Autoinducers act as biological timers in Vibrio harveyi

Quorum sensing regulates cell density-dependent phenotypes and involves the synthesis, excretion and detection of so-called autoinducers. Vibrio harveyi strain ATCC BAA-1116 (recently reclassified as Vibrio campbellii), one of the best-characterized model organisms for the study of quorum sensing, produces and responds to three autoinducers. HAI-1, AI-2 and CAI-1 are recognized by different receptors, but all information is channeled into the same signaling cascade, which controls a specific set of genes. Here we examine temporal variations of availability and concentration of the three autoinducers in V. harveyi, and monitor the phenotypes they regulate, from the early exponential to the stationary growth phase in liquid culture. Specifically, the exponential growth phase is characterized by an increase in AI-2 and the induction of bioluminescence, while HAI-1 and CAI-1 are undetectable prior to the late exponential growth phase. CAI-1 activity reaches its maximum upon entry into stationary phase, while molar concentrations of AI-2 and HAI-1 become approximately equal. Similarly, autoinducer-dependent exoproteolytic activity increases at the transition into stationary phase. These findings are reflected in temporal alterations in expression of the luxR gene that encodes the master regulator LuxR, and of four autoinducer-regulated genes during growth. Moreover, in vitro phosphorylation assays reveal a tight correlation between the HAI-1/AI-2 ratio as input and levels of receptor-mediated phosphorylation of LuxU as output. Our study supports a model in which the combinations of autoinducers available, rather than cell density per se, determine the timing of various processes in V. harveyi populations

Combinatorial quorum sensing allows bacteria to resolve their social and physical environment

Quorum sensing (QS) is a cell–cell communication system that controls gene expression in many bacterial species, mediated by diffusible signal molecules. Although the intracellular regulatory mechanisms of QS are often well-understood, the functional roles of QS remain controversial. In particular, the use of multiple signals by many bacterial species poses a serious challenge to current functional theories. Here, we address this challenge by showing that bacteria can use multiple QS signals to infer both their social (density) and physical (mass-transfer) environment. Analytical and evolutionary simulation models show that the detection of, and response to, complex social/physical contrasts requires multiple signals with distinct half-lives and combinatorial (nonadditive) responses to signal concentrations. We test these predictions using the opportunistic pathogen Pseudomonas aeruginosa and demonstrate significant differences in signal decay betweeallyn its two primary signal molecules, as well as diverse combinatorial responses to dual-signal inputs. QS is associated with the control of secreted factors, and we show that secretome genes are preferentially controlled by synergistic “AND-gate” responses to multiple signal inputs, ensuring the effective expression of secreted factors in high-density and low mass-transfer environments. Our results support a new functional hypothesis for the use of multiple signals and, more generally, show that bacteria are capable of combinatorial communication

Heterogeneous Response to a Quorum-Sensing Signal in the Luminescence of Individual Vibrio fischeri

The marine bacterium Vibrio fischeri regulates its bioluminescence through a quorum sensing mechanism: the bacterium releases diffusible small molecules (autoinducers) that accumulate in the environment as the population density increases. This accumulation of autoinducer (AI) eventually activates transcriptional regulators for bioluminescence as well as host colonization behaviors. Although V.fischeri quorum sensing has been extensively characterized in bulk populations, far less is known about how it performs at the level of the individual cell, where biochemical noise is likely to limit the precision of luminescence regulation. We have measured the time-dependence and AI-dependence of light production by individual V.fischeri cells that are immobilized in a perfusion chamber and supplied with a defined concentration of exogenous AI. We use low-light level microscopy to record and quantify the photon emission from the cells over periods of several hours as they respond to the introduction of AI. We observe an extremely heterogeneous response to the AI signal. Individual cells differ widely in the onset time for their luminescence and in their resulting brightness, even in the presence of high AI concentrations that saturate the light output from a bulk population. The observed heterogeneity shows that although a given concentration of quorum signal may determine the average light output from a population of cells, it provides far weaker control over the luminescence output of each individual cell

Identification and characterization of a direct activator of a gene transfer agent

Gene transfer agents (GTAs) are thought to be ancient bacteriophages that have been co-opted into serving their host and can now transfer any gene between bacteria. Production of GTAs is controlled by several global regulators through unclear mechanisms. In Rhodobacter capsulatus, gene rcc01865 encodes a putative regulatory protein that is essential for GTA production. Here, I show that rcc01865 (hereafter gafA) encodes a transcriptional regulator that binds to the GTA promoter to initiate production of structural and DNA packaging components. Expression of gafA is in turn controlled by the pleiotropic regulator protein CtrA and the quorum-sensing regulator GtaR. GafA and CtrA work together to promote GTA maturation and eventual release through cell lysis. Identification of GafA as a direct GTA regulator allows the first integrated regulatory model to be proposed and paves the way for discovery of GTAs in other species that possess gafA homologues

Computed tomography osteoabsorptiometry is reliable for the determination of the subchondral bone mineralization distribution in the rabbit knee

