Fanny Renois

Prevalence of rotavirus, adenovirus, norovirus, and astrovirus infections and coinfections among hospitalized children in northern France.

ABSTRACT From January to December 2007, 973 stool specimens were prospectively collected from children hospitalized for gastroenteritis signs or from neonates and premature cases who were born in two French hospital settings in the north of France. They were tested by rapid enzyme immunoassay (EIA) analyses for rotavirus and adenovirus and by two commercially available ELISA tests for the detection of norovirus and astrovirus. The overall rates of prevalence for rotavirus, norovirus, adenovirus, and astrovirus were 21, 13, 5, and 1.8%, respectively, and they did not significantly differ between the two hospital settings (P = 0.12). Mixed virus infections were detected in 32 (3.3%) of the 973 study children and were associated with norovirus in 21 (66%) infants, including 5 premature cases. From fall to spring, norovirus infections accounted for 52% of documented gastroenteritidis viral infections at a time when rotavirus was epidemic, resulting in mixed norovirus and rotavirus gastrointestinal tract infections. Of the 367 documented viral gastroenteritis cases, 15 (4.1%) were identified as nosocomial infections, 5 of which occurred in premature cases. These findings highlight the need to implement norovirus and astrovirus ELISA detection assays in association with rapid EIA rotavirus and adenovirus detection assays for the clinical diagnosis and the nosocomial prevention of gastroenteritis viral infections in pediatric departments.

Rapid Detection of Respiratory Tract Viral Infections and Coinfections in Patients with Influenza-Like Illnesses by Use of Reverse Transcription-PCR DNA Microarray Systems

ABSTRACT We prospectively tested 95 nasal swabs or nasopharyngeal aspirates taken from 56 adults and 39 children visiting the Reims University Medical Centre (northern France) for influenza-like illnesses (ILI) during the early stage of the French influenza A/H1N1v pandemic (October 2009). Respiratory samples were tested using a combination of two commercially available reverse transcription-PCR (RT-PCR) DNA microarray systems allowing rapid detection of influenza A virus strains, including the new A/H1N1v strain as well as 20 other common or newly discovered respiratory viruses. Concomitantly, a generic and classical real-time RT-PCR assay was performed to detect all circulating influenza A virus strains in the same samples. Of the 95 respiratory samples tested, 30 (31%) were positive for the detection of influenza A/H1N1v virus infection by both RT-PCR DNA microarray and classical real-time RT-PCR detection assays. Among the infections, 25 (83%) were monoinfections, whereas 5 (17%) were multiple infections associating influenza A/H1N1v virus with coronavirus (CoV), human bocavirus (HBoV), respiratory syncytial virus (RSV), or human rhinoviruses (HRVs). Of the 95 respiratory samples tested, 35 (37%) were positive for respiratory viruses other than influenza A/H1N1v virus. Among these infections, we observed 30 monoinfections (HRVs [63%], parainfluenza viruses [PIVs] [20%]), influenza A/H3N2 virus [6%], coronavirus [4%], and HBoV [4%]) and 5 multiple infections, in which HRVs and PIVs were the most frequently detected viruses. No specific single or mixed viral infections appeared to be associated significantly with secondary hospitalization in infectious disease or intensive care departments during the study period (P > 0.5). The use of RT-PCR DNA microarray systems in clinical virology practice allows the rapid and accurate detection of conventional and newly discovered viral respiratory pathogens in patients suffering from ILI and therefore could be of major interest for development of new epidemiological survey systems for respiratory viral infections.

Human Bocavirus quantitative DNA detection in French children hospitalized for acute bronchiolitis.

