Laboratory testing and rapid HIV assays: applications for HIV surveillance in hard-to-reach populations.

Most HIV surveillance has been performed through serologic surveys in relatively stable, accessible populations. Similar surveillance, with or without counseling and testing, in populations that are hard-to-reach, presents logistical challenges, including the selection of laboratory testing strategy and algorithm. The advent of rapid serologic assays for HIV now allows for on-site testing, including confirmatory testing, and rapid provision of test results and counseling. The possibility of only a single contact makes repeat sampling, which current diagnostic testing recommendations include, difficult. To address the logistical complexities in surveillance in hard-to-reach populations and the increased availability of rapid tests, we propose adapting the testing strategies for HIV of the World Health Organization/the joint United Nations Programme on HIV/AIDS in order to facilitate this surveillance, including, where carried out, the provision of test results back to individuals. The choice of enzyme-linked immunosorbent assay (ELISA) versus rapid testing for these settings is discussed, as is the choice of specimen--blood, oral fluid, or urine. Three appendices summarize: (1) test algorithms for the various testing strategies; (2) advantages and disadvantages of ELISA and of rapid test formats, and (3) the characteristics and status of currently available rapid HIV tests. We also discuss the potential application of the recently developed 'detuned' methodology for estimating HIV incidence in hard-to-reach populations.

The development and evaluation of a probe hybridization method for subtyping HIV type 1 infection in Uganda.

We developed a method for large-scale screening of HIV-1 genotypic variation based on DNA probe hybridization. Nested PCR amplifications were performed to generate fragments in the env C2-V3 region and also in the gp41 region, which encompasses the immunodominant domain. The proviral DNA sequences were derived from 68 samples and phylogenetically analyzed. For comparison, the C2-V3 fragment was used in DNA probe hybridization to rapidly determine the infecting HIV subtype. The hybridizing probes were designed on the basis of the two most prevalent subtypes in Uganda, A and D. The results were compared to evaluate the feasibility of using this hybridization method for large-scale genotypic screening. Sequence analysis of the 68 amplified PCR fragments showed that 39 were subtype A and 29 were subtype D. The results of DNA hybridization to the amplified products with A and D subtype-specific probes were more than 90% concordant with the subtypes determined by sequence analysis. Our findings suggest that probe hybridization with subtype-specific probes is effective for large-scale screening of HIV-infected populations. Application of this method will significantly reduce the time needed for large, population-based investigations.

Mucosal disruption due to use of a widely-distributed commercial vaginal product: potential to facilitate HIV transmission.

OBJECTIVE: Policresulen vaginal suppositories are a condensation product of metacresolsulfonic acid and formaldehyde. We investigated their use by female commercial sex workers (CSW) and whether such use could facilitate HIV transmission. METHODS: We interviewed female CSW in Thailand about use of the product, and we directly observed the effects of self-administration of a single suppository by each of six women. RESULTS: Of 200 CSW interviewed, 32% had used policresulen vaginal suppositories in the preceding year and 46% had used them at some time. Many used them for reasons not listed on the package insert, such as improving their male partners' sexual pleasure, and most did not abstain from vaginal sex following use. Among 36 brothel-based and 67 non-brothel-based CSW with known HIV infection, the use of the product was not associated with HIV-1 infection (adjusted relative risk 1.0, 95% confidence interval, 0.5-2.0). Exfoliation of the vaginal and cervical mucosa was observed in all six CSW 1 day after product use, and, although it could have been the result of repeated examinations, an increase in genital HIV-1 RNA shedding was also detected in all three HIV-seropositive women. CONCLUSION: Although there was no epidemiological association with HIV infection, policresulen vaginal suppository use did disrupt the genital mucosa and therefore may have the potential to facilitate HIV transmission. Drug licensing authorities may wish to reassess the safety of this product. If the product continues to be distributed, steps should be taken to limit its use to the specific conditions for which it is indicated and to ensure that women abstain from vaginal sex following its use.

Mapping and serodiagnostic application of a dominant epitope within the human herpesvirus 8 ORF 65-encoded protein.

A dominant epitope within the human herpesvirus 8 (HHV8) ORF 65-encoded protein was mapped to an 8-amino-acid (aa) sequence (RKPPSGKK [aa 162 to 169]) by an amino acid replacement method. Using a 14-aa peptide (P4) encompassing this epitope as the antigen, we developed an enzyme immunoassay for HHV8 antibodies. The presence of P4 antibodies in a panel of 61 human serum specimens was highly correlated with biopsy-confirmed Kaposi's sarcoma. The homologous Epstein-Barr virus peptide derived from BFBR3-encoded protein did not interfere with the assay, suggesting that P4 is specific for HHV8.

Presence of human immunodeficiency virus (HIV) type 1 subtype A infection in a New York community with high HIV prevalence: a sentinel site for monitoring HIV genetic diversity in North America. Centers for Disease Control and Prevention-Bronx Lebanon HIV Serosurvey Team.

