[Show abstract][Hide abstract] ABSTRACT: Infections with cytomegalovirus (CMV) and Epstein Barr Virus (EBV) remain important in solid organ transplantation. Quantitative viral nucleic acid testing is a major advance to patient management. These assays are limited by a lack of standardization, resulting in viral load measurements that differ among clinical laboratories. The variability in viral load measurements makes interpretation of multicenter clinical trials data difficult. This study compares the current practices in CMV and EBV viral load testing at four large transplant centers participating in multicenter Clinical Trials in Organ Transplantation (CTOT/CTOTC).
[Show abstract][Hide abstract] ABSTRACT: Background: Neonatal enterovirus (EV) infections have high morbidity & mortality. Antiviral therapy is not currently available. Pleconaril is an oral capsid binder with activity against EVs.
Methods: Neonates with suspected EV sepsis (hepatitis, coagulopathy, or myocarditis) were randomized 2:1 to receive oral pleconaril or placebo x 7 days. Specimens (oropharynx, rectum, urine, serum) for viral culture & polymerase chain reaction (PCR), pharmacokinetic analysis, & safety evaluations were obtained over 14 days & clinical assessments were performed over 24 months.
Results: 61 subjects were enrolled (43 treatment, 18 placebo), of whom 43 were confirmed EV-infected by culture or PCR (31 treatment, 12 placebo). Baseline characteristics were similar between EV-infected groups; median (range) age at illness onset was 4.5 (1-15) & 5.0 (1-10) days, respectively. Low culture yields precluded demonstrating a difference in the primary endpoint, day 5 oropharyngeal culture positivity (25% positive on Day 1 & 0% on Day 5 in the treatment group v. 30% on Day 1 & 0% on Day 5 in the placebo group). However, subjects in the treatment group became culture-negative from all anatomic sites combined faster than subjects in the placebo group (Fig 1; median 4.0 v. 7.0 days, p = 0.08) & fewer subjects in the treatment group remained PCR-positive from the oropharynx when last sampled (83% positive on Day 1 & 23% positive at a median of 14 days in the treatment group v. 100% positive on Day 1 & 58% positive at a median of 14 days in the placebo group, p = 0.02). By intent to treat, 10/43 (23%) of all subjects in the treatment group & 8/18 (44%) in the placebo group died (Fig 2; p = 0.02 for 2 month survival difference). Among EV-confirmed subjects, 7/31 (23%) in the treatment group died v. 5/12 (42%) in the placebo group (Fig 3; p = 0.26). Pleconaril concentrations exceeded the IC90 after the first treatment day, but 41% of subjects did not achieve this targetduring the 1st treatment day. 1 subject in the treatment group & 3 in the placebo group had treatment-related adverse events.
Conclusion: Shorter times to culture & PCR negativity & suggestion of greater survival among pleconaril recipients support potential efficacy & warrant further evaluation.
SEQ Figure * ARABIC 1. Culture positive
SEQ Figure * ARABIC 2. Survival, all subjects
SEQ Figure * ARABIC 3. Survival, EV-confirmed
IDWeek 2014 Meeting of the Infectious Diseases Society of America; 10/2014
[Show abstract][Hide abstract] ABSTRACT: Background
The Human Microbiome Project (HMP) was undertaken with the goal of defining microbial communities in and on the bodies of healthy individuals using high-throughput, metagenomic sequencing analysis. The viruses present in these microbial communities, the `human virome¿, are an important aspect of the human microbiome that is particularly understudied in the absence of overt disease. We analyzed eukaryotic double-stranded DNA (dsDNA) viruses, together with dsDNA replicative intermediates of single-stranded DNA viruses, in metagenomic sequence data generated by the HMP. 706 samples from 102 subjects were studied, with each subject sampled at up to five major body habitats: nose, skin, mouth, vagina, and stool. Fifty-one individuals had samples taken at two or three time points 30 to 359 days apart from at least one of the body habitats.ResultsWe detected an average of 5.5 viral genera in each individual. At least 1 virus was detected in 92% of the individuals sampled. These viruses included herpesviruses, papillomaviruses, polyomaviruses, adenoviruses, anelloviruses, parvoviruses, and circoviruses. Each individual had a distinct viral profile, demonstrating the high interpersonal diversity of the virome. Some components of the virome were stable over time.Conclusions
This study is the first to use high-throughput DNA sequencing to describe the diversity of eukaryotic dsDNA viruses in a large cohort of normal individuals who were sampled at multiple body sites. Our results show that the human virome is a complex component of the microbial flora. Some viruses establish long-term infections that may be associated with increased risk or possibly with protection from disease. A better understanding of the composition and dynamics of the virome may hold important keys to human health.
