[Evolution of the number of rotavirus and respiratory syncytial virus infections in children hospitalised in a French university hospital between 1998 and 2005]. / Évolution du nombre d'infections àRotavirus et à virus respiratoire syncytial chez les enfants hospitalisés au CHU de Dijon entre 1998 et 2005.

AIM: Respiratory syncytial virus (RSV) and Rotavirus infections represent up to 30% of cross infections in pediatric units. As they are a major public health problem, we studied their evolution and distribution at the Dijon University Hospital. POPULATION AND METHODS: This exhaustive retrospective study included children under 15 with a new Rotavirus or RSV infection who were hospitalised at the Dijon University Hospital between 1998 and 2005. The general trend was determined by using moving averages, and the Spearman correlation coefficient r(s) was calculated. RESULTS: From 1998 to 2005, 1886 new RSV (n=981) or Rotavirus (n=905) infections were identified in hospitalised children. The number of the infections decreased significantly, both for RSV (r(s)=-0.71 ; p<0.0001) and for Rotavirus (r(s)=-0.77 ; p<0.0001). Almost half of Rotavirus infections were nosocomial (46.3%) vs 5.3% of RSV infections, p<0.0001. There was no significant difference in the proportion of RSV nosocomial infections between the epidemic and non-epidemic period (4.9% of nosocomial infections vs 7.1% respectively, p=0.25). Rotavirus nosocomial infections were less frequent in epidemic period (41.6%) than in non-epidemic period (54.6%); p=0.0002. CONCLUSION: RSV and Rotavirus infections significantly decreased between 1998 and 2005. Proportion of RSV or Rotavirus infections didn't increase in epidemic period, which could be explained both by an increased attention from healthcare professionals and by the effectiveness of hygiene measures taken.

Recurent and de novo membranous glomerulopathy after kidney transplantation.

The aim of this study was to compare the clinical characteristics of recurrent and de novo membranous glomerulopathy (MG) among a cohort of 614 recipients transplanted between 1989 and 2006. Lupus nephritides were excluded. The diagnosis was established on protocol biopsies performed 1, 2, 4, or 8 years after transplantation or because of proteinuria/nephrotic syndrome and/or an increased serum creatinine level. HCV infection, cryoglobulinemia, monoclonal gammopathy, skin cancers, Kaposi sarcoma, diabetes mellitus, anti-HLA antibodies, and graft survival were not significantly different between the groups. Seventeen MG were diagnosed in 15 patients (2.45% of the whole group), including 6 recurrent MG (35%) and 11 de novo MG (75%). Recurrent MG occurred earlier than de novo MG (15.58 +/- 19.13 vs 49.27 +/- 32.71 months). Recipients with de novo MG were more frequently infected with HCV, which seemed to be the main etiologic factor for de novo MG, and may be linked to a Th2 polarization of the immune response.

[Severe drug rash with eosinophilia and systemic symptoms after treatment with minocycline]. / Syndrome d'hypersensibilité médicamenteuse avec manifestations systémiques sévères après traitement par minocycline.

INTRODUCTION: Lung involvement is rarely observed in the DRESS syndrome (Drug rash with eosinophilia and systemic symptoms). We report here a severe minocycline induced hypersensitivity syndrome with initial respiratory distress. CASE REPORT: A 19 year old man was admitted to the intensive care unit for acute respiratory distress with fever (400C), lymph node enlargement, hepatomegaly, splenomegaly and eosinophilia (1640/mm3). Bilateral alveolar opacities were observed on the chest x-ray. Sedation and mechanical ventilation rapidly became necessary because of severe hypoxaemia (47 mm Hg) and the sudden onset of severe aggressive behaviour. The diagnosis of DRESS was immediately suspected as the patient had been treated for acne with minocycline for 28 days, and IV corticosteroids (2 mmg/kg/day) were initiated. Skin lesions were delayed and appeared 3 days later. The outcome was uncertain for the following 6 weeks with serious disturbance of hepatic and renal function. Serology for human herpes virus (HHV6) was initially negative but became positive. One year later, after progressive withdrawal of corticosteroid therapy, the patient had made a complete recovery with no sequelae. CONCLUSION: The DRESS syndrome can cause considerable morbidity with multiple, severe visceral functional disturbances. Respiratory physicians should be aware of this syndrome as lung involvement can be serious and may precede cutaneous symptoms.

Investigation of a Case of Genotype 5a Hepatitis C Virus Transmission in a French Hemodialysis Unit Using Epidemiologic Data and Deep Sequencing.

