Characterization of viral pathogens associated with symptomatic upper respiratory tract infection in adults during a low COVID-19 transmission period.

Background: The epidemiology of respiratory tract infections (RTI) has dramatically changed over the course of the COVID-19 pandemic. A major effort in the clinical management of RTI has been directed toward diagnosing COVID-19, while the causes of other, common community RTI often remain enigmatic. To shed light on the etiological causes of RTI during a low COVID-19 transmission period in 2021, we did a pilot study using molecular testing for virologic causes of upper RTI among adults with respiratory symptoms from Almaty, Kazakhstan. Methods: Adults presenting at two public hospitals with respiratory symptoms were screened using SARS-CoV-2 PCR on nasopharyngeal swabs. A subset of RTI+, COVID-19-negative adults (n = 50) was then tested for the presence of common RTI viruses and influenza A virus (IAV). Next generation virome sequencing was used to further characterize the PCR-detected RTI pathogens. Results: Of 1,812 symptomatic adults, 21 (1.2%) tested SARS-CoV-2-positive. Within the COVID-19 negative outpatient subset, 33/50 subjects (66%) had a positive PCR result for a common community RTI virus, consisting of human parainfluenza virus 3-4 (hPIV 3-4) in 25/50 (50%), rhinovirus (hRV) in 2 (4%), hPIV4-hRV co-infection in four (8%) and adenovirus or the OCR43/HKU-1 coronavirus in two (4%) cases; no IAV was detected. Virome sequencing allowed to reconstruct sequences of most PCR-identified rhinoviruses and hPIV-3/human respirovirus-3. Conclusions: COVID-19 was cause to a low proportion of symptomatic RTI among adults. Among COVID-negative participants, symptomatic RTI was predominantly associated with hPIV and hRV. Therefore, respiratory viruses other than SARS-CoV-2 should be considered in the clinical management and prevention of adult RTI in the post-pandemic era.

SG-APSIC1054: Sputnik-V postvaccination immunologic responses in nasal mucosa: A prospective cohort study in Kazakhstan

Objectives: Sputnik-V (Gam-COVID-Vac) is a recombinant adenoviral (rAdv) vector-based, COVID-19 vaccine now used in >70 countries. Mucosal immunity is thought to be important for protection against COVID-19. We did a prospective cohort study to assess Sputnik-V–elicited mucosal SARS-CoV-2 antibody responses. Methods: We divided 82 COVID-19–free participants into prior COVID-19 and no prior COVID-19 groups and followed them at day 21 after Sputnik-V dose 1′ (rAd5) and dose 2′ (rAd26). Nasopharyngeal swabs and blood were collected to perform SARS-CoV-2 diagnostic and immunologic assays. SARS-CoV-2 spike-specific IgG and IgA ELISAs were performed on both nasal swabs and blood. SARS-CoV-2 real-time RT-PCR testing was performed to exclude infectious influencing. Results: Nasal S-IgG levels increased 25-fold after dose 1′ (P < .001) and remained high after dose 2 in all participants. Prior COVID-19 exposure was associated with both elevated baseline mucosal IgG and IgA and higher postvaccination IgG, but not IgA, boost. Nasal IgA levels increased 16.5-fold after dose 1′ (P < .001) and remained high after dose 2' in all participants. Compared to dose 1′, Sputnik-V dose 2′ did not further increase either mucosal IgG levels (P = .626) or IgA levels (P = .609). Conclusions: A single dose of Sputnik-V boosted mucosal SARS-CoV-2 immunity. The effects of Sputnik-V dose 2′ on mucosal immunity were minimal. These findings indicate (1) that intramuscularly administered adenoviral vaccines enhance SARS-CoV-2 immunity via both systemic and mucosal routes and (2) that cost-effectiveness and the efficacy of Sputnik-V vaccination could be improved by adjusting the current prime-booster regimen and extending the 21-day interval between the doses. Trial registration: Registered on ClinicalTrials.gov (no. NCT04871841).

