We investigated the value of monitoring CMV antigenemia during and after antiviral therapy for CMV disease. During the study period, 10 out of 214 renal transplant recipients were treated for CMV disease, receiving a total of 14 courses of treatment. Antigenemia decreased within 7 days after onset of treatment in eight of nine courses associated with a rapid clinical recovery. In three courses with a slow or absent response, antigenemia levels initially increased. Monitoring antigenemia was helpful in differentiating persisting CMV disease from other opportunistic infections and rejection. Relapses of CMV disease were preceded by rises in antigenemia. Viral isolation became negative within 3 days after initiation of ganciclovir, irrespective of the clinical response. Antigenemia is a marker of the effect of ganciclovir on CMV replication in vivo, and its monitoring may be valuable in the management of patients with severe CMV disease.
BackgroundCongenital cytomegalovirus (cCMV) infection is a significant, but potentially under-recognized health threat. Approximately 1 of 150 neonates in the US is born with cCMV infection, with 20% exhibiting long-term health problems due to infection. Both targeted CMV testing of newborns with failed hearing screens and universal CMV screening of all newborns have been proposed as approaches to identify infected newborns early in life. Congenital CMV infection can be diagnosed by testing a newborn’s saliva, urine, or blood by CMV qPCR or culture. Dried blood spots for use in qPCR assays have been shown to be a minimally sensitive specimen. Of the three specimens that are recommended, only saliva is simple, noninvasive and easy to collect.MethodsIn this study, we have validated a real-time (TaqMan) PCR assay for use in testing saliva samples from neonates for the presence of CMV. In conjunction with compatible clinical findings, a CMV positive PCR result forms the basis for a clinical diagnosis. The assay was shown to be specific for CMV, with no cross-reactivity detected for other human herpesviruses or for other human viral pathogens. Since CMV shedding levels from cCMV cases are known to be above the analytical limit of detection for the assay, and samples are collected in a nonsterile environment in which incidental CMV shedding may be present from other neonatal or pediatric patients, the reporting cutoff for this assay was set at 1000 IU/mL. Following analytical validation of the assay, stored (-80°C) residual de-identified clinical saliva samples were tested. The comparator assay was CMV cell culture, and the clinical diagnosis was used to resolve discrepant results.ResultsA total of 9 saliva samples, collected at approx. 1 month of age (or earlier) were tested by both assays. Two samples were negative by both assays and 6 samples were positive by both assays. A single sample was positive by qPCR but negative by cell culture; the qPCR value for this sample was 15,100 IU/mL. This infant had two positive urine cultures, a positive saliva shell vial culture and was clinically confirmed to have cCMV infection.ConclusionThis study suggests improved sensitivity of qPCR over CMV cell culture for identification of neonates congenitally infected with CMV.Disclosures M. Wissel, Viracor Eurofins Laboratories: Employee, Salary S. Cravens, Viracor Eurofins Laboratories: Employee, Salary A. Berg, Viracor Eurofins Laboratories: Employee, Salary A. Widen, Viracor Eurofins Laboratories: Employee, Salary M. Altrich, Viracor Eurofins Laboratories: Employee, Salary S. Kleiboeker, Viracor Eurofins Laboratories: Employee, Salary
Background Influenza (flu) infections affect a large subset of the population every year and have significant impacts on the health of patients, especially those with weak or compromised immune systems such as the elderly, children, cancer patients, and transplant recipients. Baloxavir marboxil was approved in October 2018 as a novel antiviral therapeutic for treating flu. During clinical trials, mutations were identified at the I28 codon of the polymerase acidic (PA) protein that greatly increased the resistance of a flu strain to this novel drug. In this study, a qPCR was developed and validated to identify these resistance mutations, allowing for guided therapy based on the resistance profile of the strain. Methods Flu A sequences (6,175) of the PA gene from the NCBI Influenza Virus Database collected over the last 5 years were compiled and aligned. Primers and probes were designed to target the I38 codon of the PA gene, and specific probes for each codon yielding a resistant amino acid mutation (I38T, -M, and -F) were designed. Locked nucleic acid (LNA) bases were used to increase the specificity of the probes. A combination of clinical flu specimens, laboratory strains, and synthetic constructs of each potential resistance mutation were used to validate the precision, sensitivity, and accuracy of the assay in nasopharyngeal swabs. Results Precision of the cycle threshold (Ct) values for each detector was determined to have a standard deviation of less than 3 for inter-assay and less than 2 for intra-assay replicates. Sensitivity was determined to be 800 copies/mL in nasopharyngeal swabs. Accuracy was found to be 92.3%. A single laboratory strain from the H1N1 2009 epidemic showed cross-reactivity with both wild-type and resistant probes, but no circulating clinical H1N1 samples tested showed this response. Conclusion The precision, sensitivity, and accuracy of a qPCR for resistance mutations to baloxavir marboxil support this assay’s utility as an aid in the treatment of flu in at-risk patient groups. This assay allows for rapid detection (<24 hours) of resistance markers to aid clinicians in improving flu case outcomes. Disclosures All authors: No reported disclosures.
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