Use of a novel medium, the Polymyxin Ceftazidime Oxford Medium, for isolation of Listeria monocytogenes from raw or non-pasteurized foods.

Polymyxin Ceftazidime Oxford Medium (PCOM), a novel selective and differential plating medium for Listeria monocytogenes was compared with Modified Oxford Agar (MOX) for efficacy to isolate L. monocytogenes and other Listeria spp. naturally present in non-pasteurized Mexican-style cheese (n = 50), non-pasteurized fresh squeezed orange juice (n = 50), raw beef chunks (n = 36), and fresh cabbage (n = 125). Samples were collected from retail markets and farms in Mexico and tested following the US Department of Agriculture enrichment technique. Listeria spp. were isolated from 23.4% of analyzed samples, and from those, 75.0% corresponded to raw beef chunks, 38.0% to non-pasteurized Mexican-style cheese, and 30.0% to fresh squeezed orange juice. No Listeria spp. were isolated from fresh cabbage samples. L. monocytogenes was recovered from 15.3% of food samples analyzed. Non-pasteurized Mexican-style cheese showed the highest proportion of L. monocytogenes positive samples (36.0%), followed by orange juice (26.0%) and raw beef (25.0%). The frequency of isolation of Listeria spp. and L. monocytogenes was not different (P > 0.05) between PCOM and MOX. The advantages of using PCOM when comparing to MOX, include the easier way to identify Listeria species, the lower cost per plate and the availability of its ingredients for Latin-American countries.

The Polymyxin Ceftazidime Oxford Medium as an alternative selective and differential medium for isolation of Listeria monocytogenes from raw or unpasteurized food.

The Polymyxin Ceftazidime Oxford Medium (PCOM) was developed to recover Listeria monocytogenes from raw or unpasteurized foods. It contains esculin-ferric ammonium citrate as indicator system for Listeria growth, and ceftazidime and polymyxin B as selective agents, which are available in several Latin American countries. Comparison of PCOM, Modified Oxford Medium (MOX) and Tryptic Soy agar with 0.6% yeast extract (TSAYE) indicated that both selective media were equally effective at recovering four individual strains of L. monocytogenes (Scott A, V7, California and broccoli), and a mixture of these strains (LMM) (P > 0.05). The ability of PCOM, MOX, TSAYE and TSAYE supplemented with 4% NaCl to recover heat, acid and freeze-damaged LMM was similar for all media (P > 0.05). The PCOM proved to be effective at isolating colonies of LMM from inoculated raw beef chunks, unpasteurized orange juice, cabbage, and Mexican-style cheese by direct plating and by the US Department of Agriculture's Food Safety and Inspection Service enrichment method. Differentiation of L. monocytogenes colonies was easier on PCOM than on MOX for foods with high levels of background microbiota. Based on the evaluations performed on foods naturally contaminated with L. monocytogenes, PCOM was a more economical alternative than MOX for selective and differential isolation of Listeria from raw or unpasteurized foods.

Effect of the use of a neutralizing step after antimicrobial application on microbial counts during challenge studies for orange disinfection.

The effects of using a neutralizer after applying antimicrobial treatments and the effect of time lapse between treatment application and subsequent recovery and enumeration of Escherichia coli O157:H7 and Salmonella were investigated in Valencia oranges. Inoculated oranges surfaces were washed with distilled water for 15 s and then sprayed with a solution containing 200 mg/liter sodium hypochlorite (pH 6.5) for 15 s; they were then dipped in L-lactic acid (2.0% at 55°C) for 1 min or in distilled water at 80°C for 1 min. Posttreatment, oranges were divided into two groups. In the first group, oranges were dipped in neutralization treatment: 270 ml of buffered peptone water for 2 min for lactic acid-treated oranges, 270 ml of Dey-Engley broth for 2 min for chlorine-treated oranges, or 3.7 liters of tap water (25°C) for 10 s for hot water-treated oranges. The second group of treated oranges was not subjected to any neutralizer. All oranges then were kept at room temperature (average 26.2°C) and sampled at 0, 7.5, and 15 min for enumeration of surviving Salmonella and E. coli O157:H7. The orange surface (30 cm(2)) was excised for pathogen enumeration. The presence of free chlorine and changes in pH and temperature on the orange surface were determined in uninoculated, treated oranges. Free chlorine was detected on oranges after treatment; the change in temperature of orange surfaces was greater during treatment with hot water than with lactic acid. Nevertheless, pathogen enumeration did not show any impact of neutralizer use on the residual activity of antimicrobials or any impact of the time elapsed between antimicrobial treatment and recovery of bacterial pathogens from inoculated oranges (P ≥ 0.05). The results of this study indicate that the lack of a neutralizing step before enumeration of pathogens is not likely to affect the accuracy of results during challenge studies to test pathogen reduction strategies on oranges.

Comparison of different washing treatments for reducing pathogens on orange surfaces and for preventing the transfer of bacterial pathogens to fresh-squeezed orange juice.

The objectives of this study were to compare the effectiveness of various washing treatments for reducing Escherichia coli O157:H7, Salmonella sp., and Listeria monocytogenes populations on orange surfaces and to measure the effect of some of these treatments in preventing the transfer of pathogens during juice extraction. Orange surfaces inoculated with L. monocytogenes or a mixture of E. coli O157:H7 and Salmonella Typhimurium were washed by water spray and then sprayed with or dipped in water at 80°C for 1 min, 70% ethanol for 15, 30, or 45 s or 1, 2, or 4 min, 2 or 4% lactic acid solution at 55°C for 15, 30, or 45 s or 1, 2, or 4 min, or 200 mg/liter hypochlorite at pH 6.5 or 10 for 15 s. The surviving populations of these pathogens on the oranges were enumerated after each treatment. In a further stage, the ability of these pathogens to be transferred to the juice during extraction was tested. Juice was obtained from inoculated oranges that were subjected to selected treatments using chlorine, lactic acid, ethanol, and hot water as described above, and then bacterial counts in orange juice were determined. The application of these treatments reduced the populations of pathogens on orange surfaces by 1.9 to >4.9 log, 1.9 to >4.6 log, and 1.4 to 3.1 log cycles for E. coli O157:H7, Salmonella Typhimurium, and L. monocytogenes, respectively. The treatments using hot water or lactic acid showed greater reductions than other treatments. The time, antimicrobial concentration, and form of application affected the bacterial reduction. All treatments resulted in undetectable counts in the juice. Nevertheless, pathogens were recovered by the enrichment-plating method. Treatment of oranges before juice extraction may reduce the risk associated with consuming orange juice.