Fatal outcome after reactivation of inherited chromosomally integrated HHV-6A (iciHHV-6A) transmitted through liver transplantation.

HHV-6A and HHV-6B are found as inherited and chromosomally integrated forms (iciHHV-6A and -6B) into all germinal and somatic cells and vertically transmitted in a Mendelian manner in about 1% of the population. They were occasionally shown to be horizontally transmitted through hematopoietic stem cell transplantation. Here, we present a clinical case of horizontal transmission of iciHHV-6A from donor to recipient through liver transplantation. Molecular analysis performed on three viral genes (7.2 kb) in the recipient and donor samples supports transmission of iciHHV-6A from the graft. Transmission was followed by reactivation, with high viral loads in several compartments. The infection was uncontrollable, leading to severe disease and death, despite antiviral treatments and the absence of resistance mutations. This case highlights the fact that physicians should be aware of the possible horizontal transmission of iciHHV-6 and its consequences in case of reactivation in immunocompromised patients.

Detection of Bartonella in cat scratch disease using a single-step PCR assay kit.

PURPOSE: Bartonella is an increasingly isolated emerging pathogen that can cause severe illness in humans, including cat scratch disease (CSD). The bacteria are difficult to grow and thus many detection methods have been developed, especially molecular. We previously developed a PCR method targeting ribC to identify Bartonella sp. A manufactured kit (RealCycler BART, Progenie Molecular) was commercialised shortly thereafter for the detection of Bartonella infection, including Bartonella henselae. METHODOLOGY: We performed a comparison between this test and our in-house PCR assay on 73 lymphadenopathy samples sent to the laboratory for suspicion of CSD.Results/Key findings. Among the 28 positive samples for Bartonella, 21 were identified by the two PCR assays, and seven by the commercial kit only. CONCLUSION: The performance of this commercial kit suggests that it could be a suitable alternative to our in-house PCR assay, highlighting the importance of the molecular methods used to diagnose CSD.

Impact on the medical decision-making process of multiplex PCR assay for respiratory pathogens.

The objective of this study was to determine how clinicians make use of the modern multiplex PCR assays (MPAs) to manage patients hospitalized for community-acquired pneumonia (CAP). We studied the use of MPAs in 1648 patients hospitalized for CAP over a 3-year period at the moment of the setup of the new PCR assay. We observed that the use of MPAs for the identification of multiple respiratory pathogens marks a radical change in the investigation of CAP etiology. Surprisingly, the contribution of MPAs to the medical decision-making process varies drastically according to the units of care.

Diagnosis of community-acquired acute respiratory illness: From conventional microbiological methods to molecular detection (multiplex).

Investigations of the etiologic agents of community-acquired acute respiratory illness may lead to better treatment decisions and patient outcomes. In a routine care setting, we assessed the diagnostic performance of a multiplex PCR assay with respect to conventional microbiological methods, in a continuous series of adult cases of community-acquired acute respiratory illness. We enrolled 279 adult patients hospitalised for community-acquired acute respiratory illness at Tours University Hospital during the winter of 2011-2012. Respiratory samples (mostly nasopharyngeal aspirates) were studied prospectively by indirect immunofluorescence assay and multiplex PCR, that enable detection of 8 viruses and 21 respiratory pathogens respectively. In total, 255 of the 279 (91.4%) samples had interpretable results by both methods. At least one respiratory pathogen was detected by multiplex PCR in 171 specimens (65%). Overall, 130 (76%) of the 171 positive samples were positive for only one respiratory pathogen, 37 (22%) samples were positive for two pathogens and four (2%) were positive for three pathogens. With indirect immunofluorescence assay, a respiratory virus was detected in 27 of the 255 (11%) specimens. Indirect immunofluorescence assay detected some of the influenza virus A (15/51, 29%) infections identified by multiplex PCR and some (7/15, 47%) human metapneumovirus and (5/12, 42%) respiratory syncytial virus infections, but it did not detect all the adenovirus infections. Thus, access to multiplex molecular assays improves the diagnostic spectrum and accuracy over conventional methods, increasing the frequency of identification of the respiratory pathogens involved in community-acquired acute respiratory illness.

Vaginal self-sampling is a cost-effective way to increase participation in a cervical cancer screening programme: a randomised trial.

