Showing papers in "Journal of Medical Virology in 2012"

Review: Viral infections and mechanisms of thrombosis and bleeding.

Abstract: Viral infections are associated with coagulation disorders. All aspects of the coagulation cascade, primary hemostasis, coagulation, and fibrinolysis, can be affected. As a consequence, thrombosis and disseminated intravascular coagulation, hemorrhage, or both, may occur. Investigation of coagulation disorders as a consequence of different viral infections have not been performed uniformly. Common pathways are therefore not fully elucidated. In many severe viral infections there is no treatment other than supportive measures. A better understanding of the pathophysiology behind the association of viral infections and coagulation disorders is crucial for developing therapeutic strategies. This is of special importance in case of severe complications, such as those seen in hemorrhagic viral infections, the incidence of which is increasing worldwide. To date, only a few promising targets have been discovered, meaning the implementation in a clinical context is still hampered. This review discusses non-hemorrhagic and hemorrhagic viruses for which sufficient data on the association with hemostasis and related clinical features is available. This will enable clinicians to interpret research data and place them into a perspective.

A diagnostic polymerase chain reaction assay for Zika virus.

Abstract: Zika virus (ZIKV) is a mosquito-borne flavivirus. Infection results in a dengue-like illness with fever, headache, malaise, and a maculopapular rash. Nearly all cases are mild and self-limiting but in 2007, a large outbreak of ZIKV was reported from the island of Yap (in Micronesia, northwest of Indonesia). Singapore is already endemic for dengue, and its impact on public health and economic burden is significant. Other dengue-like infections (e.g., Chikungunya virus) are present. Yet only 10% of reported dengue cases have laboratory confirmation. The identification and control of other dengue-like, mosquito-transmitted infections is thus important for the health of Singapore's population, as well as its economy. Given that ZIKV shares the same Aedes mosquito vector with both dengue and Chikungunya, it is possible that this virus is present in Singapore and causing some of the mild dengue-like illness. A specific and sensitive one-step, reverse transcription polymerase chain reaction (RT-PCR) with an internal control (IC) was designed and tested on 88 archived samples of dengue-negative, Chikungunya-negative sera from patients presenting to our hospital with a dengue-like illness, to determine the presence of ZIKV in Singapore. The assay was specific for detection of ZIKV and displayed a lower limit of detection (LoD) of 140 copies viral RNA/reaction when tested on synthetic RNA standards prepared using pooled negative patient plasma. Of the 88 samples tested, none were positive for ZIKV RNA, however, the vast majority of these were from patients admitted to hospital and further study may be warranted in community-based environments.

Current prevalence of HTLV-1 in Japan as determined by screening of blood donors.

Abstract: Human T-cell leukemia virus type-1 (HTLV-1), a major source of adult T-cell leukemia and related diseases, is endemic to southwestern Japan Mother-to-infant transmission via breast milk is an important route of infection, and establishing programs to prevent such transmission requires exact figures on the HTLV-1 prevalence rate and the number of carriers Therefore, the seroprevalence of HTLV-1 among 1,196,321 Japanese first-time blood donors from 2006 to 2007 was investigated A total of 3,787 of such donors were confirmed to be positive for anti-HTLV-1 antibody By applying a fitness curve to the age ranges outside the blood donor age range, the present number of HTLV-1 carriers covering ages from 0 to 99 years was estimated to be at least 108 million in Japan; this value was 10% lower than that reported in 1988 The adjusted overall prevalence rates were estimated to be 066% and 102% in men and women, respectively The peak in carrier numbers was found among individuals in their 70s, which is a shift from the previous peak observed in the 1988 database among individuals in their 50s Carriers were distributed not only in the endemic southwestern region of Japan, but throughout the country, particularly in the greater Tokyo metropolitan area By applying population projections, it was calculated that the carrier number will decrease by half in the next two decades; however, the carrier population will age over that interval, meaning that the age of patients with adult T-cell leukemia will also be higher

Identification of prohibitin as a Chikungunya virus receptor protein

Abstract: National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency,Pathumthani, ThailandChikungunya virus (CHIKV) has recently re-emerged causing millions of infections in coun-tries around the Indian Ocean. While CHIKV hasa broad host cell range and productively infectsa number of different cell types, macrophageshave been identified as a potential viral reser-voir serving to increase the duration of symp-toms. To date no CHIKV interacting protein hasbeen characterized and this study sought toidentify CHIKV binding proteins expressed ontarget cell membranes. Two-dimensional virusoverlay identified prohibitin (PHB) as a micro-glial cell expressed CHIKV binding protein. Co-localization, co-immunoprecipitation as well asantibody and siRNA mediated infection inhibi-tion studies all confirmed a role for PHB in me-diating internalization of CHIKV into microglialcells. PHB is the first identified CHIKV receptorprotein, and this study is evidence that PHBmay play a role in the internalization of multipleviruses. J. Med. Virol. 84:1757–1770,2012.

Prevalence and genotype distribution of human papillomavirus infection of the cervix in Spain: the CLEOPATRE study.

