Rapid and sensitive amplicon-based genome sequencing of SARS-CoV-2.

As SARS-CoV-2 variants of concern emerged, the genome sequencing of SARS-CoV-2 strains became more important. In this study, SARS-CoV-2 was sequenced using amplicon-based genome sequencing with MinION. The primer panel used in this study consisted of only 11 primer panels and the size of the amplicons was approximately 3 kb. Full genome sequences were obtained with a hundred copies of the SARS-CoV-2 genome, and 92.33% and 75.39% of the genome sequences were obtained with 10 copies of the SARS-CoV-2 genome. The few differences in nucleotide sequences originated from mutations in laboratory cultures and/or mixed nucleotide sequences. The quantification of the SARS-CoV-2 genomic RNA was done using RT-ddPCR methods, and the level of LoD indicated that this sequencing method can be used for any RT-qPCR positive clinical sample. The sequencing results of the SARS-CoV-2 variants and clinical samples showed that our methods were very reliable. The genome sequences of five individual clinical samples were almost identical, and the analysis of the sequence variance showed that most of these nucleotide substitutions were observed in the genome sequences of the other clinical samples, indicating this amplicon-based whole-genome sequencing method can be used in various clinical fields.

Family caregivers' responses to a visitation restriction policy at a Korean surgical intensive care unit before and during the coronavirus disease 2019 pandemic.

BACKGROUND: Since the COVID-19 pandemic, restricting family visits in the ICU has increased concerns regarding negative psychosocial consequences to patients and families. OBJECTIVES: To compare the quality of life, depressive symptoms, and emotions in family caregivers of ICU patients before and during the COVID-19 pandemic, and to explore families' perceptions and suggestions for the visitation. METHODS: A cross-sectional descriptive survey was conducted in 99 family caregivers of adult surgical ICU patients from an urban academic medical center in South Korea (February to July 2021). The WHO's Quality of Life-BREF, Center for Epidemiologic Studies Depression, and Visual Analogue Scale were used to assess quality of life, depressive symptoms, and emotions, respectively. The Family Perception Checklist was used to assess families' perceptions and suggestions about the visitation restriction. Results were compared with the data from our previous survey (n = 187) in 2017. RESULTS: Family caregivers were mostly women (n = 59), adult children (n = 43) or spouse (n = 38) of patients with mean age of 47.34 years. Family caregivers surveyed during the pandemic reported worsening sadness (54.66 ± 28.93, 45.58 ± 29.44, P = 0.005) and anxiety (53.86 ± 30.07, 43.22 ± 29.02, P = 0.001) than those who were surveyed in. While majority of families were satisfied with the visitation restrictions (86.9%), only 50.5% were satisfied with the amount of information provided on the patient's condition. CONCLUSIONS: Visitation restriction is necessary during the COVID-19 pandemic despite sadness and anxiety reported in caregivers. Hence, alternative visitation strategies are needed to mitigate psychological distress and provide sufficient information to ICU family caregivers.

Human Alphacoronavirus Universal Primers for Genome Amplification and Sequencing.

Rapid and accurate sequencing covering the entire genome is essential to identify genetic variations of viral pathogens. However, due to the low viral titers in clinical samples, certain amplification steps are required for viral genome sequencing. At present, there are no universal primers available for alphacoronaviruses and that, since these viruses have diverse strains, new primers specific to the target strain must be continuously developed for sequencing. Thus, in this study, we aimed to develop a universal primer set valid for all human alphacoronaviruses and applicable to samples containing trace amounts of the virus. To this aim, we designed overlapping primer pairs capable of amplifying the entire genome of all known human alphacoronaviruses. The selected primers, named the AC primer set, were composed of 10 primer pairs stretching over the entire genome of alphacoronaviruses, and produced PCR products of the expected size (3-5 kb) from both the HCoV-229E and HCoV-NL63 strains. After genome amplification, an evaluation using various sequencing platforms was carried out. The amplicon library sequencing data were assembled into complete genome sequences in all sequencing strategies examined in this study. The sequencing accuracy varied depending on the sequencing technology, but all sequencing methods showed a sequencing error of less than 0.01%. In the mock clinical specimen, the detection limit was 10-3 PFU/ml (102 copies/ml). The AC primer set and experimental procedure optimized in this study may enable the fast diagnosis of mutant alphacoronaviruses in future epidemics.

