Unsupervised machine learning reveals key immune cell subsets in COVID-19, rhinovirus infection, and cancer therapy.

For an emerging disease like COVID-19, systems immunology tools may quickly identify and quantitatively characterize cells associated with disease progression or clinical response. With repeated sampling, immune monitoring creates a real-time portrait of the cells reacting to a novel virus before disease-specific knowledge and tools are established. However, single cell analysis tools can struggle to reveal rare cells that are under 0.1% of the population. Here, the machine learning workflow Tracking Responders EXpanding (T-REX) was created to identify changes in both rare and common cells across human immune monitoring settings. T-REX identified cells with highly similar phenotypes that localized to hotspots of significant change during rhinovirus and SARS-CoV-2 infections. Specialized MHCII tetramer reagents that mark rhinovirus-specific CD4+ cells were left out during analysis and then used to test whether T-REX identified biologically significant cells. T-REX identified rhinovirus-specific CD4+ T cells based on phenotypically homogeneous cells expanding by ≥95% following infection. T-REX successfully identified hotspots of virus-specific T cells by comparing infection (day 7) to either pre-infection (day 0) or post-infection (day 28) samples. Plotting the direction and degree of change for each individual donor provided a useful summary view and revealed patterns of immune system behavior across immune monitoring settings. For example, the magnitude and direction of change in some COVID-19 patients was comparable to blast crisis acute myeloid leukemia patients undergoing a complete response to chemotherapy. Other COVID-19 patients instead displayed an immune trajectory like that seen in rhinovirus infection or checkpoint inhibitor therapy for melanoma. The T-REX algorithm thus rapidly identifies and characterizes mechanistically significant cells and places emerging diseases into a systems immunology context for comparison to well-studied immune changes.

Respiratory Pathogen Coinfections in SARS-CoV-2-Positive Patients in Southeastern Wisconsin: A Retrospective Analysis.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent of coronavirus disease 2019 (COVID-19), has infected all age groups and disproportionately impacted vulnerable populations globally. Polymicrobial infections may play an important role in the development of SARS-CoV-2 infection in susceptible hosts. These coinfections may increase the risk of disease severity and pose challenges to the diagnosis, treatment, and prognosis of COVID-19. There have been limited SARS-CoV-2 coinfection studies. In this retrospective study, residual nucleic acid extracts from 796 laboratory-confirmed COVID-19-positive specimens, collected between March 2020 and February 2021, were analyzed using a Luminex NxTAG respiratory pathogen panel (RPP). Of these, 745 returned valid results and were used for analysis; 53 (7.1%) were positive for one or more additional pathogens. Six different respiratory viruses were detected among the 53 SARS-CoV-2-positive patient specimens, and 7 of those specimens tested positive for more than one additional respiratory virus. The most common pathogens include rhinovirus/enterovirus (RV/EV) (n = 22, 41.51%), human metapneumovirus (hMPV) (n = 18, 33.9%), and adenovirus (n = 12, 22.6%). Interestingly, there were no SARS-CoV-2 coinfections involving influenza A or influenza B in the study specimens. The median age of the SARS-CoV-2-positive patients with coinfections was 38 years; 53% identified as female, and 47% identified as male. Based on our retrospective analysis, respiratory coinfections associated with SARS-CoV-2-positive patients were more common in young children (≤9 years old), with white being the most common race. Our findings will likely prompt additional investigation of polymicrobial infection associated with SARS-CoV-2 during seasonal respiratory pathogen surveillance by public health laboratories. IMPORTANCE This examination of respiratory pathogen coinfections in SARS-CoV-2 patients will likely shed light on our understanding of polymicrobial infection associated with COVID-19. Our results should prompt public health authorities to improve seasonal respiratory pathogen surveillance practices and address the risk of disease severity.

Diagnostic Efficacy and Tolerability of Molded Plastic Nasopharyngeal Swab (FinSwab) Compared to Flocked Nylon Swab in Detection of SARS-CoV-2 and Other Respiratory Viruses.

