Novel UL23 and UL30 substitutions in HSV1 and HSV2 viruses related to polymorphism or drug resistance.

Data on herpes simplex virus (HSV) polymorphism as well as acyclovir (ACV) and foscarnet (FOS) resistance mutations are not exhaustive and may hinder accurate diagnosis by next-generation sequencing (NGS). Here, we report novel UL23 and UL30 substitutions for HSV1 and HSV2 identified in immunocompromised patients treated for hematological malignancies during the last 6 years of HSV resistance surveillance at the University Hospital of Lyon. For HSV1, 35 novel UL23 substitutions and 52 novel UL30 substitutions were identified. For HSV2, 2 novel UL23 substitutions and 12 novel UL30 substitutions were identified. These results allow to complete the database of HSV1 and HSV2 substitutions, related either to polymorphism or to ACV and FOS resistance.

Unveiling the Ir single atoms as selective active species for the partial hydrogenation of butadiene by operando XAS.

Single-atom catalysts represent an intense topic of research due to their interesting catalytic properties for a wide range of reactions. Clarifying the nature of the active sites of single-atom catalysts under realistic working conditions is of paramount importance for the design of performant materials. We have prepared an Ir single-atom catalyst supported on a nitrogen-rich carbon substrate that has proven to exhibit substantial activity toward the hydrogenation of butadiene with nearly 100% selectivity to butenes even at full conversion. We evidence here, by quantitative operando X-ray absorption spectroscopy, that the initial Ir single atoms are coordinated with four light atoms i.e., Ir-X4 (X = C/N/O) with an oxidation state of +3.2. During pre-treatment under hydrogen flow at 250 °C, the Ir atom loses one neighbour (possibly oxygen) and partially reduces to an oxidation state of around +2.0. We clearly demonstrate that Ir-X3 (X = C/N/O) is an active species with very good stability under reactive conditions. Moreover, Ir single atoms remain isolated under a reducing atmosphere at a temperature as high as 400 °C.

Influenza-induced acute respiratory distress syndrome during the 2010-2016 seasons: bacterial co-infections and outcomes by virus type and subtype.

OBJECTIVES: We aimed to describe bacterial co-infections and acute respiratory distress (ARDS) outcomes according to influenza type and subtype. METHODS: A retrospective observational study was conducted from 2012 to 2016 in patients admitted to the respiratory intensive care unit (ICU) of Marseille university hospital for influenza-induced ARDS. Microbiological investigations, including multiplex molecular respiratory panel testing and conventional bacteriological cultures, were performed as part of the routine ICU care on the bronchoalveloar lavage collected at admission. Bacterial co-infections, ICU mortality and respiratory function were investigated according to virus type and subtype. RESULTS: Among the 45 ARDS patients included, A(H1N1)pdm09 was the most frequent influenza virus identified (28/45 A(H1N1)pdm09, eight out of 45 A(H3N2) and nine out of 45 influenza B). Bacterial co-infections involving a total of 23 bacteria were diagnosed in 16/45 patients (36%). A(H1N1)pdm09 patients presented fewer bacterial co-infections (17.9% vs. 50.0% for A(H3N2) patients and 77.8% for B patients; p < 0.01). Overall, mortality at 90 days post admission was 33.3% (15/45), and there was no significant difference between influenza type and subtype. The need for extracorporeal membrane oxygenation was more frequent for A(H1N1)pdm2009 (20/28, 71.4%) and B patients (7/9, 77.8%) than the A(H3N2) subtype (1/8, 12.5%; p < 0.01). A(H1N1)pdm09-ARDS patients were associated with fewer ventilation-free days at day 28 (median (IQR): 0 (0-8) days) compared with other influenza-ARDS patients (15 (0-25) days, p < 0.05). DISCUSSION: In a population of influenza-induced ARDS, A(H1N1)pdm09 was associated with fewer bacterial co-infections but poorer respiratory outcomes. These data underline the major role of A(H1N1)pdm09 subtype on influenza disease severity.

