Literature review > Issue_2 > Review Ikoma et al. 

Glycoprotein G is a structural protein produced by HSV-1 and HSV-2. To date, it has been the mainstay in development of tests that can accurately differentiate between antibodies to HSV-1 and those to HSV-2. Gull Laboratories received FDA approval in 1999 for ELISAs based on affinity purified gG-1 or gG-2. These tests were sold briefly by Meridian Biosciences, then withdrawn from the market early in 2001. Diagnology, (Belfast, Northern Ireland) received FDA approval in October 1999 for a novel point-of-care membrane test called "POCkit -HSV-2". In 2000, gG-based ELISAs and a strip immunoblot format developed by MRL received FDA approval. MRL is now Focus Technologies and its kits are sold in the United States and worldwide under the HerpeSelect brand name. A number of gG-based tests are available in the European market, including POCkit-HSV-2 and HerpeSelect ELISA.

The authors of this paper constitute a European powerhouse of expertise in the area of human immune response to glycoprotein G. Much of the practical knowledge about this largely type specific protein is due to their work. The paper offers a concise review of current knowledge of the structure, processing, and function of gG. The study itself was designed to test performance characteristics of two glycoprotein fragments; one containing epitopes specific for gG-1 and the other containing most of the known epitopes of gG-2. The fragments are shown to be highly type specific by their selective reactivity with monoclonal antibodies. Preliminary Western blot analyses demonstrated type specificity with human sera from patients of known HSV serostatus.

ELISAs based on the gG-1 and gG-2 fragments were then tested with human sera. A total of 267 sera from a variety of sources were used including 20 sera with previously published discordant type specific antibody testing results. One of the subsets consisted of 64 sera from culture-documented HSV-2 patients in Sweden. Against this gold standard, the gG-2 fragment-ELISA was 100% sensitive. A second set of 64 sera from culture-documented HSV-2 patients in Amsterdam showed a much lower sensitivity (50%). Further testing suggests that some of the "false-negative" sera were from patients with new HSV-2 infections. The discrepancy between the two sets may be explained if the Swedish patients were pre-selected to have histories of recurrent herpes whereas the Amsterdam group were "all comers." Unfortunately, clinical data are lacking on both groups.

The performance data are mainly in the form of comparison against the Gull gG-2 ELISA. Although this test was probably the best single comparator candidate at the time of the study; it is unfortunate that the considerable body of data on the Gull test is no longer immediately applicable. The protocol may have been underway when a 2001 publication by Whittington and colleagues reported low sensitivity of the Gull assay relative to Western blot in a very large, well characterized STD population. The insensitivity of the Gull assay almost certainly skewed the specificity calculation for the fragment gG-2 ELISA.

The authors do not speculate how these antigens might augment the existing recombinant-based Focus assay. The HerpeSelect test is, by default, the major gG-based assay in the United States and is available through nearly every major testing lab. There is no widely available test to sort out HerpeSelect equivocal results or to definitively tie down a low positive result in the occasional patient who needs the reassurance of a second test. One can hope that the authors, with their skill in recombinant technology, will follow on this study with one that shows how their test can fit into the current diagnostic toolbox.

The fragment-gG tests have been thoughtfully developed and carefully tested by this group. The tests discriminate between HSV-1 and HSV-2 antibodies in most cases. The authors did an admirable job in collecting and presenting data. But good as this study is, it points up some real problems in the development and testing of new diagnostics for genital herpes [1]. The development and application of diagnostic tests for genital herpes is complicated by the unique biologic aspects of the disease: subclinical acquisition, involvement of both HSV-1 and HSV-2, and high seroprevalance of both HSV types, to name a few. Test development has been expensive and slow because of the high cross reactivity of HSV-1 and HSV-2.

The lack of a clear gold standard for new tests has led to a great deal of confusion in this field. New diagnostic tests can look wonderful against a comparator test in premarket studies [2,3], then turn out to have substantial flaws in wider practice [4,5], when more focused patient groups are identified. In any population, comparing one test to another is fundamentally limited by the summed inadequacies of the two tests. Even Western blot may miss a diagnosis [6]! The ideal gold standard is a virological test for infection status. But this limits performance information to symptomatic patients and, practically, restricts studies of new tests to large centers with substantial resources for identifying, characterizing, and consenting participants

References:

1. Ashley RL. Performance and use of HSV type-specific test kits. Herpes 2002;9:38-45.

2. Ashley RL, Wald A, Eagleton M. Premarket evaluation of the POCkit HSV-2 type specific serologic test in culture-documented cases of genital herpes simplex virus type 2. Sex Trans Dis 2000;27:266-269.

3. Ashley RL, Wu L, Pickering JW, Tu M-C, Schnorenberg L. Premarket evaluation of a commercial glycoprotein-G based enzyme immunoassay for herpes simplex virus type-specific antibodies. J Clin Microbiol 1998;36:294-295.

4. Saville M, Brown D, Burgess C, Perry K, Barton S. Cowan F, et al. An evaluation of near patient tests for detecting herpes simplex virus type-2 antibody. Sex Transm Infect 2000; 76:381-382.

5. Whittington WLH, Celum C, Cent A, Ashley R. Use of a glycoprotein G-based type-specific assay to detect antibodies to herpes simplex virus type 2 among persons attending sexually transmitted disease clinics. Sex Trans Dis 2001;28:99-104.

6. Ashley RL, Krantz E, Wald A. Time course of seroconversion by HerpeSelect ELISA after acquisition of genital herpes simplex virus type 1 (HSV-1) or HSV-2. Sex Trans Dis 2003; 30:310-314

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