SAMENVATTING BACTERIOLOGIE
inleiding___________________________________________________________________ 5
TAXONOMIE BINNEN DE BACTERIOLOGIE ________________________________________ 5
Leerdoelen _____________________________________________________________________________ 5
Terrein van de medische microbiologie ______________________________________________________ 5
Classificatie van micro-organismen __________________________________________________________ 6
Morfologische classificatie ______________________________________________________________ 6
Biochemische classificatie ______________________________________________________________ 10
Numerische classificatie _______________________________________________________________ 11
Fylogenetische classificatie _____________________________________________________________ 11
Opdrachten ___________________________________________________________________________ 12
pathogeniciteit ________________________________________________________________ 13
DE MICROSCOOP __________________________________________________________ 13
De onderdelen van de microscoop _________________________________________________ 13
De helderveldmicroscoop ________________________________________________________________ 13
De donkerveldmicroscoop________________________________________________________________ 13
De fasecontrastmicroscoop _______________________________________________________________ 13
Fluorescentiemicroscopie ________________________________________________________________ 14
VERS ONDERZOEK _____________________________________________________________ 14
DE GRAMKLEURING ____________________________________________________________ 14
Uitvoering ____________________________________________________________________________ 14
Het principe van de Gram-kleuring _________________________________________________________ 15
DE ZIEHL-NEELSEN-KLEURING ____________________________________________________ 16
DE AURAMINEKLEURING =NL______________________________________________________ 16
IMMUNOFLUORESCENTIETECHNIEKEN _____________________________________________ 16
FLAGELLAKLEURING ____________________________________________________________ 17
DE GIËMSAKLEURING =3 De jaar __________________________________________________ 17
DE CALCOFLUORKLEURING ______________________________________________________ 17
OPDRACHTEN _________________________________________________________________ 17
WEEKETHODEN ____________________________________________________________ 18
LEERDOELEN __________________________________________________________________ 18
ALGEMENE INLEIDING __________________________________________________________ 18
ISOLATIEBODEMS ______________________________________________________________ 18
Universele vaste/vloeibare voedingsbodems _________________________________________________ 19
Bloedagar ___________________________________________________________________________ 19
BHI: Brain Heart infusion _______________________________________________________________ 19
Chocolade-agar ______________________________________________________________________ 19
Universele vloeibare voedingsbodems ____________________________________________________ 20
Bacteriologie 1 1
, Selectieve voedingsbodems ______________________________________________________________ 20
MacConkey agar (McC): _______________________________________________________________ 20
MSA (mannitol salt agar): ______________________________________________________________ 21
Hectoen Enteric Agar (HE) ______________________________________________________________ 21
Eosine Methyleenblauw Agar = EMB _____________________________________________________ 22
CLED: Cystine Lactose Electrolyte Deficient Agar ____________________________________________ 22
Andere te kennen VB’s ________________________________________________________________ 22
Aanrijkingsbodems _____________________________________________________________________ 22
Specifieke isolatiebodems ________________________________________________________________ 23
ENTINGSTECHNIEKEN ___________________________________________________________ 23
INCUBATIE-OMSTANDIGHEDEN ___________________________________________________ 24
Temperatuur __________________________________________________________________________ 24
Omgeving _____________________________________________________________________________ 24
OPDRACHTEN _________________________________________________________________ 25
Biochemische identificatiemethodes ___________________________________________ 25
LEERDOELEN __________________________________________________________________ 25
BIOCHEMISCHE TESTEN IN EEN STRATEGIE ______________________________________ 26
PRIMAIRE KENMERKEN _________________________________________________________ 26
Morfologische kenmerken _______________________________________________________________ 26
Groeivoorwaarden _____________________________________________________________________ 26
Omgevingsatmosfeer _________________________________________________________________ 27
Temperatuurtolerantie ________________________________________________________________ 27
Zouttolerantie _______________________________________________________________________ 27
Galtolerantie en galstimulatie ___________________________________________________________ 27
