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1、The Genomic and Environmental Stressors that Impact Microbial PathogenesisAlvin LeeFood Safety Summit 2024Food Environment Physical/chemical environments ProductionProcessingPreservationStorageTransportationConsumption Microbiological quality evaluationsStandardized proceduresAntaris Consulting Adap

2、tedUn-adaptedpH 5.8pH 3.3pH 7.6pH 7.6pH 3.3Classic acid stress example by Foster&Hall in 90s on Salmonella ATRStress Adaptation“Asituationwherebyabriefexposuretoasuboptimal physical or chemical environment thatenables the cells to resist subsequent exposure tothe same or other types of harsher treat

3、ment towhich the species is normally susceptible.”Ray et alStress AdaptationRay et alTypes of StressBacterial cells alter lipid composition(membrane)Maintained fluid stateProportion of cyclopropane fatty acids in the cell membraneregulated by cyclopropane fatty acid synthaseLeads to Gram negative pr

4、essure,acid and oxidativeresistanceMechanisms of Stress AdaptationMechanisms of Stress AdaptationCurrent Concept:Gene Response Shock ProteinsStress ProteinsSpecific or nonspecific Inducible or constitutive Aid for adaptation to other stressorsRay et al Bacteria contain a primary factor that is respo

5、nsible for transcription ofhousekeeping genes necessary for growth and survival Many bacteria encode multiple alternative factorsThe level and activity of the alternative factors are highly regulated and can varydepending on environmental or developmental signals Example:rpo systems Usually regulate

6、d by a regulatory gene Gram(-)rpoH and rpoE for heat response and rpoS for general stress,celldensity and starvationStress-Related GenesSigma Factors E.coli can choose between 7 factors to fine tune its transcriptional outputJ.Helmann.Sigma factors in gene expression.2005;10.1038/npg.els.0003829Emer

7、ging or Re-Emerging Novel etiological agents that have been recently introduced in a population Often zoonotic in origin Microorganisms include Gram+,Gram bacteria,parasites and viruses Acquisition of new virulence factors Acquisition of antibiotic resistance Genetic assortment Factors human host,en

8、vironment,processing,travel,novel companion animals etcSpanish flu 19181957-Asian flu1968 Hong Kong flu1977 Russian flu2009 swine fluSeveral other flu outbreaksCirculation of bovine G6 and G10 strains in calves and humansAdaptation to Stress A Good Idea?Beneficial bacteria Fermentation bacteria new

9、and novel foods Pathogens and spoilage bacteria Antibiotic resistance More virulent strains Impact on characteristics of food New flavors starter cultures Food as medicine?Control Measures Clean and disinfect environmental surfaces Hand washing and personal hygiene(asymptomatic transmission control)

10、Interventions for reduction of pathogen load Improve diagnostics for detection and characterization Understand and evaluate food safety systemsMicrobial Goal?Adapt&SurviveQuanta MagazinePerishalee33iit.eduOREvolution and Pathogenesis:What time has done to microorganisms-what we know and dont knowPur

11、nendu C.VasavadaProfessor Emeritus of Food ScienceUniversity of Wisconsin-River FallsRiver Falls,WI.54022AgendaIntroduction:Emerging,Reemerging Pathogens-What is in the name?Mechanism of Bacterial PathogenicityEvolution of microbial PathogensWhat we know and dont knowSummaryEmerging,Reemerging Patho

12、gens-What is in the name?The Challenge of Emerging PathogensClin Infect Dis.2001;32:67585.Nature,volume 430,pages 242249(2004)Emerging,Reemerging and Opportunistic Pathogens Emerging pathogens are New,reemerging,or drug-resistant infections whose incidence in humans have increased within the past tw

13、o decades or threatened to increase in the near futureReemerging Pathogens-involved in the reappearance of a known disease following a decline in incidence including newly recognized pathogens,new diseases caused by known organisms,and the extension of the geographic or host range of a pathogenOppor

14、tunistic Pathogens-are microbes that usually do not cause disease in healthy people,but may become virulent with immunocompromised and unhealthy individualsEmerging,Reemerging and Opportunistic PathogensEarly 1900 1940s-1960s1960s-1990s 2000-s Typhoid fever Tuberculosis Septic sore throat Diphtheria

