MFD Theme 1 Flashcards
1
Q
what is evolution
A
process of change over time that results in new varieties and species of organisms
2
Q
what is phylogeny
A
Evolutionary relationships between organisms
3
Q
how can relationships be deduced in different specimens
A
by comparing the genetic information
4
Q
what is excellent for determining phylogeny
A
ribosomal RNA (rRNA)
5
Q
where is rRNA present
A
in all cellular life forms
6
Q
what is contained in rRNA that can be used to differentiate between species
A
highly variable regions
7
Q
what is the benefit of DNA sequence analysis
A
it provides an insight into evolution and diversity of the organisms
8
Q
how can you measure evolution and diversity
A
- isolate DNA from each organism
- make copies of rRNA gene by PCR
- sequence DNA
- analyse sequence
- generate phylogenetic tree
9
Q
what dominates the tree of life
A
microorganisms
v few macroorganisms
10
Q
what are the classifications in the tree of life
A
Domain Kingdom Phylum Class Order Family Genus Species Strain
11
Q
what are the top taxonomic levels
A
Archaea Bacteria Eukaryota Viruses Viroids
12
Q
are prions living or non living
A
non-living proteinaceous infectious agents
13
Q
what are Protozoa and fungi
A
groups within the eukaryotes
14
Q
what are prions
A
Misfolded proteins that cause correctly folded proteins to gradually fold abnormally
15
Q
when does disease occur as a result of a prion
A
when a host-encoded a-helical protein (PRPC) is converted to b-pleated sheet form (PRPSc)
16
Q
what does Exogenous PRPSc do
A
triggers disease
17
Q
what is further conversion of PRPC
A
autocatalytic
18
Q
what can prions cause
A
transmissible spongiform encephalopathies (e.g. vCJD)
19
Q
what do transmissible spongiform encephalopathies do
A
Loss of voluntary and autonomic function
Degeneration leads to death
20
Q
what is PRPSc is extremely resistant to
A
heat (including autoclaving), proteases etc
21
Q
why is extreme care needed in the case of PRPSc in regards to dentists/medics
A
iatrogenic transmission
22
Q
what are some characteristics of viruses
A
Can only replicate inside cells using cells’ machinery
Small genomes
Evolve rapidly
23
Q
what are some examples of viruses
A
HIV, herpes simplex, influenza, cytomegalovirus
24
Q
what is Archaea
A
Previously thought to be a type of bacteria but actually are closer to eukaryotes
25
what features do archaea share with eukaryotes
DNA replication is initiated at multiple origins
26
what is the role of archaea in the mouth
unknown
27
which patients are archaea particularly found in
patients with severe periodontal disease
28
what is protozoa
Single celled eukaryotes
29
where are protozoa found
Single celled eukaryotes
30
what do protozoa do
Graze on bacteria
31
what are common examples of protozoa
amoeba, plasmodium
32
what are the Oral protozoa
Trichomonas tenax and Enamoeba gingivalis
33
what is the only serious pathogenic oral protozoan
Leishmania
34
what are fungi
Simple eukaryotes
35
which fungi can form multicellular structures
mushrooms, pin moulds
36
what forms do some fungi transition between
yeast and hyphal forms
37
how are microbial processes related to the environment
```
Creation of the Earth’s atmosphere
Nitrogen cycle
Decomposition
Metal ore recovery
Sewage digestion
```
38
how are microbial processes related to disease
Man, animals, plants
39
how are microbial processes related to food and drink
Beer, bread, wine, vinegarm cheese, yoghurt
40
how are microbial processes related to Biotechnology
```
Antibiotics
Therapeutics
Genetic engineering
Human gene therapy
Production of drugs, e.g. insulin
```
41
what are prions
misfolded proteins which form aggregates: proteinaceous infectious particles
42
why are prions 'infectious'
they spread from cell to cell and in between individuals via contaminated food, blood and surgical instruments
43
what neurodegenerative diseases can prions cause
Cretzfeldt-Jakob Disease (CJD), transmissable spongiform encephalopathies (TSEs)
44
how are prion diseases caused
by proteins whose misfolding is infectious
45
outline what happens after a protein undergoes a rare change to give an abnormally folded prion
- the protein form of the protein can bind to the normal form making it an abnormal prion
- abnormal prion proteins propagate and aggregate to form amyloid fibrils
46
what can amyloid fibrils do
they disrupt brain cell function causing neurodegenerative disorder
47
what is significant about the amyloid fibres
they're protease resistant and resistant to autoclaving for long periods
48
what does stacking of Beta sheets allow for
some misfolded proteins to aggregate into amyloid fibres
49
what are some protein diseases in man
spontaneous- sporadic CJD
inherited- familial CJD
acquired- vCJD, Kuru
50
what are the consequences of of protein diseases in man
- very long incubation periods- slow replication
- symptoms progress rapidly and disease is always fatal
- form insoluble aggregates visible as plaques in tissue
- neuronal dysfunction and tissue death visible as holes
51
what don't prion diseases do
induce an inflammatory response
52
what is the most common prion disease
sporadic CJD
| occurs at a freq of 1-2 per million population worldwide
53
what can other misfolded proteins e.g. amyloid proteins contribute to
non-infective tissue degenerative disorders e.g. alzheimer's, parkinson's, huntington's, atherosclerosis, type 2 diabetes
54
what are enveloped proteins
have a nucleocaspid containing the viral genome lipid layer from host with envelope proteins
55
what are non-enveloped proteins
nucleocaspid proteins only
56
what do viral genomes encode
viral structural proteins
proteins that interact with the host:
-proteases, DNA/RNA polyermase, reverse transcriptase, immune system inhibitors-see IAH
57
*what are the structural properties of viral genomes RNA and DNA
single stranded
| double stranded
58
*what shape are viral genomes
linear, circular or segmented
59
why do viruses have a variety of replication mechanisms
have many different nucleic acid structures
60
what are most viruses that cause human disease
double stranded DNA (e.g. herpes viruses) or single stranded RNA (e.g. retroviruses such as HIV)
61
what is the life cycle of a virus with a dsDNA genome
- The virus first enters the cell by binding to receptors on the surface of the cell
- The virus’ DNA is released into the cell
- The DNA is replicated to produce multiple copies
- The copies are transcribed and translated to produce new viral particles (coat protein)
- Assembly of the new virus particles occurs
- Viral particles then break free out of the host cell by causing cell lysis
62
what is the life cycle of retrovirus (ssRNA genome)
- Once the retrovirus has entered the host cell, reverse transcriptase enzyme is used to converted the viral RNA to DNA
- The reverse transcriptase then synthesises a complementary DNA strand to make dsDNA
- The DNA is then integrated with the host DNA
- Transcription and translation occur, producing new viral particles (coat proteins, envelope proteins, reverse transcriptase)
- The new viruses are assembled and then break free from the host by causing lysis
63
what can viruses be classified into
DNA
RNA
Subcategories
64
what are the types of RNA and and examples
enveloped- HIV/influenza A/B
| non enveloped- Rhino
65
what are the types of DNA and and examples
enveloped- Herpes
| Non-enveloped- Human Papilloma Virus (HPV)
66
what are the subcategories for viruses
SS/DD - nucleocaspid shape
67
how are viruses involved in oral health and disease
some have directly infect the mouth and affect oral health
| others colonise the mouth without directly causing oral disease (following release from other tissues)
68
what viruses have systemic effects but may have oral symptoms
measles (rash-Koplik's spots)
mumps (salivary gland inflammation/swelling of the jaws and neck)
rubella (rash)
69
what are the viral infections of the mouth
- Hand, foot and mouth disease
- Human herpes viruses (HHV)
- Human papilloma viruses (HPV)
70
what are the consequences of hand, foot and mouth disease
enterovirus
(coxsackie A virus) which causes oral blisters
(not the same as foot and mouth disease in cattle)
71
what is the Human herpes viruses (HHV)
```
large class of
structurally related viruses
```
72
what does the human herpes virus family consist of and what are the diseases
HHV1- fever blisters (cold sores)
HHV3- chicken pox/shingles
HHV4- Glandular fever, Hairy leukoplakia, B-cell Lymphoma
HHV5- Congenital defects/pneumonia in AIDS
HHV8- Kaposi’s sarcoma
73
what are the characteristics of HHV1 (HSV-1)
-Infects epithelial cells (oral mucosa) and
neuronal cells
-Exhibits latency
-Causes stomatitis on initial infection, then
fever blisters on reactivation
-Transmission during childhood contact via
saliva
74
what are different primary oral HSV-1 infections
primary palatal herpes
| primary herpetic gingivo-stomatitis
75
what are different recurrent oral HSV-1 infections
herpes labialis
76
what are the characteristics of HHV3 (varicella zoster virus, VZV)
```
-Very contagious-transmitted via aerosols and
direct contact
-Can cause blisters in the oral mucosa
-Primary disease is chicken pox-can be
reactivated as shingles
-Shingles appears in skin along track of nerve
usual on trunk but can be facial (30% of
shingles cases)
```
77
what is an example of primary HHV3 (VZV)
chicken pox
78
what is an example of recurrent HHV3 (VZV)
shingles
| - follows trigeminal nerve (mandibular divison)
79
how many types of HPV are there
over 150 types
| some cause pathology, others are part of the virome of the skin
80
what can some HPV cause
warts of the genital or oral mucosa
81
which HPV's associated with papillomas are also associated with cancers e.g. cervical/oral cancer
HPV-16
| HPV-18
82
which HPV's cause focal epithelial hyperplasia are restricted to oral cavity
HPV-13
| HPV-32
83
what are the viruses present in the mouth that do not directly cause oral disease
Hepatitis B virus (HBV)
| HIV
84
what are the consequences of Hep B
Blood and saliva transmission
- Causes severe liver disease, including hepatocellular carcinoma
- Acute and chronic infection phases
- Carriers often asymptomatic
- Complex pathogenesis
85
what is HIV associated with
- Oral candidiasis
- Necrotising ulcerative gingivitis
- Viral infections
e. g.Kaposi sarcoma
86
what is necrotising ulcerative gingivitis associated with
stress, mixed bacterial infection, smoking and HIV
| very painful
87
what are the treatments for necrotising ulcerative gingivitis
```
plaque control (CHX, ultrasonics, OHI)
metronidazole
```
88
what are the steps of diagnosis of viral infection
1. clinical experience
2. lab tests immunoassay (ELISA) and sero-conversion (detect antibodies)
3. microbiology: viral culture
4. histology of tissues: biopsy (cellular morphology and immunohistochemistry)
89
how does antiviral health care prevent infection
cross infection control and vaccination
90
what are the treatments of antiviral health care
antiviral drugs, e.g. nucleoside analogues (inhibit replication as they are the building blocks of DNA): acyclovir and azidothymidine
91
outline the features of prokaryotes
no internal membranes (organelles)
single chromosomes, no histones
no fusion of gametes (can transfer DNA)
92
what lengths do bacteria vary from
0.2-700 micrometers
93
what are the major morphologies of bacterial cells?
