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International audience; Introduction: Bronchiolitis is the leading cause of hospitalization for infants but its economic burden is not well documented. Our objective was to describe the clinical evolution and to assess the 1-month cost of a first episode of acute bronchiolitis presenting to the emergency department (ED).Methods: Our study was an epidemiologic analysis and a cost study of the cohort drawn from the clinical trial GUERANDE, conducted in 24 French pediatric EDs. Infants of 6 weeks to 12 months of age presenting at pediatric EDs with a first episode of bronchiolitis were eligible. The costs considered were collected from a societal viewpoint, according to the recommendations of the French National Health Authority.Results: A total of 777 infants were included with a median age of 4 months. A total of 57% were hospitalized during the month following the first consultation in the ED, including 28 (3.6%) in an intensive care unit. The mean length of stay was 4.2 days (SD = 3.7). The average time to relief of all symptoms was 13 days (SD = 7). Average total cost per patient was €1919 (95% confidence interval: 1756-2138) from a societal perspective, mostly due to hospitalization cost. The estimated annual cost of bronchiolitis in infants was evaluated to be between €160 and €273 million in France.Discussion: Bronchiolitis represent a high cost for the health care system and broadly for society, with hospitalizations costs being the main cost driver. Thus significant investments should be made to develop innovative therapies, to reduce the number of hospitalizations and length of stay.

Resume Introduction Le diagnostic etiologique des infections bronchopulmonaires ne peut se faire sur les seules donnees cliniques, radiologiques et epidemiologiques. La part importante des infections virales chez l’adulte et chez l’enfant est demontree. Etat des connaissances Le diagnostic des infections bronchopulmonaires virales repose sur l’analyse d’un prelevement respiratoire, de preference profond, par des techniques qui detectent soit les virus grippaux et le virus respiratoire syncytial, soit un large panel d’agents susceptibles d’etre responsables de la pathologie. Ces dernieres, appelees techniques de PCR multiplex, permettent une approche syndromique de l’infection respiratoire. Leur cout eleve pour le laboratoire pose la question de leur place dans la prise en charge des patients en termes d’epargne antibiotique et d’isolement. Devant l’absence de recommandation, la strategie et l’equipement sont tres heterogenes. Perspectives La place de ces tests dans la prise en charge des patients doit etre evaluee par des etudes medicoeconomiques. Le role des differents virus, souvent detectes en coinfection, en particulier chez les enfants, merite egalement l’attention des virologues et des cliniciens. Conclusions Les nouvelles technologies de diagnostic, l’emergence du coronavirus SARS-CoV-2 et la mise sur le marche de nouveaux antiviraux vont impacter les recommandations de la prise en charge de ces infections.

International audience; Over two years (2012–2014), 719 nasopharyngeal samples were collected from 6-week- to 12-month-old infants presenting at the emergency department with moderate to severe acute bronchiolitis. Viral testing was performed, and we found that 98% of samples were positive, including 90% for respiratory syncytial virus, 34% for human rhino virus, and 55% for viral co-detections, with a predominance of RSV/HRV co-infections (30%). Interestingly, we found that the risk of being infected by HRV is higher in the absence of RSV, suggesting interferences or exclusion mechanisms between these two viruses. Conversely, coronavirus infection had no impact on the likelihood of co-infection involving HRV and RSV. Bronchiolitis is the leading cause of hospitalizations in infants before 12 months of age, and many questions about its role in later chronic respiratory diseases (asthma and chronic obstructive pulmonary disease) exist. The role of virus detection and the burden of viral codetections need to be further explored, in order to understand the physiopathology of chronic respiratory diseases, a major public health issue.

CERVOXY CLIN; International audience

International audience; We report a fatal case of measles inclusion-body encephalitis occurring in a woman from Romania with AIDS. After an extensive but unsuccessful diagnostic evaluation, a pan-pathogen shotgun metagenomic approach revealed a measles virus infection. We identified no mutations previously associated with neurovirulence.

Background Measles remains endemic worldwide, despite current vaccination recommendations, and is associated with high morbidity and mortality rates. We describe all cases hospitalized in Bordeaux University Hospital (BUH), the starting point of a national significant measles outbreak in 2017–2018. Methods In this retrospective study, we included all patients hospitalized in BUH from September 1, 2017, to May 31, 2018. Inclusion criteria were age >1 year, clinical symptoms, and biological confirmation by measles immunoglobulin M or measles reverse transcription polymerase chain reaction positivity. Results We included 171 patients. Most patients were immunocompetent; only 19% had preexisting medical histories. Most patients had rash and fever (97%), but some cases were atypical and difficult to diagnose. Köplik’s spots were reported in 66 cases (38%). The most frequent biological markers were blood inflammation markers (96%) and lymphopenia (81%). Unexpectedly, we found hyponatremia ( Conclusions During the BUH outbreak, measles was often complicated and sometimes atypical. Vaccination coverage was dramatically insufficient. We also describe vaccination failure cases that must be better investigated.

Prolonged measles virus detection in maternal saliva and blood was evidenced in 6 pregnant women. Maternal-fetal transmission was evidenced in 2 of 4 infants who were asymptomatic at birth, 21–24 weeks after maternal infection. Whereas peripartum congenital measles is severe, asymptomatic measles virus vertical transmission can occur earlier in pregnancy.

Résumé Le spectre des virus à tropisme respiratoire s’élargit et des maladies émergentes sont régulièrement décrites depuis une quinzaine d’années. L’origine de ces virus respiratoires émergents peut être zoonotique (par franchissement de barrière d’espèce, après mutations pour les virus à ARN tels que les virus aviaires influenza de type A ou les coronavirus), ou liée aux nouvelles techniques d’identification (métapneumovirus, bocavirus). Les relations entre bronchiolite et asthme sont désormais mieux appréhendées grâce au suivi prospectif de cohortes de nouveau-nés. Le rôle des rhinovirus devient prépondérant par rapport au virus respiratoire syncytial. L’identification de facteurs de prédispositions immunologiques, fonctionnelles, atopiques et génétiques, pour la survenue de l’asthme après une infection à rhinovirus suggère que l’infection virale révèle un terrain prédisposé plutôt qu’elle est à l’origine de l’asthme. Le rôle des bactéries dans l’histoire naturelle de l’asthme commence également à être mieux cerné. Les résultats de la cohorte danoise COPSAC ont montré la fréquence des identifications bactériennes au cours des épisodes sifflants avant 3 ans, ainsi que l’impact de la colonisation bactérienne à l’âge de un mois sur la survenue de l’asthme à l’âge de 5 ans. Le rôle des infections bactériennes dans l’asthme sévère du jeune enfant est également discuté.

