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Flashcards in T1 Deck (296):
1

Genera of Gram + bacteria

Actinomyces
Bacillus
Bifidobacterium
Clostridium
Corynebacterium, Propionibacterium (& other diphtheroids)
Enterococcus
Gardnerella
Lactobacillus
Listeria
Mobiluncus
Peptostreptococcus
Staphylococcus
Streptococcus

2

med-important G+ cocci

Strep
Staph
Enterococcus

3

general of bacterial endospore-formers

Clostridium
Bacillus

4

Genera of acid-fast + pink bacteria

Mycobacteria
Nocardia (usually)

5

cell shaped determined by

murein sacculus and cytoskeleton

6

cocci arrangements

diplococcus (pairs)
G- : (kidney bean)
G+: (lancet)
chains (strep)
clusters (staph)

7

bacillus/rod

v. short: G-
short, thin: G-
short, thin, needle-like: fusiforms, G-
long, thick: G+
short, thick: G+
clubbed-shaped: G+
-thin, branching filamentous rods w. club ends
*arranged as single cells, pairs (doublet) or chains

8

helicoidal

curved, comma: (G-)
curved, comma: (G+)
spirochetes
*arranged as single cells, pairs, chains

9

pleomorphic

can vary in size/shape:

10

flagella

H-Ag, protein composition
plain: extend out from cell surface
endoflagella: internal structure (spirochetes)
motility, sensory system (chemotactic), surface translocation, aids in identification, virulence factor (chemotactic)

11

Pili (Fimbriae)

protein composition
normal (I-IV) and sex pili extend out into environment
adherence (virulence factor), antiphagocytic, surface translocation (v.f.)
sex pilus: conjugation

12

capsule

glycocalyx, exopolysaccharides (EPS)
mucoid-like coat around cell, slime layer
polysacch. polymers: K-Ags: single, complex, D-glutamic acid repeating units OR O-Ag: LPS (G-)

13

biofilms

communal, protected, complex, 3-D structure (bac/yeast)
in exopolysacch. film, sometimes protein amyloid fibers
CDC: 50% human bac inf. involved biofilms
phenotypic changes: become sessile in biofilm, MDR, quorum sensing-->activation?
can revert to normal/planktonic form-->recurrent inf.
D-A.A. can disrupt amyloid protein connections-->release bad
*are a capsule

14

clin sig of EPS/biofilms

adherence, anti-phagocytic, anti-antibiotic, anti-dessication, Ag used to ID some human pathogen

15

HKO Ag

H: flagella
K: capsular
O: LPS (acts as capsule)
G+: capsule or no capsule
G-: O+K-, O+K+, O+,K- (again)

16

S-layers

A- or T-layers, (glyco)protein composed, 10-20% cell mass
present on some human normal flora AND pathogens
on cell wall, rigid layer w. pores of fixed diameter
virulence factors: anti-complement (C3b), anti-phago(PMN)

17

translocation

thru human cells to new site
paracellular, sliding motility (biofilms)

18

cell membrane contains no sterols except

Mycoplasma
Helicobacter
Ehrlichia
Anaplasma spp.

19

Type III secretion system

injectosome; conserved multiprotein system used by G- bac to insert protein toxins into human cells

20

cell membrane contains respiratory system

ETC, ATP synthase, PMF/ion current

21

cell membrane systems

perm/transport, respiration, cell wall synthesis components, cellular replication components, osmoreg/sensory (chemotactic) mechanisms (hypertonic to ext. environ.)

22

G+ or G-: polysacchs are covalently linked to peptidoglycan layer in cell wall and lipoteichoic acid polymers are anchored in cell membrane

G+

23

G+ or G-: contains an outer membrane with LPS, peptidoglycan layer located in periplasm

G-

24

cell wall functions

sieve, prevents bursting, mech. strength, vir/tox factors, Ag-comps, rec. for Abs, sex pilus, bacteriophages; anchors external bac structures (flag, pili, caps)

25

peptidoglycan layer

"fabric" of crosslinking, covalently bound threads: N-acetylmuramic acid (NAM) and N-actelyglucosamine (NAG)
-cleavage by lysozyme (human tears/saliva)-->bac lysis
-all peptide stems possess some D-a.a.s (i.e. D-alanine)

26

lytic transglycosylases

cell wall enzyme, causes cell wall turnover during exponential phase growth
product recognized by TLRs-->SIRS!

27

pathogenic bacteria WITHOUT peptidoglycan

Mycoplasma
Rickettsia
Ehrlichia
Anaplasma

28

G+ or G-: contain periplasm and an outer membrane

G- (thin pep. layer)
*confers resistance to dyes, hydrolytic enzymes, detergents

29

periplasm functions

osmotic protection (for thin pep. layer), nutrient uptake from OM-->CM, chemotactic sens. mech, degradative enzyms, osmoreg

30

OM functions

bac-environ interaction site, vir/tox factors, Ag-comps, ref for Abs, sex pilus, bacteriaphages, anchors ext. structures of bac, shield against dyes, hydrolytic enzymes, detergents

31

OM comp

lipopolysacc, phoslip, proteins (OMP)

32

OM structure

lipid bilayer (NOT phoslip bilayer)
LPS as outer leaflet, phoslip inner
porins: OMP, allows hydrophilics to pass thru

33

LPS

aka: endotoxin, exogenous pyrogen
Lipid A + core + O-Ag

34

Lipid A

disacch, phos grps, fatty acids, toxic factor

35

Core oligosacch of LPS

sugars, aminosugars, sugar acids, or sugar alcohols
-ketodeoxyoctulonate (KDO): common Ag in enteric bac

36

terminal polysacch.

aka O-Ag, repeating unit, contains sugars (like core), highly specific btw genera AND species