Abstract: Human papillomavirus (HPV) infection is a necessary cause of cervical cancer. The aim of this study was to estimate the prevalence of cervical HPV infection and HPV type-specific distribution among women attending cervical cancer screening in Spain during 2007 and 2008. Women aged 18-65 years were recruited according to an age-stratified sampling method. Liquid-based cervical samples were collected and analyzed for cytology, HPV detection, and genotyping. HPV genotyping was determined using the INNO-LiPA HPV Genotyping Extra Reverse Hybridization Line Probe Assay. Prevalence estimates were age-standardized using 2001 Spanish census data. The present study included 3,261 women. Age-standardized HC2-based HPV prevalence was 14.3% (95% CI, 13.1-15.5) among women aged 18-65 years, and 28.8% (26.6-31.1) among women aged 18-25 years. High-risk HPV types were detected in 12.2% (95% CI, 11.1-13.4) of HPV-tested women, representing 84.0% of HPV-positive samples. Multiple infections were present in 4.1% (95% CI, 3.4-4.8) of HPV-tested women (25.0% of HPV-positive samples). The most common high-risk HPV-types among HPV-tested women were 16 (2.9%), 52 (1.8%), 51 (1.6%), 31 (1.3%), and 66 (1.2%). HPV-type 16 was present in 16.9% of HPV-positive samples. One or more of the HPV vaccine types 6/11/16/18 were detected in 3.8% of HPV-tested women (22.1% of HPV-positive samples). Though not a true population-based survey, this study provides valuable baseline data for future assessment of the impact of current HPV vaccination programs in Spain. The high prevalence of HPV infection among young women may reflect recent changes in sexual behavior.

Comprehensive analysis of the prevalence of hepatitis B virus escape mutations in the major hydrophilic region of surface antigen

Abstract: Escape mutations in the major hydrophilic region (MHR) of hepatitis B surface antigen (HBsAg) are reported widely worldwide; these mutations lead to diagnostic problems, emergence of vaccine-escape mutants, and hepatitis B immunoglobulin (HBIG) therapy failure. However, the prevalence of these mutations in different genotypes remains to be studied systematically. In the current study, 11,221 non-redundant hepatitis B virus (HBV) sequences of 8 genotypes (from A to H), obtained from the National Center for Biotechnology Information (NCBI), were analyzed to determine the prevalence of HBsAg escape mutations that were previously described. Eight important mutations associated with diagnostic failure, P120T, T126S, Q129H, G130N, S143L, D144A, and G145A/R, were prevalent in one or more genotypes, with the frequency of no less than 1%. With regard to escape variants that evade vaccine or immunoglobulin therapy, mutations were located mainly at positions 120, 126, 129, 130, 133, 134, 137, 140, 143, 144, and 145. The majority of such mutations showed genotypic heterogeneity, indicating the different distribution of the escape mutations. Most of the escape mutations clustered in the "a" determinant, indicating that this region was more likely to be affected by immune selection or antiviral therapy than other regions. Understanding the prevalence and heterogeneity of escape mutations could provide useful guidance for the improvement of diagnostic assays, design of new vaccines, and prevention of failure of HBIG therapy.

Molecular epidemiology of rotavirus and norovirus in Ile‐Ife, Nigeria: High prevalence of G12P[8] rotavirus strains and detection of a rare norovirus genotype

Abstract: Rotavirus (RV) and norovirus (NoV) are considered the most common causes of viral gastroenteritis in children. In this study, the prevalence of RV and NoV infection in 55 children with diarrhea fro ...

Reactivation of herpes simplex virus type 1 is associated with cytomegalovirus and age.

Abstract: Recent studies have shown that Cytomegalovirus (CMV) may be an emerging marker of immunosenescence. CMV can affect the immune system by directly infecting leukocytes and hematopoietic cells or by eliciting an expansion of oligoclonal CD8+ T cells/contraction of the naive T cell compartment that may reduce the host’s ability to fight other pathogens. To investigate further CMV-associated changes in immunity, a study was conducted with 1,454 adults (ages 25–91) to determine the association between CMV and reactivation of another latent herpesvirus, Herpes simplex virus type 1 (HSV-1), as indexed by antibody titers. Elevated antibody titers to latent HSV-1 were significantly associated with both CMV seropositivity and high CMV antibody levels. Evaluation by specific age groups (<45, 45–64, and 65+ years old) revealed that this association was detectable early in life (<45 years of age). Increases in HSV-1 antibodies by age occurred in CMV seropositive individuals but not CMV seronegative subjects. Within CMV seropositive subjects, increases in HSV-1 antibodies by age were only found in individuals with low CMV antibody levels as those with high CMV antibodies already exhibited elevated HSV-1 antibodies. These associations remained significant after accounting for body mass index, gender, and socioeconomic status. These results suggest that CMV can influence the immune response to another pathogen and support the concept that CMV may accelerate immunosenescence.

Distribution of hepatitis C virus genotypes in a diverse US integrated health care population.

Abstract: Hepatitis C virus (HCV) genotypes influence response to therapy, and recently approved direct-acting antivirals are genotype-specific. Genotype distribution information can help to guide antiviral development and elucidate infection patterns. HCV genotype distributions were studied in a diverse cross-section of patients in the Northern California Kaiser Permanente health plan. Associations between genotype and race/ethnicity, age, and sex were assessed with multivariate logistic regression models. The 10,256 patients studied were median age 56 years, 62% male, 55% White non-Hispanic. Overall, 70% were genotype 1, 16% genotype 2, 12% genotype 3, 1% genotype 4, <1% genotype 5, and 1% genotype 6. Blacks (OR 4.5 [3.8-5.5]) and Asians (OR 1.2 [1.0-1.4]) were more likely to have genotype 1 than 2/3 versus non-Hispanic Whites. Women less likely had genotype 1 versus 2/3 than did men (OR 0.86 [0.78-0.94]). Versus non-Hispanic Whites, Asians (OR 0.38 [0.31-0.46]) and Blacks (OR 0.73 [0.63-0.84]) were less likely genotype1a than 1b; Hispanics (OR 1.3 [1.1-1.5]) and Native Americans (OR 1.9 [1.2-2.8]) more likely had genotype 1a than 1b. Patients age ≥65 years less likely had genotype 1a than 1b versus those age 45-64 (OR 0.34 [0.29-0.41]). The predominance of genotype 1 among all groups studied reinforces the need for new therapies targeting this genotype. Racial/ethnic variations in HCV genotype and subtype distribution must be considered in formulating new agents and novel strategies to successfully treat the diversity of hepatitis C patients.