In this study, we report a complete (clinical, radiological, and virological) sustained (1 year) response after nivolumab salvage therapy in a progressive multifocal leukoencephalopathy patient. Analyses of the cells infiltrate in a pretreatment brain biopsy suggest that parenchymal programmed cell death-L1+ macrophages could be the T-cells partnership in immune exhaustion and virus escape., PD-1 and its ligand are pivotal in the T-cell-exhaustion pathway involved in viral infections. In this paper, we report a complete response after nivolumab salvage therapy in a progressive multifocal leukoencephalopathy patient. Analyses of brain biopsy suggest that PD-L1+ macrophages could be the T-cells partnership in immune exhaustion and virus escape.

We described nosocomially clustered cases of measles secondary to a nonvaccinated index case occurring in a teenage psychiatric unit despite optimum vaccine coverage. Surveillance of this fully vaccinated closed cohort showed a 7% attack rate. Vaccination limited the risk of complicated measles and the onset of a large outbreak.

Background Seasonal influenza has a major individual and collective impact, especially among the elderly living in nursing homes. To prevent infection by influenza viruses, vaccination of residents and professionals is an essential measure. However, while the vaccination rates of residents are generally high (>85%), rates among professionals are generally approximately 20%. To evaluate the effectiveness of an intervention campaign on the improvement of the influenza vaccination rate of professionals, a regional intervention study was proposed for nursing homes during the 2014–15 season. Methods Cluster-randomized controlled trial (with a nursing home representing a cluster). In the intervention group, a campaign on influenza vaccination was offered to staff, combining different teaching aids in a multimodal approach. In the control group, no intervention was proposed. The primary endpoint was the rate of influenza vaccination among staff. Before and after the study, professionals were asked to complete short questionnaires on their perceptions of influenza vaccination. A multilevel analysis was carried out to compare the vaccination rates between the 2 groups and their evolution before/after the winter period. Results A total of 32 nursing homes were randomized, and 6 were excluded. Initial vaccination rates were 27.6% in the intervention group and 24.2% in the control group (p = 0.16). After the study, these rates increased to 33.7% and 22.9%, respectively, which was a relative difference of +22.1% in the intervention group compared to −5.4% in the control group, p = 0.0025. Conclusions Despite professionals’ reluctance to be vaccinate, participation in a promotional campaign with a pragmatic approach has increased the rate of influenza vaccination. The approach will be offered to all nursing homes in the region after revision of the tools to enhance their ease of use and pedagogical messages focused on the direct benefits to professionals.

International audience

International audience

International case definitions recommended by the Centers for Disease Control and Prevention (CDC), the European Centre for Disease Prevention and Control (ECDC), and the World Health Organization (WHO) are commonly used for influenza surveillance. We evaluated clinical factors associated with the laboratory-confirmed diagnosis of influenza and the performance of these influenza case definitions by using a complete dataset of 14,994 patients with acute respiratory infection (ARI) from whom a specimen was collected between August 2009 and April 2014 by the Groupes Régionaux d’Observation de la Grippe (GROG), a French national influenza surveillance network. Cough and fever ≥ 39 °C most accurately predicted an influenza infection in all age groups. Several other symptoms were associated with an increased risk of influenza (headache, weakness, myalgia, coryza) or decreased risk (adenopathy, pharyngitis, shortness of breath, otitis/otalgia, bronchitis/ bronchiolitis), but not throughout all age groups. The WHO case definition for influenza-like illness (ILI) had the highest specificity with 21.4%, while the ECDC ILI case definition had the highest sensitivity with 96.1%. The diagnosis among children younger than 5 years remains challenging. The study compared the performance of clinical influenza definitions based on outpatient surveillance and will contribute to improving the comparability of data shared at international level.

Measles outbreaks are regularly reported in European countries despite efforts to improve vaccination coverage. In January 2016, an outbreak occurred in a refugee settlement in Calais, France. We report here the complete genome sequence of a wild-type measles virus isolated from a health care worker (MVi/Calais. FRA/01.16) infected during this outbreak.

Recommendations for the use of diagnostic testing in low respiratory infection in children older than 3 months were produced by the Groupe de Recherche sur les Avancees en Pneumo-Pediatrie (GRAPP) under the auspices of the French Paediatric Pulmonology and Allergology Society (SP(2)A). The Haute Autorite de sante (HAS) methodology, based on formalized consensus, was used. A first panel of experts analyzed the English and French literature to provide a second panel of experts with recommendations to validate. Only the recommendations are presented here, but the full text is available on the SP(2)A website.

Background. No information is available on the viral etiology of upper respiratory tract infections in Cameroon. Methods. We prospectively enrolled outpatients with influenza-like illness (ILI) presenting at 14 sentinel clinics located across the country from January through December 2009. The specimens were tested using real-time and multiplex reverse-transcription polymerase chain reaction methods for the detection of 15 RNA respiratory viruses. Results. We detected at least 1 respiratory virus in 365 of 561 specimens (65.1%). Overall, influenza virus was the most commonly detected virus (28.2% of specimens), followed by human rhinovirus (17.8%); parainfluenza virus (PIV) types 1–4 (7.5%); enterovirus (5.9%); respiratory syncytial virus (RSV; 5.7%); human coronavirus (HCoV) OC43, 229E, NL63, and HKU1 (5.3%); and human metapneumovirus (HMPV; 5.0%). RSV (26 of 31 specimens [83.9%]), PIV (30 of 39 [76.9%]), and HRV (64 of 99 [64.6%]) were most common among children

Measles is one of the most contagious diseases implicated in an outbreak in Western Europe for a few years. In France, the outbreak began in 2008, especially in the southern regions. Most of the cases are mild but deleterious effects can occur in infants, during the pregnancy, or in immunosuppressive patients. Pneumonia and encephalitis are the major complications in these patients. Measles illness during pregnancy results in a higher risk of premature labor, spontaneous abortion, and low-birthweight infants. Death from measles was reported in approximately 0.3% of cases. The National Reference Center for measles and paramyxoviridae is implicated in biological confirmation of cases and epidemiologic surveillance. Virologic diagnosis consists in a direct detection of viral RNA (real time PCR), or indirect detection of the immune response (IgM and IgG) in saliva samples. The measles genotype is determined by sequencing a 450 nucleotids fragment of the N gene, as the World Health Organization (WHO) recommendation. This molecular analysis helps determine the geographic origin of the virus and the viral strains circulating in the country. In 2011, 3105 specimens were received. PCR was positive for 1990 patients (64%), with a large majority of children younger than 2 years of age and young adults. The most important cause of measles resurgence is low vaccination coverage. The role of pediatricians is important in order to achieve the goal of 95% of vaccination coverage established by the WHO.

