Multiple plating from enrichment media as an aid to salmonella isolation

Abstract

SUMMARYFaeces samples enriched for salmonellas in selenite F broth incubated at 43°C were subcultured to brilliant green MacConkey agar after 6 h. Four platings were made. As a control technique, each sample was enriched in selenite F incubated at 37°C for 24 h and subcultured once to brilliant green MacConkey agar. Of the specimens plated at 6 h, out of a total of 25 positive samples, 6 required more than a single subculture to reveal the presence of salmonellas. Twenty-four positive isolations were made from the selenite broths incubated at 37°C for 24 h and subcultured once only. All faecal samples positive by enrichment were also positive by direct plating. Nine samples seeded by 6 h produced only 1-2 salmonella colonies per plate. Convalescent specimens are, therefore, unlikely to be efficiently diagnosed by early subculture even if it is combined with multiple plating.River Taff samples, after pre-enrichment in buffered peptone water, were enriched in Rappaport's medium at 37°C for 6 h. Subcultures were made to four plates of brilliant green MacConkey agar. A control subculture was made at 24 h to a single plate of brilliant green MacConkey. Although multiple subculture at 6 h improved salmonella isolation, the method was not as efficient as a single plating from the same enrichment culture at 24 h. There was no evidence that subculture at 6 h combined with multiple plating had a value either for faecal examination or for the isolation of salmonellas from sewage-polluted natural water.Samples of other natural waters submitted routinely by Environmental Health Officers were pre-enriched in buffered peptone water before subculture to Rappaport's medium. The enrichment cultures were incubated at 37°C for 24 h and duplicate platings were made to brilliant green MacConkey agar. By double plating at 24 h, a gain of 16 % in salmonella recovery was made. This compares favourably with the gain obtained by subculture from Rappaport after 24 and 48 h incubation. It also saves 24 h in examination time.

A comparison of two modifications of Rappaport's enrichment medium (R25 and RV) for the isolation of salmonellas from sewage polluted natural water

“…Salmonellas are isolated more often at the 24 h subculture with R25 than with RV. Early recovery of salmonellas is an advantage and has prompted several studies in the past (Dixon, 1961) and more recently (Price, 1983;van Schothorst & Renaud, 1983). Time of isolation of salmonellas from enrichment appears to be related to the numbers present in the original inoculum (McCoy, 1962) and also the concentration ofthe selective agent in the fluid medium .…”