Microbiome in the nasopharynx: Insights into the impact of COVID-19 severity.

Background: The respiratory tract harbors a variety of microbiota, whose composition and abundance depend on specific site factors, interaction with external factors, and disease. The aim of this study was to investigate the relationship between COVID-19 severity and the nasopharyngeal microbiome. Methods: We conducted a prospective cohort study in Mexico City, collecting nasopharyngeal swabs from 30 COVID-19 patients and 14 healthy volunteers. Microbiome profiling was performed using 16S rRNA gene analysis. Taxonomic assignment, classification, diversity analysis, core microbiome analysis, and statistical analysis were conducted using R packages. Results: The microbiome data analysis revealed taxonomic shifts within the nasopharyngeal microbiome in severe COVID-19. Particularly, we observed a significant reduction in the relative abundance of Lawsonella and Cutibacterium genera in critically ill COVID-19 patients (p < 0.001). In contrast, these patients exhibited a marked enrichment of Streptococcus, Actinomyces, Peptostreptococcus, Atopobium, Granulicatella, Mogibacterium, Veillonella, Prevotella_7, Rothia, Gemella, Alloprevotella, and Solobacterium genera (p < 0.01). Analysis of the core microbiome across all samples consistently identified the presence of Staphylococcus, Corynebacterium, and Streptococcus. Conclusions: Our study suggests that the disruption of physicochemical conditions and barriers resulting from inflammatory processes and the intubation procedure in critically ill COVID-19 patients may facilitate the colonization and invasion of the nasopharynx by oral microorganisms.

Longitudinal dynamics of the nasopharyngal microbiome in response to SARS-CoV-2 Omicron variant and HIV infection in Kenyan women and their infants.

The nasopharynx and its microbiota are implicated in respiratory health and disease. The interplay between viral infection and the nasopharyngeal microbiome is an area of increased interest and of clinical relevance. The impact of SARS-CoV-2, the etiological agent of the Coronavirus Disease 2019 (COVID-19) pandemic, on the nasopharyngeal microbiome, particularly among individuals living with HIV, is not fully characterized. Here we describe the nasopharyngeal microbiome before, during and after SARS-CoV-2 infection in a longitudinal cohort of Kenyan women (21 living with HIV and 14 HIV-uninfected) and their infants (18 HIV-exposed, uninfected and 18 HIV-unexposed, uninfected), followed between September 2021 through March 2022. We show using genomic epidemiology that mother and infant dyads were infected with the same strain of the SARS-CoV-2 Omicron variant that spread rapidly across Kenya. Additionally, we used metagenomic sequencing to characterize the nasopharyngeal microbiome of 20 women and infants infected with SARS-CoV-2, 6 infants negative for SARS-CoV-2 but experiencing respiratory symptoms, and 34 timepoint matched SARS-CoV-2 negative mothers and infants. Since individuals were sampled longitudinally before and after SARS-CoV-2 infection, we could characterize the short- and long-term impact of SARS-CoV-2 infection on the nasopharyngeal microbiome. We found that mothers and infants had significantly different microbiome composition and bacterial load (p-values <.0001). However, in both mothers and infants, the nasopharyngeal microbiome did not differ before and after SARS-CoV-2 infection, regardless of HIV-exposure status. Our results indicate that the nasopharyngeal microbiome is resilient to SARS-CoV-2 infection and was not significantly modified by HIV.

SARS-CoV-2 Infection is Associated with Age- and Gender-Specific Changes in the Nasopharyngeal Microbiome.

