The Pseudomonas quinolone signal (PQS), and its precursor 2-heptyl-4-quinolone (HHQ), play a key role in coordinating virulence in the important cystic fibrosis pathogen Pseudomonas aeruginosa. The discovery of HHQ analogues in Burkholderia and other microorganisms led us to investigate the possibility that these compounds can influence interspecies behaviour. We found that surface-associated phenotypes were repressed in Gram-positive and Gram-negative bacteria as well as in pathogenic yeast in response to PQS and HHQ. Motility was repressed in a broad range of bacteria, while biofilm formation in Bacillus subtilis and Candida albicans was repressed in the presence of HHQ, though initial adhesion was unaffected. Furthermore, HHQ exhibited potent bacteriostatic activity against several Gram-negative bacteria, including pathogenic Vibrio vulnificus. Structure-function analysis using synthetic analogues provided an insight into the molecular properties that underpin the ability of these compounds to influence microbial behaviour, revealing the alkyl chain to be fundamental. Defining the influence of these molecules on microbial-eukaryotic-host interactions will facilitate future therapeutic strategies which seek to combat microorganisms that are recalcitrant to conventional antimicrobial agents.

Signal-mediated interactions between the human opportunistic pathogens Pseudomonas aeruginosa and Candida albicans affect virulence traits in both organisms. Phenotypic studies revealed that bacterial supernatant from four P. aeruginosa strains strongly reduced the ability of C. albicans to form biofilms on silicone. This was largely a consequence of inhibition of biofilm maturation, a phenomenon also observed with supernatant prepared from non-clinical bacterial species. The effects of supernatant on biofilm formation were not mediated via interference with the yeast-hyphal morphological switch and occurred regardless of the level of homoserine lactone (HSL) produced, indicating that the effect is HSL-independent. A transcriptome analysis to dissect the effects of the P. aeruginosa supernatants on gene expression in the early stages of C. albicans biofilm formation identified 238 genes that exhibited reproducible changes in expression in response to all four supernatants. In particular, there was a strong increase in the expression of genes related to drug or toxin efflux and a decrease in expression of genes associated with adhesion and biofilm formation. Furthermore, expression of YWP1, which encodes a protein known to inhibit biofilm formation, was significantly increased. Biofilm formation is a key aspect of C. albicans infections, therefore the capacity of P. aeruginosa to antagonize this has clear biomedical implications.

The rice blast fungus Magnaporthe grisea develops specialized infection structures known as appressoria, which develop enormous turgor pressure to bring about plant infection. Turgor is generated by accumulation of compatible solutes, including glycerol, which is synthesized in large quantities in the appressorium. Glycogen, trehalose and lipids represent the most abundant storage products in M. grisea conidia. Trehalose and glycogen are rapidly degraded during conidial germination and it is known that trehalose synthesis is required for virulence of the fungus. Lipid bodies are transported to the developing appressoria and degraded at the onset of turgor generation, in a process that is cAMP-dependent. A combined biochemical and genetic approach is being used to dissect the process of turgor generation in the rice blast fungus.

Periodontal disease (PD) is a significant problem in dogs affecting between 44% and 63.6% of the population. The main etiological agent for PD is plaque, a microbial biofilm that colonizes teeth and causes inflammation of the gingiva. Understanding how this biofilm initiates on the tooth surface is of central importance in developing interventions against PD. Although the stages of plaque development on human teeth have been well characterized little is known about how canine plaque develops. Recent studies of the canine oral microbiome have revealed distinct differences between the canine and human oral environments and the bacterial communities they support, particularly with respect to healthy plaque. These differences mean knowledge about the nature of plaque formation in humans may not be directly translatable to dogs. The aim of this study was to identify the bacterial species important in the early stages of canine plaque formation in vivo and then use isolates of these species in a laboratory biofilm model to develop an understanding of the sequential processes which take place during the initial colonization of enamel. Supra-gingival plaque samples were collected from 12 dogs at 24 and 48 hour time points following a full mouth descale and polish. Pyrosequencing of the 16S rDNA identified 134 operational taxonomic units after statistical analysis. The species with the highest relative abundance were Bergeyella zoohelcum, Neisseria shayeganii and a Moraxella species. Streptococcal species, which tend to dominate early human plaque biofilms, had very low relative abundance. In vitro testing of biofilm formation identified five primary colonizer species, three of which belonged to the genus Neisseria. Using these pioneer bacteria as a starting point, viable two and three species communities were developed. Combining in vivo and in vitro data has led us to construct novel models of how the early canine plaque biofilm develops.

