Procedures For Detection And Enumeration

The selection of preenrichment, enrichment, and/or direct plating media, as well as conditions for incubation and procedures for confirmation of isolates will differ, depending on the microorganism or group of microorganisms targeted for detection or enumeration. Media, incubation conditions, and confirmation techniques selected for each microorganism or group of microorganisms that may be inoculated onto or naturally present in produce should be the same across laboratories. Optimum protocols for retrieving pathogens and nonpathogens from fruits and vegetables may differ, depending upon whether analysis of the surface, tissue, or a composite of both is desirable. Washing, rubbing, blending, homogenizing, stomaching, macerating, and grinding, or a combination of one or more of these procedures, are among the choices to process samples for preenrichment, enrichment, or direct plating. One piece of fruit or vegetable, several pieces, or only a portion of the whole or cut produce may be selected for analysis, but the procedure needs to be standardized in terms of sample weight and/or excision technique. The composition and pH of the diluent and ratio of diluent to sample need to be consistent, at least within each type of fruit and vegetable. The time and temperature for processing samples for preenrichment, enrichment, or direct plating should be standardized. The likelihood of stressed or injured microbial cells being present on or in fruits and vegetables should be recognized, and appropriate resuscitation conditions should be considered and applied. Repair of cells on the surface of produce that, for example, may be debilitated by desiccation or as a result of exposure to a harsh chemical environment, is important if these cells are to be detected or enumerated. Adjustment of the pH of homogenates of highly acidic fruits and vegetables may be necessary to protect microorganisms against exposure to potentially lethal conditions during preparation of samples for inoculation of recovery media.

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