15、 BrucellosisSalmonellaClostridium perfringensVibrio parahaemolyticus Bacillus cereus Pathogenic E.coli Campylobacter jejuni Salmonella Spp.E.coli 057:H7 Non 0157 EHEC/STECs Listeria monocytogenes*Yersinia enterocolitica*Bacillus cereus*Staphylococcus aureus Cronobacter sakazakii Vibrio parahemolytic

16、us V.vulnificus Cryptosporidium,Cyclospora,Toxoplasma gondii Norovirus,Hep A.Nipah Virus Clostridium botulinum Salmonella Spp.Staphylococcus aureus StreptococciFactors Contributing to the Emergence of Foodborne Pathogens and Diseases Microbial adaptation and change Human susceptibility to infection

17、Climate and weather Changing ecosystems Human demographics and behavior Economic development and land use International travel and commerce Technology and industry Breakdown of public health measures Other social,political and economic factors e.g.War and famine,Lack of political will,and Intent to

18、harmMechanisms of Bacterial PathogenicityPathogenesis and Pathogenicity Pathogenesis refers to the development of a diseasePathogenicity refers to the ability of an organism to cause disease.Commensals and opportunistic pathogens lack this inherent ability to cause diseaseVirulence refers to the deg

19、ree of pathology caused by the organism.Virulence genes are involved in horizontal(lateral)transferThe extent of the virulence is usually correlated with the ability of the pathogen to multiply within the host and may be affected by other factors.Pathogenicity is used as a qualitative term,virulence

20、 is used more as a quantitative term.Mechanisms of Bacterial Pathogenicity 4 Stages of PathogenesisExposure(contact)Adhesion(colonization)InvasionInfectionSurvival and replication after invasionAdaptation to the new environmentAntibiotic resistanceBiofilmWilson,Schurr,LeBlanc,et al.2002 Postgrad Med

21、 J:78:216224 Evolution of microbial PathogensEvolutionDarwinian Principles of Evolution Genetic variability Phenotype formation Selection,and IsolationPathogens evolve over time following natural selection.MorschhIuser,Kohler,Ziebuhr et al.2000.Phil.Trans.R.Soc.Lond.B:355,695-704 Evolution of Microb

22、ial PathogensBacteria can evolve from non-pathogens to become pathogens through acquisition of new genetic material that enables them to colonize a host species and have detrimental effects on the host Bacterial evaluation occurs through mechanisms including natural selection and genetic drift and c

23、an result in adaptations to environmental change or host immunityThe genetic makeup of bacterial genomes is subject to rapid and dramatic change through a variety of processes collectively referred to as“horizontal gene transfer”(HGT),which plays a principal part in the molecular evolution of novel

24、bacterial pathogens Pathogenic bacteria can undergo further genetic modification that leads to altered virulence and changes in their genomeGenetic Mechanisms in Bacterial EvolutionMacro evaluation-Long-term processes leading to the development of new species or subspeciesMicroevolution-Short-term d

25、evelopments,which occur during days or weeksBoth processes,macro-and microevolution need horizontal gene transfer,which is particularly important for the development of pathogenic microorganisms.Horizontal or Lateral gene transfer(HGT)and Mobile Genetic Elements(MGE)are important in evolution of pat

26、hogen from non-pathogenic ancestorGenetic Mechanisms in Bacterial EvolutionTransfer of foreign DNA Horizontal or Lateral gene transfer(HGT)TransformationTransduction,ConjugationMobile genetic elements(MGE)Genetic Mechanisms in Bacterial EvolutionHorizontal Gene Transfer(HGT)Mobile Genetic Elements(M

27、GE)-the DNA mobilized into the host bacteriumPlasmids,bacteriophages,Integrative and conjugative element,prophages,and Pathogenicity islands(PAIs)play a crucial role in the evolution of pathogens.Virulence genes transferred via HGT include genes for bacterial adherence to host cells,type 3 secretion

