Analysis of 16S rRNA Gene Sequence of Nasopharyngeal Exudate Reveals Changes in Key Microbial Communities Associated with Aging.

Functional or compositional perturbations of the microbiome can occur at different sites, of the body and this dysbiosis has been linked to various diseases. Changes in the nasopharyngeal microbiome are associated to patient's susceptibility to multiple viral infections, supporting the idea that the nasopharynx may be playing an important role in health and disease. Most studies on the nasopharyngeal microbiome have focused on a specific period in the lifespan, such as infancy or the old age, or have other limitations such as low sample size. Therefore, detailed studies analyzing the age- and sex-associated changes in the nasopharyngeal microbiome of healthy people across their whole life are essential to understand the relevance of the nasopharynx in the pathogenesis of multiple diseases, particularly viral infections. One hundred twenty nasopharyngeal samples from healthy subjects of all ages and both sexes were analyzed by 16S rRNA sequencing. Nasopharyngeal bacterial alpha diversity did not vary in any case between age or sex groups. Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidetes were the predominant phyla in all the age groups, with several sex-associated. Acinetobacter, Brevundimonas, Dolosigranulum, Finegoldia, Haemophilus, Leptotrichia, Moraxella, Peptoniphilus, Pseudomonas, Rothia, and Staphylococcus were the only 11 bacterial genera that presented significant age-associated differences. Other bacterial genera such as Anaerococcus, Burkholderia, Campylobacter, Delftia, Prevotella, Neisseria, Propionibacterium, Streptococcus, Ralstonia, Sphingomonas, and Corynebacterium appeared in the population with a very high frequency, suggesting that their presence might be biologically relevant. Therefore, in contrast to other anatomical areas such as the gut, bacterial diversity in the nasopharynx of healthy subjects remains stable and resistant to perturbations throughout the whole life and in both sexes. Age-associated abundance changes were observed at phylum, family, and genus levels, as well as several sex-associated changes probably due to the different levels of sex hormones present in both sexes at certain ages. Our results provide a complete and valuable dataset that will be useful for future research aiming for studying the relationship between changes in the nasopharyngeal microbiome and susceptibility to or severity of multiple diseases.

Nasopharyngeal microbiome of COVID-19 patients revealed a distinct bacterial profile in deceased and recovered individuals.

The bacterial co-infections in SARS-CoV-2 patients remained the least explored subject of clinical manifestations that may also determine the disease severity. Nasopharyngeal microbial community structure within SARS-CoV-2 infected patients could reveal interesting microbiome dynamics that may influence the disease outcomes. Here, in this research study, we analyzed distinct nasopharyngeal microbiome profile in the deceased (n = 48) and recovered (n = 29) COVID-19 patients and compared it with control SARS-CoV-2 negative individuals (control) (n = 33). The nasal microbiome composition of the three groups varies significantly (PERMANOVA, p-value <0.001), where deceased patients showed higher species richness compared to the recovered and control groups. Pathogenic genera, including Corynebacterium (LDA score 5.51), Staphylococcus, Serratia, Klebsiella and their corresponding species were determined as biomarkers (p-value <0.05, LDA cutoff 4.0) in the deceased COVID-19 patients. Ochrobactrum (LDA score 5.79), and Burkholderia (LDA 5.29), were found in the recovered group which harbors ordinal bacteria (p-value <0.05, LDA-4.0) as biomarkers. Similarly, Pseudomonas (LDA score 6.19), and several healthy nasal cavity commensals including Veillonella, and Porphyromonas, were biomarkers for the control individuals. Healthy commensal bacteria may trigger the immune response and alter the viral infection susceptibility and thus, may play important role and possible recovery that needs to be further explored. This research finding provide vital information and have significant implications for understanding the microbial diversity of COVID-19 patients. However, additional studies are needed to address the microbiome-based therapeutics and diagnostics interventions.

Age-Related Changes in the Nasopharyngeal Microbiome Are Associated With Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infection and Symptoms Among Children, Adolescents, and Young Adults.