SG-APSIC1053: Detection of SARS-COV-2 in nasopharyngeal swags with MALDI-TOF MS and machine learning

Objectives: The widespread distribution of SARS-CoV-2 and its high contagiousness pose a challenge for researchers seeking to develop a rapid and cost-effective screening method to identify carriers of this virus. RT-PCR is considered the gold standard for detecting viral RNA in nasopharyngeal swabs, but it is time-consuming and requires constant changes in the primer composition due to the mutation of SARS-CoV-2 strains. We propose a method for the detection of SARS-CoV-2 in nasopharyngeal swabs using MALDI-TOF MS and machine learning. Methods: Nasopharyngeal swabs from patients with PCR-confirmed COVID-19 and control participants were tested (130 and 80 swabs, respectively) with MALDI-TOF MS MicroFlex LT using the HCCA matrix. MALDI spectra were preprocessed in R version 4.1.2 software with the MALDIquant R package using the workflow: sqrt transformation, wavelet smoothing, SNIP-based base removal, and PQN intensity calibration. Peaks were detected with MAD algorithms with following Peak alignment on the following parameters: minFreq 70% and tolerance 0.005. Machine learning was performed with the rtemis r package on GLM, random forest, and XGBoost models. Results: These models were characterized by specificity, sensitivity, and F1 score. GLM models (specificity 1 and sensitivity 0.5) showed a low F1 score of 0.71. However, the random forest and XGBoost models demonstrated sensitivity, specificity, and F1 score equaling 1. Conclusions: We propose a screening method for SARS-CoV-2 detection (sensitivity 1 and specificity 1). This methodology combines the analysis of nasopharyngeal swab samples using MALDI-TOF-MS with machine learning. It is suitable for screening patients with COVID-19 at the first stages of diagnosis. Random forest and XGBoost models demonstrated sensitivity, specificity, and F1 scores equaling 1.

Characterization of viral pathogens associated with symptomatic upper respiratory tract infection in adults during a low COVID-19 transmission period

Background The epidemiology of respiratory tract infections (RTI) has dramatically changed over the course of the COVID-19 pandemic. A major effort in the clinical management of RTI has been directed toward diagnosing COVID-19, while the causes of other, common community RTI often remain enigmatic. To shed light on the etiological causes of RTI during a low COVID-19 transmission period in 2021, we did a pilot study using molecular testing for virologic causes of upper RTI among adults with respiratory symptoms from Almaty, Kazakhstan. Methods Adults presenting at two public hospitals with respiratory symptoms were screened using SARS-CoV-2 PCR on nasopharyngeal swabs. A subset of RTI+, COVID-19-negative adults (n = 50) was then tested for the presence of common RTI viruses and influenza A virus (IAV). Next generation virome sequencing was used to further characterize the PCR-detected RTI pathogens. Results Of 1,812 symptomatic adults, 21 (1.2%) tested SARS-CoV-2-positive. Within the COVID-19 negative outpatient subset, 33/50 subjects (66%) had a positive PCR result for a common community RTI virus, consisting of human parainfluenza virus 3-4 (hPIV 3-4) in 25/50 (50%), rhinovirus (hRV) in 2 (4%), hPIV4-hRV co-infection in four (8%) and adenovirus or the OCR43/HKU-1 coronavirus in two (4%) cases;no IAV was detected. Virome sequencing allowed to reconstruct sequences of most PCR-identified rhinoviruses and hPIV-3/human respirovirus-3. Conclusions COVID-19 was cause to a low proportion of symptomatic RTI among adults. Among COVID-negative participants, symptomatic RTI was predominantly associated with hPIV and hRV. Therefore, respiratory viruses other than SARS-CoV-2 should be considered in the clinical management and prevention of adult RTI in the post-pandemic era.

Next Generation Sequencing Approaches to Characterize the Respiratory Tract Virome.

The COVID-19 pandemic and heightened perception of the risk of emerging viral infections have boosted the efforts to better understand the virome or complete repertoire of viruses in health and disease, with a focus on infectious respiratory diseases. Next-generation sequencing (NGS) is widely used to study microorganisms, allowing the elucidation of bacteria and viruses inhabiting different body systems and identifying new pathogens. However, NGS studies suffer from a lack of standardization, in particular, due to various methodological approaches and no single format for processing the results. Here, we review the main methodological approaches and key stages for studies of the human virome, with an emphasis on virome changes during acute respiratory viral infection, with applications for clinical diagnostics and epidemiologic analyses.

