Comparison of the Avidity Index Method and the Serologic Testing Algorithm for Recent Human Immunodeficiency Virus (HIV) Seroconversion, Two
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微生物临床杂志 2005年第12期
Center for Epidemiological Studies on HIV/AIDS of Catalonia (CEESCAT), Hospital Universitari Germans Trias i Pujol. Badalona, Spain
Microbiology Service, Hospital Universitari Germans Trias i Pujol, Departament de Genetica i Microbiologia, Universitat Autònoma de Barcelona, Badalona, Spain
Department of Infectious Diseases, Istituto Superiore di Sanita, Rome, Italy
Laboratory of Virology, Ospedale Amedeo di Savoia, Turin, Italy
Newborn Screening Quality Assurance Program, Centers for Disease Control and Prevention, Atlanta, Georgia
ABSTRACT
A study was designed to compare an avidity index method to the serologic testing algorithm for recent human immunodeficiency virus (HIV) seroconversion (STARHS) for the detection of recent HIV infection. One hundred sixty HIV-positive sera were tested. Both techniques performed similarly in identifying recent infections, although STARHS tended to misclassify more individuals that had long-standing infection as being recently infected.
TEXT
Increasing interest in estimating the incidence of human immunodeficiency virus (HIV) infection by using cross-sectional specimens has led to the development of several laboratory methods. A sensitive/less-sensitive serologic strategy (the serologic testing algorithm for recent HIV seroconversion [STARHS]) was developed in 1998 (4) at the Centers for Disease Control and Prevention (CDC), Atlanta, Ga., to identify recently infected individuals and has been used in numerous studies (3, 9-11, 13-15). In Catalonia, Spain, STARHS has been introduced as part of enhanced surveillance of HIV and sexually transmitted infections since 2002 (1). Antibody avidity has long been used as a marker for recent infection (2, 5, 8), and in 2002, an automated technique was developed for HIV (17, 18) based on the rationale that in the early phase of infection antibodies show a low avidity for the antigen. In this study, we report the first comparison of these two methods for the detection of recent HIV infection.
A total of 160 serum specimens from HIV-positive subjects (identified according to standard HIV testing algorithms for screening and confirmatory testing) were included in the study. For 66 specimens, the seroconversion date or clinical stage of infection was not available. For 70 specimens, a seroconversion date could be estimated as the midpoint between the last HIV-negative test and the first HIV-positive test (interval between the two tests of <6 months). These specimens were sorted into two groups. Group 1 included 13 specimens from 12 recently infected individuals (sera drawn 6 months after the seroconversion date). Group 2 comprised 57 specimens from 26 individuals with long-standing infection (sera drawn >6 months after the diagnosis of HIV infection). Group 3 consisted of 24 individuals with a diagnosis of AIDS.
STARHS was implemented whereby all specimens were first tested using Biotest HIV-TETRA HIV 1 + 2 (Biotest; Diasorin Diagnostics S.R.L., Italy) as the sensitive enzyme-linked immunosorbent assay (ELISA). The Vironostika HIV-1 Microelisa System (bioMerieux, Durham, N.C.) was used as the less-sensitive ELISA after a 1:20,000 three-step dilution as previously described (12). Appropriate controls and calibrator provided by the CDC were used in each run. First, a screening assay using one well per specimen was performed to allow for the identification of probable recent seroconverters according to standardized optical density (SOD), calculated as (patient OD – negative control OD)/(calibrator OD – negative control OD). Specimens with an SOD of <2.000 were then retested in triplicate (confirmatory assay). Specimens with an SOD of <1.000 in the confirmatory assay were considered as belonging to subjects who had seroconverted within the previous 170 days (95% confidence interval, 145 to 200).
For the avidity method, two aliquots of each sample were subjected to a pretest 1:10 dilution. One aliquot was diluted in phosphate-buffered saline (PBS) and the other one in 1 M guanidine hydrochloride (G; Pierce Chemicals, Dallas, Tex.). After incubation at room temperature, both aliquots were tested by an automated microparticle immunoassay (AxSYM HIV 1/2 gO; Abbott Diagnostics Division, Delkenheim, Germany) as recommended by the manufacturer. Sample/cutoff (S/CO) ratios were calculated, and the avidity index of HIV antibodies was computed as (S/CO ratio of the G aliquot)/(S/CO ratio of the PBS aliquot). A cutoff of 0.85 was selected based on previous results (16).