Antiviral activity of arbidol, a broad-spectrum drug for use against respiratory viruses, varies according to test conditions.

Abstract: The therapeutic activity of arbidol was investigated against representatives of seven different virus families. Its 50% median effective concentration (EC(50) ) was 0.22-11.8 µg/ml (0.41-22 nM). Therapeutic indices of 91 were obtained for type 1 poliovirus and 1.9-8.5 for influenza A and B, human paramyxo-3, avian infectious bronchitis-, and Marek's disease viruses. Arbidol was more inhibitory for influenza A/Aichi/2/68 (H3N2) virus than rimantadine or amantadine (EC(50) 10 vs. >15 and >31.6 µg/ml); greater inhibition occurred when end-points were expressed as TCID(50) s. For respiratory syncytial virus (RSV), a reduction in plaque size but not number was observed. However, when the drug was added to infected cultures (≥5 µg/ml), a 3-log reduction in titer occurred. Arbidol did not inhibit directly influenza A/Aichi/2/68 hemagglutinin (HA) or neuraminidase (NA) activity, but inhibition of fusion between the viral envelope and chicken red blood cells occurred when added at 0.1 µg/ml prior to infection. Arbidol induced changes to viral mRNA synthesis of the PB2, PA, NP, NA, and NS genes in MDCK cultures infected with influenza A/PR/8/34. There was no indirect evidence of enhancement of interferon-α by arbidol following infection with A/Aichi/2/68. Arbidol neither reduced lung viral titers nor caused a significant reduction of lung consolidation in BALB/c mice after administration by the oral and intraperitoneal (i.p.) routes and intranasal challenge with influenza A/Aichi/2/68. A small reduction in lung consolidation, but not viral titer, occurred after i.p. administration and subsequent challenge with RSV. The results indicate the potential of arbidol as a broad-spectrum respiratory antiviral drug.

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

Abstract: 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 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.

Characterization of putative Japanese encephalitis virus receptor molecules on microglial cells

Abstract: Japanese encephalitis virus (JEV) a mosquito-borne flavivirus is a major cause of viral encephalitis in Asia. While the principle target cells for JEV in the central nervous system are believed to be neurons, microglia are activated in response to JEV and have been proposed to act as a long lasting virus reservoir. Viral attachment to a host cell is the first step of the viral entry process and is a critical mediator of tissue tropism. This study sought to identify molecules associated with JEV entry to microglial cells. Virus overlay protein-binding assay (VOPBA) and liquid chromatography-mass spectrometry (LC/MS/MS) identified the 37/67 kDa high-affinity laminin receptor protein and nucleolin as a potential JEV-binding proteins. These proteins were subsequently investigated for a contribution to JEV entry to mouse microglial BV-2 cells together with other possible candidate receptor molecules including Hsp70, Hsp90, GRP78, CD14, and CD4. In antibody mediated inhibition of infection experiments, both anti-laminin receptor and anti-CD4 antibodies significantly reduced virus entry while anti-Hsp70 and 90 antibodies produced a slight reduction. Significant inhibition of virus entry (up to 80%) was observed in the presence of lipopolysaccharide (LPS) which resulted in a complete down-regulation of CD4 and moderate down-regulation of CD14. These results suggest that multiple receptor proteins may mediate the entry of JEV to microglial cells, with CD4 playing a major role.

Monitoring the HTLV-1 proviral load in the peripheral blood of asymptomatic carriers and patients with HTLV-associated myelopathy/tropical spastic paraparesis from a Brazilian cohort: ROC curve analysis to establish the threshold for risk disease.

Abstract: Human T-lymphotropic virus 1 (HTLV-1) infection is associated with HTLV-associated myelopathy/tropical spastic paraparesis (HAM/TSP), which affects approximately 5% of carriers. High proviral load is a risk marker for HAM/TSP, although there is an overlap of proviral load levels in peripheral blood between asymptomatic carriers and HAM/TSP patients. In this study, receiver operating characteristic curve analysis was used to define a set point of HTLV-1 proviral load that better indicates an increased risk for HAM/TSP. Proviral load was quantified in 75 asymptomatic carriers and 78 HAM/TSP patients in a Brazilian cohort. The cut-off of proviral load was defined as 114 copies/104 cells, with 78.2% sensitivity to identify true HAM/TSP patients. The mean proviral load levels were not significantly different between males and females with the same clinical status, and there was no significant correlation between proviral load and age at blood sampling, age at the onset of illness, or duration of disease. In HAM/TSP patients, proviral load was significantly higher in wheelchair-bound patients than in individuals able to walk without support and in those with the worst spinal cord injuries. Follow-up of HTLV-1-infected individuals showed that proviral load was more stable in asymptomatic carriers than in HAM/TSP patients. In a cohort study, periodically quantifying proviral load in asymptomatic carriers is necessary to identify those at risk for developing neurological disease, and it is necessary for HAM/TSP patients to monitor spinal injury and progression to walking disability. The measure of proviral load in clinical practice implicates the definition of the cut-off of proviral load and its validation during follow-up. J. Med. Virol. 84:664–671, 2012. © 2011 Wiley Periodicals, Inc.

