Véronique Lemée

A new human immunodeficiency virus derived from gorillas.

We have identified a new human immunodeficiency virus in a Cameroonian woman. It is closely related to gorilla simian immunodeficiency virus (SIVgor) and shows no evidence of recombination with other HIV-1 lineages. This new virus seems to be the prototype of a new HIV-1 lineage that is distinct from HIV-1 groups M, N and O. We propose to designate it HIV-1 group P.

HIV-1 resistance genotyping on dried serum spots.

Objective:to assess the feasibility of HIV-1 group M resistance genotyping on dried serum spots, by testing samples from previously untreated patients, patients on treatment, and patients having stopped treatment, representing a wide genetic diversity panel. Methods:serum samples from 62 HIV-1-infected Caucasian and African patients, with viral load values from 715 copies/ml to more than 750 000 copies/ml, were deposited on filter paper. After elution and RNA extraction, nested RT-PCR was used to amplify the protease and RT regions of the pol gene. Resistance sequencing was performed on all the protease and RT amplicons. The sequences obtained for resistance genotyping were used for subtyping by phylogenetic analysis. Results:amplification was successful in the protease region in 53/62 cases (85.5%) and in the RT region in 51/62 cases (82.3%). All samples with viral loads of at least 5 Log (17 of 62) were successfully amplified in both the RT and protease regions. Of the 29 samples with viral loads between 4 Log and 5 Log, 28 (97%) were amplified in the RT region and 25 (86%) in the protease region. The detected mutations were in keeping with the treatment status. Marked natural polymorphism was observed in the protease region, but no major consequences were deduced in terms of resistance. The results showed a broad diversity of the panel, including subtype B (n = 36) and non B or recombinant forms (n = 20). Conclusion:Our results show the feasibility of this dried serum spot method for monitoring resistance to antiretroviral drugs and the molecular epidemiology of HIV diversity. The simplicity of sample preparation, storage and transport potentially makes this an importance tool for individual and epidemiological monitoring throughout the world.

Census and analysis of persistent false-negative results in serological diagnosis of human immunodeficiency virus type 1 group O infections.

ABSTRACT Human immunodeficiency viruses (HIV) have a high level of genetic diversity. The outlier variants of HIV type 1 (HIV-1) group O are distantly related to HIV-1 group M. Their divergence has an impact on serological diagnosis, with a risk of false-negative results. In this study, we report 20 failure cases, involving patients with primary or chronic infection, in France and Cameroon between 2001 and 2008. Our results indicate that some assays detected group O infection much less efficiently than others. Two major reasons for these false-negative results were identified: the presence or absence of a group O-specific antigen (and the designed sequence) for the detection of antibodies and the greater envelope variability of group O than of group M strains. This study highlights the complexity of screening for these divergent variants and the need to evaluate test performance with a large panel of strains, due to the extensive diversity of group O variants.

The variable sensitivity of HIV Ag/Ab combination assays in the detection of p24Ag according to genotype could compromise the diagnosis of early HIV infection

BACKGROUND In France, HIV infection diagnosis was modified by a decree, published in 2010, that requires the use of HIV Ag/Ab assays able to detect at least 2 IU/ml of p24Ag. This measure raises the concern of the capacity of these assays to equally detect all HIV variants. OBJECTIVES To assess the performance of HIV Ag/Ab assays for the detection of p24Ag from diverse HIV isolates. STUDY DESIGN Ten HIV Ag/Ab assays were compared using two p24Ag reference standards, 297 samples from 99 HIV-1 and HIV-2 cell-culture derived isolates including various subtypes and groups, and 9 native specimens from subjects with primary HIV infection. RESULTS The p24Ag limit of detection (LOD) ranged from 0.505 IU/ml to 1.90 1 IU/ml and, from 11.9 pg/ml to 33.5 pg/ml when using WHO and French national standards, respectively. The overall percentage of positive samples ranged from 26.8% to 74.5%. Five assays failed to detect all dilutions of at least one group M subtype, three missed all group O and six all the group P samples. Three assays were able to detect 2-10 of the 30 HIV-2 samples. The distribution of LODs for each group M isolate showed a wide dispersion between the assays. Percentage of isolates detected at a p24Ag level less than 2 IU/ml varied from 22% to 98.7%. CONCLUSION This study demonstrated that, even though their analytical sensitivity fulfills the requirements, many of HIV Ag/Ab assays could fail to detect HIV primary infection due to HIV-1 non-B, non-M and HIV-2 strains.

Differences in proviral DNA load between HIV-1- and HIV-2-infected patients.