Gevoeligheid voor chemische stoffen en antibiotica ___________________________________________ 27
Pigmentproductie ______________________________________________________________________ 28
Hemolyse _____________________________________________________________________________ 28
- en -hemolyse ____________________________________________________________________ 28
De CAMP-test__________________________________________________________________________ 29
De omgekeerde CAMP-test _______________________________________________________________ 30
Snelle enzymatische testen _______________________________________________________________ 31
KatalasetestReactie: (principe) __________________________________________________________ 31
Cytochroomoxidasetest _______________________________________________________________ 31
Coagulasetest _______________________________________________________________________ 32
Latexagglutinatietesten __________________________________________________________________ 33
SECUNDAIRE KENMERKEN _____________________________________________________ 33
GECOMBINEERDE TESTMEDIA __________________________________________________ 33
Kligler Iron Agar (KIA)____________________________________________________________________ 33
Doel _______________________________________________________________________________ 33
Principe ____________________________________________________________________________ 34
Werkwijze ______________________________________________________________________ 34
tSI en MIU ____________________________________________________________________________ 37
Mobiliteit-Indol-Ornithine medium (MIO) ___________________________________________________ 37
Lysine Iron Agar (LIA) ____________________________________________________________________ 38
Bacteriologie 1 2
, COMMERCIËLE BIOCHEMISCHE IDENTIFICATIESYSTEMEN _____________________________ 38
OPDRACHTEN _________________________________________________________________ 39
MALDI-TOF- MS _______________________________________________________________ 39
Doel _________________________________________________________________________________ 39
Principe ______________________________________________________________________________ 39
MOLECULAIRE TECHNIEKEN IN DE DIAGNOSTIEK BIJ BACTERIËLE INFECTIES ___________ 40
LEERDOELEN __________________________________________________________________ 40
INLEIDING ____________________________________________________________________ 40
NUCLEÏNEZUREN EN HUN INTERACTIES ___________________________________________ 41
TOEPASSING VAN MOLECULAIRE TECHNIEKEN IN het MICROBIOLOGIE lab _____________ 41
Aantonen aanwezigheid van een species i.e. klinisch staal ______________________________________ 41
PCR ________________________________________________________________________________ 41
qPCR en melting curve analysis (MCA) ____________________________________________________ 44
Western blotting : niet in ppt ___________________________________________________________ 46
Genotypering van stammen via fingerprinting ________________________________________________ 46
RFLP _______________________________________________________________________________ 46
Southern blotting => enkel int kort in boek ________________________________________________ 47
Amplified fragment length polymorphism _______________________________________________ 48
Pulsed Field Gelelektroforese _________________________________________________________ 49
Northern blot = nl? ___________________________________________________________________ 50
Random amplification of polymorphic DNA (RAPD) _________________________________________ 50
PCR: short tandem repeats _____________________________________________________________ 51
Genotypering van bacteriën ______________________________________________________ 51
HUIDIGE DIAGNOSTIEK IN HET LAB MICROBIOLOGIE In het routinelabo bacteriologie 52
OPDRACHTEN _________________________________________________________________ 52
6 ENTEROBACTERIACEAE _______________________________________________ 52
LEERDOELEN __________________________________________________________________ 52
DE GRAM-NEGATIEVE BACTERIËN _______________________________________________ 52
GROEPEN bij de Gram-negatieve bacteriën: _________________________________________________ 53
GESLACHTEN EN ALGEMENE EIGENSCHAPPEN _____________________________________ 55
CULTUUR EN BIOCHEMISCHE IDENTIFICATIE ______________________________________ 56
DE ENTEROPATHOGENEN _______________________________________________________ 57
Salmonella enterica _____________________________________________________________________ 57
Specifieke kenmerken _________________________________________________________________ 57
6.5.1.2 Pathogenese ____________________________________________________________ 58
Pathogeen vermogen _________________________________________________________________ 58
Gastro‐enteritis ____________________________________________________________________ 58
Darmkoorts (tyfus) _________________________________________________________________ 59
Sepsis ____________________________________________________________________________ 59
Staal ________________________________________________________ Error! Bookmark not defined.