The supply of testing equipment is vital in controlling the spread of SARS-CoV-2. We compared the diagnostic efficacy and tolerability of molded plastic (FinSwab; Valukumpu, Finland) versus flocked nylon (FLOQSwab; Copan, Italy) nasopharyngeal swabs in a clinical setting. Adults (n = 112) with suspected symptomatic COVID-19 infection underwent nasopharyngeal sampling with FinSwab and FLOQSwab from the same nostril at a drive-in coronavirus testing station. In a subset of 36 patients the samples were collected in a randomized order to evaluate the discomfort associated with sampling. SARS-CoV-2 and 16 other respiratory viruses, as well as human ß-actin mRNA were analyzed by using reverse transcriptase PCR (RT-PCR) assays. Among the 112 patients (mean age, 38 [standard deviation (SD), 14] years) ß-actin mRNA was found in all samples. There was no difference in the ß-actin mRNA cycle threshold (CT) values between FinSwab (mean, 22.3; SD, 3.61) and FLOQSwab (mean, 22.1; SD, 3.50; P = 0.46) swabs. There were 31 virus-positive cases (26 rhinovirus, 4 SARS-CoV-2, and 1 coronavirus-OC43), 24 of which were positive in both swabs; 3 rhinovirus positives were only found in the FinSwab, and similarly 4 rhinovirus positives were only found in the FLOQSwab. Rhinovirus CT values were similar between swab types. Of the 36 patients, 22 (61%) tolerated the sampling with the FinSwab better than with the FLOQSwab (P = 0.065). The molded plastic nasopharyngeal swab (FinSwab) was comparable to the standard flocked swab in terms of efficacy for respiratory virus detection and tolerability of sampling. IMPORTANCE We demonstrate that a molded plastic swab is a valid alternative to conventional brush-like swabs in collection of a nasopharyngeal sample for virus diagnostics.

The Reemergence of Seasonal Respiratory Viruses in Houston, Texas, after Relaxing COVID-19 Restrictions.

Measures intended to limit the spread of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus at the start of the coronavirus disease 2019 (COVID-19) pandemic resulted in a rapid decrease in other respiratory pathogens. Herein, we describe the trends of respiratory pathogens in a major metropolitan health care system central microbiology reference laboratory before and during the COVID-19 pandemic, with attention to when COVID-19 mitigation measures were implemented and relaxed. During the initial lockdown period, COVID-19 was the primary respiratory pathogen detected by multiplex respiratory panels. As COVID-19 containment measures were relaxed, the first non-COVID respiratory viruses to return to prepandemic levels were members of the rhinovirus/enterovirus family. After the complete removal of COVID-19 precautions at the state level, including an end to mask mandates, we observed the robust return of seasonal coronaviruses, parainfluenza virus, and respiratory syncytial virus. Inasmuch as COVID-19 has dominated the landscape of respiratory infections since early 2020, it is important for clinicians to recognize that the return of non-COVID respiratory pathogens may be rapid and significant when COVID-19 containment measures are removed. IMPORTANCE We describe the return of non-COVID respiratory viruses after the removal of COVID-19 mitigation measures. It is important for the public and physicians to recognize that, after months of COVID-19 being the primary driver of respiratory infection, more typical seasonal respiratory illnesses have returned, and this return is out of the normal season for some of these pathogens. Thus, clinicians and the public must now consider both COVID-19 and other respiratory illnesses when a patient presents with symptomatic respiratory illness.

The Positive Rhinovirus/Enterovirus Detection and SARS-CoV-2 Persistence beyond the Acute Infection Phase: An Intra-Household Surveillance Study.