[Show abstract][Hide abstract] ABSTRACT: Background
Human Herpes Virus type-6 (HHV-6) is a significant cause of the febrile illness roseola infantum in young children. Infection with HHV-6 typically causes a self-limited febrile illness, but occasionally is associated with central nervous system manifestations, including febrile seizures and encephalitis. Host factors associated with severe manifestations of HHV-6-associated neurologic disease remain poorly characterized.
We report two cases of previously healthy young boys with HHV-6-associated encephalitis who developed a progressive, and ultimately fatal, encephalopathy with refractory movement disorder concurrent with acquisition of acute HHV-6 infection. Both children were treated with the antiviral ganciclovir without improvement in their neurologic symptoms, although quantitative HHV-6 PCR of CSF and/or blood confirmed a decline in viral load with treatment. The clinical course in both cases was most consistent with Alpers-Huttenlocher syndrome, given the intractable seizures, developmental regression and, ultimately, death due to liver and renal failure. In support of this, post-mortem analysis identified both children to be compound heterozygotes for mutations in the mitochondrial polymerase γ gene, POLG.
POLG mutations are associated with Alpers-Huttenlocher syndrome, however no prior studies have examined the role of acute HHV-6 infection in these patients presenting with severe neurologic disease. It is possible the POLG mutation phenotype was unmasked and/or exacerbated by HHV-6 infection in these two patients, potentially contributing to a more rapid clinical deterioration. This report provides new insight into a previously unrecognized association between POLG mutations and poor neurologic outcome following HHV-6 infection.
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to identify best practice for the detection of Shiga toxin-producing Escherichia coli (STEC) in children with diarrheal illness treated at a tertiary care center: sorbitol-MacConkey (SMAC) agar culture, enzyme immunoassay (EIA) for Shiga toxin, or the use of both methodologies simultaneously. STEC were detected in 100 of 14,997 stool specimens submitted for enteric culture (0.7%), of which 65 were E. coli O157. Among E. coli O157, 57 (88%) were identified by both SMAC and EIA, 6 (9%) by SMAC alone, and 2 (3%) by EIA alone. Of the 62 individuals with diarrheal hemolytic uremic syndrome (HUS) seen at our institution during the study period, 16 (26%) had STEC isolated from culture at our institution and 15 (24%) had STEC isolated at other institutions. No STEC was recovered in 31 cases (50%). Of the HUS cases in which an STEC was isolated, 28 (90%) were attributable to E. coli O157 and 3 (10%) were attributable to non-O157 STEC. Consistent with previous studies, we have determined that a subset of E. coli O157 STEC will not be detected if an agar based method is excluded from the enteric culture workup; this has both clinical and public health implications. Best practice would be concomitant use of an agar based method and a Shiga toxin EIA, but a Shiga toxin EIA should not be considered to be an adequate stand-alone test for detection of E. coli O157 in clinical samples.