Isolatie en identificatie ________________________________________________________________ 59
Bacteriologie 1 3
, Serotypering van Salmonella ___________________________________________________________ 60
Shigella spp. ___________________________________________________________________________ 61
Specifieke kenmerken _________________________________________________________________ 61
Pathogenese ________________________________________________________________________ 61
Pathogeen vermogen _________________________________________________________________ 61
Staal _______________________________________________________________________________ 61
Isolatie en identificatie ________________________________________________________________ 62
Yersinia spp. ___________________________________________________________________________ 62
Specifieke kenmerken _________________________________________________________________ 62
Pathogenese ________________________________________________________________________ 62
Pathogeen vermogen _________________________________________________________________ 62
Staal _______________________________________________________________________________ 63
Isolatie en identificatie ________________________________________________________________ 63
E. coli ________________________________________________________________________________ 64
Specifieke kenmerken _________________________________________________________________ 64
Pathogenese ________________________________________________________________________ 64
Pathogeen vermogen _________________________________________________________________ 64
- EHEC: enterohemorragische E. coli stammen ______________________________________ 65
Staal _______________________________________________________________________________ 66
Isolatie en identificatie ________________________________________________________________ 66
OPPORTUNISTISCHE PATHOGENEN ______________________________________________ 67
Enterobacter spp. ______________________________________________________________________ 67
Klebsiella spp. _________________________________________________________________________ 67
Proteus-Providencia-Morganella __________________________________________________________ 68
Serratia spp. ___________________________________________________________________________ 69
Citrobacter spp. ________________________________________________________________________ 69
Hafnia alvei ___________________________________________________________________________ 69
Edwarsiella tarda ________________________________________________ Error! Bookmark not defined.
Andere Enterobacteriaceae ________________________________________ Error! Bookmark not defined.
7 OXIDASE-POSITIEVE ____________________________ Error! Bookmark not defined.
LEERDOELEN _____________________________________________ Error! Bookmark not defined.
Bacteriologie 1 4
inleiding___________________________________________________________________ 5
TAXONOMIE BINNEN DE BACTERIOLOGIE ________________________________________ 5
Leerdoelen _____________________________________________________________________________ 5
Terrein van de medische microbiologie ______________________________________________________ 5
Classificatie van micro-organismen __________________________________________________________ 6
Morfologische classificatie ______________________________________________________________ 6
Biochemische classificatie ______________________________________________________________ 10
Numerische classificatie _______________________________________________________________ 11
Fylogenetische classificatie _____________________________________________________________ 11
Opdrachten ___________________________________________________________________________ 12
pathogeniciteit ________________________________________________________________ 13
DE MICROSCOOP __________________________________________________________ 13
De onderdelen van de microscoop _________________________________________________ 13
De helderveldmicroscoop ________________________________________________________________ 13
De donkerveldmicroscoop________________________________________________________________ 13
De fasecontrastmicroscoop _______________________________________________________________ 13
Fluorescentiemicroscopie ________________________________________________________________ 14
VERS ONDERZOEK _____________________________________________________________ 14
DE GRAMKLEURING ____________________________________________________________ 14
Uitvoering ____________________________________________________________________________ 14
Het principe van de Gram-kleuring _________________________________________________________ 15
DE ZIEHL-NEELSEN-KLEURING ____________________________________________________ 16
DE AURAMINEKLEURING =NL______________________________________________________ 16
IMMUNOFLUORESCENTIETECHNIEKEN _____________________________________________ 16
FLAGELLAKLEURING ____________________________________________________________ 17
DE GIËMSAKLEURING =3 De jaar __________________________________________________ 17
DE CALCOFLUORKLEURING ______________________________________________________ 17
OPDRACHTEN _________________________________________________________________ 17
WEEKETHODEN ____________________________________________________________ 18
LEERDOELEN __________________________________________________________________ 18
