Comparing Nasopharyngeal and Midturbinate Nasal Swab Testing for the Identification of Severe Acute Respiratory Syndrome Coronavirus 2.

Testing of paired midturbinate (MT) nasal and nasopharyngeal (NP) swabs, collected by trained personnel from 40 patients with coronavirus disease 2019 (COVID-19), showed that more NP (76/95 [80%]) than MT swabs tested positive (61/95 [64%]) (P = .02). Among samples collected a week after study enrollment, fewer MT than NP samples were positive (45% vs 76%; P = .001).

Clinical Diagnostic Testing for Human Cytomegalovirus Infections.

Human cytomegalovirus (HCMV) infections are among the most common complications arising in transplant patients, elevating the risk of various complications including loss of graft and death. HCMV infections are also responsible for more congenital infections worldwide than any other agent. Congenital HCMV (cCMV) infections are the leading nongenetic cause of sensorineural hearing loss and a source of significant neurological disabilities in children. While there is overlap in the clinical and laboratory approaches to diagnosis of HCMV infections in these settings, the management, follow-up, treatment, and diagnostic strategies differ considerably. As yet, no country has implemented a universal screening program for cCMV. Here, we summarize the issues, limitations, and application of diagnostic strategies for transplant recipients and congenital infection, including examples of screening programs for congenital HCMV that have been implemented at several centers in Japan, Italy, and the United States.

Cytomegalovirus Genetic Diversity Following Primary Infection.

BACKGROUND: Infection with multiple cytomegalovirus (CMV) strains (mixed infection) was reported in a variety of hosts. As the virus genetic diversity in primary CMV infection and the changes over time remain incompletely defined, we examined CMV diversity and changes in diversity over time in healthy adolescent females who participated in a phase 2 CMV gB/MF59 vaccine trial. METHODS: CMV genetic diversity was determined by genotyping of 5 genes-gB (UL55), gH (UL75), gN (UL73), US28, and UL144-in urine, saliva, and plasma samples from 15 study subjects. RESULTS: At the time of primary infection, 5 of 12 (42%) urine samples had multiple virus strains, and 50% of vaccine recipients were infected with gB1 genotype (vaccine strain). Mixed infection was documented in all 15 subjects within 3 months after primary infection, and the majority had different CMV genotypes in different compartments. Changes in genotypes over time were observed in all subjects. CONCLUSIONS: Infection with multiple CMV genotypes was common during primary infection and further diversification occurred over time. Infection with gB1 genotype in vaccine recipients suggests a lack of strain-specific protection from the vaccine. As only 5 polymorphic genes were assessed, this study likely underestimated the true genetic diversity in primary CMV infection.

Contribution of Congenital Cytomegalovirus Infection to Permanent Hearing Loss in a Highly Seropositive Population: The Brazilian Cytomegalovirus Hearing and Maternal Secondary Infection Study.

BACKGROUND: The exact contribution of congenital cytomegalovirus infection (cCMVI) to permanent hearing loss (HL) in highly seropositive populations is unknown. We determined the contribution of cCMVI to HL and estimated the effectiveness of newborn hearing screening (HS) in identifying neonates with CMV-related HL. METHODS: A total of 11 900 neonates born from a population with ≥97% maternal seroprevalence were screened for cCMVI and HL. cCMVI was confirmed by detection of CMV-DNA in saliva and urine at age <3 weeks. RESULTS: Overall, 68 (0.6%; 95% confidence interval [CI], 0.4-0.7) neonates were identified with cCMVI. Of the 91 (0.8%) newborns who failed the HS, 24 (26.4%) were confirmed with HL, including 7 (29.2%; 95% CI, 17.2-59.3) with cCMVI. Another newborn with cCMVI passed the HS but was confirmed with HL at age 21 days. Of the 62 neonates with cCMVI who underwent a complete hearing evaluation, 8 (12.9%; 95% CI, 6.7-23.4) had HL and most (7/8; 87.5%; 95% CI, 46.6-99.7) were identified by HS. The rate of CMV-related HL was 8 per 11 887 neonates (0.7 per 1000 live births). The prevalence ratio of HL among neonates with cCMVI compared to CMV-uninfected neonates was 89.5 (95% CI, 39.7-202.0). No late-onset cCMVI-related HL was detected during a median follow-up of 36 months. CONCLUSIONS: cCMVI is an important cause of HL in childhood in all settings. Integrating targeted cCMVI screening among neonates who fail a HS could be a reasonable, cost-effective strategy to identify newborns with early-onset cCMVI-related HL.