BACKGROUND: The recent Coronavirus Disease 2019 (COVID-19) pandemic has dramatically exposed our gap in understanding the pathogenesis of airborne infections. Within such a context, it is increasingly clear that the nasal cavity represents a critical checkpoint not only in the initial colonization phase but also in shaping any infectious sequelae. This is particularly relevant to COVID-19 in that the nasal cavity is characterized by high-level expression of the Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS-CoV-2) receptor, Angiotensin-Converting Enzyme 2 (ACE2), all along the respiratory tract. As part of the nasal mucosa, commensal microbes harbored by the nasal cavity likely are far more than just innocent bystanders in the interaction between SARS-CoV-2 and the local microenvironment. Yet the role of the qualitative composition of the nasal microbiome is unclear, as is its function, whether protective or not. METHODS: In this study, individuals undergoing SARS-CoV-2 molecular testing at the Hospital of Perugia (Italy) were recruited, with their residual material from the nasopharyngeal swabs being collected for microbiome composition analysis and short-chain fatty acid (SCFA) measurements (by 16S rRNA sequencing and gas chromatography-mass spectrometry), respectively. RESULTS: After stratification by age, gender, and viral load, the composition of the nasopharyngeal microbiome appeared to be influenced by age and gender, and SARS-CoV-2 infection further determined compositional changes. Notwithstanding this variability, a restricted analysis of female subjects-once SARS-CoV-2-infected-unraveled a shared expansion of Lachnospirales-Lachnospiraceae, irrespective of the viral load and age. This was associated with a reduction in the branched SCFA isobutanoic acid, as well as in the SCFAs with longer chains. CONCLUSIONS: Our results indicate that the nasopharyngeal microbiome is influenced by age, gender, and viral load, with consistent patterns of microbiome changes being present across specific groups. This may help in designing a personalized medicine approach in COVID-19 patients with specific patterns of nasal microbial communities.

Draft genome sequence of novel Candidatus Ornithobacterium hominis carrying antimicrobial resistance genes in Egypt.

BACKGROUND: Candidatus Ornithobacterium hominis (O. hominis), which was identified in nasopharyngeal swabs from Egypt, has been associated with respiratory disorders in humans. O. hominis, a recently identified member of the Flavobacteriaceae family, belongs to the largest family within the Bacteroidetes phylum. This family includes hundreds of species and 90 genera, including major human pathogens such as Capnocytophaga canimorsus and Elizabethkingia meningoseptica. Herein, we presented two draft genome assemblies of O. hominis that were extracted from metagenomic data using the Illumina sequencing method. The alignment of reads against the O. hominis genome was accomplished using BLASTN, and the reads with significant hits were extracted using Seqtk and assembled using SPAdes. The primary goal of this study was to obtain a more profound understanding of the genomic landscape of O. hominis, with an emphasis on identifying the associated virulence, antimicrobial genes, and distinct defense mechanisms to shed light on the potential role of O. hominis in human respiratory infections. RESULTS: The genome size was estimated to be 1.84 Mb, including 1,931,660 base pairs (bp), with 1,837 predicted coding regions and a G+C content of 35.62%. Genes encoding gliding motility, antibiotic resistance (20 genes), and the toxA gene were all included in the genome assembly. Gliding motility lipoproteins (GldD, GldJ, GldN, and GldH) and the gliding motility-associated ABC transporter substrate-binding protein, which acts as a crucial virulence mechanism in Flavobacterium species, were identified. The genome contained unique genes encoding proteins, such as the ParE1 toxin that defend against the actions of quinolone and other antibiotics. The cobalt-zinc-cadmium resistance gene encoding the protein CzcB, which is necessary for metal resistance, urease regulation, and colonization, was also detected. Several multidrug resistance genes encoding proteins were identified, such as MexB, MdtK, YheI, and VanC. CONCLUSION: Our study focused on identifying virulence factors, and antimicrobial resistance genes present in the core genome of O. hominis. These findings provide valuable insights into the potential pathogenicity and antibiotic susceptibility of O. hominis.

A cross-sectional study on the nasopharyngeal microbiota of individuals with SARS-CoV-2 infection across three COVID-19 waves in India.