37

Lipooligiosaccharide (LOS)

lipid A + extended core, NO O-Ag
syn. instead of LPS, assoc. w/ Neisseria meningitidis and N. gonorrhoeae; Haemophilus influenza and H. ducreyi

38

props of LPS/LOS

part of OM, chromosome encoded, broad sp.(effects many org sys in susc. host) i.e. induction of IL-1 (endo. pyro), also acts directly on hypothalamus itself (exogenous pyrogen)
act. alt compl pathway, activator of Hageman factor (XII), induction release of endo mediators, heat stable, does not form toxoids

39

endogenous mediators induced/released by LPS/LOS

TNF-a, IL-1, IL-6
arch acid metals, bradykinin, histamine, NO, free radicals

40

only ways to disrupt primary structure of LPS/LOS

burning, oxidation -->detoxifies the endotoxin

41

Endotoxin/LPS/Lipid A is a potent immunomodulatory substance

SIRS-->distributive shock (DS) (G-)
systemic w. macros, PMNs, endothelial cells

42

SIRS activated by

50%: infection (usually G- (contain LPS))
50%: non-inf. etiology

43

initiation of SIRS

LPS binds LBP-->LPS-LBP complex interacts with mem-bound CD14 rec on PMS, macros/monos-->LPS-CD14 binds TLR-->signal transduction-->cytoplasm-->nucleus
-->induction/release of endogenous mediators
soluble CD14 rec in serum-->binds ECs-->dysfunction/leakiness>hypotension/vascular leak syndrome (ARDS: feature of DS, not SIRS)

44

why endotoxin (LPS) is not a true toxin

human response to presence is what causes fatality, not toxin itself

45

SIRS classification

2+:
temp >38 or =90 bpm
tachypnea >=20
leukocytosis: >12000 or

46

3 pathophys processes of sepsis etc.

systemic inflammation ("hyperinflammatory state")
coagulation activation
fibrinolysis inhibition

47

Disseminated Intravascular Coagulation (DIC)

LPS activates Hageman factor (XII)-->activation of coag (fibrin deposition)-->fibrinolysis activated by this but inhibited by plasminogen activator inhibitor-->accumulation of undiss. thrombin in microcirculation
DIC-->MOF +/- purpura fulminans

48

if survived hyperinflammatory state of sepsis, pt enters

"immunoparalysis"; hypoinflammatory state:
loss of type IV hypersn, failure to clear primary infection, dev. of new secondary infections, dormant viruses may "awaken" (HSV-1, EBV, CMV, HHV-7)

49

sepsis classification

2+ of SIRS criteria PLUS proven infection: i.e. pneumonia, UTI, bacteremia

50

severe sepsis

sepsis criteria PLUS organ failure (OD, MOD, MOF) most: heart, lungs, liver, kidneys

51

septic shock

severe sepsis criteria PLUS refractory hypotension

52

shock

inadequate profusion of tissues; 3 forms:
cardiogenic, vascular obstructiv, hypovolemic (DS)

53

distributive shock (DS)

"warm shock" (dilation)
-EC dysfunction/leakiness-->loss of plasma into tissues-->hypotension
-loss of vasc resistance-->hypotension
-coagulopathy-->DIC
-septic cardiomyopathy: rev., does not damage heart structure-->reduces CO (LPS, C5A, IL-1B, TNF-a, IL-6)

54

clinical manifestations of DS

fever or hypothermia, chills,
leukopenia/cytosis
tachy x2
DIC
hypotnsn, shock, DIC-->OD, MOD, MOF

55

Early Goal-Directed Therapy (EGDT) for SIRS/DS

-sepsis resuscitation bundle: measure serum lactate, obtain blood spec for culture, admin broad spec Ab, if hypotnsv: admin fluids: iconic crystalloid or iso-onccotic colloid (4% albumin) or vasopressin, achieve O2 sat goals
-sepsis management bundle: corticosteroids, tight glycemic control

56

failed SIRS/DS tx

eritoran tetrasodium: anti-TLR-4 compound
Drotecogin alfa: act. recombinant protein C, approved, no proof of benefit

57

detection of LPS in pharm industry

Limulus Amebocyte lysate test and/or monoclonal Abs (MoAb) against LPS (detects nanogram amounts of endotoxin)

58

G+ peptidoglycan layer

larger (50% cell wall) and more cross linked than G-
can induce TNF-a, IL-6
can lead to SIRS/shock/DS (not as much as LPS)

59

Lipoteichoic acid structure

polymer of glycerol-PO4 or ribitol-PO4, covalently bound to glycolipid, integrated/NON-COVALENTLY bonded into outer leaflet of CM, extends thru cell wall/pep layer into environ
*adhesion of Strep. pyogenes to fibronectin on surface of pharyngeal epithelial cells

60

Teichoic acids (and other polymers)

polymer of glycerol-PO4 or ribitol-PO4, covalently bound to peptidoglycan, extends thru cell wall/pep layer into environ
*peptidoglycan + teichoic acids-->can produce endotoxin-like shock in pts with Staph aureus infections
*interacts with CRP-->activates alternative comp pathway-->inflammatory response

61

PAMPs (pathogen assoc. molecular patterns)

on pathogen, recognized by PARs/PRRs-->can initiate release of endogenous mediators that cause SIRS/DIC/DS
*caused DIC/DS to occur in absence of endotoxin: G+ bac inf., fungal inf., viral inf.