BACKGROUND: Children are less susceptible to SARS-CoV-2 infection and typically have milder illness courses than adults, but the factors underlying these age-associated differences are not well understood. The upper respiratory microbiome undergoes substantial shifts during childhood and is increasingly recognized to influence host defense against respiratory pathogens. Thus, we sought to identify upper respiratory microbiome features associated with SARS-CoV-2 infection susceptibility and illness severity. METHODS: We collected clinical data and nasopharyngeal swabs from 285 children, adolescents, and young adults (<21 years) with documented SARS-CoV-2 exposure. We used 16S ribosomal RNA gene sequencing to characterize the nasopharyngeal microbiome and evaluated for age-adjusted associations between microbiome characteristics and SARS-CoV-2 infection status and respiratory symptoms. RESULTS: Nasopharyngeal microbiome composition varied with age (PERMANOVA, P < .001; R2 = 0.06) and between SARS-CoV-2-infected individuals with and without respiratory symptoms (PERMANOVA, P  = .002; R2 = 0.009). SARS-CoV-2-infected participants with Corynebacterium/Dolosigranulum-dominant microbiome profiles were less likely to have respiratory symptoms than infected participants with other nasopharyngeal microbiome profiles (OR: .38; 95% CI: .18-.81). Using generalized joint attributed modeling, we identified 9 bacterial taxa associated with SARS-CoV-2 infection and 6 taxa differentially abundant among SARS-CoV-2-infected participants with respiratory symptoms; the magnitude of these associations was strongly influenced by age. CONCLUSIONS: We identified interactive relationships between age and specific nasopharyngeal microbiome features that are associated with SARS-CoV-2 infection susceptibility and symptoms in children, adolescents, and young adults. Our data suggest that the upper respiratory microbiome may be a mechanism by which age influences SARS-CoV-2 susceptibility and illness severity.

NGS analysis of nasopharyngeal microbiota in SARS-CoV-2 positive patients during the first year of the pandemic in the Campania Region of Italy.

Since its first appearance, the SARS-CoV-2 has spread rapidly in the human population, reaching the pandemic scale with >280 million confirmed infections and more than 5 million deaths to date (https://covid19.who.int/). These data justify the urgent need to enhance our understanding of SARS-CoV-2 effects in the respiratory system, including those linked to co-infections. The principal aim of our study is to investigate existing correlations in the nasopharynx between the bacterial community, potential pathogens, and SARS-CoV-2 infection. The main aim of this study was to provide evidence pointing to possible relationships between components of the bacterial community and SARS-CoV-2 in the nasopharynx. Meta-transcriptomic profiling of the nasopharyngeal microbial community was carried out in 89 SARS-Cov-2 positive subjects from the Campania Region in Italy. To this end, RNA extracted from nasopharyngeal swabs collected at different times during the initial phases of the pandemic was analyzed by Next-Generation Sequencing (NGS). Results show a consistently high presence of members of the Proteobacteria (41.85%), Firmicutes (28.54%), and Actinobacteria (16.10%) phyla, and an inverted correlation between the host microbiome, co-infectious bacteria, and super-potential pathogens such as Staphylococcus aureus, Klebsiella pneumoniae, Streptococcus pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, and Neisseria gonorrhoeae. In depth characterization of microbiota composition in the nasopharynx can provide clues to understand its potential contribution to the clinical phenotype of Covid-19, clarifying the interaction between SARS-Cov-2 and the bacterial flora of the host, and highlighting its dysbiosis and the presence of pathogens that could affect the patient's disease progression and outcome.

Concordance in RT-PCR detection of SARS-CoV-2 between samples preserved in viral and bacterial transport medium.

BACKGROUND: While the detection of SARS-CoV-2 in samples preserved in viral transport medium (VTM) by RT-PCR is a standard diagnostic method, this may preclude the study of bacterial respiratory pathogens from the same specimen. It is unclear if the use of skim milk, tryptone, glucose, and glycerin (STGG) transport media, used for study of respiratory bacteria, allows an efficient and concurrent study of SARS-CoV-2 infections. OBJECTIVES: To determine the concordance in SARS-CoV-2 detection by real time RT-PCR between paired nasopharyngeal (NP) swabs preserved in STGG and nasal (NS) swabs preserved in VTM. STUDY DESIGN: Paired samples of NP and NS swabs were collected between December 2020 and March 2021 from a prospective longitudinal cohort study of 44 households and 132 participants from a peri-urban community (Lima, Peru). NP and NS swabs were taken from all participants once and twice per week, respectively, independent of respiratory symptoms. STGG medium was used for NP samples and VTM for NS samples. Samples were analyzed for SARS-CoV-2 by RT-PCR for N, S and ORF1ab targets. We calculated the concordance in detections between sample types and compared the RT-PCR cycle thresholds (Ct). RESULTS: Among the 148 paired samples, we observed a high concordance in detections between NP and NS samples (agreement = 94.59%; Kappa = 0.79). Median Ct values were statistically similar between sample types for each RT-PCR target: N, S and ORF1ab (p = 0.11, p = 0.71 and p = 0.11, respectively). CONCLUSIONS: NP swabs collected in STGG medium are reliable alternatives to nasal swabs collected in VTM for the study of SARS-CoV-2.