BACKGROUND: Cervical cancer screening coverage remains insufficient in most countries. Our objective was to assess whether in-home vaginal self-sampling with a dry swab for high-risk human papillomavirus (HR-HPV) testing is effective and cost-effective in increasing participation in cervical cancer screening. METHODS: In March 2012, 6000 unscreened women aged 30-65 years, living in a French region covered by a screening programme, who had not responded to an initial invitation to have a Pap smear were equally randomised to three groups: 'no intervention'; 'recall', women received a letter to have a Pap smear; and 'self-sampling', women received a self-sampling kit to return to a centralised virology laboratory for PCR-based HPV testing. RESULTS: Participation was higher in the 'self-sampling' than in the 'no intervention' group (22.5% vs 9.9%, P<0.0001; OR 2.64) and 'recall' group (11.7%, P<0.0001; OR 2.20). In the 'self-sampling' group, 320 used the self-sampling kit; for 44 of these women with positive HR-HPV test results, 40 had the recommended triage Pap smear. The ICER per extra screened woman was 77.8[euro ] and 63.2[euro ] for the 'recall' and 'self-sampling' groups, respectively, relative to the 'no intervention' group. CONCLUSIONS: Offering an in-home, return-mail kit for vaginal self-sampling with a dry swab is more effective and cost-effective than a recall letter in increasing participation in cervical cancer screening.

Accuracy of dry vaginal self-sampling for detecting high-risk human papillomavirus infection in cervical cancer screening: a cross-sectional study.

OBJECTIVE: Cervical cancer screening coverage remains insufficient in most countries. Testing self-collected samples for high-risk human papillomavirus (HR-HPV) could be an alternative to the Pap smear, but costs, sampling methods and transport issues hamper its wide use. Our objective was to compare diagnostic accuracy of 2 vaginal self-collection methods, a dry swab (vsc-DRY) or swab in liquid medium (vsc-LIQ), for detecting HR-HPV cervical infection assessed by a cervical clinician-collected sample in liquid medium (ccc-LIQ). METHODS: Women 20 to 65 years attending a Pap smear were recruited between September, 2009 and March, 2011. Each sample (3 per woman) underwent HPV DNA testing. Samples were classified as HR-HPV+ with detection of at least one HR-HPV or probable HR-HPV type. RESULTS: Of 734 women included, 722 had complete HPV data. HR-HPV was detected in 20.9% of ccc-LIQ samples. Estimated sensitivity and specificity to detect HR-HPV in vsc-DRY samples were 88.7% and 92.5%, respectively, and in vsc-LIQ samples, 87.4% and 90.9%. Cytology findings were abnormal for 79 women (10.9%): among 27 samples of low-grade squamous intraepithelial lesions, 25 were HR-HPV+ in vsc-DRY, vsc-LIQ and ccc-LIQ samples. Among 6 samples of high-grade squamous intraepithelial lesions, all were HR-HPV+ in vsc-DRY samples, 1 was HR-HPV- in vsc-LIQ samples and 1 was HR-HPV- in ccc-LIQ samples. CONCLUSIONS: Vaginal self-sampling with a dry swab is accurate to detect HR-HPV infection as compared with cervical clinician-collection and accurate as compared with cytology results. This cheap and easy-to-ship sampling method could be widely used in a cervical cancer screening program.

CMV infection in the donor and increased kidney graft loss: impact of full HLA-I mismatch and posttransplantation CD8(+) cell reduction.

Despite a large body of literature, the impact of chronic cytomegalovirus (CMV) infection in donor on long-term graft survival remains unclear, and factors modulating the effect of CMV infection on graft survival are presently unknown. In this retrospective study of 1279 kidney transplant patients, we analyzed long-term graft survival and evolution of CD8(+) cell population in donors and recipients by CMV serology and antigenemia status. A positive CMV serology in the donor was an independent risk factor for graft loss, especially among CMV-positive recipients (R(+) ). Antigenemia was not a risk factor for graft loss and kidneys from CMV-positive donors remained associated with poor graft survival among antigenemia-free recipients. Detrimental impact of donor's CMV seropositivity on graft survival was restricted to patients with full HLA-I mismatch, suggesting a role of CD8(+) cells. In R(+) patients with positive CMV antigenemia during the first year, CD8(+) cell count did not increase at 2 years posttransplantation, in contrast to R(-) recipients. In addition, marked CD8(+) -cell decrease was a risk factor of graft failure in these patients. This study identifies HLA-I full mismatch and a decrease of CD8(+) cell count at 2 years as important determinants of CMV-associated graft loss.

