6/19/2014

1
BIOL2P98D3Spring2014
PrinciplesofMicrobiology
Dr.CarolynnEPietrangeliPhD
Lecture2June20,2014
CHSC2P98PrinciplesofMicrobiology
SpringD3June16July182014
INSTRUCTOR: Dr.CarolynnE.Pietrangeli,PhD
CONTACT: email: cpietrangeli@brocku.ca
Office:MCF212
OFFICEHOURS: Monday/Friday12PM1PM
Andbyappointment email
request
FORMAT: Lectures,3hourstwiceweekly
Monday/Friday9AM12PM
AS216
6/19/2014
2
Coccus
Spirochete
ProkaryoteMorphology:HowdoIlook?...
Rod
Stalk Hypha
Buddingand
appendaged bacteria
Width:0.54.0m
Length:<15m
Figure2.11
Spirillum
Filamentousbacteria
2012 Pearson Education, Inc.
Eukaryoticcells:Diameter
10to>200m
WhySmallMatters
Highsurfacetovolume(S/V)ratiosupports
Fastergrowththaninlargercells
Greaternutrientexchangeperunitcellvolume
Whenissmalltoosmall?:<0.15 m
6/19/2014
3
Nutrientand
wastetransport
Respiration
Plasma
membrane
Ribosome
Protein
synthesis
Pili
Attachment
Conjugation
Motility
Flagellum
membrane
Plasmids
Extrachromosomal DNA
Chromosome Chromosome
DNA;Geneticarsenal
CellWall
Shape
Protection
fromosmotic
stress
BacterialCell
Ultrastructure
Fattyacids
Glycerol
Phosphate
ThePlasma
Membrane
Phosphate
Ethanolamine
Fattyacids
Hydrophilic
region
region
Hydrophobic
Figure3.4
Glycerophosphates
g
region
Hydrophilic
Fattyacids
2012PearsonEducation,Inc.
6/19/2014
4
Phospholipids
groups
Hydrophobic
OutsideCell
FluidMosaicModelof
ThePlasma(Cytoplasmic)Membrane
Integral
68nm Hydrophilic
groups
g p
InsideCell
Figure2.15
Integral
membrane
proteins
Phospholipid
molecule
2012PearsonEducation,Inc.
FunctionsofThePlasmaMembrane
Permeabilitybarrier:
Preventsleakageandfunctions
asagatewayfortransportof
nutrientsinto,andwastesout
of,thecell
Proteinanchor:
Siteofmanyproteinsthat
participateintransport,
bioenergetics,andchemotaxis
Energyconservation:
Siteofgenerationanduseofthe
protonmotiveforce
2012 Pearson Education, Inc.
Figure2.18
6/19/2014
5
Out In
Transported
substance
Simpletransport:
Drivenbytheenergy
intheprotonmotive
force
substance
Grouptranslocation:
Chemicalmodification
ofthetransported
substancedrivenby
phosphoenolpyruvate
1
2
Figure2.20
ABCtransporter:
Periplasmicbinding
proteinsareinvolved
andenergycomes
fromATP
2
3
2012 Pearson Education, Inc.
Outside
Cell
Figure2.21
2012 Pearson Education, Inc.
Inside
Cell
Uniporter Antiporter Symporter
6/19/2014
6
DistinctCellWallStructureinGrampositive
andGramnegativeBacteria
Murein =peptidoglycan
CellWall
Shape
Protection
fromosmotic
stress
Opolysaccharide Corepolysaccharide
LipidA Protein
OutsideCell
Lipopolysaccharide
(LPS)
TheOuterMembraneofGramnegative
BacteriaContainsLipopolysaccharide(LPS)
Peptidoglycan
Porin
8nm
Phospholipid
(LPS)
Lipoprotein
I id C ll
Outer
membrane
Periplasm
Cytoplasmic
membrane
Cell
wall
Figure2.29
InsideCell
membrane
2012 Pearson Education, Inc.
6/19/2014
7
SoWhat?WhydowecareaboutLPS?...
