Objectives Transmitted HIV strains may harbour drug resistance mutations. HIV‐1 drug resistance mutations are currently detected in plasma viral RNA. HIV‐1 proviral DNA could be an alternative marker, as it persists in infected cells. Methods This was a prospective study assessing the prevalence and persistence of HIV‐1 drug resistance mutations in DNA from CD4 cells before and after protease inhibitor (PI)‐ or nonnucleoside reverse transcriptase inhibitor (NNRTI)‐based therapy initiation in 69 drug‐naïve patients. Results Before therapy, 90 and 66% of detected mutations were present in CD4 cells and plasma, respectively. We detected seven key mutations, and four of these (M184M/V, M184M/I, K103K/N and M46M/I) were only found in the cells. When treatment was started, 40 patients were followed; the mutations detected at the naïve stage remained present for at least 1 year. Under successful treatment, new key mutations emerged in CD4 cells (M184I, M184M/I and Y188Y/H). Conclusions The proportion of mutations detected in the DNA was statistically significantly higher than that detected in standard RNA genotyping, and these mutations persisted for at least 1 year irrespective of therapy. The pre‐existence of resistance mutations did not jeopardise treatment outcome when the drug concerned was not included in the regimen. Analysis of HIV‐1 DNA could be useful in chronic infections or when switching therapy in patients with undetectable viraemia.

ABSTRACT A panel of new cytomegalovirus (CMV) assays for use on the Architect instrument has been developed, including a CMV avidity assay based on a new technology. The purpose of this study was to compare the performance characteristics of the fully automated CMV immunoglobulin M (IgM), IgG, and IgG avidity tests on the Architect instrument with those of other available assays. A total of 503 consecutive fresh patient serum specimens (routine serum specimens) and 96 serum specimens from 33 pregnant women with a recent CMV primary infection (seroconversion serum specimens) were tested for CMV IgM and IgG by the Architect (Abbott), Vidas (BioMérieux), and Enzygnost (Siemens) assays. The seroconversion sera and 100 preselected serum specimens IgM negative and IgG positive by the AxSYM assay were also tested by the IgG avidity tests on the Architect and Vidas instruments. The relative agreements for CMV IgM determination with routine sera between the Architect assay and the Vidas, Enzygnost, and AxSYM assays were 97%, 94%, and 93%, respectively, for the CMV IgM tests and 99%, 98%, and 98%, respectively, for the CMV IgG tests. The specificities of the CMV IgG avidity test were 98% for the Architect assay and 76% for the Vidas assay. No high CMV IgG avidity test results were found within the first 3 months after seroconversion by either of those assays. The correlation between the results of the newly developed CMV IgM and IgG tests on the Architect instrument with the Vidas and Enzygnost assays was excellent (≥94%). The CMV IgG avidity test reliably excluded patients with recent infections and showed an excellent specificity (98%).

BACKGROUND:pp65 antigenaemia and real-time PCR are two methods that are used to diagnose CMV infection in its early stages and, thereby, to facilitate initiation of pre-emptive therapy.OBJECTIVES:Firstly, to compare PCR with antigenaemia and clinical outcome in order to define a clinical threshold for starting pre-emptive therapy. Secondly, to study the impact of the transplant recipient's serological status on the viral load and on the cut-offs.STUDY DESIGN:Sixty-two patients were analysed using antigenaemia (APAAP method) and real-time PCR. ROC curves were established with antigenaemia or clinical outcome as reference. Patients were divided into primo-infection or reactivation on the basis of the serological status.RESULTS:PCR correlated better with the clinical data (AUC closer to 1 and best sensitivity, PPV and NPV) than antigenaemia. Furthermore, the performance of qPCR was even better in the reactivation patients.CONCLUSIONS:This work suggests that transplant recipients should be divided according to their serological status. Indeed, replacing antigenaemia by real-time PCR for decisions regarding initiation of pre-emptive therapy is of particular appeal in patients with positive serology. As a result of this work, we have set our clinical threshold at 1500 copies/ml for reactivation.

