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Journal of Clinical Microbiology, June 2000, p. 2459-2461, Vol. 38, No. 6
0095-1137/00/$04.00+0
LETTERS TO THE EDITOR
Contribution of Combined Detection Assays of p24 Antigen and
Anti-Human Immunodeficiency Virus (HIV) Antibodies in Diagnosis of
Primary HIV Infection by Routine Testing
 |
LETTER |
In reference to the paper of Weber et al. (1), we
report here the contribution of a fourth-generation human
immunodeficiency virus (HIV) diagnosis assay, the VIDAS HIV DUO
(bioMérieux), to diagnosis of primary HIV infection earlier than
the third generation.
During a recent 9-month survey, from April to December 1998, a study
was conducted in five French public and private laboratories that had
reported diagnosis of primary HIV infections. In France, the use of two
different HIV screening assays is mandatory for testing for
non-blood-donation HIV. In four of the five laboratories, part of the
routine testing was done for an anonymous high-risk population. A total
of 29,657 consecutive and unselected samples were tested for the
presence of anti-HIV antibodies systematically with the VIDAS HIV DUO
plus one of the following third-generation detection assays: Ortho
Capture HIV-1/HIV-2 (Ortho Diagnostic Systems), Genscreen HIV1/2
(Sanofi Diagnostics Pasteur), Axsym HIV-1/HIV-2 (Abbott), Vironostika
HIV Uniform II plus 0 v 3.3 (Organon Teknika), or COBAS Core
anti-HIV-1/HIV-2 EIA DAGS (Roche Diagnostics).
Of the 29,657 samples tested, 29,002 were found to be negative by both
the VIDAS HIV DUO and a third-generation assay. A total of 453 samples
were reactive with both assays. Of these 453 samples, 449 were
confirmed to be positive by Western blotting (New Lav Blot I and II;
Sanofi Diagnostics Pasteur) and four were found to be negative by the
same technology. Follow-up samples were reactive with both the VIDAS
HIV DUO and Western blotting, suggesting that the patients from whom
these samples came were infected at the time the initial samples were
collected. The positive prevalence rate of this tested population was
1.52%.
With the VIDAS HIV DUO, 160 samples were found to be repeatedly
reactive only by this assay. Both an HIV antigen assay and Western
blotting were performed on these 160 samples. All 160 samples were
shown to be negative by Western blotting. Seventeen of the 160 samples
were confirmed to show primary HIV infection with a positive p24
antigen test. In addition, the HIV type 1 (HIV-1) RNA viral load was
determined for 11 of the 17 samples and the samples were also found to
be positive. Finally, HIV infection was confirmed for all 17 patients
by determining complete Western blot profiles on follow-up samples.
These additional tests allowed us to confirm that the 17 patients who
were negative, by both a third-generation assay and Western blotting,
were positive with the VIDAS HIV DUO due to a primary HIV infection.
With the third-generation tests, there were 42 samples that were found
to be repeatedly reactive only with these assays. Western blotting was
performed for these 42 samples, and none was confirmed to be positive
when follow-up samples were used.
For comparison of the sensitivity and specificity of the VIDAS HIV
DUO with those of the third-generation tests, samples were considered
to be either HIV positive or negative according to the following
decision trees. Samples were considered positive if (i) at least one
screening assay was positively reactive, and it was confirmed by
Western blotting; (ii) if the Western blot was negative, then a p24
antigen test was performed (only in the case of positive samples for
the VIDAS HIV DUO); (iii) if either the p24 antigen and/or RNA assay
was positive, then Western blotting was performed on a follow-up
sample; (iv) in all cases, there was a final positive result by Western
blotting. Samples were considered negative (i) if both screening assays
were negative or (ii) a negative confirmation result followed an enzyme
immunoassay-positive reaction using Western blotting and/or a p24
antigen test.
When both the sensitivity and specificity of the VIDAS HIV DUO were
compared with those of the third-generation tests, there were some
differences to be noted. According to the protocol of the study
(referred above), the sensitivity of the VIDAS HIV DUO was shown to be
100%, which was statistically significantly higher than the 96.4% of
the third-generation tests (McNemar test P value < 0.01). Therefore, the use of this assay enabled the diagnosis of
infection in 3.6% additional HIV-infected patients. These results agree with the data of Weber et al. (1), demonstrating
earlier diagnosis of HIV infection by fourth-generation assays. In
contrast, the specificity of the third-generation tests were more
specific at 99.86% and statistically significant when compared with
the 99.51% specificity of the VIDAS HIV DUO (McNemar test
P value < 0.01).