28、 systems,toxins,iron acquisition,and antimicrobial resistanceMobile Genetic Elements in Selected Bacterial Pathogens OrganismMobile ElementVirulence mechanismEHEC,EPEC,ETEC Plasmids,Phage,Pathogenic IslandAdherence,type III secretions,stxSalmonella entericaPlasmid,Pathogenic IslandInvasion of nonpha

29、gocytic cells,intracellular survival and replicationClostridium perfringensPlasmid,Pathogenic IslandToxinsListeria monocytogenesPlasmid,Pathogenic IslandsAdherence,Invasion,enzymesEnterococcus spp.Plasmid,Pathogenic IslandsBiofilm,toxins,piliStaphylococcus aureusPathogenic Islands,PhagesSuperantigen

30、,leukocidinGyles and Boerlin.2014.Veterinary Pathology.Vol.51(2)328-340Genomic Islands and Pathogenicity IslandsGenomic islands blocks of DNA containing mobile genetic elements transferred from the donor organisms to recipient.Widely distributed in GM+and GM bacteria and known to encode a variety of

31、 functionsDepending on the functions which are encoded by genomic islands,they may also be called symbiosis islands,metabolic or resistance islands and pathogenic islandsGenomic Islands and Pathogenicity IslandsOccur as distinct units on the core chromosome with a general genetic structures characte

32、rized by a set of unifying features.Present in the genomes of many bacteria but absent from the genomes of closely related strains Often large(10200 kb);however,smaller inserts(110 kb)can occurThey differ in GC content and in their codon usage from the rest of the chromosomeThey are flanked by speci

33、fic sequences(direct repeats)They are usually associated with tRNA loci.Pathogenicity IslandsBest known Genomic islandsRepresent compact,distinct genetic units,often flanked by direct repeatsEncode clusters of genes whose products contribute to virulenceDifferent G+C content in comparison to DNA of

34、host bacteriaOccupy large chromosomal regions(often 30 kb).Found in GM and GM+bacteria and known to encode a variety of functionsThey are present in the genome of pathogenic strains of a given species but absent or only rarely present in those of non-pathogenic variants of the same or related specie

35、s.Functions Coded by Pathogenic IslandsFunctionOrganismIncreased pathogenicityiron uptakeSalmonella enterica Klebsiella spp.Yersinia spp.,Bacillus cereus+toxin productionVibrio cholerae+Adhesinsurinary E.coliMethicillin resistanceStaphylococcus aureustype IIIsystemSalmonella entericaShigella flexner

36、iYersinia spp.+type IVsystemHelicobacter pylori+Hacker and Carmiel(2001What we know and dont knowSummaryEmerging,re-emerging and opportunistic pathogens are important threat to food safety and public healthBacterial pathogenesis involve ability of pathogen to invade and infect the host,survival and

37、replication after invasion and adaptation to the new environment as well as dealing with host immunity,antibiotic resistance,and biofilmHorizontal gene transfer via transformation,transduction and combination as well as Mobile Genetic Elements(MGE)are particularly important for the development of pa

38、thogenic microorganisms from non-pathogenic ancestorPathogenic islands,the best known Genomic islands are found in GM and GM+bacteria and encode clusters of genes whose products contribute to virulenceThey are present in the genome of pathogenic strains of a given species but absent or only rarely p

39、resent in those of non-pathogenic variants of the same or related speciesAvailability of numerous complete genome sequences of bacterial pathogens and the use of genomic techniques have given us new tools to study and understand microbial pathogenesisOur knowledge on mechanisms of pathogenesis is in

40、creasing but much is still unknownFinal Thoughts“Almost any bacterial species is capable of producing intestinal symptoms if swallowed in sufficient numbers”DuPont and Pickering 1980“Expect the unexpected”Swerdlow and Altekruse,1998“The future of microbes and mankind will probably unfold as episodes

41、 of a suspense thriller that could be entitled Our Wits Versus Their Genes”.-Joshua Lederberg.2000.New Era,Old Problems:Emerging and Re-Emerging PathogensMeet Our SpeakersBrendan Niemira,Ph.D.U.S.Department of AgricultureAlvin Lee,Ph.D.,MASMIllinois Institute of TechnologyPurnendu C.Vasavada,Ph.DUni