Le JC virus (JCV) peut être responsable de leucoencéphalopathie multifocale progressive chez le sujet infecté par le VIH, alors que le BK virus (BKV) peut induire des cystites hémorragiques et des néphropathies avec risque de rejet de greffe chez les transplantés rénaux. L'étude a consisté à évaluer la PCR en temps réel publiée par notre laboratoire (Sehbani et al., 2006) pour la quantification plasmatique de BKV et JCV chez des patients greffés rénaux, en confrontant les résultats obtenus à la cytologie (decoy cells), l'histologie (immunoperoxydase) et la clinique, dans le cadre de présomption de rejet de greffe. Nous avons également recherché l'impact de JCV dans les néphropathies des transplantés rénaux. Les résultats de PCR, cytologie et histologie de 256 échantillons d'une cohorte de 119 patients greffés rénaux susceptibles de faire un rejet de greffe, ont été analysés et comparés sur une période de 2 ans. La PCR JCV était faiblement positive pour un seul sérum (< 1 250 copies/ml). Sur 8 patients présentant une ou plusieurs PCR BKV positives et une clinique évoquant un rejet du greffon, 7 avaient au moins un résultat cytologique concordant. 3 d'entre eux (sur les 5 ayant pu bénéficier d'une immunohistochimie) avaient parallèlement une immunoperoxydase positive. JCV ne semblait pas être responsable de néphropathies chez les transplantés rénaux. Les résultats de PCR plasmatique et cytologie urinaire étaient concordants en début de suivi post-greffe. La charge virale fluctuait de façon importante et reste longtemps positive, alors que la cytologie se négativait plus rapidement. La fréquence de PCR plasmatique dans le suivi post-greffe reste à établir.

The use of livers from anti-hepatitis B core (HBc) positive donors can alleviate donor shortage. Nineteen of 367 (6%) adults receiving anti-HBc positive allografts [three were hepatitis B antigen (HBsAg) negative, hepatitis B antibody (HBsAb) positive; four were HBsAg positive and 12 were not exposed to hepatitis B viral (HBV) infection] were retrospectively reviewed. In HBsAg negative recipients, immunoprophylaxis (IP) was guided by viral serology and immunohistochemistry (IH) of day 0 and day 7 liver biopsies. If IH was negative, IP was stopped. None of three HBsAg negative, HBsAb positive recipients infected; one (replicating) of four HBsAg positive recipients reinfected and seven of eight (87.5%) HBsAg, HBsAb negative recipients, who did not receive long-term IP, infected after a median time of 2 years (range 1-5); one patient died of liver failure. Four HBsAg, HBsAb negative recipients, receiving life-long IP, remained infection free. Anti-HBc positive donor livers must be directed selectively first to HBsAg positive recipients, next to recipients having HBV antibodies and finally to HBV-naive recipients. Identification of both donor and recipient risk factors for HBV infection before transplantation allows indiscriminate use of antiviral prophylaxis. The necessity for IP therapy should be guided by HBV-DNA testing of donor liver tissue and serum. IH of early liver biopsies is an unreliable marker for predicting antiviral treatment requirements.

We report an outbreak of gastroenteritis due to Norovirus in a care unit in a Belgian hospital involving thirty-three people. The origin of the outbreak was traced to one nursing assistant. The virus strain identified by reverse transcription polymerase chain reaction and electron microscopy belonged to the genogroup II.

Objective: The objective of this study was to study the prevalence of herpes simplex virus (HSV) type 2 in pregnant women in Belgium. Study Design: The serum of 1000 consecutive women was collected. HSV-1 and HSV-2 control sera were added to the study. HSV-2 antibodies were tested with the HerpeSelect 2 enzyme-linked immunosorbent assay (ELISA; Focus) based on the use of the recombinant gG-2 antigen. Results: The 21 HSV-2 control subjects were positive. Among the HSV-1 control subjects, 18 were negative and 4 were positive. Among the pregnant women, 80.3% were negative, 1.5% had equivocal results, and 18.2% were positive. No statistical difference was observed according to the origin (European or African) of the women. Conclusions: The results obtained with the control sera indicate a high sensitivity of the Focus ELISA, as well as a capacity to discriminate between HSV-1/HSV-2 infection. The HSV-2 prevalence in the studied population raises the question of the possible benefit of a specific preventive program in pregnant women.