Background: Multiple variants of the SARS-CoV-2 virus have plagued the world through successive waves of infection over the past three years. Independent research groups across geographies have shown that the microbiome composition in COVID-19 positive patients (CP) differs from that of COVID-19 negative individuals (CN). However, these observations were based on limited-sized sample-sets collected primarily from the early days of the pandemic. Here, we study the nasopharyngeal microbiota in COVID-19 patients, wherein the samples have been collected across the three COVID-19 waves witnessed in India, which were driven by different variants of concern. Methods: The nasopharyngeal swabs were collected from 589 subjects providing samples for diagnostics purposes at the Centre for Cellular and Molecular Biology (CSIR-CCMB), Hyderabad, India and subjected to 16s rRNA gene amplicon - based sequencing. Findings: We found variations in the microbiota of symptomatic vs. asymptomatic COVID-19 patients. CP showed a marked shift in the microbial diversity and composition compared to CN, in a wave-dependent manner. Rickettsiaceae was the only family that was noted to be consistently depleted in CP samples across the waves. The genera Staphylococcus, Anhydrobacter, Thermus, and Aerococcus were observed to be highly abundant in the symptomatic CP patients when compared to the asymptomatic group. In general, we observed a decrease in the burden of opportunistic pathogens in the host microbiota during the later waves of infection. Interpretation: To our knowledge, this is the first analytical cross-sectional study of this scale, which was designed to understand the relation between the evolving nature of the virus and the changes in the human nasopharyngeal microbiota. Although no clear signatures were observed, this study shall pave the way for a better understanding of the disease pathophysiology and help gather preliminary evidence on whether interventions to the host microbiota can help in better protection or faster recovery.

Specific nasopharyngeal Corynebacterium strains serve as gatekeepers against SARS-CoV-2 infection.

The SARS-CoV-2 virus is still causing a worldwide problem. The virus settles primarily on the nasal mucosa, and the infection and its course depend on individual susceptibility. Our aim was to investigate the nasopharynx composition's role in the individual susceptibility. During the first phase of SARS-CoV-2 pandemic, nasopharyngeal microbiome samples of close contact unvaccinated patients were investigated by 16S rRNA analysis and by culturing. The whole genome of cultured Corynebacteria was sequenced. The relative expression of ACE2, TMPRSS2, and cathepsin L on Caco-2 cells and the strength of S1-ACE2 binding were determined in the presence of Corynebacteria. From 55 close contacts exposed to identical SARS-CoV-2 exposure, 26 patients became infected and 29 remained uninfected. The nasopharyngeal microbiome analysis showed significantly higher abundance of Corynebacteria in uninfected group. Corynebacterium accolens could be cultivated only from uninfected individuals and Corynebacterium propinquum from both infected and uninfected. Corynebacteria from uninfected patient significantly reduced the ACE2 and cathepsin L expression. C. accolens significantly reduced the TMPRSS2 expression compared to other Corynebacteria. Furthermore, Corynebacterium spp. weakened the binding of the S1-ACE2. Most C. accolens isolates harbored the TAG lipase LipS1 gene. Based on these results, the presence of Corynebacterium spp. in the nasopharyngeal microbiota, especially C. accolens strains, could reduce the individual susceptibility to SARS-CoV-2 infection by several mechanisms: by downregulation the ACE2, the TMPRSS2 receptors, and cathepsin L in the host; through the inhibition of S1-ACE2 binding; and lipase production. These results suggest the use of C. accolens strains as probiotics in the nasopharynx in the future.

Analysis of the nasopharyngeal microbiome and respiratory pathogens in COVID-19 patients from Saudi Arabia.

BACKGROUND: Infection with SARS-CoV-2 may perturb normal microbiota, leading to secondary infections that can complicate the viral disease. The aim of this study was to probe the alteration of nasopharyngeal (NP) microbiota in the context of SARS-CoV-2 infection and obesity and to identify other respiratory pathogens among COVID-19 cases that may affect patients' health. METHODS: A total of 107 NP swabs, including 22 from control subjects and 85 from COVID-19 patients, were processed for 6S amplicon sequencing. The respiratory pathogens causing secondary infections were identified by RT-PCR assay, using a kit that contained specific primers and probes combinations to amplify 33 known respiratory pathogens. RESULTS: No significant (p > 0.05) difference was observed in the alpha and beta diversity analysis, but specific taxa differed significantly between the control and COVID-19 patient groups. Genera of Sphingomonas, Kurthia, Microbacterium, Methylobacterium, Brevibacillus, Bacillus, Acinetobacter, Lactococcus, and Haemophilus was significantly abundant (p < 0.05) in COVID-19 patients compared with a healthy control group. Staphylococcus was found in relatively high abundance (35.7 %) in the COVID-19 patient groups, mainly those treated with antibiotics. A relatively high percentage of Streptococcus was detected in COVID-19 patient groups with obesity or other comorbidities. Respiratory pathogens, including Staphylococcus aureus, Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, and Salmonella species, along with Pneumocystis jirovecii fungal species were detected by RT-PCR mainly in the COVID-19 patients. Klebsiella pneumoniae was commonly found in most of the samples from the control and COVID-19 patients. Four COVID-19 patients had viral coinfections with human adenovirus, human rhinovirus, enterovirus, and human parainfluenza virus 1. CONCLUSIONS: Overall, no substantial difference was observed in the predominant NP bacterial community, but specific taxa were significantly changed between the healthy control and COVID-19 patients. Comparatively, an increased number of respiratory pathogens were identified in COVID-19 patients, and NP colonization by K. pneumoniae was probably occurring in the local population.

