Nasopharyngeal microbiome reveals the prevalence of opportunistic pathogens in SARS-CoV-2 infected individuals and their association with host types
Authors: Yogesh Shouche
Rajesh Karyakarte
Abhishek Gupta
Avinash Sharma
Suvarna Joshi
Rashmita Das
Kunal Jani
Journal: Microbes and Infection (2021)
Discipline: IMMUNOLOGY
Keywords: Asymptomatic; Host types; Nasopharyngeal microbiome; SARS-CoV-2; Symptomatic.
Affiliations: DBT-National Centre for Cell Science, Pune, Maharashtra, 411007, India., Byramjee Jeejeebhoy Government Medical College, Pune, Maharashtra, 411001, India., DBT-National Centre for Cell Science, Pune, Maharashtra, 411007, India. Electronic address: .

Abstract:  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 &lt; 0.05) decreased in the SARS-CoV-2 infected individuals in comparison to non-infected individuals. Pairwise Wilcoxon test showed a significant (p &lt; 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. 

Retracted: No

PubPeer comments:
 We read the article from Gupta et al. with great interest. This article reports alterations in nasopharyngeal microbiota in SARS-CoV-2 infections. Nasopharyngeal samples are low biomass and well knownu00a0 to be affected by reagent contamination. Therefore, it is surprising the article does not describe any sequencing controls for both DNA extraction and PCR amplification step (1,2). It is not surprising to see some *Stenotrophomonas* and *Ralstonia* OTUs reported as key microbes in low biomass samples because they mainly originate from kit reagents (2,3,4). Fig3, f3 panel at least one sample seems to be mostly comprising of *Stenotrophomonas*? The colors are very difficult to differentiate. What are the DNA concentrations in all the samples (mean, median, minimum)? Do you see any correlation of specific bacterial abundance with DNA concn? The legend for fig3 says 11 taxa but figure has many more. Could the authors clarify this better in the legend section on how these were selected? In addition, it is difficult to confirm if OTUs related to *Acinetobacter* and *Pseudomonas* are of nasopharyngeal origin or from reagents i.e. the kitome. Sequencing of negative and positiveu00a0 controls with these samples could have allowed the authors to identify sources of these microbes. We believe that it is important for authors to address these issues because this relates to an important research area in the on-going pandemic with implications for global public health. Reference: 1. Olomu, I.N., Pena-Cortes, L.C., Long, R.A.u00a0et al.u00a0Elimination of u201ckitomeu201d and u201csplashomeu201d contamination results in lack of detection of a unique placental microbiome.u00a0BMC Microbiolu00a020,u00a0157 (2020). https: doi.org 10.1186 s12866-020-01839-y 2. Salter SJ, Cox MJ, Turek EM, Calus ST, Cookson WO, Moffatt MF, Turner P, Parkhill J, Loman NJ, Walker AW. Reagent and laboratory contamination can critically impact sequence-based microbiome analyses. BMC Biol. 2014;12:87. 3. Glassing A, Dowd SE, Galandiuk S, Davis B, Chiodini RJ. Inherent bacterial DNA contamination of extraction and sequencing reagents may affect interpretation of microbiota in low bacterial biomass samples. Gut Pathog. 2016;8:24. 4. Marsh RL, Nelson MT, Pope CE, Leach AJ, Hoffman LR, Chang AB, Smith-Vaughan HC. How low can we go? The implications of low bacterial load in respiratory microbiota studies. Pneumonia (Nathan). 2018;10:7 ,