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Pathogenic differences of Type 1 restriction-modification allele variants in experimental Listeria monocytogenes meningitis


Zbinden, Florian R; De Ste Croix, Megan; Grandgirard, Denis; Haigh, Richard D; Vacca, Irene; Zamudio, Roxana; Goodall, Emily C A; Stephan, Roger; Oggioni, Marco R; Leib, Stephen L (2020). Pathogenic differences of Type 1 restriction-modification allele variants in experimental Listeria monocytogenes meningitis. Frontiers in Cellular and Infection Microbiology, 10:590657.

Abstract

Background: L. monocytogenes meningoencephalitis has a mortality rate of up to 50% and neurofunctional sequelae are common. Type I restriction-modification systems (RMS) are capable of adding methyl groups to the host genome. Some contain multiple sequence recognition (hsdS) genes that recombine, resulting in distinct DNA methylation patterns and patterns of gene expression. These phenotypic switches have been linked to virulence and have recently been discovered in multiple clonal complexes of L. monocytogenes. In the present study, we investigated the significant of RMS on L. monocytogenes virulence during the acute phase of experimental meningitis. Methods: L. monocytogenes strains containing RMS systems were identified, and purified clones enriched for single hsdS alleles were isolated. In vivo, 11-day old Wistar rats were infected with an inoculum containing (a) one of 4 single RMS allele variants (A, B, C, D) treated with amoxicillin (AMX 50 mg/kg/dosis, q8h), (b) a mixture of all 4 variants with or without AMX treatment, or (c) different mixtures of 2 RMS allele variants. At selected time points after infection, clinical and inflammatory parameters, bacterial titers and brain damage were determined. Changes in the relative frequency of the occurring RMS alleles in the inoculum and in CSF or cerebellum of infected animals were analyzed by capillary electrophoresis. Results: We have identified a phase variable RMS locus within L. monocytogenes CC4 and generated stocks that stably expressed each of the possible hsdS genes within that loci. Generation of these allele variants (A, B, C, D) allowed us to determine the methylation pattern associated with each hsdS through SMRT sequencing. In vivo infections with these single allele variants revealed differences in disease severity in that C induced the worst clinical outcome and more pronounced hippocampal apoptosis; D showed the most pronounced weight loss and the highest bacterial titer in the cerebellum. A caused the least severe disease. Conclusion: We identified that L. monocytogenes expressing hsdS (A) causes less damage than when other hsdS genes are expressed. While expression of hsdSC and D worsened the outcome in L. monocytogenes meningitis. We also demonstrate a competitive advantage of variants C and B over variant A in this model. Phenotypical switching may therefore represent a mechanism of virulence regulation during the acute phase of CNS infections with L. monocytogenes.

Abstract

Background: L. monocytogenes meningoencephalitis has a mortality rate of up to 50% and neurofunctional sequelae are common. Type I restriction-modification systems (RMS) are capable of adding methyl groups to the host genome. Some contain multiple sequence recognition (hsdS) genes that recombine, resulting in distinct DNA methylation patterns and patterns of gene expression. These phenotypic switches have been linked to virulence and have recently been discovered in multiple clonal complexes of L. monocytogenes. In the present study, we investigated the significant of RMS on L. monocytogenes virulence during the acute phase of experimental meningitis. Methods: L. monocytogenes strains containing RMS systems were identified, and purified clones enriched for single hsdS alleles were isolated. In vivo, 11-day old Wistar rats were infected with an inoculum containing (a) one of 4 single RMS allele variants (A, B, C, D) treated with amoxicillin (AMX 50 mg/kg/dosis, q8h), (b) a mixture of all 4 variants with or without AMX treatment, or (c) different mixtures of 2 RMS allele variants. At selected time points after infection, clinical and inflammatory parameters, bacterial titers and brain damage were determined. Changes in the relative frequency of the occurring RMS alleles in the inoculum and in CSF or cerebellum of infected animals were analyzed by capillary electrophoresis. Results: We have identified a phase variable RMS locus within L. monocytogenes CC4 and generated stocks that stably expressed each of the possible hsdS genes within that loci. Generation of these allele variants (A, B, C, D) allowed us to determine the methylation pattern associated with each hsdS through SMRT sequencing. In vivo infections with these single allele variants revealed differences in disease severity in that C induced the worst clinical outcome and more pronounced hippocampal apoptosis; D showed the most pronounced weight loss and the highest bacterial titer in the cerebellum. A caused the least severe disease. Conclusion: We identified that L. monocytogenes expressing hsdS (A) causes less damage than when other hsdS genes are expressed. While expression of hsdSC and D worsened the outcome in L. monocytogenes meningitis. We also demonstrate a competitive advantage of variants C and B over variant A in this model. Phenotypical switching may therefore represent a mechanism of virulence regulation during the acute phase of CNS infections with L. monocytogenes.

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Item Type:Journal Article, refereed, original work
Communities & Collections:05 Vetsuisse Faculty > Institute of Food Safety and Hygiene
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Scopus Subject Areas:Life Sciences > Microbiology
Life Sciences > Immunology
Health Sciences > Microbiology (medical)
Health Sciences > Infectious Diseases
Uncontrolled Keywords:Immunology, Microbiology (medical), Microbiology, Infectious Diseases
Language:English
Date:30 October 2020
Deposited On:16 Feb 2021 16:27
Last Modified:18 Feb 2021 12:00
Publisher:Frontiers Research Foundation
ISSN:2235-2988
OA Status:Gold
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.3389/fcimb.2020.590657

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