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.

Temporal Dynamics of the Nasopharyngeal Microbiome and its Relationship with Childhood Asthma Exacerbation.

Despite distinct nasopharyngeal microbiome (NPM) profiles between asthmatics and healthy subjects, little is known about the NPM dynamics and its relation to childhood asthma exacerbation (AE). We investigated NPM changes by longitudinally collecting 135 flocked nasopharyngeal swabs (FNPSs) from 33 school-age asthmatic children at six time points (2 to 4-week intervals) from September to December 2017 in Hong Kong. Subjects were categorized into AE and stable asthma (AS) groups according to whether they experienced any exacerbation during follow-up. One-off FNPSs from nine nonasthmatic children were included as controls. Microbiota profiles were analyzed using 16S rRNA gene sequencing. All 144 NPMs were classified into six microbiome profile groups (MPGs), each dominated by Moraxella, Corynebacterium 1, Dolosigranulum, Staphylococcus, Streptococcus, or Anoxybacillus. The microbial diversity and compositions of NPM in exacerbation samples were different from both baseline samples and those from healthy controls. Moraxella and Dolosigranulum-dominated NPM exhibited high temporal stability revealed by MPG transition analysis. NPM diversity decreased whereas microbial composition remained similar over time. The relative abundances of Moraxella increased while Corynebacterium 1, Anoxybacillus, and Pseudomonas decreased longitudinally. However, these temporal patterns did not differ between AE and AS groups, suggesting that short-term dynamic patterns were not sufficient to predict AE occurrence. Asthmatic NPM underwent Moraxella expansion during AE and presented a high microbiome resilience (recovery potential) after AE resolution. Microbial pathways involved in methane, ketone bodies, and vitamin B3 metabolisms were enhanced during AE and primarily contributed by Moraxella. IMPORTANCE Evidence on the dynamic changes of NPM in asthmatic patients remains limited. Here, we present that asthmatic NPMs deviating from a healthy status still showed resilience after disturbance. Our data imply from a longitudinal perspective that Moraxella increase is closely related to AE occurrence. The finding of functional dysbiosis (imbalance) during AE offers a plausible explanation for the known association between nasopharyngeal Moraxella expansion and increased AE risk. This work serves as a basis for future long-term prospective studies leveraging multiomics approaches to elucidate the temporal association between NPM and pediatric AE.

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.

Nasopharyngeal microbiome reveals the prevalence of opportunistic pathogens in SARS-CoV-2 infected individuals and their association with host types.

The novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is causing a severe global health emergency owing to its highly infectious nature. Although the symptoms of SARS-CoV-2 are well known but its impact on nasopharyngeal microbiome is poorly studied. The present cross-sectional study was intended to understand the perturbation in the nasopharyngeal microbiome composition within the infected (n = 63) and non-infected (n = 26) individuals using 16S rRNA gene based targeted amplicon sequencing and their association with host types and the prevalence of opportunistic pathogens at the stage of infection. The results confirmed that number of OTUs were significantly (p < 0.05) decreased in the SARS-CoV-2 infected individuals in comparison to non-infected individuals. Pairwise Wilcoxon test showed a significant (p < 0.05) increase in the abundance of Proteobacteria in infected individuals compared to non-infected ones and vice-versa for Fusobacteria and Bacteroidetes. Similarity percentage (SIMPER) analysis showed the increment in the abundance of opportunistic pathogens (Haemophilus, Stenotrophomonas, Acinetobacter, Moraxella, Corynebacterium 1, Gemella, Ralstonia, and Pseudomonas) involved in secondary infection. Furthermore, this study highlighted the microbial community structure of individuals within and across the families. In this study, we also performed the assesment of microbiome associated with host types (age and genders) and COVID-19 conditions (symptomatic and asymptomatic). The data suggested that the host types/conditions during the COVID-19 infection are potential factors in enrichment of specific bacterial communities in upper respiratory tract.

