Staphylococcus epidermidis is a normal constituent of the healthy human microflora, but it is also the most common cause of nosocomial infections associated with the use of indwelling medical devices. Isolates from device-associated infections are known for their pronounced phenotypic and genetic variability, and in this study we searched for factors that might contribute to this flexibility. We show that mutator phenotypes, which exhibit elevated spontaneous mutation rates, are rare among both pathogenic and commensal S. epidermidis strains. However, the study revealed that, in contrast to those of commensal strains, the genomes of clinical S. epidermidis strains carry multiple copies of the insertion sequence IS256, while other typical staphylococcal insertion sequences, such as IS257 and IS1272, are distributed equally among saprophytic and clinical isolates. Moreover, detection of IS256 was found to be associated with biofilm formation and the presence of the icaADBC operon as well as with gentamicin and oxacillin resistance in the clinical strains. The data suggest that IS256 is a characteristic element in the genome of multiresistant nosocomial S. epidermidis isolates that might be involved in the flexibility and adaptation of the genome in clinical isolates.Staphylococcus epidermidis is a normal constituent of the healthy human skin and mucosal microflora. In recent decades, however, the bacterium has emerged as a nosocomial multiresistant pathogen and is now the most common cause of deviceassociated infections. Little is known of the factors that have contributed to this development, but the increasing number of immunocompromised patients, the use of indwelling medical devices, and a high selective pressure by antibiotics offer bacteria a novel ecological niche. It is unclear why just staphylococci were able to occupy this niche and by which factors pathogenic S. epidermidis differ from their commensal counterparts. In recent years, it has been shown that the ability to form biofilms on medical devices is a characteristic feature of nosocomial S. epidermidis isolates. Moreover, clinical S. epidermidis isolates exhibit an extraordinarily high phenotypic and genotypic flexibility. Thus, variants of the same parent strain can differ in terms of colony morphology, growth rate, hemolysis, biofilm formation, and antibiotic susceptibility (4, 7). The molecular mechanisms involved in this phenomenon are poorly understood, but it is assumed that the generation of phenotypic and genotypic variants is an evolutionary advantage that helps staphylococci to adapt to changing environmental conditions. The purpose of this study was therefore to search for genetic factors and mechanisms in clinical S. epidermidis that might contribute to this process. Previous studies have shown that staphylococcal biofilm formation is a highly variable factor which is influenced by both regulatory processes and genetic mechanisms such as phase variations, mutations, and chromosomal rearrangements (5,10,26,(32)(33)(34). The observation t...

IS256 is a highly active insertion sequence (IS) element of multiresistant staphylococci and enterococci. Here we show that, in a Staphylococcus epidermidis clinical isolate, as well as in recombinant Staphylococcus aureus and Escherichia coli carrying a single IS256 insertion on a plasmid, IS256 excises as an extrachromosomal circular DNA molecule. First, circles were identified that contained a complete copy of IS256. In this case, the sequence connecting the left and right ends of IS256 was derived from flanking DNA sequences of the parental genetic locus. Second, circle junctions were detected in which one end of IS256 was truncated. Nucleotide sequencing of circle junctions revealed that (i) either end of IS256 can attack the opposite terminus and (ii) the circle junctions vary significantly in size. Upon deletion of the IS256 open reading frame at the 3 end and site-directed mutageneses of the putative DDE motif, circular IS256 molecules were no longer detectable, which implicates the IS256-encoded transposase protein with the circularization of the element.IS256 is an insertion sequence (IS) element of staphylococci and enterococci that was originally identified as a bordering component of the composite aminoglycoside resistance-mediating transposon Tn4001 (2,8,9). IS256, together with 32 other elements, forms a family of ISs that also includes eukaryotic relatives (3). Information about IS256 is very limited (10), despite its wide distribution. Recently, it was demonstrated that the flanking IS256 copies of Tn4001 form tandem dimers and IS circles in an Escherichia coli genetic background (17). In previous studies we have been able to show that IS256 is associated with genetic rearrangements, causing phenotypic changes in its natural host Staphylococcus epidermidis. Multiresistant S. epidermidis as well as Staphylococcus aureus are the most common causes of nosocomial infections associated with implanted biomaterials. In these clinical staphylococcal isolates IS256 proved to be highly active and caused a variety of genetic aberrations, such as reversible gene inactivations, DNA rearrangements, and large chromosomal deletions that affected the expression of virulence-and resistance-associated genes (12,(27)(28)(29)(30). Specifically, biofilm formation, which is one of the major factors in S. epidermidis pathogenesis, is influenced by IS256. The element causes phase variation of biofilm expression in S. epidermidis by inactivation of the icaADBC operon, which encodes enzymes responsible for biofilm synthesis (28). Apparently, the icaC gene represents a hot spot for IS256 insertions. It was shown that the element creates 8-and 7-bp target site duplications during transposition (28; S. H. Cho and W. Ziebuhr, unpublished data). The spontaneous ica:: IS256 insertions are reversible and, accordingly, nucleotide sequencing of biofilm-forming revertants confirmed the precise excision of the element, including the initially duplicated target sequences. In these initial studies, however, the molecular mechanisms th...

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This paper presents the design and control of an actuated bivalve robot, which has been developed to study the burrowing locomotion of bivalves in sediment. The setup consists of a tank filled with sand and water, plastic models of bivalve shells capable of expelling water and an external actuation mechanism simulating the rocking burrowing motion typically used by these animals. The realistic shell shapes have been realized using three-dimensional plotting techniques allowing testing influences of different shell shapes and surface structures (sculptures) on the burrowing efficiency. Based on the experimental setup, the burrowing process has been reproduced. The results show that this setup can be used to identify correlations in the burrowing process. Further experimental work will investigate the influence of factors such as shell shape and sculpture or the motion sequence on the burrowing performance. Keywords: biorobotics; biomimetics; burrowing locomotion; bivalves Actuated Bivalve Robot Study of the Burrowing Locomotion in SedimentAgathe Koller-Hodac, Daniel P. Germann, Alexander Gilgen, Katja Dietrich, Maik Hadorn, Wolfgang Schatz and Peter Eggenberger HotzAbstract-This paper presents the design and control of an actuated bivalve robot, which has been developed to study the burrowing locomotion of bivalves in sediment. The setup consists of a tank filled with sand and water, plastic models of bivalve shells capable of expelling water and an external actuation mechanism simulating the rocking burrowing motion typically used by these animals. The realistic shell shapes have been realized using three-dimensional plotting techniques allowing testing influences of different shell shapes and surface structures (sculptures) on the burrowing efficiency.Based on the experimental setup, the burrowing process has been reproduced. The results show that this setup can be used to identify correlations in the burrowing process. Further experimental work will investigate the influence of factors such as shell shape and sculpture or the motion sequence on the burrowing performance.

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