Background: Microbiota from different niches within the canine oral cavity were profiled and compared. Supragingival plaque and stimulated saliva, were collected alongside samples from the buccal and tongue dorsum mucosa, from 14 Labrador retrievers at three timepoints within a 1 month timeframe. The V3-V4 region of the 16S rRNA gene was sequenced via Illumina MiSeq. Results: Supragingival plaque microbiota had the highest bacterial diversity and the largest number of significant differences in individual taxa when compared to the other oral niches. Stimulated saliva exhibited the highest variability in microbial composition between dogs, yet the lowest bacterial diversity amongst all the niches. Overall, the bacteria of the buccal and tongue dorsum mucosa were most similar. Conclusions: The bacterial community profiles indicated three discrete oral niches: soft tissue surfaces (buccal and tongue dorsum mucosa), hard tissue surface (supragingival plaque) and saliva. The ability to distinguish the niches by their microbiota signature offers the potential for microbial biomarkers to be identified in each unique niche for diagnostic use.

Periodontal disease is a frequent problem seen in veterinary practices. Primary‐care veterinary practices, where diagnosis of periodontal disease is predominantly based on visual oral assessment of conscious dogs, report an average prevalence of 9.3 to 18.2% within the dog population. Detailed examinations of anaesthetised dogs report much higher prevalence of between 44 and 100%. The prevalence and severity of periodontitis varies between sizes of dogs, breeds and individuals. The lack of robust evaluation of these variances opens the opportunity for a review of the topic. A literature search was therefore undertaken to determine the frequency and extent (severity and oral location) of periodontal disease in the dog population. Both clinical and retrospective data based surveys of veterinary records, show there is an increased likelihood of developing periodontitis as age increases and with bodyweight decreases. The maxillary and mandibular incisors, fourth premolars and first molars are likely to develop the disease first although in some breeds the canines may also be involved. Poor oral care is a significant risk factor for the development of periodontal disease although diet, behaviour, environment and genetics are also likely to play a role. The information provided in this review can be used by veterinarians to improve the effectiveness of their diagnosis, treatments and interventions by focussing their efforts on the sizes, breeds and individuals most at risk. Given the potential link between periodontal disease and systemic health, veterinarians have the opportunity to positively impact the quality of life of a large number of dogs and therefore their owners.

The filamentous fungus Magnaporthe oryzae is the causal agent of rice blast disease. Here we show that glycogen metabolic genes play an important role in plant infection by M. oryzae. Targeted deletion of AGL1 and GPH1, which encode amyloglucosidase and glycogen phosphorylase, respectively, prevented mobilisation of glycogen stores during appressorium development and caused a significant reduction in the ability of M. oryzae to cause rice blast disease. By contrast, targeted mutation of GSN1, which encodes glycogen synthase, significantly reduced the synthesis of intracellular glycogen, but had no effect on fungal pathogenicity. We found that loss of AGL1 and GPH1 led to a reduction in expression of TPS1 and TPS3, which encode components of the trehalose-6-phosphate synthase complex, that acts as a genetic switch in M. oryzae. Tps1 responds to glucose-6-phosphate levels and the balance of NADP/NADPH to regulate virulence-associated gene expression, in association with Nmr transcriptional inhibitors. We show that deletion of the NMR3 transcriptional inhibitor gene partially restores virulence to a Δagl1Δgph1 mutant, suggesting that glycogen metabolic genes are necessary for operation of the NADPH-dependent genetic switch in M. oryzae.

Plant disease resistance (R) gene products recognize pathogen avirulence (Avr) gene products and induce defense responses. It is not known if an R gene can function in different plant families, however. The Arabidopsis thaliana R genes RPW8.1 and RPW8.2 confer resistance to the powdery mildew pathogens Erysiphe orontii, E. cichoracearum, and Oidium lycopersici, which also infect plants from other families. We produced transgenic Nicotiana tabacum, N. benthamiana, and Lycopersicon esculentum plants containing RPW8.1 and RPW8.2. Transgenic N. tabacum plants had increased resistance to E. orontii and O. lycopersici, transgenic N. benthamiana plants had increased resistance to E. cichoracearum, but transgenic L. esculentum plants remained susceptible to these pathogens. The defense responses induced in transgenic N. tabacum and N. benthamiana were similar to those mediated by RPW8.1 and RPW8.2 in Arabidopsis. Apparently, RPW8.1 and RPW8.2 could be used to control powdery mildew diseases of plants from other families.