The overall agreement between both techniques was good ( = 0.67, P < 0.001), where both techniques were concordant in classifying 25 specimens as recent infections and 117 as long-standing infections. Fourteen specimens were considered as recent infections by STARHS while 4 specimens were considered recent infections by the avidity index. Although the avidity technique showed better intra- and interday coefficients of variation (<10% versus 10 to 30%) (17), a number of specimens were close to the cutoff (Fig. 1). Results according to clinical information are shown in Table 1. Both techniques misclassified one out of 13 specimens corresponding to recent infections as being long-standing (the misclassified specimens were different from each other). Six specimens from long-standing infections were misclassified as recent infections by STARHS; one of them had been collected 7 months after the first HIV-positive test and was from a patient on highly active antiretroviral treatment (HAART), while the other five were obtained from subjects who had been infected with HIV for over a year. The only specimen misclassified by the avidity method had been collected 1 year after the first HIV-positive test. Another two specimens from subjects with long-standing infection were from subjects on HAART and were correctly identified as old infections by both techniques. Among the 24 AIDS patients, one was misclassified by STARHS only and another two were misclassified by both assays. Neither of the two tests seemed to be affected by a low CD4 count.
While STARHS has a proven value, several drawbacks limit its use. First, even small errors in the three-step specimen dilution magnify differences in SOD values, leading to a high variability among replicates and requiring additional testing (6). Secondly, a common calibrator and quality control material, along with a software package, were developed and must be provided by the CDC. Third, STARHS results vary according to HIV subtype, requiring the adoption of different cutoffs (19). Finally, problems with the availability of the commercialized test used for this technique have occurred in the past and will arise again in the near future.
The avidity index method shows several advantages over STARHS, as previously suggested (18). After a single 1:10 dilution, the technique can be fully automated using the AxSYM platform, which leads to better reproducibility. This instrument is available among clinical laboratories in Europe and in most countries worldwide. While STARHS uses a second-generation ELISA based on a virus lysate (HIV-1 subtype B), the avidity index assay is a third-generation enzyme immunoassay that is more sensitive and detects antibodies earlier (it is able to recognize all antibody classes). Moreover, this assay allows the detection of HIV-1, groups M and O, and HIV-2. In addition, the selection of different avidity index cutoffs offers different sensitivities and specificities, depending on the study purpose. Moreover, the avidity index method showed more accurate results than STARHS among patients with long-standing HIV infection or AIDS. In our study there were too few subjects on antiretroviral therapy to evaluate its effect on test performance. However, STARHS tends to misclassify the subjects on therapy as being recently infected, since long-term HAART may lead to a decrease in antibody titers (4, 7, 14). Recently, a longitudinal study that compared STARHS and the avidity index assay for patients who had started antiretroviral therapy (G. Murphy, A. Charlett, G. L. Dean, M. Fisher, D. McElborough, and J. V. Parry, Abstr. XV Int. AIDS Conf., abstr. C11390, 2004) concluded that this therapy had a marked effect on STARHS but had only minor effects on the avidity assay. The avidity index method is easily exportable and does not require any specific training other than that provided by the manufacturer. Finally, the avidity index method does not need official approval by the regulatory authority in any country because it is not a new assay or a modification of an already existing assay and only a preparation of the sample is needed.
It is appropriate to stress that currently the results of both of these tests cannot be interpreted at an individual level. The discrimination between recent and long-standing HIV infection is based on a cutoff that defines the time period from seroconversion to either high antibody titers (STARHS) or antibody maturation (avidity index). However, this time period may vary among individuals, causing misclassification in some cases. Only at the population level will the false-negative and false-positive results balance each other.
ACKNOWLEDGMENTS
This work was partially supported by a grant from the Istituto Superiore di Sanita, V Progetto Nazionale di ricerca sull'AIDS 2003, and grant from Fundacio Barcelona SIDA 2002.
We thank Rafael Muoz for management and storage of specimens and Meritxell Granell for the revision of medical charts. We also thank Robert H. Byers for helpful discussion. Joanne V. Mei's role in this project was limited to providing technical assistance and advice.