LipidAcomponent=Endotoxin
LipidA TolllikeReceptor TNF /IL1
On E.coli OnMacrophage MacrophageCytokines
LipidA potent
immunostimulant
Immunesystem
overproducesinflammatory
cytokines: y
Fever
Respiratorydistress
Lowbloodpressure
Endotoxic shock
CapsulesandSlimeLayers:
Attachment/Immuneevasion
Fimbriae andFlagellae
Cell Capsule
Flagellae
Fimbriae
Adh i / I i
Locomotion
Figure2.32;Figure2.33
2012 Pearson Education, Inc.
Adhesion/Invasion
Pellicleformation
Biofilm formation
6/19/2014
8
MicrobialTaxis
Directedmovementinresponsetochemical
orphysicalgradients
Chemotaxis: response to chemicals Chemotaxis:responsetochemicals
Phototaxis:responsetolight
Aerotaxis:responsetooxygen
Osmotaxis:responsetoionicstrength
Hydrotaxis:responsetowater
ConcentrationGradient NoConcentration
Gradient
Repellent
Control
Attractant
MeasuringBacterialTaxis
Control
Attractant
e
l
l
s

p
e
r

t
u
b
e

Figure2.58
Time
Repellent
C
e
2012PearsonEducation,Inc.
Aerotaxis:Attractiontooxygen
6/19/2014
9
Pili
Viruscoveredpilus
(Bacteriophage)
Conjugation exchangeofgeneticmaterial
Adhesiontohosttissues/invasion
Figure2.34
ThePilus MediatesPlasmidTransfer
http://bio3400.nicerweb.net/Locked/media/ch06/
Plasmid extrachromosomal loopofDNAthatcan
replicateindependentlyofthecellandbetransferred
fromonebacteriumtoanother
6/19/2014
10
StormingOutside?GotoSleep!
Endospores andSporulation
Terminalspores Subterminal spores Centralspores
Figure2.42
2012 Pearson Education, Inc.
Endospore:Survivalstructurerenderingacellresistanttoheat,
chemicals,radiation
DormantstageoflifecycleinsomeGrampositive bacteriaonly
Figure3.33
Vegetativecell
Developing spore
Sporulation
Developingspore
Sporulatingcell
Maturespore
2012 Pearson Education, Inc.
6/19/2014
11
Exosporium
Sporecoat
Corewall
Cortex Cortex
DNA
Figure2.45
2012 Pearson Education, Inc.
Nutrientand
wastetransport
Respiration
Plasma
membrane
Ribosome
Protein
synthesis
Pili
Attachment
Conjugation
Motility
Flagellum
membrane
Plasmids
Extrachromosomal DNA
Chromosome Chromosome
DNA;Geneticarsenal
CellWall
Shape
Protection
fromosmotic
stress
BacterialCell
Ultrastructure
6/19/2014
12
Purinebases Pyrimidine bases
Cytosine
(C)
Thymine
(T)
Uracil
(U)
Adenine
(A)
Guanine
(G)
5 position
Componentsofthe
NucleicAcids
(C) (T) (U) (A) (G)
Phosphate
3 position
Phosphodiester
bond
Nitrogenbase
attachedto
1 position
Deoxyribose
Figure4.1
Ribose
Honly
inDNA
Originof
replication
lac operon
(lactose
degradation)
100/0
90
10
HfrH
P804
HfrC
t
Not1restriction
sites,inkbp
4,639,679bp
EscherichiacoliK12
80
70
30
20
KL14
Figure4.8
trp operon
(tryptophan
biosynthesis)
his operon
(histidine
biosynthesis)
60
50
40
Hfr44
2012 Pearson Education, Inc.
6/19/2014
13
Plasmids:Extrachromosomal ProkaryoticDNA
Replication
functions
mer
sul
str
cat
IS1
94.3/0kbp
SmallcircularorlineardsDNA
molecules;oftensupercoiled
tra
IS1
IS10
IS10
75kbp 25kbp
50kbp
Figure 4 9
tet
oriT
Tn10
Figure4.10
GeneralProperties
Size:1kbpto>1Mbp;<5%sizeofhostchromosome
Replicateindependentlyofhostchromosome
Genes:Nonessentialbut
Highlyinfluentialtohostphysiology
Mayprovidegrowthadvantageunderadverseconditions(antibioticpresence)
Figure4.9
FeaturesandSignificanceofPlasmids
Feature Bacterial
chromosome
Plasmid
Size:Numberof
Genes
Base pairs
Uptothousands
10005000 kbp
Up to30
500kbp
Replicative
potential
Yes Yes
Persistence Yes Curing lossofplasmidsunder
laboratory conditions laboratoryconditions
(spontaneous/deliberate)
Essential Yes No
6/19/2014
14
AMomentintheLab
Wecaninducecuringbytreatingbacterialcultures
with radiation or mutagens including acridine orange withradiationormutagensincludingacridine orange
Thistreatmentwillnotaffectbacterialreproduction
Whywouldwewanttodothis?