BACKGROUND Hepatitis C virus (HCV) is a major cause of chronic liver disease worldwide. A patient was recently found to be HCV seropositive during hemodialysis follow-up. OBJECTIVE To determine whether nosocomial transmission had occurred and which viral populations were transmitted. DESIGN HCV transmission case. SETTING A dialysis unit in a French hospital. METHODS Molecular and epidemiologic investigations were conducted to determine whether 2 cases were related. Risk analysis and auditing procedures were performed to determine the transmission pathway(s). RESULTS Sequence analyses of the NS5b region revealed a 5a genotype in the newly infected patient. Epidemiologic investigations suggested that a highly viremic genotype 5a HCV-infected patient who underwent dialysis in the same unit was the source of the infection. Phylogenetic analysis of NS5b and hypervariable region-1 sequences revealed a genetically related virus (>99.9% nucleotide identity). Deep sequencing of hypervariable region-1 indicated that HCV quasispecies were found in the source whereas a single hypervariable region-1 HCV variant was found in the newly infected patient, and that this was identical to the major variant identified in the source patient. Risk analysis and auditing procedures were performed to determine the transmission pathway(s). Nosocomial patient-to-patient transmission via healthcare workers' hands was the most likely explanation. In our dialysis unit, this unique incident led to the adjustment of infection control policy. CONCLUSIONS The data support transmission of a unique variant from a source with a high viral load and genetic diversity. This investigation also underlines the need to periodically evaluate prevention and control practices.

[Eye and cat scratch disease: A case series]. / Œil et maladie des griffes du chat : à propos de 7 cas.

INTRODUCTION: Cat scratch disease is a pleiomorphic condition, sometimes with isolated ophthalmic involvement. We report the clinical observations of seven cases with ophthalmologic manifestations of cat scratch disease. OBSERVATIONS: There were seven patients, with a median age of 52 years, of whom five were women and three had unilateral involvement. Six exhibited Leber's stellate neuroretinitis, an incomplete syndrome in two cases, and one associated with chorioretinal foci. One patient had isolated retinal infiltrates. The diagnosis of cat scratch disease was confirmed by Bartonella henselae serology, positive in all cases. All patients received treatment with doxycycline. Ocular complications (with optic atrophy and macular retinal pigment epithelial changes) were noted in five cases. DISCUSSION: Ocular bartonellosis is an atypical clinical form. It requires a directed ancillary work-up with serology or PCR, which has the peculiarity of being highly specific if not very sensitive. Treatment is above all preventive. Antibiotics may be initiated. CONCLUSION: Cat scratch disease must be excluded in the work-up of posterior uveitis.

Identification of a duplicated V3 domain in NS5A associated with cirrhosis and hepatocellular carcinoma in HCV-1b patients.

BACKGROUND: The NS5A protein of the hepatitis C virus has been shown to be involved in the development of hepatocellular carcinoma. OBJECTIVES: In a French multicenter study, we investigated the clinical and epidemiological features of a new HCV genotype 1b strain bearing a wide insertion into the V3 domain. STUDY DESIGN: We studied NS5A gene sequences in 821 French patients infected with genotype 1b HCV. RESULTS: We identified an uncharacterized V3 insertion without ORF disruption in 3.05% of the HCV sequences. The insertion comprised 31 amino-acids for the majority of patients; 3 patients had 27 amino-acids insertions and 1 had a 12 amino-acids insertion. Sequence identity between the 31 amino-acids insertions and the V3 domain ranged from 48 to 96% with E-values above 4e(-5), thus illustrating sequence homology and a partial gene duplication event that to our knowledge has never been reported in HCV. Moreover we showed the presence of the duplication at the time of infection and its persistence at least during 12 years in the entire quasispecies. No association was found with extrahepatic diseases. Conversely, patients with cirrhosis were two times more likely to have HCV with this genetic characteristic (p=0.04). Moreover, its prevalence increased with liver disease severity (from 3.0% in patients without cirrhosis to 9.4% in patients with both cirrhosis and HCC, p for trend=0.045). CONCLUSIONS: We identified a duplicated V3 domain in the HCV-1b NS5A protein for the first time. The duplication may be associated with unfavorable evolution of liver disease including a possible involvement in liver carcinogenesis.

Value of a ligase chain reaction assay for detection of ganciclovir resistance-related mutation 594 in UL97 gene of human cytomegalovirus.