Sustained Delivery of a Monoclonal Antibody against SARS-CoV-2 by Microencapsulated Cells: A Proof-of-Concept Study.

BACKGROUND: Monoclonal antibody (mAb) therapy is a promising antiviral intervention for Coronovirus disease (COVID-19) with a potential for both treatment and prophylaxis. However, a major barrier to implementing mAb therapies in clinical practice is the intricate nature of mAb preparation and delivery. Therefore, here, in a pre-clinical model, we explored the possibility of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mAb delivery using a mAb-expressing encapsulated cell system. METHODS: Murine G-8 myoblasts were transfected with plasmids coding for the heavy and light chains of CR3022, a well-characterized SARS-CoV-2 mAb that targets the Spike receptor binding domain (RBD), and then encapsulated into alginate microcapsules. The microcapsules were then intraperitoneally implanted into immunocompetent (C57/BL6J) mice and changes in circulating CR3022 titres were assessed. The in vitro and ex vivo characterization of the mAb was performed using western blotting, RBD ELISA, and microscopy. RESULTS: Transfected G-8 myoblasts expressed intact CR3022 IgG at levels comparable to transfected HEK-293 cells. Cell encapsulation yielded microcapsules harbouring approximately 1000 cells/capsule and sustainably secreting CR3022 mAb. Subsequent peritoneal G-8 microcapsule implantation into mice resulted in a gradual increase of CR3022 concentration in blood, which by day 7 peaked at 1923 [1656-2190] ng/mL and then gradually decreased ~4-fold by day 40 post-implantation. Concurrently, we detected an increase in mouse anti-CR3022 IgG titers, while microcapsules recovered by day 40 post-implantation showed a reduced per-microcapsule mAb production. SUMMARY: We demonstrate here that cell microencapsulation is a viable approach to systemic delivery of intact SARS-CoV-2 mAb, with potential therapeutic applications that warrant further exploration.

Sputnik-V reactogenicity and immunogenicity in the blood and mucosa: a prospective cohort study.

Sputnik-V (Gam-COVID-Vac) is a heterologous, recombinant adenoviral (rAdv) vector-based, COVID-19 vaccine now used in > 70 countries. Yet there is a shortage of data on this vaccine's performance in diverse populations. Here, we performed a prospective cohort study to assess the reactogenicity and immunologic outcomes of Sputnik-V vaccination in Kazakhstan. COVID-19-free participants (n = 82 at baseline) were followed at day 21 after Sputnik-V dose 1 (rAd5) and dose 2 (rAd26). Self-reported local and systemic adverse events were captured using questionnaires. Blood and nasopharyngeal swabs were collected to perform SARS-CoV-2 diagnostic and immunologic assays. We observed that most of the reported adverse events were mild-to-moderate injection site or systemic reactions, no severe or potentially life-threatening conditions were reported, and dose 1 appeared to be more reactogenic than dose 2. The seroconversion rate was 97% post-dose 1, remaining the same post-dose 2. The proportion of participants with detectable virus neutralization was 83% post-dose 1, increasing to 98% post-dose 2, with the largest relative increase observed in participants without prior COVID-19 exposure. Dose 1 boosted nasal S-IgG and S-IgA, while the boosting effect of dose 2 on mucosal S-IgG, but not S-IgA, was only observed in subjects without prior COVID-19. Systemically, vaccination reduced serum levels of growth regulated oncogene (GRO), which correlated with an elevation in blood platelet count. Overall, Sputnik-V dose 1 elicited both blood and mucosal SARS-CoV-2 immunity, while the immune boosting effect of dose 2 was minimal. Thus, adjustments to the current vaccine dosing regimen are necessary to optimize immunization efficacy and cost-effectiveness. While Sputnik-V reactogenicity is similar to that of other COVID-19 vaccines, the induced alterations to the GRO/platelet axis warrant investigation of the vaccine's effects on systemic immunology.

High SARS-CoV-2 seroprevalence in Karaganda, Kazakhstan before the launch of COVID-19 vaccination.