Characterization of longitudinal nasopharyngeal microbiome patterns in maternally HIV-exposed Zambian infants.

Background: Previous studies of infants born to HIV-positive mothers have linked HIV exposure to poor outcomes from gastrointestinal and respiratory illnesses, and to overall increased mortality rates. The mechanism behind this is unknown, but it is possible that differences in the nasopharyngeal (NP) microbiome between infants who are HIV-unexposed or HIV-exposed could play a role in perpetuating some outcomes. Methods: We conducted a longitudinal analysis of 170 NP swabs of healthy infants who are HIV-exposed (n=10) infants and their HIV(+) mothers, and infants who are HIV-unexposed, uninfected (HUU; n=10) .and their HIV(-) mothers. These swabs were identified from a sample library collected in Lusaka, Zambia between 2015 and 2016. Using 16S rRNA gene sequencing, we characterized the maturation of the microbiome over the first 14 weeks of life to determine what quantifiable differences exist between HIV-exposed and HUU infants, and what patterns are reflected in the mothers' NP microbiomes. Results: In both HIV-exposed and HUU infants, Staphylococcus and Corynebacterium began as primary colonizers of the NP microbiome but were in time replaced by Dolosigranulum, Streptococcus, Moraxella and Haemophilus. When evaluating the interaction between HIV exposure status and time of sampling among infants, the microbe Staphylococcus haemolyticus showed a distinctive high association with HIV exposure at birth. When comparing infants to their mothers with paired analyses, HIV-exposed infants' NP microbiome composition was only slightly different from their HIV(+) mothers at birth or 14 weeks, including in their carriage of S. pneumoniae, H. influenzae, and S. haemolyticus. Conclusions: Our analyses indicate that the HIV-exposed infants in our study exhibit subtle differences in the NP microbial composition throughout the sampling interval. Given our results and the sampling limitations of our study, we believe that further research must be conducted in order to confidently understand the relationship between HIV exposure and infants' NP microbiomes.

Nasopharyngeal Dysbiosis Precedes the Development of Lower Respiratory Tract Infections in Young Infants, a Longitudinal Infant Cohort Study.

Background: Infants suffering from lower respiratory tract infections (LRTIs) have distinct nasopharyngeal (NP) microbiome profiles that correlate with severity of disease. Whether these profiles precede the infection or are a consequence of it, is unknown. In order to answer this question, longitudinal studies are needed. Methods: We conducted a retrospective analysis of NP samples collected in a longitudinal birth cohort study of Zambian mother-infant pairs. Samples were collected every two weeks from 1-week through 14-weeks of age. Ten of the infants in the cohort who developed LRTI were matched 1:3 with healthy comparators. We completed 16S rRNA gene sequencing on the samples each of these infants contributed and compared the NP microbiome of the healthy infants to infants who developed LRTI. Results: The infant NP microbiome maturation was characterized by transitioning from Staphylococcus dominant to respiratory-genera dominant profiles during the first three months of life, similar to what is described in the literature. Interestingly, infants who developed LRTI had distinct NP microbiome characteristics before infection, in most cases as early as the first week of life. Their NP microbiome was characterized by the presence of Novosphingobium, Delftia, high relative abundance of Anaerobacillus, Bacillus, and low relative abundance of Dolosigranulum, compared to the healthy controls. Mothers of infants with LRTI also had low relative abundance of Dolosigranulum in their baseline samples compared to mothers of infants that did not develop an LRTI. Conclusions: Our results suggest that specific characteristics of the NP microbiome precede LRTI in young infants and may be present in their mothers as well. Early dysbiosis may play a role in the causal pathway leading to LRTI or could be a marker of underlying immunological, environmental, or genetic characteristics that predispose to LRTI.