We aimed to assess the duration of nasopharyngeal severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA persistence in adults self-confined at home after acute infection; and to identify the associations of SARS-CoV-2 persistence with respiratory virus co-detection and infection transmission. A cross-sectional intra-household study was conducted in metropolitan Barcelona (Spain) during the time period of April to June 2020. Every adult who was the first family member reported as SARS-CoV-2-positive by reverse transcription polymerase chain reaction (RT-PCR) as well as their household child contacts had nasopharyngeal swabs tested by a targeted SARS-CoV-2 RT-PCR and a multiplex viral respiratory panel after a 15 day minimum time lag. Four-hundred and four households (404 adults and 708 children) were enrolled. SARS-CoV-2 RNA was detected in 137 (33.9%) adults and 84 (11.9%) children. Rhinovirus/Enterovirus (RV/EV) was commonly found (83.3%) in co-infection with SARS-CoV-2 in adults. The mean duration of SARS-CoV-2 RNA presence in adults' nasopharynx was 52 days (range 26-83 days). The persistence of SARS-CoV-2 was significantly associated with RV/EV co-infection (adjusted odds ratio (aOR) 9.31; 95% CI 2.57-33.80) and SARS-CoV-2 detection in child contacts (aOR 2.08; 95% CI 1.24-3.51). Prolonged nasopharyngeal SARS-CoV-2 RNA persistence beyond the acute infection phase was frequent in adults quarantined at home during the first epidemic wave; which was associated with RV/EV co-infection and could enhance intra-household infection transmission.

Respiratory viruses in the healthy middle ear and middle ear with otitis media with effusion.

To investigate the presence of respiratory viruses in the middle ear cavity of the individuals with a healthy middle ear and the children with otitis media with effusion (OME). A total of 72 middle ear samples were collected from 25 children with OME (Group 1) and 47 individuals with no middle ear disease (Group 2). Multiplex real-time polymerase chain reaction was used to investigate the presence of 20 different respiratory viruses. Virus results were compared with bacteriomes of the same populations. At least one respiratory virus was detected in 56% of the patients in Group 1 and 12.8% of the individuals in Group 2. The viral co-infection rate for Group 1 and 2 was 8% and 2.1%, respectively. In Group 1, adenovirus was the most frequently detected virus with a rate of 24%, either alone (16%) or concurrent with other viruses (8%), followed by influenza B (12%), rhinovirus, and bocavirus (8%) each. Parainfluenza 4, coronavirus OC43, and RSV A/B were detected in 4% of the sample each. In Group 2, rhinovirus was detected in two samples (4.3%) followed by adenovirus, coronavirus OC43, coronavirus E299, and coronavirus NL63 with a rate of 2.1% each. The detection rate of respiratory viruses was significantly higher in children aged 6 to 11 years. There was no positive association between virus and bacteria found in the middle ear cavity. The current study has provided comprehensive data indicating the presence of diverse respiratory viruses in the healthy middle ear cavity. Our results also suggest that respiratory viruses might have a contribution to OME pathogenesis.

Low circulation of Influenza A and coinfection with SARS-CoV-2 among other respiratory viruses during the COVID-19 pandemic in a region of southern Brazil.

With the arrival of coronavirus disease 2019 (COVID-19) in Brazil in February 2020, several preventive measures were taken by the population aiming to avoid severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection including the use of masks, social distancing, and frequent hand washing then, these measures may have contributed to preventing infection also by other respiratory viruses. Our goal was to determine the frequencies of Influenza A and B viruses (FLUAV/FLUBV), human mastadenovirus C (HAdV-C), Enterovirus 68 (EV-68), and rhinovirus (RV) besides SARS-CoV-2 among hospitalized patients suspect of COVID-19 with cases of acute respiratory disease syndrome (ARDS) in the period of March to December 2020 and to detect possible coinfections among them. Nucleic acid detection was performed using reverse-transcription quantitative polymerase chain reaction (RT-qPCR) in respiratory samples using naso-oropharyngeal swabs and bronchoalveolar lavage. A total of 418 samples of the 987 analyzed (42.3%) were positive for SARS-CoV-2, 16 (1.62%) samples were positive for FLUAV, no sample was positive for FLUBV or EV-68, 67 (6.78%) samples were positive for HAdV-C, 55 samples were positive for RV 1/2 (26.3%) and 37 for RV 2/2 (13.6%). Coinfections were also detected, including a triple coinfection with SARS-CoV-2, FLUAV, and HAdV-C. In the present work, a very low frequency of FLUV was reported among hospitalized patients with ARDS compared to the past years, probably due to preventive measures taken to avoid COVID-19 and the high influenza vaccination coverage in the region in which this study was performed.

