Increased viral variants in children and young adults with impaired humoral immunity and persistent SARS-CoV-2 infection: A consecutive case series.

BACKGROUND: There is increasing concern that persistent infection of SARS-CoV-2 within immunocompromised hosts could serve as a reservoir for mutation accumulation and subsequent emergence of novel strains with the potential to evade immune responses. METHODS: We describe three patients with acute lymphoblastic leukemia who were persistently positive for SARS-CoV-2 by real-time polymerase chain reaction. Viral viability from longitudinally-collected specimens was assessed. Whole-genome sequencing and serological studies were performed to measure viral evolution and evidence of immune escape. FINDINGS: We found compelling evidence of ongoing replication and infectivity for up to 162 days from initial positive by subgenomic RNA, single-stranded RNA, and viral culture analysis. Our results reveal a broad spectrum of infectivity, host immune responses, and accumulation of mutations, some with the potential for immune escape. INTERPRETATION: Our results highlight the potential need to reassess infection control precautions in the management and care of immunocompromised patients. Routine surveillance of mutations and evaluation of their potential impact on viral transmission and immune escape should be considered.

High Prevalence of SARS-CoV-2 Genetic Variation and D614G Mutation in Pediatric Patients With COVID-19.

BACKGROUND: The full spectrum of the disease phenotype and viral genotype of coronavirus disease 2019 (COVID-19) have yet to be thoroughly explored in children. Here, we analyze the relationships between viral genetic variants and clinical characteristics in children. METHODS: Whole-genome sequencing was performed on respiratory specimens collected for all SARS-CoV-2-positive children (n = 141) between March 13 and June 16, 2020. Viral genetic variations across the SARS-CoV-2 genome were identified and investigated to evaluate genomic correlates of disease severity. RESULTS: Higher viral load was detected in symptomatic patients (P = .0007) and in children <5 years old (P = .0004). Genomic analysis revealed a mean pairwise difference of 10.8 single nucleotide variants (SNVs), and the majority (55.4%) of SNVs led to an amino acid change in the viral proteins. The D614G mutation in the spike protein was present in 99.3% of the isolates. The calculated viral mutational rate of 22.2 substitutions/year contrasts the 13.5 substitutions/year observed in California isolates without the D614G mutation. Phylogenetic clade 20C was associated with severe cases of COVID-19 (odds ratio, 6.95; P = .0467). Epidemiological investigation revealed major representation of 3 of 5 major Nextstrain clades (20A, 20B, and 20C) consistent with multiple introductions of SARS-CoV-2 in Southern California. CONCLUSIONS: Genomic evaluation demonstrated greater than expected genetic diversity, presence of the D614G mutation, increased mutation rate, and evidence of multiple introductions of SARS-CoV-2 into Southern California. Our findings suggest a possible association of phylogenetic clade 20C with severe disease, but small sample size precludes a definitive conclusion. Our study warrants larger and multi-institutional genomic evaluation and has implications for infection control practices.

Comparison of Upper Respiratory Viral Load Distributions in Asymptomatic and Symptomatic Children Diagnosed with SARS-CoV-2 Infection in Pediatric Hospital Testing Programs.

The distribution of upper respiratory viral loads (VL) in asymptomatic children infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is unknown. We assessed PCR cycle threshold (Ct) values and estimated VL in infected asymptomatic children diagnosed in nine pediatric hospital testing programs. Records for asymptomatic and symptomatic patients with positive clinical SARS-CoV-2 tests were reviewed. Ct values were (i) adjusted by centering each value around the institutional median Ct value from symptomatic children tested with that assay and (ii) converted to estimated VL (numbers of copies per milliliter) using internal or manufacturer data. Adjusted Ct values and estimated VL for asymptomatic versus symptomatic children (118 asymptomatic versus 197 symptomatic children aged 0 to 4 years, 79 asymptomatic versus 97 symptomatic children aged 5 to 9 years, 69 asymptomatic versus 75 symptomatic children aged 10 to 13 years, 73 asymptomatic versus 109 symptomatic children aged 14 to 17 years) were compared. The median adjusted Ct value for asymptomatic children was 10.3 cycles higher than for symptomatic children (P < 0.0001), and VL were 3 to 4 logs lower than for symptomatic children (P < 0.0001); differences were consistent (P < 0.0001) across all four age brackets. These differences were consistent across all institutions and by sex, ethnicity, and race. Asymptomatic children with diabetes (odds ratio [OR], 6.5; P = 0.01), a recent contact (OR, 2.3; P = 0.02), and testing for surveillance (OR, 2.7; P = 0.005) had higher estimated risks of having a Ct value in the lowest quartile than children without, while an immunocompromised status had no effect. Children with asymptomatic SARS-CoV-2 infection had lower levels of virus in their nasopharynx/oropharynx than symptomatic children, but the timing of infection relative to diagnosis likely impacted levels in asymptomatic children. Caution is recommended when choosing diagnostic tests for screening of asymptomatic children.