Performance of the Alethia CMV Assay for Detection of Cytomegalovirus by Use of Neonatal Saliva Swabs.

Congenital cytomegalovirus (cCMV) infection is a major cause of childhood hearing loss and neurodevelopmental delay. Identification of newborns with cCMV infection allows provision of beneficial interventions. However, most infants with cCMV infection have subclinical infection and go undiagnosed. Thus, expanded neonatal CMV testing is increasingly recommended. Saliva is an attractive sample type for CMV testing of newborns, because it is easier to collect than urine and more sensitive for CMV detection than dried blood spots. We evaluated the Alethia CMV assay, a rapid, easy-to-use loop-mediated isothermal amplification method for qualitative detection of CMV DNA in neonatal saliva samples. Saliva swabs were collected prospectively from newborns <21 days old and tested by the Alethia assay according to the manufacturer's instructions. Archived saliva swabs from newborns with cCMV infection were also tested retrospectively. A composite reference method (CRM; two validated PCR assays followed by bidirectional sequencing of amplicons) was performed on all samples as the reference standard comparator. Of 1,480 prospectively collected saliva swabs, 1,472 (99.5%) were negative by both the Alethia assay and CRM, 5 (0.34%) were positive by both the Alethia assay and CRM, and 3 (0.20%) were positive only by the Alethia assay. All 34 (100%) archived swabs from newborns with cCMV infection were positive by both the CRM and the Alethia assay. Overall, the Alethia assay showed 100% and 99.8% positive and negative agreement with the CRM, respectively. The Alethia CMV assay is an accurate method for identifying neonates with cCMV infection and, given its simplicity, appears suitable for CMV testing using neonatal saliva outside a reference laboratory, including remote and resource-limited settings.

Correction to: Association of CMV genomic mutations with symptomatic infection and hearing loss in congenital CMV infection.

After publication of the original article [1], we were notified that Fig. 3 has "Fig. 1" posted on the top of it and Figs. 4 and 5 have "Genomic Position" in a different spot.

Association of CMV genomic mutations with symptomatic infection and hearing loss in congenital CMV infection.

BACKGROUND: Congenital cytomegalovirus (cCMV) infection is the most common congenital infection and a leading cause of long-term neurological and sensory sequelae, the most common being sensorineural hearing loss (SNHL). Despite extensive research, clinical or laboratory markers to identify CMV infected children with increased risk for disease have not been identified. This study utilizes viral whole-genome next generation-sequencing (NGS) of specimens from congenitally infected infants to explore viral diversity and specific viral variants that may be associated with symptomatic infection and SNHL. METHODS: CMV DNA from urine specimens of 30 infants (17 asymptomatic, 13 symptomatic) was target enriched and next generation sequenced resulting in 93% coverage of the CMV genome allowing analysis of viral diversity. RESULTS: Variant frequency distribution was compared between children with symptomatic and asymptomatic cCMV and those with (n = 13) and without (n = 17) hearing loss. The CMV genes UL48A, UL88, US19 and US22 were found to have an increase in nucleotide diversity in symptomatic children; while UL57, UL20, UL104, US14, UL115, and UL35 had an increase in diversity in children with hearing loss. An analysis of single variant differences between symptomatic and asymptomatic children found UL55 to have the highest number, while the most variants associated with SNHL were in the RL11 gene family. In asymptomatic infants with SNHL, mutations were observed more frequently in UL33 and UL20. CONCLUSION: CMV genomes from infected newborns can be mapped to 93% of the genome at a depth allowing accurate and reproducible analysis of polymorphisms for variant and gene discovery that may be linked to symptomatic and hearing loss outcomes.

