never detected. Following an extended period of immunization of over 80 weeks, the animals were challenged with a pathogenic simian-human immunodeficiency virus (SHIV) isolate, SIV89. 6PD, by intravenous injection. All of the SIV239Deltanef-immunized animals became infected with the SHIV isolate; two of five animals eventually controlled the challenge and three of five animals, which failed to check the immunizing virus, progressed to disease state before the unvaccinated controls. One of five animals immunized with SIVPBj6.6Deltanef totally resisted infection by the challenge virus, while three others limited its growth and the remaining animal became persistently infected and eventually died of a pulmonary thrombus. These data indicate that vaccination with attenuated SIV can protect macaques from disease and in some cases from infection by a divergent SHIV. However, if animals are unable to control the immunizing virus, potential damage that can accelerate the disease course of a pathogenic challenge virus may occur.

Phylogenetic analysis of gag genes from 70 international HIV-1 isolates provides evidence for multiple genotypes.

OBJECTIVE: To determine the extent of genetic variation among internationally collected HIV-1 isolates, to analyse phylogenetic relationships and the geographic distribution of different variants. DESIGN: Phylogenetic comparison of 70 HIV-1 isolates collected in 15 countries on four continents. METHODS: To sequence the complete gag genome of HIV-1 isolates, build multiple sequence alignments and construct phylogenetic trees using distance matrix methods and maximum parsimony algorithms. RESULTS: Phylogenetic tree analysis identified seven distinct genotypes. The seven genotypes were evident by both distance matrix methods and maximum parsimony analysis, and were strongly supported by bootstrap resampling of the data. The intra-genotypic gag distances averaged 7%, whereas the inter-genotypic distances averaged 14%. The geographic distribution of variants was complex. Some genotypes have apparently migrated to several continents and many areas harbor a mixture of genotypes. Related variants may cluster in certain areas, particularly isolates from a single city collected over a short time. CONCLUSIONS: The genetic variation among HIV-1 isolates is more extensive than previously appreciated. At least seven distinct HIV-1 genotypes can be identified. Diversification, migration and establishment of local, temporal 'blooms' of particular variants may all occur concomitantly.

Genetic variants of HIV-1 in Thailand.

Serosurveys conducted prior to 1988 indicated a very low level of HIV-1 infection in Thailand, even among high-risk groups. The Ministry of Health has reported a dramatic increase in HIV-1 infection during the last three years. The geographic and demographic distribution of the epidemic is broad, involving multiple provinces and risk groups. Foci of higher incidence and prevalence have been noted in the urban center of Bangkok and in the northern provinces of Chiang Mai and Chiang Rai. Here we report the results of genetic characterization of 16 HIV-1 isolates from Thailand using a combination of polymerase chain reaction (PCR) typing and DNA sequencing. The complete sequence of gp160 (env) of five isolates, partial env sequence of six additional isolates, and the gag gene of two isolates were determined. Two highly distinct HIV-1 variants were found. One variant resembled those prevalent in North America and Europe; five of the isolates were of this type. The remaining eleven isolates were very similar to one another and represented a variant unlike any previously described. Phylogenetic tree analysis of complete env and gag genes placed the two variants on widely separated branches. Protein sequence comparisons indicate both general and specific features that distinguish the Northern Thailand variant both from the Bangkok variant and from virtually all previously sequenced HIV-1 isolates. A simple PCR test for distinguishing the two variants has been developed for use in epidemiologic surveys.

Constraints to DNA unwinding near radiation-induced strand breaks in Ewing's sarcoma cells.

Ewing's sarcoma cell lines were compared to other cell lines for induction of DNA strand breaks by ionizing radiation and their ability to repair those breaks. The alkali-unwinding assay and alkaline sucrose gradient analysis were used for these studies. The alkali-unwinding assay revealed that the amount of DNA unwound per strand break in Ewing's sarcoma cells was less than for other cells and was not influenced by high-salt denaturation conditions. Ewing's sarcoma cells had similar induction and repair rates for strand breaks compared with other cell lines. The kinetics of unwinding suggests there are constraints to DNA unwinding in the chromatin of Ewing's sarcoma cells, possibly related to high levels of poly(ADP-ribose) polymerase in these cells.