The Recent HIV Infections Study Group consists of Diana Puente, Claudio Galli, M. Jesús Casado, Daniela Bernasconi, Mauro Sciandra, Laura Camoni, Lurdes Matas, Giovanni Rezza, Vicen Ausina, and Jordi Casabona.
REFERENCES
Center for Epidemiological Studies on HIV/AIDS of Catalonia (CEESCAT). 2005. SIVES 2004, Integrated HIV/STI Surveillance System of Catalonia (SIVES) Annual Report. Generalitat de Catalunya, Departament de Salut, Barcelona, Spain.
Eggers, M., U. Bader, and G. Enders. 2000. Combination of microneutralization and avidity assays: improved diagnosis of recent primary human cytomegalovirus infection in single serum sample of second trimester pregnancy. J. Med. Virol. 60:324-330.
Gouws, E., B. G. Williams, H. W. Sheppard, B. Enge, and S. A. Karim. 2002. High incidence of HIV-1 in South Africa using a standardized algorithm for recent HIV seroconversion. J. Acquired Immune Defic. Syndr. 29:531-535.
Janssen, R. S., G. A. Satten, S. L. Stramer, B. D. Rawal, T. R. O'Brien, B. J. Weiblen, F. M. Hecht, N. Jack, F. R. Cleghorn, J. O. Kahn, M. A. Chesney, and M. P. Busch. 1998. New testing strategy to detect early HIV-1 infection for use in incidence estimates and for clinical and prevention purposes. JAMA 280: 42-48.
Jenum, P. A., B. Stray-Pedersen, and A. G. Gundersen. 1997. Improved diagnosis of primary Toxoplasma gondii infection in early pregnancy by determination of antitoxoplasma immunoglobulin G avidity. J. Clin. Microbiol. 35:1972-1977.
Kothe, D., R. H. Byers, S. P. Caudill, G. A. Satten, R. S. Janssen, W. H. Hannon, and J. V. Mei. 2003. Performance characteristics of a new less sensitive HIV-1 enzyme immunoassay for use in estimating HIV seroincidence. J. Acquired Immune Defic. Syndr. 33:625-634.
Machado, D. M., E. L. Delwart, R. S. Diaz, C. F. de Oliveira, K. Alves, B. D. Rawal, M. Sullivan, M. Gwinn, K. A. Clark, and M. P. Busch. 2002. Use of the sensitive/less-sensitive (detuned) EIA strategy for targeting genetic analysis of HIV-1 to recently infected blood donors. AIDS 16:113-119.
Matter, L., K. Kogelschatz, and D. Germann. 1997. Serum levels of rubella virus antibodies indicating immunity: response to vaccination of subjects with low or undetectable antibody concentrations. J. Infect. Dis. 175:749-755.
McFarland, W., M. P. Busch, T. A. Kellogg, B. D. Rawal, G. A. Satten, M. H. Katz, J. Dilley, and R. S. Janssen. 1999. Detection of early HIV infection and estimation of incidence using a sensitive/less-sensitive enzyme immunoassay testing strategy at anonymous counseling and testing sites in San Francisco. J. Acquired Immune Defic. Syndr. 22:484-489.
Murphy, G., A. Charlett, L. F. Jordan, N. Osner, O. N. Gill, and J. V. Parry. 2004. HIV incidence appears constant in men who have sex with men despite widespread use of effective antiretroviral therapy. AIDS 18:265-272.
Murphy, G., J. V. Parry, S. B. Gupta, C. Graham, L. F. Jordan, A. N. Nicoll, and O. N. Gill. 2001. Test of HIV incidence shows continuing HIV transmission in homosexual/bisexual men in England and Wales. Commun. Dis. Public Health 4:33-37.
Rawal, B. D., A. Degula, L. Lebedeva, R. S. Janssen, F. M. Hecht, H. W. Sheppard, and M. P. Busch. 2003. Development of a new less-sensitive enzyme immunoassay for detection of early HIV-1 infection. J. Acquired Immune Defic. Syndr. 33:349-355.