Contribution of Breastfeeding to False-Positive Saliva Polymerase Chain Reaction for Newborn Congenital Cytomegalovirus Screening.

Real-time polymerase chain reaction (PCR) of saliva is highly sensitive for newborn congenital cytomegalovirus (CMV) screening. This study uses nationally published CMV seroprevalence and breastfeeding rates to estimate the contribution of CMV DNA in breast milk to false-positive saliva PCR results. The false-positive rates adjusted for breastfeeding ranged from 0.03% in white Hispanic persons to 0.14% in white non-Hispanic persons. Saliva CMV PCR for newborn screening is highly sensitive, and the low false-positive rates in this study suggest that saliva PCR results are unlikely to be significantly influenced by breastfeeding or other perinatal exposures.

Breast Milk Human Cytomegalovirus (CMV) Viral Load and the Establishment of Breast Milk CMV-pp65-Specific CD8 T Cells in Human CMV Infected Mothers.

The role of human cytomegalovirus (HCMV)-specific T-cell responses in breast milk of HCMV-seropositive mothers is not well defined. In these studies, we demonstrate that the frequency of cytomegalovirus (CMV)-pp65-specific T-cell responses in peripheral blood mononuclear cells (PBMCs) and breast milk cells (BMCs) is increased for CD8+ T cells in both sample sources when compared with CD4+ T cells. The frequency of pp55-specific CD8 T cells producing interferon γ (IFN-γ) alone or dual IFN-γ/granzyme rB producers is increased in breast milk compared with PBMCs. Lastly, we observed a positive correlation between breast milk viral load and the CD8 pp65-specific response, suggesting that local virus replication drives antigen-specific CD8 T cells into the breast.

Pathogenesis of Non-Zika Congenital Viral Infections.

A widespread epidemic of Zika virus (ZIKV) infection was reported in 2015 from South and Central America and the Caribbean. Although the full spectrum of ZIKV infection of the newborn has yet to be determined, other maternal viral infections resulting in transmission to the fetus provide instructive lessons that can be applied to the prospective evaluation of individuals with ZIKV infection. This review focuses on those other congenital infections, including rubella, congenital cytomegalovirus, human immunodeficiency virus, hepatitis B virus, and neonatal herpes simplex virus, from which lessons for the evaluation of ZIKV in the newborn can be applied.

Newborn Dried Blood Spot Polymerase Chain Reaction to Identify Infants with Congenital Cytomegalovirus-Associated Sensorineural Hearing Loss.

OBJECTIVE: To determine the utility of dried blood spot (DBS) polymerase chain reaction (PCR) in identifying infants with cytomegalovirus (CMV) infection-associated sensorineural hearing loss (SNHL). STUDY DESIGN: Newborns at 7 US hospitals between March 2007 and March 2012 were screened for CMV by saliva rapid culture and/or PCR. Infected infants were monitored for SNHL during the first 4 years of life to determine sensitivity, specificity, and positive and negative likelihood ratios of DBS PCR for identifying CMV-associated SNHL. RESULTS: DBS at birth was positive in 11 of 26 children (42%) with SNHL at age 4 years and in 72 of 270 children (27%) with normal hearing (P = .11). The sensitivity (42.3%; 95% CI, 23.4%-63.1%) and specificity (73.3%; 95% CI, 67.6%-78.5%) was low for DBS PCR in identifying children with SNHL at age 4 years. The positive and negative likelihood ratios of DBS PCR positivity to detect CMV-associated SNHL at age 4 years were 1.6 (95% CI, 0.97-2.6) and 0.8 (95% CI, 0.6-1.1), respectively. There was no difference in DBS viral loads between children with SNHL and those without SNHL. CONCLUSIONS: DBS PCR for CMV has low sensitivity and specificity for identifying infants with CMV-associated hearing loss. These findings, together with previous reports, demonstrate that DBS PCR does not identify either the majority of CMV-infected newborns or those with CMV-associated SNHL early in life.

The "silent" global burden of congenital cytomegalovirus.

