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Listeria and Omics Approaches for Understanding Its Biology
Janet R. Donaldson, Ph.D.Mississippi State University
Department of Biological [email protected]
Listeria monocytogenesOne of the deadliest
food-borne pathogensCantaloupe outbreak
in 2011 was the third most deadly outbreak since 1924; killed 30 people 1st: E. coli O104:H4
outbreak of 2011; Killed 53
2nd: Listeria outbreak in 1985; Killed 52
Soil bacteriumInfects both humans and animals
Low pH, intracellular growth, high bile salts5-10% of population are asymptomatic carriers
Grows in a wide range of temperatures (2°C to 39°C) and environmental conditions (salt, mildly acidic)
Major problems with ready to eat foods, such as smoked fish industry, deli meats, and cheese
Survives a variety of environments
It’s complicated…Many factors contribute to
the complications of studying Listeria monocytogenes13 different serotypes, 4
genetic lineagesMost food-borne outbreaks are related to serotype 4b, but 1/2a, 1/2b, and 4a are also important for human and animal health
Conversion to L-forms Formation of biofilms
Advances in molecular biology allowed for huge advancements to be made in terms of monitoring strain persistence in food processing plants
GOLDEN AGE OF OMICS
Advances in OmicsVast amount of
informationTranscriptomicsProteomicsSecretomicsMetabolomicsInteractomics
Functional Proteomics Approach to Answer Complicated Questions How do basic metabolic processes differ
between different serotypes?What are the mechanisms that allow for
survival of Listeria monocytogenes in stressful environments?
How do these strategies for survival differ between different serotypes of Listeria monocytogenes?
Proteomics AnalysisPreparation of protein samplesAnalyzed by HPLC coupled with nano-LTQ
linear ion trap mass spectrometerSpectra analyzed against trypsin digested
FASTA database with converted file Protein modifications need to be analyzed;
analysis run against decoy databaseProtein expression comparisons made
based on spectral intensitySignificantly expressed proteins
assigned to classification category based on ListiList
Variations in proteome expressionEGDe (1/2a
serotype) and F2365 (4b) proteomes differ in stationary phase
F2365total: 1427(350)
EGDtotal: 1754(677)
Common(1077)
Donaldson et al, 2009 Appl Environ Microbiol
F2365EGDe
Expression of 322 proteins was significantly increased in EGDe in comparison to F2365 Mostly in cell wall physiology and cell wall anchor
proteinsStress response proteins UvrC, UvrA, MutS, MutY,
RecN, GroEL, GroES, DnaKExpression of 91 proteins was significantly
decreased in F2365 in comparison to EGDeFlagellar biosynthesis protein Chemotaxis protein CheY Catalase Thiol peroxidase
Proteins Differentially Expressed
Donaldson et al, 2009 Appl Environ Microbiol
Functional Proteomics Approach to Answer Complicated Questions How do basic metabolic processes differ
between different serotypes?What are the mechanisms that allow for
survival of Listeria monocytogenes in stressful environments?
How do these strategies for survival differ between different serotypes of Listeria monocytogenes?
Mechanisms that allow for survival in stressful environmentsMinimal exposure to one stressor can
influence resistance to another stressorMethods used can increase resistance of
Listeria monocytogenes to other stressors encountered and also influence its persistence in facilities
Studies have suggested overlap in adaptive mechanisms used by L. monocytogenes to cold stress and osmotic stress
Cold Smoking ProcessProcess involves:
Freeze/ thaw of fish to remove parasitesBrining with salt Exposure to smoking conditions for less than
24 hPackaging and storage
Can exposure to cold influence survival under osmotic stress?
Functional Proteomics Approach to Answer Complicated Questions How do basic metabolic processes differ
between different serotypes?What are the mechanisms that allow for
survival of Listeria monocytogenes in stressful environments?
How do these strategies for survival differ between different serotypes of Listeria monocytogenes?
1 3 5 7
F2365
EGD-e
HCC23
Time (hr) post infection
Intr
acell
ula
r C
FU
/mL
Serotype variability in response to stressful environments
Responses to intracellular growthAvirulent: decrease in lipid metabolism,
increase in cell wall proteinsVirulent strains: increase in oxidative
repair proteins2x10
^7
1x10^5
Donaldson et al, 2011 J Proteomics
Other studies…Variations in the mechanism by which
avirulent and virulent strains of Listeria monocytogenes respond to bile salt environmentsEfficient repair mechanisms are essential for
long term survivalVariations in intracellular v. extracellular
growth between virulent and avirulent strains
Payne et al, 2013 J Med Microbiol
ConclusionsProteomics has advanced our understanding of
the biology of Listeria monocytogenesVariations between serotypes: provide insight
into ecological niches Mechanisms for survival in presence of stressors:
Cross adaptation between cold and osmotic stress: Proteins involved re-modeling of the cell wall and cell
membrane allow for the integrity and fluidity of the cell to be maintained in response to osmotic stress
Variations between different serotypes in response to stressorsAvirulent strains possess different mechanism to try
to survive in presence of bile salts than virulent strains
Thank You!MSU
Joseph PittmanKendrick Currie Jessica Grissett-WilsonGabe PosadasStephanie VamentaAngela PayneSusan BridgesChamali Thanthiriwatte Justin ThorntonMark LawrenceBindu NanduriTibor Pechan
Shane Burgess, Univ. AZKen Pendarvis, Univ. AZTodd Callaway, USDA-ARS Jeff Carroll, USDA-ARS Joe Buntyn, UNLTy Schmidt, UNL Tjakko Abee, Wageningen
University
Funding: MSU ORED
MSU Dept. of Biological SciencesDOE Sustainable Energy Research Center
MAFES Special Research InitiativesNIH (T35RR007071)