The objective of this study is primarily to compare the performance of the VIDAS(®) Measles immunoglobulin (Ig)G assay to that of two other serological assays using an immunoassay technique, Enzygnost(®) Anti-measles Virus/IgG (Siemens) and Measles IgG CAPTURE EIA(®) (Microimmune). The sensitivity and the agreement of the VIDAS(®) Measles IgG assay compared to the Enzygnost(®) Anti-measles Virus/IgG assay and the Measles IgG CAPTURE EIA(®) assay are 100%, 97.2% and 99.0%, 98.4%, respectively. The very low number of negative sera for IgG antibodies does not allow calculation of specificity. As a secondary objective, we have evaluated the ability of the VIDAS(®) Measles IgG assay to measure anti-measles virus IgG antibody avidity with the help of the VIDAS(®) CMV IgG Avidity reagent, using 76 sera from subjects with measles and 238 other sera. Different groups of populations were analyzed. In the primary infection measles group, the mean IgG avidity index was 0.16 (range of 0.07 to 0.93) compared to 0.79 (range of 0.25 to 1) in the serum group positive for IgG antibodies and negative for IgM. These data allow to define a weak anti-measles virus IgG antibody avidity as an avidity index (AI)0.3 and a strong avidity as an AI0.6. The VIDAS(®) Measles IgG assay has a performance equivalent to that of other available products. Its use, individual and quick, is well adapted to testing for anti-measles immunity in exposed subjects.

International audience; BACKGROUND: Severe acute respiratory illness (SARI) is recognized as an important cause of morbidity, mortality, and hospitalization among children in developing countries. Little is known, however, in tropical countries like Cameroon about the cause and seasonality of respiratory infections, especially in hospitalized settings.OBJECTIVES: Our study investigates the viral etiology and seasonality of SARI in hospitalized children in Yaounde, Cameroon.METHODS: Prospective clinic surveillance was conducted to identify hospitalized children aged ≤15 years presenting with respiratory symptoms ≤5-day duration. Demographic and clinical data, and respiratory specimens were collected. Nasopharyngeal samples were tested for 17 respiratory viruses using a multiplex polymerase chain reaction. The viral distribution and demographic data were statistically analyzed.RESULTS: From September 2011 through September 2013, 347 children aged ≤15 years were enrolled. At least one virus was identified in each of 65·4% children, of which 29·5% were coinfections; 27·3% were positive for human adenovirus (hAdV), 13·2% for human respiratory syncytial virus (hRSV), 11·5% for rhinovirus/enterovirus (RV/EV), 10·6% for human bocavirus (hBoV), 9·8% for influenza virus (Inf), 6·6% for human parainfluenza virus (hPIV), 5·7% for human coronavirus (hCoV), and 2·3% for human metapneumovirus (hMPV). While hRSV showed seasonal patterns, hAdV and RV/EV were detected throughout the year and no evident temporal patterns were observed for the remaining viruses.CONCLUSION: Respiratory viruses were associated with a high burden of hospitalizations among children in Cameroon. Nevertheless, additional studies evaluating asymptomatic Cameroonian children will be important in understanding the relationship between viral carriage and disease.

Background: The disease burden caused by recently identified respiratory viruses like HCoV-NL63 is unknown. Objectives: We determined the burden of disease due to HCoV-NL63 infections using the population-based PRI.DE cohort of children under the age of 3 with lower respiratory tract infections (LRTIs). Study design: In total 1756 respiratory samples, from hospitalized children or children who visited the outpatient clinic, were tested for HCoV-NL63. Sampling covered a period of 2 years and the frequency of infection in different years was compared to other Western European studies that tested for this virus in 2 or more consecutive years. Results: Sixty-nine samples were HCoV-NL63 positive, 35 were with high loads, and of these 25 were single HCoV-NL63 infections. Based on the number of children with high HCoV-NL63 infection and no additional infection, the overall annual incidence in outpatients was 7 per 1000 children per year (95% confidence interval (CI) 3-13 per 1000 children per year), which can be extrapolated to an absolute number of 16,929 visits to the physician due to an HCoV-NL63 infection in Germany per year. The estimated hospitalization rate is 22 per 100,000 children (95% CI: 7-49 per 100,000 children per year). This number reflects 522 HCoV-NL63 children in Germany per year. A large year-to-year difference in HCoV-NL63 infection frequency was observed. Combining these data with those of other studies in Western Europe revealed that HCoV-NL63 infections follow a 2-year inter-epidemic period with peaks of infection in the winters of 2000/2001, 2002/2003 and 2004/2005 (p

Acute respiratory infection is one of the leading causes of child morbidity, especially in developing countries. Viruses are recognized as the predominant causative agents of acute respiratory infections. In Senegal, few data concerning the causes of respiratory infections are available, and those known relate mainly to classical influenza infections. Clinical and virological surveillance of acute respiratory infections was carried out in a rural community in children less than 5 years old. A standardized questionnaire was used and a nasopharyngeal swab sample was collected from each patient. These samples were tested for the detection of 20 respiratory viruses by multiplex RT‐PCR or by viral culture. A total of 82 acute respiratory episodes were included, and 48 (58.5%) were found to be positive, with a total of 55 viral detections; several samples were positive for two (n = 5) or 3 (n = 1) viruses. Ten different viruses were identified: influenza viruses A, B, and C (n = 25), human respiratory syncytial virus type A (n = 13), rhinoviruses (n = 8), human coronaviruses type 229E and NL63 (n = 6), parainfluenza viruses 3 and 4 (n = 2), and bocavirus (n = 1). These results provide evidence on the importance and the diversity of viruses as causative agents of acute respiratory infections in children living in a rural community in Senegal. The establishment of sentinel surveillance sites could help estimate the burden of acute respiratory infection in the pediatric population and should help prepare the health care systems to identify and respond to new viral respiratory emergencies. J. Med. Virol. 82:866–872, 2010. © 2010 Wiley‐Liss, Inc.