Human cytomegalovirus (CMV) is a leading cause of congenital infections worldwide. In the developed world, following the virtual elimination of circulating rubella, it is the commonest nongenetic cause of childhood hearing loss and an important cause of neurodevelopmental delay. The seroprevalence of CMV in adults and the incidence of congenital CMV infection are highest in developing countries (1 to 5% of births) and are most likely driven by nonprimary maternal infections. However, reliable estimates of prevalence and outcome from developing countries are not available. This is largely due to the dogma that maternal preexisting seroimmunity virtually eliminates the risk for sequelae. However, recent data demonstrating similar rates of sequelae, especially hearing loss, following primary and nonprimary maternal infection have underscored the importance of congenital CMV infection in resource-poor settings. Although a significant proportion of congenital CMV infections are attributable to maternal primary infection in well-resourced settings, the absence of specific interventions for seronegative mothers and uncertainty about fetal prognosis have discouraged routine maternal antibody screening. Despite these challenges, encouraging results from prototype vaccines have been reported, and the first randomized phase III trials of prenatal interventions and prolonged postnatal antiviral therapy are under way. Successful implementation of strategies to prevent or reduce the burden of congenital CMV infection will require heightened global awareness among clinicians and the general population. In this review, we highlight the global epidemiology of congenital CMV and the implications of growing knowledge in areas of prevention, diagnosis, prognosis, and management for both low (50 to 70%)- and high (>70%)-seroprevalence settings.

Detection of congenital cytomegalovirus infection by real-time polymerase chain reaction analysis of saliva or urine specimens.

Viral culture of urine or saliva has been the gold standard technique for the diagnosis of congenital cytomegalovirus (CMV) infection. Results of rapid culture and polymerase chain reaction (PCR) analysis of urine and saliva specimens from 80 children were compared to determine the clinical utility of a real-time PCR assay for diagnosis of congenital CMV infection. Results of urine PCR were positive in 98.8% of specimens. Three PCR-positive urine samples were culture negative. Results of saliva PCR and culture were concordant in 78 specimens (97.5%). Two PCR-positive saliva samples were culture negative. These findings demonstrate that PCR performs as well as rapid culture of urine or saliva specimens for diagnosing congenital CMV infection and saliva specimens are easier to collect. Because PCR also offers more rapid turnaround, is unlikely to be affected by storage and transport conditions, has lower cost, and may be adapted to high-throughput situations, it is well suited for targeted testing and large-scale screening for CMV.

Saliva polymerase-chain-reaction assay for cytomegalovirus screening in newborns.

BACKGROUND: Congenital cytomegalovirus (CMV) infection is an important cause of hearing loss, and most infants at risk for CMV-associated hearing loss are not identified early in life because of failure to test for the infection. The standard assay for newborn CMV screening is rapid culture performed on saliva specimens obtained at birth, but this assay cannot be automated. Two alternatives--real-time polymerase-chain-reaction (PCR)-based testing of a liquid-saliva or dried-saliva specimen obtained at birth--have been developed. METHODS: In our prospective, multicenter screening study of newborns, we compared real-time PCR assays of liquid-saliva and dried-saliva specimens with rapid culture of saliva specimens obtained at birth. RESULTS: A total of 177 of 34,989 infants (0.5%; 95% confidence interval [CI], 0.4 to 0.6) were positive for CMV, according to at least one of the three methods. Of 17,662 newborns screened with the use of the liquid-saliva PCR assay, 17,569 were negative for CMV, and the remaining 85 infants (0.5%; 95% CI, 0.4 to 0.6) had positive results on both culture and PCR assay. The sensitivity and specificity of the liquid-saliva PCR assay were 100% (95% CI, 95.8 to 100) and 99.9% (95% CI, 99.9 to 100), respectively, and the positive and negative predictive values were 91.4% (95% CI, 83.8 to 96.2) and 100% (95% CI, 99.9 to 100), respectively. Of 17,327 newborns screened by means of the dried-saliva PCR assay, 74 were positive for CMV, whereas 76 (0.4%; 95% CI, 0.3 to 0.5) were found to be CMV-positive on rapid culture. Sensitivity and specificity of the dried-saliva PCR assay were 97.4% (95% CI, 90.8 to 99.7) and 99.9% (95% CI, 99.9 to 100), respectively. The positive and negative predictive values were 90.2% (95% CI, 81.7 to 95.7) and 99.9% (95% CI, 99.9 to 100), respectively. CONCLUSIONS: Real-time PCR assays of both liquid- and dried-saliva specimens showed high sensitivity and specificity for detecting CMV infection and should be considered potential screening tools for CMV in newborns. (Funded by the National Institute on Deafness and Other Communication Disorders.).

