BACKGROUNDEnteroviruses are highly diverse with a wide spectrum of genotypes and clinical manifestations. Coxsackievirus A22 (CVA22) has been detected globally from sewage surveillance; however, ...currently there is limited information on its prevalence in patients, as well as available genomic data. OBJECTIVEWe aimed to provide genomic and relative frequency data on CVA22 from a regional hospital perspective between 2013-2020. STUDY DESIGNSanger sequencing was performed on all samples with a positive enterovirus RT-qPCR result (≤Ct 32). Viral targeted sequence capture (ViroCap) and next-generation sequencing (NGS) (Illumina NextSeq 500) was used to characterize and generate near-complete CVA22 genomes for enteroviruses without genotyping results from Sanger sequencing. Epidemiological and phylogenetic analysis was performed using maximum likelihood trees on MEGA-11. RESULTSA total of twenty detections derived from fecal material from sixteen patients were observed between 2013- 2020. One transplant recipient had five different CVA22 infection episodes over five years, with phylogenetic analysis indicating possible reinfection with an additional prolonged infection (>3 weeks). Furthermore, we report the first two near-complete CVA22 sequences from Europe, which grouped with a strain previously isolated from Bangladesh in 1999. CONCLUSIONSWe show a highly diverse enterovirus genotype which causes infections annually, typically in autumn and winter, and is capable of recurrent infection in an immunocompromised patient. Furthermore, we highlight the use of NGS to complement conventional targeted Sanger sequencing.
Hepatitis B virus (HBV) genotyping has become important in epidemiological and clinical diagnoses, given the relationship between the viral genotype and the progression of disease or the appearance ...of antiviral resistance. Since genotyping by sequence and phylogenetic analyses is not convenient in the clinical setting, we evaluated InnoLipa HBV genotyping (Innogenetics, Belgium) and an HBV DNA-Chip (bioMerieux, France) prototype assay and compared their sequencing of the gold standard S gene, using a cohort of 275 individual patient samples. All but two samples, belonging to distant and individual subgroups within a single genotype, were detected by InnoLipa HBV assay. Four samples with dual infections belonging to genotypes A and G were identified only by InnoLipa HBV assay. Using an HBV DNA-Chip assay, one sample could not be amplified due to a low viral load. Four samples were identified as genotype C and two as genotype D by sequencing but were classified as genotype A (two samples) and D (two samples) and as A (one sample) and G (one sample) by the DNA-Chip assay. In conclusion, the InnoLipa HBV genotyping strip assay detected dual infections and was an easy and quick tool for genotyping, with a sensitivity of 99.3% and a specificity of 100% compared to sequence analysis. HBV DNA-Chip assay showed a sensitivity and specificity of 97.5 and 97.8%, respectively.
Torquetenovirus (TTV), a small, single stranded anellovirus, is currently being explored as a marker of immunocompetence in patients with immunological impairment and inflammatory disorders. TTV has ...an extremely high prevalence and is regarded as a part of the human virome, the replication of which is controlled by a functioning immune system. The viral load of TTV in plasma of individuals is thought to reflect the degree of immunosuppression. Measuring and quantifying this viral load is especially promising in organ transplantation, as many studies have shown a strong correlation between high TTV loads and increased risk of infection on one side, and low TTV loads and an increased risk of rejection on the other side. As clinical studies are underway, investigating if TTV viral load measurement is superior for gauging antirejection therapy compared to medication-levels, some aspects nevertheless have to be considered. In contrast with medication levels, TTV loads have to be interpreted bearing in mind that viruses have properties including transmission, tropism, genotypes and mutations. This narrative review describes the potential pitfalls of TTV measurement in the follow-up of solid organ transplant recipients and addresses the questions which remain to be answered.