From November 2004 to April 2007, specimens were obtained from 2,281 patients with acute respiratory tract illness in Normandy, France. Eighteen strains of influenza C virus were detected in these samples using a combined tissue culture/RT‐PCR diagnostic method. Most patients with influenza C virus infection (13/18) were infants or young children (

The relationships between host factors, virus strain, viral load, and illness severity in respiratory syncytial virus (RSV)-induced bronchiolitis are poorly defined. These relationships were evaluated prospectively in 81 previously healthy infants hospitalized with RSV bronchiolitis. Disease severity was determined by the respiratory rate, the duration of hospitalization, and whether patients during their hospitalization required pediatric intensive care unit admission or mechanical ventilation. RSV typing into subgroup A and B was obtained by RT-PCR-hybridization assay. The nasopharyngeal RSV viral loads were measured by real-time quantitative RT-PCR. Disease severity correlated significantly with the presence of risk factor (estimated gestational age

Background Detection of enteroviral nucleic acid in cerebrospinal fluid (CSF) specimens has been demonstrated to improve the management of patients with aseptic meningitis. Objective To develop on the LightCycler (LC) instrument a real-time RT-PCR assay based on TaqMan technology for the detection of enteroviruses (EV) in cerebrospinal fluid (CSF) specimens. Study design After evaluation of the analytical performances, seventy-four CSF samples collected prospectively from patients who have been suspected for a clinical diagnosis of meningitis were evaluated by two LC real-time RT-PCR assays and one conventional RT-PCR assay. Results Our assay detected all 30 different EV species tested, whereas no reactivity was observed with other neurotropic viruses. The analytical sensitivity of both LC RT-PCR real-time assays was 1 TCID50 for LC one-step and two-step RT-PCR assays. Results for LC one-step and LC two-step RT-PCR were compared to results of the conventional RT-PCR: of the 74 CSF specimens tested, 11 were positive and 56 were negative by all methods. Four other specimens were positive for EV by at least two of the methods (including the LC two-step RT-PCR and the conventional RT-PCR), two other CSF specimens were positive by the LC two-step RT-PCR assay only, and another one CSF specimen was positive by the LC one-step RT-PCR assay only. No CSF specimens were negative by the LC two-step RT-PCR assay and positive by the conventional RT-PCR assay. The sensitivity, specificity, positive and negative predictive values of both LC RT-PCR assays by using conventional RT-PCR as the “gold standard” were, respectively, 73.3, 98.3, 91.7, 93.5% for the LC one-step RT-PCR and 100, 96.6, 88.2, 100% for the LC two-step RT-PCR. There was substantial agreement between the three assays (k = 0.80). Conclusions The LC two-step RT-PCR assay is a rapid, sensitive and reliable method which can be routinely performed with CSF samples for diagnosis of EV infection and is an important improvement for optimal patient management.

Coronavirus NL63 was found in hospitalized children with upper and lower respiratory infections., The human coronavirus NL63 (HCoV-NL63) was first identified in the Netherlands, and its circulation in France has not been investigated. We studied HCoV-NL63 infection in hospitalized children diagnosed with respiratory tract infections. From November 2002 to April 2003, we evaluated 300 respiratory specimens for HCoV-NL63. Of the 300 samples, 28 (9.3%) were positive for HCoV-NL63. The highest prevalence was found in February (18%). The main symptoms were fever (61%), rhinitis (39%), bronchiolitis (39%), digestive problems (33%), otitis (28%), pharyngitis (22%), and conjunctivitis (17%). A fragment of the spike protein gene was sequenced to determine the variety of circulating HCoV-NL63. Phylogenetic analysis indicated that strains with different genetic markers cocirculate in France.

Objective To determine the incidence, risk factors, and clinical relevance of viral ventilator-associated pneumonia (VAP) in an adult intensive care unit (ICU). Design Prospective observational study. Setting A 22-bed adult medical ICU in a university hospital. Patients All consecutive adult patients ventilated more than 48 h in a 9-month period including regular seasonal viral infections. Interventions A tracheobronchial aspirate upon enrollment and at the time of VAP suspicion. Measurements and results All respiratory specimens were tested in culture, indirect immunofluorescence assay, and PCR or RT-PCR for virological assessment. Patients were followed until ICU discharge or death. One hundred thirty-nine patients were included. Upon enrollment, a respiratory virus was detected in the tracheobronchial aspirate in 25% of patients (35 of 139). The incidence of VAP, defined according to clinical daily evaluation, was 28% (39 of 139 patients). A bacteria was documented in 74% of cases, whereas no case of a causative viral infection was encountered among VAP patients; however, herpes simplex virus type-1 (HSV 1) infection was detected in respiratory specimens of 31% of VAP (12 of 39). Conclusions We found a high incidence of HSV-1 infection in VAP patients; however, nosocomial viral VAP is likely to be rare in ICU, as assessed by the absence of respiratory virus-induced VAP identified in this prospective cohort study.

Asthma prevalence has increased significantly; this increase appears to be relatively recent. The observed increases in this prevalence cannot be genetic in origin; on the other hand lifestyles have changed in many ways for environment or feeding. The hygiene hypothesis must be distinguished in terms whether one is addressing asthma or atopy. Although there are associations between asthma and atopy, they are not interchangeable. The pathogenic links between childhood infections and asthma require an undisputed definition of asthma. But many wheezing syndromes have been recognized in the pediatric age group, some overlap exists between these various phenotypes. The age at first severe infection seems to be crucial in determining the outcome of reinfection and suggests that the environment of the neonatal lung is a major determinant of disease and allergic patterns in later life.

Background: Preemptive antiviral treatment of Human Cytomegalovirus (HCMV) disease is a major goal in the management of organ transplant patients . It requires sensitive diagnostic methods. Automated real-time PCR systems have been recently proposed to monitor HCMV infection in such patients. Objective: Objectives of this study was to compare a real-time quantitative PCR on whole blood with the HCMV pp65 antigenemia assay in renal transplant recipients, and also to evaluate two different DNA extraction methods. Study design: A total of 248 specimens from 21 patients were tested by quantitative pp65 antigenemia and quantitative real-time PCR. DNA was extracted from whole blood samples using two different methods: a conventional column manual assay and an automated system. Results: Quantification of HCMV DNA using the two extraction methods showed highly similar results (Spearman rank test, r =0.863). We found a significant correlation between DNA quantification by real-time PCR in whole blood and pp65 antigenemia test (Spearman rank test, r =0.767). This correlation was not modified when the HCMV DNA results were normalized by quantification of the albumin cellular gene. In eight patients, HCMV infection was detected earlier with quantitative PCR than with the antigenemia test (mean delay of 11.25 days). HCMV DNA load equivalent of 50 pp65 positive cells/200 000 polymorphonuclear leukocytes (PMNLs) is log 4.095 copies per ml of blood. Conclusions: Real-time PCR in whole blood is a sensitive method for estimating the HCMV genome load in renal transplant patients, and is more rapid and practicable than using PMNLs for pp65 antigenemia tests.