Introduction:The lesser pathogenicity of HIV-2 relative to HIV-1 is generally attributed to its slower replication. To compare the amounts of total HIV DNA during human HIV-1 and HIV-2 infection, we developed a quantitative real-time PCR method with a unique external quantification standard based on a single plasmid harboring both the HIV-1 and the HIV-2 LTR. Methods:Viral DNA load was compared between 40 HIV-1-infected and 42 HIV-2-infected antiretroviral-naive patients. Results:The difference between HIV-1 and HIV-2 proviral DNA load was highly significant in patients with CD4 cell counts > 500 cells/μl [HIV-1: n = 14; median, 2.5; interquartile range (IQR), 2.1–2.7; HIV-2: n = 22, median, 1.6; IQR, 1.0–2.0] and in patients with CD4 cell counts between 300 cells/μl and 500 cells/μl (HIV-1: n = 12; median, 2.7; IQR, 2.3–2.8; HIV-2: n = 11; median, 2.0; IQR, 1.0–2.4). Too few HIV-2-infected patients had CD4 cell counts < 300 cells/μl to detect a significant difference but DNA values were again lower in HIV-2-infected patients (HIV-1: n = 14; median, 2.9; IQR, 2.2–3.2; HIV-2: n = 9; median, 2.7; IQR, 2.2–3.3). Conclusions:These differences are in line with the natural histories of the two infections and show that HIV-2 infection is a valid model for studying the pathophysiology of HIV infection in general.

First evidence of a HIV-1 M/O recombinant form circulating outside Cameroon.

HIV-1 are now classified into four groups: M (major), O (outlier), N (non-M – non-O), and a new group P, the prototype of which is the divergent strain recently characterized in a Cameroonian patient [1]. Recombination between HIV strains is a very frequent phenomenon, greatly increasing genetic div

Unequal Detection of HIV Type 1 Group O Infection by Simple Rapid Tests

possibility of transmission of this pathogen and the need for precautions against airborne pathogen transmission that are associated with the use of high-risk respiratory procedures in patient care. An outbreak of Ad14 infection among military trainees at Lackland Air Force Base, Texas, that started in February 2007 has been described elsewhere [2]. In April and May 2007, 4 trainees required admission to the intensive care unit, including 1 who presented in severe respiratory distress and required immediate intubation, prolonged bag-mask ventilation, and subsequent transition to HFOV. Many HCWs were involved in the patient’s resuscitation and initial care in the intensive care unit; within a week, 7 HCWs reported respiratory illness and/or conjunctivitis. As part of our facility’s outbreak investigation, active surveillance for respiratory symptoms among this patient’s HCWs was undertaken, and 23 reported symptoms. Eight were tested for adenovirus by respiratory viral culture or PCR, and 6 had positive results. Although these specimens were not serotyped, given the exposure to this patient with Ad14 and the high proportion of Ad14 respiratory illnesses among trainees at the time (60 [92.3%] of 65 serotyped isolates [3]), it is most likely that the 8 adenoviruses were Ad14 as well. This cluster of respiratory illness among HCWs involved in the initial resuscitation of this patient was thought to be related to the high-risk respiratory procedures involved, including endotracheal intubation, prolonged bag-mask ventilation, and the use of HFOV. High-risk respiratory procedures were linked to severe acute respiratory syndrome (SARS) transmission among HCWs, and HCW use of N95 masks may have conferred additional protection to those involved with such procedures [3–5]. Given the ability of HFOV to generate aerosols, there is the potential for transmission of adenoviral and other communicable infections. Although we have not observed transmission of Ad14 from one hospitalized patient to another, we recently cared for one of our own fellows who was admitted to the hospital with lobar pneumonia, delirium, sepsis, and acute renal failure and who received a diagnosis of Ad14 infection. He had not had duties at Lackland Air Force Base for several months and had no contact with patients at this hospital or with children or trainees on the base. His only epidemiologic link to the Ad14 outbreak was through his girlfriend, a nurse on our facility’s internal medicine ward, who reported a febrile respiratory illness during the preceding week. In summary, we describe an outbreak of apparently occupational adenovirus infection among HCWs that was associated with high-risk respiratory procedures, including HFOV, and we describe a severe secondary case in a household contact of an HCW. We encourage aggressive implementation of appropriate isolation precautions, including precautions against airborne transmission, during high-risk respiratory procedures in patients infected with Ad14.