•A duplex of RT-PCRs was developed and validated on the VP1 gene to differentiate the four BKV genotypes. These assays are internally controlled, precise and are specific for the main BKV genotypes, ...B19, CMV, EBV, HHV6, HHV8, HSV 1, HSV 2, JCV and VZV.•In 194 (91.5%) of 212 samples genotyping was possible with the developed RT-PCRs. In 18 (8.5%) samples genotyping was not successful due to a low viral load.•By sequence analysis, the genotype of 46 out of 55 and 2 out of 4 samples with double infection could be confirmed.•The developed RT-PCRs for the detection of BKV genotypes, proved to be rapid, cheap and more sensitive compared to sequencing.•It is possible to detect double infections.
Polyomavirus BK (BKV) may cause nephropathy in renal transplant recipients and hemorrhagic cystitis in bone marrow recipients. We developed real-time PCRs (RT-PCR) to determine easily and rapidly the different BKV genotypes (BKGT) (I–IV).
On the VP1 gene a duplex of RT-PCRs was developed and validated to differentiate the four main BKGT. 212 BKV positive samples (21 plasma, 191 urine) were tested with these specific PCRs. Of these 212 samples, 55 PCR results were additionally confirmed by sequencing a VP1 gene fragment (nucleotide 1630–1956).
For every genotype, a highly specific, precise and internally controlled assay was developed with a limit of detection of log 3 copies per ml. In 18 (8.5%) of these samples genotyping was not successful due to a low viral load. By sequence analysis, the genotype of 46 out of 55 and 2 out of 4 samples with double infection could be confirmed.
This study describes RT-PCRs for detection of the main BKGT. It proved to be rapid, cheap and sensitive compared to sequencing. Double infections can also be detected. This method will be of value to investigate the role of BKV infection in relation to the genotype.
ObjectiveCurrent evidence on the effectiveness of SARS-CoV-2 prophylaxis is inconclusive. We aimed to systematically evaluate published studies on repurposed drugs for the prevention of ...laboratory-confirmed SARS-CoV-2 infection and/or COVID-19 among healthy adults.DesignSystematic review.EligibilityQuantitative experimental and observational intervention studies that evaluated the effectiveness of repurposed drugs for the primary prevention of SARS-CoV-2 infection and/or COVID-19 disease.Data sourcePubMed and Embase (1 January 2020–28 September 2022).Risk of biasCochrane Risk of Bias 2.0 and Risk of Bias in Non-Randomised Studies of Interventions tools were applied to assess the quality of studies.Data analysisMeta-analyses for each eligible drug were performed if ≥2 similar study designs were available.ResultsIn all, 65 (25 trials, 40 observational) and 29 publications were eligible for review and meta-analyses, respectively. Most studies pertained to hydroxychloroquine (32), ACE inhibitor (ACEi) or angiotensin receptor blocker (ARB) (11), statin (8), and ivermectin (8). In trials, hydroxychloroquine prophylaxis reduced laboratory-confirmed SARS-CoV-2 infection (risk ratio: 0.82 (95% CI 0.74 to 0.90), I2=48%), a result largely driven by one clinical trial (weight: 60.5%). Such beneficial effects were not observed in observational studies, nor for prognostic clinical outcomes. Ivermectin did not significantly reduce the risk of SARS-CoV-2 infection (RR: 0.35 (95% CI 0.10 to 1.26), I2=96%) and findings for clinical outcomes were inconsistent. Neither ACEi or ARB were beneficial in reducing SARS-CoV-2 infection. Most of the evidence from clinical trials was of moderate quality and of lower quality in observational studies.ConclusionsResults from our analysis are insufficient to support an evidence-based repurposed drug policy for SARS-CoV-2 prophylaxis because of inconsistency. In the view of scarce supportive evidence on repurposing drugs for COVID-19, alternative strategies such as immunisation of vulnerable people are warranted to prevent the future waves of infection.PROSPERO registration numberCRD42021292797.
To explore the mechanism of horizontal transmission of hepatitis B virus (HBV) among children, we investigated the quantitative relationship between HBV in saliva and blood from 46 children with ...chronic hepatitis B. We found high levels of HBV DNA in saliva of HBeAg (+) children, suggesting saliva as a vehicle for horizontal transmission of HBV among children.