Single treatment with ethanol hand rub is ineffective against human rhinovirus--hand washing with soap and water removes the virus efficiently.

Abstract: Ethanol-containing hand rubs are used frequently as a substitute for hand washing with water and soap. However, not all viruses are inactivated by a short term rubbing with alcohol. The capacity of a single round of instructed and controlled hand cleaning with water and soap or ethanol-containing hand rub, respectively, was tested for removal of human rhinovirus administered onto the skin of healthy volunteers on the back of the hands. Hand washing with soap and water appeared to be much more efficient for removing rhinoviruses from skin than rubbing hands with an ethanol-containing disinfectant. After washing with soap and water the virus was detected in 3/9 (33.3%) test persons from the left hand and 1/9 (11.1%) cases from the right hand, whereas the virus was detected invariably by real-time RT-PCR from both hands after cleaning with alcohol hand rub (P-value <0.01). Both substances evaluated clinically were also tested in vitro for virucidal efficacy against Human rhinovirus2 (HRV2) using a standardized assay. Both tested substances were poor within the contact time used in the hand-cleaning test. In conclusion, thorough and conventional hand washing with water and soap can clean efficiently hands contaminated with the virus responsible for an extensive share of common cold episodes.

Prevalence, risk factors, and molecular epidemiology of hepatitis B and hepatitis delta virus in pregnant women and in patients in Mauritania.

Abstract: No recent data are available on hepatitis B virus (HBV) and hepatitis Delta virus (HDV) prevalence in Mauritania. One thousand twenty pregnant women and 946 patients visiting for routine checkups were screened for HBV and HDV infection. Demographic, epidemiological, ethnic, clinical, and biological data were recorded. HBV and HDV genotypes were determined by sequencing and phylogenetic analyses. In the pregnant women and patients cohorts, respectively, the prevalence of HBsAg (10.7% and 18.3%) and anti-HBcAb (66.3% and 76.5%) indicated high HBV endemicity. In pregnant women, exposure to HBV was significantly associated in multivariate analysis with education level, ethnicity, blood transfusion, and occupation. HDV antibodies (HDVAb) were found in 14.7% of pregnant women. In patients, HBsAg was found less frequently in females than in males. Again in multivariate analysis, exposure to HBV was significantly correlated with gender (males), and HDVAb positivity with age and gender. The HBV DNA viral load was >3 log IU/ml in only 10.1% of pregnant women and in 17.3% of patients. HDV-RNA was detectable in 21 (67.7%) of the 31 patients positive for HDVAb, and in 11 of the 16 pregnant women positive for HDVAb (68.8%). The most frequent HBV genotypes were: HBV/D, 53%; HBV/E, 35%; and HBV/A, 12%. Sub-genotyping revealed HBV/D1,/D7, and the recently described/D8. HDV genotypes were: HDV-1, 90.3% and HDV-5, 9.7%. This study confirms the high prevalence of HBV and HDV infections in Mauritania and demonstrates the high genetic diversity of HBV in this country. J. Med. Virol. 84: 1186–1198, 2012. © 2012 Wiley Periodicals, Inc.

Clinical evaluation of multiplex real-time PCR panels for rapid detection of respiratory viral infections.

Abstract: Respiratory viral infections are one of the leading causes of morbidity and mortality, particularly in children, the elderly and immunocompromised persons. Rapid identification of viral etiology is critical in ruling out non-viral infections, initiating antiviral treatment and limiting the spread of the infection. Multiplex assays of more than one viral gene target in a single tube have the advantage of rapid screening of a large number of potential viral pathogens in a short time. A multiplex real-time PCR assay was used in this study for detection of respiratory RNA and DNA viral infections in 728 specimens received from 585 adult and pediatric patients comprised of symptomatic and asymptomatic organ transplant recipients and non-recipients for diagnosis of respiratory illnesses and for routine clinical monitoring. Multiplex PCR was more sensitive than the multiplex immunofluoresence culture assay (R-mix) and also detected additional respiratory viruses that were not covered by the R-mix panel. The number of respiratory viruses detected in symptomatic patients was significantly higher than asymptomatic patients in both adult and pediatric patients. Herpesviral infections were the predominant cause of lower respiratory tract infection in the organ transplant recipients, whereas respiratory syncytial virus was the most common pathogen in non-transplant patients particularly children. Multiplex real-time PCR for detection of respiratory viruses has the potential for rapid identification of viral pathogens. In this era of emerging viral infections, addition of newer viral targets to the multiplex PCR panels will be beneficial in determining both patient management and public health epidemiology.

Molecular detection and epidemiology of astrovirus, bocavirus, and sapovirus in Italian children admitted to hospital with acute gastroenteritis, 2008–2009

Abstract: Although a number of enteric viruses have been identified in children with acute gastroenteritis, the majority of cases of gastroenteritis remain undiagnosed. In order to provide more insights into the epidemiology of enteric viruses that are not included usually in routine diagnostic tests, cases of childhood sporadic gastroenteritis of unknown etiology requiring hospital admission in Parma, Italy, during 2008–2009, were screened for astrovirus (AstV), sapovirus (SaV), and bocavirus (BoV). The stools of 712 children, negative for rotavirus, norovirus, adenovirus, enterovirus, and reovirus, were examined by PCR or RT-PCR for AstV, BoV, and SaV. The prevalence of AstV, BoV, and SaV in the patients examined was 2.1%, 3.2%, 2.4%, respectively, with the viruses being detected mostly in children <3 years of age. AstV strains were characterized by sequencing as types 1, 2, and 4, with a AstV-1 peak occurring in the 2008 fall–winter season. BoV strains were characterized as types 1, 2, and 3, with BoV-3 circulating more frequently in the 2008 autumn and winter season and BoV-2 during March–April 2009. The most common SaVs were GI.2 and GII.1 while GIV and GV SaVs were detected sporadically. Overall, AstV, BoV, and SaV infections accounted for 7.7% of the sporadic cases of acute gastroenteritis with unknown etiology selected for the study. Different virus types and lineages were found to circulate and temporal peaks of virus activity were also demonstrated, suggesting either small clusters of infections or small outbreaks or epidemics in local population. J. Med. Virol. 84:643–650, 2012. © 2011 Wiley Periodicals, Inc.

