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Detection and quantification of GMO in food and raw material
Detection of food and cosmetics pathogens
Acredited laboratory
Since April 1th 2006 (CIA)
usage of highly sensitive system PCR a Real on Time PCR
based on standard normative methods
based on moleculár genetics methods
PCR – Polymerase chain reaction
DNA chips
-Salmonella sp. -Listeria monocytogenes -Campylobacter sp. -Cronobacter sakazakii -Staphylococcus auresus
based on imunochemical methods - Listeria monocytogenes - Staphylococcus auresus
Tests of microbial cosmetics contamination
member of ENGL (European Network of GMO laboratories)
member of GMO laboratories network in CR and SR
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Topics for presentations
1. Mechanism of penicillin +vancomycin and bacitracine effect on bacteria and resistence.
2. Antibiotics interfering with bacterial proteosynthesis, and mechanism of resistence
3. Effect of various types of thermal treatment on food – impact on microorganisms.
4. Effect of physical (excluding thermal) on food - impact on microorganisms.
5. Impact of bacterial sporulation on food.
6. Biofilm control strategies in food industry.
7. Typical spoilage microorganisms of cereals.
8. Means of disinfection and sanitation in food industry.
9. Survey of major food-born epidemies in the last 5 years in Europe.
10. Typical spoilage microorganism of beverages
11. Principle of continuous cultivation – production of biomass
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• Avetian David
• Guilbault Helene
• Le Stum Sophie
• Olshtrem Anastasiya
• Pasquelins Anais
• Ras Amandine
• von Korff Pauline
• Ilpars Emre
• Jordao Leal Cátia Daniela
• Simoes Cadete Mylene
• Ballon Aurélie 5 (6)
• García Cases Ananai 10
• Jiyoon Kang
• Yeoseung Chot
• Collet Felix
• Lozano Rios Blanka 10
• Betbeder Marie-Hélène 5 (6)
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• 22.2. Microbial growth curve, thermal inactivation, primary and secondary products Pazlarová
• 1.3.Factors influencing growth of bacteria Demnerová
• 8.3. Survey of food-borne pathogens– part 1
• 15.3. Survey of food-borne pathogens – part 2
• 22.3. Classic methods of detection and quantification – Purkrtová
• 29.3. Rapid methods of detection and quantification + EU legislative– Purkrtová
• 5.4. Microbial fermentative processes, biomass, enzymes, etc. – Pazlarová
• 12.4. Micromycetes,mycotoxines – Demnerová
• 19.4. Lactic acid bacteria – probiotics Demnerová
• 26.4. Spoilage bacteria, biogenous amines - Pazlarová
• 3.5. Presentations
• 10.5. Presentations
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Microorganisms on the Earth
Some numbers for beginning……
• Bacteria 3,6 mil. years
• Eucaryota 2,1 mil. years
• 99% of microorganisms (bacteria) present on the planet we are not able to cultived
• In the human body there are together 1013 cells
• In the human body there are 1014 microorganisms
• 7-10 thousands species???
• Human MICROBIOM – „second human genom“
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Perception of risks from intake foreign compounds
The public Experts
Rezidua of pesticides
Contaminants of environments
Food aditives
Microbiological contamination
Natural toxins
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Perception of risks from intake foreign compounds
The public Experts
Rezidua of pesticides Microbial contamination
Contaminants of environments Natural toxins
Food aditives Contaminants of environments
Microbial contamination Rezidua of pesticides
Natural toxins Food aditives
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How we can define safe and healthy food?
• Eggs?????
• Vegetables????
• Meat and their products????
• Flour and their products????
• Milk and dairy products????
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Microorganisms important in food
industry
atractive undesirable
Natural microflora
Pure cultures used in food industry
Pathogenic microorganisms
Microorganisms having unwanted enzymic activity
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Which are sources of spoilage?
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FAT TOM
explains what allows foodborne pathogens to grow. You'll also learn some important food
safety terms.