SARS-CoV-2 infection reduces human nasopharyngeal commensal microbiome with inclusion of pathobionts.

The microbiota of the nasopharyngeal tract (NT) play a role in host immunity against respiratory infectious diseases. However, scant information is available on interactions of SARS-CoV-2 with the nasopharyngeal microbiome. This study characterizes the effects of SARS-CoV-2 infection on human nasopharyngeal microbiomes and their relevant metabolic functions. Twenty-two (n = 22) nasopharyngeal swab samples (including COVID-19 patients = 8, recovered humans = 7, and healthy people = 7) were collected, and underwent to RNAseq-based metagenomic investigation. Our RNAseq data mapped to 2281 bacterial species (including 1477, 919 and 676 in healthy, COVID-19 and recovered metagenomes, respectively) indicating a distinct microbiome dysbiosis. The COVID-19 and recovered samples included 67% and 77% opportunistic bacterial species, respectively compared to healthy controls. Notably, 79% commensal bacterial species found in healthy controls were not detected in COVID-19 and recovered people. Similar dysbiosis was also found in viral and archaeal fraction of the nasopharyngeal microbiomes. We also detected several altered metabolic pathways and functional genes in the progression and pathophysiology of COVID-19. The nasopharyngeal microbiome dysbiosis and their genomic features determined by our RNAseq analyses shed light on early interactions of SARS-CoV-2 with the nasopharyngeal resident microbiota that might be helpful for developing microbiome-based diagnostics and therapeutics for this novel pandemic disease.

Bacterial microbiota in upper respiratory tract of COVID-19 and influenza patients.

The upper respiratory tract is inhabited by diverse range of commensal microbiota which plays a role in protecting the mucosal surface from pathogens. Alterations of the bacterial community from respiratory viral infections could increase the susceptibility to secondary infections and disease severities. We compared the upper respiratory bacterial profiles among Thai patients with influenza or COVID-19 by using 16S rDNA high-throughput sequencing based on MiSeq platform. The Chao1 richness was not significantly different among groups, whereas the Shannon diversity of Flu A and Flu B groups were significantly lower than Non-Flu & COVID-19 group. The beta diversity revealed that the microbial communities of influenza (Flu A and Flu B), COVID-19, and Non-Flu & COVID-19 were significantly different; however, the comparison of the community structure was similar between Flu A and Flu B groups. The bacterial classification revealed that Enterobacteriaceae was predominant in influenza patients, while Staphylococcus and Pseudomonas were significantly enriched in the COVID-19 patients. These implied that respiratory viral infections might be related to alteration of upper respiratory bacterial community and susceptibility to secondary bacterial infections. Moreover, the bacteria that observed in Non-Flu & COVID-19 patients had high abundance of Streptococcus, Prevotella, Veillonella, and Fusobacterium. This study provides the basic knowledge for further investigation of the relationship between upper respiratory microbiota and respiratory disease which might be useful for better understanding the mechanism of viral infectious diseases.

In-vitro cytokine production and nasopharyngeal microbiota composition in the early stage of COVID-19 infection.