42、versity of Wisconsin-River Falls Alvin Lee The genomic and environmental stressors that impact microbial pathogenesis Purnendu Vasavada Evolution and Pathogenesis:What time has done to microorganisms-what we know and dont know.Brendan Niemira Risk management options for preparedness and response to

43、emerging pathogens Conclusions,Q&ASession OverviewThe genomic and environmental stressors that impact microbial pathogenesis Alvin Lee,IITEvolution and Pathogenesis:What time has done to microorganisms-what we know and dont know.Purnendu C.Vasavada,UW River FallsRisk management options for preparedn

44、ess and response to emerging pathogensBrendan Niemira,USDARisk management options Pathogens and food safety incidents are part of the landscape Risk avoidance comes from preparedness Quantitative and qualitative risk assessment as part of SOP Risk avoidance and mitigation strategies should be baked

45、in Understand risks and potential impacts Risks of familiar and conventional pathogens Risks of unfamiliar and emerging pathogens May require fresh analysis of risk matrix Organisms cause disease when present in the right environment,in sufficient numbers New products,new processes new pathogens?Wha

46、t makes an organism a pathogen:Genes for harmful toxins Circumstances that promote expression of those genes Exposure/transfer to food and people Controls for a pathogen General,broadly effective Specific,targetedChange causes changeHazards vs.risks,with a side of analysisHazard:a bad thing that cou

47、ld happenHazard analysis:A list of all*the bad things that COULD happen How bad*it would be if they DID happenRisk:the likelihood that a hazard WILLhappenRisk analysis:(likelihood)(how bad it would be)Risk assessment and calculation Models focus on primary risks,significant hazards Can safely ignore

48、 risk of alien invasions,giant asteroids,etc.Potential food safety hazards:microbiological tests on raw ingredients are out of spec;failure of process control;break in cold chain;glass fragments in finished product;etc.,etc.Assign numerical probabilities to each risk based on statistics,experience,r

49、ecord-keeping,etc.Derive relationships among various risks,some of which you can mitigate,some of which you cant.Pathogen incidence:potential responsesAvoidanceContainmentEradicationDo NothingPathogen incidence:Avoidance Take steps to understand risks so as to avoidthem What comes into the plant Sup

50、plier verification,ingredient testing,land usage records,animal health records,carcass testing Whats in the plant already Water quality metrics,sanitary equipment design What happens in the plant Process controls,worker trainingPathogen incidence:Containment Once an issue has been identified,keep it

51、 from spreading HACCP:control points,limits,corrective actions,testing HARPC:hazard analysis and risk-based preventative controls Process validation,test and hold,recalls,redirection Record keeping,historical incident analysis,retrospectives,root cause analysis Regulatory compliance and process chan

52、ge plansPathogen incidence:Eradication Treatments applied to foods and food contact surfaces to kill pathogens which might be present Interventions,processing technologies,antimicrobial steps Conventional end-of-shift cleaning Disassembly,clean-in-place Hot water,sanitizers,scrubs Newer intervention

53、s Thermal or nonthermal interventions Peracids,quats,hypochlorites,surfactants Cold plasma,PEF,peroxidesPathogen incidence:Do Nothing Is this ever a good idea?Not the automatic first option May be appropriate to consider if Impacts(immediate and long term)are very low or trivial Actions(preemptive o

54、r responsive)are high cost or very difficult Instead of“Do Nothing”,may choose to“Do Little”Even if a limited response seems warranted,a full analysis will repay efforts by avoiding future occurrence BiofilmsBiofilms a complex challenge Bacteria live in communities Multiple species Multiple strains

55、Adherent extracellular matrix of proteins,carbohydrates,biopolymers Exchange of metabolites,DNA,RNA Protection from sanitizers,desiccation,freezing,heatAhmed et al.2019.Materials Today Bio.2.100017.Biofilms a complex challenge Pathogens can participate in mostly non-pathogenic biofilms Toxigenic str

56、ain may be weak,sensitive,poor biofilm former.Not much risk?In partnership with a non-toxigenic,non-pathogen which forms a strong,durable biofilm,risk is increased.Does the pathogen actively participate in biofilm formation?Or is it just along for the ride?Are genes traded?Are new genes expressed in