62

PARs (PRRs)

NOD1/2: internal/cytoplasmic *NOD2 def. related to Crohn's
TLR rec: extracellular, 11 total, bind to sp. PAMPs: peptidoglycan, teichoic acid polymers, N-f-met-leu-phe, CpG nucs, LPS

63

bacterial spores

-most bac do not form spores, help survive adverse conditions, forms inside mother cell (dies), complete but inactive cell in protective shell, germination when conditions are favorable

64

bacterial endospores have increased

longevity, resistances to heat/temp, desiccation, chem agents *req special disinfectants/sterilization

65

medically important spore-forming bacteria

Bacillus
Clostridium

66

small colony variants (SCV)

growth-def variants, form colonies 1/10 normal size
enhanced resistance to Ab (*aminoglycosides)
formed by both G+/G-: Staph aureus, Pseudomonas aeruginosa, E.coli, UTIs
phenotype switching: able to revert to normal size

67

SCVs are associated with chronic, recurrent infections of

bones, heart, lungs, urinary tract

68

bac are good metabolizers b/c

large surface:volume ratio, close contact w. environ, accumulate nutrients quickly, grow rapidly

69

autotrophs

"fix CO2", cell energy from redox of inorganic ions (chemoautotroph) OR harvesting light energy (photoautotroph)

70

heterotrophs

oxidize organic molecules for cellular energy
*all bac which cause disease in humans

71

heterotrophs utilize..

carbs, proteins, then lipids

72

fastidious bacteria

will not grow on blood agar (complex growth req)
non-fastidious do

73

optimal growth occurs at temps..

closer to maximum
min determine primarily by reduced enzyme activity and red. mem fluidity
max: protein denaturation

74

mesophile growth occurs btw

20-50 degrees C
*most pathogens are mesophiles: 35-36 C

75

thermophiles

(obl. or fac) >55 C

76

psychrophiles/cryophiles

(obl, fac)

77

microaerophilic organisms

grow in presence of red. O2 conc.

78

facultative anaerobe

*most pathogens
aerobic respiration when O2, otherwise fermentation
-early on use aerobic, consume all O2, switch to anerobic

79

aerotolerant anaerobe

grow best in absence of O2 (only fermentation) but can grow in O2

80

obligate anaerobe

only grow in absence of O2 (only fermentation)

81

bacteria become O2 tolerant when..

they produce enzymes (SOD, catalase) to detoxify the byproducts of O2 metab (O2-, H2O2, respectively)

82

if bac are O2 intolerant

they lack nec. enzymes, so toxic agents (O2-, H2O2) kill them

83

special ionic requirements

non-halophile vs halophiles (need Na+)
eg:
low Fe+++: C. diph-->produces diphtheria toxin
low Ca++: plague bac-->produces exotoxin
low Mg++: S. aureus strains-->TSST-1

84

glycolysis

catabolic pathway; partially oxidizes organic matter-->end products are substrates for other pathways; phosphorylation generates ATP
*additional energy if end-products enter:
-TCA(gen red power (NADH2)-->respiration (gen ATP, recyc. NADH2)
-fermentation pathways

85

TCA

completes oxidization of organic carbon into CO2
gen. intermed. for anabolic pathways
gen. reducing power (NADH2) for ana/catabolic pathways
FADH2 and/or NADH2 is recycled in resp/anabolism

86

fermentation pathways

produce shorter C chain-org comps +/-CO2
recycle NADH2 for ana/catabolic pathways
may gen ATP via substrate level phosphorylation* sole source energy

87

respiration must occur

in mem vesicle/sack, generates energy: ion current (PMF) for ATP synthesis, occurs during recycling of NADH2-->NAD+(oxidized) *Abs exist that collapse the gradient

88

1st part of respiration: ETC..

transfers e-s and H+ from NADH2 to TEA-->generates both PMF and reduced TEA

89

oxidase test

determines presence of ETC comp (cytochrome C) in some bac that can oxidize derivatives of P-phenylenediamine to a colored product
-used by lab to ID bac: enterobacteria (oxidase -)
other G- rods (oxidase +)

90

2nd part or respiration: ATP synthetase...

uses PMF to synthesize ATP from ADP and inorg. phos

91

aerobic respiration

(oxidative phosphorylation)
TEA is O2-->red. to H2O by ETS
*common pathway among pathogens and humans

92

anaerobic respiration

TEAs are inorganic comps
med sig:
-methemoglobinemia (MetHb) happens when elevated levels of NO3 are in drinking water
-GI tract NF convert NO3-->NO2-->if absorbed in blood-->MetHb (risk esp to fetus)

93

fermentation

simpler than respiration, incomplete oxidation of C substrate, utilizes substrates less efficiently (NO respiration), but still allows growth, occurs in cytosol (NOT vesicle), does NOT directly produce PMF

94

fermentation substrates are..

partially oxidized to 1-4 C compounds and some CO2, these serve as TEA (accept e- from NADH2, H+) during recycling of NADH2-->NAD+
-->then excreted from cell (pyruvate-->ethanol +CO2)

95

microbial end products of fermentation cause..

dental caries; end prod. is lactic acid from homolactic fermentation (like hum. musc.)

96

microbial end products of fermentation lead to...

acidic pH of vagina and skin (lactic acid, again, and propionic acid, acetic acid, CO2)

97

microbial end products of fermentation cause abscesses..

that are acidic and anaerobic
-many Abs not effective at low pH
-many Abs bind free NAs and render them unavailable
-low pH kills surrounding viable human cells -->rel. compounds that bac req for growth (para-aminobenzoid acid) -->Abs like sulfas are ineffective

98

microbial end products of fermentation help..

ID bac
-mixed acids: lactic, acetic, formic, succinic, etOH, CO2, H2
-typical of enteric bac of human gut (coliforms)

99

certain bac can only grow fermentatively

-lack cytochrome/ETC OR cannot use it for energy prod.
-do not use NAD+ or NADP+ as e- and H+ ion carrier, rather ferredoxin is used-->must be recycled to oxidized form

100

aerotolerant anaerobes

(Strep and Lactobacillus)
-produce lactic acid and H2O2 (from ferredoxin recycle)
H2O2 detoxed by human host's peroxidase (otherwise can't grow)

101

fermentation by Clostridium

end products include H2, CO2 and 4 C compounds; recycling of ferredoxin catalyzed by hydrogen lyase (not aerotolerant?) -->H2 byproduct

102

H2 gas produced by Clostridium's fermentation can cause..

