Chapter 8 Flashcards
Mention and explain methods of surface sampling
- Surface slides
- thin slices, scalpels and forceps - Rinse and wash
- sterile diluents (ten part by weight), washings initial 10^-1 dilution) - Swab
quantitative results, area to be examined defined by previously sterilized template - Impression method
- transfer mo directly - Adhesive tape
- investigation of microflora on human skin/surfaces
- tapes and labels self sterilizing - Agar sausage
- agar is cut, exposed part is pressed and removed, impressed, incubate
Impression techniques and adhesive tape transfer do not allow dilution series (small microbes load)
impression tech, adhesive tape, contact slides, swabs = non destructive
How to transport and store frozen samples?
- solid co2
- insulated containers
- deep freezer
How to transport and store unfrozen samples?
1, shouldn’t be frozen
2. if unavoidable, chilled and kept @ 4 degree c
qualitative analysis:
biochemical reaction, enzymatic reaction, redox potential reaction. ex: proteolytic activity, carbs, lipids
proteolytic activity:
hydrolysis of gelatin, casein, coagulated serum, protein2, deamination, nitrate reduction
Test for presence of active enzymes:
catalase, oxidase, coagulase test
Reactions involving carbs & other compounds
starch, sugar, litmus milk
Reactions involving lipids and phospolipids
hydrolysis of tributyrin, butter fat, tween, lecithin
Quantitative analysis:
- Direct microscopic count
- no incubation, simplest method
- rapid, stained n read later, ez equipment, morphological and/or gram strain identified
disadvantages:
- harus banyak mo, small qty ga precise, debris, analyst fatigue - Colony count method:
- plating tech: pour plate & surface/spread plate
Semi-quantitative method
Most probable number
DMC Standardization of equipment for food films:
calibrated by the breed DMC< MF 300,000-600,000
Explain method of DMC step by step
- loop
- draw loop sample vertically, transfer n spread 0.01 ml portion over 1 cm^2 area, prepared films should be dried without delay over the 1 cm area 40-45 degree c, dried within 5 mins, place a drop of immersion oil
- count separate fields, start midway at the top/bottom. kalo banyak, repeat 2 ,m away dri awal
What’s the function of counting chamber?
enumerate bacteria, yeast, and mold spores or hyphae fragments. = hymacytometer
cb sbutkan rumus2 DMC ceunah
THOMA, petroff hauser, neubauer
ga premium sih gbs paste gambar…
CCM
- TPC
aerobic mesophilic count: general viable count, indication of standard of hygiene/plant sanitation, potential health hazard - MYC
mould and yeast counts -> similar problems to bacterial counts (unicellular), difficult to interpret mycelia, depends on the number of colonies would depend on degree of homogenization and extent of consequent fragmentation of hyphae
CCM method gmn kak
- liquid: pipet
- fine particulate solid: blending + sterile diluents,
inoculation: 15-30 mins
CCM choice of diluents
a) general purpose:
ISO standard methods: 0.1% peptone plus 0.85% sodium chloride
b) diluents for anaerobic
- low redox
- very oxygen sensitive, hungate/anaerobic
c) diluents for osmophiles and halophiles:
- osmophilic: sterile 20% sucrose
- halophilic: sterile up to 15% nacl
Incubation temperatures
0-10: psychothrophs and psychrophiles
20-32: saprophytic mesophiles
35-37 or 45: parasitics and commensal of homoiothermic animals
55-63: higher count of thermophile
many marine bacteria are able to grow if distilled water in the medium is replaced by sea water
bacterial in the presence of moulds n yeast may be determined by using media containing the antifungal…
antibiotic cycloheximide to 10 ppm
food sample is analysed by TMC and viable count, high total count in conjunction with a low viable count…
doesn’t indicate the majority is dead, cm unable to multiply in particular incubation environment
Rumus pour plate method
BAM: 25-250
ISO: 10-300
TPC: 10-150 (yeast + mold)
Surface plate method: spreading spatula and spiral plate
gampang dibedain soalnya morphology is easily observed
Surface plate method
miles and misra surface colony count: placing drops of serial dilutions on the surface of poured agar plates and counting the colonies that develop on incubation of the plates. best: opaque medium and surface culture
Surface plate method: spiral plater
diputer2 liquid across the surface of an agar plate. counts colonies on a part of the track
Dry rehydratable films
used in surface plate method: petrifilm, small amount of space taken up by the films
Most probable number
estimating the density of viable organism in sample
general consideration in most probable number
low level of bacteria: enumeration using liquid media (MPN) to improve statistical reliability of the results.
MPN can be used for
enumeration: enterbocateriaceae coliform, e. coli
isolation/or numeration pf staphyloccoci, streptococci, vibrio parahaemolytocus, salmonella
MPN: detection of positive tubes
- turbidity
- metabolic end products (gas production, acid/base, reduction, other).
Determination of MPN values:
- calculation using mathematical formulae
- consultation of MPN tables
- utilization of specific computer programs
