Tijmen Lourens Summary Microbes and Infection
Innate and adaptive immune responses to micro-organisms
Immune responses to bacterial infection
1) Recognition / sensing
a. TLRs
b. NLRs
c. CDS (cytosolic DNA sensor)
2) Innate response
a. Phagocytosis
b. Complement
c. inflammation
3) Adaptive response
a. Antibodies
i. Binding of bacterial toxins
ii. Killing (by complement activation)
b. CD4 T cells
i. Production of cytokines
ii. Activation of macrophages
c. CD8 T cells
i. Killing of infected cells
ii. Production of cytokines
LPS in gram-negative bacteria can be fatal → septic shock
Bacteria and LPS in blood → cytokines storm: (TNF, IL-6, IL-1) →
Hemodynamic shock by:
- Heart failure
- Loss of blood pressure
Bacterial infection:
Extracellular bacteria:
Staphylococcus aureus and clostridium tetani are pathogenic
Pathogenic mechanisms:
- Production of toxins
- Induction of inflammation
Intracellular bacteria:
- Mycobacterium laprae
- Mycobacterium tuberculosis
- Legionella pneumophila
- Porphyromonas gingivalis
,Tijmen Lourens Summary Microbes and Infection
Location: live/hide in phagocytes
Pathogenic mechanisms:
- Host immune response (granulomas, cytokines)
- Induction of epigenetic changes leading to gene silencing or activation
The innate immune mechanisms that are inhibited by a staphylococcal immune evasion:
1) Neutrophil extravasation
2) Priming, chemotaxis, and activation of neutrophils
3) Opsonization and phagocytosis
4) NET formation
5) Bacterial killing by neutrophils
Lecture 4: infections
Infectious diseases are acute problems that need fast answer for clinician to save patients
life. Therefore fast diagnostics is crucial.
How to find out what pathogen makes you sick? → 2 ways
- Look for pathogen specific DNA
- Look for pathogen specific protein
How to identify and type the bacterial pathogens?
- Looking at them through a microscope
- Growing them on a proper media, looking at their colour, form of grown colonies and
the smell. (You need to wait at least 24h!)
- Antibiotic sensitivity testing → to know which antibiotic will work for the patient
treatment (you wait another 24h)
Classic culture is being called the ‘gold standard’.
Bacterial DNA:
- Every bacterium has unique DNA sequence
- Specific bacterial genes are being used for quick identification of pathogens as well as
their resistance, virulence and typing
- Specific genes are used for fast diagnostic tests where the results are known within
couple of hours
Molecular typing is a way of identifying specific strains of bacteria or viruses, by looking at
their genetic material.
,Tijmen Lourens Summary Microbes and Infection
For bacterial typing there are 3 methods used:
1) Pulsed-field gel electrophoresis (PFGE)
2) Multiple locus variable-number tandem repeat analysis (MLVA)
3) Multilocus sequence typing (MLST)
Why did we integrate next generation sequencing (NGS) in the routine workflow?
- To type with the highest possible discriminatory power (hospital outbreaks)
- To reveal antibiotics resistance and virulence genes
- To identify novel bacterial species
Metagenomics: high-throughput sequencing for in the future (not yet applied)
- Profiling microbial communities (shotgun genomics)
- Universal pathogen detection without culture
- Detection of virulence and resistance genes in sample
- Determining host/pathogen response by sequencing DNA and RNA from the same
sample (prognosis for patient outcome)
, Tijmen Lourens Summary Microbes and Infection
Summary:
- There are different techniques available for molecular diagnostics of microbial
pathogens
- Which technique is being chosen depends on:
o Sample type
o Time you have to get the answer (if there is a sceptic patient in the hospital
you need to be really fast)
- Infection prevention is becoming more and more important in hospitals to control
potential spread of the pathogens between hospitalized patients
- Also, the use of molecular typing of pathogens is crucial for epidemiology to properly
monitor spread of pathogen on local, national, international, and global level
- In the recent years importance of NGS in routine diagnostics, infection prevention
and epidemiology are enormously increased. This method with so far the highest
discriminatory power is still getting improved to shorten time to the results and
reduce the costs.
Read chapter 5.
Understand PCR, real-time PCR and MALDI-TOF.
How to detect presence of bacteria or virus in patient material
- Take a blot and use (16S) PCR.
Understand difference between sequencing techniques: sanger sequencing, illumine
sequencing, IonTorrent sequencing and Minlon sequencing. (nanopore sequencing)
Lecture 5: respiratory infections (Streptococcus pneumoniae and Mycobacterium
tuberculosis)
Upper respiratory tract containing the nasal and oral cavity is not sterile. Bacteria can easily
get into your body via breathing.
Infections in the upper respiratory tract are common cold, sinusitis or otitis media.
The lower respiratory tract is thought to be sterile. They are actually not fully sterile but
there are a lot less bacteria and viruses in your lungs.
Infections in the lower respiratory tract are bronchitis, pneumonia, tuberculosis,
legionnaires’ disease.
