# Biofilm-Facilitated Dissemination of Integrative Antibiotic Resistance Elements in Streptococcus pneumoniae

> **NIH NIH F31** · EMORY UNIVERSITY · 2021 · $46,036

## Abstract

I. PROJECT SUMMARY
Streptococcus pneumoniae (Spn) colonizes the human nasopharynx during early childhood. This opportunistic
pathogen causes ~15 million cases of invasive pneumococcal disease (IPD) and ~500,000 deaths in children
worldwide each year. While pneumococcal conjugate vaccine (PCV) development has impacted disease
incidence, the emergence and spread of antimicrobial resistance in Spn, an NIAID priority, has complicated IPD
treatment regimens. Consequently, the CDC and WHO list Spn amongst the antibiotic-resistant priority
pathogens. In particular, antibiotic resistance in Spn is increasingly due to acquisition of >20 kb, Tn916-like
integrative and conjugative elements (ICEs), most notably Tn2009, Tn6002, and Tn2010. The specific
mechanisms facilitating the wide spread of these large ICEs among Spn have not been elucidated. In other
bacteria, ICEs are transferred via transposition with the formation of circular intermediates and a conjugative
type 4 secretion system. Although conjugative genes are conserved within Spn ICEs, their roles in Spn ICE
transference have not been established. ICEs are unusually large elements for efficient in vitro transformation of
naturally competent Spn, which typically acquire ~2-6 kb DNA fragments. Accordingly, we confirmed that
competent Spn strains did not uptake ICE DNA (<10-9) under in vitro conditions. Yet, when two Spn strains were
co-inoculated in an ex vivo bioreactor system, forming a mixed biofilm on human nasopharyngeal cells, the
transference of the entire ICE occurred at frequencies up to 10-4, which likely recapitulates the scenario during
in vivo host colonization. In Aim 1, I propose to use the bioreactor system and a collection of various mutant
strains to decipher the mechanism(s) responsible for this five-orders of magnitude increase in dissemination of
large ICEs among naturally competent Spn strains. Specifically, we will assess the roles of ICE-encoded
conjugation genes, competence factors, and the DNA uptake apparatus. Conjugative and competence gene
expression will be measured via qRT-PCR and ICE circular intermediates, if present, will also be quantified using
qPCR. Conjugative protein expression will be investigated by Western blots. The role of extracellular membrane
vesicles (EVs) in the delivery of >20 kb ICE DNA will also be evaluated. Whether EVs carry ICE DNA will be
examined by ICE-targeting qPCR and purified biofilm-derived EVs will be tested via in vitro transformations. In
Aim 2, genomic characteristics of multi-drug resistance-conferring ICE elements will be defined in the global Spn
population. Using innovative bioinformatics tools and approaches, publicly available whole genome sequences
from global Spn isolates will be examined to identify new ICEs and novel ICE insertion sites in addition to
previously defined sites from a regional isolate collection. Analyses will be based on pre- and post-vaccine
periods, vaccine or non-vaccine serotypes, and clonal complexes. Pr...

## Key facts

- **NIH application ID:** 10235365
- **Project number:** 1F31AI154792-01A1
- **Recipient organization:** EMORY UNIVERSITY
- **Principal Investigator:** Brenda Stephanie Antezana
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $46,036
- **Award type:** 1
- **Project period:** 2021-09-01 → 2024-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10235365

## Citation

> US National Institutes of Health, RePORTER application 10235365, Biofilm-Facilitated Dissemination of Integrative Antibiotic Resistance Elements in Streptococcus pneumoniae (1F31AI154792-01A1). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10235365. Licensed CC0.

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