Association between the nasopharyngeal microbiome and metabolome in patients with COVID-19.

SARS-CoV-2, the causative agent for COVID-19, infect human mainly via respiratory tract, which is heavily inhabited by local microbiota. However, the interaction between SARS-CoV-2 and nasopharyngeal microbiota, and the association with metabolome has not been well characterized. Here, metabolomic analysis of blood, urine, and nasopharyngeal swabs from a group of COVID-19 and non-COVID-19 patients, and metagenomic analysis of pharyngeal samples were used to identify the key features of COVID-19. Results showed lactic acid, l-proline, and chlorogenic acid methyl ester (CME) were significantly reduced in the sera of COVID-19 patients compared with non-COVID-19 ones. Nasopharyngeal commensal bacteria including Gemella morbillorum, Gemella haemolysans and Leptotrichia hofstadii were notably depleted in the pharynges of COVID-19 patients, while Prevotella histicola, Streptococcus sanguinis, and Veillonella dispar were relatively increased. The abundance of G. haemolysans and L. hofstadii were significantly positively associated with serum CME, which might be an anti-SARS-CoV-2 bacterial metabolite. This study provides important information to explore the linkage between nasopharyngeal microbiota and disease susceptibility. The findings were based on a very limited number of patients enrolled in this study; a larger size of cohort will be appreciated for further investigation.

Comparative Analysis of Microbiome in Nasopharynx and Middle Ear in Young Children With Acute Otitis Media.

Acute otitis media (AOM) is the most common pediatric infection for which antibiotics are prescribed in the United States. The role of the respiratory tract microbiome in pathogenesis and immune modulation of AOM remains unexplored. We sought to compare the nasopharyngeal (NP) microbiome of children 1 to 3 weeks prior to onset of AOM vs. at onset of AOM, and the NP microbiome with the microbiome in middle ear (ME). Six children age 6 to 24 months old were studied. Nasal washes (NW) were collected at healthy visits 1 to 3 weeks prior to AOM and at onset of AOM. The middle ear fluids (MEF) were collected by tympanocentesis at onset of AOM. Samples were stored in Trizol reagents or phosphate-buffered saline (PBS) at -80°C until use. The microbiome was characterized by 16S rRNA gene sequencing. Taxonomic designations and relative abundance of bacteria were determined using the RDP classifier tool through QIIME. Cumulative sum scaling normalization was applied before determining bacterial diversity and abundance. Shannon diversity index was calculated in Microsoft excel. The relative abundance of each bacteria species was compared via Mann-Whitney U test. We found that the NW microbiome of children during healthy state or at baseline was more diverse than microbiome during AOM. At AOM, no significant difference in microbiome diversity was found between NW and MEF, although some bacteria species appear to differ in MEF than in NW. The microbiome of samples stored in PBS had significant greater diversity than samples stored in Trizol reagent.

Antibiotic treatment in feedlot cattle: a longitudinal study of the effect of oxytetracycline and tulathromycin on the fecal and nasopharyngeal microbiota.

BACKGROUND: Beef cattle in North America frequently receive an antibiotic injection after feedlot placement to control and manage bovine respiratory disease. The potential collateral effect of these antibiotics on the bovine microbiome is largely unknown. Therefore, we determined the longitudinal impact of two commonly administered veterinary antibiotics, oxytetracycline and tulathromycin, on the fecal and nasopharyngeal (NP) microbiota of beef cattle that were transported to a feedlot. We also report the effect these antibiotics have on several antibiotic resistance determinants in both the fecal and NP microbiome. RESULTS: Oxytetracycline and tulathromycin perturbation of the bovine fecal and NP microbiota was greatest at days 2 and 5. Although the NP microbiota of the tulathromycin-treated cattle had recovered by day 12, the NP microbiota of the oxytetracycline-treated group remained altered through day 34. Overall, the NP microbiota appeared to be more sensitive to antibiotic treatment than the fecal microbiota. Members of the bacterial Microbacteriaceae family were most notably affected by antibiotic administration in the NP microbiota. Both antibiotics protected against Pasteurella spp. in the nasopharynx at days 2 and 5. Despite very similar diets at both locations, the largest shift in the fecal and NP microbiota occurred after transport to the feedlot (P < 0.05). Antibiotic resistance determinants in the NP microbiome were also affected more strongly by antibiotic treatment than those in the fecal microbiome. Oxytetracycline increased the proportion of erm(X), sul2, tet(H), tet(M), and tet(W) in NP samples and tet(M) and tet(W) in fecal samples, at day 12 (P < 0.05). The effect of tulathromycin on the relative abundance of resistance genes in the NP microbiome was greatest at day 34 as erm(X), sul2, and tet(M) were enriched (P < 0.05). CONCLUSIONS: Administration of a single injection of oxytetracycline and tulathromycin resulted in significant changes in the NP and fecal microbiota during the first 5 days after treatment. Antibiotic treatment also increased the relative abundance of several antibiotic resistance determinants in the fecal and NP microbiome at either day 12 or 34.