To determine whether US residents are infected with subtypes of human immunodeficiency virus (HIV) type 1 other than subtype B (Western), the predominant North American subtype with a unique GPGR genetic sequence in the V3 loop, viruses from 22 HIV-infected adults were serotyped and subtyped. Twenty patients had subtype B (Western), of whom 15 had serotype B (Western), 3 had serotype A/C, 1 had serotype B (Thai), and 1 had a nontypeable serotype. Two had subtype A, both serotype A/C. Both subtype A-infected patients, only 1 of whom had been outside the United States, reported sex with persons traveling abroad, suggesting possible acquisition in the United States. Because US residents are infected with non-subtype B (Western) strains, US surveillance for HIV-1 diversity is needed to elucidate subtype-specific transmission patterns and pathogenesis and to guide evaluation and development of HIV diagnostic tests and vaccines.

Detection of genetically diverse human immunodeficiency virus type 1 group M and O isolates by PCR.

A panel of 136 genetically diverse group M and 5 group O adult isolates from outside the United States and Europe were evaluated by PCR with the Roche AMPLICOR HIV-1 test, a modified version of the AMPLICOR HIV-1 test, and a new primer pair/probe system. Detection of some of these isolates was less efficient with the AMPLICOR HIV-1 test; however, the assay was significantly improved by reducing the sample input and lowering the annealing temperature. The new primer pair/probe set detected 140 of 141 isolates, including the 5 group O isolates that were not detected with either of the AMPLICOR HIV-1 test formats.

Cross-reactivity between herpes simplex virus glycoprotein B and a 63,000-dalton varicella-zoster virus envelope glycoprotein.

Cross-reactive monoclonal antibodies recognizing both herpes simplex virus (HSV) glycoprotein B and a major 63,000-dalton varicella-zoster virus (VZV) envelope glycoprotein were isolated and found to neutralize VZV infection in vitro. None of the other VZV glycoproteins was recognized by any polyclonal anti-HSV serum tested. These results demonstrate that HSV glycoprotein B and the 63,000-dalton VZV glycoprotein share antigenic epitopes and raise the possibility that these two proteins have a similar function in infection.

A rapid procedure for the enrichment of undenaturated, antigenically active herpes simplex virus glycoproteins.

The usefulness of lentil lectin affinity chromatography for the rapid enrichment of HSV glycoproteins in an undenatured state for both research and clinical purposes was investigated. In order to compare the lentil lectin-binding characteristics and immunologic specificities of undenatured HSV-1 and HSV-2 glycoproteins, [35S]methionine-labelled extracts of virus-infected HEp-2 cells were subjected to lentil lectin affinity chromatography. Individual HSV-1 and HSV-2 glycoproteins in bound and unbound fractions were identified using monoclonal antibodies. With the exception of a portion of pgD and gD, all major viral glycoprotein species (gA, gB, gC, gD, gE and gF) and their glycosylated processive intermediates bound to lentil lectin indicating that all possess predominantly mannosyl and/or glucosyl carbohydrate moieties. Although the unbound pgD and gD species were glycosylated, no gD and only a portion of pgD bound to lentil lectin when reapplied to the column indicating that these subspecies possess alterations in factors required for efficient lectin binding. Immunoprecipitation of undenatured lectin-bound glycoproteins from infected cells using HSV-1 and HSV-2-specific rabbit and human antisera confirmed previous findings that the predominant type-specific glycoproteins of HSV-1 and HSV-2 are gC and gE/gF, respectively.

Aminoacyl fucosides as possible biochemical markers at tumorigenic and metastatic potential in herpes simplex virus type 2-transformed rat cells.

Two classes of aminoacyl fucosides termed FL3 and FL4 were studied as possible markers of tumorigenic and metastatic potential in herpes simplex virus type 2 transformed rat cells. In the present study, clonal cell lines of transformed highly tumorigenic and metastatic (t-REF-G-1.1), weakly tumorigenic and nonmetastatic (t-REF-G-2.1), nontumorigenic (t-REF-G-2.0), and secondary nontransformed rat embryo fibroblast cells were labeled with [3H]fucose, and cell extracts were analyzed for ratio of radioactivity incorporated into FL3 and FL4. Results indicated that, in extracts from t-REF-G-2.0 and nontransformed rat embryo fibroblast cells, the ratios of FL4/FL3 were 5.78 and 5.71, respectively. In contrast, t-REF-G-2.1 cells exhibited a FL4/FL3 ratio of 1.45, while t-REF-G-1.1 cells exhibited a FL4/FL3 ratio of 0.74. In subclonal cell lines isolated from TPA-treated and mock-treated t-REF-G-2.1 cells, the FL4/FL3 ratios correlated with the tumorigenic and metastatic potential of these subclones in newborn syngeneic White Buffalo rats. These data suggested that alterations in fucose-labeled components can be used to predict the tumorigenic and metastatic potential of herpes simplex virus type 2-transformed rat cells.