Bacteria swim towards improving conditions by controlling flagellar activity via signals (CheY) sent from chemosensory protein clusters, which respond to changing stimuli. The best studied chemotactic bacterium, E. coli, has one transmembrane chemosensory protein cluster controlling flagellar behaviour. R. sphaeroides has two clusters, one transmembrane and one cytoplasmic. The roles of the two clusters in regulating swimming and chemosensory behaviour are explored here. Newly-developed software was used to measure the effect of deleting or mutating each chemotaxis protein on unstimulated swimming and on the chemosensory response to dynamic change. New behaviours were identified by using much larger sample sizes than previous studies. R. sphaeroides chemotaxis mutants were classified as (i) stoppy unresponsive; (ii) smooth unresponsive or (iii) stoppy inhibited compared to wildtype swimming and chemosensory behaviour. The data showed that the ability to stop during free-swimming is not necessarily connected to the ability to respond to a chemotaxis challenge. The data suggested a new model of connectivity between the two chemosensory pathways. CheY₃ and CheY₄ are phosphorylated by the transmembrane polar cluster in response to external chemoeffector concentrations. CheY₆-P produced by the cytoplasmic cluster is a requirement for chemotaxis, whether or not the polar cluster is able to produce CheY₆-P. CheY₆-P stops the motor, whereas CheY_3,4-P allow smooth swimming. When chemoeffector levels fall, the signals through CheY_3,4 fall, allowing CheY₆-P to bind and stop the motor. As the polar cluster adapts to the fall by the action of the adaptation proteins CheB₁ and CheR₂, the concentration of CheY_3,4-P increases again, to compete with CheY₆-P and allow periods of smooth swimming. Under aerobic conditions, the cytoplasmic cluster controls the basal stopping frequency and does not appear to respond to external chemoeffector changes. The role of the adaptation proteins in resetting the signalling state in R. sphaeroides is unclear, particularly the roles of the proteins associated with the cytoplasmic cluster, CheB₂ and CheR₃. Tandem mass spectrometry was used to identify glutamate and glutamine (EQ) sites on the cytoplasmic R. sphaeroides chemoreceptor TlpT that are deamidated and methylated by the R. sphaeroides adaptation homologues. In E. coli, adaptation sites are usually EQ/EQ pairs. However the sites reported in TlpT vary at the first residue in the pair. Mutation of the putative EQ adaptation sites caused changes in adaptation, suggesting that CheY₆-P levels are controlled and reset by CheB₂ and CheR₃ controlling the adaptation state of TlpT.

Bacterial genomes provide records of the molecular processes associated with emergence and evolution of different bacterial lifestyles. This thesis is based on whole-genome comparisons within the genus Bartonella, an excellent model system for studies of host- and vector-specificity and infection outcome in animal-associated bacteria. The louse-borne human specialist and trench fever agent Bartonella quintana was contrasted to the flea-borne generalist relatives Bartonella henselae and Bartonella grahamii, which cause asymptomatic infection in cat and mouse respectively. While B. henselae is commonly isolated from humans, and causes cat scratch disease, there is only one reported case of B. grahamii human infection. The gene complements of the three species are nested like Russian dolls with the smaller genome (B. quintana) being entirely contained in the medium sized (B. henselae), which in turned is contained in the largest (B. grahamii). Size differences reflect differences in the horizontally and vertically acquired gene content, and in the number of genus- and species- specific genes, owing to differential impact of bacteriophages and plasmids, and to different degrees of genome decay. These processes can be attributed to the three distinct lifestyles. Comparisons with other alpha-proteobacteria suggest that the Bartonella genus as a whole evolved from plant-associated species, and that horizontal transfer, in particular of genes involved in interaction with the host, played a key role in the transition to animal intracellular lifestyle. The long-term genome decay associated with this lifestyle is most advanced in the host-restricted B. quintana. The broad host-range species B. grahamii has the largest genome and the largest proportion of auxiliary DNA of the three, probably because it has access to a larger gene pool. In encodes all the known pathogenicity determinants found in the genomes of B. henselae and B. quintana, suggesting that these genes primarily evolved to facilitate colonization in the reservoir host.

Gene Transfer Agents (GTAs) are phage-like pthesiss that are produced by many alpha proteobacteria in late stationary growth phase and are capable of transferring chromosomal genes (termed "constitutive transduction"). Examination of alpha proteobacterial genomic sequences indicated widespread occurrence of GTA-like elements. The goal of this study was to investigate gene transfer potential of GTAs of marine alpha proteobacteria in culture as well as in natural marine environments. Another goal was to determine the potential of bacterial symbionts from zooxanthellae and coral to genetically transfer beneficial properties between symbionts. Ruegeria mobilis (ID 45A6) was isolated from cultures of the coral endosymbiotic dinoflagellate, Symbiodinium spp. A goal of the research was to determine if GTAs from this isolate have the capability of transferring genes to environmental recipients and have an impact on settlement of coral larvae. Little is known about coral settlement cues, yet there may be contributions from the extensive symbiotic relationship of coral reef-associated bacteria. Several gene transfer experiments in different environments were performed using transformed isolates of Ruegeria mobilis containing a transposon marker gene. Experiments were also performed using GTAs from the Ruegeria mobilis isolate to observe any impact GTAs have on coral larval settlement, using larvae from the brooding coral, Porites astreoides, and from the reef building coral, Montastraea faveolata. Gene transfer frequencies from statistically significant gene transfer experiments resulted in an average of 2.92 × 10-1 (transfer recipients to total viable population). Coral settlement experiments resulted in a statistically significant increase in larval settlement with the addition of GTAs for 80% of the executed experiments. The entire study has demonstrated that GTA-mediated gene exchange is much higher than any other mode of horizontal gene transfer and it has been established that these genes can be exchanged between bacterial taxa. GTAs can also have an impact on coral larval settlement mechanisms that are not yet completely understood. GTA-mediated beneficial gene exchange may be an important driver in adaptation to an evolving planet.