Significant impact of nationwide SARS-CoV-2 lockdown measures on the circulation of other respiratory virus infections in Austria.

BACKGROUND: Since the worldwide spread of SARS-CoV-2, different European countries reacted with temporary national lockdowns with the aim to limit the virus transmission in the population. Also Austria started a lockdown of public life in March 2020. OBJECTIVES: In this study we investigated whether the circulation of different respiratory virus infections in Austria, as assessed by the established respiratory virus surveillance system, is affected by these measures as well and may reflect the success of the lockdown in limiting respiratory virus transmission. STUDY DESIGN: Sentinel data obtained for influenza virus, respiratory syncytial virus, human metapneumovirus and rhinovirus cases were analyzed and compared between the season 2019/2020 and the five previous seasons. RESULTS: We observed a rapid and statistically significant reduction of cumulative cases for all these viruses within short time after the lockdown in March 2020, compared to previous seasons (each p < 0.001). Also, sentinel screening for SARS-CoV-2 infections was performed and a decrease of SARS-CoV-2 was seen after the lockdown. While for the seasonally occurring viruses as influenza, respiratory syncytial virus or human metapneumovirus the lockdown led to the end of the annual epidemics, a re-increase of rhinovirus infections was observed after liberalization of numerous lockdown measures. CONCLUSIONS: Our data provide evidence that occurrence of different respiratory virus infections reflect not only the efficiency of lockdown measures taken against SARS-CoV-2 but it shows also the effects of lockdown releases on the transmission of respiratory viruses.

Increased risk of rhinovirus infection in children during the coronavirus disease-19 pandemic.

BACKGROUND: Coronavirus disease (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was first detected in Japan in January 2020 and has spread throughout the country. Previous studies have reported that viral interference among influenza virus, rhinovirus, and other respiratory viruses can affect viral infections at the host and population level. METHODS: To investigate the impact of COVID-19 on influenza and other respiratory virus infections, we analyzed clinical specimens collected from 2244 patients in Japan with respiratory diseases between January 2018 and September 2020. RESULTS: The frequency of influenza and other respiratory viruses (coxsackievirus A and B; echovirus; enterovirus; human coronavirus 229E, HKU1, NL63, and OC43; human metapneumovirus; human parainfluenza virus 1, 2, 3, and 4; human parechovirus; human respiratory syncytial virus; human adenovirus; human bocavirus; human parvovirus B19; herpes simplex virus type 1; and varicella-zoster virus) was appreciably reduced among all patients during the COVID-19 pandemic except for that of rhinovirus in children younger than 10 years, which was appreciably increased. COVID-19 has not spread among this age group, suggesting an increased risk of rhinovirus infection in children. CONCLUSIONS: Rhinovirus infections should be continuously monitored to understand their increased risk during the COVID-19 pandemic and viral interference with SARS-CoV-2.

Systematic review and meta-analysis of the prevalence of common respiratory viruses in children < 2 years with bronchiolitis in the pre-COVID-19 pandemic era.

INTRODUCTION: The advent of genome amplification assays has allowed description of new respiratory viruses and to reconsider the role played by certain respiratory viruses in bronchiolitis. This systematic review and meta-analysis was initiated to clarify the prevalence of respiratory viruses in children with bronchiolitis in the pre-COVID-19 pandemic era. METHODS: We performed an electronic search through Pubmed and Global Index Medicus databases. We included observational studies reporting the detection rate of common respiratory viruses in children with bronchiolitis using molecular assays. Data was extracted and the quality of the included articles was assessed. We conducted sensitivity, subgroups, publication bias, and heterogeneity analyses using a random effect model. RESULTS: The final meta-analysis included 51 studies. Human respiratory syncytial virus (HRSV) was largely the most commonly detected virus 59.2%; 95% CI [54.7; 63.6]). The second predominant virus was Rhinovirus (RV) 19.3%; 95% CI [16.7; 22.0]) followed by Human bocavirus (HBoV) 8.2%; 95% CI [5.7; 11.2]). Other reported viruses included Human Adenovirus (HAdV) 6.1%; 95% CI [4.4; 8.0]), Human Metapneumovirus (HMPV) 5.4%; 95% CI [4.4; 6.4]), Human Parainfluenzavirus (HPIV) 5.4%; 95% CI [3.8; 7.3]), Influenza 3.2%; 95% CI [2.2; 4.3], Human Coronavirus (HCoV) 2.9%; 95% CI [2.0; 4.0]), and Enterovirus (EV) 2.9%; 95% CI [1.6; 4.5]). HRSV was the predominant virus involved in multiple detection and most codetections were HRSV + RV 7.1%, 95% CI [4.6; 9.9]) and HRSV + HBoV 4.5%, 95% CI [2.4; 7.3]). CONCLUSIONS: The present study has shown that HRSV is the main cause of bronchiolitis in children, we also have Rhinovirus, and Bocavirus which also play a significant role. Data on the role played by SARS-CoV-2 in children with acute bronchiolitis is needed. REVIEW REGISTRATION: PROSPERO, CRD42018116067.