Microorganisms associated with respiratory syncytial virus pneumonia in the adult population.

Respiratory syncytial virus (RSV) has been recognized as responsible for severe respiratory illness in adults, especially in the elderly. While pneumonia is commonly observed during RSV infection, the burden and epidemiology of bacterial superinfection is poorly understood. The aim of this study was to identify microorganisms associated with RSV-positive pneumonia in adults. A retrospective study was conducted during three consecutive winters (October to April 2013-2016) in the University Hospital of Lyon, France. During RSV circulation periods, a systematic RSV screening was performed by reverse-transcription PCR on all respiratory samples collected from adults. Records of RSV-positive patients were subsequently analyzed to identify radiologically confirmed pneumonia cases. Bacteria were identified by standard bacteriology cultures or urinary antigen screening and classified as potentially causative of pneumonia if quantification was above the specific threshold as defined by the European Manual of Clinical Microbiology. Overall, 14,792 adult respiratory samples were screened for RSV detection by PCR. In total, 292 had a positive RSV detection (2.0%) among which 89 presented with pneumonia including 27 bacterial superinfections (9.3%) with Streptococcus pneumonia, Haemophilus influenza, Staphylococcus aureus, Pseudomonas aeruginosa, and Moraxella catarrhalis. Most patients were elderly (55.6%) and patients with comorbidities (77.8%). A more severe outcome was observed for RSV-bacteria-associated pneumonia compared with RSV pneumonia: length of stay was significantly longer (16 days vs 10 days) and ICU hospitalization more frequent (66.7% vs 21.0%) (p < 0.05). In conclusion, we did not observe major differences in the epidemiology of bacterial superinfections in RSV-positive pneumonia compared to reports on post-influenza pneumonia.

Quality control implementation for universal characterization of DNA and RNA viruses in clinical respiratory samples using single metagenomic next-generation sequencing workflow.

BACKGROUND: In recent years, metagenomic Next-Generation Sequencing (mNGS) has increasingly been used for an accurate assumption-free virological diagnosis. However, the systematic workflow evaluation on clinical respiratory samples and implementation of quality controls (QCs) is still lacking. METHODS: A total of 3 QCs were implemented and processed through the whole mNGS workflow: a no-template-control to evaluate contamination issues during the process; an internal and an external QC to check the integrity of the reagents, equipment, the presence of inhibitors, and to allow the validation of results for each sample. The workflow was then evaluated on 37 clinical respiratory samples from patients with acute respiratory infections previously tested for a broad panel of viruses using semi-quantitative real-time PCR assays (28 positive samples including 6 multiple viral infections; 9 negative samples). Selected specimens included nasopharyngeal swabs (n = 20), aspirates (n = 10), or sputums (n = 7). RESULTS: The optimal spiking level of the internal QC was first determined in order to be sufficiently detected without overconsumption of sequencing reads. According to QC validation criteria, mNGS results were validated for 34/37 selected samples. For valid samples, viral genotypes were accurately determined for 36/36 viruses detected with PCR (viral genome coverage ranged from 0.6 to 100%, median = 67.7%). This mNGS workflow allowed the detection of DNA and RNA viruses up to a semi-quantitative PCR Ct value of 36. The six multiple viral infections involving 2 to 4 viruses were also fully characterized. A strong correlation between results of mNGS and real-time PCR was obtained for each type of viral genome (R2 ranged from 0.72 for linear single-stranded (ss) RNA viruses to 0.98 for linear ssDNA viruses). CONCLUSIONS: Although the potential of mNGS technology is very promising, further evaluation studies are urgently needed for its routine clinical use within a reasonable timeframe. The approach described herein is crucial to bring standardization and to ensure the quality of the generated sequences in clinical setting. We provide an easy-to-use single protocol successfully evaluated for the characterization of a broad and representative panel of DNA and RNA respiratory viruses in various types of clinical samples.

Hospitalization in double-occupancy rooms and the risk of hospital-acquired influenza: a prospective cohort study.

