Biological Methods of Preservation Flashcards
Define Biopreservation:
Give some examples:
Use of Microorganisms, their metabolic products, or both to preserve food (excluding fermentation)
- controlled acidification
- bacteriocins
- bacteriophages
What is controlled acidification?
acidification of food for preservation:
done by direct addition of organic acid, fermentation, or use LAB to produce in situ
What is MicroGard?
form of controlled acidification:
contains fermentable carb + customized culture (depending on product)
acts as safeguard: if food temperature abused -> MicroGard microbes will grow and make acid, prevent pathogens
What are the main benefits of MicroGard?
- protect shelf life/safety
- maintain organoleptic qualities
- “natural” product
What is the Winsconsin process, and what are the advantages?
add less nitrite, 0.7% sucrose, LAB: to prevent C. botulinum in bacon
less nitrite needed (less potential carcinogens)
What are bacteriocins?
Bacteriocins produced by ____ are of particular interest.
antimicrobial peptides produced by bacteria that are lethal to others, but not the producer (or humans)
LAB
What are the different categories of bacteriocins?
Class I, Class II, Class III, Class IV
What are the Class I bacteriocins and how are they made?
unusual AA; made by posttranslational modification
ex: dehydroalanine, dehydrobutyrine, lanthionine, methyllanthione
What is a specific subcategory of Class I bacteriocins? How is it produced?
Lantibiotics (containing lanthionine ring)
rxn of dehydro-amino acids with cysteine -> thioether lanthionine
What is the best characterized LAB bacteriocin? What category does it belong in, and how is it produced industrially?
NISIN class I bacteriocin
made by culturing L. lactis (not synthesized chemically) -> for cheese/meat/beverages
True/False: Nisin is produced by lactic acid bacteria
True
What is an example of a lantibiotic not made by LAB? What is it used for?
subtilin (made by Bacillus subtilis)
antibacterial action; also protease (food, laundry detergent, lens cleaners)
What are 2 examples of lantibiotics?
subtilin, nisin
What are class II bacteriocins?
small heat stable proteins
with consensus leader sequence that signals to cell to export protein
What are the subclasses of Class II bacteriocins?
IIa: active against L. monocytogenes
IIb: requires 2 different peptides for activity
IIc: requires reduced cysteine for activity
What are class III bacteriocins?
> 30kDa (large) heat-liable proteins
What are class IV bacteriocins?
with lipid or carb components (unknown function)
What method is used to discover new bacteriocins?
Place colony of bacteriocin producer onto agar inoculated with lawn of other bacterium -> incubate, observe for ZONE OF INHIBITION (circle of no growth)
True/False: a zone of inhibition indicates there is definitely bacteriocin production
False: could be other inhibitory substance (organic acid, H peroxide, phage contamination)
How can you differentiate between the potential causes of a zone of inhibition?
bacteriocin: sharp edges, colony in center
organic acid: fuzzy edges
Phage: no colony n center
Perform test: remove colony, apply protease -> if it was bacteriocin, then the bacteria should now be able to grow over
What is the general mechanism of all LAB bacteriocins?
disrupt integrity of cytoplasmic membrane
- bind and create pore (hole)
or - membrane solubilization
Bacteriocins are most effects against cells in what state?
vegetative
What is the effect of bacteriocins on vegetative cells?
disrupt membrane -> ions, AA, ATP will spill out -> ruins chemical/electrical gradient -> cannot generate energy, cannot protect from outer environment -> inhibition or death
What property of bacteriocins allows for its effects against the membrane?
amphiphilic cationic peptides -> can insert into membrane
True/False: Nisin will encourage germination of spores, making it risky to use as a bacteriocin
False: it DOES encourage germination, BUT: will attack pre-emergent spore (before starts growing)
(germinated spore is more vulnerable than dormant)
What are Colicins?
bacteriocins made by some strains of E. coli against other strains
What do E. coli strains need to be able to produce colicin?
