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DANIELA VERGARA YANEZ
MOLECULAR BIOLOGY
2018-2
INTRODUCTION
ESCHERICHIA COLI
 Gram-negative, bacillus-like bacterium of the enterobacteria family
 It is found in the gastrointestinal tract of humans and warm-blooded
animals
 The most abundant commensal facultative anaerobic bacterium in
the microbiota.
 It is one of the most relevant pathogenic organisms in humans.
 Produce gastrointestinal infections in other systems like: urinary,
sanguineous, nervous
 It has forms without mobility and mobile.
INTRODUCTION
SHIGA
TOXIN
uremic-hemolytic
síndrome
Enterohemorrhagic
diarrhea:
S. dysenteriae bacteria
Shigatoxigenic group of
Escherichia coli (STEC)
Serotype
O157:H7
Inhibit protein
synthesis
The A subunit injures
the eukaryotic
ribosome,B block
GB3 receptor
GENERAL OBJECTIVE
To prove shiga toxin producing escherichia coli O103:H2 outbreak in
Germany after school trip to Austria due to raw cow milk,2017.
MÉTODOS
Epidemiologia
Grupo de 22
estudiantes
con 9
intoxicados
en hotel de
Austria,2017
Diarrea,
vomito y dolor
abdominal, 10
días luego del
viaje
Cuestionario a
alumnos sobre
menú del hotel.
Huéspedes
pasados
presentaron
síntomas similares
Se descubre en
el desayuno
leche de vaca
cruda
MÉTODOS
PCR: amplificación de un gen o fragmento de DNA
DNA a
amplificar(heces
humanas)
Desnatulizar DNA a 90-97℃
Hibridación hebras
sencillas de DNA
Agregar
primer/oligonucleótido y
DNTp´s
37-65 ℃
DNA polimerasa (taq
polimerasa) inicia
replicación desde
primer
Se realiza a 72-75 ℃
Dirección 5´3`
PCR
RFLP
Uso enzimas de
restricción para cortar
secuencias especificas
de DNA(e.coli),
variabilidad, luego
electroforesis
Usado para
amplificación y
detección de genes de
toxina shiga
MÉTODOS
ELISA
 Técnica donde se identifica un anticuerpo o
antígeno utilizando los mismos. Para revelar la
reacción antígeno-anticuerpo se usa anti
gammaglobulina unida a una enzima,
 Intensidad leído en
espectrofotómetro=cantidad de anticuerpo o
antígeno
AGLUTINACION POR LATEX
 Se agrega a la muestra gotas de latex, si se
unen aglutinan las partículas la sustancia
estudiada esta presente.
Para la detección
de genes de
toxinas similares a
Shiga (stx1, stx2)
MÉTODOS
METODO QUIMICO
 Hidrolisis química y diseño original
aplicable a secuencias de DNA de
menos de 250 nucleótidos
METODO ENZIMATICO
 Interrupción controlada de síntesis de
una hebra complementaria en
replicación in vitro
 Catalizado por DNA polimerasa
Marcaje DNA
Hidrolisis química
selectiva DNA
Análisis de los
productos
Síntesis enzimática
Análisis de los
fragmentos
Automatización:
fluorocromo
Se realizo en
muestras humanas
positivas para la
bacteria, para
determinar el lugar de
acción de la bacteria
nivel genético
SECUENCIACIÓN DEL GENOMA
RESULTADOS
RESULTADOS
RESULTADOS
RESULTADOS
DISCUSSION
AUTHOR WHAT HE SAID YES OR
NOT
Headrick et
al.,1998;Mungai
et al.,2015
Both epidemiological and microbiological results identified the
raw cow milk provided by the hotel as the vehicle of the
outbreak. The environmental and microbiological
investigations involving PFGE and WGS-based cgMLST
analysis detected closely related STEC strains in humans
and cattle at the milk-delivering dairy farm. It is well known
that raw or unpasteurized milk contains a broad range of
pathogens, e.g. Campylobacter, Listeria, Mycobacterium
bovis, STEC, Salmonella or Norovirus.
Allerberger et
al.,2003
Two cases of HUS in small children occurred in Austria in
2003, when an 11-monthold boy and a 28-month-old girl both
had consumed unpasteurized cow's milk which was offered
unlabeled at the breakfast buffet of a hotel, similar to the
present outbreak. Laboratory analyses later revealed the
presence of E. coli O26:H- in the milk, confirming it as the
vehicle of infection.
¡Recordar
que solo
hay dos
autores!
CONCLUSIONS
 This articule contributed to decrese the cases of intoxication with shiga
toxin included in raw cow milk, which is very common in hotel´s menú,
specifically in Austria, because the hotels stopped offering this milk in
breakfast for guests.