BACKGROUND: To determine and compare nasopharyngeal microbiota (NM) composition, in vitro basal (Nil tube), provoked (Mitogen tube) production of cytokines at the early stage of COVID-19. METHODS: This cross-sectional study included 4 age and sex-matched study groups; group 1 (recovered COVID-19) (n = 26), group 2 (mild COVID-19) (n = 24), group 3 (severe COVID-19) (n = 25), and group 4 (healthy controls) (n = 25). The study parameters obtained from the COVID-19 (group 2, and 3) at the early phase of hospital admission. RESULTS: The results from the reaserch deoicted that the Mean ± SD age was 53.09 ± 14.51 years. Some of the in vitro cytokines production was significantly different between the study groups. Some of the findinggs on cytokines depicted a significant differences between study groups were interleukin (IL)-1ß Nil, IL-1ß Mitogen, and their subtraction (i.e Mitogen-Nil). Regarding IL-10, and IL-17a levels, Mitogen, and Mitogen-Nil tube production levels were significantly different between the groups. Surprisingly, most of these measures were lowest in the severe COVID-19 patients' group. Using discriminant analysis effect size (LEfSe), Taxa of NM with significant abundance was determined. About 20 taxa with an LDA score > 4 were identified as candidate biomarkers. Some of these taxa showed a significant correlation with IL-1ß and IL-10 Mitogen and Mitogen- Nil levels (R > 0.3 or < -0.3, p < 0.05). CONCLUSIONS: The findings of this perticular study regarting the early stage of COVID-19 showed that in vitro cytokines production, studies might be more useful than the ordinary cytokines' blood level measurement. Besides, the study identified some NM species that could be candidate biomarkers in managing this infection. However, further detailed studies are needed in these fields.

Oral Microbiome Alterations and SARS-CoV-2 Saliva Viral Load in Patients with COVID-19.

Bacterial-viral interactions in saliva have been associated with morbidity and mortality for respiratory viruses such as influenza and SARS-CoV. However, such transkingdom relationships during SARS-CoV-2 infection are currently unknown. Here, we aimed to elucidate the relationship between saliva microbiota and SARS-CoV-2 in a cohort of newly hospitalized COVID-19 patients and controls. We used 16S rRNA sequencing to compare microbiome diversity and taxonomic composition between COVID-19 patients (n = 53) and controls (n = 59) and based on saliva SARS-CoV-2 viral load as measured using reverse transcription PCR (RT-PCR). The saliva microbiome did not differ markedly between COVID-19 patients and controls. However, we identified significant differential abundance of numerous taxa based on saliva SARS-CoV-2 viral load, including multiple species within Streptococcus and Prevotella. IMPORTANCE Alterations to the saliva microbiome based on SARS-CoV-2 viral load indicate potential biologically relevant bacterial-viral relationships which may affect clinical outcomes in COVID-19 disease.

Comparison of 11 respiratory pathogens among hospitalized children before and during the COVID-19 epidemic in Shenzhen, China.

BACKGROUND: The effect of SARS-CoV-2 on existing respiratory pathogens in circulation remains uncertain. This study aimed to assess the impact of SARS-CoV-2 on the prevalence of respiratory pathogens among hospitalized children. METHODS: This study enrolled hospitalized children with acute respiratory infections in Shenzhen Children's Hospital from September to December 2019 (before the COVID-19 epidemic) and those from September to December 2020 (during the COVID-19 epidemic). Nasopharyngeal swabs were collected, and respiratory pathogens were detected using multiplex PCR. The absolute case number and detection rates of 11 pathogens were collected and analyzed. RESULTS: A total of 5696 children with respiratory tract infection received multiplex PCR examination for respiratory pathogens: 2298 from September to December 2019 and 3398 from September to December 2020. At least one pathogen was detected in 1850 (80.5%) patients in 2019, and in 2380 (70.0%) patients in 2020; the detection rate in 2020 was significantly lower than that in 2019.The Influenza A (InfA) detection rate was 5.6% in 2019, but 0% in 2020. The detection rates of Mycoplasma pneumoniae, Human adenovirus, and Human rhinovirus also decreased from 20% (460), 8.9% (206), and 41.8% (961) in 2019 to 1.0% (37), 2.1% (77), and 25.6% (873) in 2020, respectively. In contrast, the detection rates of Human respiratory syncytial virus, Human parainfluenza virus, and Human metapneumovirus increased from 6.6% (153), 9.9% (229), and 0.5% (12) in 2019 to 25.6% (873), 15.5% (530), and 7.2% (247) in 2020, respectively (p < 0.0001). CONCLUSIONS: Successful containment of seasonal influenza as a result of COVID-19 control measures will ensure we are better equipped to deal with future outbreaks of both influenza and COVID-19.Caused by virus competition, the detection rates of Human respiratory syncytial virus, Human parainfluenza virus, and Human metapneumovirus increased in Shenzhen,that reminds us we need to take further monitoring and preventive measures in the next epidemic season.

