توضیحات
The ALOA culture medium is used for the isolation and identification of Listeria monocytogenes in food and clinical samples. this culture media is prepared based on the Agosti & Ottavianu culture medium. This medium is used for confirmation after using the Fraser enrichment medium. This is also the recommended medium in ISO-11290-1 for the identification and enumeration of Listeria monocytogenes.
Using chromogenic substrate for the identification of Listeria causes all Listeria strains to be identified with blue color. Other organisms that possess this enzyme such as Enterococci can be inhibited by growth inhibitors present in the medium as well as antibiotics.
Listeria monocytogenes, unlike other strains of this bacterium, has phospholipase activity. Another differential activity is also demonstrated by the use of lipase C substrate. Lipase activity creates a cloudy halo around Listeria monocytogenes colonies.
Simultaneous use of these two substrates distinguishes Listeria monocytogenes colonies from other strains. Because although all strains appear blue, the monocytogenes strain is identified by a cloudy halo around the colony. Listeria ivanovii strain, which is often pathogenic for both animals and humans, also exhibits lipase activity.
STORAGE
- Store the powder at a temperature of 15-30 degrees Celsius and the prepared medium at a temperature of 2-8 degrees Celsius in a dark and dry place.
Test
Test
- Weigh 35.2 grams of the powder and dissolve it in 500 ml of distilled water. Mix well.
- Autoclave at 121°C for 15 minutes.
- Under sterile conditions, add an iChrome Listeria supplement vial to a sterile and cooled medium at a temperature of 50-45 degrees Celsius.
- Under sterile conditions, add 10 milligrams of nalidixic acid, 38350 units of polymyxin B, 10 milligrams of Ceftazidime, and 5 milligrams of Amphotericin B to the medium.
- Mix well and pour into sterile Petri dishes.
Identification and enumeration of Listeria monocytogenes and listeria strains according to ISO 11290.
Test
- Weigh 25 grams (25 milliliters) of the sample and add it to 225 milliliters of Fraser broth at a concentration of one-tenth.
- Mix well and incubate in 30 degree Celsius for 25 ± 1 hours.
- Add 1 ml of this culture medium to 10 ml of Fraser broth medium. Incubate for 24 ± 2 hours in 37 degree Celsius under aerobic condition.
- Inoculate the sample onto the primary enrichment medium, ALOA surface (lacking chromogenic substrates), and another selective medium for laboratory diagnosis to obtain separate colonies.
- Repeat the same process using the second enrichment medium and inoculate the surface of one ALOA plate (lacking chromogenic substrate) and another selective medium.
- Incubate inoculated plate for at least 48 ± 2 hours.
- Please select probable colonies and perform confirmatory tests for Listeria monocytogenes as well as other strains.
Enumeration method
- Prepare an initial suspension with a ratio of 1 from the sample an 10 from buffered peptone water. If you are performing identification and counting simultaneously, Fraser broth with a dilution of one-third can be used as a diluent.
- Inoculate 1 ml of suspension on the surface on ALOA chromogenic plate. Incubate for 24 + 2 hours at 37 degree Celsius. If no microbial growth is observed, continue incubating for another 24 hours.
- select probable colonies and perform confirmatory tests for Listeria monocytogenes as well as other strains.
- Calculate the number of Listeria monocytogenes or other strains from the confirmed colonies.