Rutherford, G. W., S. K. Schwarcz, and W. McFarland. 2000. Surveillance for incident HIV infection: new technology and new opportunities. J. Acquired Immune Defic. Syndr. 25(Suppl 2):S115-S119.
Schwarcz, S., T. Kellogg, W. McFarland, B. Louie, R. Kohn, M. Busch, M. Katz, G. Bolan, J. Klausner, and H. Weinstock. 2001. Differences in the temporal trends of HIV seroincidence and seroprevalence among sexually transmitted disease clinic patients, 1989-1998: application of the serologic testing algorithm for recent HIV seroconversion. Am. J. Epidemiol. 153:925-934.
Schwarcz, S. K., T. A. Kellogg, W. McFarland, B. Louie, J. Klausner, D. G. Withum, and M. H. Katz. 2002. Characterization of sexually transmitted disease clinic patients with recent human immunodeficiency virus infection. J. Infect. Dis. 186:1019-1022.
Suligoi, B., V. Bossi, M. Massi, and C. Galli. 2004. Determinazione sierologica per l'identificazione delle infezioni recenti da HIV. STD News 22:6-9. (In Italian.)
Suligoi, B., C. Galli, M. Massi, F. Di Sora, M. Sciandra, P. Pezzotti, O. Recchia, F. Montella, A. Sinicco, and G. Rezza. 2002. Precision and accuracy of a procedure for detecting recent human immunodeficiency virus infections by calculating the antibody avidity index by an automated immunoassay-based method. J. Clin. Microbiol. 40:4015-4020.
Suligoi, B., M. Massi, C. Galli, M. Sciandra, F. Di Sora, P. Pezzotti, O. Recchia, F. Montella, A. Sinicco, and G. Rezza. 2003. Identifying recent HIV infections using the avidity index and an automated enzyme immunoassay. J. Acquired Immune Defic. Syndr. 32:424-428.
Young, C. L., D. J. Hu, R. Byers, S. Vanichseni, N. L. Young, R. Nelson, P. A. Mock, K. Choopanya, R. Janssen, T. D. Mastro, and J. V. Mei. 2003. Evaluation of a sensitive/less sensitive testing algorithm using the bioMerieux Vironostika-LS assay for detecting recent HIV-1 subtype B' or E infection in Thailand. AIDS Res. Hum. Retroviruses 19:481-486.(Elisa Martro, Barbara Sul)
Microbiology Service, Hospital Universitari Germans Trias i Pujol, Departament de Genetica i Microbiologia, Universitat Autònoma de Barcelona, Badalona, Spain
Department of Infectious Diseases, Istituto Superiore di Sanita, Rome, Italy
Laboratory of Virology, Ospedale Amedeo di Savoia, Turin, Italy
Newborn Screening Quality Assurance Program, Centers for Disease Control and Prevention, Atlanta, Georgia
ABSTRACT
A study was designed to compare an avidity index method to the serologic testing algorithm for recent human immunodeficiency virus (HIV) seroconversion (STARHS) for the detection of recent HIV infection. One hundred sixty HIV-positive sera were tested. Both techniques performed similarly in identifying recent infections, although STARHS tended to misclassify more individuals that had long-standing infection as being recently infected.
TEXT
Increasing interest in estimating the incidence of human immunodeficiency virus (HIV) infection by using cross-sectional specimens has led to the development of several laboratory methods. A sensitive/less-sensitive serologic strategy (the serologic testing algorithm for recent HIV seroconversion [STARHS]) was developed in 1998 (4) at the Centers for Disease Control and Prevention (CDC), Atlanta, Ga., to identify recently infected individuals and has been used in numerous studies (3, 9-11, 13-15). In Catalonia, Spain, STARHS has been introduced as part of enhanced surveillance of HIV and sexually transmitted infections since 2002 (1). Antibody avidity has long been used as a marker for recent infection (2, 5, 8), and in 2002, an automated technique was developed for HIV (17, 18) based on the rationale that in the early phase of infection antibodies show a low avidity for the antigen. In this study, we report the first comparison of these two methods for the detection of recent HIV infection.