Gas gangrene! (myonecrosis)
H2 is insoluble in tissues, tracks along fascial planes (sep muscles, collapses blood vessels, impeding perfusion (anaerobic)

103

alkaline end products are NOT

fermentative or oxidative phosphorylation end products
i.e. Proteus spp. (cause UTIs, kidney stones) rel. urease-->hydrolyzes urea in urine-->ammonia and CO2-->raises pH to above 7, allows Proteus to grow-->Ca++ and NH4+ ions form salts, precip. at alkaline pH-->renal calculi

104

renal calculi are composed of

triphosphate: Struvite: Mg ammon phos)and poorly crystalline form of apatite (hydroxylated Ca phos (some repl by carbonated)

105

H. pylori cause..

type b and duodenal ulcers
produce urease-->cleaves urea to CO2, NH4+-->raises microenvironment pH (stomach mucous lining) so bac can grow

106

primase

synthesizes short ssRNA primers for DNA synthesis

107

DNA gyrase (topoisomerase II)

negatively supercoils bac genome and plasmid DNA
relieves torsional stress cause by helicase: "unwinding"

108

topoisomerase IV

required for decatenation (separation of 2 daughter chromosomes (rings))
*both topo. II and IV are essential for bac DNA sun

109

DNA synthesis originates..

at one origin of replication in prokaryotes
is bidirectional (as is eukaryotic)

110

DNA synthesis must be primed with..

RNA
primes synthesizes primers

111

partitioning of daughter strands

-in prok: req membrane attachment
-in euk: utilize spindle fibers and centromere to sep. each chromosome

112

DNA synthesis occurs when

prok: lag and exponential phase
euk: S-phase

113

RNA synthesis produces

mRNA, tRNA, rRNA

114

why antimicrobial protein synthesis inhibitors are so effective

mRNA in bac has a v. short 1/2 life

115

some gene transcription req. DNA gyrase

to neg. supercoil DNA

116

protein synthesis occurs in the

cytoplasm

117

bac ribosomes

free, 70S (30S + 50S)
rRNA + proteins + accessory comps

118

charged tRNAs accomplished by

aminoacyl-tRNA synthetase: covalently bond sp. aa's to appropriate tRNA (mRNA is template)
mRNA is codon, tRNA is anticodon (base-pair)

119

what catalyzes peptide bond formation during which ATP is consumed

rRNA (not proteins)
-->polypeptides are product

120

protein turnover is v. short, so

Abs that inhib port sun are effective

121

biosynthesis of peptidoglycan(PTG) backbone of cell wall:
1. synthesis of amino sugars/PTG subunits

uracil diphosphate (UDP) is tag for directed synthesis of amino sugars/PTG subunits
NAG and NAM syn. occurs while cov. bonded to UDP
peptide side chain of NAM syn. by ind. enzymes

122

biosynthesis of peptidoglycan backbone of cell wall:
2. assembly and translocation of PTG subunit thru CM

PTG subunit formed by transfer of NAM and then NAG from UDP-->bactoprenol (Lipid P (carrier)) w/ release of UMPs
PTG subunit then shuttled thru CM to growing end of PTG chain

123

addition of subunit to PTG chain occurs via..

transglycolase enzyme (transglycosylation)

124

biosynthesis of PTG backbone of cell wall:
final cross linking (transpeptidation)

done by transpeptidases (penicillin binding proteins (PBPs)
2 aa subunits from each peptide side chain covalently bonded-->"fabric shell": mech. strength and rigidity to PTG

125

G+ bacteria use unique enzymatic pathway to make heme

HemQ, used in final step
MRSA, enterococcus, listeria, Mtb
*selective toxicity

126

lag phase

cell volume and mass increase, chromosome replication (DNA syn) begins
*NO cell division/change in #

127

exponential/log phase

balanced growth occurs
cell number, mass, volume, and cell comp. amounts increase by same exponential factor

128

exp./log phase expressed as

generation (doubling/replication) time (GT): time req for one bacterium to divide into 2 cells (replicate)

129

stationary phase

no net increase in cell numbers occur

130

death phase

cell death occurs at logarithmic rate
most bac autolyse (everything is gone)

131

rapidly growing organism produces

acute disease (fulminant) -in general
short mean GT
high Ag dose, strong IR
anti-infective tx usually 8-10 days

132

slow growing organism produces

chronic disease (insidious) -in general
long mean GT
low Ag dose, weak IR
anti-infective tx prolonged

133

chemotherapy that inhibits protein and/or PTG synthesis is most effective against

fast growing organisms

134

what DOES NOT determine GT/growth rate

Gram stain, metabolism, ext. cell structures, spore production

135

genome

chromosomes PLUS any extrachromosomal elements (plasmids) deemed crucial to the organism
-in bac: sing or doub stranded, covalently closed, circular

136

replicon

DNA or RNA molecule that controls its own replication, can self-duplicate

137

extra chromosomal elements (ECE)

replicons in cell, except host DNA

138

genomic islands are

horizontally transferred?...

139

plasmids

typ. ds, much smaller, code for ancillary genes, replicons
control their own DNA replication and copy number

140

plasmids DO NOT code for

housekeeping genes (req for viability)

141

plasmids replicate in ..

the cytoplasm, utilize host bacterial cell DNA replication machinery

142

conjugative plasmids

encode for mech to transfer a copy of itself from donor cell to recipient cell

143

resistance plasmids

possess Ab resistance determinants

144

plasmids can be..

acquired or lost from bac cells (non essential info) and are a metabolic burden on the host

145

bacteriophage

virus replicons (DNA, RNA) which infect bac cells
*can exist latently in bac cells as a prophage

146

prophage can be..