Saliva Is a Promising Alternative Specimen for the Detection of SARS-CoV-2 in Children and Adults.

Testing efforts for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been burdened by the scarcity of testing materials and personal protective equipment for health care workers. The simple and painless process of saliva collection allows for widespread testing, but enthusiasm is hampered by variable performance compared to that of nasopharyngeal swab (NPS) samples. We prospectively collected paired NPS and saliva samples from a total of 300 unique adult and pediatric patients. SARS-CoV-2 RNA was detected in 32.2% (97/300) of the individuals using the TaqPath COVID-19 Combo kit (Thermo Fisher). Performance of saliva and NPS was compared against the total number of positives regardless of specimen type. The overall concordances for saliva and NPS were 91.0% (273/300) and 94.7% (284/300), respectively. The values for positive percent agreement (PPA) for saliva and NPS were 81.4% (79/97) and 89.7% (87/97), respectively. Saliva yielded detection of 10 positive cases that were negative by NPS. For symptomatic and asymptomatic pediatric patients not previously diagnosed with COVID-19, the performances of saliva and NPS were comparable (PPA, 82.4% versus 85.3%). The overall values for PPA for adults were 83.3% and 90.7% for saliva and NPS, respectively, with saliva yielding detection of 4 fewer cases than NPS. However, saliva performance for symptomatic adults was identical to NPS performance (PPA of 93.8%). With lower cost and self-collection capabilities, saliva can be an appropriate sample choice alternative to NPS for detection of SARS-CoV-2 in children and adults.

Saliva is a promising alternative specimen for the detection of SARS-CoV-2 in children and adults.

Testing efforts for SARS-CoV-2 have been burdened by the scarcity of testing materials and personal protective equipment for healthcare workers. The simple and painless process of saliva collection allows for widespread testing, but enthusiasm is hampered by variable performance compared to nasopharyngeal swab (NPS) samples. We prospectively collected paired NPS and saliva samples from a total of 300 unique adult and pediatric patients. SARS-CoV-2 RNA was detected in 32.2% (97/300) of the individuals using the TaqPath COVID-19 Combo Kit (Thermo Fisher). Performance of saliva and NPS were compared against the total number of positives regardless of specimen type. The overall concordance for saliva and NPS was 91.0% (273/300) and 94.7% (284/300), respectively. The positive percent agreement (PPA) for saliva and NPS was 81.4% (79/97) and 89.7% (87/97), respectively. Saliva detected 10 positive cases that were negative by NPS. In symptomatic and asymptomatic pediatric patients not previously diagnosed with COVID-19, the performances of saliva and NPS were comparable (PPA: 82.4% vs 85.3%). The overall PPA for adults were 83.3% and 90.7% for saliva and NPS, respectively, with saliva detecting 4 cases less than NPS. However, saliva performance in symptomatic adults was identical to NPS (PPA of 93.8%). With lower cost and self-collection capabilities, saliva can be an appropriate alternative sample choice to NPS for detection of SARS-CoV-2 in children and adults. SUMMARY: Saliva is an acceptable alternative specimen compared to nasopharyngeal swabs for detection of SARS-CoV-2. Specifically, saliva demonstrated comparable performance to nasopharyngeal swabs in symptomatic and asymptomatic pediatric patients and in symptomatic adults.