A total of 160 serum specimens from HIV-positive subjects (identified according to standard HIV testing algorithms for screening and confirmatory testing) were included in the study. For 66 specimens, the seroconversion date or clinical stage of infection was not available. For 70 specimens, a seroconversion date could be estimated as the midpoint between the last HIV-negative test and the first HIV-positive test (interval between the two tests of <6 months). These specimens were sorted into two groups. Group 1 included 13 specimens from 12 recently infected individuals (sera drawn 6 months after the seroconversion date). Group 2 comprised 57 specimens from 26 individuals with long-standing infection (sera drawn >6 months after the diagnosis of HIV infection). Group 3 consisted of 24 individuals with a diagnosis of AIDS.
STARHS was implemented whereby all specimens were first tested using Biotest HIV-TETRA HIV 1 + 2 (Biotest; Diasorin Diagnostics S.R.L., Italy) as the sensitive enzyme-linked immunosorbent assay (ELISA). The Vironostika HIV-1 Microelisa System (bioMerieux, Durham, N.C.) was used as the less-sensitive ELISA after a 1:20,000 three-step dilution as previously described (12). Appropriate controls and calibrator provided by the CDC were used in each run. First, a screening assay using one well per specimen was performed to allow for the identification of probable recent seroconverters according to standardized optical density (SOD), calculated as (patient OD – negative control OD)/(calibrator OD – negative control OD). Specimens with an SOD of <2.000 were then retested in triplicate (confirmatory assay). Specimens with an SOD of <1.000 in the confirmatory assay were considered as belonging to subjects who had seroconverted within the previous 170 days (95% confidence interval, 145 to 200).
For the avidity method, two aliquots of each sample were subjected to a pretest 1:10 dilution. One aliquot was diluted in phosphate-buffered saline (PBS) and the other one in 1 M guanidine hydrochloride (G; Pierce Chemicals, Dallas, Tex.). After incubation at room temperature, both aliquots were tested by an automated microparticle immunoassay (AxSYM HIV 1/2 gO; Abbott Diagnostics Division, Delkenheim, Germany) as recommended by the manufacturer. Sample/cutoff (S/CO) ratios were calculated, and the avidity index of HIV antibodies was computed as (S/CO ratio of the G aliquot)/(S/CO ratio of the PBS aliquot). A cutoff of 0.85 was selected based on previous results (16).
The overall agreement between both techniques was good ( = 0.67, P < 0.001), where both techniques were concordant in classifying 25 specimens as recent infections and 117 as long-standing infections. Fourteen specimens were considered as recent infections by STARHS while 4 specimens were considered recent infections by the avidity index. Although the avidity technique showed better intra- and interday coefficients of variation (<10% versus 10 to 30%) (17), a number of specimens were close to the cutoff (Fig. 1). Results according to clinical information are shown in Table 1. Both techniques misclassified one out of 13 specimens corresponding to recent infections as being long-standing (the misclassified specimens were different from each other). Six specimens from long-standing infections were misclassified as recent infections by STARHS; one of them had been collected 7 months after the first HIV-positive test and was from a patient on highly active antiretroviral treatment (HAART), while the other five were obtained from subjects who had been infected with HIV for over a year. The only specimen misclassified by the avidity method had been collected 1 year after the first HIV-positive test. Another two specimens from subjects with long-standing infection were from subjects on HAART and were correctly identified as old infections by both techniques. Among the 24 AIDS patients, one was misclassified by STARHS only and another two were misclassified by both assays. Neither of the two tests seemed to be affected by a low CD4 count.
While STARHS has a proven value, several drawbacks limit its use. First, even small errors in the three-step specimen dilution magnify differences in SOD values, leading to a high variability among replicates and requiring additional testing (6). Secondly, a common calibrator and quality control material, along with a software package, were developed and must be provided by the CDC. Third, STARHS results vary according to HIV subtype, requiring the adoption of different cutoffs (19). Finally, problems with the availability of the commercialized test used for this technique have occurred in the past and will arise again in the near future.