Clinical and molecular epidemiology of norovirus infection in childhood diarrhea in China

Abstract: A prospective investigation was carried out among pediatric outpatients and inpatients with acute non-dysenteric diarrhea between August, 2008 and July, 2009 in Shanghai, Hangzhou, Chongqing, and Tianjin, China One step real-time RT-PCR was used for detection of norovirus (NoV) genogroups I and II (GI, GII) The NoV genotypes were classified based on partial capsid sequences Rotavirus (RV) was detected in parallel Among 4,123 fecal samples from outpatients, 1,067 (259%) were NoV-positive, of which 1,051 (985%) belonged to GII and 1,309 (317%) were RV-positive In the inpatient group (n = 317), 256% were NoV-positive and 416% were RV-positive Four hundred and fifty-one out of 1,067 NoV-positive strains were sequenced and genotyped and 6 typed strains were GI (3 GI3, 2 GI5, 1 GI4) and 445 typed strains were GII GII strains clustered into nine genotypes including GII4 2006b (692%), the only GII4 variant identified in this study, followed by GII3 (238%), GII6 (36%), GII12 (13%), GII2 (09%), GII13 (04%), GII14 (02%), GII7 (02%), and GII16 (02%) A peak of NoV infections was observed during the cold season in Tianjin, while NoV activity was higher between late summer and autumn and lower during winter in Shanghai, Hangzhou, and Chongqing NoV is a common causative agent of childhood diarrhea in China and the seasons of NoV-associated diarrhea varies between regions The results show that NoV GII4 2006b was the predominant strain circulating in China between 2008 and 2009 J Med Virol 84:145–151, 2011 © 2011 Wiley Periodicals, Inc

Increase of GII.2 norovirus infections during the 2009–2010 season in Osaka City, Japan

Abstract: During the 2009–2010 season, a significant numerical increase of genotype GII.2 norovirus (NoV)-associated outbreaks was observed in Osaka City, Japan. The most common genotype in that season was GII.2 (44.6%), followed by GII.4 (39.2%). Mostly, GII.2 strains were associated with outbreaks in children and with person-to-person contact. The National Infectious Disease Surveillance Center reported that GII.2 NoV infections were widespread in Japan in that season. Comparative phylogenetic analysis of RNA-dependent RNA polymerase (RdRp) and capsid sequences revealed that this GII.2 epidemic resulted from two genetic strains. The first, GII.2p2 strains, had an identical genotype in the RdRp and capsid genes. GII.2p2 strains in the 2009–2010 season were a different genetic cluster from the strains of spring 2004, the previous epidemic of GII.2 NoV, but showed no unique amino acid change. The second, GII.2 chimera virus (GII.2p16), had GII.16 RdRp and GII.2 capsid genotypes, suggesting prior recombination at the junction of ORF1 and ORF2. GII.2p16 strains had four significant amino acid changes in the P2 subdomain, suggesting antigenic changes. Before the 2009–2010 season, GII.2 chimera viruses had been observed only sporadically. This spreading of GII.2p16 strains in the 2009–2010 season might be the first epidemic of GII.2 chimera virus. This study revealed that the NoV epidemic in the 2009–2010 season differed considerably from the prior season, when GII.4 was predominant. Furthermore, GII.2 strains persisted in human populations by drastic recombination and gradual accumulation of mutations, indicating a prevalent pattern of non-GII.4 genotypes with genetic evolution. J. Med. Virol. 84:517–525, 2012. © 2011 Wiley Periodicals, Inc.

Tumor-specific and gender-specific pre-vaccination distribution of human papillomavirus types 6 and 11 in anogenital warts and laryngeal papillomas: a study on 574 tissue specimens.

Abstract: Anogenital warts and laryngeal papillomas are two most important benign tumors etiologically linked with HPV. In the study, which included both the largest number of laryngeal papilloma tissue specimens (152 specimens from 152 patients) to date and the largest number of prospectively collected and histologically confirmed tissue specimens of anogenital warts obtained from both genders (422 specimens from 315 patients), HPV DNA was detected in 413/422 (97.9%) of anogenital warts and 139/152 (91.4%) of laryngeal papillomas. HPV-6 and/or HPV-11 were detected in 291/315 (92.4%) patients with anogenital warts and in 138/152 (90.8%) patients with laryngeal papillomas, indicating that the great majority of both tumors could be prevented with prophylactic quadrivalent vaccine. The HPV-6 gender-specific distribution in both anogenital warts and laryngeal papillomas was not statistically significant. In contrast, HPV-11 was found almost three times more often in males than in females with anogenital warts (16.5% vs. 6.3%; P = 0.008), with a gender neutral HPV-11 type distribution in laryngeal papillomas. The overall HPV DNA prevalence in anogenital warts was significantly different from that in laryngeal papillomas (97.1% vs. 91.4%; P = 0.01). In the first comparison of the HPV-6/HPV-11 type-specific distribution between patients suffering from anogenital warts and laryngeal papillomas with the same geographic and ethnic background, a significant imbalance in tumor-specific distribution of HPV-6 and HPV-11 was identified: HPV-6 was statistically more often present in anogenital warts than in laryngeal papillomas (79.0% vs. 59.2%; P = 0.000013), whereas HPV-11 was statistically more frequent in laryngeal papillomas than in anogenital warts (28.9% vs. 12.4%; P = 0.00003). J. Med. Virol. 84: 1233–1241, 2012. © 2012 Wiley Periodicals, Inc.

