Modified McBride Agar Research Articles

Occurrence of Listeria monocytogenes in food in Chile

Out of 2145 food samples analysed 77 were found contaminated with Listeria monocytogenes in Santiago, Chile. Samples were: 603 ice-cream (3.5% contaminated), 256 soft cheese (0.8%), 155 hard cheese (0%), 229 baby milk bottles (0%), 634 processed meat products (3.6%) and 268 crustaceous shellfish (11.6%). Three different isolation media were used: for 318 samples, Modified McBride Agar (MMA), Lithium chloride Phenylethanol Moxalactam agar, and Polymyxin Acriflavine Lithium chloride Ceftazidime Aesculin Mannitol agar; for 1827 samples MMA was replaced by Listeria Selective Agar Oxford Formulation. Isolates were classified as follow: serovar 1/2a (25 isolates), serovar 4b (20), serovar 1/2b (19), serovar 3b (7), serovar 1/2c (2), untypable (4). A high variety of phagovars was detected although 52% of strains was untypable.

RECOVERY LIMITS OF HEAT‐INJURED LISTERIA MONOCYTOGENES FROM ENRICHMENT BROTH AND ENRICHED CULTURES OF INOCULATED FOODS

ABSTRACT Recovery of heat‐injured Listeria monocytogenes strain LM82 was evaluated quantitatively in Listeria enrichment broth (LEB) and in enriched cultures of cooked shrimp and Brie cheese. LM82 cells [108 colony forming units (CFU)/ml] were heated for 60 min at 52C in phosphate‐buffered saline. After 24 and 48 h enrichment, injured LM82 (6 replicates at each of 5 inoculation levels) were isolated on 3 selective media: lithium chloride‐phenylethanol‐moxalactam agar (LPMA), modified McBride agar (MMA) and Oxford agar (OXA). The recovery limit was expressed as a 50% end point value (RL50), which is the calculated inoculation value necessary to recover LM82 on half of the replicates of each type of isolation agar plate after streaking from the enrichment of measured inoculum. The RL50 values for injured cells were comparable to those of uninjured cells after 48 h enrichment in LEB without food. The type of isolation agar did not affect the RL50 value, although with food, MMA gave consistently but not significantly higher values, i.e., recovery inferior to that of LPMA and OXA. RL50 values were higher in Brie and cooked shrimp, presumably because of the competitive microflora in those foods. Addition of lactose or pyruvate to LEB improved recovery but had little or no effect when foods were present.

Occurrence of Listeria species in raw milk and dairy products produced in Northern Ireland.

The overall incidence of Listeria spp. in raw milk samples surveyed was found to be 25.0% (Listeria monocytogenes 15.3%), with the incidence in samples from processing centres 54.0% (L. monocytogenes 33.3%); this was higher than that in samples from dairy farms (Listeria spp. 8.8%; L. monocytogenes 5.3%). The FDA enrichment procedure was much more productive than cold enrichment and Oxford agar was superior to modified McBride agar for isolation of Listeria. Listeria monocytogenes was never isolated by direct plating of raw milk samples on Oxford agar at a detection level of 1.0 cfu/ml. Listeria spp. were isolated from 1 of 95 pasteurized milk samples (L. monocytogenes) and 1 of 33 soft cheese samples (L. seeligeri). Restriction fragment length polymorphism was more useful than sero- or phage-typing for typing of L. monocytogenes strains, and results suggest that specific L. monocytogenes strains may persist in both farm and processing environments.

Evaluation of FDA and USDA procedures for enumerating Listeria monocytogenes in ground beef

The suitability of FDA and USDA-FSIS methods for isolating Listeria monocytogenes from meat samples was evaluated and examined. The enrichment broths tested were Fraser broth (FB), Listeria enrichment broth (LEB), University of Vermont (UVM) broth, and UVM with extra acriflavine (UVMA). The agar media used included modified McBride agar (MMA), LiCl-phenylethanol-moxalactam (LPM) agar, modified Oxford (MOX) agar, and McBride agar (MA) as a control. The performance of 16 combinations of enrichment broth and plating agar media was examined to recover three strains of L. monocytogenes from pure culture suspension and a slurry of inoculated fresh ground beef (10 2–10 4 cfu ml −1). All media performed well; however, FDA-LEB combined with USDA-MOX agar was the most efficient for the detection of L. monocytogenes strains.

Initial Cell Concentration and Selective Media Effects on the Isolation of Listeria monocytogenes from Enrichment Cultures of Inoculated Foods

To facilitate rapid quantitation of Listeria monocytogenes in artificially inoculated dairy products, seafoods, and other foods, the recently revised U.S. Food and Drug Administration methodology for Listeria in foods was further abbreviated. The ability to isolate L. monocytogenes was measured in terms of the number of inoculated colony forming units (cfu)/g of food. Isolation values for the foods tested, using modified McBride's agar (MMA), were <1, 1–10, and >11 inoculated cfu/g in 30, 44, and 26% of test samples, respectively. The corresponding percentages with lithium chloride-phenylethanol-moxalactam (LPM) agar were 41, 51, and 8%, respectively. In eight (21%) instances, isolation values on LPM agar were significantly superior to those on MMA. The superiority of LPM was most dramatic when MMA isolation values were >10 initial cfu/g. However, LPM was not an infallible remedy for MMA deficiencies even at >10 cfu/g. Ease of isolation was apparently not related to food type. For 10 different test portions of Brie cheese, isolation values ranged from 0.3 to > 10,000 of inoculated cfu/g with MMA and 0.3 to 2500 of inoculated cfu/g with LPM agar.