1. a plasmid in bac cytoplasm (ECE)
2. integrated into bac cell chromosome
3. can encode genes that confer a new phenotype to their host bac
*most abundant bio entity on earth!

147

recombination

exchange of recipient DNA w/ donor DNA
-breakage and joining of replicon DNA molecules to form hybrid, recombinant molecules
-can only occur in cell

148

homologous/legitimant recombination (prok)

-donor DNA integrated into rec. chrom. and excised rec. DNA fragment is degraded
-allows gene transfer/exchange, esp. w/ transformation or abortive transduction

149

homologous/legitimant recombination requires

RecA, an enzyme which func. when donor and rec. DNA segments share sig. homologous sequences

150

site-specific/DNA seq specific
(AKA illegitimate/non-homologous recombination)

insertion of or replication of genetic elements in DNA w/ recombination restricted to identical sites at 2 locations on one replicon or at identical locations on 2 replicons

151

site-specific/DNA seq specific does NOT require

RecA

152

several types of site-sp recombination mediated by mobile genetic elements

insertion sequences
transposons

153

since genetic elements cannot move themselves from donor to recipient in site-sp recombination, this must occur

Horizontal gene transfer (HGT) w. genetic element carried on transferred DNA

154

genetic apparatus of bacteria

chromosome: structure, size, number
ECEs: bacteriophages (prophages in bac cells), plasmids

155

phenotypic variation

an event in which all cells in a population respond to environmental stimulus in the same what which produces a new/altered phenotype via gene expression, without change in genotype**no genotypic variation is involved!

156

the total potential phenotype is limited by

the organism's genotype
*expressed phenotype is usually LESS than the full genotype potential (genotype can encode for more traits then the phenotypic traits currently express)

157

phenotypic variation occurs b/c

microorgs are exposed to radically diff environments, req different phenotypes

158

microorg does not express its entire phenotype at one time b/c

it would require enormous energy expenditure and cell mass (would be out-competed by more reserving, environ-appropriate cells)
i.e. fungus switches from minimal capsule-->max capsule when in human host-->cause meningoencephalitis

159

phenotypic variation occurs via

-reg proteins controlling transcr/translation of sp. operons
-2 component signal transduction via sensor kinase and response regulator
-quorum sensing
-Ag variation
(and other)

160

genotypic variation

event in which genome of 1+ cells in population is altered
(acquire new genetic information)

161

2 mechanisms of genotypic variation

-internal by mutation (NO foreign/donated DNA involved)
or
-external: trasformation, conjugation, transduction
-->transduction: generalized/abortive or lysogenic conver

162

mutation (internal alteration of genotype)

-internal alteration of genotype
rare, but happens due to so many bacteria

163

mutations create new genes, but do not transfer them; require this mechanism to transfer

Horizontal gene transfer
(dynamic duo!)

164

horizontal gene transfer

transformation, transduction, conjugation

165

transformation

DNA fragments released by donor cell autolysis and accumulated by recipient cell (donor dies!)

166

transduction

abortive phages carrying donor cell chromosomal fragments transfer their DNA to the recipient cell

167

conjugation

donor cell plasmid encodes for mechanism to transfer a self-copy of itself to recipient cell which lacks the plasmid

168

genetic info is either..

carried/contained:
-plasmid: conjugation, transformation
-gen/abortive bacteriophage: transduction
-lysogenic bacteriophage: transduction
OR "naked" :transformation

169

external genetic information exchanged when..

-highly homologous genes form donor replace (recomb) the corresp. chromosomal or plasmid genes in the rec cell
-plasmid carrying new/altered genes makes a home in the rec. cell
-latent bacteriophage/prophage (carrying new/altered genes) makes a home in the rec cell by 2 mechanism..

170

2 ways a latent bacteriophage/prophage can make a home in the rec cell

-integrating into rec cell's chromosome
-functioning as a plasmid in the rec. cell's cytoplasm

171

clin sig of HGT

can transfer Ab resistance
-Acinetobacter baumannii: major cause of hops-acq inf

172

transformation

uptake of "naked" EC DNA (fragment) by rec cell by mech encoded by rec cell's chromosomal genes
-donor cell autolyses
-can be accomplished in vitro (HSSN) and imp. to certain genera: Strep

173

only what types of cells can acquire "naked" extracellular DNA

competent:
-naturally competent: chromosome of rec. bac cell contains genes that encode for acquisition of extracellular DNA
-"forced" competent: chem/phys tx which "force"/induce bac cell to acquire EC DNA by unknown mech*
*used in recomb DNA technology (G- enteric bac E. coli)

174

transformation part 1

rec accumulates EC DNA of any origin (bac chromosome or plasmid)

175

transformation part 2

integration of acquired DNA requires RecA (i.e. mediates homologous recombination)
any accum. DNA fragments not integrated into host (rec) cell replicon (host genome/plasmid) cannot replicate and is degraded and consumed for carbon and energy

176

HGT mechs that DO require RecA

transformation
generalized/abortive transduction (SOMETIMES-if homolog.)