Innate and adaptive immune responses to micro-organisms
Immune responses to bacterial infection
1) Recognition / sensing
a. TLRs
b. NLRs
c. CDS (cytosolic DNA sensor)
2) Innate response
a. Phagocytosis
b. Complement
c. inflammation
3) Adaptive response
a. Antibodies
i. Binding of bacterial toxins
ii. Killing (by complement activation)
b. CD4 T cells
i. Production of cytokines
ii. Activation of macrophages
c. CD8 T cells
i. Killing of infected cells
ii. Production of cytokines
LPS in gram-negative bacteria can be fatal → septic shock
Bacteria and LPS in blood → cytokines storm: (TNF, IL-6, IL-1) →
Hemodynamic shock by:
- Heart failure
- Loss of blood pressure
Bacterial infection:
Extracellular bacteria:
Staphylococcus aureus and clostridium tetani are pathogenic
Pathogenic mechanisms:
- Production of toxins
- Induction of inflammation
Intracellular bacteria:
- Mycobacterium laprae
- Mycobacterium tuberculosis
- Legionella pneumophila
- Porphyromonas gingivalis
,Tijmen Lourens Summary Microbes and Infection
Location: live/hide in phagocytes
Pathogenic mechanisms:
- Host immune response (granulomas, cytokines)
- Induction of epigenetic changes leading to gene silencing or activation
The innate immune mechanisms that are inhibited by a staphylococcal immune evasion:
1) Neutrophil extravasation
2) Priming, chemotaxis, and activation of neutrophils
3) Opsonization and phagocytosis
4) NET formation
5) Bacterial killing by neutrophils
Lecture 4: infections
Infectious diseases are acute problems that need fast answer for clinician to save patients
life. Therefore fast diagnostics is crucial.
How to find out what pathogen makes you sick? → 2 ways
- Look for pathogen specific DNA
- Look for pathogen specific protein
How to identify and type the bacterial pathogens?
- Looking at them through a microscope
- Growing them on a proper media, looking at their colour, form of grown colonies and
the smell. (You need to wait at least 24h!)
- Antibiotic sensitivity testing → to know which antibiotic will work for the patient
treatment (you wait another 24h)
Classic culture is being called the ‘gold standard’.
Bacterial DNA:
- Every bacterium has unique DNA sequence
- Specific bacterial genes are being used for quick identification of pathogens as well as
their resistance, virulence and typing
- Specific genes are used for fast diagnostic tests where the results are known within
couple of hours
Molecular typing is a way of identifying specific strains of bacteria or viruses, by looking at
their genetic material.
,Tijmen Lourens Summary Microbes and Infection
For bacterial typing there are 3 methods used:
1) Pulsed-field gel electrophoresis (PFGE)
2) Multiple locus variable-number tandem repeat analysis (MLVA)
3) Multilocus sequence typing (MLST)
Why did we integrate next generation sequencing (NGS) in the routine workflow?
- To type with the highest possible discriminatory power (hospital outbreaks)
- To reveal antibiotics resistance and virulence genes
- To identify novel bacterial species
Metagenomics: high-throughput sequencing for in the future (not yet applied)
- Profiling microbial communities (shotgun genomics)
- Universal pathogen detection without culture
- Detection of virulence and resistance genes in sample
- Determining host/pathogen response by sequencing DNA and RNA from the same
sample (prognosis for patient outcome)
, Tijmen Lourens Summary Microbes and Infection
Summary:
- There are different techniques available for molecular diagnostics of microbial
pathogens
- Which technique is being chosen depends on:
o Sample type
o Time you have to get the answer (if there is a sceptic patient in the hospital
you need to be really fast)
- Infection prevention is becoming more and more important in hospitals to control
potential spread of the pathogens between hospitalized patients
- Also, the use of molecular typing of pathogens is crucial for epidemiology to properly
monitor spread of pathogen on local, national, international, and global level
- In the recent years importance of NGS in routine diagnostics, infection prevention
and epidemiology are enormously increased. This method with so far the highest
discriminatory power is still getting improved to shorten time to the results and
reduce the costs.
Read chapter 5.
Understand PCR, real-time PCR and MALDI-TOF.
How to detect presence of bacteria or virus in patient material
- Take a blot and use (16S) PCR.
Understand difference between sequencing techniques: sanger sequencing, illumine
sequencing, IonTorrent sequencing and Minlon sequencing. (nanopore sequencing)
Lecture 5: respiratory infections (Streptococcus pneumoniae and Mycobacterium
tuberculosis)
Upper respiratory tract containing the nasal and oral cavity is not sterile. Bacteria can easily
get into your body via breathing.
Infections in the upper respiratory tract are common cold, sinusitis or otitis media.
The lower respiratory tract is thought to be sterile. They are actually not fully sterile but
there are a lot less bacteria and viruses in your lungs.
Infections in the lower respiratory tract are bronchitis, pneumonia, tuberculosis,
legionnaires’ disease.