We gathered the clinical data of these 17 patients to draw a picture
corresponding to various cases of primary HIV infection (Table
1). One of the patients showed no
symptoms because he was tested for HIV in the context of a presurgical
procedure. In this case, the VIDAS HIV DUO detected a completely
unexpected primary HIV infection that would have been missed with the
third-generation tests. Primary HIV infection was suspected in 8 of
these 17 patients; therefore, a p24 antigen detection assay was
originally prescribed. The remaining nine patients were later diagnosed
as HIV infected by using a p24 antigen test due to the repeatedly
positive results of the VIDAS HIV DUO test.
In conclusion, these results clearly demonstrate that by using the most
up-to-date diagnostic tools, such as these fourth-generation tests, we
can reduce the risk of missing any unsuspected primary HIV infections
in routine testing. Since the viral load is very high during primary
HIV infection, we now have the capability of diagnosing the HIV
infection earlier, allowing for the administration of antiretroviral
therapy as early as possible. This earlier diagnosis gives us the
possibility of increasing the efficiency of the treatment and
especially helps to decrease the risk of HIV transmission.
| |
REFERENCE |
| 1.
|
Weber, B.,
E. H. Mbargane Fall,
A.-M. Berger, and H. W. Doerr.
1998.
Reduction of diagnostic window by new fourth-generation human immunodeficiency virus screening assays.
J. Clin. Microbiol.
36:2235-2239[Abstract/Free Full Text].
|
| | | | |
Thoai Doung Ly
Institut Alfred Fournier
Paris, France
|
| | | | |
Catherine Edlinger
Hôpital Bichat-Claude Bernard
Paris, France
|
| | | | |
Astrid Vabret
Hôpital de Caen
Caen, France
|
| | | | |
Odile Morvan
Hôpital La Beauchée
St Brieuc,
France
|
| | | | |
Béryl Greuet
Laboratoire
d'Analyses
Medicales Greuet-Dessus
Neuilly-Plaisance, France
|
| |
AUTHOR'S REPLY |
Ly et al. report the results of a study which was performed in five
French laboratories with the aim of comparing the performance of a new
fourth-generation HIV assay for the combined detection of antigen and
antibody to that of diverse third-generation assays under routine
conditions. The new assay showed a higher sensitivity for the diagnosis
of primary HIV infection in comparison to that of conventional tests in
urban centers with high HIV incidence and prevalence.
Actually, four different assays for combined antigen and antibody
detection are available for laboratory diagnosis and blood donor
screening for HIV in Europe. Although this new assay generation represents a major improvement in terms of sensitivity in comparison to
the former generation through a mean reduction of 4 days in the
diagnostic window, optimization of the performance characteristics is
required for the following reasons.
(i) Combined assays for antigen and antibody detection cannot replace
single antigen tests for blood donor screening. The detection limit of
fourth-generation assays is higher (20 to >100 pg of antigen p24/ml)
than that of antigen assays (3.5 to 15 pg of p24 antigen/ml). (iii)
Highly sensitive antigen assays detect primary infection on average 1 to 2 days earlier than fourth-generation enzyme immunoassays (EIA)
(5).
(ii) The antigen detection module of fourth-generation assays shows a
variable sensitivity for detection of different HIV-1 non-B subtypes,
including group O and HIV-2 (1). Some assays may fail to
detect low levels of antigen of HIV-1 non-B subtype strains, although
monoclonal antibody is directed against conserved epitopes of p24
antigen (unpublished results). Since the genetic diversity of HIV is
rapidly increasing worldwide, including in industrialized countries,
fourth-generation assays need to be optimized in order to detect
accurately all HIV-1 subtypes and HIV-2.