Plasma virion reverse transcriptase activity and heat dissociation-boosted p24 assay for HIV load in Burkina Faso, West Africa

Background:In resource-limited settings, the requirement for inexpensive, easy-to-perform viral load monitoring has increased with greater antiretroviral drug availability. Objectives:To evaluate feasibility, in Burkina Faso, of a simple assay for plasma HIV reverse transcriptase (RT) activity quantification compared to heat dissociation-boosted (HDB) p24 antigen and RNA-based quantifications in plasma samples from HIV-infected patients. Methods:Plasma viraemia was quantified by RT activity, HDB-p24 and RNA copies in 84 samples from 70 HIV-1 group M-infected patients (82% non-B subtype, 93% treatment naive), including serial samples from nine patients. Results:RT activity detected 86% of plasma samples containing measurable RNA copies; corresponding to 0, 93 and 100% of samples with 1.7–4.0 log10, 4.1–4.8 log10 and 4.9–6.7 log10 RNA copies/ml, respectively. HDB-p24 detected 77% of plasma samples containing measurable RNA copies; corresponding to 27, 80 and 86% of samples with 1.7–4.0 log10, 4.1–4.8 log10 and 4.9–6.7 log10 RNA copies/ml, respectively. Measurement error based on one-way analysis of variance between RT activity and HDB-p24 values with RNA copies showed good agreement with RT activity (ME, <10%), however poorer agreement was obtained with HDB-p24 values (ME, >10%). Patient follow up showed a similar pattern of viraemia with RNA and RT activity assays. Conclusion:Field trials in Burkina Faso support the practical use of plasma RT activity assay as an affordable alternative for HIV viral load determination in regions where RNA detection remains difficult to perform. HDB-p24 use requires further evaluation before being considered as an alternative method in African HIV-infected patient follow up.

A New Real-Time Quantitative PCR for Diagnosis and Monitoring of HIV-1 Group O Infection

ABSTRACT The correct diagnosis and monitoring of HIV-1 group O (HIV-O) infection are essential for appropriate patient management, for the prevention of mother-to-child transmission, and for the detection of dual HIV-M/HIV-O infections. HIV-O RNA quantification is currently possible with two commercial kits (from Abbott and Roche), which quantify HIV-M and HIV-O strains indifferently; therefore, they cannot be used for the specific identification of HIV-O infection. We designed a new real-time quantitative reverse transcription PCR (RT-qPCR assay) (INT-O), which we compared with our previous version, LTR-O, and with the Abbott RealTime HIV-1 kit. Specificity was assessed with 27 HIV-1 group M strains and the prototype strain of group P. Clinical performances were analyzed by using 198 stored plasma samples, representative of HIV-O genetic diversity. Analytical sensitivity, repeatability, and reproducibility were also determined. The detection limit of the INT-O assay was 40 copies/ml, and its specificity was 100%. The repeatability and reproducibility were excellent. Analysis of clinical samples showed a good correlation between the INT-O and LTR-O assays (r = 0.8240), with an improvement of analytical sensitivity. A good correlation was also obtained between the INT-O and Abbott assays (r = 0.8599) but with significantly higher values (0.19 logs) for the INT-O method, due to marked underquantifications for some patients. These results showed that HIV-O genetic diversity still has an impact on RNA quantification. The new assay, INT-O, allows both the specific diagnosis of HIV-O infection and the quantification of diverse HIV-O strains. Its detection limit is equivalent to that of commercial kits. This assay is cheap and suitable for use in areas in which strains of HIV-1 groups M and O cocirculate.

Field evaluation of the Abbott ARCHITECT HIV Ag/Ab Combo immunoassay

BACKGROUND Fourth generation assays for HIV diagnosis are progressively being introduced into routine services, due to their improvement of diagnosis. In spite of this, HIV diagnosis remains a challenge in sub-Saharan Africa, due to false positive reactivity. There is a continuous need for field evaluations and routine validations of fourth generation HIV tests in African populations. OBJECTIVES Evaluate the performances of the ARCHITECT HIV Ag/Ab kit (Abbott) in a population living in an African setting-Cameroon compared to a population living in a European setting-France. STUDY DESIGN 645 HIV samples from both France and Cameroon were evaluated. The positive panel (378 samples) included a diverse series of HIV-1 variants (groups M, N, O, and P) as well as HIV-2 samples. Results were compared to original diagnosis done with other 4th generation assays (AxSYM HIV Ag/Ab (Abbott) and Vidas HIV DUO QUICK) (bioMérieux). RESULTS Sensitivity of the ARCHITECT was 100% in both sites. It diagnosed all variants of the panel with different reactivity profiles following strain diversity. A wider range of reactivity was observed for group O. Specificity was slightly lower (97.6%) in Cameroon than in France (98.6%), probably due to a higher rate of false positive reactivity. ARCHITECT HIV Ag/Ab assay had high performances in clinical sensitivity and specificity and is adapted to the wide genetic diversity of viruses circulating in West Central Africa. CONCLUSION Our results further highlight the need to evaluate HIV diagnostic tests before introduction into routine diagnostic services both in the North and in the South.