Mutational (genetic) robustness is phenotypic constancy in the face of mutational changes to the genome. Robustness is critical to the understanding of evolution because phenotypically expressed genetic variation is the fuel of natural selection. Nonetheless, the evidence for adaptive evolution of mutational robustness in biological populations is controversial. Robustness should be selectively favored when mutation rates are high, a common feature of RNA viruses. However, selection for robustness may be relaxed under virus co-infection because complementation between virus genotypes can buffer mutational effects. We therefore hypothesized that selection for genetic robustness in viruses will be weakened with increasing frequency of co-infection. To test this idea, we used populations of RNA phage φ6 that were experimentally evolved at low and high levels of co-infection and subjected lineages of these viruses to mutation accumulation through population bottlenecking. The data demonstrate that viruses evolved under high co-infection show relatively greater mean magnitude and variance in the fitness changes generated by addition of random mutations, confirming our hypothesis that they experience weakened selection for robustness. Our study further suggests that co-infection of host cells may be advantageous to RNA viruses only in the short term. In addition, we observed higher mutation frequencies in the more robust viruses, indicating that evolution of robustness might foster less-accurate genome replication in RNA viruses.

Co-infection may be beneficial in large populations of viruses because it permits sexual exchange between viruses that is useful in combating the mutational load. This advantage of sex should be especially substantial when mutations interact through negative epistasis. In contrast, co-infection may be detrimental because it allows virus complementation, where inferior genotypes profit from superior virus products available within the cell. The RNA bacteriophage φ6 features a genome divided into three segments. Co-infection by multiple φ6 genotypes produces hybrids containing reassorted mixtures of the parental segments. We imposed a mutational load on φ6 populations by mixing the wild-type virus with three single mutants, each harboring a deleterious mutation on a different one of the three virus segments. We then contrasted the speed at which these epistatic mutations were removed from virus populations in the presence and absence of co-infection. If sex is a stronger force, we predicted that the load should be purged faster in the presence of co-infection. In contrast, if complementation is more important we hypothesized that mutations would be eliminated faster in the absence of co-infection. We found that the load was purged faster in the absence of co-infection, which suggests that the disadvantages of complementation can outweigh the benefits of sex, even in the presence of negative epistasis. We discuss our results in light of virus disease management and the evolutionary advantage of haploidy in biological populations.

Human hands are an important source of microbial contamination of foods. However, published data on the effectiveness of handwashing and glove use in a foodservice setting are limited. Bacterial transfer through foodservice quality gloves was quantified using nalidixic acid-resistant Enterobacter aerogenes (a nonpathogenic surrogate with attachment characteristics similar to Salmonella). Five transfer rates were determined: chicken to bare hand, chicken to hand through gloves, bare hand to lettuce, hand to lettuce through gloves (with low inoculum on hands), and hand to lettuce through gloves (with high inoculum on hands). At least 30 observations were made for each percent transfer rate using 30 individual volunteers. The logarithm of percent transfer data were then fit to distributions: chicken to bare hand, normal (0.71, 0.42); chicken to hand through gloves, gamma (5.91, 0.40, -5.00); bare hand to lettuce, logistic (1.16, 0.30); hand to lettuce through gloves (low inoculum), normal (0.35, 0.88); hand to lettuce through gloves (high inoculum), normal (-2.52, 0.61). A 0.01% transfer was observed from food to hands and from hands to food when subjects wore gloves and a 10% transfer was observed without a glove barrier. These results indicate that gloves are permeable to bacteria although transfer from hands to food through a glove barrier was less than without a glove barrier. Our results indicate that gloves may reduce both bacterial transfer from food to the hands of foodservice workers and in subsequent transfer from hands back to food.

Many factors have been shown to influence bacterial transfer between surfaces, including surface type, bacterial species, moisture level, pressure, and friction, but the effect of inoculum size on bacterial transfer has not yet been established. Bacterial cross contamination rates during performance of common food service tasks were previously determined in our laboratory using nalidixic acid-resistant Enterobacter aerogenes. Eight different transfer rates were determined, each involving a minimum of 30 volunteers. The influence of source inoculum level on the percentage of bacteria transferred (percent transfer rates) and log 10 CFU per recipient surface was determined using statistical analysis. The effect of inoculum size on transfer rate was highly statistically significant (P < 0.0001) for all transfer rate data combined (352 observations) and for each individual cross contamination rate, except for data on contamination via transfer from chicken to hand through a glove barrier (P ‫؍‬ 0.1643). Where inoculum size on the source was greater, transfer rates were lower, and where inoculum size on the source was less, transfer rates were higher. The negative linear trend was more obvious for activities that had a larger range of inoculum sizes on the source surface. This phenomenon has serious implications for research seeking to determine bacterial cross contamination rates, since the different transfer efficiencies that were previously shown to be associated with certain activities may actually be the result of differing initial inoculum levels. The initial inoculum size on the source and the amount of bacteria transferred must both be considered to accurately determine bacterial transfer rates.Microbial concentration plays an essential role in many microbial systems. It plays an essential role in regulating bioluminescence (8), antibiotic biosynthesis (1), virulence determination (20), catalase activity (6), and initiation of chromosomal replication (29). Bacillus megaterium spores germinate faster when present at higher concentrations (4). The inoculum size of Clostridium botulinum affects time to detection and the fraction of samples that show growth (30). A threshold inoculum size for Listeria monocytogenes to initiate growth at suboptimal conditions has been established (19,23). Whether due to interaction between cells, statistical effects, or sensitivity of microbiological methods, initial inoculum levels can drastically affect experimental results.Many factors that influence the transfer of bacteria from surface to surface have been identified. Type of bacteria (14, 24), source and destination surfaces (5, 10, 24), time postinoculation (26), and moisture level (10, 25) have all been shown to affect cross contamination rates. However, the effect of the initial inoculum level on transfer efficiency has not been established.Research conducted in our laboratory to determine the effectiveness of gloves as a barrier to cross contamination identified inoculum size as a possible factor influencing the percent t...

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