Human respiratory syncytial virus (RSV) is the major viral cause of lower respiratory tract disease in children. Infections with the virus occur as annual winter epidemics in temperate climates, placing considerable pressure on the provision of hospital beds. Most molecular epidemiological studies have, until now, focused on isolates from infants in industrialised countries. No data have been available with regard to RSV strains from northern Africa.In this report, a recent RSV outbreak in Tunisia was studied and results showed that 176 of 815 (21.6%) nasopharyngeal aspirates collected from hospitalised children were RSV-positive by immunofluorescence assay. This RSV outbreak showed a temperature-dependant pattern (P = 0.026) but no significant association with rainfall. A total of 73 RSV-positive samples were tested by two reverse-transcription-polymerase chain reaction assays (RT-PCR): RT-PCR-1, which amplifies the RNA of all RSV strains, and RT-PCR-2, which allows subgroup classification of RSV. Analysis by hybridisation assay of RT-PCR-2-amplified 1B protein gene products showed a higher prevalence of group B RSV than that of group A (82.5% vs. 17.5% among the typed strains). Knowledge of the variants is important in terms of both diagnosis and definition of a vaccine composition. J. Med. Virol. 72:683–687, 2004. © 2004 Wiley-Liss, Inc.

Respiratory syncytial virus (RSV) is rarely searched for in respiratory infections in adults. This study assessed its frequency and diagnosis.Three separate studies were conducted in adults presenting with (1) a flu-like illness, (2) a lower respiratory tract infection in the community, and (3) a severe pneumonia requiring hospitalisation. The diagnosis of RSV infection was sought by PCR in all cases, and compared to antigen detection and culture in two studies.RSV was identified in 20 (11.7%) of 170 influenza-vaccinated adults suffering from flu-like symptoms. In the 270 cases of non-severe lower respiratory tract illnesses in the community, viruses were identified in 86 (31.8%) cases, with RSV accounting for 13 (4.8%). In the 164 cases of acute bronchitis, a virus was detected in 64 (36.7%) of which 11 (6.3%) were RSV, 37 (21.3%) rhinovirus, 5 influenza viruses A and B, and 12 other viruses. In the 60 cases of infective exacerbations of chronic bronchitis, rhinovirus was detected in 9 (15%) and para-influenza 3 virus in 2 cases. In the 21 acute pneumonia's, 1 RSV, 1 influenza virus A and 2 rhinovirus cases were detected as well as 1 RSV, 1 parainfluenza 3 viruses and 4 rhinovirus cases in the 11 lower respiratory tract illnesses in patients with pre-existing lung disease. There were overall 19 viral and bacterial associated infections. Finally, in the 51 acute pneumonias hospitalised with respiratory distress syndrome, a virus was identified in 17 (33.3%) cases, including 3 (5.5%) RSV, 6 influenza A, 3 rhinovirus, 2 adenovirus, 2 herpes simplex virus and 1 cytomegalovirus. There were 6 bacterial-associated infections, and 4 were hospital-acquired. All RSV-infected patients were old people and had chronic pulmonary or cardiac disease.In adults, RSV is a frequent cause of flu-like symptoms. It can sometimes cause lower respiratory tract illness, which can be severe, and should be considered in the differential diagnosis in such cases. The PCR method is a particularly effective diagnostic test, but as yet is not routinely available.

Resume Objectifs. – Les rhinovirus sont les principaux agents du rhume, mais la frequence et la gravite de leurs autres atteintes sont mal connues. Le but de ce travail est d’etudier l’expression clinique et la gravite potentielle des infections a rhinovirus de l’enfant hospitalise. Methodes. – De septembre 1998 a octobre 2000, nous avons etudie retrospectivement une cohorte hospitaliere de 211 enfants infectes par un rhinovirus, celui-ci etant recherche par culture ou RT-PCR. Resultats. – Chez les enfants infectes par un rhinovirus, 60 cas (28,4 %) d’atteintes ORL, 81 cas (38,4 %) de bronchiolites, 25 cas (11,9 %) de pneumopathies et 12 cas (4,7 %) de crises d’asthme aigues ont ete identifies. Parmi les signes cliniques on releve 32 % de râles sibilants, 37 % de râles bronchiques et 29 % de signes de detresse respiratoire ; 40 % des radiographies pulmonaires sont anormales. Huit enfants ont ete transferes en reanimation et deux sont decedes. Vingt-cinq enfants (11,8 %) avaient une infection a rhinovirus d’origine nosocomiale. Dans 19 cas le rhinovirus (9 %) etait associe a un autre virus : 3 VRS, 2 virus influenza, 8 virus para-influenza, 2 adenovirus, 4 enterovirus ; 19 enfants (9 %) presentent une surinfection bacterienne. L’identification virologique des 211 cas d’infections a rhinovirus a ete faite par isolement en culture dans 112 cas (53 %) et lorsque celui-ci etait negatif par RT-PCR dans 99 cas (47 %). Conclusion . – En eliminant les cas de co-infections virales et bacteriennes, les tableaux cliniques des infections a rhinovirus isolees de l’enfant sont dans notre etude les suivants : 25,6 % d’infections ORL, 25,6 % de bronchiolites ou bronchites, 6,2 % de pneumopathies et 5,7 % de crises d’asthme. Le diagnostic virologique est beaucoup ameliore par les outils moleculaires.

The 2 groups of human coronaviruses (HCoVs) represented by the prototype strains HCoV 229E and HCoV OC43 are mostly known as viruses responsible for common cold syndrome. HCoVs are difficult to detect, and epidemiological data are rare. From October 2000 through April 2001, we tested 1803 respiratory samples for HCoV by reverse-transcriptase polymerase chain reaction. From 8 February through 27 March 2001, HCoV OC43 was detected in samples obtained from 30 (6%) of 501 patients. The other viruses detected were respiratory syncytial virus (6.1%), parainfluenza virus 3 (1%), influenza virus A (7.8%), influenza virus B (7.2%), rhinovirus (6.4%), enterovirus (1%), and adenovirus (2%). Infection with HCoV OC43 was detected in patients of all age groups. The following clinical symptoms were noted: fever (in 59.8% of patients), general symptoms (in 30%), digestive problems (in 56.8%), rhinitis (in 36.6%), pharyngitis (in 30%), laryngitis (in 3.3%), otitis (in 13.3%), bronchitis (in 16.6%), bronchiolitis (in 10%), and pneumonia (in 6.6%). This study shows that an outbreak of HCoV OC43 respiratory infection was responsible for the lower respiratory tract symptoms observed in nearly one-third of patients identified by active surveillance for coronavirus infection.