Abstract Background Human Bocavirus (HBoV) is a newly discovered parvovirus whose role as a causative agent of respiratory disease remains unclear. Study design We investigated the presence of HBoV by quantitative PCR in the nasopharyngeal samples of 192 French children consecutively hospitalized for acute bronchiolitis. Other common respiratory viruses were detected using immunofluorescence assays, cell culture detection, or RT-PCR assays. Results HBoV was detected in 24 (12.5%) of 192 study children. In 14/192 cases (7%) HBoV was the sole isolate and in 10/192 (5%) it was part of a mixed viral infection. HBoV was the third most common pathogen detected after respiratory syncytial virus (45/192; 23%) and rhinovirus (24/192; 12%). It occurred more often in infants aged 1–12 months (P =0.002). Median levels of HBoV DNA genome in respiratory samples were significantly higher in patients with single HBoV infection than in patients with mixed respiratory viral infection with HBoV (4×108 copies/ml vs. 2×103 copies/ml, P <0.001). Conclusions Our data suggest that HBoV at a high viral load could be an etiologic agent of respiratory tract disease, whereas the exact role of HBoV at a low viral load, as etiological cause or as pathophysiological co-factor of respiratory diseases, remains to be determined.

Low frequency of cytomegalovirus infection during exacerbations of inflammatory bowel diseases

Although numerous reports have described inflammatory bowel diseases (IBDs) complicated with cytomegalovirus (CMV) infection, the virus participation as an exacerbating factor remains unclear. The aim of this study was thus to clarify the clinical significance of CMV infection complicating exacerbation and to correlate CMV detection with various characteristics in IBD patients. Sixty‐seven colonic biopsies obtained from 53 patients admitted for IBD exacerbation were retrospectively analyzed by real‐time PCR assay. The CMV genome was detected in seven (10.4%) colonic biopsies related to seven patients (three ulcerative colitis and four Crohns diseases). Among the patients with IBD studied, patients with evidence of CMV infection were older (P = &!thinsp;0.047), were more likely male gender (relative risk [RR] 4.48; 95% confidence interval [CI] 0.94–21.36), received corticosteroids (RR 3.2; CI 0.79–13.02) or azathioprine (RR 3.17; CI 0.80–12.57) treatments, presented more extended lesions (RR for rectum‐sigmoid‐left colon 3.75 (0.0–69.37) and for pancolitis 2.45 (0.36–16.23)), and had a more severe disease (RR 3.3; CI 0.87–12.48) than those without CMV infection. Viral loads measured in the colonic mucosa of infected patient ranged from 5 to 236961 genome copies by microgram of total extracted DNA. No relationship was observed between the severity of the disease and the viral load level. Furthermore, CMV disappeared in five infected IBD patients in remission without antiviral agents. In conclusion, these results showed infrequent CMV detection in colonic biopsies of IBD patients during exacerbation leaving open the question of the relationship between CMV reactivation and the onset or the severity of IBD exacerbation. J. Med. Virol. 82:1694–1700, 2010.

Broad respiratory virus detection in infants hospitalized for bronchiolitis by use of a multiplex RT-PCR DNA microarray system†

Newly available molecular tools allow a sensitive detection of a broad panel of viruses in respiratory tract specimens. In the present study, the application of a multiplex RT‐PCR DNA microarray in diagnosis and epidemiological survey of viral infections in infants hospitalized for bronchiolitis was assessed. One hundred and thirty‐eight nasopharyngeal aspirates collected from October 2007 to September 2008 were tested by direct immunofluorescence and viral culture, a combination of referenced RT‐PCRs and the DNA microarray. One or more viruses were detected in 96, 126 and 126 of the specimens by direct immunofluorescence and viral culture, RT‐PCRs and DNA microarray, respectively (70 vs. 91 vs. 91%, P < 10−3). The RT‐PCRs and the DNA microarray yielded concordant results for 99% of specimens and identified mixed viral infections in 85 (62%). The most common associations were: human bocavirus and respiratory syncytial virus (32%), adenovirus and respiratory syncytial virus (30%), and parainfluenza virus type 3 and respiratory syncytial virus (23%). None of the bronchiolitis severity parameters including intensive care unit admission, O2 supply, O2 saturation percentage, O2 length and length of stay at the hospital appeared to be significantly increased in multiple viral infections compared to single viral infections (P > 0.1). In conclusion, the use of this DNA microarray in clinical virology practice allows rapid and accurate identification of common and uncommon viral respiratory pathogens in infants hospitalized for bronchiolitis. It should improve the clinical management, the epidemiological survey, and the prevention of the nosocomial transmission of respiratory viruses in pediatric wards. J. Med. Virol. 84:979–985, 2012.

