A MIXED CULTURE RECOVERY METHOD INDICATES THAT ENTERIC BACTERIA DO NOT ENTER THE VIABLE BUT NONCULTURABLE STATE

A new method, called the mixed culture recovery (MCR) method, has been developed to determine whether recovery of culturable bacterial cells from a population of largely nonculturable cells is due to resuscitat ion of the nonculturable cells from a viable but nonculturable state o r simply to growth of residual culturable cells. The MCR method addres ses this issue in that it involves the mixing of two easily distinguis hable strains (e.g., lactose positive and negative) in such a way that large numbers of nonculturable cells of both strains are present toge ther with a small number of culturable cells of only one strain, perfo rming a nutrient addition resuscitation procedure, and then plating th e cells to determine whether both cell types are recoverable. In repea ted experiments with strains of Escherichia coli, Klebsiella pneumonia e, Enterococcus faecalis, Enterobacter aerogenes, and Salmonella chole raesuis, only cells of the culturable strain were recovered after appl ication of various resuscitation techniques. These results suggest tha t the nonculturable cells were dead and that the apparent resuscitatio n was merely due to the growth of the remaining culturable cells.