FunctionsofPlasmids
Plasmidsencodeproteinsthat
Conferantibiotic/poisonresistance
Conferpathogenicity
Scavengepollutantsandrecycle
6/19/2014
15
TypesofPlasmids
RPlasmids Conferantibioticandtoxicmetal
resistance resistance
FPlasmids Conjugative
VirulencePlasmids Hemolysin /Enterotoxin
BacteriocinencodingPlasmids Narrow
spectrumantibioticstoxictocloselyrelated
bacterial species/strains of the same species bacterialspecies/strainsofthesamespecies
(bacteriocins:e.g.colicin,nicin)
Episomes andCuring
Curing Eliminationofthe
plasmid p
Spontaneously
Treatmentinduced
affectsplasmidsurvival
withoutaffectinghostcell
survival
Episome plasmidincorporatedinto
bacterialchromosome
6/19/2014
16
FPlasmidsare
Episomes: p
Canexistwithor
withoutintegration
intothehostcell
chromosome
Prescottetal.
8thed.Fig.14.20
7thed.Fig.13.25
PlasmidRulesofEngagement
Asinglebacteriummayharbour multiple
plasmids of various types as long as the plasmids ofvarioustypes aslongasthe
plasmidsfallintodifferentIncompatibility
Groups
Copynumber numberofplasmidsofthe
sametypefoundwithinasinglecell
C t ll d b l id Controlledbyplasmidgenes
Controlledbyinteractionbetweenplasmidand
host
6/19/2014
17
PlasmidIncompatibilityGroups
Group Host Range Plasmid Original
Host
Phenotype
I FI
E h i hi li
F E li T + D (F1 IncFI
Escherichiacoli
Proteusvulgaris
Salmonellatyphimurium
Proteusmirabilis
F
R453
E. coli
P. morganii
Tra+ Dps (F1,
F2, etc.), Phi
(T3, T7, Q11)
Tra+ Dps (F1,
F2, MS2) Phi (),
Ap, Cm, Sm, etc.
IncFII R100 Shigella
flexineri
Tra+ Fi+ (F) Dps
(F1, F2, MS,
flexineri
(
M13) Cm, Fa,
Sm, etc.
IncP Gram negative bacteria RP1 Pseudomonas
aeruginosa
Tra+ Fi- (F) Dps
(PRD1, PRR1)
Ap, Km, Tc
ReplicationoftheCircularBacterialChromosome
6/19/2014
18
Nucleocapsid
Envelope
Capsid
Nucleic
acid
Capsid
(composedof
Capsid
Nucleic
acid
Figure8.1
( p
capsomeres)
Nakedvirus
Envelopedvirus
2012 Pearson Education, Inc.
6/19/2014
19
HepatitisCVirus(HCV) HumanImmunodeficiencyVirus1(HIV1)
ViralClass
ViralGenome
DNAviruses RNAviruses
RNA DNA
viruses
ssDNA dsDNA ssRNA dsRNA
ssRNA
(Retroviruses)
dsDNA
(Hepadnaviruses)
Group Viral Nucleic Acid Examples
BaltimoreClassificationSystem
Group ViralNucleic Acid Examples
DNAViruses
I dsDNA Adenovirus
II ssDNA Parvovirus (B19)
RNAViruses
III dsRNA Rotavirus
IV + Sense/strand ssRNA Poliovirus HCV
Figure8.2
2012PearsonEducation,Inc.