Detection of multiple viral and bacterial infections in acute exacerbation of chronic obstructive pulmonary disease: A pilot prospective study

Few studies have evaluated the contribution of multiple virus and bacterial infections in acute exacerbation of chronic obstructive pulmonary disease. This study estimated the burden of multiple viral and bacterial respiratory infections in moderate to very severe chronic obstructive pulmonary disease patients that were prospectively followed‐up during a 12‐month pilot study. Clinical data were collected monthly and sputum was collected at the time of each acute exacerbation event. Classical culture techniques for bacteria and multiplex polymerase chain reaction (PCR) and microarray detection assays were performed to identify viral and atypical bacterial pathogens in the sputum. Overall, 51 patients were included and 45 acute exacerbation events were investigated clinically and microbiologically. Among the 45 acute exacerbation events, 44% had evidence of viral infection involving human rhinovirus (HRV) and metapneumovirus (hMPV) in 20% and 18%, respectively. Intracellular bacteria were not found in sputum by PCR. Common bacterial pathogens were identified in 42% of acute exacerbation patients, most frequently Branhamella catarrhalis, Streptococcus pneumoniae and Haemophilus influenzae. Viral or virus and bacteria co‐infections were detected in 27% of acute exacerbation events (n = 12) with HRV and hMPV involved in 92% of cases. Patients with co‐infections did not present greater clinical severity scores at exacerbation and more recurrence of acute exacerbation events at 3 and 6 months than those with single infections (P > 0.4). These results suggest that HRV and hMPV may be contributors or cofactors of AECOPD. These findings indicate that viral or virus and bacterial co‐infections do not impact significantly on the clinical severity of acute exacerbation of chronic obstructive pulmonary disease and recurrence at 3 and 6 months. J. Med. Virol. 85:866–873, 2013.

Enterovirus 68 in Pediatric Patients Hospitalized for Acute Airway Diseases

ABSTRACT Enterovirus 68 was detected in 10 respiratory specimens from pediatric patients hospitalized for acute wheezing or bronchitis during 2009 in the northeast of France. Viral loads ranged from 2 × 105 to 7.2 × 107 copies/ml. Alignment of 5′ nontranslated regions and phylogenetic analysis of partial VP1 gene sequences show that these viruses clustered and belonged to clade C.

Rapid Virological Diagnosis of Central Nervous System Infections by Use of a Multiplex Reverse Transcription-PCR DNA Microarray

ABSTRACT Viruses are the main etiological cause of central nervous system (CNS) infections. A rapid molecular diagnosis is recommended to improve the therapeutic management of patients. The aim of this study was to evaluate the performances of a DNA microarray, the Clart Entherpex kit (Genomica, Coslada, Spain), allowing the rapid and simultaneous detection of 9 DNA and RNA neurotropic viruses: herpes simplex virus 1 (HSV-1), HSV-2, varicella-zoster virus (VZV), cytomegalovirus (CMV), Epstein-Barr virus (EBV), human herpesvirus 6 (HHV-6), HHV-7, HHV-8, and the human enteroviruses (HEVs). This evaluation was performed with 28 samples from the European proficiency panels (Quality Control for Molecular Diagnostics [QCMD]; Glasgow, Scotland) and then with 78 cerebrospinal fluid (CSF) specimens. The majority of the QCMD results obtained by the DNA microarray were similar to those recorded by the overall QCMD participants. The main discrepant results were observed for low concentrations of HSV-2 and HEVs. From the clinical samples, the kit detected 27 of the 28 herpesvirus CNS infections and all of the 30 HEV-positive CSF samples. No false-positive result was observed among the 20 virus-negative CSF samples. The clinical sensitivity, specificity, and negative and positive predictive values of the assay were 98.3, 100, 95.2, and 100%, respectively, when the results were compared to those of commercially available PCR assays. Interestingly, HHV-7 was detected in 11 (37%) of the 30 HEV-positive CSF samples from children suffering from aseptic meningitis causing significantly longer lengths of stay at the hospital than infection with HEVs alone (2.4 versus 1.4 days; P = 0.038). In conclusion, this preliminary study showed that this DNA microarray could be a valuable molecular diagnostic tool for single and mixed DNA and RNA virus infections of the CNS.