BACKGROUND:Up to 20 million persons are infected with the human retroviruses human T-lymphotropic virus (HTLV)-I and HTLV-II globally. Most data on the seroprevalence of HTLV-I and HTLV-II in Europe are from studies of low-risk blood donors or high-risk injection drug users (IDUs). Little is known about the general population.METHODS:A prospective anonymous study of HTLV-I and HTLV-II seroprevalence among 234,078 pregnant women in Belgium, France, Germany, Italy, Portugal, Spain, and the United Kingdom was conducted. Maternal antibody status was determined by standard methods using sera obtained for routine antenatal infection screens or eluted from infant heel prick dried blood spots obtained for routine neonatal metabolic screens.RESULTS:Anti-HTLV-I/II antibodies were detected and confirmed in 96 pregnant women (4.4 per 10,000, 95% confidence interval [CI]: 3.5-5.2). Of these, 73 were anti-HTLV-I, 17 were anti-HTLV-II, and 6 were specifically anti-HTLV but untyped. The seroprevalence ranged from 0.7 per 10,000 in Germany to 11.5 per 10,000 in France.CONCLUSIONS:Pregnant women better reflect the general population than blood donors or IDUs. The seroprevalence of HTLV-I and HTLV-II in Western Europe is 6-fold higher among pregnant women (4.4 per 10,000) than among blood donors (0.07 per 10,000). These data provide a robust baseline against which changes in HTLV-I and HTLV-II seroprevalence in Europe can be measured.

L’infection à CMV est l’infection congénitale la plus fréquente, avec une incidence moyenne de 1 %. L’infection peut se transmettre in utero lors d’une primo-infection maternelle ou lors d’une réactivation de l’infection chez une mère séropositive avant grossesse. La fréquence et la gravité de l’infection congénitale sont très différentes suivant le cas et il est donc essentiel de pouvoir faire le diagnostic différentiel entre infection primaire et réactivation. Dans ce contexte, il n’existe aucun test de référence et la présence d’IgM est encore trop souvent considérée comme un critère d’infection récente. Différentes techniques ont été développées afin d’améliorer le diagnostic. Parmi ces nouvelles approches, la plus utilisée est la mesure de l’avidité des IgG qui permet d’exclure une infection récente dans bon nombre de cas. Si le risque est élevé pour le fœtus (séroconversion maternelle ou IgG de faible avidité) le diagnostic de l’infection congénitale est réalisé par la recherche de CMV dans le liquide amniotique en culture et/ou en PCR ; les performances de ces 2 techniques en termes de sensibilité et de spécificité sont comparables. Il faut cependant garder à l’esprit, que si la mise en évidence de virus dans le liquide amniotique signe une infection congénitale, elle ne permet pas d’en évaluer la gravité.

