Spatiotemporal modelling of CheY complexes in Escherichia coli chemotaxis.

The chemotaxis pathway of Escherichia coli is one of the best studied and modelled biological signalling pathways. Here we extend existing modelling approaches by explicitly including a description of the formation and subcellular localization of intermediary complexes in the phosphotransfer pathway. The inclusion of these complexes shows that only about 60% of the total output response regulator (CheY) is uncomplexed at any moment and hence free to interact with its target, the flagellar motor. A clear strength of this model is its ability to predict the experimentally observable subcellular localization of CheY throughout a chemotactic response. We have found good agreement between the model output and experimentally determined CheY localization patterns.

Overview of mathematical approaches used to model bacterial chemotaxis II: bacterial populations.

We review the application of mathematical modeling to understanding the behavior of populations of chemotactic bacteria. The application of continuum mathematical models, in particular generalized Keller-Segel models, is discussed along with attempts to incorporate the microscale (individual) behavior on the macroscale, modeling the interaction between different species of bacteria, the interaction of bacteria with their environment, and methods used to obtain experimentally verified parameter values. We allude briefly to the role of modeling pattern formation in understanding collective behavior within bacterial populations. Various aspects of each model are discussed and areas for possible future research are postulated.

Overview of mathematical approaches used to model bacterial chemotaxis I: the single cell.

Mathematical modeling of bacterial chemotaxis systems has been influential and insightful in helping to understand experimental observations. We provide here a comprehensive overview of the range of mathematical approaches used for modeling, within a single bacterium, chemotactic processes caused by changes to external gradients in its environment. Specific areas of the bacterial system which have been studied and modeled are discussed in detail, including the modeling of adaptation in response to attractant gradients, the intracellular phosphorylation cascade, membrane receptor clustering, and spatial modeling of intracellular protein signal transduction. The importance of producing robust models that address adaptation, gain, and sensitivity are also discussed. This review highlights that while mathematical modeling has aided in understanding bacterial chemotaxis on the individual cell scale and guiding experimental design, no single model succeeds in robustly describing all of the basic elements of the cell. We conclude by discussing the importance of this and the future of modeling in this area.

TlpC, a novel chemotaxis protein in Rhodobacter sphaeroides, localizes to a discrete region in the cytoplasm.

TlpC is encoded in the second chemotaxis operon of Rhodobacter sphaeroides. This protein shows some homology to membrane-spanning chemoreceptors of many bacterial species but, unlike these, is essential for R. sphaeroides chemotaxis to all compounds tested. Genomic replacement of tlpC with a C-terminal gfp fusion demonstrated that TlpC localized to a discrete cluster within the cytoplasm. Immunogold electron microscopy also showed that TlpC localized to a cytoplasmic electron-dense region. Correct TlpC-GFP localization depended on the downstream signalling proteins, CheW3, CheW4 and CheA2, and was tightly linked to cell division. Newly divided cells contained a single cluster but, as the cell cycle progressed, a second cluster appeared close to the initial cluster. As elongation continued, these clusters moved apart so that, on septation, each daughter cell contained a single TlpC cluster. The data presented suggest that TlpC is either a cytoplasmic chemoreceptor responding to or integrating global signals of metabolic state or a novel and essential component of the chemotaxis signalling pathway. These data also suggest that clustering is essential for signalling and that a mechanism may exist for targeting and localizing proteins within the bacterial cytoplasm.

CheR- and CheB-dependent chemosensory adaptation system of Rhodobacter sphaeroides.

