Lecture 27: Biofilms and Chronic Bacterial Infections

Biofilms are _____ of bacteria attached to a _____ and or _____ and are encased in a self produced _____

aggregates; surface; each other; matrix

a wide variety of _____ are caused by bacterial biofilms

infections/diseases

what are two main bacterial infections that we talk about in this class?

bacterial endocarditis and cystic fibrosis

Bacterial endocarditis can be caused by a range of bacterial _____, but they are all caused by gram _____ bacteria

species; positive

what are two common bacteria that cause bacterial endocarditis?

streptococcus and staphylococcus

colonization of bacterial causing bacterial endocarditis occurs at an _____

injured heart valve

where does the bacterial come from that colonizes the heart valve?

low levels in blood

rabbit infection model studies have show that high levels of _____ can be delivered to _____, but will fail to _____ the biofilm bacteria

antibiotics; vegetations; kill

True or False: antibiotics can’t kill biofilm bacteria because it can’t get to the bacteria

FALSE: it can get there, just can’t kill them

why are biofilms untreatable?

can’t be cleared by immune system or antibiotics

the primary infection lesion in endocarditis is a complex _____ composed of both _____ and _____ components located on a cardiac valve

biofilm; bacterial; host

what are the three reasons why biofilms are traditionally more resistant to antimicrobial agents than planktonic cells?

EPS matrix = diffusion barrier, slow growing subpopulation, biofilm phenotype

what are planktonic cells?

free living cell (unattached from biofilm)

antibiotics are not as effective against _____ bacteria

slow growing

what is a “biofilm phenotype”

traits to support biofilm growth and maintenance (special genes are expressed)

how does the “biofilm phenotype” increase antimicrobial resistance?

genes expressed for biofilms may be helpful for inactivating/removing antibiotics (ex. efflux pumps)

the biofilm state is _____ resistant to antibiotics

inherently

when testing for slow growing populations; a colony was spotted on top of a _____ on an agar plate. Another _____ was place _____ the colony followed by an _____

filter; filter; above; absorption disk

after growing the colony on a filter on a normal agar plate, the colony was moved to a _____ plate

antibiotic

after the colony was moved to an antibiotic plate, researchers measured the levels of _____ that moved through the colony

antibiotics

what was the absorption disk for?

to absorb the antibiotics at the top → proves that antibiotics were able to fully penetrate the colony!

researchers also measured the concentration of _____ at various _____ throughout the colony

oxygen; depths

research found that all you reach greater depths, the concentration of oxygen _____

decreases

the anaerobic portions of the biofilm metabolically _____

inactive

research found that all you reach greater depths, the antibiotics were increasingly _____ effective

LESS

Why can’t chronic CF airway infections be eliminated by the immune system or antibiotics?

it is a biofilm

existing evidence support the involvement of _____ in chronic CF airway infections.

biofilm infection

biolfikms are a surface-attached _____

microbial community

_____ is the primary cause of morbidity and mortality in CF

Pseudomonas aeruginosa

colonization of the host occurs _____ in lfie

early

CF is an _____ disease

autosomal recessive

in order for an individual to have CF, they must have _____ copy/ies of the CF gene

TWO

if an individual has one normal gene and one CF gene, will they have CF?

NO → carrier (its is recessive!)

does the autosomal disease occur more in females or males?

neither → equal!

what are the chances that child will develop CF if each parent is a carrier?

25%

what is the gene that causes CF called?

CFTR

CFTR encodes for an _____ found in _____ cells

ion transporter; epithelial

CFTR is a _____ transporter found in the _____ of _____ cells

chloride; membrane; epithelial

in normal cells, the CTFR protein allows the _____ of _____ from the cell

RELEASE; Cl

those with CF will have a _____ version of CFTR which causes a _____

nonfunctional; buildup of ions

True or False: Cystic fibrosis only causes symptoms in the lungs and airways

FALSE: All epithelial cells are effective → problems all over the body

what two issues are associated with the airways in CF patients?

infection and inflammation

what causes the MOST problems for patients with CF? Why?

airway inflammation → big immune response causes collateral damage to body

CF patients have a large _____ against the _____ in their airways. This causes damage to the _____ in the process, eventually leading to a gradual deterioration of _____

immune response; bacteria; body; lung function

what are 5 other locations where CF patients often experience adverse symptoms?