The serologically negative window period at the time of HIV seroconversion may be reduced by HIV p24 antigen or viral nucleic acid assays. Recently combined antigen/antibody assays have been developed in the enzyme-linked immunosorbent assay format [1]. A 32-year-old man was admitted with febrile pharyngitis. He had been in a stable homosexual relationship for 9 years, and was subjected to piercing during a trip to Greece. As shown in Table 1, the routine detection of anti-HIV-1 and anti-HIV-2 antibodies with Axsym HIV 1/2 g O (Abbott, Illinois, USA) was negative on the first specimen. The tests performed 3 days later with two combined antigen/antibody assays are also shown in Table 1 : the Genscreen plus HIV antigen/antibody (Ag/Ab) (Bio-Rad, Marnes la Coquette, France) was positive with low reactivity, whereas the Vironostika Uni-form HIV Ag/Ab (Organon Teknika, Fresnes, France) was negative. The sample was not confirmed positive by Western blotting. A primary HIV infection was confirmed by testing the HIV-1-RNA viral load (HIV-1 Amplicor Monitor 1.5, Roche, Branchburg, NJ, USA) at 678 000 copies/ml. HIV-1 proviral polymerase chain reaction was also positive in the gag, pol and env genes.Table 1: Results of HIV seroconversion and antigen detection. We tested the consecutive samples from this HIV-1 seroconversion case with three HIV antigen/antibodies combined screening assays and one antibody assay. The three combined assays were Genscreen plus HIV Ag/Ab (Bio-Rad), and Vironostika Uni-form HIV Ag/Ab (Organon Teknika) used for routine HIV diagnostic screening, and the Vidas HIV Duo (BioMérieux, Lyon, France). The antibody assay was the Axsym HIV 1/2 gO assay (Abbott). HIV-1 p24 antigen was also assayed using the Genetic Systems HIV-1 antigen enzyme immunoassay (Bio-Rad). We compared the HIV-1 p24 antigen limit of detection of the three combined assays and the single antigen assay with three dilutions of HIV-1 p24 antigen National Institute for Biological Standards and Control (NIBSC) reference (1000 arbitrary units; AU) (Table 1). The HIV-1 p24 antigen/anti-HIV antibodies combined assays theoretically offer an earlier diagnosis of HIV infection than assays based only on the detection of specific antibodies, even if the yield in terms of a reduction of the window period may vary. Surprisingly in this seroconversion case, the Axsym antibodies assay was positive 3 days after the first sample, whereas the Vironostika combined assay remained negative. The other two combined assays were already reactive with the first sample. Looking at the results with the HIV-1 p24 antigen NIBSC reference dilutions (Table 1), the Genetic Systems immunoassay remained positive with all dilutions. The Vidas Duo assay seems to be the more sensitive combined assay for antigen detection because it remained positive with HIV-1 p24 antigen diluted 200 times. The Vironostika Ag/Ab assay and the Genscreen Ag/Ab assay did not detect the HIV-1 p24 antigen diluted 200 times. The observed HIV-1 p24 antigen sensitivity is approximately 100 or 30 pg/ml for Vironostika Uni-form HIV Ag/Ab with or without 1 h of agitation, respectively [2], 140 pg/ml for Genscreen plus HIV Ag/Ab, 18 pg/ml for Vidas HIV Duo and 8 pg/ml for Genetic Systems HIV-1 antigen enzyme immunoassay (manufacturer's package inserts). A second similar case was recently observed in our laboratory: we received a serum sample, positive using the Axsym HIV 1/2 g O and Vidas HIV Duo. In our hands the Vidas HIV Duo remained positive but the Vironostika Uni-form HIV Ag/Ab was negative on the same specimen. A diagnosis of primary HIV infection was confirmed by a high viral load. Combined assays appear less sensitive than HIV p24 antigen tests for the detection of this marker, and our results indicate that they may also be less sensitive than single assays for HIV antibodies, particularly in seroconversion cases. Combining the detection of both markers seems to lead to less sensitive assays than single antibody or antigen assays [3]. Combined assays are theoretically capable of reducing the diagnostic window at the start of infection, but their differences in sensitivity necessitate individual evaluation of these tests for both their antigen and their antibody component. Benoît Kabamba Mukadia Bernard Vandercamb Monique Bodeusa Michel Moreaub Patrick Goubaua

ABSTRACT Fetal damage following cytomegalovirus (CMV) intrauterine infection is mostly linked to primary infection. To differentiate primary infection from nonprimary infection, immunoglobulin M (IgM) tests are not reliable enough, and measurement of the IgG avidity appears to be the method that is the most widely used at present. In the present study the performance of the Vidas (bioMérieux) avidity assay was compared with that of a new enzyme immunoassay based on the use of a recombinant CMV glycoprotein B protein (Biotest).

