HIV type 1 subtypes in Malaysia include B, C, and E

Peripheral blood mononuclear cell specimens were collected from 13 HIV-1-infected IV drug users in Kuala Lumpur, Malaysia, as well as one HIV-infected baby, between 1992 and 1993. DNA was then amplified by nested polymerase chain reaction and a 345-bp fragment of the C2V3 region of the env gene was sequenced. 11 of the 14 Malaysian sequences clustered with the B' subtype, one different from the typical subtype B US strains HIVMN and HIVSF2. Two sequences grouped in the C subtype and had sister taxa closer to the Indian C subtype sequences than those from Zambia. The sequence from the infant was identified as a subtype E virus, grouped more closely with subtype E strains from Thailand than subtype E viruses from the Central African Republic.     

HIV type 1 subtypes, coreceptor usage, and CCR5 polymorphism

Identification of the chemokine receptors CCR5 and CXCR4 as the major coreceptors for HIV-1 entry has greatly assisted our understanding of HIV-1 pathogenesis, transmission, and tropism. However, most of our current knowledge on coreceptor usage comes from studies using HIV-1 strains or env genes derived from the genetic subtype B predominant in North America and western Europe. In this report, the coreceptor usage of 20 primary viral isolates representative of genetic subtypes A, B, C, D, E, and group O was examined. Thirty-nine full-length CCR5 sequences from individuals of diverse geographic origins were also obtained to examine the possible effect of CCR5 polymorphism on HIV-1 subtype distribution. Our results indicate that (1) CCR5 and CXCR4 serve as the two major coreceptors for viruses belonging to HIV-1 subtypes A, B, C, D, E, and group O, whereas other chemokine receptors such as CCR2b and CCR3 play only a minor role in facilitating viral entry into stimulated PBMCs; (2) the coreceptor usage is determined by the viral phenotype rather than its genotype because all NSI strains, irrespective of their subtype classification, utilize CCR5, whereas all SI strains are able to use CXCR4; and (3) there is no geographic clustering of CCR5 polymorphism in different ethnic populations, suggesting that CCR5 diversity is not the underlying explanation for differences in the spread of different HIV-1 subtypes. Therefore, the uneven worldwide distribution of HIV-1 subtypes is more likely the result of stochastic dissemination.     

Diversity among clinical isolates of Helicobacter pylori in Korea

Eleven strains of Helicobacter pylori were isolated from biopsy specimens obtained at the A-San Medical Center from December, 1995 to February, 1996. Every H. pylori positive patient was diagnosed to have chronic, erosive, or mild superficial gastritis. To determine the diversity in clinical isolates, the following were studied: total protein profile, plasmid profile, presence of cagA, and variation in DNA sequence. Protein profiles of nine isolates were similar to each other while two isolates had a 35 kDa protein which did not appear in others. The presence of cagA was detected with PCR in seven isolates (63%). Among eleven isolates, seven (63%) carried plasmids. Each isolate showed a big diversity with a PCR-based randomly amplified polymorphic DNA method. Even though all H. pylori isolates used in this study were isolated from gastritis patients at the same hospital, their molecular and biological characteristics were quite different from another showing a big diversity in H. pylori.     

HIV type 1 subtypes in Malaysia, determined with serologic assays: 1992-1996

We investigated the molecular epidemiology of HIV-1 subtypes in Malaysia among injecting drug users (IDUs) and sexual transmission risk groups, using serologic and genetic techniques. Frozen sera collected at a general hospital, a blood bank, several drug treatment centers, and an STD clinic in Kuala Lumpur, between 1992 and 1996, were investigated retrospectively. V3 peptide serotyping and monomeric gp120 capture serotyping were used to study 89 known HIV-1-infected subjects. The methods differentiate subtypes B, E, and C. V3 peptide and gp120 capture results were comparable. No subtype C-specific reactive sera were found; one specimen was dually reactive for subtypes C and B, using the V3 peptide ELISA; and four were durally reactive for subtypes E and C using this assay. Genotypic analysis of HIV-1 gag RNA in serum was done on a subset of subjects and confirmed serologic findings. HIV-1 subtypes differed significantly by risk category: of 53 IDUs, 29 (55%) were infected with subtype B and 19 (36%) were infected with subtype E, 3 (6%) were dually reactive, and 2 (4%) were not typable. Of 36 persons with heterosexual risks, 29 (81%) were infected with subtype E, 5 (14%) were infected with subtype B, and 2 (5%) were not typable. Persons with IDU risks were significantly more likely to be infected with subtype B than were those with sexual risks (OR 5.89; 95% CI, 1.94-18.54; p < 0.001). Subtypes B and E of HIV-1 appear to predominate in Malaysia; subtype B was more prevalent among IDUs; subtype E was more prevalent among all other groups. These results may have important HIV-1 vaccine implications.     

