Listeria, Salmonella Flashcards
Listeria monocytogenes
- resistance
- multiplies in soil and water
- multiplies up to 10% NaCl concentration
- multiplies til pH 4.4 (more sensitive to organic acids)
- heat resistance: 0-42C (multiplication); 60C few minutes (destruction)
Listeria monocytogenes
- which media to use
Fraser broth, ALOA agar, Oxford agar, Palcam agar
Listeria monocytogenes
- Fraser broth
- liquid medium
- Inhibitory components: LiCl, nalidixic acid, akriflavine-hidrochloride
- reaction: olive green, black complex
Listeria monocytogenes
- ALOA
- Solid media
- ALOA: Agar Listeria Ottavani and Agosti
- Inhibitors: LiCl, nalidixic acid, ceftazimide, cycloheximide, plymyxin B, L-a-phosphatodyl-inositol
- reaction: Listeria spp produces blue-green colonies and pathogenic species (L. monocytogenes and L. ivanovii) produce similar color colonies with opaque halo
Listeria monocytogenes
- oxford-agar
- solid media
- inhibitors: LiCl, nalidixic acid, akriflavine-hidrochoride, aesculin, ammonium-iron(III)-citrate
- reaction: brownish green colonies with black halo
Listeria monocytogenes
- Palcam agar
- solid media
- Palcam: Polymyxin-B, Akriflavine, Lithium-chloride, Na-cetazimid, Aesculin, Mannit
- Inhibitors: Plymyxin-B, Acriflavin, LiCl, Ceftazimide sodium
- Reaction: greyish green colonies with black halo
Listeria monocytogenes
- conformation test to use
- Catalase test, Motility test, Haemolysis test, CAMP-test, acid production from carbohydrates
Listeria monocytogenes
- Catalase test
Listeria monocytogenes is catalase positive (gas formation/bubbles are seen)
- Catalase is an enzyme that mediates the breakdown of hydrogen peroxide (H2O2) into oxygen (O2) and water (H2O)
Listeria monocytogenes
- motility test
- inoculation into a triptone-soy-yeast broth, 25*C for 8-24hr –> typical somersaulting/tumbling motility under microscope
- deep inoculation into soft agar, 25*C, 8-24hr –> growth in an umbrella form
Listeria monocytogenes
- Haemolysis test
- method: hemolysis on sheep-blood agar, 37*C, 24hr
- L. monocytogenes: smooth, bright, narrow b-hemolysis
- L. ivanovii: wide, bright b-hemolysis
- L. innocua: no hemolysis
- L. seeligeri: weak b-hemolysis
Listeria monocytogenes
- CAMP test
- CAMP (Christine, atkins, munch, peterson)
- L. monocytogenes strengthens the hemolysis of Staph. aureus –> strong, arrow-shaped hemolysis
- L. ivanovii does the same but with Rhodococcus equi
Listeria monocytogenes
- acid production from carbohydrates
- L. monocytogenes can produce acid from rhamnose (positive reaction) –> pH decrease and the color turns yellow
- cannot produce acid from xylose (negative reaction) –> stays purple
Salmonella enterica
- characteristics
- multiplying: 6-47C, optimum: 35-37C (body temp)
- pH resistance: 3,8-9,5. Optimum pH: 7-7,5
Salmonella enterica
- resistance
- freezing
- at low water activity
- 7D values: 60C 2-6min. 70C < 1min. Low water activity - extremely high resistance: 7D value at 70*C: 1050 min
- 0,1% acetic acid (pH 5.1) can prevent growth
Salmonella enterica
- which media to use for detection
- RVS (Rappaport-Vassiliadis broth), MKTTn, XLD agarm BP agar, BPLS agar, MSRV agar
Salmonella enterica
- RVS: Rappaport-Vassiliadis broth
- Composition: soy peptone, malachite green, magnesium chloride, NaCl, potassium dihydrogen phosphate, pH 5,2+-0.2
- Blue color when negative
- light blue/transparent when positive
Salmonella enterica
- MKTTn
- MKTTn (Muller-Kauffman, tetrathionate, novobiocin)
- composition: Na-tiosulphate, cattle bile, brilliant green, lugol solution, CaCO3, Novobiocin, pH: 8.2 +- 0.2
- Light green color
Salmonella enterica
- XLD agar
- XLD (Xylose, lysine, desoxycholate agar)
- composition: xylose, lactose, sucrose (do not form acid from any of them), L-lysine hydrochloride, sodium thiosulphate, ferric ammonium citrate, sodium desoxycholate
- original color of the medium is red
- decompose xylose, lactose, sucrose to acid –> yellow color (NOT salmonella)
- H2S production –> ferrous sulphide –> black color (salmonella)
- Lysine –> cadaverine (decarboxylation) –> form purple color/ring around colonies
- postive: salmonella gives reddish purple colonies (like the medium), sometimes with black center and sometimes a purple ring
Salmonella enterica
- MSRV agar
- MSRV agar: Modified Semi-solid Rappaport-Vassiliadis agar
- composition: soy peptone, malachite green, MgCl, NaCl, potassium dihydrogen phosphate, pH = 5.2 +- 0.2
- detection is based on ability of Salmonella to move: a growing opalescent zone in the semi liquid media
Salmonella enterica
- BPL agar
- BPL (Brillian green, phenol red, lactose)
- composition: Lactose, protein, NaCl, phenol red, brilliant green, pH= 6.5 +- 0.2
- original color of medium is red
- small, purple colonies are formed on the agar because the pH does not increase
Salmonella enterica
- BPLS agar
- BPLS (brilliant green, phenol red, lactose, sucrose agar)
- composition: same as BPL + sucrose
- purple red colonies are formed
Salmonella enterica
- confirmation methods
- Biochemical conformation: TSI agar, Urea medium, tryptone broth, lysine broth, buffered glucose broth, Voges-Proskauer test, Methyl red test, ONPG test
- Serological examination
Salmonella enterica
- TSI slang agar
TSI (Triple-sugar-iron agar)
Composition: lactose, sucrose, glucose, iron citrate, sodium thiosulphate
Reaction:
- the original color of medium is orange
- fermentation of glucose: yellow color at the bottom of the tube: acid production + (with or without gas) –> glucoe positive
- fermentation of lactose: - (but! Salmonella enterica subzp. arizonae +/-) –> TSI top: red –> lactose and sugrose negative
- H2S production (from Na-thiosulphate): + black color (black spots in the sample)
Salmonella enterica
- decomposition of urea (urease test)
- Salmonella is urea negative, cannot produce ammonia from urea, the sample will not turn purple
- original color of the medium is orange/yellow
- Positive reaction: turns pink then purple (e.g Proteus)
- negative reaction: stays yellow (salmonella)