Abstract Background Hepatitis E virus (HEV) has long been known as a major cause of acute hepatitis in developing countries with occasional travel-related cases in developed countries, most of them ...belonging to genotype 1. Currently, genotype 3 HEV is recognized as an emerging public health issue in developed countries and can cause a chronic hepatitis in immunocompromised patients. Objectives The aim of this study was to get an overview of the clinical course of HEV infection, from July 2007 to December 2012, and further characterize HEV in patients of the University Medical Center Groningen (UMCG) over a 5-year time period. Methods Since the second half of 2007, patients in the UMCG with unexplained hepatitis were screened for HEV and clinical data were collected. HEV was characterized by sequencing of the ORF1 and ORF2 regions. Results In total, 34 patients of the 1129 tested patients showed HEV viremia. The majority of the infected patients were immunocompromised; 18 were solid organ transplant (SOT) patients and 9 were patients immunocompromised for other reasons. Seven patients diagnosed with HEV were immunocompetent. Viral genotyping revealed genotype 3 isolates, mostly genotype 3c. Conclusion Non-travel related HEV hepatitis is an important diagnosis. In immunocompromised patients HEV infection often has major clinical impact, necessitating medical intervention including antiviral treatment. In immunocompetent patients, the detection could expand our understanding about the route of transmission and the relation with the zoonotic origin. Therefore, besides an increasing awareness for HEV among clinicians and medical microbiologists, diagnostics should be routinely incorporated into standard patients care.
Highlights • This report describes the enterovirus D68 epidemic in Europe, 2014. • EU strains were genetically similar to those of the United States outbreak, 2014. • We showed that enterovirus can ...be frequently detected in respiratory samples. • EV-D68 mostly in children with respiratory illness and in immune-compromised adults. • Cross-border collaboration is essential for an adequate response on emerging viruses.
Antiviral resistance in cytomegalovirus (CMV) may result from mutations in the molecular targets of antiviral agents. The aim of this study was to investigate both the prevalence of ...resistance-associated mutations and the factors associated with antiviral resistance in solid organ transplant (SOT) patients with repeated high CMV loads during antiviral treatment.
SOT patients were selected retrospectively, based on CMV loads of >30000 IU/mL at least twice in a period during which treatment was given. Patient samples were tested for antiviral resistance by Sanger sequencing the UL97 and UL54 genes of CMV, which code for the viral kinase and polymerase. Factors predisposing to and resulting from the development of antiviral resistance mutations were analysed.
Multiple samples from 113 SOT patients were tested, showing resistance-associated mutations in 25 patients (22%). A further 20 (18%) patients showed mutations that were not known to be associated with antiviral resistance. Several factors were associated with development of resistance-associated mutations in UL97 as well as UL54, including human leucocyte antigen (HLA) mismatch, which occurred more frequently in the group of patients with resistance mutations. High-level resistance mutations were most frequently seen in UL97.
This study shows that by selecting patients solely on the basis of virological response to treatment, more patients with antiviral resistance mutations are identified. In this study we confirm findings by other groups that primary infections are associated with resistance development. Moreover, we show that HLA mismatch is associated with the development of antiviral resistance, which suggests a role for host immunity in the development of resistance.
For some well-known pathogens like influenza or RSV, diagnostic and epidemiological data is available and continuously complement each other. For most other pathogens however, data is not always ...available or severely delayed. Furthermore, clinical data is needed to assess the burden of disease, which will enhance awareness and help to gain knowledge on emerging pathogens. In this position paper, we discuss the interdependence of diagnostics and epidemiology from a European perspective. In 2004, the European Centre for Disease Prevention and Control (ECDC) was founded to coordinate European wide surveillance and control. At present however, the ECDC still relies on university hospitals, public health institutions and other diagnostic institutions. Close collaboration between all stakeholders across Europe is therefore complex, but necessary to optimize the system for the individual patient. From the diagnostic side, data on detected pathogens should be shared with relevant health institutions in real-time. From the public health side, collected information should be made accessible for diagnostic and clinical institutions in real-time. Subsequently, this information needs to be disseminated across relevant medical disciplines to reach its full potential.
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