Human cytomegalovirus (HCMV) isolates resistant to ganciclovir were found in patients undergoing therapy. Therefore, we have developed a new specific and sensitive method--a ligase chain reaction (LCR) assay--for detection of frequently encountered 594 mutated codon in ganciclovir (GCV) resistant virus. Previous studies characterized an alanine to valine change on codon 594 in resistant strains. A novel substitution in 594, alanine to glycine, is described which is also capable of conferring ganciclovir resistance. LCR products were analyzed on polyacrylamide gel- and the mutant was detected using a non radioactive method. The LCR product detection was then adapted to a microtitre plate format with a colorimetric detection. This method allowed the distinction of mutated GCV-resistant strains from sensitive strains with a high sensitivity, and the detection of a low percentage of mutated DNA in virus load. This assay could be useful in following the evolution of mutated DNA compared to viral infection.

HCV infection and diabetes mellitus: influence of the use of finger stick devices on nosocomial transmission.

An increased prevalence of hepatitis C virus (HCV) infection in patients with diabetes mellitus has suggested a link between these two conditions and the possibility of patient-to-patient HCV transmission during hospital admissions in diabetes units. We investigated the prevalence of HCV antibodies in 259 patients with diabetes mellitus consecutively admitted to our diabetic unit in 1998. The control group was composed of 14,100 volunteer blood donors. We divided the diabetic patients into two groups according to their HCV antibody status and also analysed patients for the following variables: age, disease duration, diabetes treatment, previous hospital admissions in a diabetes unit and use of finger stick devices. Anti-HCV antibodies were detected in 8 diabetic patients and 6 blood donors (3.09% vs 0.04%, p < 0.001). No differences were observed between anti-HCV-positive and anti-HCV-negative diabetic patients in terms of mode of treatment, previous hospital admissions in a diabetic unit and use of finger stick devices for capillary blood sampling. Our findings indicate that these medical practices play no role in nosocomial transmission of HCV in diabetic patients.

Rapid and sensitive detection of metapneumovirus in clinical specimens by indirect fluorescence assay using a monoclonal antibody.

Human metapneumovirus, with two known genotypes named A and B, is associated with mild respiratory symptoms to severe LRTI in children, high-risk adults and the elderly. Rapid and reliable methods of hMPV detection in clinical samples are essential to implement appropriate care, to better understand the pathology of hMPV and to determine its epidemiology. Respiratory samples from 1,386 patients collected during 2 consecutive years were screened for hMPV using indirect immunofluorescence (IFA) assay with a monoclonal antibody. Forty-three patients tested positive for hMPV by the IFA method. In parallel, the samples were examined with RT-PCR on the F gene. Of these, 41 specimens were RT-PCR positive. The remaining two IF positives were cultured and the cultures were subsequently RT-PCR positive. IFA showed therefore a sensitivity of 100%. No false positive signals were obtained with the influenza virus, respiratory syncytial virus or parainfluenza. When tested by RT-PCR, all IFA-negative samples (n = 204)were found negative. Therefore the specificity of IFA was 100%, IC95 [98-100%], with a negative predictive value of 100%. Based upon phylogenetic analysis of the fusion gene, both subgroups of hMPV were efficiently detected by IFA, and the viral aetiology could be given in 2 hr. These results demonstrate the potential usefulness of immunofluorescence with our monoclonal antibody for the rapid detection of hMPV in clinical specimens in the management of therapy and the control of nosocomial diffusion.

Adolescent varicocele: objective indications for treatment.

We measured the gonadotropin response pattern to the intravenous injection of luteinizing hormone-releasing hormone in 53 male adolescents 11 to 17 years old (average age 14.2 years) with a grade II to III varicocele. Volume loss of the testis ipsilateral to the varicocele of 2 cc or more was noted in 32 patients. Baseline serum levels of follicle-stimulating hormone, luteinizing hormone and testosterone were determined, and measured again at 15, 30 and 60 minutes following intravenous injection of luteinizing hormone-releasing hormone. Of the patients 23 had an exaggerated response pattern following luteinizing hormone-releasing hormone stimulation, including abnormally high post-stimulation levels of luteinizing hormone only in 6, follicle-stimulating hormone only in 4, and follicle-stimulating and luteinizing hormones in 13. No significant difference in mean age, testosterone levels or loss of testicular volume was noted between the abnormal and normal response groups. This study indicates that approximately 45 per cent of adolescents with a varicocele have evidence of testicular injury. The physical examination was of no value in predicting which patient with a varicocele would display evidence of testicular dysfunction. We suggest that the luteinizing hormone-releasing hormone stimulation test become a routine part of the evaluation of an adolescent with a varicocele.

Use of synthetic peptides to locate neutralizing antigenic domains on the fusion protein of respiratory syncytial virus.

