Lecture 6 - bacterial persistence and surviving an environment in flux

Question 1

Environments that bacteria can grow in

Answer 1

Bacteria can pretty much grow in any environment (soil, hot, dry, cold, ice, within and on us)

Question 2

oxygen is one of the most important ______ factors in both medical and environmental microbiology

Answer 2

Abiotic

Abiotic factors refer to non-living physical and chemical elements in the ecosystem.

Question 3

Aerobic environment

Answer 3

Aerobic = high oxygen (atmospheric, 21%)

Aerobes prefer oxic environments

Question 4

Microaerobic environment

Answer 4

Microaerobic = trace oxygen (2-10% or less)

Microaerophiles prefer hypoxic environments

Question 5

Anaerobic environment

Answer 5

Anaerobic = no oxygen (strictly 0%)

Anaerobes prefer anoxic environments
Facultative = change metabolism under oxia (they can adapt)

Question 6

Host associated environments

Answer 6

internal or bacteria dense environments are anaerobic

Question 7

Soils and oceans as environments

Answer 7

decreasing oxygen with increasing depth (oxygen can’t diffuse as far and because of bacterial activity)

Question 8

Eutrophication

Answer 8

High local activity of aerobes can lead to oxygen minimum zones, eutrophication (Eutrophication is when the environment becomes enriched with nutrients. The microbes may use up all the oxygen in the water, leaving none for other marine life. This results in the death of many aquatic organisms such as fish, which need the oxygen in the water to live.)

Question 9

Flexibility in terms of microbes means

Answer 9

the ability to adapt to a wide range of environments

Question 10

What environment does most microbes inhabit?

Answer 10

Relatively few microbes are lucky enough to live in nutrient-rich eutrophic environments. Rather, most inhabit oligotrophic environments - ones in which nutrient levels are low and some live in feast-and-famine environments.

Question 11

Higher eukaryotes in terms of flexibility

Answer 11

Higher eukaryotes (which are hosts for example) = inflexible 
Obligately heterotrophic (e.g. sugars, amino acids, fats) 
Obligately aerobic (oxygen for respiration) 
Ferments transiently and products must be recycled

Question 12

Bacteria in terms of flexibility

Answer 12

Bacteria = flexible
Diverse organic and inorganic electron donors
Oxygen and anaerobic electron acceptors (facultative anaerobe)
Fermentation as terminal, sustained mode of energy generation (even when there is limited processes for respiration to occur)

Question 13

E. coli in terms of flexibility

Answer 13

Numerous substrates for energy generation

Can use greater than 6 alternatives to oxygen for respiration

Can live on trace gases (H2) if no nutrients are available

Flexiblity allows ATP generation when environment changes

Question 14

Bacteria as recyclers

Answer 14

Bacteria are the ultimate recyclers which is part of their flexibility, there is a vast amount of carbon and energy sources that they harness - plant tissue, animal tissue, excretory products - end products, cells of dead bacteria, antibiotics, plastics, heavy metals

Question 15

Bacteria have diverse _____ _____ for breaking down several polymers into monomers

Answer 15

digestive enzymes

Question 16

Bacteria digestive enzymes are typically

Answer 16

Typically bound-in or associated with cell surface to prevent losses to competitors

Releases monomers or shorter oligomers that conserved pathways can use such as glucose, amino acids, fatty acids

Question 17

Polymers and monomers

Answer 17

Polymers are molecules made of monomers such as proteins (monomer =amino acid), DNA (monomer = nucleotides, fats (monomers = glycerol)

Question 18

What happens if there is not food for bacteria?

Answer 18

Bacteria are specialists at growing slowly or not at all - persistence

Still metabolically active

Different to true dormancy i.e. being an endospore (endospore is not only resistant to a variety of stressors but are also metabolically dormant)

Most environments are too nutrient limited to reflect batch culture

Persistence most likely normal state of bacteria in environment

In reality in the environment, bacteria can enter a long term stationary phase which is also known as long term resistance where the bacteria that have survived the death phase have now adapted to growing slowly/if not at all and in this case there is very little cell death but what we can have is spontaneous mutations occur that can create lots of different kinds of strains

Question 19

R type strategist examples

Answer 19

e.g. E.coli, B.subtilis

Question 20

R type strategist

Answer 20

Fast growing (TD= minutes)

May use food wastefully

Last ditch strategies during stress: sporulation, “bust and boom”

Dynamic persistence of small subpopulation

Overproduce and those that survive are those that are likely to have entered the persistent state

Question 21

R type strategist TD

Answer 21

Fast growing (TD= minutes)

Question 22

K type strategist examples

Answer 22

e.g. M.tuberculosis, soil bacteria

Question 23

K type strategist

Answer 23

e.g. M.tuberculosis, soil bacteria

Slow growing (TD= days)

Uses food efficiently therefore there is less waste

Survives stress for months-years with less drastic metabolic changes

Bulk of population persists

Question 24

K type strategist TD

Answer 24

Slow growing (TD= days)

Question 25

Gram negative vs gram positive

Answer 25

Gram negative cell wall has two plasma membranes separated by a periplasmic space, gram positive has one inner membrane and peptidoglycan layer that is thick

Question 26

current antibiotics

Answer 26

are only effective when a bacterium is growing (log phase)

e.g. cell wall synthesis inhibitors and protein synthesis inhibitors

Question 27

Starvation responses induce ….