Objectives: To analyze the practical use of the anticytomegalovirus IgG avidity and its impact on the follow-up of pregnancy. To evaluate the performance of IgG avidity to exclude the risk of congenital infection. Methods: 409 IgM-positive women without a documented seroconversion were prospectively followed. Data concerning the follow-up of the pregnancies were collected (amniotic fluid puncture and samples from the offspring). These observations were compared to those of 76 seroconversions during the same period. Results: High avidity excluding a primary infection within the past 3 months was observed in 270 women. As the gestational age was less than 3 months for 121 women, exclusion of a primary infection was achieved in 30% of the cases. The rate of amniotic fluid puncture was influenced by the serological result: high avidity (9%), low avidity (42%) and seroconversion (65%). Conclusions: A high avidity index during the first trimester of pregnancy could reasonably be considered as a good indicator of past infection and invasive prenatal diagnosis is not necessary. Nearly 70% of the IgM-positive women could be reassured if the first serology was systematically performed before 12 weeks of gestation.

1 A.W. Liley Medal 2001 Awarded to Prof. Wolfgang Holzgreve Chervenak, F.A. (New York, N.Y.) 3 Prenatal and Perinatal Aspects of a Giant Fetal Cervicothoracal Lymphangioma Axt-Fliedner, R.; Hendrik, H.J.; Schwaiger, C.; Ertan, A.K.; Friedrich, M.; Schmidt, W. (Homburg/Saar) 8 Gelatin Sponge Embolization. A Method for the Management of Iatrogenic Preterm Premature Rupture of the Membranes O’Brien, J.M.; Milligan, D.A.; Barton, J.R. (Lexington, Ky.) 11 Diagnostic and Therapeutic Dilemma with Large Prenatally Detected Cystic Adrenal Masses de Luca, J.L.; Rousseau, T.; Durand, C.; Sagot, P.; Sapin, E. (Dijon) 17 The Influence of Smoking and Parity on Serum Markers for Down’s Syndrome Screening Tišlarić, D. (Zagreb); Brajenović-Milić, B.; Ristić, S. (Rijeka); Latin, V. (Zagreb); ZZ uvić-Butorac, M.; BacZ ić, J. (Rijeka); Petek, M. (Zagreb); Kapović, M. (Rijeka) 22 Erythropoietin as Treatment for Late Hyporegenerative Anemia in Neonates with Rh Hemolytic Disease after in utero Exchange Transfusion Nicaise, C.; Gire, C.; Casha, P.; d’Ercole, C.; Chau, C.; Palix, C. (Marseille) 25 Alpha-Fetoprotein Values in Amniotic Fluid Obtained during Early Amniocentesis (11–13 Weeks) Nwebube, N.I.; Lockitch, G.; Halstead, C. (Vancouver); Johnson, J. (Toronto); Wilson, R.D. (Vancouver) 29 Prenatal Diagnosis of Arthrogryposis multiplex congenita with Increased Nuchal Translucency but without Any Underlying Fetal Neurogenic or Myogenic Pathology Madazl}, R.; Tüysüz, B.; Aksoy, F.; Barbaros, M.; UludagE , S.; Ocak, V. (Istanbul) 34 Prenatal Diagnosis of Lissencephaly (Type II) by Ultrasound and Fast Magnetic Resonance Imaging Kojima, K.; Suzuki, Y.; Seki, K.; Yamamoto, T.; Sato, T.; Tanaka, T.; Suzumori, K. (Nagoya) 37 Expression of C-Type Natriuretic Peptide in Human Placenta and Myometrium in Normal Pregnancies and Pregnancies Complicated by Intrauterine Growth Retardation. Preliminary Results Stepan, H.; Faber, R.; Stegemann, S. (Leipzig); Schultheiss, H.-P.; Walther, T. (Berlin) 42 Early Transvaginal Biometry of Fetal Orbits: A Cross-Sectional Study Guariglia, L.; Rosati, P. (Rome) 48 Uterine Artery Doppler Velocimetry in Patients with Idiopathic Hydramnios Hershkovitz, R.; Sheiner, E.; Furman, B.; Smolin, A.; Hallak, M.; Mazor, M. (Beer-Sheva) 52 First-Trimester Ductus venosus Velocimetry in Relation to Nuchal Translucency Thickness and Fetal Karyotype Zoppi, M.A.; Putzolu, M.; Ibba, R.M.; Floris, M.; Monni, G. (Cagliari) 58 MR Autopsy in Fetuses Huisman, T.A.G.M.; Wisser, J.; Stallmach, T.; Krestin, G.P.; Huch, R.; Kubik-Huch, R.A. (Zürich) No. 2

