RNA and DNA Sanger sequencing versus next-generation sequencing for HIV-1 drug resistance testing in treatment-naive patients.

Background: Sanger sequencing of plasma RNA is the standard method for HIV-1 drug resistance testing in treatment-naive patients, but is limited by the non-detection of resistance-associated mutations (RAMs) with prevalence below approximately 20%. Objectives: We compared RNA and DNA Sanger sequencing (RSS and DSS) with RNA next-generation sequencing (NGS) for RAM detection in HIV-1 reverse transcriptase (RT), protease (PR) and integrase (IN) genes. Methods: Sanger sequencing was performed on RNA and DNA, following the recommendations of the French Agency for AIDS Research (ANRS). NGS was performed on RNA using the HIV-1 Drug Resistance Assay, v. 3.0 (Roche) on the 454 GS Junior sequencer. The IAS-USA list was used to identify RAMs. ANRS, Rega and Stanford algorithms were used for drug resistance interpretation. Results: The study included 48 ART-naive patients. The number of patients with at least one major RAM was 3, 3, 4 and 8 when using RSS, DSS, NGS 20% and NGS 5%, respectively. Numerous minor mutations were detected in patients, especially in the protease gene. None of the methods detected any major mutation in the integrase gene. Overall, the mutation detection rate was similar between RSS and DSS, and higher with NGS 20%. Differences in drug resistance interpretation were found between algorithms. No impact of the minority RAMs detected by NGS was found on the short-term treatment outcome. Conclusions: DSS does not clearly improve the detection of RAMs in ART-naive patients, as compared with RSS. NGS allows detection of additional minority RAMs; however, their clinical relevance requires further investigation.

Factors associated with virological response to a switch regimen containing maraviroc for antiretroviral-experienced HIV-1-infected patients.

OBJECTIVES: There are few data on clinical and virological factors associated with maraviroc virological response (VR) in clinical practice. This study aimed to identify factors associated with VR in 94 treatment-experienced, but CCR5 inhibitor-naive, HIV-1 patients switched to maraviroc-containing regimens. METHODS: Patients with HIV-1 RNA viral load (VL) <50 copies/mL switching to an antiretroviral treatment containing maraviroc were followed. VR was defined at month 3 as VL <50 copies/mL. The impact of age, baseline tropism, zenith VL, nadir CD4 cell count and CD4 cell count, HIV subtype (B versus non-B), genotypic susceptibility score of treatment, once- or twice-daily treatment and presence of raltegravir in optimized background therapy on VR was investigated. RESULTS: Baseline characteristics were: median age 49 years (range 25-73 years), median CD4 cell count 481 cells/mm(3) (range 57-1830 cells/mm(3)) and median nadir CD4 cell count 99 cells/mm(3) (range 3-585). Maraviroc was administered twice daily in 88 of 94 patients and once daily in 6 of 94 patients (300 mg/day for 4 of 6 and 150 mg/day for 2 of 6). At month 3, 89.4% of patients were responders. A better VR to a switch regimen containing maraviroc was associated with the B subtype (P = 0.0216) and a lower zenith VL (median of 5.24 and 5.70 log10 copies/mL for patients in success or in failure, respectively) in univariate analysis. Only B subtype was associated with a better VR in multivariate analysis. CONCLUSIONS: This study evidenced the efficacy of a switch regimen containing maraviroc in clinical practice. VR was better for patients with a lower zenith VL and B subtype.

Auditing HIV Testing Rates across Europe: Results from the HIDES 2 Study.

