Intracellular microbial shifts during COVID-19 infection and longitudinal recovery revealed by single-cell RNA sequencing

Host-microbe dynamics during SARS-CoV-2 Omicron variant infection and recovery remain poorly understood, particularly regarding intracellular microbial communities in immune cells. We performed single-cell RNA sequencing of 191,417 peripheral blood mononuclear cells (PBMCs) from 57 individuals (9 healthy, 24 Omicron-infected, 16 recently recovered, 8 long-recovered) using the BD Rhapsody platform. Microbial signatures identified with PathogenTrack revealed elevated alpha diversity in acutely infected and recently recovered groups, driven by opportunistic pathogens such as <i>Escherichia coli</i> and <i>Providentia stuartii.</i> In contrast, healthy and long-recovered individuals displayed commensal-dominated profiles, notably <i>Streptomyces sviceus</i>, indicative of restored balance. Functional analysis showed persistent microbial signatures, including <i>Clostridium botulinum's rpoB</i> in B cells of long-recovered individuals, broad expression of <i>E. coli</i> stress gene <i>sgrR</i>, and <i>Mycoplasma hyopneumoniae's rpsO</i> across 12 immune subsets. Notably, <i>S. sviceus dnaK</i> was exclusive to healthy monocytes. These results suggest enduring intracellular microbial influences, implicating them in long-COVID pathophysiology with potential therapeutic relevance.