Chemical and enzymic cleavages of the F1 subunit of the fusion (F) protein of respiratory syncytial (RS) virus showed that the sequence 184-Gly to 314-Trp reacted with neutralizing monoclonal antibodies (MAbs). Twelve synthetic peptides covering a part of this sequence were analysed for their immunoreactivity with neutralizing MAbs and anti-RS virus rabbit serum. Two sequential antigenic domains corresponding to amino acids 200 to 225 and 255 to 278 were defined with anti-RS virus rabbit serum. The peptides 205-225 and 259-278, belonging to these antigenic domains, inhibited binding to the F protein and the neutralizing activity of the anti-RS virus rabbit serum. One MAb (RS-348) reacted with peptides containing amino acids 200 to 225. Moreover, the peptide 205-225 induced an anti-peptide rabbit serum neutralizing RS virus in vitro. These results indicate that the sequence from residues 200 to 225 was present in one of the immunodominant sites of the F protein.

Identification of T-cell epitopes adjacent to neutralizing antigenic domains on the fusion protein of respiratory syncytial virus.

F glycoprotein has been identified as an important target structure in the immunological response following respiratory syncytial virus (RSV) infection. Two sequential B epitopes corresponding to amino acids 200 to 225 and 255 to 278 have already been defined with anti-RSV rabbit serum. The T helper response to peptides which belong to these sequences was investigated in this study. Proliferative T-cell responses to these peptides were analysed in BALB/c mice (H-2d) and others strains: SJL (H-2s), C3H/He (H-2k), B10.BR (H-2k) and C57BL/6 (H-2b). By using various strategies, two T-cell epitopes were identified in the amino acid 200 to 225 and 255 to 278 regions, close to a neutralizing epitope. The T-cell responses to these peptides were H-2- restricted. In addition, the peptide 255-278 was able to stimulate T cells that responded to a subsequent immunization with F glycoprotein in vitro.

Epitope mapping of the major inner capsid protein of group A rotavirus using peptide synthesis.

Three hundred and ninety-one consecutive heptapeptides derived from the VP6 protein of bovine rotavirus (397 AA) were synthesized using the "pepscan" method and were assayed on the synthesis pins with monoclonal antibodies to VP6. Heptapeptides reactive with MAbs were located in four main regions: regions AA 32-64, AA 155-167, AA 208-274, and a fourth region at the C-terminal, from AA 380 to AA 397. Among these regions, two sequences were also reactive with the MAbs when longer peptides were assayed. The sequence located between AA 58 and AA 62 (NWNFD), recognized by MAbs RV-1026, RV-50, and RV-443, was previously reported. A new site was defined in the region essential for trimerization, between AA 159 and AA 165 (PYSASFT), which was recognized by MAbs RV-133 and RV-138.

Prophylactic administration of a complementarity-determining region derived from a neutralizing monoclonal antibody is effective against respiratory syncytial virus infection in BALB/c mice.

Immunotherapy with antibodies against respiratory syncytial virus (RSV) is a treatment option given the absence of any vaccine or other available satisfactory treatment. We selected one of our monoclonal antibodies, RS-348, that is highly neutralizing. We showed that a single peptide (PEP3H) derived from complementarity-determining region 3 (CDR3) of its heavy chain was capable of neutralizing the virus in vitro. When intranasally administered 24 h before challenge, this peptide protected BALB/c mice against RSV lung infection. These results indicate that a single CDR can be effective against RSV infection.

Heparin-like structures on respiratory syncytial virus are involved in its infectivity in vitro.

Addition of heparin to the virus culture inhibited syncytial plaque formation due to respiratory syncytial virus (RSV). Moreover, pretreatment of the virus with heparinase or an inhibitor of heparin, protamine, greatly reduced virus infectivity. Two anti-heparan sulfate antibodies stained RSV-infected cells, but not noninfected cells, by immunofluorescence. One of the antibodies was capable of neutralizing RSV infection in vitro. These results prove that heparin-like structures identified on RSV play a major role in early stages of infection. The RSV G protein is the attachment protein. Both anti-heparan sulfate antibodies specifically bound to this protein. Enzymatic digestion of polysaccharides in the G protein reduced the binding, which indicates that heparin-like structures are on the G protein. Such oligosaccharides may therefore participate in the attachment of the virus.

[Epidemiology of viral nosocomial infections in pediatrics]. / Epidémiologie des infections nosocomiales virales en pédiatrie.