Hospitalization in double-occupancy rooms and the risk of hospital-acquired influenza were assessed prospectively. The incidence was 2.0 for 100 patient-days in double- vs. 0.7 in single-occupancy rooms (p 0.028). The adjusted hazard ratio of hospital-acquired influenza was 2.67 (95% confidence interval 1.05-6.76) in patients hospitalized in double- compared to single-occupancy rooms.

Nanoalloying bulk-immiscible iridium and palladium inhibits hydride formation and promotes catalytic performances.

The hydrogen sorption properties of oxide-supported Ir-Pd nanoalloys have been determined for the first time, and correlated with their catalytic behavior. The addition of Ir to Pd suppresses hydride formation and leads to improved catalytic performances with respect to pure metals in the preferential oxidation of CO in H2 excess (PROX).

Increased detection of Mycoplasma pneumoniae infection in children, Lyon, France, 2010 to 2011.

Recent reports from several northern European countries indicate an increase in detection of Mycoplasma pneumoniae infection in the past two years, notably in children aged 5­15 years. Analysis of our laboratory database showed a similar pattern, with a higher proportion of respiratory samples positive for M. pneumonia by real-time PCR in paediatric patients aged 5­15 years. Our data indicate that in 2010 and 2011, France experienced the first epidemic peak of M. pneumonia infection since 2005.

Pediatric neurological complications associated with the A(H1N1)pdm09 influenza infection.

BACKGROUND: Influenza-related neurological complications (INC) have been reported during seasonal flu in children. OBJECTIVES: To investigate the types, outcomes and incidence of INC occurring during the 2009 A(H1N1) pandemic, a retrospective analyze was conducted in the single French pediatric hospital of Lyon from October 2009 to February 2010. STUDY DESIGN: All children presenting with fever, influenza-like illness, respiratory distress or neurological symptoms were tested for influenza A(H1N1)pdm09 infection from respiratory specimens using real time RT-PCR. RESULTS: INC occurred in 14 A(H1N1)pdm09 positive children (7.7% of A(H1N1)pdm09 positive children admitted to hospital) with a median age of 5.1 years. Admission to the intensive care unit (ICU) was required for nine children (64.3%). Half of the children with INC had comorbidity and three had coinfection, both characteristics mainly found in children requiring the ICU. All children received oral oseltamivir treatment. Febrile seizures were observed in eight children, half of them having a chronic comorbidity (2 epilepsy, 1 nonketotic hyperglycinemia, 1 anoxic encephalopathy). Other INC, less commonly reported, included 2 cases of encephalitis, 1 encephalopathy, 1 basilar artery thrombosis, 1 myasthenic crisis and 1 coma. Eleven of the 14 children (78.6%) recovered, one had a minor disability, one child developed a locked-in syndrome and one died from complications of an acute necrotizing encephalopathy. DISCUSSION: INC can be observed even in children with no underlying disorder. It may lead to dramatic issue in a significant number of cases.

Zidovudine (AZT) has a bactericidal effect on enterobacteria and induces genetic modifications in resistant strains.

The spread of multiresistant bacteria increases the need for new antibiotics. The observation that some nucleoside analogues have antibacterial activity led us to further investigate the antimicrobial activity and resistance of zidovudine (AZT). We determined the minimum inhibition concentration (MIC), studied time-kill curves, induced resistant bacteria and sequenced the gene for thymidine kinase. We demonstrate that AZT has a bactericidal effect on some enterobacteria. However, AZT could induce resistance in Escherichia coli. These resistances were associated with various modifications in the thymidine kinase gene. In particular, we observed the presence in this gene of an insertion sequence (IS) similar to IS911 of Shigella dysenteriae in two resistant clones. No cross-resistance with classical antibiotics in strains with modified thymidine kinase gene was observed. Finally, an additive or synergistic activity between AZT and the two aminoglycoside antibiotics amikacin and gentamicin was observed. We demonstrate the bactericidal activity of AZT and show synergy in association with gentamicin. Genetic modifications in resistant bacteria were identified. Our results indicate that AZT could potentially be added in the treatment of infections with enterobacteria or represent the basis for the development of derivatives with better activity and inducing less resistance.