Racial and Neighborhood-Level Disparities in COVID-19 Incidence among Patients on Hemodialysis in New York City.

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic has disproportionately affected socially disadvantaged populations. Whether disparities in COVID-19 incidence related to race/ethnicity and socioeconomic factors exist in the hemodialysis population is unknown. METHODS: Our study involved patients receiving in-center hemodialysis in New York City. We used a validated index of neighborhood social vulnerability, the Social Vulnerability Index (SVI), which comprises 15 census tract-level indicators organized into four themes: socioeconomic status, household composition and disability, minority status and language, and housing type and transportation. We examined the association of race/ethnicity and the SVI with symptomatic COVID-19 between March 1, 2020 and August 3, 2020. COVID-19 cases were ascertained using PCR testing. We performed multivariable logistic regression to adjust for demographics, individual-level social factors, dialysis-related medical history, and dialysis facility factors. RESULTS: Of the 1378 patients on hemodialysis in the study, 247 (17.9%) developed symptomatic COVID-19. In adjusted analyses, non-Hispanic Black and Hispanic patients had significantly increased odds of COVID-19 compared with non-Hispanic White patients. Census tract-level overall SVI, modeled continuously or in quintiles, was not associated with COVID-19 in unadjusted or adjusted analyses. Among non-Hispanic White patients, the socioeconomic status SVI theme, the minority status and language SVI theme, and housing crowding were significantly associated with COVID-19 in unadjusted analyses. CONCLUSIONS: Among patients on hemodialysis in New York City, there were substantial racial/ethnic disparities in COVID-19 incidence not explained by neighborhood-level social vulnerability. Neighborhood-level socioeconomic status, minority status and language, and housing crowding were positively associated with acquiring COVID-19 among non-Hispanic Whites. Our findings suggest that socially vulnerable patients on dialysis face disparate COVID-19-related exposures, requiring targeted risk-mitigation strategies.

A consensus guideline of herbal medicine for coronavirus disease 2019.

BACKGROUND: The Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2), which originated in Wuhan, Hubei Province, China in late December 2019, is the cause of ongoing pandemic. We analyzed the symptoms of SARS-CoV-2, a classification of the Chinese medicine dialectic and treatment regimen, and promptly enacted the recommendation of Korean medicine preparations in herbal medicine covered under domestic medical insurance benefits depending on the circumstances in our country. METHOD: The clinical practice guideline (CPG) for the treatment of SARS-CoV-2 was developed based on consensus from a group of experts. RESULTS: Two kinds of herbal medicines (HM) were recommended for the prevention of SARS-CoV-2; Youngyopaedoc-san plus Bojungikgitang, and Youngyopaedoc-san plus Saengmaek-san. Two herbal preparations were recommended for people with a history of exposure to SARS-CoV-2; Youngyopaedoc-san plus Bulhwangeumjeonggi-san, and Youngyopaedoc-san plus Bojungikgi-tang. Three herbal preparations were recommended for mildly symptomatic COVID-19 patients; Youngyopaedoc-san plus Galgunhaegui-tang was recommended for those without pneumonia with wind-warmth disease invading the lungs; Sosiho-tang plus Bulhwangeumjeonggi-san was recommended for those with dampness-heat disease in the lungs. For the recovery stage, Samchulkunbi-tang plus Saengmaek-san, or Samchulkunbi-tang plus Chungseuiki-tang was recommended. CONCLUSION: The CPG was developed to guide the use of Korean herbal medicine in the treatment of SARS-CoV-2, and it is expected that this will be the basis for providing proper treatment of similar infectious diseases in the future.