The isolation and characteristics of monoclonal antibodies to recombinant proteins of HIV-1 and HIV-2--the env and gag gene products

Ten hybridomas secreting monoclonal antibodies (Mab) against recombinant HIV-1 and HIV-2 antigens were produced (3 Mab anti-gag protein, 2 anti-env1, and 5 anti-env2). In the immunoblotting assay all the anti-gag Mabs reacted with HIV capsid protein p24, whereas one of them reacted also with p55 protein and with 7 other polypeptides. Another anti-gag Mab cross-reacted with the antigen of subpopulation of human peripheral blood lymphocytes. The third one interacted with the antigen of both HIV-1 and HIV-2. All the 10 Mabs interacted with natural HIV antigens and can be used for identification and differentiation of HIV-1 and HIV-2.     

Reduced infectivity of human immunodeficiency virus type 1 produced in the presence of a truncated Gag protein containing p7 gag and p6 gag

The C-terminal portion of human immunodeficiency virus type 1 p55gag protein, p15gag, contains two functional proteins; p6gag which is required for incorporation of Vpr into the virion, and p7gag which binds to viral RNA and is necessary for packaging of genomic RNA into virions. p7gag protein overexpressed in trans may compete with wild type p55gag for binding to genomic viral RNA, thereby inhibiting incorporation of RNA into the virions. To investigate if overexpression of the C-terminal portion of p55gag could interfere with generation of infectious virus, a plasmid producing a protein consisting of p2gag, p7gag and p6gag, termed p15gag*, was generated and cotransfected with an infectious proviral human immunodeficiency virus type 1 clone. Cells overexpressing p15gag* in trans produced approximately 40 fold less infectious virus than cells lacking exogenous p15gag*. These results demonstrated that expression of the C-terminal portion of p55gag efficiently reduced virus infectivity.     

Identification of immunoreactive epitopes in proteins coded by gag, env, and pol genes of the type I human T-lymphotropic virus (HTLV-I) using synthetic peptides

Reactivity of 26 synthetic peptides that comprise 12 to 26 amino acid residues corresponding to segments of the gag p19, env gp46, and pol proteins of human T-lymphotropic virus type I toward 31 positive sera was studied using enzyme-linked immunosorbent assay. Specific reactivity with high titers of antibodies (presented in reciprocal dilution values) was detected for the synthetic peptides corresponding to fragments 110-130 and 100-130 (titers up to 4050) of p19, 174-197 (up to 800), 186-201 (up to 4050), 191-215 (up to 1350), 242-257 (up to 800), and 272-292 (up to 450) of gp46. Immunoreactivity of seven peptides, fragments of pol-proteins, was weak. New linear epitopes in the regions 145-158, 272-277, and 292-300 of gp46 were detected. In addition, location of the known linear epitopes in p19 and gp46 was refined on the basis of comparative study of overlapping peptides from these proteins.     

Genetic localization of the type I trimethoprim resistance gene and its dissemination in urinary tract isolates in Taiwan

In a total of 425 urinary isolates of E. coli, Enterobacter spp., and Klebsiella spp. selected, there were 169 (45.4%) isolates harbouring type I dihydrofolate reductase (DHFR) gene among 374 trimethoprim-resistant isolates. In these 169 isolates, only 17.2% hybridized with the Tn7 probe. According to another probe specific for the integrase gene of integron, 87.6% showed a positive reaction. Further analysis by restriction mapping proved that the type I DHFR gene was inserted into a integron-like structure. These results indicate that the type I DHFR gene that was initially observed in association with transposable element Tn7 is becoming associated with an integrase function similar to integrons in most instances. Further analysis of the distribution of Tn21-like integrase gene in clinical isolates indicated that the prevalence rates were 86.4%, 84.8%, and 76.7% respectively in E. coli, Enterobacter spp., and Klebsiella spp.. Furthermore, the integrase gene found in our clinical isolates proved to be mediated by a plasmid, demonstrated by Southern hybridization. Thus, the trimethoprim-resistant gene that developed under selective pressure from the double drug trimethoprim and sulphonamide was transmitted by insertion into integron-like structure and then mediated by plasmid transfer for dissemination.     

Diversification of subtype E human immunodeficiency virus type 1 env in heterosexual seroconverters from Northern Thailand

The C2-V3 region of the human immunodeficiency virus (HIV)-1 env was determined from 15 northern Thailand seroconverters between 1993 and 1995. Similar sequences were also determined from 18 seroconverting injection drug users in Baltimore. All seroconverters from northern Thailand were infected with subtype E HIV-1 on the basis of env sequences. Intersubject viral DNA distances increased from 2.3% in asymptomatic HIV-1-infected subjects characterized between 1990 and 1992 to 7.8% in these more recent seroconverters from Thailand. On the other hand, sequences from 18 seroconverters from Baltimore had a mean intersubject distance of 13.2%. The genetic diversity within HIV-1 subtype E in seroconverters in Thailand has increased significantly but is still less than that observed in HIV-1 from seroconverters in the United States, where the epidemic of HIV-1 infection is more mature. These results suggest that continued monitoring of the molecular epidemiology of HIV-1 infection in Thailand will be important for HIV vaccine development and evaluation.