Clinical and virological features of an aseptic meningitis outbreak in North-Eastern France, 2005

BACKGROUND Enteroviruses (EVs) are considered as a major viral etiological cause of aseptic meningitis in children. OBJECTIVES We assessed the clinical and virological features of an aseptic meningitis outbreak in North-East of France, 2005. STUDY DESIGN Classical bacteriological analysis, Herpesviridae and EV PCR assays had been prospectively performed on cerebrospinal fluid (CSF) samples taken from 80 children hospitalized for aseptic meningitis. For each EV strain identified as etiological agent, a phylogenetic comparison of partial EV VP1 capsid protein coding gene was performed. RESULTS The children older than 12 months (n=75) presented a typical aseptic meningitis syndrome, whereas the children aged less than 1 year (n=5) demonstrated only fever and hypotonia. Among the 80 studied children, EV was identified as the etiological cause of aseptic meningitis in 73 (91%) cases. Echovirus 30 (E30) was the most common isolated serotype (84% of 51 EV strains). VP1 phylogenetic analysis revealed that E30 strains were genetically closer to those isolated during 2000 aseptic meningitis outbreak comparatively to those identified during 2003 and 2006 non-epidemic years. Moreover, the genetic study demonstrated the co-circulation of four distinct lineages without any difference in temporal distribution or clinical features during the 2005 outbreak. CONCLUSIONS The present report demonstrates the co-circulation of distinct E30 lineages during the same aseptic meningitis outbreak season. This E30 genetic diversity may be a prerequisite for the emergence of new strains potentially responsible for further aseptic meningitis outbreaks.

Enterovirus-related activation of the cardiomyocyte mitochondrial apoptotic pathway in patients with acute myocarditis

AIMS We examined the impact of enterovirus (EV) cardiac replication activity on the endomyocardial mitochondrial pathway in patients with acute myocarditis. METHODS AND RESULTS Levels of apoptotic cardiomyocytes were determined by TUNEL and ligation-mediated polymerase chain reaction (PCR) assays and EV replication activity was assessed by immunostaining of EV VP1 capsid protein in ventricular myocytes of patients with acute myocarditis (n = 25), and healthy heart controls (n = 15). Ratio of cytosolic/mitochondrial cytochrome c concentrations was determined by ELISA assay, levels of active caspase-9 were determined by western blot analysis and Bax/Bcl2 mRNA ratio was assessed by real-time reverse transcription-polymerase chain reaction (RT-PCR) in the same cardiac tissues. Patients with EV-associated acute myocarditis (n = 15) exhibited a significantly higher number of apoptotic cardiomyocytes than those with non-EV-associated acute myocarditis (n = 10) and controls (n = 15) (P < 0.001). Endomyocardial ratio of cytosolic/mitochondrial cytochrome c concentrations and levels of active caspase-9 protein were significantly increased in EV than in non-EV-related myocarditis patients (P < 0.001). Moreover, Bax/Bcl2 mRNA ratio was significantly increased in EV than in non-EV-related myocarditis patients (P < 0.001). CONCLUSION Our findings evidence an EV-related activation of the cardiomyocyte mitochondrial apoptotic pathway in patients with acute myocarditis. Moreover, our results indicate that this EV-induced pro-apoptotic mechanism could be partly related to an up-regulation of Bax expression, and suggest that inhibition of this cell death process may constitute the basis for novel therapies.