177

HGT mechs that do NOT require RecA

conjugation
generalized/abortive transduction (SOMETIMES)
lysogenic conversion

178

conjugation

donor cell plasmid encodes for mechanism to transfer copy of itself (the plasmids) to rec cell
-incidental transfer of donor cell chrom DNA v. rare, may not be possible
-involves ECE (plasmid)

179

1st mechanism of conjugation: in G- bac

ex: transfer of resistance (R-factors)
-"mating": cell to cell contact is essential- sex pili functions to make contact in some, not all conditions
-ssDNA copy of R factor transferred through mem pore from donor to recipient cell during replication of R factor
-transferred DNA made double stranded in rec cell
-NO RecA OR site sp. recombination involved

180

limitation of conjugation

limited bacterial host range

181

limitation of transformation

not all bac cells are naturally competent

182

transduction

horizontal transfer of information (chromosomal, plasmid DNA, latent virus bearing a particular gene) btw bacteria mediated by bacteriophage

183

lytic phage

infects bacterial host cell to generate a productive infection(new virions) by binding to a sp. comp. on cell surface-->penetration of phage NA (DNA, RNA) into cytoplasm-->replication of NA-->transcription & translation of phage core and coat genes-->phage components self-assemble into infectious particles-->host cell releases new inf. particles (virus)

184

new infectious particles produced in lytic phage transduction are release into medium via

virus encoded cell wall hydrolase(PTG hydrolase)->cell lysis
OR slow release without lysis

185

temperate phage

infects host cell by binding to sp comp on cell surface-->penetration of phage NA into cytoplasm--> then 2 options for phage..

186

temperate phage after penetration

1. phage can undergo normal lytic cycle as above OR
2. phage can become latent

187

latency

involves repression of phage genes which code for lytic (productive) cycle of phage replication-->will reside as plasmid or is integrated into host cell DNA-->replicates in synchrony with host cell DNA & passed on to daughter cells-->are now "lysogenized"-->latency ends when phage genes become productive again: replicate and lyse host cell

188

lysogenized

bac strains with prophage DNA

189

2 options for temperate bacteriophage replication (again):

1. lytic cycle will result in a productive viral infection with lysis of host cell
2. temperate cycle viral reproduction habbpens via replication of lysogenic virus genome and distribution to each daughter cell

190

mechanisms of transduction (HGT)

generalized/abortive transduction
lysogenic conversion

191

which form of transduction occurs is determined by

the vector (bacteriophage)

192

generalized (abortive) transduction

occurs with defective phage particles of both lytic and temperate phages and may require RecA

193

gen (abort) transduction: during viral lytic replication cycle..

the donor genome is sheared into fragments-->v. rarely; the DNA or intact plasmid is randomly "packaged" into virus particles: pseudovirions/abortive phages-->function as normal infectious virion: attach to rec on uninf. rec cell-->pkgd DNA is introduced into the rec cell cytoplasm-->any DNA frag not integrated or unable to replicate is degraded and consumed

194

in generalized transduction, the pseudovirion/abortive phage DOES NOT

carry viral genome, so no lytic or temperate life cycle can occur in rec cell
*NO viral reproduction can occur!

195

unlike a typical virus, the packaged DNA of a pseudovirion is a..

-plasmid that can replicate in rec cell
-chromosomal fragment that shares homology with a portion of the recipient chromosome and is integrated into that chromosome via RecA
-plasmid fragment that shares homology w. portion of rec plasmid and is integrated via RecA
*no site sp. recombination is involved!

196

lysogenic conversion is mediated ONLY by

a temperate bacteriophage (rec. is inf by temperate virus)

197

lysogenic conversion

when rec cell possess a new phenotype/trait due to acquisition of a prophage (latent temp. phage) which encodes for the new phenotype *can be expressed without activation of the temp. virus genome!*

198

does lysogenic require RecA

No

199

lysogenic conversion process

temp. phage encodes for an exotoxin gene- not req for viral replication or any part of viral infectious cycle

200

the exotoxin gene

is reg sep&ind of the viral genes
can be expressed without altering repression of the phages genes which code for the lytic cycle (does not affect latency)
-changes the cell phenotype from exotoxin negative to exotoxin positive

201

clin sig of transduction

transfer of drug resistance
lysogenic conversion: prophages can carry genes for toxin production

202

bacterial genomes consists of a conserved core gene pool

most encode "housekeeping" proteins
exhibit homogen. G+C contents/codon usage

203

bacterial genomes also consist of flexible gene pools

204

the functional (flexible) gene pool consists of

genomic islands (GEIs, >10kb) related to mobile genetic elements
genomic islets (

205

genomic islands (aka fitness islands) traits

presence of:
-residual material from mobile genetic elements
-flanking direct repeats
-genes that aid in an org's adaptation (put/virulence funcs)
carry fragments (transfer genes) of other mobile elements (phages, plasmids)
large: >10Kb up to 100Kb

206

genomic islands are inserted in

the vicinity of tRNA sequences or other small RNA genes, which leads to instability, risk for excisement

207

genomic islands can be differentiated from native genome

has evidence of horizontal/lateral origins:
-abnormal %GC index, dinuc frequency diff, codon usage bias

208

GEIs help an org survive a certain ecological niche, i.e.

PAIs: pathogenicity islands carry type 3 and 4 sec sys
Ab resistance islands

209

DNA polymorphism answers

what/whom is the SOURCE of the infection, does NOT identify the agent present
ex: plasmid analysis (GE), RFLP/ribotyping, PFGE

210

in plasmid analysis, plasmid DNA is

isolated, purified, and may be treated with restriction endonucleases

211

gel electrophoresis

separates plasmid DNA/fragments by size-->stain, take pic
*bac must possess plasmids!

212

restriction fragment length polymorphism (RFLP) and Ribotyping

bac chromosome (DNA) is isolated to fragments and purified-->treated with restriction endonucleases to fragment (again?)-->gel e-phoresis used to sep DNA fragments-->compare genomes (not used for banding patterns-too many!)