liver, pancreas, intestine, reproductive tract, sweat glands

CF lung disease is a _____ defect

local host defense

_____ is a hallmark of CF lung disease

bacterial infection of the airways

other organs as well as _____ non-CF lungs are _____ infected with bacteria causing CF lung disease

transplanted; NOT

_____ of the airway infection is the most effective therapy for CF lung disease

antibiotic treatment

antibiotic treatment, while it will NOT _____ the bacteria, it WILL _____

eliminate; reduce its numbers

airway disease in cystic fibrosis is a _____ bacterial infection with many organisms (particularly _____ and _____) as well as lots of _____

chronic; P. auroginosa, S. aureus; inflammation

the lungs of patients with airway disease in CF are flooded with _____ indicating _____

neutrophils; inflammation

newborn lungs are _____

sterile

early in life, the CF patients acquires a bacterial strain from the _____. Over time the strain will _____ to the body and increase its _____

environment; adapt; pathogenesis

after newborns are colonized, there is a _____ that causes _____ infection

host defense defect; intermittent

After intermittent infection _____ occurs which eventually leads to _____ colonization

bacterial adaption; permanent

P. aeruginosa is a gram _____, _____-shaped bacteria that is very _____ in nature

negative; rod; ubiquitous

True or False: P. aeruginosa is a primary pathogen

FALSE: its opportunistic!

P. aeruginosa is an _____ human pathogen that can cause both _____ AND _____ infections

opportunistic; acute; chronic

P. aeruginosa in also a pathogen to many _____, _____, and _____ (apart from humans)

plants; insects; nematodes

P. aeruginosa is a model organism for the study of _____ and _____

biofilms; quorum sensing

what is an example of P. aeruginosa causing ACUTE infection?

burn infection, pneumonia

what is an example of P. aeruginosa causing CHRONIC infection?

catheter infection

Chronic and acute infections are fundamentally _____

different

acute infections _____ and _____, but are also _____ susceptible to treatment

disseminate; invade; MORE

acute infections are caused by _____ bacteria, while chronic infections are caused by _____ bacteria

planktonic; biofilm

chronic infections are _____ and _____ but are also _____ susceptible to treatment

localized; persist; LESS

in healthy cells, the mucus layer is much _____, which _____ for cilia function

thinner; ALLOWS

_____ caused by difunctional ion channels results in impaired _____

dehydration; mucociliary clearance

in CF patients there is a _____ of mucus that _____ cilia function

buildup; PREVENTS

CF causes persistent mucus _____ by _____ cells, which creates a very _____ mucus layer

hyper secretion; goblet; thick

P. aeruginosa is able to _____ the thick mucus and initiate _____ formation

colonize; biofilm

Because many longitudinal isolates are _____, we know that P. aeruginosa _____ to the lung environment

clonal; adapts

what are the 4 main adaptions that P. aeruginosa makes to adapt to the lung environment?

loss of motility, loss of LPS O-antigen, alginate overproduction, amino acid auxotrophy

Why does P. aeruginosa loose its motility function when adapting to the CF lungs?

no need to swim in thick mucus

Alginate is a type of _____ produced by P. aeruginosa

EPS

P. aeruginosa colonies that produce Alg appear much _____ and more _____ than Alg- colonies on agar

larger; mucoid

what does amino acid auxotrophy mean?

the organisms CANNOT make amino acids

What is MucA?

antisigma factor that prevents alginate production

What is the sigma factor that turns ON Alginate production?

AlgU sigma

when a mutation occurs in mucA, this activates the _____ sigma factor and _____ alginate production gene expression

AlgU sigma; increases

a number of CF isolated exhibit the _____ phenotype

RSCV

the CF airways select for _____ variants

RSCV

RSCV variants have enhanced _____ and _____, reduced _____, and increased _____

biofilm formation; antibiotic resistance; motility; cyclic di-GMP

which of these organisms is an RSCV variant?

left

What causes the RSCV phenotype in P. aeruginosa?

overproduction of cyclic di-GMP

RSCV variants will _____ in liquid culture

auto aggregate

Cyclic-diGMP is involved in a switch to a _____ lifestyle

biofilm

Cyclic di-GMP is an _____ signaling molecule that affects _____

intracellular; surface attachment

what is the protein that MAKES cyclic-di-GMP

cyclase

what is the protein that BREAKS DOWN cyclic-di-GMP

phosphodiesterase

what is the active site domain of Cyclase?

GGDEF

what is the active site domain for phosphodiesterase?

EAL

Cyclase takes _____ and removes _____ to makes cyclic-di-GMP

2 GTPs; 2 PPi

Phosphodiesterases convert cyclic di-GMP into _____

2 GMP

Cyclases and Phosphodiesterases are sensitive to a number of _____ factors

environmental

in many species _____ cyclic-di-GMP is linked to increased _____

ELEVATED; cell surface adhesiveness (biofilm formation)

what are the 2 main loci that play a role in RSCV variant types?

pel and psl

what happens if you delete both a pel and psl loci from an RSCV variant?

no aggregation at all!

what happens if you remove EITHER pel or psl loci from an RSCV variant?

partial relief → some aggregation, but much less