IV +Sense/strandssRNA Poliovirus, HCV
V Sense/strandssRNA Influenza, ebola
Virusesutilizingreversetranscriptase
VI Retroviruses ssRNA DNA HIV
VII Hepadnaviruses dsDNA HBV
6/19/2014
20
http://classes.midlandstech.com/carterp/Courses/bio225/chap13/lecture3.htm
Laboratories
Schedule:AsperpostingonSakai
LaboratoryDemonstrators
ChristeneCarpenterClelandMCF210
Extension5788ccarpenterclela@brocku.ca
MarkLukewich MCF213
Extension3398mlukewich@brocku.ca
6/19/2014
21
Evaluation
Laboratory 35% SeeLaboratoryManual
MidtermExamination 20% FridayJuly49:00AM10:30AM
Discussion Forum will follow DiscussionForumwillfollow
FinalExamination 20% 2hourduration
DiscussionForum 25% 5exercises/5markseach
SeeSakaiforInstructions
20%content
5%participation p p
ThereareNOmakeuporsupplemental examinationsinBIOL2P98
ThisappliestoLectureExaminations
(MidtermANDFinalExamination)ANDtothe
LaboratoryExamination
BIOL2P98SPLaboratorySchedule
LaboratorySchedule
Weekof LaboratoryExerciseTitle
Jun 16 Lab 1 Lab Safety Techniques & Enumeration
LabsareinIH309.
Jun16 Lab1 LabSafety,Techniques&Enumeration
Jun23 Lab2 BacterialGrowth&Identification
Jun30 CanadaDayweek NOLABS Assignment2onSakai
Jul07 Lab3 TheControlofMicrobialGrowth
Jul14 LabExam Basedonalllabmaterial
Labsections IH309
Section5 Monday 2:00pm 5:00pm
Section3 Tuesday 10:00am 1:00pm
Section1 Tuesday 2:00pm 5:00pm
Section2 Wednesday 2:00pm 5:00pm
ReadallAppendices&Lab1onSakai
CreateaflowchartforLab1
Bringalabcoat&safetyglasses/goggles,Lab1,
flowchartandPhotographicAtlas
6/19/2014
22
DiscussionForums
OccureachFridayduringlastlecturehour(11AM12PM)
Eachweek,weevaluateacaserelevanttolecture/laboratory
content in Microbiology in assigned Small Groups contentinMicrobiologyinassignedSmallGroups
Eachcasewillbeaccompaniedbysetsofquestionstobediscussed
andworkedonbyeachGroup
EachGroupsubmitsaSINGLEelectronicReport(nopaper
submission)respondingtothecasequestionssetforthatGroup
ELECTRONICReport(nopapersubmission)dueby5PMonthe
T d f th f ll i k ( C O tli ) Tuesdayofthefollowingweek(seeCourseOutline)
TheGroupwillreportthespecificcontributionofeachmember
EachmemberoftheGroupearnstheSAMEGRADEfortheReport
DiscussionForumInstructionspostedonSakai includesGroup
listings
DiscussionForumParticipation
20%ofthetotalgradeearnedforthe
Discussion Forums (5% of the final grade in DiscussionForums(5%ofthefinalgradein
BIOL2P98)isderivedfromparticipationin
DiscussionForumexercises
Attendancewillberecordedbysigninsheet
atthebeginningofeachDiscussionForum
Di i G ill b k d t d ib DiscussionGroupswillbeaskedtodescribe
thespecificcontributionofeachmemberto
eachGroupReport
6/19/2014
23
Resources
RequiredTextbook
BrockBiologyofMicroorganisms.FourteenthEdition.MadiganMT,Martinko
JM,BenderKS,BuckleyDH,StahlDA.PearsonBoston2012
Providesabasicreviewofconceptstobediscussedduringthecourse
DoesNOTsubstituteforcourselecturesorDiscussionForumcontent
Textbookchapterswillnotbeassigned;studentswillmatchthelecture
contenttorelevantchaptersinthetextbook
TextbookcontentwillbesupplementedbyreadingsmadeavailableonSakai.
Postingsofadditionalreadingswillbeannouncedduringlecturesandon
Sakai
RecommendedTextbookforLaboratory
APhotographicAtlasfortheMicrobiologyLaboratory.Leboffe MJ,PierceBE
4thEdition.MortonPublishing2011
GeneralBackgroundMicrobiology
http://www.mhhe.com/biosci/cellmicro/talaro/links.mhtml
AcademicIntegrity
AReflectionofPersonalIntegrity
Manythingsmaybestolenfromusbyother
peopleoverthecourseofourlives
Integrityisonethingwecanstealonlyfrom
ourselves
Integrity
Respecting it Respecting yourself Respectingit=Respectingyourself
Preserveit
Beproudofupholdingit