Journal of Clinical Virology

Editorial Board

2010-04-01

Febrile illness in a returned traveller from Thailand

Bjørn Waagsbø, Anders Sundøy, Liv Ragnhild Høyvoll — 2009-12-09

A previously healthy female in her fifties, returned from Phuket, Thailand after a 3-week holiday stay. She was an experienced traveller with multiple earlier journeys to the south-east Asia. On day 3 after returning home to Norway, she fell ill and contacted her primary health care physician by telephone complaining over headache, myalgia, joint pain, vomiting and fever of 38.7°C. Over the next days she experienced transient improvement, although high-peeking fever reaching 39°C persisted. On day 7 she once more phoned her physician and was advised to stay home in town, instead of spending the weekend at her cabin as planned. She went anyway.

Human polyoma viruses and disease with emphasis on clinical BK and JC

Raghavender Boothpur, Daniel C. Brennan — 2010-01-08

Abstract: Polyoma viruses are ubiquitous infecting many different mammalian species including humans. There are five known human polyoma viruses. JC virus and BK virus are two polyoma viruses identified nearly three decades ago. Recently WU, KI and Merkel cell polyoma viruses have been isolated from humans. The exact role of these three newly discovered viruses in human disease is not known. Most human polyoma disease is caused by BK and JC viruses which are usually acquired in childhood. Approximately 50–80% of humans have seropositivity to these viruses. Clinically apparent diseases in immunocompetent hosts are extremely rare. These viruses remain latent possibly in the lymphoid organs, neuronal tissue, and kidney and under the circumstances of severe immunosuppression both these viruses reactivate. Neurotropic JC virus reaches the brain and causes progressive multifocal leukoencephalopathy, a demyelinating disease of the central nervous system with a high mortality rate. BK virus is urotheliotropic and its reactivation causes a form of interstitial nephritis, known as BK or polyoma virus associated nephropathy which is associated with high graft loss if not recognized early. There are no known effective antiviral agents for any of the polyoma viruses.

Microsatellite analysis of HSV-1 isolates: From oropharynx reactivation toward lung infection in patients undergoing mechanical ventilation

2010-02-22

Abstract: Background: According to recent reports, herpes simplex virus type 1 (HSV-1) induces bronchopneumonitis (BPn) in immunocompetent patients undergoing prolonged mechanical ventilation (MV), whose respiratory functions deteriorate with a poor outcome. HSV-1 BPn is associated with HSV symptomatic or symptomless reactivation in the oropharynx.Objectives: We sought to systematically and genetically characterize HSV-1 strains isolated from immunocompetent patients receiving prolonged MV and to characterize the genetic relationship of strains sequentially isolated from oropharyngeal samples (OPS) and broncho-alveolar liquids (BAL) to determine the natural course of HSV BPn.Study design: In this molecular epidemiological study, microsatellite technology was used to determine genetic relationships between 211 HSV-1 strains isolated from OPS and/or BAL from 106 patients receiving MV.Results: Microsatellite haplotypes of HSV-1 strains sequentially isolated from the same individual were identical, and HSV-1 isolates from the lung were genetically indistinguishable from strains isolated from the oral cavity. Each patient was characterized by their own HSV-1 microsatellite haplotype, and no nosocomial transmission of strains between patients was observed.Conclusion: Our results demonstrate that, in patients who receive MV, the HSV-1 pulmonary infection results from the reactivation of genetically related HSV-1 in the oropharynx, which progressively infects the lower respiratory tract.