```
coccus
rod
spirillum
spirochete
budding and appendaged bacteria
filamentous bacteria
```
94
what are the major structures of bacteria
cell wall
cell membrane
nucleoid/cytoplasm
95
what are the surface appendages of bacteria
flagella, fimbriae, capsule
96
what are the specialised structures of bacteria?
spores
| inclusion of bodies
97
how is light microscopy carried out
1. prepare smear- spread culture in thin film over slide and then dry in air
2. heat fixing and staining- pass slide through flame and flood slide with stains, rinse and dry
3. microscopy- place drop oil on slide and examine with 100x lens
98
what is the procedure and results for staining
1. flood the heat-fixed smear with crystal violet for 1 min- all cells purple
2. add iodine solution for 1 min- all still purple
3. decolorize with alcohol briefly -about 20 sec- gram+ purple, gram- colourless
4. counterstain with safranin for 1-2 min - G+ purple, G- pink to red
99
what does the gram stain distinguish between
major groups of pathogenic organisms
100
whats is the gram stain bases on
presence of outer membrane in gram negatives
101
*what are some other methods for the gram stain
gram - rods
| gram + cocci
102
what is the difference in the gram stain between + and -
+ thick cell wall, largely peptidoglycan (retains crystal violet-iodide complex)
- thinner peptidoglycan layer
(permeable to crystal violet-iodide)
103
what are acid fast bacterial cell walls
carbol fuchsin
drive with heat
destain with 3% acid-alcohol
high lipid/wax content (mycolic acids)
104
what is an example of acid fast bacterial cell walls
myobacterium tuberculosis (bacterial cell which causes TB)
105
what is an example of gram positive cocci
streptococcus pyogenes
106
what % of humans carry streptococcus pyogenes in their respiratory tract
5-15%
107
what acute infections are as a result of streptococcus pyogenes
Pharyngitis (severe sore throat) spread by droplets of saliva or nasal secretions
108
what are the clinical manifestations of Streptococcus pyogenes
– 2 to 4 day incubation
| – pharyngitis, tonsillitis, malaise, fever, headache, and redness, and lymph node enlargement in throat
109
what is an example of gram negative rods
salmonella enterica
110
what is the most common foodborne infection
salmonellosis
111
what is the most common form of salmonella enterica, and how long does it like
entercocolitis
| lasts 2-5 days
112
what types of salmonella are extremely dangerous
S. enterica serovar Typhi - typhoid fever
113
what are the characteristics of cytoplasmic membrane
phospholipid bilayer
thin barrier
6-8nm
114
what are the characteristics of the outer membrane (gram negative)
asymmetrical- phospholipid/LPS
| exposed on cell surface
115
what are the characteristics of flagella
```
motility
rotates
long and thin
can be used for classifications as different patterns
helical
protein= flagellin subunits
```
116
what is the flagellum
a highly complex molecular motor. rotation is coupled to the flow of protons across the membrane
117
what is fimbriae
protein filamentous structure that enables cells to stick to surfaces. it aids biofilm production
118
what is pilli
similar to fimbriae but longer and there are fewer on the cell
119
why are pilli important
```
genetic exchange (conjugation)- important in antibiotic resistance
adhesion of pathogens to host cells
```
120
example of pilli
neissera spp, s pyogenes
121
importance of polysaccharide capsule and slime layers
aid adherence
evade phagocytosis
protect against desiccation
122
what can be capsules and slime layers be composed of
polysaccaride or protein
123
what are cell inclusions
carbon storage polymers (poly B-hydroxybutyric acid, glycogen)
polyphosphate and sulfur
124
what are endospores
highly differentiated cell- cell almost becomes dormant and resistant to the environment
125
examples of endospores
bacillus anthracis, clostridium
126
whats needed to grow bacteria
anabolic and catabolic
micro and macro nutrients
H, O2, C, N, P, S and selenium pivotal
70% water
127
energy source for heterotrophs
obtain C from organic chemicals
128
energy source for autotrophs
obtain C from CO2
129
how are autotrophs different from auxotrophs
auxotrophs cannot synthesise an essential organic compound
130
what is fermentation
(anaerobic): organic compound used as both electron donor and electron acceptor
131
what is Respiration
(aerobic or anaerobic): electron donor is oxidised with O2 or O2 substitute.
132
what is the first step in fermentation and respiration
glycolysis (glucose --> pyruvate)
133
how is ATP synthesised in fermentation
substrate level phosphorylation
134
how is ATP synthesised in respiration
driven by oxidative phosphorylation
135
what is a major product from h the metabolism of streptococcal bacteria
lactic acid
136
what is the F0F1 ATPase
Highly abundant protein complex in actively growing bacterial cell walls
137
how does F0F1 ATPase generate ATP
from flow of H+ into cells
138
why do oral bacteria (s.mutans) use ATPase to pump H+ out of cells
too much H+ reverses ATP generation so protons pumped out to reduce acidic environment
139
what is bacterial growth
Irreversible increase in biomass and also, usually, numbers
140
how can bacterial growth be measured
microscopy, colony forming units (CFU), absorbance, nitrogen, carbon, DNA, protein
141
what are the methods of measuring bacterial growth
1. microscopic counts
2. viable counts
3. turbidimetric methods
142
what is recombinant DNA technology
joining together of DNA molecules from two different species
143
what is e.coli
harmless resident of the human gut, lives on our skin
144
what is the ecoli K12 genome
4.6 million base pairs
encodes 4,400 proteins
40% genes are of unknown function
many genes encoding enzymes clustered into operons
average protein contains approx 300 amino acids
145
what are bacterial plasmids
• Small circular DNA present in varying copy number per cell
146
what genes do bacterial plasmids carry
genes that are useful but not essential to survival: e.g. genes which make bacteria resistant to antibiotics
147
what are plasmids released by
dead bacteria and absorbed by those still living thus genetic information is exchanged.