European guidelines recommend the routine offer of an HIV test in patients with a number of AIDS-defining and non-AIDS conditions believed to share an association with HIV; so called indicator conditions (IC). Adherence with this guidance across Europe is not known. We audited HIV testing behaviour in patients accessing care for a number of ICs. Participating centres reviewed the case notes of either 100 patients or of all consecutive patients in one year, presenting for each of the following ICs: tuberculosis, non-Hodgkins lymphoma, anal and cervical cancer, hepatitis B and C and oesophageal candidiasis. Observed HIV-positive rates were applied by region and IC to estimate the number of HIV diagnoses potentially missed. Outcomes examined were: HIV test rate (% of total patients with IC), HIV test accepted (% of tests performed/% of tests offered) and new HIV diagnosis rate (%). There were 49 audits from 23 centres, representing 7037 patients. The median test rate across audits was 72% (IQR 32-97), lowest in Northern Europe (median 44%, IQR 22-68%) and highest in Eastern Europe (median 99%, IQR 86-100). Uptake of testing was close to 100% in all regions. The median HIV+ rate was 0.9% (IQR 0.0-4.9), with 29 audits (60.4%) having an HIV+ rate >0.1%. After adjustment, there were no differences between regions of Europe in the proportion with >0.1% testing positive (global p = 0.14). A total of 113 patients tested HIV+. Applying the observed rates of testing HIV+ within individual ICs and regions to all persons presenting with an IC suggested that 105 diagnoses were potentially missed. Testing rates in well-established HIV ICs remained low across Europe, despite high prevalence rates, reflecting missed opportunities for earlier HIV diagnosis and care. Significant numbers may have had an opportunity for HIV diagnosis if all persons included in IC audits had been tested.

[Catch-up vaccination of worldwide newcoming (adopted, refugee or migrant) children in France]. / Mise à jour des vaccinations de l'enfant arrivant de l'étranger (adopté, réfugié ou migrant) en France.

In France, international adoption includes around to 90,000 children since 1980 and near 300,000 immigrant children were counted in 2008. This population is heterogeneous, according to age and country of origin, and its large number. It is not easy to completely and surely assess the vaccine status of the child. Due to a great variability of individual situations, it is not possible to have systematic and unchangeable rules. This article aims to give an update of catch-up vaccination of internationally adopted or refugee or migrant children in France. The vaccination status of a child who recently arrived in France is complex and has to be adapted to his country of origin. Some of them were never vaccinated whereas the vaccine status of others is uncertain or unknown. Three parameters have to be considered: the age of the child, the country of origin, and sometimes serology in the case of doubts of his vaccine status. Catch-up vaccination of foreign children has to be adapted to French vaccine recommendations, as a reference, and to vaccines already administered to the child.

[Cryptococcal osteomyelitis in a patient with a lymphocytic leukemia treated with fludarabine-cyclophosphamide-rituximab]. / Cryptococcose osseuse chez une patiente porteuse d'une leucémie lymphocytique traitée par fludarabine-cyclophosphamide-rituximab.

INTRODUCTION: Cryptococcosis is a serious invasive fungal infection mostly described in patients with cell-mediated immunosuppression. Cryptococcus neoformans osteomyelitis is a rare infection that occurs mainly during disseminated forms. OBSERVATION: A 72-year-old diabetic patient, treated with fludarabine-cyclophosphamide-rituximab (since 10 months) for lymphocytic leukemia presented with osteolysis of the fourth left hand metacarpien the histological examination of which revealed C. neoformans. This bone involvement was associated with costal osteolytis and pulmonary cryptococcosis but central nervous system (CNS) was spared. Fluconazole was administered intravenously for 15 days, then switched to oral route for 6 months with favorable clinical course. This case describes an unusual clinical presentation of disseminated cryptococcosis without CNS involvement with multiple osseous metastases. A review of cryptococcal osteomyelitis cases reported in adult from 2000 to 2011 is also discussed. CONCLUSION: Cryptococcosis is a rare infection that should be discussed in seriously immunocompromized patients presenting with osteomyelitis even in the absence of CNS involvement.

Vaccinal issues between Belgian and French travelers.