To the Editor: The reemergence of measles in Europe is a reminder of the forgotten risk for severe illness and death associated with this disease in industrialized countries. Since 2008, >20,000 measles cases and 9 measles-associated deaths (in 7 immunocompromised and 2 immunocompetent persons) have been reported to the French Institute for Public Health. Among these cases, the reported causes of death were pneumonia and/or acute respiratory distress syndrome (ARDS) (n = 7) and encephalitis (n = 2). All patients except 1, an immunocompromised patient, had the typical morbillous rash. We report another fatal case of measles, with intractable ARDS but no rash, in an apparently immunocompetent adult. The patient was a 29-year-old woman in Grenoble, France, who smoked but had no relevant medical history except an episode of depression. In 2011, she sought care for fever, cough, coryza, diarrhea, and a 10-kg weight loss over 10 days. A general practitioner empirically prescribed pristinamycin and oral prednisone (60 mg/d for 5 d) for sinusitis. Five days later, the patient was admitted to the hospital because of persistent signs and symptoms. Physical examination at admission (day 1) detected fever (38.5°C), dyspnea, and a low body mass index of 17.5 kg/m2. Hematologic tests showed nonregenerative anemia (hemoglobin concentration 9 g/dL) and leukopenia (2.2 × 109 leukocytes/L) with profound lymphopenia (0.2 × 109 lymphocytes/L) and mild thrombocytopenia (135.0 × 109 platelets/L). A chest radiograph showed bilateral diffuse interstitial infiltrates. Antimicrobial therapy with levofloxacin and ceftriaxone was started. On day 2, several examinations were conducted to explore the possibility of underlying immunosuppressive disease. Body scans showed no adenopathy or lesions suggestive of cancer. HIV test result was negative. General immunologic test results were within normal limits (immunoglobulin quantification, autoantibody testing) or consistent only with an acute viral infection (serum protein electrophoresis). A bone marrow biopsy sample indicated isolated erythroblastopenia with no abnormality of other cell lineages (PCR for parvovirus B19 was negative). On day 3, because of severe respiratory failure, the patient was transferred to the intensive care unit, where the diagnosis of ARDS was confirmed and mechanical ventilation was started. Treatment with tazocillin/tazobactam, ciprofloxacin, amphotericin B, and acyclovir was also started. Microbiological findings from bronchoalveolar lavage (BAL) samples were repeatedly negative for bacteria, mycobacteria, fungi, and Pneumocystis jirovecii. Cytology of BAL samples showed an acute inflammatory response with atypical epithelial cells, supporting a diagnosis of viral infection. However, none of 14 respiratory viruses or human herpesviruses type 1, 3, 4, 5, or 6 were recovered from BAL samples by PCR. Blood and urine culture results were repeatedly negative, as were serologic test results for Legionella spp., Mycoplasma pneumoniae, and Chlamydia pneumoniae. On day 5, because of refractory ARDS, venoarterial extracorporal membrane oxygenation was started. On day 6, the results of a broad serologic investigation demonstrated isolated IgM against measles virus. The patient was additionally given ribavirin, corticosteroids, and intravenous immunoglobulin. On day 10, the lymphocyte level had returned to reference range and the anemia had become regenerative. However, the patient’s respiratory condition did not improve, and after 2 weeks of the oxygenation therapy, the patient died of hemorrhagic shock. Her parents declined an autopsy. PCR testing of the patient’s saliva by the French National Reference Center confirmed the presence of measles virus. Retrospective testing of serum, bone marrow, and BAL specimens collected during days 2–20 of hospitalization demonstrated measles virus RNA. The strain was identified as genotype D4, which is the epidemic strain circulating in France and elsewhere in Europe (1). The patient had no history of enanthem (Koplik spots) or morbilliform rash before or after symptom onset and no documented history of measles vaccination. Deaths from measles with pneumonia or ARDS but without rash have been reported but mostly in patients with deficient cell-mediated immunity (2–6). Despite all our testing, we found no indications of an underlying immunosuppressive disease in this patient; however, we cannot categorically rule out this possibility, especially that of a primary immunodeficiency. The initial therapy with corticosteroids and the patient’s weight loss could also have interfered with her cellular immune response. The diagnosis of ARDS caused by measles was supported by detection of the measles genome in BAL samples and body fluids in the absence of any other pathogen, but pulmonary superinfection with unidentified pathogens could not be ruled out. Detection of the measles genome and isolated erythroblastopenia in the bone marrow biopsy sample is consistent with reports that measles virus can infect erythroid progenitors and interfere indirectly with hematopoiesis (7,8). Although ribavirin and passive immunotherapy have been reported to aid in recovery from severe measles pneumonia, their clinical efficacy is still unproven (9,10); and for the patient reported here, they were probably used too late. This unusual case underscores the need for physicians to consider the diagnosis of measles, even in the absence of classical clinical features, during measles outbreaks. It also reemphasizes the insufficient vaccination coverage against measles in France.

The aims of this study were to assess the sociodemographic, epidemiological, clinical, and biological characteristics of French patients co-infected with human immunodeficiency virus-hepatitis C virus (HIV-HCV), as well as the management of their HCV infection. Data on 509 HIV-HCV co-infected patients, followed up at six French University Hospitals, were collected using a questionnaire. Student's t-test, Pearson's chi-square, Fisher's exact, and Fisher-Freeman-Halton's exact tests were used. The mean age of the patients was 38.3 years, and the male to female sex ratio 2.08; 88% of patients were born in Metropolitan France, and 20% were dependent on health benefits; 74% were intravenous drug users and 14% blood or blood product recipients. Forty-seven percent were in CDC classification stage A, 18% had a CD4+ count of200, and 79% were undergoing current antiretroviral treatment. HCV RNA was positive in 84% (50% type 1, 13% untypable). Forty-four percent had normal alanine aminotransferase (ALT) levels, 24% alcohol consumption15 g/day, and 51% had undergone liver biopsy (10% of which had cirrhosis). Histological grade was not related to ALT level or CD4+ count. Overall, 40% of patients had been treated for HCV infection. HCV treatment was significantly associated with performance of liver biopsy, histological grade, ALT level, CD4+ count, Centers for Disease Control (CDC) classification, but not with age or alcohol consumption. Rate of early response to treatment was fifty percent among patients treated with bitherapy. Eighty-nine percent of all patients with previous or current anti-HCV treatment had undergone liver biopsy. In conclusion, despite the difficulties in managing hepatitis C in HIV-infected patients, almost one-half of all patients in this study had received anti-HCV treatment.