148
how does horizontal gene transfer occur in bacteria
Occurs naturally by transformation, transduction or conjugation
149
what is bacterial sex in dental plaque
Genes transferred between bacteria in dental plaque
| Occurs naturally by transformations, transduction or conjugation
150
which bacteria in dental plaque can increase its 'fitness' after bacterial sex
streptococcus mutans
151
how does dna exchange take place between bacteria
dna uptake (transformation)
phage-mediated dna transfer (transduction)
mating (pilus-mediated transfer; conjugation)
152
what is dna transformation
dna incorporated into the chromosome of the recipient by homologous recombination
153
what the goals of genetic manipulation
Biology: To understand the function of a gene
- Knock-¬‐out genes
- Change individual amino acids in protein
- Clone and over-¬‐express proteins
- Construct fusions with reporter genes
Biotechnology:
-- Production of recombinant proteins eg insulin
- Develop vaccines eg hepatitis B
- Generate improved probiotic bacteria
154
what is recombinant dna technology
1. creation of recombinant dna- new combinations of unrelated genes in test tube
2. cloning of recombinant dna
- amplifying the new dna by introducing it into living cells
3. using recombinant dna
- expressing a cloned gene to produce a protein
155
what is in a molecular biologists 'toolbox'
* Enzymes- to manipulate the DNA/RNA
* Vectors- act as a vehicle to carry recombinant DNA into host cells
* DNA/RNA- the raw material- usually purified from tissue/cell culture
* Cells- to amplify- and sometimes express the recombinant DNA
156
what enzymes are important in the creation of recombinant dna
• Restriction enzymes: Cut double-stranded DNA
• DNA Ligase: Sticks fragments of DNA together
• Taq polymerase: PCR
- POLYMERASE CHAIN REACTION, Creates multiple copies of DNA fragment
• Reverse transcriptase: Copies RNA into DNA
157
what are restriction enzymes
molecular scissors
Cleave DNA, usually at very specific sequences
Naturally produced by bacteria: self-defence mechanism
many recognise 4-8 base pair palindromic sequences
158
what are the general properties of vectors
contain unique restriction sites for insertion of new dna
introduced into host easily
Contain a gene to allow selection of cells which contain the plasmid – eg an antibiotic resistance gene
regulatory sequences
159
what is can be used as a vector
plasmid (most common)
bacteriophage- virus which infects bacteria
cosmids/phagemids- genetically engineered hybrids which replicate as a plasmid but can be packaged as a bacteriophage
160
how would we clone a human gene in e.coli
1. create recombinant dna
| 2. clone recombinant dna
161
how would we create recombinant dna to clone a human gene in e.coli
Obtain DNA of interest
| Insert DNA of interest into vector (eg.plasmid)
162
how would we clone recombinant dna to clone a human gene in ecoli
- Recombinant plasmid is inserted into E.coli host
By transformation
Formation of pores in bacterial membrane to aid uptake of DNA, using CaCl2 on ice + heat shock (42°C, 2 mins) or Electroporation
- E.coli containing plasmid are selectively cultured
Each bacterial colony arises from a single transformed E coli cell
Each cell in the colony contains copies of the SAME plasmid
The cells in the colony are therefore clones
- Recombinant plasmid is purified or expressed
Technique may be used in industry to produce large quantities of the recombinant DNA
163
what are the problems with hormones from animal sources
animal protein not identical to human- side effects
difficult to purify
possible contamination with pathogens
164
what was the first recombinant drug licensed for human use
insulin
165
how is recombinant human insulin produced
- The gene for proinsulin contains 2 introns
- mRNA is reverse-‐transcribed to produce pro-‐insulin cDNA (no introns)
- joined to plasmid to produce recombinant plasmid and then infected into ecoli
166
what is the SMaRT replacement therapy technology
Creation of a genetically altered strain of Streptococcus mutans, called SMaRT, which does not produce lactic acid.