PACIFIC: a lightweight deep-learning classifier of SARS-CoV-2 and co-infecting RNA viruses.

Viral co-infections occur in COVID-19 patients, potentially impacting disease progression and severity. However, there is currently no dedicated method to identify viral co-infections in patient RNA-seq data. We developed PACIFIC, a deep-learning algorithm that accurately detects SARS-CoV-2 and other common RNA respiratory viruses from RNA-seq data. Using in silico data, PACIFIC recovers the presence and relative concentrations of viruses with > 99% precision and recall. PACIFIC accurately detects SARS-CoV-2 and other viral infections in 63 independent in vitro cell culture and patient datasets. PACIFIC is an end-to-end tool that enables the systematic monitoring of viral infections in the current global pandemic.

Pneumonia in children admitted to the national referral hospital in Bhutan: A prospective cohort study.

OBJECTIVES: The study aim was to describe the etiological profile and clinical characteristics of pneumonia among children hospitalized in Thimphu, Bhutan. METHODS: This prospective study enrolled children aged 2-59 months admitted to the Jigme Dorji Wangchuck National Referral Hospital with World Health Organization (WHO)-defined clinical pneumonia. Demographic and clinico-radiological data were collected through questionnaires, physical examination, and chest radiography. Blood samples and nasopharyngeal washing were collected for microbiological analysis including culture and molecular methods. RESULTS: From July 2017 to June 2018, 189 children were enrolled, of which 53.4% were infants. Pneumonia-related admissions were less frequent over the winter. Chest radiographies were obtained in 149 children; endpoints included pneumonia in 39 cases (26.2%), other infiltrates in 31 (20.8%), and were normal in 79 children (53.0%). Non-contaminated bacterial growth was detected in 8/152 (5.3%) blood cultures, with only two cases of Streptococcus pneumoniae. Viral detection in upper respiratory secretions was common, with at least one virus detected in 103/115 (89.6%). The three most-commonly isolated viruses were respiratory syncytial virus (52/115; 45.2%), rhinovirus (42/115; 36.5%), and human parainfluenza virus (19/115; 16.5%). A third of patients with viral infections showed mixed infections. Case fatality rate was 3.2% (6/189). CONCLUSION: Respiratory viral infections predominated among this cohort of WHO-defined clinical pneumonia cases, whereas bacterial aetiologies were uncommon, highlighting the epidemiologic transition that Bhutan seems to have reached.

Identification of SARS-CoV-2 and additional respiratory pathogens cases under the investigation of COVID-19 initial phase in a Brazilian reference laboratory.

Coronavirus disease 2019 (COVID-19) surveillance, in Brazil, initiated shortly after its description, in China. Our aim was to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and additional pathogens in samples from the initial phase of the outbreak in Brazil, from late February to late March. From 707 samples analysed, 29 (4.1%) were SARS-CoV-2 positive. Fever and cough were their most prevalent symptoms. Co-detection of rhinovirus was observed in 2 (6.9%) cases. Additional pathogens were identified in 66.1% of the SARS-CoV-2 negative cases, mainly rhinovirus and influenza A(H1N1)pdm09. Thus, we emphasise the importance of differential diagnosis in COVID-19 suspected cases.