Molecular Epidemiology of Human Rhinovirus Infections in Kilifi, Coastal Kenya

Abstract: This study reports pediatric surveillance over 3 years for human rhinovirus (HRV) at the District Hospital of Kilifi, coastal Kenya. Nasopharyngeal samples were collected from children presenting at outpatient clinic with no signs of acute respiratory infection, or with signs of upper respiratory tract infection, and from children admitted to the hospital with lower respiratory tract infection. Samples were screened by real-time reverse transcriptase polymerase chain reaction (real-time RT-PCR) and classified further to species by nucleotide sequencing of the VP4/VP2 junction. Of 441 HRV positives by real-time RT-PCR, 332 were classified to species, with 47% (155) being HRV-A, 5% (18) HRV-B, and 48% (159) HRV-C. There was no clear seasonal pattern of occurrence for any species. The species were present in similar proportions in the inpatient and outpatient sample sets, and no significant association between species distribution and the severity of lower respiratory tract infection in the inpatients could be determined. HRV sequence analysis revealed multiple but separate clusters in circulation particularly for HRV-A and HRV-C. Most HRV-C clusters were distinct from reference sequences downloaded from GenBank. In contrast, most HRV-A and HRV-B sequences clustered with either known serotypes or strains from elsewhere within Africa and other regions of the world. This first molecular epidemiological study of HRV in the region defines species distribution in accord with reports from elsewhere in the world, shows considerable strain diversity and does not identify an association between any species and disease severity.

Utility of IgM ELISA, TaqMan real-time PCR, reverse transcription PCR, and RT-LAMP assay for the diagnosis of Chikungunya fever.

Abstract: Chikungunya fever a re-emerging infection with expanding geographical boundaries, can mimic symptoms of other infections like dengue, malaria which makes the definitive diagnosis of the infection important. The present study compares the utility of four laboratory diagnostic methods viz. IgM capture ELISA, an in house reverse transcription PCR for the diagnosis of Chikungunya fever, TaqMan real-time PCR, and a one step reverse transcription-loop mediated isothermal amplification assay (RT-LAMP). Out of the 70 serum samples tested, 29 (41%) were positive for Chikungunya IgM antibody by ELISA and 50 (71%) samples were positive by one of the three molecular assays. CHIKV specific nucleic acid was detected in 33/70 (47%) by reverse transcription PCR, 46/70 (66%) by TaqMan real-time PCR, and 43/70 (62%) by RT-LAMP assay. A majority of the samples (62/70; 89%) were positive by at least one of the four assays used in the study. The molecular assays were more sensitive for diagnosis in the early stages of illness (2-5 days post onset) when antibodies were not detectable. In the later stages of illness, the IgM ELISA is a more sensitive diagnostic test. In conclusion we recommend that the IgM ELISA be used as an initial screening test followed one of the molecular assays in samples that are collected in the early phase of illness and negative for CHIKV IgM antibodies. Such as approach would enable rapid confirmation of the diagnosis and implementation of public health measures especially during outbreaks.

Genetic characterization of enterovirus 71 isolated from patients with severe disease by comparative analysis of complete genomes

Abstract: Enterovirus 71 (EV71) which causes mild illness in children is also associated with severe neurological complications. This study analyzed the complete genomes of EV71 strains derived from mild and severe diseases in order to determine whether the differences of EV71 genomes were responsible for different clinical presentations. Compared to complete genomes of EV71 strains derived from mild cases (less virulent strains), nucleotide differences in EV71 strains isolated from severe cases (more virulent strains) were observed primarily in the internal ribosomal entry site (IRES) of the 5'-untranslated region (UTR), which is vital for the cap-independent translation of viral proteins. In the protein-coding region, an E-Q substitution at amino acid position 145 of structural protein VP1 that occurred in more than one of more virulent strains was observed. This site is known to be related functionally to receptor binding and virulence in mice. Overall, strains (Group III) isolated from patients with fatal or severe sequelae outcomes had greater sequence substitutions in the 5'-UTR and/or protein-coding region and exhibited a relatively low-average homology to less virulent strains across the entire genome, indicating the possibility of significant genomic diversity in the most virulent EV71 strains. Further studies of EV71 pathogenesis should examine the significance of genomic diversity and the effects of multiple mutations in a viral population.

Specific increase in local IL-17 production during recovery from primary RSV bronchiolitis.