167
what are planktonic cells
homogenous, single species, nutrient rich
168
what are surface associated cells
limited heterogeneity, single species, nutrient rich. cells at the edge of the colony have access to more nutrients
169
what are natural cells
structured community, heterogenous, mixed species, nutrient limited
170
what is a biofilm
a community of microbial ells encased within a matrix of polymers and associated with an interface
171
what is the difference between a biofilm and a broth culture
broth cultures have no matrix
172
where do most bacteria grow
in biofilms
173
what are the characteristics of planktonic culture
```
homogenous
single species
uniform growth rate in chemostat
even distribution of O2, nutrients and waste etc
cells well-separated
no ECM
```
174
what are the characteristics of a biofilm (heterogenous)
```
mixed species which all interact
mixed growth rate
uneven distribution of O2, nutrients, waste
cells are in close contact
ECM present
```
175
what is the difference between biofilms and pure broth cultures
- biofilm bacteria are up to 1000 times more resistant to antibiotics than planktonic cells
- biofilms difficult to remove
- bacterial communities more pathogenic
- bacteria in biofilms can hide from host immunity
- biofilms provide a range of niches
176
where are biofilms found
anywhere there is an:
| interface, moisture, nutrients/energy source
177
what is an example of biofilms at air liquid interfaces
surfaces of ponds, oceans
178
what is an example of biofilms at air solid interfaces
tree trunks, rocks, solids particles
179
what is an example of biofilms at solid-liquid interfaces
internal surfaces of pipes (including dental unit waterlines), river beds, indwelling catheters, teeth
180
what is the structure of a biofilm
3 dimensional, mushroom like structures, contain serum, open structures often with channels permeating the matrix, heterogenous (cell distribution is not uniform), nutrients and wase products non-uniformly distributed
181
outline the growth of a biofilm
- attachment- adhesion of few motile cells to suitable solid surface
- colonisation- intercellular communication, growth, and polysaccharide formation
- development- more growth and polysaccharide
- active dispersal- triggered by environmental factors such as nutrient availability
182
what is the structure of a biofilm- heterogenicity
range of different niches
anaerobic bacteria can grow in parts of the biofilm that contain lower levels of O2
may be pockets in biofilm with aerated areas and those with low nutrients
183
what are the structures in the biofilm matrix
macromolecules form the basic structure
small molecules trapped in the matrix
biofilm growth and resistance to antimicrobials
184
what are the diagnostic criteria for biofilm infections
- associated with a surface
- aggregated cells in cell clusters incased in matrix
- infection confined to particular site in host
- recalcitrance to antibiotic treatment
- catheter infections often recalcitrant to antibiotics
- used a model containing artificial urine flowing over discs of catheter infections
- inoculated with pseudomonas aeruginosa and tested antibiotic sensitivity of biofilm bacteria and compared them with planktonic cells.
185
what were the results of the diagnostic criteria test for biofilms
results: biofilms reduced with tobramycin
when you treat a planktonic culture with 50mg per mil you wipe it out and when you treat a biofilm with 1000 mg per mil you dont wipe it out
186
why are biofilm bacteria so resistant
```
slow growing
persister cells present
sequestration from immune system
elevated expression of efflux pumps
poor penetration of antibiotics
```
187
what does antibiotic sensitivity vary with
stage of growth
188
are antibiotics that target cell wall more effective against biofilms or planktonic cells
planktonic cells
189
are antibiotics that target protein or RNA synthesis more effective against biofilms or planktonic cells
equally effective against both
190
what are persister cells
inactive/dormant cells
191
why are persister cells dangerous
they are very difficult to kill with antibiotics
192
what happens when dormant cells become a significant proportion of biofilm bacteria
can become responsible for the reactivation of infections
193
what are SCVs and what are they responsible for
small colony variants- formed on agar by persister cells
| may be responsible for infections on replacement hips 15-20 years after the operation
194
what is the difference between slow growth and persisters
in persister cells there are one or two highly resistant persister cells in the population whereas in slow growth there are many cells
195
why are biofilms highly resistant to phagocytosis
inflammatory cells cannot penetrate the matrix and if they do, they do not ingest the bacteria effectively
196
what adaptive responses in biofilms or communities can protect bacteria
- pseudomonas spp. produce rhamnolipids in biofilms, which can kill neutrophils.
- interactions with oral streptococci induce production of a complement-evading protein by aggregatibacter actinomycetemcomitans
- up regulation of efflux pumps
197
what do efflux pumps do
kick out drugs e.g. cancer cells kicking out drugs
198
in which biofilms are antibiotic efflux pumps up-regulated
biofilms formed by E.coli, P.aeruginosa, Candida albicans
199
are efflux pumps effective against all antibiotics
no
| up-regulation of efflux pumps is not universal
200
why are biofilm bacteria so resistant
```
slow growing
persister cells
sequestration from the immune system
more efflux pumps
poor penetration of antibiotics
```
201
how is mass transfer measured in biofilms
using fluorescently labelled latex beads, dyes or labelled proteins
202
what is the consequence of diffusion being impeded by the biofilm matrix
not v significant for small molecules
even large molecules are weakly limited
molecules are carried through water channels
203
how are electrostatic and vdw interactions important in biofilms
the matrix acts as an ion channel resin e.g. metal cations become trapped in the anionic matrix
204
is tobramycin impeded by the matrix in biofilms
yes as its positively charged
205
is ciprofloxacin impeded by the matrix in biofilms
no as its neutral
206
when does reduced penetration of the biofilms occur
in non-mucoid strains (don't produce the carbohydrate alginate)
207
how can protection against tobramycin be achieved
adding extracellular DNA to biofilms enhances protection
208
what are the bacterial cell-cell interactions in oral biofilms
signalling, synergism, antagonism
209
what is the human microbiome
microorganisms within the human body- colonised by 10 trillion bacteria (10% human)
210
what was the first microbial gene sequenced
haemophilus influenza
211
what is the ORF
the part of the reading frame which has the potential to be translated
212
how is a list of ORFs obtained from the reading frame
1. computer finds possible start codons then stop codons
2. computer counts codons between start and stop