Respiratory viruses in children admitted to hospital intensive care units: evaluating the CLART® Pneumovir DNA array.

Viruses play a significant part in children's respiratory infections, sometimes leading to hospitalization in cases of severe respiratory distress. The aim of this study was to investigate respiratory infections in children treated in a hospital intensive care unit (ICU). Assays were performed using the CLART® Pneumovir DNA array assay (Genomica, Coslada, Madrid, Spain), which makes it possible to detect 11 genus of respiratory viruses simultaneously. During the winter of 2008-2009, 73 respiratory specimens collected from 53 children under 2 years of age and admitted to an ICU were tested. At least one virus was detected in 78% (57/73) of the samples. The virological diagnosis was based on single infections in 65% (37/57) and on multiple infections in 35% (20/57) of cases. The array assay revealed respiratory syncytial virus (RSV) in 73.6% (42/57) of the samples and rhinovirus in 24.6% (14/57), either on their own or in co-infections. All viruses identified in single and multiple infections were tested, taking into account clinical features, risk factors, and severity criteria. Children with no risk factors presented more multiple infections, up to 42% of cases, than children with at least one risk factor. RSV seemed to induce severe symptoms by itself as no difference in intubation needs was observed when RSV was detected on its own or in co-infection. The CLART® Pneumovir DNA array was useful for examining severe viral respiratory infections, when other viruses than those detected by conventional methods could be involved, particularly in an ICU.

[Role of neuraminidase inhibitors for the treatment of influenza A virus infections]. / Intérêts des inhibiteurs de la neuraminidase dans la prise en charge des infections dues aux virus influenza.

Oseltamivir and zanamivir are two neuraminidase inhibitors (NAIs) active on A and B influenza viruses. These analogues have been developed from the structure of sialic acid, the neuraminidase (NA) substrate. Resistance to NAIs have been detected. They are mainly associated to mutations located on the NA gene. The use of these antiviral drugs remains low in the context of seasonal flu, even the duration of symptoms can be reduced of one day if an antiviral treatment is started within 48 hours after disease onset. NAIs also present a significant effect when used in postexposition prophylaxis. Resistance, mainly to oseltamivir, have been detected but remained rare until the spontaneous emergence in 2007-2008 winter of a seasonal A(H1N1) variant resistant to this drug. NAIs are also interesting for the treatment of severe flu infections, specially those associated to A(H5N1). Finally, because of the pandemic A(H1N1)2009 virus, NAIs use has largely increased for prophylactic and therapeutic treatment of severe and non severe infections. This large use may be associated to an increased risk of selection of resistant viruses. Up to now, this phenomenon remains fortunately limited but has to be closely monitored.

Pandemic A(H1N1)2009 influenza virus detection by real time RT-PCR: is viral quantification useful?

The emergence of the influenza A(H1N1) 2009 virus prompted the development of sensitive RT-PCR detection methods. Most are real time RT-PCRs which can provide viral quantification. In this manuscript, we describe a universal influenza A RT-PCR targeting the matrix (M) gene, combined with an RNaseP RT-PCR. These PCRs allow the detection of all influenza A virus subtypes, including A(H1N1)2009, together with a real-time assessment of the quality of the specimens tested. These PCR procedures were evaluated on 209 samples collected from paediatric patients. Viral loads determined through Ct values were corrected according to the RNaseP Ct value. The mean viral load in the collected samples was estimated to be 6.84 log RNA copies/mL. For poor quality samples (RNaseP Ct > 27), corrections resulted in +3 to +8 Ct values for the M gene RT-PCR. Corrected influenza Ct values were lower in late samples. No correlation was established between viral loads and clinical severity or duration of disease.This study shows that real time RT-PCR targeting the matrix gene is a reliable tool for quantification of type A influenza virus but emphasises the need for sample quality control assessment through cellular gene quantification for reliable estimation of the viral load. This method would be useful for disease management when repeated specimens are collected from an infected individual.