Incidence and characterization of cytomegalovirus resistance mutations among pediatric solid organ transplant patients who received valganciclovir prophylaxis

Mélanie Martin, Nathalie Goyette, Jane Ives, Guy Boivin — 2010-02-08

Abstract: Background: Drug-resistant cytomegalovirus (CMV) infections can cause significant morbidity among high-risk transplant recipients.Objectives: The aims of this study were to determine the incidence and clinical consequences of CMV mutations conferring ganciclovir resistance in pediatric solid organ transplant (SOT) patients who received valganciclovir oral solution or tablets for prophylaxis of CMV disease. Recombinant CMV mutants were also generated to assess the role of two UL97 mutations of unknown significance.Study design: Genotypic resistance mutations and CMV viral load were sought in blood samples from pediatric SOT recipients who received valganciclovir prophylaxis for 100 days. Recombinant viruses containing novel CMV UL97 mutations were generated using a bacterial artificial chromosome containing the CMV genome to assess ganciclovir susceptibility.Results: Overall, four known resistance UL97 mutations were observed in blood samples from 2 of 46 patients during the study with no development of CMV disease. Two UL97 changes (M615V and V466G) of unknown significance and one UL97 mutation (C603R) associated with ganciclovir resistance, but not yet confirmed by marker transfer, were also detected. Recombinant viruses containing these novel mutations were generated to assess ganciclovir susceptibility. The M615V recombinant virus was susceptible to ganciclovir while the V466G and C603R mutant viruses displayed 3.5-fold and 3.6-fold decreases in susceptibility, respectively.Conclusions: The low incidence of ganciclovir resistance-associated mutations and the absence of clinical consequences associated with drug-resistant viruses observed in this pilot study should encourage the design of larger clinical trials aimed at evaluating the efficacy of valganciclovir prophylaxis and treatment in the pediatric setting.

Disease burden of herpes zoster in Korea

2010-02-25

Abstract: Background: The occurrence of herpes zoster can deteriorate the quality of life considerably, resulting in high disease burden. While Korea is assumed to have high disease burden of herpes zoster, there has been no researches analyzing this.Objectives: We performed this study to investigate the disease burden of herpes zoster in the Korean population as a whole.Study design: We used the database of the Health Insurance Review & Assessment Service of Korea and analyzed the data of patients who had herpes zoster as a principal diagnosis during the period from 2003 to 2007. We investigated the annual prevalence, rate of clinical visits, rate of hospitalization, and the pattern of medical services use. The socioeconomic burden of herpes zoster was calculated by a conversion into cost.Results: Rates of clinic visits and hospitalizations due to herpes zoster during the 5-year period from 2003 to 2007 were 7.93–12.54 per 1000 population and 0.22–0.32 per 1000 population, respectively. Prevalence rates according to age increased sharply after 50 years and reached a peak at 70 years. The total socioeconomic cost of herpes zoster was $75.9–143.8 million per year, increasing every year by 14–20%.Conclusions: There is a heavy socioeconomic burden due to herpes zoster in Korea and indicate that appropriate policies need to be established to reduce this burden. Additional researches are also necessary to assess the safety, efficacy and cost-effectiveness of a herpes zoster vaccine in the Korean population.

Using of nevirapine is associated with intermediate and reduced response to etravirine among HIV-infected patients who experienced virologic failure in a resource-limited setting

2010-02-26

Abstract: Background: Non-nucleoside reverse transcriptase inhibitor (NNRTI)-based regimens have been extensively used for treatment of HIV infection in resource-limited settings. Treatment options after failing an initial regimen are limited because of cross-resistance of NNRTIs.Objective: To determine the factors associated with reduced response to etravirine among patients with virological failure.Study design: A retrospective study was conducted. We stratified patients into two groups by the total weighted scores of etravirine-resistance-associated mutations (ETV-RAMs), highest response (score 0–2, N=123) and intermediate and reduced response (score ≥2.5, N=61). Factors associated with a score of ≥2.5 were evaluated.Results: There were 184 patients with mean (SD) age of 42 (9) years old and 60% were males. Of all, 68% used NNRTI in the failing regimen and 51% used stavudine/lamivudine as a backbone. Common ETV-RAMs included Y181C (27%), G190A (17%), and K101E (10%). Higher proportion of K101E, K101P, Y181C, G190S, and M230L were found in patients with a score of ≥2.5 (p<0.05, all). By univariate logistic regression, using protease inhibitor (OR 0.22, 95% CI 0.07–0.67), nevirapine (OR 10.56, 95% CI 4.04–27.74), and efavirenz (OR 2.91, 95% CI 1.01–2.51) in the current regimen were associated with a score of ≥2.5. By multiple logistic regression, only using nevirapine was associated with a score of ≥2.5 (OR 7.61, 95% CI 2.40–24.06).Conclusions: Using nevirapine in the failing regimen was associated with intermediate and reduced response to ETV. The recommendation of using nevirapine as a preferred NNRTI should be re-considered in resource-limited settings where efavirenz is accessible.