3. computer finds possible RBS
4. computer calculates codon bias in ORF
5. computer decides if ORF is likely to be genuine
6. List of probable ORFs
213
what are the 2 components of the bacterial genome
core genome
| pan genome
214
what is the core genome
shared by all strains of the species
215
what is the pan genome
all optional extras in some but not all strains of the species
216
what are chromosomal islands
region of bacterial chromosome of foreign origin that contains clustered genes for some extra property such as virulence or symbiosis (thought to be acquired by horizontal gene transfer)
217
what are pathogenicity islands
chromosomal islands containing genes for virulence (disease) acquired by horizontal gene transfer
218
what can transposons do
jump to other places in the chromosome
219
how is the human microbiome important in health
its essential for healthy immune system
predictions can be made from analysis of microbial community types
oral community types correlate with gut communities
220
which community type is least stable
oral is less stable than microbial community types in the gut or vagina
221
how is the human microbiome important in disease
analysis of microbiome is used to identify biomarkers for disease
many diseases result from shifts in microbial community types rather than infection by a single agent
222
what do probiotics do
introduce a microbe into the human body for its beneficial qualities
223
what is the difference between chromosomal islands and the rest of the genome
- chromosomal islands believed to have a foreign origin
| - base composition and codon usage varies in chromosomal islands from the rest of the genome
224
what is a microbiome
the total compliment of microorganisms in an environment
225
how can microbial communities be analysed
metagenomic sequences- obtains information about all DNA present
226
how can bacterial pathogens be identified
sequencing of 16S rRNA gene
227
what is the 16S rRNA gene
component of the 30S small sub unit of the bacterial ribosome -provides indication of the bacterial species present
228
why doesn't the sequence analysis of the 16S rRNA gene include viruses or fungi
viruses lack rRNA genes
| fungi have the 18s rRNA gene
229
what is the diversity of human microbiome
- concordant among measures
- unique to everyone
- determined by microbial habitat
- diversity between individuals at a single body site is generally lower than diversity within individuals at different body sites
230
what is the downside of antibiotics
they kill good bacteria as well as those that cause disease
231
what influences the core human microbiome
```
host lifestyle
host genotype
host physiology
host immune system
transient community members
host environment
host pathobiology
```
232
how is the human microbiome important in health
its essential for a healthy immune system
233
what are the diseases of different systems related to human microbial symbiosis
```
obesity
IBD
liver disease
emerging infectious disease
diabetes mellitus
atherosclerosis
metabolic syndrome
```
234
what is the relationship between mouthwash and blood pressure
it can raise blood pressure due to oral microbiota converting nitrate to nitrite
235
what is disease aetiology
the study of causation
236
what is the germ theory of disease
the idea that disease is caused by 'invisible' creatures dates back to roman times
237
what bacteria is anthrax caused by
gram-positive, rod-shaped bacteria known as Bacillus anthracis
238
what are the characteristics of Bacillus anthracis
spore forming
highly resistant
resilient
239
what was Koch's proof that B.anthracis causes infections
- spleen tissue from infected farm animal was given to the mouse and the mouse died
- proliferated organisms in mouse after the infected tissue had been cultured in a growth medium in the eye
240
what are Koch-Henle Postulates
- the organism must be present in all cases of disease and not present in healthy individuals
- the organisms must be isolated in pure in culture
- the isolated organisms must cause disease in a suitable animal
- the organism must be reisolated from infected animal (shows proliferation)
241
where are legionella bacteria found
naturally in the environment, usually in water
242
where do legionella bacteria grow best
in warm water e.g.
| hot tubs, cooling towers, hot water tanks, large plumbing systems, air conditioning systems
243
how are legionella able to survive in harsh environments
they survive within amoobae
244
what harbours legionella pneumophila
Protist (Tethymena)
245
when is it not possible to fulfil Koch's postulate of: The organism must be present in all cases of disease, and not present in healthy individuals
there may be asymptomatic carriers (cholera, Neisseria meningitidis)
246
when is it not possible to fulfil Koch's postulate of: The organism must be isolated in pure culture
- viruses could not be cultured in 1900s. Prions are still difficult/impossible to culture
- Some infections are polymicrobial (no single aetiological agent)
247
when is it not possible to fulfil Koch's postulate of: The isolated organism must cause disease in a suitable animal
animal models may not be available/may not be representative of human disease
248
when can Koch's postulates not be applied precisely
- If no infecting organism can be detected
- If the bacteria cannot be grown in culture
- If no suitable animal model is available
- If more than one species of bacterium is involved
- If it is the level rather than just the presence of the infecting bacterium that is important
249
how do organisms cause disease
```
Transmission (the 6 ‘I’s)
- Implantation
- In utero
- Ingestion
- Inhalation
- Injection
- Inunction
Pathogenesis
```
250
how is dna analysis useful in the molecular biology era to detect infectious agents
circumvents the isolation of an infectious agent
very sensitive (detects agetns in the absence of disease)
may be quantitative-i.e. can determine load of infectious agents
251
what are many infectious diseases now regarded as
imbalances in the microbiota
252
what is meant by commensal
an organism that lives in harmless association with its host ('normal flora')
253
what is meant by carriage (carrier state)
colonisation with a pathogen i.e. this host can act as a source of infection for others, but shows no symptoms themselves
254
what is an opportunistic pathogen
an organisms which requires the host to have a pre-existing defect in its defences before it can cause diseases- exists in the body but doesnt cause disease
255
what is virulence
a measure of the capacity of an organism to cause disease
256
what are virulence factors
properties of a bacterium which contribute to its virulence i.e. toxins created by bacteria
257
what are endogenous infections
caused by an infectious agent that is already present in the body, but has previously been inapparent or dormant
258
what are exogenous infections
acquired from sources outside the patient (i.e. bacteria which causes tetanus infection).