COVID-19 exposure in Central Asia appears underestimated and SARS-CoV-2 seroprevalence data are urgently needed to inform ongoing vaccination efforts and other strategies to mitigate the regional pandemic. Here, in a pilot serologic study we assessed the prevalence of SARS-CoV-2 antibody-mediated immunity in a multi-ethnic cohort of public university employees in Karaganda, Kazakhstan. Asymptomatic subjects (n = 100) were recruited prior to their first COVID-19 vaccination. Questionnaires were administered to capture a range of demographic and clinical characteristics. Nasopharyngeal swabs were collected for SARS-CoV-2 RT-qPCR testing. Serological assays were performed to detect spike (S)-reactive IgG and IgA and to assess virus neutralization. Pre-pandemic samples were used to validate the assay positivity thresholds. S-IgG and -IgA seropositivity rates among SARS-CoV-2 PCR-negative participants (n = 100) were 42% (95% CI [32.2-52.3]) and 59% (95% CI [48.8-69.0]), respectively, and 64% (95% CI [53.4-73.1]) of the cohort tested positive for at least one of the antibodies. S-IgG titres correlated with virus neutralization activity, detectable in 49% of the tested subset with prior COVID-19 history. Serologically confirmed history of COVID-19 was associated with Kazakh ethnicity, but not with other ethnic minorities present in the cohort, and self-reported history of respiratory illness since March 2020. Overall, SARS-CoV-2 exposure in this cohort was ~15-fold higher compared to the reported all-time national and regional COVID-19 prevalence, consistent with recent studies of excess infection and death in Kazakhstan. Continuous serological surveillance provides important insights into COVID-19 transmission dynamics and may be used to better inform the regional public health response.

Epidemiology, clinical characteristics, and virologic features of COVID-19 patients in Kazakhstan: A nation-wide retrospective cohort study.

BACKGROUND: The earliest coronavirus disease-2019 (COVID-19) cases in Central Asia were announced in March 2020 by Kazakhstan. Despite the implementation of aggressive measures to curb infection spread, gaps remain in the understanding of the clinical and epidemiologic features of the regional pandemic. METHODS: We did a retrospective, observational cohort study of patients with laboratory-confirmed COVID-19 hospitalized in Kazakhstan between February and April 2020. We compared demographic, clinical, laboratory and radiological data of patients with different COVID-19 severities on admission. Logistic regression was used to assess factors associated with disease severity and in-hospital death. Whole-genome SARS-CoV-2 analysis was performed in 53 patients. FINDINGS: Of the 1072 patients with laboratory-confirmed COVID-19 in March-April 2020, the median age was 36 years (IQR 24-50) and 484 (45%) were male. On admission, 683 (64%) participants had asymptomatic/mild, 341 (32%) moderate, and 47 (4%) severe-to-critical COVID-19 manifestation; 20 in-hospital deaths (1•87%) were reported by 5 May 2020. Multivariable regression indicated increasing odds of severe disease associated with older age (odds ratio 1•05, 95% CI 1•03-1•07, per year increase; p<0•001), the presence of comorbidities (2•34, 95% CI 1•18-4•85; p=0•017) and elevated white blood cell count (WBC, 1•13, 95% CI 1•00-1•27; p=0•044) on admission, while older age (1•09, 95% CI 1•06-1•13, per year increase; p<0•001) and male sex (5•63, 95% CI 2•06-17•57; p=0•001) were associated with increased odds of in-hospital death. The SARS-CoV-2 isolates grouped into seven phylogenetic lineages, O/B.4.1, S/A.2, S/B.1.1, G/B.1, GH/B.1.255, GH/B.1.3 and GR/B.1.1.10; 87% of the isolates were O and S sub-types descending from early Asian lineages, while the G, GH and GR isolates were related to lineages from Europe and the Americas. INTERPRETATION: Older age, comorbidities, increased WBC count, and male sex were risk factors for COVID-19 disease severity and mortality in Kazakhstan. The broad SARS-CoV-2 diversity suggests multiple importations and community-level amplification predating travel restriction. FUNDING: Ministry of Education and Science of the Republic of Kazakhstan.