Viral load assay sensitivity and low level viremia in HAART treated HIV patients

2010-02-08

Abstract: Background: The recent introduction of highly sensitive viral load assays resulted in a significant increase in number of treated HIV-infected patients with a detectable viral load. The significance of a viral load between 20 and 50copies/mL remains unclear.Objectives: To compare the performance of three viral load assays, with special attention for specificity and sensitivity at the lowest level of quantification.Study design: Samples (n=181) were selected from 62 HIV-positive individuals that experience viral blips or episodes of low but detectable viremia under antiretroviral treatment, and from 216 HIV-negative individuals. Each sample was tested in at least two of three assays: the Cobas Amplicor HIV-1 Monitor (CAP/CA), the Cobas Ampliprep/Cobas TaqMan HIV-1 version 1 (CAP/CTM1) and the Cobas Ampliprep/Cobas TaqMan HIV-1 version 2 (CAP/CTM2).Results: No false positive results were recorded. Kappa statistics revealed fair to moderate agreement between the results of the three assays, but important differences in sensitivity were observed, with the highest sensitivity reported for CAP/CTM2 followed by CAP/CTM1 and CAP/CA. The differences in sensitivity remained after equalization of the detection limit for all assays at 50copies/mL. Analysis of samples collected over time showed that patients with single blips in CAP/CA present with recurrent blips in CAP/CTM1 and persistent detectable viremia in CAP/CTM2.Conclusions: Viral load results between 20 and 50copies/mL in either CAP/CTM1 or CAP/CTM2, indicate true viremia. The availability of highly sensitive assays force reconsideration of the terms ‘undetectable’ viral load and ‘virological success’ of antiretroviral treatment.

Identification of G8 rotavirus strains determined as G12 by rotavirus genotyping PCR: Updating the current genotyping methods

Farah Aladin, Sameena Nawaz, Miren Iturriza-Gómara, Jim Gray — 2010-02-08

Abstract: Background: Rotaviruses are classified into G- and P-types, which are determined by the reactivity with antibodies to the outer viral proteins, VP7 and VP4, respectively, or sequence variation in the genes encoding these proteins. There are presently a number of different rotavirus strains co-circulating within the UK, with the common human strains G1P[8], G2P[4] and G9P[8] being the most prevalent. As part of strain surveillance for the European Rotavirus Network (EuroRotaNet) a cluster (n=29) of G8 strains was detected in the UK between February and May 2009.Objectives: G8 strains were initially mistyped as G12 through multiplex RT-PCR, therefore further investigation was performed to ascertain the reasons behind this mistyping.Study design: The genes encoding the VP7 of these G8 strains were sequenced and aligned with the existing G8- and G12-specific oligonucleotide primers.Results: Multiple alignment of sequences derived from these strains and the G8- and G12-specific oligonucleotide primers revealed a series of point mutations which resulted in mismatches at the 3′ end of the G8-specific primer binding site that prevented amplification with the G8-specific primer, whilst a close homology with the G12-specific primer allowed mis-priming. Both the G8 and G12 primers were redesigned and their ability to correctly identify G8 and G12 strains was evaluated and confirmed.Conclusion: These findings highlight the importance of monitoring the specificity and sensitivity of the genotyping methods in order to detect changes in the genotype distribution and changes associated with genetic drift of common or uncommon genotypes.

