When Campylobacter jejuni NCTC 11351 was grown microaerobically in rich medium at 39°C, entry into stationary phase was followed by a rapid decline in viable numbers to leave a residual population of 1% of the maximum number or less. Loss of viability was preceded by sublethal injury, which was seen as a loss of the ability to grow on media containing 0.1% sodium deoxycholate or 1% sodium chloride. Resistance of cells to mild heat stress (50°C) or aeration was greatest in exponential phase and declined during early stationary phase. These results show that C. jejuni does not mount the normal phenotypic stationary-phase response which results in enhanced stress resistance. This conclusion is consistent with the absence of rpoS homologues in the recently reported genome sequence of this species and their probable absence from strain NCTC 11351. During prolonged incubation of C. jejuni NCTC 11351 in stationary phase, an unusual pattern of decreasing and increasing heat resistance was observed that coincided with fluctuations in the viable count. During stationary phase of Campylobacter coli UA585, nonmotile variants and those with impaired ability to form coccoid cells were isolated at high frequency. Taken together, these observations suggest that stationary-phase cultures of campylobacters are dynamic populations and that this may be a strategy to promote survival in at least some strains. Investigation of two spontaneously arising variants (NM3 and SC4) of C. coli UA585 showed that a reduced ability to form coccoid cells did not affect survival under nongrowth conditions.

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When stationary phase batch cultures of Csmpylobacter jejuni were stored in sealed flasks under static conditions, viable numbers declined from 2 x lo9 c.f.u. ml'' to around 103-106 c.f.u. ml'l within 4-6 weeks. When the aged cultures were sparged with a microaerobic gas mixture, there was a rapid increase in viable numbers accompanied by a change from predominantly coccoid t o vibrioid morphology. The most probable number (MPN) technique was used to distinguish resuscitation of injured or dormant cells from multiplication of residual viable cells. MPN estimates using fresh Brucella broth containing 0.2 O/ O mucin revealed that plate counts underestimated the true viable count by up to 23-fold. The experiments clearly demonstrated that a proportion of surviving cells in aged cultures were in an injured or latent state that prevented growth on agar plates. It is possible that the size of this fraction is greater than was demonstrated and that much higher recoveries would be obtained under other recovery conditions. Nevertheless, from presently available evidence, it must be concluded that the size of the latent fraction is quite small and that most of the increase in count that occurs on regassing a spent culture comes from multiplication of residual viable cells.

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The fluorogenic redox indicator 5-cyano-2,3-ditolyltetrazolium chloride (CTC) was compared with the chromogenic p-iodonitrotetrazolium violet (INT) and conventional methods to assess cellular viability. Mild heat treatment was used as well-controlled method for producing non-viable and sub-lethally injured cells. CTC gave an underestimation of the viability of Listeria monocytogenes cells when compared with classical plating methods whereas INT gave an overestimation. However, CTC proved to be a sensitive indicator of uninjured cells. The difference between the total count and the CTC count was equivalent to the injured cell population. The fluorescent formazan formed on reduction of CTC was readily detected with a charge coupled device and cells enumerated automatically using image analysis.