Abstract: Although Respiratory syncytial virus (RSV) bronchiolitis is the most important cause of hospital admission for infants during the winter season, the pathogenesis is largely unknown. Interleukin-17 (IL-17) concentrations were studied in nasopharyngeal aspirates from 21 non-ventilated and 17 ventilated infants admitted to hospital with RSV bronchiolitis at time of admission and discharge from the hospital. On admission, nasopharyngeal concentrations of most cytokines and chemokines were lower in non-ventilated infants than in ventilated infants, reaching statistical significance for Eotaxin, IL-1a, and IL-6. During course of disease, nasopharyngeal concentrations of most cytokines and chemokines decreased, reaching statistical significance for IL-6 and IP-10. However, nasopharyngeal IL-17 concentrations were higher at discharge than at admission in children with non-ventilated RSV disease (209101?pg/ml, P?=?0.008), a response pattern not observed in ventilated RSV patients nor for other cytokines or chemokines. It is speculated that local IL-17 production may be involved during convalescence from RSV bronchiolitis in non-ventilated patients by facilitating innate and adaptive antiviral immune responses. The role of IL-17 in the pathogenesis of RSV bronchiolitis is to be explored further. J. Med. Virol. 84: 10841088, 2012. (C) 2012 Wiley Periodicals, Inc.

High detection frequency and viral loads of human rhinovirus species A to C in fecal samples; diagnostic and clinical implications.

Abstract: Human rhinoviruses (HRVs) can be divided into three species; HRV-A to HRV-C Up to 148 different HRV (sero)types have been identified to date Because of sequence similarity between 5'-NCR of HRVs and enteroviruses (EVs), it is problematic to design EV-specific RT-PCR assays The aims of this study were to assess the rate of false-detection of different rhinoviruses by EV RT-PCR, and to evaluate the diagnostic and clinical significance of such cross-reactivity In vitro RNA transcripts of HRV A-C created from cDNA templates were quantified spectrophotometrically Six hundred twenty-one stool samples screened as part of routine diagnostic for EV, 17 EV-positive stool samples referred for typing, 288 stool samples submitted for gastroenteritis investigations, and 1,500 CSF samples were included in the study EV-specific RT-PCR detected RNA transcripts of HRV-A1b, HRV-B14, and HRV-Crpat18 but with 10-1,000 reduced sensitivity compared to EV transcripts Screening fecal samples by EV RT-PCR identified 13 positive samples identified subsequently as rhinoviruses; a further 26 HRV-positive samples were identified by nested HRV RT-PCR All individuals were hospitalized and presented mostly with diarrhea A total of 26 HRV types were identified (HRV-A: 46%; HRV-B: 13%; HRV-C: 41%) Results confirm that EV-specific RT-PCR can detect HRVs, and at a practical level, identify potential problems of interpretation if fecal samples are used for surrogate screening in cases of suspected viral meningitis High detection frequencies (10%) and viral loads in stool samples provide evidence for enteric replication of HRV, and its association with enteric disease requires further etiological studies

Human bocavirus infection diagnosed serologically among children admitted to hospital with community-acquired pneumonia in a tropical region.

Abstract: Human bocavirus (HBoV) is a human virus associated with respiratory disease in children. Limited information is available on acute infection with HBoV among children admitted to hospital with community-acquired pneumonia in tropical regions and the current diagnosis is inadequate. The aims were to diagnose and describe acute HBoV infections among children hospitalized for community-acquired pneumonia. In Salvador, Brazil, 277 children with community-acquired pneumonia were prospectively enrolled. Paired serum samples were tested by IgG, IgM, and IgG-avidity enzyme immunoassays (EIAs) using recombinant HBoV VP2. HBoV DNA was detected in nasopharyngeal aspirates and serum by a quantitative polymerase-chain reaction (PCR). HBoV DNA was detected in nasopharyngeal aspirates of 62/268 (23%) children and 156/273 (57%) were seropositive. Acute primary HBoV infection was reliably diagnosed (bearing at least two acute markers: Positive IgM, a fourfold increase/conversion of IgG, low IgG avidity or viremia) in 21 (8%) of 273 patients, 90% of 20 had HBoV DNA in nasopharyngeal aspirates, 83% with a high DNA load. The median age of infection with HBoV was 16 months, range 5–36. Community-acquired pneumonia was confirmed radiographically in 85% of 20 patients with acute HBoV infection diagnosed serologically. HBoV DNA was found in nasopharyngeal aspirates of 42/246(17%) children without an acute primary HBoV infection and available nasopharyngeal aspirate. Four children with HBoV secondary immune responses were detected, lacking both IgM and viremia. HBoV infection was diagnosed accurately in children aged 5–36 months with community-acquired pneumonia confirmed radiographically. PCR of nasopharyngeal aspirates is not a reliable marker of acute HBoV infection. J. Med. Virol. 84:253–258, 2012. © 2011 Wiley Periodicals, Inc.

Detection of non-polio enteroviruses from 17 years of virological surveillance of acute flaccid paralysis in the Philippines.

Abstract: Acute flaccid paralysis (AFP) surveillance has been conducted as part of the World Health Organization (WHO) strategy on poliomyelitis eradication. Aside from poliovirus, which is the target pathogen, isolation, and identification of non-polio enteroviruses (NPEVs) is also done by neutralization test using pools of antisera which can only identify limited number of NPEVs. In the Philippines, despite the significant number of isolated NPEVs, no information is available with regard to its occurrence, diversity, and pattern of circulation. In this study, a total of 790 NPEVs isolated from stool samples submitted to the National Reference Laboratory from 1992 to 2008 were analyzed; neutralization test was able to type 55% (442) of the isolates. Of the remaining 356 isolates, which were untyped by using neutralization test, 348 isolates were analyzed further by RT-PCR targeting the VP1 gene. A total of 47 serotypes of NPEV strains were identified using neutralization test and molecular typing, including 28 serotypes of human enterovirus B (HEV-B), 12 serotypes of HEV-A, and 7 of HEV-C. The HEV-B group (625/790; 79%) constituted the largest proportion of isolates, followed by HEV-C (108/790; 13.7%), HEV-A (57/790; 7.2%), and no HEV-D. Coxsackievirus (CV) B, echovirus (E)6, E11, and E13 were the most frequent isolates. E6, E11, E13, E14, E25, E30, E33, CVA20, and CVA24 were considered as endemic strains, some NPEVs recurred and few serotypes existed only for 1–3 years during the study period. Despite some limitations in this study, plural NPEVs with multiple patterns of circulation in the Philippines for 17 years were identified. J. Med. Virol. 84:624–631, 2012. © 2011 Wiley Periodicals, Inc.