 Microbiology techqniques day a day improve a lot of diagnoses, in this
case STEC and HUS,beacuse it´s posible look specifics gen where is the
toxin codificated and how it acts with the final purpose to understand and
treat the pathology that causes.
Seminario biología molecular 2018-2

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Seminario biología molecular 2018-2

  • 2. INTRODUCTION ESCHERICHIA COLI  Gram-negative, bacillus-like bacterium of the enterobacteria family  It is found in the gastrointestinal tract of humans and warm-blooded animals  The most abundant commensal facultative anaerobic bacterium in the microbiota.  It is one of the most relevant pathogenic organisms in humans.  Produce gastrointestinal infections in other systems like: urinary, sanguineous, nervous  It has forms without mobility and mobile.
  • 3. INTRODUCTION SHIGA TOXIN uremic-hemolytic síndrome Enterohemorrhagic diarrhea: S. dysenteriae bacteria Shigatoxigenic group of Escherichia coli (STEC) Serotype O157:H7 Inhibit protein synthesis The A subunit injures the eukaryotic ribosome,B block GB3 receptor
  • 4. GENERAL OBJECTIVE To prove shiga toxin producing escherichia coli O103:H2 outbreak in Germany after school trip to Austria due to raw cow milk,2017.
  • 5. MÉTODOS Epidemiologia Grupo de 22 estudiantes con 9 intoxicados en hotel de Austria,2017 Diarrea, vomito y dolor abdominal, 10 días luego del viaje Cuestionario a alumnos sobre menú del hotel. Huéspedes pasados presentaron síntomas similares Se descubre en el desayuno leche de vaca cruda
  • 6. MÉTODOS PCR: amplificación de un gen o fragmento de DNA DNA a amplificar(heces humanas) Desnatulizar DNA a 90-97℃ Hibridación hebras sencillas de DNA Agregar primer/oligonucleótido y DNTp´s 37-65 ℃ DNA polimerasa (taq polimerasa) inicia replicación desde primer Se realiza a 72-75 ℃ Dirección 5´3` PCR RFLP Uso enzimas de restricción para cortar secuencias especificas de DNA(e.coli), variabilidad, luego electroforesis Usado para amplificación y detección de genes de toxina shiga
  • 7. MÉTODOS ELISA  Técnica donde se identifica un anticuerpo o antígeno utilizando los mismos. Para revelar la reacción antígeno-anticuerpo se usa anti gammaglobulina unida a una enzima,  Intensidad leído en espectrofotómetro=cantidad de anticuerpo o antígeno AGLUTINACION POR LATEX  Se agrega a la muestra gotas de latex, si se unen aglutinan las partículas la sustancia estudiada esta presente. Para la detección de genes de toxinas similares a Shiga (stx1, stx2)
  • 8. MÉTODOS METODO QUIMICO  Hidrolisis química y diseño original aplicable a secuencias de DNA de menos de 250 nucleótidos METODO ENZIMATICO  Interrupción controlada de síntesis de una hebra complementaria en replicación in vitro  Catalizado por DNA polimerasa Marcaje DNA Hidrolisis química selectiva DNA Análisis de los productos Síntesis enzimática Análisis de los fragmentos Automatización: fluorocromo Se realizo en muestras humanas positivas para la bacteria, para determinar el lugar de acción de la bacteria nivel genético SECUENCIACIÓN DEL GENOMA
  • 13. DISCUSSION AUTHOR WHAT HE SAID YES OR NOT Headrick et al.,1998;Mungai et al.,2015 Both epidemiological and microbiological results identified the raw cow milk provided by the hotel as the vehicle of the outbreak. The environmental and microbiological investigations involving PFGE and WGS-based cgMLST analysis detected closely related STEC strains in humans and cattle at the milk-delivering dairy farm. It is well known that raw or unpasteurized milk contains a broad range of pathogens, e.g. Campylobacter, Listeria, Mycobacterium bovis, STEC, Salmonella or Norovirus. Allerberger et al.,2003 Two cases of HUS in small children occurred in Austria in 2003, when an 11-monthold boy and a 28-month-old girl both had consumed unpasteurized cow's milk which was offered unlabeled at the breakfast buffet of a hotel, similar to the present outbreak. Laboratory analyses later revealed the presence of E. coli O26:H- in the milk, confirming it as the vehicle of infection. ¡Recordar que solo hay dos autores!
  • 14. CONCLUSIONS  This articule contributed to decrese the cases of intoxication with shiga toxin included in raw cow milk, which is very common in hotel´s menú, specifically in Austria, because the hotels stopped offering this milk in breakfast for guests.  Microbiology techqniques day a day improve a lot of diagnoses, in this case STEC and HUS,beacuse it´s posible look specifics gen where is the toxin codificated and how it acts with the final purpose to understand and treat the pathology that causes.