259
what are some virulence factors that pathogens possess
adhesins- stick to the body
invasins- can invade cells
toxins- harm cell types an cause disease
extracellular enzymes
260
```
what are the pathogens in
1- whooping cough
2-gonorrhoea
3-black death
4-anthrax
```
1- bordetella pertussis
2-neisseria gonorrhoeae
3-yersinia pestis
4- bacillus anthracis
261
what are opportunist pathogens in
1- catheters/endocarditis
2- burns
3-diarrhoea
1-staphylococcus epidermis
2-pseudomonas aeruginosa
3-escherichia coli
262
what are the stages of infection
```
1- adherence- to skin/mucosa
2- invasion- through epithelium
3- infection/growth-virulence factors and toxins
4- invasiveness toxicity
5- damage to host
```
effect: tissue damage, disease
263
what interactions occur between surface structures of bacteria and the host tissue
pilli, fimbrae and surface proteins of bacteria bind to host tissue receptors including to glycoproteins and tissue-specific antigens
flagella is an important virulence factor
264
what are examples of bacteria adhering to the host
E.coli with type I pili adgere to epithelial cells in the uniary tract
vibrio cholerae is highly motile, swims through intestinal mucus to attach to receptors in gut wall
265
what are some innate host defences
- Skin (keratinised, shedding of outer layers)
- Flushing (saliva, urine, peristalsis in gut)
- Secretions (lysozyme, stomach acid, bile salts)
- Mucous membranes (cilia)- antibacterial peptides
- Normal bacterial population (colonisation resistance
266
what are some adaptive host defences
- Phagocytosis
- Inflammation, fever
- Antibodies
267
why may you be more vulnerable to infection if you've taken omeporazol
it increases stomach acid so less bacteria needed to cause disease.
268
how have pathogens overcome the skin as a host defence
keratinases, skin penetration
269
how have pathogens overcome flushing
adherence, motility
270
how have pathogens overcome secretions
lysozyme resistant walls, tolerance to salts, bile, pH
271
how have pathogens overcome mucous membranes
degrade or swim through mucin
272
how have pathogens overcome commensal flora
metabolic end products, bacteriocins, bacteriophage
273
how have pathogens overcome phagocytosis
capsules
274
how have pathogens overcome inflammation/fever
heat shock response
275
how have pathogens overcome antibodies
breakdown of immunoglobulins kill host defence cells, intracellular growth, antigenic variation
276
what is bacteraemia
invasion of bacteria into the blood stream
277
what is septicaemia
infection in the blood stream
| -release of large number of cytokines which kills the host in the end when its meant to kill the pathogen
278
what is an infection
a situation when a microorganism that is not a natural coloniser grows within the host, whether or not it causes damage
279
when does a disease arise
when there is damage to the host
280
how do bacteria spread
- Through lungs, gut, urinary tract, blood via sinuses and body cavities
- Tracking along nerve pathways
- Direct cell‐to-¬cell
- Tissue breakdown
281
what are the microbial virulence factors
Toxins
1. endotoxins- lipopolysaccharides cause septic shock
2. exotoxins - damage the host cells
Tissue degrading enzymes
collagenase/haemolysin/hyaluronidase
282
what are endotoxins, where are they found, when are they released and what do they cause
- Lipopolysaccharide (LPS) is an endotoxin
- Found only in Gram-negative bacteria
- Present in the outer layer of the outer membrane
- Released during growth or when bacteria die and lyse
- Causes fever, septic shock and local inflammation
283
what are exotoxins and what are they produced by
- Secreted by bacteria
- Produced by some Gram positive and Gram-negative species
- Relatively unstable (often heat‐sensitive)
- Highly toxic - 1 nanogram (10^-9g) of botulism toxin can be fatal
284
why are antibiotics useless to treat e.g. food poisoning by enterotoxin. what should be used to treat it
bc damage by exotoxin does not always require bacterial infection. vaccines are needed to target the toxin rather than the infectious agent
285
what do tissue degrading enzymes do and what are some examples
Breaks down junctions between skin cells and destroys skin cells
S.pyogenes, hyaluronidase
286
what bacteria causes tetanus and what are its characteristics
Clostridium tetani
Gram positive, spore forming rod
Secretes a powerful neurotoxin that blocks the release of neurotransmitters from the pre-synaptic membranes of inhibitory nerve synapses
Found in soil and animal faeces
287
what are the symptoms of tetanus
- ‘lockjaw’, muscular contraction
| - can Bend over backwards- snap spine and die
288
what bacteria causes botulism and what are its characteristics
- Clostridium botulinum
- Found in soil, water, foodstuffs (including honey)
- Extremely potent toxin (like tetanus) inactivated by cooking.
289
what is the botulism toxin used for
botox
Treat muscle conditions
Cosmetics- remove wrinkles, temporary relax muscles
Poison-tip umbrella assassination
290
what bacteria causes diphtheria and what are its characteristics
Corynebacterium diphtheriae
- Makes toxin that stops protein synthesis in cells
- Toxin gene carried by bacteriophage
- Inhibits protein synthesis
- Causes pseudo membranes to be formed in the posterior pharynx
291
how to bacteria cause damage to the host (step 5)
Tissue breakdown (Necrotising Fasciitis)
overactive immune response
most common mono infections are Streptococcus pyogenes and MRSA
292
what are factors that influence susceptibility to the disease
- trauma
- underlying disease or other infections
- Age
- Genetic constitution (tissue type etc)
- Nutrition
- Hormonal factors and stress
- Pollutants
293
what does targeted microbiome analysis involve
sequencing the 16S rRNA gene for bacteria
tells you what OTU (generally what species it is, doesn’t exactly correlate)
294
what does metagenomics involve
sequencing all DNA present in a sample, then using bioinformatics to predict the source/function of the DNA sequences
295
what is the SPH
diseases are caused by a single microorganism.