Human parechovirus infections in patients admitted to hospital in Northern Italy, 2008–2010

Abstract: Human parechoviruses (HPeVs) infection is associated with a wide range of clinical syndromes such as respiratory, gastrointestinal, neurologic diseases, and neonatal sepsis-like illness. The main objective of this study was to investigate the epidemiology of HPeVs infection in hospitalized patients in a period of 2 years. Respiratory samples from 3,525 patients with respiratory syndrome, cerebrospinal fluid (CSF) from 340 patients with neurologic syndrome as well as CSF and plasma samples from five neonatal patients with sepsis-like illness collected from October 2008 to 2010 were tested retrospectively using HPeV-specific real-time RT-PCR. Phylogenetic analysis of VP3/VP1 region was performed on the positive samples. Fourteen out of 3,525 (0.4%) patients with respiratory syndrome and five out of five patients with sepsis-like illness were positive for HPeV. In 3/5 patients with sepsis-like illness multiple samples (e.g., stool, plasma, CSF, or respiratory samples) were available, and HPeV was found in all specimens. In contrast, no positive CSF was detected among the 340 patients with neurologic syndromes. Eleven patients (57.9%) were infected with HPeV1 strain, 7 (36.8%) with HPeV3, and 1 (5.3%) with HPeV6 strains. Ten of the 14 HPeV patients with respiratory syndrome were co-infected with other respiratory viruses (eight with rhinovirus and two with coronavirus OC43). All five patients with sepsis-like illness were less than 1 month of age and were infected with HPeV3. Although not circulating at high frequency and unlikely to cause respiratory syndrome, HPeV was associated with severe clinical syndromes in a minority of newborns.

Toll-like receptor 3 polymorphism and its association with hepatitis B virus infection in Saudi Arabian patients.

Abstract: Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi ArabiaHepatitis B virus (HBV) is the major causativeagent of chronic liver complications includingcirrhosis and hepatocellular carcinoma (HCC).Individuals infected with HBV show a widespectrum of disease manifestations rangingfrom asymptomatic carriers to HCC. TLR3 ispart of the innate immune system thatrecognizes double-stranded RNA (dsRNA) andprovides early immune response to exogenousantigens. The genetic polymorphisms such assingle nucleotide polymorphisms (SNPs) in theTLR3 could be considered as factors for thesusceptibility to viral pathogens including HBV.Due to lack of knowledge on the role of TLR3polymorphisms in HBV infection, this studyinvestigated the distribution of nine SNPs inthe TLR3 gene and its association with SaudiArabian patients infected with HBV. A total of707 patients and 600 uninfected controls wereexamined for different parameters includingthe nine SNPs (rs5743311, rs5743312, rs1879026,rs5743313, rs5743314, rs5743315, rs111611328,rs78726532 and a newly identified SNP locatedat position 184322913 of chr4). The associationanalysis confirmed that only one SNP,rs1879026 (G/T), showed a significant difference(P ¼ 0.0480; OR ¼ 0.809, 95% CI ¼ 0.655–0.999)in the distribution between HBV carriers anduninfected controls. While, the rest of the SNPsshowed no significant association with regardsto HBV infection or in the progression tocirrhosis of the liver and HCC. Furthermore,haplotype analysis revealed that one haplotypeGCGA (rs1879026, rs5743313, rs5743314, andrs5743315, respectively), was associated signifi-cantly with HBV infection in this population.These findings indicate that genetic variationsin the TLR3 gene could affect the outcome ofHBV infection among Saudis. J. Med. Virol. 84:1353–1359, 2012.

Emergence of a new norovirus GII.6 variant in Japan, 2008-2009.

Abstract: Norovirus (NoV) is recognized as one of the most common causative agents of diarrhea disease in young children. A total of 187 fecal specimens collected from non-hospitalized children with acute gastroenteritis in Shizuoka, Japan during July 2008 to June 2009 were investigated for the presence of diarrhea viruses by a multiplex RT-PCR. Diarrhea viruses were overall detected in 158 of 187 (84.5%). Of the viruses detected, NoV was the most prevalent (55.6%). Most of the NoV sequences belonged to GII.4 (53.8%). NoV GII.6 emerged as the second most common strain (40.4%). The full-length capsid sequences of five representative Shizuoka GII.6 strains were compared with all 12 GII.6 strains available in GenBank database between 1990 and 2009. At least three distinct GII.6 subclusters (a–c) appeared in different parts of the world. Shizuoka GII.6 strains formed their own subcluster c, distinct from other complete GII.6 reference sequences. The Shizuoka strains had significant amino acid divergence, particularly in the P2 domain up to 10.9–17.5% and contained eight unique mutations in the P domains, compared with subcluster a and b viruses. The homology model showed that the eight mutations were predicted to be located at the surface-exposed P1 and P2 domains. The data suggest the emergence of a new NoV GII.6 variant in Shizuoka, with a high level of genetic variation. J. Med. Virol. 84: 1089–1096, 2012. © 2012 Wiley Periodicals, Inc.