The avidity index method shows several advantages over STARHS, as previously suggested (18). After a single 1:10 dilution, the technique can be fully automated using the AxSYM platform, which leads to better reproducibility. This instrument is available among clinical laboratories in Europe and in most countries worldwide. While STARHS uses a second-generation ELISA based on a virus lysate (HIV-1 subtype B), the avidity index assay is a third-generation enzyme immunoassay that is more sensitive and detects antibodies earlier (it is able to recognize all antibody classes). Moreover, this assay allows the detection of HIV-1, groups M and O, and HIV-2. In addition, the selection of different avidity index cutoffs offers different sensitivities and specificities, depending on the study purpose. Moreover, the avidity index method showed more accurate results than STARHS among patients with long-standing HIV infection or AIDS. In our study there were too few subjects on antiretroviral therapy to evaluate its effect on test performance. However, STARHS tends to misclassify the subjects on therapy as being recently infected, since long-term HAART may lead to a decrease in antibody titers (4, 7, 14). Recently, a longitudinal study that compared STARHS and the avidity index assay for patients who had started antiretroviral therapy (G. Murphy, A. Charlett, G. L. Dean, M. Fisher, D. McElborough, and J. V. Parry, Abstr. XV Int. AIDS Conf., abstr. C11390, 2004) concluded that this therapy had a marked effect on STARHS but had only minor effects on the avidity assay. The avidity index method is easily exportable and does not require any specific training other than that provided by the manufacturer. Finally, the avidity index method does not need official approval by the regulatory authority in any country because it is not a new assay or a modification of an already existing assay and only a preparation of the sample is needed.
It is appropriate to stress that currently the results of both of these tests cannot be interpreted at an individual level. The discrimination between recent and long-standing HIV infection is based on a cutoff that defines the time period from seroconversion to either high antibody titers (STARHS) or antibody maturation (avidity index). However, this time period may vary among individuals, causing misclassification in some cases. Only at the population level will the false-negative and false-positive results balance each other.
ACKNOWLEDGMENTS
This work was partially supported by a grant from the Istituto Superiore di Sanita, V Progetto Nazionale di ricerca sull'AIDS 2003, and grant from Fundacio Barcelona SIDA 2002.
We thank Rafael Muoz for management and storage of specimens and Meritxell Granell for the revision of medical charts. We also thank Robert H. Byers for helpful discussion. Joanne V. Mei's role in this project was limited to providing technical assistance and advice.
The Recent HIV Infections Study Group consists of Diana Puente, Claudio Galli, M. Jesús Casado, Daniela Bernasconi, Mauro Sciandra, Laura Camoni, Lurdes Matas, Giovanni Rezza, Vicen Ausina, and Jordi Casabona.
REFERENCES
Center for Epidemiological Studies on HIV/AIDS of Catalonia (CEESCAT). 2005. SIVES 2004, Integrated HIV/STI Surveillance System of Catalonia (SIVES) Annual Report. Generalitat de Catalunya, Departament de Salut, Barcelona, Spain.
Eggers, M., U. Bader, and G. Enders. 2000. Combination of microneutralization and avidity assays: improved diagnosis of recent primary human cytomegalovirus infection in single serum sample of second trimester pregnancy. J. Med. Virol. 60:324-330.
Gouws, E., B. G. Williams, H. W. Sheppard, B. Enge, and S. A. Karim. 2002. High incidence of HIV-1 in South Africa using a standardized algorithm for recent HIV seroconversion. J. Acquired Immune Defic. Syndr. 29:531-535.
Janssen, R. S., G. A. Satten, S. L. Stramer, B. D. Rawal, T. R. O'Brien, B. J. Weiblen, F. M. Hecht, N. Jack, F. R. Cleghorn, J. O. Kahn, M. A. Chesney, and M. P. Busch. 1998. New testing strategy to detect early HIV-1 infection for use in incidence estimates and for clinical and prevention purposes. JAMA 280: 42-48.
Jenum, P. A., B. Stray-Pedersen, and A. G. Gundersen. 1997. Improved diagnosis of primary Toxoplasma gondii infection in early pregnancy by determination of antitoxoplasma immunoglobulin G avidity. J. Clin. Microbiol. 35:1972-1977.
Kothe, D., R. H. Byers, S. P. Caudill, G. A. Satten, R. S. Janssen, W. H. Hannon, and J. V. Mei. 2003. Performance characteristics of a new less sensitive HIV-1 enzyme immunoassay for use in estimating HIV seroincidence. J. Acquired Immune Defic. Syndr. 33:625-634.