Distinct systemic and mucosal immune responses during acute SARS-CoV-2 infection.

Coordinated local mucosal and systemic immune responses following severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection either protect against coronavirus disease 2019 (COVID-19) pathologies or fail, leading to severe clinical outcomes. To understand this process, we performed an integrated analysis of SARS-CoV-2 spike-specific antibodies, cytokines, viral load and bacterial communities in paired nasopharyngeal swabs and plasma samples from a cohort of clinically distinct patients with COVID-19 during acute infection. Plasma viral load was associated with systemic inflammatory cytokines that were elevated in severe COVID-19, and also with spike-specific neutralizing antibodies. By contrast, nasopharyngeal viral load correlated with SARS-CoV-2 humoral responses but inversely with interferon responses, the latter associating with protective microbial communities. Potential pathogenic microorganisms, often implicated in secondary respiratory infections, were associated with mucosal inflammation and elevated in severe COVID-19. Our results demonstrate distinct tissue compartmentalization of SARS-CoV-2 immune responses and highlight a role for the nasopharyngeal microbiome in regulating local and systemic immunity that determines COVID-19 clinical outcomes.

Targeted Hybridization Capture of SARS-CoV-2 and Metagenomics Enables Genetic Variant Discovery and Nasal Microbiome Insights.

The emergence of novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genetic variants that may alter viral fitness highlights the urgency of widespread next-generation sequencing (NGS) surveillance. To profile genetic variants of the entire SARS-CoV-2 genome, we developed and clinically validated a hybridization capture SARS-CoV-2 NGS assay, integrating novel methods for panel design using double-stranded DNA (dsDNA) biotin-labeled probes, and built accompanying software. This test is the first hybrid capture-based NGS assay given Food and Drug Administration (FDA) emergency use authorization for detection of the SARS-CoV-2 virus. The positive and negative percent agreement (PPA and NPA, respectively) were defined in comparison to the results for an orthogonal real-time reverse transcription polymerase chain reaction (RT-PCR) assay (PPA and NPA, 96.7 and 100%, respectively). The limit of detection was established to be 800 copies/ml with an average fold enrichment of 46,791. Furthermore, utilizing the research-use-only analysis to profile the variants, we identified 55 novel mutations, including 11 in the functionally important spike protein. Finally, we profiled the full nasopharyngeal microbiome using metagenomics and found overrepresentation of 7 taxa and evidence of macrolide resistance in SARS-CoV-2-positive patients. This hybrid capture NGS assay, coupled with optimized software, is a powerful approach to detect and comprehensively map SARS-CoV-2 genetic variants for tracking viral evolution and guiding vaccine updates. IMPORTANCE This is the first FDA emergency-use-authorized hybridization capture-based next-generation sequencing (NGS) assay to detect the SARS-CoV-2 genome. Viral metagenomics and the novel hybrid capture NGS-based assay, along with its research-use-only analysis, can provide important genetic insights into SARS-CoV-2 and other emerging pathogens and improve surveillance and early detection, potentially preventing or mitigating new outbreaks. Better understanding of the continuously evolving SARS-CoV-2 viral genome and the impact of genetic variants may provide individual risk stratification, precision therapeutic options, improved molecular diagnostics, and population-based therapeutic solutions.

Mycotic infection prevalence among patients undergoing bronchoalveolar lavage with search of SARS-CoV-2 after two negative nasopharyngeal swabs.