Nosocomial viral infections account for at least 5% of the total of NI and reach 23% in pediatric wards. The nosocomial infection (NI) incidence rate varies from 0.59 to 0.72 per 100 patients in pediatric wards. Many viruses have been associated with NI in pediatric wards. Rotavirus and respiratory syncytial virus (RSV) are the most frequent. Other viruses frequently identified are: calicivirus, adenovirus, astrovirus, influenza et para-influenza, rhinovirus and coronavirus. Asymptomatic infections occur frequently. The period of communicability varies and depends on the virus. It often begins before the clinical signs appear and ends after the healing. Viral shedding may be intermittent. Children and hospital environment and less frequently hospital staff are the main source for the virus. Poor handwashing results in direct spread to patient or self-inoculation even for respiratory viruses like RSV and rhinovirus. The main risk factors for NI are prolonged hospital stay, past history of prematurity and low age. Immunocompromised patients constitute a special high-risk group. Understaffing is also a risk factor. Minimal infective doses depend on the route of inoculation and the kind of virus. Low doses are for example sufficient for rotavirus, adenovirus and calicivirus. Viral inactivation is all the more easy when there is an envelope. Handwashing and appropriate isolation (technical and geographical) are the mainstay of prevention of viral NI. Vaccines are promising, especially for rotavirus.

Localization of group-specific epitopes on the major capsid protein of group A rotavirus.

Chemical cleavage of the VP6 protein of bovine rotavirus showed that VP6-specific monoclonal antibodies (MAbs) reacted with the amino acid sequence between glycine 48 and asparagine 107. Furthermore, three synthetic peptides (amino acids 48 to 64, 60 to 75 and 91 to 108) containing part of this sequence and 22 consecutive overlapping heptapeptides corresponding to the region between amino acids 48 and 75 were analysed for their immunoreactivity using group-specific MAbs. The MAbs recognized peptides 48-64 and/or 60-75, and a set of overlapping heptapeptides located between residues 53 (asparagine) and 67 (glycine), which have two short sequences in common: IRNW (residues 56 to 59), recognized by MAb RV-133, and (NW)NFD (residues 58/60 to 62), recognized by MAbs RV-50, -1026 and -443. These results indicate that the sequence between amino acid residues 48 and 75 is present in one of the immunodominant sites of VP6.

Severe fetal cytomegalic inclusion disease after documented maternal reactivation of cytomegalovirus infection during pregnancy.

Recurrent cytomegalovirus infection during pregnancy is considered less dangerous for the fetus than primary infection. We present a case of severe fetal cytomegalic inclusion disease after maternal reactivation of cytomegalovirus during the first trimester of pregnancy. The possibility of such fetal injury is an argument for prenatal diagnosis in seropositive pregnant women when ultrasonographic findings suggest cytomegalovirus infection.

Prevalence of hepatitis C virus infection in patients with rheumatoid arthritis.

BACKGROUND: Various viruses have been implicated in the cause and pathogenesis of rheumatoid arthritis (RA). Hepatitis C virus (HCV) infection, which has been recognised as a cause of some autoimmune diseases, and which has been described as sometimes presenting with rheumatic manifestations indistinguishable from RA, might be a candidate. OBJECTIVE: To evaluate the prevalence of HCV infection in patients with RA. METHODS: Consecutive patients with RA admitted to hospital in two departments of rheumatology were prospectively studied. Patients' serum samples were screened for the presence of anti-HCV antibodies. Patients with positive serology were further evaluated for the presence of HCV ribonucleic acid by reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: 309 patients (232 women, 77 men, mean age (SD) 54.1 (14.8) years) were studied. Their mean (SD) disease duration was 74.1 (91) months. Tests for rheumatoid factors and antinuclear antibodies were positive in 213 (69%) and 114 (37%) of the patients respectively. Systemic vasculitis was found in 12 (4%) of the patients. Mean erythrocyte sedimentation rate was 36.4 (SD 30.5) mm at the first hour (normal <10 mm) and C reactive protein was 36.8 (SD 45.8) mg/l (normal range <5 mg/l), respectively, with 181(58.6%) of patients considered as having active disease. Aspartate transaminases were increased in 14 (4%) patients, and alkaline phosphatase in 14 (4%). A positive anti-HCV serology was found in two (0.65%) patients, including one with a previously diagnosed HCV infection. HCV RNA was positive by RT-PCR in one of those two patients. CONCLUSION: A 0.65% prevalence of past or active HCV infection was found in patients with RA, which did not differ from the prevalence of HCV in the general French population. This result does not support the participation of HCV infection in the pathogenesis of RA.