THE EFFECT OF AEROBIC MESOPHILIC MICROFLORAL LEVELS ON THE ISOLATION OF INOCULATED LISTERIA MONOCYTOGENES STRAIN LM82 FROM SELECTED FOODS

ABSTRACT The effect of aerobic mesophilic microfloral concentration on the isolation of Listeria monocytogenes LM82 was studied in 31 (18 cheeses and 7 noncheese) retail foods having standard plate counts of 101 to 108 colony forming units (CFU)/g. Foods were spiked with L. monocytogenes and enriched at 30°C for 24 h in a selective enrichment broth used in a U.S. Food and Drug Administration method. Inoculum levels for isolation on modified McBride agar ranged from 0.1 to &gt; 5 × 103 with a geometric mean value of 5 inoculated CFU/g or 1.4 CFU/g. Pure Enterococcus (Streptococcus) faecalis (0 to 6 × 106 inoculated CFU/mL) in the absence of food matrix had no effect on the enrichment of L. monocytogenes. Ease of isolation of LM82 was independent of the food microflora concentration both generally and in the specific food type of 9 Brie cheeses. Competition, when it occurs, therefore, may be due to specific bacterial competitors rather than bacterial numbers.

A technique for the direct identification of haemolytic-pathogenic listeria on selective plating media

A technique based on the addition of a red cells top layer to a selective plating medium after listeria growth is proposed in order to detect directly the haemolytic activity of pathogenic listeria colonies. It was applied to different selective plating media (modified McBride agar, lithium chloride-phenylethanol-moxalactam, listeria selective medium–Oxford formulation, polymyxin-acriflavine-lithium chloride-ceftazidime-aesculin-mannitol and LSAMM). The haemolytic activity of listeria colonies was more easily detected with the top layer than when red cells were incorporated in the selective plating medium. The LSAMM was the best medium for the recovery and identification of Listeria monocytogenes colonies by this technique (three Listeria monocytogenes colonies were distinguished among 2520 Listeria innocua colonies in raw milk).

Comparison of Four Procedures to Detect Listeria spp. in Foods

A comparison was made of four procedures to detect Listeria spp. in two food categories. The study comprised 309 assays, 71 on milk from both infected and uninfected cows, and 238 on ten types of fresh vegetables. A sample was considered positive if it could be detected by at least a single method and if isolates could be confirmed as Listeria spp. The procedures detected 98–100% of the positive milk samples. Recovery from vegetable samples ranged from 45 to 86%, probably because of low levels of Listeria spp. in the presence of mixed flora. The ELISA procedure of the Organon Teknika® corporation detected 68% of the 44 positive vegetable samples; the GENE-TRAK®DNA probe, 45%; the U.S. Food and Drug Administration (FDA) culture procedure, 75%; and the FDA probe procedure, 86%. Recovery was higher with LiCl-phenylethanol-moxalactam agar (FDA probe procedure) than with modified McBride Agar (FDA culture procedure).

The rapid demonstration and presumptive identification of Listeria monocytogenes in food using monoclonal antibodies in a direct immunofluorescence test (DIFT)

Two monoclonal antibodies conjugated to fluorescein isothiocyanate (FITC) were successfully used in a direct immunofluorescence test (DIFT) to demonstrate listeria in seven samples of soft cheese where Listeria monocytogenes had been cultured by conventional techniques. Using DIFT, listeria was not detected in 20 cheese samples from which L. monocytogenes had not been isolated, or was present in low numbers (< 102/g). The DIFT was also used to presumptively identify > 90% of strains of L. monocytogenes isolated from food and cultured on Modified McBride agar or Blood agar. Less than 10% of strains of other species of listeria would be misidentified when grown on these media. All tests were carried out within 2 h.

EXTINCTION OF LISTERIA MONOCYTOGENES IN SINGLE‐STRENGTH ORANGE JUICE: COMPARISON OF METHODS FOR DETECTION IN MIXED POPULATIONS1

ABSTRACTThe FDA method of selective enrichment followed by selective plating on modified McBride agar was capable of detecting the presence of L. monocytogenes inoculated into a typical, commerical, reconstituted, single‐strength orange juice at the 100 cfu/mL level. The KOH shock treatment, formerly recommended in the FDA protocol, negatively affected recovery of Listeria at 100 and 101 levels in juice which also had a low background microflora (102 cfu/mL total count); however, KOH treatment was required for Listeria detection in juice which had high background microflora (108 cfu/mL). Two other protocols for detection of Listeria which utilized cold enrichment or different selective enrichment media were less effective than the FDA procedure due to heavy growth of background microflora. No Listeria was detected in 100 commerical orange juice samples from dairy and nondairy processors in geographically distinct areas of North America using the FDA method.

A new selective medium for isolating Listeria spp. from heavily contaminated material.

Food-associated outbreaks of human listeriosis have emphasized the importance and necessity of screening food for the presence of Listeria isolates-selective agar medium combining acriflavine (10 mg/liter) with ceftazidime (50 mg/liter) was developed. A total of 1,099 cheese production specimens were cultured, from which 157 Listeria isolates. (14.3%) grew. When compared with modified McBride agar, the acriflavine-ceftazidime agar recovered more Listeria isolates (98 versus 65%, P less than 0.001) more rapidly (57% after 48 h of incubation of the enrichment broth versus 35%, P less than 0.01) and in greater amounts. Acriflavine-ceftazidime selective agar medium proved to be a highly sensitive medium to recover Listeria spp. from heavily contaminated food products.

Improved Listeria monocytogenes selective agar.

By increasing the LiCl concentration to 5 g/liter and adding 20 mg of moxalactam per liter to modified McBride agar base, it was possible to inhibit the growth of many bacteria which interfered with the recovery of Listeria monocytogenes from beef.