[Study of hospitalizations for pneumococcal pneumoniae in Centre region, 2004-2008]. / Étude des séjours pour pneumopathie à Streptococcus pneumoniae entre 2004 et 2008 en région Centre.

BACKGROUND: Streptococcus pneumoniae is a significant cause of morbidity-mortality: leading agent of community-acquired pneumonia and the first cause of death due to infectious diseases in France. Vaccines are available for children and adults, avoiding serious complications. We studied hospitalizations for pneumococcal pneumonia in Centre region in 2004-2008, using the 2004-2008 national hospital discharges database and assessed vaccine coverage of a sample population. METHODS: Hospital discharges with diagnosis of pneumococcal pneumonia were selected, using the corresponding code (J13) in the French Diagnosis-Related Group coding system. We analysed hospitalization and patient data using linkage of the inpatient stays with their anonymous identification number. We analysed hospitalization and patient data: number and length of stay/patient, co-morbid factors and pneumococcal immunisation status. RESULTS: One thousand five hundred and forty-one hospitalisations were found for pneumococcal pneumonia in Centre Region, in 2004-2008. The time pattern showed an annual increase in hospital stays: winter hospitalisations were most frequent. The median age was 58 years, range: 2 months-106 years. The sex ratio M/F of the 1417 distinct inpatients was 1.43. The hospital impact was substantial, with prolonged length of stay (mean=9.9 days), frequent stays in intensive care unit (20%) and high death rate (6%). The vaccine coverage of the population with pneumococcal pneumonia was only 23%. CONCLUSION: Using the national hospital discharges database, this study presents a snapshot of pneumococcal pneumonia in one French region and demonstrates the local major clinical impact, as found in France. It shows that the hospital discharge database is a potential tool for epidemiology despite its possible bias. This type of study could be useful for organizing a regional vaccination campaing due to the better knowledge of the disease.

[1997-2007, 10 years of monitoring the evolution of resistance of Streptococcus pneumoniae to antibiotics in the region Centre; review of the Pneumococcus network]. / 1997-2007, dix ans de suivi de l'évolution de la résistance de Streptococcus pneumoniae aux antibiotiques en région Centre; bilan de l'observatoire du pneumocoque.

Regional pneumococcal observatories in region Centre, created in 1997, participate with the others pneumococcal observatories alongside the National Reference Center for Pneumococci and the Institut de Veille Sanitaire at the monitoring of the evolution of resistance of pneumococci to antibiotics in France. Between 1997 and 2007, 2427 strains of Streptococcus pneumoniae were isolated in part from cerebrospinal fluids, blood and middle ear fluid, from children and adults. The prevalence of pneumococci with a decreased susceptibility to penicillin (PDSP) decreased strongly in region Centre: 56.8 % in 2001, 39.6 % en 2007. These data are similar to the French national data over the same period.

[Prevalence of extended-spectrum beta-lactamases of the CTX-M type producing Escherichia coli and Klebsiella pneumoniae in Bretonneau hospitals (CHRU Tours)]. / Prévalence des souches de Escherichia coli et de Klebsiella pneumoniae productrices de beta-lactamases à spectre étendu de type CTX-M à l'hôpital Bretonneau (CHRU de Tours).

The CTX-M type extended spectrum beta-lactamases constitute a rapidly growing cluster of enzymes that have disseminated geographically. This study evaluates the prevalence of these enzymes in the university hospital in Tours, in 2007. Twenty-eight strains were studied: 21 Escherichia coli and seven Klebsiella pneumoniae. The gene bla(CTX-M) was detected by real-time PCR for 27 strains. The CTX-M-1 group, including CTX-M-15, was the most frequent CTX-M-type enzyme.

[Pediatric osteoarticular infections caused by Kingella kingae from 1995 to 2006 at CHRU de Tours]. / Infections ostéo-articulaires pédiatriques à Kingella kingae de 1995 à 2006 au CHRU de Tours.

Use of molecular biology shows that Kingella kingae is a pathogen frequently involved in osteoarticular infections in young children. This study describes the cases of osteoarticular infections due to K. kingae which happened from 1995 to 2006 in the CHRU of Tours. The description is based on clinical and biological features. A K. kingae specific polymerase chain reaction was performed in our laboratory in order to improve K. kingae osteoarticular infections diagnosis, and is detailed here.