INTRODUCTION: The yellow-fever vaccination center of the Tourcoing Hospital (France) has been accessible to Belgian travelers since its opening in 1994. METHOD: The authors reported the specificities of these consultations during the year 2010, by retrospectively analyzing electronic medical records. RESULTS: Some medical issues encountered during the consultation were due to differences in vaccination schedules: for the polio vaccine, since the last dose is administered between 5 and 7 years of age in Belgium; and for the measles vaccine since a late two-dose schedule (second dose between 12 and 13 years of age) is recommended in this country. Moreover, some specific vaccines are available only in Belgium: a diphtheria-tetanus bivalent vaccine, and a live attenuated oral typhoid vaccine. DISCUSSION: The specificities of the Belgian border traveler consultation in our French yellow-fever center are due to a difference in European vaccination schedules; the physician must be aware of these. CONCLUSION: The physician has to propose updates on vaccination schedules, and be aware of yellow-fever vaccine compatibility with vaccines recently administered in Belgium.

Long-term efficacy of darunavir/ritonavir monotherapy in patients with HIV-1 viral suppression: week 96 results from the MONOI ANRS 136 study.

OBJECTIVES: Long-term results at week 96 are needed to evaluate the capacity of the darunavir/ritonavir monotherapy strategy to maintain a sustained control of the HIV-1 viral load. METHODS: MONOI is a prospective, open-label, non-inferiority, randomized, 96 week trial comparing darunavir/ritonavir monotherapy versus a darunavir/ritonavir triple-therapy strategy to maintain HIV-1 viral load suppression in HIV-1-infected patients. CLINICAL TRIAL REGISTRATION: NCT00412551. RESULTS: From 225 randomized patients, 219 patients reached the 48 week follow-up and 211 reached the 96 week follow-up (106 patients in the darunavir monotherapy arm and 105 in the darunavir triple-therapy arm). Baseline characteristics were well balanced between the two treatment groups. At week 96, in intent-to-treat analysis, 91/103 patients (88%, 95% CI 81-94) allocated to the darunavir/ritonavir monotherapy arm and 87/104 patients (84%, 95% CI 75-90) allocated to the darunavir triple-therapy arm achieved an HIV-1 viral load <50 copies/mL, with no statistical difference between the two groups. Throughout the 96 week follow-up, 66/112 patients (59%, 95% CI 49-68) and 79/113 patients (70%, 95% CI 61-78) consistently had HIV-1 RNA <50 copies/mL with darunavir/ritonavir monotherapy and darunavir/ritonavir triple therapy, respectively. CONCLUSIONS: The MONOI study establishes darunavir/ritonavir monotherapy as durable and efficacious for maintaining virological suppression in HIV-1 patients. Darunavir/ritonavir monotherapy should be considered as a (tailored) treatment option for standard triple-therapy patients who have had a substantial period of viral suppression.

Acute upper gastro-intestinal bleeding in morocco: what have changed?

Objective. In the present study, we aimed to investigate epidemiological, clinical, and etiological characteristics of acute upper gastro-intestinal bleeding. Materials and Methods. This retrospective study was conducted between January 2003 and December 2008. It concerned all cases of acute upper gastroduodenal bleeding benefited from an urgent gastro-intestinal endoscopy in our department in Morocco. Characteristics of patients were evaluated in terms of age, gender, medical history, presenting symptoms, results of rectal and clinical examinations, and endoscopy findings. Results. 1389 cases were registered. As 66% of the patients were male, 34% were female. Mean age was 49. 12% of patients had a history of previous hemorrhage, and 26% had a history of NSAID and aspirin use. Endoscopy was performed in 96%. The gastroduodenal ulcer was the main etiology in 38%, followed by gastritis and duodenitis in 32.5%. Conclusion. AUGIB is still a frequent pathology, threatening patients' life. NSAID and aspirin are still the major risk factors. Their impact due to peptic ulcer remains stable in our country.

[Study of new HIV infection between 2000 and 2007 in the north and east of France]. / Étude des infections à VIH nouvellement découvertes entre 2000 et 2007 dans le Nord et l'Est de la France.