Phylogenetic analysis of human P[8]G9 rotavirus strains circulating in Brazil reveals the presence of a novel genetic variant

2010-02-05

Abstract: Background: Group A rotavirus (RV-A) genotype P[8]G9 has emerged as one of the leading causes of gastroenteritis in children worldwide and currently is recognized as one of the five most common genotypes detected in humans. High intragenotype diversity in G9 RV-A has been observed, and nowadays, based on the genetic variability of the VP7 gene, six different phylogenetic lineages and eleven sublineages were described.Objectives: To study the degree of genetic variation and evolution of Brazilian P[8]G9 RV-A strains.Study design: Phylogenetic analysis of 19 P[8]G9 RV-A strains isolated from 2004 to 2007 in five different Brazilian states was conducted using the NSP1, NSP3, NSP5, VP4 and VP7 genes. For the VP4 and VP7 genes, 3D protein structure predictions were generated to analyze the spatial distribution of amino acid substitutions observed in Brazilian strains.Results: Based on the phylogenetic analyses, all Brazilian strains clustered within lineage G9-III and P[8]-3 for VP7 and VP4, respectively, and were classified as genotype A1, T1 and H1 for the NSP1, NSP3 and NSP5 genes, respectively. Interestingly, all the strains isolated in Acre State (Northern Brazil) formed a closely related cluster clearly separated from the other Brazilian and prototype strains with regard to the five genes studied. Unique amino acid substitutions were observed in Acre strains in comparison with the prototype and Brazilian strains.Conclusion: Inclusion of Acre strains in the phylogenetic analysis revealed the presence of a novel genetic variant and demonstrated a diversification of P[8]G9 rotaviruses in Brazil.

Are hepatitis B virus “subgenotypes” defined accurately?

2010-02-15

Abstract: Background: Recently, several novel hepatitis B virus (HBV) subgenotypes have been introduced that do not meet proper definition of “subgenotypes”. In particular for HBV genotype A, such novel subgenotypes have been reported.Objective: To comprehensively reanalyse all HBV subgenotypes A, and to propose a novel, consistent alternative for HBV classification.Study design: All HBV full-length genome subgenotypes A1–A6 were reanalysed using phylogenetic reconstruction and genetic distance calculation in order to study their evolutionary relationships.Results: Phylogenetic analysis based on the complete genome sequence of subgenotype A strains revealed four distinct clusters supported by high bootstrap values, whereas only the three groups A1, A2 and A6 could be assigned as subgenotypes. Previously introduced subgenotype A3, “tentative A4” and A5 clustered together in one main branch and were designated as “quasi-subgenotypes”. Also genetic distances failed to classify these three groups as definite subgenotypes. These results advocate for a new classification of HBV genotype A into subgenotype A1, A2, “quasi-subgenotype A3” and A4.Conclusion: Detailed phylogenetic analysis of the complete genome sequences demonstrates that some of available HBV genotype A strains may not be considered as definite “subgenotypes”. These strains, which are mainly of African origin, could be considered as “quasi-subgenotypes” which puts them in between the “clade” and “subgenotype” definition. Geographical origin may have a key role in further classification of HBV subgenotypes.

The effectiveness of routine laboratory findings in determining disease severity in patients with Crimean-Congo hemorrhagic fever: Severity prediction criteria

Gurdal Yilmaz, Iftihar Koksal, Murat Topbas, Hulya Yilmaz, Firdevs Aksoy — 2010-02-10