296
what is the N-SPH
diseases are caused by the amount of plaque
297
what is the ecological plaque hypothesis
disease is caused by interactions of specific microorganisms with the microbial community
298
what are keystone species
they disproportionately large effects on their communities given their abundance
299
what does a keystone pathogen do
supports and stabilises the dysbiotic state, and triggers an immune reaction even when present at relatively low numbers
300
what is periodontitis initiate by
a synergistic and dysbiotic microbiota, within which different members, or specific gene combinations thereof , fulfil distinct roles that converge to shape and stabilise a disease-provoking microbiota
301
what is the gut microbiome
of the most diverse microbial communities in the body and is the totality of microorganisms, bacteria, viruses, protozoa, and fungi, and their collective genetic material present in the gastrointestinal tract (GIT).
302
what are differences in the microbiota between individuals associated with
obesity, colorectal cancer, inflammatory bowel disease & many systemic conditions
303
what is IBS caused by
Host genetics play a key role
| Gut microbiome is gaining increasing attention.
304
which is clinical microbiology
In vitro diagnosis and management of diseases caused by bacteria, viruses, fungi and parasites
305
what is the role of the clinical microbiology lab
accurate information about the presence or absence of microorganisms in a patient sample
aid in the diagnosis and management of diseases caused by pathogenic microorganisms
provide information regarding the antimicrobial susceptibility of the microorganisms detected
Do all this is a clinically relevant timeframe
306
What makes a good diagnostic test in clinical microbiology?
Specificity (NO FALSE POSITIVES!)
Sensitivity
Time taken to produce a result
Ease of use
307
what are there different Types of lab tests
Identification of a specific microbial product
Detection of specific antibodies to a pathogen
Detection of genes (DNA or RNA) specific to a pathogen in a patient sample
308
what are the Types of lab tests used
Direct microscopy
| isolation and culture
309
what are the drawbacks of direct microscopy
Lacks sensitivity & specificity for many organisms
You needs lots of the pathogen
Usually requires culture confirmation
310
when would you used direct microscopy
e.g. Detection of tuberculosis
311
what is required for the identification of bacteria
Microscopy: Gram stain and morphology
Growth requirements
Biochemical Identification tests
Antibody-based tests
MALDI-TOF mass spectrometry
PCR/Nucleic acid-based tests (gene specific)
312
what does the oxidase test test for
Oxidase Positive bacteria contain cytochrome C oxidase and can therefore use oxygen for energy production with an electron transfer chain
313
what is good about Biochemical Identification tests
metabolise sugars oxidatively or fermentatively
produce enzymes which can be easily detected
use a range of substrates for growth.
314
what are limitations of culture-based methods
Some organisms grow very slowly in vitro
Some organisms are difficult to grow in vitro
Some organisms don’t grow in vitro
Organisms don’t always grow if the patient has already been treated with antibiotics
315
how can antigens be detected
Detection of soluble carbohydrate antigens by antibody-coated latex
316
what can examination of blood serum detect for
* Measles
* Mumps
* Mycoplasma
* Parvovirus
* Rubella
* Syphilis
* Toxoplasma
* Varicella-zoster
317
when is Nucleic acid-based tests (NATS) used
Mainly used in virology
Some application in bacteriology
Very sensitive and specific
Mainly polymerase chain reaction (PCR
318
what are advantages of PCR
Rapid testing:
Multiplex detection
Increased sensitivity
Specific detection
319
how is Bordetella pertussis (whooping cough) detected
PCR
More rapid and sensitive than culture
320
what is Legionnaires disease
Nosocomial or community acquired pulmonary infection
Often causes outbreaks of infection
Transmitted through the inhalation of Legionella from a contaminated environmental source
321
how is Legionellosis detected
culture of the organism from clinical samples
Rapid diagnosis by PCR (3-5 hrs) improves the prognosis
322
how is bacterial meningitis detected (N. meningitidis and Streptococcus pneumoniae)
Culture doesn’t work
PCR testing used- Sample types CSF, blood
323
what is syndromic testing
testing for multiple pathogens at the same time
324
what are PCR arrays
Up to +40 targets per sample
Gastrointestinal panel (viral/bacterial/parasitic
Respiratory panel (viral/bacterial)
Viral haemorrhagic fever panel (inc. malaria)
325
what does dysbiosis show
shows association, not necessarily disease causing organism
326
what is one condition that differences in the microbiota can be associated with
irritable bowel disease
| crowns disease and ulcerative colitis
327
how does a healthy microbiome interplay with disease
protects agains pathogens
train/stimulates immune function
supplies nutrients, energy, vitamins
328
how does a diseased microbiome affect the individual
inflammation, oxidative stress, increase in gram negative bacteria, infection, altered metabolite production
329
what are the treatments used to target the microbiome
Careful selection of antibiotics to minimise unwanted shifts in the microbiota
e.g. avoid repeated use of a single antibiotic.
repopulation of the gut with healthy bacteria
(prebiotics and probiotics)
330
what are prebiotics
feed the good organisms in gut/mouth (encourage good bacteria to restore balance)
331
what are probiotics
deliver the microorganisms into the gut/mouth (e.g. yoghurt)
332
what is Faecal microbiota transplantation
separate microorganisms from faeces and give back to someone with the disease (encouraging results for relapsing C. diff. infection but not so clear for IBD).
333
what are the stages of ecological plaque hypothesis and the aetiology of dental caries
excess sugar induces acid production (stress)
shift from neutral pH to low pH (environmental change)
s.sanguinis and s.gordinii to s.mutains, lactobacillus spp, and scardovia wiegsiae (ecological shift)
shift from remineralisation to demineralisation (disease)