Wheezing associated with upper respiratory tract infections is common in children. Using conventional techniques (viral culture and immunofluorescence) and molecular techniques (PCR), we studied the prevalence of viral, Chlamydia pneumoniae (CP) and Mycoplasma pneumoniae (MP) infections in 118 children hospitalised for acute asthma exacerbation. A virus was identified by conventional techniques in 40 of the 118 nasal swabs (34%), while PCR allowed identification of virus CP and MP in 80 samples (68%). Combination of both techniques allowed identification of an infectious agent in 91 cases (77%). More than one agent was isolated in 15 cases (23%). Rhinovirus (RV) (45%) were prevalent, followed by respiratory syncytial virus (RSV) (28%) and enterovirus (8.5%). RV and RSV have a similar prevalence (42% and 36% respectively) before two years of age, as compared with 66% and 27% respectively in older children. CP and MP were identified by PCR in only 6 cases. Molecular techniques of identification demonstrated a clear advantage in sensitivity compared with conventional techniques. The high prevalence of RV and RSV infections is remarkable, while CP and MP do not seem particularly involved in children acute asthma exacerbation.

Importance Acute bronchiolitis is the leading cause of hospitalization among infants. Previous studies, underpowered to examine hospital admission, have found a limited benefit of nebulized hypertonic saline (HS) treatment in the pediatric emergency department (ED). Objective To examine whether HS nebulization treatment would decrease the hospital admission rate among infants with a first episode of acute bronchiolitis. Design, Setting, and Participants The Efficacy of 3% Hypertonic Saline in Acute Viral Bronchiolitis (GUERANDE) study was a multicenter, double-blind randomized clinical trial on 2 parallel groups conducted during 2 bronchiolitis seasons (October through March) from October 15, 2012, through April 15, 2014, at 24 French pediatric EDs. Among the 2445 infants (6 weeks to 12 months of age) assessed for inclusion, 777 with a first episode of acute bronchiolitis with respiratory distress and no chronic medical condition were included. Interventions Two 20-minute nebulization treatments of 4 mL of HS, 3%, or 4 mL of normal saline (NS), 0.9%, given 20 minutes apart. Main Outcomes and Measures Hospital admission rate in the 24 hours after enrollment. Results Of the 777 infants included in the study (median age, 3 months; interquartile range, 2-5 months; 468 [60.2%] male), 385 (49.5%) were randomized to the HS group and 387 (49.8%) to the NS group (5 patients did not receive treatment). By 24 hours, 185 of 385 infants (48.1%) in the HS group were admitted compared with 202 of 387 infants (52.2%) in the NS group. The risk difference for hospitalizations was not significant according to the mixed-effects regression model (adjusted risk difference, –3.2%; 95% CI, –8.7% to 2.2%; P = .25). The mean (SD) Respiratory Distress Assessment Instrument score improvement was greater in the HS group (–3.1 [3.2]) than in the NS group (–2.4 [3.3]) (adjusted difference, –0.7; 95% CI, –1.2 to –0.2; P = .006) and similarly for the Respiratory Assessment Change Score. Mild adverse events, such as worsening of cough, occurred more frequently among children in the HS group (35 of 392 [8.9%]) than among those in the NS group (15 of 384 [3.9%]) (risk difference, 5.0%; 95% CI, 1.6%-8.4%; P = .005), with no serious adverse events. Conclusions and Relevance Nebulized HS treatment did not significantly reduce the rate of hospital admissions among infants with a first episode of acute moderate to severe bronchiolitis who were admitted to the pediatric ED relative to NS, but mild adverse events were more frequent in the HS group. Trial Registration clinicaltrials.gov Identifier:NCT01777347

The role for Mycoplasma pneumoniae and Chlamydophila pneumoniae in lower and upper respiratory tract infections in childhood increased by use of specialised diagnostic techniques, more and more performant for the early diagnosis of these infections. However, the prevalence of M. pneumoniae and C. pneumoniae as a cause of severe pneumoniae among hospitalized children has been rarely described. We report a case of M. pneumoniae et C. pneumoniae coinfection in a 10-year-old child hospitalized with a respiratory distress.

The aim of this study was to evaluate residual viral replication by assessing the HIV load of circulating infected cells in patients given an effective antiprotease-containing treatment for 1 year. PBMC HIV RNA and HIV DNA was quantified by techniques standardized and evaluated by interlaboratory quality control testing. Viral markers identified in a multicenter study were validated in a cross-sectional study of 121 patients beginning treatment. A longitudinal study of 3 viral markers was carried out in 18 patients, each of whom had fewer than 200 copies of HIV RNA per milliliter of plasma after 12 months of treatment. The cross-sectional study showed that viral replication in PBMCs was correlated with the number of circulating infected cells (Spearman rank correlation; p = 0.0001, r = 0.35) and the concentration of virus particles in the plasma (Spearman; p = 0.0001, r = 0.54). The longitudinal study showed that the decrease in HIV RNA levels was smaller in PBMCs than in the plasma. The largest decrease in HIV DNA levels after 12 months of treatment was recorded in patients with low levels of intracellular replication (Spearman; p = 0.005, r = 0.69). PBMC HIV RNA and HIV DNA levels were very informative markers, complementary to plasma HIV RNA levels. They should be used in future trials evaluating the long-term efficacy of new associations of highly active antiretroviral treatments.

Resume Les infections respiratoires virales sont une cause majeure de sifflements chez le nourrisson. Un virus peul etre identifie dans 80 % a 85 % des crises d'asthme chez l'enfant. Afin de mieux comprendre le retentissement des infections respiratoires sur l'asthme, les recherches se sont portees sur la Symptomatologie clinique, la fonction respiratoire et l'inflammation generee. Les cytokines ont un role cle dans ce processus et l'atteinte de l'epithelium respiratoire par le virus est probablement la source principale de leur production lors de la phase aigue de l'infection. Les cellules lymphocytaircs T sont les cellules organisatrices de la reponse immunitaire tant visa-vis des allergenes que des virus; elles regulent les cellules effectrices cytotoxiques et pro-inflammaloires. L'immunopalhogenie de l'atteinte pulmonaire lors des infections virales fait vraisemblablement intervenir les cellules T. mais leur role dans la genese de l'asthme reste discute. Les infections survenant durant la prime enfance ont une importance dans la mise en place des phenomenes d'immunoregulalion et probablement dans la survenue d'une allergie ou d'un asthme. L'atopie est caracterisee par une reponse lymphocvlaire Th-2 importante face aux allergenes usuels avec une secretion de cytokines. notamment l'IL-4 et l'IL-5 qui favorisent respectivement la synthese d'IgE et la multiplication des polynucleaires eosinophiles. a l'oppose, les infections durant l'enfance induisent habituellement une reponse de type Th-1 caracterisee par la secretion de l'interferon % qui favorise I aclie antivirale des cellules effectrices. Il existe une reponse mutuellement antagoniste entre ces deux types cellulaires.