The evidence that severe coronavirus disease 2019 (COVID-19) is a risk factor for development of mycotic respiratory infection with an increased mortality is rising. Immunosuppressed are among the most susceptible patients andAspergillusspecies is the most feared superinfection. In this study we evaluated mycotic isolation prevalence on bronchoalveolar lavage (BAL) of patients who underwent bronchoscopy in search of severe acute respiratory coronavirus 2 (SARS-CoV-2) RNA. Moreover, we described the clinical characteristics and main outcomes of these patients. We included 118 patients, 35.9% of them were immunosuppressed for different reasons: in 23.7% we isolated SARS-CoV-2 RNA, in 33.1% we identified at least one mycotic agent and both in 15.4%. On BAL we observed in three casesAspergillusspp, in six casesPneumocystisand in 32Candidaspp. The prevalence of significant mold infection was 29.3% and 70.7% of cases were false positive or clinically irrelevant infections. In-hospital mortality of patients with fungal infection was 15.3%. The most frequent computed tomography (CT) pattern, evaluated with the Radiological Society of North America consensus statement, among patients with a mycotic pulmonary infection was the atypical one (p< 0.0001). Mycotic isolation on BAL may be interpreted as an innocent bystander, but its identification could influence the prognosis of patients, especially in those who need invasive investigations during the COVID-19 pandemic; BAL plays a fundamental role in resolving clinical complex cases, especially in immunosuppressed patients independently from radiological features, without limiting its role in ruling out SARS-CoV-2 infection.

Signatures of COVID-19 Severity and Immune Response in the Respiratory Tract Microbiome.

Viral infection of the respiratory tract can be associated with propagating effects on the airway microbiome, and microbiome dysbiosis may influence viral disease. Here, we investigated the respiratory tract microbiome in coronavirus disease 2019 (COVID-19) and its relationship to disease severity, systemic immunologic features, and outcomes. We examined 507 oropharyngeal, nasopharyngeal, and endotracheal samples from 83 hospitalized COVID-19 patients as well as non-COVID patients and healthy controls. Bacterial communities were interrogated using 16S rRNA gene sequencing, and the commensal DNA viruses Anelloviridae and Redondoviridae were quantified by qPCR. We found that COVID-19 patients had upper respiratory microbiome dysbiosis and greater change over time than critically ill patients without COVID-19. Oropharyngeal microbiome diversity at the first time point correlated inversely with disease severity during hospitalization. Microbiome composition was also associated with systemic immune parameters in blood, as measured by lymphocyte/neutrophil ratios and immune profiling of peripheral blood mononuclear cells. Intubated patients showed patient-specific lung microbiome communities that were frequently highly dynamic, with prominence of Staphylococcus. Anelloviridae and Redondoviridae showed more frequent colonization and higher titers in severe disease. Machine learning analysis demonstrated that integrated features of the microbiome at early sampling points had high power to discriminate ultimate level of COVID-19 severity. Thus, the respiratory tract microbiome and commensal viruses are disturbed in COVID-19 and correlate with systemic immune parameters, and early microbiome features discriminate disease severity. Future studies should address clinical consequences of airway dysbiosis in COVID-19, its possible use as biomarkers, and the role of bacterial and viral taxa identified here in COVID-19 pathogenesis. IMPORTANCE COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection of the respiratory tract, results in highly variable outcomes ranging from minimal illness to death, but the reasons for this are not well understood. We investigated the respiratory tract bacterial microbiome and small commensal DNA viruses in hospitalized COVID-19 patients and found that each was markedly abnormal compared to that in healthy people and differed from that in critically ill patients without COVID-19. Early airway samples tracked with the level of COVID-19 illness reached during hospitalization, and the airway microbiome also correlated with immune parameters in blood. These findings raise questions about the mechanisms linking SARS-CoV-2 infection and other microbial inhabitants of the airway, including whether the microbiome might regulate severity of COVID-19 disease and/or whether early microbiome features might serve as biomarkers to discriminate disease severity.

Coinfection in SARS-CoV-2 Infected Children Patients.

INTRODUCTION: The aim of this study is to determine the coinfections with other respiratory pathogens in SARS-CoV-2 infected children patients in a pediatric unit in Istanbul. METHODOLOGY: This retrospective descriptive study was conducted in a 1000-bedded tertiary education and research hospital in Istanbul. All children hospitalized with the diagnosis of SARS-CoV-2 infection had been investigated for respiratory agents in nasopharyngeal secretions. Laboratory confirmation of SARS-CoV-2 and the other respiratory pathogens were performed using reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: A total of 209 hospitalized children with suspected SARS-CoV-2 infection between March 2020-May 2020 were enrolled in this study. Among 209 children, 93 (44.5%) were RT-PCR positive for SARS-CoV-2 infection, and 116 (55.5%) were RT-PCR negative. The most common clinical symptoms in all children with SARS-CoV-2 infection were fever (68.8%) and cough (57.0%). The other clinical symptoms in decreasing rates were headache (10.8%), myalgia (5.4%), sore throat (3.2%), shortness of breath (3.2%), diarrhea (2.2%) and abdominal pain in one child. In 7 (7.5%) patients with SARS-CoV-2 infection, coinfection was detected. Two were with rhinovirus/enterovirus, two were with Coronavirus NL63, one was with adenovirus, and one was with Mycoplasma pneumoniae. In one patient, two additional respiratory agents (rhinovirus/enterovirus and adenovirus) were detected. There was a significantly longer hospital stay in patients with coinfection (p = 0.028). CONCLUSIONS: Although the coinfection rate was low in SARS-CoV-2 infected patients in our study, we found coinfection as a risk factor for length of hospital stay in the coinfected patient group.