Journal of Clinical Microbiology 07/2014; · 4.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A current controversy is whether patients with sepsis progress to an immunosuppressed state. We hypothesized that reactivation of latent viruses occurred with prolonged sepsis thereby providing evidence of clinically-relevant immunosuppression and potentially providing a means to serially-monitor patients' immune status. Secondly, if viral loads are markedly elevated, they may contribute to morbidity and mortality. This study determined if reactivation of herpesviruses, polyomaviruses, and the anellovirus TTV occurred in sepsis and correlated with severity. Serial whole blood and plasma samples from 560 critically-ill septic, 161 critically-ill non-septic, and 164 healthy age-matched patients were analyzed by quantitative-polymerase-chain-reaction for cytomegalovirus (CMV), Epstein-Barr (EBV), herpes-simplex (HSV), human herpes virus-6 (HHV-6), and TTV. Polyomaviruses BK and JC were quantitated in urine. Detectable virus was analyzed with respect to secondary fungal and opportunistic bacterial infections, ICU duration, severity of illness, and survival. Patients with protracted sepsis had markedly increased frequency of detectable virus. Cumulative viral DNA detection rates in blood were: CMV (24.2%), EBV (53.2%), HSV (14.1%), HHV-6 (10.4%), and TTV (77.5%). 42.7% of septic patients had presence of two or more viruses. The 50% detection rate for herpesviruses was 5-8 days after sepsis onset. A small subgroup of septic patients had markedly elevated viral loads (>104-106 DNA copies/ml blood) for CMV, EBV, and HSV. Excluding TTV, DNAemia was uncommon in critically-ill non-septic patients and in age-matched healthy controls. Compared to septic patients without DNAemia, septic patients with viremia had increased fungal and opportunistic bacterial infections. Patients with detectable CMV in plasma had higher 90-day mortality compared to CMV-negative patients; p<0.05. Reactivation of latent viruses is common with prolonged sepsis, with frequencies similar to those occurring in transplant patients on immunosuppressive therapy and consistent with development of an immunosuppressive state. Whether reactivated latent viruses contribute to morbidity and mortality in sepsis remains unknown.
PLoS ONE 06/2014; 9(6):e98819. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Multiplex molecular assays now make it possible for clinical laboratories to detect human coronaviruses (HCoV). We investigated the clinical characteristics of HCoV-OC43 and HCoV-NL63 in patients <5 years of age during a recent coronavirus season.
Respiratory viruses were detected using a multiplex molecular assay at St. Louis Children`s Hospital starting in November 2012. We analyzed demographic and clinical data from all patients <5 years old with solo detection of HCoV-OC43 (n=52) and HCoV-NL63 (n=44) and for comparison, samples of children with respiratory syncytial virus (RSV), parainfluenza virus (PIV), and picornaviruses.
During the study period, HCoV-OC43 (4%) was the 5 and HCoV-NL63 the 8 (2%) most common respiratory virus. Coinfections were detected in 35% and 38% of children with HCoV-OC43 and HCoV-NL63, respectively. Croup was more common with HCoV-NL63 (30%) than with HCoV-OC43 (2%). Lower respiratory tract infection occurred in 33% of children with HCoV-OC43 and 25% of children with HCoV-NL63. Severe illness was less common in HCoV-NL63, HCoV-OC43, and PIV (14%, each) compared with RSV (30%) and picornaviruses (26%) (p=0.055 for HCoVs combined compared with the other respiratory viruses), and occurred mainly in those with underlying medical conditions.
Infections caused by HCoV-OC43 and HCoV-NL63 are common and include some with lower respiratory tract involvement and severe disease, especially in children with underlying medical conditions. Overall, a substantial burden of disease associated with both HCoV-OC43 and HCoV-NL63 was observed for hospitalized children younger than 5 years old.
The Pediatric Infectious Disease Journal 02/2014; · 3.14 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Viral infections are common causes of fever without an apparent source in young children. Despite absence of bacterial infection, many febrile children are treated with antibiotics. Virus and bacteria interact with different pattern recognition receptors in circulating blood leukocytes, triggering specific host transcriptional programs mediating immune response. Therefore, unique transcriptional signatures may be defined that discriminate viral from bacterial causes of fever without an apparent source. Gene expression microarray analyses were conducted on blood samples from 30 febrile children positive for adenovirus, human herpesvirus 6, or enterovirus infection or with acute bacterial infection and 22 afebrile controls. Blood leukocyte transcriptional profiles clearly distinguished virus-positive febrile children from both virus-negative afebrile controls and afebrile children with the same viruses present in the febrile children. Virus-specific gene expression profiles could be defined. The IFN signaling pathway was uniquely activated in febrile children with viral infection, whereas the integrin signaling pathway was uniquely activated in children with bacterial infection. Transcriptional profiles classified febrile children with viral or bacterial infection with better accuracy than white blood cell count in the blood. Similarly accurate classification was shown with data from an independent study using different microarray platforms. Our results support the paradigm of using host response to define the etiology of childhood infections. This approach could be an important supplement to highly sensitive tests that detect the presence of a possible pathogen but do not address its pathogenic role in the patient being evaluated.