Analysis of Covid-19 and non-Covid-19 viruses, including influenza viruses, to determine the influence of intensive preventive measures in Japan.

BACKGROUND: Severe acute respiratory coronavirus 2 (SARS-CoV-2) has spread and caused death worldwide. Preventive measures and infection control are underway, and some areas show signs of convergence. Other viruses in addition to SARS-CoV-2 cause cold-like symptoms and spread in the winter. However, the extent to which SARS-CoV-2, influenza viruses and other causative viruses have prevailed since implementing preventive measures is unclear. OBJECTIVES: We aim to investigate the incidence of causative viruses and pathogens in patients. STUDY DESIGN: We collected 191 nasopharyngeal swabs from patients with cold-like symptoms in Japan. All samples were subjected to multiplex PCR with the FilmArray Respiratory Panel and reverse transcription PCR (RT-PCR) to detect SARS-CoV-2. RESULTS: FilmArray Respiratory Panel analysis detected at least one virus in 32 of 191 patients with cold-like symptoms (21 %). Of these, we frequently identified human rhinoviruses/enteroviruses (5.8 %, n=11), human metapneumovirus (3.7 %, n=7), coronavirus 229E (2.1 %, n=4) and coronavirus OC43 (1.6 %, n=3); while no influenza viruses were detected. RT-PCR analysis detected SARS-CoV-2 (4.2 %, n=8) in patients who were not infected with the aforementioned respiratory viruses. CONCLUSIONS: Co-infection with SARS-CoV-2 and other viruses was not observed. Causative viruses remain prevalent after implementing preventive measures. SARS-CoV-2 differs from influenza viruses in its infectivity.

Inside the lungs of COVID-19 disease.

In the setting of the coronavirus disease 2019 (COVID-19) pandemic, only few data regarding lung pathology induced by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is available, especially without medical intervention interfering with the natural evolution of the disease. We present here the first case of forensic autopsy of a COVID-19 fatality occurring in a young woman, in the community. Diagnosis was made at necropsy and lung histology showed diffuse alveolar damage, edema, and interstitial pneumonia with a geographically heterogeneous pattern, mostly affecting the central part of the lungs. This death related to COVID-19 pathology highlights the heterogeneity and severity of central lung lesions after natural evolution of the disease.

An investigation into respiratory tract viruses in children with acute lower respiratory tract infection or wheezing.

BACKGROUND: This study aimed to determine the frequencies of respiratory tract viruses in patient (acute lower respiratory tract infection [LRTI] or wheezing) and control (history of asthma without symptoms) groups. METHODS: Using multiplex-polymerase chain reaction (PCR), respiratory tract viruses were investigated in the respiratory tract specimens from patient and control groups followed in the Pediatric Clinic. RESULTS: The viruses detected in the patient and control groups (P=0.013) were as follows, respectively: rhinoviruses A, B, C (25.6% and 36.7%), influenza virus A (21.1% and 0.0%), parainfluenza virus type 1 (7.8% and 1.7%), parainfluenza virus type 4 (5.6% and 0.0%), adenoviruses A, B, C, D, E (4.4% and 1.7%), parainfluenza virus type 3 (4.4% and 1.7%), coronaviruses 229E and NL63 (4.4% and 1.7%), coronavirus OC43 (3.3% and 0.0%), respiratory syncytial virus A (3.3% and 0.0%), parainfluenza virus type 2 (2.2% and 0.0%), influenza virus B (2.2% and 0.0%), and respiratory syncytial virus B (1.1% and 1.7%). No bocavirus, metapneumovirus or enterovirus was found in any specimen. Statistically significant differences in the detection of influenza virus A (P=0.000), the total detection of parainfluenza viruses (P=0.008) and coinfection (P=0.004) were observed between the patient and control groups. CONCLUSIONS: The advantage of our study compared with other studies is the inclusion of not only wheezing patients but also children with asthma without symptom. The higher detection of rhinoviruses both in patient and control groups give rise to thought that these viruses may be responsible for asthma exacerbations and may be related with long duration of virus shedding.