Although signs and symptoms may become severe, most viral respiratory infections of infancy are self-limited and improvement usually occurs within several days. Patients hospitalized with viral pneumonia usually require supportive therapy, including oxygen and fluids, and eventually mechanical ventilation. Bacterial superinfection can occur, accompanied by purulent sputum production and isolation of pathogenic bacteria from sputum. Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis and Staphylococcus aureus are the most common secondary invaders. Appropriate antibiotherapy must be administrated after cultures. There is no evidence that prophylactic antibiotherapy is of any use to prevent bacterial superinfection in viral pneumonia.

International audience; During the 2011 measles outbreak in Paris (France), patients with clinical suspicion of measles were tested for virological confirmation of measles virus (MV) infection. To assess the practical value of molecular diagnosis in an epidemic setting, 171 oral fluid samples and 235 serum samples collected from 270 patients were tested prospectively for MV-RNA using a novel one-step real-time RT-PCR assay including an internal control. Serum samples were also tested for MV-specific IgG and IgM antibodies. MV infection was confirmed by detection of MV-RNA and/or MV-IgM for 229 of the 270 patients. The results for the 102 cases with both serum and oral fluid samples available were used to compare the techniques. The detection rate of MV-RNA by RT-PCR was 98% (100/102) for oral fluid and 95% (97/102) for serum samples. The detection rate of MV-IgM was 85% (87/102). Negative MV-IgM results were observed mostly for serum samples collected early after the onset of the rash. A MV-RNA standard of known concentration obtained by in vitro transcription was used to quantify MV-RNA in samples. MV-RNA copy numbers were significantly higher in oral fluid than in serum samples, but did not correlate with time of sampling (within 1 week after the onset of the rash), patient age, or vaccination status. During the early stage of infection, the MV-RNA viral load in serum was lower in patients positive than in those negative for MV-IgG. In conclusion, the one-step real-time RT-PCR assay is a simple and sensitive tool suitable for MV diagnosis within hours. J. Med. Virol. © 2013 Wiley Periodicals, Inc.

Human cytomegalovirus (HCMV) is the most common cause of viral intrauterine infection and fetal damage largely due to maternal primary infection. Virological procedures which are able to detect HCMV fetal infection were evaluated. HCMV IgG antibodies were detected in 62.5% of the pregnant women and 1.47% had a primary infection. From March, 1992 to August, 1995, 29 seroconversions were observed, and in 64 other cases. HCMV IgM antibodies were detected in the first serological test. The mean IgG antibody avidity test (AI) was 31% for the 11 seroconversions tested and 74% in 32 cases where IgG and IgM HCMV antibodies were detected in the first serum. In the 29 HCMV seroconversions, 19 amniocentesis were carried out and 12 fetuses (41.4%) were infected in utero. In four amniotic fluids positive in culture and PCR, the fetus or newborns were infected and in one out of the two cordocentesis undertaken, hepatitis, anemia, and thrombocytopenia were noted. In four other cases, investigations seeking HCMV in amniotic fluid were negative whereas infants were infected at birth. Among the 64 cases with positive HCMV IgM and IgG antibodies detected in the first serological test, three fetuses were infected in utero, but no amniotic fluid was available in these cases. Amniotic fluids were studied in 39 cases, and HCMV detection by culture and PCR-hybridization was negative. HCMV DNA was detected in the maternal sera of five out of 21 pairs of seroconversions and in two cases on the first negative serum. The assay was also carried out on 50 of the 64 HCMV IgM positive sera. Two had detectable HCMV DNA. © 1996 Wiley-Liss, Inc.

Les bronchiolites a VRS sont frequentes, epidemiques et necessitent parfois l'hospitalisation des nourrissons, voire une reanimation.Le determinisme des lesions pulmonaires est complexe, lie au virus et a la reaction immunitaire. Le traitement est symptomatique, et l'utilisation de la ribavirine est limitee aux formes severes. Le diagnostic virologique doit etre rapide, et la recherche immunologique directe des proteines virales est la technique la plus utilisee.

We developed a polymerase chain reaction (PCR) to detect human metapneumovirus (hMPV) in French children hospitalized for acute respiratory tract disease during the winter of 2001 to 2002. This assay showed that 19 (6.6%) of 337 respiratory specimens that were negative for other respiratory viruses were positive for hMPV. Seven hMPV infections were also detected in the previous winter. The patients had the following clinical syndromes: bronchiolitis, 16; acute exacerbation of asthma, 4; bronchitis, 1; laryngitis, 1; high fever with diarrhea, 2; high fever with rhinopharyngitis, 1; rhinopharyngitis with conjunctivitis and otitis, 1. The sequences of the seven amplified products were similar for all cases and had 99% homology with the previously described N gene sequence.

Rapid and specific diagnosis of influenza A/B and respiratory syncytial virus (RSV) viruses is needed for optimal management of patients with acute respiratory infections. In this study, a one‐step triplex real‐time RT‐PCR assay was developed for rapid diagnosis of influenza A/B and RSV infections to optimize diagnosis efficiency of acute respiratory infections. Cell‐culture supernatants and clinical samples were used to evaluate specificity and sensitivity of the assay. The assay was used routinely during two winter epidemics for testing respiratory specimens from 2,417 patients. The limit of detection in cell‐culture supernatant was 1–10 plaque forming units/input (influenza A/B) and 2 × 10−2 50% tissue culture infectious dose/input (RSV). In clinical samples, the assay was as sensitive as commercial molecular assays for the detection of each influenza A/B and RSV (Flu‐A/B and RSV‐A/B r‐gene™) individually, and far more sensitive than antigen detection. During the winter 2008–2009, the assay identified 145 RSV, 42 influenza A, and one mixed RSV‐influenza A infections among 298 patients. The next winter, the assay was used in two independent hospital laboratory settings. 776 patients were tested in one hospital and 1,343 in the other, resulting in 184 and 501 RSV, 133 and 150 influenza A, and 1 and 11 mixed RSV‐influenza A infections, respectively, being detected. This new user‐friendly assay allows rapid (within hours), effective molecular diagnosis of single or mixed infections involving influenza A (including seasonal A H1N1 and H3N2, and A(H1N1) 2009), influenza B, and RSV(A/B). The assay is very valuable for managing patients during winter epidemics when influenza and respiratory syncytial viruses co‐circulate. J. Med. Virol. 83:695–701, 2011. © 2011 Wiley‐Liss, Inc.