Proceedings of the National Academy of Sciences 07/2013; · 9.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: High throughput, deep sequencing assays are powerful tools for gaining insights into virus-host interactions. Sequencing assays can discover novel viruses and describe the genomes of novel and known viruses. Genomic information can predict viral proteins that can be characterized, describe important genes in the host that control infections, and evaluate gene expression of viruses and hosts during infection. Sequencing can also describe variation and evolution of viruses during replication and transmission. This review recounts some of the major advances in the studies of virus-host interactions from the last two years, and discusses the uses of sequencing technologies relating to these studies.
Current opinion in microbiology 05/2013; · 7.87 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The Anelloviridae family consists of non-enveloped, circular, single-stranded DNA viruses. Three genera of anellovirus are known to infect humans, named TTV, TTMDV, and TTMV. Although anelloviruses were initially thought to cause non-A-G viral hepatitis, continued research has shown no definitive associations between anellovirus and human disease to date. Using high-throughput sequencing, we investigated the association between anelloviruses and fever in pediatric patients 2-36 months of age. We determined that although anelloviruses were present in a large number of specimens from both febrile and afebrile patients, they were more prevalent in the plasma and nasopharyngeal (NP) specimens of febrile patients compared to afebrile controls. Using PCR to detect each of the three species of anellovirus that infect humans, we found that anellovirus species TTV and TTMDV were more prevalent in the plasma and NP specimens of febrile patients compared to afebrile controls. This was not the case for species TTMV which was found in similar percentages of febrile and afebrile patient specimens. Analysis of patient age showed that the percentage of plasma and NP specimens containing anellovirus increased with age until patients were 19-24 months of age, after which the percentage of anellovirus positive patient specimens dropped. This trend was striking for TTV and TTMDV and very modest for TTMV in both plasma and NP specimens. Finally, as the temperature of febrile patients increased, so too did the frequency of TTV and TTMDV detection. Again, TTMV was equally present in both febrile and afebrile patient specimens. Taken together these data indicate that the human anellovirus species TTV and TTMDV are associated with fever in children, while the highly related human anellovirus TTMV has no association with fever.
PLoS ONE 11/2012; 7(11):e50937. · 3.53 Impact Factor
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[Show abstract][Hide abstract] ABSTRACT: OBJECTIVE:Fever without an apparent source is common in young children. Currently in the United States, serious bacterial infection is unusual. Our objective was to determine specific viruses that might be responsible.METHODS:We enrolled children aged 2 to 36 months with temperature of 38°C or greater without an apparent source or with definite or probable bacterial infection being evaluated in the St Louis Children's Hospital Emergency Department and afebrile children having ambulatory surgery. Blood and nasopharyngeal swab samples were tested with an extensive battery of virus-specific polymerase chain reaction assays.RESULTS:One or more viruses were detected in 76% of 75 children with fever without an apparent source, 40% of 15 children with fever and a definite or probable bacterial infection, and 35% of 116 afebrile children (P < .001). Four viruses (adenovirus, human herpesvirus 6, enterovirus, and parechovirus) were predominant, being detected in 57% of children with fever without a source, 13% of children with fever and definite or probable bacterial infection, and 7% of afebrile children (P < .001). Thirty-four percent of 146 viral infections were detected only by polymerase chain reaction performed on blood. Fifty-one percent of children with viral infections and no evidence of bacterial infection were treated with antibiotics.CONCLUSIONS:Viral infections are frequent in children with fever without an apparent source. Testing of blood in addition to nasopharyngeal secretions expanded the range of viruses detected. Future studies should explore the utility of testing for the implicated viruses. Better recognition of viruses that cause undifferentiated fever in young children may help limit unnecessary antibiotic use.
[Show abstract][Hide abstract] ABSTRACT: Background: Respiratory viruses have been linked to bronchiolitis obliterans syndrome in lung transplant recipients. Prospective epidemiology of respiratory viral infections has not been reported in pediatric lung transplant recipients (PLTRs).
Methods: As part of the NIAID-sponsored Clinical Trials in Organ Transplantation in Children (CTOT-C) study of PTLRs, prospective serial nasopharyngeal (NP) and bronchoalveolar lavage (BAL) specimens were interrogated by multiplex PCR (Luminex xTAG®) that identifies 17 viruses. Rhinoviruses and enteroviruses are not differentiated from one another and are referred to here as picornavirus. Sequencing of the picornavirus 5’ non-translated region was performed on samples from patients with multiple picornavirus positive specimens.