Strain Level Streptococcus Colonization Patterns during the First Year of Life.

Pneumococcal pneumonia has decreased significantly since the implementation of the pneumococcal conjugate vaccine (PCV), nevertheless, in many developing countries pneumonia mortality in infants remains high. We have undertaken a study of the nasopharyngeal (NP) microbiome during the first year of life in infants from The Philippines and South Africa. The study entailed the determination of the Streptococcus sp. carriage using a lytA qPCR assay, whole metagenomic sequencing, and in silico serotyping of Streptococcus pneumoniae, as well as 16S rRNA amplicon based community profiling. The lytA carriage in both populations increased with infant age and lytA+ samples ranged from 24 to 85% of the samples at each sampling time point. We next developed informatic tools for determining Streptococcus community composition and pneumococcal serotype from metagenomic sequences derived from a subset of longitudinal lytA-positive Streptococcus enrichment cultures from The Philippines (n = 26 infants, 50% vaccinated) and South African (n = 7 infants, 100% vaccinated). NP samples from infants were passaged in enrichment media, and metagenomic DNA was purified and sequenced. In silico capsular serotyping of these 51 metagenomic assemblies assigned known serotypes in 28 samples, and the co-occurrence of serotypes in 5 samples. Eighteen samples were not typeable using known serotypes but did encode for capsule biosynthetic cluster genes similar to non-encapsulated reference sequences. In addition, we performed metagenomic assembly and 16S rRNA amplicon profiling to understand co-colonization dynamics of Streptococcus sp. and other NP genera, revealing the presence of multiple Streptococcus species as well as potential respiratory pathogens in healthy infants. A range of virulence and drug resistant elements were identified as circulating in the NP microbiomes of these infants. This study revealed the frequent co-occurrence of multiple S. pneumoniae strains along with Streptococcus sp. and other potential pathogens such as S. aureus in the NP microbiome of these infants. In addition, the in silico serotype analysis proved powerful in determining the serotypes in S. pneumoniae carriage, and may lead to developing better targeted vaccines to prevent invasive pneumococcal disease (IPD) in these countries. These findings suggest that NP colonization by S. pneumoniae during the first years of life is a dynamic process involving multiple serotypes and species.

The mechanism or mechanisms driving atopic asthma initiation: The infant respiratory microbiome moves to center stage.

Developments over the last 5 to 10 years, principally from studies on comprehensively phenotyped prospective birth cohorts, have highlighted the important role of viral respiratory tract infections during infancy and early childhood, particularly those occurring against a background of pre-existing sensitization to perennial aeroallergens, in driving the development of early-onset atopic asthma. Although debate surrounding the mechanism or mechanisms governing this causal pathway remains intense, demonstration of the capacity of pretreatment with anti-IgE antibody to blunt seasonal virus-associated asthma exacerbations in children provides strong support for the underlying concept. However, emerging data appear set to further complicate this picture. Notably, a combination of culture-based studies and complementary population-wide bacterial metagenomic data suggests that parallel host-bacteria interactions during infancy might play an additional role in modulating this causal pathway, as well as contributing independently to pathogenesis. These and related issues surrounding development of immune competence during the crucial early postnatal period, when these pathways are maximally active, are discussed below.