57、 a biofilm which are NOT expressed by free-living bacteria?Davey and OToole,2000.Microb Mol Biol Rev.64(4)Biofilms quorum sensing Bacterial populations grow accumulation,reproduction Intercellular communication,signaling.Sensing of environment and each other.Which critical thresholds are reached,new

58、 genes activate Changes to the behavior of individual participants and the biofilm as a whole Inhibition and/or deregulation of QS is a strategy for advanced antimicrobial interventions Challenge:need to know what happening before you can effectively interfere with itBiofilms quorum sensing,microbia

59、l ecology Biofilm microbial ecology remains only partially understood Multispecies interactions,dynamically changing environment Persistent,resistant Sanitation programs use combination of conventional tools and novel interventions Variable efficacy against biofilms and biofilm-associated organismsR

60、esources and further reading FDA:Foodborne Pathogenic Microorganisms and Natural Toxins Handbook,2ndedition “The Bad Bug Book”Sorted by type of organism.Relevant information on food vectors,common contamination pathways,illness symptoms and treatments Downloadable PDF www.fda.gov/food/foodborne-path

61、ogens/bad-bug-book-second-edition CDC Current Outbreaks:www.cdc.gov/outbreaks/index.html CDCs“Solve the Outbreak”online game“Become a disease detective”learn about dozens of infectious organisms and diseases while investigating outbreaks,foodborne and otherwise www.cdc.gov/mobile/applications/sto/we

62、b-app.htmlEvolution and Pathogenesis:What time has done to microorganisms-what we know and dont knowPurnendu C.VasavadaProfessor Emeritus of Food ScienceUniversity of Wisconsin-River FallsRiver Falls,WI.54022AgendaIntroduction:Emerging,Reemerging Pathogens-What is in the name?Mechanism of Bacterial

63、PathogenicityEvolution of microbial PathogensWhat we know and dont knowSummaryEmerging,Reemerging Pathogens-What is in the name?The Challenge of Emerging PathogensClin Infect Dis.2001;32:67585.Nature,volume 430,pages 242249(2004)Emerging,Reemerging and Opportunistic Pathogens Emerging pathogens are

64、New,reemerging,or drug-resistant infections whose incidence in humans have increased within the past two decades or threatened to increase in the near futureReemerging Pathogens-involved in the reappearance of a known disease following a decline in incidence including newly recognized pathogens,new

65、diseases caused by known organisms,and the extension of the geographic or host range of a pathogenOpportunistic Pathogens-are microbes that usually do not cause disease in healthy people,but may become virulent with immunocompromised and unhealthy individualsEmerging,Reemerging and Opportunistic Pat

66、hogensEarly 1900 1940s-1960s1960s-1990s 2000-s Typhoid fever Tuberculosis Septic sore throat Diphtheria BrucellosisSalmonellaClostridium perfringensVibrio parahaemolyticus Bacillus cereus Pathogenic E.coli Campylobacter jejuni Salmonella Spp.E.coli 057:H7 Non 0157 EHEC/STECs Listeria monocytogenes*Y

67、ersinia enterocolitica*Bacillus cereus*Staphylococcus aureus Cronobacter sakazakii Vibrio parahemolyticus V.vulnificus Cryptosporidium,Cyclospora,Toxoplasma gondii Norovirus,Hep A.Nipah Virus Clostridium botulinum Salmonella Spp.Staphylococcus aureus StreptococciFactors Contributing to the Emergence

68、 of Foodborne Pathogens and Diseases Microbial adaptation and change Human susceptibility to infection Climate and weather Changing ecosystems Human demographics and behavior Economic development and land use International travel and commerce Technology and industry Breakdown of public health measur

69、es Other social,political and economic factors e.g.War and famine,Lack of political will,and Intent to harmMechanisms of Bacterial PathogenicityPathogenesis and Pathogenicity Pathogenesis refers to the development of a diseasePathogenicity refers to the ability of an organism to cause disease.Commen

70、sals and opportunistic pathogens lack this inherent ability to cause diseaseVirulence refers to the degree of pathology caused by the organism.Virulence genes are involved in horizontal(lateral)transferThe extent of the virulence is usually correlated with the ability of the pathogen to multiply wit