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Conditions influencing the growth of microorganisms in food
• Food
• Acidity
• Time
• Temperature
• Oxygen
• Moisture
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The growth of microorganisms in food
•water
•pH
•structure
•oxygen
•temperature
•time
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Food
Microorganisms need nutrients to survive. Food provides these nutrients in the form of proteins and carbohydrates.
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Acidity x alkalinity
• Bacteria grow best in food that contains little or no acid, measured as pH.
• On the pH scale of 0 to 14, a value of 0 is highly acidic, while a value of 14 is not acidic (alkaline) and a value of 7.0 is neutral.
• Bacteria grow best in food that is neutral to slightly acidic, represented by a pH of 6.5 -7.5
• Yeasts 4.5 – 5.5 • Molds a widerange of pH
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Time and environment
• Bacteria are able to double within 20 to 30 minutes.
• Food should be removed from "the danger zone" within two-four hours, either by cooling or heating. While most guidelines state two hours, a few indicate four hours is still safe.
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Temperature Food-borne pathogens grow best in temperatures between 5 to 60 °C, a range referred to as the temperature danger zone (TDZ). They thrive in temperatures that are between 21 to 40 °C.[
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Atmosphera/Oxygen
Almost all foodborne pathogens are aerobic, that is requiring oxygen to grow. Some pathogens, such as Clostridium botulinum, the source of botulism, are anaerobic
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Moisture - water activity
• Water is for growth and reproduction of microorganisms a limiting factor.
• Moisture is measured in %
• Water activity (aw) is measured on a scale of 0 to 1.0. Foodborne pathogens grow best in foods that have aw between 0.95 and 1.0. FDA regulations for canned foods require aw of 0.85 or below
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Food safety
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Commission Regulation (EC) No 2073/2005 on microbiological criteria for foodstuffs
• Foodstuffs should not contain micro-organisms or their toxins or metabolites in quantities that present an unacceptable risk for human health
• ‘micro-organisms’ means bacteria, viruses, yeasts, moulds, algae, parasitic protozoa, microscopic parasitic helminths, and their toxins and metabolites
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Microbial criteria for foodstuffs • Food safety criteria
• Process hygiene criteria
a) Meat and products thereof
b) Milk and dairy products
c) Egg products
d) Fishery products
e) Vegetables, fruits and products thereof
• Rules for sampling and preparation of test samples
a) General rules for sampling and preparation of test samples
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Food safety criteria Commission Regulation (EC) No 1441/2007 on
microbiological criteria for foodstuffs
• Listeria monocytogenes
• Salmonella
• Enterobacter sakazakii Cronobacter
• Escherichia coli
• Biogenic amines
• Staphylococcal enterotoxins
• Bacillus cereus
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Process hygiene criteria Meat and products thereof: Aerobic colony
count, Enterobacteriaceae, Salmonella, Escherichia coli
Milk and dairy products: Enterobacteriaceae, Coagulase-positive staphylococci, E.coli, Bacillus cereus
Egg products: Enterobacteriaceae
Fishery products: E.coli, Coagulase-positive staphylococci
Vegetables, fruits and products thereof: E.coli
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Staphylococcus aureus
Elektronový mikroskop Světelný mikroskop (po obarvení dle Grama) Typické kolonie na selektivním médiu
• growth in range of 7 – 48 °C • growth in range of pH 4 - 10 • limit concentration of NaCl 15 % • limit value of aw 0,85 (production of toxinu aw 0,98)
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Escherichia coli
Elektronový mikroskop Světelný mikroskop (po obarvení dle Grama) Typické kolonie na selektivním médiu TBX
• Growth in range of 10 – 47 °C • Growth in range of pH 3,8 - 9,5 • limit concentration of NaCl
8,5% • limit value of aw 0,93
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Bacillus cereus
Electron microskop Light microskop y Typické kolonie na selektivním médiu
• growth in range of 7 – 50 °C • growth in range of pH 4,3 - 9,3 • limit concentration of NaCl 7,5% • limit value of aw 0,95 • sporforming bacteria
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Salmonella
Elektronový mikroskop Světelný mikroskop (po obarvení dle Grama) Typické kolonie na selektivním médiu
• growth in range of 5 – 47 °C • growth in range of pH 3,8 - 9,5 • limit concentration of NaCl 9 % • limit value of aw 0,92
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Enterobacter sakazakii / Cronobacter spp.