UNLABELLED: In France, since 2003, all new HIV infection must be reported. Data collected with the declaration system is not exhaustive and only concerns epidemiological data. OBJECTIVE: The authors' aim was to study the epidemiologic evolution of new HIV cases between January 1, 2000 and December 31, 2007 in North and East of France, to compare them with national and local data, to complete them, and to identify local specificities. METHOD: A retrospective observational study was made, with a standardized questionnaire completed by any volunteer HIV care center in the North and the East of France. RESULTS: Three thousand and thirty questionnaires were analyzed. The main trends over these eight years were similar to those observed in the rest of France: a decreasing number of women and patients of foreign origin, a decreasing number of patients with a late diagnosis, an increasing number of primary infections, and a higher CD4 count on initiation of antiretroviral treatment. However, local specificities appeared, such as: increasing proportion of men having sex with men and a less important proportion of co-infected patients with hepatitis B and/or C than on the national level. The therapeutic regimen is adequate according to expert recommendations, with, however, a marked "center effect" concerning prescription habits. DISCUSSION: Such a local epidemiological study, even if it confirms observed trends in the rest of France, allows detailing them and suggesting prevention measures more specifically adapted to local settings.

Vaccination in adults with auto-immune disease and/or drug related immune deficiency: results of the GEVACCIM Delphi survey.

INTRODUCTION: There are insufficient data regarding the efficacy and safety of vaccination in patients with auto-immune disease (AID) and/or drug-related immune deficiency (DRID). The objective of this study was to obtain professional agreement on vaccine practices in these patients. METHODS: A Delphi survey was carried out with physicians recognised for their expertise in vaccinology and/or the caring for adult patients with AID and/or DRID. For each proposed vaccination practice, the experts' opinion and level of agreement were evaluated. RESULTS: The proposals relating to patients with AID specified: the absence of risk of AID relapse following vaccination; the possibility of administering live virus vaccines (LVV) to patients not receiving immunosuppressants; the pertinence of determining protective antibody titre before vaccination; the absence of need for specific monitoring following the vaccination. The proposals relating to patients with DRID specified that a 3-6 month delay is needed between the end of these treatments and the vaccination with LVV. There is no contraindication to administering LVV in patients receiving systemic corticosteroids prescribed for less than two weeks, regardless of their dose, or at a daily dose not exceeding 10mg of prednisone, if this involves prolonged treatment. Out of 14 proposals, the level of agreement between the experts was "very good" for eleven, and "good" for the remaining three. CONCLUSION: Proposals for vaccine practices in patients with AID and/or DRID should aid with decision-making in daily medical practice and provide better vaccine coverage for these patients.

[Are there differences in immunogenicity and safety of vaccines according to the injection method?]. / Existe-t-il des différences d'immunogénicité et de tolérance des vaccins en fonction du mode d'injection ?

OBJECTIVE: A literature review was made to answer the following question: are there differences in immunogenicity and safety of vaccines according to the administration route (intramuscular or subcutaneous) and the length of needles used for injection? DESIGN: The search strategy included electronic searching (Medline database via PubMed) and cross-references. Articles were selected by reading abstracts, guided by the clinical question. A total of 18 articles were selected and analyzed; 13 answered the question. RESULTS: Nine articles compared the immunogenicity and/or the safety of a given vaccine administered via both intramuscular and subcutaneous routes. All the results showed that immunogenicity and systemic safety of the intramuscular route was at least as good as that of the subcutaneous route. Local tolerance was usually better via intramuscular route (lower risk of developing erythema or edema). Four articles compared the immunogenicity and the safety of vaccines administered with short (16mm) and long (25mm) needles in children. Long needles induced fewer local reactions, probably because they can reach more vascularized muscle, especially in overweight and obese patients. CONCLUSIONS: Immunogenicity and safety results are in favor of intramuscular vaccination. The appropriate needle length must to be adapted according to the morphological aspects (subcutaneous tissue and muscle thickness).

[Non-cutaneous manifestations of secondary syphilis]. / Manifestations extra-cutanéomuqueuses de la syphilis secondaire.