71、hin the host and may be affected by other factors.Pathogenicity is used as a qualitative term,virulence is used more as a quantitative term.Mechanisms of Bacterial Pathogenicity 4 Stages of PathogenesisExposure(contact)Adhesion(colonization)InvasionInfectionSurvival and replication after invasionAda

72、ptation to the new environmentAntibiotic resistanceBiofilmWilson,Schurr,LeBlanc,et al.2002 Postgrad Med J:78:216224 Evolution of microbial PathogensEvolutionDarwinian Principles of Evolution Genetic variability Phenotype formation Selection,and IsolationPathogens evolve over time following natural s

73、election.MorschhIuser,Kohler,Ziebuhr et al.2000.Phil.Trans.R.Soc.Lond.B:355,695-704 Evolution of Microbial PathogensBacteria can evolve from non-pathogens to become pathogens through acquisition of new genetic material that enables them to colonize a host species and have detrimental effects on the

74、host Bacterial evaluation occurs through mechanisms including natural selection and genetic drift and can result in adaptations to environmental change or host immunityThe genetic makeup of bacterial genomes is subject to rapid and dramatic change through a variety of processes collectively referred

75、 to as“horizontal gene transfer”(HGT),which plays a principal part in the molecular evolution of novel bacterial pathogens Pathogenic bacteria can undergo further genetic modification that leads to altered virulence and changes in their genomeGenetic Mechanisms in Bacterial EvolutionMacro evaluation

76、-Long-term processes leading to the development of new species or subspeciesMicroevolution-Short-term developments,which occur during days or weeksBoth processes,macro-and microevolution need horizontal gene transfer,which is particularly important for the development of pathogenic microorganisms.Ho

77、rizontal or Lateral gene transfer(HGT)and Mobile Genetic Elements(MGE)are important in evolution of pathogen from non-pathogenic ancestorGenetic Mechanisms in Bacterial EvolutionTransfer of foreign DNA Horizontal or Lateral gene transfer(HGT)TransformationTransduction,ConjugationMobile genetic eleme

78、nts(MGE)Genetic Mechanisms in Bacterial EvolutionHorizontal Gene Transfer(HGT)Mobile Genetic Elements(MGE)-the DNA mobilized into the host bacteriumPlasmids,bacteriophages,Integrative and conjugative element,prophages,and Pathogenicity islands(PAIs)play a crucial role in the evolution of pathogens.V

79、irulence genes transferred via HGT include genes for bacterial adherence to host cells,type 3 secretion systems,toxins,iron acquisition,and antimicrobial resistanceMobile Genetic Elements in Selected Bacterial Pathogens OrganismMobile ElementVirulence mechanismEHEC,EPEC,ETEC Plasmids,Phage,Pathogeni

80、c IslandAdherence,type III secretions,stxSalmonella entericaPlasmid,Pathogenic IslandInvasion of nonphagocytic cells,intracellular survival and replicationClostridium perfringensPlasmid,Pathogenic IslandToxinsListeria monocytogenesPlasmid,Pathogenic IslandsAdherence,Invasion,enzymesEnterococcus spp.

81、Plasmid,Pathogenic IslandsBiofilm,toxins,piliStaphylococcus aureusPathogenic Islands,PhagesSuperantigen,leukocidinGyles and Boerlin.2014.Veterinary Pathology.Vol.51(2)328-340Genomic Islands and Pathogenicity IslandsGenomic islands blocks of DNA containing mobile genetic elements transferred from the

82、 donor organisms to recipient.Widely distributed in GM+and GM bacteria and known to encode a variety of functionsDepending on the functions which are encoded by genomic islands,they may also be called symbiosis islands,metabolic or resistance islands and pathogenic islandsGenomic Islands and Pathoge

83、nicity IslandsOccur as distinct units on the core chromosome with a general genetic structures characterized by a set of unifying features.Present in the genomes of many bacteria but absent from the genomes of closely related strains Often large(10200 kb);however,smaller inserts(110 kb)can occurThey