Elektronový mikroskop Světelný mikroskop (po obarvení dle Grama) Typické kolonie na selektivním médiu
• growth in range of 5 – 45 °C • growth in range of pH 3,8 - 9,5 • limit value of aw 0,93
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Listeria monocytogenes
Elektronový mikroskop Světelný mikroskop (po obarvení dle Grama) Typické kolonie na selektivním médiu
• growth in range of 7 – 48 °C • growth in range of pH 4 - 10 • limit concentration of NaCl 15 % • limit value of aw 0,85 (production of toxinu aw 0,98)
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Campylobacte jejuni/coli
Elektronový mikroskop Světelný mikroskop (po obarvení dle Grama) Typické kolonie na selektivním médiu
• growth in range of 32- 45°C • growth in range of pH 4.9 - 9 • limit concentration of NaCl 1.5 % • limit value of aw 0.98 (microaerophilic)
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What about molds??? • They spoiling food and producing mycotoxins
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Aspergillus brassiliensis
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Trends of kampylobacteriozis in ČR
0
5000
10000
15000
20000
25000
30000
35000
2004 2005 2006 2007 2008 2009 2010 2011 2012 2013 2014*
poče
t hlá
šených
pří
pad
ů
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Trends of salmonelozis in ČR
0
5000
10000
15000
20000
25000
30000
35000
2004 2005 2006 2007 2008 2009 2010 2011 2012 2013 2014*
poče
t hlá
šených
pří
pad
ů
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Occurence of salmonellas in plant products – epidemiologic cases in EU
Year Country Vehikulum Serotype Num. of cases
2000 Netherland (NL) sprounts of mungo Enteritidis 27
2002 Germany anise tea Agona 42
2006 Danmark (D) basil pesto Anatum >200
2006 Norway (N) wild rocket Thompson 21
2007 D, UK, NL basil pesto Senftenberg 51
2007 N, D, Finland sprouts of alfaalfa Weltverden 45
2008 Finland iceberg lettuce Newport/Reading 107
2009 Finland sprouts of alfaalfa Bovismorbificans 42
2010 UK sprouts of alfaalfa Bareilly 213
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Occurence of salmonellas in plant products – epidemiologic cases in USA
Year Number of
states Vehikulum Serotype Number of cases
2007 44 peanat butter Tennessee
2008 43 tomatoes Saintpaul 1407
2009 14 alfa alfa sprouts Saintpaul 235
2009 ? White pepper Rissen 87
2010 26 alfa alfa sprouts 4,5,12:i.- 140
2010 11 alfa alfa sprouts Newport 44
2010 44 pepper Montevideo 272
2011 25 papaya Agona 106
2011 5 alfa alfa sprouts Enteritidis 25
2011 10 melon Panama 20
2011 5 pine nuts Enteritidis 43
2012 20 peanat butter Bredeney 42
2012 15 mango Braenderup 127
2012 24 melon Typhimurium 261
2012 ? germinating seeds Cubana ?
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Trends of listeriózis in ČR
0
10
20
30
40
50
60
70
80
90
2004 2005 2006 2007 2008 2009 2010 2011 2012 2013 2014*
poče
t hlá
šených
pří
pad
ů
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Trends of virus hepatitidis in CR
0
200
400
600
800
1000
1200
1400
1600
1800
2004 2005 2006 2007 2008 2009 2010 2011 2012 2013 2014*
poče
t hlá
šených
pří
pad
ů
VHA VHE
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How we can fight with microorganisms???
• HACCP
• Improvement of hygiene and sanitation
The need of modern, fast methods:
• chromogenic media
• molecular biology methods
• imunochemical methods
• mass spectrometry, MS
• nanobiosensors
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HACCP Hazard Analysis and
Critical Control Point
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Nowadays BIG problems
*Resistence of bacteria to antimicrobial compounds *Biofilm formation
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Louis Pasteur (1822-1895)
„Sirs, they are microbes, whose have the last word“
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Thank you for your attention