BACKGROUND: The usual presentation of secondary syphilis is with cutaneous and mucosal symptoms. However, systematic symptoms can also occur. The purpose of this study was to describe non-mucocutaneous manifestations of secondary syphilis. PATIENTS AND METHODS: Patients from the Infectious Diseases Department of Tourcoing Hospital in whom secondary syphilis was diagnosed between January 2000 and December 2006 were enrolled in this study. Patients with secondary syphilis had the typical cutaneous and mucosal symptoms and a VDRLgreater than or equal to one quarter (or a fourfold increase in the VDRL if previously positive). RESULTS: Seventy-seven patients presenting a total of 80 cases of secondary syphilis were enrolled, 50 of whom were HIV-positive. Of these patients, 21 (26.3 p. 100) had neurological symptoms with three cases (3.8 p. 100) of uveitis, four (5 p. 100) of papillitis, two (2.5 p. 100) of retinitis and one (1.25 p. 100) of otosyphilis. In 14 of these 21 patients (67 p. 100), lumbar puncture was performed, confirming the diagnosis of neurosyphilis in six cases. Three patients (3.8 p. 100) had diarrhoea, four (5 p. 100) had abdominal pain and six (7.5 p. 100) had hepatomegaly. Seven (11.5 p. 100) patients had alanine aminotransferase levels above twice the normal upper limit and two above 10 times the normal upper limit. Three patients had bone pain and in one patient, osteitis was confirmed by technetium and gallium scintigraphy (osteolysis). CONCLUSION: In patients with secondary syphilis, clinicians should search for non-mucocutaneous symptoms. In the presence of these symptoms, appropriate syphilis treatment should be initiated.

ABCB1 allele polymorphism is associated with virological efficacy in naïve HIV-infected patients on HAART containing nonboosted PIs but not boosted PIs.

OBJECTIVE: To assess the effect of the multidrug resistance-1 single nucleotide polymorphism (ABCB1 SNP) C3435T in exon 26 on the virological responses to first-line protease inhibitor (PI)-containing HAART regimens. METHOD: A cohort of 182 HIV-infected patients with a PI-containing HAART regimen initiated from 1997 to 2004 was enrolled. Time to the first indetectable viral load (VL) was determined in patients with the CC, CT, or TT genotype. RESULTS: There were 37%, 44%, and 19% of patients who had the CC, CT and TT genotypes, respectively. The median estimated times to VL indetectability in the CC, CT, and TT groups were respectively 5.9, 3.9, and 4.8 months (p= .06). In patients on a non-boosted PI regimen, ABCB1 genotype was associated with time to VL indetectability that was shorter in patients with the CT than CC genotype (CT vs. CC, hazard ratio [HR]=0.62, p= .02; TT vs. CC, HR= 0.72, p= .21). This association was not found in patients with first-generation boosted PI-containing regimens and especially not with second-generation boosted PI-containing regimens. CONCLUSION: Our results show that the ABCB1 SNP in exon 26 is associated with virological efficacy in HIV-infected patients treated with non-boosted PI-containing regimens but not with those containing boosted PIs, particularly of the second generation.

Development of a PCR assay followed by nonradioactive hybridization using oligonucleotides covalently bound to CovaLink NH microwells for detection of four Plasmodium species in blood samples from humans.

We developed and evaluated a PCR-based assay to detect four Plasmodium species in 79 blood samples from 56 travelers returning from areas where malaria is endemic. DNA amplification targeting a small region of the 18S rRNA gene was performed with Plasmodium genus-specific primers. The biotinylated PCR products were then identified by PCR-colorimetric Covalink NH microwell plate hybridization (CMPH) using species-specific phosphorylated probes covalently bound to a pretreated polystyrene surface. The results from PCR-CMPH showed high specificity, and for 47 of the 56 patients (84%), microscopy and PCR-CMPH results were in agreement. Discordant results were reevaluated with microscopy examination, other molecular methods, and DNA sequencing. Except for one patient, discrepancies were resolved in favor of PCR-CMPH: three mixed infections were detected, four species identification errors were corrected, and two negative results were shown to be positive. Our results indicate that PCR-CMPH is a simple, rapid, and specific method for malaria diagnosis. It employs stable reagents and inexpensive equipment, making it suitable for routine epidemiological use.