Results: 289 NP and 229 BAL specimens from 39 patients were tested. Considering a viral infection to be detection of viral nucleic acid in NP, BAL, or both, 89 infections were detected, of which 73(82%) were picornaviruses. Other viruses detected included: coronaviruses (4); parainfluenza viruses (4); respiratory syncytial virus (3); human metapneumovirus (2); influenza viruses (2); and adenovirus (1). Of the 39 patients, 27(69%) had a least one picronavirus-positive specimen. Of 289 NP specimens, 79(27%) were positive, of which 67(85%) were picornaviruses. Of 229 BAL specimens, 25(11%) were virus-positive, of which 18(72%) were picornaviruses. Of 193 paired NP and BAL specimens, 42 were positive for picornaviruses in one or both specimens. Picornaviruses were detected in the NP only in 26(62%), in BAL only in 4(10%), and in both in 12(29%).
Of note, two patients were picornavirus positive every time they were sampled over a period of time ranging from 13 to 16 months. Sequencing of the 5’ non-translated region of 23 picornoviruses revealed them all to be rhinoviruses. Results revealed both persistent infection with one serotype as well as reinfection with different serotypes.
Conclusion: Picornavirus infections were very common in our population of PLTRs, comprising 82% of all respiratory viral infections detected. Although picornaviruses were the most commonly detected virus, the role of these infections in determining the outcome of pediatric lung transplantation is still uncertain.
IDWeek 2012 Meeting of the Infectious Diseases Society of America; 10/2012
[Show abstract][Hide abstract] ABSTRACT: Unexplained fever (UF) is a common problem in children under 3 years old. Although virus infection is suspected to be the cause of most of these fevers, a comprehensive analysis of viruses in samples from children with fever and healthy controls is important for establishing a relationship between viruses and UF. We used unbiased, deep sequencing to analyze 176 nasopharyngeal swabs (NP) and plasma samples from children with UF and afebrile controls, generating an average of 4.6 million sequences per sample. An analysis pipeline was developed to detect viral sequences, which resulted in the identification of sequences from 25 viral genera. These genera included expected pathogens, such as adenoviruses, enteroviruses, and roseoloviruses, plus viruses with unknown pathogenicity. Viruses that were unexpected in NP and plasma samples, such as the astrovirus MLB-2, were also detected. Sequencing allowed identification of virus subtype for some viruses, including roseoloviruses. Highly sensitive PCR assays detected low levels of viruses that were not detected in approximately 5 million sequences, but greater sequencing depth improved sensitivity. On average NP and plasma samples from febrile children contained 1.5- to 5-fold more viral sequences, respectively, than samples from afebrile children. Samples from febrile children contained a broader range of viral genera and contained multiple viral genera more frequently than samples from children without fever. Differences between febrile and afebrile groups were most striking in the plasma samples, where detection of viral sequence may be associated with a disseminated infection. These data indicate that virus infection is associated with UF. Further studies are important in order to establish the range of viral pathogens associated with fever and to understand of the role of viral infection in fever. Ultimately these studies may improve the medical treatment of children with UF by helping avoid antibiotic therapy for children with viral infections.
PLoS ONE 06/2012; 7(6):e27735. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The human virome is the collection of all viruses that are found in or on humans, including both eukaryotic and prokaryotic viruses. Eukaryotic viruses clearly have important effects on human health, ranging from mild, self-limited acute or chronic infections to those with serious or fatal consequences. Prokaryotic viruses can also influence human health by affecting bacterial community structure and function. Therefore, definition of the virome is an important step toward understanding how microbes affect human health and disease. We review progress in virome analysis, which has been driven by advances in high-throughput, deep sequencing technology. Highlights from these studies include the association of viruses with clinical phenotypes and description of novel viruses that may be important pathogens. Together these studies indicate that analysis of the human virome is critical as we aim to understand how microbial communities influence human health and disease. Descriptions of the human virome will stimulate future work to understand how the virome affects long-term human health, immunity, and response to coinfections. Analysis of the virome ultimately may affect the treatment of patients with a variety of clinical syndromes.
Translational research : the journal of laboratory and clinical medicine. 04/2012; 160(4):283-90.