Spectrum of disease and outcome in children with symptomatic congenital cytomegalovirus infection.

OBJECTIVE: To evaluate differences in presentation and outcomes in children with symptomatic congenital cytomegalovirus (cCMV) identified on newborn screening (screened group) and those identified based on clinical findings at birth (referred group). STUDY DESIGN: Data on 178 infants with symptomatic cCMV were analyzed. Demographic characteristics, clinical and laboratory findings documented in the nursery, and sequelae data were compared between the screened and the referred groups using χ(2) or Fisher exact test. RESULTS: Two or more clinical findings were detected at birth in 91% of referred infants, and only 58% of screened infants (P < .001). Significantly more children in the referred group had hearing loss compared with screened infants (P = .009). Fifty-one percent of screened children were free of sequelae compared with only 28% of the referred group (P < .003). CONCLUSIONS: Infants with symptomatic cCMV identified based on clinical suspicion have more severe disease at birth and more commonly have sequelae than those identified on newborn screening. Inclusion of referral infants in many previous reports may have overestimated the severity of disease because of selection bias. Defining the complete spectrum of symptomatic disease due to cCMV and providing precise estimates of disease burden can only be gathered from large newborn screening studies.

Congenital cytomegalovirus infection: clinical outcome.

Congenital cytomegalovirus (CMV) infection is a leading cause of hearing loss and neurologic disabilities in children worldwide. Infants with symptomatic congenital CMV infection at birth are at significantly increased risk for developing adverse long-term outcomes. The vast majority of infants with congenital CMV infection have no clinical findings at birth (asymptomatic infants), and about 10%-15% of these children develop long-term sequelae. Currently, predictors of adverse outcome in asymptomatic congenital CMV infection are not known, and it is important that future studies address this issue.

Increasing Prevalence of Pediatric Community-acquired UTI by Extended Spectrum ß-Lactamase-producing E. coli: Cause for Concern.

BACKGROUND: Antimicrobial resistance and emerging spectrum-ß-lactamase (ESBL) infections are a rising concern in public health. Despite the increasing prevalence of community-acquired (CA) ESBL-E. coli UTIs, there is little data on the antibiotic resistance profiles of this bacterial strain in the pediatric population. We review antibiotic resistance profile and rising trend in pediatric ESBL-E. coli UTI presentation at our pediatric hospital. METHODS: This retrospective study reviewed data drawn from the infectious disease database at our pediatric hospital for all patients whose urine culture grew ESBL-E. coli from 01/2015 to 01/2021. Demographic information and antimicrobial susceptibility test results for ESBL-E. coli isolates from CA-UTIs were collected. Annual changes in resistance to antimicrobial agents and average annual percent change in ESBL-E. coli UTI presentation over the study period are reported. RESULTS: From 01/2015 to 01/2021, 6403 urine cultures at our hospital grew E. coli. Of these, 169 urine cultures from 135 children grew ESBL-E. coli. The study population was 57% male (77) with a mean age of 6.9 ± 6.2 years and multiethnic. CA-UTI by ESBL-producing E. coli accounted for 2.62% of total E. coli UTIs within the study period and increased from 0.97% in 2015 to 3.54% in 2020 by an average of 0.51% each year. CONCLUSIONS: These findings demonstrate an increase in CA-ESBL E. coli UTIs in children. We observed most isolates demonstrated multidrug resistance. As CA-ESBL E. coli UTIs are associated with prolonged hospitalization and increased morbidity, our findings highlight the rising trend in pediatric CA-ESBL E. coli UTI.