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Crohn's Disease-Associated Bacteria Tolerate Antibiotics

By Labmedica International staff writers
Posted on 26 Nov 2019
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Image: Axio Vert.A1 inverted microscope (Photo courtesy of Carl Zeiss)
Image: Axio Vert.A1 inverted microscope (Photo courtesy of Carl Zeiss)
Crohn's disease is a chronic disease that causes inflammation and irritation in the digestive tract. The disease is characterized by an imbalance in the intestinal microbiome. In particular, adherent-invasive Escherichia coli (AIEC) strains have been implicated in the disease.

The diversity of virulence factors displayed by multiple AIEC strains suggests that members of this pathovar have evolved different strategies to colonize their hosts. AIEC ability to persist, and in some cases replicate within macrophages is particularly intriguing. A reference strain for this pathovar, AIEC LF82, forms micro-colonies within phagolysosomes, an environment that prevents commensal E. coli multiplication.

Microbiologists at the Université Paris Sciences et Lettres (Paris, France) and their associates used single-cell analysis, genetic dissection and mathematical models to monitor the growth status and cell cycle regulation of intracellular AIEC LF82. The investigators infected cells resulting in the observation of three LF82 bacteria per macrophage on average at 1 hour. Imaging was performed on an inverted Zeiss Axio Imager (Jena, Germany) with a spinning disk CSU W1 (Yokogawa, Tokyo, Japan).

The team also performed antibiotic challenge and viable bacterial count using the gentamycin protection assay, fluorescence quantification, live and dead assays using the Live and Dead BacLight Viability kit (Thermo Fisher Scientific, Waltham, MA, USA), measurement of gene expression by RT-qPCR using a MyiQ real-time qPCR machine (Bio-Rad, Hercules, CA, USA).

The scientists reported that they found that within macrophages, bacteria may replicate or undergo non-growing phenotypic switches. This switch results from stringent response firing immediately after uptake by macrophages or at later stages, following genotoxic damage and SOS induction during intracellular replication. Importantly, non-growers resist treatment with various antibiotics. Thus, intracellular challenges induce AIEC LF82 phenotypic heterogeneity and non-growing bacteria that could provide a reservoir for antibiotic-tolerant bacteria responsible for relapsing infections. Importantly, non-growers resist treatment with various antibiotics.

The authors concluded that intracellular challenges induce AIEC LF82 phenotypic heterogeneity and non-growing bacteria that could provide a reservoir for antibiotic-tolerant bacteria responsible for relapsing infections. The study was published on November 14, 2019 in the journal PLOS Pathogens.

Related Links:
Université Paris Sciences et Lettres
Carl Zeiss
Yokogawa
Thermo Fisher Scientific
Bio-Rad



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