Rhodobacter sphaeroides has multiple homologues of most of the Escherichia coli chemotaxis genes, organized in three major operons and other, unlinked, loci. These include cheA(1) and cheR(1) (che Op(1)) and cheA(2), cheR(2), and cheB(1) (che Op(2)). In-frame deletions of these cheR and cheB homologues were constructed and the chemosensory behaviour of the resultant mutants examined on swarm plates and in tethered cell assays. Under the conditions tested, CheR(2) and CheB(1) were essential for normal chemotaxis, whereas CheR(1) was not. cheR(2) and cheB(1), but not cheR(1), were also able to complement the equivalent E. coli mutants. However, none of the proteins were required for the correct polar localization of the chemoreceptor McpG in R. sphaeroides. In E. coli, CheR binds to the NWETF motif on the high-abundance receptors, allowing methylation of both high- and low-abundance receptors. This motif is not contained on any R. sphaeroides chemoreceptors thus far identified, although 2 of the 13 putative chemoreceptors, McpA and TlpT, do have similar sequences. This suggests that CheR(2) either interacts with the NWETF motif of E. coli methyl-accepting chemotaxis proteins (MCPs), even though its native motif may be slightly different, or with another conserved region of the MCPs. Methanol release measurements show that R. sphaeroides has an adaptation system that is different from that of Bacillus subtilis and E. coli, with methanol release measurable on the addition of attractant but not on its removal. Intriguingly, CheA(2), but not CheA(1), is able to phosphorylate CheB(1), suggesting that signaling through CheA(1) cannot initiate feedback receptor adaptation via CheB(1)-P.

Fine tuning bacterial chemotaxis: analysis of Rhodobacter sphaeroides behaviour under aerobic and anaerobic conditions by mutation of the major chemotaxis operons and cheY genes.

Rhodobacter sphaeroides chemotaxis is significantly more complex than that of enteric bacteria. Rhodobacter sphaeroides has multiple copies of chemotaxis genes (two cheA, one cheB, two cheR, three cheW, five cheY but no cheZ), controlling a single 'stop-start' flagellum. The growth environment controls the level of expression of different groups of genes. Tethered cell analysis of mutants suggests that CheY(4) and CheY(5) are the motor-binding response regulators. The histidine protein kinase CheA(2) mediates an attractant ('normal') response via CheY(4), while CheA(1) and CheY(5) appear to mediate a repellent ('inverted') response. CheY(3) facilitates signal termination, possibly acting as a phosphate sink, although CheY(1) and CheY(2) can substitute. The normal and inverted responses may be initiated by separate sets of chemoreceptors with their relative strength dependent on growth conditions. Rhodobacter sphaeroides may use antagonistic responses through two chemosensory pathways, expressed at different levels in different environments, to maintain their position in a currently optimum environment. Complex chemotaxis systems are increasingly being identified and the strategy adopted by R.sphaeroides may be common in the bacterial kingdom.

Identification of a fourth cheY gene in Rhodobacter sphaeroides and interspecies interaction within the bacterial chemotaxis signal transduction pathway.

The Escherichia coli chemotaxis signal transduction pathway has: CheA, a histidine protein kinase; CheW, a linker between CheA and sensory proteins; CheY, the effector; and CheZ, a signal terminator. Rhodobacter sphaeroides has multiple copies of these proteins (2 x CheA, 3 x CheW and 3 x CheY, but no CheZ). In this study, we found a fourth cheY and expressed these R. sphaeroides proteins in E. coli. CheA2 (but not CheA1) restored swarming to an E. coli cheA mutant (RP9535). CheW3 (but not CheW2) restored swarming to a cheW mutant of E. coli (RP4606). R. sphaeroides CheYs did not affect E. coli lacking CheY, but restored swarming to a cheZ strain (RP1616), indicating that they can act as signal terminators in E. coli. An E. coli CheY, which is phosphorylated but cannot bind the motor (CheY109KR), was expressed in RP1616 but had no effect. Overexpression of CheA2, CheW2, CheW3, CheY1, CheY3 and CheY4 inhibited chemotaxis of wild-type E. coli (RP437) by increasing its smooth-swimming bias. While some R. sphaeroides proteins restore tumbling to smooth-swimming E. coli mutants, their activity is not controlled by the chemosensory receptors. R. sphaeroides possesses a phosphorelay cascade compatible with that of E. coli, but has additional incompatible homologues.

Dealing with infectious and parasitic diseases in safari parks, roadside menageries, exotic animal auctions and rehabilitation centres.

Safari parks, roadside menageries, exotic animal auctions and wildlife rehabilitation centres exist world-wide, and serve as sites of wild animal concentrations. When these animals are brought together, the potential is great for the spread and eventual outbreak of many different infectious and parasitic diseases. It is therefore necessary that procedures be developed in these facilities to minimize the risk of occurrence and spread of these diseases. This will require financial resources for professional personnel, diagnostic testing and appropriate facility design.