Laboratory strains of Bacillus anthracis exhibit pervasive alteration in expression of proteins related to sporulation under laboratory conditions relative to genetically related wild strains

PLOS ONE, Dec 2018

The spore forming pathogen Bacillus anthracis is the etiologic agent of anthrax in humans and animals. It cycles through infected hosts as vegetative cells and is eventually introduced into the environment where it generates an endospore resistant to many harsh conditions. The endospores are subsequently taken up by another host to begin the next cycle. Outbreaks of anthrax occur regularly worldwide in wildlife and livestock, and the potential for human infection exists whenever humans encounter infected animals. It is also possible to encounter intentional releases of anthrax spores, as was the case in October 2001. Consequently, it is important to be able to rapidly establish the provenance of infectious strains of B. anthracis. Here, we compare protein expression in seven low-passage wild isolates and four laboratory strains of B. anthracis grown under identical conditions using LC-MS/MS proteomic analysis. Of the 1,023 total identified proteins, 96 had significant abundance differences between wild and laboratory strains. Of those, 28 proteins directly related to sporulation were upregulated in wild isolates, with expression driven by Spo0A, CodY, and AbrB/ScoC. In addition, we observed evidence of changes in cell division and fatty acid biosynthesis between the two classes of strains, despite being grown under identical experimental conditions. These results suggest wild B. anthracis cells are more highly tuned to sporulate than their laboratory cousins, and this difference should be exploited as a method to differentiate between laboratory and low passage wild strains isolated during an anthrax outbreak. This knowledge should distinguish between intentional releases and exposure to strains in nature, providing a basis for the type of response by public health officials and investigators.

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Laboratory strains of Bacillus anthracis exhibit pervasive alteration in expression of proteins related to sporulation under laboratory conditions relative to genetically related wild strains

December Laboratory strains of Bacillus anthracis exhibit pervasive alteration in expression of proteins related to sporulation under laboratory conditions relative to genetically related wild strains Owen P. LeiserID 0 1 Jason K. Blackburn 1 Ted L. Hadfield 1 Helen W. Kreuzer 0 1 David S. Wunschel 0 1 Cindy J. Bruckner-Lea 0 1 0 Chemical and Biological Signature Science, Pacific Northwest National Laboratory, Richland, Washington, United States of America, 2 Emerging Pathogens Institute, University of Florida, Gainesville, Florida, United States of America, 3 Spatial Epidemiology & Ecology Research Laboratory, Department of Geography, University of Florida , Gainesville, Florida , United States of America 1 Editor: Adam Driks, Loyola University Chicago , UNITED STATES - Funding: This work was supported by internal laboratory directed research and development funding through PNNL to OPL, HWK, CJBL, and DSW in collaboration with JKB and TLH. The funder had no role in study design, data collection The spore forming pathogen Bacillus anthracis is the etiologic agent of anthrax in humans and animals. It cycles through infected hosts as vegetative cells and is eventually introduced into the environment where it generates an endospore resistant to many harsh conditions. The endospores are subsequently taken up by another host to begin the next cycle. Outbreaks of anthrax occur regularly worldwide in wildlife and livestock, and the potential for human infection exists whenever humans encounter infected animals. It is also possible to encounter intentional releases of anthrax spores, as was the case in October 2001. Consequently, it is important to be able to rapidly establish the provenance of infectious strains of B. anthracis. Here, we compare protein expression in seven low-passage wild isolates and four laboratory strains of B. anthracis grown under identical conditions using LC-MS/MS proteomic analysis. Of the 1,023 total identified proteins, 96 had significant abundance differences between wild and laboratory strains. Of those, 28 proteins directly related to sporulation were upregulated in wild isolates, with expression driven by Spo0A, CodY, and AbrB/ ScoC. In addition, we observed evidence of changes in cell division and fatty acid biosynthesis between the two classes of strains, despite being grown under identical experimental conditions. These results suggest wild B. anthracis cells are more highly tuned to sporulate than their laboratory cousins, and this difference should be exploited as a method to differentiate between laboratory and low passage wild strains isolated during an anthrax outbreak. This knowledge should distinguish between intentional releases and exposure to strains in nature, providing a basis for the type of response by public health officials and investigators. and analysis, decision to publish, or preparation of the manuscript. Introduction Bacteria growing in the laboratory experience dramatically different selective pressures than those found in the environment. Bacillus anthracis cells respond to conditions outside of mammalian hosts by forming a metabolically dormant endospore, capable of surviving extended periods of harsh conditions [ 1 ]. Cells must overcome interspecies competition and nutrient-limiting conditions to infect new hosts. In contrast to growth in the environment, growth conditions in the laboratory are often stable, with abundant nutrients?conditions tailored for optimum growth. Intuitively, adaptation to different selective pressures between laboratory and environmental conditions will result in measurable genotypic or phenotypic changes. Indeed, long-term evolution has been studied extensively in an ongoing experiment in Escherichia coli [ 2?5 ], in which cultures have been maintained for over 60,000 generations with pervasive genomic and phenotypic changes observed. Additionally, Mikkola and Kurland [6], Eydallin et al. [ 7 ] and Saxer et al. [ 8 ] examined genomic signatures of adaptation of wild E. coli to laboratory conditions. However, far less is known about the mechanisms of wild pathogen adaptation to laboratory conditions: Sjo?din et al. [ 9 ] investigated naturally occurring and laboratory strains of Francisella tularensis using whole-genome sequencing, and Leiser et al. [ 10 ] investigated the proteomic and genomic indicators of wild Y. pestis adaptation to laboratory conditions. Sjo?din et al. examined very closely related strains of F. tularensis [ 9 ], and the laboratory-adapted strains of Y. pestis examined by Leiser et al. [ 10 ] were direct descendants of the respective starting wild strains. Systemic differences in gene/protein expression between wild and laboratoryadapted strains can be elucidated using genetically similar (same clade) but distinct (wild type or laboratory adapted) strains. Previous work in our laboratory demonstrated the utility of proteomics to study mechanisms of Y. pestis adaptation to laborato (...truncated)


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Owen P. Leiser, Jason K. Blackburn, Ted L. Hadfield, Helen W. Kreuzer, David S. Wunschel, Cindy J. Bruckner-Lea. Laboratory strains of Bacillus anthracis exhibit pervasive alteration in expression of proteins related to sporulation under laboratory conditions relative to genetically related wild strains, PLOS ONE, 2018, Volume 13, Issue 12, DOI: 10.1371/journal.pone.0209120