colicinogenic plasmic
genetic info for synthesis, immunity, and release
Colicin expression is under the ____ regulatory system
SOS
Describe the mechanism of action for colicin:
binds to specific receptors (outer membrane proteins for nutrient uptake) -> enter, transported by Tol or TonB -> disrupt inner membrane or attack DNA, rRNA, or tRNA
True/False: colicin must enter the bacterial cell in order to be lethal
true (attacks inner mechanisms)
The 2 classes of colicins:
enzymatic
pore-forming
What are the lethal modes of action of enzymatic colicins?
- block synth of peptidoglycans
- cleave tRNA -> stop protein synth
- cleave 16S rRNA -> stop protein synth
- DNA degradation
What are the approved ways to add bacteriocins to food?
- add purified bacteriocin directly
- add producing culture (make in situ)
- choose bacteriocin making culture to include for fermentation
Give an example of a bacteriocin that is added to food:
What are its main purposes?
Nisin: for milk, cheese, dairy, canned food, smoked fish, mayo, baby food (GRAS)
anti-listerial agent; sensitize spores to heat (prevent botulism, lessen thermal treatment time needed)
Is Nisin used alone or in conjunction with other treatments?
Together - part of a multiple barrier inhibitory system
ex: with mod. atmosphere storage (fresh fish, ground pork)
What is a bacteriocin that is produced in situ in food? What does it protect against?
Pediocins (in wieners, dressings, cottage cheese, meat, RTE salad)
effective against vegetative L. monocytogenes (not good for spores)
The protective effect of pediocin is more effective with (higher/lower) temperatures, and better in (aerobic/anaerobic) conditions
lower
anaerobic
Why is a bacteriocin producing strain beneficial for the making of fermented foods? Give an example
helps inactivate existing bacteria -> allow for culture to grow, better chance of success
ex: adding pediocin producers to sausage -> reduces L. monocytogenes
how does bacteriocin producers affect shelf life in fermented foods? give an example:
Increase shelf life
ex: add nisin producing LAB to cheese -> shelf life increased from 14 to 87 days
True/False: unlike antibiotics, bacterial resistance is not a concern with bacteriocins.
False: resistant strains exist and can develop!
What are some developed resistance mechanisms to bacteriocins that have been observed? How does specificity vary?
- destruction of bacteriocin (specific or general)
- modification (specific)
- Altered receptors (specific)
- Membrane composition (general)
Can bacteriocin resistance be overcome by applying a combo of different bacteriocins? Why or why not?
Yes, if it is a specific resistance mechanism
No, if it is a general mechanism (need a different functioning bacteriocin)
Is cross-resistance with antibiotics an issue for bacteriocin resistance?
Usually no;
Multidrug resistant bacteria can still be sensitive to bacteriocin
What is a resistance mechanism observed in L. monocytogenes against nisin?
modified membrane composition - more straight chain FA (less fluidity) -> nisin can’t insert as easily
What is a specific defense mechanism against dehydroreductase bacteriocins?
Modification
What is important in the food industry to combat bacteriocin resistance?
Apply multiple hurdle technology (use many diff steps/methods to kill bacteria)
Bacteriocin resistant cells can be sensitive to other factors (heat, cold, etc)
True/False; we normally consume bacteriophages
True (part of natural food microbiota)
What are the main challenges with using phages for microbial control in food?
- lack of consumer acceptance
- possible resistance
- low numbers of bacteria in food (phage cannot effectively grow)
- targets specific species/strain
Phages need _____ actively growing bacteria to be effective
10^5 - 10^6
There are commercial phages used to control growth of ___, ___ and ____.
Salmonella, E. coli, Listeria
How is bacteriophage resistance created? How can we overcome this?
changes in bacteriophage receptor site (proteins, LPS, lipoproteins on surface)
use mix of different phages (if one is useless, other still are functional)