Answer 27

protective responses to some antibiotics

Question 28

Some antibiotics need…

Answer 28

Some antibiotics need energy to be taken up by bacteria, stationary vs log
- Adding nutrients can recover activity

However since 1940s its known that some log phase bacteria survive in clinic

Question 29

Persisters

Answer 29

Persisters are bacterial cells identified by their ability to survive exposure to an antibiotic (often multiple antibiotics) even though they do not harbour antibiotic resistance genes. They are non-growing cells that are though to be dormant. Dormancy is central to the hypotheses regatding their antibiotic-tolerant phenotype; in dormant cells, the target of an antibiotic is unavailable to inactive and would be unaffected by the antibiotic. Persisters are of considerable interest for several reasons, including their potential to cause recurrent infections after major antibiotic treatment

Question 30

Antibiotic resistance

Answer 30

Antibiotic resistance = heritable genetic changes allow bacteria to grow

Mutate their genes or acquire enzymes but there are changes in DNA therefore are heritable

Question 31

Antibiotic persistance

Answer 31

Antibiotic persistance = non-heritable physiological changes allow survival of a few slow or non-growing cells

Something happens during the course of treatment, most of them die off but there is a dynamic subpopulation of cells that grow very slowly can then regrow back to the beginning point when conditions become favourable again

Question 32

What keeps the resistance genes in a population and contributes to the drug-resistant bacteria strains is …

Answer 32

exposure to the drug

Question 33

What makes treatment time longer in terms of antibiotics?

Answer 33

inability of antibiotics to kill non-growing cells makes treatment time longer

Question 34

Persistence can lead to

Answer 34

resistance

Question 35

Wiping out populations with antibiotics …

Answer 35

Unlike environmental persistence, although we have the R and K type selection in the environment and population level strategies, when we just wipe out populations with antibiotics it is environmental catastrophe, their responses tend to look a lot more like an R type selection response regardless of the organism

Question 36

Environmental persistence features

Answer 36

Response to fluctuating food and nutrient supplies

Different population-level strategies (r/K)

Adaptation for long-term survival

More time to respond

Question 37

Antibiotic persistance features

Answer 37

Response to an environmental catastrophe (antibiotic killing)

Similar population-level strategies

Subpopulation of cells stochastically (randomly) preadapted

Less time to response

Question 38

responses to stress (3)

Answer 38

stringent response, SOS response, Toxin-antitoxin systems

Question 39

methods of persistance (5)

Answer 39

mutations, stationary phase, antibiotic exposure, vacuoles and biofilms

Question 40

responses to stress purpose

Answer 40

Evidence suggests that in some cases, small sub­ sets of cells in a population of growing cells spontaneously be­ come dormant persister cells, even when nutrients are readily available. These persisters can be thought of as the population’s attempt to prepare for future possible starvation conditions. There is also increasing evidence that persisters arise in response to various triggers, with starvation probably being the most im­ portant.

Question 41

Stringent response

Answer 41

shift in the expression the genes to a survival based response

Mediated by molecule (p)ppGpp

Stops expensive biosynthetic processes

ppGpp is a signaling molecule that functions in a regulatory network called the stringent response. ppGpp is often referred to as an alarmone because it is synthesized in response to many stressors and regulates the activity of numerous genes and proteins so that the cell can survive.

Question 42

SOS response

Answer 42

stress induces DNA repair, also has survival features

Both repair and survival responses

Mediated by protein RecA

Question 43

Toxin-antitoxin systems

Answer 43

Balancing act with diverse cellular functions

Disruption of antitoxin leads to growth inhibition by toxin

The toxin when functioning disrupts normal cellular functions, causing growth arrest and the persister phenotype

inter­nal, nonsecreted toxin and a cognate antitoxin that prevents the toxin from exerting its effects on the cell. The toxin, when func­tioning, disrupts normal cellular functions, causing growth arrest and the persister phenotype.

Question 44

Hypoxia in terms of persistance and example

Answer 44

gradual depletion of oxygen leads to a stationary phase, slowed growth and persistent phenotypes

M.tuberculosis frequently encounters hypoxia in host and intracellular environments
Macrophage phaglysosome
Granuloma; immune structures forming wall

Question 45

Biofilms

Answer 45

Slimy matrix of various bacteria produced polymers

Local depletion of nutrients/oxygen leads to persister cell formation

Physical protection from environment combined with antibiotic persistence

A mature biofilm is a complex, dynamic community of microorganisms. It exhibits considerable heterogeneity due to differences in the metabolic activity of microbes at various locations within the biofilm; some are persister cells

While in the biofilm, microbes are protected from numerous harmful agents such s UV light and antibiotics

Question 46

All forms of persistance cause a shift to form of

Answer 46

energy metabolism that permits survival without growth