The human microbiome and COVID-19: A systematic review.

BACKGROUND: Human microbiotas are communities of microorganisms living in symbiosis with humans. They play an important role in the host immune response to respiratory viral infection. However, evidence on the human microbiome and coronavirus disease (COVID-19) relationship is insufficient. The aim of this systematic literature review was to evaluate existing evidence on the association between the microbiome and COVID-19 in humans and summarize these data in the pandemic era. METHODS: We conducted a systematic literature review on the association between the microbiome and COVID-19 in humans by searching PubMed, Embase, and the Cochrane Library, CINAHL, and Web of Science databases for articles in English published up to October 31, 2020. The results were analyzed qualitatively. This study is registered with PROSPERO (CRD42020195982). RESULTS: Of the 543 articles identified by searching databases, 16 in line with the research objectives were eligible for qualitative review: eight sampled the microbiome using stool, four using nasopharyngeal or throat swab, three using bronchoalveolar lavage fluid, and one using lung tissue. Fecal microbiome dysbiosis and increased opportunistic pathogens were reported in COVID-19 patients. Several studies suggested the dysbiosis in the lung microbiome of COVID-19 patients with an abundance of opportunistic pathogens using lower respiratory tract samples. The association between COVID-19 severity and the human microbiome remains uncertain. CONCLUSION: The human fecal and respiratory tract microbiome changed in COVID-19 patients with opportunistic pathogen abundance. Further research to elucidate the effect of alternation of the human microbiome in disease pathogenesis is warranted.

Influenza co-infection associated with severity and mortality in COVID-19 patients.

BACKGROUND: In COVID-19 patients, undetected co-infections may have severe clinical implications associated with increased hospitalization, varied treatment approaches and mortality. Therefore, we investigated the implications of viral and bacterial co-infection in COVID-19 clinical outcomes. METHODS: Nasopharyngeal samples were obtained from 48 COVID-19 patients (29% ICU and 71% non-ICU) and screened for the presence of 24 respiratory pathogens using six multiplex PCR panels. RESULTS: We found evidence of co-infection in 34 COVID-19 patients (71%). Influenza A H1N1 (n = 17), Chlamydia pneumoniae (n = 13) and human adenovirus (n = 10) were the most commonly detected pathogens. Viral co-infection was associated with increased ICU admission (r = 0.1) and higher mortality (OR 1.78, CI = 0.38-8.28) compared to bacterial co-infections (OR 0.44, CI = 0.08-2.45). Two thirds of COVID-19 critically ill patients who died, had a co-infection; and Influenza A H1N1 was the only pathogen for which a direct relationship with mortality was seen (r = 0.2). CONCLUSIONS: Our study highlights the importance of screening for co-infecting viruses in COVID-19 patients, that could be the leading cause of disease severity and death. Given the high prevalence of Influenza co-infection in our study, increased coverage of flu vaccination is encouraged to mitigate the transmission of influenza virus during the on-going COVID-19 pandemic and reduce the risk of severe outcome and mortality.

SARS-CoV-2 does not have a strong effect on the nasopharyngeal microbial composition.