213

RFLPs and Ribotyping uses this as detection

Southern or Northern blots-->labeled probe is hybridized to sep. DNA fragments-->detection

214

in RFLP analysis, the probe detects

a particular sequence
*used for cellular life-forms: prok and euk

215

in ribotyping, the probe detects

DNA encoding for rRNA; which is unique for each genera/spp
*used only for cellular life-forms: bacteria

216

pulse field gel electrophoresis

variation of RFLP BUT inf. agent's cells are gently lysed to rel. their chrome. DNA INTACT (vs. fragments)-->dig w/ rester endonuclease reg rare sites so large DNA frags-->sep by special agarose GE where the e- field orientation is changed often ("pulsing")

217

PFGE produces restriction fragments profile of

5-->20 bands, 10-->800kb, visualized by staining
*restricted to cellular life forms: prok and euk

218

clin sig of DNA polymorph technologies

epidemiological; compare strains of partic pathogens
-typically multiple strains for one infectious agent in a host population
-determine if there is a common source

219

detection of etiologic agents in human specimens:
detection of target NA AND amplification of target NA genome answer..

what AGENT is making the patient ill?
NOT used to answer what/whom is the source (DNA polymorphisms)

220

detection of target DNA

Southern blot

221

detection of target RNA

Northern blot

222

to detect target, use

a single probe for each nat. occurring in vivo target, labeled with either radioactive or non radioactive marker

223

tissue sample prep (w/ target NA)

in situ prep: specimen treated so NA is single stranded and accessible to probe

224

blotting

NA extracted-->dig by restriction endonuclease*-->electrophoresed*-->transferred to support (blotted to membrane)-->denatured (made ss)
*may or may not occur

225

RNA is often chosen as target NA b/c

1000x more (r)RNA than DNA in bac cells
possess unique signature of sequences sp. for genus/spp

226

hybridization

ss, labeled (DNA/RNA) prob anneals w/ ss NA in sample
wash off non-spec. bound probe

227

detection

label probe-->anneals to target NA-->detection!

228

since target DNA may be rare in certain human infections this helps detection

amplification: signal or target

229

signal amplification

produces multiple signals vs. one signal per in vivo target
-mult. non-radioactive marker bind to each target so SIGNAL not target is amplified
-->developed so no need to employ heat stable DNA polymerase/PCR, save $

230

PCR

on form of in vitro or in situ amplification of target DNA or RNA
amplifies NA sequences unable to be detect. by N/S blots directly (10-100x more sensitive, more in vitro targets!)

231

PCR: 3 step cycling process

(if RNA, amp then converted to DNA (cDNA) by reverse transcriptase)
1. denature (heat) ds-->ss DNA
2. anneal primers to ss DNA (cDNA) after cooling
3. extension of primer: amp w/ heat stable DNA polymerase
then detect with Northern or Southern blot

232

PCR clinical applications:

-screen blood
-detect viruses, bac, genetic defect, genetic marker (neoplasm), infectious agent (not successfully cultured)

233

other amplification techniques

LCR: ligase chain reaction
TMA: transcription mediated amplification
*do NOT employ heat stable DNA polymerase

234

do viruses require living cells

YES (some bacteria do not)

235

do viruses divide by binary fission

NO (bac do)

236

do viruses have both DNA and RNA

NO (bac do)

237

are viruses susceptible to Ab

NO (bac are)

238

exception: mimivirus and CMV

contain BOTH RNA and DNA

239

viruses are

submicroscopic entities
obligate intracellular parasites
infect specific living cells and reproduce in said cells
consist of DNA OR RNA and protein, often an envelope
have a definite structure

240

although relatively simple replicons, viruses..

can take over host cell, utilize for own replication and self-assembly, do not always kill the host they infect

241

viruses are small

15-->300 nm, observed by e- microscope not light

242

virion

complete viral particle, able to infect another host cell and repeat the replicative cycle

243

for any one virus, each infectious virion possess the same NA..


-dsDNA (w/ ss regions)
-ssDNA:
-+ssRNA (positive sense polarity)
--ssRNA (negative) (seg/nonseg genome)
-dsRNA: 2 identical or complimentary strands

244

the NA genome is either...

linear or circular
usually condensed by histones or histone-like proteins

245

protomer

protein subunit of capsid, repetitive polypep subunits arranged in symm patterns, either:
1. protomers-->capsomers-->capsid (i.e. icosahedral capsid)
2. protomers-->capsid (i.e. helical capsid)

246

capsomers

oligomeric clusters of protomers
form the capsid
(in orgs w/ icosahedral symmetry)

247

capsid

protein shell or coat of protomers that self-assemble by non covalent bonds to enclose the core of NA genome + associated proteins

248

3 architectural structures of capsids

complex, cubic/icosahedral, helical

249

complex symmetry

viruses w/ unresolved structures b/c too complex

250

cubic symmetry of all animal viruses is icosahedral which

consists of 20 faces, each an equilat triangle, in most protomers are arranged into capsomers

251

helical symmetry

capsid consists of multiple copies of single species of protomer that bind to each other and ssRNA genome

252

helical structure of non-enveloped virus

individual protomers assembled in helical structure around NA genome of virus-->forming the nucleocapsid
-RNA genome extends the entire length
-the core is hollow
-each helical virus has sp diameter/length

253

nucleocapsid

capsid + NA genome + associated proteins
*stable structure; resistant to drying, mild acids, detergents

254

some virus consist solely of a..

nucleocapsid

255

envelope

viral membrane that covers/encloses the nucleocapsid (not always present)

256

envelop composition

virus-encoded proteins
host-derived lipids and carbs

257

2 important proteins in viral envelope:

peplomer= spikes: viral glycoproteins, play a role in virion-rec attachment
layer btw envelope and nucleocapsid that mediates interaction btw capsid and envelope:
-tegument: amorphous layer, complex funds
-matrix protein: structure-providing lattice

258

viral membrane derived from

nuclear membrane, sub-cellular organelle membranes, cytoplasmic membrane
-obtained from host during "budding" of some viruses

259

enveloped viruses are...

inherently fragile, sn/inac by drying, detergents
(the NUCLEOCAPSID is more stable)