Genetic diversity of the hepatitis C virus: impact and issues in the antiviral therapy.

The hepatitis C Virus (HCV) presents a high degree of genetic variability which is explained by the combination of a lack of proof reading by the RNA dependant RNA polymerase and a high level of viral replication. The resulting genetic polymorphism defines a classification in clades, genotypes, subtypes, isolates and quasispecies. This diversity is known to reflect the range of responses to Interferon therapy. The genotype is one of the predictive parameters currently used to define the antiviral treatment strategy and the chance of therapeutic success. Studies have also reported the potential impact of the viral genetic polymorphism in the outcome of antiviral therapy in patients infected by the same HCV genotype. Both structural and non structural genomic regions of HCV have been suggested to be involved in the Interferon pathway and the resistance to antiviral therapy. In this review, we first detail the viral basis of HCV diversity. Then, the HCV genetic regions that may be implicated in resistance to therapy are described, with a focus on the structural region encoded by the E2 gene and the non-structural genes NS3, NS5A and NS5B. Both mechanisms of the Interferon resistance and of the new antiviral drugs are described in this review.

Application of the French guidelines for preventing neonatal group B streptococcal disease in a university hospital.

This study evaluated the application of the French guidelines for prevention of neonatal group B streptococcus (GBS) infections. The prevalence of GBS vaginal carriage by pregnant women during the study period was 6%. Less than 50% of pregnant women testing positive for GBS were treated with at least two doses of antibiotics during labour, and most received only one dose or no antibiotics. In addition, several neonates were colonised or infected by GBS although their mothers were GBS-negative. These results are consistent with vaginal screening having a poor sensitivity, as suggested by the low prevalence of GBS carriage.

Is hepatitis C virus NS3 protease quasispecies heterogeneity predictive of progression from cirrhosis to hepatocellular carcinoma?

We investigated whether an HCV NS3 protease quasispecies heterogeneity was associated with progression from viral cirrhosis to hepatocellular carcinoma (HCC). The NS3 protease quasispecies structure of 10 HCV-1b cirrhotic patients (controls) was compared with that of 10 paired HCV-1b cirrhotic patients who displayed progression to HCC (cases). NS3 protease genetic complexity and diversity did not differ significantly between cases and controls. Amino acid substitutions were detected at 20 (11%) and 25 (14%) sites in at least two variants of the NS3 protease in cases and controls, respectively. Significant differences in the percentage of substituted clones were observed for 10 NS3 sites. Mutations Y56F, I71V, T72I, Q86P, P89S, S101G/D, R117H, S122G/T/N, V132I and V170I were more frequently observed in the NS3 protease sequences of controls than in those of cases. Residue V107 was substituted in NS3 cases but not in controls. However, these differences did not allow the definition of a specific NS3 profile related to HCC occurrence. The NS3 secondary structure B1-1 previously identified as potentially predictive of HCC was identified with a higher frequency in cases quasispecies (84.2%) than in controls (55.9%; P < 0.05). Our results suggest that there may be a relationship to fibrosis progression when diversity parameters are considered together with secondary structure profiles. Further investigations are required to determine the cellular interactions of HCV NS3 protease in the context of carcinogenesis.

[Molecular identification of species within the Mycobacterium tuberculosis complex using regions of difference]. / Identification moléculaire des espèces de mycobactéries du groupe de la tuberculose par étude des régions de différence.

The differentiation within the Mycobacterium tuberculosis complex (MTBC) based on phenotypic methods is long and does not give an unambiguous result in every case whereas the advance in genetic knowledge leads to new views. Thus, regions of difference (RD), that seem to characterize the species of the MTBC, have been identified. Amplification methods, targeted on these zones, have been developed then. The study of four regions (RD1, RD5, RD9, RD10) has been done on 64 isolates formerly identified thanks to phenotypic methods. Genotypic results confirm phenotypic identifications except in one case. This strain initially identified as M. tuberculosis and isolated from a Gabonese patient, corresponds, according to genotypic identification, to M. africanum. Since phenotypic characterization of M. africanum is difficult, this method would allow to accurately determine the true prevalence of this specie. Moreover, the study of RD10 doesn't seem to be informative. The amplification of only two RD, RD1 and RD9 carries out the identification of all M. tuberculosis and M. bovis BCG isolates. M. bovis and M. africanum will be then identified thanks to RD5. Thus, this easy and rapid method of identification of the major species of MTBC seems to be appropriated for a routine use.