The coronavirus disease 2019 (COVID-19) has rapidly spread around the world, impacting the lives of many individuals. Growing evidence suggests that the nasopharyngeal and respiratory tract microbiome are influenced by various health and disease conditions, including the presence and the severity of different viral disease. To evaluate the potential interactions between Severe Acute Respiratory Syndrome Corona 2 (SARS-CoV-2) and the nasopharyngeal microbiome. Microbial composition of nasopharyngeal swab samples submitted to the clinical microbiology lab for suspected SARS-CoV-2 infections was assessed using 16S amplicon sequencing. The study included a total of 55 nasopharyngeal samples from 33 subjects, with longitudinal sampling available for 12 out of the 33 subjects. 21 of the 33 subjects had at least one positive COVID-19 PCR results as determined by the clinical microbiology lab. Inter-personal variation was the strongest factor explaining > 75% of the microbial variation, irrespective of the SARS-CoV-2 status. No significant effect of SARS-CoV-2 on the nasopharyngeal microbial community was observed using multiple analysis methods. These results indicate that unlike some other viruses, for which an effect on the microbial composition was noted, SARS-CoV-2 does not have a strong effect on the nasopharynx microbial habitants.

Etiological and epidemiological characteristics of severe acute respiratory infection caused by multiple viruses and Mycoplasma pneumoniae in adult patients in Jinshan, Shanghai: A pilot hospital-based surveillance study.

BACKGROUND: Severe acute respiratory infection (SARI) results in a tremendous disease burden worldwide. Available research on active surveillance among hospitalized adult patients suffering from SARI in China is limited. This pilot study aimed to identify associated etiologies and describe the demographic, epidemiological and clinical profiles of hospitalized SARI patients aged over 16 years in Jinshan, Shanghai. METHODS: Active surveillance was conducted at 1 sentinel hospital in Jinshan district, Shanghai, from April 2017 to March 2018. Hospitalized SARI patients aged over 16 years old were enrolled, and nasopharyngeal swabs were collected within 24 hours of admission and tested for multiple respiratory viruses (including 18 common viruses) and Mycoplasma pneumoniae with real-time polymerase chain reaction. Demographic, epidemiological and clinical information was obtained from case report forms. RESULTS: In total, 397 SARI patients were enrolled; the median age was 68 years, and 194 (48.9%) patients were male. A total of 278 (70.0%) patients had at least one underlying chronic medical condition. The most frequent symptoms were cough (99.2%) and sputum production (88.4%). The median duration of hospitalization was 10 days. A total of 250 infection patients (63.0%) were positive for at least one pathogen, of whom 198 (49.9%) were positive for a single pathogen and 52 (13.1%) were positive for multiple pathogens. The pathogens identified most frequently were M. pneumoniae (23.9%, 95/397), followed by adenovirus (AdV) (11.6%, 46/397), influenza virus A/H3N2 (Flu A/H3N2) (11.1%, 44/397), human rhinovirus (HRhV) (8.1%, 32/397), influenza virus B/Yamagata (Flu B/Yamagata) (6.3%, 25/397), pandemic influenza virus A/H1N1 (Flu A/pH1N1) (4.0%, 16/397), parainfluenza virus (PIV) type 1 (2.0%, 8/397), human coronavirus (HCoV) type NL63 (2.0%, 8/397), HCoV 229E (1.5%, 6/397), HCoV HKU1 (1.5%, 6/397), PIV 3 (1.5%, 6/397), human metapneumovirus (HMPV) (1.5%, 6/397), PIV 4 (1.3%, 5/397), HCoV OC43 (1.0%, 4/397), influenza virus B/Victoria (Flu B/Victoria) (0.5%, 2/397), respiratory syncytial virus (RSV) type B (0.5%, 2/397), and human bocavirus (HBoV) (0.3%, 1/397). The seasonality of pathogen-confirmed SARI patients had a bimodal distribution, with the first peak in the summer and the second peak in the winter. Statistically significant differences were observed with respect to the rates of dyspnea, radiographically diagnosed pneumonia and the presence of at least one comorbidity in patients who were infected with only M. pneumoniae, AdV, HRhV, Flu A/H3N2, Flu A /pH1N1 or Flu B/Yamagata. The differences in the positivity rates of the above 6 pathogens among the different age groups were nonsignificant. CONCLUSIONS: M. pneumoniae, AdV and Flu A/H3N2 were the main pathogens detected in hospitalized SARI patients aged over 16 years old in Jinshan district, Shanghai. Our findings highlight the importance of sustained multipathogen surveillance among SARI patients in sentinel hospitals, which can provide useful information on SARI etiologies, epidemiology, and clinical characteristics.