260

disruption of envelope of enveloped virus leads to

virus inactivation, no longer infectious

261

nearly all enveloped viruses are transmitted via

arthropods
respiratory droplets
bodily fluids: semen, saliva, secretions
*non-env. viruses may also be transmitted these ways

262

how to classify animal viruses

type of nucleic acid with polarity
type of capsid symmetry
+/- envelope
quantitative: # of capsomeres for icosahedrals, diameter of helix for helicals

263

clade

group containing all descendants from a given common ancestor, grouped based on seq similarity among members and seq diversity w.in total viral population
*NOT a serotype
**the clinical importance is immune evasion

264

serotypes are based on

antigenic diversity

265

all human viruses that successfully replicate in humans result in ? infections but only some also produce ? infections

productive
persistent

266

non-productive

no infectious virions:
-abortive (lack infectious viral synthesis post-absorption?)
-interference: virus interferes w. growth of other viruses in same cell

267

productive infections comprised of..

1. non-lethal alteration of cell & functions
2. cell damage/death: lytic infections: via viral replication
3. persistent infection without cell death

268

persistent infections without cell death..

may persist for years
-latent: intermittent acute episodes of disease, periods of NO infectious particles (HSV, varicella-zoster)
-chronic: continued viral presence, disease may be absent (CMV, HHV, HBV) or is assoc. with late immunopathologic disease: HBV-->cirrhosis of liver or primary hepatocellular cancer
-slow: long incubation period, slowly progressive, lethal (SSPE, measles) no infectious virion may be detected!

269

the most common outcome of viral infection is

asymptomatic infection with seroconversion

270

mechanisms which cause injury in viral infections

-immunopathology
-autoimmune induction
-cell damage/death (lytic infections)

271

lytic infections cause cell death

viral replication modifies and damages host cell:
-inhibits/shuts down macromolecular sun
-cytopathic effect (CPR); toxic
-inclusion bodies and cell fusion (syncytia formation-multinuc giant cells)
-induce apoptosis
-chromosomal alterations-->oncogenesis

272

characteristics of tumors

-transformation to unrestricted cell growth
-loss of contact inhibition/senescence (unreg growth)
-appearance of new Ags (tumor sp Ags)
-other changes: metabolic, genetic

273

viral agents assoc with malig neoplasms

RNA tumor viruses HTLV
DNA tumor viruses: HPV, EBV, HBV

274

viral agents of benign neoplasms

(somewhat organized growth, does not invade)
human wart viruses: verruca lesions, condyloma acuminatum
poxvirus: molluscum contagiosum

275

6 stages of replication

attachement-->penetration-->uncoating-->macromolecular synthesis-->viral genome replication-->translation of viral transcripts-->synthesis of other/non-protein comps

276

virus-rec interactions are major determinant of

infectivity
-specificity determined by:
host range (low affinity rec)
tissue tropism (high affinity rec)

277

HIV therapy target (Fusion inhibitors)

inhibit HIV virion fusion w/ human cell surface receptors-->prevent infection of individual cells

278

penetration

direct
surface eclipse
rec-mediated endocytosis

279

direct penetration

non-env. viruses, nucleocapsid attaches cell surface-->release viral NA genome into host cytoplasm

280

surface eclipse penetration

pH-independent cell entry
enveloped virus membrane fuses with cell membrane releasing viral genome into cytoplasm (*penetration and uncoating happen in 1 mechanism)

281

rec-mediated endocytosis

viroplexis: pH-DEPENDENT cell entry
attachment of enveloped virion to cell rec-->endocytosis of virion-->enclosed in endosome-->acidification-->virus env. fuses with vesicle membrane-->releasing viral genome into cytoplasm

282

uncoating

removal of protective coats with release of NA
*infectivity lost here!!*
-some virus AND host encoded mechanism to uncoat
-target of antiviral therapy

283

macromolecular synthesis

need to make more genome and more viral proteins:
1. DNA viruses replicate in nuc and RNA in cytoplasm
-exp. pox (DNA) virus in cytoplasm, influenze (RNA) in
nuc
2. +ssRNA, -ssRNA, and dsRNA virions all must encode for an RNA-dep RNA polymerase (RNA POL)
-exp. retroviruses: reverse transcriptase: RNA
dependent-DNA polymerase
3. both -ssRNA and dsRNA infectious virus must also carry function RNA POL (the protein) to make the mRNA along with possessing the gene
-+ssRNA viruses DON'T carry RNA POL

284

DNA viruses replicate genome utilizing

host cell DNA replication machinery

285

RNA viruses

+ sense or - sense RNA genome replication produces many new viral genomes and produces RI (req. RNA POL, virus encoded)

286

retroviruses are the exception

-encode for reverse transcriptase (RT; RNA dep-DNA POL)
transcribes +ssRNA-->DNA-->incorp. into host genome-->viral DNA transcribed into mRNa

287

RT is target for many anti-HIV drugs

-nucleoside analogue RT inhibitors
-non-nucleoside RT inhibitors
integrase enzyme also target (integrate inhibitor prevents insertion of HIV DNA genome into human genome)

288

primary replication

site or sites in host where 1st replication occurs
-is it near the portal of entry? (POE), related to incubation
period

289

secondary replication

where replication occurs after spread from primary site (not present in all viruses)

290

some not all viruses spread from

the POE, primary site

291

local spread

always occurs

292

when does spread occur?

before, during, after entry, or only local
before, during or after primary replication

293

spread/replication within host

cell to cell within tissues
in bloodstream/lymphatics
in nerves: peripheral-->CNS
transplacentally

294

tissue tropism

specificity of a virus for a particular host tissue
*major determinant of infectivity!

295

signs and symptoms may only occur..

during the primary and/or secondary replication

296

immune response to viral infection

outcome related to capacity of virus to damage host and disrupt host defense mechs

Decks in Micro Class (61):