Isolation was performed according to the ISO 11290–1:2017 method with two-stage enrichment. First, 25 g of each food sample was inoculated in 225 mL half Fraser broth (Basingstoke, Oxoid, UK) for initial selective enrichment and homogenized in a stomacher (Seward 400, Radnor, PA, USA). Incubation followed at 30 ± 1 °C for 25 ± 1 h and 0.1 mL of the broth culture was inoculated in 10 mL of full-strength Fraser broth for the second enrichment, which was cultured at 37 °C for 24 ± 2 h. A loopful (1 mL) of each of the half- and full-strength Fraser broths were plated on the chromogenic Listeria agar Ottaviani and Agosti (ALOA agar) (Merck, Darmstadt, Germany). The plates were incubated at 37 °C for 24 to 48 h. Five typical colonies from each ALOA agar were restreaked on tryptic soy agar supplemented with 0.6% yeast extract (TSA-YE) (Sigma, Darmstadt, Germany) as a nonselective medium and incubated at 37 °C for 24 to 48 h. The colonies from TSA-YE were verified by Gram staining, catalase reactions, oxidase tests, carbohydrate utilization, CAMP tests, and motility at 20 to 25 °C. Listeria monocytogenes was enumerated in the samples according to the methodology described in the ISO 11,290–2:2017 Microbiology of the food chain standard—Horizontal method for the detection and enumeration of L. monocytogenes and Listeria spp.—Part 2: Enumeration method [17 ,18 ].
Aloa agar
Manufactured by Merck Group
Sourced in Germany
ALOA agar is a microbiological culture medium primarily used for the detection and enumeration of Listeria monocytogenes in food samples. It provides a selective and differential environment for the growth of this pathogenic bacterium.
Automatically generated - may contain errors
3 protocols using aloa agar
1
Isolation and Enumeration of Listeria monocytogenes
2
Listeria monocytogenes Detection Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
One portion of sponge from each environmental sample was cultured in 200 ml of half Fraser broth (Merck, Germany) for 24 h at 30 °C in accordance to ISO 11290-1 (Anonymous, 2017b) . After the pre-enrichment step, 0.1 ml was put onto 10 ml Fraser broth (Merck, Germany) for 48 h at 37 °C. Ten microliters were plated into ALOA agar (Merck, Germany), another 10 μl were plated into Oxford Agar Base (Scharlau Chemie, Spain) and incubated during 24-48 h at 37 °C. The presumptive colonies were identified by real-time PCR with the commercial kit PATHfinder Listeria monocytogenes/IPC Detection Assay (Generon, Italy).
3
Listeria spp. Detection in Food Samples
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Twenty-five mL of each sample were homogenized in 225 mL of listeria enrichment broth (LEB, from Oxoid ® ) and incubated at 30°C for 4 hours. Subsequently, 0.5% (1.8mL) nalidixic acid, 1% cycloheximide (1.15mL) and 0.5% acriflavine (0.455mL) were added as selective agents and the samples were incubated at 30°C for 48 hours. After 24 and 48 hours, the incubated samples were seeded onto ALOA agar (Sigma ® ) and PALCAM agar (Oxoid ® ) using a 10μl disposable inoculating loop, and incubated at 35ºC for up to 48 hours. After that, up to five characteristic colonies (blue, ALOA and black agar with black halos on PALCAM agar) were place in a tube with tryptone soya yeast extract agar (TSYEA) (TSA plus 0.6% yeast extract) and incubated at 35ºC for 24 hours. After this, preliminary identification tests were conducted, such as Gram staining (Gram positive rods), catalase proof (catalase positive), esculin hydrolysis or agar motility in order to observe isolated "umbrella-like" growth patterns.
To evaluate the development of Listeria in selective medium using possibly contaminated samples, as well as the behavior of the bacterium at different culture temperatures, all the samples of one of the samplings were cultured in duplicate. After culturing in LEB and adding selective agents, these samples were also cultured on PALCAM and ALOA agars and incubated at 10°C for 15 days.
To evaluate the development of Listeria in selective medium using possibly contaminated samples, as well as the behavior of the bacterium at different culture temperatures, all the samples of one of the samplings were cultured in duplicate. After culturing in LEB and adding selective agents, these samples were also cultured on PALCAM and ALOA agars and incubated at 10°C for 15 days.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!