Potentially pathogenic bacteria, such as Escherichia coli and Vibrio cholerae, become non-culturable during stasis. The analysis of such cells has been hampered by difficulties in studying bacterial population heterogeneity. Using in situ detection of protein oxidation in single E. coli cells, and using a density-gradient centrifugation technique to separate culturable and non-culturable cells, we show that the proteins in non-culturable cells show increased and irreversible oxidative damage, which affects various bacterial compartments and proteins. The levels of expression of specific stress regulons are higher in non-culturable cells, confirming increased defects relating to oxidative damage and the occurrence of aberrant, such as by amino-acid misincorporation, proteins. Our data suggest that non-culturable cells are produced due to stochastic deterioration, rather than an adaptive programme, and pinpoint oxidation management as the 'Achilles heel' of these cells.

Objective. To compare quality of life (QOL) scores 3 and 10 years after total hip arthroplasty (THA) or total knee arthroplasty (TKA) for osteoarthritis with QOL scores in a general population, and to determine factors associated with QOL after surgery. Methods. Data were obtained from 2 multicenter cohorts of patients with THA or TKA: 232 patients were recruited during 2003 (3-year cohort) and 221 patients were recruited during 1994 (10-year cohort). Preoperative data (QOL, radiograph results) and followup data (demographics, comorbidities, pain locations, environmental factors, and QOL) were collected. QOL data for the general population were obtained from a 2003 population-based survey. Results. A total of 195 and 89 patients for the 3-and 10-year cohorts, respectively, were followed up; the mean age at followup was 73 years. For both of the cohorts, physical functioning and role-physical or role-emotional QOL scores were lower than those for a general population with comparable age. Scores for pain, mental health, and social dimensions were lower than those for the reference population for only the 10-year cohort. For both cohorts, increased number of comorbidities, painful locations other than THA or TKA location, and unfavorable environmental factors were associated with impaired QOL. Low preoperative QOL scores were predictive of impaired QOL at followup for only the 3-year cohort. Conclusion. THA or TKA can improve QOL, but the benefits may be time limited. Addressing environmental factors and treating comorbidities and pain in locations other than the arthroplasty location could have mid-and long-term effects on the QOL of patients with THA or TKA.

We have investigated the first events that occur when exponentially grown cells are transferred from a liquid medium (Luria-Bertani [LB]) to a solid medium (LB agar [LBA]). We observed an initial lag phase of 180 min for the wild type MG1655 without any apparent growth. This lack of growth was independent of the bacterial physiological state (either the stationary or the exponential phase), the solid medium composition, or the number of cells on the plate, but it was dependent on the bacterial genotype. Using lacZ-reporter fusions and two-dimensional electrophoresis analysis, we observed that when cells from exponential-phase cultures were plated on LBA, several global regulons, like heat shock regulons (RpoH, RpoE, CpxAR) and oxidative-stress regulons (SoxRS, OxyR, Fur), were immediately induced. Our results indicate that in order to grow on plates, bacteria must not only adapt to new conditions but also perceive a real stress.Growth and division form the foundation of standard microbiological methods for testing samples for viable bacteria, and viability is equated with culturability. By consequence, plate count remains the method of choice for obtaining a "total viable count," i.e., the total number of bacteria capable of yielding a population discernible by the observer, usually a visible colony on the surface of a nutrient agar plate. However, it has been proposed that some readily culturable species of bacteria, when subjected to starvation or other stress (5,6,14,22), may enter a long-term survival state in which they are imperceptible by culturability tests but in which they have characteristics compatible with but indirectly related to viability, like conservation of membrane integrity or metabolic activity (24). These cells have been named "viable but nonculturable" (24). It is expected that, for readily culturable species of bacteria, viable but nonculturable cells could be capable of regaining culturability, which has frequently generated a sharp debate (3,7,11).With the plate count method, microbiologists have made one important assumption, that this method has no deleterious effects on bacteria. Since the 1950s, however, it has been reported that apparently dead cells can be reactivated when scavengers of reactive oxygen species, naturally produced during aerobic respiration, are added to agar plates (11, 12, 16-18, 23, 26). These cells have been named "injured cells." Injury in bacteria is defined as an increased sensitivity to components of growth media that are not normally inhibitory (16,23). The injured state is transient, resulting from cumulative cellular damage, and can be reversed under appropriate conditions to enable the injured cells to resume growth. For instance, various stresses, like starvation, hypochlorous acid, heat shock, and desiccation, may leave cells in a vulnerable physiological state in which atmospheric oxygen, during the recovery period, increases the toxic effect of the primary stressor, which could in part explain the phenomenon of cells being viable but nonculturable...

In previous experiments we were able to separate, using a nondestructive separation technique, culturable and nonculturable bacteria, from a Luria-Bertani (LB) medium culture of Escherichia coli incubated for 48 h. We observed in the nonculturable bacterial population an increase in oxidative damage and up-induction of most defenses against reactive oxygen species (ROS), along with a decrease in cytoplasmic superoxide dismutases. In this study, using the same separation technique, we separated into two subpopulations a 10-h LB medium culture containing only culturable bacteria. For the first time, we succeeded in associating physical separation with physiological differences. Although the levels of defense against ROS (RpoS, RpoH, OxyR, and SoxRS regulons) and oxidative damage (carbonyl contents) were apparently the same, we found that bacteria in one subpopulation were more sensitive to LB medium starvation and to various stresses, such as phosphate buffer starvation, heat shock, and hydrogen peroxide exposure. Based on these results, we suggest that these physiological differences reflect uncharacterized bacterial modifications which do not directly involve defenses against ROS.Biological aging could be defined as the gradual decline in the capacity of an organism to resist stress, damage, and disease. In 1956, Denham Harman (10) postulated that this ubiquitous progressive decay in the functional capacity of aging eukaryotes is a consequence of the accumulation of oxidative damage caused by reactive oxygen species (ROS) (12); this was called the free radical theory (10). A small percentage of oxygen is chemically reduced by addition of single electrons, and the products are sequentially converted into ROS, including the superoxide anion, hydrogen peroxide, and the hydroxyl radical (8). ROS have been shown to cause molecular damage relatively indiscriminately to proteins, lipids, and nucleic acids (3, 9).In cells of prokaryotes, such as Escherichia coli, entering a nonproliferating state (stationary phase) due to nutrient depletion, the bacteria gradually lose the ability to divide and reproduce (21). Similar to eukaryotes, the life span of a starved bacterium appears to be limited by the cell's ability to combat ROS. Indeed, Dukan and Nyström demonstrated the existence of an accumulation of oxidized proteins during starvation of an E. coli population (6). Moreover, the life span of growtharrested wild-type E. coli can be increased Ͼ100% by omitting oxygen during stasis (7). This process has been referred to as conditional senescence elicited by growth arrest (17). Given that one of the criteria for defining senescence is an increase in the mortality rate over time (12), it appears that prokaryotes such as E. coli also senesce (20). More recently, using an ultracentrifugation separation technique, we (4) isolated a nonculturable subpopulation from a Luria-Bertani (LB) medium culture of E. coli incubated for 48 h. We suggested that the main reason for the loss of culturability observed after 48 h was a decrease ...