84、 differ in GC content and in their codon usage from the rest of the chromosomeThey are flanked by specific sequences(direct repeats)They are usually associated with tRNA loci.Pathogenicity IslandsBest known Genomic islandsRepresent compact,distinct genetic units,often flanked by direct repeatsEncode

85、 clusters of genes whose products contribute to virulenceDifferent G+C content in comparison to DNA of host bacteriaOccupy large chromosomal regions(often 30 kb).Found in GM and GM+bacteria and known to encode a variety of functionsThey are present in the genome of pathogenic strains of a given spec

86、ies but absent or only rarely present in those of non-pathogenic variants of the same or related species.Functions Coded by Pathogenic IslandsFunctionOrganismIncreased pathogenicityiron uptakeSalmonella enterica Klebsiella spp.Yersinia spp.,Bacillus cereus+toxin productionVibrio cholerae+Adhesinsuri

87、nary E.coliMethicillin resistanceStaphylococcus aureustype IIIsystemSalmonella entericaShigella flexneriYersinia spp.+type IVsystemHelicobacter pylori+Hacker and Carmiel(2001What we know and dont knowSummaryEmerging,re-emerging and opportunistic pathogens are important threat to food safety and publ

88、ic healthBacterial pathogenesis involve ability of pathogen to invade and infect the host,survival and replication after invasion and adaptation to the new environment as well as dealing with host immunity,antibiotic resistance,and biofilmHorizontal gene transfer via transformation,transduction and

89、combination as well as Mobile Genetic Elements(MGE)are particularly important for the development of pathogenic microorganisms from non-pathogenic ancestorPathogenic islands,the best known Genomic islands are found in GM and GM+bacteria and encode clusters of genes whose products contribute to virul

90、enceThey are present in the genome of pathogenic strains of a given species but absent or only rarely present in those of non-pathogenic variants of the same or related speciesAvailability of numerous complete genome sequences of bacterial pathogens and the use of genomic techniques have given us ne

91、w tools to study and understand microbial pathogenesisOur knowledge on mechanisms of pathogenesis is increasing but much is still unknownFinal Thoughts“Almost any bacterial species is capable of producing intestinal symptoms if swallowed in sufficient numbers”DuPont and Pickering 1980“Expect the une

92、xpected”Swerdlow and Altekruse,1998“The future of microbes and mankind will probably unfold as episodes of a suspense thriller that could be entitled Our Wits Versus Their Genes”.-Joshua Lederberg.2000.New Era,Old Problems:Emerging and Re-Emerging PathogensMeet Our SpeakersBrendan Niemira,Ph.D.U.S.D

93、epartment of AgricultureAlvin Lee,Ph.D.,MASMIllinois Institute of TechnologyPurnendu C.Vasavada,Ph.DUniversity of Wisconsin-River Falls Alvin Lee The genomic and environmental stressors that impact microbial pathogenesis Purnendu Vasavada Evolution and Pathogenesis:What time has done to microorganis

94、ms-what we know and dont know.Brendan Niemira Risk management options for preparedness and response to emerging pathogens Conclusions,Q&ASession OverviewThe genomic and environmental stressors that impact microbial pathogenesis Alvin Lee,IITEvolution and Pathogenesis:What time has done to microorgan

95、isms-what we know and dont know.Purnendu C.Vasavada,UW River FallsRisk management options for preparedness and response to emerging pathogensBrendan Niemira,USDARisk management options Pathogens and food safety incidents are part of the landscape Risk avoidance comes from preparedness Quantitative a

96、nd qualitative risk assessment as part of SOP Risk avoidance and mitigation strategies should be baked in Understand risks and potential impacts Risks of familiar and conventional pathogens Risks of unfamiliar and emerging pathogens May require fresh analysis of risk matrix Organisms cause disease w

97、hen present in the right environment,in sufficient numbers New products,new processes new pathogens?What makes an organism a pathogen:Genes for harmful toxins Circumstances that promote expression of those genes Exposure/transfer to food and people Controls for a pathogen General,broadly effective S

98、pecific,targetedChange causes changeHazards vs.risks,with a side of analysisHazard:a bad thing that could happenHazard analysis:A list of all*the bad things that COULD happenHow bad*it would be if they DID happenRisk:the likelihood that a hazard WILLhappenRisk analysis:(likelihood)(how bad it would