ALGEMENE INLEIDING __________________________________________________________ 18
ISOLATIEBODEMS ______________________________________________________________ 18
Universele vaste/vloeibare voedingsbodems _________________________________________________ 19
Bloedagar ___________________________________________________________________________ 19
BHI: Brain Heart infusion _______________________________________________________________ 19
Chocolade-agar ______________________________________________________________________ 19
Universele vloeibare voedingsbodems ____________________________________________________ 20
Bacteriologie 1 1
, Selectieve voedingsbodems ______________________________________________________________ 20
MacConkey agar (McC): _______________________________________________________________ 20
MSA (mannitol salt agar): ______________________________________________________________ 21
Hectoen Enteric Agar (HE) ______________________________________________________________ 21
Eosine Methyleenblauw Agar = EMB _____________________________________________________ 22
CLED: Cystine Lactose Electrolyte Deficient Agar ____________________________________________ 22
Andere te kennen VB’s ________________________________________________________________ 22
Aanrijkingsbodems _____________________________________________________________________ 22
Specifieke isolatiebodems ________________________________________________________________ 23
ENTINGSTECHNIEKEN ___________________________________________________________ 23
INCUBATIE-OMSTANDIGHEDEN ___________________________________________________ 24
Temperatuur __________________________________________________________________________ 24
Omgeving _____________________________________________________________________________ 24
OPDRACHTEN _________________________________________________________________ 25
Biochemische identificatiemethodes ___________________________________________ 25
LEERDOELEN __________________________________________________________________ 25
BIOCHEMISCHE TESTEN IN EEN STRATEGIE ______________________________________ 26
PRIMAIRE KENMERKEN _________________________________________________________ 26
Morfologische kenmerken _______________________________________________________________ 26
Groeivoorwaarden _____________________________________________________________________ 26
Omgevingsatmosfeer _________________________________________________________________ 27
Temperatuurtolerantie ________________________________________________________________ 27
Zouttolerantie _______________________________________________________________________ 27
Galtolerantie en galstimulatie ___________________________________________________________ 27
Gevoeligheid voor chemische stoffen en antibiotica ___________________________________________ 27
Pigmentproductie ______________________________________________________________________ 28
Hemolyse _____________________________________________________________________________ 28
- en -hemolyse ____________________________________________________________________ 28
De CAMP-test__________________________________________________________________________ 29
De omgekeerde CAMP-test _______________________________________________________________ 30
Snelle enzymatische testen _______________________________________________________________ 31
KatalasetestReactie: (principe) __________________________________________________________ 31
Cytochroomoxidasetest _______________________________________________________________ 31
Coagulasetest _______________________________________________________________________ 32
Latexagglutinatietesten __________________________________________________________________ 33
SECUNDAIRE KENMERKEN _____________________________________________________ 33
GECOMBINEERDE TESTMEDIA __________________________________________________ 33
Kligler Iron Agar (KIA)____________________________________________________________________ 33
Doel _______________________________________________________________________________ 33
Principe ____________________________________________________________________________ 34
Werkwijze ______________________________________________________________________ 34
tSI en MIU ____________________________________________________________________________ 37
Mobiliteit-Indol-Ornithine medium (MIO) ___________________________________________________ 37
Lysine Iron Agar (LIA) ____________________________________________________________________ 38
Bacteriologie 1 2
, COMMERCIËLE BIOCHEMISCHE IDENTIFICATIESYSTEMEN _____________________________ 38
OPDRACHTEN _________________________________________________________________ 39
MALDI-TOF- MS _______________________________________________________________ 39
Doel _________________________________________________________________________________ 39
Principe ______________________________________________________________________________ 39
MOLECULAIRE TECHNIEKEN IN DE DIAGNOSTIEK BIJ BACTERIËLE INFECTIES ___________ 40
LEERDOELEN __________________________________________________________________ 40
INLEIDING ____________________________________________________________________ 40
NUCLEÏNEZUREN EN HUN INTERACTIES ___________________________________________ 41
TOEPASSING VAN MOLECULAIRE TECHNIEKEN IN het MICROBIOLOGIE lab _____________ 41
Aantonen aanwezigheid van een species i.e. klinisch staal ______________________________________ 41