Rhinoviruses delayed the circulation of the pandemic influenza A (H1N1) 2009 virus in France.

In contrast to the experience in other European countries, the onset of the A(H1N1)2009 influenza virus epidemic was unexpectedly slow in France during the first part of autumn 2009. Our objective was to test the hypothesis that intense circulation of rhinoviruses might have reduced the probability of infection by A(H1N1)2009 virus at the beginning of autumn 2009. Systematic analysis for the detection of A(H1N1)2009 (H1N1) and human rhinovirus (HRV) was performed by RT-PCR from week 36 to week 48 on respiratory samples sent to the diagnostic laboratory by the paediatric hospital (n = 2121). Retrospective analysis of the obtained data, using 2 x 2 contingency tables with Fisher's exact test, revealed evidence of an inverse relationship between HRV and H1N1 detection. Between weeks 36 and 48 of 2009, both HRV and H1N1 were detected but in different time frames. HRV dispersed widely during early September, peaking at the end of the month, whereas the H1N1 epidemic began during mid-October and was still active at the end of this survey. During the co-circulation period of these two respiratory viruses (weeks 43-46), HRV detection appeared to reduce the likelihood of H1N1 detection in the same sample (OR = 0.08-0.24 p <0.0001). These results support the hypothesis that HRV infections can reduce the probability of A(H1N1) infection. This viral interference between respiratory viruses could have affected the spread of the H1N1 viruses and delayed the influenza pandemic at the beginning of autumn in France.

Impact of the 2009 influenza A(H1N1) pandemic wave on the pattern of hibernal respiratory virus epidemics, France, 2009.

This short report based on clinical surveillance and laboratory data describes the circulation of rhinoviruses, influenza viruses and respiratory syncytial viruses (RSV) in France during the 2009-10 season compared with the previous winter season. The delayed circulation of RSV observed in 2009-10 compared with 2008-09 suggests that the early circulation of the 2009 pandemic influenza A(H1N1) viruses had an impact on the RSV epidemic.

Anti N1 cross-protecting antibodies against H5N1 detected in H1N1 infected people.

The A(H5N1) influenza virus pandemic may be the result of avian H5N1 adapting to humans, leading to massive human to human transmission in a context of a lack of pre-existing immunity. As A(H1N1) and A(H5N1) share the same neuraminidase subtype, anti-N1 antibodies subsequent to H1N1 infections or vaccinations may confer some protection against A(H5N1). We analysed, by microneutralization assay, the A/Vietnam/1194/04 (H5N1) anti-N1 cross-protection acquired either during A/New-Caledonia/20/99 (H1N1) infection or vaccination. In cases with documented H1N1 infection, H5N1 cross-protection could be observed only in patients born between 1930 and 1950. No such protection was detected in the sera of vaccinated individuals.

[Avian influenza in children]. / Grippe aviaire chez l'enfant.

The emerging A (H5N1) influenza virus is a clear pandemic threat. This avian virus is responsible for the largest epizootic event described so far. To date, 423 humans have been infected. In humans, this virus is responsible for a rapidly developing pneumonia, with an acute respiratory failure leading to death in 60% of the infected cases. The multi-organ failure seems to result from the cytokine storm. A (H5N1) infections are mainly reported in children and young adults. Different hypotheses have been proposed to explain this specific feature, including the lack of control of cytokine release during infection, or the presence of alternative cellular receptors. The specific susceptibility of children is more likely to be related with exposure to infected birds than to specific immune or physiological factors. The proper and efficient management of A (H5N1) infection is possible nowadays. Today, the pandemic threat is real and may be imminent because of the circulation of new A (H1N1). However, an active surveillance of A (H5N1) virus remains very important to monitor the genetic evolution of this changing and virulent virus.