99、be)Risk assessment and calculation Models focus on primary risks,significant hazards Can safely ignore risk of alien invasions,giant asteroids,etc.Potential food safety hazards:microbiological tests on raw ingredients are out of spec;failure of process control;break in cold chain;glass fragments in

100、finished product;etc.,etc.Assign numerical probabilities to each risk based on statistics,experience,record-keeping,etc.Derive relationships among various risks,some of which you can mitigate,some of which you cant.Pathogen incidence:potential responsesAvoidanceContainmentEradicationDo NothingPathog

101、en incidence:AvoidanceTake steps to understand risks so as to avoid themWhat comes into the plant Supplier verification,ingredient testing,land usage records,animal health records,carcass testingWhats in the plant already Water quality metrics,sanitary equipment designWhat happens in the plant Proce

102、ss controls,worker trainingPathogen incidence:Containment Once an issue has been identified,keep it from spreading HACCP:control points,limits,corrective actions,testing HARPC:hazard analysis and risk-based preventative controls Process validation,test and hold,recalls,redirection Record keeping,his

103、torical incident analysis,retrospectives,root cause analysis Regulatory compliance and process change plansPathogen incidence:Eradication Treatments applied to foods and food contact surfaces to kill pathogens which might be present Interventions,processing technologies,antimicrobial steps Conventio

104、nal end-of-shift cleaning Disassembly,clean-in-place Hot water,sanitizers,scrubs Newer interventions Thermal or nonthermal interventions Peracids,quats,hypochlorites,surfactants Cold plasma,PEF,peroxidesPathogen incidence:Do NothingIs this ever a good idea?Not the automatic first optionMay be approp

105、riate to consider if Impacts(immediate and long term)are very low or trivial Actions(preemptive or responsive)are high cost or very difficultInstead of“Do Nothing”,may choose to“Do Little”Even if a limited response seems warranted,a full analysis will repay efforts by avoiding future occurrence Biof

106、ilmsBiofilms a complex challenge Bacteria live in communities Multiple species Multiple strains Adherent extracellular matrix of proteins,carbohydrates,biopolymers Exchange of metabolites,DNA,RNA Protection from sanitizers,desiccation,freezing,heatAhmed et al.2019.Materials Today Bio.2.100017.Biofil

107、ms a complex challenge Pathogens can participate in mostly non-pathogenic biofilms Toxigenic strain may be weak,sensitive,poor biofilm former.Not much risk?In partnership with a non-toxigenic,non-pathogen which forms a strong,durable biofilm,risk is increased.Does the pathogen actively participate i

108、n biofilm formation?Or is it just along for the ride?Are genes traded?Are new genes expressed in a biofilm which are NOT expressed by free-living bacteria?Davey and OToole,2000.Microb Mol Biol Rev.64(4)Biofilms quorum sensing Bacterial populations grow accumulation,reproduction Intercellular communi

109、cation,signaling.Sensing of environment and each other.Which critical thresholds are reached,new genes activate Changes to the behavior of individual participants and the biofilm as a whole Inhibition and/or deregulation of QS is a strategy for advanced antimicrobial interventions Challenge:need to

110、know what happening before you can effectively interfere with itBiofilms quorum sensing,microbial ecology Biofilm microbial ecology remains only partially understood Multispecies interactions,dynamically changing environment Persistent,resistant Sanitation programs use combination of conventional to

111、ols and novel interventions Variable efficacy against biofilms and biofilm-associated organismsConclusions and QuestionsResources and further reading FDA:Foodborne Pathogenic Microorganisms and Natural Toxins Handbook,2ndedition “The Bad Bug Book”Sorted by type of organism.Relevant information on fo

112、od vectors,common contamination pathways,illness symptoms and treatments Downloadable PDF www.fda.gov/food/foodborne-pathogens/bad-bug-book-second-edition CDC Current Outbreaks:www.cdc.gov/outbreaks/index.html CDCs“Solve the Outbreak”online game“Become a disease detective”learn about dozens of infectious organisms and diseases while investigating outbreaks,foodborne and otherwise www.cdc.gov/mobile/applications/sto/web-app.html