Abstract: Background: Crimean-Congo hemorrhagic fever (CCHF) is a potentially fatal disease caused by a tick-borne virus from the Bunyaviridae family.Objectives: To determine the predictive criteria for severity among patients with CCHF based on clinical and laboratory findings.Study design: This retrospective study was conducted on patients with CCHF and hospitalized between June 2004 and August 2008 at Karadeniz Technical University, Turkey. Demographic characteristics, clinical findings and laboratory tests on admission of all patients with CCHF were investigated.Results: A total of 152 patients with confirmed CCHF were investigated. Sixty-three (41.4%) of these patients were in the severe group. Laboratory findings using the ROC curve method and optimum diagnostic cut-off points for specific laboratory parameters in the severe group were; PLT: 90,000, Hb: 13.5g/dL, PT: 13.1s, aPTT: 34s, INR: 1, AST: 117U/L, ALT: 71U/L, AST/ALT: 1.62, LDH: 508U/L, CK: 267U/L and CRP: 0.59mg/dL. At multivariable analysis, the risk for a severe clinical course in CCHF patients increased 2.59 and 3.93 times in the presence of platelet count and Hb below cut-off values, whereas the same risk increased 2.95, 2.92 and 3.47 times when the results for INR, AST and CRP, respectively, were above the predetermined cut-off values.Conclusions: A number of laboratory findings that can easily be measured at routine examination of patients hospitalized with a suspicion of CCHF are valuable and sensitive predictors. These parameters will contribute considerably to the design, practice and management of supportive treatment, blood and blood products replacement and intensive care services.

Human adenoviruses in respiratory infections: Sequencing of the hexon hypervariable region reveals high sequence variability

Barbara Biere, Brunhilde Schweiger — 2010-02-11

Abstract: Background: In respiratory infections, human adenoviruses (hAdV) of species B1 and C are frequently detected, but severe or even fatal disease outbreaks are predominantly caused by only few serotypes. The molecular typing of hAdV hexon sequences can help to speed up the discrimination of serotypes, thus improving on-time epidemiological examinations and patient care.Objectives: We aimed to develop a molecular method for the rapid species B1 and C serotype identification in respiratory samples based on sequence generation of the hexon hypervariable region (HVR).Study design: We developed two PCR-based genotyping systems for a generic HVR amplification and sequence determination of species B1 and C viruses. The assays were applied to hAdV prototypes and 106 samples.Results: The primer sets proved to be capable of amplifying all B1 and C serotypes. The viruses detected in clinical samples belong to serotypes 1, 2, 3, 5 and 6. The obtained sequences of serotypes 2, 3 and 5 form 2–3 phylogenetic clusters that are based on the characteristic amino acid changes within the variable HVR sites.Conclusions: Our assay can significantly speed up the time-span needed for serotype identification and will improve epidemiological surveillance and patient care. The obtained hexon sequences of field viruses vary significantly and form multiple genetic lineages. The variability is focussed on the HVR sites and can be interpreted as the ongoing evolutionary process. Further research is needed on the hexon sequence variability of other (respiratory) hAdV serotypes.

Impact of extensive HIV-1 variability on molecular diagnosis in the Congo basin

2010-02-22

Abstract: Background: Previous studies have shown a high HIV-1 genetic variability in the Republic of Congo. This can greatly influence the performance of molecular assays for HIV-1 diagnosis.Objectives: To define a reliable test for detection of HIV-1 DNA in this area.Study design: We compared a commercial nested PCR (C-PCR) assay and an in house nested PCR (H-PCR) assay for the detection of HIV-1 DNA in the first 30 seropositive pregnant women enrolled into the ongoing “Kento-Mwana” project, for the prevention of HIV mother-to-child transmission in the city of Pointe Noire, Republic of Congo, Africa. Sequencing and phylogenetic analysis of partial HIV-1 pol sequences were also performed.Results: C-PCR detected HIV-1 DNA in only 15/30 samples from seropositive women (50.0%), as opposed to 29/30 (96.6%) by H-PCR (P<0.0001). Phylogenetic analysis and bootscanning showed that only 10 sequences could be assigned to known clades (seven pure subtypes and three circulating recombinant forms), whereas the other 20 sequences were unique recombinant forms.Conclusions: The great genetic variability of HIV-1 in this area strongly advises to for using molecular methods only after local validation to avoid false negative results.