(iii) A further potential risk for impaired sensitivity is that a more
limited area of the solid phase can be used for antibody detection since about one-third of the binding sites are occupied by anti-p24 antibody for HIV antigen detection; the antibody
detection module may be less sensitive than single third-generation
antibody assays. Antibody detection may be delayed in seroconversion
panels without antigenemia, and a second diagnostic window may be
observed in the early seroconversion phase when low antibody titers are present and antigenemia declines (1).
(iv) Since fourth-generation EIAs combine two different test principles
in one assay, the potential risk for nonspecific reactivity may be
higher than for second- and third-generation antibody assays. The rate
of false-positive results obtained with blood donors and interfering
samples varies from 0.3 to 0.8% (in comparison to a maximum of 0.2%
for third-generation EIAs), depending on the donor background
(1).
Fourth-generation assays demand a special algorithm for the analysis of
reactive samples. For the anti-HIV part of the assay, confirmation of
reactivity should be done first with an assay that lacks the p24
antigen detection module and when reactivity persists subsequently, as
determined by immunoblotting. For the p24 antigen part, confirmation of
reactivity should be analyzed by an assay that lacks the anti-HIV
detection part and when reactivity persists, as determined by a nucleic
acid-based assay. Confirmation of this part of reactivity is hampered
by the fact that actually none of the commercially available nucleic
acid-based assays is able to detect HIV-1 subtype O and HIV-2 genome.
Finally, the most efficient way to reduce the diagnostic window will be
in future screening of blood donations by nucleic acid amplification
techniques since HIV RNA is detected about 11 days before antibody to
HIV is (4) and, in the chimpanzee model, no demonstrable
infectivity in either plasma or peripheral blood mononuclear cells is
obtained before molecular markers are detectable (2).
Recently, Roth et al. (3) demonstrated the feasibility and
efficacy of routine PCR screening of pooled blood donations for HIV-1
in a blood bank setting. However, for efficient nucleic acid
amplification testing under routine conditions, assays that are not
affected by genetic variability of HIV and that include rapid automated
nucleic acid extraction procedures need to be developed.
| |
REFERENCES |
| 1.
|
Gürtler, L.,
A. Mühlbacher,
U. Michl,
H. Hofmann,
G. G. Paggi,
V. Bossi,
R. Thorstensson,
R. G. Villaescusa,
A. Eiras,
J. H. Hernandez, and B. Weber.
1998.
Reduction of the diagnostic window with a new combined p24 antigen and human immunodeficiency virus antibody screening assay.
J. Virol. Methods
75:27-38[CrossRef][Medline].
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Murthy, K. K.,
D. R. Henrard,
J. W. Eichberg,
K. E. Cobb,
M. P. Busch,
J. P. Allain, and H. J. Alter.
1999.
Redefining the HIV-infectious window period in the chimpanzee model: evidence to suggest that viral nucleic acid testing can prevent blood-borne transmission.
Transfusion
39:688-693[CrossRef][Medline].
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|
Roth, W. K.,
H. Weber, and E. Seifried.
1999.
Feasibility and efficacy of routine PCR screening of blood donations for hepatitis C virus, hepatitis B virus, and HIV-1 in a blood bank setting.
Lancet
353:359-363[CrossRef][Medline].
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Schreiber, G. B.,
M. P. Busch,
S. H. Kleinman, and J. J. Koreliz for the Retrovirus Epidemiology Donor Study.
1996.
The risk of transfusion-transmitted viral infections.
N. Engl. J. Med.
334:1685-1690[Abstract/Free Full Text].
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Weber, B.,
A. Mühlbacher,
U. Michl,
G. Paggi,
V. Bossi,
C. Sargento,
R. Camacho,
F. M. B. Fall,
A. Berger,
U. Schmitt, and W. Melchior.
1999.
Multicenter evaluation of a new rapid automated HIV antigen detection assay.
J. Virol. Methods
78:61-70[CrossRef][Medline].
|
| | | | |
Bernard Weber
Laboratoires Réunis Kutter-Lieners-Hastert
Centre Langwies
L-6131 Junglinster, Germany
Phone: (352) 78 02 90 1
Fax: (352) 78 88 94
E-mail: labor klh{at}pt.lu.
|
Journal of Clinical Microbiology, June 2000, p. 2459-2461, Vol. 38, No. 6
0095-1137/00/$04.00+0
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