[Systematic screening tests for Chlamydia trachomatis, Mycoplasma hominis and Ureaplasma urealyticum in urogenital specimens of infertile couples]. / Existe-t-il un bénéfice au dépistage systématique de Chlamydia trachomatis, Mycoplasma hominis et Ureaplasma urealyticum dans les prélèvements génito-urinaires réalisés au cours d'un bilan d'infertilité?

We conducted a prospective study on 100 couples consulting for infertility at the teaching Hospital of Tours, with the scope to determine if there is a benefit for systematic screening of Chlamydia trachomatis, Mycoplasma hominis and Ureaplasma urealyticum among genito-urinary specimen when exploring couples infertility. C. trachomatis was detected by PCR on sperm, endocervix and urine specimen. M. hominis and U. urealyticum were detected by culture on A7 agar medium and with minigaleries on sperm and endocervix specimen. Standard cultures were also performed on sperm, endocervix, vaginal and urine specimen. Only one specimen (sperm) was positive for C. trachomatis. Three percent of the specimen were positive for U. urealyticum (from which 2,5% of the sperm specimen). No specimen was positive for M. hominis. Our results show that screening of C. trachomatis, M. hominis and U. urealyticum is not systematically required for among check up of infertile couples, given the prevalence of chlamydiosis among the population studied. However, it would be interesting to perform it on a targeted population, according to anamnestic or clinical criteria. In addition, an important modification of vaginal flora was observed in 12% of cases, and 2 vaginosis were diagnosed; the putative consequences of this disequilibrium has to be further investigated.

[The genetic diversity of hepatitis C virus and its implications for pathogenesis and resistance to treatment]. / La diversité du virus de l'hépatite C : méthodes d'étude, implication dans la pathogenèse et la résistance au traitement.

Hepatitis C virus (HCV) is characterized by a high genetic variability mainly due to the lack of proofreading activity of its RNA polymerase. Noncoding regions located on 5' and 3' ends of the HCV genome are conserved, whereas the hypervariable regions of the E2 envelope gene are highly mutated. This genetic heterogeneity is observed both between patients and between strains isolated at different time from a given patient. Identification of six major genotypes and of a large number of subtypes has resulted from the phylogenetic analysis of HCV strains isolated from different continents. In infected individuals, HCV exists as pools of related genetic variants, referred to as quasispecies. The role of the genetic variability of HCV in primary resistance to treatment or in the emergence of escape mutants has been described but remains controversial.

[Mutation analysis of ISDR and V3 domains of hepatitis C virus NS5A region before interferon therapy with or without ribavirin]. / Analyse des mutations des domaines ISDR et V3 de la protéine NS5A du virus de l'hépatite C avant le traitement par l'interféron avec ou sans ribavirine.

AIM OF THE STUDY: The hepatitis C virus (HCV) non-structural NS5A protein has been controversially implicated in the resistance of HCV to interferon therapy in clinical studies. In Japan, mutations in the interferon sensitivity-determining region (ISDR) in the NS5A gene were associated with response to interferon therapy in patients infected with genotype 1b. In contrast, studies from Europe did not confirm such association. More recently, it has been suggested that the V3 domain outside the putative ISDR might also have amino acids changes that may be involved in the resistance to IFN. In this study, the relationship between NS5A mutations in ISDR and V3 domains and virological response to therapy were investigated. MATERIALS AND METHODS: The NS5A gene was sequenced from 35 HCV genotype 1b infected patients at D0 of a prospective clinical trial of interferon therapy and interferon plus Ribavirin combination therapy. RESULTS: In the ISDR domain, we did not observe any significant differences in amino acids changes between responders (1.7 +/- 1.8, n = 19, range 0-6) and non-responders (1.1 +/- 0.8, n = 14, range: 0-3), (P = 0.483), to therapy before the beginning of treatment. In the V3 domain, we found more mutations in responders (6.5 +/- 1.9, range: 2-11) than in non-responders (4.7 +/- 1.2, range: 3-8) (P = 0.0013), before the beginning of treatment. CONCLUSION: Our results confirm that, in Europe, the ISDR domain is not predictive for treatment success but suggest that the V3 domain have greater variability in responders than non-responders.