HIV-1 elite controllers: Beware of super-infections

Olivier Clerc, Sara Colombo, Sabine Yerly, Amalio Telenti, Matthias Cavassini — 2010-02-15

Abstract: Super- and co-infection with HIV-1 are generally associated with accelerated disease progression. We report on the outcome of super-infection in two HIV-1 infected individuals previously known as elite controllers. Both presented an acute retroviral syndrome following super-infection and showed an immuno–virological progression thereafter. Host genotyping failed to reveal any of the currently recognized protective factors associated with slow disease progression. This report indicates that elite controllers should be informed of the risk of super-infection, and illustrates the complexity of mounting broad anti-HIV immunity.

Detection and characterization of human parechoviruses in archived cell cultures, in Hungary

Ákos Boros, Mária Új, Péter Pankovics, Gábor Reuter — 2010-02-15

Abstract: Background: Human parechoviruses (HPeV) belonging to the family Picornaviridae are widespread enteric pathogens and associated with various clinical symptoms in humans.Objectives: There is no report for detection of the circulating parechoviruses in Central Europe. The aim of this retrospective study was to detect and characterize human parechoviruses in cell cultures with “enterovirus”-like cytophatic effect (CPE) archived between 1990 and 2000, in Hungary.Study design: Fecal samples from children with symptoms of gastroenteritis under age of 10 were cultured as a previous routine diagnostic laboratory protocol for “enterovirus”. Cell cultures indicating CPE were archived and deeply chilled (−80°C) from minimum 2 individuals (2–12 patients) in each year. Specimens were tested retrospectively, in 2009, for HPeV by reverse transcription-PCR (RT-PCR) method using 5′UTR conserved primers. Specific primer pairs were designed to determine the complete nucleotide sequence of the structural region (VP0–VP3–VP1) of HPeV.Results: Nine (9.1%) of 66 archived samples were found to be HPeV-positive. Six (67%) samples were identified as HPeV1, 2 (22%) were HPeV4 and 1 could not be determined. Three HPeV1 clusters were identified according to the isolation date originated from years 1990/1991, 1992/1995 and 1998.Conclusions: This is the first detection of human parechoviruses in Central Europe. Detection and genetic characterization of HPeV in available historical samples infected with previously unidentifiable agents with “enterovirus-like” cytopathogenic effect help to understand the genetic diversity and evolution of these viruses.

Emergence of a genomic variant of the recombinant 2k/1b strain during a mixed Hepatitis C infection: A case report

2010-02-15

Abstract: To date, few natural intergenotypic recombinant hepatitis C virus (HCV) strains have been characterized. A recombinant strain 2k/1b was detected for one HCV RNA-positive individual who had just completed therapy for HCV 3a genotype infection. In the present report, five serum samples collected over the pre- and post-treatment periods were used to investigate all the present HCV strains and the change over time of the infection pattern.Interestingly, the 2k/1b strain was already present during the genotype 3a infection and persisted during treatment. In the specimen collected three months post-treatment, two distinct strains, 2k/1b and type 1, were found and then one 2k/1b strain in the subsequent ones. A genomic variant of the HCV RF1_2k/1b strain was identified. It was part of a mixed HCV infection and persisted and re-emerge after eradication of the dominant subtype 3a. This case indicates that HCV co-infection screening after relapse should be an alternative to explain the lack of response to treatment and the necessity to carefully study the epidemic spreading of this recombinant strain.

Pooling the wrong conclusion

Da-Dong Huang — 2010-02-18

I read with interest the proof-of-concept study by Woo et al. on the use of high-density resequencing microarray for the detection of viruses from bacterial culture-negative conjunctival swabs.

Relative lymphopenia and monocytosis may be considered as a surrogate marker of pandemic influenza a (H1N1)

2010-02-04

A novel influenza A (H1N1) [pandemic influenza A (PI)] virus has spread rapidly across the globe. In March and April 2009, an outbreak of respiratory illness was first noted in Mexico, which was eventually identified as being related to PI. The outbreak disseminated rapidly to the other countries. Turkey is among the mostly affected countries.

ESCV Membership

2010-04-01