At present, the measurement of IgG avidity appears to be the best method for differentiating primary from nonprimary cytomegalovirus (CMV) infection. This study compared the performances of three denaturation assays for the measurement of CMV IgG avidity: an in-house method and the commercially available assays Enzygnost (Dade-Behring, Germany) and Vidas (bioMérieux, France). The ability of these assays to exclude or to detect a recent CMV infection were calculated according to the results obtained in two control groups of pregnant women: 49 who had seroconverted and 80 with past infections. All three assays demonstrated a good ability to detect a recent infection (98–100%). The Dade-Behring test, in its present form, appears to be ineffective in excluding a recent CMV infection (exclusion ability: 30%), while the in-house method (exclusion ability: 96%) and the bioMérieux method (exclusion ability: 82%) performed better. The practical use of the in-house and the bioMérieux assays was evaluated in 80 women with CMV-specific IgG and a positive IgM result but without documented seroconversion. At the recommended diagnostic thresholds, the concordance between these two tests was 70%. Larger studies will allow more precise determination of the capacities of both assays and specification of the diagnostic thresholds or grey areas to be used.

The hepatitis C virus genotype distribution was studied among age groups in 501 referred patients with chronic hepatitis C by INNO-LiPA HCV II (Innogenetics, Belgium). Ten patients had coinfection with several genotypes. Two hundred seventy of 491 singly infected individuals (57%) had 1b, 66 (13.4%) 3a, 57 (11.6%) 1a. HCV subtype 1b was predominant but its prevalence increased with age (76.5% of patients born in the '20s, 39.3% in the '70s) (P < 0.0001). Three possibilities could explain the shift towards a wider variety of genotypes in younger age. (1) 1b could be the original subtypes in this population, (2) the non-1b subtypes could give less chronic carriers, (3) the non-1b subtypes could have a higher mortality, which seems improbable. The 1b genotype seems the oldest subtype in our country while others were imported later through increased population movements and changing habits.

ABSTRACT We evaluated the reactivities of sera against p52 and CM2 recombinant antigens of human cytomegalovirus (HCMV), coated on microparticles, for the differentiation of primary HCMV infection from an established infection. Two different test formats of the CMV Multiplex Copalis assay were evaluated. The 214 serum samples tested were immunoglobulin M (IgM) positive or equivocal by our reference assay. Reactivities against p52 and CM2 antigens were tested for sera from 37 patients with a well-documented seroconversion within the preceding 3 months (119 serum specimens), 31 patients known to have had a seroconversion at least 8 months earlier (31 serum specimens), and 57 patients without a documented seroconversion (64 serum specimens). The assay had a sensitivity for the detection of a primary infection of 70 or 86% by the first test format and a sensitivity of 88 or 94% by the second test format, according to the criteria used to indicate a primary infection by this test. A good correlation of the results of the assay with our in-house avidity index was found. The specificity of the assay warrants further evaluation. With IgM-positive sera, the assay was not sufficiently specific to make a distinction between a primary infection and an established infection.

The diagnosis of acute cytomegalovirus (CMV) infection is frequently based on a positive IgM result. False-positive reactions due to interfering infections may exist. Between August 1998 and May 1999, 62 patients were found to be IgM positive and IgG negative with the Axsym assay (Abbott, Germany). Serological testing for Epstein-Barr virus (EBV) was performed in these patients to detect any cross-reactivity due to acute mononucleosis. Additionally, the results of the CMV Axsym was evaluated in 40 patients with acute EBV infection. The results suggest that the CMV-IgM Axsym assay shows a lack of specificity due to acute EBV infection. Precautions must be taken when CMV-IgM Axsym results are interpreted. It seems necessary to confirm equivocal results with another technique and to take into account other clinical and biological observations.