Rabbits are among the most frequently used animals in osteoarthritis research. It is meanwhile accepted that the subchondral bone plate (SBP) plays a key role in the development of osteoarthritis. The most suitable technique for analyzing subchondral bone mineralization is computed tomography osteoabsorptiometry (CT-OAM). Because CT-OAM has not yet been applied to smaller animals, the purpose of this study is to test the reliability of CT-OAM in the rabbit knee. Another important task in animal experiments is the intra- and interindividual difference of the measurement parameters. Our hypothesis is that there is no difference regarding both the position of the density maxima and the bone mineral density (BMD) of the SBP comparing right and left tibial plateaus of rabbits. For evaluating the reliability, a rabbit knee was examined by computed tomography 6 times at weekly intervals. The subchondral mineralization distribution was measured by means of CT-OAM. Positions of the density maxima and BMD of the SBP were determined in a standardized procedure. Furthermore, both parameters were evaluated in 6 female White New Zealand rabbits. Positions of density maxima and BMD in the SBP in left tibial plateaus were compared with right tibial plateaus. The relative coefficient of variation as a parameter for reproducibility was 1.6% for determining the position of the density maxima and 1.2% for measuring the BMD. The positions of density maxima and relative BMD between right and left tibial plateaus varied only about 2% intraindividually, whereas interindividual variance was about 10%. In conclusion, determination of the position of density maxima as well as BMD of the SBP by means of CT-OAM is reliable and reproducible in the rabbit knee. We recommend using the contralateral limb as control, because intraindividual accordance of the mineralization patterns and of the BMD of the SBP was higher than interindividual accordance

Meniscectomy: indications, procedure, outcomes, and rehabilitation

Hermann Anetzberger,1 Christof Birkenmaier,2 Stephan Lorenz3 1Orthopädische Gemeinschaftspraxis am OEZ, Munich, Germany; 2Department of Orthopedics, Ludwig-Maximilian-University Munich, Munich, Germany; 3Department of Orthopedic Sports Medicine, Technical University of Munich, Klinikum rechts der Isar, Munich, Germany Abstract: Meniscal injuries are among the most frequent reasons for knee problems. The goal of this manuscript is to review the biomechanical relevance of the human knee's menisci in relation to surgical indications, surgical techniques, rehabilitation, and outcomes. In order to identify the relevant literature, we performed a PubMed search for the years ranging from 1980–2013 using the following search terms: meniscus; biomechanical function; meniscectomy; meniscal repair; and clinical outcome. The meniscus helps to distribute the forces between the tibial and femoral articular cartilage layers in a load-sharing capacity. Meniscus damage or meniscectomy intuitively leads to an overloading of the cartilage and, hence, to the development of osteoarthrosis. Precise knowledge of meniscal shape and function, of the type of injury, of surgical techniques, as well as of postsurgical rehabilitative care are of decisive importance for an individually-adjusted treatment strategy. Other underlying coexisting knee pathologies also need to be considered. The diagnosis of a meniscal injury is based upon clinical history, physical examination, and imaging studies. The treatment of a meniscal lesion includes conservative, as well as operative, procedures. The goals of surgery are to reduce pain and disability, as well as to preserve meniscal function without causing additional cartilage damage. The resection of meniscal tissue should be restricted to as much as is necessary, and as little as is reasonably possible. Postoperative rehabilitation serves the purpose of improving functional deficits and pain, as well as of restoring a good range of motion and preventing secondary damage. It is the surgical therapy that dictates the aftercare, and with regards to the latter, there are clear differences between meniscectomy and meniscal repair. Keywords: meniscectomy, osteoarthritis, meniscus pathology, meniscus functio

Meniscectomy leads to early changes in the mineralization distribution of subchondral bone plate

PURPOSE: It is generally recognized that the subchondral bone plate (SBP) is involved in development of osteoarthritis (OA). However, the pathophysiological significance is not yet clear. The goal of this study is to investigate the extent of the changes that occur in SBP of the tibial plateau in the early stages of experimental OA. METHODS: Forty-three female rabbits were assigned to 5 experimental (n = 8 each group) and one sham group (n = 3). OA was induced by medial meniscectomy in the right knee, the left knee served as control. 2, 4, 8, 12, and 24 weeks after meniscectomy, cartilage damage was evaluated, and bone mineral density (BMD) and mineralization distribution of the SBP was measured by computed tomography osteoabsorptiometry (CT-OAM). RESULTS: Cartilage damage started 2 weeks after meniscectomy with surface roughening. Cartilage defects increased over time. 24 weeks postoperatively, subchondral bone was exposed. As early as 2 weeks after meniscectomy, BMD in the medial tibial plateau decreased significantly. BMD increased again and reached the values of the non-operated knee 12 weeks postoperatively. In addition, already 4 weeks after meniscectomy a significant shift of the densitiy maximum on the medial tibial plateau, which is normally centrally located toward the margin was observed. CONCLUSIONS: In conclusion, the results of this study contribute to the concept of early involvement of the SBP in the development of OA. The hypothesis that changes in the SBP occur simultaneously to cartilage damage was confirmed