Machado, D. M., E. L. Delwart, R. S. Diaz, C. F. de Oliveira, K. Alves, B. D. Rawal, M. Sullivan, M. Gwinn, K. A. Clark, and M. P. Busch. 2002. Use of the sensitive/less-sensitive (detuned) EIA strategy for targeting genetic analysis of HIV-1 to recently infected blood donors. AIDS 16:113-119.
Matter, L., K. Kogelschatz, and D. Germann. 1997. Serum levels of rubella virus antibodies indicating immunity: response to vaccination of subjects with low or undetectable antibody concentrations. J. Infect. Dis. 175:749-755.
McFarland, W., M. P. Busch, T. A. Kellogg, B. D. Rawal, G. A. Satten, M. H. Katz, J. Dilley, and R. S. Janssen. 1999. Detection of early HIV infection and estimation of incidence using a sensitive/less-sensitive enzyme immunoassay testing strategy at anonymous counseling and testing sites in San Francisco. J. Acquired Immune Defic. Syndr. 22:484-489.
Murphy, G., A. Charlett, L. F. Jordan, N. Osner, O. N. Gill, and J. V. Parry. 2004. HIV incidence appears constant in men who have sex with men despite widespread use of effective antiretroviral therapy. AIDS 18:265-272.
Murphy, G., J. V. Parry, S. B. Gupta, C. Graham, L. F. Jordan, A. N. Nicoll, and O. N. Gill. 2001. Test of HIV incidence shows continuing HIV transmission in homosexual/bisexual men in England and Wales. Commun. Dis. Public Health 4:33-37.
Rawal, B. D., A. Degula, L. Lebedeva, R. S. Janssen, F. M. Hecht, H. W. Sheppard, and M. P. Busch. 2003. Development of a new less-sensitive enzyme immunoassay for detection of early HIV-1 infection. J. Acquired Immune Defic. Syndr. 33:349-355.
Rutherford, G. W., S. K. Schwarcz, and W. McFarland. 2000. Surveillance for incident HIV infection: new technology and new opportunities. J. Acquired Immune Defic. Syndr. 25(Suppl 2):S115-S119.
Schwarcz, S., T. Kellogg, W. McFarland, B. Louie, R. Kohn, M. Busch, M. Katz, G. Bolan, J. Klausner, and H. Weinstock. 2001. Differences in the temporal trends of HIV seroincidence and seroprevalence among sexually transmitted disease clinic patients, 1989-1998: application of the serologic testing algorithm for recent HIV seroconversion. Am. J. Epidemiol. 153:925-934.
Schwarcz, S. K., T. A. Kellogg, W. McFarland, B. Louie, J. Klausner, D. G. Withum, and M. H. Katz. 2002. Characterization of sexually transmitted disease clinic patients with recent human immunodeficiency virus infection. J. Infect. Dis. 186:1019-1022.
Suligoi, B., V. Bossi, M. Massi, and C. Galli. 2004. Determinazione sierologica per l'identificazione delle infezioni recenti da HIV. STD News 22:6-9. (In Italian.)
Suligoi, B., C. Galli, M. Massi, F. Di Sora, M. Sciandra, P. Pezzotti, O. Recchia, F. Montella, A. Sinicco, and G. Rezza. 2002. Precision and accuracy of a procedure for detecting recent human immunodeficiency virus infections by calculating the antibody avidity index by an automated immunoassay-based method. J. Clin. Microbiol. 40:4015-4020.
Suligoi, B., M. Massi, C. Galli, M. Sciandra, F. Di Sora, P. Pezzotti, O. Recchia, F. Montella, A. Sinicco, and G. Rezza. 2003. Identifying recent HIV infections using the avidity index and an automated enzyme immunoassay. J. Acquired Immune Defic. Syndr. 32:424-428.
Young, C. L., D. J. Hu, R. Byers, S. Vanichseni, N. L. Young, R. Nelson, P. A. Mock, K. Choopanya, R. Janssen, T. D. Mastro, and J. V. Mei. 2003. Evaluation of a sensitive/less sensitive testing algorithm using the bioMerieux Vironostika-LS assay for detecting recent HIV-1 subtype B' or E infection in Thailand. AIDS Res. Hum. Retroviruses 19:481-486.(Elisa Martro, Barbara Sul)