PCR ________________________________________________________________________________ 41
qPCR en melting curve analysis (MCA) ____________________________________________________ 44
Western blotting : niet in ppt ___________________________________________________________ 46
Genotypering van stammen via fingerprinting ________________________________________________ 46
RFLP _______________________________________________________________________________ 46
Southern blotting => enkel int kort in boek ________________________________________________ 47
Amplified fragment length polymorphism _______________________________________________ 48
Pulsed Field Gelelektroforese _________________________________________________________ 49
Northern blot = nl? ___________________________________________________________________ 50
Random amplification of polymorphic DNA (RAPD) _________________________________________ 50
PCR: short tandem repeats _____________________________________________________________ 51
Genotypering van bacteriën ______________________________________________________ 51
HUIDIGE DIAGNOSTIEK IN HET LAB MICROBIOLOGIE In het routinelabo bacteriologie 52
OPDRACHTEN _________________________________________________________________ 52
6 ENTEROBACTERIACEAE _______________________________________________ 52
LEERDOELEN __________________________________________________________________ 52
DE GRAM-NEGATIEVE BACTERIËN _______________________________________________ 52
GROEPEN bij de Gram-negatieve bacteriën: _________________________________________________ 53
GESLACHTEN EN ALGEMENE EIGENSCHAPPEN _____________________________________ 55
CULTUUR EN BIOCHEMISCHE IDENTIFICATIE ______________________________________ 56
DE ENTEROPATHOGENEN _______________________________________________________ 57
Salmonella enterica _____________________________________________________________________ 57
Specifieke kenmerken _________________________________________________________________ 57
6.5.1.2 Pathogenese ____________________________________________________________ 58
Pathogeen vermogen _________________________________________________________________ 58
Gastro‐enteritis ____________________________________________________________________ 58
Darmkoorts (tyfus) _________________________________________________________________ 59
Sepsis ____________________________________________________________________________ 59
Staal ________________________________________________________ Error! Bookmark not defined.
Isolatie en identificatie ________________________________________________________________ 59
Bacteriologie 1 3
, Serotypering van Salmonella ___________________________________________________________ 60
Shigella spp. ___________________________________________________________________________ 61
Specifieke kenmerken _________________________________________________________________ 61
Pathogenese ________________________________________________________________________ 61
Pathogeen vermogen _________________________________________________________________ 61
Staal _______________________________________________________________________________ 61
Isolatie en identificatie ________________________________________________________________ 62
Yersinia spp. ___________________________________________________________________________ 62
Specifieke kenmerken _________________________________________________________________ 62
Pathogenese ________________________________________________________________________ 62
Pathogeen vermogen _________________________________________________________________ 62
Staal _______________________________________________________________________________ 63
Isolatie en identificatie ________________________________________________________________ 63
E. coli ________________________________________________________________________________ 64
Specifieke kenmerken _________________________________________________________________ 64
Pathogenese ________________________________________________________________________ 64
Pathogeen vermogen _________________________________________________________________ 64
- EHEC: enterohemorragische E. coli stammen ______________________________________ 65
Staal _______________________________________________________________________________ 66
Isolatie en identificatie ________________________________________________________________ 66
OPPORTUNISTISCHE PATHOGENEN ______________________________________________ 67
Enterobacter spp. ______________________________________________________________________ 67
Klebsiella spp. _________________________________________________________________________ 67
Proteus-Providencia-Morganella __________________________________________________________ 68
Serratia spp. ___________________________________________________________________________ 69
Citrobacter spp. ________________________________________________________________________ 69
Hafnia alvei ___________________________________________________________________________ 69
Edwarsiella tarda ________________________________________________ Error! Bookmark not defined.
Andere Enterobacteriaceae ________________________________________ Error! Bookmark not defined.
7 OXIDASE-POSITIEVE ____________________________ Error! Bookmark not defined.
LEERDOELEN _____________________________________________ Error! Bookmark not defined.
Bacteriologie 1 4
